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1
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Forgioni A, Watanabe M, Goto R, Harada T, Ota T, Shimamura T, Taketomi A. Anti-Inflammatory Effects of Ex Vivo-Generated Donor Antigen-Specific Immunomodulatory Cells on Pancreatic Islet Transplantation. Cell Transplant 2025; 34:9636897251317887. [PMID: 39981681 PMCID: PMC11843686 DOI: 10.1177/09636897251317887] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/15/2024] [Revised: 01/01/2025] [Accepted: 01/13/2025] [Indexed: 02/22/2025] Open
Abstract
Pancreatic islet transplantation (PITx) is a promising treatment option for patients with type 1 diabetes mellitus. Previously, we demonstrated that therapy with alloantigen-specific immunomodulatory cells (IMCs) generated ex vivo in the presence of anti-CD80 and CD86 monoclonal antibodies (mAbs), successfully induced tolerance following clinical liver transplantation. To extend IMC therapy to PITx, it is crucial to address the strong inflammatory and innate immune responses that occur immediately after PITx. In this study, we investigated the efficacy of IMCs in modulating macrophage activation and mitigating inflammatory damage of pancreatic islets. IMCs were induced using mouse splenocytes in the presence of anti-mouse anti-CD80 (RM80) and anti-CD86 (GL-1) mAbs. IMCs exerted donor-specific immunosuppressive effects in a mixed lymphocyte reaction. During lipopolysaccharide (LPS) stimulation, the addition of IMCs suppressed conversion to the M1 phenotype and promoted a shift toward the M2 phenotype, particularly under direct cell-cell contact conditions. Nitric oxide production, a hallmark of M1 polarized macrophages, was significantly reduced in LPS-stimulated RAW264 macrophages by IMC treatment. These findings were associated with reduced secretion of pro-inflammatory cytokines, tumoral necrosis factor α, and interleukin-6, and increased interleukin-10 production by macrophages. IMCs effectively prevented macrophage-mediated islet destruction after 12 h of co-culture with LPS-stimulated macrophages and significantly inhibited macrophage migration toward allogeneic islets in vitro. Intraportal co-infusion of IMCs with syngeneic islets in a mouse PITx model resulted in reduced messenger RNA (mRNA) expression of pro-inflammatory cytokines in the recipient liver. Immunohistochemical staining revealed a significantly lower number of F4/80+ macrophages at the transplantation site in IMCs-treated mice. These results demonstrate that IMCs modulate macrophage polarization, promoting a shift toward the M2 phenotype and protecting islets from macrophage-mediated damage. These effects combined with its intrinsic donor antigen-specific immunosuppressive capacity make IMC therapy a promising strategy for improving outcomes after PITx.
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Affiliation(s)
- Agustina Forgioni
- Department of Gastroenterological Surgery I, Hokkaido University, Sapporo, Japan
| | - Masaaki Watanabe
- Department of Transplant Surgery, Hokkaido University, Sapporo, Japan
| | - Ryoichi Goto
- Department of Gastroenterological Surgery I, Hokkaido University, Sapporo, Japan
| | - Takuya Harada
- Department of Gastroenterological Surgery I, Hokkaido University, Sapporo, Japan
| | - Takuji Ota
- Department of Gastroenterological Surgery I, Hokkaido University, Sapporo, Japan
| | - Tsuyoshi Shimamura
- Department of Organ Transplantation Medicine, Hokkaido University Hospital, Sapporo, Japan
| | - Akinobu Taketomi
- Department of Gastroenterological Surgery I, Hokkaido University, Sapporo, Japan
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Katano T, Inagaki A, Imura T, Yamana H, Saito R, Endo Kumata Y, Suzuki S, Hagiwara Y, Ohashi K, Watanabe K, Tabata Y, Goto M. A novel approach for hepatocyte transplantation at the liver surface. Cell Transplant 2025; 34:9636897251329308. [PMID: 40208805 PMCID: PMC12032460 DOI: 10.1177/09636897251329308] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/17/2024] [Revised: 03/02/2025] [Accepted: 03/05/2025] [Indexed: 04/12/2025] Open
Abstract
Hepatocyte transplantation (HTx) is a promising alternative to liver transplantation; however, poor engraftment remains a major challenge. Although co-transplantation with adipose tissue-derived stromal cells (ADSCs) or islets improves engraftment, exposure of these cells to the portal vein enhances innate immune responses, resulting in a significant loss of hepatocytes. Therefore, we investigated HTx at the liver surface as a novel approach that does not involve the portal vein. Hepatocytes were transplanted onto the liver surface of syngeneic analbuminemic rats with or without ADSCs and/or islets. Serum albumin levels and immunohistochemical staining of the transplanted hepatocytes were evaluated. Hepatocyte engraftment was compared between the liver surface and intraportal groups. To examine the detailed mechanisms behind co-transplantation, co-cultured supernatants were analyzed using multiplex assays, and inhibition tests using neutralizing antibodies were performed. Results showed that islet and ADSC co-transplantation markedly enhanced hepatocyte engraftment at the liver surface (P < 0.01), and its efficiency was comparable to that of intraportal transplantation (P = 0.35). In the co-transplantation group, cells were not necessarily in proximity, suggesting that humoral factors are important. In an in vitro study, hepatocyte function was significantly improved by co-culturing with islets and ADSCs (P < 0.01). Multiplex assays and inhibition tests revealed several important humoral factors, most notably insulin, which promoted hepatocyte engraftment. These findings suggest that HTx at the liver surface, together with crucial factors, may be a novel alternative strategy for intraportal transplantation.
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Affiliation(s)
- Takumi Katano
- Division of Transplantation and Regenerative Medicine, Tohoku University School of Medicine, Sendai, Japan
| | - Akiko Inagaki
- Division of Transplantation and Regenerative Medicine, Tohoku University School of Medicine, Sendai, Japan
| | - Takehiro Imura
- Division of Transplantation and Regenerative Medicine, Tohoku University School of Medicine, Sendai, Japan
| | - Hiroki Yamana
- Department of Surgery, Tohoku University School of Medicine, Sendai, Japan
| | - Ryusuke Saito
- Department of Surgery, Tohoku University School of Medicine, Sendai, Japan
| | - Yukiko Endo Kumata
- Department of Surgery, Tohoku University School of Medicine, Sendai, Japan
| | - Shoki Suzuki
- Department of Surgery, Tohoku University School of Medicine, Sendai, Japan
| | - Yoshiya Hagiwara
- Department of Surgery, Tohoku University School of Medicine, Sendai, Japan
| | - Kazuo Ohashi
- Graduate School and School of Pharmaceutical Sciences, Osaka University, Suita, Japan
| | - Kimiko Watanabe
- Division of Transplantation and Regenerative Medicine, Tohoku University School of Medicine, Sendai, Japan
| | - Yasuhiko Tabata
- Department of Plastic and Reconstructive Surgery, Graduate School of Medicine, Kyoto University, Kyoto, Japan
| | - Masafumi Goto
- Division of Transplantation and Regenerative Medicine, Tohoku University School of Medicine, Sendai, Japan
- Department of Surgery, Tohoku University School of Medicine, Sendai, Japan
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Shi H, Ding Y, Sun P, Lv Z, Wang C, Ma H, Lu J, Yu B, Li W, Wang C. Chemical approaches targeting the hurdles of hepatocyte transplantation: mechanisms, applications, and advances. Front Cell Dev Biol 2024; 12:1480226. [PMID: 39544361 PMCID: PMC11560891 DOI: 10.3389/fcell.2024.1480226] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/13/2024] [Accepted: 10/21/2024] [Indexed: 11/17/2024] Open
Abstract
Hepatocyte transplantation (HTx) has been a novel cell-based therapy for severe liver diseases, as the donor livers for orthotopic liver transplantation are of great shortage. However, HTx has been confronted with two main hurdles: limited high-quality hepatocyte sources and low cell engraftment and repopulation rate. To cope with, researchers have investigated on various strategies, including small molecule drugs with unique advantages. Small molecules are promising chemical tools to modulate cell fate and function for generating high quality hepatocyte sources. In addition, endothelial barrier, immune responses, and low proliferative efficiency of donor hepatocytes mainly contributes to low cell engraftment and repopulation rate. Interfering these biological processes with small molecules is beneficial for improving cell engraftment and repopulation. In this review, we will discuss the applications and advances of small molecules in modulating cell differentiation and reprogramming for hepatocyte resources and in improving cell engraftment and repopulation as well as its underlying mechanisms.
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Affiliation(s)
- Huanxiao Shi
- Department of Cell Biology, Naval Medical University, Shanghai, China
| | - Yi Ding
- Experimental Teaching Center, Naval Medical University, Shanghai, China
| | - Pingxin Sun
- Department of Cell Biology, Naval Medical University, Shanghai, China
| | - Zhuman Lv
- Department of Cell Biology, Naval Medical University, Shanghai, China
| | - Chunyan Wang
- Department of Cell Biology, Naval Medical University, Shanghai, China
| | - Haoxin Ma
- Department of Cell Biology, Naval Medical University, Shanghai, China
| | - Junyu Lu
- Department of Cell Biology, Naval Medical University, Shanghai, China
| | - Bing Yu
- Department of Cell Biology, Naval Medical University, Shanghai, China
| | - Wenlin Li
- Department of Cell Biology, Naval Medical University, Shanghai, China
- Shanghai Key Laboratory of Cell Engineering, Naval Medical University, Shanghai, China
| | - Chao Wang
- Department of Cell Biology, Naval Medical University, Shanghai, China
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Liu S, Yang H, Heng X, Yao L, Sun W, Zheng Q, Wu Z, Chen H. Integrating Metabolic Oligosaccharide Engineering and SPAAC Click Chemistry for Constructing Fibrinolytic Cell Surfaces. ACS APPLIED MATERIALS & INTERFACES 2024; 16:35874-35886. [PMID: 38954798 DOI: 10.1021/acsami.4c07619] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 07/04/2024]
Abstract
To effectively solve the problem of significant loss of transplanted cells caused by thrombosis during cell transplantation, this study simulates the human fibrinolytic system and combines metabolic oligosaccharide engineering with strain-promoted azide-alkyne cycloaddition (SPAAC) click chemistry to construct a cell surface with fibrinolytic activity. First, a copolymer (POL) of oligoethylene glycol methacrylate (OEGMA) and 6-amino-2-(2-methylamido)hexanoic acid (Lys) was synthesized by reversible addition-fragmentation chain transfer (RAFT) copolymerization, and the dibenzocyclooctyne (DBCO) functional group was introduced into the side chain of the copolymer through an active ester reaction, resulting in a functionalized copolymer DBCO-PEG4-POL with ε-lysine ligands. Then, azide functional groups were introduced onto the surface of HeLa model cells through metabolic oligosaccharide engineering, and DBCO-PEG4-POL was further specifically modified onto the surface of HeLa cells via the SPAAC "click" reaction. In vitro investigations revealed that compared with unmodified HeLa cells, modified cells not only resist the adsorption of nonspecific proteins such as fibrinogen and human serum albumin but also selectively bind to plasminogen in plasma while maintaining good cell viability and proliferative activity. More importantly, upon the activation of adsorbed plasminogen into plasmin, the modified cells exhibited remarkable fibrinolytic activity and were capable of promptly dissolving the primary thrombus formed on their surfaces. This research not only provides a novel approach for constructing transplantable cells with fibrinolytic activity but also offers a new perspective for effectively addressing the significant loss of transplanted cells caused by thrombosis.
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Affiliation(s)
- Shengjie Liu
- College of Chemistry, Chemical Engineering and Materials Science, Soochow University, Suzhou 215123, P. R. China
| | - He Yang
- College of Chemistry, Chemical Engineering and Materials Science, Soochow University, Suzhou 215123, P. R. China
| | - Xingyu Heng
- College of Chemistry, Chemical Engineering and Materials Science, Soochow University, Suzhou 215123, P. R. China
| | - Lihua Yao
- College of Chemistry, Chemical Engineering and Materials Science, Soochow University, Suzhou 215123, P. R. China
| | - Wei Sun
- College of Chemistry, Chemical Engineering and Materials Science, Soochow University, Suzhou 215123, P. R. China
| | - Qing Zheng
- College of Chemistry, Chemical Engineering and Materials Science, Soochow University, Suzhou 215123, P. R. China
| | - Zhaoqiang Wu
- College of Chemistry, Chemical Engineering and Materials Science, Soochow University, Suzhou 215123, P. R. China
| | - Hong Chen
- College of Chemistry, Chemical Engineering and Materials Science, Soochow University, Suzhou 215123, P. R. China
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Shu Y, Otake M, Seta Y, Hori K, Kuramochi A, Ohba Y, Teramura Y. Activation of cellular antioxidative stress and migration activities by purified components from immortalized stem cells from human exfoliated deciduous teeth. Sci Rep 2024; 14:15340. [PMID: 38961142 PMCID: PMC11222459 DOI: 10.1038/s41598-024-66213-8] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/25/2024] [Accepted: 06/28/2024] [Indexed: 07/05/2024] Open
Abstract
Although stem cell-based regenerative medicine has been extensively studied, it remains difficult to reconstruct three dimensional tissues and organs in combination with vascular systems in vitro. One clinically successful therapy is transplantation of mesenchymal stem cells (MSC) into patients with graft versus host disease. However, transplanted cells are immediately damaged and destroyed because of innate immune reactions provoked by thrombogenic inflammation, and patients need to take immunosuppressive drugs for the immunological regulation of allogeneic cells. This reduces the benefits of stem cell transplantation. Therefore, alternative therapies are more realistic options for clinical use. In this study, we aimed to take advantage of the therapeutic efficacy of MSC and use multiple cytokines released from MSC, that is, stem cells from human exfoliated deciduous teeth (SHEDs). Here, we purified components from conditioned media of immortalized SHED (IM-SHED-CM) and evaluated the activities of intracellular dehydrogenase, cell migration, and antioxidative stress by studying the cells. The immortalization of SHED could make the stable supply of CM possible. We found that the fractionated component of 50-100 kD from IM-SHED-CM had higher efficacy than the original IM-SHED-CM in terms of intracellular dehydrogenase and cell migration in which intracellular signal transduction was activated via receptor tyrosine kinases, and the glutathione peroxidase and reductase system was highly active. Although antioxidative stress activities in the fractionated component of 50-100 kD had slightly lower than that of original IM-SHE-CM, the fraction still had the activity. Thus, the use of fractionated components of 50-100 kD from IM-SHED-CM could be an alternative choice for MSC transplantation because the purified components from CM could maintain the effect of cytokines from SHED.
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Affiliation(s)
- Yujing Shu
- U-Factor Co., Ltd, 1F, ESCALIER Rokubancho, 7-11, Rokubancho, Chiyoda, Tokyo, 102-0085, Japan
| | - Masato Otake
- U-Factor Co., Ltd, 1F, ESCALIER Rokubancho, 7-11, Rokubancho, Chiyoda, Tokyo, 102-0085, Japan
| | - Yasuhiro Seta
- Hitonowa Medical, K.PLAZA 2F, 1-7 Rokubancho, Chiyoda, Tokyo, 102-0085, Japan
| | - Keigo Hori
- U-Factor Co., Ltd, 1F, ESCALIER Rokubancho, 7-11, Rokubancho, Chiyoda, Tokyo, 102-0085, Japan
| | - Akiko Kuramochi
- Cellular and Molecular Biotechnology Research Institute (CMB), National Institute of Advanced Industrial Science and Technology (AIST), AIST Tsukuba Central 5, 1-1-1 Higashi, Tsukuba, Ibaraki, 305-8565, Japan
| | - Yoshio Ohba
- Cellular and Molecular Biotechnology Research Institute (CMB), National Institute of Advanced Industrial Science and Technology (AIST), AIST Tsukuba Central 5, 1-1-1 Higashi, Tsukuba, Ibaraki, 305-8565, Japan
| | - Yuji Teramura
- Cellular and Molecular Biotechnology Research Institute (CMB), National Institute of Advanced Industrial Science and Technology (AIST), AIST Tsukuba Central 5, 1-1-1 Higashi, Tsukuba, Ibaraki, 305-8565, Japan.
- Department of Immunology, Genetics and Pathology (IGP), Uppsala University, Dag Hammarskjölds Väg 20, 751 85, Uppsala, Sweden.
- Master's/Doctoral Program in Life Science Innovation (T-LSI), University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki, 305-8577, Japan.
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6
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Abbas N, You K, Getachew A, Wu F, Hussain M, Huang X, Chen Y, Pan T, Li Y. Kupffer cells abrogate homing and repopulation of allogeneic hepatic progenitors in injured liver site. Stem Cell Res Ther 2024; 15:48. [PMID: 38378583 PMCID: PMC10877762 DOI: 10.1186/s13287-024-03656-w] [Citation(s) in RCA: 1] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/14/2023] [Accepted: 02/05/2024] [Indexed: 02/22/2024] Open
Abstract
BACKGROUND Allogeneic hepatocyte transplantation is an emerging approach to treat acute liver defects. However, durable engraftment of the transplanted cells remains a daunting task, as they are actively cleared by the recipient's immune system. Therefore, a detailed understanding of the innate or adaptive immune cells-derived responses against allogeneic transplanted hepatic cells is the key to rationalize cell-based therapies. METHODS Here, we induced an acute inflammatory regenerative niche (3-96 h) on the surface of the liver by the application of cryo-injury (CI) to systematically evaluate the innate immune response against transplanted allogeneic hepatic progenitors in a sustained micro-inflammatory environment. RESULTS The resulting data highlighted that the injured site was significantly repopulated by alternating numbers of innate immune cells, including neutrophils, monocytes and Kupffer cells (KCs), from 3 to 96 h. The transplanted allo-HPs, engrafted 6 h post-injury, were collectively eliminated by the innate immune response within 24 h of transplantation. Selective depletion of the KCs demonstrated a delayed recruitment of monocytes from day 2 to day 6. In addition, the intrasplenic engraftment of the hepatic progenitors 54 h post-transplantation was dismantled by KCs, while a time-dependent better survival and translocation of the transplanted cells into the injured site could be observed in samples devoid of KCs. CONCLUSION Overall, this study provides evidence that KCs ablation enables a better survival and integration of allo-HPs in a sustained liver inflammatory environment, having implications for rationalizing the cell-based therapeutic interventions against liver defects.
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Affiliation(s)
- Nasir Abbas
- Center for Health Research, Guangdong Provincial Key Laboratory of Biocomputing, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530, China
- Centre for Regenerative Medicine and Health (CRMH), Hong Kong Institute of Science and Innovation, Chinese Academy of Sciences, Hong Kong, Hong Kong SAR, China
| | - Kai You
- Center for Health Research, Guangdong Provincial Key Laboratory of Biocomputing, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530, China
| | - Anteneh Getachew
- Department of Biomedical Engineering, The University of Alabama at Birmingham, Birmingham, USA
| | - Feima Wu
- Center for Health Research, Guangdong Provincial Key Laboratory of Biocomputing, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530, China
| | - Muzammal Hussain
- Department of Biochemistry and Molecular Pharmacology, New York University Grossman School of Medicine, New York, NY, 10016, USA
| | - Xinping Huang
- Center for Health Research, Guangdong Provincial Key Laboratory of Biocomputing, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530, China
| | - Yan Chen
- Center for Health Research, Guangdong Provincial Key Laboratory of Biocomputing, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530, China
- Key Laboratory of Stem Cell and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530, China
- CAS Key Laboratory of Regenerative Biology, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530, China
| | - Tingcai Pan
- Department of Hepatobiliary Surgery II, Zhujiang Hospital, Southern Medical University, Guangzhou, 510280, Guangdong Province, China
| | - Yinxiong Li
- Center for Health Research, Guangdong Provincial Key Laboratory of Biocomputing, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530, China.
- Key Laboratory of Stem Cell and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530, China.
- CAS Key Laboratory of Regenerative Biology, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530, China.
- State Key Laboratory of Respiratory Disease, Guangzhou, 510000, China.
- China-New Zealand Joint Laboratory of Biomedicine and Health, Guangzhou, 510530, China.
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Udagawa D, Nagata S, Yagi H, Nishi K, Morisaku T, Adachi S, Nakano Y, Tanaka M, Hori S, Hasegawa Y, Abe Y, Kitago M, Kitagawa Y. A Novel Approach to Orthotopic Hepatocyte Transplantation Engineered With Liver Hydrogel for Fibrotic Livers, Enhancing Cell-Cell Interaction and Angiogenesis. Cell Transplant 2024; 33:9636897241253700. [PMID: 38770981 PMCID: PMC11110510 DOI: 10.1177/09636897241253700] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/08/2023] [Revised: 04/22/2024] [Accepted: 04/22/2024] [Indexed: 05/22/2024] Open
Abstract
Hepatocyte transplantation (HCT) is a potential bridging therapy or an alternative to liver transplantation. Conventionally, single-cell hepatocytes are injected via the portal vein. This strategy, however, has yet to overcome poor cell engraftment and function. Therefore, we developed an orthotopic HCT method using a liver-derived extracellular matrix (L-ECM) gel. PXB cells (flesh mature human hepatocytes) were dispersed into the hydrogel solution in vitro, and the gel solution was immediately gelated in 37°C incubators to investigate the affinity between mature human hepatocyte and the L-ECM gel. During the 3-day cultivation in hepatocyte medium, PXB cells formed cell aggregates via cell-cell interactions. Quantitative analysis revealed human albumin production in culture supernatants. For the in vivo assay, PXB cells were encapsulated in the L-ECM gel and transplanted between the liver lobes of normal rats. Pathologically, the L-ECM gel was localized at the transplant site and retained PXB cells. Cell survival and hepatic function marker expression were verified in another rat model wherein thioacetamide was administered to induce liver fibrosis. Moreover, cell-cell interactions and angiogenesis were enhanced in the L-ECM gel compared with that in the collagen gel. Our results indicate that L-ECM gels can help engraft transplanted hepatocytes and express hepatic function as a scaffold for cell transplantation.
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Affiliation(s)
- Daisuke Udagawa
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
| | - Shogo Nagata
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
| | - Hiroshi Yagi
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
| | - Kotaro Nishi
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
| | | | - Shungo Adachi
- Fundamental Innovative Oncology Core, National Cancer Center Research Institute, Tokyo, Japan
| | - Yutaka Nakano
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
| | - Masayuki Tanaka
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
| | - Shutaro Hori
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
| | - Yasushi Hasegawa
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
| | - Yuta Abe
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
| | - Minoru Kitago
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
| | - Yuko Kitagawa
- Department of Surgery, Keio University School of Medicine, Tokyo, Japan
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8
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Sun Z, Yuan X, Wu J, Wang C, Zhang K, Zhang L, Hui L. Hepatocyte transplantation: The progress and the challenges. Hepatol Commun 2023; 7:e0266. [PMID: 37695736 PMCID: PMC10497249 DOI: 10.1097/hc9.0000000000000266] [Citation(s) in RCA: 8] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/13/2023] [Accepted: 07/26/2023] [Indexed: 09/13/2023] Open
Abstract
Numerous studies have shown that hepatocyte transplantation is a promising approach for liver diseases, such as liver-based metabolic diseases and acute liver failure. However, it lacks strong evidence to support the long-term therapeutic effects of hepatocyte transplantation in clinical practice. Currently, major hurdles include availability of quality-assured hepatocytes, efficient engraftment and repopulation, and effective immunosuppressive regimens. Notably, cell sources have been advanced recently by expanding primary human hepatocytes by means of dedifferentiation in vitro. Moreover, the transplantation efficiency was remarkably improved by the established preparative hepatic irradiation in combination with hepatic mitogenic stimuli regimens. Finally, immunosuppression drugs, including glucocorticoid and inhibitors for co-stimulating signals of T cell activation, were proposed to prevent innate and adaptive immune rejection of allografted hepatocytes. Despite remarkable progress, further studies are required to improve in vitro cell expansion technology, develop clinically feasible preconditioning regimens, and further optimize immunosuppression regimens or establish ex vivo gene correction-based autologous hepatocyte transplantation.
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Affiliation(s)
- Zhen Sun
- State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
- School of Life Science and Technology, ShanghaiTech University, Shanghai, China
| | - Xiang Yuan
- State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
| | - Jingqi Wu
- State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
| | - Chenhua Wang
- State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
| | - Kun Zhang
- State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
| | - Ludi Zhang
- State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
| | - Lijian Hui
- State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
- School of Life Science and Technology, ShanghaiTech University, Shanghai, China
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9
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Ma H, Wang C, Liang S, Yu X, Yuan Y, Lv Z, Zhang J, Jin C, Zhu J, Wang C, Sun P, Li W. ROCK inhibition enhanced hepatocyte liver engraftment by retaining membrane CD59 and attenuating complement activation. Mol Ther 2023; 31:1846-1856. [PMID: 36860134 PMCID: PMC10277888 DOI: 10.1016/j.ymthe.2023.02.018] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/16/2022] [Revised: 01/19/2023] [Accepted: 02/23/2023] [Indexed: 03/03/2023] Open
Abstract
Hepatocyte transplantation can be an effective treatment for patients with certain liver-based metabolic disorders and liver injuries. Hepatocytes are usually infused into the portal vein, from which hepatocytes migrate into the liver and integrate into the liver parenchyma. However, early cell loss and poor liver engraftment represent major hurdles to sustaining the recovery of diseased livers after transplantation. In the present study, we found that ROCK (Rho-associated kinase) inhibitors significantly enhanced in vivo hepatocyte engraftment. Mechanistic studies suggested that the isolation of hepatocytes caused substantial degradation of cell membrane proteins, including the complement inhibitor CD59, probably due to shear stress-induced endocytosis. ROCK inhibition by ripasudil, a clinically used ROCK inhibitor, can protect transplanted hepatocytes by retaining cell membrane CD59 and blocking the formation of the membrane attack complex. Knockdown of CD59 in hepatocytes eliminates ROCK inhibition-enhanced hepatocyte engraftment. Ripasudil can accelerate liver repopulation of fumarylacetoacetate hydrolase-deficient mice. Our work reveals a mechanism underlying hepatocyte loss after transplantation and provides immediate strategies to enhance hepatocyte engraftment by inhibiting ROCK.
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Affiliation(s)
- Haoxin Ma
- Department of Cell Biology, Naval Medical University, Shanghai 200433, China
| | - Chao Wang
- Department of Cell Biology, Naval Medical University, Shanghai 200433, China
| | - Shulong Liang
- Department of Cell Biology, Naval Medical University, Shanghai 200433, China
| | - Xinlu Yu
- Department of Cell Biology, Naval Medical University, Shanghai 200433, China
| | - Yuan Yuan
- Department of Cell Biology, Naval Medical University, Shanghai 200433, China
| | - Zhuanman Lv
- Department of Cell Biology, Naval Medical University, Shanghai 200433, China
| | - Jiqianzhu Zhang
- Department of Health Toxicology, Naval Medical University, Shanghai 200433, China
| | - Caixia Jin
- Department of Regenerative Medicine, College of Medicine, Tongji University, Shanghai 200433, China
| | - Jiangbo Zhu
- Department of Health Toxicology, Naval Medical University, Shanghai 200433, China
| | - Chao Wang
- Department of Cell Biology, Naval Medical University, Shanghai 200433, China
| | - Pingxin Sun
- Department of Cell Biology, Naval Medical University, Shanghai 200433, China
| | - Wenlin Li
- Department of Cell Biology, Naval Medical University, Shanghai 200433, China
- Shanghai Key Laboratory of Cell Engineering, Naval Medical University, Shanghai 200433, China
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10
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Cotransplantation With Adipose Tissue-derived Stem Cells Improves Engraftment of Transplanted Hepatocytes. Transplantation 2022; 106:1963-1973. [PMID: 35404871 PMCID: PMC9521584 DOI: 10.1097/tp.0000000000004130] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/04/2023]
Abstract
BACKGROUND Hepatocyte transplantation is expected to be an alternative therapy to liver transplantation; however, poor engraftment is a severe obstacle to be overcome. The adipose tissue-derived stem cells (ADSCs) are known to improve engraftment of transplanted pancreatic islets, which have many similarities to the hepatocytes. Therefore, we examined the effects and underlying mechanisms of ADSC cotransplantation on hepatocyte engraftment. METHODS Hepatocytes and ADSCs were cotransplanted into the renal subcapsular space and livers of syngeneic analbuminemic rats, and the serum albumin level was quantified to evaluate engraftment. Immunohistochemical staining and fluorescent staining to trace transplanted cells in the liver were also performed. To investigate the mechanisms, cocultured supernatants were analyzed by a multiplex assay and inhibition test using neutralizing antibodies for target factors. RESULTS Hepatocyte engraftment at both transplant sites was significantly improved by ADSC cotransplantation ( P < 0.001, P < 0.001). In the renal subcapsular model, close proximity between hepatocytes and ADSCs was necessary to exert this effect. Unexpectedly, ≈50% of transplanted hepatocytes were attached by ADSCs in the liver. In an in vitro study, the hepatocyte function was significantly improved by ADSC coculture supernatant ( P < 0.001). The multiplex assay and inhibition test demonstrated that hepatocyte growth factor, vascular endothelial growth factor, and interleukin-6 may be key factors for the abovementioned effects of ADSCs. CONCLUSIONS The present study revealed that ADSC cotransplantation can improve the engraftment of transplanted hepatocytes. This effect may be based on crucial factors, such as hepatocyte growth factor, vascular endothelial growth factor, and interleukin-6, which are secreted by ADSCs.
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11
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Kizhakkedathu JN, Conway EM. Biomaterial and cellular implants: foreign surfaces where immunity and coagulation meet. Blood 2022; 139:1987-1998. [PMID: 34415324 DOI: 10.1182/blood.2020007209] [Citation(s) in RCA: 27] [Impact Index Per Article: 9.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/10/2021] [Accepted: 08/05/2021] [Indexed: 11/20/2022] Open
Abstract
Exposure of blood to a foreign surface in the form of a diagnostic or therapeutic biomaterial device or implanted cells or tissue elicits an immediate, evolutionarily conserved thromboinflammatory response from the host. Primarily designed to protect against invading organisms after an injury, this innate response features instantaneous activation of several blood-borne, highly interactive, well-orchestrated cascades and cellular events that limit bleeding, destroy and eliminate the foreign substance or cells, and promote healing and a return to homeostasis via delicately balanced regenerative processes. In the setting of blood-contacting synthetic or natural biomaterials and implantation of foreign cells or tissues, innate responses are robust, albeit highly context specific. Unfortunately, they tend to be less than adequately regulated by the host's natural anticoagulant or anti-inflammatory pathways, thereby jeopardizing the functional integrity of the device, as well as the health of the host. Strategies to achieve biocompatibility with a sustained return to homeostasis, particularly while the device remains in situ and functional, continue to elude scientists and clinicians. In this review, some of the complex mechanisms by which biomaterials and cellular transplants provide a "hub" for activation and amplification of coagulation and immunity, thromboinflammation, are discussed, with a view toward the development of innovative means of overcoming the innate challenges.
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Affiliation(s)
- Jayachandran N Kizhakkedathu
- Centre for Blood Research
- Department of Pathology and Laboratory Medicine
- School of Biomedical Engineering, Life Sciences Institute
- Department of Chemistry; and
| | - Edward M Conway
- Centre for Blood Research
- School of Biomedical Engineering, Life Sciences Institute
- Department of Chemistry; and
- Department of Medicine, University of British Columbia, Vancouver, BC, Canada
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12
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Short-term inhalation of isoflurane improves the outcomes of intraportal hepatocyte transplantation. Sci Rep 2022; 12:4241. [PMID: 35273344 PMCID: PMC8913608 DOI: 10.1038/s41598-022-08237-6] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/12/2021] [Accepted: 03/04/2022] [Indexed: 11/25/2022] Open
Abstract
Clinical hepatocyte transplantation (HTx) is only performed without general anesthesia, while inhalation anesthetics are usually used in animal experiments. We hypothesized that isoflurane may be a possible reason for the discrepancy between the results of animal experiments and the clinical outcomes of HTx. Syngeneic rat hepatocytes (1.0 × 107) were transplanted to analbuminemic rats with (ISO group) and without (AW group) isoflurane. The serum albumin, AST, ALT, LDH levels and several inflammatory mediators were analyzed. Immunohistochemical staining and ex vivo imaging were also performed. The serum albumin levels of the ISO group were significantly higher in comparison to the AW group (p < 0.05). The serum AST, ALT, LDH levels of the ISO group were significantly suppressed in comparison to the AW group (p < 0.0001, respectively). The serum IL-1β, IL-10, IL-18, MCP-1, RNTES, Fractalkine and LIX levels were significantly suppressed in the ISO group. The ischemic regions of the recipient livers in the ISO group tended to be smaller than the AW group; however, the distribution of transplanted hepatocytes in the liver parenchyma was comparable between the two groups. Isoflurane may at least in part be a reason for the discrepancy between the results of animal experiments and the clinical outcomes of HTx.
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13
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Laner-Plamberger S, Oeller M, Rohde E, Schallmoser K, Strunk D. Heparin and Derivatives for Advanced Cell Therapies. Int J Mol Sci 2021; 22:12041. [PMID: 34769471 PMCID: PMC8584295 DOI: 10.3390/ijms222112041] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/21/2021] [Revised: 11/03/2021] [Accepted: 11/05/2021] [Indexed: 12/27/2022] Open
Abstract
Heparin and its derivatives are saving thousands of human lives annually, by successfully preventing and treating thromboembolic events. Although the mode of action during anticoagulation is well studied, their influence on cell behavior is not fully understood as is the risk of bleeding and other side effects. New applications in regenerative medicine have evolved supporting production of cell-based therapeutics or as a substrate for creating functionalized matrices in biotechnology. The currently resurgent interest in heparins is related to the expected combined anti-inflammatory, anti-thrombotic and anti-viral action against COVID-19. Based on a concise summary of key biochemical and clinical data, this review summarizes the impact for manufacturing and application of cell therapeutics and highlights the need for discriminating the different heparins.
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Affiliation(s)
- Sandra Laner-Plamberger
- Department of Transfusion Medicine, Paracelsus Medical University of Salzburg, 5020 Salzburg, Austria; (S.L.-P.); (M.O.); (E.R.)
- Spinal Cord Injury and Tissue Regeneration Center Salzburg, Paracelsus Medical University of Salzburg, 5020 Salzburg, Austria;
| | - Michaela Oeller
- Department of Transfusion Medicine, Paracelsus Medical University of Salzburg, 5020 Salzburg, Austria; (S.L.-P.); (M.O.); (E.R.)
- Spinal Cord Injury and Tissue Regeneration Center Salzburg, Paracelsus Medical University of Salzburg, 5020 Salzburg, Austria;
| | - Eva Rohde
- Department of Transfusion Medicine, Paracelsus Medical University of Salzburg, 5020 Salzburg, Austria; (S.L.-P.); (M.O.); (E.R.)
- Spinal Cord Injury and Tissue Regeneration Center Salzburg, Paracelsus Medical University of Salzburg, 5020 Salzburg, Austria;
| | - Katharina Schallmoser
- Department of Transfusion Medicine, Paracelsus Medical University of Salzburg, 5020 Salzburg, Austria; (S.L.-P.); (M.O.); (E.R.)
- Spinal Cord Injury and Tissue Regeneration Center Salzburg, Paracelsus Medical University of Salzburg, 5020 Salzburg, Austria;
| | - Dirk Strunk
- Spinal Cord Injury and Tissue Regeneration Center Salzburg, Paracelsus Medical University of Salzburg, 5020 Salzburg, Austria;
- Cell Therapy Institute, Paracelsus Medical University of Salzburg, 5020 Salzburg, Austria
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14
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Ogasawara H, Inagaki A, Fathi I, Imura T, Yamana H, Saitoh Y, Matsumura M, Fukuoka K, Miyagi S, Nakamura Y, Ohashi K, Unno M, Kamei T, Goto M. Preferable Transplant Site for Hepatocyte Transplantation in a Rat Model. Cell Transplant 2021; 30:9636897211040012. [PMID: 34525872 PMCID: PMC8450989 DOI: 10.1177/09636897211040012] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022] Open
Abstract
Intraportal injection is regarded as the current standard procedure of hepatocyte transplantation (HTx). In islet transplantation, which shares many aspects with HTx, recent studies have clarified that instant blood-mediated inflammatory reaction (IBMIR), characterized by strong innate immune responses, can cause poor engraftment, so other transplant sites to avoid such a reaction have been established. Although IBMIR was reported to occur in HTx, few reports have evaluated alternative transplant sites for HTx. In this study, we sought to determine the optimum transplant site for HTx. Rat hepatocytes (1.0 × 107) were transplanted at the 9 transplant sites (intraportal (IPO), intrasplenic (IS), liver parenchyma, subcutaneous, intraperitoneal, renal subcapsular, muscle, inguinal subcutaneous white adipose tissue, and omentum) of analbuminemic rats. The serum albumin levels, immunohistochemical staining (albumin, TUNEL, and BrdU), and in vivo imaging of the grafts were evaluated. The serum albumin levels of the IPO group were significantly higher than those of the other groups (p < .0001). The BrdU-positive hepatocyte ratio of liver in the IS group (0.9% ± 0.2%) was comparable to that of the IPO group (0.9% ± 0.3%) and tended to be higher than that of the spleen in the IS group (0.5% ± 0.1%, p = .16). Considering the in vivo imaging evaluation and the influence of splenectomy, the graft function in the IS group may be almost entirely achieved by hepatocytes that have migrated to the liver. The present study clearly showed that the intraportal injection procedure is more efficient than other procedures for performing HTx
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Affiliation(s)
- Hiroyuki Ogasawara
- Department of Surgery, Tohoku University Graduate School of Medicine, Sendai, Japan
| | - Akiko Inagaki
- Division of Transplantation and Regenerative Medicine, Tohoku University Graduate School of Medicine, Sendai, Japan
| | - Ibrahim Fathi
- Division of Transplantation and Regenerative Medicine, Tohoku University Graduate School of Medicine, Sendai, Japan
| | - Takehiro Imura
- Division of Transplantation and Regenerative Medicine, Tohoku University Graduate School of Medicine, Sendai, Japan
| | - Hiroki Yamana
- Department of Surgery, Tohoku University Graduate School of Medicine, Sendai, Japan
| | - Yoshikatsu Saitoh
- Department of Surgery, Tohoku University Graduate School of Medicine, Sendai, Japan
| | - Muneyuki Matsumura
- Department of Surgery, Tohoku University Graduate School of Medicine, Sendai, Japan
| | - Kengo Fukuoka
- Department of Surgery, Tohoku University Graduate School of Medicine, Sendai, Japan
| | - Shigehito Miyagi
- Department of Surgery, Tohoku University Graduate School of Medicine, Sendai, Japan
| | - Yasuhiro Nakamura
- Division of Pathology, Faculty of Medicine, Tohoku Medical and Pharmaceutical University, Sendai, Japan
| | - Kazuo Ohashi
- Laboratory of Drug Development and Science, Graduate School of Pharmaceutical Sciences, Osaka University, Osaka, Japan
| | - Michiaki Unno
- Department of Surgery, Tohoku University Graduate School of Medicine, Sendai, Japan
| | - Takashi Kamei
- Department of Surgery, Tohoku University Graduate School of Medicine, Sendai, Japan
| | - Masafumi Goto
- Department of Surgery, Tohoku University Graduate School of Medicine, Sendai, Japan.,Division of Transplantation and Regenerative Medicine, Tohoku University Graduate School of Medicine, Sendai, Japan
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15
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Bram Y, Nguyen DHT, Gupta V, Park J, Richardson C, Chandar V, Schwartz RE. Cell and Tissue Therapy for the Treatment of Chronic Liver Disease. Annu Rev Biomed Eng 2021; 23:517-546. [PMID: 33974812 PMCID: PMC8864721 DOI: 10.1146/annurev-bioeng-112619-044026] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/04/2022]
Abstract
Liver disease is an important clinical problem, impacting 600 million people worldwide. It is the 11th-leading cause of death in the world. Despite constant improvement in treatment and diagnostics, the aging population and accumulated risk factors led to increased morbidity due to nonalcoholic fatty liver disease and steatohepatitis. Liver transplantation, first established in the 1960s, is the second-most-common solid organ transplantation and is the gold standard for the treatment of liver failure. However, less than 10% of the global need for liver transplantation is met at the current rates of transplantation due to the paucity of available organs. Cell- and tissue-based therapies present an alternative to organ transplantation. This review surveys the approaches and tools that have been developed, discusses the distinctive challenges that exist for cell- and tissue-based therapies, and examines the future directions of regenerative therapies for the treatment of liver disease.
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Affiliation(s)
- Yaron Bram
- Division of Gastroenterology and Hepatology, Department of Medicine, Weill Cornell Medicine, New York, NY 10065, USA;
| | - Duc-Huy T Nguyen
- Division of Gastroenterology and Hepatology, Department of Medicine, Weill Cornell Medicine, New York, NY 10065, USA;
| | - Vikas Gupta
- Division of Gastroenterology and Hepatology, Department of Medicine, Weill Cornell Medicine, New York, NY 10065, USA;
| | - Jiwoon Park
- Department of Physiology, Biophysics and Systems Biology, Weill Cornell Medical College, New York, NY 10065, USA
| | - Chanel Richardson
- Department of Pharmacology, Weill Cornell Medical College, New York, NY 10065, USA
| | - Vasuretha Chandar
- Division of Gastroenterology and Hepatology, Department of Medicine, Weill Cornell Medicine, New York, NY 10065, USA;
| | - Robert E Schwartz
- Division of Gastroenterology and Hepatology, Department of Medicine, Weill Cornell Medicine, New York, NY 10065, USA; .,Department of Physiology, Biophysics and Systems Biology, Weill Cornell Medical College, New York, NY 10065, USA
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16
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Shibuya K, Watanabe M, Goto R, Zaitsu M, Ganchiku Y, Taketomi A. The Efficacy of the Hepatocyte Spheroids for Hepatocyte Transplantation. Cell Transplant 2021; 30:9636897211000014. [PMID: 33900126 PMCID: PMC8085376 DOI: 10.1177/09636897211000014] [Citation(s) in RCA: 9] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022] Open
Abstract
The safety and short-term efficacy of hepatocyte transplantation (HCTx) have been widely proven. However, issues such as reduced viability and/or function of hepatocytes, insufficient engraftment, and lack of a long-term effect have to be overcome for widespread application of HCTx. In this study, we evaluated hepatocyte spheroids (HSs), formed by self-aggregation of hepatocytes, as an alternative to hepatocytes in single-cell suspension. Hepatocytes were isolated from C57BL/6 J mice liver using a three-step collagenase perfusion technique and HSs were formed by the hanging drop method. After the spheroids formation, the HSs showed significantly higher mRNA expression of albumin, ornithine transcarbamylase, glucose-6-phosphate, alpha-1-antitrypsin, low density lipoprotein receptor, coagulation factors, and apolipoprotein E (ApoE) than 2 dimensional (2D)-cultured hepatocytes (p < 0.05). Albumin production by HSs was significantly higher than that by 2D-cultured hepatocytes (9.5 ± 2.5 vs 3.5 ± 1.8 μg/dL, p < 0.05). The HSs, but not single hepatocytes, maintained viability and albumin mRNA expression in suspension (92.0 ± 2.8% and 1.03 ± 0.09 at 6 h). HSs (3.6 × 106 cells) or isolated hepatocytes (fSH, 3.6 × 106 cells) were transplanted into the liver of ApoE knockout (KO-/-) mice via the portal vein. Following transplantation, serum ApoE concentration (ng/mL) of HS-transplanted mice (1w: 63.1 ± 56.7, 4w: 17.0 ± 10.9) was higher than that of fSH-transplanted mice (1 w: 33.4 ± 13.0, 4w: 13.7 ± 9.6). In both groups, the mRNA levels of pro-inflammatory cytokines (IL-6, IL-1β, TNF-α, MCP-1, and MIP-1β) were upregulated in the liver following transplantation; however, no significant differences were observed. Pathologically, transplanted HSs were observed as flat cell clusters in contact with the portal vein wall on day 7. Additionally, ApoE positive cells were observed in the liver parenchyma distant from the portal vein on day 28. Our results indicate that HS is a promising alternative to single hepatocytes and can be applied for HCTx.
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Affiliation(s)
- Kazuaki Shibuya
- Department of Gastroenterological surgery I, 12810Hokkaido university graduate school, kita-ku, Sapporo, Japan
| | - Masaaki Watanabe
- Transplant surgery, 163693Hokkaido University Hospital, kita-ku, Sapporo, Japan
| | - Ryoichi Goto
- Department of Gastroenterological surgery I, 12810Hokkaido university graduate school, kita-ku, Sapporo, Japan
| | - Masaaki Zaitsu
- Department of Gastroenterological surgery I, 12810Hokkaido university graduate school, kita-ku, Sapporo, Japan
| | - Yoshikazu Ganchiku
- Department of Gastroenterological surgery I, 12810Hokkaido university graduate school, kita-ku, Sapporo, Japan
| | - Akinobu Taketomi
- Department of Gastroenterological surgery I, 12810Hokkaido university graduate school, kita-ku, Sapporo, Japan
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17
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Demaret T, Evraerts J, Ravau J, Roumain M, Muccioli GG, Najimi M, Sokal EM. High Dose Versus Low Dose Syngeneic Hepatocyte Transplantation in Pex1-G844D NMRI Mouse Model is Safe but Does Not Achieve Long Term Engraftment. Cells 2020; 10:cells10010040. [PMID: 33396635 PMCID: PMC7823729 DOI: 10.3390/cells10010040] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/08/2020] [Revised: 12/21/2020] [Accepted: 12/23/2020] [Indexed: 12/17/2022] Open
Abstract
Genetic alterations in PEX genes lead to peroxisome biogenesis disorder. In humans, they are associated with Zellweger spectrum disorders (ZSD). No validated treatment has been shown to modify the dismal natural history of ZSD. Liver transplantation (LT) improved clinical and biochemical outcomes in mild ZSD patients. Hepatocyte transplantation (HT), developed to overcome LT limitations, was performed in a mild ZSD 4-year-old child with encouraging short-term results. Here, we evaluated low dose (12.5 million hepatocytes/kg) and high dose (50 million hepatocytes/kg) syngeneic male HT via intrasplenic infusion in the Pex1-G844D NMRI mouse model which recapitulates a mild ZSD phenotype. HT was feasible and safe in growth retarded ZSD mice. Clinical (weight and food intake) and biochemical parameters (very long-chain fatty acids, abnormal bile acids, etc.) were in accordance with ZSD phenotype but they were not robustly modified by HT. As expected, one third of the infused cells were detected in the liver 24 h post-HT. No liver nor spleen microchimerism was detected after 7, 14 and 30 days. Future optimizations are required to improve hepatocyte engraftment in Pex1-G844D NMRI mouse liver. The mouse model exhibited the robustness required for ZSD liver-targeted therapies evaluation.
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Affiliation(s)
- Tanguy Demaret
- Laboratoire d’Hépatologie Pédiatrique et Thérapie Cellulaire, Unité PEDI, Institut de Recherche Expérimentale et Clinique (IREC), Université Catholique de Louvain (UCLouvain), 1200 Brussels, Belgium; (J.E.); (J.R.); (M.N.); (E.M.S.)
- Correspondence:
| | - Jonathan Evraerts
- Laboratoire d’Hépatologie Pédiatrique et Thérapie Cellulaire, Unité PEDI, Institut de Recherche Expérimentale et Clinique (IREC), Université Catholique de Louvain (UCLouvain), 1200 Brussels, Belgium; (J.E.); (J.R.); (M.N.); (E.M.S.)
| | - Joachim Ravau
- Laboratoire d’Hépatologie Pédiatrique et Thérapie Cellulaire, Unité PEDI, Institut de Recherche Expérimentale et Clinique (IREC), Université Catholique de Louvain (UCLouvain), 1200 Brussels, Belgium; (J.E.); (J.R.); (M.N.); (E.M.S.)
| | - Martin Roumain
- Bioanalysis and Pharmacology of Bioactive Lipids Research Group (BPBL), Louvain Drug Research Institute (LDRI), Université Catholique de Louvain (UCLouvain), 1200 Brussels, Belgium; (M.R.); (G.G.M.)
| | - Giulio G. Muccioli
- Bioanalysis and Pharmacology of Bioactive Lipids Research Group (BPBL), Louvain Drug Research Institute (LDRI), Université Catholique de Louvain (UCLouvain), 1200 Brussels, Belgium; (M.R.); (G.G.M.)
| | - Mustapha Najimi
- Laboratoire d’Hépatologie Pédiatrique et Thérapie Cellulaire, Unité PEDI, Institut de Recherche Expérimentale et Clinique (IREC), Université Catholique de Louvain (UCLouvain), 1200 Brussels, Belgium; (J.E.); (J.R.); (M.N.); (E.M.S.)
| | - Etienne M. Sokal
- Laboratoire d’Hépatologie Pédiatrique et Thérapie Cellulaire, Unité PEDI, Institut de Recherche Expérimentale et Clinique (IREC), Université Catholique de Louvain (UCLouvain), 1200 Brussels, Belgium; (J.E.); (J.R.); (M.N.); (E.M.S.)
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18
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Salminen AT, Allahyari Z, Gholizadeh S, McCloskey MC, Ajalik R, Cottle RN, Gaborski TR, McGrath JL. In vitro Studies of Transendothelial Migration for Biological and Drug Discovery. FRONTIERS IN MEDICAL TECHNOLOGY 2020; 2:600616. [PMID: 35047883 PMCID: PMC8757899 DOI: 10.3389/fmedt.2020.600616] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/30/2020] [Accepted: 10/20/2020] [Indexed: 12/13/2022] Open
Abstract
Inflammatory diseases and cancer metastases lack concrete pharmaceuticals for their effective treatment despite great strides in advancing our understanding of disease progression. One feature of these disease pathogeneses that remains to be fully explored, both biologically and pharmaceutically, is the passage of cancer and immune cells from the blood to the underlying tissue in the process of extravasation. Regardless of migratory cell type, all steps in extravasation involve molecular interactions that serve as a rich landscape of targets for pharmaceutical inhibition or promotion. Transendothelial migration (TEM), or the migration of the cell through the vascular endothelium, is a particularly promising area of interest as it constitutes the final and most involved step in the extravasation cascade. While in vivo models of cancer metastasis and inflammatory diseases have contributed to our current understanding of TEM, the knowledge surrounding this phenomenon would be significantly lacking without the use of in vitro platforms. In addition to the ease of use, low cost, and high controllability, in vitro platforms permit the use of human cell lines to represent certain features of disease pathology better, as seen in the clinic. These benefits over traditional pre-clinical models for efficacy and toxicity testing are especially important in the modern pursuit of novel drug candidates. Here, we review the cellular and molecular events involved in leukocyte and cancer cell extravasation, with a keen focus on TEM, as discovered by seminal and progressive in vitro platforms. In vitro studies of TEM, specifically, showcase the great experimental progress at the lab bench and highlight the historical success of in vitro platforms for biological discovery. This success shows the potential for applying these platforms for pharmaceutical compound screening. In addition to immune and cancer cell TEM, we discuss the promise of hepatocyte transplantation, a process in which systemically delivered hepatocytes must transmigrate across the liver sinusoidal endothelium to successfully engraft and restore liver function. Lastly, we concisely summarize the evolving field of porous membranes for the study of TEM.
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Affiliation(s)
- Alec T. Salminen
- Biomedical Engineering, University of Rochester, Rochester, NY, United States
| | - Zahra Allahyari
- Biomedical Engineering, Rochester Institute of Technology, Rochester, NY, United States
| | - Shayan Gholizadeh
- Biomedical Engineering, Rochester Institute of Technology, Rochester, NY, United States
| | - Molly C. McCloskey
- Biomedical Engineering, University of Rochester, Rochester, NY, United States
| | - Raquel Ajalik
- Biomedical Engineering, University of Rochester, Rochester, NY, United States
| | - Renee N. Cottle
- Bioengineering, Clemson University, Clemson, SC, United States
| | - Thomas R. Gaborski
- Biomedical Engineering, University of Rochester, Rochester, NY, United States
- Biomedical Engineering, Rochester Institute of Technology, Rochester, NY, United States
| | - James L. McGrath
- Biomedical Engineering, University of Rochester, Rochester, NY, United States
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19
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Teramura Y, Ekdahl KN, Fromell K, Nilsson B, Ishihara K. Potential of Cell Surface Engineering with Biocompatible Polymers for Biomedical Applications. LANGMUIR : THE ACS JOURNAL OF SURFACES AND COLLOIDS 2020; 36:12088-12106. [PMID: 32927948 DOI: 10.1021/acs.langmuir.0c01678] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/11/2023]
Abstract
The regulation of the cellular surface with biomaterials can contribute to the progress of biomedical applications. In particular, the cell surface is exposed to immunological surveillance and reactions in transplantation therapy, and modulation of cell surface properties might improve transplantation outcomes. The transplantation of therapeutic cells, tissue, and organs is an effective and fundamental treatment and has contributed to saving lives and improving quality of life. Because of shortages, donor cells, tissues, and organs are carefully transplanted with the goal of retaining activity and viability. However, some issues remain to be resolved in terms of reducing side effects, improving graft survival, managing innate and adaptive immune responses, and improving transplant storage and procedures. Given that the transplantation process involves multiple steps and is technically complicated, an engineering approach together with medical approaches to resolving these issues could enhance success. In particular, cell surface engineering with biocompatible polymers looks promising for improving transplantation therapy and has potential for other biomedical applications. Here we review the significance of polymer-based surface modification of cells and organs for biomedical applications, focusing on the following three topics: Cell protection: cellular protection through local immune regulation using cell surface modification with biocompatible polymers. This protection could extend to preventing attack by the host immune system, freeing recipients from taking immunosuppressive drugs, and avoiding a second transplantation. Cell attachment: cell manipulation, which is an important technique for delivery of therapeutic cells and their alignment for recellularization of decellularized tissues and organs in regenerative therapy. Cell fusion: fusion of different cells, which can lead to the formation of new functional cells that could be useful for generating, e.g., immunologically competent or metabolically active cells.
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Affiliation(s)
- Yuji Teramura
- Department of Bioengineering, School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo, 113-8656, Japan
- Department of Immunology, Genetics and Pathology (IGP), Uppsala University, Dag Hammarskjölds väg 20, SE-751 85, Uppsala, Sweden
| | - Kristina Nilsson Ekdahl
- Department of Immunology, Genetics and Pathology (IGP), Uppsala University, Dag Hammarskjölds väg 20, SE-751 85, Uppsala, Sweden
- Linnaeus Center of Biomaterials Chemistry, Linnaeus University, SE-391 82 Kalmar, Sweden
| | - Karin Fromell
- Department of Immunology, Genetics and Pathology (IGP), Uppsala University, Dag Hammarskjölds väg 20, SE-751 85, Uppsala, Sweden
| | - Bo Nilsson
- Department of Immunology, Genetics and Pathology (IGP), Uppsala University, Dag Hammarskjölds väg 20, SE-751 85, Uppsala, Sweden
| | - Kazuhiko Ishihara
- Department of Material Engineering, School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo, 113-8656, Japan
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20
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Exposure of von Willebrand Factor on Isolated Hepatocytes Promotes Tethering of Platelets to the Cell Surface. Transplantation 2020; 103:1630-1638. [PMID: 30896677 DOI: 10.1097/tp.0000000000002707] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022]
Abstract
BACKGROUND Hepatocyte transplantation (Hctx) is a potentially attractive method for the treatment of acute liver failure and liver-based metabolic disorders. Unfortunately, the procedure is hampered by the instant blood-mediated inflammatory reaction (IBMIR), a thromboinflammatory response elicited by the vascular innate immune system, causing activation of the coagulation and complement systems and clearance of transplanted cells. Observations have also revealed platelets adhered to the surface of the hepatocytes (Hc). To establish Hctx as a clinical treatment, all factors that trigger IBMIR need to be identified and controlled. This work explores the expression of von Willebrand factor (VWF) on isolated Hc resulting in tethering of platelets. METHODS VWF on Hc was studied by flow cytometry, confocal microscopy, immunoblot, and real-time polymerase chain reaction. Interaction between Hc and platelets was studied in a Chandler loop model. Adhesion of platelets to the hepatocyte surface was demonstrated by flow cytometry and confocal microscopy. RESULTS Isolated Hc constitutively express VWF on their cell surface and mRNA for VWF was found in the cells. Hc and platelets, independently of coagulation formed complexes, were shown by antibody blocking studies to be dependent on hepatocyte-associated VWF and platelet-bound glycoprotein Ibα. CONCLUSIONS VWF on isolated Hc causes, in contact with blood, adhesion of platelets, which thereby forms an ideal surface for coagulation. This phenomenon needs to be considered in hepatocyte-based reconstitution therapy and possibly even in other settings of cell transplantation.
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Coppin LCF, Smets F, Ambroise J, Sokal EEM, Stéphenne X. Infusion-related thrombogenesis by liver-derived mesenchymal stem cells controlled by anticoagulant drugs in 11 patients with liver-based metabolic disorders. Stem Cell Res Ther 2020; 11:51. [PMID: 32028991 PMCID: PMC7006410 DOI: 10.1186/s13287-020-1572-7] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/27/2019] [Revised: 12/17/2019] [Accepted: 01/26/2020] [Indexed: 02/07/2023] Open
Abstract
BACKGROUND Mesenchymal stem cell (MSC) transplantation is a fast-developing therapy in regenerative medicine. However, some concerns have been raised regarding their safety and the infusion-related pro-coagulant activity. The aim of this study is to analyze the induced thrombogenic risk and the safety of adding anticoagulants during intraportal infusions of liver-derived MSCs (HepaStem), in patients with Crigler-Najjar (CN) and urea cycle disorders (UCD). METHODS Eleven patients (6 CN and 5 UCD patients) were included in this partially randomized phase 1/2 study. Three cell doses of HepaStem were investigated: low (12.5 × 106 cells/kg), intermediate (50 × 106 cells/kg), and high doses (200 × 106 cells/kg). A combination of anticoagulants, heparin (10 I.U./5 × 106cells), and bivalirudin (1.75 mg/kg/h) were added during cell infusions. The infusion-related thrombogenic risk and anticoagulation were evaluated by clinical monitoring, blood sampling (platelet and D-dimer levels, activated clotting time, etc.) and liver Doppler ultrasound. Mixed effects linear regression models were used to assess statistically significant differences. RESULTS One patient presented a thrombogenic event such as a partial portal vein thrombus after 6 infusions. Minor adverse effects such as petechiae, epistaxis, and cutaneous hemorrhage at the site of catheter placement were observed in four patients. A significant decrease in platelet and increase in D-dimer levels were observed at the end of the infusion cycle, normalizing spontaneously after 7 days. No significant and clinically relevant increase in portal vein pressure could be observed once the infusion cycle was completed. CONCLUSIONS The safety- and the infusion-related pro-coagulant activity remains a concern in MSC transplantation. In our study, a combination of heparin and bivalirudin was added to prevent the thrombogenic risk induced by HepaStem infusions in 11 patients. A significant decrease in platelet and increase in D-dimer levels were observed, suggesting the activation of coagulation in these patients; however, this was spontaneously reversible in time. We can conclude that adding this combination of anticoagulants is safe and limits infusion-related thrombogenesis to subclinical signs in most of the patients. TRIAL REGISTRATION ClinicalTrials.gov identifier: NCT01765283-January 10, 2013.
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Affiliation(s)
- Louise C F Coppin
- Service de Gastro-Entérologie et Hépatologie Pédiatrique, Cliniques Universitaires Saint-Luc, Université Catholique de Louvain, Av Hippocrate 10, B-1200, Brussels, Belgium.
| | - Françoise Smets
- Service de Gastro-Entérologie et Hépatologie Pédiatrique, Cliniques Universitaires Saint-Luc, Université Catholique de Louvain, Av Hippocrate 10, B-1200, Brussels, Belgium
| | - Jérome Ambroise
- Institut de Recherche Expérimentale et Clinique, Université Catholique de Louvain, Brussels, Belgium
| | - Etienne E M Sokal
- Service de Gastro-Entérologie et Hépatologie Pédiatrique, Cliniques Universitaires Saint-Luc, Université Catholique de Louvain, Av Hippocrate 10, B-1200, Brussels, Belgium
| | - Xavier Stéphenne
- Service de Gastro-Entérologie et Hépatologie Pédiatrique, Cliniques Universitaires Saint-Luc, Université Catholique de Louvain, Av Hippocrate 10, B-1200, Brussels, Belgium
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Tokodai K, Kumagai-Braesch M, Karadagi A, Johansson H, Ågren N, Jorns C, Ericzon BG, Ellis E. Blood Group Antigen Expression in Isolated Human Liver Cells in Preparation for Implementing Clinical ABO-Incompatible Hepatocyte Transplantation. J Clin Exp Hepatol 2020; 10:106-113. [PMID: 32189925 PMCID: PMC7068001 DOI: 10.1016/j.jceh.2019.07.001] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/18/2019] [Accepted: 07/05/2019] [Indexed: 12/12/2022] Open
Abstract
BACKGROUND ABO blood group antigens in the liver are expressed mainly on endothelial cells or biliary epithelial cells but not on hepatocytes. This suggests that ABO-incompatible hepatocyte transplantation (ABOi-HTx) is theoretically feasible. However, the effects of stress on ABO blood group antigen expression caused by isolation and intraportal infusion require thorough investigation before ABOi-HTx can be implemented in clinical settings. METHODS Human hepatocytes were isolated from liver tissue obtained from liver resection or deceased donor livers. The expression of blood group antigens on cryopreserved human liver tissues and isolated hepatocyte smear specimens were examined by immunofluorescent staining. The effect of proinflammatory cytokines on blood group antigen expression of hepatocytes was evaluated by flow cytometry. Instant blood-mediated inflammatory reaction after hepatocyte incubation with ABO-incompatible whole blood was examined using the tubing loop model. RESULTS Blood group antigens were mainly expressed on vessels in the portal area. In hepatocyte smear specimens, isolated hepatocytes did not express blood group antigens. In contrast, a subset of cells in the smear specimens of nonparenchymal liver cells stained positive. In the flow cytometry analysis, isolated hepatocytes were negative for blood group antigens, even after 4-h incubation with cytokines. Platelet counts and complement activation were not significantly different in ABO-identical versus ABO-incompatible settings in the tubing loop model. CONCLUSION Our study showed that blood group antigens were not expressed on hepatocytes, even after isolation procedures or subsequent incubation with cytokines. This finding is an important step toward removing the restriction of ABO matching in hepatocyte transplantation. Our results suggest that ABOi-HTx is a feasible therapeutic option, especially in patients who require urgent treatment with freshly isolated hepatocytes, such as those with acute liver failure.
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Key Words
- ABO-incompatible transplantation
- ABOi, ABO-incompatible
- ABOi-HTx, ABO-incompatible hepatocyte transplantation
- C3a, Complement 3a
- CCA, Cholangiocarcinoma
- CRC, Colorectal cancer metastasis
- DCD, Donation after death
- DSA, Donor-specific anti-HLA antibody
- HCC, Hepatocellular carcinoma
- HTx, Hepatocyte transplantation
- IBMIR, Instant blood-mediated inflammatory reaction
- ICH, Intracranial hemorrhage
- IQR, Interquartile range
- MSCs, Mesenchymal stem cells
- blood group antigens
- hepatocytes
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Affiliation(s)
| | | | | | | | | | | | | | - Ewa Ellis
- Address for correspondence: Ewa Ellis Ph.D., Department of Clinical Science, Intervention and Technology (CLINTEC), Division of Transplantation Surgery, Karolinska University Hospital Huddinge, Karolinska Institute, SE-141 86, Stockholm, Sweden.
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Ha DH, Thi PM, Chaudhary P, Jeong JH. Efficient Formation of Three Dimensional Spheroids of Primary Hepatocytes Using Micropatterned Multi-Well Plates. Macromol Res 2019. [DOI: 10.1007/s13233-019-7103-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/27/2022]
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Coppin L, Sokal E, Stéphenne X. Thrombogenic Risk Induced by Intravascular Mesenchymal Stem Cell Therapy: Current Status and Future Perspectives. Cells 2019; 8:cells8101160. [PMID: 31569696 PMCID: PMC6829440 DOI: 10.3390/cells8101160] [Citation(s) in RCA: 82] [Impact Index Per Article: 13.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/15/2019] [Revised: 09/20/2019] [Accepted: 09/25/2019] [Indexed: 12/11/2022] Open
Abstract
Mesenchymal stem cells (MSCs) are currently studied and used in numerous clinical trials. Nevertheless, some concerns have been raised regarding the safety of these infusions and the thrombogenic risk they induce. MSCs express procoagulant activity (PCA) linked to the expression of tissue factor (TF) that, when in contact with blood, initiates coagulation. Some even describe a dual activation of both the coagulation and the complement pathway, called Instant Blood-Mediated Inflammatory Reaction (IBMIR), explaining the disappointing results and low engraftment rates in clinical trials. However, nowadays, different approaches to modulate the PCA of MSCs and thus control the thrombogenic risk after cell infusion are being studied. This review summarizes both in vitro and in vivo studies on the PCA of MSC of various origins. It further emphasizes the crucial role of TF linked to the PCA of MSCs. Furthermore, optimization of MSC therapy protocols using different methods to control the PCA of MSCs are described.
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Affiliation(s)
- Louise Coppin
- Laboratoire d'Hépatologie Pédiatrique et Thérapie Cellulaire, Unité PEDI, Institut de Recherche Expérimentale et Clinique, Université catholique de Louvain (UCLouvain), 1200 Brussels, Belgium.
| | - Etienne Sokal
- Laboratoire d'Hépatologie Pédiatrique et Thérapie Cellulaire, Unité PEDI, Institut de Recherche Expérimentale et Clinique, Université catholique de Louvain (UCLouvain), 1200 Brussels, Belgium.
| | - Xavier Stéphenne
- Laboratoire d'Hépatologie Pédiatrique et Thérapie Cellulaire, Unité PEDI, Institut de Recherche Expérimentale et Clinique, Université catholique de Louvain (UCLouvain), 1200 Brussels, Belgium.
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25
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Khan RS, Newsome PN. A Comparison of Phenotypic and Functional Properties of Mesenchymal Stromal Cells and Multipotent Adult Progenitor Cells. Front Immunol 2019; 10:1952. [PMID: 31555259 PMCID: PMC6724467 DOI: 10.3389/fimmu.2019.01952] [Citation(s) in RCA: 24] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/29/2019] [Accepted: 08/02/2019] [Indexed: 12/15/2022] Open
Abstract
Both Multipotent Adult Progenitor Cells and Mesenchymal Stromal Cells are bone-marrow derived, non-haematopoietic adherent cells, that are well-known for having immunomodulatory and pro-angiogenic properties, whilst being relatively non-immunogenic. However, they are phenotypically and functionally distinct cell types, which has implications for their efficacy in different settings. In this review we compare the phenotypic and functional properties of these two cell types, to help in determining which would be the superior cell type for different applications.
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Affiliation(s)
- Reenam S Khan
- National Institute for Health Research (NIHR), Birmingham Biomedical Research Centre, University Hospitals Birmingham NHS Foundation Trust, University of Birmingham, Birmingham, United Kingdom.,Centre for Liver Research, Institute of Immunology and Immunotherapy, University of Birmingham, Birmingham, United Kingdom.,Liver Unit, University Hospitals Birmingham NHS Foundation Trust, Birmingham, United Kingdom
| | - Philip N Newsome
- National Institute for Health Research (NIHR), Birmingham Biomedical Research Centre, University Hospitals Birmingham NHS Foundation Trust, University of Birmingham, Birmingham, United Kingdom.,Centre for Liver Research, Institute of Immunology and Immunotherapy, University of Birmingham, Birmingham, United Kingdom.,Liver Unit, University Hospitals Birmingham NHS Foundation Trust, Birmingham, United Kingdom
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26
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Clinical Protocol to Prevent Thrombogenic Effect of Liver-Derived Mesenchymal Cells for Cell-Based Therapies. Cells 2019; 8:cells8080846. [PMID: 31394759 PMCID: PMC6721739 DOI: 10.3390/cells8080846] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/04/2019] [Revised: 08/03/2019] [Accepted: 08/06/2019] [Indexed: 12/11/2022] Open
Abstract
The efficacy of mesenchymal stem cell infusion is currently tested in numerous clinical trials. However, therapy-induced thrombotic consequences have been reported in several patients. The aim of this study was to optimize protocols for heterologous human adult liver-derived progenitor cell (HHALPC) infusion, in order to eliminate acute thrombogenesis in liver-based metabolic or acute decompensated cirrhotic (ADC) patients. In rats, thrombotic effects were absent when HHALPCs were infused at low cell dose (5 × 106 cells/kg), or at high cell dose (5 × 107 cells/kg) when combined with anticoagulants. When HHALPCs were exposed to human blood in a whole blood perfusion assay, blocking of the tissue factor (TF) coagulation pathway suppressed fibrin generation and platelet activation. In a Chandler tubing loop model, HHALPCs induced less explosive activation of coagulation with blood from ADC patients, when compared to blood from healthy controls, without alterations in coagulation factor levels other than fibrinogen. These studies confirm a link between TF and thrombogenesis, when TF-expressing cells are exposed to human blood. This phenomenon however, could be controlled using either a low, or a high cell dose combined with anticoagulants. In clinical practice, this points to the suitability of a low HHALPC dose infusion to cirrhotic patients, provided that platelet and fibrinogen levels are monitored.
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Caplan H, Olson SD, Kumar A, George M, Prabhakara KS, Wenzel P, Bedi S, Toledano-Furman NE, Triolo F, Kamhieh-Milz J, Moll G, Cox CS. Mesenchymal Stromal Cell Therapeutic Delivery: Translational Challenges to Clinical Application. Front Immunol 2019; 10:1645. [PMID: 31417542 PMCID: PMC6685059 DOI: 10.3389/fimmu.2019.01645] [Citation(s) in RCA: 210] [Impact Index Per Article: 35.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/23/2019] [Accepted: 07/02/2019] [Indexed: 12/12/2022] Open
Abstract
For several decades, multipotent mesenchymal stromal cells (MSCs) have been extensively studied for their therapeutic potential across a wide range of diseases. In the preclinical setting, MSCs demonstrate consistent ability to promote tissue healing, down-regulate excessive inflammation and improve outcomes in animal models. Several proposed mechanisms of action have been posited and demonstrated across an array of in vitro models. However, translation into clinical practice has proven considerably more difficult. A number of prominent well-funded late-phase clinical trials have failed, thus calling out for new efforts to optimize product delivery in the clinical setting. In this review, we discuss novel topics critical to the successful translation of MSCs from pre-clinical to clinical applications. In particular, we focus on the major routes of cell delivery, aspects related to hemocompatibility, and potential safety concerns associated with MSC therapy in the different settings.
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Affiliation(s)
- Henry Caplan
- Department of Pediatric Surgery, McGovern Medical School, University of Texas Health Science Center at Houston, Houston, TX, United States
| | - Scott D. Olson
- Department of Pediatric Surgery, McGovern Medical School, University of Texas Health Science Center at Houston, Houston, TX, United States
| | - Akshita Kumar
- Department of Pediatric Surgery, McGovern Medical School, University of Texas Health Science Center at Houston, Houston, TX, United States
| | - Mitchell George
- Department of Pediatric Surgery, McGovern Medical School, University of Texas Health Science Center at Houston, Houston, TX, United States
| | - Karthik S. Prabhakara
- Department of Pediatric Surgery, McGovern Medical School, University of Texas Health Science Center at Houston, Houston, TX, United States
| | - Pamela Wenzel
- Department of Pediatric Surgery, McGovern Medical School, University of Texas Health Science Center at Houston, Houston, TX, United States
| | - Supinder Bedi
- Department of Pediatric Surgery, McGovern Medical School, University of Texas Health Science Center at Houston, Houston, TX, United States
| | - Naama E. Toledano-Furman
- Department of Pediatric Surgery, McGovern Medical School, University of Texas Health Science Center at Houston, Houston, TX, United States
| | - Fabio Triolo
- Department of Pediatric Surgery, McGovern Medical School, University of Texas Health Science Center at Houston, Houston, TX, United States
| | - Julian Kamhieh-Milz
- Department of Transfusion Medicine, Charité Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health (BIH), Berlin, Germany
| | - Guido Moll
- BIH Center for Regenerative Therapies (BCRT), Charité Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health (BIH), Berlin, Germany
| | - Charles S. Cox
- Department of Pediatric Surgery, McGovern Medical School, University of Texas Health Science Center at Houston, Houston, TX, United States
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Iansante V, Chandrashekran A, Dhawan A. Cell-based liver therapies: past, present and future. Philos Trans R Soc Lond B Biol Sci 2019; 373:rstb.2017.0229. [PMID: 29786563 DOI: 10.1098/rstb.2017.0229] [Citation(s) in RCA: 36] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 12/23/2017] [Indexed: 12/16/2022] Open
Abstract
Liver transplantation represents the standard treatment for people with an end-stage liver disease and some liver-based metabolic disorders; however, shortage of liver donor tissues limits its availability. Furthermore, whole liver replacement eliminates the possibility of using native liver as a possible target for future gene therapy in case of liver-based metabolic defects. Cell therapy has emerged as a potential alternative, as cells can provide the hepatic functions and engraft in the liver parenchyma. Various options have been proposed, including human or other species hepatocytes, hepatocyte-like cells derived from stem cells or more futuristic alternatives, such as combination therapies with different cell types, organoids and cell-biomaterial combinations. In this review, we aim to give an overview of the cell therapies developed so far, highlighting preclinical and/or clinical achievements as well as the limitations that need to be overcome to make them fully effective and safe for clinical applications.This article is part of the theme issue 'Designer human tissue: coming to a lab near you'.
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Affiliation(s)
- Valeria Iansante
- Dhawan Lab, Paediatric Liver GI and Nutrition Center and MowatLabs, Institute of Liver Studies, King's College London at King's College Hospital, London SE5 9PJ, UK
| | - Anil Chandrashekran
- Dhawan Lab, Paediatric Liver GI and Nutrition Center and MowatLabs, Institute of Liver Studies, King's College London at King's College Hospital, London SE5 9PJ, UK
| | - Anil Dhawan
- Dhawan Lab, Paediatric Liver GI and Nutrition Center and MowatLabs, Institute of Liver Studies, King's College London at King's College Hospital, London SE5 9PJ, UK
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Noiri M, Asawa K, Okada N, Kodama T, Murayama Y, Inoue Y, Ishihara K, Ekdahl KN, Nilsson B, Teramura Y. Modification of human MSC surface with oligopeptide‐PEG‐lipids for selective binding to activated endothelium. J Biomed Mater Res A 2019; 107:1779-1792. [DOI: 10.1002/jbm.a.36697] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/14/2019] [Revised: 04/04/2019] [Accepted: 04/10/2019] [Indexed: 01/12/2023]
Affiliation(s)
- Makoto Noiri
- Department of Bioengineering School of Engineering, The University of Tokyo 7‐3‐1 Hongo, Bunkyo‐ku, Tokyo, 113‐8656 Japan
| | - Kenta Asawa
- Department of Bioengineering School of Engineering, The University of Tokyo 7‐3‐1 Hongo, Bunkyo‐ku, Tokyo, 113‐8656 Japan
| | - Naoya Okada
- Department of Bioengineering School of Engineering, The University of Tokyo 7‐3‐1 Hongo, Bunkyo‐ku, Tokyo, 113‐8656 Japan
| | - Tomonobu Kodama
- Department of Neurosurgery The Jikei University Hospital Tokyo Japan
| | - Yuichi Murayama
- Department of Neurosurgery The Jikei University Hospital Tokyo Japan
| | - Yuuki Inoue
- Department of Material Engineering School of Engineering, The University of Tokyo 7‐3‐1 Hongo, Bunkyo‐ku, Tokyo, 113‐8656 Japan
| | - Kazuhiko Ishihara
- Department of Bioengineering School of Engineering, The University of Tokyo 7‐3‐1 Hongo, Bunkyo‐ku, Tokyo, 113‐8656 Japan
- Department of Material Engineering School of Engineering, The University of Tokyo 7‐3‐1 Hongo, Bunkyo‐ku, Tokyo, 113‐8656 Japan
| | - Kristina N Ekdahl
- Linnaeus Center of Biomaterials Chemistry Linnaeus University SE‐391 82, Kalmar Sweden
- Department of Immunology, Genetics, and Pathology (IGP) Uppsala University Dag Hammarskjölds väg 20, SE‐751 85, Uppsala Sweden
| | - Bo Nilsson
- Department of Immunology, Genetics, and Pathology (IGP) Uppsala University Dag Hammarskjölds väg 20, SE‐751 85, Uppsala Sweden
| | - Yuji Teramura
- Department of Bioengineering School of Engineering, The University of Tokyo 7‐3‐1 Hongo, Bunkyo‐ku, Tokyo, 113‐8656 Japan
- Department of Immunology, Genetics, and Pathology (IGP) Uppsala University Dag Hammarskjölds väg 20, SE‐751 85, Uppsala Sweden
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Lee C, Dhawan A, Iansante V, Filippi C, Mitry R, Tang J, Walker S, Fernandez DaCosta R, Sinha S, Hughes RD, Koulmanda M, Fitzpatrick E. Improving engraftment of hepatocyte transplantation using alpha-1 antitrypsin as an immune modulator. J Mol Med (Berl) 2019; 97:563-577. [PMID: 30820592 PMCID: PMC6440943 DOI: 10.1007/s00109-019-01747-3] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/31/2018] [Revised: 01/18/2019] [Accepted: 01/22/2019] [Indexed: 01/11/2023]
Abstract
Abstract For patients with non-cirrhotic liver-based metabolic disorders, hepatocyte transplantation can be an effective treatment. However, long-term function of transplanted hepatocytes following infusion has not been achieved due to insufficient numbers of hepatocytes reaching the liver cell plates caused by activation of the instant blood-mediated inflammatory reaction (IBMIR). Our aim was to determine if the natural immune modulator, alpha-1 antitrypsin (AAT), could improve engraftment of transplanted hepatocytes and investigate its mechanism of action. A tubing loop model was used to analyse activation of the IBMIR when human hepatocytes were in contact with ABO-matched blood and 4 mg/ml AAT. Platelet and white cell counts, complement and cytokine expression were analysed. To determine if AAT could improve short-term engraftment, female rats underwent tail vein injection of AAT (120 mg/kg) or water (control) prior to the intrasplenic transplantation of 2 × 107 male hepatocytes. At 48 h and 1 week, livers were collected for analysis. In our loop model, human hepatocytes elicited a significant drop in platelet count with thrombus formation compared to controls. Loops containing AAT and hepatocytes showed no platelet consumption and no thrombus formation. Further, AAT treatment resulted in reduced IL-1β, IL-6 and IFN-γ and increased IL-1RA compared to untreated loops. In vivo, AAT significantly improved engraftment of rat hepatocytes compared to untreated at 48 h. AAT infusion may inhibit the IBMIR, thus improving short-term engraftment of donor hepatocytes and potentially improve the outcomes for patients with liver-based metabolic disease. Key messages • Alpha-1 antitrypsin (AAT) acts as an immune modulator to improve the efficacy of hepatocyte transplantation. • Treatment with AAT decreased thrombus formation and pro-inflammatory cytokine expression in a tubing loop model. • AAT significantly improved engraftment of donor hepatocytes within the first 48 h post transplantation. Electronic supplementary material The online version of this article (10.1007/s00109-019-01747-3) contains supplementary material, which is available to authorized users.
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Affiliation(s)
- Charlotte Lee
- Dhawan Group at Mowat Labs, Institute of Liver Studies, King's College London at King's College Hospital, London, UK
| | - Anil Dhawan
- Paediatric Liver, GI and Nutrition Centre, King's College London School of Medicine at King's College Hospital, Denmark Hill, London, UK.
| | - Valeria Iansante
- Dhawan Group at Mowat Labs, Institute of Liver Studies, King's College London at King's College Hospital, London, UK
| | - Celine Filippi
- Dhawan Group at Mowat Labs, Institute of Liver Studies, King's College London at King's College Hospital, London, UK
| | - Ragai Mitry
- Dhawan Group at Mowat Labs, Institute of Liver Studies, King's College London at King's College Hospital, London, UK
| | - Joanne Tang
- Dhawan Group at Mowat Labs, Institute of Liver Studies, King's College London at King's College Hospital, London, UK
| | - Simon Walker
- Paediatric Liver, GI and Nutrition Centre, King's College London School of Medicine at King's College Hospital, Denmark Hill, London, UK
| | - Raquel Fernandez DaCosta
- Paediatric Liver, GI and Nutrition Centre, King's College London School of Medicine at King's College Hospital, Denmark Hill, London, UK
| | - Siddharth Sinha
- Dhawan Group at Mowat Labs, Institute of Liver Studies, King's College London at King's College Hospital, London, UK
| | - Robin D Hughes
- Dhawan Group at Mowat Labs, Institute of Liver Studies, King's College London at King's College Hospital, London, UK
| | - Maria Koulmanda
- Departments of Medicine and Surgery, The Transplant Institute, Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, MA, USA
| | - Emer Fitzpatrick
- Paediatric Liver, GI and Nutrition Centre, King's College London School of Medicine at King's College Hospital, Denmark Hill, London, UK.
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Song G, Hu Y, Liu Y, Jiang R. Layer-by-Layer Heparinization of the Cell Surface by Using Heparin-Binding Peptide Functionalized Human Serum Albumin. MATERIALS 2018; 11:ma11050849. [PMID: 29783776 PMCID: PMC5978226 DOI: 10.3390/ma11050849] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 04/27/2018] [Revised: 05/17/2018] [Accepted: 05/18/2018] [Indexed: 02/07/2023]
Abstract
Layer-by-layer heparinization of therapeutic cells prior to transplantation is an effective way to inhibit the instant blood-mediated inflammatory reactions (IBMIRs), which are the major cause of early cell graft loss during post-transplantation. Here, a conjugate of heparin-binding peptide (HBP) and human serum albumin (HSA), HBP-HSA, was synthesized by using heterobifunctional crosslinker. After the first heparin layer was coated on human umbilical vein endothelial cells (HUVECs) by means of the HBP-polyethylene glycol-phospholipid conjugate, HBP-HSA and heparin were then applied to the cell surface sequentially to form multiple layers. The immobilization and retention of heparin were analyzed by confocal microscopy and flow cytometry, respectively, and the cytotoxity of HBP-HSA was further evaluated by cell viability assay. Results indicated that heparin was successfully introduced to the cell surface in a layer-by-layer way and retained for at least 24 h, while the cytotoxity of HBP-HSA was negligible at the working concentration. Accordingly, this conjugate provides a promising method for co-immobilization of heparin and HSA to the cell surface under physiological conditions with improved biocompatibility.
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Affiliation(s)
- Guowei Song
- College of Life and Health Sciences, Northeastern University, Shenyang, 110169, China.
| | - Yaning Hu
- College of Life and Health Sciences, Northeastern University, Shenyang, 110169, China.
| | - Yusheng Liu
- College of Life and Health Sciences, Northeastern University, Shenyang, 110169, China.
| | - Rui Jiang
- College of Life and Health Sciences, Northeastern University, Shenyang, 110169, China.
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Lee CA, Dhawan A, Iansante V, Lehec S, Khorsandi SE, Filippi C, Walker S, Fernandez-Dacosta R, Heaton N, Bansal S, Mitry RR, Fitzpatrick E. Cryopreserved neonatal hepatocytes may be a source for transplantation: Evaluation of functionality toward clinical use. Liver Transpl 2018; 24:394-406. [PMID: 29356341 DOI: 10.1002/lt.25015] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/31/2017] [Revised: 12/13/2017] [Accepted: 12/15/2017] [Indexed: 12/27/2022]
Abstract
Neonatal livers are a potential source of good-quality hepatocytes for clinical transplantation. We compared viability and function of neonatal hepatocytes (NHs) and adult hepatocytes (AHs) and report their clinical use both intraportally and in alginate microbeads. Following isolation from donor livers, hepatocyte function was assessed using albumin, alpha-1-antitrypsin, and factor VII. Metabolic function was investigated by measuring resorufin conjugation, ammonia metabolism, uridine diphosphate glucuronosyltransferase enzyme activity, and cytochrome P450 (CYP) function following induction. Activation of the instant blood-mediated inflammatory reaction by NHs and AHs was investigated using an in vitro blood perfusion model, and tissue factor expression was analyzed using real-time polymerase chain reaction (RT-PCR). Clinical hepatocyte transplantation (HT) was undertaken using standard protocols. Hepatocytes were isolated from 14 neonatal livers, with an average viability of 89.4% ± 1.8% (mean ± standard error of the mean) and average yield of 9.3 × 106 ± 2.0 × 106 cells/g. Hepatocytes were isolated from 14 adult livers with an average viability of 78.6% ± 2.4% and yield 2.2 × 106 ± 0.5 × 105 cells/g. NHs had significantly higher viability after cryopreservation than AHs, with better attachment efficiency and less plasma membrane leakage. There were no differences in albumin, alpha-1-antitrypsin, and factor VII synthesis between NHs and AHs (P > 0.05). Neonatal cells had inducible phase 1 enzymes as assessed by CYP function and functional phase 2 enzymes, in which activity was comparable to AHs. In an in vitro blood perfusion model, AHs elicited increased thrombus formation with a greater consumption of platelets and white cells compared with NHs (28.3 × 109 versus 118.7 × 109 and 3.3 × 109 versus 6.6 × 109 ; P < 0.01). Intraportal transplantation and intraperitoneal transplantation of alginate encapsulated hepatocytes was safe, and preliminary data suggest the cells may activate the immune response to a lesser degree than adult cells. In conclusion, we have shown NHs have excellent cell viability, function, and drug metabolism making them a suitable alternative source for clinical HT. Liver Transplantation 24 394-406 2018 AASLD.
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Affiliation(s)
- Charlotte A Lee
- Dhawan Lab, Institute of Liver Studies and, King's College London at King's College Hospital, London, UK
| | - Anil Dhawan
- Pediatric Liver, GI and Nutrition Centre, King's College London at King's College Hospital, London, UK
| | - Valeria Iansante
- Dhawan Lab, Institute of Liver Studies and, King's College London at King's College Hospital, London, UK
| | - Sharon Lehec
- Dhawan Lab, Institute of Liver Studies and, King's College London at King's College Hospital, London, UK
| | - Shirin E Khorsandi
- Institute of Liver Studies, King's College Hospital National Health Service Foundation Trust, London, UK
| | - Celine Filippi
- Dhawan Lab, Institute of Liver Studies and, King's College London at King's College Hospital, London, UK
| | - Simon Walker
- Dhawan Lab, Institute of Liver Studies and, King's College London at King's College Hospital, London, UK
| | - Raquel Fernandez-Dacosta
- Dhawan Lab, Institute of Liver Studies and, King's College London at King's College Hospital, London, UK
| | - Nigel Heaton
- Institute of Liver Studies, King's College Hospital National Health Service Foundation Trust, London, UK
| | - Sanjay Bansal
- Pediatric Liver, GI and Nutrition Centre, King's College London at King's College Hospital, London, UK
| | - Ragai R Mitry
- Dhawan Lab, Institute of Liver Studies and, King's College London at King's College Hospital, London, UK
| | - Emer Fitzpatrick
- Pediatric Liver, GI and Nutrition Centre, King's College London at King's College Hospital, London, UK
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Iansante V, Mitry RR, Filippi C, Fitzpatrick E, Dhawan A. Human hepatocyte transplantation for liver disease: current status and future perspectives. Pediatr Res 2018; 83:232-240. [PMID: 29149103 DOI: 10.1038/pr.2017.284] [Citation(s) in RCA: 150] [Impact Index Per Article: 21.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/15/2017] [Accepted: 10/02/2017] [Indexed: 12/16/2022]
Abstract
Liver transplantation is the accepted treatment for patients with acute liver failure and liver-based metabolic disorders. However, donor organ shortage and lifelong need for immunosuppression are the main limitations to liver transplantation. In addition, loss of the native liver as a target organ for future gene therapy for metabolic disorders limits the futuristic treatment options, resulting in the need for alternative therapeutic strategies. A potential alternative to liver transplantation is allogeneic hepatocyte transplantation. Over the last two decades, hepatocyte transplantation has made the transition from bench to bedside. Standardized techniques have been established for isolation, culture, and cryopreservation of human hepatocytes. Clinical hepatocyte transplantation safety and short-term efficacy have been proven; however, some major hurdles-mainly concerning shortage of donor organs, low cell engraftment, and lack of a long-lasting effect-need to be overcome to widen its clinical applications. Current research is aimed at addressing these problems, with the ultimate goal of increasing hepatocyte transplantation efficacy in clinical applications.
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Affiliation(s)
- V Iansante
- DhawanLab, Paediatric Liver GI and Nutrition Center and MowatLabs, Institute of Liver Studies, King's College London, Faculty of Life Sciences and Medicine, King's College London, King's College Hospital, London, UK
| | - R R Mitry
- DhawanLab, Paediatric Liver GI and Nutrition Center and MowatLabs, Institute of Liver Studies, King's College London, Faculty of Life Sciences and Medicine, King's College London, King's College Hospital, London, UK
| | - C Filippi
- DhawanLab, Paediatric Liver GI and Nutrition Center and MowatLabs, Institute of Liver Studies, King's College London, Faculty of Life Sciences and Medicine, King's College London, King's College Hospital, London, UK
| | - E Fitzpatrick
- DhawanLab, Paediatric Liver GI and Nutrition Center and MowatLabs, Institute of Liver Studies, King's College London, Faculty of Life Sciences and Medicine, King's College London, King's College Hospital, London, UK
| | - A Dhawan
- DhawanLab, Paediatric Liver GI and Nutrition Center and MowatLabs, Institute of Liver Studies, King's College London, Faculty of Life Sciences and Medicine, King's College London, King's College Hospital, London, UK
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Tatsumi K, Okano T. Hepatocyte Transplantation: Cell Sheet Technology for Liver Cell Transplantation. CURRENT TRANSPLANTATION REPORTS 2017; 4:184-192. [PMID: 28932649 PMCID: PMC5577064 DOI: 10.1007/s40472-017-0156-7] [Citation(s) in RCA: 45] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
Purpose of Review We will review the recent developments of cell sheet technology as a feasible tissue engineering approach. Specifically, we will focus on the technological advancement for engineering functional liver tissue using cell sheet technology, and the associated therapeutic effect of cell sheets for liver diseases, highlighting hemophilia. Recent Findings Cell-based therapies using hepatocytes have recently been explored as a new therapeutic modality for patients with many forms of liver disease. We have developed a cell sheet technology, which allows cells to be harvested in a monolithic layer format. We have succeeded in fabricating functional liver tissues in mice by stacking the cell sheets composed of primary hepatocytes. As a curative measure for hemophilia, we have also succeeded in treating hemophilia mice by transplanting of cells sheets composed of genetically modified autologous cells. Summary Tissue engineering using cell sheet technology provides the opportunity to create new therapeutic options for patients with various types of liver diseases.
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Affiliation(s)
- Kohei Tatsumi
- Department of Physiology and Regenerative Medicine, Kindai University Faculty of Medicine, 377-2 Ohno-higashi, Osaka-sayama, Osaka 589-8511 Japan
| | - Teruo Okano
- Institute of Advanced Biomedical Engineering and Science, Tokyo Women's Medical University, 8-1 Kawada-cho, Shinjuku-ku, Tokyo, 162-8666 Japan.,Cell Sheet Tissue Engineering Center and Department of Pharmaceutics and Pharmaceutical Chemistry, University of Utah, Salt Lake City, UT 84112 USA
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Ekdahl KN, Teramura Y, Hamad OA, Asif S, Duehrkop C, Fromell K, Gustafson E, Hong J, Kozarcanin H, Magnusson PU, Huber-Lang M, Garred P, Nilsson B. Dangerous liaisons: complement, coagulation, and kallikrein/kinin cross-talk act as a linchpin in the events leading to thromboinflammation. Immunol Rev 2017; 274:245-269. [PMID: 27782319 DOI: 10.1111/imr.12471] [Citation(s) in RCA: 109] [Impact Index Per Article: 13.6] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
Innate immunity is fundamental to our defense against microorganisms. Physiologically, the intravascular innate immune system acts as a purging system that identifies and removes foreign substances leading to thromboinflammatory responses, tissue remodeling, and repair. It is also a key contributor to the adverse effects observed in many diseases and therapies involving biomaterials and therapeutic cells/organs. The intravascular innate immune system consists of the cascade systems of the blood (the complement, contact, coagulation, and fibrinolytic systems), the blood cells (polymorphonuclear cells, monocytes, platelets), and the endothelial cell lining of the vessels. Activation of the intravascular innate immune system in vivo leads to thromboinflammation that can be activated by several of the system's pathways and that initiates repair after tissue damage and leads to adverse reactions in several disorders and treatment modalities. In this review, we summarize the current knowledge in the field and discuss the obstacles that exist in order to study the cross-talk between the components of the intravascular innate immune system. These include the use of purified in vitro systems, animal models and various types of anticoagulants. In order to avoid some of these obstacles we have developed specialized human whole blood models that allow investigation of the cross-talk between the various cascade systems and the blood cells. We in particular stress that platelets are involved in these interactions and that the lectin pathway of the complement system is an emerging part of innate immunity that interacts with the contact/coagulation system. Understanding the resulting thromboinflammation will allow development of new therapeutic modalities.
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Affiliation(s)
- Kristina N Ekdahl
- Department of Immunology, Genetics and Pathology (IGP), Rudbeck Laboratory C5:3, Uppsala University, Uppsala, Sweden.,Linnaeus Center of Biomaterials Chemistry, Linnaeus University, Kalmar, Sweden
| | - Yuji Teramura
- Department of Immunology, Genetics and Pathology (IGP), Rudbeck Laboratory C5:3, Uppsala University, Uppsala, Sweden.,Department of Bioengineering, The University of Tokyo, Tokyo, Japan
| | - Osama A Hamad
- Department of Immunology, Genetics and Pathology (IGP), Rudbeck Laboratory C5:3, Uppsala University, Uppsala, Sweden
| | - Sana Asif
- Department of Immunology, Genetics and Pathology (IGP), Rudbeck Laboratory C5:3, Uppsala University, Uppsala, Sweden
| | - Claudia Duehrkop
- Department of Immunology, Genetics and Pathology (IGP), Rudbeck Laboratory C5:3, Uppsala University, Uppsala, Sweden
| | - Karin Fromell
- Department of Immunology, Genetics and Pathology (IGP), Rudbeck Laboratory C5:3, Uppsala University, Uppsala, Sweden
| | - Elisabet Gustafson
- Department of Women's and Children's Health, Uppsala University Hospital, Uppsala, Sweden
| | - Jaan Hong
- Department of Immunology, Genetics and Pathology (IGP), Rudbeck Laboratory C5:3, Uppsala University, Uppsala, Sweden
| | - Huda Kozarcanin
- Department of Immunology, Genetics and Pathology (IGP), Rudbeck Laboratory C5:3, Uppsala University, Uppsala, Sweden
| | - Peetra U Magnusson
- Department of Immunology, Genetics and Pathology (IGP), Rudbeck Laboratory C5:3, Uppsala University, Uppsala, Sweden
| | - Markus Huber-Lang
- Department of Orthopedic Trauma, Hand, Plastic and Reconstructive Surgery, University of Ulm, Ulm, Germany
| | - Peter Garred
- Laboratory of Molecular Medicine, Department of Clinical Immunology, Section 7631, Faculty of Health and Medical Sciences, Rigshospitalet, University of Copenhagen, Copenhagen, Denmark
| | - Bo Nilsson
- Department of Immunology, Genetics and Pathology (IGP), Rudbeck Laboratory C5:3, Uppsala University, Uppsala, Sweden.
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Abstract
Complement is a major contributor to inflammation and graft injury. This system is especially important in ischemia-reperfusion injury/delayed graft function as well as in acute and chronic antibody-mediated rejection (AMR). The latter is increasingly recognized as a major cause of late graft loss, for which we have few effective therapies. C1 inhibitor (C1-INH) regulates several pathways which contribute to both acute and chronic graft injuries. However, C1-INH spares the alternative pathway and the membrane attack complex (C5–9) so innate antibacterial defenses remain intact. Plasma-derived C1-INH has been used to treat hereditary angioedema for more than 30 years with excellent safety. Studies with C1-INH in transplant recipients are limited, but have not revealed any unique toxicity or serious adverse events attributed to the protein. Extensive data from animal and ex vivo models suggest that C1-INH ameliorates ischemia-reperfusion injury. Initial clinical studies suggest this effect may allow transplantation of donor organs which are now discarded because the risk of primary graft dysfunction is considered too great. Although the incidence of severe early AMR is declining, accumulating evidence strongly suggests that complement is an important mediator of chronic AMR, a major cause of late graft loss. Thus, C1-INH may also be helpful in preserving function of established grafts. Early clinical studies in transplantation suggest significant beneficial effects of C1-INH with minimal toxicity. Recent results encourage continued investigation of this already-available therapeutic agent.
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Wang J, Sun Z, Gou W, Adams DB, Cui W, Morgan KA, Strange C, Wang H. α-1 Antitrypsin Enhances Islet Engraftment by Suppression of Instant Blood-Mediated Inflammatory Reaction. Diabetes 2017; 66:970-980. [PMID: 28069642 PMCID: PMC5360304 DOI: 10.2337/db16-1036] [Citation(s) in RCA: 60] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/24/2016] [Accepted: 01/03/2017] [Indexed: 12/19/2022]
Abstract
Islet cell transplantation has limited effectiveness because of an instant blood-mediated inflammatory reaction (IBMIR) that occurs immediately after cell infusion and leads to dramatic β-cell death. In intraportal islet transplantation models using mouse and human islets, we demonstrated that α-1 antitrypsin (AAT; Prolastin-C), a serine protease inhibitor used for the treatment of AAT deficiency, inhibits IBMIR and cytokine-induced inflammation in islets. In mice, more diabetic recipients reached normoglycemia after intraportal islet transplantation when they were treated with AAT compared with mice treated with saline. AAT suppressed blood-mediated coagulation pathways by diminishing tissue factor production, reducing plasma thrombin-antithrombin complex levels and fibrinogen deposition on islet grafts, which correlated with less graft damage and apoptosis. AAT-treated mice showed reduced serum tumor necrosis factor-α levels, decreased lymphocytic infiltration, and decreased nuclear factor (NF)-κB activation compared with controls. The potent anti-inflammatory effect of AAT is possibly mediated by suppression of c-Jun N-terminal kinase (JNK) phosphorylation. Blocking JNK activation failed to further reduce cytokine-induced apoptosis in β-cells. Taken together, AAT significantly improves islet graft survival after intraportal islet transplantation by mitigation of coagulation in IBMIR and suppression of cytokine-induced JNK and NF-κB activation. AAT-based therapy has the potential to improve graft survival in human islet transplantation and other cellular therapies on the horizon.
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Affiliation(s)
- Jingjing Wang
- Department of Surgery, Medical University of South Carolina, Charleston, SC
| | - Zhen Sun
- Department of Surgery, Medical University of South Carolina, Charleston, SC
| | - Wenyu Gou
- Department of Surgery, Medical University of South Carolina, Charleston, SC
| | - David B Adams
- Department of Surgery, Medical University of South Carolina, Charleston, SC
| | - Wanxing Cui
- MedStar Georgetown University Hospital, Washington, DC
| | - Katherine A Morgan
- Department of Surgery, Medical University of South Carolina, Charleston, SC
| | - Charlie Strange
- Department of Medicine, Medical University of South Carolina, Charleston, SC
| | - Hongjun Wang
- Department of Surgery, Medical University of South Carolina, Charleston, SC
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Gustafson E, Asif S, Kozarcanin H, Elgue G, Meurling S, Ekdahl KN, Nilsson B. Control of IBMIR Induced by Fresh and Cryopreserved Hepatocytes by Low Molecular Weight Dextran Sulfate Versus Heparin. Cell Transplant 2016; 26:71-81. [PMID: 27452808 DOI: 10.3727/096368916x692609] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022] Open
Abstract
Rapid destruction of hepatocytes after hepatocyte transplantation has hampered the application of this procedure clinically. The instant blood-mediated inflammatory reaction (IBMIR) is a plausible underlying cause for this cell loss. The present study was designed to evaluate the capacity of low molecular weight dextran sulfate (LMW-DS) to control these initial reactions from the innate immune system. Fresh and cryopreserved hepatocytes were tested in an in vitro whole-blood model using ABO-compatible blood. The ability to elicit IBMIR and the capacity of LMW-DS (100 μg/ml) to attenuate the degree of activation of the cascade systems were monitored. The effect was also compared to conventional anticoagulant therapy using unfractionated heparin (1 IU/ml). Both fresh and freeze-thawed hepatocytes elicited IBMIR to the same extent. LMW-DS reduced the platelet loss and maintained the cell counts at the same degree as unfractionated heparin, but controlled the coagulation and complement systems significantly more efficiently than heparin. LMW-DS also attenuated the IBMIR elicited by freeze-thawed cells. Therefore, LMW-DS inhibits the cascade systems and maintains the cell counts in blood triggered by both fresh and cryopreserved hepatocytes in direct contact with ABO-matched blood. LMW-DS at a previously used and clinically applicable concentration (100 μg/ml) inhibits IBMIR in vitro and is therefore a potential IBMIR inhibitor in hepatocyte transplantation.
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Davies LC, Alm JJ, Heldring N, Moll G, Gavin C, Batsis I, Qian H, Sigvardsson M, Nilsson B, Kyllonen LE, Salmela KT, Carlsson PO, Korsgren O, Le Blanc K. Type 1 Diabetes Mellitus Donor Mesenchymal Stromal Cells Exhibit Comparable Potency to Healthy Controls In Vitro. Stem Cells Transl Med 2016; 5:1485-1495. [PMID: 27412884 DOI: 10.5966/sctm.2015-0272] [Citation(s) in RCA: 44] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/30/2015] [Accepted: 04/01/2016] [Indexed: 12/13/2022] Open
Abstract
: Bone marrow mesenchymal stromal cells (BM-MSCs) have been characterized and used in many clinical studies based on their immunomodulatory and regenerative properties. We have recently reported the benefit of autologous MSC systemic therapy in the treatment of type 1 diabetes mellitus (T1D). Compared with allogeneic cells, use of autologous products reduces the risk of eliciting undesired complications in the recipient, including rejection, immunization, and transmission of viruses and prions; however, comparable potency of autologous cells is required for this treatment approach to remain feasible. To date, no analysis has been reported that phenotypically and functionally characterizes MSCs derived from newly diagnosed and late-stage T1D donors in vitro with respect to their suitability for systemic immunotherapy. In this study, we used gene array in combination with functional in vitro assays to address these questions. MSCs from T1D donors and healthy controls were expanded from BM aspirates. BM mononuclear cell counts and growth kinetics were comparable between the groups, with equivalent colony-forming unit-fibroblast capacity. Gene microarrays demonstrated differential gene expression between healthy and late-stage T1D donors in relation to cytokine secretion, immunomodulatory activity, and wound healing potential. Despite transcriptional differences, T1D MSCs did not demonstrate a significant difference from healthy controls in immunosuppressive activity, migratory capacity, or hemocompatibility. We conclude that despite differential gene expression, expanded MSCs from T1D donors are phenotypically and functionally similar to healthy control MSCs with regard to their immunomodulatory and migratory potential, indicating their suitability for use in autologous systemic therapy. SIGNIFICANCE The potential for mesenchymal stromal cells (MSCs) as a cell-based therapy in the treatment of immunologic disorders has been well established. Recent studies reported the clinical potential for autologous MSCs as a systemic therapy in the treatment of type I diabetes mellitus (T1D). The current study compared the genotypic and phenotypic profiles of bone marrow-derived MSCs from T1D and healthy donors as autologous (compared with allogeneic) therapy provides distinct advantages, such as reduced risk of immune reaction and transmission of infectious agents. The findings of the current study demonstrate that despite moderate differences in T1D MSCs at the gene level, these cells can be expanded in culture to an extent corresponding to that of MSCs derived from healthy donors. No functional difference in terms of immunosuppressive activity, blood compatibility, or migratory capacity was evident between the groups. The study findings also show that autologous MSC therapy holds promise as a T1D treatment and should be evaluated further in clinical trials.
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Affiliation(s)
- Lindsay C Davies
- Center for Hematology and Regenerative Medicine, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden
- Divisions of Clinical Immunology and Transfusion Medicine, Department of Laboratory Medicine, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden
| | - Jessica J Alm
- Center for Hematology and Regenerative Medicine, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden
- Divisions of Clinical Immunology and Transfusion Medicine, Department of Laboratory Medicine, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden
| | - Nina Heldring
- Center for Hematology and Regenerative Medicine, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden
- Divisions of Clinical Immunology and Transfusion Medicine, Department of Laboratory Medicine, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden
| | - Guido Moll
- Center for Hematology and Regenerative Medicine, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden
- Divisions of Clinical Immunology and Transfusion Medicine, Department of Laboratory Medicine, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden
| | - Caroline Gavin
- Center for Hematology and Regenerative Medicine, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden
- Divisions of Clinical Immunology and Transfusion Medicine, Department of Laboratory Medicine, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden
| | - Ioannis Batsis
- Center for Hematology and Regenerative Medicine, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden
| | - Hong Qian
- Center for Hematology and Regenerative Medicine, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden
| | - Mikael Sigvardsson
- Institution for Clinical and Experimental Medicine, Linköping University, Linköping, Sweden
| | - Bo Nilsson
- Rudbeck Laboratory, Department of Immunology, Genetics and Pathology, Uppsala University, Sweden
| | - Lauri E Kyllonen
- Division of Transplantation, Helsinki University Hospital, Helsinki, Finland
| | - Kaija T Salmela
- Division of Transplantation, Helsinki University Hospital, Helsinki, Finland
| | - Per-Ola Carlsson
- Department of Medical Cell Biology, Uppsala University, Sweden
- Department of Medical Sciences, Uppsala University, Sweden
| | - Olle Korsgren
- Rudbeck Laboratory, Department of Immunology, Genetics and Pathology, Uppsala University, Sweden
| | - Katarina Le Blanc
- Center for Hematology and Regenerative Medicine, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden
- Divisions of Clinical Immunology and Transfusion Medicine, Department of Laboratory Medicine, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden
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Ekdahl KN, Huang S, Nilsson B, Teramura Y. Complement inhibition in biomaterial- and biosurface-induced thromboinflammation. Semin Immunol 2016; 28:268-77. [PMID: 27211838 PMCID: PMC7129373 DOI: 10.1016/j.smim.2016.04.006] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/14/2016] [Revised: 04/25/2016] [Accepted: 04/26/2016] [Indexed: 12/20/2022]
Abstract
Therapeutic medicine today includes a vast number of procedures involving the use of biomaterials, transplantation of therapeutic cells or cell clusters, as well as of solid organs. These treatment modalities are obviously of great benefit to the patient, but also present a great challenge to the innate immune system, since they involve direct exposure of non-biological materials, cells of non-hematological origin as well as endothelial cells, damaged by ischemia-perfusion in solid organs to proteins and cells in the blood. The result of such an exposure may be an inappropriate activation of the complement and contact/kallikrein systems, which produce mediators capable of triggering the platelets and PMNs and monocytes, which can ultimately result in thrombotic and inflammatory (i.e., a thrombo-inflammatory) response to the treatment modality. In this concept review, we give an overview of the mechanisms of recognition within the innate immunity system, with the aim to identify suitable points for intervention. Finally, we discuss emerging and promising techniques for surface modification of biomaterials and cells with specific inhibitors in order to diminish thromboinflammation and improve clinical outcome.
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Affiliation(s)
- Kristina N Ekdahl
- Department of Immunology, Genetics and Pathology (IGP), Rudbeck Laboratory C5:3, Uppsala University, SE-751 85 Uppsala, Sweden; Linnæus Center of Biomaterials Chemistry, Linnæus University, SE-391 82 Kalmar, Sweden
| | - Shan Huang
- Linnæus Center of Biomaterials Chemistry, Linnæus University, SE-391 82 Kalmar, Sweden
| | - Bo Nilsson
- Department of Immunology, Genetics and Pathology (IGP), Rudbeck Laboratory C5:3, Uppsala University, SE-751 85 Uppsala, Sweden
| | - Yuji Teramura
- Department of Immunology, Genetics and Pathology (IGP), Rudbeck Laboratory C5:3, Uppsala University, SE-751 85 Uppsala, Sweden; Department of Bioengineering, The University of Tokyo, 7-3-1Hongo, Bunkyo-Ku, Tokyo, 113-8656, Japan.
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Heparinization of cell surfaces with short peptide-conjugated PEG-lipid regulates thromboinflammation in transplantation of human MSCs and hepatocytes. Acta Biomater 2016; 35:194-205. [PMID: 26876877 DOI: 10.1016/j.actbio.2016.02.018] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/02/2015] [Revised: 02/05/2016] [Accepted: 02/11/2016] [Indexed: 12/14/2022]
Abstract
Infusion of therapeutic cells into humans is associated with immune responses, including thromboinflammation, which result in a large loss of transplanted cells. To address these problems, heparinization of the cell surfaces was achieved by a cell-surface modification technique using polyethylene glycol-conjugated phospholipid (PEG-lipid) derivatives. A short heparin-binding peptide was conjugated to the PEG-lipid for immobilization of heparin conjugates on the surface of human mesenchymal stem cells (hMSCs) and human hepatocytes. Here three kinds of heparin-binding peptides were used for immobilizing heparin conjugates and examined for the antithrombogenic effects on the cell surface. The heparinized cells were incubated in human whole blood to evaluate their hemocompatibility by measuring blood parameters such as platelet count, coagulation markers, complement markers, and Factor Xa activity. We found that one of the heparin-binding peptides did not show cytotoxicity after the immobilization with heparin conjugates. The degree of binding of the heparin conjugates on the cell surface (analyzed by flow cytometer) depended on the ratio of the active peptide to control peptide. For both human MSCs and hepatocytes in whole-blood experiments, no platelet aggregation was seen in the heparin conjugate-immobilized cell group vs. the controls (non-coated cells or control peptide). Also, the levels of thrombin-antithrombin complex (TAT), C3a, and sC5b-9 were significantly lower than those of the controls, indicating a lower activation of coagulation and complement. Factor Xa analysis indicated that the heparin conjugate was still active on the cell surface at 24h post-coating. It is possible to immobilize heparin conjugates onto hMSC and human hepatocyte surfaces and thereby protect the cell surfaces from damaging thromboinflammation. STATEMENT OF SIGNIGFICANCE We present a promising approach to enhance the biocompatibility of therapeutic cells. Here we used short peptide-conjugated PEG-lipid for cell surface modification and heparin conjugates for the coating of human hepatocytes and MSCs. We screened the short peptides to find higher affinity for heparinization of cell surface and performed hemocompatibility assay of heparinized human hepatocytes and human MSCs in human whole blood. Using heparin-binding peptide with higher affinity, not only coagulation activation but also complement activation was significantly suppressed. Thus, it was possible to protect human hepatocytes and human MSCs from the attack of thromboinflammatory activation, which can contribute to the improvement graft survival.
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Oldhafer F, Bock M, Falk CS, Vondran FWR. Immunological aspects of liver cell transplantation. World J Transplant 2016; 6:42-53. [PMID: 27011904 PMCID: PMC4801804 DOI: 10.5500/wjt.v6.i1.42] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/31/2015] [Revised: 10/21/2015] [Accepted: 12/08/2015] [Indexed: 02/05/2023] Open
Abstract
Within the field of regenerative medicine, the liver is of major interest for adoption of regenerative strategies due to its well-known and unique regenerative capacity. Whereas therapeutic strategies such as liver resection and orthotopic liver transplantation (OLT) can be considered standards of care for the treatment of a variety of liver diseases, the concept of liver cell transplantation (LCTx) still awaits clinical breakthrough. Success of LCTx is hampered by insufficient engraftment/long-term acceptance of cellular allografts mainly due to rejection of transplanted cells. This is in contrast to the results achieved for OLT where long-term graft survival is observed on a regular basis and, hence, the liver has been deemed an immune-privileged organ. Immune responses induced by isolated hepatocytes apparently differ considerably from those observed following transplantation of solid organs and, thus, LCTx requires refined immunological strategies to improve its clinical outcome. In addition, clinical usage of LCTx but also related basic research efforts are hindered by the limited availability of high quality liver cells, strongly emphasizing the need for alternative cell sources. This review focuses on the various immunological aspects of LCTx summarizing data available not only for hepatocyte transplantation but also for transplantation of non-parenchymal liver cells and liver stem cells.
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Lee CA, Dhawan A, Smith RA, Mitry RR, Fitzpatrick E. Instant Blood-Mediated Inflammatory Reaction in Hepatocyte Transplantation: Current Status and Future Perspectives. Cell Transplant 2016; 25:1227-36. [PMID: 26996786 DOI: 10.3727/096368916x691286] [Citation(s) in RCA: 33] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/30/2022] Open
Abstract
Hepatocyte transplantation (HT) is emerging as a promising alternative to orthotopic liver transplantation (OLT) in patients with certain liver-based metabolic disease and acute liver failure. Hepatocytes are generally infused into the portal venous system, from which they migrate into the liver cell plates of the native organ. One of the major hurdles to the sustained success of this therapy is early cell loss, with up to 70% of hepatocytes lost immediately following infusion. This is largely thought to be due to the instant blood-mediated inflammatory reaction (IBMIR), resulting in the activation of complement and coagulation pathways. Transplanted hepatocytes produce and release tissue factor (TF), which activates the coagulation pathway, leading to the formation of thrombin and fibrin clots. Thrombin can further activate a number of complement proteins, leading to the activation of the membrane attack complex (MAC) and subsequent hepatocyte cell death. Inflammatory cells including granulocytes, monocytes, Kupffer cells, and natural killer (NK) cells have been shown to cluster around transplanted hepatocytes, leading to their rapid clearance shortly after transplantation. Current research aims to improve cell engraftment and prevent early cell loss. This has been proven successful in vitro using pharmacological interventions such as melagatran, low-molecular-weight dextran sulphate, and N-acetylcysteine (NAC). Effective inhibition of IBMIR would significantly improve hepatocyte engraftment, proliferation, and function, providing successful treatment for patients with liver-based metabolic diseases.
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Affiliation(s)
- Charlotte A Lee
- Institute of Liver Studies, King's College London, School of Life Sciences and Medicine, King's College Hospital, London, UK
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Dhawan A. Clinical human hepatocyte transplantation: Current status and challenges. Liver Transpl 2015; 21 Suppl 1:S39-44. [PMID: 26249755 DOI: 10.1002/lt.24226] [Citation(s) in RCA: 41] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/28/2015] [Revised: 07/27/2015] [Accepted: 07/29/2015] [Indexed: 02/07/2023]
Affiliation(s)
- Anil Dhawan
- Department of Pediatric Hepatology, Cell Therapy Unit, National Institute for Health Research/Wellcome Trust King's Clinical Research Facility, King's College Hospital, London, UK
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Parys M, Nelson N, Koehl K, Miller R, Kaneene JB, Kruger JM, Yuzbasiyan-Gurkan V. Safety of Intraperitoneal Injection of Adipose Tissue-Derived Autologous Mesenchymal Stem Cells in Cats. J Vet Intern Med 2015; 30:157-63. [PMID: 26512713 PMCID: PMC4913639 DOI: 10.1111/jvim.13655] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/29/2015] [Revised: 07/17/2015] [Accepted: 09/29/2015] [Indexed: 12/18/2022] Open
Abstract
Background Chronic inflammatory diseases are common in cats and mesenchymal stem cells (MSC) are a promising therapeutic approach for management of these disorders. The purpose of this study was to evaluate the safety of intraperitoneal injection of MSC in cats. Hypothesis Intrapertioneal injection of autologous MSC in cats is safe. Animals Ten healthy adult purpose‐bred cats. Methods Mesenchymal stem cells were isolated from subcutaneous adipose tissue collected during ovariohysterectomy and characterized for expression of CD90, CD105 and CD44 and trilineage differentiation. Three weeks postoperatively a complete blood count, serum chemistry profile, urinalysis, and abdominal ultrasound were performed. Five cats then received 1 × 106 of autologous MSC/kg of body weight intraperitoneally with ultrasound guidance; 5 additional cats were sham injected. Cats were monitored for 6 weeks with daily physical examinations and weekly clinicopathological evaluations. Abdominal ultrasonography was repeated at weeks 1 and 5 after injection. Results Serious adverse effects were not observed in any MSC‐injected cat. Two animals developed transient lethargy and decreased activity. Jejunal lymph node size was increased in MSC‐injected cats compared to controls at weeks 1 (1.38 ± 0.25 versus 0.88 ± 0.25 cm2; P = .036) and 5 (1.75 ± 0.82 versus 0.79 ± 0.12 cm2; P = .047). A hyperechoic renal segmental cortical lesion was observed in 1 MSC‐injected cat. Conclusions and Clinical Relevance Intraperitoneal MSC injection was well tolerated with only mild, self‐limiting adverse effects being observed in 2 cats. This route provides a safe means of administration for cell‐based treatment in cats.
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Affiliation(s)
- M Parys
- Department of Small Animal Clinical Sciences, College of Veterinary Medicine, Michigan State University, East Lansing, MI
| | - N Nelson
- Department of Small Animal Clinical Sciences, College of Veterinary Medicine, Michigan State University, East Lansing, MI
| | - K Koehl
- Department of Small Animal Clinical Sciences, College of Veterinary Medicine, Michigan State University, East Lansing, MI
| | - R Miller
- Department of Large Animal Clinical Sciences, College of Veterinary Medicine, Michigan State University, East Lansing, MI.,Center for Population, College of Veterinary Medicine, Michigan State University, East Lansing, MI
| | - J B Kaneene
- Department of Large Animal Clinical Sciences, College of Veterinary Medicine, Michigan State University, East Lansing, MI.,Center for Population, College of Veterinary Medicine, Michigan State University, East Lansing, MI
| | - J M Kruger
- Department of Small Animal Clinical Sciences, College of Veterinary Medicine, Michigan State University, East Lansing, MI
| | - V Yuzbasiyan-Gurkan
- Department of Small Animal Clinical Sciences, College of Veterinary Medicine, Michigan State University, East Lansing, MI
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Hårdstedt M, Lindblom S, Karlsson-Parra A, Nilsson B, Korsgren O. Characterization of Innate Immunity in an Extended Whole Blood Model of Human Islet Allotransplantation. Cell Transplant 2015; 25:503-15. [PMID: 26084381 DOI: 10.3727/096368915x688461] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022] Open
Abstract
The instant blood-mediated inflammatory reaction (IBMIR) has been studied in whole blood models of human allo-islet transplantation for short periods (<6 h). Beyond this time frame the innate response to intraportally transplanted islets is less well described. A novel whole blood model was applied to study blood-islet-graft interactions up to 48 h. Heparinized polyvinyl chloride tubing was sealed into small bags containing venous blood together with allogeneic human islets and exocrine tissue, respectively. The bags were attached to a rotating wheel (37°C). Concentrated glucose and sodium hydrogen carbonate were added every 12 h to maintain physiological limits for sustained immune cell functions. Plasma was collected at repeated time points for analyses of coagulation/complement activation and chemokine/cytokine production. Immune cell infiltration was analyzed using immunohistochemistry. Coagulation and platelet activation markers, thrombin-antithrombin complex (TAT) and soluble CD40 ligand (sCD40L) showed early high concentrations (at 6-12 h). sC5b-9 steadily increased over 48 h. At 6 h neutrophils and monocytes surrounded the clotted cellular grafts with a following massive infiltration of neutrophils. High and increasing concentrations of CXCR1/2 ligands [IL-8 and growth-regulated oncogene α/β/γ (Gro-α/β/γ)] and IL-6 were produced in response to human islets and exocrine tissue. The CCR2 ligand monocyte chemoattractant protein 1 (MCP-1) exhibited increasing concentrations in response to exocrine tissue. The CXCR3 ligand interferon-inducible T cell α chemoattractant (I-TAC) was produced in response to both human islets and exocrine tissue from 6 h. Monokine induced by γ interferon (Mig) and interferon γ-induced protein 10 (IP-10) showed a later response, preferentially to exocrine tissue and with larger variations among preparations. An extended blood model of clinical islet transplantation allowed characterization of early immune activation in response to human islets and exocrine tissue. Increased production of chemokines targeting CXCR1/2, CCR2, and CXCR3 was observed, accompanied by massive intraislet neutrophil infiltration over 48 h. The model proved to be useful in exploring early blood-mediated reactions to cellular transplants and has relevance for evaluation of pharmacological interventions to prevent graft loss.
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Affiliation(s)
- Maria Hårdstedt
- Department of Immunology, Genetics and Pathology, Clinical Immunology, The Rudbeck Laboratory, Uppsala University, Uppsala, Sweden
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Hårdstedt M, Lindblom S, Hong J, Nilsson B, Korsgren O, Ronquist G. A novel model for studies of blood-mediated long-term responses to cellular transplants. Ups J Med Sci 2015; 120:28-39. [PMID: 25322825 PMCID: PMC4389005 DOI: 10.3109/03009734.2014.965290] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/02/2023] Open
Abstract
AIMS Interaction between blood and bio-surfaces is important in many medical fields. With the aim of studying blood-mediated reactions to cellular transplants, we developed a whole-blood model for incubation of small volumes for up to 48 h. METHODS Heparinized polyvinyl chloride tubing was cut in suitable lengths and sealed to create small bags. Multiple bags, with fresh venous blood, were incubated attached to a rotating wheel at 37°C. Physiological variables in blood were monitored: glucose, blood gases, mono- and divalent cations and chloride ions, osmolality, coagulation (platelet consumption, thrombin-antithrombin complexes (TAT)), and complement activation (C3a and SC5b-9), haemolysis, and leukocyte viability. RESULTS Basic glucose consumption was high. Glucose depletion resulted in successive elevation of extracellular potassium, while sodium and calcium ions decreased due to inhibition of energy-requiring ion pumps. Addition of glucose improved ion balance but led to metabolic acidosis. To maintain a balanced physiological environment beyond 6 h, glucose and sodium hydrogen carbonate were added regularly based on analyses of glucose, pH, ions, and osmotic pressure. With these additives haemolysis was prevented for up to 72 h and leukocyte viability better preserved. Despite using non-heparinized blood, coagulation and complement activation were lower during long-term incubations compared with addition of thromboplastin and collagen. CONCLUSION A novel whole-blood model for studies of blood-mediated responses to a cellular transplant is presented allowing extended observations for up to 48 h and highlights the importance of stringent evaluations and adjustment of physiological conditions.
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Affiliation(s)
- Maria Hårdstedt
- Department of Immunology, Genetics and Pathology, Clinical Immunology, The Rudbeck Laboratory, Uppsala University, Uppsala, Sweden
- Center for Clinical Research Dalarna-Uppsala University, Falun, Sweden
| | - Susanne Lindblom
- Department of Immunology, Genetics and Pathology, Clinical Immunology, The Rudbeck Laboratory, Uppsala University, Uppsala, Sweden
| | - Jaan Hong
- Department of Immunology, Genetics and Pathology, Clinical Immunology, The Rudbeck Laboratory, Uppsala University, Uppsala, Sweden
| | - Bo Nilsson
- Department of Immunology, Genetics and Pathology, Clinical Immunology, The Rudbeck Laboratory, Uppsala University, Uppsala, Sweden
| | - Olle Korsgren
- Department of Immunology, Genetics and Pathology, Clinical Immunology, The Rudbeck Laboratory, Uppsala University, Uppsala, Sweden
| | - Gunnar Ronquist
- Department of Medical Sciences, Clinical Chemistry, Uppsala University, Uppsala, Sweden
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Thromboinflammation in Therapeutic Medicine. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2015; 865:3-17. [DOI: 10.1007/978-3-319-18603-0_1] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
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Kopec AK, Luyendyk JP. Coagulation in liver toxicity and disease: role of hepatocyte tissue factor. Thromb Res 2014; 133 Suppl 1:S57-9. [PMID: 24759146 DOI: 10.1016/j.thromres.2014.03.023] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
The liver is the primary source of a number of circulating coagulation factors, and acute liver injury and chronic liver disease are each associated with alterations in blood coagulation. Current views of the connection between liver injury and coagulation extend beyond the impact of liver disease on synthesis of coagulation factors to include a role for coagulation factor activity in the initiation and progression of liver disease. Mechanisms of coagulation initiation in liver disease are not completely understood. Compared to other tissues, liver expresses very low levels of tissue factor (TF). Recent studies indicate that expression of TF by hepatocytes comprises the majority of liver procoagulant activity, and that hepatocyte TF activates coagulation induced by liver injury. This review will briefly cover the expression and regulation of TF by hepatocytes, the role of TF in coagulation triggered by liver toxicity, and the contribution of coagulation activity to the progression of liver disease.
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Affiliation(s)
- Anna K Kopec
- Department of Pathobiology & Diagnostic Investigation, Michigan State University, East Lansing, MI 48824
| | - James P Luyendyk
- Department of Pathobiology & Diagnostic Investigation, Michigan State University, East Lansing, MI 48824.
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