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Fibrosis of Peritoneal Membrane as Target of New Therapies in Peritoneal Dialysis. Int J Mol Sci 2022; 23:ijms23094831. [PMID: 35563220 PMCID: PMC9102299 DOI: 10.3390/ijms23094831] [Citation(s) in RCA: 40] [Impact Index Per Article: 13.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/06/2022] [Revised: 04/25/2022] [Accepted: 04/25/2022] [Indexed: 01/27/2023] Open
Abstract
Peritoneal dialysis (PD) is an efficient renal replacement therapy for patients with end-stage renal disease. Even if it ensures an outcome equivalent to hemodialysis and a better quality of life, in the long-term, PD is associated with the development of peritoneal fibrosis and the consequents patient morbidity and PD technique failure. This unfavorable effect is mostly due to the bio-incompatibility of PD solution (mainly based on high glucose concentration). In the present review, we described the mechanisms and the signaling pathway that governs peritoneal fibrosis, epithelial to mesenchymal transition of mesothelial cells, and angiogenesis. Lastly, we summarize the present and future strategies for developing more biocompatible PD solutions.
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Awonuga AO, Chatzicharalampous C, Thakur M, Rambhatla A, Qadri F, Awonuga M, Saed G, Diamond MP. Genetic and Epidemiological Similarities, and Differences Between Postoperative Intraperitoneal Adhesion Development and Other Benign Fibro-proliferative Disorders. Reprod Sci 2021; 29:3055-3077. [PMID: 34515982 DOI: 10.1007/s43032-021-00726-9] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/08/2021] [Accepted: 08/22/2021] [Indexed: 12/11/2022]
Abstract
Intraperitoneal adhesions complicate over half of abdominal-pelvic surgeries with immediate, short, and long-term sequelae of major healthcare concern. The pathogenesis of adhesion development is similar to the pathogenesis of wound healing in all tissues, which if unchecked result in production of fibrotic conditions. Given the similarities, we explore the published literature to highlight the similarities in the pathogenesis of intra-abdominal adhesion development (IPAD) and other fibrotic diseases such as keloids, endometriosis, uterine fibroids, bronchopulmonary dysplasia, and pulmonary, intraperitoneal, and retroperitoneal fibrosis. Following a literature search using PubMed database for all relevant English language articles up to November 2020, we reviewed relevant articles addressing the genetic and epidemiological similarities and differences in the pathogenesis and pathobiology of fibrotic diseases. We found genetic and epidemiological similarities and differences between the pathobiology of postoperative IPAD and other diseases that involve altered fibroblast-derived cells. We also found several genes and single nucleotide polymorphisms that are up- or downregulated and whose products directly or indirectly increase the propensity for postoperative adhesion development and other fibrotic diseases. An understanding of the similarities in pathophysiology of adhesion development and other fibrotic diseases contributes to a greater understanding of IPAD and these disease processes. At a very fundamental level, blocking changes in the expression or function of genes necessary for the transformation of normal to altered fibroblasts may curtail adhesion formation and other fibrotic disease since this is a prerequisite for their development. Similarly, applying measures to induce apoptosis of altered fibroblast may do the same; however, apoptosis should be at a desired level to simultaneously ameliorate development of fibrotic diseases while allowing for normal healing. Scientists may use such information to develop pharmacologic interventions for those most at risk for developing these fibrotic conditions.
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Affiliation(s)
- Awoniyi O Awonuga
- Division of Reproductive Endocrinology and Infertility, Department of Obstetrics and Gynecology, Wayne State University School of Medicine, Detroit, MI, 48201, USA.
| | - Charalampos Chatzicharalampous
- Division of Reproductive Endocrinology and Infertility, Department of Obstetrics and Gynecology, Wayne State University School of Medicine, Detroit, MI, 48201, USA
| | - Mili Thakur
- Reproductive Genomics Program, The Fertility Center, Grand Rapids, MI, USA.,Department of Obstetrics, Gynecology and Reproductive Biology, College of Human Medicine, Michigan State University, Grand Rapids, MI, USA
| | - Anupama Rambhatla
- Division of Reproductive Endocrinology and Infertility, Department of Obstetrics and Gynecology, Wayne State University School of Medicine, Detroit, MI, 48201, USA
| | - Farnoosh Qadri
- Division of Reproductive Endocrinology and Infertility, Department of Obstetrics and Gynecology, Wayne State University School of Medicine, Detroit, MI, 48201, USA
| | - Modupe Awonuga
- Division of Neonatology, Department of Pediatrics and Human Development, Michigan State University, 1355 Bogue Street, East Lansing, MI, USA
| | - Ghassan Saed
- Division of Reproductive Endocrinology and Infertility, Department of Obstetrics and Gynecology, Wayne State University School of Medicine, Detroit, MI, 48201, USA
| | - Michael P Diamond
- Department of Obstetrics and Gynecology, Augusta University, 1120 15th Street, CJ-1036, Augusta, GA, 30912, USA
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3
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Is There a Genetic Predisposition to Postoperative Adhesion Development? Reprod Sci 2020; 28:2076-2086. [PMID: 33090376 PMCID: PMC7579853 DOI: 10.1007/s43032-020-00356-7] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/27/2020] [Accepted: 10/11/2020] [Indexed: 12/11/2022]
Abstract
Adhesions are permanent fibrovascular bands between peritoneal surfaces, which develop following virtually all body cavity surgeries. The susceptibility to develop, and the severity, of adhesions following intra-abdominal surgery varies within and between individuals, suggesting that heritable factors influence adhesion development. In this manuscript, we discuss the pathophysiology of adhesion development from the perspective of genetic susceptibility. We restrict our discussion to genes and single-nucleotide polymorphisms (SNPs) that are specifically involved in, or that cause modification of, the adhesion development process. We performed a literature search using the PubMed database for all relevant English language articles up to March 2020 (n = 186). We identified and carefully reviewed all relevant articles addressing genetic mutations or single-nucleotide polymorphisms (SNPs) that impact the risk for adhesion development. We also reviewed references from these articles for additional information. We found several reported SNPs, genetic mutations, and upregulation of messenger RNAs that directly or indirectly increase the propensity for postoperative adhesion development, namely in genes for transforming growth factor beta, vascular endothelial growth factor, interferon-gamma, matrix metalloproteinase, plasminogen activator inhibitor-1, and the interleukins. An understanding of genetic variants could provide insight into the pathophysiology of adhesion development. The information presented in this review contributes to a greater understanding of adhesion development at the genetic level and may allow modification of these genetic risks, which may subsequently guide management in preventing and treating this challenging complication of abdominal surgery. In particular, the information could help identify patients at greater risk for adhesion development, which would make them candidates for anti-adhesion prophylaxis. Currently, agents to reduce postoperative adhesion development exist, and in the future, development of agents, which specifically target individual genetic profile, would be more specific in preventing intraperitoneal adhesion development.
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Abudukeyoumu A, Li MQ, Xie F. Transforming growth factor-β1 in intrauterine adhesion. Am J Reprod Immunol 2020; 84:e13262. [PMID: 32379911 DOI: 10.1111/aji.13262] [Citation(s) in RCA: 50] [Impact Index Per Article: 10.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/25/2020] [Revised: 04/30/2020] [Accepted: 05/01/2020] [Indexed: 02/06/2023] Open
Abstract
Intrauterine adhesion (IUA), led by trauma to the basal layer, can prevent the endometrium from growing, resulting in complications in females, such as infertility and amenorrhea. Transforming growth factor-β1 (TGF-β1) plays a crucial role in inducing and promoting the differentiation and proliferation of mesenchymal cells, in the secretion of extracellular matrix-associated components, and is a major cytokine in initiating and terminating tissue repair downstream of the TGF-β/Smad signaling pathway. Some evidence supports that TGF-β1 is closely associated with the occurrence and development of IUA, and is regarded as an early risk factor of disease recurrence. Furthermore, the role of TGF-β1 has been demonstrated to be potentially regulated by a variety of cytokines, hormones, enzymes, and microRNAs. This review provides an overview of the expression, function, and regulation of TGF-β1 in IUA, with a brief discussion and perspectives on its future clinical implications on the diagnosis and treatment of IUA.
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Affiliation(s)
- Ayitila Abudukeyoumu
- Laboratory for Reproductive Immunology, Institute of Obstetrics and Gynecology, Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, China.,Medical Center of Diagnosis and Treatment for Cervical Diseases, Obstetrics and Gynecology Hospital of Fudan University, Shanghai, China
| | - Ming-Qing Li
- Laboratory for Reproductive Immunology, Institute of Obstetrics and Gynecology, Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, China.,NHC Key Lab of Reproduction Regulation (Shanghai Institute of Planned Parenthood Research), Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, China.,Shanghai Key Laboratory of Female Reproductive Endocrine Related Diseases, Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, China
| | - Feng Xie
- Laboratory for Reproductive Immunology, Institute of Obstetrics and Gynecology, Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, China.,Medical Center of Diagnosis and Treatment for Cervical Diseases, Obstetrics and Gynecology Hospital of Fudan University, Shanghai, China.,Shanghai Key Laboratory of Female Reproductive Endocrine Related Diseases, Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, China
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Shukal D, Bhadresha K, Shastri B, Mehta D, Vasavada A, Johar K. Dichloroacetate prevents TGFβ-induced epithelial-mesenchymal transition of retinal pigment epithelial cells. Exp Eye Res 2020; 197:108072. [PMID: 32473169 DOI: 10.1016/j.exer.2020.108072] [Citation(s) in RCA: 20] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/25/2019] [Revised: 04/30/2020] [Accepted: 05/04/2020] [Indexed: 12/13/2022]
Abstract
Proliferative retinopathies are associated with formation of fibrous epiretinal membranes. At present, there is no pharmacological intervention for the treatment of retinopathies. Cytokines such as TGFβ are elevated in the vitreous humor of the patients with proliferative vitro-retinopathy, diabetic retinopathy and age-related macular degeneration. TGFβ isoforms lead to epithelial-mesenchymal transition (EMT) or trans-differentiation of the retinal pigment epithelial (RPE) cells. PI3K/Akt and MAPK/Erk pathways play important roles in the EMT of RPE cells. Therefore, inhibition of EMT by pharmacological agents is an important therapeutic strategy in retinopathy. Dichloroacetate (DCA) is shown to prevent proliferation and EMT of cancer cell lines but its effects are not explored on the prevention of EMT of RPE cells. In the present study, we have investigated the role of DCA in preventing TGFβ2 induced EMT of RPE cell line, ARPE-19. A wound-healing assay was utilized to detect the anti-EMT effect of DCA. The expressions of EMT and cell adhesion markers were carried out by immunofluorescence, western blotting, and quantitative real-time PCR. The expression of MAPK/Erk and PI3K/Akt pathway members was carried out using western blotting. We found that TGFβ2 exposure leads to an increase in the wound healing response, expression of EMT markers (Fibronectin, Collagen I, N-cadherin, MMP9, S100A4, α-SMA, Snai1, Slug) and a decrease in the expression of cell adhesion/epithelial markers (ZO-1, Connexin 43, E-cadherin). These changes were accompanied by the activation of PI3K/Akt and MAPK/Erk pathways. Simultaneous exposure of DCA along with TGFβ2 significantly inhibited wound healing response, expression of EMT markers and cell adhesion/epithelial markers. Furthermore, DCA and TGFβ2 effectively attenuated the activation of MAPK/Erk/JNK and PI3K/Akt/GSK3β pathways. Our results demonstrate that DCA has a strong anti-EMT effect on the ARPE-19 cells and hence can be utilized as a therapeutic agent in the prevention of proliferative retinopathies.
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Affiliation(s)
- Dhaval Shukal
- Department of Cell and Molecular Biology, Iladevi Cataract and IOL Research Centre, Ahmedabad, Gujarat, India; Manipal Academy of Higher Education, Manipal, Karnataka, India.
| | - Kinjal Bhadresha
- Department of Cell and Molecular Biology, Iladevi Cataract and IOL Research Centre, Ahmedabad, Gujarat, India.
| | - Bhoomi Shastri
- Department of Cell and Molecular Biology, Iladevi Cataract and IOL Research Centre, Ahmedabad, Gujarat, India.
| | - Deval Mehta
- Department of Cell and Molecular Biology, Iladevi Cataract and IOL Research Centre, Ahmedabad, Gujarat, India.
| | - Abhay Vasavada
- Department of Cell and Molecular Biology, Iladevi Cataract and IOL Research Centre, Ahmedabad, Gujarat, India.
| | - Kaid Johar
- Department of Zoology, BMTC, Human Genetics, USSC, Gujarat University, Ahmedabad, Gujarat, India.
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Fletcher NM, Awonuga AO, Abusamaan MS, Saed MG, Diamond MP, Saed GM. Adhesion phenotype manifests an altered metabolic profile favoring glycolysis. Fertil Steril 2016; 105:1628-1637.e1. [PMID: 26920255 DOI: 10.1016/j.fertnstert.2016.02.009] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/15/2015] [Revised: 02/03/2016] [Accepted: 02/03/2016] [Indexed: 10/22/2022]
Abstract
OBJECTIVE To determine whether metabolic markers are differentially expressed in normal and adhesion fibroblasts with and without hypoxia exposure. DESIGN Prospective experimental study. SETTING University research laboratory. PATIENT(S) Fibroblasts established from normal peritoneum and adhesion tissues from the same patients. INTERVENTION(S) In vitro experiments on normal peritoneal and adhesion fibroblasts under normal and hypoxic (2% O2) conditions. MAIN OUTCOME MEASURE(S) Expression of metabolic markers, glyceraldehyde 3-phosphate dehydrogenase (GAPDH), glucose transporter 1 (GLUT1), hypoxia inducible factor (HIF)-1α, hexokinase 2 (HK2), lactose dehydrogenase A (LDHA), and pyruvate dehydrogenase alpha 1 (PDHA1) were measured using real-time reverse transcription polymerase chain reaction; adenosine triphosphate (ATP), HIF-1α, and lactate levels were assessed with ELISAs. RESULT(S) Baseline mRNA levels of GAPDH and HIF-1α were increased, while GLUT1 and PDHA1 were decreased in adhesion as compared with in normal peritoneal fibroblasts. There was no change in baseline levels of HK2 or LDHA between the cell lines. Hypoxia increased protein levels of HIF-1α and mRNA levels of GAPDH, GLUT1, and HK2 and decreased levels of PDHA1 in both cell lines. Hypoxia increased LDHA mRNA levels in normal peritoneal fibroblasts. Baseline levels of lactate and ATP were lower in adhesion as compared with in normal peritoneal fibroblasts. In response to hypoxia, there was an increase in lactate in both cell lines and a decrease in ATP in normal fibroblasts. CONCLUSION(S) Adhesion fibroblasts manifested an altered metabolic profile, which favors the glycolytic pathway, and is further altered by hypoxia. Targeting these specific metabolic markers during surgery can be an important therapeutic intervention minimizing the development of postoperative adhesions.
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Affiliation(s)
- Nicole M Fletcher
- Department of Obstetrics and Gynecology, C.S. Mott Center for Human Growth and Development, Wayne State University School of Medicine, Detroit, Michigan
| | - Awoniyi O Awonuga
- Department of Obstetrics and Gynecology, C.S. Mott Center for Human Growth and Development, Wayne State University School of Medicine, Detroit, Michigan
| | - Mohammed S Abusamaan
- Department of Obstetrics and Gynecology, C.S. Mott Center for Human Growth and Development, Wayne State University School of Medicine, Detroit, Michigan
| | - Mohammed G Saed
- Department of Obstetrics and Gynecology, C.S. Mott Center for Human Growth and Development, Wayne State University School of Medicine, Detroit, Michigan
| | - Michael P Diamond
- Department of Obstetrics and Gynecology, Georgia Regents University, Augusta, Georgia
| | - Ghassan M Saed
- Department of Obstetrics and Gynecology, C.S. Mott Center for Human Growth and Development, Wayne State University School of Medicine, Detroit, Michigan.
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7
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Fletcher NM, Awonuga AO, Neubauer BR, Abusamaan MS, Saed MG, Diamond MP, Saed GM. Shifting anaerobic to aerobic metabolism stimulates apoptosis through modulation of redox balance: potential intervention in the pathogenesis of postoperative adhesions. Fertil Steril 2015. [DOI: 10.1016/j.fertnstert.2015.06.041] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/23/2022]
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Ambler DR, Fletcher NM, Diamond MP, Saed GM. Effects of hypoxia on the expression of inflammatory markers IL-6 and TNF-a in human normal peritoneal and adhesion fibroblasts. Syst Biol Reprod Med 2012; 58:324-9. [PMID: 23043632 DOI: 10.3109/19396368.2012.713439] [Citation(s) in RCA: 38] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
Inflammation is known to be involved in the postoperative adhesion development. Interleukin (IL)-6 and tumor necrosis factor (TNF)-α are cytokines that stimulate the acute-phase reaction, which leads to a systemic reaction including inflammation, fever, and activation of the complement and clotting cascades. The goal of this study was to examine the expression of these inflammatory markers, under normal and hypoxic conditions, in normal and adhesion fibroblasts. Primary cultures of fibroblasts were established from normal peritoneum and adhesion tissues from the same patient(s) and cultured under 20% O(2) or hypoxic 2% O(2) conditions for 24 hours. Cells were harvested and total RNA was isolated. Complimentary DNA was generated by reverse transcription and subjected to real-time RT-PCR using specific primers for IL-6 and TNF-α. Both normal peritoneal and adhesion fibroblasts expressed IL-6 and TNF-α. Adhesion fibroblasts exhibited significantly higher levels of IL-6 and TNF-α mRNA as compared to normal peritoneal fibroblasts (p < 0.05). Both IL-6 and TNF-α mRNA levels were upregulated in response to hypoxia in both normal peritoneal and adhesion fibroblasts. The increase in IL-6 and TNF-α mRNA levels of normal fibroblasts reached the levels observed in adhesion fibroblasts. Our results suggest that hypoxia promotes the development of the adhesion phenotype by the induction of inflammatory markers, which may contribute to the development of postoperative adhesions. The inhibition of inflammation may be a potential therapeutic approach in the prevention and/or reduction of postoperative adhesion development.
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Affiliation(s)
- Dana R Ambler
- Division of Reproductive Endocrinology and Infertility Department of Obstetrics and Gynecology, Wayne State University, Detroit, MI, USA
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9
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Uncoupling oxidative phosphorylation with 2,4-dinitrophenol promotes development of the adhesion phenotype. Fertil Steril 2012; 97:729-33. [DOI: 10.1016/j.fertnstert.2011.12.009] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/25/2011] [Revised: 12/02/2011] [Accepted: 12/06/2011] [Indexed: 12/24/2022]
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10
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Imai A, Suzuki N. Topical non-barrier agents for postoperative adhesion prevention in animal models. Eur J Obstet Gynecol Reprod Biol 2010; 149:131-5. [DOI: 10.1016/j.ejogrb.2009.12.026] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/06/2009] [Revised: 12/08/2009] [Accepted: 12/21/2009] [Indexed: 11/28/2022]
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11
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Imai A, Takagi H, Matsunami K, Suzuki N. Non-barrier agents for postoperative adhesion prevention: clinical and preclinical aspects. Arch Gynecol Obstet 2010; 282:269-75. [DOI: 10.1007/s00404-010-1423-3] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/01/2010] [Accepted: 03/01/2010] [Indexed: 11/24/2022]
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12
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Shavell VI, Saed GM, Diamond MP. Review: cellular metabolism: contribution to postoperative adhesion development. Reprod Sci 2009; 16:627-34. [PMID: 19293132 DOI: 10.1177/1933719109332826] [Citation(s) in RCA: 29] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/27/2022]
Abstract
Postoperative adhesions are a significant source of morbidity, including contributions to pelvic pain, bowel obstruction, and infertility. While the mechanisms of postoperative adhesion development are complex and incompletely understood, hypoxia appears to trigger a cascade of intracellular responses involving hypoxia-inducible factors, lactate, reactive oxygen species, reactive nitrogen species, and insulin-like growth factors that results in manifestation of the adhesion phenotype. Thus, substantial evidence exists to implicate the direct role of cellular metabolism in wound repair and adhesion development.
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Affiliation(s)
- Valerie I Shavell
- Department of Obstetrics and Gynecology, Division of Reproductive Endocrinology and Infertility, Wayne State University School of Medicine, Detroit, Michigan 48201, USA
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13
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Abstract
Peritoneal adhesions are a near inevitable occurrence after laparotomy and a major cause of both patient and physician misery. To date, clinical attempts at their amelioration have concentrated on manipulating the physical factors that affect their development despite a wealth of experimental data elucidating the molecular mechanisms that underlie their initiation, development and maturation. However, the advent of targeted, specific anti-cytokine agents as directed therapy for inflammatory and neoplastic conditions raises the prospect of a new era for anti-adhesion strategies. To harness this potential will require considerable cross-disciplinary collaboration and that surgeon-scientists propel themselves to the forefront of this emerging field.
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14
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Diamond MP, Saed G. Modulation of the expression of peroxisome proliferators-activated receptors in human fibroblasts. Fertil Steril 2006; 87:706-9. [PMID: 17156782 DOI: 10.1016/j.fertnstert.2006.07.1513] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/24/2005] [Revised: 07/06/2006] [Accepted: 07/06/2006] [Indexed: 11/28/2022]
Abstract
To determine the levels of peroxisome proliferators-activated receptors (PPARs) in normal and adhesion fibroblasts, we utilized real-time reverse transcription-polymerase chain reaction to measure messenger RNA (mRNA) levels in fibroblasts from normal peritoneum and adhesions from five patients in both the presence or absence of dichloroacetic acid (DCA) and a cyclooxygenase-2 (COX-2) inhibitor, NS-398. Peroxisome proliferators-activated receptor alpha, PPARbeta, PPARgamma1, and PPARgamma2 mRNA are all present in normal peritoneal and adhesion fibroblasts, and selectively rose in response to hypoxia and either DCA or NS-398.
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15
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Luo Y, Kobler JB, Zeitels SM, Langer R. Effects of Growth Factors on Extracellular Matrix Production by Vocal Fold Fibroblasts in 3-Dimensional Culture. ACTA ACUST UNITED AC 2006; 12:3365-74. [PMID: 17518673 DOI: 10.1089/ten.2006.12.3365] [Citation(s) in RCA: 44] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022]
Abstract
Culturing cells in 3-dimensional (3D) systems is important in tissue engineering and in fundamental studies of cellular mechanisms that are sensitive or specific to the 3D environment. To guide the engineering of artificial vocal fold lamina propria tissue, we developed 3D cultures containing human vocal fold fibroblasts (hVFFs) dispersed in a synthetic peptide hydrogel matrix. Growth factors were added to the culture to examine their influence on extracellular matrix (ECM) synthesis, cell proliferation, and matrix contraction. The hVFF-hydrogel constructs were treated with transforming growth factor-beta 1 (TGF-beta1), basic fibroblast growth factor (bFGF), or hepatocyte growth factor (HGF), and the culture was maintained for 21 days. TGF-beta1 induced matrix contraction and enhanced collagen and sulfated glycosaminoglycan production, bFGF effectively increased cell proliferation, and HGF stimulated synthesis of hyaluronic acid and elastin with less collagen accumulation than other conditions. Of the growth factors tested, HGF appears to be most useful for stimulating essential tissue components for restoring vocal fold pliability. The results also suggest that multiple growth factors might be employed sequentially or in combination to program the makeup of cell-hydrogel constructs for vocal fold tissue repair.
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Affiliation(s)
- Ying Luo
- Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA
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16
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Fujita T, Kambe N, Uchiyama T, Hori T. Type I Interferons Attenuate T Cell Activating Functions of Human Mast Cells by Decreasing TNF-α Production and OX40 Ligand Expression While Increasing IL-10 Production. J Clin Immunol 2006; 26:512-8. [PMID: 16988887 DOI: 10.1007/s10875-006-9043-1] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/20/2006] [Accepted: 08/22/2006] [Indexed: 11/29/2022]
Abstract
Recent studies have demonstrated that mast cells not only mediate inflammatory reactions in type I allergy but also play an important role in adaptive immunity. In the present study, we investigated the effects of interferon-alpha, which shares the same receptor as IFN-beta, on human cord blood-derived mast cells. Mast cells produced TNF-alpha, and IL-10, and expressed OX40 ligand upon activation by crosslinking of FcepsilonRI. When treated with interferon-alpha, TNF-alpha production was decreased while IL-10 and TGF-beta productions were increased. Furthermore, flow cytometric analysis revealed that interferon-alpha downregulated expression OX40 ligand on mast cells which is crucial for mast cell-T cell interaction. We confirmed that the viability of mast cells was not affected by interferon-alpha treatment. Accordingly, interferon-alpha-treated mast cells induced lower levels of CD4+ T cell proliferation compared with those without interferon-alpha treatment. These results suggest that type I interferons suppress T cell immune responses through their regulatory effects on mast cells.
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Affiliation(s)
- Tomoko Fujita
- Department of Hematology and Oncology, Graduate School of Medicine, Kyoto University, Kyoto 606-8507, Japan
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Christner PJ, Ayitey S. Extracellular matrix containing mutated fibrillin-1 (Fbn1) down regulates Col1a1, Col1a2, Col3a1, Col5a1, and Col5a2 mRNA levels in Tsk/+ and Tsk/Tsk embryonic fibroblasts. Amino Acids 2006; 30:445-51. [PMID: 16583319 DOI: 10.1007/s00726-005-0265-y] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/27/2005] [Accepted: 09/29/2005] [Indexed: 10/24/2022]
Abstract
It is known that the extracellular matrix (ECM) is able to signal to cells and thereby direct or modulate the transcription of certain mRNAs. This signaling plays an important role in tumor invasion and metastasis, wound healing, remodeling of the ECM and cell differentiation. There are several mechanisms whereby the ECM signals cells to change their metabolism: (1) receptor molecules binding to specific domains in the ECM, (2) direct phagocytosis of the ECM molecules or domains into the cell, (3) structural changes of the ECM domains. We report the effect of an ECM containing either mutant or normal Fbn1 on the transcription levels of several collagen mRNAs. Tsk/Tsk, Tsk/+ and +/+ mouse embryonic fibroblast cell lines were used. Tsk/Tsk cells produce only mutated fibrillin-1 which arises from mRNA containing an in-frame duplication of exons 17-40. To test the effect of the ECM containing mutant Fbn1, cells of the Tsk/Tsk, Tsk/+ and the wild-type (+/+) genotype were each grown on an ECM produced by either Tsk/Tsk, Tsk/+ cells or by wild-type cells (+/+). The embryonic cells were genotyped by Northern analyses for Fbn1 and grown to confluence. The cultures were then harvested and the cells removed, leaving the matrix in the flasks. Matrices produced from Tsk/Tsk, Tsk/+ and from +/+ cells were reseeded with Tsk/Tsk cells, Tsk/+ cells or +/+ cells. The cells were plated at a confluent concentration and incubated on the matrices for 48 h, after which total RNA was harvested and cDNA generated. Real-time PCR using cDNA or Northern analyses using RNA were performed for Fbn1 and Types I, III and V collagens. The PCR and Northern results were normalized using beta-actin and GAPDH, respectively. The Northern analyses showed that the steady state levels of mRNA for Col1a1 were depressed in both Tsk/Tsk and +/+ cells when grown on the matrix produced by Tsk/Tsk cells. Real-time PCR was then performed with primers specific for Col1a2, Col3a1, Col5a1 and Col5a2. The results showed that cells with the Tsk/Tsk, Tsk/+, and +/+ genotype all had lower steady-state levels of the above 4 collagen mRNAs when grown on the matrix produced by homozygous Tsk/Tsk cells or the matrix produced by heterozygous Tsk/+ cells compared with those grown on a matrix produced by +/+ cells. We hypothesize that the mutated Fbn1 molecules with many additional EGF-calcium binding regions and TGF-beta binding domains may (1) change the homeostasis of the ECM by binding additional growth factors and/or (2) present a radically different ECM 3-dimensional architecture. Either or both of these changes could signal the cell to produce less collagen.
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Affiliation(s)
- P J Christner
- Division of Rheumatology, Department of Medicine, Thomas Jefferson University, Philadelphia, Pennsylvania 19107-5541, USA.
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Funahashi H, Okada Y, Sawai H, Takahashi H, Matsuo Y, Takeyama H, Manabe T. The role of glial cell line-derived neurotrophic factor (GDNF) and integrins for invasion and metastasis in human pancreatic cancer cells. J Surg Oncol 2005; 91:77-83. [PMID: 15999351 DOI: 10.1002/jso.20277] [Citation(s) in RCA: 36] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/08/2022]
Abstract
BACKGROUND AND OBJECTIVES It is generally accepted that the malignancy of pancreatic cancer is dependent upon the extent of invasion as well as metastasis. However, the factors and mechanisms are incompletely understood. We investigated whether glial cell lined-derived neurotrophic factor (GDNF) enhances the invasive and adhesive behaviors of pancreatic cancer cells by altering of the expression of integrins. METHODS The expression of the GDNF receptor in pancreatic cancer cell lines (SW1990 and Capan-2) was confirmed by RT-PCR. Then we determined the expression of integrin subunits and the alteration of their expression by GDNF using flow-cytometric analysis and a cellular enzyme-linked immunosorbent assay (CELISA). Adhesion and invasion assay were performed to investigate whether increased integrin expression affected the interaction between cancer cells and ECM proteins. RESULTS The GDNF receptor subunits were expressed in pancreatic cancer cells. GDNF enhanced the expression of some of the integrin subunits and increased their adhesive and invasive abilities. The enhanced expression and associated increase in adhesive and invasive abilities were inhibited by blocking the GDNF receptor or the integrin beta1 subunit. CONCLUSION The enhancement of integrin expression by GDNF signaling through the GDNF receptor strongly influences invasion and adhesion to ECM proteins by pancreatic cancer cells.
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Affiliation(s)
- Hitoshi Funahashi
- Department of Gastroenterological Surgery, Nagoya City University Graduate School of Medical Sciences, Nagoya, Japan.
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Diamond MP, El-Hammady E, Munkarah A, Bieber EJ, Saed G. Modulation of the expression of vascular endothelial growth factor in human fibroblasts. Fertil Steril 2005; 83:405-9. [PMID: 15705382 DOI: 10.1016/j.fertnstert.2004.06.073] [Citation(s) in RCA: 48] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/16/2004] [Revised: 06/18/2004] [Accepted: 06/18/2004] [Indexed: 01/28/2023]
Abstract
OBJECTIVE To examine the up-regulation of vascular endothelial growth factor (VEGF) expression by hypoxia, a crucial event leading to neovascularization, as the reduction in VEGF expression may facilitate minimization of adhesion development. DESIGN Prospective experimental study. SETTING University medical center. PATIENT(S) Five patients with adhesions undergoing laparotomy with excision of adhesions and normal peritoneum. INTERVENTION(S) Adhesion and normal peritoneal fibroblasts were treated with dichloroacetic acid (DCA) or NS-398 (a cyclooxygenase-2 [COX-2] inhibitor) for 24 to 48 hours. MAIN OUTCOME MEASURE(S) A real-time reverse transcriptase polymerase chain reaction (RT-PCR) to quantify relative changes in mRNA levels of VEGF from each treatment. RESULT(S) In both normal peritoneal and adhesion fibroblasts, VEGF mRNA was present with statistically significantly higher levels in adhesion fibroblasts (32%). The DCA treatment resulted in a statistically significant decrease in VEGF mRNA levels in adhesion (20%) and normal peritoneal (18%) fibroblasts. The NS-398 treatment resulted in a statistically significant decrease in VEGF mRNA levels in adhesion (25%) and normal peritoneal (16%) fibroblasts. CONCLUSION(S) Stimulation of aerobic metabolism by DCA or inhibition of COX-2 by NS-398 reduces VEGF expression. Angiogenesis, which is an integral component in the development of dense vascular adhesions, may be reduced by either COX-2 inhibitors or stimulation of aerobic metabolism by DCA.
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Affiliation(s)
- Michael P Diamond
- Division of Reproductive Endocrinology and Infertility, Department of Obstetrics and Gynecology, Wayne State University/Detroit Medical Center, Detroit, MI 48201, USA.
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Wang YL, Pan CE, Yang PL, Tian Y, Pei SW, Dong M. Effects of Antiadhesion preparation on free fibrinogen and fibrin degrading products in abdominal exudates of rabbits postoperatively. World J Gastroenterol 2004; 10:2762-6. [PMID: 15309738 PMCID: PMC4572212 DOI: 10.3748/wjg.v10.i18.2762] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/15/2022] Open
Abstract
AIM: To observe effects of ACOL on fibrinogen (FIB), fibrin degrading products (FDP) and changes of FIB and FDP concentration in rabbits with intro-abdominal exudates during 7 d after major abdominal surgery.
METHODS: Sixty New Zealand rabbits were randomly divided into 4 groups: ACOL group, the control group, DCT group and the normal group. After being modeled, except the normal group, the other 3 groups were treated with different ways for a week; the intro-abdominal exudates of rabbits in the 4 groups were drawn for FIB and FDP measurement once daily during 7 d after major abdominal surgery.
RESULTS: FIB and FDP in the intro-abdominal exudates altered in a regular way and ACOL could change the concentration of FIB and FDP in the intra-abdominal exudates after major abdominal surgery.
CONCLUSION: ACOL can prevent intestinal adhesion by reducing the concentration of FIB and raising that of FDP in the intro-abdominal exudates after major abdominal surgery.
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Affiliation(s)
- You-Li Wang
- Department of Hepatobiliary Surgery, Xi'an Jiaotong University First Hospital, Xi'an 710061, Shaanxi Providence, China.
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Diamond MP, El-Hammady E, Wang R, Kruger M, Saed G. Regulation of expression of tissue plasminogen activator and plasminogen activator inhibitor-1 by dichloroacetic acid in human fibroblasts from normal peritoneum and adhesions. Am J Obstet Gynecol 2004; 190:926-34. [PMID: 15118614 DOI: 10.1016/j.ajog.2004.02.009] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/28/2022]
Abstract
OBJECTIVE As part of our ongoing studies to understand the biologic mechanisms of wound repair that lead to postoperative adhesions, we have identified characteristics of an adhesion phenotype that differs between fibroblasts that are obtained from human normal peritoneum and adhesions. In this study, we sought to examine whether stimulation of aerobic metabolism would alter differential expression of tissue plasminogen activator and plasminogen activator inhibitor-1, thereby creating a milieu likely to be less favorable to postoperative adhesion development. To examine this issue, we used a compound, dichloroacetic acid, that stimulates the pyruvate dehydrogenase complex, which causes pyruvate to be metabolized in the Kreb's cycle rather than being converted into lactate, thereby switching anaerobic to aerobic metabolism. STUDY DESIGN Human fibroblasts from normal peritoneum and adhesions were cultured in the absence or presence of dichloroacetic acid (100 microg/mL) for 24 hours, under normal and hypoxic (2% O(2)) conditions. Real-time reverse transcriptase-polymerase chain reaction of tissue plasminogen activator, plasminogen activator inhibitor-1, and a housekeeping gene beta-actin was performed with messenger RNA that was extracted from all treatment points. RESULTS Dichloroacetic acid stimulated normal peritoneal fibroblast tissue plasminogen activator messenger RNA expression under hypoxic conditions. In adhesion fibroblasts, dichloroacetic acid treatment enhanced tissue plasminogen activator messenger RNA expression under both normoxic and hypoxic conditions. Plasminogen activator inhibitor-1 messenger RNA expression was unaltered by dichloroacetic acid in normoxic normal peritoneal fibroblasts; but during culture under hypoxic conditions, dichloroacetic acid reduced plasminogen activator inhibitor-1 messenger RNA expression. Similarly, in adhesion fibroblasts, dichloroacetic acid reduced plasminogen activator inhibitor-1 messenger RNA expression under both normoxic and hypoxic conditions. As a result, in normal peritoneal fibroblasts under hypoxic conditions and in adhesion fibroblasts under normoxic and hypoxic conditions, dichloroacetic acid greatly increased the tissue plasminogen activator/plasminogen activator inhibitor-1 ratios. CONCLUSION These findings confirm that fibroblasts from adhesions are characterized by reduced tissue plasminogen activator and increased plasminogen activator inhibitor-1 production. These observations are extended to show the stimulation of oxidative metabolism by dichloroacetic acid increases tissue plasminogen activator expression under hypoxic conditions. Dichloroacetic acid reduces plasminogen activator inhibitor-1 production by hypoxic normal peritoneal fibroblasts and adhesion fibroblasts under hypoxic conditions. The resultant increases in the tissue plasminogen activator/plasminogen activator inhibitor-1 ratios would favor the development of a fibrinolytic milieu, which would be expected potentially to limit postoperative adhesion development. Thus, regulation of metabolic activity of peritoneal cells may provide a target for future interventions for the reduction of the development of postoperative adhesions, particularly as intervention relates to the healing of peritoneal sites that previously had adhesions. (eg, sites of potential adhesion reformation).
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Affiliation(s)
- Michael P Diamond
- Department of Obstetrics and Gynecology, Division of Reproductive Endocrinology and Infertility, Wayne State University/Detroit Medical Center, Hutzel Hospital, Detroit, MI 48201, USA.
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Diamond MP, El-Hammady E, Wang R, Saed G. Regulation of matrix metalloproteinase-1 and tissue inhibitor of matrix metalloproteinase-1 by dichloroacetic acid in human fibroblasts from normal peritoneum and adhesions. Fertil Steril 2004; 81:185-90. [PMID: 14711564 DOI: 10.1016/j.fertnstert.2003.05.006] [Citation(s) in RCA: 17] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/17/2022]
Abstract
OBJECTIVE To examine the role of stimulation of aerobic metabolism on the differential expression of matrix metalloproteinase-1 (MMP-1) and tissue inhibitor of matrix metalloproteinase-1 (TIMP-1), which are differentially regulated in fibroblasts isolated from normal human peritoneum and adhesions. DESIGN Tissue culture study. SETTING University research laboratory. PATIENT(S) Human fibroblasts cultures from normal peritoneum and adhesions that were exposed to dichloroacetic acid (DCA; 0 and 100 microg/mL) for 24 hours under normal and hypoxic conditions. MAIN OUTCOME MEASURE(S) Real-time reverse-transcription polymerase chain reaction of MMP-1, TIMP-1, and beta-actin. RESULT(S) Dichloroacetic acid stimulated peritoneal fibroblast MMP-1 mRNA expression under normoxic conditions; this stimulation was lost during hypoxia. In adhesion fibroblasts, DCA increased MMP-1 mRNA expression; this effect was reversed by hypoxia. Expression of TIMP-1 mRNA was insignificantly increased by DCA in normal peritoneal and adhesion fibroblasts under normoxic conditions; however under hypoxic conditions, DCA reduced TIMP-1 mRNA expression from both. CONCLUSION(S) Regulation of metabolic activity of peritoneal cells may provide a target for future interventions for reduction of development of postoperative adhesions, particularly as it relates to healing of peritoneal sites that did not previously have adhesions as opposed to sites that underwent lysis of preexistent adhesions.
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Affiliation(s)
- Michael P Diamond
- Department of Obstetrics and Gynecology, Wayne State University/Detroit Medical Center, Hutzel Hospital, Detroit, Michigan 48201, USA.
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