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1
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Dong XQ, Zhang YH, Luo J, Li MJ, Ma LQ, Qi YT, Miao YL. Keratin 1 modulates intestinal barrier and immune response via kallikrein kinin system in ulcerative colitis. World J Gastroenterol 2025; 31:102070. [PMID: 39958441 PMCID: PMC11752705 DOI: 10.3748/wjg.v31.i6.102070] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/08/2024] [Revised: 12/06/2024] [Accepted: 12/19/2024] [Indexed: 01/10/2025] Open
Abstract
BACKGROUND External factors in ulcerative colitis (UC) exacerbate colonic epithelial permeability and inflammatory responses. Keratin 1 (KRT1) is crucial in regulating these alterations, but its specific role in the progression of UC remains to be fully elucidated. AIM To explore the role and mechanisms of KRT1 in the regulation of colonic epithelial permeability and inflammation in UC. METHODS A KRT1 antibody concentration gradient test, along with a dextran sulfate sodium (DSS)-induced animal model, was implemented to investigate the role of KRT1 in modulating the activation of the kallikrein kinin system (KKS) and the cleavage of bradykinin (BK)/high molecular weight kininogen (HK) in UC. RESULTS Treatment with KRT1 antibody in Caco-2 cells suppressed cell proliferation, induced apoptosis, reduced HK expression, and increased BK expression. It further downregulated intestinal barrier proteins, including occludin, zonula occludens-1, and claudin, and negatively impacted the coagulation factor XII. These changes led to enhanced activation of BK and HK cleavage, thereby intensifying KKS-mediated inflammation in UC. In the DSS-induced mouse model, administration of KRT1 antibody mitigated colonic injury, increased colon length, alleviated weight loss, and suppressed inflammatory cytokines such as interleukin (IL)-1, IL-6, tumor necrosis factor-α. It also facilitated repair of the intestinal barrier, reducing DSS-induced injury. CONCLUSION KRT1 inhibits BK expression, suppresses inflammatory cytokines, and enhances markers of intestinal barrier function, thus ameliorating colonic damage and maintaining barrier integrity. KRT1 is a viable therapeutic target for UC.
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Affiliation(s)
- Xiang-Qian Dong
- Department of Gastroenterology, The First Affiliated Hospital of Kunming Medical University, Yunnan Province Clinical Research Center for Digestive Diseases, Kunming 650032, Yunnan Province, China
| | - Ying-Hui Zhang
- Department of Gastroenterology, Affiliated Hospital of Yunnan University, Kunming 650021, Yunnan Province, China
| | - Juan Luo
- Department of Gastroenterology, The First Affiliated Hospital of Kunming Medical University, Yunnan Province Clinical Research Center for Digestive Diseases, Kunming 650032, Yunnan Province, China
| | - Mao-Juan Li
- Department of Gastroenterology, The First Affiliated Hospital of Kunming Medical University, Yunnan Province Clinical Research Center for Digestive Diseases, Kunming 650032, Yunnan Province, China
| | - Lan-Qing Ma
- Department of Gastroenterology, The First Affiliated Hospital of Kunming Medical University, Yunnan Province Clinical Research Center for Digestive Diseases, Kunming 650032, Yunnan Province, China
| | - Ya-Ting Qi
- Department of Gastroenterology, The First Affiliated Hospital of Kunming Medical University, Yunnan Province Clinical Research Center for Digestive Diseases, Kunming 650032, Yunnan Province, China
| | - Ying-Lei Miao
- Department of Gastroenterology, The First Affiliated Hospital of Kunming Medical University, Yunnan Province Clinical Research Center for Digestive Diseases, Kunming 650032, Yunnan Province, China
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Wang X, Huang X, Gao P, Ren Y, Li X, Diao Y. Kallistatin attenuates inflammatory response in rheumatoid arthritis via the NF-κB signaling pathway. Eur J Pharmacol 2023; 943:175530. [PMID: 36690053 DOI: 10.1016/j.ejphar.2023.175530] [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: 10/18/2022] [Revised: 01/18/2023] [Accepted: 01/19/2023] [Indexed: 01/22/2023]
Abstract
Cartilage degeneration and inflammation are important features of rheumatoid arthritis (RA). Chondrocyte inflammation and apoptosis have been increasingly demonstrated to be related to cartilage decomposition. In this study, we analyzed the protective role of kallistatin against RA and its associated mechanisms. We obtained in vitro and in vivo RA models using IL-1β and heat-inactivated Mycobacterium tuberculosis, respectively. Our results showed that kallistatin mitigated IL-1β-mediated chondrocyte apoptosis and inhibited the synthesis of ECM-degrading generation, like matrix metalloproteinase (MMP)-3/13 and a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS)-4/5, in IL-1β-mediated chondrocytes. Furthermore, kallistatin markedly suppressed IL-1β-mediated inflammation while decreasing the levels of inflammatory factors and mediators via the NF-κB pathway. Daily administration of kallistatin reduced the expression levels of PGE2, TNF-α, IL-1β, and IL-6. Histochemical analysis revealed that the kallistatin-treated rats exhibited reduced RA severity compared with control mice. In summary, kallistatin suppressed IL-1β-mediated inflammation in chondrocytes via the NF-κB pathway. Administration of kallistatin remarkably inhibited RA development, accompanied by reduced inflammation and apoptosis. Therefore, kallistatin administration can be used as a candidate therapeutic strategy for RA.
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Affiliation(s)
- Xiao Wang
- School of Medicine, Huaqiao University, Quanzhou, 362021, China
| | - Xiaoping Huang
- College of Chemical Engineering and Materials Sciences, Quanzhou Normal University, Quanzhou, 326000, China
| | - Pingzhang Gao
- College of Chemical Engineering and Materials Sciences, Quanzhou Normal University, Quanzhou, 326000, China
| | - Yanxuan Ren
- School of Medicine, Huaqiao University, Quanzhou, 362021, China
| | - Xiaokun Li
- School of Medicine, Huaqiao University, Quanzhou, 362021, China
| | - Yong Diao
- School of Medicine, Huaqiao University, Quanzhou, 362021, China.
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3
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Pépin D, Shao ZQ, Huppé G, Wakefield A, Chu CW, Sharif Z, Vanderhyden BC. Kallikreins 5, 6 and 10 differentially alter pathophysiology and overall survival in an ovarian cancer xenograft model. PLoS One 2011; 6:e26075. [PMID: 22102857 PMCID: PMC3216928 DOI: 10.1371/journal.pone.0026075] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/23/2011] [Accepted: 09/19/2011] [Indexed: 12/31/2022] Open
Abstract
Human tissue kallikreins (KLKs) are members of a multigene family of serine proteases aberrantly expressed in many cancer types. In ovarian cancer, 12 KLKs are upregulated, and of those KLK5, 6 and 10 have been the focus of investigations into new diagnostic and prognostic biomarkers. However, little is known about the contributions of KLK5, 6 and 10 to ovarian cancer pathophysiology. In this study, a panel of 13 human ovarian cancer cell lines was screened by ELISA for secretion of KLK5, 6, 8, 10, 13, and 14. The ES-2 cell line, devoid of these kallikreins, was transfected with expression vectors of KLK5, 6 and 10 individually or in pairs. Co-expression of KLK5, 6 and 10 was correlated with lessened aggressivity of ovarian cancer cell lines as defined by reduced colony formation in soft agar and tumorigenicity in nude mice. ES-2 clones overexpressing KLK5, 10/5, 10/6, 5/6 made significantly fewer colonies in soft agar. When compared to control mice, survival of mice injected with ES-2 clones overexpressing KLK10, 10/5, 10/6, 5/6 was significantly longer, while KLK6 was shorter. All groups displaying a survival advantage also differed quantitatively and qualitatively in their presentation of ascites, with both a reduced incidence of ascites and an absence of cellular aggregates within those ascites. The survival advantage conferred by KLK10 overexpression could be recapitulated with the exogenous administration of a recombinant KLK10. In conclusion, these findings indicate that KLK5, 6 and 10 may modulate the progression of ovarian cancer, and interact together to alter tumour pathophysiology. Furthermore, results support the putative role of KLK10 as a tumour suppressor and suggest it may hold therapeutic potential in ovarian cancer.
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Affiliation(s)
- David Pépin
- Department of Cellular and Molecular Medicine, University of Ottawa, Ottawa, Ontario, Canada
- Centre for Cancer Therapeutics, Ottawa Health Research Institute, Ottawa, Ontario, Canada
| | | | | | | | - Chee-Wui Chu
- Ibex Pharmaceuticals Inc., Montreal, Quebec, Canada
| | - Zahra Sharif
- Department of Cellular and Molecular Medicine, University of Ottawa, Ottawa, Ontario, Canada
- Centre for Cancer Therapeutics, Ottawa Health Research Institute, Ottawa, Ontario, Canada
| | - Barbara C. Vanderhyden
- Department of Cellular and Molecular Medicine, University of Ottawa, Ottawa, Ontario, Canada
- Centre for Cancer Therapeutics, Ottawa Health Research Institute, Ottawa, Ontario, Canada
- * E-mail:
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4
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Liu Y, Pixley R, Fusaro M, Godoy G, Kim E, Bromberg ME, Colman RW. Cleaved high-molecular-weight kininogen and its domain 5 inhibit migration and invasion of human prostate cancer cells through the epidermal growth factor receptor pathway. Oncogene 2009; 28:2756-65. [PMID: 19483730 PMCID: PMC2861141 DOI: 10.1038/onc.2009.132] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/23/2008] [Revised: 03/06/2009] [Accepted: 04/18/2009] [Indexed: 12/20/2022]
Abstract
Upregulation and activation of epidermal growth factor receptor and/or urokinase-type plasminogen activator receptor in a variety of cancers have been shown to be associated with poor prognosis. High-molecular-weight kininogen can be hydrolysed by plasma kallikrein to bradykinin and cleaved high-molecular-weight kininogen (HKa). HKa and its domain 5 (D5) both have been shown to have potent anti-angiogenic activity. We now show that HKa blocks human prostate cancer cell (DU145) migration by 76.0+/-2.4% at 300 nM and invasion by 78.0+/-12.9% at 11.1 nM. D5 inhibits tumor migration and invasion in a concentration-dependent manner. Stimulation by basic fibroblast growth factor (bFGF) or vascular endothelial growth factor results in clustering of urokinase-type plasminogen activator receptor (uPAR) and epidermal growth factor receptor (EGFR) on the surface of DU145 cells. The co-localization of uPAR and EGFR is prevented by HKa. Immunoprecipitation suggests that uPAR, EGFR and alpha5beta1 integrin formed a ternary complex. Immunoblotting shows that HKa significantly decreases the bFGF-transactivated phosphorylation of EGFR at Tyr 1173 between 30 min and 4 h. The phosphorylation of extracellular signal-regulated kinase (ERK) and AKT, which are downstream effectors of EGFR, is also inhibited by HKa. These novel data indicate that HKa and D5 inhibit migration and invasion of human prostate cancer cells through an EGFR/uPAR pathway, suggesting the therapeutic potential of HKa and D5 to decrease metastasis of human prostate cancer.
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Affiliation(s)
- Y Liu
- The Sol Sherry Thrombosis Research Center, Temple University School of Medicine, Philadelphia, PA 19140, USA.
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5
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Bryant J, Shariat-Madar Z. Human plasma kallikrein-kinin system: physiological and biochemical parameters. Cardiovasc Hematol Agents Med Chem 2009; 7:234-50. [PMID: 19689262 PMCID: PMC4905712 DOI: 10.2174/187152509789105444] [Citation(s) in RCA: 97] [Impact Index Per Article: 6.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
The plasma kallikrein-kinin system (KKS) plays a critical role in human physiology. The KKS encompasses coagulation factor XII (FXII), the complex of prekallikrein (PK) and high molecular weight kininogen (HK). The conversion of plasma prekallikrein to kallikrein by the activated FXII and in response to numerous different stimuli leads to the generation of bradykinin (BK) and activated HK (HKa, an antiangiogenic peptide). BK is a proinflammatory peptide, a pain mediator and potent vasodilator, leading to robust accumulation of fluid in the interstitium. Systemic production of BK, HKa with the interplay between BK bound-BK receptors and the soluble form of HKa are key to angiogenesis and hemodynamics. KKS has been implicated in the pathogenesis of inflammation, hypertension, endotoxemia, and coagulopathy. In all these cases increased BK levels is the hallmark. In some cases, the persistent production of BK due to the deficiency of the blood protein C1-inhibitor, which controls FXII, is detrimental to the survival of the patients with hereditary angioedema (HAE). In others, the inability of angiotensin converting enzyme (ACE) to degrade BK leads to elevated BK levels and edema in patients on ACE inhibitors. Thus, the mechanisms that interfere with BK liberation or degradation would lead to blood pressure dysfunction. In contrast, anti-kallikrein treatment could have adverse effects in hemodynamic changes induced by vasoconstrictor agents. Genetic models of kallikrein deficiency are needed to evaluate the quantitative role of kallikrein and to validate whether strategies designed to activate or inhibit kallikrein may be important for regulating whole-body BK sensitivity.
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Affiliation(s)
- J.W. Bryant
- Pfizer Global Research and Development, CVMED Exploratory, Groton, CT 06340
| | - z Shariat-Madar
- School of Pharmacy, Department of Pharmacology, University of Mississippi, University, MS 38677-1848
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6
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Physiological responses to protein aggregates: Fibrinolysis, coagulation and inflammation (new roles for old factors). FEBS Lett 2009; 583:2691-9. [DOI: 10.1016/j.febslet.2009.06.013] [Citation(s) in RCA: 39] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/31/2009] [Revised: 06/10/2009] [Accepted: 06/10/2009] [Indexed: 01/06/2023]
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7
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Young WB, Rai R, Shrader WD, Burgess-Henry J, Hu H, Elrod KC, Sprengeler PA, Katz BA, Sukbuntherng J, Mordenti J. Small molecule inhibitors of plasma kallikrein. Bioorg Med Chem Lett 2006; 16:2034-6. [PMID: 16413183 DOI: 10.1016/j.bmcl.2005.12.060] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/10/2005] [Revised: 12/13/2005] [Accepted: 12/15/2005] [Indexed: 11/25/2022]
Abstract
Plasma kallikrein is a serine protease that is involved in pathways of inflammation, complement fixation, coagulation, and fibrinolysis. Herein, we describe the SAR and structural binding modes of a series of inhibitors of plasma kallikrein as well as the pharmacokinetics of a lead analog 11 in rat.
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Affiliation(s)
- Wendy B Young
- Celera Genomics, 180 Kimball Way, South San Francisco, CA 94080, USA.
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8
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Tang J, Yu CL, Williams SR, Springman E, Jeffery D, Sprengeler PA, Estevez A, Sampang J, Shrader W, Spencer J, Young W, McGrath M, Katz BA. Expression, crystallization, and three-dimensional structure of the catalytic domain of human plasma kallikrein. J Biol Chem 2005; 280:41077-89. [PMID: 16199530 DOI: 10.1074/jbc.m506766200] [Citation(s) in RCA: 59] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Plasma kallikrein is a serine protease that has many important functions, including modulation of blood pressure, complement activation, and mediation and maintenance of inflammatory responses. Although plasma kallikrein has been purified for 40 years, its structure has not been elucidated. In this report, we described two systems (Pichia pastoris and baculovirus/Sf9 cells) for expression of the protease domain of plasma kallikrein, along with the purification and high resolution crystal structures of the two recombinant forms. In the Pichia pastoris system, the protease domain was expressed as a heterogeneously glycosylated zymogen that was activated by limited trypsin digestion and treated with endoglycosidase H deglycosidase to reduce heterogeneity from the glycosylation. The resulting protein was chromatographically resolved into four components, one of which was crystallized. In the baculovirus/Sf9 system, homogeneous, crystallizable, and nonglycosylated protein was expressed after mutagenizing three asparagines (the glycosylation sites) to glutamates. When assayed against the peptide substrates, pefachrome-PK and oxidized insulin B chain, both forms of the protease domain were found to have catalytic activity similar to that of the full-length protein. Crystallization and x-ray crystal structure determination of both forms have yielded the first three-dimensional views of the catalytic domain of plasma kallikrein. The structures, determined at 1.85 A for the endoglycosidase H-deglycosylated protease domain produced from P. pastoris and at 1.40 A for the mutagenically deglycosylated form produced from Sf9 cells, show that the protease domain adopts a typical chymotrypsin-like serine protease conformation. The structural information provides insights into the biochemical and enzymatic properties of plasma kallikrein and paves the way for structure-based design of protease inhibitors that are selective either for or against plasma kallikrein.
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Affiliation(s)
- Jie Tang
- Department of Structural Chemistry, Celera Genomics, South San Francisco, California 94080, USA
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9
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Isordia-Salas I, Pixley RA, Sáinz IM, Martínez-Murillo C, Colman RW. The role of plasma high molecular weight kininogen in experimental intestinal and systemic inflammation. Arch Med Res 2005; 36:87-95. [PMID: 15900628 DOI: 10.1016/j.arcmed.2005.02.001] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/28/2023]
Abstract
Inflammation is accompanied by activation of the plasma kallikrein-kinin system (KKS). KKS activation has been demonstrated in a variety of inflammatory human diseases. To further explore the participation of KKS in arthritis and inflammatory bowel disease, we used two experimental animal models in arthritis and enterocolitis. We found that activation of KKS is associated with arthritis induced by intraperitoneal injection of peptidoglycan-polysaccharide polymers (PG-PS) as well as the enterocolitis and systemic inflammation induced also by PG-PS when injected into the intestinal wall of genetically susceptible Lewis rats. We postulated that KKS participates in the pathogenesis of inflammatory reactions involved in cellular injury, coagulation, fibrinolysis, kinin formation, complement activation, cytokine secretion, and release of proteases. We demonstrated that therapy with a specific plasma kallikrein inhibitor modulated the experimental enterocolitis, arthritis, and systemic inflammation. The fact that deficiency of plasma high molecular weight kininogen in the genetically susceptible Lewis rat results in decreased chronic enterocolitis and systemic inflammation also supports our hypothesis. We suggest that KKS plays a similar role in idiopathic human intestinal inflammatory disease and arthritis, making kallikrein-kinin system proteins appealing targets for drug therapy in chronic inflammatory diseases such as rheumatoid arthritis and Crohn's disease.
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Affiliation(s)
- Irma Isordia-Salas
- The Sol Sherry Thrombiosis Research Center, Temple University School of Medicine, Philadelphia, PA 19140, USA
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10
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Isordia-Salas I, Pixley RA, Sáinz IM, Martínez-Murillo C, Colman RW. The role of plasma high molecular weight kininogen in experimental intestinal and systemic inflammation. Arch Med Res 2005; 35:369-77. [PMID: 15610905 DOI: 10.1016/j.arcmed.2004.05.004] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/01/2004] [Accepted: 05/27/2004] [Indexed: 11/23/2022]
Abstract
Inflammation is accompanied by activation of the plasma kallikrein-kinin system (KKS). KKS activation has been demonstrated in a variety of inflammatory human diseases. To further explore the participation of KKS in arthritis and inflammatory bowel disease, we used two experimental animal models in arthritis and enterocolitis. We found that activation of KKS is associated with arthritis induced by intraperitoneal injection of peptidoglycan-polysaccharide polymers (PG-PS) as well as the enterocolitis and systemic inflammation induced also by PG-PS when injected into the intestinal wall of genetically susceptible Lewis rats. We postulated that KKS participates in the pathogenesis of inflammatory reactions involved in cellular injury, coagulation, fibrinolysis, kinin formation, complement activation, cytokine secretion, and release of proteases. We demonstrated that therapy with a specific plasma kallikrein inhibitor modulated the experimental enterocolitis, arthritis, and systemic inflammation. The fact that deficiency of plasma high molecular weight kininogen in the genetically susceptible Lewis rat results in decreased chronic enterocolitis and systemic inflammation also supports our hypothesis. We suggest that KKS plays a similar role in idiopathic human intestinal inflammatory disease and arthritis, making kallikrein-kinin system proteins appealing targets for drug therapy in chronic inflammatory diseases such as rheumatoid arthritis and Crohn's disease.
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Affiliation(s)
- Irma Isordia-Salas
- The Sol Sherry Thrombosis Research Center, Temple University School of Medicine, Philadelphia, PA 19140, USA
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11
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Keith JC, Sainz IM, Isordia-Salas I, Pixley RA, Leathurby Y, Albert LM, Colman RW. A monoclonal antibody against kininogen reduces inflammation in the HLA-B27 transgenic rat. Arthritis Res Ther 2005; 7:R769-76. [PMID: 15987478 PMCID: PMC1175023 DOI: 10.1186/ar1728] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/31/2005] [Accepted: 03/03/2005] [Indexed: 01/12/2023] Open
Abstract
The human leukocyte antigen B27 (HLA-B27) transgenic rat is a model of human inflammatory bowel disease, rheumatoid arthritis and psoriasis. Studies of chronic inflammation in other rat models have demonstrated activation of the kallikrein–kinin system as well as modulation by a plasma kallikrein inhibitor initiated before the onset of clinicopathologic changes or a deficiency in high-molecular-mass kininogen. Here we study the effects of monoclonal antibody C11C1, an antibody against high-molecular-mass kininogen that inhibits the binding of high-molecular-mass kininogen to leukocytes and endothelial cells in the HLA-B27 rat, which was administered after the onset of the inflammatory changes. Thrice-weekly intraperitoneal injections of monoclonal antibody C11C1 or isotype IgG1 were given to male 23-week-old rats for 16 days. Stool character as a measure of intestinal inflammation, and the rear limbs for clinical signs of arthritis (tarsal joint swelling and erythema) were scored daily. The animals were killed and the histology sections were assigned a numerical score for colonic inflammation, synovitis, and cartilage damage. Administration of monoclonal C11C1 rapidly decreased the clinical scores of pre-existing inflammatory bowel disease (P < 0.005) and arthritis (P < 0.001). Histological analyses confirmed significant reductions in colonic lesions (P = 0.004) and synovitis (P = 0.009). Decreased concentrations of plasma prekallikrein and high-molecular-mass kininogen were found, providing evidence of activation of the kallikrein–kinin system. The levels of these biomarkers were reversed by monoclonal antibody C11C1, which may have therapeutic potential in human inflammatory bowel disease and arthritis.
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Affiliation(s)
- James C Keith
- Department of Cardiovascular and Metabolic Diseases Research, Wyeth Research, Cambridge, Massachusetts, USA
| | - Irma M Sainz
- The Sol Sherry Thrombosis Research Center, Temple University School of Medicine, Philadelphia, Pennsylania, USA
| | - Irma Isordia-Salas
- The Sol Sherry Thrombosis Research Center, Temple University School of Medicine, Philadelphia, Pennsylania, USA
| | - Robin A Pixley
- The Sol Sherry Thrombosis Research Center, Temple University School of Medicine, Philadelphia, Pennsylania, USA
| | - Yelena Leathurby
- Department of Cardiovascular and Metabolic Diseases Research, Wyeth Research, Cambridge, Massachusetts, USA
| | - Leo M Albert
- Department of Cardiovascular and Metabolic Diseases Research, Wyeth Research, Cambridge, Massachusetts, USA
| | - Robert W Colman
- The Sol Sherry Thrombosis Research Center, Temple University School of Medicine, Philadelphia, Pennsylania, USA
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12
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Okerberg ES, Wu J, Zhang B, Samii B, Blackford K, Winn DT, Shreder KR, Burbaum JJ, Patricelli MP. High-resolution functional proteomics by active-site peptide profiling. Proc Natl Acad Sci U S A 2005; 102:4996-5001. [PMID: 15795380 PMCID: PMC555687 DOI: 10.1073/pnas.0501205102] [Citation(s) in RCA: 80] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022] Open
Abstract
Characterization and functional annotation of the large number of proteins predicted from genome sequencing projects poses a major scientific challenge. Whereas several proteomics techniques have been developed to quantify the abundance of proteins, these methods provide little information regarding protein function. Here, we present a gel-free platform that permits ultrasensitive, quantitative, and high-resolution analyses of protein activities in proteomes, including highly problematic samples such as undiluted plasma. We demonstrate the value of this platform for the discovery of both disease-related enzyme activities and specific inhibitors that target these proteins.
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Affiliation(s)
- Eric S Okerberg
- ActivX Biosciences, 11025 North Torrey Pines Road, Suite 120, La Jolla, CA 92037, USA
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13
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Brooks SPJ, McAllister M, Sandoz M, Kalmokoff ML. Culture-independent phylogenetic analysis of the faecal flora of the rat. Can J Microbiol 2004; 49:589-601. [PMID: 14663493 DOI: 10.1139/w03-075] [Citation(s) in RCA: 57] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022]
Abstract
The dominant faecal flora of the rat was determined using randomly cloned 16S rDNA comparative sequence analysis. A total of 109 near full-length 16S rDNA clones were sequenced, representing 69 unique 16S rRNA phylotypes or operational taxonomic units (OTUs). Estimates of species richness indicated that approximately 338 species were present in the faeces, suggesting that only 20% of species were identified. Only two of 39 Gram-negative clones aligned with previously cultured species, the remainder fell into a separate lineage within the Bacteroides-Cytophaga phylum. Several clones within this new group were related to 16S rDNA sequences previously identified from mouse faeces. Lactobacilli were the most abundant Gram-positive species, representing 23% of the total clones but only 7% of OTUs. The remaining Gram-positive clones were distributed among the Clostridium coccoides group (9%), the Clostridium leptum subgroup (18%), and throughout the low GC Gram-positive bacteria (13%). The majority of OTUs (63/69 or 91%) were less than 97% homologous to previously cultured bacteria. Faecal samples were also cultured using a variety of anaerobic media. With the exception of the lactobacilli, the cultured isolates demonstrated low species diversity and poorly reflected the population, as defined through comparative sequence analysis.
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Affiliation(s)
- S P J Brooks
- Nutrition Research Division, Food Directorate, Health Products and Foods Branch, Health Canada, Ottawa, ON
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14
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Abstract
OBJECTIVES To review the literature for conditions, diseases, and disorders that affect activity of the contact factors, and further to review the literature for evidence that less than normal activity of any of the contact factors may be associated with thrombophilia. DATA SOURCES MEDLINE search for English-language articles published from 1988 to 2001 and pertinent references contained therein, as well as search of references in recent relevant articles and reviews. STUDY SELECTION Relevant clinical and laboratory information was extracted from selected articles. Meta-analysis was not feasible because of heterogeneity of reports. DATA EXTRACTION AND SYNTHESIS Evidence for association of altered levels of the contact factors and thrombophilia was sought. A wide variety of disorders is associated with decreased activity of the contact factors; chief among these disorders are liver disease, hepatic immaturity of newborns, the antiphospholipid syndrome, and, for factor XII, being of Asian descent. These disorders are more common than homozygous deficiency. The few series and case reports of thrombophilic events in patients homozygous for deficiency of contact factors are not persuasive enough to support causality. The apparent association between levels consistent with heterozygosity (40%-60% of normal) of any of the contact factors (but especially factor XII) in persons with antiphospholipid antibodies appears to be due to falsely decreased in vitro activity levels of these factors, which are normal on antigenic testing. The apparent association with thrombosis is better explained by the antiphospholipid syndrome than by the modest reduction of the levels of contact factors. CONCLUSIONS Presently, it is not recommended to measure activity of contact factors during routine evaluation of patients who have suffered venous or arterial thromboembolism or acute coronary syndromes.
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Affiliation(s)
- Craig S Kitchens
- Division of Hematology/Oncology, Department of Medicine, University of Florida, Gainesville, USA.
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15
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Kamat K, Hayashi I, Mizuguchi Y, Arai K, Saeki T, Ohno T, Saigenji K, Majima M. Suppression of dextran sulfate sodium-induced colitis in kininogen-deficient rats and non-peptide B2 receptor antagonist-treated rats. JAPANESE JOURNAL OF PHARMACOLOGY 2002; 90:59-66. [PMID: 12396028 DOI: 10.1254/jjp.90.59] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 10/27/2022]
Abstract
Various proinflammatory mediators are believed to be involved in the processes and symptoms of ulcerative colitis (UC). To determine whether endogenous kinin enhances the severity of UC, we induced experimental colitis (EC) in kininogen-deficient mutant rats and tested the effect of a non-peptide B2 receptor antagonist. EC was induced in male kininogen-deficient Brown Norway-Katholiek rats (BN-Ka) and normal Brown Norway-Kitasato rats (BN-Ki) with 5% dextran sulfate sodium (DSS). Sprague-Dawley rats (SD) were also used. Colon length, body weight and hematocrit were determined for 7 days. Effects of FR173657, an orally active B2 antagonist, were tested. The colon length was shortened in BN-Ki with DSS treatment, but not in BN-Ka, and the difference between their lengths was significant. The hematocrit value was also reduced in BN-Ki, and the difference in hematocrit between BN-Ki and BN-Ka was significant. In SD, shortening of the colon and reduction in hematocrit were also observable, and both were blunted by FR173657. The survival rate in SD given DSS for 7 days was 68%, but FR173657 treatment restored it significantly to 100%. These results suggest that the endogenous kinins generated from the kallikrein-kinin system have a significant role in the development of EC.
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Affiliation(s)
- Kazuhisa Kamat
- Department of Internal Medicine, Kitasato University Graduate School of Medical Sciences, Sagamihara, Kanagawa, Japan
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Sharma JN, Zubaid M, Khan BZSJ, Yusof APM, Asmawi MZ. Pathophysiological activities of the kallikrein-kinin system with emphasis on the cardiovascular disorders. Inflammopharmacology 2000. [DOI: 10.1163/156856000750260487] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/19/2022]
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