Complement Biosynthesis in Human Synovial Tissue

Moffat, Graeme John (1989) Complement Biosynthesis in Human Synovial Tissue. PhD thesis, University of Glasgow.

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The complement system, which comprises a group of at least 20 plasma proteins, plays a major role in host defence against bacterial infections and in immunologically-mediated inflammation. The pro-inflammatory biological activities generated as a result of complement activation include increased vascular permeability, chemotaxis of neutrophils and mononuclear phagocytes, enzyme secretion by phagocytic cells and cytolysis. Complement activation occurs within inflamed synovial joints of patients with rheumatoid arthritis and is thought to play a role in the pathogenesis of the condition. Although plasma levels of most complement components are maintained by hepatic synthesis, the cell responsible being the hepatocyte, it is now well established that other cells including monocytes, macrophages, fibroblasts and epithelial cells of the gastrointestinal and genitourinary tracts synthesise complement components when cultured in vitro. Such cells are potential sources of extrahepatic synthesis of complement components in vivo. Synovial fluid macrophages and adherent synovial membrane cells have been shown to synthesise complement components when cultured in vitro, as have isolated fragments of synovial membrane from patients with either rheumatoid arthritis or osteoarthritis. However, the only evidence for extrahepatic synthesis occurring in vivo comes from metabolic studies with radiolabelled (125I) C3, which indicated that up to 50% of C3 present in the rheumatoid joint was synthesised locally. The principal aim of this study was to provide conclusive evidence that a large number of complement components including C4, C2, C3, C5, factor B and C1-inhibitor, were synthesised in synovial tissue in vivo. The availability of cDNA probes for these complement components allowed the use of molecular biological technology, in addition to immunochemical techniques, to study complement biosynthesis by synovial tissue from patients with rheumatoid arthritis or osteoarthritis and by normal synovial tissue from a patient undergoing patellectomy. Using Northern and dot-blot analyses, mRNAs coding for C1-inhibitor (2.1 kb), factor B (2.6 kb), C1q B-chain (1.4 kb), C2 (2.9 kb), C3 (5.2 kb) and C4 (5.5 kb) were detected in RNA isolated from both rheumatoid synovium and synovial membrane from osteoarthritis patients. Although the presence of C5 mRNA was also investigated, it was not detected in any sample. Dot-blot analysis showed the presence of mRNA coding for C1-inhibitor in RNA isolated from normal synovial membrane. Quantitative analyses of the data using the students t-test did not show any statistically significant difference between the steady state levels of C3, C1-inhibitor or factor B mRNA in synovium from rheumatoid arthritis and osteoarthritis patients. Although ' the levels of C2 mRNA were significantly higher in rheumatoid synovium, and C4 mRNA levels were higher in osteoarthritis synovium, the P value was only marginally significant in both cases (0. 05). When fragments of normal synovium or synovial membrane fragments from patients with rheumatoid arthritis or osteoarthritis, were cultured in vitro, synthesis of C1-inhibitor, C2, C3, C4 and factor B was detected by ELISA and C2, C3 and factor B were shown to be functionally active. Following the demonstration of complement biosynthesis in synovial tissue in vivo, the next step would be to identify the cells within the synovium reponsible for complement biosynthesis. A number of cell-types that are present in synovial membrane have previously been shown to be capable of synthesising complement components in vitro. These include mononuclear phagocytes and possibly the B-cells of the synovial membrane lining, fibroblasts and endothelial cells. However, the cells responsible for complement biosynthesis in vivo remain unidentified. This question may only be answered with the use of in situ hybridisation on frozen sections of synovial tissue. As an initial step in this study, cells were isolated from synovial tissue of patients with rheumatoid arthritis and osteoarthritis and cultured in vitro, ELISA analyses of the culture supernatants showed synthesis of C4, C3, factor B and C1-inhibitor by both the adherent and non-adherent synovial cells. Immunohistochemistry showed that in both cell populations, fibroblasts were present in by far the greatest abundance (>95%), although a small proportion (5%) of macrophages was identified. This study thus provides conclusive evidence that synthesis of complement components occurs locally within normal and inflamed synovial tissue in vivo. The local synthesis of complement within normal synovial joints may be of importance in their defence against infection, whereas in inflamed joints it may contribute to the inflammatory response. Further work is required to define the regulation of complement synthesis within the synovial joint as well as to identify the cells responsible for complement biosynthesis in vivo.

Item Type: Thesis (PhD)
Qualification Level: Doctoral
Keywords: Medicine, Pathology
Date of Award: 1989
Depositing User: Enlighten Team
Unique ID: glathesis:1989-77841
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 14 Jan 2020 11:53
Last Modified: 14 Jan 2020 11:53

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