Functional analysis of plasminogen related genes in cartilage metabolism

本研究的目的是阐明PRGB和PRPB在类风湿关节炎等关节炎关节中的作用。我们研究了PRGB在类风湿关节炎中的表达及其在滑膜和软骨主要成分中的作用，并与血管生成密切相关。在体外正常条件下，滑膜细胞不表达PRGB，即使在细胞因子(IL-1和bFGF)刺激滑膜细胞以模拟类风湿关节炎的情况下，也不能诱导表达。在软骨中，PRGB在正常情况下不在软骨细胞中表达，但在细胞因子刺激下表达增强。在ISH中，PRGB在滑膜中不表达，但在RA关节软骨中有部分表达。这些发现表明，PRGB在正常关节软骨中不表达，但当受到刺激时，例如在炎症期间，其表达增强，尽管其机制尚不清楚。为了研究pRGB的作用，我们制备了重组P-…更多的RGB蛋白(PRPB)。在RA中，由于滑膜生长参与了RA的发病过程，因此我们研究了PRPB对滑膜细胞的影响。滑膜细胞在细胞因子(IL-1和bFGF)刺激下增殖。然而，PRPB并不能抑制其增殖。因此，我们重点研究了血管内皮细胞生长因子能强烈诱导类风湿关节炎炎症滑膜新生血管形成的假说，并探讨了pRPB对血管内皮生长因子表达的影响。首先，我们在细胞因子(IL-1和bFGF)刺激的滑膜细胞和软骨细胞中加入PRPB，并从基因和蛋白水平研究了VEGF表达的变化。PRPB在基因和蛋白水平上均能显著抑制滑膜细胞中血管内皮生长因子的表达，在基因水平上也能显著抑制软骨中血管内皮生长因子的表达。在活体研究中，使用了被认为代表类风湿性关节炎病理的CIA小鼠。与接受PBS治疗的对照组相比，PRPB治疗组的爪厚随着时间的推移而显著减少。此外，在Micro CT上，骨质侵蚀和破坏减少。关节滑膜组织中CD31抗体免疫组织化学染色显示，关节滑膜组织中CD31阳性的血管内皮细胞数量减少。提示PRPB治疗组可抑制关节炎，抑制血管生成。关于PRPB对血管内皮细胞的作用，其抑制作用是通过整合素发挥的。也有人认为，抑制滑膜组织产生的血管内皮生长因子抑制了血管生成。PRPB抑制血管内皮细胞生长因子的作用机制尚不清楚。这项研究证实了PRPB通过抑制血管生成来抑制关节炎，但其作用机制的许多细节尚不清楚，仍需进一步研究。较少

The purpose of this study is to clarify the role of PRGB and PRPB in arthritis joint such as RA. We investigated the expression of PRGB and the role of PRPB in the main components, synovium and cartilage, in rheumatoid arthritis closely involving angiogenesis. PRGB was not expressed in synoviocytes under normal conditions in vitro, and expression was not induced, even when the cells were stimulated with cytokines (IL-1 and bFGF) to mimic a condition similar to rheumatoid arthritis. In cartilage, PRGB was not expressed in chondrocytes under normal conditions, but the expression was enhanced by cytokine stimulation. On ISH, PRGB was not expressed in the synovial membrane, but was partially expressed in joint cartilage with RA. These findings suggested that PRGB is not expressed in normal joint cartilage, but that the expression is enhanced when stimulated, such as during inflammation, although the mechanism remains unclear. To investigate the action of PRGB, we prepared the recombinant P … More RGB protein (PRPB). In RA, the influence of PRPB on synoviocytes was investigated because synovium growth is involved in the pathogenesis. Synoviocytes proliferated when stimulated with cytokines (IL-1 and bFGF). However, PRPB did not inhibit the proliferation. Thus, we focused on the hypothesis that VEGF could strongly induce new blood vessel formation in inflammatory synovium in RA, and investigated the influence of PRPB on VEGF expression. Firstly, we added PRPB to synoviocytes and chondrocytes stimulated with cytokines (IL-1 and bFGF), and investigated the changes in VEGF expression at the gene and protein levels. PRPB significantly inhibited VEGF expression in synoviocytes at both the gene and protein levels, and in cartilage at the gene level. For in vivo study, CIA mice that are presumed to represent the pathology of rheumatoid arthritis were used. The paw thickness was significantly reduced over the time in the PRPB treatment group, compared to that in the control group treated with PBS. In addition, bone erosion and destruction were reduced on micro CT. Histopathologically, synovium growth and bone destruction were both reduced, and the number of CD31-positive vascular endothelial cells was also decreased on immunostaining of vascular endothelial cells in the articular synovial tissue with anti-CD31 antibody. It was suggested that arthritis was inhibited and angiogenesis was suppressed in the PRPB treatment group. Regarding the action of PRPB on vascular endothelial cells, the inhibitory effect was exerted via integrin. It was also suggested that inhibition of VEGF produced by the synovial tissue suppressed angiogenesis. The mechanism of action of the inhibition of VEGF by PRPB remains to be investigated. This study confirmed that PRPB inhibits arthritis by suppressing angiogenesis, but many of the details of the mechanism of action are unclear and remain to be investigated. Less