The Effect Of PACE4on The Founction Of RA And Breast Cancer Cells And The Research Of Mechanism

Background： Rheumatoid arthritis (rheumatoid arthritis, RA) is a chronic systemicautoimmune disease, the main pathological characteristics of RA reflected in the synovitis,synovial inflammation cell infiltration, abnormal proliferation of synovial cells, pannusformation, cartilage and bone injury. Synovial fibroblasts (rheumatoid arthritis synovialfibroblasts, RASFs) play a very vital role in the pathological process of rheumatoid cartilageand bone injuries with invasive and proliferative abilities. RASFs uncontrolled proliferationpromote a large number of inflammatory cell infiltration, which in turn results RASFshyperplasia, such a vicious circle further leads to irreversible damage in RA. Thepathogenesisi of RA is not yet clear, genetic and environmental factors have been confirmedby many studies. There are more women than men in the population of RA, statistics about2-4:1, the incidence of menopausal women is significantly higher than that in male andfemale elderly age, and the level of androgen and metabolism products in patients decreasedobviously, suggest that it may be related with the male and female hormone in RA patients.The synovial tissues of RA have a common characteristic with tumor tissues which representsat angiogenesis, fibrin deposition, abnormal proliferation and high thrombin activity. Weexplore the pathogenesis of RA and breast cancer at the same time. PACE4is aCa2+-dependent serine endoprotease which involves in a variety of activation and has theability to increase cell invasion and proliferation, Therefore, abnormal expression of PACE4in RA and breast cancer may be associated with the occurrence or progression of the disease. Objective: With si-RNA technology, we study the founction of PACE4in RASFs andMDA-MB-231breast cancer cells. While re-combinant protein of PACE4was used to inducecells of breast cancer cells to observe the effects of biological functions and mechanisms.Methods: Real-time PCR and western-blotting were used for detecting the expression ofPACE4in RA, OA and AS pathology tissues, and si-RNA technology was used to silence theexpression of PACE4in RASFs and MDA-MB-231cells respectively. MTT, scratches andtranswell methods were used to determinate the cell proliferation, migration and invasion.While we detected the cell cycle and inflammatory cytokines with flow cytometry and ELISA.Real-time PCR was also used for detecting PACE4expression after cell induced with estrogen.In addition, we performed re-combinant protein of PACE4to induce MDA-MB-231cells anddetected the biological founction and the effect on Erk, Wnt3a and stat3cell sinallingpathway.Results: Real-time PCR and western-blotting methods showed that PACE4expression in RAsynovium was significantly higher than that in OA and AS synovial tissues. Transientlytransfected siRNA technology in RASFs and MDA-MB-231cells was able to successfullysilence the expression of PACE4. When PACE4expression was silenced, both RASFs andMDA-MB-231cells reduced cell proliferation, decreased cell migration and invasion werefound obviously. Cell cycle experiment found that following PACE4silencing, cell cycle wasarrest in G0-G1phase. In addition, ELISA was used to detect the inflammatory cytokinesIL-1α, IL-1β, IL-17and TNF-α and found that TNF-α was significantly reduced withdecreasing expression of PACE4. When estrogen was used to induce RASFs andMDA-MB-231cells, the Mrna level of PACE4was detected and the results suggested thatestrogen acted no effect on the expression of PACE4. PACE4recombinant protein was used toinduce breast cancer cells and it was observed that could promote cell proliferation, cellmigration and enhance cell invasion. In addition, the added re-combinant protein couldactivate the Erk and Wnt3a ignaling pathways in MDA-MB-231breast cancer cells.Conlusion: The high expression of PACE4in RA synovial tissue and breast cancer indicatedthat the PACE4may be involved in the occurrence or development of rheumatoid arthritis andbreast cancer. Si-RNA technology could reduce the Mrna and protein levels of PACE4,indicated that transient transfection can specifically silence PACE4expression. MTT, cell scratch and transwell methods respectively used to detecte RASFs and MDA-MB-231cellsproliferation, migration and invasion, and re-PACE4recombinant protein was used to inducebreast cancer cells, the results showed that PACE4has great effect on cell biologicalfounction. Also PACE4involved in the regulation of the cell cycle, affecting the expression ofinflammatory factors on RASFs TNF-α, but the relationship between the expression ofestrogen in breast cancer and RA is not obvious. After induced by recombinant protein ofPACE4in breast cancer cells we found that it could activate Erk and Wnt3a ignalingpathway, suggesting that PACE4might activate Erk and Wnt3a ignaling pathways toregulate biological functions of the deseases.