An Experimental Study Of The Role Of Protein P27 In Cardiac Fibroblasts Proliferation Modulated By Arginine VasopressiiK Interleukin-1 3 And Interl

Essential hypertension (EH) is a common and reccurrent disease, which does great harm to people, but detailed etiological mechanisms have remained unidentified and there have been no effective prevention and treatment measures so far. It has been accepted that EH is a general disease resulted from multiple factors, with cell abnormal proliferation as a prominent pathological character Arginine vasopressin (AVP), a vasoconstrictive peptide secreted from hypothalamus, is believed to be involved in EH and its pathological leisons. Many studies show that AVP can induce hypertrophy of mycardial cells (MC) and proliferation of cardiac fibroblasts (CFs), thus result in left ventricular hypertrophy (LVH) and cardiac fibrosis. But the underlying precise mechamisms remain obscure. p27, a negative regulatory molecular of cell cycle engine which is involved in cell proliferation/differentiation and tumor development, can prevent most cells from entering S stage to proliferation by inhibiting cyclin-Cdks-7-(cyclin-dependent kinase) complexs. This study was therefore designed to investigate the role of p27 in CFs proliferation modulated by AVP and its intracelluar signal transduction mechanisms, study the effects of IL-1 ?and IL-6 on CFs proliferation and p27 expression, aiming to elucidate the cellular and molecular mechanisms of AVP on CFs proliferation, provide new experimental and theoritical evidences and treatment methods for cardiac fibrosis and LVH of EH. Experimental contents:As a model, CFs of neonatal Sprague-Dawley (SD) rats were cultured with enzymatic dissoiation. CFs proliferation or cell number was measured with MTT assay. Protein content of CFs was determined according to the modified Bradford's method. To identify the cell cycle profile, propidium iodide was used to lable DNA. p27 was labled with primary monoclone antibody and secondary antibody labled with FITC. Cell cycle distribution and positive rate of p27 expression was determined with flow cytometer (FCM). This study was to dynamically observe. (1) the modulation effects of AVP on CFs proliferation and hypertrophy; (2) the role of p27 in CFs proliferation induced by AVP; (3) the intracellular signal transduction mechanisms in CFs proliferation hypertrophy and p27 expression modulated by AW; (4) Effects of IL-1 ?on CFs proliferation^ hypertrophy and p27 expression under basal and AVP treatment conditions; (5) Effects of IL-6 intervention on CFs proliferation > hypertrophy and p27 expression under basal condition or induced by AVP. Experimental results:(1) A value of CFs by MTT assay increased graduately with the elevation of AVP concentrations. A value of 10'7M and 10"*M AVP groups were significantly higher than that of control (P<0.05> 0.01, respectively), as contrast,there was no-8-obvious difference between 10~8M, 10"9M AVP groups and control (both P>0.05); A value of CFs also increased with the lasting time of AW treatment. The result of 48h group under 10~7M AVP treatment condition was much higher than that of 24h group (P<0.01). (2) Protein content of CFs was measured with accordance to modified Bradford's method. AVP could promote protein content of CFs in a concentration and time dependent manner. Protein content of CFs inlO~7M and lO^M AVP groups were significantly higher than that of control (both P<0.01); And 48h group of 10"7 M AVP was also obviously higher than that of 24h group (P<0.01). (3) AVP could also increase the percentage of S stage cells and proliferation index (PI) in a concentration and time dependent manner, commitment with a decrease of Go/Gi stage cells percent. There was great difference in cell cycle distribution between 10"7Mx \Q*M AVP groups and control group (/><().01); Percentage of S stage cells and PI in CFs of 48h group were significantly higher than those of 24h group,while Go/Gi stage cells percent of 48h group was much lower than that of 24h group (both P<0.01). (4) p27 expression level in CFs decreased gradulately with the elevation of AVP concentrations. Positive rate of p27 expression in...