| Renin angiotensin system (RAS) is composed of renin, angiotensinogen,angiotensin I ,angiotensin II (Ang II ),angiotensin-converting enzyme and angiotensin receptor(ATR). Ang II is the main bioactive peptide in RAS, Ang II produces biological effects by combined angiotensin receptor on target cells. Traditionally, RAS belongs to endocrine system, it primarily regulates the balance of blood pressure and water-electrolyte. Now people gradually realize that the expression of RAS component presents in many tissues, RAS can be divided into circular RAS (cRAS) and tissue RAS (tRAS). Some investigation shows cRAS primarily plays short-term effct, it keeps homeostasis; tRAS regulates long-term function of an organ,and is the main cause of organ structure alteration. Utilizing molecular biological techniques, people have discovered that all RAS components almost present in all parts of kidney. As a kind of renal endocrine system, paracrine system and cellular endocrine system, its important role in renal pathophysiology is gradually known.It is known that the traditional function of Ang II was mediated by angiotensin II type 1 receptors. Multiple evidences in animal models and clinical trials have indicated that ACEI and AT,RA not only effectly depressed blood pressure, but also ameliorated gromerular hemodynamics, decreased proteinuria, raised insuline sensibitity of tissues, and attenuated the development of diabetic nephropathy and other kinds of renal glomerular disease. Although evidences in animal models and clinical trials show that ACEI and AT,RA can produce effects of protecting renal function not depending on their depressing BP by blockading RAS, but their precise mechanisms are not understood. The promoting effect of Ang II on the proliferation of mesangial cells is essentially alleged on the basis of above trials. In vitro, the effect of Ang II and AT,RA on cell proliferation and expression of type IV collagen and TGF P , in cultured human mesangial cells (HMC) has few investigated.In present study, we mainly studied the effect of Ang II on cell proliferation and protein biosynthesis of type IV collagen and TGF P ,and the expression of TGF P , mRNA in primary cultured human fetal mesangial cells, observed the effect of AT,RA on cell proliferation, biosynthesis of ColIV and TGF P l and expression of TGF P ,mRNA in Ang II-stimulated HMC. The purpose of above experiments was to profoundly comprehend the effect of RAS on renal cells and the mechanism of AT,RA protecting renal function.Human fetal mesangial cells were originally cultured as previously described, which were assessed by immunohistochemistry method, using anti-vitronectin, anti-desmin and anti-cytokeratin monoclonal antibodies. We observed the effects of Ang II and AT,RA(losartan) on cell proliferation and cell cycle of HMC, using MTT incorporation and flow cytometry : HMC were transfered of culture and stick on wall for 24 hours; HMC were synchronized for 24 hours; adding Ang II and /or losartan to HMC and incubated for 24 or 48 hours; Removing supernatant, adding MTT, estimating cell proliferation of HMC using MTT incorporation;or harvesting and fixing HMC, adding PI DNA dye, detecting cell cycle of HMC. The effects of Ang II and losartan on biosynthsis of ColIV and TGF P , ofHMC were measured by ELISA: HMC were transfered of culture, synchronized and affected by Ang II and /or losartan, extracting supernatant of HMC, detecting the concentration of ColIV and TGF P , using ELISA. At the same time, we observed the effect of losartan on the expression of TGF P ,mRNA of Ang II -stimulated HMC, using RT-PCR: HMC were transfered of culture, synchronized and affected by Ang II and /or losartan for 8 hours, harvesting HMC, detecting the expression of TGF P ,mRNA of HMC by RT-PCR.The results showed: compared with control group, the cell numbers of HMC which were treated by different concentrations Ang II for 24 hours did not obviously changes; the cell numbers of HMC which were treated by different concentrations Ang II for 48 hours...
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