The MRNA-Stabilizing Factor Human-Antigen R Takes Part In Renal CyclinD1Expression Post-Transcriptionally Stimulated By Angiotensin Ⅱ In Human Mesangial Cells

Objective:The morphological and functional abnormality of human mesangial cells is one of important pathophysiological alterations in chronic renal diseases. AngⅡ over-expression can exist in injured kidney of chronic renal diseases, which can stimulate proliferation in human mesangial cells and increase in various intracellular protein synthesis. That can lead to morphological and functional changes in human mesangial cells. Therefore, AngⅡ has important stimulative roles in the starting and developmental course of chronic renal diseases. CyclinD1protein over-expression has an important role in cell proliferation, so that it can be regarded as effect protein of proliferation of human mesangial cells. MRNA-stabilizing factor HuR protein is mRNA binding protein, which can bind with mRNA of effect protein post-transcriptionally. It can strengthen mRNA stability and increase relevant protein expression to exert important pathophysiological effects. A survey suggests that HuR protein can bind with post-transcriptional mRNA of cyclinDl protein and strengthen its mRNA stability and increase relevant protein expression in a physiological state. In this survey Ang Ⅱ stimulation is applied to imitate a pathological state in chronic renal diseases, and investigate the role of mRNA stability factor HuR in angiotensin Ⅱ-stimulated cyclinD1over-expression. Methods:1.Experimental group:Human mesangial cells cultured in vitro were divided into OA group,3A group,6A group,9A group and24A group based on how long AngⅡ has stimulated.2. Cell cycle of OA group and24A group could be detected by flow cytometry.3.Cellular localization of HuR protein in each group could be detected by immunocytochemistry.4.Detection of protein expression:Total and cytoplasmic HuR protein and total cyclinD1protein in each group could be observed by western blot.5.Detection of RNA level:CyclinD1RNA level could be determined by RT-PCR.6. Angll protective effect on total cyclinD1protein expression when suppression of HuR protein could be surveyed by RNA interference techonology.Results:Cell proliferation trend of24A group could be more significant than that of OA group (P<0.05); HuR protein could shift from nucleus to cytoplasm, especially in3A group; Expression of total HuR protein remained unchanged, cytoplasmic HuR protein increased especially in3A group (P<0.05) and total cyclinD1protein in24A group (P<0.05), but no change in the RNA level. Compared with transfection before, expression of cytoplasmic HuR protein decreased clearly in3A group (P<0.05) and total cyclinDl protein in24A group (P<0.05), but no change in RNA level.Conclusion:1. AngⅡ induces proliferation of human mesangial cells.2.HuR protein shifts from nucleus to cytoplasm induced by Ang Ⅱ.3. HuR protein is involved in expression of cyclinD1protein induced by AngⅡ.