Analysis Of Gene Expression Profile In Renal Tissue Of Hypertensive Mice And The Study Of Renoprotective Effect Of Aliskiren On Its Kidney

Objectives Through gene expression microarray screening of male human angiotensinogen-renin(AGT-REN) double transgenic hypertensive mice in kidney, then through further study of AT1, Mas and transcription regulation factors Ets1, osteopontin(OPN) and NADPH oxidase(NOX2 and NOX4) and induced nitric oxide synthase(i NOS) effect of aliskiren on AGT-REN double hypertensive transgenic mice kidney, to investigate the mechanism of effect of aliskiren on hypertensive renal damage.Methods Part 1: 1 Animal grouping: three10-month-old AGT-RENdouble transgenic hypertensive mice(HTgroup) and three C57B6 mice with the same background were used as wild tipe control group(WT group). 2 Arterial blood pressure monitoring: MAP was surveyed under sober and quiet state by non- invasive tail cuff method;Blood pressure of mice in each group for 29 days changes were observedevery other day. 3 Screening of differentially expressed genes in AGT-REN hypertensive mice and WT mice by mouse whole genome c DNA microarray. 4 Verify the results of gene chip by western blot(WB) method. Part 2: 1 Animal grouping and drug delivery methods: twelve 10-month-old male human AGT-Ren transgenic mice were randomly divided into two groups(n=6/group): HT group and HT+Aliskiren group; Six C57B6 mice with the same background were selected as wild type control group(WT group).By intraperitoneal injection, Aliskiren were given to HT+Aliskiren group with a dosage of 20mg/kg/d for 28 days, the rest of the two groups were given the same amount of normal saline as the controls. 2 The pathological changes of renal tissues were observed by light microscope and electron microscope. 3 Expression of kidney Ets1, OPN and AT1, Mas receptor and i NOS, NOX2, NOX4 were determined by WB.Results Part 1: 1 Blood pressure of AGT-Ren double transgenic hypertensive mice: Compared with WT group, the MAP of HT group was significantly higher. 2 Data analysis of chip result:(1)Screening of differentially expressed genes:The mouse genome c DNA microarray was used to find the differentially expressed genes from 33696 genes in the kidney of the mice of HT group and WTgroup.There are 20 genes up-regulated and 16 genes down-regulated in three groups.(2)Biological function of gene chip:There are 16 genes up-regulated, 9 genes down-regulated in molecular functional analysis; 14 genes upregulated, 7 genes down-regulated in biological pathway analysis and cell component analysis respectively.(3)Pathway analysis of gene chip: The differentially expressed genes mainly involved in arachidonic acid metabolism, fatty acid metabolism, renal cell carcinoma and other biological processes in KEGG sphingomyelin pathway analysis. 3 The validation of gene chip results: Western blot showed that the expression of Ets1 and OPN was significantly increased, and the expression was consistent with the results of gene chip. Part 2: 1 Blood pressure: compared with group WT mice, the MAP of HT group was significantly increased;After Aliskiren treatment, the MAP of HT+Aliskiren group was significantly lower than that in the HT group. 2 Pathological results of renal tissue: Under light microscope in renal tissue HE staining, glomerular mesangial matrix without hyperplasia, no inflammatory cell infiltration in interstitial tissue, tubular structure was normal, and no protein tube type; Glomerular sclerosis in HT group, most of the capillary vessel wall thickening and hyalinization, renal tubular atrophy off, visible protein tube type, inflammatory cell infiltration in renal interstitial tissue; the lesions improved significantly in HT+Aliskiren group.Under the electron microscope, the glomerular structure of WT group was complete and clear, the mitochondria of renal tubular epithelial cells were rich, and the arrangement was orderly; In HT group,the glomerular basement membrane thickening, fusion or disappearenceof foot process, the renal tubular epithelial cells mitochondria edema and vacuole degeneration, and the formation of a large number of collagen fibersin the stroma.Above renal pathological changes was significantly reduced in HT+Aliskiren group. 3 WB results: Compared with WT group, the expression of Ets1, OPN and AT1 receptor was significantly increased,Mas receptor expression was significantly decreased; i NOS, NOX2, NOX4 were increased in HT group. Compared with the HT group, the expression of Ets1, OPN and AT1 receptors and i NOS, NOX2, NOX4 were reduced, the expression of Mas receptor was raised apparently in HT+Aliskiren group.Conclusions 1 The activation of the renin angiotensin system can induce multi gene expression and the genes involved in the occurrence and development of hypertensive renal injury. 2 AngⅡ-Ets1-OPN pathway and AngⅡ-NADPH oxidase signaling pathway participate in the development of hypertension and hypertensive nephropathy. 3 Aliskiren plays a protective role in hypertensive renal injury byinhibiting the expression of Ets1, OPN and oxidative stress.