| Objective: To investigate the role of tissue inhibitor of metalloproteinase-1(TIMP-1)in diabetic nephropathy and its development., we study the possible mechanism by down regulate TIMP-1 in gene expression of diabetic nephropathy in rats.Methods: In this study, model of diabetic nephropathy rat was made .The DM rats were given retrovirus with sense human TIMP-1 cDNA and antisense human TIMP-1 cDNA transfect to the kidney. To observe each group of rat's renal function, kidney pathology morphology used by PAS,PAM dyeing, RT-PCR verified the expression of human TIMP-1, rat TIMP-1 and Weston Blot detected the change of rat TIMP-1,MMP-2,MMP-9,P-ERK,P-AKT,P-BAD(Ser112,Ser136)protein level , and observed the expression of rat TIMP-1,fibronection(FN),LAMINI(LN),â…£collagen by immunohistochemistry at 2, 4, 6, 8 week's time.Result: With the extension of the time, the diabetic rat kidney function gradually began to suffer, after 8 weeks the rat model succeeds, urine protein/urine creatinine, serum creatinine, blood urea nitrogen, relative kidney weight(KW/BW), the figure in DM model group was higher than normal group's (P<0.05). In DM-T group, the figure was increase remarkably (P<0.05), compared with DM group and the treatment groups were remarkable decrease (P<0.05). Nephridial tissue pathomorphology PAS,PAM showed that: In DM group, the mesangial region expanded obviously, matrix increased, glomerular basement membrane thickening, mesangial area mesangial matrix expansion, mesangial cells increased; in the DM-T group, appeared to change focal glomerulosclerosis. But the DM-ANTI group and the DM-Los group, the above pathology changeobviously reduced, matrix index remarkable dropped, mesangial matrix area reduced and mesangial cells reduced. The human TIMP-1 mRNA was only detected amplification fragment in the DM-T group and the DM-ANTI group kidney by RT-PCR, it means that the antisense succeed transfer in rat's kidney. Respectively analysis rat TIMP-1mRNA at 2,4,6,8 weeks, with the extension of the time, the DM group's expression was higher than normal group's ( P<0.05 ), in the DM-T group, it increased evidently(P<0.05), but the DM-ANTI group and the DM-Los group were decreased conspicuous(P<0.05). In the protein level of rat TIMP-1 was consonance with the gene level; compared with control group MMP-2,MMP-9's expression was accrescence in the DM group, the DM-T group was present more than the DM group(P<0.05), yet, the DM-ANTI group and the DM-Los group descented significantly than the DM group (P<0.05). In nephridial tissue, the immunohistochemistry results of rTIMP-1,FN,LN,â…£collagen were enhanced the masc dyeing areas in the DM group, compared with the DM group, the DM-T group obviously improved(P<0.05). The DM-ANTI group and the DM-Los group extracellular matrix was diminution(P<0.05). Meanwhile, we studied the cell signaling pathway MEK/ERK and PI3K/AKT, and found that p-ERK1/2,p-AKT,inhibit the apoptosis factor p-BAD(Ser112,Ser136)were increased in the DM group compared with the control group(P<0.05), the DM-T was upgrade than the DM group(P<0.05), also the DM-ANTI group and the DM-Los group was downgrade obviously(P<0.05).Conclusions:1. TIMP-1 is playing an important role in development of diabetic nephropathy.2. The antisense TIMP-1 down regulate the gene expression of TIMP-1,and protects rat renal function.3. TIMP-1 exerts its promote glomerular mesangial cells proliferation effect in diabetic nephropathy, at least in part, by activation of MEK/ERK,PI3K/AKT signal pathway.
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