| Objective: Diabetic nephropathy (DN) is one of the commonest microangiopathies of diabetes mellitus (DM) and has become a major cause of end-stage renal failure (ESRF). There is an increasing tendency of the morbidity of DN in our country at present. It has become a very influential and powerful research topic in the field of DM and renal disease, whose purpose is to explore the pathogenesis of DN and to formulate more effective programs for it. It has been a long history for the traditional medicine's application to DM, as a result of which, the traditional medcine have accumulated much experience and a unique academic system is taking shape in it. And Chinese Herbs have shown a priority to chemical synthesis medcine with their mechanism exposed gradually. Chinese Herbs have a characteristic of different ways, different links, different targets. Much research has indicated that Chinese Herbs are able to enhance fitness, retrieve metabolic disorders of glucose, protein and fat while lowering the glucose level. Cordyceps sinensis(Berk.)Sacc is a kind of valuable, traditional and invigorating herbs in our country. It confirms that Cordyceps sinensis(Berk.)Sacc has effects of reinforcing kidney and lung, improving fragile conditions, and assisting pneuma. Basement-membrane thickening and mesangial expansion have long been recognized as pathological hallmark of DN. Type IV collagen (Col-Ⅳ) is the main component of the normal glomerular basement membrane and its thickening is the main road leading to DN. In fact, DN involves several mechanisms, among which TGF-β1 (transforming growth factor-β1), a key mesomerism, has been verificated to play an important role. TGF-β1 is a polypeptide depoted in the surface of the cells or ECM, mediating various chemical reaction to kidney induced by hyperglycaemia. Most studies suggest that free radicals mult with metabolic disorders of glucose, protein and fat in DM, which will injury kidney directly. Meanwhile, hyperglycaemia can affect the activities of many antioxidases [TSOD, CAT (catalase), GSH-PX(glutathione peroxidase)], therefore aggravating the injuries. In this study, type 1 diabetic rat model was established by intraperitoneal injection of streptozocin (STZ). The expression of Col-Ⅳand TGF-β1 and the activity of TSOD, CAT, GSH-PX, malondialdehyde(MDA) in the rat kidney were detected in order to discuss their roles in the pathogenic mechanics of DN. At the same time, Cordyceps Sinensis Cultivated by Artificial Fermentation was applied to investigate his prevention and treatment mechanisms of DN.Methods: Male SD rats were divided into two groups randomly: 12 normal control rats (group A) and 48 test rats. All rats were fed with routine diets. STZ(55mg/kg) was injected intraperitoneally to destroy some pancreas in order to induce hyperglycemia for test rats. 72 hours later, the rats whose blood glucose level was higher than 16.8 mmol/L in the subsequent 3 days were considered as diabetes. The diabetic rats were randomly divided into following groups: diabetic model rats (group B), diabetic model rats treated with 1.2g/kg/d Cordyceps Sinensis cultivated by Artificial Fermentation (group C), 2.4g/kg/d (group D) and diabetic rats treated with 3.6g/kg/d (group E). After 16 weeks of the experiment, body weight, blood glucose, serum triglyceride (TG), serum total cholesterol (TC), very low density lipoprotein cholesterol (VLDL-C), low density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C), urinary albumin excretion (UAE), blood urea nitrogen (BUN), serum creatinine (Scr), creatinine clearance (Ccr) were detected. Morphological changes were observed by microscope. Measured ECM content in the kidney by picric acid - toluidine blue staining. The expression of Col-Ⅳand TGF-β1 in rat kidney were detected by immunohistochemistry, and the results were analyzed by computer image-analysis system and the IOD of Col-Ⅳin the rat kidney tissue were counted. The activity of TSOD, MDA, CAT, GSH-PX was measured by biochemistry. All the experimental data were dealt with SPSS12.0.Results1 At the end of 4 weeks, the test rats were lighter than group A (all P<0.05). The blood glucose of test rats were significantly higher than those of control rats (all P<0.01).2 When the experiment was finished at end of the 16th week, blood glucose of group B was significantly higher than that of group A (P<0.01), and blood glucose of group C and D was significantly lower than that of group B and E(all P<0.01). Serum TG, TC, LDL-C and VLDL-C of group B were significantly higher than that of group A (P<0.01), and all the above indexes decreased obviously in group C, D and E compared with those in group B. HDL-C in group B decreased than group A (P<0.01), while increased in group C, D and E (all P<0.01). And blood glucose of group C and D was significantly lower than that of group B and E (all P<0.01), while there was no difference in blood glucose between group B and E. UAE, BUN and Scr of group B were significantly higher than that of control groups, while all the above indexes decreased obviously in group C, D and E compared with those in group B, and it seemed that it was positively correlated with the herb doze (p<0.05, p<0.05, p<0.01 respectively). But it was on the contrary for Ccr .3 The morphological examination: In group B, there were a slight mesangial cell proliferation and ECM deposition in mesangial regions. The glomerular basement membrane (GBM) was thickened and the picric acid-toluidine blue-positive material deposited in the mesangium. The area of glomerulus was increased. All the changes were attenuated in group C, D and E, and the changes were outpocketing in group E.4 The protein expression of TGF-β1 and Col-IV in renal cortex: The ratio of positive staining areas of TGF-β1 to renal tubules and matrix in group B (21.24±2.45) was significantly lower than that of group A (5.85±0.65)(P<0.01). It was lessened in group C (13.90±1.71), D (9.72±2.02) and E (7.82±2.09)(all P<0.01), so that it seemed it was positively correlated with the herb doze (p<0.05, p<0.05, p<0.01 respectively). In group B, the protein expression of Col-Ⅳ(13.02±1.74) was much higher than that of the control group (6.27±0.72)(p< 0.01). The IOD of Col-Ⅳin group C (10.24±1.32), D (9.02±1.20) and E (7.92±1.18) were much lower than that in group B(all p<0.01).The Col-Ⅳof group E was lowest than that of group C and D (p<0.01). Correlation analysis showed that the expression of TGF-β1 had negative correlation with Col-Ⅳ(r=-0.509,p< 0.01).5 The bioactivity of antioxidase in renal cortex: the bioactivity of TSOD, CAT in group B(55.25±2.46u/mgpro, 26.31±2.09u/mgpro respectively) was significantly lower than that of group A (79.55±4.67u/mgpro, 47.95±3.05u/mgpro respectively) (P<0.01). Compared with group B, the bioactivity of TSOD, CAT in group C (60.34±2.02u/mgpro, 31.03±2.04u/mgpro respectively), group D (67.26±2.01u/mgpro, 33.47±1.09u/mgpro respectively) and group E (66.22±1.09u/mgpro, 37.25±1.34u/mgpro respectively) was significantly elevated (all p< 0.01). 6 The bioactivity of MDA in renal cortex: the bioactivity of MDA in group B (18.19±1.02nmol/mgpro) was significantly higher than that of group A (7.41±0.83nmol/mgpro). Compared with group B, the bioactivity of MDA in group C (9.38±0.46nmol/mgpro), group D (8.39±0.89nmol/mgpro), group E (8.00±0.69nmol/mgpro) significantly decreased(all p< 0.01).Conclusions1 The expression of Col-Ⅳand TGF-β1 in renal cortex increased significantly in diabetic rats than that in the control group. It implies that TGF-β1 and Col-Ⅳmight contribute to the occurrence and development of DN.2 The reactive oxygen substances (ROS) in renal cortex increased significantly in diabetic rats than that in the control group, while the bioactivity of antioxidase in renal cortex decreased greatly in diabetic rats than that in the control.It implied that the oxidative stress might have a close relationship with the occurrence and development of DN.3 Cordyceps Sinensis Cultivated by Artificial Fermentation could reduce the level of glucose and albuminuria, regulate the levels of serum lipids and the expression of TGF-β1, lessen the thickening of Col-Ⅳand better the activity of the antioxidase. Thus the Chinese herb above may prevent and treat DN through those pathway.4 Cordyceps Sinensis Cultivated by Artificial Fermentation may prevent and treat DN also by means of ameliorating metabolism, reinforcing immunity, et al. And there were some different ideas about the mechanism of Cordyceps Sinensis Cultivated by Artificial Fermentation that might take action in the pathogenesis of DN needs further study.5 Within the scope of a certain doze, Cordyceps Sinensis Cultivated by Artificial Fermentation may have the function of lowering the level of blood glucose in diabetic rats. But if the doze surpasses the seeming limit, the effects will not be so obvious. |
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