| OBJECTIVE To investigated the renoprotective roles of BBR in different stages of DN,in order to clear and definite BBR at different stages of DN on renal function and thetissue of rats with lesion characteristics. Through detecting the expression of PGE2, EP4,Gαs, cAMP and regulation of BBR on them, so as to reveal the mechanism of BBRexerts a protective effect on renal function of DN rats.METHODS Diabetes was induced in mice by intraperitoneal injection of STZ, after72h in the determination of fasting blood glucose, the blood glucose value≥11.1mol/l as a successful model in experiment. Then the mice were then randomlydivided into groups: normal, diabetes, high sugar and high fat and berberine (high,median and low dose) groups, enalapril group, metformin group and simvastatin group.Daily intragastric administration (intragastric administration, ig) once and continuedhigh sugar and high fat diet, respectively at2,4,6,8weeks to observe the generalcondition of rats, weighing body weight once a week, every two weeks measurement ofblood glucose. In different periods, respectively, rats were placed in metabolic cages,collecting24h urine; collecting blood samples and get the separation of serum. UTP,UCr, BUN, SCr, TG, TC, HDL-c, LDL-c, VLDL-c were measured on different weeksthroughout the study. The rat kidney was removed and weighed, calculate the KW/BW;HE staining of renal pathological changes; the level of TGF-β1in the kidney weredetected by immunohistochemistry, the protein levels of EP4, Gαs in the kidney weredetected by western blot analysis; the level of PGE2and IL-18was detected by ELISA;the level of cAMP and IGF-1was detected by radioimmunoassay. RESULTS1Effects of BBR on FBG and BW of different stages of DN ratsCompared with the normal group, FBG of different stages of DN model group rats wassignificantly increased. At the4th week, the FBG of BBR (100mg/kg,200mg/kg) groupwas significantly decreased compared with the model group, the difference wasstatistically significant (P<0.05); BBR (50mg/kg) administration had no significanteffects on FBG in DN rats, from the2nd week, metformin treatment group can alsosignificantly reduce the FBG of model group.2Effects of BBR on biochemical and renal functional parameters in DN ratsThe results showed that the KW/BW ratio, BUN, SCr, UTP/C, TC, TG, LDL-c andVLDL-c levels of model group were significantly increased, while HDL-c wassignificantly decreased compared with those of the NC rats. From the4th week, BBR(200mg/kg) can improve the abnormal renal function and biochemical indexes of DNrats, while BBR (100mg/kg) works at the6th week. BBR (50mg/kg) have no significanteffect.3Effect of BBR on renal histopathology of DN ratsCompared with the normal group, model group rat glomerular volume was increased,basement membrane thickening, some tubular epithelial cells with a large number ofinflammatory cell infiltration. At the2nd week, BBR had no significant effect; BBR(200mg/kg) can significantly improve the pathological changes of rats from the4thweek, while BBR (100mg/kg) works at the6th week; positive drug treatment groupshad similar improvement effect on kidney of rats in the model group.4Effects of BBR on the expression of TGF-β1in kidney tissueImmunohistochemical analysis of kidney tissue sections showed that the level ofTGF-β1, as significantly increased in model group compared with normal group,suggesting positive results. BBR (100,200mg/kg) treatment can prevented theenhanced expression level of TGF-β1,while BBR (50mg/kg) have no significant effect.5Effect of BBR on the expression of IL-18in renal tissue in DN rats ELISA showed that the levels of IL-18of model group were significantly higher thanthose of the normal group. BBR(200mg/kg) could obviously reduce the IL-18levelfrom the4th week, while BBR (100mg/kg) works at the6th week. BBR (50mg/kg) haveno significant effect.6Effect of BBR on the levels of IGF-1in renal cortex of DN ratsRadioimmunoassay showed that the level of IGF-1of model group was significantlyincreased compared to the normal group. BBR (200mg/kg) can significantly down-regulate the level of IGF-1from the4th week, while BBR (100mg/kg) works at the6thweeks, and BBR (50mg/kg) have no significant effect.7Effect of BBR on the expression of PGE2in renal tissue in DN ratsELISA showed that the levels of PGE2of model group were significantly higher thanthose of the normal group. BBR (200mg/kg) could obviously reduce the PGE2levelfrom the4th week, while BBR (100mg/kg) works at the6th week. BBR (50mg/kg) haveno significant effect.8Effects of BBR on the expression of EP4, Gαs in renal cortexWestern blotting showed that, compared with normal group, the expression of EP4, Gαsin renal cortex of DN rats was significantly decreased. BBR group in different periodshave different effect on the level of EP4, Gαs: from the beginning of the4week, BBR(200mg/kg) can upregulate the expression of EP4, Gαs in DN rats, while BBR(100mg/kg) works at the6th week, and BBR (50mg/kg) have no significant effect.9Effect of BBR on the levels of cAMP in renal cortex of DN ratsRadioimmunoassay showed that the level of cAMP of model group was significantlydecreased compared to the normal group. BBR (200mg/kg) can significantlyup-regulate the level of cAMP from the4th week, while BBR (100mg/kg) works at the6th week, and BBR (50mg/kg) have no significant effect.CONCLUSIONS1. BBR (100,200mg/kg) can make the FBG of model group was significantly decreased during different stages of DN; BBR (100,200mg/kg) have differenteffect in improvement of blood biochemical indexes and renal function indexes inthe model group, suggesting that BBR should be administered for a long period oftime (at least four weeks) in order to improve the state of disease in DN rats; BBR(100,200mg/kg) can up-regulate the expression of TGF-β1, IL-18, IGF-1and PGE2in DN rats, suggesting that BBR has protective effects on kidney of DN rats may berelated to the regulation of TGF-β1, IL-18, IGF-1and PGE2.2. BBR (100,200mg/kg) can up-regulate the expression of EP4, Gαs, cAMP in renalcortex of DN rats, suggesting that BBR may play its renoprotective role in DN ratsthrough the EP4-Gαs-cAMP signaling pathways. |
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