| Objective: With the progress of medical diagnosis,more and more patients with diabetic kidney disease(DKD)have been found.But the patient's condition generally develops to the middle and late stage,while people is diagnosed with DKD.They will find it difficult to cure.Most of patients could not recover completely under the current clinical treatment.So there is an urgent need to find new drugs to treat diabetes.And the purpose of this article is to explore the effect of Cyslosorus acuminatus flavonone glycoside(CAF)on diabetic kidney disease(DKD).Methods:1.Cell experiment in vitro1.1 The method of first cell experiment,which study on the effect of C AF in the proliferation of HK-2 stimulated by high glucose,is that HK-2is randomly divided into normal group,isosmotic control group,high sugar model group and experiment group.The experiment group is also divi ded into the positive control group and six concentrations of medication administration groups which concentrations are 2.5 ?g·m L-1,5 ?g·m L-1,10?g·m L-1,25 ?g·m L-1,50 ?g·m L-1and 100 ?g·m L-1.The survival rate of the cells in each group was determined by CCK8 after 24 hours,48 hours and 72 hours of cell culture.1.2 The method of second cell experiment,which study on effect of CAF on the oxidative damage of HK-2 with high sugar stimulation,is that HK-2 is randomly divided into normal group,isosmotic control group,high sugar model group and experiment group.The experiment group is also divided into two concentrations of medication administration groups which concentrations are25 ?g/ml,50 ?g/ml.ROS in cell is detected by flow cytometry after 48 hours and 72 hours of cell culture.1.3 The method of third cell experiment,,which study on effect of CAF in EMT of HK-2 stimulated by high glucose,is that HK-2 is randomly divided into normal group,high sugar model group and medication adm-inistration group.The concentrations of medication administration group is 50?g·m L-1.The renal EMT was evaluated by immunofluorescence technique for the makers of vimentin and fibronectin.2.Experiment in rats with diabetic kidney disease The experimental rat model of DKD was induced by injected with 60 mg/kg streptozocin and given 16 weeks high-fat diet.CAF(12.5 or 25 mg/(kg·d))was orally treated for 4 weeks.After the experimental period,blood samples were collected,and renal tissue was fixed and the samples were submitted for examination.2.1 Biochemical detection:the levels of fasting glucose,serum creatinine(Scr)and bloodurea nitrogen(BUN)were measured.2.2 Histological analysis:the collagen deposition was observed by masson and sirius red staining,as well as the basement membrane enlargement was observed by PAS staining.2.3 Molecular biological detection:The renal EMT was evaluated by immunohistochemistry analysis for the makers of?-SMA,fibronectin and E-cadherin.And the renal expression of?-catenin and the phosphorylation level of GSK-3?were measured by western blot.Results:1.Results of cell experiment in vitro 1.1 The CAF groups are compared with the high glucose model group after 48 hours of cell culture,which the CAF groups concentration of 2.5 ?g·m L-1 and5 ?g·m L-1 could inhibit the proliferation of cells induced b-y high glucose(P<0.05)and the CAF groups concentration of 10 ?g·m L-1,25 ?g·m L-1,50 ?g·m L-1and 100 ?g·m L-1 could obviously inhibit the prolif-eration of cells induced by high glucose(P< 0.01).The CAF groups are c-ompared with the high glucose model group after 72 hours of cell culture,which the all CAF groups could obviously inhibit the proliferation of cells induced by high glucose(P< 0.01).1.2 The CAF groups are compared with the high glucose model group after 48 hours or 72 hours of cell culture,which the CAF groups concentration of 25?g·m L-1 and 50 ?g·m L-1 could apparently inhibit the aggregation of ROS(P<0.01).1.3 50 ?g·m L-1 CAF inhibited the expression of vimentin and fibronectin in renal by immunofluorescence technique,compared with the high glucose model group.2.Results of experiment in rats with diabetic kidney disease2.1 The levels of fasting blood glucose,serum creatinine and urinary nitrogen were obviously increased in DKD model group,when compared to the vehicle control(P<0.01).When compared to DKD model,CAF significantly attenuated the levels of fasting glucose,Scr and BUN(P<0.05).2.2 When compared to the vehicle control,the renal deposition of collagen was evident.When compared to DKD model,CAF significantly improved renal deposition of collagen.2.3 When compared to the vehicle control,the EMT was activated and the expression of ?-catenin and the phosphorylation of GSK-3?were also significantly enhanced(P<0.01).When compared to DKD model,CAF significantly inhibited the expression of ?-catenin and the phosphorylation of GSK-3?(P<0.05).Conclusion:The whole results indicated that CAF could obviously inhibit the proliferation of cells induced by high glucose and inhibit the aggregation of ROS,as well as inhibit renal EMT in DKD rats. |
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