The Correlation Research On Of MRNA Expression Of BK_Ca Channel In Renal Artery And Renal Lesion In T2DM Nephropathy Rats

Objective:To reveal the correlation research between expression change of BK_Ca channel in renal artery and renal lesion in T2DM nephropathy rats in the different course.Methods:1.The establishment of T2DM nephropathy models:60 SD rats adaptively raised for a week,were randomly divided into the control group 10 and the model group 40respectively.To remove left kidney in model group,the control group were on the left renal resection control.Model group were feed into high-fat diet and sugar?Main ingredients for 10%lard,2.5%cholesterol,20%sugar,1%cholic acid salt,66.5%normal feed?;the control group only were given ordinary feed.At 4 weeks,rats in model group were injected with 25mg·kg^-1 of streptozotocin through intraperitoneal injection;The control group only were injected with the volume of citric acid buffer.Three days later,tail blood measuring Fasting blood glucose was 7.0 or higher mmol·L^-1screening in 40rats to continue feeding.Modeling time is a total of eight weeks?only half of the 40?and 12 weeks?only half of the 40?.The day before in 8 and 12 weeks to kill the rat,collecting 24 h urine to measure trace urinary protein.At 8 and12 weeks,fistly measuring Fasting blood glucose,then measuring the rat body weight,then anesthesia to kill the rat,abdominal aortic to measure blood serum creatinine and blood urea nitrogen,getting renal artery,collecting right kidney to measure the weight.Modeling time was a total of 8 and 12 weeks.2.Identification of T2DM nephropathy model:?1?Fasting blood glucose was 7.0or higher mmol·L^-1;2h postprandial blood glucose was 11.0 or higher mmol·L^-1and insulin sensitivity was reduced;?2?had early renal hypertrophy of diabetic nephropathy;?3?had microalbuminuria,to be model.3.Detection of renal biochemical indicator:after anesthesia,open the abdominal cavity,collected blood from abdominal aorta and obtained serum by the centrifugal in rats;used automatic biochemical analyzer to test blood urea nitrogen,serum creatinine and urinary protein.4.HE staining:after anesthesia and abdominal aortic blood collected in rats at 8 and 12 weeks,taked some kidney tissues,fixed in 10%neutral formalin.conventional paraffin embedding,sectioning,dewaxing,HE staining,transparent,processing and to observe the kidney organization structure change by light microscopy.5.Electron microscope:the other part of the renal tissue fixed in 2.5%glutaraldehyde,conventional dehydration,osmosis,embedding,sectioning,to observe kidney tissue ultrastructure changes by electron microscope.6.Real-time fluorescent quantitative PCR:after anesthesia and abdominal aortic blood collected,determined?,?₁ subunits mRNA expression level of BK_Ca channel.To extract total RNA in groups,to reverse transcription into cDNA,to use the primers and SYBRgreen dyes and amplification.Results:?1?the T2DM nephropathy rat model:compared with the control group,?1?Fasting blood glucose was 7.0 or higher mmol·L^-1;2h postprandial blood glucose was 11.0 or higher mmol·L^-1and accompanied by reduced insulin sensitivity,by insulin sensitive index?P<0.05?in the model group;?2?delearly renal hypertrophy of diabetic nephropathy by the kidney hypertrophy index?P<0.05?;?3?had microalbuminuria by 24 h urine protein?P<0.05?.?2?the kidney changes and biochemical indicators:erum creatinine and blood urea nitrogen respectively were 44.00±4.07,49.73±2.98 and 6.28±1.40,9.90±1.55,compared with the control groups,concentration increased significantly?P<0.05?.Concentration increasing higher than 8 weeks.?3?kidney disease morphological indicators:?1?HE staining:compared with the control group,glomerulus and renal tubules had pathological change and had glomerular mesangial cells proliferation in the model group.?2?the electron microscope results:compared with the control group,basement membrane heterogeneity thickening;part foot process mixed together,fell off;matrix increased in mesangial area.?4?Compared with the control group,?subunit expression of BK_Caa in the model group at 8 weeks and 12 weeks were obviously unchanged.?2??₁ subunit expression of BK_Caa in the model group at 8 weeks and 12 weeks were reduced.Conclusion:?1?T2DM nephropathy rats model was succeed.?2?kidney disease of the T2DM nephropathy rats,in 8 weeks and 12 weeks,had obvious changes than the control group.And 12 weeks,kidney changes more obviously than8weeks.?3??₁ subunit expression of BK_Ca channel were lowered in the T2DM nephropathy rats in the model group than in the control group at 8 weeks and12 weeks.And 12 weeks to reduce even more significant than 8weeks.BK_Caa channel may be associated with the progress of renal lesion.