| Objective: In recent years the incidence of diabetic nephropathy(DN) isgoing up. DN is one main cause of end-stage renal disease, and has been paidmore and more attention in its prevention and treatment. In the three types ofthe glomerular’s inherent cells, the podocyte is an important composition offiltration barrier, with characteristic biological feature and physiologicalfunction. Podocyte injury contributes to proteinuria, which is the main clinicalfeature of DN, that is to say podocyte injury drives the development ofdiabetic nephropathy partly.Podocalyxin (PCX) is a sialomucin most closely related to CD34andendoglycan, and is expressed in kidney podocytes, hematopoietic progenitors,vascular endothelia, and a subset of neurons.PCX is always found along theapical surface of podocyte and foot processes.Its negative charge makesadjacent podocytes separate, which formats slit diaphragms (SD), and itsfeature of decreasing cell adhesion maintains filtration barrier opening up. Indiabetic nephropathy, podocalyxin expression is suppressed directly, andalbumin carrying negative charge can filtrate more easily. On the other hand,the reduced expression of PCX increases the intercellular adhesion, andaccelerates the occurrence rate of proteinuria.WT1is a zinc-finger transcription factor, and regulates the expression ofa number of genes. Recent studies demonstrate that podocalyxin is a directdownstream target of WT1, and WT1can combinate the conserved sequenceof PCX gene’s promoter, which makes WT1get strong transcriptionalactivation force. In addition, WT1, being one specific marker of the podocyte,itself also plays an important role in maintaining podocyte’s normal function.Desmin is found in mesangial cells and vascular smooth muscle cells,rarely in podocytes normally. Recent studies suggest that podocyte injury makes desmin upregulation, therefore desmin is an early marker of podocyteinjury in chronic glomerulonephropathy.In this study, we build diabetic Sprague Dawley rat model induced bystreptozotocin, administrate sulodexide and benazepril for three months, detectthe expression of podocalyxin, WT1and desmin in diabetic rats, observe theexpression level of podocalyxin, WT1and desmin, and explore the effectionof sulodexide, comparing with benazepril, in proteinuria and the developmentof renal failure in diabetic rats kidney disease.Method: The clean health male SD rats, altogether60,weight about180～200g,were randomly divided into normal control group (N group,n=12),diabetes model group (D group, n=12),diabetes+benazepril group (Bgroup, n=12),diabetes+sulodexide group(S group, n=12) and diabetes+benazepril+sulodexide group(B+S group, n=12).The rats in the last fourgroups were given STZ by abdominal cavity injection to build diabetic ratsmodel, at a dose of65mg/kg.The diabetic model was considered to besuccessful when the blood glucose achieved16.7mmol/L and urine glucosemaintained positive after48hours of STZ injection; rats who did not meet theabove criteria excluded the experiment. At the end of6week and12week, wecollected the blood,24-hour urine and renal tissue samples, and measured24hurinary protein excretion, serum creatinine and urea. We applied ofimmunochemistry and RT-PCR on kidney tissues, and observed the form ofpodocytes in diabetic rats by transmission electron microscope. All the dataswere expressed as mean value±SD. Analysis were carried out usingSPSS13.0. One-way ANOVA was used to compare mean values among groups.A two-tailed P-value of＜0.05was considered to be statistically significant.P-value of＜0.01was considered to be strongly statistically significant.Results:1. General situation of diabetic rats The diabetic rats appearedpolydipsia, polyphagia, polyuria and gradually emaciation, and thesesymptoms were the most significant in D group.2.24h urinary protein excretion, serum creatinine and urea comparisonAt12week, in D group, B group, S group and B+S group,24-hour urinary protein、serum creatinine and urea were significantly higher than in N group(P<0.05), the highest in B+S group(P<0.01).There was no significantdifference between B group and S group (P>0.05).3. Immunohistochemistry and RT-PCR expression of podocalyxin,WT1and desmin At the end of6and12week, the number of glomerular inherentcells in group B and S increased more significantly than in group B+S(P<0.05), but lower than in group D (P<0.05). At the end of6and12week inthe rats of group D, B, S and B+S, the expression of podocalyxin reducedshowing granular brown coloring. Both qualitative and quantitative expressionis stronger than in group N (P<0.05), the strongest in B+S group (P<0.01), andthere was no significant difference between B group and S group (P>0.05).But the expression of desmin increased, and its development trend wascontrary with other factors. In RT-Perth expression of podocalyxin and WT1reduced in the same period of group D, B, S and B+S, and the expression ofdesmin upregulated. There was a significant difference (P <0.05), the mostsignificant in B+S group (P<0.01), and there was no significant differencebetween B group and S group (P>0.05).4. Transmission electron microscope Compared with rats in the sameperiod, in Group D, B, S and B+S, glomerular basement membrane (GBM)performs uneven thickness, endothelial cells are swelling, and foot processesare fused. In the late three groups, the phenomenon was more relievedobviously than in Group D.Conclusion:1. In diabetic nephropathy, the expression of podocalyxinand WT1decrease, and the expression of desmin increase. So it suggests thatthese factors play a role in the development of diabetic nephropathy.2. Sulodexide can upregulate expression of podocalyxin and WT1,downregulate expression of desmin, therefore it can improve the electrostaticbarrier of podocytes in diabetic rats, and prevent the progress of diabeticnephropathy.3. Combined application of Sulodexide and benazepril has additive effectin improving proteinuria and delaying renal failure in diabetic rats, so it provides an effective mean of prevention and treatment in early diabeticnephropathy, and provides a scientific basis in screening of a new target fordiabetic nephropathy. |
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