Study On The Role Of Angiopoietins In Progressive Glomerulosclerosis In Rats

Changes of Angiopoietin in Plasma Levels and Glomerular Expression Involved in Daunorubicin-Induced Nephrosis in RatsObjective To study the potential pathological role of endogenous Angiopoietins in daunorubicin-induced nephrosis in rats.Methods 70 male wistar rats were randomly allocated into control group (Control, n=30) and daunorubicin group (DRB, n=40). The rats in DRB group were injected into rat`s tail with DRB(15mg·kg-1). Since then, at week 1, 2, 4, 6, 8, and 12 respectively, 5 male wistar rats in each group were taken randomly for determining 24-h urinary protein quantitative measurement(24hUPQM), plasma TNF-α, Ang1, and Ang2. Sections of kidneys were examined by electric microscope, PAS staining, immunohistochemical staining and In situ hybridization histochemistry.Results There was no significant difference in plasma Ang1 concentration between the control group and the DRB group. Rats in the DRB group exhibited a trend toward increased plasma Ang2, TNF-αand 24hUPQM from week 2 to week 12. With progression of Glomerulosclerosis, in DRB group, Expression of Ang1mRNA and protein in glomeruli decreased, while expression of TNF-αprotein, Ang2 mRNA and protein,in glomeruli increased. Expression of Ang1mRNA and protein in glomeruli had a negative correlation with 24hUPQM. Plasma Ang2, expression of Ang2mRNA and protein in glomeruli had a positive correlation with 24hUPQM. Plasma TNF-αand expression of TNF-αin glomeruli had a positive correlation with expression of Ang2mRNA and protein in glomeruli. In glomeruli, expression of Ang1 protein had a negative correlation with expression of Ang2, expression of Ang2 protein had a negative correlation with Fn protein expression and mean area of extracellular matrix (MAECM).Conclusion In daunorubicin-induced nephrosis, expression of Ang1 downregulated and expression of Ang2 upregulated in glomeruli. An increase in plasma Ang2 and expression of Ang2 may mediate an increase in permeability of the glomerular filtration barrier. In adition, Ang2 may upregulate expression of components of extracellular matrix to contribute to glomerulosclerosis.Part Two The Role that Abnormal Expression of Angiopoietins in Glomeruli Following Podocyte Injury in Progressive Glomerulosclerosis in RatsⅠ. Alteration of Angiopoietins Expression after Glomerular Podocyte Injury Relative to Progressive GlomerulosclerosisObjective To study the potential pathological role of endogenous angiopoietins that expression altered in glomeruli during progressive glomerulosclerosis.Methods Daunorubicin (DRB)-induced podocyte injury in rat were used as animal model. 100 male wistar rats were randomly allocated into the following three experimental groups, sham operation group (Sham, n=30), uninephrectomy group (UPHT, n=30) and uninephrectomy of single kidney+daunorubicin group (DRB, n=40). The rats in DRB group were injected into rat`s tail with DRB(5mg·kg-1) on the seventh and the fourteenth day after one renal excision. At week 1, 2, 4, 6, and 8 respectively following finishing animal model, 6 male wistar rats in each group were taken randomly for determining 24-h urinary protein quantitative measurement(24hUPQM), serum urea nitrogen and creatinine, whose sections of kidneys were examined by electric microscope, PAS staining, immunohistochemical staining and In situ hybridization histochemistry for semiquantitative morphological evaluation.Results 24hUPQM, Bun and Scr in DRB group is more than that in Sham group and UPHT group at same time point, morphological evaluation shows that there was a trend towards an increase on level of GSI in DRB from week 2 to week 8. Electric microscope reveals that podocyte injury present in DRB group. Expression of Ang1mRNA and protein in glomeruli of DRB group decreased, while expression of Ang2 protein in glomeruli of DRB group increased. However, there were no pathological change in glumeruli of Sham group and UPHT group. In DRB group, expression of Ang1 mRNA had a negative correlation with expression of Ang2mRNA, expression of Ang1 protein had a positive correlation with expression of Ang1 mRNA, and had a negative correlation with 24hUPQM, Bun, Scr, GSI, expression of Ang2 protein and CoIV protein. Expression of Ang2 protein had a positive correlation with expression of Ang2 mRNA, and had a positive correlation with 24hUPQM, Bun, Scr, GSI, expression of CoIV protein.Conclusion Podocyte injury may lead to glomeruli abnormally express angiopoietins. Expression of Ang1 decreased, Ang2 got rid of inhibition from Ang1, which facilitate expression of Ang2, and modulate progressive glomerulosclerosis in the rats. Ⅱ. Abnormal Expression of Angiopoietins is Associated with Loss of Glomerular Capillary in Progressive Glomerulosclerosis following Podocyte InjuryObjective To study the relationship between the change of angiopoietins in glomeruli and loss of glomerular capillary in progressive glomerulosclerosis following podocyte injury.Methods 100 Male Wistar rats were randomly allocated into sham operation group (Sham, n=30), uninephrectomy group (UPHT, n=30) and uninephrectomy+daunorubicin group (DRB, n=40). The rats in DRB group were injected into rat`s tail with DRB (5mg·kg-1) on day 7 and day 14 after one renal excision. At week 1, 2, 4, 6 and 8 respectively following finishing animal model, 6 rats in each group were taken randomly for determining creatinine cleareace (Ccr), sections of kidneys were examined by PAS staining, immunohistochemical staining and In situ hybridization histochemistry for semiquantitative morphological evaluation. Glomerular cell apoptosis was examined by terminal-deoxynucleotidyl transferase mediated nick end labeling (TUNEL) and transmission electric microscope (TEM).Results In DRB groups, there was a trend towards a decrease on level of Ccr, and glomerular sclerotic index(GSI) showed gradual elevation. TEM showed there was a apoptotic endothelial cell in glomeruli. At the same time, glomerular apotopsis index(GAI) became more and more rise with a decline in glomerular capillary density(GCD)and Ccr. The magnitude of quantitative analysis in morphology showed that there was a trend towards a decrease on expression of Ang1mRNA and protein, but expression of Ang2mRNA and protein as well as expression of Fas, FasL and Caspase-3 protein tended to an increase. Moreover, expression of Ang1 mRNA and protein had a negative correlation with GAI, GSI, and expression of Fas, FasL and Caspase-3 protein, and a positive correlation with GCD and Ccr. Expression of Ang2mRNA and protein had a positive correlation with GAI, GSI, expression of Fas, FasL and Caspase-3 protein, and had a negative correlation with GCD, Ccr.Conclusion Podocyte injury may lead to an alteration in expression of Ang1 and Ang2 within glomeruli, which correlated with upregulation in expression of Fas, FasL and Caspase-3 protein, and facilitated the apoptosis of endothelial cell in glomeruli to result in the glomerular capillary loss relative to progressive glomerulosclerosis.