The Distribution And Expression Of Protein Kinase C-α, βⅠ, βⅡ And Its Potential Role In Diabetic Nephropathy

Part I Protein kinase C-α,βI,βII in mouse diabetic nephropathy kidney and its relation to the MicardisThesis 1: Protein kinase C-α in mouse diabetic nephropathy kidney and itsrelation to the MicardisObjective: To investigate the localization and expression of protein kinase C-α (PKC-α) in diabetic nephropathy (DN) mouse kidney and its relation to micardis.Methods: DN models were induced by i.p. injected with streptozotocin. Normal mice were provided as controls (NG). The DN mice in were divided into two group: DN group and T group which were treated by micardis. Performed metabolic cage experiments on these mice at the end of 6th week,determined the localization of PKC-α by confocal immunofluorescence laser scanning, the expression of PKC-α by semi-quantitative western blotting as well as the glomerular expression of PKC-α, transforming growth factor- beta 1 (TGF- beta 1) and vascular endothelial growth factor( VEGF) by semi-quantitative immunofluorescence. Results: (1) Blood glucose, 24-hour urine protein amount, total and mononuclear cell number of glomerular, right kidney weight, body weight, right kidney weight index, glomerular areaand perimeter of DN group mice were obviously higher than that of NG group mice and these values were attenuated in T group as compared with DN group. (2) In DN and normal mice, PKC-α localizated in glomular mesangial area, podocytes, proximal tubules, intercalated cells of cortical collecting duct and principal cells of medullary collecting duct. But PKC-α transferred to brush border as compared with NG group and the expression of PKC-α in T group was lower than DN group. (3) The expression of PKC-α in the kidney cortex and the outer medulla of DN group mice were significantly higher than that in NG and T group. The expression of PKC-α in inner medulla presented no significant changes in all groups. (4) The expression of glomcrular PKC-α, TGF-β1 and VEGF in DN group were higher than NG group, but decreased in T group. The expression of PKC-α exhibited a positive correlation to VEGF(r=0.742, P=0.024) and no correlation to TGF-β1(r=0.287, P =0.085).Conclusions: (1) The localization and expression of PKC-α are changed in DN mice, PKC-α may take part in function change of promixal tubule in DN mice and it may also contribute to DN pathogenesis by mediating the expression of glomerular VEGF and the development of proteinuria. (2) In DN mice, treatment with Micardis can partly correct the abnormal expression of PKC-α which suggest that RAS is implicated in the pathogenesis of DN by regulating the expression and activation of PKC-α isoform.Thesis 2: Protein kinase C-βI,βII in mouse diabetic nephropathy kidney and its the MicardisObjective: To investigate the localization and expression of protein kinase C-βI,βII (PKC-βI,βII) in diabetic nephropathy (DN) mouse kidney and its relation to micardis.Methods: DN models were induced by i.p. injected with streptozotocin. Normal mice were provided as controls (NG). The DN mice in were divided into two group: DN group and T group which were treated by micardis. At the end of 6th week, determined the localization of PKC-βI,βII by confocal immunofluorescence laser scanning, the expression of PKC-βI,βII by semi-quantitative western blotting as well as the glomerular expression of PKC-βI,βII, transforming growth factor-betal(TGF-betal) and vascular endothelial growth factor(VEGF) by semi-quantitative immunofluorescence.Results: (1)In DN and normal mice, PKC-βi localizated in glomular podocytes, proximal tubules, cortical and medullary thick ascending limbs,intercalated cells of cortical collecting duct and principal cells of medullary collecting duct. But the expression of PKC-βI on the proximal tubules were obviously as compared with NG group and the expression of PKC-βI in T group was lower than DN group. (2) In DN and normal mice, PKC-βII localizated in glomular podocytes, interstitial cells of cortex and medulla. PKC-βII also expressed on the proximal tubules, while no expression in cortex and inner medullary collecting duct,as compared with NG group. No expression of PKC-βII on proximal tubules and having expression on inner medullary collecting duct in T group, as compared with DN group. (3) The expression of glomcrular PKC-βI, TGF-βI and VEGF in DN group were higher than NG group, but decreased in T group. The expression of glomcrular PKC-βII in DN group were lower than NG group, and have no difference between the NG group and T group. The expression of PKC-βI exhibited a positive correlation to TGF-β1(r=0.629, P=0.034) and no correlation to VEGF(r=0.360, P=0.079).The expression of PKC-βII exhibited no correlationto TGF-β1(r=0.287, P=0.085) and VEGF(r=0.341, P=0.082) . (4) The expression of PKC-βI in the kidney cortex and the outer medulla of DN group mice were significantly higher than that in NG and T group. The expression of PKC-βI in inner medulla presented no significant changes in all groups. (5)The expression of PKC-βII in the kidney cortex of DN group mice were significantly lower than that in NG. The medulla presented no significant changes in all groups.Conclusions: (1) The localization and expression of PKC-βI,βII are changed in DN mice.(2)PKC-βI,βII may take part in function change of promixal tubule in DN mice and PKC-βI may also contribute to DN pathogenesis by mediating the expression of glomerular TGF-βI and the development of glomerular hypertrophy. PKCβII may also take part in function change of collecting duct .(3) In DN mice, treatment with Micardis can partly correct the abnormal expression of PKC-βI and PKC-βII which suggest that renin-angiotensin system(RAS) is implicated in the pathogenesis of DN by regulating the expression and activation of PKC-βI isoform.Part II The expression of protein kinase C-α,βI,βII and its potential in diabetic nephropathy patientsObjective To investigate the expression of protein kinase C-α, βI,βII in kidney of health adult and diabetic nephropathy (DN) patients as well as its relation to glomerular transforming growth factor-β1(TGF-β1) and vascular endothelial growth factor( VEGF). Methods Fetch normal and DN patient biopsy kidney tissue and judge the expanding degree of glomerular extracellular matrix (ECM ) with PAS staining. The expression of glomerular PKC-α,βI,βII were detected by semi-quantitative immunohistochemical method. Results (1) PKC-α,βI,βII expressed in glomerulus of normal persons; (2) In glomerulus of DN patients, the expression of PKC-α,βI, VEGF and TGF-βI as well as the accumulation of ECM increased significantly, but the expression of PKC-βII decreased significantly. Meanwhile, PKC-α , βI presented a positive relationship to VEGF and TGF-βI separately (r=0.600, P=0.039; r=0.521, P=0.043).Conclusions: The expression of PKC-α, βI, βII in glomerulus of normal person and diabetic nephropathy (DN) patient are different; PKC-α may take part in DN pathogenesis by mediating the expression of glomerular VEGF and development of proteinuria, PKC-βI may contribute to the genesis of DN glomerular hypertrophy by mediating the expression of glomerular TGF- β1 and accumulation of ECM, but human glomerular PKC-βII may be a kind of factor providing protective response to high blood glucose.