Colchicine Ameliorates High Glucose-Induced ICAM-1 and Fibronectin Expression in Renal Cells via Inhibiting Locally-Produced Angiotensin II / 대한신장학회지

PURPOSE: A previous study has demonstrated that colchicine abrogated intercellular adhesion molecule (ICAM)-1 and fibronectin expression in renal cells exposed to high glucose media, but the underlying mechanism was not clarified. This study was undertaken to elucidate whether it was attributed to the inhibitory effect of colchicine on locally-produced angiotensin II (AII) under diabetic conditions. METHODS: Rat mesangial cells and NRK-52E cells were cultured in media containing 5.6 mM glucose (NG), NG+10(-7) M AII (NG+AII), or 30 mM glucose (HG) with or without 10(-8) M colchicine (Col) and/or 10(-6) M L-158,809, an AII type 1 receptor blocker (ARB). ICAM-1 and fibronectin mRNA and protein expressions were determined by real-time PCR (RT-PCR) and Western blot, respectively. AII levels in conditioned media were determined by ELISA. RESULTS: AII levels in conditioned media were significantly higher in HG-stimulated mesangial cells and NRK-52E cells compared to NG cells (p<0.05). ICAM-1 and fibronectin mRNA and protein expression were significantly increased in renal cells exposed to HG media (p<0.05 or p<0.01), and these increases were significantly ameliorated by colchicine or ARB treatment (p<0.05). Colchicine and ARB also significantly attenuated AII-induced ICAM-1 and fibronectin expression (p<0.05). However, there was no additive inhibitory effect of colchicine and ARB on the increases in ICAM-1 and fibronectin expression. CONCLUSION: Colchicine abrogated increased ICAM-1 and fibronectin expression in renal cells under diabetic conditions, which is partly mediated by inhibiting HG-induced locally-produced AII. These findings provide a new renoprotective mechanism of colchicine in diabetic nephropathy in addition to its impact on leukocyte functions.

The Effect of Monocyte Chemoattractant Protein-1 (MCP-1) on Epithelial-Mesenchymal Transition in Human Peritoneal Mesothelial Cells / 대한신장학회지

PURPOSE: This study was undertaken to elucidate whether CC chemokine receptor 2 (CCR2) exists in human peritoneal mesothelial cells (HPMCs) and whether monocyte chemoattractant protein-1 (MCP-1) has direct effects on epithelial to mesenchymal transition (EMT) and fibronectin expression in HPMCs. METHODS: HPMCs were isolated from a piece of human omentum and were incubated with M199 media containing 5.6 mM glucose (LG), 5.6 mM glucose+94.4 mM mannitol (LG+M), LG+10 ng/mL recombinant human MCP-1 (LG+MCP-1), or 100 mM glucose (HG) with or without a specific inhibitor of CCR2, 1.0 micrometer RS102895, for 4 days. Levels of secreted MCP-1 in culture media were determined by ELISA. Western blot was performed to determine fibronectin, E-cadherin, alpha-smooth muscle actin (alpha-SMA) and CCR2 protein expression. RESULTS: MCP-1 protein levels were significantly increased in HG-conditioned media compared to LG media (p<0.05). CCR2 protein was expressed in HPMCs, but there was no difference between LGand HG-stimulated cells. alpha-SMA protein expressions in HG and LG+MCP-1 groups were significantly higher relative to LG cells, while E-cadherin protein expressions were decreased in HG and LG+ MCP-1 groups compared to LG cells (p<0.05). In addition, there were significant increases in fibronectin mRNA and protein expression in HG and LG+MCP-1 groups (p<0.05). These HG-induced changes were significantly abrogated upon pre-treatment with RS102895. CONCLUSION: HG and MCP-1 directly induce EMT and enhance fibronectin expression in HPMCs, and these HG-induced changes were attenuated by the inhibition of MCP-1/CCR2 system, suggesting that increased MCP-1 levels by HG may contribute to the development of peritoneal fibrosis.

The Uses of Urinary betaig-h3 in Differential Diagnosis of Isolated Microscopic Hematuria / 대한신장학회지

PURPOSE: This study was undertaken to elucidate the usefulness of urinary betaig-h3 concentrations in differential diagnosis of isolated microscopic hematuria patients. METHODS: Seventy-seven patients, in whom renal biopsy was performed due to microscopic hematuria without proteinuria, were enrolled. The patients were divided into two groups, IgAN group (patients with IgA nephropathy, N=37) and NM group (patients with normal or minor change on renal biopsy, N=40), and the clinical characteristics and laboratory findings were compared between the two groups. TGF-beta and betaig-h3 concentrations in urine were determined by ELISA and were compared between the two groups. To establish the optimal cut-off value of betaig-h3/creatinine (Cr) ratio for the diagnosis of IgA nephropathy, a receiver operating characteristic curve was constructed and the sensitivity and specificity were calculated. RESULTS: A comparative analysis revealed no significant differences in age and sex ratio between the two groups. There were no differences in serum IgG, IgA, IgM, C3, and C4 levels between the two groups. The urinary betaig-h3/Cr ratio was significantly higher in the IgAN group compared to the NM group (6.632.6 vs. 4.462.6 ng/mg, p0.05). A cut-off betaig-h3/Cr ratio 4.5 has a sensitivity of 85.0% and a specificity of 77.8%. CONCLUSION: The urinary betaig-h3/Cr ratio was a good predictor for the diagnosis of IgA nephropathy. Therefore, renal biopsy should be considered in isolated microscopic hematuria patients with high urinary betaig-h3/Cr ratio.

Differential Gene Expression According to the Size of Glomeruli in Experimental Diabetic Nephropathy / 대한신장학회지

PURPOSE: Although a few gene-profiling studies with whole renal tissue have been described in experimental diabetic nephropathy, there is only one microarray study using diabetic glomeruli. Furthermore, hypertrophic glomeruli have not been explored. The purpose of this study is to elucidate gene expression profiles of hypertrophic glomeruli in early diabetic nephropathy. METHODS: Forty-male Sprague-Dawley rats were injected with diluent (N=20) or streptozotocin intraperitoneally (DM, N=20) and were sacrificed at 6- and 12-week. Glomeruli were isolated by sieving technique. Glomeruli from 125 and 75 m sieves were classified into large (hypertrophic, DM-LG) and small glomeruli (DM-SG), respectively. After RNA extraction, hybridization was performed on the Rat cDNA 5K chip in triplicate, and slides were analyzed. The significant genes were selected using significant analysis of microarray. RESULTS: At 6-week, hierarchical clustering revealed that gene expression profiles of DM-LG were different from those of DM-SG, whereas DM-SG and C glomeruli showed similar gene expression pattern. In contrast, gene expression profiles at 12-week were similar between DM-LG and DM-SG, whereas C glomeruli showed different gene expression pattern from DM glomeruli. At 6-week, a total of 207 genes showed greater than 1.5-fold differential expression. 149 genes were upregulated, whereas 58 were downregulated in DM-LG. On the other hand, differential gene expression greater than 1.4-fold was observed in 37 genes at 12-week, upregulated in 26 and downregulated in 11. CONCLUSION: These results suggest that the gene expression profiles of DM-LG are different from DM-SG, and the gene expression patterns change with the progression of diabetic nephropathy.

Angiotensin II Receptor Blocker Induced Inhibition of Cellular Hypertrophy and Differential Expression of Cyclin-dependent Kinase Inhibitors in Cultured Podocytes Stimulated by Long-term High Glucose / 대한신장학회잡지

BACKGROUND: Hypertrophy of podocytes is observed in type 2 diabetic patients. Cellular hypertrophy requires combined effects of various mitogen- induced entry into the cell cycle and subsequent cell cycle arrest at the G1/S interphase. This cell cycle arrest is mediated by various cyclin-dependent kinase inhibitors (CKIs). We investigated the effect of angiotensin II receptor blocker (ARB) treatment on podocyte hypertrophy and CKIs expression in cultured podocytes stimulated by long-term high glucose. METHODS: Immortalized mouse podocytes were cultured in media containing 5.6 mM normal glucose (NG), 30 mM high glucose (HG), or NG+angiotensin II (AII, 10(-7)M) for 7 days with or without ARB (L-158,809, 10(-6)M). Cellular hypertrophy was assessed by measurement of cellular protein/cell counts, and CKIs mRNA and protein expression were assessed by reverse-transcription polymerase chain reaction (RT-PCR) and Western blot, respectively. RESULTS: Cellular hypertrophy was induced in podocytes exposed to HG or AII compared to NG cells and this HG-induced cellular hypertrophy was inhibited with ARB treatment by 70% (p<0.05). In addition, there were 1.5-fold and 2.0 fold increases in p27Kip1 mRNA and protein expression, respectively, in HG-stimulated podocytes compared to NG- treated cells (p<0.05). p27Kip1 mRNA and protein expression were also increased in cultured podocytes stimulated by AII by 156% and 199%, respectively (p<0.05). ARB treatment ameliorated HG-induced increase in p27Kip1 mRNA by 75% and protein expression by 70% (p<0.05). In contrast, there were no significant changes in p21Cip1 and p57Kip2 protein expression in cultured podocytes exposed to HG or AII. CONCLUSION: High glucose induced significant cellular hypertrophy and increased p27Kip1 mRNA and protein expression in cultured mouse podocytes, and these changes were effectively inhibited by ARB treatment.

Angiotensin II Receptor Blocker Induced Inhibition of Cellular Hypertrophy and Differential Expression of Cyclin-dependent Kinase Inhibitors in Cultured Podocytes Stimulated by Long-term High Glucose / 대한신장학회잡지

BACKGROUND: Hypertrophy of podocytes is observed in type 2 diabetic patients. Cellular hypertrophy requires combined effects of various mitogen- induced entry into the cell cycle and subsequent cell cycle arrest at the G1/S interphase. This cell cycle arrest is mediated by various cyclin-dependent kinase inhibitors (CKIs). We investigated the effect of angiotensin II receptor blocker (ARB) treatment on podocyte hypertrophy and CKIs expression in cultured podocytes stimulated by long-term high glucose. METHODS: Immortalized mouse podocytes were cultured in media containing 5.6 mM normal glucose (NG), 30 mM high glucose (HG), or NG+angiotensin II (AII, 10(-7)M) for 7 days with or without ARB (L-158,809, 10(-6)M). Cellular hypertrophy was assessed by measurement of cellular protein/cell counts, and CKIs mRNA and protein expression were assessed by reverse-transcription polymerase chain reaction (RT-PCR) and Western blot, respectively. RESULTS: Cellular hypertrophy was induced in podocytes exposed to HG or AII compared to NG cells and this HG-induced cellular hypertrophy was inhibited with ARB treatment by 70% (p<0.05). In addition, there were 1.5-fold and 2.0 fold increases in p27Kip1 mRNA and protein expression, respectively, in HG-stimulated podocytes compared to NG- treated cells (p<0.05). p27Kip1 mRNA and protein expression were also increased in cultured podocytes stimulated by AII by 156% and 199%, respectively (p<0.05). ARB treatment ameliorated HG-induced increase in p27Kip1 mRNA by 75% and protein expression by 70% (p<0.05). In contrast, there were no significant changes in p21Cip1 and p57Kip2 protein expression in cultured podocytes exposed to HG or AII. CONCLUSION: High glucose induced significant cellular hypertrophy and increased p27Kip1 mRNA and protein expression in cultured mouse podocytes, and these changes were effectively inhibited by ARB treatment.