scholarly journals Glibenclamide Induces Collagen IV Catabolism in High Glucose-Stimulated Mesangial Cells

2012 ◽  
Vol 2012 ◽  
pp. 1-9 ◽  
Author(s):  
Liping Zhu ◽  
Pedro Cortes ◽  
Clare Hassett ◽  
David W. Taube ◽  
Jerry Yee
Keyword(s):  
Protein Expression ◽  
High Glucose ◽  
Mesangial Cells ◽  
Collagen Iv ◽  
Diabetic Rats ◽  
Protein Accumulation ◽  
Generating Capacity ◽  
Mrna And Protein Expression ◽  
Low Dosage ◽  
Pai 1

We have shown the full prevention of mesangial expansion in insulin-deficient diabetic rats by treatment with clinically-relevant dosages of glibenclamide (Glib). Studies in mesangial cells (MCs) also demonstrated reduction in the high glucose (HG)-induced accumulation of collagens, proposing that this was due to increased catabolism. In the present study, we investigated the signaling pathways that may be implicated in Glib action. Rat primary MCs were exposed to HG for 8 weeks with or without Glib in therapeutic (0.01 μM) or supratherapeutic (1.0 μM) concentrations. We found that HG increased collagen IV protein accumulation and PAI-1 mRNA and protein expression, in association with decreased cAMP generating capacity and decreased PKA activity. Low Glib increased collagen IV mRNA but fully prevented collagen IV protein accumulation and PAI-1 overexpression while enhancing cAMP formation and PKA activity. MMP2 mRNA, protein expression and gelatinolytic activity were also enhanced. High Glib was, overall, ineffective. In conclusion, low dosage/concentration Glib prevents HG-induced collagen accumulation in MC by enhancing collagen catabolism in a cAMP-PKA-mediated PAI-1 inhibition.

Glibenclamide prevents increased extracellular matrix formation induced by high glucose concentration in mesangial cells

2007 ◽  
Vol 292 (1) ◽  
pp. F57-F65 ◽  
Author(s):  
Giovanna Giannico ◽  
Pedro Cortes ◽  
Mohammed H. Baccora ◽  
Clare Hassett ◽  
David W. Taube ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
High Glucose ◽  
Mesangial Cells ◽  
Cell Layer ◽  
Collagen Iv ◽  
Collagen I ◽  
Metabolic Effects ◽  
Extracellular Matrix Components ◽  
Pai 1

Other than stimulation of cell contractility, little is known about the potential metabolic effects induced by sulfonylureas, independently of insulin action. Previous studies from our laboratory demonstrated complete abrogation of glomerulosclerosis in an experimental model of type 1 diabetes chronically (9 mo) treated with low-dose sulfonylureas (Biederman JI, Vera E, Pankhaniya R, Hassett C, Giannico G, Yee J, Cortes P. Kidney Int 67: 554–565, 2005). Therefore, the effects of glibenclamide (Glib) on net collagen I, collagen IV, and fibronectin medium net secretion and cell layer collagen I deposition were investigated in mesangial cells continuously exposed to 25 mM glucose for 8 wk and treated with predetermined increasing concentrations of Glib for the same period. Clinically relevant concentrations (0.01 μM) of Glib fully suppressed the high glucose-enhanced accumulation of collagen I, collagen IV, and fibronectin in the medium and inhibited collagen I deposition in the cell layer. These effects occurred while transforming growth factor (TGF)-β1 medium concentration remained elevated and glucose uptake was increased to levels above those in 25 mM glucose-incubated cultures. The decreased collagen I accumulation occurred simultaneously with enhanced collagen I mRNA expression in concert with marked suppression of plasminogen inhibitor type-1 (PAI-1) mRNA and protein expression. This strongly suggests an accelerated matrix turnover favoring breakdown. Glib-induced effects demonstrated a biphasic pattern, being absent or reversed in cells treated with higher Glib concentrations (0.1 or 1 μM). Therefore, chronic Glib treatment at low concentrations markedly diminishes the high glucose-induced enhanced accumulation of extracellular matrix components by suppression of steady-state PAI-1 transcriptional activity. These results and those previously reported in vivo suggest that long-term Glib treatment may prevent glomerulosclerosis in insulin-deficient diabetes.

scholarly journals Glucose Promotes the Production of Interleukine-1β and Cyclooxygenase-2 in Mesangial Cells via Enhanced (Pro)Renin Receptor Expression

Endocrinology ◽  
2009 ◽  
Vol 150 (12) ◽  
pp. 5557-5565 ◽  
Author(s):  
Jiqian Huang ◽  
Helmy M. Siragy
Keyword(s):  
Angiotensin Ii ◽  
Protein Expression ◽  
Receptor Antagonist ◽  
High Glucose ◽  
Small Interfering Rna ◽  
Mesangial Cells ◽  
Receptor Blockade ◽  
Mrna And Protein Expression ◽  
Cox 2 ◽  
Interfering Rna

Abstract (Pro)renin receptor (PRR) is present in renal glomeruli, and its expression is up-regulated in diabetes. Similarly, renal inflammation is increased in the presence of hyperglycemia. The linkage between PRR and renal inflammation is not well established. We hypothesized that glucose-induced up-regulation of PRR leads to increased production of the proinflammatory factors IL-1β and cyclooxygenase-2 (COX-2). Studies were conducted in rat mesangial cells (RMCs) exposed to 30 mmd-glucose for 2 wk followed by PRR small interfering RNA knockdown, IL-1 receptor blockade with IL-1 receptor antagonist or angiotensin II type 1 receptor blockade with valsartan. The results showed that d-glucose treatment up-regulates prorenin, renin, angiotensin II, PRR, IL-1β, and COX-2 mRNA and protein expression and increases phosphorylation of ERK1/2, c-Jun N-terminal kinase, c-Jun, and nuclear factor-κB (NF-κB) p65 (serine 276,468 and 536), respectively. PRR small interfering RNA attenuated PRR, IL-1β, and COX-2 mRNA and protein expressions and significantly decreased angiotensin II production and phosphorylation of ERK1/2 and NF-κB p65 associated with high glucose exposure. Similarly, IL-1 receptor antagonist significantly reduced COX-2 mRNA and protein expression induced by high glucose. COX-2 inhibition reduced high-glucose-induced PRR expression. We conclude that glucose induces the up-regulation of PRR and its ligands prorenin and renin, leading to increased IL-1β and COX-2 production via the angiotensin II-dependent pathway. It is also possible that PRR could enhance the production of these inflammatory cytokines through direct stimulation of ERK1/2-NF-κB signaling cascade.

Hexosamines and TGF-β1 use similar signaling pathways to mediate matrix protein synthesis in mesangial cells

2004 ◽  
Vol 286 (2) ◽  
pp. F409-F416 ◽  
Author(s):  
Lalit P. Singh ◽  
Kenneith Green ◽  
Michelle Alexander ◽  
Shira Bassly ◽  
Errol D. Crook
Keyword(s):  
High Glucose ◽  
Matrix Protein ◽  
Transforming Growth Factor ◽  
Cell Function ◽  
Mesangial Cells ◽  
Rat Kidney ◽  
Protein Accumulation ◽  
Western Blots ◽  
Matrix Production ◽  
Mes Cells

Hyperglycemia-induced alterations in mesangial (MES) cell function and extracellular matrix (ECM) protein accumulation are seen in diabetic glomerulopathy. Transforming growth factor-β1 (TGF-β1) mediates high-glucose-induced matrix production in the kidney. Recent studies demonstrated that some of the effects of high glucose on cellular metabolism are mediated by the hexosamine biosynthesis pathway (HBP) in which fructose-6-phosphate is converted to glucosamine (GlcN) 6-phosphate. We previously showed that the high-glucose-mediated fibronectin and laminin synthesis in MES cells is mediated by the HBP and that GlcN is more potent than glucose in inducing TGF-β1 promoter luciferase activity. In this study, we investigated the hypothesis that the effects of glucose on MES matrix production occur via hexosamine regulation of TGF-β1. Culturing simian virus (SV)-40-transformed rat kidney MES cells in 25 mM glucose (HG) for 48 h increases cellular fibronectin and laminin levels about twofold on Western blots compared with low glucose (5 mM). GlcN (1.5 mM) or TGF-β1 (2.5-5 ng/ml) for 24-48 h also increases ECM synthesis. However, the effects of HG or GlcN with TGF-β1 are not additive. The presence of anti-TGF-β1 antibodies (20 μg/ml) blocks both TGF-β1- and GlcN-induced fibronectin synthesis. TGF-β1 increased ECM levels via PKA (laminin and fibronectin) and PKC (fibronectin) pathways. Similarly, TGF-β1 and hexosamines led to nonadditive increases in phosphorylation of the cAMP responsive element binding transcription factor. These results suggest that the effects of excess glucose on MES ECM synthesis occur via HBP-mediated regulation of TGF-β1.

P56. Cytokines and NO upregulate prolyl hydroxylase 3 (PHD-3) mRNA and protein expression in rat renal mesangial cells

Nitric Oxide ◽  
2008 ◽  
Vol 19 ◽  
pp. 56
Author(s):  
Meike Boosen ◽  
Miriam Pleskova ◽  
Liliana Schäfer ◽  
Josef Pfeilschifter ◽  
Karl-Friedrich Beck
Keyword(s):  
Protein Expression ◽  
Mesangial Cells ◽  
Prolyl Hydroxylase ◽  
Mrna And Protein Expression

scholarly journals Influence of Acute High Glucose on Protein Abundance Changes in Murine Glomerular Mesangial Cells

2016 ◽  
Vol 2016 ◽  
pp. 1-11 ◽  
Author(s):  
Michelle T. Barati ◽  
James C. Gould ◽  
Sarah A. Salyer ◽  
Susan Isaacs ◽  
Daniel W. Wilkey ◽  
...  
Keyword(s):  
Protein Expression ◽  
Posttranslational Modifications ◽  
High Glucose ◽  
Mesangial Cells ◽  
Expression Patterns ◽  
Growth Conditions ◽  
Glomerular Mesangial Cells ◽  
Two Dimensional Gel Electrophoresis ◽  
Glucose Levels ◽  
Flight Mass Spectrometry

The effects of acute exposure to high glucose levels as experienced by glomerular mesangial cells in postprandial conditions and states such as in prediabetes were investigated using proteomic methods. Two-dimensional gel electrophoresis and matrix assisted laser desorption ionization time of flight mass spectrometry methods were used to identify protein expression patterns in immortalized rat mesangial cells altered by 2 h high glucose (HG) growth conditions as compared to isoosmotic/normal glucose control (NG⁎) conditions. Unique protein expression changes at 2 h HG treatment were measured for 51 protein spots. These proteins could be broadly grouped into two categories: (1) proteins involved in cell survival/cell signaling and (2) proteins involved in stress response. Immunoblot experiments for a protein belonging to both categories, prohibitin (PHB), supported a trend for increased total expression as well as significant increases in an acidic PHB isoform. Additional studies confirmed the regulation of proteasomal subunit alpha-type 2 and the endoplasmic reticulum chaperone and oxidoreductase PDI (protein disulfide isomerase), suggesting altered ER protein folding capacity and proteasomal function in response to acute HG. We conclude that short term high glucose induces subtle changes in protein abundances suggesting posttranslational modifications and regulation of pathways involved in proteostasis.

Downregulation of TRPC6 protein expression by high glucose, a possible mechanism for the impaired Ca2+ signaling in glomerular mesangial cells in diabetes

2007 ◽  
Vol 293 (4) ◽  
pp. F1381-F1390 ◽  
Author(s):  
Sarabeth Graham ◽  
Min Ding ◽  
Sherry Sours-Brothers ◽  
Thomas Yorio ◽  
Jian-Xing Ma ◽  
...  
Keyword(s):  
Patch Clamp ◽  
Protein Expression ◽  
High Glucose ◽  
Mesangial Cells ◽  
Membrane Currents ◽  
Maximum Effect ◽  
Ang Ii ◽  
Glomerular Mesangial Cells ◽  
Fura 2 ◽  
Fluorescence Measurements

The present study was performed to investigate whether transient receptor potential (TRPC)6 participated in Ca2+ signaling of glomerular mesangial cells (MCs) and expression of this protein was altered in diabetes. Western blots and real-time PCR were used to evaluate the expression level of TRPC6 protein and mRNA, respectively. Cell-attached patch-clamp and fura-2 fluorescence measurements were utilized to assess angiotensin II (ANG II)-stimulated membrane currents and Ca2+ responses in MCs. In cultured human MCs, high glucose significantly reduced expression of TRPC6 protein, but there was no effect on either TRPC1 or TRPC3. The high glucose-induced effect on TRPC6 was time and dose dependent with the maximum effect observed on day 7 and at 30 mM glucose, respectively. In glomeruli isolated from streptozotocin-induced diabetic rats, TRPC6, but not TRPC1, was markedly reduced compared with the glomeruli of control rats. Furthermore, TRPC6 mRNA in MCs was also significantly decreased by high glucose as early as 1 day after treatment with maximal reduction on day 4. Patch-clamp experiments showed that ANG II-stimulated membrane currents in MCs were significantly attenuated or enhanced by knockdown or overexpression of TRPC6, respectively. Fura-2 fluorescence measurements revealed that the ANG II-induced Ca2+ influxes were markedly inhibited in MCs with TRPC6 knockdown, reminiscent of the impaired Ca2+ entry in response to ANG II in high glucose-treated MCs. These results suggest that the TRPC6 protein expression in MCs was downregulated by high glucose and the deficiency of TRPC6 protein might contribute to the impaired Ca2+ signaling of MCs seen in diabetes.

scholarly journals Abundance of TRPC6 protein in glomerular mesangial cells is decreased by ROS and PKC in diabetes

2011 ◽  
Vol 301 (2) ◽  
pp. C304-C315 ◽  
Author(s):  
Sarabeth Graham ◽  
Yves Gorin ◽  
Hanna E. Abboud ◽  
Min Ding ◽  
Duck Yoon Lee ◽  
...  
Keyword(s):  
Nadph Oxidase ◽  
Protein Expression ◽  
Transient Receptor Potential ◽  
Cultured Cells ◽  
Mesangial Cells ◽  
Receptor Potential ◽  
Underlying Mechanism ◽  
Diabetic Rats ◽  
Dependent Manner ◽  
Glomerular Mesangial Cells

The present study was performed to investigate the underlying mechanism, particularly the roles of reactive oxygen species (ROS) and protein kinase C (PKC), in the diabetes-induced canonical transient receptor potential 6 (TRPC6) downregulation. We found that high glucose (HG) significantly reduced TRPC6 protein expression in cultured mesangial cells (MCs). TRPC6 protein was also significantly reduced in the glomeruli but not in the heart or aorta isolated from streptozotocin-induced diabetic rats. In the cultured MCs, H2O2 suppressed TRPC6 protein expression in a dose- and time-dependent manner, which emulated the HG effect. Catalase as well as superoxide dismutase were able to prevent the inhibitory effect of HG on TRPC6. The antioxidant effect observed in cultured cells was also observed in diabetic rats treated with tempol for 2 wk, which exhibited a preservation of TRPC6 in the glomeruli. Specific knockdown of Nox4, a component of NADPH oxidase, increased TRPC6 protein expression. Furthermore, the PKC activator phorbol 12-myristate 13-acetate (PMA), but not its analog 4α-phorbol 12, 13-didecanoate (4α-PDD), suppressed TRPC6 expression, and this PMA effect was not affected by catalase. Moreover, Gö6976, but not LY333531, attenuated the negative effect of HG on TRPC6 expression. Gö6976 also inhibited H2O2 effect on TRPC6. Furthermore, either knockdown of TRPC6 or HG treatment significantly decreased ANG II-stimulated MC contraction, and the HG-impaired MC contraction was rescued by overexpression of TRPC6. These results suggest that hyperglycemia in diabetes downregulated TRPC6 protein expression in MCs through a NADPH oxidase Nox4-ROS-PKC pathway, proving a mechanism for impaired MC contraction in diabetes.

Proteomic profile of primary isolated rat mesangial cells in high-glucose culture condition and decreased expression of PSMA6 in renal cortex of diabetic ratsThis paper is one of a selection of papers published in this special issue entitled “Second International Symposium on Recent Advances in Basic, Clinical, and Social Medicine” and has undergone the Journal's usual peer review process.

2010 ◽  
Vol 88 (4) ◽  
pp. 635-648 ◽  
Author(s):  
Zhiguo Li ◽  
Haojun Zhang ◽  
Xi Dong ◽  
Frank J. Burczynski ◽  
Patrick Choy ◽  
...  
Keyword(s):  
Diabetic Nephropathy ◽  
High Glucose ◽  
Gel Electrophoresis ◽  
Renal Cortex ◽  
Mesangial Cells ◽  
Diabetic Rats ◽  
Differentially Expressed ◽  
Differentially Expressed Proteins ◽  
Two Dimensional Gel Electrophoresis ◽  
Isolated Rat

Diabetic nephropathy (DN) is one of the most important complications of diabetic patients and is characterized histologically by an accumulation of extracellular matrix (ECM) protein in the glomerular mesangium. Therefore, mesangial cells likely play an important role in the development of diabetic nephropathy. Here, we employed proteomic techniques to investigate the protein profile of rat mesangial cells under high-glucose culture conditions. Primary isolated rat glomerular mesangial cells were cultured under different concentrations of glucose (5.4 mmol·L–1 for normal control and 30 mmol·L–1 for high glucose) for 0, 8, 16, and 72 h, as well as for 25 days. Cellular total proteins were isolated from these cells and employed for two-dimensional gel electrophoresis (2-DE). Differentially expressed proteins were identified by matrix-assisted laser desorption – ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and some of these proteins were documented in rat models of diabetes by Western blot. Rat mesangial cells were successfully isolated in the laboratory and their proliferation rates were significantly inhibited by high glucose. Two-dimensional gel electrophoresis analyses revealed 28 differentially expressed protein spots between the normal and high-glucose groups. After MALDI-TOF-MS analysis, all 28 protein spots were successfully identified with the peptide mass fingerprint (PMF) method. Representatively, SOD1, PCBP1 and PSMA6 were validated by Western blot analysis following protein extractions from the normal and high-glucose groups. Abundance of these proteins was consistent with that found in 2-DE. Moreover, expression of SOD1, PCBP1, and PSMA6 in renal cortex was further examined in two rat models of diabetes (streptozotocin-induced and spontaneous OLETF diabetic models). Abundance of SOD1 and PCBP1 proteins did not show any significant difference between normal control and diabetic rats. However, abundance of the PSMA6 protein was significantly reduced in the renal cortex of both STZ-induced and spontaneous OLETF diabetic rats. Proteomic analysis identified 28 differentially expressed proteins in primary isolated rat mesangial cells between normal and high glucose treatments. Expression of one identified protein was found to be consistent with expression in the renal cortex of two rat diabetic models. Therefore, identification of protein expression patterns in mesangial cells can be employed to develop a therapeutic target for treatment of diabetic nephropathy.

scholarly journals PO-298 Effect of exercise and Siyeshen upon hippocampal cytochrome c and caspase-3 of diabetic rats

2018 ◽  
Vol 1 (5) ◽  
Author(s):  
Dongmei Wang ◽  
Jinming Zhang ◽  
Haibin Liu ◽  
Rongmei Wang
Keyword(s):  
Cytochrome C ◽  
Protein Expression ◽  
Caspase 3 ◽  
Diabetic Control ◽  
Exercise Group ◽  
Diabetic Rats ◽  
Control Group ◽  
Diabetic Control Group ◽  
Mrna And Protein Expression ◽  
Cyt C

Objective To observe the effects of aerobic exercise and Siyeshen water extract on cytochrome c (Cyt c) and caspase-3 in hippocampus of diabetic rats and to explore the possible mechanism of improving diabetes. Methods Healthy male Wister rats fed with high fat and high sugar and combined with streptozotocin to establish type II diabetes model. They were randomly divided into 4 groups: diabetic control group, exercise group, Siyeshen group and exercise+Siyeshen group, and another normal control group, with 6 rats in each group. After aerobic exercise (15m/min, 5°slope, 60min, every other day) or/and Siyeshen (200mg/kg) gastrointestinal administration for 8w, the expression of Cyt c and caspase-3 in hippocampus of each group were detected by immunoblotting, and mRNA expressions were detected by RT-PCR. Results Compared with the normal control group, the mRNA and protein expressions of Cyt c and caspase-3 in the hippocampus of the diabetic control group were significantly increased (P<0.05). Compared with the diabetic control group, the blood glucose level of exercise group and exercise+ Siyeshen group decreased (P<0.05), the mRNA and protein expression of Cyt c and caspase-3 decreased significantly (P<0.05), but there were no significant changes in the mRNA and protein expression of Cyt c and caspase-3 between Siyeshen group and diabetic control group (P﹥0.05). Conclusions Exercise and exercise combined with Siyeshen can inhibit cytochrome c release and reduce caspase-3 protein expression, which may be related to the improvement of blood glucose levels in diabetic rats.

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