Prolonged High Glucose Treatment Increased Orai1 Protein Expression through Inhibition of Lysosomal Pathway in Human Mesangial Cells

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.

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

AJP Renal Physiology ◽

10.1152/ajprenal.00185.2007 ◽

2007 ◽

Vol 293 (4) ◽

pp. F1381-F1390 ◽

Cited By ~ 61

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.

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Knockdown of lncRNA NORAD inhibits the proliferation, inflammation and fibrosis of human mesangial cells under high‑glucose conditions by regulating the miR‑485/NRF1 axis

Experimental and Therapeutic Medicine ◽

10.3892/etm.2021.10306 ◽

2021 ◽

Vol 22 (2) ◽

Author(s):

Linna Wang ◽

Xiaoying Yuan ◽

Lifeng Lian ◽

Huali Guo ◽

Hongxia Zhang ◽

...

Keyword(s):

High Glucose ◽

Mesangial Cells ◽

Human Mesangial Cells

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Glucagon-like peptide-1 receptor pathway inhibits extracellular matrix production by mesangial cells through store-operated Ca2+ channel

Experimental Biology and Medicine ◽

10.1177/1535370219876531 ◽

2019 ◽

Vol 244 (14) ◽

pp. 1193-1201 ◽

Cited By ~ 3

Author(s):

Linjing Huang ◽

Rong Ma ◽

Tingting Lin ◽

Sarika Chaudhari ◽

Parisa Y Shotorbani ◽

...

Keyword(s):

Extracellular Matrix ◽

High Glucose ◽

Protein Production ◽

Matrix Protein ◽

Mesangial Cells ◽

Collagen Iv ◽

Extracellular Matrix Protein ◽

Human Mesangial Cells ◽

Glucagon Like Peptide ◽

Glucagon Like Peptide 1

Glomerular mesangial cell is the major source of mesangial matrix. Our previous study demonstrated that store-operated Ca2+ channel signaling suppressed extracellular matrix protein production by mesangial cells. Recent studies demonstrated that glucagon-like peptide-1 receptor (GLP-1R) pathway had renoprotective effects. However, the underlying mechanism(s) remains unclear. The present study was aimed to determine if activation of GLP-1R decreased extracellular matrix protein production by mesangial cells through upregulation of store-operated Ca2+ function. Experiments were conducted in cultured human mesangial cells. Liraglutide and exendin 9–39 were used to activate and inhibit GLP-1R, respectively. Store-operated Ca2+ function was estimated by evaluating the SOC-mediated Ca2+ entry (SOCE). We found that liraglutide treatment reduced high glucose-stimulated production of fibronectin and collagen IV. The inhibitory effects of liraglutide were not observed in the presence of exendin 9–39. Exendin-4, another GLP-1R agonist also blunted high glucose-stimulated fibronectin and collagen IV production. Treatment of human mesangial cells with liraglutide for 24 h significantly attenuated the high glucose-induced reduction of Orai1 protein. Consistently, Ca2+ imaging experiments showed that the inhibition of high glucose on SOCE was significantly attenuated by liraglutide. However, in the presence of exendin 9–39, liraglutide failed to reverse the high glucose effect. Furthermore, liraglutide effects on fibronectin and collagen IV protein abundance were significantly attenuated by GSK-7975A, a selective blocker of store-operated Ca2+. Taken together, our findings suggest that GLP-1R signaling inhibited high glucose-induced extracellular matrix protein production in mesangial cells by restoring store-operated Ca2+ function. Impact statement Diabetic kidney disease continues to be a major challenge to health care system in the world. There are no known therapies currently available that can cure the disease. The present study provided compelling evidence that activation of GLP-1R inhibited extracellular matrix protein production by glomerular mesangial cells. We further showed that the beneficial effect of GLP-1R was attributed to upregulation of store-operated Ca2+ channel function. Therefore, we identified a novel mechanism contributing to the renal protective effects of GLP-1R pathway. Activation of GLP-1R pathway and/or store-operated Ca2+ channel signaling in MCs could be an option for patients with diabetic kidney disease.

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