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Involvement of Endoplasmic Reticulum Stress, Autophagy, and Apoptosis in Advanced Glycation End Products-Induced Glomerular Mesangial Cell Injury

View Article: PubMed Central - PubMed

ABSTRACT

Advanced glycation end-products (AGEs)-induced mesangial cell death is one of major causes of glomerulus dysfunction in diabetic nephropathy. Both endoplasmic reticulum (ER) stress and autophagy are adaptive responses in cells under environmental stress and participate in the renal diseases. The role of ER stress and autophagy in AGEs-induced mesangial cell death is still unclear. Here, we investigated the effect and mechanism of AGEs on glomerular mesangial cells. AGEs dose-dependently decreased mesangial cell viability and induced cell apoptosis. AGEs also induced ER stress signals in a time- and dose-dependent manner. Inhibition of ER stress with 4-phenylbutyric acid effectively inhibited the activation of eIF2α and CHOP signals and reversed AGEs-induced cell apoptosis. AGEs also activated LC-3 cleavage, increased Atg5 expression, and decreased p62 expression, which indicated the autophagy induction in mesangial cells. Inhibition of autophagy by Atg5 siRNAs transfection aggravated AGEs-induced mesangial cell apoptosis. Moreover, ER stress inhibition by 4-phenylbutyric acid significantly reversed AGEs-induced autophagy, but autophagy inhibition did not influence the AGEs-induced ER stress-related signals activation. These results suggest that AGEs induce mesangial cell apoptosis via an ER stress-triggered signaling pathway. Atg5-dependent autophagy plays a protective role. These findings may offer a new strategy against AGEs toxicity in the kidney.

No MeSH data available.


The role of ER stress in AGEs-induced mesangial cell apoptosis.Cells were treated with AGEs (160 μg/ml) with or without ER stress inhibitor 4-phenylbutyric acid (4PBA, 1 mM) treatment for 24 h. The protein levels of phospho-eIF2α, CHOP, LC3, and cleaved caspase-3 were determined by Western blot (A-a). In some experiments, the effect of 4PBA on rapamycin-induced autophagy was tested (A-b). The percentages of apoptotic cells were determined by PI-Annexin V staining (B). Data are presented as mean ± SEM of three independent experiments performed in duplicates (A) or triplicates (B). *P < 0.05 as compared to BSA-treated group or AGEs-treated group.
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f7: The role of ER stress in AGEs-induced mesangial cell apoptosis.Cells were treated with AGEs (160 μg/ml) with or without ER stress inhibitor 4-phenylbutyric acid (4PBA, 1 mM) treatment for 24 h. The protein levels of phospho-eIF2α, CHOP, LC3, and cleaved caspase-3 were determined by Western blot (A-a). In some experiments, the effect of 4PBA on rapamycin-induced autophagy was tested (A-b). The percentages of apoptotic cells were determined by PI-Annexin V staining (B). Data are presented as mean ± SEM of three independent experiments performed in duplicates (A) or triplicates (B). *P < 0.05 as compared to BSA-treated group or AGEs-treated group.

Mentions: 4PBA is a potent ER stress inhibitor, which can attenuate ER stress-induced cell injury3334. We tried to use 4PBA to inhibit AGEs-induced ER stress and figure out the role of ER stress in AGEs-induced mesangial cell apoptosis. Mesangial cells were treated with AGEs (160 μg/ml) with or without 4PBA (1 mM) for 24 h. Co-treatment with 4PBA significantly reversed AGEs-induced CHOP protein expression and eIF2α phosphorylation and caspase-3 cleavage (Fig. 7Aa). Annexin V/PI staining also showed that 4PBA significantly reduced the percentage of AGEs-induced apoptotic mesangial cells (Fig. 7B). Moreover, co-treatment with 4PBA significantly reversed AGEs-induced LC3 cleavage (decrease of LC3-II/LC3-I ratio) and p62 protein degradation (Fig. 7Aa). 4PBA by itself did not affect the rapamycin-induced autophagy in mesangial cells (Fig. 7Ab). These results indicate that AGEs may induce caspase-3 activation and apoptosis and autophagy through an eIF2α/CHOP signaling pathway.


Involvement of Endoplasmic Reticulum Stress, Autophagy, and Apoptosis in Advanced Glycation End Products-Induced Glomerular Mesangial Cell Injury
The role of ER stress in AGEs-induced mesangial cell apoptosis.Cells were treated with AGEs (160 μg/ml) with or without ER stress inhibitor 4-phenylbutyric acid (4PBA, 1 mM) treatment for 24 h. The protein levels of phospho-eIF2α, CHOP, LC3, and cleaved caspase-3 were determined by Western blot (A-a). In some experiments, the effect of 4PBA on rapamycin-induced autophagy was tested (A-b). The percentages of apoptotic cells were determined by PI-Annexin V staining (B). Data are presented as mean ± SEM of three independent experiments performed in duplicates (A) or triplicates (B). *P < 0.05 as compared to BSA-treated group or AGEs-treated group.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5035926&req=5

f7: The role of ER stress in AGEs-induced mesangial cell apoptosis.Cells were treated with AGEs (160 μg/ml) with or without ER stress inhibitor 4-phenylbutyric acid (4PBA, 1 mM) treatment for 24 h. The protein levels of phospho-eIF2α, CHOP, LC3, and cleaved caspase-3 were determined by Western blot (A-a). In some experiments, the effect of 4PBA on rapamycin-induced autophagy was tested (A-b). The percentages of apoptotic cells were determined by PI-Annexin V staining (B). Data are presented as mean ± SEM of three independent experiments performed in duplicates (A) or triplicates (B). *P < 0.05 as compared to BSA-treated group or AGEs-treated group.
Mentions: 4PBA is a potent ER stress inhibitor, which can attenuate ER stress-induced cell injury3334. We tried to use 4PBA to inhibit AGEs-induced ER stress and figure out the role of ER stress in AGEs-induced mesangial cell apoptosis. Mesangial cells were treated with AGEs (160 μg/ml) with or without 4PBA (1 mM) for 24 h. Co-treatment with 4PBA significantly reversed AGEs-induced CHOP protein expression and eIF2α phosphorylation and caspase-3 cleavage (Fig. 7Aa). Annexin V/PI staining also showed that 4PBA significantly reduced the percentage of AGEs-induced apoptotic mesangial cells (Fig. 7B). Moreover, co-treatment with 4PBA significantly reversed AGEs-induced LC3 cleavage (decrease of LC3-II/LC3-I ratio) and p62 protein degradation (Fig. 7Aa). 4PBA by itself did not affect the rapamycin-induced autophagy in mesangial cells (Fig. 7Ab). These results indicate that AGEs may induce caspase-3 activation and apoptosis and autophagy through an eIF2α/CHOP signaling pathway.

View Article: PubMed Central - PubMed

ABSTRACT

Advanced glycation end-products (AGEs)-induced mesangial cell death is one of major causes of glomerulus dysfunction in diabetic nephropathy. Both endoplasmic reticulum (ER) stress and autophagy are adaptive responses in cells under environmental stress and participate in the renal diseases. The role of ER stress and autophagy in AGEs-induced mesangial cell death is still unclear. Here, we investigated the effect and mechanism of AGEs on glomerular mesangial cells. AGEs dose-dependently decreased mesangial cell viability and induced cell apoptosis. AGEs also induced ER stress signals in a time- and dose-dependent manner. Inhibition of ER stress with 4-phenylbutyric acid effectively inhibited the activation of eIF2&alpha; and CHOP signals and reversed AGEs-induced cell apoptosis. AGEs also activated LC-3 cleavage, increased Atg5 expression, and decreased p62 expression, which indicated the autophagy induction in mesangial cells. Inhibition of autophagy by Atg5 siRNAs transfection aggravated AGEs-induced mesangial cell apoptosis. Moreover, ER stress inhibition by 4-phenylbutyric acid significantly reversed AGEs-induced autophagy, but autophagy inhibition did not influence the AGEs-induced ER stress-related signals activation. These results suggest that AGEs induce mesangial cell apoptosis via an ER stress-triggered signaling pathway. Atg5-dependent autophagy plays a protective role. These findings may offer a new strategy against AGEs toxicity in the kidney.

No MeSH data available.