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Protective role of PGC-1α in diabetic nephropathy is associated with the inhibition of ROS through mitochondrial dynamic remodeling.

Guo K, Lu J, Huang Y, Wu M, Zhang L, Yu H, Zhang M, Bao Y, He JC, Chen H, Jia W - PLoS ONE (2015)

Bottom Line: This was associated with an increase in ROS generation and mesangial cell hypertrophy.These data suggest that PGC-1α may protect DN via the inhibition of DRP1-mediated mitochondrial dynamic remodeling and ROS production.These findings may assist the development of novel therapeutic strategies for patients with DN.

View Article: PubMed Central - PubMed

Affiliation: Department of Endocrinology and Metabolism, Shanghai Clinical Center for Diabetes, Shanghai Diabetes Institute, Shanghai Key Laboratory of Diabetes Mellitus, Shanghai Jiaotong University Affiliated Sixth People's Hospital, Shanghai, 200233, China.

ABSTRACT
The overproduction of mitochondrial reactive oxygen species (ROS) plays a key role in the pathogenesis of diabetic nephropathy (DN). However, the underlying molecular mechanism remains unclear. Our aim was to investigate the role of PGC-1α in the pathogenesis of DN. Rat glomerular mesangial cells (RMCs) were incubated in normal or high glucose medium with or without the PGC-1α-overexpressing plasmid (pcDNA3-PGC-1α) for 48 h. In the diabetic rats, decreased PGC-1α expression was associated with increased mitochondrial ROS generation in the renal cortex, increased proteinuria, glomerular hypertrophy, and higher glomerular 8-OHdG (a biomarker for oxidative stress). In vitro, hyperglycemia induced the downregulation of PGC-1α, which led to increased DRP1 expression, increased mitochondrial fragmentation and damaged network structure. This was associated with an increase in ROS generation and mesangial cell hypertrophy. These pathological changes were reversed in vitro by the transfection of pcDNA3-PGC-1α. These data suggest that PGC-1α may protect DN via the inhibition of DRP1-mediated mitochondrial dynamic remodeling and ROS production. These findings may assist the development of novel therapeutic strategies for patients with DN.

No MeSH data available.


Related in: MedlinePlus

Decreased PGC-1α expression is associated with a hyperglycemia-induced increase in mitochondrial fragmentation and ROS generation.A: Kidney mitochondrial ROS production in the control and diabetic rats (DM). ROS production was identified by the fluorescent probe Mitochondrial ROS. B: Immunofluorescent micrographs of 8-hydroxy-2-deoxyguanosine (8-OHdG) in the control and diabetic rats (DM). (Original magnification: ×400). Scale bar: 25 μm. C: Intracellular ROS production was determined by the mitochondrial fluorescent probe in RMCs exposed to normal glucose (NG), mannitol (Man), and high glucose (HG). Scale bar: 100 μm. D, E and F: Mitochondrial morphology was determined using the MitoTracker probe in RMCs exposed to NG, HG, and Man. D: Mitochondrial morphology at x200. Quantitative analysis of mitochondrial morphology was conducted using a computer-assisted morphometric analysis application for the calculation of form factor (FF) and aspect ratio (AR) values. Scale bar: 10 μm. G: Real-time RT-PCR assay for PGC-1α mRNA relative to the NG values. H: Western blotting analysis of PGC-1α expression in NG, HG, and Man. I: A correlation analysis between ROS generation and the mitochondrial morphology parameter, FF. J: A correlation analysis between ROS generation and PGC-1α expression. Data are expressed as the mean ± SD values for 5–10 rats per group or three cells per group, and the experiments were repeated independently at least three times with similar results (**P < 0.01 vs. control or vs. NG).
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pone.0125176.g002: Decreased PGC-1α expression is associated with a hyperglycemia-induced increase in mitochondrial fragmentation and ROS generation.A: Kidney mitochondrial ROS production in the control and diabetic rats (DM). ROS production was identified by the fluorescent probe Mitochondrial ROS. B: Immunofluorescent micrographs of 8-hydroxy-2-deoxyguanosine (8-OHdG) in the control and diabetic rats (DM). (Original magnification: ×400). Scale bar: 25 μm. C: Intracellular ROS production was determined by the mitochondrial fluorescent probe in RMCs exposed to normal glucose (NG), mannitol (Man), and high glucose (HG). Scale bar: 100 μm. D, E and F: Mitochondrial morphology was determined using the MitoTracker probe in RMCs exposed to NG, HG, and Man. D: Mitochondrial morphology at x200. Quantitative analysis of mitochondrial morphology was conducted using a computer-assisted morphometric analysis application for the calculation of form factor (FF) and aspect ratio (AR) values. Scale bar: 10 μm. G: Real-time RT-PCR assay for PGC-1α mRNA relative to the NG values. H: Western blotting analysis of PGC-1α expression in NG, HG, and Man. I: A correlation analysis between ROS generation and the mitochondrial morphology parameter, FF. J: A correlation analysis between ROS generation and PGC-1α expression. Data are expressed as the mean ± SD values for 5–10 rats per group or three cells per group, and the experiments were repeated independently at least three times with similar results (**P < 0.01 vs. control or vs. NG).

Mentions: The in vivo production of mitochondrial ROS was significantly increased in the cortex of diabetic rats (Fig 2A; P < 0.01), and the level of 8-OHdG, one of the predominant forms of free radical-induced oxidative lesions [18], was also significantly higher in the glomeruli of diabetic rats compared to control rats (Fig 2B; P < 0.01). Moreover, in vitro exposure of RMCs to high levels of glucose led to increased ROS production (Fig 2C) and decreased PGC-1α expression (Fig 2G and 2H). To exclude the influence of osmolarity on ROS generation and PGC-1α expression, mannitol was used to treat the cells, but had no effect (Fig 2C). Further mitochondrial morphology analyses indicated that mesangial cells had an extensive network of mitochondria, in which the shape of individual mitochondria was long, tubular, and highly branched under normal glucose conditions (Fig 2D and 2E). In contrast, when cells were grown under high glucose conditions, the mitochondrial network of mesangial cells appeared significantly disrupted, while in the osmotic control medium, there was no significant disruption (Fig 2D and 2E). Accordingly, the average form factor (FF) and aspect ratio (AR) values, which indicate the length and branching of the mitochondria, were lower in RMCs grown in HG medium compared to those grown in NG medium (2.64 vs. 3.82 [P < 0.01] and 1.75 vs. 2.51 [P < 0.01], respectively; Fig 2F). Pearson correlation analysis indicated that the increase in ROS generation was significantly correlated with the mitochondrial morphology parameters, FF and levels of PGC-1α protein (Fig 2I).


Protective role of PGC-1α in diabetic nephropathy is associated with the inhibition of ROS through mitochondrial dynamic remodeling.

Guo K, Lu J, Huang Y, Wu M, Zhang L, Yu H, Zhang M, Bao Y, He JC, Chen H, Jia W - PLoS ONE (2015)

Decreased PGC-1α expression is associated with a hyperglycemia-induced increase in mitochondrial fragmentation and ROS generation.A: Kidney mitochondrial ROS production in the control and diabetic rats (DM). ROS production was identified by the fluorescent probe Mitochondrial ROS. B: Immunofluorescent micrographs of 8-hydroxy-2-deoxyguanosine (8-OHdG) in the control and diabetic rats (DM). (Original magnification: ×400). Scale bar: 25 μm. C: Intracellular ROS production was determined by the mitochondrial fluorescent probe in RMCs exposed to normal glucose (NG), mannitol (Man), and high glucose (HG). Scale bar: 100 μm. D, E and F: Mitochondrial morphology was determined using the MitoTracker probe in RMCs exposed to NG, HG, and Man. D: Mitochondrial morphology at x200. Quantitative analysis of mitochondrial morphology was conducted using a computer-assisted morphometric analysis application for the calculation of form factor (FF) and aspect ratio (AR) values. Scale bar: 10 μm. G: Real-time RT-PCR assay for PGC-1α mRNA relative to the NG values. H: Western blotting analysis of PGC-1α expression in NG, HG, and Man. I: A correlation analysis between ROS generation and the mitochondrial morphology parameter, FF. J: A correlation analysis between ROS generation and PGC-1α expression. Data are expressed as the mean ± SD values for 5–10 rats per group or three cells per group, and the experiments were repeated independently at least three times with similar results (**P < 0.01 vs. control or vs. NG).
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pone.0125176.g002: Decreased PGC-1α expression is associated with a hyperglycemia-induced increase in mitochondrial fragmentation and ROS generation.A: Kidney mitochondrial ROS production in the control and diabetic rats (DM). ROS production was identified by the fluorescent probe Mitochondrial ROS. B: Immunofluorescent micrographs of 8-hydroxy-2-deoxyguanosine (8-OHdG) in the control and diabetic rats (DM). (Original magnification: ×400). Scale bar: 25 μm. C: Intracellular ROS production was determined by the mitochondrial fluorescent probe in RMCs exposed to normal glucose (NG), mannitol (Man), and high glucose (HG). Scale bar: 100 μm. D, E and F: Mitochondrial morphology was determined using the MitoTracker probe in RMCs exposed to NG, HG, and Man. D: Mitochondrial morphology at x200. Quantitative analysis of mitochondrial morphology was conducted using a computer-assisted morphometric analysis application for the calculation of form factor (FF) and aspect ratio (AR) values. Scale bar: 10 μm. G: Real-time RT-PCR assay for PGC-1α mRNA relative to the NG values. H: Western blotting analysis of PGC-1α expression in NG, HG, and Man. I: A correlation analysis between ROS generation and the mitochondrial morphology parameter, FF. J: A correlation analysis between ROS generation and PGC-1α expression. Data are expressed as the mean ± SD values for 5–10 rats per group or three cells per group, and the experiments were repeated independently at least three times with similar results (**P < 0.01 vs. control or vs. NG).
Mentions: The in vivo production of mitochondrial ROS was significantly increased in the cortex of diabetic rats (Fig 2A; P < 0.01), and the level of 8-OHdG, one of the predominant forms of free radical-induced oxidative lesions [18], was also significantly higher in the glomeruli of diabetic rats compared to control rats (Fig 2B; P < 0.01). Moreover, in vitro exposure of RMCs to high levels of glucose led to increased ROS production (Fig 2C) and decreased PGC-1α expression (Fig 2G and 2H). To exclude the influence of osmolarity on ROS generation and PGC-1α expression, mannitol was used to treat the cells, but had no effect (Fig 2C). Further mitochondrial morphology analyses indicated that mesangial cells had an extensive network of mitochondria, in which the shape of individual mitochondria was long, tubular, and highly branched under normal glucose conditions (Fig 2D and 2E). In contrast, when cells were grown under high glucose conditions, the mitochondrial network of mesangial cells appeared significantly disrupted, while in the osmotic control medium, there was no significant disruption (Fig 2D and 2E). Accordingly, the average form factor (FF) and aspect ratio (AR) values, which indicate the length and branching of the mitochondria, were lower in RMCs grown in HG medium compared to those grown in NG medium (2.64 vs. 3.82 [P < 0.01] and 1.75 vs. 2.51 [P < 0.01], respectively; Fig 2F). Pearson correlation analysis indicated that the increase in ROS generation was significantly correlated with the mitochondrial morphology parameters, FF and levels of PGC-1α protein (Fig 2I).

Bottom Line: This was associated with an increase in ROS generation and mesangial cell hypertrophy.These data suggest that PGC-1α may protect DN via the inhibition of DRP1-mediated mitochondrial dynamic remodeling and ROS production.These findings may assist the development of novel therapeutic strategies for patients with DN.

View Article: PubMed Central - PubMed

Affiliation: Department of Endocrinology and Metabolism, Shanghai Clinical Center for Diabetes, Shanghai Diabetes Institute, Shanghai Key Laboratory of Diabetes Mellitus, Shanghai Jiaotong University Affiliated Sixth People's Hospital, Shanghai, 200233, China.

ABSTRACT
The overproduction of mitochondrial reactive oxygen species (ROS) plays a key role in the pathogenesis of diabetic nephropathy (DN). However, the underlying molecular mechanism remains unclear. Our aim was to investigate the role of PGC-1α in the pathogenesis of DN. Rat glomerular mesangial cells (RMCs) were incubated in normal or high glucose medium with or without the PGC-1α-overexpressing plasmid (pcDNA3-PGC-1α) for 48 h. In the diabetic rats, decreased PGC-1α expression was associated with increased mitochondrial ROS generation in the renal cortex, increased proteinuria, glomerular hypertrophy, and higher glomerular 8-OHdG (a biomarker for oxidative stress). In vitro, hyperglycemia induced the downregulation of PGC-1α, which led to increased DRP1 expression, increased mitochondrial fragmentation and damaged network structure. This was associated with an increase in ROS generation and mesangial cell hypertrophy. These pathological changes were reversed in vitro by the transfection of pcDNA3-PGC-1α. These data suggest that PGC-1α may protect DN via the inhibition of DRP1-mediated mitochondrial dynamic remodeling and ROS production. These findings may assist the development of novel therapeutic strategies for patients with DN.

No MeSH data available.


Related in: MedlinePlus