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A novel cellular defect in diabetes: membrane repair failure.

Howard AC, McNeil AK, Xiong F, Xiong WC, McNeil PL - Diabetes (2011)

Bottom Line: Skeletal muscle myopathy is a common diabetes complication.Downhill running also resulted in a higher level of repair failure in diabetic mice.However, a repair defect could be induced, in the absence of high glucose, by enhancing AGE binding to RAGE, or simply by increasing cell exposure to AGE.

View Article: PubMed Central - PubMed

Affiliation: Institute of Molecular Medicine and Genetics, Georgia Health Sciences University, Augusta, Georgia, USA.

ABSTRACT

Objective: Skeletal muscle myopathy is a common diabetes complication. One possible cause of myopathy is myocyte failure to repair contraction-generated plasma membrane injuries. Here, we test the hypothesis that diabetes induces a repair defect in skeletal muscle myocytes.

Research design and methods: Myocytes in intact muscle from type 1 (INS2(Akita+/-)) and type 2 (db/db) diabetic mice were injured with a laser and dye uptake imaged confocally to test repair efficiency. Membrane repair defects were also assessed in diabetic mice after downhill running, which induces myocyte plasma membrane disruption injuries in vivo. A cell culture model was used to investigate the role of advanced glycation end products (AGEs) and the receptor for AGE (RAGE) in development of this repair defect.

Results: Diabetic myocytes displayed significantly more dye influx after laser injury than controls, indicating a repair deficiency. Downhill running also resulted in a higher level of repair failure in diabetic mice. This repair defect was mimicked in cultured cells by prolonged exposure to high glucose. Inhibition of the formation of AGE eliminated this glucose-induced repair defect. However, a repair defect could be induced, in the absence of high glucose, by enhancing AGE binding to RAGE, or simply by increasing cell exposure to AGE.

Conclusions: Because one consequence of repair failure is rapid cell death (via necrosis), our demonstration that repair fails in diabetes suggests a new mechanism by which myopathy develops in diabetes.

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An epithelium-derived cell line, BS-C-1, also develops repair failure in high glucose, and this defect is not readily reversible. A: BS-C-1 cells were cultured for 8 weeks in 7.5 mmol/L glucose (N Gluc), 30 mmol/L glucose (H Gluc), or iso-osmolar control 30 mmol/L mannitol (Man). Laser analysis revealed that cells cultured in high glucose displayed a significant increase in dye uptake when compared with normal glucose and mannitol. B: BS-C-1 cells were cultured for 10 weeks in H Gluc and then switched to L Gluc (5.5 mmol/L glucose) for 10 days (H Gluc/L Gluc), or cells were cultured for 11 weeks in L Gluc or H Gluc only. Laser analysis determined that the high glucose membrane repair defect remained 1 week after switching to low glucose (H Gluc/L Gluc). Data are presented as the mean ± SEM. *P < 0.01; 2 weeks, n = 18 for N Gluc +Ca, n = 14 for H Gluc +Ca, n = 12 for Man +Ca, and n = 13 for H Gluc –Ca; 10 weeks, n = 14 for L Gluc, n = 16 for H Gluc, and n = 15 for H Gluc/L Gluc.
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Figure 5: An epithelium-derived cell line, BS-C-1, also develops repair failure in high glucose, and this defect is not readily reversible. A: BS-C-1 cells were cultured for 8 weeks in 7.5 mmol/L glucose (N Gluc), 30 mmol/L glucose (H Gluc), or iso-osmolar control 30 mmol/L mannitol (Man). Laser analysis revealed that cells cultured in high glucose displayed a significant increase in dye uptake when compared with normal glucose and mannitol. B: BS-C-1 cells were cultured for 10 weeks in H Gluc and then switched to L Gluc (5.5 mmol/L glucose) for 10 days (H Gluc/L Gluc), or cells were cultured for 11 weeks in L Gluc or H Gluc only. Laser analysis determined that the high glucose membrane repair defect remained 1 week after switching to low glucose (H Gluc/L Gluc). Data are presented as the mean ± SEM. *P < 0.01; 2 weeks, n = 18 for N Gluc +Ca, n = 14 for H Gluc +Ca, n = 12 for Man +Ca, and n = 13 for H Gluc –Ca; 10 weeks, n = 14 for L Gluc, n = 16 for H Gluc, and n = 15 for H Gluc/L Gluc.

Mentions: To determine if other cell types develop a membrane repair defect after high glucose exposure, we applied the above analysis to BS-C-1 (monkey kidney epithelial) and HeLa (human cervical) cells. As was the case for C2C12 myoblasts, BS-C-1 cells developed a repair deficiency only after 8 weeks of high glucose treatment (Fig. 5A, H Gluc +Ca). BS-C-1 and HeLa survival of the scraping injury was also significantly decreased after 8 weeks of high glucose exposure, compared with cells grown in normal glucose or mannitol (Supplementary Fig. 2A, H Gluc HeLa and BS-C-1). This glucose-induced membrane repair defect remained even after cells were returned to a low glucose medium for over a week (Fig. 5B, H Gluc/L Gluc). Additionally, fibroblasts obtained from diabetic (INS2Akita+/−) mice also displayed a membrane repair defect after culturing in normal glucose medium for 1 week (Supplementary Fig. 2B, INS2).


A novel cellular defect in diabetes: membrane repair failure.

Howard AC, McNeil AK, Xiong F, Xiong WC, McNeil PL - Diabetes (2011)

An epithelium-derived cell line, BS-C-1, also develops repair failure in high glucose, and this defect is not readily reversible. A: BS-C-1 cells were cultured for 8 weeks in 7.5 mmol/L glucose (N Gluc), 30 mmol/L glucose (H Gluc), or iso-osmolar control 30 mmol/L mannitol (Man). Laser analysis revealed that cells cultured in high glucose displayed a significant increase in dye uptake when compared with normal glucose and mannitol. B: BS-C-1 cells were cultured for 10 weeks in H Gluc and then switched to L Gluc (5.5 mmol/L glucose) for 10 days (H Gluc/L Gluc), or cells were cultured for 11 weeks in L Gluc or H Gluc only. Laser analysis determined that the high glucose membrane repair defect remained 1 week after switching to low glucose (H Gluc/L Gluc). Data are presented as the mean ± SEM. *P < 0.01; 2 weeks, n = 18 for N Gluc +Ca, n = 14 for H Gluc +Ca, n = 12 for Man +Ca, and n = 13 for H Gluc –Ca; 10 weeks, n = 14 for L Gluc, n = 16 for H Gluc, and n = 15 for H Gluc/L Gluc.
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Figure 5: An epithelium-derived cell line, BS-C-1, also develops repair failure in high glucose, and this defect is not readily reversible. A: BS-C-1 cells were cultured for 8 weeks in 7.5 mmol/L glucose (N Gluc), 30 mmol/L glucose (H Gluc), or iso-osmolar control 30 mmol/L mannitol (Man). Laser analysis revealed that cells cultured in high glucose displayed a significant increase in dye uptake when compared with normal glucose and mannitol. B: BS-C-1 cells were cultured for 10 weeks in H Gluc and then switched to L Gluc (5.5 mmol/L glucose) for 10 days (H Gluc/L Gluc), or cells were cultured for 11 weeks in L Gluc or H Gluc only. Laser analysis determined that the high glucose membrane repair defect remained 1 week after switching to low glucose (H Gluc/L Gluc). Data are presented as the mean ± SEM. *P < 0.01; 2 weeks, n = 18 for N Gluc +Ca, n = 14 for H Gluc +Ca, n = 12 for Man +Ca, and n = 13 for H Gluc –Ca; 10 weeks, n = 14 for L Gluc, n = 16 for H Gluc, and n = 15 for H Gluc/L Gluc.
Mentions: To determine if other cell types develop a membrane repair defect after high glucose exposure, we applied the above analysis to BS-C-1 (monkey kidney epithelial) and HeLa (human cervical) cells. As was the case for C2C12 myoblasts, BS-C-1 cells developed a repair deficiency only after 8 weeks of high glucose treatment (Fig. 5A, H Gluc +Ca). BS-C-1 and HeLa survival of the scraping injury was also significantly decreased after 8 weeks of high glucose exposure, compared with cells grown in normal glucose or mannitol (Supplementary Fig. 2A, H Gluc HeLa and BS-C-1). This glucose-induced membrane repair defect remained even after cells were returned to a low glucose medium for over a week (Fig. 5B, H Gluc/L Gluc). Additionally, fibroblasts obtained from diabetic (INS2Akita+/−) mice also displayed a membrane repair defect after culturing in normal glucose medium for 1 week (Supplementary Fig. 2B, INS2).

Bottom Line: Skeletal muscle myopathy is a common diabetes complication.Downhill running also resulted in a higher level of repair failure in diabetic mice.However, a repair defect could be induced, in the absence of high glucose, by enhancing AGE binding to RAGE, or simply by increasing cell exposure to AGE.

View Article: PubMed Central - PubMed

Affiliation: Institute of Molecular Medicine and Genetics, Georgia Health Sciences University, Augusta, Georgia, USA.

ABSTRACT

Objective: Skeletal muscle myopathy is a common diabetes complication. One possible cause of myopathy is myocyte failure to repair contraction-generated plasma membrane injuries. Here, we test the hypothesis that diabetes induces a repair defect in skeletal muscle myocytes.

Research design and methods: Myocytes in intact muscle from type 1 (INS2(Akita+/-)) and type 2 (db/db) diabetic mice were injured with a laser and dye uptake imaged confocally to test repair efficiency. Membrane repair defects were also assessed in diabetic mice after downhill running, which induces myocyte plasma membrane disruption injuries in vivo. A cell culture model was used to investigate the role of advanced glycation end products (AGEs) and the receptor for AGE (RAGE) in development of this repair defect.

Results: Diabetic myocytes displayed significantly more dye influx after laser injury than controls, indicating a repair deficiency. Downhill running also resulted in a higher level of repair failure in diabetic mice. This repair defect was mimicked in cultured cells by prolonged exposure to high glucose. Inhibition of the formation of AGE eliminated this glucose-induced repair defect. However, a repair defect could be induced, in the absence of high glucose, by enhancing AGE binding to RAGE, or simply by increasing cell exposure to AGE.

Conclusions: Because one consequence of repair failure is rapid cell death (via necrosis), our demonstration that repair fails in diabetes suggests a new mechanism by which myopathy develops in diabetes.

Show MeSH
Related in: MedlinePlus