Palmitate-induced Slc2a4/GLUT4 downregulation in L6 muscle cells: evidence of inflammatory and endoplasmic reticulum stress involvement.

BACKGROUND: Obesity is strongly associated to insulin resistance, inflammation, and elevated plasma free fatty acids, but the mechanisms behind this association are not fully comprehended. Evidences suggest that endoplasmic reticulum (ER) stress may play a role in this complex pathophysiology. The aim of the present study was to investigate the involvement of inflammation and ER stress in the modulation of glucose transporter GLUT4, encoded by Slc2a4 gene, in L6 skeletal muscle cells. METHODS: L6 cells were acutely (2 h) and chronically (6 and 12 h) exposed to palmitate, and the expression of several proteins involved in insulin resistance, ER stress and inflammation were analyzed. RESULTS: Chronic and acute palmitate exposure significantly reduced GLUT4 protein (~ 39%, P < 0.01) and its mRNA (18%, P < 0.01) expression. Only acute palmitate treatment increased GRP78 (28%, P < 0.05), PERK (98%, P < 0.01), eIF-2A (35%, P < 0.01), IRE1a (60%, P < 0.05) and TRAF2 (23%, P < 0.05) protein content, and PERK phosphorylation (106%, P < 0.001), but did not elicit eIF-2A, IKK phosphorylation or increased XBP1 nuclear content. Additionally, acute and chronic palmitate increased NFKB p65 nuclear content (~ 30%, P < 0.05) and NFKB binding activity to Slc2a4 gene promoter (~ 45%, P < 0.05). CONCLUSION: Different pathways are activated in acute and chronic palmitate induced-repression of Slc2a4/GLUT4 expression. This regulation involves activation of initial component of ER stress, such as the formation of a IRE1a-TRAF2-IKK complex, and converges to NFKB-induced repression of Slc2a4/GLUT4. These results link ER stress, inflammation and insulin resistance in L6 cells.

Oleic and linoleic fatty acids downregulate Slc2a4/GLUT4 expression via NFKB and SREBP1 in skeletal muscle cells.

Oleic (OA) and linoleic (LA) fatty acids may be important regulators of Slc2a4 gene (GLUT4 protein) in skeletal muscle, thus participating in insulin resistance. We investigated the effect of OA and LA on the Slc2a4/GLUT4 expression in L6 muscle cells; as well as potential transcriptional regulators. OA and LA (50-400 µM) decreased the Slc2a4/GLUT4 expression in a dose-dependent way (maximum of ~50%, P < 0.001). OA and LA did not alter the Slc2a4-binding activity of oxysterols-receptor-LXR-alpha and peroxisome-proliferator-activated-receptor-gamma; but decreased the Slc2a4-binding activity of the sterol-regulatory-element-binding-protein-1 (SREBP1) enhancer (50%, P < 0.001), and increased (~30%, P < 0.001) the nuclear proteins binding into the Slc2a4-nuclear-factor-NF-kappa-B-binding site (repressor), and the phosphorylation of the inhibitors of nuclear-factor-kappa-B-kinase alpha/beta (150-300%, P < 0.001). In sum, OA and LA are potent inhibitors of the Slc2a4/GLUT4 expression in muscle cells; an effect involving reduced SREBP1 and increased NFKB transcriptional activity. These regulations may participate in the fatty acid-related pathophysiology of insulin resistance.

Cigarette smoke exposure severely reduces peripheral insulin sensitivity without changing GLUT4 expression in oxidative muscle of Wistar rats.

OBJECTIVE: To evaluate the effect of exposure to cigarette smoke and running training on a treadmill on the expression of glucose transporter GLUT4 in oxidative soleus muscle of rats. MATERIALS AND METHODS: Wistar rats were divided into: (C) control, (E) exercise control, (SS), sedentary smoker, and (ES) exercise smoker. Insulin Tolerance Test, Western blotting, and RT-PCR were performed for the evaluation of GLUT4 levels. RESULTS: The SS group presented lower insulin sensitivity with reduced GLUT4 protein in the plasma membrane (PM), no changes in the microsomal fraction, but increased mRNA content. Training reversed this condition. No intervention altered total GLUT4 content of the oxidative muscle. CONCLUSION: These results suggest that passive smoking stimulates GLUT4 transcription without changing total protein content, but impairs the ability of GLUT4 translocation to the PM. On the other hand, training seems to reduce the deleterious effects, even under the influence of cigarette smoking.

Cigarette smoke exposure severely reduces peripheral insulin sensitivity without changing GLUT4 expression in oxidative muscle of Wistar rats / Exposição à fumaça de cigarro reduz gravemente a sensibilidade à insulina periférica sem alterar a expressão de GLUT4 em músculo oxidativo de ratos Wistar

OBJECTIVE: To evaluate the effect of exposure to cigarette smoke and running training on a treadmill on the expression of glucose transporter GLUT4 in oxidative soleus muscle of rats. MATERIALS AND METHODS: Wistar rats were divided into: (C) control, (E) exercise control, (SS), sedentary smoker, and (ES) exercise smoker. Insulin Tolerance Test, Western blotting, and RT-PCR were performed for the evaluation of GLUT4 levels. RESULTS: The SS group presented lower insulin sensitivity with reduced GLUT4 protein in the plasma membrane (PM), no changes in the microsomal fraction, but increased mRNA content. Training reversed this condition. No intervention altered total GLUT4 content of the oxidative muscle. CONCLUSION: These results suggest that passive smoking stimulates GLUT4 transcription without changing total protein content, but impairs the ability of GLUT4 translocation to the PM. On the other hand, training seems to reduce the deleterious effects, even under the influence of cigarette smoking.

OBJETIVO: Avaliar o efeito da exposição à fumaça de cigarro e do treinamento de corrida em esteira rolante sobre a expressão do transportador de glicose GLUT4 no músculo oxidativo solear de ratos. MATERIAIS E MÉTODOS: Ratos Wistar foram divididos em: (C) controle, (E) exercitado, (SS) fumante sedentário e (ES) fumante exercitado. Foram realizados testes de tolerância à insulina, Western Blotting e RT-PCR para avaliação da expressão de GLUT4. RESULTADOS: O grupo SS apresentou menor sensibilidade à insulina, com redução de proteína GLUT4 na membrana plasmática (MP), sem alteração na fração microssomal, e conteúdo de RNAm aumentado. O treinamento reverteu esse quadro. Nenhuma intervenção alterou o conteúdo total de GLUT4 no músculo oxidativo. CONCLUSÃO: Esses resultados sugerem que o fumo estimula a transcrição de GLUT4 sem alterar o conteúdo total de proteína, porém prejudica a capacidade de translocação para a MP. Já o treinamento em esteira parece, mesmo sob influência do cigarro, reduzir seus efeitos deletérios.