Effect of miRNA-802 on PI3K/Akt pathway in insulin resistant skeletal muscle cells / 解放军医学杂志

ABSTRACT

Objective To investigate the expression of miRNA-802 (miR-802) and the relationship with glucose metabolism and insulin sensitivity in insulin resistant skeletal muscle cells. Methods L6 rat myoblasts were recovered, cultured and differentiated into L6 muscle cells, incubated with 0.4 mmol/L palmitic acid (PA) and then randomly divided into two groups: Control group (Con) and PA group (PA). After 24 hours, the glucose concentration in the culture medium was measured to determine whether the model was constructed successfully. After successful establishment of the insulin resistance L6 rat myoblasts model, then the cells were transfected with miR-802 mimic or inhibitor to regulate the expression of miR-802. The experimental groups were divided into 5 subgroups as follows: control group, PA group, PA group infected with miR-802 mimic (PA+miR-802-mimic), PA group infected with miR-802 inhibitor (PA+miR-802-inhibitor), and PA group infected with negative control (PA+negative control). The mRNA and protein expressions of the insulin signaling pathway [including phosphatidylinositol-3kinase (PI3K), protein kinase B (Akt), and glucose transporter 4 (GluT4)] were examined by RT-PCR and Western blotting. Results Compared with control group, the mRNA expression in miR-802 group increased significantly (1.458±0.264 vs. 3.108±0.513). Compared with PA group and PA+negative control group, the mRNA expression in miR-802 group increased obviously in PA+miR-802 mimic group (3.108±0.513, 3.442±0.104 vs. 11.743±0.933), and the mRNA expressions of PI3K (0.724±0.032, 0.682±0.059 vs. 0.494±0.025), Akt (0.819±0.044, 0.718±0.033 vs. 0.754±0.028) and GluT4 (0.719±0.038, 0.666±0.056 vs. 0.427±0.031) significantly decreased, and the protein expression of p-PI3K/PI3K (0.349±0.056, 0.351±0.019 vs. 0.195±0.026), p-Akt/Akt (0.639±0.002, 0.557±0.04 vs. 0.261±0.075)and GluT4 (0.648±0.028, 0.590±0.026 vs. 0.413±0.096) significantly decreased (P<0.01). Compared with PA and PA+negative control group, the mRNA expression in miR-802 group decreased in PA+miR-802 inhibitor group (3.108±0.513, 3.442±0.104 vs. 1.069±0.056), the mRNA expressions of PI3K (0.724±0.032, 0.682±0.059 vs. 0.887±0.016), Akt (0.819±0.044, 0.718±0.033 vs. 0.814±0.026) and GluT4 (0.719±0.038, 0.666±0.056 vs. 0.908±0.054) significantly increased, and the protein expressions of p-PI3K/PI3K (0.349±0.056, 0.351±0.019 vs. 0.494±0.012), p-Akt/Akt (0.639±0.002, 0.557±0.04 vs. 0.951±0.031) and GluT4 (0.648±0.028, 0.590±0.026 vs. 0.756±0.007) significantly increased (P<0.01). Conclusion miR-802 regulates insulin sensitivity and glucose metabolism in L6 muscle cells, so inhibition of miR-802 expression may be a potential therapeutic target for type 2 diabetes.