Effects of glucose and insulin on the H9c2 (2-1) cell proliferation may be mediated through regulating glucose transporter 4 expression
Authors: LIU Qian , HUANG Qing-xian , LOU Fu-chen , ZHANG Li , WANG Kun , YU Shan , XU Hua , WANG Qian , ZHANG Ying , HOU Wei-kai 
Published: January 2024

Background
The change of glucose transporter 4 (GLUT4) expression could influence glucose uptake in the myocardial cells and then effect myocardial metabolism, which maybe one of the factor for the diabetes cardiovascular disease. This study aimed to explore the influence of glucose and insulin at different concentrations on H9c2 (2-1) cell proliferation and its GLUT4 expression in vitro, and evaluate the correlation between myocardial cells proliferation and GLUT4 expression. This might be helpful for understanding the relationship between glucose metabolism and cardiovascular disease.

Methods
According to glucose concentrations in culture medium, cultured H9c2 rat myocardial cells were divided into five groups: control group (NC, glucose concentration 5.0 mmol/L), low glucose group (LG, glucose concentration 0.1 mmol/L), high glucose group 1 (HG1, glucose concentration 10 mmol/L), high glucose group 2 (HG2, glucose concentration 15 mmol/L), high glucose group 3 (HG3, glucose concentration 20 mmol/L). Then according to different insulin concentrations in culture medium, each group was further divided into two subgroups: normal insulin subgroup (INSc, insulin concentration 3.8 mU/L), high insulin subgroup (INSh, insulin concentration 7.6 mU/L). H9c2 (2-1) cells were cultured for 1, 2, 3 days, the proliferation of cells were assayed by cell counting Kit-8 assay, the expressions of GLUT4 mRNA and protein were detected with RT-PCR and Western Blotting technique, and the relation between myocardial cells proliferation and GLUT4 expression was evaluated.

Results
Compared with NC group, cell proliferation (OD value) was lower in LG, HG2, HG3 group but higher in HG1 group on the second and the third day (P <0.05). There was a negative correlation between OD value and the glucose level in HG1, HG2, HG3 groups (P <0.05). OD value in INSc subgroups was lower than that in INSh subgroups (P <0.05). GLUT4 mRNA was lower in LG, HG2, HG3 groups than that in NC group (P<0.05). Compared with NC group, GLUT4 mRNA level in HG1 group was higher on the first day but lower on the second and third day (P<0.05). In HG1, HG2 and HG3 groups, GLUT4 mRNA level had a negative correlation with the level of glucose (P <0.05). GLUT4 mRNA in INSc subgroups was lower than that in INSh subgroups (P <0.05). The expression of GLUT4 protein was similar to that of GLUT4 mRNA. There was a positive correlation between H9c2 cell proliferation and GLUT4 expression (P<0.02).

Conclusions
Glucose levels could regulate glucose uptake in myocardial cells through influencing GLUT4 expression, and thus affected the cell proliferation and cell function. Insulin levels could affect the myocardial cell function by regulating GLUT4 expression. Effects of glucose and insulin on the myocardial cells proliferation might be mediated through regulating GLUT4 expression. There may be a mechanism of hyperglycemia pre-accommodation (HGPA) in myocardial cells mediated through regulation of GLUT4 expression.