Objective: To observe the effects of insulin on the expression of epithelial sodium channel alpha subunit (ENaC-alpha) in human pulmonary adenocarcinoma cell line (A549 cells) and its impact on apoptosis of the cells.
Methods: A549 cells were cultured with insulin in a concentration of 30 U/L for 30 minutes or 60 minutes. Then ENaC-alpha, serum and glucocorticoid-inducible kinase-1 (SGK-1) protein and mRNA were determined with Western blotting and reverse transcription-polymerase chain reaction (RT-PCR). Then chelidonii, a blocking agent of protein kinase C (PKC), which could inhibit the effect of SGK-1, was added to the cells culture for 60 minutes, and ENaC-alpha protein was then determined. Cells were cultured in a hypoxia circumstance for 30 minutes, to induce apoptosis, the rate of which was detected with terminal-deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL), in cells cultured with or without insulin.
Results: Insulin could up-regulate the protein and mRNA expression of ENaC-alpha and SGK-1 in A549 cells, and there was a significant difference at 30 minutes, and it gradually increased with prolongation of time. When A549 cells were cultured without insulin, the protein expression of A549 cells was 100%. The protein expression of ENaC-alpha and SGK-1 was 124% and 135% at 30 minutes after being cultured with insulin, and 186% and 176% at 60 minutes, respectively (all P<0.05). Insulin also could inhibit apoptosis which was induced by hypoxia (10.3% vs. 21.6%, P<0.05).
Conclusion: Insulin can up-regulate expression of the ENaC-alpha in A549 cells via PKC and SGK-1 pathways, also insulin can inhibit hypoxia induced apoptosis. So insulin can be beneficial in the prognosis of acute lung injury/acute respiratory distress syndrome (ALI/ARDS).