Hyperthermia induced HIF-1a expression of lung cancer through AKT and ERK signaling pathways

Jun Wan, Wei Wu

Hyperthermia is a promising treatment for human lung cancer, but recurrence of the primary lesion is common, as the residual tumor becomes adapted to heat treatment and growth is induced by hypoxia-triggered HIF-1a expression. Here, we explored the effects of hyperthermia on HIF-1a expression, proliferation, and lung cancer angiogenesis. Human NSCLC NCI-H1650 and SCLC NCI-H446 cell lines were used to examine cell viability, apoptosis, and HIF-1a expression level under a gradient of thermal conditions (37, 42 and 47 °C for 40 min). The 47 °C heat-adapted NCI-H1650 and NCI-H446 sublines (also called NCI-H1650-b and NCI-H446-b cells) had enhanced viability and HIF-1a expression levels compared to the parental and 42 °C heat-adapted cells and were thus used for subsequent research. Concentration gradients of wortmannin and PD98095 were used to inhibit AKT and ERK expression, respectively in the NSCLC NCI-H1650-b and SCLC NCI-H446-b cell lines, and cell growth curves were drawn. Western blots were used to detect the expression of HIF-1a, extracellular signal-regulated kinase (ERK), protein kinase B (AKT), phospho-ERK, and phospho-AKT. We established a subcutaneous transplantation tumor model with wortmannin and PD98095 intervention. Immunohistochemistry was used to detect the expression of HIF-1a and the vascular specific marker CD34, and tumor growth curves were drawn. Following hyperthermia treatment, HIF-1a expression in 47 °C heat-adapted NSCLC and SCLC cell lines was regulated by the AKT pathway. However, HIF-1a expression was also regulated by the ERK pathway in NSCLCs, while SCLCs did not exhibit changes in ERK. These biological behaviors are governed by signaling pathway protein phosphorylation. Furthermore, inhibiting the AKT pathway can suppress the proliferation and angiogenesis potential of both 47 °C heat-adapted NSCLCs and SCLCs, but inhibiting the ERK pathway only affects SCLCs. Our study suggests that following hyperthermia, the proliferation and angiogenesis potential of residual NSCLCs and SCLCs is induced by HIF-1a. However, HIF-1a expression in NSCLCs is regulated by both the AKT and ERK signaling pathway, but HIF-1a expression in SCLCs is regulated only by the AKT signaling pathway. This study sheds light on the molecular regulatory mechanisms of lung cancer recurrence following hyperthermia treatment.