Effects And Mechanisms Of Piperlongumine In Hepatocellular Carcinoma Cell Migration/Invasion

The primary liver cancer is one of the most common malignant tumors and the third leading-cause of cancer death globally. Hepatocellular carcinoma (HCC) is the most common type of liver cancers. Most HCC patients are discovered at advanced stages and therefore are lack of effective treatment. Piperlongumine is the biologically active small molecular mides extracted from the pepper, which selective kills many types of tumor cells and is considered to be a potential anticancer drug. However, the anti-cancer effect and mechanisms of PL in hepatocellular carcinomas remains largely unclear. Here, PL was used to treat hepatoma cell line (HepG2、Huh7and LM3) and mouse primary cultured hepatocytes, and cell death or apoptosis was detected by MTT, PI staining and flow cytometry. The results demonstrated that PL selectively killed HCC cells but not normal hepatic cells. At low concentration (<10μM), PL only effectively inhibit migration and invasion of HCC cells, indicating that PL might have therapeutic effects on advanced HCC. Further, PL induced ROS accumulation in HCC cells, activated endoplasmic reticulum (ER) stress-response and its downstream signaling pathway. The antioxidant NAC completely reversed PL’s effects on HCC cytotoxicity and migration/invasion, while ER stress-response inhibitor4-PBA and MAPKs signaling pathway inhibitor SP600125(JNK inhibitor), SB203580(P38inhibitor) and U0126(Erk inhibitor) only reversed PL’s effect on HCC migration, suggesting that ER stress-response and MAPPKs signaling pathway mainly suppressed HCC cell migration upon PL treatment. Results of Western blot analysis showed that NAC apparently reduced PL-induced ER stress-response and MAPKs signaling pathway while suppressing ER stress-response could effectively inhibit activation of MAPKs signaling pathway. Immunofluorescence results showed that the suppression of ROS/ER stress-responses/MAPKs could apparently reduce PL-induced CHOP expression, suggesting that CHOP is a downstream signaling molecule of the ROS-ER-MAPKs axis in PL-treated HCC cells. Knocking-down of CHOP by si-RNA showed that CHOP played a role in HCC cell migration and was the target of PL. Finally, we established HCC-bearing mice via subcutaneous transplantation of mouse hepatoma cells (H22) in mice. PL was administred to HCC-bearing mice and the results demonstrated that PL could effectively inhibit the development of HCC in mice. Meanehile, the activatation of ER-MAPKs-CHOP signaling pathway was evident in HCC-bearing mice. In conclusion, piperlongumine could selectively kill HCC cells and inhibit their migration and invasion through ROS-ER-MAPKs-CHOP signaling pathway. Our results provide a new drug for the therapy of hepatocellular carcinoma in clinic.