Proliferative Inhibition, Apoptotic Induction, Migratory Suppression of Piper nigrum Extract on HeLa Cervical Cell Line

Abstract

Piper nigrum is broadly acted as a herbal medicine and anticancer activity as well. Previously, we reported that Piper nigrum extract caused induction of breast cancer cells death and reduction of cell migration in in vitro study. In this present study, we need to explore the anticancer effects of Piper nigrum on the human HeLa cervical cancer cells and underlying mechanism. Sulforhodamine B, colony formation, cell cycle arrest, acridine orange/ethidium bromide staining was performed to analyze cytotoxic effects on the cells. Tetraethylbenzimidazolylcarbocyanine iodide staining, dichlorodihydrofluorescein diacetate staining was used to study apoptotic effects and followed by wound healing method, investigated the cells migration. This Piper nigrum crude extract represented cytotoxic effect against HeLa cervical cancer cells by dose and time-dependent manner with the half-maximal inhibitory concentration values was 48.59±5.37, 36.49±4.34, 22.71±2.12 μg/ml for 24 h, 48 h and 72 h, respectively. Further, these extract caused reduction of cell viability by examining the acridine orange/ethidium bromide staining and decreased the colony formation by dose-dependent manner as well. Next, HeLa cervical cancer cells were significantly arrested in the resting/growth 1 phase and further reported with attenuating mitochondrial function at 24 h incubation, especially at the dose of 100 μg/ml of extract. Then, the molecular mechanisms of Piper nigrum extract were evaluated for reactive oxygen species formation, and data showed that these extracts increased reactive oxygen species production by a dose-dependent amount. On the other hand, the cancer cells migration was suppressed by Piper nigrum extract at 48 h observation time. These results indicated that Piper nigrum extract inhibited HeLa cervical cancer cells through the down-regulation of cell proliferation, induction of apoptosis and attenuation of cell migration. Piper nigrum may be used as anticancer agent for preventing/treating cervical cancer.