Cytotoxicity of Capsaicin on Cultured Human Skin Fibroblast

Yun, Seok Kweon; Kim, Jong Keun; Kim, Seong Jin; Won, Young Ho

Abstract

BACKGROUND
Capsaicin has been shown to have different biologic and toxic effects, depending on non-neuronal cells and several transformed cells, however no study has been reported from cultured human skin fibroblast. OBJECTIVE: Present study was aimed to evaluate the cytotoxicity and its mechanism of capsacin on the cultured human skin fibroblast. MATERIAL AND METHOD: Normal neonatal human fibroblasts were used, and changes of cell survival were measured by MTT assay after the cells were pre-treated with growth factors, receptor antagonist, antioxidants, calcium modulators were pre-treated or co-treated with capsaicin. RESULTS: Suvival of fibroblast was significantly increased by treatment with EGF (10ng/ml), bFGF (10ng/ml), capsazepine (10M) but inhibited by cycloheximide (1g/ml). When 200 M capsaicin was added to fibroblasts, chromatin condensations were observed at 12 hours and cell survival rate was reduced to 25-50% at 24 hours. Vanilloid receptor antagonists, capsazepine and ruthenium red, did not prevent the toxic effect of capsaicin, and 10M capsazepine paradoxically rather enhanced the cytotoxicity. In contrast to bFGF (10ng/ml), EGF (10, 100ng/ml) enhanced the cytotoxicity of capsaicin. Neuropeptides, substance P (1, 10nM) and CGRP (1, 10nM), and a structural analogue to capsaicin, tyrosine (0.3-1.2mM) did not affect the cytotoxicity. However, antioxidants such as trolox (100M) and ascorbic acid (0.1, 0.3 mM) reduced the capsaicin cytotoxicity. Of calcium modulating agents, nifedifine, a Ca2+ channel blocker (10, 20M) and cyclopiazonic acid, a Ca2+-ATPase inhibitor in ER (10M) did not influence the cytotoxicity, however BAPTA/AM (10M) as a chelater for cytoplasmic free calcium ion (10M) significantly decreased capsaicin cytotoxicity. Unlike cycloheximide, z-VAD-FMK, a protein synthesis inhibitor and a non-specific caspase inhibitor, prevented the capsaicin cytotoxicity. The DNA ladder and TUNEL positive cells were observed among the capsaicin treated fibroblasts and Western blot revealed caspase-3 activity. CONCLUSION: The capsaicin-induced cytotoxicity on human skin fibroblasts is likely to suggest the mechanism of an apoptotic pathway, which can possibly be prevented by antioxidants.