Anti-adipogenesis activities of Zingiber cassumunar Roxb. rhizome extracts on L929 cells evaluated by image-based analysis

Abstract

Some species in family Zingiberaceae have been reported as having anti-adipogenesis activity, but to date Zingiber cassumunar has not been investigated. Therefore, this paper investigates of Z. cassumunar extracts on mouse pre-adipocytes, L929 cell line; the first time this cell has been used as a non-inductive in vitro model. Dry rhizome was extracted by dichloromethane, absolute ethanol, and hot water obtaining the crude extracts: ZCD, ZCE and ZCW respectively. A non-cytotoxic activity was shown on the 24 h exposed cells to all extracts, evaluated by using the MTT (methyl-thiazolyldiphenyl-tetrazolium bromide) assay. An anti-proliferation assay was carried out to each of the extracts for 72 h, to determine the non-toxic inhibition-concentration (IC20). The quantity of lipid droplets in the cells, exposed to the IC20 or IC20/2 of each extract, was assessed by using photomicrographs and image-based assay under the ImageJ computer software. Cellular lipid accumulation was significantly decreased in the exposed cells to each of the extract in a concentration-dependent manner, in comparison to the unexposed cells, by which ZCE was the most effective followed by ZCW and ZCD respectively. Expression of adipogenesis-related genes of the exposed cells to each extraction was evaluated through real-time PCR. All of the extracts down-regulated the activity of PPARγ (the key regulator of adipogenesis) while ZCE up-regulated Pref-1 (the pre-adipocyte regulatory gene inhibiting lipid accumulation). Four groups of genes were down-regulated by at least one or more of the extracts. These findings verified the anti-adipogenesis potential of Z. cassumunar in mammalian adipocyte.