Earthworm, a novel in vivo system to validate antimitotic compounds

Abstract

We report here that earthworm can be used as an in vivo system to prescreen antimitotic compounds. The known antimitoticcompound colchicine and the aqueous extract of Acorus calamus L. rhizome were used to examine this system. The antimitoticactivity of colchicine and the rhizome extract was confirmed by methods with Allium cepa root tip, earthworm regeneration, andMTT (3-(4,5-dimethyl thiazolyl-2-yl)-2,5-diphenyltetrazolium bromide). An earthworm regeneration assay examined the regenerationability of tissues from amputated regions of the earthworm. All 3 assays showed that cell division is inhibited with colchicine and theaqueous extract of A. calamus rhizome. Histology studies with amputated earthworms confirmed that the development of the blastemawas observed only in control worms. Immunohistochemistry analysis with proliferation cell nuclear antigen and phospho serine 10histone H3 antibodies showed that amputated worms treated with colchicine or the aqueous extract of A. calamus did not have activecell division. MTT assay with the MCF-7 cell line (human breast carcinoma) further confirmed that the aqueous extract of rhizomeaffected cell proliferation. Our results suggest that the earthworm could be used to prescreen the antimitotic potential of plant extractsor other unknown compounds. It is simple, reproducible, and cost-efficient compared to animal cell line-based methods.