Authors: KAMARAJ RAJAMANIKKAM, SUBRAMANIAM ELAIYA RAJA, SENTHAMARAI KANNAN BALAJI, GANAPATHY NADANA RAJAVADIVU, SUDHAKAR SIVASUBRAMANIAM, KARUPPAIAH PALANICHELVAM
Abstract: We report here that earthworm can be used as an in vivo system to prescreen antimitotic compounds. The known antimitotic compound colchicine and the aqueous extract of Acorus calamus L. rhizome were used to examine this system. The antimitotic activity of colchicine and the rhizome extract was confirmed by methods with Allium cepa root tip, earthworm regeneration, and MTT (3-(4,5-dimethyl thiazolyl-2-yl)-2,5-diphenyltetrazolium bromide). An earthworm regeneration assay examined the regeneration ability of tissues from amputated regions of the earthworm. All 3 assays showed that cell division is inhibited with colchicine and the aqueous extract of A. calamus rhizome. Histology studies with amputated earthworms confirmed that the development of the blastema was observed only in control worms. Immunohistochemistry analysis with proliferation cell nuclear antigen and phospho serine 10 histone H3 antibodies showed that amputated worms treated with colchicine or the aqueous extract of A. calamus did not have active cell division. MTT assay with the MCF-7 cell line (human breast carcinoma) further confirmed that the aqueous extract of rhizome affected cell proliferation. Our results suggest that the earthworm could be used to prescreen the antimitotic potential of plant extracts or other unknown compounds. It is simple, reproducible, and cost-efficient compared to animal cell line-based methods.
Keywords: Antimitotic, Acorus calamus, colchicine, earthworm regeneration assay, Eudrilus eugeniae, immunohistochemistry
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