Modulation of atrazine-induced chromosomal aberrations and cyclin-dependent kinases by aqueous extract of Roylea cinerea (D.Don) Baillon leaves in Allium cepa

Abstract

Roylea cinerea (D.Don) Baillon an indigenous medicinal plant of Lamiaceae family used for the treatment of several diseases. In the present study, its aqueous (leaves) extract was tested for genoprotective action against atrazine-induced chromosomal aberrations in the root tip cells of Allium cepa. Atrazine is a herbicide of triazine class commonly used to inhibit the growth of broad leaf and grassy weeds. In order to find the concentration of atrazine that exhibits maximum toxicity, its different concentrations (1, 5 and 10 µg/mL) were tested. It was observed that 10 µg/mL concentration was more toxic as it reduced the mitotic index and also increased the chromosomal aberrations. Among all the tested concentrations of aqueous (leaves) extracts (0.25. 0.5, 1.0, 1.5 and 3.0 µg/mL), the3.0 µg/mL concentration in both modes of experiments i.e. pre and post showed a significant reduction in chromosomal aberrations induced by atrazine. To understand the mechanism of protection by plant extract on atrazine-induced chromosomal abnormalities the RT-qPCR studies were conducted to observe the expression of marker genes Cyclin-dependent kinases (CDKs) (CDKA:1, CDKB2:1 and CDKD1:1. For this, the RNA was extracted from root tips treated with extract along with atrazine by TRIzol^®. It was observed that aqueous extract of Roylea cinerea (D.Don) Baillon leaves upregulated the CDKs gene expression in both the modes i.e. pre and post treatments. A critical analysis of results indicated that aqueous extract ameliorated the chromosomal aberrations caused by atrazine which may be be due to the increased expression level of CDKs genes.

Figure 1

Representative profile of HPLC chromatogram of different polyphenols detected in aqueous extract of leaves of Royale cinerea (D.Don) Baillon, where (peak 1): Gallic acid , (peak 2): Chlorogenic acid, (peak 3): Epicatechin, (peak 4): Caffeic acid, (peak 5): Umbelliferone, (peak 6): Coumaric acid, (peak 7): tert-Butyl hydroquinone (peak 8), Kaempferol respectively were recorded at 280 nm.

Figure 2

Normal stages of mitosis. (a) prophase; (b) metaphase; (c) anaphase; (d) telophase; chromosomal aberrations; (e) c-metaphase; (f) stickiness at telophase; (g,k) vagrant chromosomes; (h) chromatin bridge; (i) laggard chromosomes; (j) chromosomes fragments/breaks; (l) delayed anaphase with bridge.

Figure 3

Effects of treatments with atrazine (ATZ), sodium azide (S.A), plant extract (P.E), Pre-treatment (PRE) and Post-treatment (POST) on CDKA:1, CDKB:2, CDKD1:1 fold change expression level in the root tip cells of Allium cepa. All values are relative to average expression level, of the root tips cells of Allium cepa treated with the tap water, atrazine, sodium azide, PE, PRE and POST treatment in triplicate, n = 3 and mean ± SE.