Downregulation of ROCK-I and ROCK-II gene expression in the cadmium-induced ventral body wall defect chick model.

PURPOSE:In the chick embryo, administration of the heavy metal cadmium (Cd) after 60 h incubation induces the ventral body wall defect (VBW) with similarities to the human omphalocele. Rho-associated coiled-coil-containing protein kinase (ROCK) I and ROCK-II mediate signalling from Rho to the actin cytoskeleton in the Wnt non-canonical pathway. ROCK-I knockout (KO), ROCK-II KO, and ROCK-I/ROCK-II double heterozygous mice have been shown to cause failure of closure of the VBW. The exact mechanism by which Cd acts in the Wnt signalling pathway still remains unclear. We designed this study to test the hypothesis, that the gene expression levels of ROCK-I and ROCK-II are downregulated during the critical period of embryogenesis in the Cd-induced VBW defect chick model. METHODS:Chick embryos were harvested 1 h (1H), 4 h (4H), and 8 h (8H) after treatment of cadmium and divided into two groups: control (n = 8 at each time point), and Cd (n = 8 at each time point). Real-time RT-PCR was performed to evaluate the relative mRNA levels of ROCK-I and ROCK-II expression in the Cd-induced VBW defect chick model. Differences between the two groups at each time point were tested by using Mann-Whitney's U test and statistical significance was accepted at P < 0.05. RESULTS:The relative mRNA levels of ROCK-I and ROCK-II at 4H were significantly decreased in Cd group compared to controls (P < 0.01 and P < 0.001, respectively). The expression levels of ROCK-I and ROCK-II at 1H and 8H were not significantly different between Cd group and controls. CONCLUSIONS:Our results provide evidence, for the first time, that the gene expression levels of ROCK-I and ROCK-II are significantly downregulated at 4 h after treatment of Cd in the VBW defect model of chick embryo. We speculate that the downregulation of ROCK-I and ROCK-II gene expressions during this narrow window of embryogenesis may cause VBW defect by disrupting Wnt non-canonical pathway.

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