Evaluation of Apaf-1, Caspase-3, Bax and Bcl-2 expression in peritoneal and spleen cells derived from mice adjacent to hydatid cyst fluid

Abstract

Hydatidosis is a zoonotic helminth disease that is caused by the larval stage of Echinococcus granulosus worms in humans and other intermediate hosts. Regulating the immune response is a significant strategy by which the establishment and growth of hydatid cysts in internal organs of the host can be warranted. The induction of apoptosis in peritoneal cells may be a considerable component. The current study aimed to evaluate the effect of hydatid cyst fluid (HCF) on the expression of apoptotic proteins in mice peritoneal cells.Materials and Methods: To investigate the apoptotic effect of hydatid cyst fluid on rat peritoneal cells, the liver of sheep infected with hydatid cyst was transferred from slaughterhouse to laboratory. The outer surface of the cyst was first cleaned with 70% alcohol and then sterilized in 50 ml tubes using a syringe, cyst fluid and its protoscoleces. Eight female mice were infected at 8-12 weeks of age by intraperitoneal injection. Animals were infected with 2 cc injection of hydatid cysts and the control group was also infected with 2 cc of PBS. Subsequently, peritoneal cells infected with hydatid fluid were isolated and evaluated for expression of pro-apoptotic Bax, anti-apoptotic Bcl-2 and Apaf-1 Caspase-3 genes at mRNA level. For this purpose, the cells' RNAs were first isolated with a single test kit (RNA extraction kit) and then cDNA synthesized with a YTA kit. Finally, the expression levels of Caspase-3, Bcl-2, Bax and Apaf-1 were quantified by RT-PCR using semi-quantitative GAPDH expression.Result: The results showed that the apoptosis-inducing genes Caspase-3, Bax and Apaf-1 increased significantly, while the anti-apoptotic genes Bcl-2 decreased. The highest expression of apoptosis inducer genes was observed in genes exposed to protoscoleces-containing hydatid fluid.