Anti-CASP3 monoclonal antibody, clone N488

Objective To investigate the influence of Msx2 conditional gene knockout during lens development in mice. Methods Lens-specific Msx2 knockout mice were generated using the Cre-loxP system. The eyes of Msx2 conditional knockout (Msx2CKO) and wild-type (Msx2WT) mice were examined during embryonic and early postnatal periods using histological, immunofluorescence, in situ hybridization, cell proliferation, apoptosis, and mRNA microarray analyses. ResultsMsx2CKO mice exhibited small lens formation and microphthalmia after birth, while Msx2CKO embryos exhibited a persistent lens stalk, small lens formation, and microphthalmia. Conditional deletion of Msx2 also led to an increased apoptosis rate, a significant reduction in FoxE3 expression, and an upregulation of Prox1 expression in the lens vesicle during the early embryonic period. Microarray comparison of Msx2CKO and Msx2WT lens transcriptomes identified a large number of differentially expressed genes. Real-time PCR showed that Casp8 and Casp3 expression was upregulated in Msx2CKO mice at post-natal day 1. Conclusion The activation of apoptosis through the caspase-8/caspase-3 signaling pathway, together with the downregulation of FoxE3 expression, appeared to account for the smaller lens formation in Msx2CKO mice.

Oxidative stress increases with age in multiple organ systems and is implicated in the development of age-related pathologies in them. Studies from our laboratory show that the intrinsic pathway proapoptotic proteins BAX and caspase-3 (CASP3) lie upstream of mitochondrial production of oxidative stress-inducing reactive species (RS) such as reactive oxygen and reactive nitrogen species (ROS and RNS) in apoptotic and nonapoptotic neurons in cell culture. Our objective in this study was to determine if these findings could be generalized to the development of oxidative stress in nonneuronal tissues in vivo. We first investigated the effect of genetic deletion of Bax on DNA damage in the liver, heart and kidneys of female mice of increasing ages (5, 14, 22 months). The organs of the aged mice showed increased oxidative DNA strand breaks compared to young animals (5 month). Ablation of Bax greatly reduced this damage. We next assessed lipid peroxidation, DNA oxidation, and protein tyrosine nitration to determine whether Casp3 deletion reduces oxidative stress in the hearts, livers, and kidneys of 12-month-old female mice. Lipid peroxides and 8-hydroxy-2'-deoxyguanosine (8-OHdG) levels were much lower in organs from mice with depleted Casp3 than in those of wild type animals. Nitration of protein tyrosine residues, caused by RNS, was also significantly suppressed in the tissues of Casp3 null mice compared to those in wild type mice. Our findings indicate that BAX and CASP3 have a vital role in the generation of oxidative stress in organs of aged mice.