Rat CASP9(Caspase 9) ELISA Kit

Enterovirus A71 (EV-A71) is one of the causative agents causing the hand-foot-mouth disease which associated with fatal neurological complications. Several sporadic outbreaks of EV-A71 infections have been recently reported from Asia-Pacific regions and potentially established endemicity in the area. Currently, there is no effective vaccine or antiviral drug for EV-A71 available. This may be attributable to the limited information about its pathogenesis. In this study, the recombinant nonstructural 2B protein of EV-A71 was successfully produced in human neuroblastoma SH-SY5Y cells and evaluated for its effects on induction of the cell apoptosis and the pathway involved. The EV-A71 2B-transfected SH-SY5Y cells showed significantly higher difference in the cell growth inhibition than the mock and the irrelevant protein controls. The transfected SH-SY5Y cells underwent apoptosis and showed the significant upregulation of caspase-9 (CASP9) and caspase-12 (CASP12) genes at 3- and 24-h post-transfection, respectively. Interestingly, the level of cytosolic Ca2+ was significantly elevated in the transfected SH-SY5Y cells at 6- and 12-h post-transfection. The caspase-9 is activated by mitochondrial signaling pathway while the caspase-12 is activated by ER signaling pathway. The results suggested that EV-A71 2B protein triggered transient increase of the cytosolic Ca2+ level and associated with ER-mitochondrial interactions that drive the caspase-dependent apoptosis pathways. The detailed mechanisms warrant further studies for understanding the implication of EV-A71 infection in neuropathogenesis. The gained knowledge is essential for the development of the effective therapeutics and antiviral drugs.

The aim of this study was to investigate effects of nitric oxide (NO) on steroidogenesis and apoptosis in goat luteinized granulosa cells (LGCs). We cultured goat LGCs from healthy follicles in culture medium supplemented with the NO donor sodium nitroprusside (SNP) or the NO synthase inhibitor N-omega-Nitro-Larginine methyl ester hydrochloride (L-NAME), then examined steroid synthesis, oxidative stress and apoptosis in vitro. The results showed that SNP treatment significantly increased the cGMP concentration in the LGCs (P < 0.05), whereas the L-NAME treatment significantly decreased cGMP concentration (P < 0.05). Then Inhibition of NO production significantly inhibited the expression of CYP19A1, a key gene that is involved in sex steroid hormones synthesis and is responsible for the decrease of E-2. Inhibition of NO production resulted in an increased percentage of apoptosis, which was accompanied by upregulating expression levels of apoptosis-related markers BAX, CASP3 and CASP9. These data indicate that NO is required for goat LGCs steroidogenesis and cell survival. Furthermore, Inhibition of NO production decreased the expression of mitochondrial biogenesis related genes and proteins (PPARGCIA, NRF-1 and TFAM) and the mtDNA copy number. Simultaneously, inhibition of NO production suppressed the transcription and translation of SOD, GPX1, and CAT, and decreased the glutathione level and increased the 8-OHdG level. However, SNP treatment increased the expression of genes involved in mitochondrial function and biogenesis, and elevated the anti-oxidant stress system and steroid synthesis. Together, our results indicate that NO may up-regulate the expression of PPARGCIA and its downstream factors through the cGMP pathway, thereby decreasing granulosa cell apoptosis, and may participate in the regulation of granulocyte steroid production through the mitochondrial-dependent pathway. (C) 2018 Elsevier Inc. All rights reserved.