Studies on the molecular mechanisms involved in apoptosis triggered by Canine Distemper Virus (CDV)

Abstract

Apoptosis is a form of natural or stress induced cell death that plays a pivotal role in many cellular processes. Virus induced apoptosis is of significant importance since many viruses/viral proteins have been reported to induces apoptosis in different cell types. The present study was carried out to identify genes and pathways to explain the mechanisms involved in CDV induced apoptosis. For this, HCT-15 cells were infected with CDV-SH, a Snyder Hill strain of canine distemper virus, at different time points. Viability and apoptosis studies were performed using MTT and TUNEL assays, respectively. The qPCR arrays were custom designed to study the expression profile of apoptotic genes in HCT-15 cells after 6, 12, 24 and 48 h of CDV infection. Results showed viability of cells as 100%, 88.18%, 78.34% and 74.62% at 6, 12, 24 and 48 h after infection. Expression studies revealed increased expression of TNF-a, TRAIL, Calpain, IFN-b and RIG-1 genes and down-regulation of 21 genes in all the groups. Ingenuity Pathway Analysis (IPA) showed activation of apoptosis signaling pathway in CDV infected HCT-15 cells. Immuno-cytochemistry studies using antibodies against apoptotic protein (caspase-8, caspase-9, caspase-3, MAVS, IRF-3, cytochrome C, MKK7) showed only basal level of expression indicating no dysregulation in the expression of these genes which was in consistence with the qPCR array results. In qPCR & IPA analysis, although up-regulation of caspase 8, 9 and 3 was not observed, we hypothesized that CDV induced apoptosis in HCT-15 cell line might have occurred through caspase independent pathways (like Calpain or other molecule mediated).