Zell- und Molekularbiologie

Abstrakt

Canine Distemper Virus Causes Apoptosis in HEK-293 Cells by both Extrinsic and Intrinsic Pathways

Sarabjot Singh, Dipak Deka, Barinder Singh Grewal, Mohan Jairath, Ramneek Verma

Background: 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 induce apoptosis in different cell types. The present study was carried out to identify genes and pathways to explain the mechanisms involved in Canine Distemper Virus (CDV) induced apoptosis.

Method: For this, HEK-293 cells were infected with CDV-SH, a Snyder Hill strain of canine distemper virus, at different time points. Viability and apoptotic studies were performed using MTT and DNA laddering assays, respectively. qPCR arrays were custom designed to study the expression profile of 43 apoptotic genes in HEK-293 cells after 6, 12, 24 and 48hrs of CDV infection.

Results: MTT results showed 100%, 84.78%, 79.21% & 76.95% cell viability after 6, 12, 24 and 48hrs after infection. DNA laddering showed a faint laddering pattern at 24hr and 48hr post CDV infection which indicated small amounts of DNA fragmentation. Expression studies revealed increased expression of nineteen genes and down regulation of three genes in all the groups. Ingenuity Pathway Analysis (IPA) showed activation of ‘Apoptosis’ pathway along with significant upstream and downstream regulators in CDV infected HEK-293 cells.

Conclusion: Our study demonstrates that apoptosis could be detected in HEK-293 cell lines, as revealed by DNA laddering 24hrs post CDV infection. qPCR & IPA analysis revealed upregulation of caspase-8, caspase-9 and caspase-3 which showed the involvement of both extrinsic and intrinsic pathways of apoptosis in HEK-293 cells following CDV infection.