Study of the dynamics of in vitro infection with bovine gammaherpesvirus type 4 and apoptosis markers in susceptible cells

Abstract

Bovine gammaherpesvirus type 4 (BoHV-4) shows tropism for the endometrium, in which it causes the death of epithelial and stroma cells. Despite having anti-apoptotic genes in its genome, experiments based on immortalized cell lines have shown that BoHV-4 induces cell death by apoptosis. In the present study, we evaluated BoHV-4 replication, pro-apoptotic (Bax) and anti-apoptotic (Bcl-2) mitochondrial genes expression and chromatin condensation in bovine endometrium primary culture cells (BEC) and in the Madin Darby bovine kidney (MDBK) cell line. Results showed that BoHV-4 has a preference for replication in BEC cells over the MDBK cell line, demonstrated by the high viral titer that is consistent with the tropism of the virus. In BEC cells, chromatin condensation was consistent with the values of viral kinetics at the late stage of infection, accompanied with a balance in the mRNA levels of apoptotic mitochondrial proteins. As a consequence, in those cells viral transmission would be enhanced by inhibiting apoptosis in the early stage of virus proliferation, allowing the complete production of viral progeny, and then, the induction of apoptosis in late stages would allow neighboring cells infection. In MDBK cells replication kinetics was coincident with the up-regulation of Bcl-2, which suggests that the productive infection in MDBK is associated with a lytic phase of the virus or another cell death pathway (probably autophagy mechanism) at the late stage of infection. The results agree with the study of nuclear morphology, where a constant chromatin condensation was observed over time. It is clear that the documented BoHV-4 apoptotic responses observed in the cell lines studied above are not valid in cells from primary cultures. The data presented in this study suggest that BoHV-4 could induce apoptosis in BEC cells without a leading role of the mitochondria pathway. Further studies will be necessary to characterize in detail the programmed cell death pathways involved in BoHV-4 infection in the primary cell cultures evaluated.