Visna/maedi virus-induced apoptosis involves the intrinsic mitochondrial pathway.

Visna/maedi virus (VMV) infection in sheep choroid plexus cells was associated with the appearance of apoptosis and the implication of a caspase-dependent mechanism. Sheep choroid plexus cells were mock-infected or infected with VMV to examine the time course of activation of the intrinsic pathway of apoptosis. The role of mitochondria and related apoptotic events were evaluated. A drop in mitochondrial potential was observed following mitochondrial membrane permeabilization using JC-1, a fluorescent probe, which shifted its fluorescence emission from green to red. Apoptosis Inducing Factor translocated to the nucleus of infected-cells and this translocation was concomitant with the release of cytochrome c in the cytosol of infected-cells and mitochondrial membrane permeabilization which seemed to be regulated by the p53 pathway. Following phosphorylated p53 induced downregulation of bcl-2. In addition, DNA flow cytometric analyses revealed a sub-G peak characteristic of an apoptotic population that gradually appeared as virus infection progressed. No cell cycle arrest was detected in infected cells while p21 expression increased. It was concluded that VMV apoptosis is mediated in part by the activation of p53 and the intrinsic mitochondrial apoptotic pathway.

Implication of caspases during maedi-visna virus-induced apoptosis.

Maedi-visna virus (MVV) causes encephalitis, pneumonia and arthritis in sheep. In vitro, MVV infection and replication lead to strong cytopathic effects characterized by syncytia formation and subsequent cellular lysis. It was demonstrated previously that MVV infection in vitro induces cell death of sheep choroid plexus cells (SCPC) by a mechanism that can be associated with apoptotic cell death. Here, the relative implication of several caspases during acute infection with MVV is investigated by employing diverse in vitro and in situ strategies. It was demonstrated using specific pairs of caspase substrates and inhibitors that, during in vitro infection of SCPC by MVV, the two major pathways of caspase activation (i.e. intrinsic and extrinsic pathways) were stimulated: significant caspase-9 and -8 activities, as well as caspase-3 activity, were detected. To study the role of caspases during MVV infection in vitro, specific, cell-permeable, caspase inhibitors were used. First, these results showed that both z-DEVD-FMK (a potent inhibitor of caspase-3-like activities) and z-VAD-FMK (a broad spectrum caspase inhibitor) inhibit caspase-9, -8 and -3 activities. Second, both irreversible caspase inhibitors, z-DEVD-FMK and z-VAD-FMK, delayed MVV-induced cellular lysis as well as virus growth. Third, during SCPC in vitro infection by MVV, cells were positively stained with FITC-VAD-FMK, a probe that specifically stains cells containing active caspases. In conclusion, these data suggest that MVV infection in vitro induces SCPC cell death by a mechanism that is strongly dependent on active caspases.