Coronavirus MHV-3-Induced Apoptosis in Macrophages

Abstract

Infection with mouse hepatitis virus strain 3 (MHV-3) results in lethal fulminant hepatic necrosis in fully susceptible BALB/c mice compared to the minimal disease observed in resistant strain A/J mice. Macrophages play a central role in the pathogenesis of MHV-3-induced hepatitis. In the present study we have shown that MHV-3 infection of macrophages induces these cells to undergo apoptosis. Three methods to detect apoptosis were applied: flow cytometry analysis of nuclear DNA content, fluorescence microscopic visualization of apoptotic cells labeled by the TUNEL assay, and gel electrophoresis to detect DNA laddering. Apoptosis in A/J and BALB/c macrophages was first detected at 8 h postinfection (p.i.) and reached a maximum by 12 h p.i. The degree of MHV-3-induced apoptosis was much greater in A/J-derived macrophages than in BALB/c-derived cells. Apoptosis was inversely correlated with the development of typical MHV cytopathology, namely syncytia formation. Infected macrophages from A/J mice did not form synctia in contrast to the extensive synctia formation observed in BALB/c-derived macrophages. In MHV-3-infected BALB/c macrophage cultures, apoptotic cells were not incorporated into syncytia. Apoptosis was also inversely correlated with the expression of MHV-3-induced fgl2 prothrombinase in macrophages. These results add the murine coronavirus MHV-3 to the list of RNA-containing viruses capable of inducing apoptosis.

RNase L-Independent Specific 28S rRNA Cleavage in Murine Coronavirus-Infected Cells

“…Progress in molecular biological and biochemical techniques has advanced our knowledge of the intracellular biochemical events of coronavirus replication, whereas the specific basis for the deleterious effects on host cells is not as well understood. Some progress has been made regarding the mechanism of cell death in coronavirus-infected cells; infection of coronavirus transmissible gastroenteritis virus and MHV induces apoptosis in certain cells (1,3,8). As found for some lytic viruses (9,11,20,21), host protein translation is inhibited (12,42,49,50) but not completely shut off in MHV-infected cells.…”

Expression of B and T lymphocyte attenuator (BTLA) in macrophages contributes to the fulminant hepatitis caused by murine hepatitis virus strain-3

“…The protocol for the isolation and purification of peritoneal macrophages has been described previously 17. The transcriptional message of apoptosis-associated genes in macrophages isolated from infected BTLA −/− and WT mice was detected by Apoptosis PCR Array.…”

“…The protocols of these two methods are described in online supplementary materials and methods. For in vitro infection experiments, macrophages were treated with MHV-3 (1000 PFU, multiplicity of infection of 0.001), and the virus titres were determined at 24 h as described previously 17. For macrophage transfer experiments, the mice were injected via tail veins with 1×10 5 to 3×10 5 purified macrophages at 3 h after MHV-3 infection.…”

Physiological functions and clinical implications of fibrinogen-like 2: A review

“…These BALB/c-susceptible and A/J-resistant phenotypes might be attributable to several factors. First, MHV3 induced significantly greater apoptosis of macrophages from A/J mice than from BALB/cJ mice [64] . MHV3 is propagated in macrophages [19] .…”

“…Apoptosis of macrophages might decrease the extent of viral replication and result in higher MHV3 viral load in BALB/cJ than in A/J mice. Additionally, macrophages are a source of FGL2 production [19,56,64] . Fgl2 +/+ - macrophages exhibited a robust procoagulant response to MHV3; whereas procoagulant activity from Fgl2 −/− macrophages exposed to MHV3 was comparable to the control levels [23] .…”