Release of myelin basic protein-degrading proteolytic activity from microglia and macrophages after infection with Theiler's murine encephalomyelitis virus: comparison between susceptible and resistant mice

Theiler's murine encephalomyelitis virus (TMEV) produces a chronic inflammatory demyelinating disease in its natural host, the mouse. A delayed-type hypersensitivity (DTH) response to viral antigens generally correlates with susceptibility to the disease and is thought to play an important role in the pathogenesis of demyelination in this model of human multiple sclerosis (MS). The hallmark of DTH responses is the recruitment by activated Th-1 cells of lymphoid cells and especially macrophages in infected areas. It is believed that soluble factors released by these cells would produce tissue damage, particularly myelin breakdown. In the present study, we compared TMEV-infected macrophages and microglia, isolated from both susceptible SJL/J and resistant C57BL/6 mice, for their ability to secrete proteolytic enzymes capable of degrading myelin basic protein. In addition, we studied whether supernatants from infected microglia/macrophages were also capable of killing oligodendrocytes in the same in vitro system. As detected by SDS-PAGE, MBP-degrading proteolytic activity was found only in supernatants from infected SJL/J microglia and macrophages, but not in supernatants collected from infected C57BL/6 microglia and macrophages, or in supernatants from mock-infected SJL/J and C57BL/6 cells. Similarly, incubation of E20.1 cells, an immortalized line of oligodendrocytes, with infected SJL/J, but not C57BL/6 supernatants, resulted in cytotoxic activity. When cells from resistant C57BL/6 mice were treated with LPS, they became susceptible to infection and also secreted proteolytic enzymes. The proteolytic activity released from infected microglia and macrophages was found to be dose-dependent, was inactivated by heat, and was inhibited by phenylmethylsulphonyl fluoride (PMSF). These results indicate that a serine protease is released from infected microglia and macrophages and suggest a role for proteases in TMEV-induced myelin injury.