These studies reveal a mechanism by which IFN contributes to enhanced host susceptibility to bacterial infection and show a previously unappreciated mecha nism of antagonistic cross speak concerning sort I and II IFNs. Outcomes AND DISCUSSION L. monocytogenes infection inhibits macrophage responsiveness to IFN To check if L. monocytogenes infection may possibly suppress macrophage responses to IFN, mouse BM derived macro phages have been subjected to a low multiplicity of WT L. monocytogenes 2 h in advance of remedy with IFN. twenty h later on, the infected and handle BMMs were harvested and cell surface MHCII expression on reside gated cells was analyzed by movement cytometry. Mock contaminated BMM handled with IFN showed 50 one hundred? larger MHCII staining than BMM not handled with IFN. Even so, almost 95% of this IFN induced MHCII grow was blocked in BMM cultures that had been infected with wt Lm.
These data propose the infection either exclusively impaired expression of cell surface MHCII expression or additional normally impaired macrophage responsive ness selleck to IFN. Induction of MHCII transcription by IFN involves the class II transactivator CIITA. To inves tigate the affect of wt Lm infection on IFN induced CIITA expression, VX222 VCH222 we evaluated transcription with the CIITA pIV isoform in handle and infected BMM. IFN treat ment greater transcription of CIITA pIV in mock infected BMM as judged by semiquantitative RT PCR. Nevertheless, no induction of CIITA pIV transcripts was witnessed in IFN treated BMM previously contaminated with wt Lm. Simi larly, wt Lm infection prevented IFN induced luciferase reporter activity in RAW CIITApIV reporter macrophages, which had been derived from RAW264. seven macro phages by steady transfection by using a CIITA pIV luciferase reporter construct. Hence, infection of macrophages with L.
monocytogenes suppressed induction of the two CIITA and cell surface MHCII. To even more discern whether or not the suppressive results of L. monocytogenes were particular to CIITA, we developed an

addi tional set of reporter cell lines. RAW264. seven cells had been stably transfected with pHTS Gas, a reporter construct contain ing four Fuel components upstream of the luciferase open reading frame. Reporter exercise during the resulting RAW Gasoline reporter cells was strongly induced by IFN and inhibited by pretreatment with all the STAT1 inhibitory anticancer agent fludarabine. When RAW Gas. six or more independently derived Fuel reporter cell lines have been contaminated with wt Lm ahead of IFN therapy, the induction of reporter gene action was reduced by 50%. Infection with wt Lm failed to suppress luciferase reporter exercise driven by an nfkb promoter. In contrast to wt Lm, infection using a dwell mutant L. monocytogenes strain lacking expression of your LLO hemoly sin had no effect on IFN dependent reporter gene exercise.