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Novel cell type-specific antiviral mechanism of interferon gamma action in macrophages.

Presti RM, Popkin DL, Connick M, Paetzold S, Virgin HW - J. Exp. Med. (2001)

Bottom Line: IFN-gamma inhibited MCMV replication in a signal transducer and activator of transcription (STAT)-1alpha-dependent manner much more effectively in BMMphi (approximately 100-fold) than MEF (5-10-fold).IFN-gamma inhibition of MCMV growth was independent of known mechanisms involving IFN-alpha/beta, tumor necrosis factor alpha, inducible nitric oxide synthase, protein kinase RNA activated (PKR), RNaseL, and Mx1, and did not involve IFN-gamma-induced soluble mediators.These studies in primary cells define a novel mechanism of IFN-gamma action restricted to Mphi, a cell type key for MCMV pathogenesis and latency.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Immunology and the Department of Molecular Microbiology, Washington University School of Medicine, St. Louis, Missouri 63110, USA.

ABSTRACT
Interferon (IFN)-gamma and macrophages (Mphi) play key roles in acute, persistent, and latent murine cytomegalovirus (MCMV) infection. IFN-gamma mechanisms were compared in embryonic fibroblasts (MEFs) and bone marrow Mphi (BMMphi). IFN-gamma inhibited MCMV replication in a signal transducer and activator of transcription (STAT)-1alpha-dependent manner much more effectively in BMMphi (approximately 100-fold) than MEF (5-10-fold). Although initial STAT-1alpha activation by IFN-gamma was equivalent in MEF and BMMphi, microarray analysis demonstrated that IFN-gamma regulates different sets of genes in BMMphi compared with MEFs. IFN-gamma inhibition of MCMV growth was independent of known mechanisms involving IFN-alpha/beta, tumor necrosis factor alpha, inducible nitric oxide synthase, protein kinase RNA activated (PKR), RNaseL, and Mx1, and did not involve IFN-gamma-induced soluble mediators. To characterize this novel mechanism, we identified the viral targets of IFN-gamma action, which differed in MEF and BMMphi. In BMMphi, IFN-gamma reduced immediate early 1 (IE1) mRNA during the first 3 h of infection, and significantly reduced IE1 protein expression for 96 h. Effects of IFN-gamma on IE1 protein expression were independent of RNaseL and PKR. In contrast, IFN-gamma had no significant effects on IE1 protein or mRNA expression in MEFs, but did decrease late gene mRNA expression. These studies in primary cells define a novel mechanism of IFN-gamma action restricted to Mphi, a cell type key for MCMV pathogenesis and latency.

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IFN-γ pretreatment inhibits growth of MCMV more effectively in BMMϕ than MEFs. (A and B) BMMϕ (A) or MEFs (B) were treated with or without IFN-γ for 48 h, infected at an MOI of 1, and cultured for the indicated times before freeze-thawing and plaque assay. Data shown is representative of two (BMMϕ) or three (MEF) independent experiments. Analysis of data in MEFs (B) revealed that the effects of IFN-γ were significant at 100 U/ml (48 h, P = 0.0231; 72 h, P = 0.02), and 1,000 U/ml (72 h, P = 0.0091), but that all other differences were insignificant (P > 0.05). (C) To determine the effect of MOI on IFN-γ treatment, BMMϕ or MEFs were treated with or without 100 U/ml IFN-γ for 48 h, infected at the MOIs shown, and cultured for 72 h before freeze-thawing and plaque assay (mean ± SEM from two independent experiments).
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Figure 1: IFN-γ pretreatment inhibits growth of MCMV more effectively in BMMϕ than MEFs. (A and B) BMMϕ (A) or MEFs (B) were treated with or without IFN-γ for 48 h, infected at an MOI of 1, and cultured for the indicated times before freeze-thawing and plaque assay. Data shown is representative of two (BMMϕ) or three (MEF) independent experiments. Analysis of data in MEFs (B) revealed that the effects of IFN-γ were significant at 100 U/ml (48 h, P = 0.0231; 72 h, P = 0.02), and 1,000 U/ml (72 h, P = 0.0091), but that all other differences were insignificant (P > 0.05). (C) To determine the effect of MOI on IFN-γ treatment, BMMϕ or MEFs were treated with or without 100 U/ml IFN-γ for 48 h, infected at the MOIs shown, and cultured for 72 h before freeze-thawing and plaque assay (mean ± SEM from two independent experiments).

Mentions: Pretreatment for 48 h with IFN-γ caused a dose-dependent inhibition of MCMV growth in both BMMϕ (Fig. 1 A) and MEFs (Fig. 1 B). Growth inhibition was more significant in BMMϕ than MEFs. BMMϕ showed a >100-fold decrease in viral titer by 72 h after infection (for example, at 100 U/ml of IFN-γ), whereas MEFs showed a maximum decrease of 5–10-fold even at 1,000 U/ml of IFN-γ. These results are consistent with other studies of IFN-γ effects on the growth of MCMV in fibroblasts 1113 and demonstrate an antiviral effect of IFN-γ in primary BMMϕ.


Novel cell type-specific antiviral mechanism of interferon gamma action in macrophages.

Presti RM, Popkin DL, Connick M, Paetzold S, Virgin HW - J. Exp. Med. (2001)

IFN-γ pretreatment inhibits growth of MCMV more effectively in BMMϕ than MEFs. (A and B) BMMϕ (A) or MEFs (B) were treated with or without IFN-γ for 48 h, infected at an MOI of 1, and cultured for the indicated times before freeze-thawing and plaque assay. Data shown is representative of two (BMMϕ) or three (MEF) independent experiments. Analysis of data in MEFs (B) revealed that the effects of IFN-γ were significant at 100 U/ml (48 h, P = 0.0231; 72 h, P = 0.02), and 1,000 U/ml (72 h, P = 0.0091), but that all other differences were insignificant (P > 0.05). (C) To determine the effect of MOI on IFN-γ treatment, BMMϕ or MEFs were treated with or without 100 U/ml IFN-γ for 48 h, infected at the MOIs shown, and cultured for 72 h before freeze-thawing and plaque assay (mean ± SEM from two independent experiments).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2195910&req=5

Figure 1: IFN-γ pretreatment inhibits growth of MCMV more effectively in BMMϕ than MEFs. (A and B) BMMϕ (A) or MEFs (B) were treated with or without IFN-γ for 48 h, infected at an MOI of 1, and cultured for the indicated times before freeze-thawing and plaque assay. Data shown is representative of two (BMMϕ) or three (MEF) independent experiments. Analysis of data in MEFs (B) revealed that the effects of IFN-γ were significant at 100 U/ml (48 h, P = 0.0231; 72 h, P = 0.02), and 1,000 U/ml (72 h, P = 0.0091), but that all other differences were insignificant (P > 0.05). (C) To determine the effect of MOI on IFN-γ treatment, BMMϕ or MEFs were treated with or without 100 U/ml IFN-γ for 48 h, infected at the MOIs shown, and cultured for 72 h before freeze-thawing and plaque assay (mean ± SEM from two independent experiments).
Mentions: Pretreatment for 48 h with IFN-γ caused a dose-dependent inhibition of MCMV growth in both BMMϕ (Fig. 1 A) and MEFs (Fig. 1 B). Growth inhibition was more significant in BMMϕ than MEFs. BMMϕ showed a >100-fold decrease in viral titer by 72 h after infection (for example, at 100 U/ml of IFN-γ), whereas MEFs showed a maximum decrease of 5–10-fold even at 1,000 U/ml of IFN-γ. These results are consistent with other studies of IFN-γ effects on the growth of MCMV in fibroblasts 1113 and demonstrate an antiviral effect of IFN-γ in primary BMMϕ.

Bottom Line: IFN-gamma inhibited MCMV replication in a signal transducer and activator of transcription (STAT)-1alpha-dependent manner much more effectively in BMMphi (approximately 100-fold) than MEF (5-10-fold).IFN-gamma inhibition of MCMV growth was independent of known mechanisms involving IFN-alpha/beta, tumor necrosis factor alpha, inducible nitric oxide synthase, protein kinase RNA activated (PKR), RNaseL, and Mx1, and did not involve IFN-gamma-induced soluble mediators.These studies in primary cells define a novel mechanism of IFN-gamma action restricted to Mphi, a cell type key for MCMV pathogenesis and latency.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Immunology and the Department of Molecular Microbiology, Washington University School of Medicine, St. Louis, Missouri 63110, USA.

ABSTRACT
Interferon (IFN)-gamma and macrophages (Mphi) play key roles in acute, persistent, and latent murine cytomegalovirus (MCMV) infection. IFN-gamma mechanisms were compared in embryonic fibroblasts (MEFs) and bone marrow Mphi (BMMphi). IFN-gamma inhibited MCMV replication in a signal transducer and activator of transcription (STAT)-1alpha-dependent manner much more effectively in BMMphi (approximately 100-fold) than MEF (5-10-fold). Although initial STAT-1alpha activation by IFN-gamma was equivalent in MEF and BMMphi, microarray analysis demonstrated that IFN-gamma regulates different sets of genes in BMMphi compared with MEFs. IFN-gamma inhibition of MCMV growth was independent of known mechanisms involving IFN-alpha/beta, tumor necrosis factor alpha, inducible nitric oxide synthase, protein kinase RNA activated (PKR), RNaseL, and Mx1, and did not involve IFN-gamma-induced soluble mediators. To characterize this novel mechanism, we identified the viral targets of IFN-gamma action, which differed in MEF and BMMphi. In BMMphi, IFN-gamma reduced immediate early 1 (IE1) mRNA during the first 3 h of infection, and significantly reduced IE1 protein expression for 96 h. Effects of IFN-gamma on IE1 protein expression were independent of RNaseL and PKR. In contrast, IFN-gamma had no significant effects on IE1 protein or mRNA expression in MEFs, but did decrease late gene mRNA expression. These studies in primary cells define a novel mechanism of IFN-gamma action restricted to Mphi, a cell type key for MCMV pathogenesis and latency.

Show MeSH
Related in: MedlinePlus