F. Javier Alvarez-leefmans (Committee Member), Nancy Bigley (Advisor), Barbara Hull (Committee Member)
Master of Science (MS)
Macrophages play an important role in the immune system, particularly in neutralizing pathogens via phagocytosis and the production of multiple cytokines and chemokines that control infection after exposure to specific stimuli. Macrophages exhibit two different phenotypes, M1 and M2. This study evaluated the role of Suppressors of cytokine signaling (SOCS)1 and SOCS3 on Herpes Simplex Virus (HSV-1) infection of polarized macrophages, cell viability, cell morphology, and the expression of cell surface CD markers. I hypothesized that J774A.1 murine macrophages cells in the naive state (M0), and M1 and M2 phenotypes would display differences in CD markers CD80,CD163, and CD200R, cell morphology, and viability following HSV-1infection and that the SOCS3 peptide mimetic would reduce the cytotoxicity associated with the M1 phenotype or HSV-1 infection. Unpolarized J774A.1 macrophages (M0) or polarized M1 and M2 phenotypes displayed differences in CD surface proteins at 18, 24, and 48 hours following HSV-1 infection and there was ~22% decrease in viability in M1 macrophages regardless of whether the macrophages were uninfected or infected with HSV-1. M2 macrophages demonstrated ~10% reductions in viability compared with infected M0 macrophage. Differences in cell morphology were noticed in uninfected polarized and unpolarized cells. M1 macrophages displayed irregular shapes and intracellular vacuoles were more pronounced at 24 and 48 hours compared to 18 hours. M0 appeared rounded or elongated, and M2 macrophages induced by IL-4, IL-10, or IL-13 showed elongation with the majority of cells appearing rounded. HSV-1-infected M0, M1 or M2 macrophages all became rounded and aggregated following HSV-1 infection. We found CD80 expression (by flow cytometry) was slightly deceased on uninfected M1 macrophages and M2 macrophages compared to uninfected M0 macrophages and infected M1 macrophages and IL-10-induced M2 macrophages also exhibited a minor increase in CD80. A reduction of CD163 expression occurred on M1 macrophages uninfected or infected with HSV-1. The level of CD163 on infected M2 macrophages was increased compared to uninfected M2 macrophage cells with the level being higher on infected IL-10-induced M2 macrophages. These observations were not statistically significant; however, the level of CD200R significantly increased on infected M2 macrophages polarized by IL-13 compared to CD200R expression in virus-infected M0 macrophages. Treatment with either SOCS3 peptide mimetic or pJAK2 increased the viability of polarized J774A.1 M1 macrophages. Production of pro-inflammatory cytokines TNF-a and IL-6 were reduced in polarized M1 macrophage treated with SOCS3 or pJAK2 peptide mimetic compared with polarized M1 untreated cells. Also, the SOCS3 peptide mimetic increased cell viability in M1 and HSV-1-infected M1 RAW264.7 and J774A.1 macrophages, but this increase was only significant in the RAW264.7 cells. This result suggests that SOCS3 mimetic exerts an anti-inflammatory effect by diminishing the lytic effect of M1 polarization. M1 cells treated with SOCS3 behave similar to the M2 phenotype and produce the anti-inflammatory cytokine IL-10. This study highlights the events that lead to M1 versus M2 polarization and the knowledge gained regarding SOCS3's ability to reduce inflammation may make it a potential therapeutic target for inflammatory diseases.
Department or Program
Microbiology and Immunology
Year Degree Awarded
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