Differential responses of two porcine macrophage- like cell lines to infection with Rift Valley fever virus MP-12

Authors: Smolensky, Dmitriy; Wilson, William; Cox, Victoria; Schirtzinger, Erin; Chitko-Mckown, Carol; Fawver, Zachary; Noronha, Leela

Submitted to: International Veterinary Immunology Symposium
Publication Type: Proceedings
Publication Acceptance Date: 7/11/2019
Publication Date: 8/30/2019
Citation: Smolensky, D., Wilson, W.C., Cox, V.G., Schirtzinger, E.E., Chitko-Mckown, C.G., Fawver, Z.T., Noronha, L.E. 2019. Differential responses of two porcine macrophage- like cell lines to infection with Rift Valley fever virus MP-12. Proceedings of International Veterinary Immunology Symposium. August 13-16, 2019, Seattle, Washington. P072:126-127. Available online at https://ivis2019.org/wp-content/uploads/2019/09/IVIS-2019-Abstract-book.pdf

Interpretive Summary: Rift Valley fever virus (RVFV) is a zoonotic pathogen that primarily affects domestic ruminants, but can also cause severe disease and death in humans. Symptomatic disease from RVFV infection has not been observed in domestic pigs (Sus scrofa); however, two serosurveys have detected circulating antibodies to RVFV among wild and domestic members of the pig family (Suidae). Experimentally, RVFV has been shown to replicate in some porcine cells in vitro, and induce viremia in a fraction of pigs inoculated in vivo. It is unclear how once infected, pigs are apparently able to avert clinical disease. To perform initial mechanistic investigations of porcine innate immune cell responses to infection with RVFV, we used two porcine monocyte-derived cell lines, C'2+ and C'2-, which have previously been demonstrated to exhibit phenotypic characteristics consistent with macrophages, with C'2- cells thought to be less differentiated than C'2+ cells. Following 12, 24, and 48-hour infections with MP-12, an attenuated form of RVFV, at 0.01, 0.1, and 1.0 MOI, C'2+ cells demonstrated lower viability than C'2- cells at intermediate time/dose combinations as measured by MTS assay. Apoptosis was measured in both cell lines at 24 hours post-infection by flow cytometric analysis using Annexin V and propidium iodide labeling. Apoptosis was observed in C'2+, but not C'2- cells, at 0.1 and 1.0 MOI. Transcription of inflammatory mediators was measured by RT-qPCR and the observed patterns differed between the two cell lines. IL-12 and IFNß expression was upregulated in C'2+ cells, but not C'2- cells. Expression of IL-1a increased immediately following a 1 hour virus adsorption in C'2+ cells, but only after 12 hours post-adsorption in C'2- cells. This work serves as a proof-of-concept starting point to warrant additional studies on the dynamics of RVFV infection in suids, including whether monocyte-lineage differentiation states may affect outcomes in infected cells.

Technical Abstract: Rift Valley fever virus (RVFV) is a zoonotic pathogen that primarily affects domestic ruminants, but can also cause severe disease and death in humans. Symptomatic disease from RVFV infection has not been observed in domestic pigs (Sus scrofa); however, two serosurveys have detected circulating antibodies to RVFV among wild and domestic members of the pig family (Suidae). Experimentally, RVFV has been shown to replicate in some porcine cells in vitro, and induce viremia in a fraction of pigs inoculated in vivo. It is unclear how once infected, pigs are apparently able to avert clinical disease. To perform initial mechanistic investigations of porcine innate immune cell responses to infection with RVFV, we used two porcine monocyte-derived cell lines, C'2+ and C'2-, which have previously been demonstrated to exhibit phenotypic characteristics consistent with macrophages, with C'2- cells thought to be less differentiated than C'2+ cells. Following 12, 24, and 48-hour infections with MP-12, an attenuated form of RVFV, at 0.01, 0.1, and 1.0 MOI, C'2+ cells demonstrated lower viability than C'2- cells at intermediate time/dose combinations as measured by MTS assay. Apoptosis was measured in both cell lines at 24 hours post-infection by flow cytometric analysis using Annexin V and propidium iodide labeling. Apoptosis was observed in C'2+, but not C'2- cells, at 0.1 and 1.0 MOI. Transcription of inflammatory mediators was measured by RT-qPCR and the observed patterns differed between the two cell lines. IL-12 and IFNß expression was upregulated in C'2+ cells, but not C'2- cells. Expression of IL-1a increased immediately following a 1 hour virus adsorption in C'2+ cells, but only after 12 hours post-adsorption in C'2- cells. This work serves as a proof-of-concept starting point to warrant additional studies on the dynamics of RVFV infection in suids, including whether monocyte-lineage differentiation states may affect outcomes in infected cells.