Virginia Research Day 2021

Graduate Student Research Biomedical

Zhuang Wang 1 ; Bettina Heid 1 ; JingJing Ren 1 ; Michael Edward 1 ; Ran Lu 1 , Deena Khan 1 ; David G. Kirsch 3 ; Christopher M. Reilly 1,2 ; Rujuan Dai 1 ; Ansar Ahmed 1 Corresponding author: wzhuang@vt.edu 09 Depletion Of microRNA-183-96-182 Cluster in Lymphocytes Suppresses anti-dsDNA autoantibody Production and IgG Deposition In Kidney In C57BL/6-Fas lpr/lpr Mice 1 Department of Biomedical and Veterinary Medicine, VMCVM, Virginia Tech, 2 Via College of Osteopathic Medicine-Virginia Campus, 3 Department of Radiation Oncology, Duke University Medical Center

and observed a reduction of IFN γ in anti-CD3/anti- CD28 and PMA stimulated splenocytes in both miR-183C -/- and miR-182 -/- B6- lpr mice. Interestingly, we only observed a significant reduction of IFN γ in LPS activated splenocytes from miR-182 but not miR-183C knock out B6- lpr mice. To investigate the immune cell development and differentiation, we assessed the cell population through flow cytometry and the results showed that conditional depletion of miR-182 and miR-183C did not change lymphocyte composition but increases macrophage population in the spleen of B6- lpr mice. Importantly, the Foxo1 protein expression was increased in splenic CD4+ cells in miR-182 -/- and miR-183C -/- B6- lpr mice and further study is needed to understand the mechanism of the regulatory role of miR-183C on autoantibody and cytokines production. Together, our data suggest that miR-183C might be a new potential target for lupus therapy.

The microRNA-183-96-182 cluster (miR-183C) is a highly conserved miRNA cluster located at human chromosome 7 and murine chromosome 6. miR- 183C family is initially identified as a sensory organ- specific miRNA cluster and its expression is very low in spleen and other tissues in normal C57BL/6 (B6) mice. miR-182 is highly induced in activated T and B lymphocytes. In vitro study indicated that miR-182 is critical for IL-2 induced Th cell expansion and proliferation via targeting Foxo1. Surprisingly, depletion of miR-182 in vivo had no obvious effect on adaptive immune cell development and function in B6 mice. Nevertheless, recent studies with B6 mice deficiency in whole miR-183C indicated that miR-183C are critical for Th17 cell pathogenicity and macrophage function. These data strongly suggest a functional compensation of miR-183C family members in immunity. We have previously reported a significant upregulation of miR-183C in the splenic lymphocytes of different

murine lupus models. However, the pathogenic significance of miR-183C in lupus remains unknown. In this study, we found that deliberately inhibition of miR-182 alone or miR-183C with antagomir significantly reduces the expression of lupus-related inflammatory cytokines, such as IFN γ and IL-6, in in vitro stimulated splenocytes from MRL and MRL/ lpr mice. These data suggest a potential inflammation regulatory role of miR-182 and miR-183C in lupus. To better understand the role of miR-182 and miR-183C in lupus, we developed B6- lpr mice with conditional depletion of miR-183C or miR-182 alone in CD2- lymphocytes (miR-183C -/- B6- lpr and miR-182 -/- B6- lpr ). Impressively, we found that depletion of miR- 183C, but not miR-182, along significantly suppresses the development of anti-dsDNA autoantibody in B6- lpr mice. Meanwhile, the IgG deposition in kidney was identified to be reduced in miR-183C and miR- 182 knockout mice and a trend of serum total IgG reduction. Then, we analyzed the cytokine production

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