AUTHOR=Vahkal Brett , Yegorov Sergey , Onyilagha Chukwunonso , Donner Jacqueline , Reddick Dean , Shrivastav Anuraag , Uzonna Jude , Good Sara V. 

TITLE=Immune System Effects of Insulin-Like Peptide 5 in a Mouse Model

JOURNAL=Frontiers in Endocrinology

VOLUME=11

YEAR=2021

URL=https://www.frontiersin.org/journals/endocrinology/articles/10.3389/fendo.2020.610672

DOI=10.3389/fendo.2020.610672

ISSN=1664-2392

ABSTRACT=<sec><title>Introduction</title><p>Insulin-like peptide 5 (INSL5) is a peptide hormone with proposed actions in glucose homeostasis and appetite regulation <italic>via</italic> its cognate receptor, relaxin family peptide receptor 4 (RXFP4). Here, we look for evidence for their involvement in the immune system using a mouse model.</p></sec><sec><title>Methods</title><p><italic>In silico analyses:</italic> we queried public databases for evidence of expression of INSL5-RXFP4 in immune system tissues/cells (NCBI’s SRA and GeoProfiles) and disorders (EMBO-EBI) and performed phylogenetic footprinting to look for evidence that they are regulated by immune-associated transcription factors (TFs). <italic>Experimental analyses:</italic> We characterized the expression and correlation of INSL5/RXFP4 and other immune system markers in central and peripheral immune organs from C57/bl6 mice in seven cohorts. We tested whether fluctuations in circulating INSL5 induce an immune response, by injecting mice with 30 μg/kg of INSL5 peptide in the peritoneum, and examining levels of immune markers and metabolic peptides in plasma. Lastly, we quantified the expression of <italic>Rxfp4</italic> in T-cells, dendritic cells and cell lines derived from human and mouse and tested the hypothesis that co-incubation of ANA-1 cells in INSL5 and LPS alters cytokine expression.</p></sec><sec><title>Results</title><p>We find <italic>Insl5</italic> expression only in thymus (in addition to colon) where its expression was highly correlated with <italic>Il-7</italic>, a marker of thymocyte development. This result is consistent with our <italic>in silico</italic> findings that <italic>Insl5</italic> is highly expressed in thymic DP, DN thymocytes and cortical TEC’s, and with evidence that it is regulated by thymocyte-associated TF’s. We find <italic>Rxfp4</italic> expression in all immune organs, and moderately high levels in DCs, particularly splenic DCs, and evidence that it is regulated by immune-associated TF’s, such as STAT’s and GATA. <italic>Systemic effects</italic>: We observed significantly elevated concentrations of blood GLP-1, GIP, GCG and PYY following intraperitoneal injection of INSL5, and significantly altered expression of cytokines IL-5, IL-7, M-CSF, IL-15, IL-27 and MIP-2. <italic>Immune cell effects</italic>: Incubation of ANA-1 cells with INSL5 impeded cell growth and led to a transient elevation of IL-15 and sustained reduction in IL-1β, IL-6 and TNFα.</p></sec><sec><title>Conclusion</title><p>We propose that INSL5-RXFP4 play a novel role in both central and peripheral immune cell signaling.</p></sec>