The Identification of the MCHR-1 in J774A.1 Macrophages and The Effect of Melanin-concentrating Hormone on a 3T3-L1 Adipocyte-J774A.1 Macrophage Co-Culture

Abstract

Melanin-concentrating hormone (MCH) stimulates appetite in higher-order mammals, and our lab has demonstrated previously that MCH signaling by developing 3T3-L1 preadipocytes elicits changes in gene expression of several immune regulatory genes (Cook, 2021). I hypothesize that signaling between immune cells and adipose tissue contributes to the negative health outcomes of the obese phenotype. Co-culturing of two or more cell types together provides an opportunity to study cells in a setting that better reflects intercellular communication between cells in a tissue, therefore I planned to establish a co-culture model system consisting of adipocytes and macrophages in our laboratory. First, the J774A.1 macrophage line was acquired. Eventually, successful macrophage culturing technique was learned. Then, I asked whether theses J774A.1 macrophage expressed the Mchr1 gene and MCH receptor protein as well as whether this cell line responded at all to MCH. Finally, migration was quantified via fluorescein-labeled macrophages into a pre-adipocyte culture with and without MCH treatment in a co-culture. These results demonstrate that J774A.1 cells do respond to MCH, although there was trouble in detecting Mchr1 gene and MCHR1 protein expression in these cells. It was also demonstrated that MCH treated co-culture wells facilitate migration of macrophages into the preadipocyte cell layer in our co-culture model. A further experiment with differentiated adipocytes showed no MCH influence on macrophage migration. Future studies will aim to repeat and expand this model to study macrophage migration towards developing and differentiated adipocytes.