Exploring the Unique Extracellular Matrix Composition of <i>Acomys</i> as a Potential Key to Resisting Fibrosis

Fibrosis is a dysregulated wound healing response characterized by excessive accumulation of dense scar tissue that inhibits organ function and is estimated to contribute to up to 45% of deaths in the industrialized world. In this work, we sought to uncover new ways to address fibrosis by drawing inspiration from an animal that does not develop fibrosis. The Spiny Mouse (<i>Acomys</i>) has the most extensive regenerative capabilities of any known mammal and can regenerate injuries to the skin, kidney, heart, skeletal muscle, and spine with little to no fibrosis. We hypothesize that the regenerative abilities of <i>Acomys</i> are due, in part, to altered stiffness-mediated fibroblast-to-myofibroblast transition (FMT). In this work, we interrogated stiffness-mediated FMT in <i>Acomys</i> and <i>Mus</i> dermal fibroblasts <i>in vitro</i> by performing RNA Sequencing and found no differential gene expression in <i>Acomys</i> fibroblasts cultured on soft vs stiff substrates. We further investigated the direct impact of stiffness-mediated FMT and species differences on ECM deposition by fabricating cell-derived matrices (CDMs) from <i>Acomys</i> and <i>Mus</i> fibroblasts cultured on varying stiffnesses. After assessing the composition of these CDMs using label-free quantitative proteomics, fibrosis-associated extracellular matrix proteins including fibrillin-1, ADAMTS1, SPARC, and galectin-1 were found to be significantly reduced or absent in <i>Acomys</i> CDMs compared to <i>Mus</i> CDMs. In addition, proteins that have been connected to fibrosis resolution, including Col12a1 and clusterin, were upregulated in <i>Acomys</i> CDMs. When cultured on <i>Acomys</i> CDMs, mouse macrophages downregulated MMP9 mRNA expression and maintained increased expression of iNOS in response to IL-4, a pro-fibrotic cytokine. These results indicate a direct impact of species-specific ECM compositions on macrophage phenotype and suggest that ECM produced by <i>Acomys</i> fibroblasts may impede the development of a pro-fibrotic macrophage phenotype in the presence of pro-fibrotic stimuli.