Mare stromal endometrial cells differentially modulate inflammation depending on oestrus cycle status: anstudy.

Abstract

The modulation of inflammation is pivotal for uterine homeostasis. Here we evaluated the effect of the oestrus cycle on the expression of pro-inflammatory and anti-inflammatory markers in a cellular model of induced fibrosis. Mare endometrial stromal cells isolated from follicular or mid-luteal phase were primed with 10&#x2009;ng/mL of TGF&#x3b2; alone or in combination with either IL1&#x3b2;, IL6, or TNF&#x3b1; (10&#x2009;ng/mL each) or all together for 24&#x2009;h. Control cells were not primed. Messenger and miRNA expression were analyzed using real-time quantitative PCR (RT-qPCR). Cells in the follicular phase primed with pro-inflammatory cytokines showed higher expression of collagen-related genes (, and) and mesenchymal marker (, and) genes;&#x2009;<&#x2009;0.05. Cells primed during the mid-luteal overexpressed genes associated with extracellular matrix, processing, and prostaglandin E synthase (, and;&#x2009;<&#x2009;0.05). There was a notable upregulation of pro-fibrotic miRNAs (miR17, miR21, and miR433) in the follicular phase when the cells were exposed to TGF&#x3b2; + IL1&#x3b2;, TGF&#x3b2; + IL6 or TGF&#x3b2; + IL1&#x3b2;&#x2009;+&#x2009;IL6&#x2009;+&#x2009;TNF&#x3b1;. Conversely, in cells from the mid-luteal phase, the treatments either did not or diminished the expression of the same miRNAs. On the contrary, the anti-fibrotic miRNAs (miR26a, miR29b, miR29c, miR145, miR378, and mir488) were not upregulated with treatments in the follicular phase. Rather, they were overexpressed in cells from the mid-luteal phase, with the highest regulation observed in TGF&#x3b2; + IL1&#x3b2;&#x2009;+&#x2009;IL6&#x2009;+&#x2009;TNF&#x3b1; treatment groups. These miRNAs were also analyzed in the extracellular vesicles secreted by the cells. A similar trend as seen with cellular miRNAs was noted, where anti-fibrotic miRNAs were downregulated in the follicular phase, while notably elevated pro-fibrotic miRNAs were observed in extracellular vesicles originating from the follicular phase. Pro-inflammatory cytokines may amplify the TGF&#x3b2; signal in the follicular phase resulting in significant upregulation of extracellular matrix-related genes, an imbalance in the metalloproteinases, downregulation of estrogen receptors, and upregulation of pro-fibrotic factors. Conversely, in the luteal phase, there is a protective role mediated primarily through an increase in anti-fibrotic miRNAs, a decrease in SMAD2 phosphorylation, and reduced expression of fibrosis-related genes.