Mechanical stretching promotes the differentiation of BMSC into pelvic floor ligament fibroblasts

Abstract

Pelvic floor dysfunction (PFD) is a prevalent and debilitating condition in aging women that is associated with weakened pelvic connective tissue. Fibroblasts are the main component of pelvic connective tissue and play a vital role in the maintenance of the pelvic organ. Mechanical stretching is a known modulator leading to the proliferation and differentiation of bone marrow mesenchymal stem cells (BMSCs). In this paper, we measured the lysyl oxidase (LOX), elastin and KDM4B expression in cultured BMSCs under different cyclic mechanical stretch (CMS) conditions via Western blot assays. Our findings indicated that the expression of these proteins was significantly higher in the CMS group than in the normal control group, and 10% CMS at a 0.5 Hz frequency had the best stimulatory effect. We further evaluated the influence of KDM4B on LOX and elastin expression via knockdown and overexpression of KDM4B in BMSCs. Chromatin immunoprecipitation (ChIP)-PCR analysis was performed to determine how H3K9me3 and KDM4B affect the regulatory regions of LOX and elastin in BMSCs subjected to 10% CMS combined with KDM4B knockdown. Finally, we verified the expression of KDM4B under CMS in vivo by subcutaneously transplanting KDM4B- overexpressing BMSCs into mice via immunofluorescence assays. The results showed that activating KDM4B by mechanical stretching increased the protein and gene expression levels of LOX and elastin and induced more LOX-positive cells in the BMSCs. These data suggest that CMS contributes to the differentiation of mesenchymal stem cells (MSCs) into fibroblasts, which indicates its potential use as a cell-based therapy for PFD.