Title: Low expression of selenoprotein S may prevents excessive ossification of BMSCs through negative feedback between Sp7 and Hsp47
Abstract:
Selenoprotein S (SelS), a key component of the endoplasmic reticulum stress response, has been implicated in metabolic regulation, yet its role in osteogenesis remains poorly understood. This study aimed to elucidate the regulatory mechanisms by which low SelS expression modulates osteogenic differentiation and bone formation, with a focus on collagen synthesis and transcriptional feedback networks. Bone marrow mesenchymal stem cells (BMSCs) were transfected with lentivirus-delivered shRNA to establish a stable SelS knockdown model. Osteogenic differentiation was induced in vitro, and dynamic changes in osteogenic markers (Sp7, ALP) were quantified using qPCR and Western blotting (WB). Functional mineralization capacity was assessed via ALP staining (Day 7) and Alizarin Red S staining (Day 21). To validate in vivo relevance, mandibular bone formation in SelS knockout (KO) mice was analyzed by micro-CT (bone volume/total volume, BV/TV) and histomorphometry (H&E, Masson’s trichrome). Transcriptome sequencing of SelS-low BMSCs identified 2,268 differentially expressed genes (DEGs), which were further subjected to KEGG pathway enrichment analysis. Mechanistic studies included dual-luciferase reporter assays and chromatin immunoprecipitation (ChIP) to verify direct transcriptional regulation between Sp7 and Hsp47. SelS knockdown significantly enhanced osteogenic potential, with ALP activity increasing by 2.5-fold (p < 0.01) and mineralized nodule area rising by 40% (p < 0.001) compared to controls. In SelS KO mice, micro-CT revealed a 25% increase in mandibular BV/TV (p < 0.05), corroborated by histology showing denser collagen fibers. Transcriptomics highlighted DEG enrichment in extracellular matrix organization (e.g., Col1a1, Hsp47) and Wnt signaling. Crucially, Sp7 was shown to directly bind the Hsp47 promoter (ChIP-seq fold enrichment = 8.2, p < 0.001), forming a negative feedback loop that attenuated excessive mineralization. Our findings establish SelS as a novel negative regulator of osteogenesis. Its downregulation potentiates bone formation through Sp7-Hsp47-mediated collagen synthesis, while the feedback mechanism ensures balanced ossification. This dual regulatory axis positions SelS as a promising therapeutic target for bone regeneration disorders, including osteoporosis and fracture nonunion.
