Asxl2-Deficiency Impairs Pdlscs Osteogenesis Via Epigenetic Reprogramming

This study clarifies the epigenetic role of ASXL2 in the osteogenic differentiation of human periodontal ligament stem cells (hPDLSCs) to resolve critical gaps in craniofacial regeneration mechanisms. hPDLSCs were isolated with approval from the Medical Ethics Committee (Approval No. TMUhMEC2018101) and expanded to passages 3-4 for experimentation. ASXL2 knockdown was achieved using lentivirus-mediated shRNA transduction. Cellular proliferation was quantified via CCK-8 assay, while apoptosis rates were determined by Annexin V/PI dual staining. Osteogenic differentiation potential was evaluated by alkaline phosphatase (ALP) activity quantification and Alizarin Red S staining for mineralized matrix deposition. Histone modifications were assessed by Western blotting. Transcriptome-wide analysis (RNA sequencing) combined with molecular validation (RT-qPCR/Western blotting) confirmed the downregulation of key osteogenic markers, including RUNX2, ALP, COL1A1, and osteocalcin (OCN) etc. ASXL2-deficient hPDLSCs exhibited enhanced proliferation rates (p < 0.05) and reduced apoptotic activity (p<0.05), while severely compromising osteogenic differentiation capacity, as demonstrated by decreased ALP activity, mineralized nodules, and western blotting results of osteogenic markers. The transcriptome-wide analysis identified comprehensive dysregulation of osteogenic transcriptional networks. Mechanistically, ASXL2 loss enriched repressive H2AK119ub/H3K27me3 and reduced activating H3K4me3 at osteogenic loci. ASXL2 influences osteogenic lineage commitment via histone modification reprogramming, positioning it as a therapeutic target for craniofacial regeneration. This study redefines epigenetic strategies for tissue engineering.