Identification of Key Candidate Genes Related to Cartilage development during Murine embryonic limb development by single cell RNA-sequencing and Experimental Confirmation

Background Cartilage, characterized by its limited self-repair capacity due to avascularity and low metabolic activity of chondrocytes, poses a significant challenge for regenerative medicine. Osteoarthritis (OA), a prevalent cartilage disorder, highlights the urgent need for effective cartilage regenerative therapies. Understanding the molecular mechanisms underlying cartilage development during embryonic stages is crucial for advancing regenerative strategies and identifying potential therapeutic targets. Methods This study employed single-cell RNA sequencing (scRNA-seq) to explore the transcriptional landscape of mouse embryonic limb development at various stages, focusing on identifying genes pivotal for cartilage differentiation. Differentially expressed genes (DEGs) specific to cartilage development were pinpointed through comparative analysis. Functional validation of these marker genes was conducted using immunohistochemistry and RT-qPCR to confirm their roles in chondrocyte maturation and differentiation. Results Our scRNA-seq analysis identified a set of novel marker genes, including Bgn, Ucma, Fmod, Msmp, and 1500015O10Rik, as specific indicators of cartilage development. Functional experiments supported the crucial roles of these markers in the differentiation and maturation of chondrocytes. Additionally, our findings revealed the dynamic transcriptomic alterations during cartilage development, emphasizing the significance of specific regulatory factors in guiding mesenchymal stem cells towards chondrogenesis. Conclusions The study elucidates the complex transcriptomic landscape governing cartilage development in embryonic mice, highlighting the discovery of novel marker genes crucial for chondrocyte differentiation. These insights into the molecular mechanisms of cartilage formation lay the groundwork for developing targeted regenerative therapies for OA and related musculoskeletal disorders. Our research underscores the importance of identifying reliable regulatory factors that enhance the effectiveness of regenerative treatments, providing a valuable reference for future studies on cartilage repair and regeneration.