UNLABELLED: Single-cell RNA sequencing (scRNA-seq) technology hold significant potential for advancing orthopedic research. This review examines the impact of ScRNA-seq on the future development of orthopedic research and practice. In the study of osteoarthritis, scRNA-seq can finely characterize the changes in the subsets of chondrocytes and their role in disease progression. In rheumatoid arthritis, this technique reveals the complex heterogeneity and cell-to-cell interactions between fibroblasts and immune cells. ScRNA-seq offers insights into the heterogeneity of nucleus pulposus, annulus fibrosus, and endplate cells, providing a novel perspective on the pathological mechanisms of intervertebral disc degeneration. Single-cell analysis in osteosarcoma research has uncovered the complexity of the tumor microenvironment and mechanisms of immunosuppression. Through these studies, scRNA-seq enhances insights into disease pathogenesis and offers innoviate approaches for precision medicine and personalized treatment strategies. THE TRANSLATIONAL POTENTIAL OF THIS ARTICLE: This article systematically reviews the cellular heterogeneity, molecular mechanisms and immune microenvironment of orthopedic diseases (such as osteoarthritis, rheumatoid arthritis, intervertebral disc degeneration, osteosarcoma) by single-cell RNA sequencing (scRNA-seq), which provides a theoretical basis for accurate diagnosis, new therapeutic target discovery (such as TRPV1, CXCR4) and individualized treatment strategies. The combination of multi-omics and spatial transcriptome technology is expected to accelerate clinical translation and optimize the diagnosis and treatment system of orthopedic diseases.