Advances in spatial transcriptomics and its application in the musculoskeletal system

Abstract

While bulk RNA sequencing and single-cell RNA sequencing have shed light on cellular heterogeneity and potential molecular mechanisms in the musculoskeletal system in both physiological and various pathological states, the spatial localization of cells and molecules and intercellular interactions within the tissue context require further elucidation. Spatial transcriptomics has revolutionized biological research by simultaneously capturing gene expression profiles and in situ spatial information of tissues, gradually finding applications in musculoskeletal research. This review provides a summary of recent advances in spatial transcriptomics and its application to the musculoskeletal system. The classification and characteristics of data acquisition techniques in spatial transcriptomics are briefly outlined, with an emphasis on widely-adopted representative technologies and the latest technological breakthroughs, accompanied by a concise workflow for incorporating spatial transcriptomics into musculoskeletal system research. The role of spatial transcriptomics in revealing physiological mechanisms of the musculoskeletal system, particularly during developmental processes, is thoroughly summarized. Furthermore, recent discoveries and achievements of this emerging omics tool in addressing inflammatory, traumatic, degenerative, and tumorous diseases of the musculoskeletal system are compiled. Finally, challenges and potential future directions for spatial transcriptomics, both as a field and in its applications in the musculoskeletal system, are discussed.

Fig. 1

A Concise Workflow for Integrating ST into Musculoskeletal System Research and Overview of the Main ST Technologies. The figure highlights key considerations for conducting ST research in the musculoskeletal system, such as sample types, transcriptome coverage, and species compatibility, in order to select between imaging-based or sequencing-based technologies. It also illustrates the main ST technologies, which researchers can choose based on different technical parameters and commercial availability. However, it is important to acknowledge that the choice of technology is a complex process, and this workflow may not be universally applicable to all biological questions. Researchers should carefully weigh their options when making a technological choice. Furthermore, it should be noted that some probe-based sequencing platforms, such as GeoMx DSP, Visium V2, and Visium HD, are limited to near-whole transcriptome detection in humans and mice only. cDNA complementary DNA. Created in BioRender. Wang, H. (2025) https://BioRender.com/e71m592

Fig. 2

Application of ST in Physiological Mechanisms and Inflammatory Diseases of the Musculoskeletal System. a At different time-points of human embryonic limb development, scRNA-seq and ST have been used to construct a spatiotemporal transcriptomic map. UMAP visualization of 125 955 human embryonic limb cells. In human posterior limb tissue slices at PCW6.2, spatial heatmaps of specific cell types and corresponding marker genes. Copyright © 2023, The Author(s). b Multiple canonical markers detected in the murine NP region, but Tie2 only in the CEP or AF region. © 2024 The Authors. Advanced Science published by Wiley-VCH GmbH. c H&E-stained sections of adult murine femur and heatmaps of the number of unique genes (nFeature) or unique transcripts (nCount) detected at each capture site. Copyright © 2023, The Author(s). d Morphological annotation, spatial clustering, and spatial expression pattern of FN1 of a serum-positive RA patient sample. Copyright © 2022, The Author(s). P proximal, M middle, D distal, PCW post-conception week, infiltrates leukocyte infiltration sites

Fig. 3

Application of ST in Traumatic Diseases of the Musculoskeletal System. a Spatiotemporal expression pattern of Meg3, Gm10076, Rpph1, miR − 206 − 3p and miR − 1a − 3p at different time-points before and after injury during the regeneration process of murine anterior tibial muscle. Copyright © 2022, The Author(s). b In vivo micro-CT imaging is used for micro-finite element analysis to create 3D landscapes of the mechanical environment at the tissue scale. ST analysis of explanted femurs was performed to construct 2D spatial transcriptome landscapes. Finally, gene spatial expression patterns under different mechanical strains were constructed through visual alignment. Copyright © 2025, The American Association for the Advancement of Science. c Deconvolution of Visium capture spots from different regions of the umbilical cord was performed to visualize cell type proportions. The color coding represents different cell types, while pie charts illustrate the proportion of each cell type at each spot. The proportion of spots in the fetal segment of the umbilical cord, where functional MSCs comprise over 80% of the cells, was higher compared to the maternal segment.© 2022 The Authors. Advanced Science published by Wiley-VCH GmbH. dpi day post-injury, Huc human umbilical cord

Fig. 4

Challenges and Prospects. Challenges and prospects faced by spatial transcriptomics itself and its application to the musculoskeletal system. AI Artificial intelligence, FFPE Formalin fixed and paraffin embedded, FF Fresh frozen. Created in BioRender. Wang, H. (2025) https://BioRender.com/e71m592