Abstract
Human bone marrow mesenchymal stromal cells (hBMSCs) are multipotent stromal cells capable of osteogenic differentiation, making them a promising cell source for bone tissue engineering and regenerative medicine. Identifying key factors that regulate hBMSCs osteogenic differentiation is crucial for enhancing bone regeneration strategies. This study aims to identify target genes underlying impaired osteogenic differentiation of hBMSCs under oxidative stress (OS) through integrated transcriptomic and proteomic approaches, and delineate the role of proenkephalin (PENK) in this process. OS model and impaired osteogenic differentiation model were established in hBMSCs using hydrogen peroxide (H2O2). Cellular oxidative stress levels were assessed using Dihydroethidium (DHE) fluorescent probes and JC-1 staining. Osteogenic differentiation was evaluated by alkaline phosphatase (ALP) activity and Alizarin Red staining (ARS). Key genes and proteins were predicted via integrated transcriptomic and proteomic analyses. The role of PENK in osteogenic differentiation was validated using lentiviral transfection. This study established 400 µM H2O2 as the optimal concentration for inducing impaired osteogenic differentiation in hBMSCs. Integrated transcriptomic and proteomic analysis identified 18 pivotal regulatory genes that orchestrate impaired osteogenic differentiation of hBMSCs under OS. Among these, PENK was identified as a potential therapeutic target involved in regulating oxidative stress-impaired osteogenic differentiation of hBMSCs. Functional validation confirmed that PENK overexpression promotes osteogenic differentiation in hBMSCs. OS contributes to impaired osteogenic differentiation in hBMSCs. PENK regulates osteogenic differentiation of hBMSCs under OS and holds promise as a novel therapeutic target for bone regeneration and repair.
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Data availability
The transcriptomics dataset has been deposited in the NCBI GEO repository under accession number GSE318199. The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE37 partner repository with the dataset identifier PXD067569. All data generated or analyzed during this study are included in this published article. All data and reagents are available from the corresponding author upon reasonable request.
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Acknowledgements
We thank the Clinical Medicine Research Center of the Affiliated Hospital of Guizhou Medical University for providing the experimental facilities and equipment. We also thank LetPub (www.letpub.com.cn) for its linguistic assistance during the preparation of this manuscript.
Funding
This research was supported by the National Natural Science Foundation of China (Grant No. 82460426), the Guizhou Provincial Natural Science Foundation (Grant No. Qiankehe basis-ZK [2023] general 345, Qiankehe Basic-[2024] youth 263, Qiankehe Basic-[2024] youth 251), the Wu Jieping Medical Foundation (320.6750.2023-02-11), and the Start-up Fund for Doctoral Research of the Affiliated Hospital of Guizhou Medical University (Grant No. gyfybsky-2024-23). Support was also received from the Key Advantageous Discipline Construction Project of Guizhou Provincial Health Commission in 2023 in Emergency Department.
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Wentao Dong designed the study and drafted and revised the manuscript; Yangyang Zheng performed the experiments and revised the manuscript; Yongfang Zhou and Tao Wang provided technical assistance; Jiexin Yang and Huaying Li performed the experiments; Fanchao Li, Chuan Wang, and Liang Liang assisted with the experimental procedures; Hao Li and Jian Zhang performed the experiments and collected and analyzed data; and Wuxun Peng designed and supervised the study.
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Dong, W., Zheng, Y., Zhou, Y. et al. Multi-omics identification of key targets for the osteogenic differentiation of human bone marrow mesenchymal stromal cells under oxidative stress. Sci Rep (2026). https://doi.org/10.1038/s41598-026-39818-4
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DOI: https://doi.org/10.1038/s41598-026-39818-4
