AccScience Publishing / IJB / Online First / DOI: 10.36922/IJB025240238
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RESEARCH ARTICLE

 3D-bioprinted osteocytes expressing Wnt7b protect osteoblast differentiation from microgravity

Jinling Zhang1† ,  Pengtao Wang1† ,  Xiaoling Chen1 ,  Saima Khan1 ,  Haiping Ouyang2 ,  Yangxi Liu2 ,  Bo He2 ,  Xian Li3 ,  Xing Liu2* ,  Xiaolin Tu1*
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1 Laboratory of Skeletal Development and Regeneration, Key Laboratory of Clinical Laboratory Diagnostics (Ministry of Education), College of Laboratory Medicine, Chongqing Medical University, Chongqing, China
2 Department of Orthopedics, Ministry of Education Key Laboratory of Child Development and Disorders, National Clinical Research Center for Child Health and Disorders, Children’s Hospital of Chongqing Medical University, Chongqing, China
3 College of Medical Informatics, Chongqing Medical University, Chongqing, China
†These authors contributed equally to this work.
IJB, 025240238 https://doi.org/10.36922/IJB025240238
Received: 12 June 2025 | Accepted: 3 July 2025 | Published online: 3 July 2025
© 2025 by the Author(s). This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution 4.0 International License ( https://creativecommons.org/licenses/by/4.0/ )
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Abstract

Maintaining bone formation in microgravity/weightless environments remains a major challenge. Under weightless conditions, osteocytes act as mechanosensors to inhibit Wnt canonical signaling and bone formation by secreting sclerostin. This study explores whether osteocytic Wnt7b can counteract microgravity-induced bone loss through Wnt non-canonical signaling. Unlike previous bioprinting studies that focused on structural scaffolds or generic cell types, a novel bioprinted scaffold consisting of polycaprolactone (supportive) and osteocyte (functional) hydrogels was constructed in this study. Osteocytes overexpressing Wnt7b were co-cultured with bone marrow stromal cells (ST2) in a 3D biomimetic weightless biomicroenvironmental system (3D-BWBM) to assess osteogenic and lipogenic differentiation. The results indicated that osteocytic Wnt7b enhanced osteogenic differentiation and mineralization of ST2 cells via the Wnt non-canonical pathway PKCδ, while suppressing the expression of lipogenic markers (Pparg, Cebpa) and adipogenesis. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) analysis revealed elevated expression of Sost and Mef2c, downregulation of the Wnt target gene Opg, and elevated expression of pro-osteoclastogenic cytokine Rankl and pro-inflammatory cytokines Tnfa and Il1b, thus validating the microgravity effect. Unlike conventional 2D culture of RCCS™ cells, the 3D hydrogels were printed with tunnels (500 μm) for efficient nutrient/ metabolite exchange, resulting in good cell growth, high cell viability (97%), and a six-fold increase in proliferative activity within 7 days. Wnt7b osteocytes were still able to maintain the osteogenic differentiation of ST2 cells, as evidenced by elevated alkaline phosphatase activity, mineralization (1.8-fold increase), and a decrease in osteoblast marker genes (Alpl, Runx2, Col1a1). In conclusion, Wnt7b-PCKδ signaling counteracts microgravity-induced bone loss, and further in vivo studies on osteocytic Wnt7b are warranted to confirm this causal relationship.

 

Graphical abstract
Keywords
3D bioprinting
Microgravity
Osteogenic differentiation
Wnt7b
Wnt noncanonical signaling
Funding
This work was supported by the National Natural Science Foundation of China (grant numbers: 82471909, 81672118, and 32101053) and the Chongqing Natural Science Foundation (grant numbers: CSTB2022NSCQ-LZX0048 and CSTB2023NSCQ-MSX0424).
Conflict of interest
The authors declare they have no competing interests.
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International Journal of Bioprinting, Electronic ISSN: 2424-8002 Print ISSN: 2424-7723, Published by AccScience Publishing
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