{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,26]],"date-time":"2026-08-26T04:23:31Z","timestamp":1787718211035,"version":"build-2784847793"},"reference-count":45,"publisher":"MDPI AG","issue":"9","license":[{"start":{"date-parts":[[2026,8,23]],"date-time":"2026-08-23T00:00:00Z","timestamp":1787443200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100014188","name":"Ministry of Science and ICT","doi-asserted-by":"publisher","award":["RS-2024-00334810"],"award-info":[{"award-number":["RS-2024-00334810"]}],"id":[{"id":"10.13039\/501100014188","id-type":"DOI","asserted-by":"publisher"}]},{"award":["RS-2024-00334810"],"award-info":[{"award-number":["RS-2024-00334810"]}],"id":[{"id":"https:\/\/ror.org\/01wpjm123","id-type":"ROR","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100003052","name":"Ministry of Trade, Industry and Energy","doi-asserted-by":"publisher","award":["20023762"],"award-info":[{"award-number":["20023762"]}],"id":[{"id":"10.13039\/501100003052","id-type":"DOI","asserted-by":"publisher"}]},{"award":["20023762"],"award-info":[{"award-number":["20023762"]}],"id":[{"id":"https:\/\/ror.org\/008nkqk13","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Bioengineering"],"abstract":"<jats:p>Cryopreserved mesenchymal stem cells (MSCs) are widely used as off-the-shelf therapeutics. However, conventional manufacturing workflows typically include a post-thaw 2D monolayer recovery step before 3D spheroid formation. This study evaluated whether immediate spheroid formation following thawing (Direct-3D) could replace the conventional 2D-to-3D workflow while maintaining or improving chondrogenic potential. Using passage 5 Wharton\u2019s jelly MSCs (WJ-MSCs), spheroid morphology, early chondrogenic priming, differentiation, and extracellular matrix (ECM) formation were systematically compared between the two strategies. The Direct-3D group exhibited faster and more uniform spheroid compaction and significantly increased SOX9 expression as early as 3 h, indicating enhanced early priming. This response was accompanied by higher COL2A1 expression and an increased COL2A1\/COL1A1 ratio during early differentiation while maintaining minimal collagen type X expression. These findings demonstrate that the post-thaw monolayer recovery step is not essential for producing high-quality chondrogenic spheroids from WJ-MSCs. By simplifying the manufacturing workflow while preserving\u2014and potentially enhancing\u2014early chondrogenic performance, the Direct-3D strategy represents a practical and scalable approach for WJ-MSC-based cartilage tissue engineering and cell therapy.<\/jats:p>","DOI":"10.3390\/bioengineering13090964","type":"journal-article","created":{"date-parts":[[2026,8,24]],"date-time":"2026-08-24T14:49:34Z","timestamp":1787582974000},"page":"964","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["An Efficient Spheroid Manufacturing Workflow for WJ-MSCs: Comparative Evaluation of Immediate 3D Aggregation and Conventional 2D Stabilization"],"prefix":"10.3390","volume":"13","author":[{"ORCID":"https:\/\/orcid.org\/0009-0009-5732-5838","authenticated-orcid":false,"given":"Minji","family":"Ha","sequence":"first","affiliation":[{"name":"Department of Health Sciences and Technology, SAIHST, Sungkyunkwan University, Seoul 06355, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6530-795X","authenticated-orcid":false,"given":"Joon Ho","family":"Wang","sequence":"additional","affiliation":[{"name":"Department of Health Sciences and Technology, SAIHST, Sungkyunkwan University, Seoul 06355, Republic of Korea"},{"name":"Department of Medical Device Management and Research, SAIHST, Sungkyunkwan University, Seoul 06351, Republic of Korea"},{"name":"Department of Orthopaedic Surgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul 06351, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,8,23]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"315","DOI":"10.1080\/14653240600855905","article-title":"Minimal Criteria for Defining Multipotent Mesenchymal Stromal Cells. 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