Isolation andfunctional characterization of HO-hMSCs as NK-supportive cells derived from hematopoietic organoids
[Objective] In in vitro systems for differentiating and expanding natural killer (NK) cells, feeder cells provide essential cell-cell contact and paracrine signals that drive precursor proliferation and terminal maturation. However, existing xenogeneic feeder cells or tumor-derived genetically modif...
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Institute of Blood Transfusion of Chinese Academy of Medical Sciences
2025-05-01
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| Series: | Zhongguo shuxue zazhi |
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| Online Access: | https://www.cjbt.cn/thesisDetails#10.13303/j.cjbt.issn.1004-549x.2025.05.008&lang=en |
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| _version_ | 1849325250790031360 |
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| author | TANG Shili LIN Bixuan HUANG Enxia HE Ying XUE Yuan ZHANG Yonggang |
| author_facet | TANG Shili LIN Bixuan HUANG Enxia HE Ying XUE Yuan ZHANG Yonggang |
| author_sort | TANG Shili |
| collection | DOAJ |
| description | [Objective] In in vitro systems for differentiating and expanding natural killer (NK) cells, feeder cells provide essential cell-cell contact and paracrine signals that drive precursor proliferation and terminal maturation. However, existing xenogeneic feeder cells or tumor-derived genetically modified feeder cells pose risks of residual immunogenicity and malignant transformation, limiting clinical use. This study aims to develop a humanized mesenchymal-like stromal cell (hematopoietic organoid-derived human mesenchymal stromal cells, HO-hMSCs) derived from iPSC-based hematopoietic organoids, and elucidate its mechanisms of NK-supportive activity to enable a safe, efficient platform for clinical-grade NK cell production. [Methods] Human induced pluripotent stem cells (iPSCs) were differentiated into hematopoietic organoids, from which HO-hMSCs were isolated. Flow-cytometric phenotyping and bulk RNA-sequencing were performed to compare HO-hMSCs with umbilical cord-derived MSCs (UC-hMSCs). The effect of HO-hMSCs on NK cell differentiation efficiency (CD3-CD56+) and effector maturation (CD16 expression) were assessed by co-culture experiments, using UC-hMSCs as control. [Results] 1) Hematopoietic organoid induction and NK differentiation: iPSCs were induced to form hematopoietic organoids using cytokine cocktails, which further differentiated into high-purity CD45+CD56+ NK cells [(82.8%±12.07)% efficiency on day 21]. 2) HO-hMSC characteristics: HO-hMSCs exhibited upregulated expression of Notch pathway ligands (DLL4, JAG1, 4.06-8.04-fold), homeobox genes (HOXA3, HOXA5, log2FC=1.28 and 1.44), and key regulators of NK development (GATA3, BCL11A) and cytokine receptors (IL7R, IL27RA, 6.76 to 13.34-fold increase). 3) Functional validation: Compared to UC-hMSCs, HO-hMSCs co-culture significantly enhanced NK cell proportion by 30.5% (P<0.05) and increased CD16 positivity (+20.5%). [Conclusion] This study for the first time reveals that human hematopoietic organoid-derived HO-hMSCs possess potent hematopoietic niche-supportive activity. It provides a humanized, feeder-free platform for robust clinical-grade NK cell production and expands the translational utility of organoid technologies in cell therapy. |
| format | Article |
| id | doaj-art-7fa4f272a65040e29b612ab32ad85f10 |
| institution | Kabale University |
| issn | 1004-549X |
| language | zho |
| publishDate | 2025-05-01 |
| publisher | Institute of Blood Transfusion of Chinese Academy of Medical Sciences |
| record_format | Article |
| series | Zhongguo shuxue zazhi |
| spelling | doaj-art-7fa4f272a65040e29b612ab32ad85f102025-08-20T03:48:27ZzhoInstitute of Blood Transfusion of Chinese Academy of Medical SciencesZhongguo shuxue zazhi1004-549X2025-05-0138564465110.13303/j.cjbt.issn.1004-549x.2025.05.0081004-549X(2025)5-0644-08Isolation andfunctional characterization of HO-hMSCs as NK-supportive cells derived from hematopoietic organoidsTANG Shili0LIN Bixuan1HUANG Enxia2HE Ying3XUE Yuan4ZHANG Yonggang5Institute of Blood Transfusion, Chinese Academy of Medical Sciences & Peking Union Medical College, Chengdu 610052, ChinaInstitute of Blood Transfusion, Chinese Academy of Medical Sciences & Peking Union Medical College, Chengdu 610052, ChinaInstitute of Blood Transfusion, Chinese Academy of Medical Sciences & Peking Union Medical College, Chengdu 610052, ChinaInstitute of Blood Transfusion, Chinese Academy of Medical Sciences & Peking Union Medical College, Chengdu 610052, ChinaInstitute of Blood Transfusion, Chinese Academy of Medical Sciences & Peking Union Medical College, Chengdu 610052, ChinaInstitute of Blood Transfusion, Chinese Academy of Medical Sciences & Peking Union Medical College, Chengdu 610052, China[Objective] In in vitro systems for differentiating and expanding natural killer (NK) cells, feeder cells provide essential cell-cell contact and paracrine signals that drive precursor proliferation and terminal maturation. However, existing xenogeneic feeder cells or tumor-derived genetically modified feeder cells pose risks of residual immunogenicity and malignant transformation, limiting clinical use. This study aims to develop a humanized mesenchymal-like stromal cell (hematopoietic organoid-derived human mesenchymal stromal cells, HO-hMSCs) derived from iPSC-based hematopoietic organoids, and elucidate its mechanisms of NK-supportive activity to enable a safe, efficient platform for clinical-grade NK cell production. [Methods] Human induced pluripotent stem cells (iPSCs) were differentiated into hematopoietic organoids, from which HO-hMSCs were isolated. Flow-cytometric phenotyping and bulk RNA-sequencing were performed to compare HO-hMSCs with umbilical cord-derived MSCs (UC-hMSCs). The effect of HO-hMSCs on NK cell differentiation efficiency (CD3-CD56+) and effector maturation (CD16 expression) were assessed by co-culture experiments, using UC-hMSCs as control. [Results] 1) Hematopoietic organoid induction and NK differentiation: iPSCs were induced to form hematopoietic organoids using cytokine cocktails, which further differentiated into high-purity CD45+CD56+ NK cells [(82.8%±12.07)% efficiency on day 21]. 2) HO-hMSC characteristics: HO-hMSCs exhibited upregulated expression of Notch pathway ligands (DLL4, JAG1, 4.06-8.04-fold), homeobox genes (HOXA3, HOXA5, log2FC=1.28 and 1.44), and key regulators of NK development (GATA3, BCL11A) and cytokine receptors (IL7R, IL27RA, 6.76 to 13.34-fold increase). 3) Functional validation: Compared to UC-hMSCs, HO-hMSCs co-culture significantly enhanced NK cell proportion by 30.5% (P<0.05) and increased CD16 positivity (+20.5%). [Conclusion] This study for the first time reveals that human hematopoietic organoid-derived HO-hMSCs possess potent hematopoietic niche-supportive activity. It provides a humanized, feeder-free platform for robust clinical-grade NK cell production and expands the translational utility of organoid technologies in cell therapy.https://www.cjbt.cn/thesisDetails#10.13303/j.cjbt.issn.1004-549x.2025.05.008&lang=enhematopoietic organoidmesenchymal stromal cellsfeeder layer cellsnk differentiation |
| spellingShingle | TANG Shili LIN Bixuan HUANG Enxia HE Ying XUE Yuan ZHANG Yonggang Isolation andfunctional characterization of HO-hMSCs as NK-supportive cells derived from hematopoietic organoids Zhongguo shuxue zazhi hematopoietic organoid mesenchymal stromal cells feeder layer cells nk differentiation |
| title | Isolation andfunctional characterization of HO-hMSCs as NK-supportive cells derived from hematopoietic organoids |
| title_full | Isolation andfunctional characterization of HO-hMSCs as NK-supportive cells derived from hematopoietic organoids |
| title_fullStr | Isolation andfunctional characterization of HO-hMSCs as NK-supportive cells derived from hematopoietic organoids |
| title_full_unstemmed | Isolation andfunctional characterization of HO-hMSCs as NK-supportive cells derived from hematopoietic organoids |
| title_short | Isolation andfunctional characterization of HO-hMSCs as NK-supportive cells derived from hematopoietic organoids |
| title_sort | isolation andfunctional characterization of ho hmscs as nk supportive cells derived from hematopoietic organoids |
| topic | hematopoietic organoid mesenchymal stromal cells feeder layer cells nk differentiation |
| url | https://www.cjbt.cn/thesisDetails#10.13303/j.cjbt.issn.1004-549x.2025.05.008&lang=en |
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