Shuntaro Yamada, Katerina Holomkova, Åshild Johansen, Masoumeh Jahani Kadousaraei, Niyaz Al-Sharabi, Francesco Torelli, Pierfrancesco Pagella, Ana Angelova Volponi, Hiroshi Egusa, Inge Fristad, Kamal Mustafa
{"title":"长期生物库牙髓干细胞保留血管生成潜力的血管化组织工程-实验室研究。","authors":"Shuntaro Yamada, Katerina Holomkova, Åshild Johansen, Masoumeh Jahani Kadousaraei, Niyaz Al-Sharabi, Francesco Torelli, Pierfrancesco Pagella, Ana Angelova Volponi, Hiroshi Egusa, Inge Fristad, Kamal Mustafa","doi":"10.1111/iej.70036","DOIUrl":null,"url":null,"abstract":"<p><strong>Aim: </strong>This study aimed to evaluate whether human dental pulp stem cells (DPSCs), after long-term biobanking (7-8 years), retain their pro-angiogenic properties and can be used to engineer vascularised tissues, addressing their potential for clinical translation in regenerative dentistry.</p><p><strong>Methodology: </strong>Cryopreserved DPSCs from adolescent donors were recovered from biobanking and characterised for chromosomal integrity, MSC immunophenotype and multipotency. After conditioning in pro-angiogenic conditions in vitro, gene and protein expression were analysed by RT-qPCR array, flow cytometry and high-throughput immunophenotyping. Functional angiogenic capacity was assessed via in vitro tube formation, ex ovo CAM implantation assay, organ-on-chip perfusion model and long-term culture (45 days) in clinical-grade GelMA hydrogels, with and without HUVECs.</p><p><strong>Results: </strong>Biobanked DPSCs retained MSC identity and multi-lineage differentiation potential. Pro-angiogenic/endothelial conditioning enhanced the expression of angiogenic/endothelial genes (PECAM1, VEGFR2, NRP1, ACE), yet most cells maintained a pericyte-like phenotype. Both naive and endothelial-conditioned DPSCs (i.e., naiveDPSCs and endoDPSCs, respectively) significantly enhanced vascular ingrowth in the CAM model. In the organ-on-chip system, naiveDPSCs formed perfusable vasculature with HUVECs and differentiated into perivascular cell types. Most notably, endoDPSCs alone successfully generated vascularised tissue with both CD31(+) and αSMA(+) cells present in GelMA hydrogels after prolonged stimulation.</p><p><strong>Conclusion: </strong>Long-term biobanked DPSCs preserve their angiogenic potential and, following extended endothelial induction, can independently generate vascularised tissue in 3D in vitro culture models. This is the first report demonstrating the comprehensive pro-angiogenic characterisation and the feasibility of using biobanked DPSCs for vascularised tissue engineering, highlighting their strong clinical applicability for future regenerative therapies.</p>","PeriodicalId":13724,"journal":{"name":"International endodontic journal","volume":" ","pages":""},"PeriodicalIF":7.1000,"publicationDate":"2025-09-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Long-Term Biobanked Dental Pulp Stem Cells Retain Angiogenic Potential for Vascularised Tissue Engineering-Laboratory Investigation.\",\"authors\":\"Shuntaro Yamada, Katerina Holomkova, Åshild Johansen, Masoumeh Jahani Kadousaraei, Niyaz Al-Sharabi, Francesco Torelli, Pierfrancesco Pagella, Ana Angelova Volponi, Hiroshi Egusa, Inge Fristad, Kamal Mustafa\",\"doi\":\"10.1111/iej.70036\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><strong>Aim: </strong>This study aimed to evaluate whether human dental pulp stem cells (DPSCs), after long-term biobanking (7-8 years), retain their pro-angiogenic properties and can be used to engineer vascularised tissues, addressing their potential for clinical translation in regenerative dentistry.</p><p><strong>Methodology: </strong>Cryopreserved DPSCs from adolescent donors were recovered from biobanking and characterised for chromosomal integrity, MSC immunophenotype and multipotency. After conditioning in pro-angiogenic conditions in vitro, gene and protein expression were analysed by RT-qPCR array, flow cytometry and high-throughput immunophenotyping. Functional angiogenic capacity was assessed via in vitro tube formation, ex ovo CAM implantation assay, organ-on-chip perfusion model and long-term culture (45 days) in clinical-grade GelMA hydrogels, with and without HUVECs.</p><p><strong>Results: </strong>Biobanked DPSCs retained MSC identity and multi-lineage differentiation potential. Pro-angiogenic/endothelial conditioning enhanced the expression of angiogenic/endothelial genes (PECAM1, VEGFR2, NRP1, ACE), yet most cells maintained a pericyte-like phenotype. Both naive and endothelial-conditioned DPSCs (i.e., naiveDPSCs and endoDPSCs, respectively) significantly enhanced vascular ingrowth in the CAM model. In the organ-on-chip system, naiveDPSCs formed perfusable vasculature with HUVECs and differentiated into perivascular cell types. Most notably, endoDPSCs alone successfully generated vascularised tissue with both CD31(+) and αSMA(+) cells present in GelMA hydrogels after prolonged stimulation.</p><p><strong>Conclusion: </strong>Long-term biobanked DPSCs preserve their angiogenic potential and, following extended endothelial induction, can independently generate vascularised tissue in 3D in vitro culture models. This is the first report demonstrating the comprehensive pro-angiogenic characterisation and the feasibility of using biobanked DPSCs for vascularised tissue engineering, highlighting their strong clinical applicability for future regenerative therapies.</p>\",\"PeriodicalId\":13724,\"journal\":{\"name\":\"International endodontic journal\",\"volume\":\" \",\"pages\":\"\"},\"PeriodicalIF\":7.1000,\"publicationDate\":\"2025-09-23\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"International endodontic journal\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://doi.org/10.1111/iej.70036\",\"RegionNum\":1,\"RegionCategory\":\"医学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q1\",\"JCRName\":\"DENTISTRY, ORAL SURGERY & MEDICINE\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"International endodontic journal","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.1111/iej.70036","RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"DENTISTRY, ORAL SURGERY & MEDICINE","Score":null,"Total":0}
Aim: This study aimed to evaluate whether human dental pulp stem cells (DPSCs), after long-term biobanking (7-8 years), retain their pro-angiogenic properties and can be used to engineer vascularised tissues, addressing their potential for clinical translation in regenerative dentistry.
Methodology: Cryopreserved DPSCs from adolescent donors were recovered from biobanking and characterised for chromosomal integrity, MSC immunophenotype and multipotency. After conditioning in pro-angiogenic conditions in vitro, gene and protein expression were analysed by RT-qPCR array, flow cytometry and high-throughput immunophenotyping. Functional angiogenic capacity was assessed via in vitro tube formation, ex ovo CAM implantation assay, organ-on-chip perfusion model and long-term culture (45 days) in clinical-grade GelMA hydrogels, with and without HUVECs.
Results: Biobanked DPSCs retained MSC identity and multi-lineage differentiation potential. Pro-angiogenic/endothelial conditioning enhanced the expression of angiogenic/endothelial genes (PECAM1, VEGFR2, NRP1, ACE), yet most cells maintained a pericyte-like phenotype. Both naive and endothelial-conditioned DPSCs (i.e., naiveDPSCs and endoDPSCs, respectively) significantly enhanced vascular ingrowth in the CAM model. In the organ-on-chip system, naiveDPSCs formed perfusable vasculature with HUVECs and differentiated into perivascular cell types. Most notably, endoDPSCs alone successfully generated vascularised tissue with both CD31(+) and αSMA(+) cells present in GelMA hydrogels after prolonged stimulation.
Conclusion: Long-term biobanked DPSCs preserve their angiogenic potential and, following extended endothelial induction, can independently generate vascularised tissue in 3D in vitro culture models. This is the first report demonstrating the comprehensive pro-angiogenic characterisation and the feasibility of using biobanked DPSCs for vascularised tissue engineering, highlighting their strong clinical applicability for future regenerative therapies.
期刊介绍:
The International Endodontic Journal is published monthly and strives to publish original articles of the highest quality to disseminate scientific and clinical knowledge; all manuscripts are subjected to peer review. Original scientific articles are published in the areas of biomedical science, applied materials science, bioengineering, epidemiology and social science relevant to endodontic disease and its management, and to the restoration of root-treated teeth. In addition, review articles, reports of clinical cases, book reviews, summaries and abstracts of scientific meetings and news items are accepted.
The International Endodontic Journal is essential reading for general dental practitioners, specialist endodontists, research, scientists and dental teachers.