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Isolation and prolonged expansion of oral mesenchymal stem cells under clinical-grade, GMP-compliant conditions differentially affects 'stemness' properties

التفاصيل البيبلوغرافية
العنوان: Isolation and prolonged expansion of oral mesenchymal stem cells under clinical-grade, GMP-compliant conditions differentially affects 'stemness' properties
المؤلفون: Eleni Aggelidou, Aristeidis Kritis, Evangelia Gousopoulou, Petros Koidis, Danae Anastasia Apatzidou, Werner Geurtsen, Gabriele Leyhausen, Athina Bakopoulou, Joachim Volk
المصدر: Stem Cell Research & Therapy, Vol 8, Iss 1, Pp 1-21 (2017)
Stem Cell Research & Therapy
بيانات النشر: BMC, 2017.
سنة النشر: 2017
مصطلحات موضوعية: 0301 basic medicine, Cellular differentiation, Cell Culture Techniques, Dental pulp stem cells, Bone Morphogenetic Protein 2, Gene Expression, Medicine (miscellaneous), Culture Media, Serum-Free, Osteogenesis, lcsh:QD415-436, Aggrecans, education.field_of_study, lcsh:R5-920, Adipogenesis, Alveolar bone marrow mesenchymal stem cells, Cell Differentiation, SOX9 Transcription Factor, Intercellular Signaling Peptides and Proteins, Molecular Medicine, Stem cell, lcsh:Medicine (General), Chondrogenesis, Good manufacturing practice-compliant cell preparation, Drug Industry, “Stemness” properties, Population, Prolonged expansion, Bone Marrow Cells, Biology, Biochemistry, Genetics and Molecular Biology (miscellaneous), Andrology, lcsh:Biochemistry, 03 medical and health sciences, Antigens, CD, Alveolar Process, Humans, CD90, education, Dental Pulp, Cell Proliferation, Oral mesenchymal stem cells, Research, Mesenchymal stem cell, Telomere Homeostasis, Mesenchymal Stem Cells, Cell Biology, Alkaline Phosphatase, beta-Galactosidase, PPAR gamma, Lipoprotein Lipase, Clinical-grade expansion, 030104 developmental biology, Cell culture, Immunology, Biomarkers, Fetal bovine serum
الوصف: Background Development of clinical-grade cell preparations is central to meeting the regulatory requirements for cellular therapies under good manufacturing practice-compliant (cGMP) conditions. Since addition of animal serum in culture media may compromise safe and efficient expansion of mesenchymal stem cells (MSCs) for clinical use, this study aimed to investigate the potential of two serum/xeno-free, cGMP culture systems to maintain long-term “stemness” of oral MSCs (dental pulp stem cells (DPSCs) and alveolar bone marrow MSCs (aBMMSCs)), compared to conventional serum-based expansion. Methods DPSC and aBMMSC cultures (n = 6/cell type) were established from pulp and alveolar osseous biopsies respectively. Three culture systems were used: StemPro_MSC/SFM_XenoFree (Life Technologies); StemMacs_MSC/XF (Miltenyi Biotek); and α-MEM (Life Technologies) with 15% fetal bovine serum. Growth (population doublings (PDs)), immunophenotypic (flow cytometric analysis of MSC markers) and senescence (β-galactosidase (SA-β-gal) activity; telomere length) characteristics were determined during prolonged expansion. Gene expression patterns of osteogenic (ALP, BMP-2), adipogenic (LPL, PPAR-γ) and chondrogenic (ACAN, SOX-9) markers and maintenance of multilineage differentiation potential were determined by real-time PCR. Results Similar isolation efficiency and stable growth dynamics up to passage 10 were observed for DPSCs under all expansion conditions. aBMMSCs showed lower cumulative PDs compared to DPSCs, and when StemMacs was used substantial delays in cell proliferation were noted after passages 6–7. Serum/xeno-free expansion produced cultures with homogeneous spindle-shaped phenotypes, while serum-based expansion preserved differential heterogeneous characteristics of each MSC population. Prolonged expansion of both MSC types but in particular the serum/xeno-free-expanded aBMMSCs was associated with downregulation of CD146, CD105, Stro-1, SSEA-1 and SSEA-4, but not CD90, CD73 and CD49f, in parallel with an increase of SA-gal-positive cells, cell size and granularity and a decrease in telomere length. Expansion under both serum-free systems resulted in “osteogenic pre-disposition”, evidenced by upregulation of osteogenic markers and elimination of chondrogenic and adipogenic markers, while serum-based expansion produced only minor changes. DPSCs retained a diminishing (CCM, StemPro) or increasing (StemMacs) mineralization potential with passaging, while aBMMSCs lost this potential after passages 6–7 under all expansion conditions. Conclusions These findings indicate there is still a vacant role for development of qualified protocols for clinical-grade expansion of oral MSCs; a key milestone achievement for translation of research from the bench to clinics. Electronic supplementary material The online version of this article (doi:10.1186/s13287-017-0705-0) contains supplementary material, which is available to authorized users.
اللغة: English
تدمد: 1757-6512
URL الوصول: https://explore.openaire.eu/search/publication?articleId=doi_dedup___::b86efbbb85a0608af3b1fea3d2a6df26
http://link.springer.com/article/10.1186/s13287-017-0705-0
حقوق: OPEN
رقم الأكسشن: edsair.doi.dedup.....b86efbbb85a0608af3b1fea3d2a6df26
قاعدة البيانات: OpenAIRE
الوصف
تدمد:17576512