Development of fully defined xeno-free culture system for the preparation and propagation of cell therapy-compliant human adipose stem cells

Stem Cell Research & Therapy

Susanna Miettinen²  Bettina Mannerström²  Mohan C Vemuri¹  Andrew Campbell³  Shayne Boucher¹  Miia Juntunen²  Mimmi Patrikoski² 

[1] Life Technologies, Cell Therapy Systems, Frederick, MD, USA;Science Center, Tampere University Hospital, Tampere, Finland;Life Technologies, BioProduction, Grand Island, MD, USA

关键词: Cell therapy;    Flow cytometry;    Immunophenotype;    Proliferation rate;    Multipotentiality;    Fetal bovine serum;    Human serum;    Serum-free;    Xeno-free;    Adipose stem cells;   

Others  :  848436
DOI  :  10.1186/scrt175

 received in 2012-09-21, accepted in 2013-03-04,  发布年份 2013

【 摘 要 】

Introduction

Adipose tissue is an attractive and abundant source of multipotent stem cells. Human adipose stem cells (ASCs) have shown to have therapeutic relevancy in diverse clinical applications. Nevertheless, expansion of ASCs is often necessary before performing clinical studies. Standard in vitro cell-culture techniques use animal-derived reagents that should be avoided in clinical use because of safety issues. Therefore, xeno- and serum-free (XF/SF) reagents are highly desirable for enhancing the safety and quality of the transplanted ASCs.

Methods

In the current study, animal component-free isolation and cell-expansion protocols were developed for ASCs. StemPro MSC SFM XF medium with either CELLstart™ CTS™ coating or Coating Matrix Kit were tested for their ability to support XF/SF growth. Basic stem-cell characteristics such as immunophenotype (CD3, CD11a, CD14, CD19, CD34, CD45RO, CD54, CD73, CD80, CD86, CD90, CD105, HLA-DR), proliferation, and differentiation potential were assessed in XF/SF conditions and compared with human serum (HS) or traditionally used fetal bovine serum (FBS) cultures.

Results

ASCs cultured in XF/SF conditions had significantly higher proliferation rates compared with HS/FBS cultures. Characteristic immunophenotypes of ASCs were maintained in every condition; however, cells expanded in XF/SF conditions showed significantly lower expression of CD54 (intercellular adhesion molecule 1, ICAM-1) at low passage number. Further, multilineage differentiation potential of ASCs was maintained in every culture condition.

Conclusions

Our findings demonstrated that the novel XF/SF conditions maintained the basic stem cell features of ASCs and the animal-free workflow followed in this study has great potential in clinical cell therapies.

【 授权许可】

   
2013 Patrikoski et al.; licensee BioMed Central Ltd.

【 图 表 】

Figure 1.

Figure 2.

Figure 3.

Figure 4.

Figure 5.

Figure 6.

Figure 7.

【 参考文献 】

• [1]Gimble J, Guilak F: Adipose-derived adult stem cells: isolation, characterization, and differentiation potential. Cytotherapy 2003, 5:362-369.
• [2]Zuk PA, Zhu M, Ashjian P, De Ugarte DA, Huang JI, Mizuno H, Alfonso ZC, Fraser JK, Benhaim P, Hedrick MH: Human adipose tissue is a source of multipotent stem cells. Mol Biol Cell 2002, 13:4279-4295.
• [3]Zuk PA, Zhu M, Mizuno H, Huang J, Futrell JW, Katz AJ, Benhaim P, Lorenz HP, Hedrick MH: Multilineage cells from human adipose tissue: implications for cell-based therapies. Tissue Eng 2001, 7:211-228.
• [4]Thesleff T, Lehtimaki K, Niskakangas T, Mannerstrom B, Miettinen S, Suuronen R, Ohman J: Cranioplasty with adipose-derived stem cells and biomaterial: a novel method for cranial reconstruction. Neurosurgery 2011, 68:1535-1540.
• [5]Casteilla L, Planat-Benard V, Laharrague P, Cousin B: Adipose-derived stromal cells: their identity and uses in clinical trials: an update. World J Stem Cells 2011, 3:25-33.
• [6]Mizuno H: Adipose-derived stem and stromal cells for cell-based therapy: current status of preclinical studies and clinical trials. Curr Opin Mol Ther 2010, 12:442-449.
• [7]Dominici M, Le Blanc K, Mueller I, Slaper-Cortenbach I, Marini F, Krause D, Deans R, Keating A, Prockop D, Horwitz E: Minimal criteria for defining multipotent mesenchymal stromal cells: The International Society for Cellular Therapy position statement. Cytotherapy 2006, 8:315-317.
• [8]Niemeyer P, Vohrer J, Schmal H, Kasten P, Fellenberg J, Suedkamp NP, Mehlhorn AT: Survival of human mesenchymal stromal cells from bone marrow and adipose tissue after xenogenic transplantation in immunocompetent mice. Cytotherapy 2008, 10:784-795.
• [9]Kuo YR, Chen CC, Goto S, Lee IT, Huang CW, Tsai CC, Wang CT, Chen CL: Modulation of immune response and T-cell regulation by donor adipose-derived stem cells in a rodent hind-limb allotransplant model. Plast Reconstr Surg 2011, 128:661e-672e.
• [10]Puissant B, Barreau C, Bourin P, Clavel C, Corre J, Bousquet C, Taureau C, Cousin B, Abbal M, Laharrague P, Penicaud L, Casteilla L, Blancher A: Immunomodulatory effect of human adipose tissue-derived adult stem cells: comparison with bone marrow mesenchymal stem cells. Br J Haematol 2005, 129:118-129.
• [11]Niemeyer P, Kornacker M, Mehlhorn A, Seckinger A, Vohrer J, Schmal H, Kasten P, Eckstein V, Sudkamp NP, Krause U: Comparison of immunological properties of bone marrow stromal cells and adipose tissue-derived stem cells before and after osteogenic differentiation in vitro. Tissue Eng 2007, 13:111-121.
• [12]Mackensen A, Drager R, Schlesier M, Mertelsmann R, Lindemann A: Presence of IgE antibodies to bovine serum albumin in a patient developing anaphylaxis after vaccination with human peptide-pulsed dendritic cells. Cancer Immunol Immunother 2000, 49:152-156.
• [13]Selvaggi TA, Walker RE, Fleisher TA: Development of antibodies to fetal calf serum with arthus-like reactions in human immunodeficiency virus-infected patients given syngeneic lymphocyte infusions. Blood 1997, 89:776-779.
• [14]Tateishi K, Ando W, Higuchi C, Hart DA, Hashimoto J, Nakata K, Yoshikawa H, Nakamura N: Comparison of human serum with fetal bovine serum for expansion and differentiation of human synovial MSC: potential feasibility for clinical applications. Cell Transplant 2008, 17:549-557.
• [15]Schallmoser K, Strunk D: Preparation of pooled human platelet lysate (pHPL) as an efficient supplement for animal serum-free human stem cell cultures. J Vis Exp 2009.
• [16]Bieback K, Hecker A, Kocaomer A, Lannert H, Schallmoser K, Strunk D, Kluter H: Human alternatives to fetal bovine serum for the expansion of mesenchymal stromal cells from bone marrow. Stem Cells 2009, 27:2331-2341.
• [17]Parker AM, Shang H, Khurgel M, Katz AJ: Low serum and serum-free culture of multipotential human adipose stem cells. Cytotherapy 2007, 9:637-646.
• [18]Haimi S, Moimas L, Pirhonen E, Lindroos B, Huhtala H, Raty S, Kuokkanen H, Sandor GK, Miettinen S, Suuronen R: Calcium phosphate surface treatment of bioactive glass causes a delay in early osteogenic differentiation of adipose stem cells. J Biomed Mater Res A 2009, 91:540-547.
• [19]Lindroos B, Boucher S, Chase L, Kuokkanen H, Huhtala H, Haataja R, Vemuri M, Suuronen R, Miettinen S: Serum-free, xeno-free culture media maintain the proliferation rate and multipotentiality of adipose stem cells in vitro. Cytotherapy 2009, 11:958-972.
• [20]Cristofalo VJ, Allen RG, Pignolo RJ, Martin BG, Beck JC: Relationship between donor age and the replicative lifespan of human cells in culture: a reevaluation. Proc Natl Acad Sci U S A 1998, 95:10614-10619.
• [21]Haimi S, Suuriniemi N, Haaparanta AM, Ella V, Lindroos B, Huhtala H, Raty S, Kuokkanen H, Sandor GK, Kellomaki M, Miettinen S, Suuronen R: Growth and osteogenic differentiation of adipose stem cells on PLA/bioactive glass and PLA/beta-TCP scaffolds. Tissue Eng Part A 2009, 15:1473-1480.
• [22]Denker AE, Nicoll SB, Tuan RS: Formation of cartilage-like spheroids by micromass cultures of murine C3H10T1/2 cells upon treatment with transforming growth factor-beta 1. Differentiation 1995, 59:25-34.
• [23]Maenpaa K, Ella V, Mauno J, Kellomaki M, Suuronen R, Ylikomi T, Miettinen S: Use of adipose stem cells and polylactide discs for tissue engineering of the temporomandibular joint disc. J R Soc Interface 2010, 7:177-188.
• [24]Pfaffl MW: A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res 2001, 29:e45.
• [25]Spees JL, Gregory CA, Singh H, Tucker HA, Peister A, Lynch PJ, Hsu SC, Smith J, Prockop DJ: Internalized antigens must be removed to prepare hypoimmunogenic mesenchymal stem cells for cell and gene therapy. Mol Ther 2004, 9:747-756.
• [26]Trivedi HL, Vanikar AV, Thakker U, Firoze A, Dave SD, Patel CN, Patel JV, Bhargava AB, Shankar V: Human adipose tissue-derived mesenchymal stem cells combined with hematopoietic stem cell transplantation synthesize insulin. Transplant Proc 2008, 40:1135-1139.
• [27]Tzouvelekis A, Koliakos G, Ntolios P, Baira I, Bouros E, Oikonomou A, Zissimopoulos A, Kolios G, Kakagia D, Paspaliaris V, Kotsianidis I, Froudarakis M, Bouros D: Stem cell therapy for idiopathic pulmonary fibrosis: a protocol proposal. J Transl Med 2011, 9:182. BioMed Central Full Text
• [28]Mesimaki K, Lindroos B, Tornwall J, Mauno J, Lindqvist C, Kontio R, Miettinen S, Suuronen R: Novel maxillary reconstruction with ectopic bone formation by GMP adipose stem cells. Int J Oral Maxillofac Surg 2009, 38:201-209.
• [29]Dromard C, Bourin P, Andre M, De Barros S, Casteilla L, Planat-Benard V: Human adipose derived stroma/stem cells grow in serum-free medium as floating spheres. Exp Cell Res 2011, 317:770-780.
• [30]Santos F, Andrade PZ, Abecasis MM, Gimble JM, Chase LG, Campbell AM, Boucher S, Vemuri MC, Silva CL, Cabral JM: Toward a clinical-grade expansion of mesenchymal stem cells from human sources: a microcarrier-based culture system under xeno-free conditions. Tissue Eng Part C Methods 2011, 17:1201-1210.
• [31]Naaijkens BA, Niessen HW, Prins HJ, Krijnen PA, Kokhuis TJ, de Jong N, van Hinsbergh VW, Kamp O, Helder MN, Musters RJ, van Dijk A, Juffermans LJ: Human platelet lysate as a fetal bovine serum substitute improves human adipose-derived stromal cell culture for future cardiac repair applications. Cell Tissue Res 2012, 348:119-130.
• [32]Blande IS, Bassaneze V, Lavini-Ramos C, Fae KC, Kalil J, Miyakawa AA, Schettert IT, Krieger JE: Adipose tissue mesenchymal stem cell expansion in animal serum-free medium supplemented with autologous human platelet lysate. Transfusion 2009, 49:2680-2685.
• [33]Chase LG, Lakshmipathy U, Solchaga LA, Rao MS, Vemuri MC: A novel serum-free medium for the expansion of human mesenchymal stem cells. Stem Cell Res Ther 2010, 1:8. BioMed Central Full Text
• [34]Yang S, Pilgaard L, Chase L, Boucher S, Vemuri M, Fink TD, Zachar V: Defined xenogeneic-free and hypoxic environment provides superior conditions for long-term expansion of human adipose-derived stem cells. Tissue Eng Part C Methods 2012, 18:593-602.
• [35]Konno M, Hamazaki TS, Fukuda S, Tokuhara M, Uchiyama H, Okazawa H, Okochi H, Asashima M: Efficiently differentiating vascular endothelial cells from adipose tissue-derived mesenchymal stem cells in serum-free culture. Biochem Biophys Res Commun 2010, 400:461-465.
• [36]Mirabet V, Solves P, Minana MD, Encabo A, Carbonell-Uberos F, Blanquer A, Roig R: Human platelet lysate enhances the proliferative activity of cultured human fibroblast-like cells from different tissues. Cell Tissue Bank 2008, 9:1-10.
• [37]Rebelatto CK, Aguiar AM, Moretao MP, Senegaglia AC, Hansen P, Barchiki F, Oliveira J, Martins J, Kuligovski C, Mansur F, Christofis A, Amaral VF, Brofman PS, Goldenberg S, Nakao LS, Correa A: Dissimilar differentiation of mesenchymal stem cells from bone marrow, umbilical cord blood, and adipose tissue. Exp Biol Med (Maywood) 2008, 233:901-913.
• [38]Park IS, Han M, Rhie JW, Kim SH, Jung Y, Kim IH, Kim SH: The correlation between human adipose-derived stem cells differentiation and cell adhesion mechanism. Biomaterials 2009, 30:6835-6843.
• [39]Kang JM, Han M, Park IS, Jung Y, Kim SH, Kim SH: Adhesion and differentiation of adipose-derived stem cells on a substrate with immobilized fibroblast growth factor. Acta Biomater 2012, 8:1759-1767.
• [40]Cordonnier T, Langonne A, Sohier J, Layrolle P, Rosset P, Sensebe L, Deschaseaux F: Consistent osteoblastic differentiation of human mesenchymal stem cells with bone morphogenetic protein 4 and low serum. Tissue Eng Part C Methods 2011, 17:249-259.
• [41]Bosetti M, Cannas M: The effect of bioactive glasses on bone marrow stromal cells differentiation. Biomaterials 2005, 26:3873-3879.
• [42]Marino G, Rosso F, Cafiero G, Tortora C, Moraci M, Barbarisi M, Barbarisi A: Beta-tricalcium phosphate 3D scaffold promote alone osteogenic differentiation of human adipose stem cells: in vitro study. J Mater Sci Mater Med 2010, 21:353-363.
• [43]Moutos FT, Estes BT, Guilak F: Multifunctional hybrid three-dimensionally woven scaffolds for cartilage tissue engineering. Macromol Biosci 2010, 10:1355-1364.