Encapsulation and 3D culture of human adipose-derived stem cells in an in-situ crosslinked hybrid hydrogel composed of PEG-based hyperbranched copolymer and hyaluronic acid

IntroductionCell therapy using adipose-derived stem cells has been reported to improve chronic wounds via differentiation and paracrine effects. One such strategy is to deliver stem cells in hydrogels, which are studied increasingly as cell delivery vehicles for therapeutic healing and inducing tissue regeneration. This study aimed to determine the behaviour of encapsulated adipose-derived stem cells and identify the secretion profile of suitable growth factors for wound healing in a newly developed thermoresponsive PEG–hyaluronic acid (HA) hybrid hydrogel to provide a novel living dressing system.MethodsIn this study, human adipose-derived stem cells (hADSCs) were encapsulated in situ in a water-soluble, thermoresponsive hyperbranched PEG-based copolymer (PEGMEMA–MEO2MA–PEGDA) with multiple acrylate functional groups in combination with thiolated HA, which was developed via deactivated enhanced atom transfer radical polymerisation of poly(ethylene glycol) methyl ether methacrylate (PEGMEMA, Mn = 475), 2-(2-methoxyethoxy) ethyl methacrylate (MEO2MA) and poly(ethylene glycol) diacrylate PEGDA (Mn = 258). hADSCs embedded in the PEGMEMA–MEO2MA–PEGDA and HA hybrid hydrogel system (P-SH-HA) were monitored and analysed for their cell viability, cell proliferation and secretion of growth factors (vascular endothelial growth factor, transforming growth factor beta and placental-derived growth factor) and cytokines (IFNγ, IL-2 and IL-10) under three-dimensional culture conditions via the ATP activity assay, alamarBlue® assay, LIVE/DEAD® assay and multiplex ELISA, respectively.ResultshADSCs were successfully encapsulated in situ with high cell viability for up to 7 days in hydrogels. Although cellular proliferation was inhibited, cellular secretion of growth factors such as vascular endothelial growth factor and placental-derived growth factor production increased over 7 days, whereas IL-2 and IFNγ release were unaffected.ConclusionThis study indicates that hADSCs can be maintained in a P-SH-HA hydrogel, and secrete pro-angiogenic growth factors with low cytotoxicity. With the potential to add more functionality for further structural modifications, this stem cell hydrogel system can be an ideal living dressing system for wound healing applications.

[1]  R. Auzély-Velty,et al.  Design of biomimetic cell-interactive substrates using hyaluronic acid hydrogels with tunable mechanical properties. , 2012, Biomacromolecules.

[2]  R Busse,et al.  Improvement of Postnatal Neovascularization by Human Adipose Tissue–Derived Stem Cells , 2004, Circulation.

[3]  Robert J Fisher,et al.  Dual growth factor-induced angiogenesis in vivo using hyaluronan hydrogel implants. , 2006, Biomaterials.

[4]  J. Park,et al.  Wound healing effect of adipose-derived stem cells: a critical role of secretory factors on human dermal fibroblasts. , 2007, Journal of dermatological science.

[5]  D. Mooney,et al.  Growth factor delivery-based tissue engineering: general approaches and a review of recent developments , 2011, Journal of The Royal Society Interface.

[6]  Keith L. March,et al.  Secretion of Angiogenic and Antiapoptotic Factors by Human Adipose Stromal Cells , 2004, Circulation.

[7]  H. Lorenz,et al.  Multilineage cells from human adipose tissue: implications for cell-based therapies. , 2001, Tissue engineering.

[8]  S. Werner,et al.  Regulation of wound healing by growth factors and cytokines. , 2003, Physiological reviews.

[9]  Brian H Annex,et al.  The VIVA Trial Vascular Endothelial Growth Factor in Ischemia for Vascular Angiogenesis , 2003 .

[10]  D. Yan,et al.  Synthesis, characterization, and in vitro evaluation of long-chain hyperbranched poly(ethylene glycol) as drug carrier. , 2010, Bioconjugate chemistry.

[11]  Achim Goepferich,et al.  Rational design of hydrogels for tissue engineering: impact of physical factors on cell behavior. , 2007, Biomaterials.

[12]  J. Hubbell,et al.  Synthetic biomaterials as instructive extracellular microenvironments for morphogenesis in tissue engineering , 2005, Nature Biotechnology.

[13]  A. Roberts,et al.  Full‐thickness wounding of the mouse tail as a model for delayed wound healing: accelerated wound closure in Smad3 knock‐out mice , 2004, Wound repair and regeneration : official publication of the Wound Healing Society [and] the European Tissue Repair Society.

[14]  Wenxin Wang,et al.  Thermoresponsive hyperbranched copolymer with multi acrylate functionality for in situ cross-linkable hyaluronic acid composite semi-IPN hydrogel , 2011, Journal of Materials Science: Materials in Medicine.

[15]  Bin Zhou,et al.  Stem cell engraftment and survival in the ischemic heart. , 2011, The Annals of thoracic surgery.

[16]  M. Benderitter,et al.  Cell Therapy Based on Adipose Tissue-Derived Stromal Cells Promotes Physiological and Pathological Wound Healing , 2009, Arteriosclerosis, thrombosis, and vascular biology.

[17]  D. Mooney,et al.  Hydrogels for tissue engineering: scaffold design variables and applications. , 2003, Biomaterials.

[18]  Li Yan,et al.  Controlled release of epidermal growth factor from hydrogels accelerates wound healing in diabetic rats. , 2012, Journal of the American Podiatric Medical Association (Print).

[19]  M. Mimeault,et al.  Stem Cells: A Revolution in Therapeutics—Recent Advances in Stem Cell Biology and Their Therapeutic Applications in Regenerative Medicine and Cancer Therapies , 2007, Clinical pharmacology and therapeutics.

[20]  A. Bayat,et al.  Exploring the role of stem cells in cutaneous wound healing , 2009, Experimental dermatology.

[21]  S. Thibeault,et al.  Biocompatibility of a synthetic extracellular matrix on immortalized vocal fold fibroblasts in 3-D culture. , 2010, Acta biomaterialia.

[22]  A. Nauta,et al.  Immunomodulatory properties of mesenchymal stromal cells. , 2007, Blood.

[23]  L. DiPietro,et al.  Factors Affecting Wound Healing , 2010, Journal of dental research.

[24]  Robert Langer,et al.  Hyaluronic acid hydrogel for controlled self-renewal and differentiation of human embryonic stem cells , 2007, Proceedings of the National Academy of Sciences.

[25]  Glenn D Prestwich,et al.  Injectable synthetic extracellular matrices for tissue engineering and repair. , 2006, Advances in experimental medicine and biology.

[26]  Glenn D Prestwich,et al.  Release of basic fibroblast growth factor from a crosslinked glycosaminoglycan hydrogel promotes wound healing , 2007, Wound repair and regeneration : official publication of the Wound Healing Society [and] the European Tissue Repair Society.

[27]  R. Martin,et al.  A guide to biological skin substitutes. , 2002, British journal of plastic surgery.

[28]  Won-Serk Kim,et al.  The wound-healing and antioxidant effects of adipose-derived stem cells , 2009, Expert opinion on biological therapy.

[29]  A. Schäffler,et al.  Concise Review: Adipose Tissue‐Derived Stromal Cells—Basic and Clinical Implications for Novel Cell‐Based Therapies , 2007, Stem cells.

[30]  Jason A. Burdick,et al.  Hyaluronic Acid Hydrogels for Biomedical Applications , 2011, Advanced materials.

[31]  S. Prakash,et al.  Bone Marrow Stem Cell Derived Paracrine Factors for Regenerative Medicine: Current Perspectives and Therapeutic Potential , 2010, Bone marrow research.

[32]  S. Bryant,et al.  Cell encapsulation in biodegradable hydrogels for tissue engineering applications. , 2008, Tissue engineering. Part B, Reviews.

[33]  Wenxin Wang,et al.  "One-step" preparation of thiol-ene clickable PEG-based thermoresponsive hyperbranched copolymer for in situ crosslinking hybrid hydrogel. , 2012, Macromolecular rapid communications.

[34]  Tatiana Segura,et al.  The spreading, migration and proliferation of mouse mesenchymal stem cells cultured inside hyaluronic acid hydrogels. , 2011, Biomaterials.

[35]  G. Vunjak‐Novakovic,et al.  Engineered microenvironments for controlled stem cell differentiation. , 2009, Tissue engineering. Part A.

[36]  D E Ingber,et al.  Cellular control lies in the balance of forces. , 1998, Current opinion in cell biology.

[37]  J. Isner,et al.  Potentiated angiogenic effect of scatter factor/hepatocyte growth factor via induction of vascular endothelial growth factor: the case for paracrine amplification of angiogenesis. , 1998, Circulation.