Chemically crosslinked alginate porous microcarriers modified with bioactive molecule for expansion of human hepatocellular carcinoma cells
Chunge Li
Tianjin Key Laboratory of Composite and Functional Materials, School of Materials Science and Engineering, Tianjin University, Tianjin, 300072 China
Search for more papers by this authorShuang Zhao
Tianjin Key Laboratory of Composite and Functional Materials, School of Materials Science and Engineering, Tianjin University, Tianjin, 300072 China
Search for more papers by this authorYanyan Zhao
Tianjin Key Laboratory of Composite and Functional Materials, School of Materials Science and Engineering, Tianjin University, Tianjin, 300072 China
Search for more papers by this authorYufeng Qian
Department of Chemistry and Biochemistry, University of Texas at Austin, Austin, Texas, 78712
Search for more papers by this authorCorresponding Author
Junjie Li
Tianjin Key Laboratory of Composite and Functional Materials, School of Materials Science and Engineering, Tianjin University, Tianjin, 300072 China
Department of Advanced Interdisciplinary Studies, Institute of Basic Medical Sciences and Tissue Engineering Research Center, Academy of Military Medical Sciences, Beijing, 100850 China
Correspondence to: Y. Yin (e-mail: [email protected]) or J. Li (e-mail: [email protected] or [email protected])Search for more papers by this authorCorresponding Author
Yuji Yin
Tianjin Key Laboratory of Composite and Functional Materials, School of Materials Science and Engineering, Tianjin University, Tianjin, 300072 China
Correspondence to: Y. Yin (e-mail: [email protected]) or J. Li (e-mail: [email protected] or [email protected])Search for more papers by this authorChunge Li
Tianjin Key Laboratory of Composite and Functional Materials, School of Materials Science and Engineering, Tianjin University, Tianjin, 300072 China
Search for more papers by this authorShuang Zhao
Tianjin Key Laboratory of Composite and Functional Materials, School of Materials Science and Engineering, Tianjin University, Tianjin, 300072 China
Search for more papers by this authorYanyan Zhao
Tianjin Key Laboratory of Composite and Functional Materials, School of Materials Science and Engineering, Tianjin University, Tianjin, 300072 China
Search for more papers by this authorYufeng Qian
Department of Chemistry and Biochemistry, University of Texas at Austin, Austin, Texas, 78712
Search for more papers by this authorCorresponding Author
Junjie Li
Tianjin Key Laboratory of Composite and Functional Materials, School of Materials Science and Engineering, Tianjin University, Tianjin, 300072 China
Department of Advanced Interdisciplinary Studies, Institute of Basic Medical Sciences and Tissue Engineering Research Center, Academy of Military Medical Sciences, Beijing, 100850 China
Correspondence to: Y. Yin (e-mail: [email protected]) or J. Li (e-mail: [email protected] or [email protected])Search for more papers by this authorCorresponding Author
Yuji Yin
Tianjin Key Laboratory of Composite and Functional Materials, School of Materials Science and Engineering, Tianjin University, Tianjin, 300072 China
Correspondence to: Y. Yin (e-mail: [email protected]) or J. Li (e-mail: [email protected] or [email protected])Search for more papers by this authorAbstract
Microcarrier is an essential matrix for the large-scale culture of anchorage-dependent cells. In this study, chemical cross-linked alginate porous microcarriers (AMC) were prepared using microemulsion and freeze-drying technology. Moreover, chitosan was coated on the surface of microcarriers (AMC-CS) via electrostatic interactions to improve the mechanical strength. The size of AMC can be modulated through adjusting the concentration of alginate, amount of dispersant and stirring rate. The surface chemical characteristics and morphology of AMC-CS were evaluated by Fourier transformed infrared, X-ray photoelectron spectroscopy, and scanning electron microscope. Fibronectin (Fn) or heparin/basic fibroblast growth factor (bFGF) was then immobilized on the surface of microcarriers via layer-by-layer technology to improve the cytocompatibility. Our data suggested that the size of AMC can be accurately modulated from 90 μm to 900 μm with a narrow size distribution. Micropore structures of AMC-CS were relatively disordered and the pore size ranged between 20 μm and 100 μm. Using AMC after modified with Fn or bFGF as the cell expansion microcarriers, we showed that the proliferation rates of HepG2 cells increased significantly, reaching to more than 30-fold of cell expansion after 10 days of culture, with minor cellular damage caused by the microcarriers. Moreover, the AMC microcarriers modified with Fn or bFGF can increase albumin secretion of HepG2. We suggest that our new modified AMC-based microcarriers will be an attractive candidate for the large-scale cell culture of therapeutic cells. © 2014 Wiley Periodicals, Inc. J Biomed Mater Res Part B, 102B: 1648–1658, 2014.
REFERENCES
- 1 Chen A, Reuveny S, Oh S. Application of human mesenchymal and pluripotent stem cell microcarrier cultures in cellular therapy: Achievements and future direction. Biotechnol Adv 2013; 31: 1032–1046.
- 2 Bardy J, Chen AK, Lim YM, Wu S, Wei S, Weiping H, Chan K, Reuveny S, Steve KW. Microcarrier suspension cultures for high-density expansion and differentiation of human pluripotent stem cells to neural progenitor cells. Tissue Eng Part C: Methods 2012; 19: 166–180.
- 3 Van Wezel A. Growth of cell-strains and primary cells on micro-carriers in homogeneous culture. Nature 1967; 216: 64–65.
- 4 Barrias CC, Ribeiro CC, Lamghari M, Miranda CS, Barbosa MA. Proliferation, activity, and osteogenic differentiation of bone marrow stromal cells cultured on calcium titanium phosphate microspheres. J Biomed Mater Res Part A 2005; 72A: 57–66.
- 5 Cer E, Gürpınar ÖA, Onur MA, Tuncel A. Polyethylene glycol-based cationically charged hydrogel beads as a new microcarrier for cell culture. J Biomed Mater Res Part B: Appl Biomater 2007; 80B: 406–414.
- 6 Borg DJ, Dawson RA, Leavesley DI, Hutmacher DW, Upton Z, Malda J. Functional and phenotypic characterization of human keratinocytes expanded in microcarrier culture. J Biomed Mater Res Part A 2009; 88A: 184–194.
- 7 Overstreet M, Sohrabi A, Polotsky A, Hungerford DS, Frondoza CG. Collagen microcarrier spinner culture promotes osteoblast proliferation and synthesis of matrix proteins. In Vitro Cell Dev-An 2003; 39: 228–234.
- 8 García-Giralt N, Cruz DG, Nogues X, Ivirico JE, Ribelles JG. Chitosan microparticles for “in vitro” 3D culture of human chondrocytes. RSC Adv 2013; 3: 6362–6368.
- 9 Wu X-B, Peng C-H, Huang F, Kuang J, Yu S-L, Dong Y-D, Han B-S. Preparation and characterization of chitosan porous microcarriers for hepatocyte culture. Hepatob Pancreat Dis Int 2011; 10: 509–515.
- 10 Chen AK-L, Chen X, Choo ABH, Reuveny S, Oh SKW. Critical microcarrier properties affecting the expansion of undifferentiated human embryonic stem cells. Stem Cell Res 2011; 7: 97–111.
- 11 Zhou L, Kong J, Zhuang Y, Chu J, Zhang S, Guo M. Ex vivo expansion of bone marrow mesenchymal stem cells using microcarrier beads in a stirred bioreactor. Biotechnol Bioproc Eng 2013; 18: 173–184.
- 12 Ambrosch K, Manhardt M, Loth T, Bernhardt R, Schulz-Siegmund M, Hacker MC. Open porous microscaffolds for cellular and tissue engineering by lipid templating. Acta Biomater 2012; 8: 1303–1315.
- 13 Moshaverinia A, Chen C, Akiyama K, Ansari S, Xu X, Chee WW, Schricker SR, Shi S. Alginate hydrogel as a promising scaffold for dental-derived stem cells: An in vitro study. J Mater Sci: Mater Med 2012; 23: 3041–3051.
- 14 Wilson JL, McDevitt TC. Stem cell microencapsulation for phenotypic control, bioprocessing, and transplantation. Biotechnol Bioeng 2013; 110: 667–682.
- 15 Santos E, Pedraz J, Hernández R, Orive G. Therapeutic cell encapsulation: Ten steps towards clinical translation. J Control Release 2013; 170: 1–14.
- 16 Aizawa Y, Owen SC, Shoichet MS. Polymers used to influence cell fate in 3D geometry: New trends. Prog Polym Sci 2012; 37: 645–658.
- 17 Hunt N, Smith AM, Gbureck U, Shelton R, Grover L. Encapsulation of fibroblasts causes accelerated alginate hydrogel degradation. Acta Biomater 2010; 6: 3649–3656.
- 18 Tan R, She Z, Wang M, Fang Z, Liu Y, Feng Q. Thermo-sensitive alginate-based injectable hydrogel for tissue engineering. Carbohyd Polym 2012; 87: 1515–1521.
- 19 Bidarra SJ, Barrias CC, Fonseca KB, Barbosa MA, Soares RA, Granja PL. Injectable in situ crosslinkable RGD-modified alginate matrix for endothelial cells delivery. Biomaterials 2011; 32: 7897–7904.
- 20 Zhao YY, Gao SQ, Zhao S, Li Y, Cheng L, Li JJ, Yin YJ. Synthesis and characterization of disulfide-crosslinked alginate hydrogel scaffolds. Mater Sci Eng: C 2012; 32: 2153–2162.
- 21 Hewitt CJ, Lee K, Nienow AW, Thomas RJ, Smith M, Thomas CR. Expansion of human mesenchymal stem cells on microcarriers. Biotechnol Lett 2011; 33: 2325–2335.
- 22 Wang C, Gong Y, Lin Y, Shen J, Wang D-A. A novel gellan gel-based microcarrier for anchorage-dependent cell delivery. Acta Biomater 2008; 4: 1226–1234.
- 23 Zheng J, Xie H, Yu W, Tan M, Gong F, Liu X, Wang F, Lv G, Liu W, Zheng G, Yang Y, Xie W, Ma X. Enhancement of surface graft density of MPEG on alginate/chitosan hydrogel microcapsules for protein repellency. Langmuir 2012; 28: 13261–13273.
- 24 Ai H, Meng H, Ichinose I, Jones SA, Mills DK, Lvov YM, Wang F, Lv G, Liu W, Zheng G, Yang Y, Xie W, Ma X. Biocompatibility of layer-by-layer self-assembled nanofilm on silicone rubber for neurons. J Neurosci Meth 2003; 128: 1–8.
- 25 Shen H, Hu X, Yang F, Bei J, Wang S. Cell affinity for bFGF immobilized heparin-containing poly (lactide-co-glycolide) scaffolds. Biomaterials 2011; 32: 3404–3412.
- 26 Wittmer CR, Phelps JA, Lepus CM, Saltzman WM, Harding MJ, Van Tassel PR. Multilayer nanofilms as substrates for hepatocellular applications. Biomaterials 2008; 29: 4082–4090.
- 27 Xiang Q, Xiao J, Zhang H, Zhang X, Lu M, Zhang H, Su Z, Zhao W, Lin C, Huang Y, Li X. Preparation and characterisation of bFGF-encapsulated liposomes and evaluation of wound-healing activities in the rat. Burns 2011; 37: 886–895.
- 28 Li JJ, Sun H, Zhang R, Li RY, Yin YJ, Wang H, Liu Y, Yao F, Yao K. Modulation of mesenchymal stem cells behaviors by chitosan/gelatin/pectin network films. J Biomed Mater Res Part B: Appl Biomater 2010; 95B: 308–319.
- 29 Gümüşderelioğlu M, Betül Kaya F, Beşkardeş IG. Comparison of epithelial and fibroblastic cell behavior on nano/micro-topographic PCL membranes produced by crystallinity control. J Colloid Interf Sci 2011; 358: 444–453.
- 30 Wang S, Zhang Y, Wang H, Dong Z. Preparation, characterization and biocompatibility of electrospinning heparin-modified silk fibroin nanofibers. Int J Biol Macromol 2011; 48: 345–353.
- 31 Senkevich JJ, Yang GR, Lu TM. Thermal stability of mercaptan terminated self-assembled multilayer films on SiO2 surfaces. Colloids and Surfaces A: Physicochem Eng Aspects 2002; 207: 139–145.
- 32 Bierwolf J, Lutgehetmann M, Deichmann S, Erbes J, Volz T, Dandri M, Cohen S, Nasan B, Pollok JM. Primary human hepatocytes from metabolic-disordered children recreate highly differentiated liver-tissue-like spheroids on alginate scaffolds. Tissue Eng Part A 2012; 18: 1443–1453.
- 33 Li JJ, Shu Y, Hao T, Wang Y, Qian YF, Duan CM, Sun HY, Lin QX, Wang CY. A chitosan-glutathione based injectable hydrogel for suppression of oxidative stress damage in cardiomyocytes. Biomaterials 2013; 34: 9071–9081.
- 34 Mao JS, Cui YL, Wang XH, Sun Y, Yin YJ, Zhao HM, Yao KD. A preliminary study on chitosan and gelatin polyelectrolyte complex cytocompatibility by cell cycle and apoptosis analysis. Biomaterials 2004; 25: 3973–3981.
- 35 Dolatshahi-Pirouz A, Jensen T, Kraft DC, Foss M, Kingshott P, Hansen JL, Larsen AN, Chevallier J, Besenbacher F. Fibronectin adsorption, cell adhesion, and proliferation on nanostructured tantalum surfaces. ACS Nano 2010; 4: 2874–2882.
- 36 Jin C, Ren LF Ding HZ, Shi GS, Lin HS, Zhang F. Enhanced attachment, proliferation, and differentiation of human gingival fibroblasts on titanium surface modified with biomolecules. J Biomed Mater Res Part B: Appl Biomater 2012; 100B: 2167–2177.
- 37 Lu G, Zhu L, Kong L, Zhang L, Gong Y, Zhao N, Zhang X. Porous chitosan microcarriers for large scale cultivation of cells for tissue engineering: Fabrication and evaluation. Tsinghua Sci Technol 2006; 11: 427–432.
- 38 Ha CE, Ha JS, Theriault AG, Bhagavan NV. Effects of statins on the secretion of human serum albumin in cultured HepG2 cells. J Biomed Sci 2009; 16: 32.
- 39 Taguchi T, Rao Z, Ito M, Matsuda M. Induced albumin secretion from HepG2 spheroids prepared using poly (ethylene glycol) derivative with oleyl groups. J Mater Sci: Mater Med 2011; 22: 2357–2363.
