Co-Culture of Human Endothelial Cells and Foreskin Fibroblasts on 3D Silk-Fibrin Scaffolds Supports Vascularization.

A successful strategy to enhance the in vivo survival of engineered tissues would be to prevascularize them. In this study, fabricated silk fibroin scaffolds from mulberry and non-mulberry silkworms are investigated and compared for supporting the co-culture of human umbilical vein endothelial cells and human foreskin fibroblasts. Scaffolds are cytocompatible and when combined with fibrin gel support capillary-like structure formation. Density and interconnectivity of the formed structures are found to be better in mulberry scaffolds. ELISA shows that levels of vascular endothelial growth factor (VEGF) released in co-cultures with fibrin gel are significantly higher than in co-cultures without fibrin gel. RT PCR shows an increase in VEGFR2 expression in mulberry scaffolds indicating these scaffolds combined with fibrin provide a suitable microenvironment for the development of capillary-like structures.

[1]  Shaker A. Mousa,et al.  Fibrin and Collagen Differentially but Synergistically Regulate Sprout Angiogenesis of Human Dermal Microvascular Endothelial Cells in 3-Dimensional Matrix , 2013, International journal of cell biology.

[2]  David L. Kaplan,et al.  New Opportunities for an Ancient Material , 2010, Science.

[3]  Pieter Koolwijk,et al.  Influence of fibrin structure on the formation and maintenance of capillary-like tubules by human microvascular endothelial cells , 2004, Angiogenesis.

[4]  A. Sahni,et al.  The VE‐cadherin binding domain of fibrinogen induces endothelial barrier permeability and enhances transendothelial migration of malignant breast epithelial cells , 2009, International journal of cancer.

[5]  H. Hutchings,et al.  Signal relays in the VEGF system. , 1999, Frontiers in bioscience : a journal and virtual library.

[6]  B. Mandal,et al.  Implication of silk film RGD availability and surface roughness on cytoskeletal organization and proliferation of primary rat bone marrow cells. , 2010, Tissue engineering. Part A.

[7]  Zhao Xie,et al.  Effects of Initial Cell Density and Hydrodynamic Culture on Osteogenic Activity of Tissue-Engineered Bone Grafts , 2013, PloS one.

[8]  G. G. Stokes "J." , 1890, The New Yale Book of Quotations.

[9]  Joydip Kundu,et al.  Invited review nonmulberry silk biopolymers. , 2012, Biopolymers.

[10]  Yubo Fan,et al.  Electrospun sulfated silk fibroin nanofibrous scaffolds for vascular tissue engineering. , 2011, Biomaterials.

[11]  Steven C George,et al.  Prevascularization of a fibrin-based tissue construct accelerates the formation of functional anastomosis with host vasculature. , 2009, Tissue engineering. Part A.

[12]  J. Feder,et al.  Fibrin‐enhanced endothelial cell organization , 1985, Journal of cellular physiology.

[13]  D. Wisser,et al.  Skin replacement with a collagen based dermal substitute, autologous keratinocytes and fibroblasts in burn trauma. , 2003, Burns : journal of the International Society for Burn Injuries.

[14]  L. Claesson‐Welsh,et al.  Signal transduction by vascular endothelial growth factor receptors. , 2011, The Biochemical journal.

[15]  F. O'Brien,et al.  Towards in vitro vascularisation of collagen-GAG scaffolds. , 2011, European cells & materials.

[16]  Yan‐Yeung Luk,et al.  Enhanced cell adhesion and mature intracellular structure promoted by squaramide-based RGD mimics on bioinert surfaces. , 2013, Bioorganic & medicinal chemistry.

[17]  Christian Mühlfeld,et al.  Silk protein fibroin from Antheraea mylitta for cardiac tissue engineering. , 2012, Biomaterials.

[18]  A. Perets,et al.  Vascular endothelial growth factor-releasing scaffolds enhance vascularization and engraftment of hepatocytes transplanted on liver lobes. , 2005, Tissue engineering.

[19]  S. Hsu,et al.  The effect of dynamic culture conditions on endothelial cell seeding and retention on small diameter polyurethane vascular grafts. , 2005, Medical engineering & physics.

[20]  GeunHyung Kim,et al.  Coaxial structured collagen–alginate scaffolds: fabrication, physical properties, and biomedical application for skin tissue regeneration , 2011 .

[21]  P. D’Amore,et al.  Cellular interactions in vascular growth and differentiation. , 2001, International review of cytology.

[22]  M. N. Nakatsu,et al.  The requirement for fibroblasts in angiogenesis: fibroblast-derived matrix proteins are essential for endothelial cell lumen formation , 2011, Molecular biology of the cell.

[23]  Asha Mathews,et al.  Vascular tissue construction on poly(ε‐caprolactone) scaffolds by dynamic endothelial cell seeding: effect of pore size , 2012, Journal of tissue engineering and regenerative medicine.

[24]  Lucie Germain,et al.  In vitro reconstruction of a human capillary‐like network in a tissue‐engineered skin equivalent , 1998, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[25]  Robert Langer,et al.  Tissue engineering: the design and fabrication of living replacement devices for surgical reconstruction and transplantation , 1999, The Lancet.

[26]  S. Majuru,et al.  Design and Characterization of a Silk-Fibroin-Based Drug Delivery Platform Using Naproxen as a Model Drug , 2012, Journal of drug delivery.

[27]  C James Kirkpatrick,et al.  The rapid anastomosis between prevascularized networks on silk fibroin scaffolds generated in vitro with cocultures of human microvascular endothelial and osteoblast cells and the host vasculature. , 2010, Biomaterials.

[28]  M. Engelse,et al.  Single and combined effects of αvβ3- and α5β1-integrins on capillary tube formation in a human fibrinous matrix , 2009, Angiogenesis.

[29]  David L Kaplan,et al.  In vitro evaluation of electrospun silk fibroin scaffolds for vascular cell growth. , 2008, Biomaterials.

[30]  B J Messmer,et al.  Tissue engineering: complete autologous valve conduit--a new moulding technique. , 2001, The Thoracic and cardiovascular surgeon.

[31]  A. Perets,et al.  Enhancing the vascularization of three-dimensional porous alginate scaffolds by incorporating controlled release basic fibroblast growth factor microspheres. , 2003, Journal of biomedical materials research. Part A.

[32]  Bart Landuyt,et al.  Vascular Endothelial Growth Factor and Angiogenesis , 2004, Pharmacological Reviews.

[33]  C. Tatu,et al.  Tumour-associated fibroblasts and mesenchymal stem cells: more similarities than differences , 2010, Journal of cellular and molecular medicine.

[34]  P. Carmeliet,et al.  Angiogenesis in cancer and other diseases , 2000, Nature.

[35]  Adrian Ranga,et al.  Heparin-binding domain of fibrin(ogen) binds growth factors and promotes tissue repair when incorporated within a synthetic matrix , 2013, Proceedings of the National Academy of Sciences.

[36]  C. Sorenson,et al.  PECAM-1 regulates proangiogenic properties of endothelial cells through modulation of cell-cell and cell-matrix interactions. , 2010, American journal of physiology. Cell physiology.

[37]  Stephanie Lucas,et al.  Fibroblast growth factor-1 (FGF-1) loaded microbeads enhance local capillary neovascularization. , 2010, The Journal of surgical research.

[38]  Smadar Cohen,et al.  The promotion of in vitro vessel-like organization of endothelial cells in magnetically responsive alginate scaffolds. , 2012, Biomaterials.

[39]  C. Lewis,et al.  Alphastatin, a 24-amino acid fragment of human fibrinogen, is a potent new inhibitor of activated endothelial cells in vitro and in vivo. , 2004, Blood.

[40]  Joydip Kundu,et al.  Mulberry non-engineered silk gland protein vis-à-vis silk cocoon protein engineered by silkworms as biomaterial matrices , 2008, Journal of materials science. Materials in medicine.

[41]  Thomas Schmitz-Rode,et al.  Tranexamic acid--an alternative to aprotinin in fibrin-based cardiovascular tissue engineering. , 2009, Tissue engineering. Part A.

[42]  C. Legrand,et al.  Platelets, thrombospondin-1 and human dermal fibroblasts cooperate for stimulation of endothelial cell tubulogenesis through VEGF and PAI-1 regulation. , 2007, Experimental cell research.

[43]  L. Krishnan,et al.  Effect of passage number and matrix characteristics on differentiation of endothelial cells cultured for tissue engineering. , 2005, Biomaterials.

[44]  P. Koolwijk,et al.  Role of Fibrin Matrix in Angiogenesis , 2001, Annals of the New York Academy of Sciences.

[45]  Biman B Mandal,et al.  A novel method for dissolution and stabilization of non‐mulberry silk gland protein fibroin using anionic surfactant sodium dodecyl sulfate , 2008, Biotechnology and bioengineering.