Bioengineering vascularized tissue constructs using an injectable cell-laden enzymatically crosslinked collagen hydrogel derived from dermal extracellular matrix.
Tissue engineering promises to restore or replace diseased or damaged tissue by creating functional and transplantable artificial tissues. The development of artificial tissues with large dimensions that exceed the diffusion limitation will require nutrients and oxygen to be delivered via perfusion instead of diffusion alone over a short time period. One approach to perfusion is to vascularize engineered tissues, creating a de novo three-dimensional (3D) microvascular network within the tissue construct. This significantly shortens the time of in vivo anastomosis, perfusion and graft integration with the host. In this study, we aimed to develop injectable allogeneic collagen-phenolic hydroxyl (collagen-Ph) hydrogels that are capable of controlling a wide range of physicochemical properties, including stiffness, water absorption and degradability. We tested whether collagen-Ph hydrogels could support the formation of vascularized engineered tissue graft by human blood-derived endothelial colony-forming cells (ECFCs) and bone marrow-derived mesenchymal stem cells (MSC) in vivo. First, we studied the growth of adherent ECFCs and MSCs on or in the hydrogels. To examine the potential formation of functional vascular networks in vivo, a liquid pre-polymer solution of collagen-Ph containing human ECFCs and MSCs, horseradish peroxidase and hydrogen peroxide was injected into the subcutaneous space or abdominal muscle defect of an immunodeficient mouse before gelation, to form a 3D cell-laden polymerized construct. These results showed that extensive human ECFC-lined vascular networks can be generated within 7 days, the engineered vascular density inside collagen-Ph hydrogel constructs can be manipulated through refinable mechanical properties and proteolytic degradability, and these networks can form functional anastomoses with the existing vasculature to further support the survival of host muscle tissues. Finally, optimized conditions of the cell-laden collagen-Ph hydrogel resulted in not only improving the long-term differentiation of transplanted MSCs into mineralized osteoblasts, but the collagen-Ph hydrogel also improved an increased of adipocytes within the vascularized bioengineered tissue in a mouse after 1 month of implantation.


CONCLUSIONS
We reported a method for preparing autologous extracellular matrix scaffolds, murine collagen-Ph hydrogels, and demonstrated its suitability for use in supporting human progenitor cell-based formation of 3D vascular networks in vitro and in vivo. Results showed extensive human vascular networks can be generated within 7 days, engineered vascular density inside collagen-Ph constructs can be manipulated through refinable mechanical properties and proteolytic degradability, and these networks can form functional anastomoses with existing vasculature to further support the survival of host muscle tissues. Moreover, optimized conditions of cell-laden collagen-Ph hydrogel resulted in not only improving the long-term differentiation of transplanted MSCs into mineralized osteoblasts, but the collagen-Ph hydrogel also improved an increased of adipocytes within the vascularized bioengineered tissue in a mouse.
Department of Applied Science, National Hsinchu University of Education, Hsinchu 30014, Taiwan, ROC.
Kuan-Chih
KC
Kuo
Department of Cardiac Surgery, Boston Children's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Ruei-Zeng
RZ
Lin
Department of Applied Science, National Hsinchu University of Education, Hsinchu 30014, Taiwan, ROC.
Han-Wen
HW
Tien
Department of Applied Science, National Hsinchu University of Education, Hsinchu 30014, Taiwan, ROC.
Pei-Yun
PY
Wu
Material and Chemical Research Laboratories, Industrial Technology Research Institute, Hsinchu 31040, Taiwan, ROC.
Yen-Cheng
YC
Li
Department of Cardiac Surgery, Boston Children's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Juan M
JM
Melero-Martin
Department of Applied Science, National Hsinchu University of Education, Hsinchu 30014, Taiwan, ROC. Electronic address: yingchiehchen@gmail.com.
Ying-Chieh
YC
Chen
D062088:Acellular Dermis; D000818:Animals; D021522:Bioartificial Organs; D001808:Blood Vessels / Q000166:cytology / Q000254:growth & development*; D003094:Collagen / Q000737:chemistry*; D003432:Cross-Linking Reagents / Q000737:chemistry; D042783:Endothelial Cells / Q000166:cytology / Q000637:transplantation; D019544:Equipment Failure Analysis; D005109:Extracellular Matrix / Q000737:chemistry; D006735:Horseradish Peroxidase / Q000737:chemistry; D020100:Hydrogels / Q000737:chemistry*; D007267:Injections; D008297:Male; D045164:Mesenchymal Stem Cell Transplantation / Q000295:instrumentation*; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D011474:Prosthesis Design; D012867:Skin / Q000737:chemistry; D023822:Tissue Engineering / Q000295:instrumentation*; D058017:Vascular Grafting / Q000295:instrumentation
D003432:Cross-Linking Reagents; D020100:Hydrogels; D003094:Collagen; D006735:Horseradish Peroxidase
Collagen hydrogels; Tissue engineering; Vascularization