Cell-laden hydrogels for osteochondral and cartilage tissue engineering

Abstract

Despite tremendous advances in the field of regenerative medicine, it still remains challenging to repair the osteochondral interface and full-thickness articular cartilage defects. This inefficiency largely originates from the lack of appropriate tissue-engineered artificial matrices that can replace the damaged regions and promote tissue regeneration. Hydrogels are emerging as a promising class of biomaterials for both soft and hard tissue regeneration. Many critical properties of hydrogels, such as mechanical stiffness, elasticity, water content, bioactivity, and degradation, can be rationally designed and conveniently tuned by proper selection of the material and chemistry. Particularly, advances in the development of cell-laden hydrogels have opened up new possibilities for cell therapy. In this article, we describe the problems encountered in this field and review recent progress in designing cell-hydrogel hybrid constructs for promoting the reestablishment of osteochondral/cartilage tissues. Our focus centers on the effects of hydrogel type, cell type, and growth factor delivery on achieving efficient chondrogenesis and osteogenesis. We give our perspective on developing next-generation matrices with improved physical and biological properties for osteochondral/cartilage tissue engineering. We also highlight recent advances in biomanufacturing technologies (e.g. molding, bioprinting, and assembly) for fabrication of hydrogel-based osteochondral and cartilage constructs with complex compositions and microarchitectures to mimic their native counterparts.

Statement of significance: Despite tremendous advances in the field of regenerative medicine, it still remains challenging to repair the osteochondral interface and full-thickness articular cartilage defects. This inefficiency largely originates from the lack of appropriate tissue-engineered biomaterials that replace the damaged regions and promote tissue regeneration. Cell-laden hydrogel systems have emerged as a promising tissue-engineering platform to address this issue. In this article, we describe the fundamental problems encountered in this field and review recent progress in designing cell-hydrogel constructs for promoting the reestablishment of osteochondral/cartilage tissues. Our focus centers on the effects of hydrogel composition, cell type, and growth factor delivery on achieving efficient chondrogenesis and osteogenesis. We give our perspective on developing next-generation hydrogel/inorganic particle/stem cell hybrid composites with improved physical and biological properties for osteochondral/cartilage tissue engineering. We also highlight recent advances in biomanufacturing and bioengineering technologies (e.g. 3D bioprinting) for fabrication of hydrogel-based osteochondral and cartilage constructs.

Keywords: Cartilage tissue engineering; Cell-laden hydrogels; Osteochondral tissue engineering; Stem cells.

Fig. 1

Tissue engineering strategy for treatment of osteochondral interface and full-thickness cartilage defects with cell-laden hydrogel constructs.

Fig. 2

Chondrocytes cultured in various hydrogels for OTE and CTE. (A) Fluorescence microscopy images showing the chondrocyte morphology inside chitosan-based hydrogels after 3 and 14 days in culture. The upper panels show low-magnification views and lower panels display the close-ups. Scanning electron microscopy (SEM) image shows the morphology of a single chondrocyte. Reproduced with permission [50] Copyright © 2009 Elsevier B.V. (B) Cartilage matrix generation of chondrocytes encapsulated in PEG-LA hydrogels (28 days). (i) Proteoglycan deposition: Chondroitin-6-sulfate (red), aggrecan (red), link protein (red), and ell nuclei (blue). (ii) Collagen deposition: collagen II (green), collagen VI (green), decorin (red), and cell nuclei (blue). Reproduced with permission [194] Copyright © 2011 The Association of Bone and Joint Surgeons. (C) Chondrocytes encapsulated in cartilaginous ECM-modified chitosan hydrogels (MeGC = methacrylated glycol chitosan; RF = riboflavin; VBL = visible blue light). (i) Chondrocyte encapsulation and expression of cartilage-related proteins. (ii) Interior microstructure of various chitosan based hydrogels. (iii) Cell adhesion onto the hydrogels. Reproduced with permission [51] Copyright © 2014 American Chemical Society.

Fig. 3

MSCs cultured in different hydrogels for OTE and CTE. Histological and immunohistochemical staining results showing the cartilaginous and osseous ECM formation after an 8-week culture of MSCs in alginate, chitosan, and fibrin hydrogels. Constructs are stained for aggrecan (Alcian blue), collagen type II, and calcium (Alizarin red). Reproduced with the permission [101] Copyright © 2015 Elsevier.

Fig. 4

Chondrogenesis and osteogenesis of chondrocytes and MSCs in multi-layered hydrogel osteochondral constructs. (A) Co-culture of chondrocytes and MSCs in PCL-PEG composite hydrogels. (i) Schematic of 3D encapsulation. (ii) In vitro cartilage-related biomarker expression (aggrecan stained by Safranin O) at 4 weeks. (iii) Histological results showing in vivo cartilage formation. Reproduced with permission [196] Copyright © 2013 John Wiley & Sons, Inc. (B) A structured bilayered co-culture of chondrocytes and MSCs in agarose hydrogels for chondrogenesis and endochondral ossification. Alcian blue and Alizarin red staining are used to characterize cartilage ECM formation. Alizarin red staining and micro-computed tomography (micro-CT) scanning are employed to examine bone ECM formation. (i) After a 49-day culture in chondrogenic medium. (ii) After a 21-day culture in chondrogenic medium and a 28-day culture in hypertrophic medium with β -glycerophosphate supplement. (iii) After a 21-day culture in chondrogenic medium and a 28-day subcutaneous implantation in nude mice. Reproduced with permission [218] Copyright © 2013 Elsevier. (C) Mechanical loading regulated MSCs differentiation that were encapsulated in layered PEG hydrogel for controlled chondrogenesis and osteogenesis. (i) Finite elemental modeling results for multi-layered hydrogel constructs under compression conditions. Negative values mean compressive strain. (ii) Expression of cartilage and bone biomarkers in hydrogel layers with different mechanical loading. Green indicates collagen II or I, and blue indicates nuclei. (iii, iv) Quantitative study of chondrogenic- and osteogenic-differentiated cells induced by mechanical property change. Reproduced with permission [182] Copyright © 2015 Elsevier.

Fig. 5

Hybrid hydrogel composites with inorganic particles for OTE and CTE. (A) MSCs encapsulation in silicate-hydrogel nanocomposites. Reproduced with permission [213] Copyright © 2015 American Chemical Society. (B) Osteogenic differentiation of bone morrow derived MSCs induced by nanosilicate platelets. Reproduced with permission [270] Copyright © 2013 John Wiley & Sons, Inc. (C) Hydrogel composites with microsilicate particles as osteogenic inducers. (i) Optical photomicrographs of human dermal fibroblasts (HDFs) cultured on silicate-incorporated hydrogels. (ii) ALP activity of MSCs cultured with different ionic extracts from inorganic particles for 7 days. Reproduced with permission [273] Copyright © 2013 Elsevier. (D) 3D printing of free-standing GelMA-nanosilicate composite hydrogel constructs. Reproduced with permission [213] Copyright © 2015 American Chemical Society.

Fig. 6

3D printing and assembling of cell-laden hydrogel constructs for OTE and CTE. (A) Microscale mixing and 3D printing for fabrication of gradient constructs. Reproduced with permission [282] Copyright © 2015 National Academy of Science. (B) 3D bioprinting of human-scale tissue constructs with structural integrity. Reproduced with permission [283] Copyright © 2016 Nature America, Inc. (C) A 3D puzzle assembly strategy for fabrication of large engineered hydrogel based cartilage and osteochondral tissue constructs. Reproduced with permission [286] Copyright © 2016 Elsevier.