Microcavitary Hydrogel-Mediating Phase Transfer Cell Culture for Cartilage Tissue Engineering

Abstract

Hydrogels have been widely used as cell-laden vehicles for therapeutic transplantation in regenerative medicine. Although the advantages of biocompatibility and injectability for in situ grafting have made hydrogel a superior candidate in tissue engineering, there remain challenges in long-term efficacy of tissue development using hydrogel, especially when more sophisticated applications are demanded. The major bottleneck lies in environmental constraints for neo-tissue generation in the gel bulk such as proliferation of encapsulated cells (colonies) per se and also accommodation of their endogenously produced extracellular matrices. In this study, we endeavor to develop an innovative tissue engineering system to overcome these drawbacks through a novel microcavitary hydrogel (MCG)-based scaffolding technology and a novel phase transfer cell culture (PTCC) strategy to enable phenotypically bona fide neo-tissue formation in an injectable artificial graft. For this purpose, microspherical cavities are created in cell-encapsulating hydrogel bulk via a retarded dissolution of coencapsulated gelatin microspheres. Based on proliferation and affinity selection, the encapsulated cell colonies adjacent to the gel-cavity interface will spontaneously outgrow the hydrogel phase and sprout into cavities, enabling neo-tissue islets to fill up the voids and further expand throughout the whole system for full tissue regeneration. The design of MCG-PTCC strategy was elicited from an observation of a spontaneous dynamic outgrowth of chondrocytes from the edge of a cell-laden hydrogel construct over prolonged cultivation--a phenomenon named edge flourish. This MCG-PTCC strategy potentially introduce a new application to hydrogels in the field of regenerative medicine through elevation of its role as a cell vehicle to a three-dimensional transplantable growth-guiding platform for further development of newly generated tissues that better fulfill the demanding criteria of scaffolds in therapeutic tissue regeneration.

Recent advances in the use of gelatin in biomedical research

“…Wang's group invented a phase transfer cell culture (PTCC) system, adopting gelatin as delayed dissoluble porogen in either agarose or alginate hydrogels (Gong et al 2010;Su et al 2012). In terms of gelatin as a capsule containing chondrocytes followed by dissolution leaving cavity filled by cells and their ECM, a phenomenon of directed cell proliferation and sprout out as isogenic groups was observed (Ng et al 2012).…”

A preclinical evaluation of an autologous living hyaline-like cartilaginous graft for articular cartilage repair: a pilot study

“…Contrastingly, the chondrocytes used for LhCG fabrication experienced only one passage of monolayer expansion followed by a 35-day culture (PTCC) in 3D alginate-based MCG constructs ( Fig. 1 (b) Step iii~viii) 22 . This strategy allows cells to maintain their differentiated phenotype, as shown through the retention of their characteristic rounded morphology and capacity to synthesize cartilage-specific molecules, which is in agreement with other reports [37][38][39] .…”

“…Additionally, a three-dimensional (3D) environment is crucial to prevent chondrocyte dedifferentiation, a process that causes them to lose their differentiated phenotype. In view of these requirements, a unique methodology that allows chondrocytes to form neocartilage through an intermediate 3D scaffolding system was developed 22 . The transient use of scaffolds prevents the long-term involvement of exogenous scaffold materials, thus preserving tissue purity.…”

“…It is constructed by culturing chondrocytes in a 3D interim scaffolding system made up of removable, non-cell adhesive micro-cavitary hydrogel (MCG) 22 . This environment promotes vigorous chondrocyte growth and abundant cartilage-specific extracellular matrix (ECM) production, hence resulting in maximal maintenance of the desired hyaline phenotype.…”

Supplementation of growth differentiation factor-5 increases proliferation and size of chondrogenic pellets of human umbilical cord-derived perivascular stem cells

“…Therefore, the repair of cartilage injuries is considered one of the most important clinical and scientific challenges. Surgical techniques including multiple drilling, microfracture, and autologous chondrocyte transfer have been developed and clinically applied for the repair of damaged cartilage [3,4]. However, these techniques are faced with problems including limited proliferative and de-differentiation potential of chondrocytes, invasive procedures, and the sacrifice of normal cartilage [5,6].…”