In this study, chondrocytes were encapsulated into an injectable, in situ forming type II collagen/hyaluronic acid (HA) hydrogel cross-linked with poly(ethylene glycol) ether tetrasuccinimidyl glutarate (4SPEG) and supplemented with the transforming growth factor β1 (TGFβ1). The chondrocyte–hydrogel constructs were cultured in vitro for 7 days and studied for cell viability and proliferation, morphology, glycosaminoglycan production, and gene expression. Type II collagen/HA/4SPEG formed a strong and stable hydrogel, and the chondrocytes remained viable during the encapsulation process and for the 7-day culture period. In addition, the encapsulated cells showed spherical morphology characteristic for chondrocytic phenotype. The cells were able to produce glycosaminoglycans into their extracellular matrix, and the gene expression of type II collagen and aggrecan, genes specific for differentiated chondrocytes, increased over time. The results indicate that the studied composite hydrogel with incorporated chondrogenic growth factor TGFβ1 is able to maintain chondrocyte viability and characteristics, and thus, it can be regarded as potential injectable cell delivery vehicle for cartilage tissue engineering.
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The authors would like to acknowledge Hannu Kautiainen (MedCare Ltd., Äänekoski, Finland) for assistance with statistical analysis. This study was supported by the Finnish Funding Agency for Technology and Innovation (projects Novel biomaterials for cartilage tissue engineering and PrinCell II) and by the Emil Aaltonen foundation.
Conflict of interest
All the authors declare that they have no conflict of interest.
The experiments comply with the current laws of Finland.
No animal or human studies were carried out by the authors for this article.
Electronic supplementary material
Below is the link to the electronic supplementary material.
Taqman primer/probe assays for the used genes: type I collagen (COL1A1), type II collagen (COL2A1), aggrecan (ACAN) and glyceralgehyde-3-phosphatase (GAPDH) (PDF 199 kb)
A confocal image of LIVE/DEAD stained chondrocytes in type II collagen/HA/4SPEG/TGFβ1 hydrogel showing the 3D structure of the imaged segment. Living cells are stained green and dead cells red. The image shows distribution of cells inside the segment. 10x magnification (PDF 308 kb)
An animation of a segment of LIVE/DEAD stained chondrocytes in type II collagen/HA/4SPEG/TGFβ1 hydrogel imaged with confocal microscope. Living cells are stained green and dead cells red. The image shows distribution of cells inside the segment. 10x magnification (MPG 63,128 kb)
Viability of chondrocytes in type II collagen/HA/4SPEG/TGFβ1 hydrogel during the 30-day culture period. Viability is reported as alamarBlue fluorescence as a function of time. The results are average values calculated from 2 repeats with 3 parallel samples. The error bars represent standard deviations between repeats (PDF 84 kb)
Phase contrast microscope image of chondrocytes encapsulated in type II collagen/HA/4SPEG/TGFβ1 hydrogel 30 days after encapsulation. 20x magnification, scale bar = 200 μm (PDF 84 kb)
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Kontturi, L., Järvinen, E., Muhonen, V. et al. An injectable, in situ forming type II collagen/hyaluronic acid hydrogel vehicle for chondrocyte delivery in cartilage tissue engineering. Drug Deliv. and Transl. Res. 4, 149–158 (2014). https://doi.org/10.1007/s13346-013-0188-1
- Cartilage tissue engineering
- Injectable hydrogel
- Type II collagen
- Hyaluronic acid
- Transforming growth factor β1