High throughput generated micro-aggregates of chondrocytes stimulate cartilage formation in vitro and in vivo.
Cell-based cartilage repair strategies such as matrix-induced autologous chondrocyte implantation (MACI) could be improved by enhancing cell performance. We hypothesised that micro-aggregates of chondrocytes generated in high-throughput prior to implantation in a defect could stimulate cartilaginous matrix deposition and remodelling. To address this issue, we designed a micro-mould to enable controlled high-throughput formation of micro-aggregates. Morphology, stability, gene expression profiles and chondrogenic potential of micro-aggregates of human and bovine chondrocytes were evaluated and compared to single-cells cultured in micro-wells and in 3D after encapsulation in Dextran-Tyramine (Dex-TA) hydrogels in vitro and in vivo. We successfully formed micro-aggregates of human and bovine chondrocytes with highly controlled size, stability and viability within 24 hours. Micro-aggregates of 100 cells presented a superior balance in Collagen type I and Collagen type II gene expression over single cells and micro-aggregates of 50 and 200 cells. Matrix metalloproteinases 1, 9 and 13 mRNA levels were decreased in micro-aggregates compared to single-cells. Histological and biochemical analysis demonstrated enhanced matrix deposition in constructs seeded with micro-aggregates cultured in vitro and in vivo, compared to single-cell seeded constructs. Whole genome microarray analysis and single gene expression profiles using human chondrocytes confirmed increased expression of cartilage-related genes when chondrocytes were cultured in micro-aggregates. In conclusion, we succeeded in controlled high-throughput formation of micro-aggregates of chondrocytes. Compared to single cell-seeded constructs, seeding of constructs with micro-aggregates greatly improved neo-cartilage formation. Therefore, micro-aggregation prior to chondrocyte implantation in current MACI procedures, may effectively accelerate hyaline cartilage formation.
Department of Developmental BioEngineering, MIRA Institute for Biomedical Technology and Technical Medicine, Faculty of Science and Technology, University of Twente, Enschede, The Netherlands.
L S
LS
Moreira Teixeira
J C H
JC
Leijten
J
J
Sobral
R
R
Jin
A A
AA
van Apeldoorn
J
J
Feijen
C
C
van Blitterswijk
P J
PJ
Dijkstra
M
M
Karperien
D053674:Aggrecans / Q000378:metabolism; D000818:Animals; D002356:Cartilage / Q000254:growth & development* / Q000378:metabolism; D002417:Cattle; D002449:Cell Aggregation; D017690:Cell Transplantation / Q000379:methods; D019902:Chondrocytes / Q000166:cytology* / Q000378:metabolism; D024042:Collagen Type I / Q000235:genetics / Q000378:metabolism; D024043:Collagen Type II / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation; D057166:High-Throughput Screening Assays; D006801:Humans; D020782:Matrix Metalloproteinases / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008819:Mice, Nude; D046228:Microarray Analysis; D059010:Single-Cell Analysis
C508319:ACAN protein, human; D053674:Aggrecans; D024042:Collagen Type I; D024043:Collagen Type II; D020782:Matrix Metalloproteinases