Chondrogenic Differentiation of Immortalized Human Mesenchymal Stem Cells on Zirconia Microwell Substrata

Abstract

Human mesenchymal stem cells (hMSCs) that can differentiate into chondrocytes are a potential autologous cell source for repair of damaged tissue. Current methods usually induce the formation of all three chondrocyte phenotypes, hyaline, fibrous, and elastic, without the ability to selectively induce only one of them. By controlling the size of hMSC cell clusters, it may be possible to direct differentiation more uniformly toward hyaline chondrocytes. We designed new cell culture platforms containing microwells of different diameters. The platforms and wells were composed of a zirconia ceramics substratum. hMSCs briefly adhered to the substratum before releasing and entering the microwells. The physical restraints imposed by the microwells enabled hMSC clusters to homogenously differentiate into hyaline chondrocyte-like cells. Chondrogenic aggregates in microwells expressed the hyaline chondrocyte-specific genes Col II, aggrecan (ACAN), and cartilage oligomeric protein (COMP). The cultures also produced hyaline chondrocyte-specific matrix proteins Col II and ACAN homogenously throughout the aggregates. In contrast, chondrogenesis in pellet cultures was heterogeneous with the expression of nonhyaline chondrocyte genes CD105, Col X, and Col I. In these pellet cultures, hyaline and nonhyaline chondrocyte-specific matrix proteins were distributed heterogeneously. Thus, this novel ceramic microwell substratum technology efficiently directed the differentiation of hyaline chondrocyte-like cells from hMSCs. These results indicate that there is a close relationship between hMSC cluster size regulation in the microwells and differentiation tendency. This microwell culture differentiation method will provide a valuable experimental system for both experimental and potential clinical studies.

Redifferentiation of High-Throughput Generated Fibrochondrocyte Micro-Aggregates: Impact of Low Oxygen Tension

“…5 ). In contrast, conventional pellet culture is reported to lead to heterogeneous matrix deposition [Mackay et al, 1998;Babur et al, 2013;Kitagawa et al, 2013], but it remains a widely accepted gold standard for studying redifferentiation.…”

Proliferation and osteogenic differentiation of human mesenchymal stem cells on zirconia and titanium with different surface topography

“…It was reported that nanostructure surface enhanced the proliferation and osteogenic differentiation of the MSCs on titanium [23][24][25][26] . However, few studies were reported about the hMSCs behavior on the TZP with different surfaces 27,28) . We assumed that the microand nano-topography may have suitable configurations for hMSCs behavior.…”

Rapid formation of multicellular spheroids in double-emulsion droplets with controllable microenvironment

“…For microscale tissue engineering, the spheroid size and microenvironmental cues such as extracellular matrix (ECM) play an important role in directing stem cell behavior1213. The size of mesenchymal stem cell (MSC) spheroid has been shown to influence its differentiation potential, with smaller spheroid directing more homogeneous chondrogenic differentiation towards hyaline chondrocytes whereas larger pellet producing more heterogeneous tissue14. Further, external microenvironmental cues in the form of matrix scaffold are often applied to direct stem cell differentiation.…”