Mesenchymal condensation to mimic the native cell density during cartilage development
L. Mecchi, T. Serra, M. Alini, A. G. Guex
Mesenchymal stem cells can be patterned in a variety of hydrogels.
Cells at high cell condensation have an increased metabolic activity.
qRT-PCR analysis of chondrogenic genes indicates enhanced chondrogenesis of patterned cells.
Our aging society and concomitant cartilage wear, onset of osteoarthritis, pathologies, or injuries have a significant impact on cartilage lining the joints associated with pain and ultimately reduced quality of life.
Different to other tissues within the body, cartilage has very limited self-repair capacity, owned to its avascular nature.
State of the art approaches in tissue engineering and biofabrication might overcome this unsolved clinical challenge by providing in vitro engineered cartilage constructs for implantation.
Pellet cultures are among the gold standard to induce chondrogenesis of mesenchymal stem cells (MSC) in vitro(1).
While this model provides a good basis to evaluate new culture conditions to induce chondrogenesis, this simplified method fails to achieve large-scale constructs of clinically relevant size and the engineered cartilage is often hypertrophic and rich in collagen type X.
Sound-induced MSC condensation has the potential to overcome these limitations and we provide evidence that hyaline cartilage, with an increased collagen II to collagen X ratio is formed in patterned constructs.
Figure 1 A) Multiple technical and experimental repeats are highly important in several experimental settings. With cymatiX, cells can be patterned in standard multi-well culture plates with custom-made frames. Here 12-well plates were used to simultaneously pattern 4 wells at the time, followed by subsequent patterning of additional wells.
B) Representative image of MSC patterned in LUMPA, aDextran-based hydrogel available through mimiX that is gelled upon irradiation at 405nm. Concentric rings are reproducibly formed in different hydrogels, such as Fibrin and Agarose.
Figure 2 MSC were patterned in Agarose and cultured in chondropermissive medium (without TGFβ addition) for 21days. Cell metabolic activity was assessed on day 7, day 14 and day 21 by Cell Titer Blue. Values are normalised to the control group on day1 (randomly dispersed MSC in Agarose).
Figure 3 Expression of cartilage specific genes was quantified after 21 days by qRT- PCR. A) In patterned constructs, the expression of collagen type I, collagen type II and collagen type X was increased compared to the non patterned condition.
In particular collagen type II. B) In patterned cells, the ratio of collagen type II collagen type X is increased. An elevated col II to col X ratio is indicative for the development of hyaline cartilage, as opposed to hypertrophic cartilage. Sound induced MSC condensation is thus a promising method to engineer in vitro cartilage. (N=1 individual experiment/donor with n=3 replicates).
Cell Type: Human Mesenchymal Stem Cells
Labware: mimiX frames for multi-well plates
(1) Johnstone, et al. In vitro chondrogenesis of bone marrow-derived mesenchymal progenitor cells, Exp. Cell Res. 238, 265–272