FIG. 3.

Quantitative real-time polymerase chain reaction of chondrogenic marker mRNA expression at days 7, 14, and 21 in the iPSC micromass culture model. COL2A1 encodes two main alternatively spliced forms (IIA and IIB) of this cartilage-specific extracellular matrix protein, type II collagen. In panel A, COL2A1 form IIA is shown; while in panel B, COL2A1 form IIB expression levels are shown. In panel C, the expression of SOX9 is shown and aggrecan is seen in panel D. The two bottom panels demonstrate the expression of later chondrogenic stage markers, COL10A1 of type X collagen (E), and RUNX2 (F). Cultures were grown in mTeSR (mT) or CM, with BMP2 (CMB) or with TGFβ1 (CMT). The patterns of expression over time differ when comparing control iPSCs (solid dark bars) with mutant (hatched bars). Note differences in scale among panels representing fold expression relative to the internal control GAPDH. The bars represent data averaged from experiments performed in triplicate using control (83i-n1 and/or 14i-n6) and TRPV4 (23i-n12 and/or 23i-n14) iPSC clones. Y-axis values represent fold difference. Asterisks indicate statistically significant differences (P<0.05) between TRPV4 compared with control iPSCs in chondrogenic conditions (CM, CMB, and CMT).