FIG. 2.

Alcian blue and Alizarin red tissue-specific staining of control and TRPV4-iPSC micromass cultures and neonatal cartilage from control human and TRPV4 patient samples. (A, B) Phase-contrast microscopy images (4×) of iPSC lines grown in micromass culture and stained with Alcian blue for sulfated proteoglycans of cartilage. Micromasses were grown in control iPSC medium (mTeSR; mT), chondrogenic medium (CM), or in CM supplemented with 100 ng/mL of BMP2 (CMB) or with 10 ng/mL of TGFβ1 (CMT) for 7, 14, or 21 days before whole mount staining. One representative clone for each control (83i-n1) and SD TRPV4 mutant (23i-n14) iPSC line is shown. Focal stains are indicated by arrows (B). Experiments were repeated at least thrice, using the control (14i-n6) and SD TRPV4 mutant (23i-n12). (C, D) Phase-contrast images of control neonatal human cartilage sections (C) and SD TRPV4 patient cartilage (D) stained with Alizarin red for developing bone or Alcian blue for cartilage. Boxed outline areas shown at 20× in adjacent photos (C, D). BMP2, bone morphogenetic protein 2; TGFβ1, transforming growth factor beta 1. Color images available online at www.liebertpub.com/scd