In this study scientists are able to create growth plate-like chondrocytes from embryonic stem cells with a specific set of gene inductions.
“Pluripotent embryonic stem cells (ESCs) generate rostral paraxial mesoderm-like progeny in 5-6 days of differentiation induced by Wnt3a and Noggin (Nog). canonical Wnt signaling introduced either by forced expression of activated β-catenin, or the small-molecule inhibitor of Gsk3, CHIR99021, satisfied the need for Wnt3a signaling, and that the small-molecule inhibitor of BMP type I receptors, LDN193189, was able to replace Nog{so abnormal methods are able to replicate the bodies processes}. Mesodermal progeny generated using such small molecules were chondrogenic in vitro, and expressed trunk paraxial mesoderm markers such as Tcf15 and Meox1, and somite markers such as Uncx, but failed to express sclerotome markers such as Pax1. Induction of the osteochondrogenically committed sclerotome from somite requires sonic hedgehog and Nog. Consistently, Pax1 and Bapx1 expression was induced when the isolated paraxial mesodermal progeny were treated with SAG1 (a hedgehog receptor agonist) and LDN193189, then Sox9 expression was induced, leading to cartilaginous nodules and particles in the presence of BMP, indicative of chondrogenesis via sclerotome specification. By contrast, treatment with TGFβ also supported chondrogenesis and stimulated Sox9 expression, but failed to induce the expression of Pax1{Pax1 is upregulated by LSJL} and Bapx1. On ectopic transplantation to immunocompromised mice, the cartilage particles developed under either condition became similarly mineralized and formed pieces of bone with marrow. Thus, the use of small molecules led to the effective generation from ESCs of paraxial mesodermal progeny, and to their further differentiation in vitro through sclerotome specification into growth plate-like chondrocytes, a mechanism resembling in vivo somitic chondrogenesis that is not recapitulated with TGFβ. ”
“The osteochondro-progenitors that develop during embryogenesis are limb bud mesenchyme (derived from lateral plate mesoderm) responsible for limb bone and cartilage generation, sclerotome (from somite/rostral paraxial mesoderm) responsible for rib, vertebral joint, intervertebral disc and vertebral body formation, and ectomesenchyme (from cranial neural crest) responsible for craniofacial bone and cartilage generation.”
“The Flk1−Pdgfrα+ rostral paraxial mesoderm from ESCs consistently show chondrogenic activity in vitro ”
“SAG+LDN (PSL) stimulation during the first 6 days of micromass culture was crucial for inducing Pax1 and Bapx1 expression from the isolated E-cadherin−Flk1−Pdgfrα+ rostral paraxial mesoderm, as was the Shh+Nog stimulation”
“BMP signaling counteracts Shh+Nog and inhibits sclerotome induction”
