Developmental analysis of Limb development after temporal blocking of Hedgehog signaling

Abstract

[한글]

[영문]The vertebrate limb skeleton develops through the endochondral bone formation, which involves in the substitution a cartilage primordium with the calcified bone matrix. The cartilage formation, called chondrogenesis, is crucial in determining the shape and size of definitive bones in vertebrate. During mouse limb chondrogenesis, mesenchymal progenitor cells are condensed differentiates into chondrocytes to form the cartilage primordium, and chondrocytes undergo sequential proliferation and differentiation along with secretion of extracellular matrix. Many studies have reported extensively the molecular mechanism on the endochondral bone formation rather than chondrogenesis. Especially, Indian hedgehog (Ihh) signaling is known to control both hypertrophy of chondrocytes and bone replacement which is particularly important in postnatal endochondral bone formation. In this study, maternal transfer of 5E1 (a hedgehog-blocking antibody) to E12 mouse embryo causes an attenuation of the Indian hedgehog activity, and causes malformation of the mouse limb. We analysed genetic relationship during chondrogenesis in limb buds with mRNA microarray. Furthermore, we treated exogenous IHH protein into mesenchymal cells of limb bud in the micromass culture system. As a result, we found some genes interacting with Ihh ; Glo1, Dkk4, Mapk8, Ltbp3, Col1a2, Wasl, Dmkn, Flnc, Eltd1, Egfl7, Col22a1, Krt1, Col22a1, Krt1, Col19a1, Krt10, Hspa8, Fbxo2, Ocrl, map3k4, Garnl4, Fgf7, Krt4, Klf7 were up-regulated, and Hhip, Hoxd13, Casp1, Nrg4, Alpl, Rassf8, Foxc2, Bmp5, Ddef1, Ptch1, Sdc4, Tbx5, Runx2, Ikkbkg, Ubc, Nog, Runx3, Fgfr1op2, Nfia, Gh were down-regulated. We suggested that Ihh increased condensation of mesenchymal cells and proliferation of prechondrocytes in early chondrogenesis. In conclusion, Ihh signaling positively regulates early chondrogenesis, as well as hypertrophy of chondrocytes in the mouse limb development.