defining the earliest transcriptional steps of chondrogenic progenitor specification during the formation of the digits in the embryonic limb定义最早转录chondrogenic祖规范的步骤在数字的形成胚胎肢.pdfVIP

Defining the Earliest Transcriptional Steps of Chondrogenic Progenitor Specification during the Formation of the Digits in the Embryonic Limb Carlos I. Lorda-Diez., Juan A. Montero., Manuel J. Diaz-Mendoza, Juan A. Garcia-Porrero, Juan M. Hurle* ´ ´ ´ ´ ´ Departamento de Anatomıa y Biologıa Celular and Instituto de Formacion e Investigacion Marques de Valdecilla, Universidad de Cantabria, Santander, Spain Abstract The characterization of genes involved in the formation of cartilage is of key importance to improve cell-based cartilage regenerative therapies. Here, we have developed a suitable experimental model to identify precocious chondrogenic events in vivo by inducing an ectopic digit in the developing embryo. In this model, only 12 hr after the implantation of a Tgfb bead, in the absence of increased cell proliferation, cartilage forms in undifferentiated interdigital mesoderm and in the course of development, becomes a structurally and morphologically normal digit. Systematic quantitative PCR expression analysis, together with other experimental approaches allowed us to establish 3 successive periods preceding the formation of cartilage. The ‘‘pre-condensation stage’’, occurring within the first 3 hr of treatment, is characterized by the activation of connective tissue identity transcriptional factors (such as Sox9 and Scleraxis) and secreted factors (such as Activin A and the matricellular proteins CCN-1 and CCN-2) and the downregulation of the galectin CG-8. Next, the ‘‘condensation stage’’ is characterized by intense activation of Smad 1/5/8 BMP-signaling and increased expression of extracellular matrix components. During this period, the CCN matricellular proteins promote the expression of extracellular matrix and cell adhesion