Besides cell death, caspase-9 participates in non-apoptotic events, including cell differentiation. To evaluate a possible impact on the expression of chondrogenic/osteogenic factors, a caspase-9 inhibitor was tested in vitro. For this purpose, mouse forelimb-derived micromass cultures, the most common chondrogenic in vitro model, were used. The following analyses were performed based on polymerase chain reaction (PCR) arrays and real-time PCR. The expression of several chondrogenesis-related genes was shown to be altered, some of which may impact chondrogenic differentiation (Bmp4, Bmp7, Sp7, Gli1), mineral deposition (Alp, Itgam) or the remodelling of the extracellular matrix (Col1a2, Mmp9) related to endochondral ossification. From the cluster of genes with altered expression, Mmp9 showed the most significant decrease in expression, of more than 50-fold. Additionally, we determined the possible impact of caspase-9 downregulation on the expression of other Mmp genes. A mild increase in Mmp14 was observed, but there was no change in the expression of other studied Mmp genes (-2, -3, -8, -10, -12, -13). Interestingly, inhibition of Mmp9 in micromasses led to decreased expression of some chondrogenic markers related to caspase-9. These samples also showed a decreased expression of caspase-9 itself, suggesting a bidirectional regulation of these two enzymes. These results indicate a specific impact of caspase-9 inhibition on the expression of Mmp9. The localisation of these two enzymes overlaps in resting, proliferative and pre-hypertrophic chondrocytes during in vivo development, which supports their multiple functions, either apoptotic or non-apoptotic. Notably, a coincidental expression pattern was identified in Pik3cg, a possible candidate for Mmp9 regulation.

• Klíčová slova
• Caspase-9, Chondrogenic differentiation, Micromasses, Mmp-9, Non-apoptotic functions,
• MeSH
• buněčná diferenciace MeSH
• chondrocyty * MeSH
• chondrogeneze * fyziologie   MeSH
• inhibitory kaspas metabolismus   farmakologie   MeSH
• kaspasa 9 genetika   metabolismus   MeSH
• kultivované buňky MeSH
• myši MeSH
• osteogeneze MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• inhibitory kaspas MeSH
• kaspasa 9 MeSH

Objective. Caspases, cysteine proteases traditionally associated with apoptosis and inflammation, have recently been identified as important regulators of autophagy and reported within the growth plate, a cartilaginous part of the developing bone. The aim of this research was to identify novel autophagy-related molecules affected by inhibition of pro-apoptotic caspases in chondrocytes. Design. Chondrocyte micromasses derived from mouse limb buds were treated with pharmacological inhibitors of caspases. Autophagy-related gene expression was examined and possible novel molecules were confirmed by real-time polymerase chain reaction and immunocytofluorescence. Individual caspases inhibitors were used to identify the effect of specific caspases. Results. Chondrogenesis accompanied by caspase activation and autophagy progression was confirmed in micromass cultures. Expression of several autophagy-associated genes was significantly altered in the caspases inhibitors treated groups with the most prominent decrease for Pik3cg and increase of Tnfsf10. The results showed the specific pro-apoptotic caspases that play a role in these effects. Importantly, use of caspase inhibitors mimicked changes triggered by an autophagy stimulator, rapamycin, linking loss of caspase activity to an increase in autophagy. Conclusion. Caspase inhibition significantly affects regulation of autophagy-related genes in chondrocytes cultures. Detected markers are of importance in diagnostics and thus the data presented here open new perspectives in the field of cartilage development and degradation.

• Klíčová slova
• autophagy, caspase, gene expression, inhibition, limb,
• MeSH
• autofagie MeSH
• chondrocyty * metabolismus   MeSH
• chondrogeneze MeSH
• inhibitory kaspas metabolismus   farmakologie   MeSH
• kaspasy * metabolismus   farmakologie   MeSH
• myši MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• inhibitory kaspas MeSH
• kaspasy * MeSH

Meckel's cartilage was first described by the German anatomist Johann Friedrich Meckel the Younger in 1820 from his analysis of human embryos. Two hundred years after its discovery this paper follows the development and largely transient nature of the mammalian Meckel's cartilage, and its role in jaw development. Meckel's cartilage acts as a jaw support during early development, and a template for the later forming jaw bones. In mammals, its anterior domain links the two arms of the dentary together at the symphysis while the posterior domain ossifies to form two of the three ear ossicles of the middle ear. In between, Meckel's cartilage transforms to a ligament or disappears, subsumed by the growing dentary bone. Several human syndromes have been linked, directly or indirectly, to abnormal Meckel's cartilage formation. Herein, the evolution, development and fate of the cartilage and its impact on jaw development is mapped. The review focuses on developmental and cellular processes that shed light on the mechanisms behind the different fates of this cartilage, examining the control of Meckel's cartilage patterning, initiation and maturation. Importantly, human disorders and mouse models with disrupted Meckel's cartilage development are highlighted, in order to understand how changes in this cartilage impact on later development of the dentary and the craniofacial complex as a whole. Finally, the relative roles of tissue interactions, apoptosis, autophagy, macrophages and clast cells in the removal process are discussed. Meckel's cartilage is a unique and enigmatic structure, the development and function of which is starting to be understood but many interesting questions still remain.

• Klíčová slova
• chondrogenesis, congenital birth defects, craniofacial, jaw development, mammal evolution,
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH