Caspase-8 is the key component of the receptor-mediated (extrinsic) apoptotic pathway. Immunological localization of active caspase-8 showed its presence in osteoblasts, including non-apoptotic ones. Further in vivo exploration of caspase-8 functions in the bone is hindered by the fact that the caspase-8 knock-out is lethal prenatally. Examinations were thus performed using individual cell populations in vitro. In this study, caspase-8 was eliminated by the CRISPR/cas9 technology in MC3T3-E1 cells, the most common in vitro model of osteoblastic populations. The aim of the work was to specify the consequences of caspase-8 deficiency on non-apoptotic pathways. The impact on the osteogenic gene expression of the osteoblastic cells along with alterations in proliferation, caspase cascades and rapamycin induced autophagy response were evaluated. Osteogenic differentiation of caspase-8 deficient cells was inhibited as these cells displayed a decreased level of mineralization and lower activity of alkaline phosphatase. Among affected osteogenic genes, based on the PCR Array, major changes were observed for Ctsk, as down-regulated, and Gdf10, as up-regulated. Other significantly down-regulated genes included those coding osteocalcin, bone morphogenetic proteins (-3, -4 and -7), collagens (-1a1, -14a1) or Phex. The formation of autophagosomes was not altered in rapamycin-treated caspase-8 deficient cells, but expression of some autophagy-related genes, including Tnfsf10, Cxcr4, Dapk1 and Igf1, was significantly downregulated. These data provide new insight into the effects of caspase-8 on non-apoptotic osteogenic pathways.

• Keywords
• CRISPR/cas9, MC3T3-E1, apoptosis, autophagy, bone, osteogenesis,
• Publication type
• Journal Article MeSH

Within the mandible, the odontogenic and osteogenic mesenchymes develop in a close proximity and form at about the same time. They both originate from the cranial neural crest. These two condensing ecto-mesenchymes are soon separated from each other by a very loose interstitial mesenchyme, whose cells do not express markers suggesting a neural crest origin. The two condensations give rise to mineralized tissues while the loose interstitial mesenchyme, remains as a soft tissue. This is crucial for proper anchorage of mammalian teeth. The situation in all three regions of the mesenchyme was compared with regard to cell heterogeneity. As the development progresses, the early phenotypic differences and the complexity in cell heterogeneity increases. The differences reported here and their evolution during development progressively specifies each of the three compartments. The aim of this review was to discuss the mechanisms underlying condensation in both the odontogenic and osteogenic compartments as well as the progressive differentiation of all three mesenchymes during development. Very early, they show physical and structural differences including cell density, shape and organization as well as the secretion of three distinct matrices, two of which will mineralize. Based on these data, this review highlights the consecutive differences in cell-cell and cell-matrix interactions, which support the cohesion as well as mechanosensing and mechanotransduction. These are involved in the conversion of mechanical energy into biochemical signals, cytoskeletal rearrangements cell differentiation, or collective cell behavior.

• Keywords
• condensation, development, mandible, mesenchyme, mouse, odontogenesis, osteogenesis,
• Publication type
• Journal Article MeSH
• Review MeSH

The mandible is a tooth-bearing structure involving one of the most prominent bones of the facial region. Mesenchymal cell condensation is the first morphological sign of osteogenesis, and several studies have focused on this stage also in the mandible. Little information is available about the early post-condensation period, during which avascular soft condensation turns into vascularized bone, and all three major bone cell types, osteoblasts, osteocytes, and osteoclasts, differentiate. In the mouse first lower molar region, the post-condensation period corresponds to the prenatal days 13-15. If during this critical period, when osteogenesis reaches the point of major bone cell differentiation, vascularization already has to play a critical role, one should be able to show molecular changes which support both types of cellular events. The aim of the present report was to follow in organ context the expression of major osteogenic and angiogenic markers and identify those that are up- or downregulated during this period. To this end, PCR Array was applied covering molecules involved in osteoblastic cell proliferation, commitment or differentiation, extracellular matrix (ECM) deposition, mineralisation, osteocyte maturation, angiogenesis, osteoclastic differentiation, and initial bone remodeling. From 161 analyzed osteogenic and angiogenic factors, the expression of 37 was altered when comparing the condensation stage with the bone stage. The results presented here provide a molecular survey of the early post-condensation stage of mandibular/alveolar bone development which has not yet been investigated in vivo.

• Keywords
• PCR Array, angiogenesis, intramembranous ossification, mandibular bone, osteogenesis,
• Publication type
• Journal Article MeSH