The Myb locus encodes the c-Myb transcription factor involved in controlling a broad variety of cellular processes. Recently, it has been shown that c-Myb may play a specific role in hard tissue formation; however, all of these results were gathered from an analysis of intramembranous ossification. To investigate a possible role of c-Myb in endochondral ossification, we carried out our study on the long bones of mouse limbs during embryonic development. Firstly, the c-myb expression pattern was analyzed by in situ hybridization during endochondral ossification of long bones. c-myb positive areas were found in proliferating as well as hypertrophic zones of the growth plate. At early embryonic stages, localized expression was also observed in the perichondrium and interdigital areas. The c-Myb protein was found in proliferating chondrocytes and in the perichondrium of the forelimb bones (E14.5-E17.5). Furthermore, protein was detected in pre-hypertrophic as well as hypertrophic chondrocytes. Gain-of-function and loss-of-function approaches were used to test the effect of altered c-myb expression on chondrogenesis in micromass cultures established from forelimb buds of mouse embryos. A loss-of-function approach using c-myb specific siRNA decreased nodule formation, as well as downregulated the level of Sox9 expression, a major marker of chondrogenesis. Transient c-myb overexpression markedly increased the formation of cartilage nodules and the production of extracellular matrix as detected by intense staining with Alcian blue. Moreover, the expression of early chondrogenic genes such as Sox9, Col2a1 and activity of a Col2-LUC reporter were increased in the cells overexpressing c-myb while late chondrogenic markers such as Col10a1 and Mmp13 were not significantly changed or were downregulated. Taken together, the results of this study demonstrate that the c-Myb transcription factor is involved in the regulation and promotion of endochondral bone formation.
- MeSH
- biologické markery metabolismus MeSH
- buněčná diferenciace MeSH
- chondrogeneze fyziologie MeSH
- hybridizace in situ MeSH
- končetiny embryologie MeSH
- myši MeSH
- protoonkogenní proteiny c-myb genetika fyziologie MeSH
- umlčování genů MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Laser capture microdissection (LCM) uniquely allows the selection of specific cell populations from histological sections. These selected cells are then catapulted into a test tube without any contamination from surrounding tissues. During the last ten years, many significant results have been achieved, particularly at the level of DNA and RNA where amplification techniques are available. However, where amplification procedures are difficult, the benefits of LCM diminish. To overcome such difficulties, a novel approach, combining laser capture microdissection and flow cytometry, has been tested here for detection of apoptosis and proliferation in tissue bound cell populations without any amplification steps. The mouse cap stage molar tooth germ was used as a model. At the centre of the inner enamel epithelium, the primary enamel knot is a clearly defined apoptotic population with minimal proliferation, flanked by the highly proliferative cervical loops on each side. Thus within the tooth germ epithelium at this stage, two distinct populations of cells are found side by side. These populations were selected by laser capture microdissection and then analysed by flow cytometry for apoptosis and proliferation. Flow cytometric results correlated well with immunohistochemical findings, demonstrating the success and sensitivity of this combined procedure.
- MeSH
- apoptóza fyziologie MeSH
- epitelové buňky cytologie MeSH
- gestační stáří MeSH
- imunohistochemie MeSH
- koncové značení zlomů DNA in situ MeSH
- kryoprezervace MeSH
- laserová terapie metody MeSH
- mikrodisekce metody MeSH
- moláry embryologie MeSH
- myši MeSH
- orgán skloviny cytologie MeSH
- počet buněk MeSH
- proliferace buněk MeSH
- proliferační antigen buněčného jádra analýza MeSH
- průtoková cytometrie MeSH
- senzitivita a specificita MeSH
- zubní krček cytologie embryologie MeSH
- zubní zárodek cytologie MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- MeSH
- deprese diagnóza MeSH
- dospělí MeSH
- epilepsie diagnóza MeSH
- finanční podpora výzkumu jako téma MeSH
- lidé středního věku MeSH
- lidé MeSH
- neuropsychologické testy MeSH
- posttraumatická stresová porucha MeSH
- příznaky a symptomy diagnóza MeSH
- psychologické testy metody MeSH
- somatoformní poruchy diagnóza MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- přehledy MeSH
- srovnávací studie MeSH
- Klíčová slova
- LEVOPROTILIN,
- MeSH
- anorektika terapeutické užití MeSH
- bulimia farmakoterapie MeSH
- dospělí MeSH
- lidé MeSH
- maprotilin analogy a deriváty terapeutické užití MeSH
- mentální anorexie farmakoterapie MeSH
- Check Tag
- dospělí MeSH
- lidé MeSH
- ženské pohlaví MeSH
