Skeletogenesis occurs through either intramembranous or endochondral ossification. In addition, some parts of the skeletal components maintain their cartilaginous characteristics throughout life without mineralization. Runx2 is known to be a pivotal transcription factor for all skeletogenic processes. In this study, we examined the expression patterns of two major isoforms of Runx2 in early skeletogenesis. During intramembranous bone formation, Runx2-type I (Runx2-I) was widely expressed in osteoprogenitor cells and active osteoblasts, while Runx2-type II (Runx2-II) expression was stringently restricted to cells lining mineralized bones. Cells in permanent cartilage expressed collagen type II (Col-II) but never expressed Runx2 or Col-X. These permanent cartilages were well circumscribed by Runx2-I positive cells, in which Runx2-II was negative. In endochondral bone formation, Runx2 expression temporarily disappeared in Col-II-positive proliferating chondrocytes, but a secondary surge of Runx2-I expression occurred in the prehypertrophic zone before the mineralization of cartilage. Collectively, both Runx2 isoforms showed very similar expression patterns in active bone forming areas; however, Runx2-I has an exclusive role in the early commitment stage of intramembranous or endochondral bone forming processes or in cells surrounding permanent cartilage.
Animals ; *Bone Development ; Cartilage/cytology/growth & development/metabolism ; Embryo and Fetal Development/genetics ; *Gene Expression Profiling ; *Gene Expression Regulation, Developmental ; In Situ Hybridization ; Mice ; Mice, Inbred ICR ; Protein Isoforms/genetics/metabolism ; Time Factors ; Transcription Factors/*genetics/metabolism

Astrocytes are ubiquitous in the brain and have multiple functions. It is becoming clear that they play an important role in monitoring the neuromicroenvironment, information processing, and signaling in the central nervous system (CNS) in normal conditions and that they respond to CNS injuries. During the development of the CNS, astrocytes play a key role as a substrate for neuronal migration and axonal growth. To identify genes that could participate in astrocyte maturation, we used the differential display reverse transcription-PCR (DDRT-PCR) method. Human fetal astrocytes were cultured and total RNAs were isolated at intervals of 5 days for 50 days. Using 24 primer combinations, we identified a set of 18 candidate cDNAs deriving from the excised DDRT-PCR bands. DNA sequencing revealed 16 genes that have been described already. We found that RTP, TG, hTM-alpha, SPARC, TRIP7, and RPL7 genes were expressed increasingly, while HMGCR, RPL27a, NACA, NPM, and TARBP2 genes were expressed decreasingly, according to their culture stages. We also found two unidentified genes, A3 and C8, which were expressed differently in culture stages; the former was expressed decreasingly and the latter increasingly. These two genes were found in the same amount in genomic DNA from various human cells such as astrocytes, astrocytoma, trophoblasts and lymphocytes. The A3 gene was found only in human genomic DNA, but not in rat (ATr5), mouse (RAW264.7), or monkey (Vero) cells, whereas the C8 gene was found in human genomic DNA and monkey cells, but not in rat or mouse cells. We analysed these two genes for identification. There was > 92% nucleotide sequence identity between the A3 gene (3, 626 bp) and the Homo sapiens general transcription factor 3 (GTF3), and > 96% nucleotide sequence identity between the C8 gene (2, 401 bp) and the transmembrane receptor Unc5h2. These findings suggest that these two genes may participate in some functional roles within the cells.
Animals ; Astrocytes/*physiology ; Cell Aging/genetics ; Cells, Cultured ; Cercopithecus aethiops ; Embryo and Fetal Development ; Fetus/*physiology ; *Gene Expression ; Gene Expression Profiling ; Human ; Mice ; Rats ; Reverse Transcriptase Polymerase Chain Reaction ; Support, Non-U.S. Gov't ; Vero Cells