3-Methylcholanthrene (3MC) is a ligand for arylhydrocarbon receptor (AhR), which binds dioxin. We examined the effects of 3MC on the proliferation and differentiation of osteoblasts using cultures of rat calvarial osteoblast-like cells (ROB cells) and mouse calvarial clonal preosteoblastic cells (MC3T3-E1 cells). Analysis by RT-PCR revealed that the mRNAs for AhR and AhR nuclear translocators were expressed in both ROB and MC3T3-E1 cells. Cell proliferation and the synthesis of DNA by ROB cells and MC3T3-E1 cells were markedly inhibited on exposure of cells to 3MC. Furthermore, 3MC reduced the activity of alkaline phosphatase and the rate of deposition of calcium by cells. The level of expression of mRNA for osteocalcin, which is a marker of osteoblastic differentiation, was also depressed by 3MC. Moreover, when 3MC (1 mg/kg body weight) was administered sc to pregnant mice at 10.5, 12.5, and 14.5 d post coitus, fetuses examined subsequently at 15.5 or 17.5 d post coitus revealed evidence of inhibition of appropriate calcification of bones. The treated metacarpals showed no subperiosteal bone matrix histologically. Our findings indicate that 3MC might have critical effects on the formation of bone both in vivo and in vitro.
The rotational movement of a spermatozoon around its longitudinal axis was investigated by two methods: by observing a spermatozoon attached vertically to a coverslip by the tip of its head, and by observing a spermatozoon freely swimming in a medium by means of ‘double-focal microscopy’, which yielded simultaneous images at two different focal planes. Similar results were obtained by these two methods. Sea urchin, starfish, medaka, human, golden hamster and bull spermatozoa rolled in both clockwise and counterclockwise directions, although there was a large difference in the proportion of spermatozoa rolling in each direction in the different species. The majority of sea urchin and starfish spermatozoa rolled in a clockwise direction when an observer viewed the cell from its anterior end, whereas the majority of medaka, golden hamster, human and bull spermatozoa rolled in a counterclockwise direction relative to the same observer. Moreover, some spermatozoa occasionally changed their rotational direction. These results suggest that the mechanism regulating the direction of rotation of the spermatozoa is lax. As rotational movement of a spermatozoon around its longitudinal axis is due to the three-dimensional component of the beat of the flagellum, the direction of the three-dimensional movement presumably changes as the spermatozoa swim.
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