Piper K, Boyde A, Jones S J
Department of Anatomy and Developmental Biology, University College London, UK.
Calcif Tissue Int. 1995 May;56(5):382-9. doi: 10.1007/BF00301607.
We have examined the relationship between the number of nuclei of an osteoclast and its volume. Chick and rat cells were released from long bones by chopping the shafts and flushing the fragments in Eagle's Minimum Essential Medium with added 10% fetal calf serum. The bone cell suspension was seeded onto glass coverslips. In Experiment 1, rat and chick cells were allowed to settle for 15 minutes, more medium was then added, and the cells were cultured in 5% CO2 at 37 degrees C for 4 hours. In Experiment 2, only rat cells were used, and the cells were cultured in the presence or absence of 10(-6) M 3-amino-1-hydroxypropylidene-1,1-bisphosphonate (APD) in the medium for 4 or 6 hours. The coverslips were washed in 37 degrees C phosphate-buffered saline and fixed for 24 hours in 2.5% glutaraldehyde in isotonic cacodylate buffer (initially 37 degrees C). The chick cells were critical point dried (CPD) or freeze dried (FD); all rat cells were FD. After drying, cells were coated with gold by vacuum evaporation. The volumes and areas of osteoclasts were measured using a video-rate, line-confocal reflection laser scanning microscope and the number of nuclei in each cell was counted. The volumes and volumes per nucleus of the FD cells were larger than those of the CPD cells but there was no significant difference in plan-areas. Rat osteoclasts were larger than chick cells in all the measured parameters except the mean number of nuclei/cell. The correlation coefficients for the areas, volumes, and the numbers of nuclei for rat and chick cells were all high (r > 0.725). The volumes and volumes per nucleus, but not the areas or areas per nucleus, of the osteoclasts cultured with APD were significantly smaller than control cells. We conclude that FD causes less shrinkage than CPD; chick osteoclasts are about two-thirds the size of rat osteoclasts; and 10(-6) M APD caused a reduction of rat osteoclast volume and volume per nucleus of 21%.
我们研究了破骨细胞核数量与其体积之间的关系。通过切碎长骨骨干并在添加10%胎牛血清的伊格尔氏最低必需培养基中冲洗碎片，从鸡和大鼠的长骨中分离出细胞。将骨细胞悬液接种到玻璃盖玻片上。在实验1中，让大鼠和鸡的细胞沉降15分钟，然后添加更多培养基，并将细胞在37℃、5%二氧化碳条件下培养4小时。在实验2中，仅使用大鼠细胞，并在培养基中添加或不添加10(-6)M 3-氨基-1-羟丙基亚丙基-1,1-二膦酸（APD）的情况下将细胞培养4或6小时。将盖玻片在37℃磷酸盐缓冲盐水中洗涤，并在等渗二甲胂酸钠缓冲液（最初为37℃）中的2.5%戊二醛中固定24小时。鸡的细胞进行临界点干燥（CPD）或冷冻干燥（FD）；所有大鼠细胞均进行FD。干燥后，通过真空蒸发给细胞镀上金。使用视频速率、线共焦反射激光扫描显微镜测量破骨细胞的体积和面积，并计算每个细胞中的核数量。FD细胞的体积和每核体积大于CPD细胞，但平面面积没有显著差异。除了每个细胞的平均核数量外，在所有测量参数中，大鼠破骨细胞都比鸡的细胞大。大鼠和鸡细胞的面积、体积和核数量的相关系数都很高（r>0.725）。用APD培养的破骨细胞的体积和每核体积，而不是面积或每核面积，显著小于对照细胞。我们得出结论，FD引起的收缩比CPD小；鸡破骨细胞的大小约为大鼠破骨细胞的三分之二；10(-6)M APD导致大鼠破骨细胞体积和每核体积减少21%。
