Abstract
The purpose of this study was to investigate the aspects of proliferation and bone nodule formation of osteogenic precursor cells. To determine the effects of ascorbic acid and dexamethasone upon capacity of osteoblast proliferation and bone nodule formation, cells were maintained in the presence of one or some of these additives for up to 30 days. Group I culture was maintained in standard medium(DMEM plus 10% plus antibiotics), group II was maintained in supplemented medium containing dexamethasone, group III was maintained in supplemented medium containing ascorbic acid and sodium-${\beta}$-glycerophosphate, and group IV was maintained in supplemented containing ascorbic acid, sodium-${\beta}$-glycerophosphate and dexamethasone. Morphology of bone nodules was observed with light microscope and electron microscope. The results were as follows: ${\bullet}$ Proliferation capacity of osteoblasts was not affected by single use of dexamethasone, but it was chiefly affected by ascorbic acid. ${\bullet}$ Cellular morphology was fibroblastic appearance initially, but, it was gradually changed to polygonal shape accompanied by confluency stage. ${\bullet}$ Pluripotent mesenchymal cells existed during primary culture, they were differentiated to adipocyte, chondrocyte, osteocyte according to culture condition. ${\bullet}$ Dexamethasone increased bone nodule formation under the condition that the culture was maintained with supplemented medium ascorbic acid and sodium-${\beta}$-glycerophosphate. ${\bullet}$ when the cultures were stained with alizarin red, the group supplemented with dexamethasone, ascorbic acid and sodium-${\beta}$-glycerophosphate showed the marked increase of bone nodule formation, but the group supplemented with ascorbic acid and sodium-${\beta}$-glycerophosphate revealed only small amounts of bone nodules. And the groups cultured without ascorbic acid showed no observed any of bone-like mass independent of dexamethasone addition.
백서 태자 두개관을 효소처리하여 얻은 조골세포 유사세포군의 증식능 및 골결절의 형성에 미치는 dexamethasone의 영향과 형성된 골결절의 조직학적 형태 관찰을 위하여, 12 well 배양접시의 각 well 당 $4{\times}10^4cell$ 을 접종하였으며, 30일간 표준배양액으로만 배양한 군, 표준배양액에 ascorbic acid와 Na-${\beta}$-glycerophosphate를 첨가한 군, 그리고 각각에 dexa methasone을 첨가한 군등 4군으로 분리 배양하여 다음과 같은 결과를 얻었다. 1. 골세포의 증식율은 표준용액으로 배양한 군에 비해 dexamethasone 을 단독으로 투여한 군에서 약간 증가하는 경향을 보였으나 통계적인 유의성은 없었다 (p<0.05). 2. 골세포의 증식율은 ascorbic acid 에 의해 영향을 많이 받았으며, 특히 ascorbic acid와 dexamethasone을 동시에 투여했을때 더욱 현저히 세포증식율이 감소됨을 볼 수 있었다(p<0.05). 3. 배양 초기에는 세포들이 주로 섬유아세포 양으로 나타났으나, 밀생상태에 이르면서 점차 다각형 형태로 바뀌었다. 4. 배양 9일후 지방세포와 연골세포들이 관찰되었으며, 이들은 모두 dexamethasone이 첨가된 군에서 발견되었다. 5. 골결절은 ascorbic acid와 Na-${\beta}$-glycerophosphate가 첨가된 군에서만 형성되었으며, dexamethasone 을 병용첨가한 군에서 광화된 골결절이 현저히 증가됨을 관찰할 수 있었다. 6. 골결절 형성이 되지 않은 부위의 세포들은 평면형상을 이루고 있었으나, 골결절 부위를 덮고있는 세포들은 많은 세포돌기를 가진 조골세포양 세포들로 나타났으며, ascorbic acid, Na-${\beta}$-glycerophosphate 그리고 dexamethasone 을 첨가한 군에서는 침상의 돌기를 가진 광화물질들이 다발의 교원섬유들 사이에 현저히 많이 존재하였으며, 골결절 내부에는 매몰된 골세포(osteocyte)양 세포들이 관찰되었다.