分離肝細胞의 蛋白合成能에 關하여 = Studies on Protein Synthesis of Isolated Liver Cells

국문 초록 (Abstract)

흰쥐간의 肝細胞分離方法을 檢討하여 分離한 肝細胞를 amino 酸 및 其他營養素組成이 豊富한 CMRL-1066培養液 또는 이에 흰쥐 血淸을 첨가한 培養液에 incubation하여 放射能追跡法으로 蛋白合成能을 檢討한바 다음과 같은 성적을 얻었다.
1. 肝細胞를 機能的으로 分離함에 있어 肝을 미리 citrate을 含有하는 Ca^++ free Locke液으로 還流했을때 肝細胞의 完全分離가 용이하였으며 이에 比해 citrate를 含有하지 않은 Ca^++ free Locke液으로 還流했을때는 小數의 肝細胞가 含有된 細胞群을 分離할 수 있었다. 이들 肝細胞의 分離에 sieve pipet를 考案하여 使用하였드니 肝細胞의 分離 또는 分散에 效果的이었다.
2. 肝組織에서 肝細胞를 完全分離했을때와 肝細胞群으로 不完全分離했을때의 各 細胞를 CMRL-1066培養液에서 incubation하여 amino acid mixed-^14C의 肝蛋白內 편입속도를 比較하였든바 細胞群으로 分離한 肝細胞에서 편입속도가 훨씬 높았다.
3. 分離한 肝細胞를 CMRL-1066液에 흰쥐血淸 15% 첨가한 培養液에 incubation하면서 肝細胞를 培養液中에서 再凝集시켜 보았으나 amino acid mixed-^14C의 肝蛋白內 편입속도는 증가되지 않았으며 肝細胞를 培養器內벽에 부착시켜 固定된 狀態에서 배양했을때 편입속도가 상승되었다.
4. 肝細胞培養器內의 酸素量을 증가시켜 보았으나 편입속도는 상승되지 않았다.

다국어 초록 (Multilingual Abstract)

Parenchymal cells obtained by dispersion of perfused rat liver in 0.25M sucrose solution were incubated in CMRL-1066 medium and their protein synthesis were studied by measuring the incorporation of amino acid mixed-^14C into liver protein.
Adult rat livers were perfused with Ca^++ free Locke solution with 0.027M citrate(37℃) and liver cells were separated by passing the minced liver tissue through nylon sieves of increasing fine mesh. The operation was performed by finger press in 0.25M surcose solution and a sieve pipet of 180 mesh, designed in our laboratory, was used for final suspension of liver cells. Majority of the hepatic cord cells recovered in the suspension existed as single cells and the yields were 5~6 × 10 exp (7) cells from 1 g of wet liver tissue. Liver cells clumps mostly consisted of a few but less than ten cells were also prepared by incomplete separation and centrifuge method for the comparison of the function with the completly separated liver cells.
Liver cells, suspended in CMRL-1066 medium 15 percent rat serum, were transfered to the incubation apparatus (Fig 1) and the cells were reaggregated in the medium or fixed on the inner surface of incubation tubes by controling the speed of tube rotation within 30 minutes after initiation of cell incubations.
The liver cells of two different condition during 4 hours incubation showed a considerable difference in the incorporation study. In fixed liver cells more increase of the incorporation was noted after 2 hours whereas the incorporation was almost stopped is reaggregated cells after the time.
Considering the possible effect of exposure of liver cells in to air phase in the fixed state according to the tube rotation, experiement replaced the air phase partialy with oxygen was also carried out, but no noticeable effect of additional oxygen supply on the incorporation was noted.