胶原和二甲亚砜对原代培养大鼠肝细胞存活和功能状态的影响~1

Influence of collagen and DMSO on the survival and functional state of primary cultured rat hepatocytes

培养皿铺胶原和培养基中加入2％二甲亚砜(DMSO)均可延长培养肝细胞的稳定存活时间,并能改善其储存糖原和维持细胞色素P450含量的能力。DMSO还能基本维持细胞呈立方球形。这个培养体系有利于观察药物对肝细胞的长期效应。

We found that hepatocytes cultured in collagen coated dishes (CCD) and / or in the medium containing 2% dimethyl sulfoxide (DMSO) survived for a longer time and maintained better functional state than those cultured in control culture-conditions:The determination of LDH in the medium showed that CCD prolonged the time of stable survival up to 72 h and DMSO up to 96 h, i.e. about 24 h and 48 h longer than the control respectively.The determination showed that the DNA amount of the attached cells cultured for 3 h in CCD equaled to that cultured for 8 h in noncoated glass dishes. The number of the attached cells in CCD was about 75% of the total cells seeded after 8 h culture.The number of the attached (viable) cells (indi-ressed.cated by DNA determination) cultured in CCD plus DMSO-containing medium for 96 h was still about 86% of the total seeded cells. This percentage decreased to 78% after 120 h culture.With the first 48 h of culture, DMSO increased the glycogen content of hepatocytes. The glycogen content of cells in CCD and in DMSO-containing medium cultured for 96 h was a two-fold increase as compared with that of the control, i.e. CCD and DMSO deferred and diminished the loss of glycogen-synthetic function of cells in a long-term culture.Within 72 h, DMSO and CCD plus DMSO deferred the decrease of the cytochrome P450 content of hepatocytes. The cytochrome P450 content of the cells cultured for 24 h in these two conditions maintained up to about 50-60% of the original value, about a two- fold as much as that in the control. Hepatocytes in CCD plus DMSO-containing medium maintained essentially the cubico-spheric shape even after a 120 h culture, indicating that DMSO lightened the morphological changes of the cells after a long-term culture.These results indicate that coating the culture dishes with collagen and / or the addition of DMSO into the medium are the two factors helpful to pro-longing the survival time and improving the functional state of hepatocytes in the monolayer culture. They are useful in the investigation of those drug effects which only manifested after a long-term exposure.