DeVito W J, Stone S, Shamgochian M
Division of Endocrinology, University of Massachusetts Medical Center, Worcester 01655, USA.
Alcohol Clin Exp Res. 2000 Jan;24(1):82-92.
The central nervous system is particularly sensitive to the cytotoxic effect of ethanol. In vivo and in vitro studies indicate that ethanol decreases cell proliferation in a number of cells types, including neurons and glial cells in the central nervous system. The cellular mechanisms involved in ethanol-induced cell toxicity, however, are unclear. In this study, we examined the effect of ethanol on tumor necrosis factor-alpha (TNFalpha)-induced cell death in a homogeneous population of cultured rat astrocytes.
Flow cytometric and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenytetrazolium bromide (MTT) dye reduction analyses were performed on cultured rat astrocytes to determine the effect of alcohol on TNFalpha-induced cell death.
Flow cytometric analysis revealed that, in quiescent astrocytes, high concentrations of ethanol were required to increase DNA fragmentation and decrease cell viability. Preexposure of astrocytes to low concentrations of ethanol (10 to 50 mM), however, increased the sensitivity of astrocytes to TNFalpha with low TNFalpha concentrations (25 to 50 ng/ml) resulting in increased DNA fragmentation. Furthermore, MTT dye reduction analysis revealed that exposure of astrocytes to 5 mM ethanol was sufficient to increase the susceptibility of astrocytes to the cytotoxic effect of ethanol. In a number of cell types, TNFalpha receptor binding results in the activation of specific signal transduction cascades, including the hydrolysis of sphingomyelin to ceramide. We show that preexposure of astrocytes to a low concentration of ethanol increased the sensitivity of astrocytes to sphingomyelinase, and C2-ceramide resulting in increased DNA fragmentation and decreased cell viability. More importantly, astrocytes prepared from rats exposed to ethanol prenatally showed increased susceptibility to TNFalpha-induced cell death.
These studies suggest that ethanol increases the susceptibility of astrocytes to TNFalpha-induced cell death by shifting the balance of sphingolipid metabolism in favor of a pathway that increases the susceptibility of astrocytes to the cytotoxic effect of TNFalpha.
中枢神经系统对乙醇的细胞毒性作用特别敏感。体内和体外研究表明，乙醇会降低多种细胞类型的细胞增殖，包括中枢神经系统中的神经元和神经胶质细胞。然而，乙醇诱导细胞毒性的细胞机制尚不清楚。在本研究中，我们检测了乙醇对培养的大鼠星形胶质细胞纯系中肿瘤坏死因子-α（TNFα）诱导的细胞死亡的影响。
对培养的大鼠星形胶质细胞进行流式细胞术和3-（4,5-二甲基噻唑-2-基）-2,5-二苯基四氮唑溴盐（MTT）比色法分析，以确定乙醇对TNFα诱导的细胞死亡的影响。
流式细胞术分析显示，在静止的星形胶质细胞中，需要高浓度的乙醇才能增加DNA片段化并降低细胞活力。然而，将星形胶质细胞预先暴露于低浓度乙醇（10至50 mM），会增加星形胶质细胞对低浓度TNFα（25至50 ng/ml）的敏感性，导致DNA片段化增加。此外，MTT比色法分析显示，将星形胶质细胞暴露于5 mM乙醇足以增加星形胶质细胞对乙醇细胞毒性作用的敏感性。在多种细胞类型中，TNFα受体结合会导致特定信号转导级联反应的激活，包括鞘磷脂水解为神经酰胺。我们发现，将星形胶质细胞预先暴露于低浓度乙醇会增加其对鞘磷脂酶和C2-神经酰胺的敏感性，导致DNA片段化增加和细胞活力降低。更重要的是，产前暴露于乙醇的大鼠制备的星形胶质细胞对TNFα诱导的细胞死亡的敏感性增加。
这些研究表明，乙醇通过改变鞘脂代谢平衡，使其有利于增加星形胶质细胞对TNFα细胞毒性作用敏感性的途径，从而增加星形胶质细胞对TNFα诱导的细胞死亡的敏感性。
