白介素2和肿瘤坏死因子α对大鼠C6胶质瘤细胞产生一氧化氮的影响

Effects of interleukin 2 and tumor necrosis factor α on nitric -oxide production from rat C6 glioma cells

用Ｇｒｉｅｓｓ法检测细胞培养上清中ＮＯ÷２含量作为反映细胞产生一氧化氮（ＮＯ）量的指标，观察细菌脂多糖（ＬＰＳ），干扰素γ（ＩＦＮ－γ），肿瘤坏死因子α（ＴＮＦ－α）及白介素２（ＩＬ－２）对大鼠Ｃ６胶质瘤细胞产生ＮＯ的影响．结果Ｃ６细胞于体外培养８ｈ时可自发产生ＮＯ，２４ｈ达峰值（１７．５±１．９ｖｓ溶剂对照组６．５±１．９μｍｏｌ·Ｌ－１），４８ｈ仍有产生．在所用剂量范围，上述因素单独作用２４ｈ均不影响Ｃ６产生ＮＯ，而５－５００ｋＵ·Ｌ－１ＩＬ－２与０．５ｍｇ·Ｌ－１ＬＰＳ或５０ｋＵ·Ｌ－１ＩＦＮ－γ合用可增强Ｃ６产生ＮＯ（１０．６－１３．４ｖｓ７．２μｍｏｌ·Ｌ－１），ＬＰＳ，ＩＦＮ－γ及ＴＮＦ－α三者合用亦有一定促进Ｃ６产生ＮＯ作用．提示炎性刺激与炎性细胞因子共同作用可促进中枢神经胶质细胞产生ＮＯ．

Activated microglia and astrocytes synthesize nitric oxide (NO) and preinflammatory cytokines such as IL-1, IL-2, IL-6 and TNFα. To investigate which one of these mediators plays a key role in the proposed inflammatory cascade in Alzheimer′s disease, NO production represented by nitrite concentration in supernatants of cultured rat C6 glioma cells was assayed with Griess reaction, and the effects of IL-2 and TNFα on NO production from C6 glioma cells were studied. The results showed that when cultured without stimulation, C6 glioma cells produced NO which started at 8 h and reached peak concentration at 24 h(17.5±1.9 vs medium control 6.5±1.9 μmol·L1). The NO production was not affected by the addition of 0.01-1 mg·L1 LPS, 10-1000 kU·L1 IFNγ, IL2 or TNFα. In the presence of 0.5 mg·L1 LPS or 50 kU·L1 IFNγ, IL2 at 5-500 kU·L1 enhanced NO production (10.6-13.4 vs 7.2 μmol·L1). In the presence of 0.33 mg·L1 LPS+33 kU·L1 IFNγ, C6 glioma cells cultured for 6 h with 33 kU·L1 TNFα produced more NO than the control (17.5 vs 10.4 μmol·L1). The results suggest that the combination of preinflammatory cytokines and infectious stimuli enhance NO production from CNS glial cells.