摘要
目的探讨外源性c8.神经酰胺诱导肺泡Ⅱ型上皮细胞凋亡的作用。方法取妊娠18dICR孕鼠,分离、培养胎鼠肺泡Ⅱ型上皮细胞(alveolartype1Iepithelialcell,AEC1I),分别加入0、20、40、80μmol/L的c8-神经酰胺溶液,继续培养24h,MTF法检测c8-神经酰胺对AECII增殖的影响,使用80μmol/LC8.神经酰胺溶液培养AECII3、6、12、24h,MTF法检测AECⅡ活力。使用0、20、40、80μmo]/LC8-神经酰胺溶液对AECII进行体外干预12和24h,分别使用流式细胞术和TUNEL法检测细胞凋亡率。结果c8-神经酰胺对AECII增殖具有明显抑制作用,并且具有浓度效应和时间效应。c8-神经酰胺溶液浓度为0、20、40、80μmol/L时,12和24h细胞总凋亡率分别为(20.634-0.86)%、(22.67±1.72)%、(42.03±1.34)%、(50.404±1.30)%和(20.93±2.51)%、(28.934±3.19)%、(47.004±1.08)%、(57.774±3.04)%(除12h20pμmol/L组外,相同时间点,其他各组与对照组相比,差异均有统计学意义,均P〈0.05)。c8-神经酰胺溶液浓度为0、20、40、80μmol/L时,12和24h的凋亡率分别为(5.514±1.41)%、(13.24±2.62)%、(35.104±4.59)%、(75.07±4.37)%和(5.564±1.36)%、(21.344±2.09)%、(37.12±2.11)%、(91.334±0.72)%(相同时间点,各浓度组与对照组相比,差异有统计学意义,均P〈0.05)。结论c8一神经酰胺可降低肺泡上皮细胞生存率,并且具有明显的诱导细胞凋亡作用。
Objective To investigate the effects of exogenous C8-ceramide on primary alveolar type Ⅱ epithelial cells (AEC Ⅱ ). Methods AEC 11 were isolated from 18 day gestational age ICR mice, and cultured in Dulbecco' s Modified Eagle Medium with 95% air and 5% CO2 at 37 ℃. The cells were treated with different concentrations (0, 20, 40 and 80 μmol/L) of C8-ceramide for 24 h and treated with 80 μmol/L of C8-ceramide for different time (3, 6, 12, and 24 h) to observe the viability of cells by MTT assay. Then, the annexin V externalization assay and TUNEL assay were performed to determine apoptosis after the cells were treated with different concentrations (0, 20, 40, and 80 μmol/L) of C8-ceramide for different time ( 12 and 24 h). Results The results of MTT assay showed that cell viability decreased in a dose- and time- dependent manner. In Annexin V externalization assay, the cells showed a distribution of (20.63 ±0.86)%, (22.67±1.72)%, (42.03 ±1.34)%, (50.40±1;30)% and (20.93 ±2.51)%, (28. 93 ± 3.19 ) % , (47.00 ± 1.08 ) %, ( 57. 77 ± 3.04 ) % cells in apoptosis after treated with 0, 20, 40, 80 txmol/L of C8-ceramide for 12 and 24 h ( It is significant difference from untreated cells except for 20 μmol/L at 12 h point, all P 〈 0. 05). In TUNEL assay, the cells showed a distribution of (5.51 ± 1.41)%, (13.24±2.62)%, (35.10±4.59)%, (75.07±4.37)% and (5.56±1.36)%, (21.34± 2. 09 ) %, ( 37. 12 ± 2. 11 ) %, (91.33 ± 0. 72) % cells in apoptosis after treated with 0, 20, 40, 80 μmol/L of C8-ceramide for 12 and 24 h (It is significant difference from untreated cells at the same time point, all P 〈 0. 05 ). Conelusion The results demonstrate that C8-eeramide can decrease the viability of AEC Ⅱ andinduce apoptosis.
出处
《中华结核和呼吸杂志》
CAS
CSCD
北大核心
2015年第6期445-450,共6页
Chinese Journal of Tuberculosis and Respiratory Diseases
基金
国家自然科学基金(81400058)
安徽省自然科学基金(1208085QH165)
关键词
肺泡
上皮细胞
神经酰胺类
细胞凋亡
Pulmonary alveoli
Epithelial cells
Ceramides
Apoptosis