DAPT suppresses the proliferation of human glioma cell line SHG-44

Objective:To explore the suppressing effect ofγ-secretase inhibitor DAPT on proliferation of human glioma cell line SHG-44 in vitro and its mechanism.Methods:The SHG-44 cell was treated by DAPT with different concentration.The proliferation of cells was detected by MTT assay;cell cycle and TSC of CD133^+were determined by flow cytometry analysis technique;the key factor in Notch signaling pathway(Notch-1,Delta-1,Hes-1)was measured by reverse transcrip tase-polymerase chain reaction and western blotting.Results:DAPT inhibited the growth and proliferation of SHG-44 cells significantly(P<0.05).And the inhibiting effect on SHG-44 cells produced by DAPT showed a dose-dependent manner.DAPT increased the rate of cells in G_0/G_1 phase of SHG-44 cells,while it decreased the rate of cells in S phase.TSC of CD133^+was significantly reduced after DAPT treated SHC-44 cells.The expression of protein and mRNA of Notch-1,Delta-1 and Hes-1 were gradually downregulated with the increase of DAPT doses.Conclusions:DAPT can downregulate these key factor in Notch signaling pathway,reduce the TSC of CD133+and inhibit the proliferation of SHC-44 cells.

芳香化酶在胶质母细胞瘤细胞系SHG-44细胞中的表达及调控

目的 研究芳香化酶细胞色素P45 0 (AROM)及雌激素受体 (ER α)在胶质母细胞瘤细胞系SHG 4 4细胞中的基因表达。 方法 细胞培养、免疫细胞化学染色、原位杂交染色及RT PCR技术。 结果 在SHG 4 4细胞中分别检测到AROM及ER α的表达 ,进一步发现SHG 4 4细胞中AROM的表达是由多个组织特异性启动子驱动基因的转录。 结论 可能为中枢神经系统肿瘤发生的激素调节提供新的资料。

Growth and radiosensitivity of irradiated human glioma cell progeny

BACKGROUND： Progenitors of the immortalized human glioma cell line, SHG-44, are significantly less sensitive to irradiation. Two hypotheses regarding the mechanism of this effect exist： several studies have suggested that there is a subgroup with different radiosensitivities in identical cell group, and the progenitors of irradiate is a adaptive response subgroup, so its radiosensitivity is descend. A second hypothesis suggests that irradiated glioma progeny have a stronger ability to repair DNA damage. This would suggest that when progeny are continuously irradiated, resistance to irradiation-induced DNA increases, and radiosensitivity decreases. OBJECTIVE： To investigate radiosensitivity and growth features after irradiation to progeny of the human glioma cell line SHG-44. DESIGN, TIME AND SETTING： A randomized, controlled experiment, which was performed at the Department of Radiology Laboratory, the First Hospital Affiliated to Soochow University, between September 2004 and January 2006. MATERIALS： The glioma cell line SHG-44 was provided by the Institute of Neuroscience, First Affiliated Hospital of Suzhou University. Propidium iodide reagent was provided by Coulter Corporation. A linear accelerator, KD-2 type, was provided by Siemens, Germany. The flow cytometer EPICS-XL was provided by Coulter Corporation. METHODS： Brain glioma SHG-44 cells were divided into four groups： SHG-44, SHG-44-2, SHG-44-6, and SHG-44-10 . The SHG-44-2, SHG-44-6, and SHG-44-10 cells were vertically irradiated with varying doses of 2, 6 and 10 Gy by a linear accelerator （6 MVX）. The cells were passaged for 15 generations and cultured in RPMI-1640 culture media. MAIN OUTCOME MEASURES： Community re-double time, mean lethal dose （D0）, extrapolation number （N）, fraction surviving fraction irradiated by 2 Gy dose （SF2）, quasi-threshold dose （Dq）, and cell cycle. RESULTS： The Population doubling time （PDT） of SHG-44-2, SHG-44-6, and SHG-44-10 cell groups was not significant （P = 0.052）. Compared to these three groups, the PDT of the SHG-44 cell group was significantly difference （F = 7.878, P 〈 0.002）. SHG-44 cell clone ratewas 26.5%, and SHG-44-10 cell group was 15.5%. The SHG-44-10 cell group also exhibited radiosensitivity, but was less than the radiosensitivity of the SHG-44 cell group. Compared to the SHG-44 cell group, the ratio of the G2/M phase was decreased in the SHG-44-10 cell group, and the radio of S phase was increased. The SHG-44 and SHG-44-10 cell groups were irradiated with 8 Gy. After 12 hours, the G2/M ratio was compared to pre-irradiation times, indicating a significantly higher ratio in the pre-irradiated groups （P 〈 0.01）. The cells between S HG-44 and SHG-44-10 groups were harvested 12 hours after irradiation： G2 phase of SHG-44-10 cells was arrested and the G2/M ratio was increased, which was intensified with increasing irradiation doses. CONCLUSION： In the present study, the proliferation delay and decreased radiosensitivity were confirmed in progeny of irradiated human glioma cells, and radiosensitivity was dose-dependent.

他莫昔芬对SHG-44人胶质瘤细胞钠通道的抑制作用

目的:研究他莫昔芬对SHG-44胶质瘤细胞钠通道电流的作用。方法:采用全细胞膜片钳方法记录SHG-44细胞的钠通道电流,并观察施用不同浓度的他莫昔芬后电流的变化。结果:该钠通道电流特性为内向电流、快速激活失活,他莫昔芬能够明显阻断该电流,该阻断具有剂量依赖性及电压依赖性。在0 mV时,8μmol/L他莫昔芬对钾电流抑制率为69%。半数抑制浓度(IC50)为5.54μmol/L。结论:他莫昔芬可明显阻断SHG-44胶质瘤细胞上的钠通道,这可能是他莫昔芬抑制胶质瘤细胞增殖的机制之一。

他莫昔芬抑制胶质瘤细胞系SHG-44增殖及钠通道电流

目的研究他莫昔芬对SHG-44胶质瘤细胞的增殖及其细胞膜上钠通道电流的作用。方法用四唑盐比色试验分析细胞活性,通过流式细胞仪检测细胞增殖和凋亡。以全细胞膜片钳法记录SHG-44细胞的钠通道电流。结果加入他莫昔芬后,SHG-44细胞变老、脱落,细胞总数减少。他莫昔芬组G2/M期细胞较对照组增多,凋亡细胞比例增加。钠通道电流特性为内向电流、快速激活失活。他莫昔芬可剂量依赖性及电压依赖性阻断该电流。在0mV时,8μmol/L他莫昔芬对钠通道电流抑制率为69%。半数抑制浓度(IC50)为5.54μmol/L。结论他莫昔芬可明显阻断SHG-44胶质瘤细胞上的钠通道,这可能是其抑制胶质瘤细胞增殖的机制之一。

他莫昔芬对SHG-44胶质瘤细胞氯通道的抑制作用

目的研究氯通道阻断剂他莫昔芬(tamoxifen)对SHG-44胶质瘤细胞电压依赖性氯通道电流的作用。方法采用全细胞膜片钳方法记录SHG-44细胞的电压依赖性氯通道电流,并观察施用不同浓度的他莫昔芬后电流的变化。结果该电流特性为外向整流、不失活,他莫昔芬能够明显阻断该电流,该阻断具有剂量依赖性及电压依赖性。在+100 mV时,μM及5μM他莫昔芬对氯电流抑制率分别为48%及89%。结论他莫昔芬可明显阻断SHC-44胶质瘤细胞上的电压依赖性氯通道。

小分子干扰RNA(siRNA)沉默CD147基因对脑胶质瘤细胞系SHG-44生长及凋亡的影响研究

目的探讨小分子干扰RNA(siRNA)沉默CD147基因对脑胶质瘤细胞系SHG-44生长及凋亡的影响。方法根据CD147 cDNA序列设计具有短发夹结构的两条DNA序列,与载体pSilencer4.1-CMV neo构建重组表达载体,鉴定后转染至SHG-44细胞,western blotting检测抑制效果,MTT检测细胞增殖情况,流式细胞术检测肿瘤细胞凋亡情况。结构成功构建了针对CD147基因表达的干扰质粒,有效抑制了SHG-44细胞增殖,促进了SHG-44细胞凋亡。结论 CD147靶向RNA干扰重组表达载体为肝癌的基因治疗提供了可能。