X射线对人肺癌A549细胞CCR7表达影响的研究

目的 研究不同剂量X射线照射及照射后不同时间点对人肺癌A549细胞CC-趋化因子受体7(CCR7)表达的影响.方法 体外培养A549细胞,实验组采用直线加速器X射线一次性照射,细胞吸收剂量分别为2、4、6和8 Gy(源皮距100 cm;剂量率442.89 cGy/min),照射后4、12、24、48和72 h分别采用实时荧光定量PCR技术及Western blot方法分别进行CCR7 mRNA及蛋白质表达水平检测;对照组A549细胞除不接受x射线照射外,余处理同实验组.结果 A549细胞经2、4、6和8 Gy的X射线照射后,CCR7 mRNA及蛋白质在照射4 h后开始表达升高,达到高峰后相继出现下降;72 h后6和8 Gy组mRNA表达量仍高于对照组水平(t=6.75～7.26,P＜0.01),2和6 Gy组蛋白质表达量高于对照组(t=11.13～14.17,P＜0.01),而4和8 Gy组蛋白质表达量在48和72 h已降至对照组水平.结论 2、4、6和8 Gy的X射线照射A549细胞后,A549细胞CCR7mRNA及蛋白质的表达量明显增加,可能与一定剂量X射线辐射促进A549细胞增殖和转移有关.

Abstract：

Objective To study the effects of X-ray radiation on CC-chemokine receptor 7(CCR7) expression in human non-small cell lung cancer (NSCLC) cells.Methods Humanadenocarcinoma cells of the line A549 were cultured and irradiated by X-ray at the absorbed doses of 2,4,6,and 8 Gy respectively by linear accelerator (with the source skin distance of 100 cm and dose rate of 442.89 cGy/min).The relative levels of CCR7 mRNA and protein expression in the A549 cells were respectively detected by real time-PCR and Western blotting 4,12,24,48,and 72 h after radiation.Untreated A549 cells were used as control group.Results The expression levels of CCR7 mRNA and protein in the A549 cells began to increase since 4 h after radiation and then decreased gradually after they reached the peak.The CCR7 mRNA expression levels 72 h after radiation of the 6 and 8 Gy groups were still significantly higher than those of the control group (t = 6.75-7.26,both P < 0.01),and the CCR7 protein expression levels of the 2 and 6 Gy group were still significantly higher than those of the control group(t=11.13-14.17,both P <0.01).Then the CCR7 protein expression levels of the 4 and 8 Gy groups decreased to the control group level 48 and 72 h after radiation respectively.Conclusions The CCR7 mRNA and protein expression levels in the NSCLC cells increase after X-ray irradiation,which may be correlated with the promotion of proliferation and metastasis of NSCLC cells by X-ray irradiation at a certain dose.     

X射线对人肺腺癌A549细胞Pokemon基因表达的影响

目的 研究不同剂量X射线照射及照射后不同时间点对人肺腺癌A549细胞Pokemon基因表达的影响.方法 用吸收剂量分别为2、4、6和8 Gy的X射线照射体外堵养的人肺腺癌A549细胞,2、4、8、12、24、48和72 ha,用实时定量PCR技术检测其中的Pokemon mRNA表达水平,以未照射组为对照.结果 在2、4、6、8 Gy X射线照射后的早期(除2 Gy照射后的2和4 h外)Pokemon mRNA的表达降低,但在晚期(48 h以后)呈升高趋势,在大部分时间点实验组与对照组的差异有统计学意义(t=3.40～154.76,P<0.05).结论 较大剂量的X射线在早期可下调A549细胞Pokemon基因mRNA的表达,诱导肿瘤细胞凋亡;但在晚期又可诱导A549细胞高表达PokemonmRNA,这可能与辐射所致A549细胞的DNA损伤修复和细胞周期调控有关.

Abstract：

Objective To study the dose-and time-effects of X-ray irradiation on the expression of Pokemon gene in A549 cells of human lung adenocarcinoma.Methods A549 cells were cultured in vitro and exposed to X-rays with the doses of 2,4,6 and 8 Gy,respectively.Untreated A549 cells were used as control group.The relative levels of Pokemon mRNA expression in the cells were detected by using quantitative real-time PCR at 2,4,8,12,24,48 and 72 h after irradiation.Results The Pokemon mRNA expression levels decreased in the early period after irradiation(except 2 and 4 h after irradiation in 2 Gy group)and then increased in the later stage(48 h after irradiation)with significant statistical differences at the most time points in comparison with the control group(t=3.40-154.76,P<0.05).Conclusions Higher doses of X-rays may degrade the expression of Pokemon mRNA in the human A549 cells and induce apoptosis in the early period,hut also may upgrade its expression in the later period, which might be correlated with the cell cycle regulation and DNA damage repair in the A549 cells.     

塞来昔布对人肺腺癌细胞株A549的放射增敏效应及细胞迁移力的影响

Objective To investigate the effects of radiosensitivity enhancement and inhibition of migration ability of human lung adenocarcinoma cells by celecoxib,a selective cyclooxygenase (COX)-2 inhibitor.Methods Human lung adenocarcinoma cells of the line A549 were cultured and then inoculated into six-well plates and randomly divided into 4 groups:control group,celecoxib group administered with celecoxib at the subtoxic doses 30 and 50 μmol/L,irradiated group exposed to 0,1,2,4,6,or 8 Gy by linear accelerator,and combined treatment (celecoxib + irradiation) group.The radiosensitizing effect of celecoxib was assessed by clonogenic cell survival test.The migration ability of the A549 cells was measured by scratch-wound test and the content of metalloproteinase-2 (MMP-2) in culture supernatant was detected with ELISA.Results The sensitization enhancement ratio of the celexib group was increased dosedependently.The values of D0 ,Dq,SF2 and D0.01 of the celecoxib + irradiation group were all significantly lower than those of the irradiated group.Scratch-wound test showed that the no-scratch area of the celecoxib + irradiation group and celecoxib group were all significantly wider than those of the mere irradiation and control groups and there was a dose-dependent manner,and the no-scratch area of the celecoxib + irradiation group was wlider than that of the celecoxib group.ELISA showed that the MMP-2 levels in the supernatant of the celecoxib group and celecoxib + irradiation group were respectively significantly lower than those of the control group and mere irradiated group (t = 3.78,5.79、3.15,P ＜ 0.05),however,there was not significant difference between the mere irradiation and control groups (t = 2.73,2.38,P ＞ 0.05).Conclusions Celecoxib enhances concentration-dependently the radiosensitivity of human lung carcinoma cell and inhibits the secretion of MMP-2 of the carcinoma cells,thus inhibiting their migration ability.     

电离辐射对小鼠脾细胞中调节性T细胞及其相关分子表达的影响

Objective To observe the changes in expressions of spleen regulatory T cells (Tregs)and the related factor forkhead box protein-3 (Foxp3) after irradiation with different doses of X-ray in mice at different times,and to elaborate the effects of X-rays on regulatory T cells and Foxp3.Methods 112male ICR mice were randomly divided into 2 groups and irradiated by X-rays at the doses of 0.075 and 2 Gy,respectively.The mice were killed at 0,4,8,16,24,48,and 72 h post-irradiation and the spleens removed.Flow cytometry was used to detect the percentage of CD4 + CD25 + Treg and protein expression of (Foxp3),and RT-PCR was used to exmiamine the mRNA expression of Fox3.Results Compared with those before irradiation,the CD4 + CD25 + Treg positive rates began to increase and peaked at 8 h post-irradiation with 0.075 Gy at 8,16,24,72 h(t = 8.73,10.55,4.21,4.65 ,P ＜ 0.05) and 2 Gy at 8,16,48,72 h(t = 4.65,4.28,3.71,2.88,P ＜ 0.05),and then slightly decreased,but still remained at high levels.The mRNA protein levels of Fox3 did not change significantly after exposure to the dose of 0.075 Gy,but began to significantly increase at 8 h after exposure to the dose of 2 Gy.However,the Foxp3 protein level began to increase 4 h post-irradiation,peaked at 16 h,and then slightly decreased,but still ramained at high levels (t =2.59,3.37,3.70,3.20,P＜0.05).Conclusions The changes in expressions of Tregs and Foxp3 after high- and low-dose X-ray irradiation may be used to explain the differences in immune effects induced by ionizing radiation at different doses.     

当归多糖对小鼠肝细胞辐照后Bcl-2和Bax蛋白表达的影响

Objective To investigate the effect of polysaccharides from Angelica sinensis (ASP3) on Bcl-2 and Bax protein expression of irradiated liver cells from mice.Methods Bcl-2 and Bax protein expression of liver cells in vitro exposed to 2.0 Gy rays were examined by using immunohistochemistry method.Results The expression of apoptosis-accelerating protein Bax in the irradiation group was enhanced obviously(70.83%),while apoptosis inhibiting protein Bcl-2 tended to decline(55.60%),with the statistically significant difference(P＜0.01) compared with that of the control.ASP3 pretreatmcnt could regulate Bcl-2 and Bax protein expression of liver cells,inhibiting Bax protein expression(64.14/58.37%) and increasing Bcl-2 protein expression( 59.21%/67.45%).The differences between the high dosage (100 mg/L of ASP3) and the irradiation group were statistically significant( P＜0.05 ).Conclusions ASP3 pretreatment could prohibit the apoptosis of radiationdamaged liver cells due to abnormal expression of Bcl-2 and Bax,and reduce the cell apoptosis by increasing Bcl-2/Bax protein expression so as to enhance the radiation endurance of liver cells.     

18F-FLT体外监测结肠癌细胞早期放射反应

Objective To evaluate whether 18F-fluorothymidine(FLT) can be used to monitor early response to irradiation in colorectal cancer (CRC).Methods SW480 cells were cultured and irradiated with 0, 10, and 20 Gy.Twenty-four hours later, morphological changes, apoptosis, necrosis, proliferation,and cell cycle phases were observed.Uptake of 18F-FLT was measured in these tumors in vitro from 24 h to 72 h after irradiation.The one-way analysis of variance was used to analyze the data.Results Apoptotic and necrotic cells were detected 24 h after radiotherapy.SW480 cells proliferation was significantly delayed after irradiation in 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenylte-trazolium bromide (MTI) assay.Cell cycle analysis showed that SW480 cells had a decreased fraction of cells in S phase( from 33.23% to 9.24%,then to 5.43% ) and an arrested fraction in G0-G1.After SW480 cells were cultured for60 min, the uptake of 18F-FLT was (5.21 ± 1.60) %; and 24 h after irradiation of 10 Gy, the uptake decreased significantly to (4.27±0.48)% (F=8.253, P=0.009).And 72 h after irradiation, the uptake further decreased significantly to (3.39 ± 0.59) % ( F = 36.715, P＜0.001 ).In tumor tissue, the uptake of 18F-FLT reduced significantly 72 h after radiotherapy (10 Gy:F = 12.388, P = 0.007; 20 Gy:F = 16.744, P = 0.004) and the attenuation degree increased with the radiation dose.Conclusion The uptake of 18F-FLT in SW480 cells or in CRC could reflect the changes of SW480 cells in proliferation, cell cycle re-distribution, cell apoptosis and necrosis.The results suggest that 18F-FLT may be used for monitoring early response to irradiation of CRC.     

18F-FLT体外监测结肠癌细胞早期放射反应

Objective To evaluate whether 18F-fluorothymidine(FLT) can be used to monitor early response to irradiation in colorectal cancer (CRC).Methods SW480 cells were cultured and irradiated with 0, 10, and 20 Gy.Twenty-four hours later, morphological changes, apoptosis, necrosis, proliferation,and cell cycle phases were observed.Uptake of 18F-FLT was measured in these tumors in vitro from 24 h to 72 h after irradiation.The one-way analysis of variance was used to analyze the data.Results Apoptotic and necrotic cells were detected 24 h after radiotherapy.SW480 cells proliferation was significantly delayed after irradiation in 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenylte-trazolium bromide (MTI) assay.Cell cycle analysis showed that SW480 cells had a decreased fraction of cells in S phase( from 33.23% to 9.24%,then to 5.43% ) and an arrested fraction in G0-G1.After SW480 cells were cultured for60 min, the uptake of 18F-FLT was (5.21 ± 1.60) %; and 24 h after irradiation of 10 Gy, the uptake decreased significantly to (4.27±0.48)% (F=8.253, P=0.009).And 72 h after irradiation, the uptake further decreased significantly to (3.39 ± 0.59) % ( F = 36.715, P＜0.001 ).In tumor tissue, the uptake of 18F-FLT reduced significantly 72 h after radiotherapy (10 Gy:F = 12.388, P = 0.007; 20 Gy:F = 16.744, P = 0.004) and the attenuation degree increased with the radiation dose.Conclusion The uptake of 18F-FLT in SW480 cells or in CRC could reflect the changes of SW480 cells in proliferation, cell cycle re-distribution, cell apoptosis and necrosis.The results suggest that 18F-FLT may be used for monitoring early response to irradiation of CRC.     

γ射线照射对肺腺癌细胞迁移能力的影响及其分子机制

目的探讨辐射对肺腺癌A549细胞迁移能力的影响及其所依赖的信号通路。方法划痕实验检测2和4 Gyγ射线照射后A549细胞的迁移能力,Western印迹实验检测STAT3及磷酸化STAT3水平,ELISA法检测IL-6的分泌水平,间接免疫荧光染色观察STAT3在细胞内定位情况。结果 2或4 Gyγ射线照射均能显著增强A549细胞的迁移能力,并能激活STAT3的磷酸化,促进STAT3入核以及IL-6的分泌。JAK-2激酶特异性抑制剂AG490能阻断辐射诱导的STAT3磷酸化并进一步抑制辐射诱导的A549细胞迁移。结论本研究结果表明辐射能通过激活JAK-2/STAT3信号通路促进A549细胞的迁移,而IL-6可能参与了辐射诱导的JAK-2/STAT3信号通路的激活。     

塞来昔布对人肺腺癌细胞株A549的放射增敏效应及细胞迁移力的影响

目的 观察选择性COX-2抑制剂塞来昔布(celecoxib)对人肺腺癌细胞株A549的放射增敏效应及抑制细胞迁移力的作用。方法 选用人肺腺癌细胞株A549作为研究对象。选择适当浓度的塞来昔布,设置对照组、药物组、单纯放射组和放射加药组,成克隆分析法测定塞来昔布对细胞的放射增敏效应,细胞划痕试验观察A549细胞的迁移力,酶联免疫吸附实验(ELISA)检测细胞培养上清液中基质金属蛋白酶(MMP)-2的含量。结果 细胞划痕试验结果显示,放射加药组比单纯放射组和药物组的无细胞划痕区均增宽,细胞迁移力均明显下降,且随着药物浓度的增加,抑制细胞迁移的能力增强。30和50 μmol/L 塞来昔布对A549细胞显示出浓度依赖性放射增敏作用,放射加药与单纯放射组相比, SF₂、D₀、D\-q出现不同程度的下调。ELISA检测发现,细胞培养上清液中MMP-2的含量,药物组较对照组,放射加药组较单纯放射组均明显下降(t=3.78、5.79,P<0.05),但单纯放射组和对照组对细胞分泌MMP-2的抑制效应不明显(t=2.73、2.38,P>0.05)。结论 塞来昔布在体外试验中对肺腺癌细胞株A549具有浓度依赖性放射敏感性,机制可能为降低了细胞对放射的亚致死损伤修复能力。塞来昔布可能通过抑制A549细胞分泌MMP-2,从而抑制细胞的迁移能力。     

人肺腺癌A549细胞低剂量辐射超敏感性及其机制的研究

目的 观察A549细胞的低剂量辐射超敏感性现象,探讨其发生的机制。方法 A549细胞接受0~2 Gy的⁶⁰Coγ射线照射后,流式细胞仪对其分选计数,克隆形成法检测细胞存活分数,Western blot法检测ATM1981Ser-P蛋白表达,Hoechst 33258荧光染色法、AnnexinⅤ-FITC/PI双染流式细胞仪检测细胞凋亡,PI单染流式细胞仪检测细胞周期。结果 细胞在0~0.3 Gy表现出单位剂量杀伤增强,在0.3~0.5 Gy表现出一定的辐射抗性,0.5 Gy后的区域存活分数随辐射剂量的增加而降低。照射后1 h,ATM激酶在0.2 Gy时开始活化,0.5 Gy时活化达高峰(t=7.96,P<0.05);与0.5 Gy相比1.0和2.0 Gy的活化水平无明显变化(t=0.69、0.55,P>0.05)。照射后24 h,部分细胞发生凋亡,其凋亡曲线与存活曲线相吻合。与未照射组相比,0.1和0.2 Gy组在各时间点(照射后6、12和24 h)的细胞周期无明显变化,而0.3、0.4和0.5 Gy 组,照射后6和12 h细胞发生G₂/M期阻滞(t＝2.87、2.88、4.92和3.70、3.12、8.11,P<0.05),照射后24 h G₂/M期细胞比例下降(t＝3.87、4.77、3.01,P<0.05)。结论 A549细胞存在HRS/IRR现象,其发生可能与ATM激酶、细胞周期变化有关,凋亡是细胞死亡的主要方式。     

咖啡酸苯一酯对人宫颈癌HeLa细胞的放射增敏作用

目的 探讨咖啡酸苯一酯(CAPE)对人宫颈癌HeLa细胞的放射增敏作用。方法 将宫颈癌HeLa细胞经不同浓度的CAPE作用24 h,四甲基偶氮唑盐比色法(MTT)法检测细胞抑制效应与CAPE浓度的关系。将HeLa细胞设对照组和药物组,两组均经⁶⁰Coγ射线照射0、2、4、6和8 Gy,计数细胞克隆;另将HeLa细胞设对照组、CAPE组、单纯照射组、照射+CAPE组,流式细胞检测技术分析CAPE对细胞周期的影响。结果 CAPE对HeLa细胞的抑制率呈剂量依赖性增加(F=126.49~3654.88,P<0.01);细胞经⁶⁰Coγ射线照射后,HeLa细胞克隆存活率随着照射剂量的增加而降低(F=174.42~9422.81,P<0.01);相同剂量下,药物组的HeLa细胞克隆存活率低于对照组(F=120.14~251.91,P<0.01);药物组和对照组HeLa细胞的平均致死剂量(D₀)为1.45和1.82 Gy、准阈剂量(D_q)为1.89和3.21 Gy, 药物组较小,放射增敏比(SER)为1.26>1;与对照组相比, CAPE组及单纯照射组G₂/M期的细胞比例升高(P<0.01),而在照射+CAPE组则降低(P<0.01)。结论CAPE通过对人宫颈癌HeLa细胞G₂/M期的阻滞及可能抑制细胞亚致死性损伤修复能力,发挥放射增敏作用。     

肺癌细胞A549和H460对137Cs γ射线辐射敏感性差异的研究

目的研究肺癌细胞A549和H460对137Cs γ射线的辐射敏感性差异及核因子E2相关因子2（Nrf2）蛋白含量的差异。方法使用2、4、6 Gy 137Cs γ射线照射A549和H460细胞;1、2、4、6 Gy 137Cs γ射线照射H460细胞,用克隆形成法检测细胞增殖能力,单细胞凝胶电泳检测细胞DNA损伤修复情况,casp_1.2.3b1彗星分析软件分析olive尾距值和尾部DNA含量,蛋白质印迹法检测Nrf2蛋白表达量。克隆形成率、olive尾距值和尾部DNA含量采用独立样本t检验进行比较。结果经2、4、6 Gy 137Cs γ射线照射后,肺癌A549细胞的克隆形成率分别为（73.78±14.69）%、（42.26±3.19）%、（17.50±2.18）%;H460细胞的克隆形成率分别为（56.38±6.28）%、（23.82±8.25）%、（4.66±0.87）%,肺癌A549细胞克隆形成率高于H460细胞,且差异均有统计学意义（t=7.99,P=0.015;t=6.75,P=0.019;t=12.03,P=0.005）。4 Gy照射后2 h,肺癌H460细胞的olive尾距值（1.27±0.05）和尾部DNA含量（4.51±0.19）%明显高于A549细胞[0.68±0.04、（2.12±0.14）%],且差异均有统计学意义（t=8.69、10.30,均P < 0.05）。蛋白质印迹实验结果显示,肺癌A549比H460细胞系的Nrf2蛋白丰度高,照射后两种细胞中的Nrf2蛋白水平均升高,但肺癌A549细胞明显高于H460细胞。结论肺癌A549细胞系对137Cs γ射线的辐射抗性强于H460细胞系,这种辐射抗性差异可能与两种细胞系内Nrf2蛋白的含量相关。     

MiR-148a对肺癌细胞放射敏感性的影响

目的探讨miR-148a对肺癌A549、H460以及H1299细胞放射敏感性的影响。方法根据不同的处理方法,分别按以下方式将肺癌A549、H460和H1299细胞进行分组。（1）将肺癌A549和H460细胞分为3组:空白对照组、4 Gy γ射线照射组和8 Gy γ射线照射组。采用实时定量PCR检测miR-148a的表达水平;（2）将肺癌A549细胞分为2组:空白对照组、miR-148a转染组;同时,将肺癌H460细胞分为2组:空白对照组、anti-miR-148a转染组。分别对转染2组细胞进行不同剂量的γ射线照射,并采用克隆形成实验方法来检测细胞增殖;（3）将肺癌A549和H1299细胞分为3组:空白对照组、miR-148a单独处理组、miR-148a和雌激素受体（ER）共转染组。分别对3组细胞进行不同剂量的γ射线照射,并采用克隆形成实验方法检测细胞生长情况。采用Student t-test对数据进行统计学分析,P < 0.05表示差异有统计学意义。结果实时定量PCR实验结果显示,γ射线照射能够显著下调肺癌A549和H460细胞中miR-148a的表达水平。克隆形成实验结果显示,与空白对照组相比,miR-148a转染能够显著增强肺癌A549细胞的放射敏感性,差异有统计学意义（t=12.16,P < 0.01）,而anti-miR-148a转染能够显著降低肺癌H460细胞的放射敏感性,差异有统计学意义（t=11.93,P < 0.01）。同时,miR-148a过表达可以明显下调肺癌A549细胞中ER的mRNA及蛋白表达水平;而anti-miR-148a能够显著上调肺癌H460细胞中ER的mRNA及蛋白表达水平。另外,与miR-148a单独处理组相比,miR-148a和ER共转染组中miR-148a对肺癌A549和H1299细胞增殖的影响显著降低,差异有统计学意义（t=11.34、12.68,均P < 0.01）。结论照射可以诱导miR-148a的表达水平降低,人为过表达miR-148a能够抑制ER的蛋白表达水平,进而降低肺癌A549和H1299细胞的增殖能力,同时增强其放射敏感性。     

严重腹腔感染大鼠JAK/STAT通路活化与多器官功能损害的关系

目的　观察Janus激酶 /信号转导子和转录激活因子 (JAK/STAT )通路在腹腔感染所致脓毒症大鼠组织中的活化规律及其与多器官功能损害的关系。方法　采用盲肠结扎穿孔 (CLP)法制作大鼠脓毒症模型。大鼠随机分为正常对照组、CLP组、JAK 2抑制剂 (AG4 90 )组和STAT 3抑制剂 (雷帕霉素 ,RPM)组。留取肝、肺、肾、肠组织测定STAT1/ 3活性 ,同时测定血清AST、BUN含量及肺组织髓过氧化物酶 (MPO)和肠组织二胺氧化酶 (DAO)活性。结果　CLP后 2h肝、肺、肠组织中STAT1均迅速活化 ,6～ 2 4h达高峰 ,而肾组织STAT1活化相对迟缓。肝、肺组织中STAT3活性亦明显升高 ,但肾、肠组织中未检测到活化的STAT 3。AG4 90、RPM早期处理后 ,除肾组织外 ,其他组织STAT1活性均有不同程度下降 (P <0 0 5或 0 0 1) ,肝、肺组织STAT3活性也显著降低。同时 ,AG4 90、RPM处理组血清AST、BUN水平和肺组织MPO活性在 2 4～ 4 8h均有不同程度下降 (P <0 0 5或 0 0 1) ,但小肠DAO活性无明显改变。结论　STAT1、STAT3在脓毒症时高度活化 ,并参与了严重腹腔感染所致脓毒症的病理过程。抑制JAK/STAT通路活化有助于多器官功能损害的防治。     

电离辐射诱导NF-κB-HIF-1α通路在人淋巴瘤细胞中的活化

目的 观察电离辐射对恶性淋巴瘤细胞中NF-κB-HIF-1α通路的活化状态的影响,探讨恶性淋巴瘤放射抗拒的可能机制.方法 采用Western blot免疫印迹分析方法,检测5 Gy X射线照射1、4、10和20 h后,Namalwa、Ramos和Raji 3种恶性淋巴瘤细胞系中NF-κB、HIF-1α及其相关蛋白IKK的表达水平,观察1、10和50 nmol/L NF-κB抑制剂QNZ对HIF-1α等蛋白表达的影响.结果 照射4 h后,3种淋巴瘤细胞中IKK蛋白表达增加(t=8.01、5.14和5.42,P＜0.01).10 h后,3种淋巴瘤细胞核p-P65蛋白表达增加(t=11.25、17.43和22.09,P＜0.01),至20 h时表达开始下降.10～20 h时,3种淋巴瘤细胞HIF-1α蛋白表达增加,与0 h相比,差异均有统计学意义(t=19.05、12.44和12.88,P＜0.01).与单纯照射组相比,采用NF-κB抑制剂预处理3种肿瘤细胞24 h后,HIF-1α蛋白表达下降,且随药物浓度的增加而下降(t=18.69、19.35和12.26,P＜0.01).结论 电离辐射能够活化恶性淋巴瘤细胞中的NF-κB-HIF-1α通路,可能是肿瘤放射抗拒的机制之一.

Abstract：

Objective To observe the effect of X-ray radiation on NF-κB-HIF-1α pathway activation in three malignant lymphoma and to explore the mechanism of radioresistance mediated by this pathway.Methods Expression levels of p-P65,IKK and HIF-1α in Namalwa,Ramos and Raji cells were determined by Western blot 1,4,10 and 20 h after 5 Gy X-ray irradiation.The effect of QNZ on HIF-1α expression was also observed.Results Expression level of IKK protein was up-regulated 4 h after irradiation in the 3 lymphoma cell strains( t = 8.01,5.14,5.42,P ＜ 0.01 ),followed by up-regulation of p-P65 protein expression level at 10 h and HIF-1α at 10-20 h after X-ray irradiation (t = 11.25,17.43,22.09,P ＜ 0.01 ).Pretreatment of QNZ 24 h before X-ray irradiation significantly reduced the upregulation of HIF-1α protein expression induced by simple ion radiation ( t = 18.69,19.35,12.26,P ＜0.01 ).Conclusion NF-κB - HIF-1α pathway was activated in human lymphoma cells after ion radiation and may be involved in the mechanism of radioresistance.     

HIF-1α基因沉默对A549肺癌细胞放射敏感性的影响

目的 探讨RNA干扰技术致HIF-1α基因沉默对裸鼠肺癌种植瘤放射敏感性的影响。方法 采用以慢病毒为载体的RNA干扰技术获得HIF-1α基因沉默的A549肺癌细胞株,用Western blot方法检测HIF-1α蛋白的表达。将A549/HIF-1α(-)细胞及A549细胞接种于4～6周BALB/C雄性裸小鼠,观察肿瘤生长情况。肿瘤体积达200～350 mm³时分别给予局部单次5、10、15和20 Gy X射线照射,观察对照组和照射组肿瘤体积和肿瘤生长延缓时间。结果 在常氧和乏氧状态下,A549/HIF-1α(-)细胞HIF-1α蛋白表达水平均显著下降,A549/HIF-1α(-)组裸鼠成瘤潜伏期较长,肿瘤生长慢于A549组(P<0.05),X射线照射对两种裸鼠种植瘤的生长均有抑制作用,但A549/HIF-1α(-)组肿瘤生长延缓显著。结论 RNA干扰技术能稳定阻断人肺腺癌A549细胞HIF-1α表达,HIF-1α基因沉默的肿瘤细胞对X射线照射更敏感。