微波与γ射线联合的细胞生物学效应
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摘要
目的:以人神经胶质瘤细胞株(SHG44)为生物模型,研究900MHz微波单独及联合γ射线照射对细胞的生物学效应。
方法:将细胞随机分为对照组、2mW/cm2、4mW/cm2和6mW/cm2微波暴露组(各组记为C,M2,M4和M6)。细胞辐照2h/d,连续照射3d后,联合组再接受5Gy的γ射线照射。共分为单独γ射线组、2mW/cm2+γ射线组、4mW/cm2+γ射线组和6mW/cm2+γ射线组(记为I,I+M2,I+M4和I+M6)。γ射线(由苏州大学辐照中心提供)采用60Co辐照,照射剂量5Gy,剂量率为1Gy/min。(1)采用MTT法和克隆法测定细胞的生长增殖情况。(2)利用流式细胞仪检测细胞的周期和凋亡情况。(3)测定各组细胞超氧化物歧化酶(superoxide dismutase,SOD)活力和丙二醛(malondialdehyde,MDA)的含量。(4)用RT-PCR方法和western blot法检测各组细胞中HSP70在mRNA和蛋白水平的表达。(5)以正常大鼠神经胶质细胞为研究对象,选取部分上述指标(MTT法和凋亡)进行检测。(6)用BPCL微弱发光测量仪检测细胞在各生长阶段,微波处理后超微弱发光的变化。
结果:(1)单独微波辐射中,M4和M6组的细胞增殖活性显著下降。联合照射组中,I+M6组细胞的增殖活性与单独γ射线组相比显著下降。各单独微波组细胞的克隆形成率均下降。联合照射组中,与I组相比,I+M4与I+M6组的克隆率显著下降。微波和γ射线对增殖活性和克隆率有协同抑制作用。(2)与单独微波组相比,联合组G1期百分比显著升高,而G2期和S期比例均有所下降,但无统计学差异。单独微波辐射后,细胞凋亡率随着微波强度的增加有上升的趋势且呈直线相关,但各组间无统计学差异。与I组相比,I+M4和I+M6组的凋亡率显著升高。与单独微波组相比,各联合组凋亡率显著上升。微波与γ射线对细胞凋亡率存在协同促进作用。(3)单独微波组中,M6组的MDA含量上升,各单独微波组SOD活性也有上升趋势,但各组间没有统计学差异。与I组相比,各联合组MDA含量均显著上升,I+M6组的SOD活性显著下降。微波与γ射线对MDA含量有协同促进作用,对SOD活性的抑制有加强作用。(4)单独微波照射后,HSP70的表达有上升的趋势,但各组间无统计学差异。γ射线照射后HSP70表达显著下降。微波和γ射线对细胞HSP70的表达无交互作用。(5)大鼠原代神经胶质细胞的增殖活性经微波、γ射线和联合照射处理后,与对照组相比均没有统计学差异,仅γ射线组和联合组有下降的趋势。各处理组均无明显凋亡发生。γ射线使ATP酶的活性显著升高,微波和γ射线对ATP酶活性无交互作用。(6)细胞的超微弱发光在接种后第1d即显著增强,一直维持到第5d后显著下降。微波辐射后第1d发光强于对照组,第2d恢复到对照水平。H_2O_2处理后细胞超微弱发光显著增强。
结论:(1)微波和γ射线均能抑制人神经胶质瘤细胞的增殖,且两者之间有协同抑制作用。微波和γ射线还可对细胞氧化损伤产生协同和加强的作用。(2)联合照射未明显抑制大鼠原代神经胶质细胞的增殖和促进其凋亡的发生,但可使ATP酶活性增高。(3)细胞增殖和氧化剂处理能使细胞的超微弱发光明显增强,微波引起细胞发光增强为早期一过性的。
Objective: To study biological effects on SHG44 cells combined by 900MHz microwave andγ-ray radiation.
Methods: SHG44 cells were divided into four groups according to the power density of microwave(0 mw/cm2,2mw/cm2,4mw/cm2 and 6mw/cm2),named as C,M2,M4 and M6,respectively.The cells were exposed to microwave 2h/d for 3d,and then were irradiated byγ-ray (5Gy),whit four groups named as I,I+M2,I+M4 and I+M6,respectively.(1) Cellular proliferation was measured with MTT test and cloning efficiency.(2) Cell cycle and apoptosis were detected by flow cytometer.(3) SOD activity and MDA level were tested by commercial kits.(4) Expression of HSP70 gene was evaluated by RT-PCR and western blot.(5) Normal rat glial cells were cultured and cellular proliferation , apoptosis and ATPase activity were measured.(6) Ultraweak luminescence of cells was measured by the Ultraweak Luminescence Analyzer.
Results: (1) Cell proliferation activity in Group M4 and M6 and cloning efficiency in all exposed groups were decreased by the microwave radiation,with a significant decrease in Group I+M4 and I+M6.Combination of Microwave andγ-ray showed synergistic effect on both cell proliferation activity and the cloning efficiency.(2) Compared to the microwave radiation groups,the percentage of G1 phase cells obviously increased in the combination groups,while that of G2 and S phase decreased.The rate of apoptosis increased with the does of microwave in the microwave radiation groups and so did the I+M4 and I+M6 Group,indicating a synergistic effect ofγ-ray and microwave.(3) The MDA level increased in Group M6 and in all combination groups,and the SOD activity decreased in Group I+M6.(4) Expression of HSP70 at both mRNA and protein levels increased slightly in the microwave exposure groups,but no interaction ofγ-ray and microwave on HSP70 expression was found.(5) Proliferation of the rat glial cell in Group I and I+M6 decreased slightly,and the ATPase activity was significantly increased.(6) Ultraweak luminescence of the exposed cells was stronger than the control cells on the first day,and returned to normal on the second day.
Conclusion: Microwave andγ-ray could inhibit the proliferation of SHG44 cells with a synergistic effect,but such effect was not observed in primary rat glial cells.Ultraweak luminescence could be strengthened by microwave at early stages of exposure.
方法:将细胞随机分为对照组、2mW/cm2、4mW/cm2和6mW/cm2微波暴露组(各组记为C,M2,M4和M6)。细胞辐照2h/d,连续照射3d后,联合组再接受5Gy的γ射线照射。共分为单独γ射线组、2mW/cm2+γ射线组、4mW/cm2+γ射线组和6mW/cm2+γ射线组(记为I,I+M2,I+M4和I+M6)。γ射线(由苏州大学辐照中心提供)采用60Co辐照,照射剂量5Gy,剂量率为1Gy/min。(1)采用MTT法和克隆法测定细胞的生长增殖情况。(2)利用流式细胞仪检测细胞的周期和凋亡情况。(3)测定各组细胞超氧化物歧化酶(superoxide dismutase,SOD)活力和丙二醛(malondialdehyde,MDA)的含量。(4)用RT-PCR方法和western blot法检测各组细胞中HSP70在mRNA和蛋白水平的表达。(5)以正常大鼠神经胶质细胞为研究对象,选取部分上述指标(MTT法和凋亡)进行检测。(6)用BPCL微弱发光测量仪检测细胞在各生长阶段,微波处理后超微弱发光的变化。
结果:(1)单独微波辐射中,M4和M6组的细胞增殖活性显著下降。联合照射组中,I+M6组细胞的增殖活性与单独γ射线组相比显著下降。各单独微波组细胞的克隆形成率均下降。联合照射组中,与I组相比,I+M4与I+M6组的克隆率显著下降。微波和γ射线对增殖活性和克隆率有协同抑制作用。(2)与单独微波组相比,联合组G1期百分比显著升高,而G2期和S期比例均有所下降,但无统计学差异。单独微波辐射后,细胞凋亡率随着微波强度的增加有上升的趋势且呈直线相关,但各组间无统计学差异。与I组相比,I+M4和I+M6组的凋亡率显著升高。与单独微波组相比,各联合组凋亡率显著上升。微波与γ射线对细胞凋亡率存在协同促进作用。(3)单独微波组中,M6组的MDA含量上升,各单独微波组SOD活性也有上升趋势,但各组间没有统计学差异。与I组相比,各联合组MDA含量均显著上升,I+M6组的SOD活性显著下降。微波与γ射线对MDA含量有协同促进作用,对SOD活性的抑制有加强作用。(4)单独微波照射后,HSP70的表达有上升的趋势,但各组间无统计学差异。γ射线照射后HSP70表达显著下降。微波和γ射线对细胞HSP70的表达无交互作用。(5)大鼠原代神经胶质细胞的增殖活性经微波、γ射线和联合照射处理后,与对照组相比均没有统计学差异,仅γ射线组和联合组有下降的趋势。各处理组均无明显凋亡发生。γ射线使ATP酶的活性显著升高,微波和γ射线对ATP酶活性无交互作用。(6)细胞的超微弱发光在接种后第1d即显著增强,一直维持到第5d后显著下降。微波辐射后第1d发光强于对照组,第2d恢复到对照水平。H_2O_2处理后细胞超微弱发光显著增强。
结论:(1)微波和γ射线均能抑制人神经胶质瘤细胞的增殖,且两者之间有协同抑制作用。微波和γ射线还可对细胞氧化损伤产生协同和加强的作用。(2)联合照射未明显抑制大鼠原代神经胶质细胞的增殖和促进其凋亡的发生,但可使ATP酶活性增高。(3)细胞增殖和氧化剂处理能使细胞的超微弱发光明显增强,微波引起细胞发光增强为早期一过性的。
Objective: To study biological effects on SHG44 cells combined by 900MHz microwave andγ-ray radiation.
Methods: SHG44 cells were divided into four groups according to the power density of microwave(0 mw/cm2,2mw/cm2,4mw/cm2 and 6mw/cm2),named as C,M2,M4 and M6,respectively.The cells were exposed to microwave 2h/d for 3d,and then were irradiated byγ-ray (5Gy),whit four groups named as I,I+M2,I+M4 and I+M6,respectively.(1) Cellular proliferation was measured with MTT test and cloning efficiency.(2) Cell cycle and apoptosis were detected by flow cytometer.(3) SOD activity and MDA level were tested by commercial kits.(4) Expression of HSP70 gene was evaluated by RT-PCR and western blot.(5) Normal rat glial cells were cultured and cellular proliferation , apoptosis and ATPase activity were measured.(6) Ultraweak luminescence of cells was measured by the Ultraweak Luminescence Analyzer.
Results: (1) Cell proliferation activity in Group M4 and M6 and cloning efficiency in all exposed groups were decreased by the microwave radiation,with a significant decrease in Group I+M4 and I+M6.Combination of Microwave andγ-ray showed synergistic effect on both cell proliferation activity and the cloning efficiency.(2) Compared to the microwave radiation groups,the percentage of G1 phase cells obviously increased in the combination groups,while that of G2 and S phase decreased.The rate of apoptosis increased with the does of microwave in the microwave radiation groups and so did the I+M4 and I+M6 Group,indicating a synergistic effect ofγ-ray and microwave.(3) The MDA level increased in Group M6 and in all combination groups,and the SOD activity decreased in Group I+M6.(4) Expression of HSP70 at both mRNA and protein levels increased slightly in the microwave exposure groups,but no interaction ofγ-ray and microwave on HSP70 expression was found.(5) Proliferation of the rat glial cell in Group I and I+M6 decreased slightly,and the ATPase activity was significantly increased.(6) Ultraweak luminescence of the exposed cells was stronger than the control cells on the first day,and returned to normal on the second day.
Conclusion: Microwave andγ-ray could inhibit the proliferation of SHG44 cells with a synergistic effect,but such effect was not observed in primary rat glial cells.Ultraweak luminescence could be strengthened by microwave at early stages of exposure.
引文
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