成骨细胞促进急性放射损伤小鼠骨髓造血系统及血管恢复的研究

目的 探讨成骨细胞对放射损伤小鼠骨髓造血系统及血管恢复的影响.方法 取18只雄性BALB/c小鼠股骨制备成骨细胞,其余42只小鼠随机分成健康对照组、成骨细胞组和生理盐水组3组.健康对照组不做任何处理;成骨细胞组和生理盐水组小鼠于6.0 Gy 60 Co γ射线一次性全身均匀照射后,分别经尾静脉输入成骨细胞(2×106个/只)和等体积的生理盐水.于照射后第7、14和21天计数小鼠外周血细胞,骨髓单个核细胞并作骨髓组织学观察.采用流式细胞仪检测骨髓单个核细胞中CD34+细胞百分比,采用免疫组织化学方法检测小鼠骨髓微血管密度.结果 照射后第7、14和21天成骨细胞组小鼠外周血细胞和骨髓单个核细胞计数,骨髓单个核细胞中CD34+细胞百分比,骨髓组织造血面积及骨髓微血管密度均明显高于生理盐水组(t=2.46～64.51,P＜0.05).结论 成骨细胞能促进放射损伤小鼠骨髓造血系统及血管的恢复.

Abstract：

Objective To explore the effects of osteoblasts on the recovery of hematopoiesis and angiogenesis in acute irradiation injury mice.Methods The femurs of 18 male BALB/c mice were used to prepare the bone marrow osteoblasts, and the rest mice were divided into 3 groups as normal group, saline group and osteoblast group.The mice in normal group received no treatment, and the other two groups were received 6.0 Gy 60Co γ-ray irradiation.After irradiation each mouse of osteoblast group was administered with 2 × 106 osteoblasts through tail vein injection, and equal volume saline was given to each mouse of saline group by the same way.The following factors were measured at 7, 14, 21 d after irradiation, they were the counts of peripheral blood cells and bone marrow mononuclear cells ( BMMNC ) , the percentage of CD34 + cells in BMMNC, the histology changes and micro vascular density (MVD) of bone marrow tissue.Results The counts of peripheral blood cells, BMMNC and hematopoietic tissue area in osteoblast group were higher than those in saline group.The percentage of CD34 + cells in BMMNC and the MVD of bone marrow in osteoblast group were also higher than those in saline group at 7, 14, 21 d after irradiation ( t = 2.46 - 64.51, P ＜ 0.05 ).Conclusions Osteoblasts could significantly promote the recovery of hematopoiesis and angiogenesis in mice after acute irradiation injury.     

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.     

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.     

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.     

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.     

血管内皮抑素联合照射对肺癌H-520细胞周期及信号传导通路的影响

Objective To investigate the mechanism of radiosensitizing effects of endostatin on H-520 human lung squamous cancer cells.Methods H-520 cells was treated with endostatin and/or radiation.Colony-forming assays were used to indicate the radiosensitising effects.Cell cycle distribution and expression of phosphor-p38-MAPK were assayed by FCM,and cyclin D1,cdk2,cdk4 and survivin mRNA leveh were assayed by RT-PCR.Phosphor-Akt was evaluated by Western-blotting.Results Combination of endostatin and irradiation inhibited the proliferation of H-520 cells.According to the colony-forming assays,the D0,Dq,D10 and SF2 values of the combination groups were much lower than those of irradiation groups.The sensitization enhancement ratio(SER)was 1.51.G2/M arrest occurred after 4 Gy irradiation.The gene expression of cyclin D1,cdk2,ckd4 and survivin and phosphor-Akt protein were down-regulated after treatment.The expression of phosphor-p38-MAPK protein was also down-regulated after treatment with 200 μg/ml endostar.Conclusions Endostatin inhibits the growth of H-520 cells and radiosensitizes the cells by induction of G0/G1 arrest,cell apoptosis and down-regulation of gene expression of cyelin D1,cdk2,cdk4 and reduces the phosphorylation of Akt and p38-MAPK.     

18氟-氟代脱氧葡萄糖对Lewis肺癌细胞增殖的影响

Objective To investigate the influence of 18F-FDG on the proliferation of Lewis lung cancer cell line,and to elucidate its possible mechanism.Methods Morphological changes of cells after culture for 24 h at different concentrations of 0,0.37,1.85,3.70 and 7.4 (×106) Bq/ml of 18F-FDG were observed by using inverted microscopy and electron microscopy.The apoptosis and phase distribution of cell cycle of irradiated cells were analyzed with flow cytometry.DNA synthesis of irradiated cells was assayed by 3H-TdR incorporation.Lipid peroxidation was measured by chromometry and expression of Bcl-2 and Bax protein was measured by immunohistochemical technique.Results Exposed to (0-7.40) × 106Bq/ml of 18F-FDG for 24 h,the cumulative absorbed doses delivered to cells in five groups were 0,0.11,0.55,1.10 and 2.20 Gy,respectively.Irradiated cells showed morphological changes of apoptosis.The apoptosis rate of irradiated cells was increased from (4.05 ± 0.01)% to (25.6 ± 0.28) % (t = 188,P＜0.01).3H-TdR incorporation rate was decreased from 100% to(22.0 ± 0.51)% (t =27.6,P ＜0.05).The levels of M DA in cells were augmented from (0.08 ± 0.03) to (0.67 ± 0.12) μmol/L (t =11.7,P ＜ 0.01).Cell cycle arrest was found in G2/M phase with the increasing doses from 0 to 2.20 Gy.The expression of Bcl-2 protein was decreased while that of Bax protein increased.Conclusions 18F-FDG could induce the apoptosis of cells and inhibit the proliferation of cells.     

血管内皮抑素联合照射对肺癌H-520细胞周期及信号传导通路的影响

Objective To investigate the mechanism of radiosensitizing effects of endostatin on H-520 human lung squamous cancer cells.Methods H-520 cells was treated with endostatin and/or radiation.Colony-forming assays were used to indicate the radiosensitising effects.Cell cycle distribution and expression of phosphor-p38-MAPK were assayed by FCM,and cyclin D1,cdk2,cdk4 and survivin mRNA leveh were assayed by RT-PCR.Phosphor-Akt was evaluated by Western-blotting.Results Combination of endostatin and irradiation inhibited the proliferation of H-520 cells.According to the colony-forming assays,the D0,Dq,D10 and SF2 values of the combination groups were much lower than those of irradiation groups.The sensitization enhancement ratio(SER)was 1.51.G2/M arrest occurred after 4 Gy irradiation.The gene expression of cyclin D1,cdk2,ckd4 and survivin and phosphor-Akt protein were down-regulated after treatment.The expression of phosphor-p38-MAPK protein was also down-regulated after treatment with 200 μg/ml endostar.Conclusions Endostatin inhibits the growth of H-520 cells and radiosensitizes the cells by induction of G0/G1 arrest,cell apoptosis and down-regulation of gene expression of cyelin D1,cdk2,cdk4 and reduces the phosphorylation of Akt and p38-MAPK.     

Artemis蛋白磷酸化对紫外线C引发的复制检测点的调控

Objective To investigate Artemis phosphorylation on S516 and S645 in response to stalled replication forks and its role in regulation of cell cycle replication checkpoint.Methods Western-blotting was used to measure the expression of phosphorylation of Artemis on S516 and S645 after UVC irradiation.The nonphosphorylatable double mutant (S516-645A) and the mimicking phosphorylation mutant (S516-645D) plasmids were constructed.HEK 293 cells with stable expression of wild type Artemis and the corresponding mutants were established by transfection.Cell cycles of the cells treated with UVC irradiation were analyzed by flow cytometry,Western-blotting was used to measure the expression of Chk1,γ-H2AX and Cdk2.IP-kinase assay was used to measure the kinase activity of Cdk2 2,6 and 12 h after UVC irradiation.Results Artemis got rapid and prolonged pbosphorylation on S516 and S645 after treatment with UVC irradiation and the major responsible kinase was ATR.The S516-645A mutant caused prolonged arrest in replication checkpoint in S phase.The Cdk2 IP kinase activity was inhibited in S516-645A mutant ceils,but the expression levels of Chk1,Cdk2 and γ-H2AX were not affected.Conclusion The ATR phosphorylation on S516 and S645 of Artemis promotes cell cycle recovery from UVC induced replication checkpoint.     

电离辐射对原代成骨细胞M-CSF表达的影响

目的 研究辐射对于原代成骨细胞巨噬细胞集落刺激因子（M-CSF）表达的影响,进而研究辐射导致骨损伤的分子机理。 方法 采用原代骨髓间充质干细胞诱导分化为成骨细胞,经0、2、4 Gy 137Cs γ射线照射后,采用实时定量PCR以及Western blot方法检测辐射对M-CSF mRNA和蛋白表达水平的影响。 结果 2 Gy和4 Gy γ射线照射均能引起成骨细胞M-CSF mRNA（t=-17.329, P < 0.01;t=-3.841, P < 0.05）和蛋白表达水平上调。4 Gy照射后则会导致成骨前体细胞M-CSF mRNA表达水平上调（t=-4.478, P < 0.05）,但与对照组比较,两者的蛋白表达水平未见显著差异。 结论 2 Gy和4 Gy γ射线照射后,成骨细胞M-CSF表达水平上调能增强核因子κB受体活化因子配体对破骨细胞分化、成熟的促进作用,有助于增强破骨细胞的骨吸收能力。     

NICD表达下调对辐射损伤小鼠成骨细胞系MC3T3-E1增殖和功能基因表达的影响

目的利用RNA干扰抑制小鼠成骨细胞系MC3T3-E1表达Notch信号通路胞内结构域（NICD）,探讨靶向抑制NICD表达对辐射损伤MC3T3-E1细胞的增殖和相关功能基因表达的影响。方法建立抑制NICD表达的MC3T3-E1细胞株,利用实时定量PCR（qRT-PCR）和Western blot法检测其NICD基因的表达。MC3T3-E1细胞和NICD RNA干扰MC3T3-E1细胞经2 Gy γ射线照射后,用BrdU掺入法和qRT-PCR法检测上述细胞的增殖及相关功能基因的表达水平。使用Student-Newman-Keuls进行组间差异分析,两组间比较采用t检验。结果用RNA干扰技术可靶向抑制MC3T3-E1细胞表达NICD。抑制NICD表达可干扰前体成骨细胞和成骨细胞的增殖。2 Gy照射后,前体成骨细胞和成骨细胞以及NICD RNA干扰的成骨细胞的增殖明显下降,各靶细胞的相关功能基因与照射前相比的变化如下:①2 Gy照射后的前体成骨细胞成骨特导性转录因子（Runx2）表达上调,差异有统计学意义（t=2.353,P < 0.05）,NICD RNA干扰的前体成骨细胞Runx2表达下调,差异有统计学意义（t=2.353,P < 0.05）;②2 Gy照射后的前体成骨细胞和成骨细胞以及NICD RNA干扰的前体成骨细胞碱性磷酸酶（ALP）表达上调,差异有统计学意义（t=3.182、3.345、3.555,均P < 0.05）,NICD RNA干扰的成骨细胞ALP表达下调,差异有统计学意义（t=5.045,P < 0.01）;③2 Gy照射后前体成骨细胞核因子κB受体活化因子配体（RANKL）表达下调,差异有统计学意义（t=2.541,P < 0.05）,成骨细胞和NICD干扰的前体成骨细胞RANKL表达上调,差异有统计学意义（t=3.299,P < 0.05;t=10.212,P < 0.01）,而抑制NICD表达则发生相反变化,差异无统计学意义（t=0.765,P>0.05）;④2 Gy照射后的前体成骨细胞和成骨细胞骨保护素（OPG）表达下调,差异有统计学意义（t=2.994、2.782,均P < 0.05）,抑制NICD表达使前体成骨细胞OPG表达上调,差异有统计学意义（t=5.841,P < 0.01）,成骨细胞OPG表达下调,差异有统计学意义（t=2.544,P < 0.05）;⑤2 Gy照射后各靶细胞巨噬细胞集落刺激因子（M-CSF）表达变化趋势与RANKL表达变化情况一致。结论在不同阶段的成骨细胞中抑制NICD表达对辐射损伤表现出的作用是不同的:①可降低前体成骨细胞和成骨细胞的增殖,对辐射损伤后的前体成骨细胞的增殖有保护作用;②可通过调节Runx2从而明显抑制辐照后前体成骨细胞分化,减少骨质丢失;③辐照后各成骨细胞不会通过RANKL/OPG/RANK系统表现出对破骨细胞功能的调节作用;④成骨细胞经过调节M-CSF表现出对破骨细胞的功能抑制作用。     

电离辐射对大鼠海马区神经发生及TrkA和TrkB表达的影响

目的 探讨电离辐射对大鼠海马区神经发生以及TrkA、TrkB蛋白表达的影响。方法 将56只SD大鼠按随机数字表法分为照射组（28只）和健康对照组（28只）,照射组给予单次10 Gy全脑照射。分别于照后1、3 d,2周以及1个月取海马组织,应用免疫荧光染色观察神经元增殖变化,Golgi染色观察海马树突棘形态变化,Western blot及RT-PCR分别检测TrkA、TrkB蛋白及RNA水平变化。结果 与健康对照组比较,免疫荧光染色显示照射组双皮质素（DCX）数量明显减少（t=6.49,P<0.05）。照射组海马树突棘明显减少,且树突棘的形态变化明显。与健康对照组相比,照后不同时间照射组TrkA蛋白表达明显升高（t=2.64、3.06、4.80、2.64, P<0.05）,而TrkB的表达则显著下降（t=4.59、3.06、2.81、2.57, P<0.05）;TrkA mRNA表达水平明显升高（t=4.57、3.06、5.39、5.86, P<0.05）,TrkB的表达显著下降（t=14.87、11.69、4.98,P<0.05）。结论 作为神经生长因子、脑源性神经因子重要的下游信号通路分子,全脑照射后TrkA、TrkB的表达变化,可能在电离辐射所致海马神经发生损伤中发挥重要作用。     

富氢水减轻造血干祖细胞的辐射损伤

目的 探讨富氢水对辐射诱导的造血干祖细胞（HSPCs）损伤的保护作用。方法 32只C57BL/6小鼠根据体重分层随机区组法分为健康对照组、富氢水组、照射组、照射+富氢水组,共4组,每组8只。富氢水组和照射+富氢水组小鼠于照射前5 min至照后7 d,每天灌胃给予0.5 ml富氢水,其余小鼠每天灌胃给予0.5 ml蒸馏水,照射组和照射+富氢水组小鼠接受2 Gy的¹³⁷Cs γ射线全身照射。照后15 d取小鼠骨髓,检测骨髓中HSPCs比例、骨髓细胞的克隆形成和移植重建能力、骨髓中LSK细胞的活性氧（ROS）水平和细胞凋亡情况。结果 与照射组相比,照射+富氢水组小鼠骨髓中造血祖细胞和LSK细胞比例升高（t=-4.935、-7.898,P<0.05）,骨髓细胞形成克隆的数目增加（t=5.488,P<0.05）,竞争性骨髓移植后受体小鼠的供体嵌合率升高（t=-12.769,P<0.05）,骨髓中LSK细胞的ROS水平和细胞凋亡比例降低（t=4.380、3.954,P<0.05）。结论 富氢水对2 Gy电离辐射诱导的HSPCs损伤具有一定的保护作用。     

电离辐射对不同放射敏感性细胞中高迁移率簇蛋白B1的诱导表达作用

目的 探讨电离辐射对已建立的宫颈癌放射抗拒细胞对比模型中高迁移率簇蛋白B1（HMGB1）和mRNA诱导表达的差异性,分析HMGB1对宫颈癌放射敏感性调控的可能性。方法人宫颈癌细胞系HeLa重复照射12次后,传代培养调整细胞状态至细胞增殖稳定,筛选得抗拒细胞系HeLaR。以2、5、10 Gy X射线分别照射亲代HeLa和HeLaR细胞,于照射后0、0.5、2、4、6、12、18、24、36、48 h收集细胞,提取蛋白质和RNA,采用Western blot和实时荧光定量PCR法分别检测样本中HMGB1蛋白和mRNA的表达情况。结果 在蛋白水平,2、5、10 Gy X射线照射后,HeLaR细胞在48 h内均表现为HMGB1表达量下降,在48 h达到未照射水平,后有增加趋势,与照射后0 h比较,2、5、10 Gy照射后6~36 h各时间点,差异具有统计学意义（t=3.574~9.754,P < 0.05）;相反,HeLa细胞在照射后6 h,其HMGB1表达逐渐增多,尤其在5和10 Gy表现明显,与照射后0 h比较,2 Gy照射后6、12、48 h（t=3.945~4.864,P < 0.05）、5 Gy照射后6、36、48 h（t=-2.875~3.295,P < 0.05）及10 Gy照射后36、48 h（t=-4.480、-4.517,P < 0.05）,差异具有统计学意义。在mRNA水平其趋势与蛋白水平基本一致。结论 不同剂量X射线照射后可诱导人宫颈癌细胞中HMGB1的表达变化,且其变化在人宫颈癌放射敏感细胞及放射抗拒细胞中不同。HMGB1可能参与人宫颈癌放射抗拒机制。     

抑制FOXD1基因表达增强结直肠癌细胞HCT116的放射敏感性

目的 研究抑制FOXD1基因的表达对结直肠癌细胞放射敏感性的影响。方法 采用实时荧光定量聚合酶链反应（qRT-PCR）和Western blot检测人结直肠癌组织和细胞中FOXD1 mRNA和蛋白的表达。对结直肠癌HCT116细胞行梯度剂量（0、2、4、6 Gy）X射线照射,qRT-PCR和Western blot检测各组细胞中FOXD1的表达。将siRNA阴性对照和FOXD1 siRNA转染至结直肠癌细胞中,分别记为si-NC组和si-FOXD1组,经4 Gy的X射线照射处理后记为si-NC+4 Gy组和si-FOXD1+4 Gy组,Western blot检测各组细胞中FOXD1的表达,四甲基偶氮唑盐（MTT）法检测各组细胞的增殖活性,克隆形成实验检测各组细胞存活率,采用TECT DNA-PK试剂盒检测各组细胞中DNA-PK活性,将转染的结直肠癌细胞接种于BALB/c裸鼠建立移植瘤模型,进行射线照射后,检测各组肿瘤的体积和质量变化。结果 与癌旁正常组织相比,结直肠癌组织中FOXD1 mRNA和蛋白的表达均显著增加（t=5.579、4.816,P<0.05）,与结肠黏膜上皮细胞NCM460相比,结直肠癌细胞株中FOXD1 mRNA（t=5.85~17.62,P<0.05）和蛋白（t=9.04~11.42,P<0.05）表达均显著升高。结直肠癌细胞HCT116中FOXD1的表达量随着放射剂量增加而升高,呈现剂量依赖性,差异有统计学意义（t=9.13~44.15,P<0.05）。转染si-FOXD1能够有效地抑制结直肠癌细胞中FOXD1的表达（t=10.51,P<0.05）,FOXD1敲低后能够抑制结直肠癌细胞的增殖活性（t=10.41,P<0.05）,提高结直肠癌细胞的放射敏感性,放射增敏比为1.797,降低放射诱导的DNA-PK的活性（t=6.20,P<0.05）。抑制FOXD1的表达经射线照射后,裸鼠种植瘤体积和重量明显减小（t=11.29、3.69,P<0.05）。结论 抑制FOXD1基因的表达能够提高结直肠癌细胞的放射敏感性,抑制结直肠癌裸鼠移植瘤的生长,可为改善放射治疗对结直肠癌患者的治疗效果提供潜在的靶向基因。     

缝隙连接蛋白43在受照人血管内皮细胞中的改变及其对受照细胞刚性的作用

目的:研究X射线对人脐静脉内皮细胞(HUVEC)中缝隙连接蛋白43(Cx43)的表达、分布和细胞刚性的影响,初步探讨Cx43对受照细胞刚性的调控作用。方法:采用Western blot方法检测10 Gy X射线照射后不同时间(0、6、12、24和48 h)和不同剂量X射线(0、2.5、5、10和20 Gy)照射后12 ...     

缝隙连接蛋白43在X射线诱导的人脐静脉内皮细胞凋亡中的作用及机制研究

目的 研究缝隙连接蛋白43（Cx43）在X射线致人脐静脉内皮细胞（HUVEC）凋亡中的作用并探讨其机制。方法 采用流式细胞术检测10 Gy X射线照射后48~96 h以及不同剂量X射线照射后72 h HUVEC细胞凋亡的变化。Western blot检测0、5、10、20 Gy X射线照射后72 h HUVEC中Cx43和cleaved caspase-3蛋白的表达。采用RNA干扰技术使细胞Cx43表达沉默,或转染Cx43高表达质粒使Cx43过表达。流式细胞术和Western blot分别检测Cx43沉默和过表达对受照HUVEC凋亡的影响以及cleaved caspase-3蛋白表达水平的变化。结果 X射线照射后48~96 h HUVEC凋亡增加并且呈现剂量依赖性。在0~20 Gy范围内,Cx43表达随剂量增加而降低,cleaved caspase-3表达随剂量增加而增高。Cx43沉默组HUVEC细胞早期凋亡及凋亡死亡细胞比例明显高于无意义序列组（t=3.674、6.375,P<0.05）;Cx43过表达组HUVEC细胞早期凋亡及凋亡死亡细胞比例明显低于空载体组（t=9.399、11.190,P<0.05）;Cx43沉默组较无意义序列组cleaved caspase-3表达增强,而过表达组低于空载体组。结论 Cx43通过调控cleaved caspase-3活化,抑制HUVEC细胞凋亡,从而对X射线照射的HUVEC细胞发挥保护作用。     

丰富环境对辐射诱导小鼠认知障碍的保护作用及可能机制

目的 研究丰富环境对辐射诱导小鼠认知功能障碍的保护作用及其可能机制。方法 将45只2月龄雌性昆明小鼠采用随机数表法分为对照组、照射组和照射丰富环境组,每组15只。照射组和照射丰富环境组予以单次4 Gy全身¹³⁷Cs γ射线照射,照射丰富环境组辐射后连续35 d给予丰富环境刺激。新旧事物识别实验检测小鼠认知功能;免疫组织化学方法检测小鼠海马区小胶质细胞标记物IBA-1的表达;Western blot方法检测小胶质细胞激活标记物CD68及突触囊泡素（SYP）的表达。结果 与对照组相比,照射组小鼠在新旧事物识别实验中新事物分辨率降低,海马区IBA-1阳性细胞数目增加,CD68蛋白表达升高,SYP蛋白表达降低（t=3.66、6.83、5.79、6.84,P<0.05）。与照射组相比,照射丰富环境组小鼠新事物分辨率升高,海马区IBA-1阳性细胞数目减少,CD68蛋白表达降低,SYP蛋白表达增加（t=3.56、7.69、4.59、4.06,P<0.05）。结论 4 Gy单次全身¹³⁷Cs γ射线照射可构建放射性认知功能障碍模型,丰富环境可改善模型小鼠认知功能,其机制可能与抑制海马区小胶质细胞激活以及减少神经元突触丢失有关。