松胞素B对辐射诱导淋巴细胞核质桥水平影响的研究

目的 探索胞质分裂阻滞分析中,松胞素B终浓度和加入时间对辐射诱导淋巴细胞核质桥水平的影响。方法 本研究按照不同松胞素B终浓度分为2、4、6、8、10 μg/ml 5个终浓度组,此外,按照不同松胞素B加入时间分为0、28、40、44 h组4个加入时间组。用2 Gy（吸收剂量率为1 Gy/min）⁶⁰Co γ射线照射正常人离体外周血（设0 Gy对照组）,采用胞质分裂阻滞分析法,进行细胞培养、收获、制片、染色。在光学显微镜下分析单核细胞、双核细胞、多核细胞的比例,以及双核细胞中核质桥率及微核率。结果 各终浓度组和加入时间组（0 h组除外）的核分裂指数、双核细胞比例均随松胞素B终浓度的升高而升高,随松胞素B加入时间的推迟而降低。2 Gy射线照射后,各终浓度组和加入时间组（0 h组除外）核质桥率差异无统计学意义（P>0.05）;5个终浓度组间微核率具有随松胞素B终浓度的增加而下降的趋势,4个加入时间组间微核率差异无统计学意义（P>0.05）。结论 不同松胞素B终浓度和加入时间可使辐射诱导的核质桥率在一定范围内变化,但差异无统计学意义。适当增加松胞素B终浓度、提前加入时间可使双核细胞比例升高,有助于提高分析效率。     

低剂量放疗所致离体外周血单核细胞凋亡呈现不连续剂量依赖性

目的 :观察在抗炎放疗剂量范围内低剂量辐射对外周血单核细胞 ( PBMC)凋亡的诱导及其可能的免疫调节机制。方法 :1标本 :PBMC取自 10名健康志愿者 ( 8男、2女 ,2 6～ 39岁 )。 2照射 :PBMC/ RPMI(标准培养基 )为 1× 10 6 /m l于 37℃照射 ,2 5 0 k V、15 m A,球靶距 40 cm,其中 9份样品以 0 .1～ 3.0 Gy单次照射 ,另 1份在 0 .1～ 1.0 Gy间以 0 .1Gy梯度增加的单次照射 ,剂量率均为 1.15 Gy/ m in,照射后培养12～ 78小时进行流式细胞分析。 3流式细胞仪检测 :用碘化丙锭 ( PI) / Triton染胞核 ,测定凋亡胞核 ;用膜联蛋白 V( Ax …     

低剂量照射TK6和U937细胞的效应:诱导p53及其对细胞周期延迟和适应性反应的作用

目的 :研究小剂量辐射对 TK6 (含有野生型 p5 3的淋巴母细胞 )和 U937(p5 3突变的单核粒细胞白血病细胞 )细胞周期的影响和诱导对随后大剂量的适应性反应。方法 :TK6和 U 937细胞系在 RPMI 16 40培养液中悬浮培养。所有实验接种密度为 2× 10 5 / ml的细胞悬液 ,在实验前 2 0 h照射 ,用剂量率为 0 .75和 0 .88Gy min- 1 的 6 0 Coγ射线 ,第一次照射诱导剂量 ,随后在不同时间给与攻击剂量。细胞周期分布和周期素用双色流式细胞仪测定。用 Western-blot方法测量 p5 3和 p2 1。用胞质分裂阻滞技术测定微核。按核质破碎和浓缩等核形态学…     

低剂量60Coγ射线诱导人淋巴细胞核质桥适应性反应的研究

目的 以核质桥(NPB)为主要指标,探索低剂量⁶⁰Co γ射线是否能诱导人外周血淋巴细胞的适应性反应及诱导剂量范围。方法 用⁶⁰Co γ射线照射健康成年男子离体外周血,照射剂量分别为0、20、50、75、100、150和200 mGy(吸收剂量率为25 mGy/min),照射后间隔6 h后再给予2 Gy照射(吸收剂量率为1 Gy/min)。采用胞质分裂阻滞法(CBMN)进行细胞培养,观察NPB及微核(MN)的发生情况。结果 0~200 mGy剂量范围内,NPB和MN数目随吸收剂量的增加而增多,并拟合出NPB的线性平方模型y=(1.5×10^-4)x²-(5.67×10^-3)x+0.598 (R²=0.893 8)。提前给予75~100 mGy照射比直接受到2 Gy照射产生的NPB及MN数目均有所减少(U=2.66、2.97、3.96、5.89,P<0.05),在100 mGy照射后NPB减少最多(43.2%)。结论 低剂量⁶⁰Co γ射线可以诱导人外周血淋巴细胞的适应性反应,诱导剂量范围为75~100 mGy。     

60Co γ射线全身均匀照射大鼠早期血浆辐射敏感脂质筛选研究

目的探索大鼠受到全身照射后血浆脂质代谢特征, 为辐射生物标志物研究提供科学依据。方法非靶向脂质组学研究中, 将50只SD大鼠分为6组, 用0、1、2、3、5、8 Gy钴60 γ射线进行全身照射;靶向脂质组学研究中, 将25只大鼠分为5组, 用0、0.5、2.5、4、6 Gy射线进行照射。照射后4 h, 采集静脉血并分离血浆, 筛选辐射敏感脂质并测定其浓度, 进行受试者工作特征曲线(ROC)和剂量-效应关系分析。结果非靶向脂质组学研究中, 共筛选出15个辐射差异脂质;经靶向脂质组学验证, 其中7个可作为辐射敏感脂质。辐射敏感脂质ROC区分0 Gy组与> 0 Gy组、< 2 Gy组与≥ 2 Gy组样本曲线下面积(AUC)均> 0.75;将其组合后进行ROC分析, AUC值提高为0.96和0.94。在0～6 Gy范围内, LysoPC(18:2)、LysoPC(22:0)、PC(18:0/18:2)、PE(18:2/16:0)和PE(18:2/18:0)浓度随照射剂量的上升而下降。结论大鼠受照后4 h, 共筛选出7个血浆辐射敏感脂质, 其组合可用于特定照射剂量分类, 其中5...     

高LET辐射诱导共济失调性毛细血管扩张症患者成淋巴类细胞凋亡的研究

目的 :探讨和比较共济失调性毛细血管扩张症 (A- T)患者和正常人 E- B病毒转化成淋巴类细胞 (L CL )在高传能线密度 (L ET)照射后所致细胞凋亡的特性。方法 :1细胞株 :采用 4种 A- T患者 L CL (ARO、BMA、CSA和 RJO)及 2种正常人 L CL (JAC和 KKB3)株。 2照射条件 :采用 32～ 45 Me V/u(8～ 12 Gy/min)快中子照射 ,并以低 L ET1.36 Gy/min的 X射线照射作比较 ,照射剂量为 1～4Gy。 3在照射后 0～ 48h,采用倒转脉冲电场凝胶电泳(FIGE)及荧光染色两种方法检测 L CL 凋亡。 4应用流式细胞仪测定细胞周期分布。 5 Western…     

c-myc和c-fos基因表达可能是肾晚期辐射损伤的预兆

目的 :测定 c- myc和 c- fos基因在肾照射 2 4h后的表达情况 ,确定该表达与肾晚期辐射损伤之间是否相关。方法 :实验采用了 15周龄、雌性 C3H/ He Slc小鼠作为研究对象 ,线性加速器 ML - 2 0 MDX产生 4MV X射线 ,以约1.0 Gy/ m in的剂量率进行左肾单侧照射 ,总剂量分别为 9、12和 15 Gy,用逆转录酶 -多聚酶链反应 (RT- PCR)技术测定 c-myc和 c- fos基因在肾接受 9、12和 15 Gy照射后 2 4h的表达 ,以及接受 15 Gy照射后 2 d和 7d的表达 ;以 12 Gy照射小鼠右肾和左肾的下半部 ,2 4h后进行左肾单侧切除 ,采用尿酶 -靛酚方法测定照射后…     

小剂量γ射线全身照射对小鼠脑组织氨基酸类神经递质含量的影响

目的 探讨小剂量γ射线全身照射后对小鼠中枢神经系统的影响.方法 将50只C57小鼠按随机表法分为健康对照组及0.5和1 Gy照射组.用60Co γ射线全身照射,吸收剂量为0.5和1 Gy,吸收剂量率为19.2 cGy/min,连续照射3d,1次/d,照射后24和48 h取小鼠脑组织并匀浆,用高效液相色谱荧光法(HPLC)测定匀浆上清中氨基酸类神经递质(谷氨酸、天冬氨酸、γ-氨基丁酸及甘氨酸)含量.结果 与健康对照组相比,0.5Gyγ射线照射后48 h以及1 Gy照射后24和48 h,小鼠脑内谷氨酸和天冬氨酸含量表达增高(t=- 4.080、- 3.935、-4.416、- 3.630、-4.831和- 4.656,P＜0.05);而0.5Gyγ射线照射后24 h小鼠脑内谷氨酸、天冬氨酸含量无明显改变.各照射组小鼠脑内γ-氨基丁酸和甘氨酸含量与健康对照组比较,差异无统计学意义.结论 小剂量γ射线短期全身照射对小鼠中枢神经系统具有轻度的兴奋作用.     

60Coγ射线诱导人外周血淋巴细胞核质桥的剂量-效应关系研究

目的 确定核质桥判定标准,建立 ⁶⁰Co γ 射线诱导正常人外周血淋巴细胞中核质桥（NPB）的剂量-效应曲线。 方法 用 ⁶⁰Co γ 射线照射3名健康男性离体外周血,照射剂量分别为0、1、2、3、4、5和6 Gy,剂量率为1 Gy/min,采用胞质分裂阻滞微核（CBMN）法进行细胞培养、收获、制片、染色。在光学显微镜下分析双核细胞中NPB及微核（MN）。 结果 在0~6 Gy ⁶⁰Co γ 射线照射后,人外周血双核淋巴细胞中的NPB符合泊松分布,且NPB频率随吸收剂量增加而增加（H=19.51,P<0.01）,拟合回归方程为线性平方模式y=（1.39×10^-3）x² + （4.94×10^-3）x （R²=0.981,P < 0.01）。 结论 成功建立 0~6 Gy ⁶⁰Co γ 射线诱导正常人外周血淋巴细胞中NPB的剂量-效应曲线。     

电离辐射剂量率、照后培养时间和生长因子对人外周血淋巴细胞克隆存活和以凋亡为特征的分裂间期细胞死亡的影响

目的 :通过人外周血淋巴细胞凋亡和增殖细胞死亡观察剂量率、照后恒温培养时间和生长因子对电离辐射诱导的分裂间期细胞死亡的作用。方法 :将健康成人静脉血淋巴细胞在 RPMI 16 40培养基中培养 16 d。开始培养 6 d换半液 ,随后每隔 2 d换液 1次 ,经流式细胞仪检测 96 %～ 97%的细胞处于 G0 期。用高剂量率 (HDR,45 Gy/ h)或低剂量率 (L DR,0 .0 2 4Gy/ h)的 1～3Gy1 3 7　 Csγ射线照射 G0 期淋巴细胞。在 3Gy L DR组开始照射的第 1天 ,同时进行 HDR组照射 (HDR1) ,并在 3Gy L DR组结束照射的第 6天进行另一组 HDR照射 (HDR6…     

胞质分裂阻滞微核法对辐射事故受照射人员的生物剂量估算

目的 探讨胞质分裂阻滞微核(CBMN)分析在估算辐射事故受照射者的生物剂量中的应用价值.方法 2008年山西太原辐射事故发生后16 h收集5名受照射者(1、2、3、4和5号)的外周血及I号的骨髓,进行CBMN分析,以微核(MN)频率估算生物剂量.对较严重的受照射者(1号)结合体外"co 1射线大剂量照射实验获得的核分裂...     

Biological dose estimation for two severely exposed patients in a radiation accident in Shandong Jining,China, in 2004

Purpose:?To estimate the biological doses for two severely exposed subjects (A and B) in a radiation accident in Shandong Jining, China in 2004.

Materials and methods:?Conventional chromosome aberration analysis and cytokinesis-block micronuclei (CBMN) assay were performed in peripheral blood and bone marrow samples on two subjects after the accident. A new dose-effect curve and the nuclear division index (NDI) obtained from in vitro irradiation experiments using high dose of ⁶⁰Co γ-rays were used to estimate the exposed doses.

Results:?No metaphases or binucleated cells were observed in the peripheral blood cultures from either of the subjects. However, metaphases and binucleated cells were obtained from both subjects after bone marrow cultures. Both dicentric/ring and micronuclei yields were very high. The dose estimated for A and B were 20.0 Gy and 8.8 Gy, respectively, by dicentric/ring scoring, similar to the data by combination of the CBMN and NDI (CBMN?+?NDI) assay. The estimated doses by the two methods were in accordance with the clinical symptoms.

Conclusion:?The new curve, together with the CBMN?+?NDI assay, are reliable for estimating higher doses of irradiation. In future radiation accidents, the accuracy and significance of these methods can be further tested.     

Differential radiation effects in smokers--culture time dependence of the yield of gamma ray-induced chromosome damage in first division metaphases

PURPOSE: Telomeric associations (TA) and unstable chromosomal aberration (CA) transmission through M1-M4 metphases (first to fourth division) in gamma-ray irradiated G0 lymphocytes in 2 smokers were examined, since TA in conventionally stained chromosomes were reported earlier as a sensitive cytogenetic marker in mutagen-exposed populations. The purpose of the present study is an extension of our earlier studies on unstable CA transmission through successive mitotic divisions. MATERIALS AND METHODS: The bromodeoxyuridine (BrdU) incorporation and fluorescence plus giemsa (FPG) method for M1-M5 metaphase analysis was carried out at 50, 72, 96 h to analyse TA and CA in conventionally and FPG stained chromosomes after irradiation of human blood samples with 3 Gy of gamma-rays. In situ hybridization (ISH) with enzymatic/fluorescence detection was used to analyse radiation-induced aneuploidy and TA. Analysis was carried out on sister chromatid exchanges (SCE) in M2 cells at 72 h and micronuclei (MN) at 24, 50, 72, 96 h. RESULTS: TA, corroborated by the absence of acentric fragments, were not detected in conventional/FPG stained/ISH chromosomes. Chromosome 21 aneuploidy was observed. Significant differences in mean frequencies of dicentrics/micronuclei (MN)/SCE with high frequency cells (HFC) were found in smokers after irradiation compared to non-smokers. Higher radiation induced CA in M1 cells were found with extended culture time. Induction of giant cells with mirror dicentrics, tricentrics and rings were found. CONCLUSION: TA in conventional or FPG stained metaphase chromosomes is not a sensitive cytogenetic marker for mutagen exposed population screening. Higher radiation induced CA frequencies in M1 cells with extended culture time were indicative of a delay in cell cycle progression of aberrant cells or different lymphocyte subset populations. Bridge-breakage-fusion (BBF) events due to dicentrics may be instrumental in the perpetuation of chromosomal instability. Differential effects were noted in radiation-induced dicentric, SCE and MN frequencies in smokers compared to non-smokers. Heavy smoking could be a confounding variable in chromosome-based biodosimetry and biomonitoring studies. Giant cells may denote a switch to amitotic modes of cell survival, providing additional mechanisms of genotoxic resistance.     

离体人血受照后染色体培养中若干问题的探讨

探讨了人离体血经X线照射后，染色体培养过程中松胞素B（Cyt-B)及培养时间对分裂指数（MI）的影响。结果表明：（1）0-15Gy照射，培养体系中加入Gyt-B，MI均有所增加；10Gy以上影响较在，25Gy不再有影响。（2）6-25Gy，延长培养时间MI明显增加，在6-10G 间仅靠延长时间即可获足够分析用的分裂像。（3）加Cyt-B的同时延长培养时间，其MI高于单纯加Gyt-B或延长时间者，特别是15Gy，昭后只有加Gyt-B同时延长培养时间才能使MI增多至可供分析，以上结果提示：（1）辐射事故中当以染色体畸变估算大于6Gy受昭者的剂量时，必须延长培养时间，有条件者加Cyt-B方可增加培养成功的机会，提供足够的分裂像。（2）改变培养条件，加Cyt-B并延长培养时间有可能制定出剂量上限高于5Gy的剂量效应刻度曲线，从而进一步提高其作为生物剂量计的临床应用价值。     

参芪扶正液对肺腺癌细胞GLC-82X射线照射后TGF-β1表达调控的研究

目的 阐明参芪扶正液对肺腺癌细胞株GLC-82 X射线照射后TGF-β₁表达的调控,探讨中药防治放射性肺损伤的分子机制。方法选取人低分化肺腺癌细胞株GLC-82,加入终浓度为1.563 mg/ml参芪扶正液后随机分组:1照射剂量关系组:分别予0、20、30和40 Gy剂量的X射线照射,于照射后6 h提取细胞总RNA。 2时间关系组:接受20 Gy的X射线照射,分别在照射后12、24和48 h提取细胞总RNA。2组均采用RT-PCR法检测TGF-β₁mRNA表达。结果剂量关系组:GLC-82细胞接受X射线照射后TGF-β₁mRNA表达量明显增高,在20 Gy照射后达到高峰,为基础未照射水平细胞的5倍。参芪扶正干预组与单纯照射组相比,其TGF-β ₁mRNA相对表达量均有不同程度下调,在吸收剂量达20 Gy时差异有统计学意义(P＜0.05)。时间关系组:GLC-82细胞经20 Gy X射线照射后在不同时间点参芪扶正干预组与单纯照射组相比,其TGF-β₁ mRNA相对表达量均有不同程度下调,受照后48 h 差异有统计学意义(P＜0.01)。结论TGF-β₁在放射性肺损伤发生、发展过程中起着重要作用,而参芪扶正液能明显降低该过程中TGF-β₁表达水平,提示参芪扶正液可能通过调控该细胞因子来起到辐射防护之作用。     

The use of cryopreserved lymphocytes in assessing inter-individual radiosensitivity with the micronucleus assay

PURPOSE: The feasibility of using cryopreserved lymphocytes to detect inter-individual differences in chromosomal radiosensitivity was investigated. Typically, such studies are conducted with fresh blood samples but, in a clinical setting, when availability of samples is unpredictable, this is not always convenient. The sensitivity of 23 normal healthy donors, 11 breast cancer patients who had shown severe acute skin reactions to radiotherapy and seven ataxia telangiectasia (A-T) heterozygotes was determined. MATERIALS AND METHODS: Thawed lymphocytes were exposed to high (HDR) or low dose rate (LDR) gamma irradiation (3.5 Gy) in Go, stimulated with PHA, treated with cytochalasin-B 24 h later and then harvested at 90 h for the determination of micronucleus (MN) yields in binucleate cells. RESULTS: Each normal donor was tested one to three times. Mean MN yields were 76.1 +/- 9.3/100 cells at HDR and 44.5 +/- 5.3 at LDR, giving an LDR sparing effect of 39.6 +/- 9.3%. A relatively high proportion of tests failed to yield sufficient binucleate cells for analysis. Inter-experimental variability was also high and it was not possible to demonstrate inter-individual differences in sensitivity in spite of the use of an internal control sample from a single normal donor in each experiment. There was a small but significant increase in radiation-induced MN in the breast cancer patients compared with the normals at LDR (but not at HDR), but a complete overlap with the normal range. There was no increase in sensitivity in the A-T heterozygotes at HDR. The LDR samples failed because the LDR protocol reduced proliferation rates, and radiation-induced mitotic inhibition in this group was higher than in normals. CONCLUSIONS: In comparison with previous experience with fresh blood samples, the use of frozen lymphocytes is not as satisfactory because: (1) experimental failures are higher; (2) inter-experiment variability is higher: (3) dose-rate sparing is lower, suggesting poorer repair; and (4) the ability to discriminate between breast cancer cases and normals is probably lower.     

Intra- and inter-individual variation of background and radiation-induced micronucleus frequencies in human lymphocytes.

Serial blood samples were taken from four healthy individuals (three males, one female, aged between 26 and 51 years) in 3-monthly intervals during 1 year. Leucocyte suspensions were prepared and exposed to 3 Gy of 137Cs gamma-rays or left unirradiated as controls. In a cytokinesis-blocked (CB) micronucleus (MN) assay significant inter- and intra-donor variations of background and radiation-induced MN incidences became apparent. The two sources of variation lead to an extra variance sigma I2, in addition to the sample variance sigma e2 of MN incidences. The contributions of the different components to the total variance were estimated by means of a variance component model. The deviation sigma I for the mean background MN level of 1.53 x 10(-2) MN/CB cell was +/- 0.67 x 10(-2) and for the mean radiation-induced MN level of 0.53 MN/CB cell it was +/- 0.10. The contribution of the intra-individual variance to sigma I2 was about 50% for background MN levels and 75% for radiation-induced MN frequencies. With respect to the application of the CB-MN assay as a biological dosimetry system, the consequences of the present findings for calibration purposes and low-dose estimation are discussed. The calculation of the variance components is explained in an appendix, which serves also as an example for the adaptation of analysis of variance techniques to the evaluation of data derived from scoring of MN, as well as from scoring of metaphase chromosomal aberrations.     

Chromosomal radiosensitivity in secondary-progressive multiple sclerosis patients

PURPOSE: To investigate chromosomal radiosensitivity of secondary progressive (SP) multiple sclerosis (MS) patients in comparison to a group of healthy individuals. MATERIAL AND METHODS: Chromosomal radiosensitivity was assessed in vitro with the G2 assay and the G0-micronucleus (MN) assay. For the G2 assay phytohaemagglutinin (PHA) stimulated blood cultures were irradiated with a dose of 0.4 Gy 60Co gamma rays in the G2 phase of the cell cycle. For the MN assay unstimulated diluted blood samples were exposed to 3.5 Gy 60Co gamma rays delivered at a high dose-rate (HDR = 1 Gy/min) or low dose-rate (LDR = 4 mGy/min). RESULTS: No significant differences in the number of chromatid breaks were observed between MS patients and healthy individuals. With the G0-MN assay a higher spontaneous MN yield was found in MS patients. At HDR irradiation no significant differences were shown, while at LDR irradiation, MS patients were found less sensitive than healthy controls. The dose-rate sparing index was higher for MS patients, pointing to a better repair capacity. CONCLUSIONS: MS patients are not characterised by an enhanced in vitro chromosomal radiosensitivity. The radioresistant response, which was only observed with the MN assay after LDR irradiation, may point to an adaptive response induced by in vivo oxidative stress in SPMS patients.     

褐藻糖胶对60Co-γ射线照射引起的人外周血淋巴细胞微核率的影响

目的 本研究通过60Co-γ射线体外照射人外周血淋巴细胞观察微核率,研究褐藻糖胶的辐射防护作用.方法 抽取4名从未接触过有害化学物质的健康男性志愿者的静脉血,按常规微量血培养方法体外培养外周血淋巴细胞以及微核细胞.实验分为对照组、照射组及加药组(照射前lh、照射后1h加入20、40、80、160、320μg/ml褐藻糖...     

盐酸川芎嗪透皮贴剂药代动力学研究

目的建立SD大鼠盐酸川芎嗪（TMPH）贴剂和灌胃给药后体内血药浓度的测定方法,计算贴剂组和灌胃组的药代动力学参数并进行比较。方法贴剂组：自制贴剂,SD大鼠背部给药（1.04 g/kg）,分别于给药后0.5、1.5、3、4、5、6、7、8、9、12、17、24 h从尾部静脉取血;灌胃组：SD大鼠灌胃给药（1.6 mg/kg）,分别在给药后5、8、10、13、15、21、34、45、61 min从尾部静脉取血;用HPLC测定两组大鼠体内TMPH的血药浓度,用DAS软件计算。结果灌胃组药代动力学参数：Tmax=0.167 h,Cmax=1.97 mg/L,T1/2=0.076 h,Auc0-t=0.597[（μg·h）/ml],Auc0-∞=0.650[（μg·h）/ml];贴剂组药代动力学参数：Tmax=5.917 h,Cmax=253 mg/L,T1/2=5.95 h,Auc0-t=3547.56[（μg.h）/ml],Auc0-∞=4132.47[（μg.h）/ml]。结论 TMPH灌胃给药时,达峰时间和半衰期都很短,说明TMPH普通制剂在体内吸收代谢非常迅速,难以长时间维持有效血药浓度;TMPH贴剂给药的达峰时间和半衰期大大延长,能够在较长时间内维持较高的血药浓度。