四君子汤 对电离辐射损伤 的防治作用

目的：观察四君子汤对电离辐射所致免疫和造血系统损伤的防治作用,探讨其可能的机制。方法：采用CCK-8试剂盒检测不同终浓度（0、60、120、600、1 200 μg/mL）的四君子汤作用48 h对淋巴母细胞AHH-1的毒性作用,以及不同终浓度（0、0.04、0.2、1、25、120 μg/mL）的四君子汤预处理2 h对4 Gy ⁶⁰Co γ射线照射后AHH-1细胞存活率的影响。选择6~8周的BALB/c小鼠160只,用随机数字表法分为阴性对照组、照射对照组以及四君子汤0.75、2.25、6.75 g/kg剂量组。3.5 Gy γ射线全身单次照射造成小鼠电离辐射损伤模型。照射后第3和第7天,观察四君子汤对小鼠外周血淋巴细胞数及百分率、胸腺系数的影响。各组小鼠经5.5 Gy γ射线全身单次照射后,观察30 d并记录死亡小鼠数和死亡时间。结果：与照射对照组细胞（0 μg/mL）相比,四君子汤在0~120 μg/mL范围内对AHH-1细胞未见明显毒性（P > 0.05）,经120 μg/mL四君子汤处理的AHH-1细胞存活率显著提高（P < 0.05）。与照射对照组比较,照后3 d,四君子汤2.25 g/kg剂量组小鼠的外周血淋巴细胞数及百分率均显著升高（P均 < 0.05）;四君子汤6.75 g/kg剂量组的淋巴细胞百分率显著升高（P < 0.05）;照后第3和第7天,四君子汤6.75 g/kg剂量组的胸腺系数均显著升高（P均 < 0.05）。与照射对照组比较,四君子汤6.75 g/kg剂量组小鼠的30 d内平均存活时间及30 d生存率均显著增加（P均 < 0.05）。结论：四君子汤通过提高外周血淋巴细胞数、减少胸腺损伤,可缓解电离辐射所致损伤,促进小鼠免疫和造血功能恢复,并可促进γ射线照射小鼠的存活,可考虑作为电离辐射损伤防治药物作进一步的研究。     

辐射损伤骨髓造血细胞的DNA含量及AgNOR定量分析

LACA小鼠经60 Coγ射线全身一次照射 ,对骨髓组织切片和骨髓细胞悬液涂片 ,分别采用改良的Feulgen染色及核仁组成区相关蛋白 (AgNOR)染色 ,利用图像分析技术对在不同照射剂量、不同时间点的造血细胞核内DNA及AgNOR含量进行了定量分析。结果显示各照射组在2 5 - 7 0Gy剂量、 6h～ 4周范围内 ,骨髓均出现明显损伤及损伤后重建现象 ,且剂量越大 ,损伤越重 ,恢复亦较慢 ;其中以 5 5Gy组骨髓细胞核内DNA和AgNOR含量变化最为典型 ,在损伤早期进行性减少 ,而在恢复期出现持续性增加 ,直至恢复正常。结果提示 :骨髓造血细胞核内AgNOR及DNA含量的变化可以作为反映骨髓辐射损伤与修复程度的定量指标。     

不同剂量X射线照射对小鼠免疫系统的影响

目的：观察不同剂量X射线照射对小鼠免疫系统的影响。方法将24只小鼠随机分为对照组（无照射）,低剂量照射组（每次2 Gy）和高剂量照射组（每次5 Gy）,每组各8只。比较不同剂量X射线对小鼠进行单次全身照射后,胸腺、脾脏变化及对外周血白细胞,淋巴细胞及血小板数量的影响。结果高剂量照射组胸腺指数较对照组降低,差异具有统计学意义（P﹤0.05）,而低剂量照射组胸腺指数与对照组比较,差异无统计学意义（P﹥0.05）;随着X线照射剂量的增大,脾脏指数降低,差异均有统计学意义（P﹤0.05）;与对照组比较,低、高剂量照射组小鼠外周血白细胞,淋巴细胞及血小板数量均减少,差异均有统计学意义（P﹤0.05）。结论不同免疫器官对X射线敏感度不同,X射线对免疫功能的损伤随着射线剂量的增加而增大,因此应该加强对放射治疗患者免疫器官的保护。     

大蒜提取物S-烯丙基-L-半胱氨酸对小鼠放疗损伤保护作用的研究

目的:研究大蒜提取物S-烯丙基-L-半胱氨酸（S-allyl-L-cysteine,SAC）对小鼠放射治疗所致损伤的保护作用。方法:采用小鼠30 d存活率实验及对造血系统、免疫系统影响实验,即小鼠受7.5 Gy剂量全身照射后,观察其存活情况及测外周血白细胞数、股骨有核细胞数、骨髓DNA含量、胸腺和脾指数的变化情况。结果:不同剂量的给药照射组小鼠30 d存活率比单纯照射组均有提高,对造血系统、免疫系统均有保护作用,特别是中剂量组存活率提高了50%,白细胞数、股骨有核细胞数、骨髓DNA含量和脾结节数分别从0.17±0.05、0.56±0.07、0.21±0.14和1.02±0.30提高到0.67±0.08、4.45±1.76、0.58±0.12和2.71±0.21,数据均有统计学意义(P＜0.05),并且SAC对胸腺、脾也有一定的保护作用。结论:大蒜提取物S-烯丙基-L-半胱氨酸对放射治疗所致损伤具有一定的保护作用。     

E838对CTX化疗损伤小鼠的保护作用

 目的 观察E838对环磷酰胺所致小鼠损伤的保护作用。 方法 对小鼠采用连续5天腹腔注射E838药物,从第3天同时给予环磷酰胺（CTX）,阳性对照为茜草双酯,观察外周血白细胞、骨髓有核细胞数、内源性脾结节形成（CFU-S）、脾脏指数及胸腺指数的变化。 结果 E838可明显减轻化疗所致小鼠免疫功能的抑制,并可使化疗后小鼠的体重恢复增长, 白细胞、骨髓有核细胞和CFU-S明显高于对照组,差异有统计学意义（P<0.05）。中、高剂量组白细胞与茜草双酯组比较,差异有统计学意义（P<0.05）。 结论 提示E838具有对抗化疗损伤的作用。     

茜草醇提物对辐射损伤小鼠造血系统保护作用的研究

目的 研究茜草醇提物对辐射损伤小鼠造血系统的保护作用。方法 建立辐射损伤小鼠模型,将50只小鼠随机分为5组,每组10只。给药组小鼠予以不同剂量［低剂量0.25 g/（kg·d）、中剂量0.49 g/（kg·d）、高剂量0.99 g/（kg·d）］茜草醇提物灌胃,空白对照组和单纯照射组分别予1 mL生理盐水灌胃,各组持续灌胃13 d,正常饲养7 d后处死小鼠。于灌胃第7天,不同剂量给药组和单纯照射组小鼠均接受60Co γ射线6 Gy一次性全身照射。观察茜草醇提物对辐射损伤小鼠体重以及外周血白细胞、红细胞、血小板和骨髓有核细胞含量的影响。结果 与单纯照射组比较,高剂量茜草醇提物给药+照射组小鼠外周血白细胞、红细胞、骨髓有核细胞计数均升高,差异均有统计学意义（P＜0.05）,但体重和血小板计数差异无统计学意义（P＞0.05）;而低剂量、中剂量茜草醇提物给药+照射组与单纯照射组比较差异均无统计学意义（P＞0.05）。结论 高剂量茜草提取物对辐射损伤小鼠造血系统有促进恢复作用。     

不同照射剂量对BRCA基因突变及其非突变的乳腺癌细胞DNA损伤和凋亡的影响

目的 探讨不同照射剂量对BRCA基因突变及BRCA基因非突变的乳腺癌细胞DNA损伤和凋亡的影响。方法 BRCA基因突变的乳腺癌细胞株MDA-MB-436及BRCA基因非突变的乳腺癌细胞株MDA-MB-231分别按0 Gy、2 Gy、4 Gy、6 Gy、8 Gy、10 Gy照射剂量进行X射线照射。以流式细胞术仪检测细胞凋亡率,在各照射剂量中,取γH2AX免疫荧光焦点最明显的时间点（30 min）检测细胞DNA双链损伤情况。结果 乳腺癌细胞DNA损伤及细胞凋亡率均随照射剂量增加而加重,8 Gy照射剂量时两个细胞株的DNA损伤及细胞凋亡率均达到最高,MDA-MB-436细胞分别为（47±1.802）个和（21.245±1.325）%,MDA-MB-231细胞分别为（45±1.779）个和（19.220±1.220）%。不同照射剂量下,MDA-MB-436细胞较MDA-MB-231细胞的DNA损伤严重及细胞凋亡率增加,两者比较差异均有统计学意义（P均＜0.05）。两个细胞株8 Gy照射剂量的细胞焦点增长数及细胞凋亡增长率与10 Gy照射剂量比较,差异均无统计学意义（P均＞0.05）。 结论 随着X射线照射剂量由0 Gy至10 Gy不断增强,乳腺癌细胞损伤不断增加,BRCA基因突变的乳腺癌细胞DNA损伤及细胞凋亡率均较BRCA基因非突变的乳腺癌细胞明显增加,具有更高的放射敏感性。     

BALB／c小鼠重度骨髓型急性放射病模型的建立

 目的　建立BALB／c小鼠重度骨髓型急性放射病模型,为重度骨髓型急性放射病的实验研究提供依据。方法　BALB／c小鼠给予60Coγ射线6.0 Gy一次全身照射。观察小鼠照射后一般临床表现、外周血细胞计数、股骨病理组织学及骨髓细胞集落生成情况。结果　小鼠照射后第3天活动量即有不同程度的减少,但均无呕吐、稀便,照射后第11天白细胞降至最低值（基础水平的3.0 %）,照射后第28天恢复至基础水平的53.7 %;照射后第14天血小板降至最低值（基础水平的8.1 %）,照射后第28天恢复至基础水平的60.4 %;照射后第14天骨髓病理示骨髓腔呈空虚状态,骨髓有核细胞集落培养亦提示粒细胞-巨噬细胞集落形成单位（CFU-GM）、混合集落形成单位（CFU-Mix）明显下降;照射后第28天骨髓病理、CFU-GM及CFU-Mix未完全恢复;照射后2个月全部小鼠存活。结论　6.0 Gy 60Coγ射线一次全身照射BALB／c小鼠可成功构建重度骨髓型急性放射病模型,可用于骨髓型急性放射病的实验研究。     

转染人突变dhfr基因的小鼠骨髓细胞对小鼠造血功能的长期保护作用

目的:将转染了人突变dhfr基因的第二代小鼠骨髓,移植给经致死剂量照射的第三代小鼠,观察该基因对小鼠造血功能的长期保护作用.方法:分离存活的第二代小鼠骨髓有核细胞,直接移植给经致死剂量照射的同系小鼠,以MTX筛选,观察小鼠血象和生存率变化,PCR和Southem印迹杂交分析目的基因在小鼠染色体DNA中的整合与表达情况.结果:在大剂量MTX筛选下,实验组小鼠造血功能逐渐恢复,对照组3周内全部死亡.实验组生存率和生存期明显高于对照组,但较前两代生存率低.PCR和Southern印迹分析结果提示,实验组脾脏和肝脏组织中均检测到前病毒标志基因neo~R和dhfr基因的特异务带.结论:转染了人突变dhfr基因的第二代小鼠骨髓,能有效地重建经致死剂量照射的第三代小鼠造血功能.保护骨髓免遭大剂量MTX所致的严重骨髓抑制,dhfr基因在小鼠基因组DNA中的稳定整合是这种长期保护作用的物质基础.     

苞叶雪莲水提物对辐射损伤小鼠防护作用的研究

目的：初步探讨苞叶雪莲水提物对辐射损伤小鼠的防护作用。方法：观察苞叶雪莲水提物对小鼠体重、外周血白细胞、红细胞、血小板计数、骨髓有核细胞数、内源性脾结节数、骨髓DNA含量的影响。结果：照射后给予一定浓度的苞叶雪莲水提物,可使小鼠外周血白细胞、红细胞、血小板计数及骨髓有核细胞计数高于照射对照组（P〈0．05）,三个照射给药组骨髓DNA含量高于照射对照组（P〈0．05）,高剂量照射给药组内源性脾结节数高于照射对照组（P〈0．01）。结论：苞叶雪莲水提物对辐射损伤小鼠造血系统有促损伤恢复作用,可能与促进骨髓细胞DNA损伤修复有关。     

放射导致小鼠味觉细胞损伤与放射模式关系研究

目的 探讨放射导致小鼠舌组织中增殖细胞和味觉细胞损伤与放射模式的关系。方法 对成年C57/bl小鼠头颈部分别进行1、2、3次8Gy放射,分别在照后第2、4、7、14天处死;取舌轮廓乳头组织,4μm厚度冷冻切片,使用不同特异标记分子行免疫组化法标记舌上皮细胞。观察小鼠受照后的增殖细胞、Ⅱ型味觉细胞数量在不同放射模式下不同时间的变化情况,探讨不同放射剂量模式对味觉细胞的影响。结果 增殖细胞数量在照射后第2天明显下降,第4天迅速回升至正常水平,与剂量模式无关。Ⅱ型味觉细胞数量在8Gy 1次照射后第2天出现低谷,8Gy 2次、8Gy 3次照射后第4天降至最低,第7天缓慢回升。结论 放射可使增殖细胞先降后快速恢复正常,可使Ⅱ型味觉细胞损伤后缓慢修复。高剂量放射后由于具有增殖功能的前体细胞的减少,导致味觉功能细胞数量的同步下降,可能是放疗后味觉功能障碍长久不能恢复的原因。     

Study of the relationship between radiation-induced taste cell injury and radiation mode in mice

Objective To investigate the relationship between radiation mode and radiation-induced taste cell injury in mouse models. Methods The head and neck of adult C57/bl mice were exposed to 8 Gy irradiation for 1, 2 and 3 times and sacrificed at 2, 4, 7 and 14 d after irradiation. Frozen sections of taste papilla tissues were treated with specific markers by immunohistochemical staining. The changes of proliferative cells and the number of type Ⅱtaste cells at different time points under different radiation modes were observed. The effect of different radiation dose patterns upon the taste cells was assessed. Results The count of proliferative cells was decreased significantly on the 2^nd day after radiation, and rapidly recovered to normal level on the 4^th day after radiation, which was not correlated with the dose mode. The number of type Ⅱ taste cells was decreased on the 2^nd day of the first 8 Gy radiation, decreased to the lowest on the 4^th day of the second and third 8 Gy radiation, and rose slowly on the 7^th day. Conclusions Radiation can initially decrease and subsequently increase the number of proliferative cells to normal range. Besides, it can gradually repair the type Ⅱ taste cell injury. After high-dose irradiation, the decrease of progenitor cells with proliferative function leads to a synchronous decrease in the number of taste function cells, which may be the reason why taste dysfunction cannot be recovered for a long time after radiotherapy.     

Effect of irradiation on thermal sensitivity of bone marrow progenitors

The heat sensitivity of murine CFU-GM and CFU-E following 2.5 Gy of total body irradiation (TBI) was studied. C3H f/Sed female mice were treated with 2.5 Gy TBI and femoral bone marrow was heated in vitro at 43 degrees C. CFU-GM show heat radiosensitization when bone marrow was heated immediately following irradiation. There was a brief decline in heat and radiation interaction when cells were heated 3 hours following 2.5 Gy of TBI, but heat radiosensitization returned to its maximum from 1 to 2 days following irradiation and remained significantly different from the control on days 5 and 7 following irradiation. The heat and radiation interaction disappeared by 30 days. CFU-E shows significant heat radiosensitization only on day 2 following 2.5 Gy of TBI. Total nucleated cells per femur showed a decrease by 70 per cent in days 1 to 2 following TBI, recovered to control values by day 5, and did not correlate with the changes in heat radiosensitization. Cell cycle analysis of CFU-GM using hydroxyurea showed no significant changes in cell cycle parameters on days 1 and 2 following 2.5 Gy, when maximum heat sensitization was observed. It is concluded that bone marrow progenitors may respond in a different way from other normal tissues to heat and irradiation sequencing, and that these differences must be considered when designing clinical trials.     

Cyclophosphamide 24 hours before or after total body irradiation: effects on lung and bone marrow

Preparative regimens for bone marrow transplantation (BMT) use a sequence of drugs, such as cyclophosphamide, in combination with radiation. However, the optimum sequencing of the two agents that will maximize tumor cell kill and minimize normal tissue damage is unknown and controversial. The studies presented here were done in order to determine the effect of cyclophosphamide on bone marrow and lung damage in mice when given 24 h before or after total body irradiation (TBI). A range of single doses of TBI was given before or after a single sublethal dose of 180 mg/kg of cyclophosphamide. The bone marrow of all mice intended for lung damage assessment was reconstituted with 5 x 10(6) syngeneic bone marrow cells. Lung damage was assessed by breathing rate and lethality; bone marrow damage by lethality at 30 days. LD50 values for pneumonitis were obtained between 30 and 84 days after cyclophosphamide and radiation and between 80 and 180 days after radiation alone. Dose modifying factors were obtained as the ratio of LD50s for mice given only TBI compared to those for mice given cyclophosphamide and TBI. Cyclophosphamide enhanced radiation pneumonitis when given before or after TBI, giving DMFs of 1.4 and 1.2 (1.1-1.4, 95% c.l.) respectively. The effect of cyclophosphamide on radiation pneumonitis was drug dose-dependent. The LD50 for death from bone marrow damage was reduced when cyclophosphamide was given either before or after TBI but the effect was greater, i.e. the LD50 was lower when cyclophosphamide was given after TBI. These data show that cyclophosphamide given 24 h after TBI causes less lung damage but more bone marrow damage in this mouse model.     

Cell survival kinetics in peripheral blood and bone marrow during total body irradiation for marrow transplantation

Cell survival kinetics in both peripheral blood and in bone marrow have been studied over the time course of hyperfractionated total body irradiation (TBI) for bone marrow transplantation. Our unique TBI regimen allows the study of the in vivo radiation effect uncomplicated by prior cyclophosphamide, since this agent is given after TBI in our cytoreduction scheme. Peripheral blood cell concentrations were monitored with conventional laboratory cell counts and differentials. Absolute bone marrow cell concentrations were monitored by measuring cell concentrations in an aspirate sample and correcting for dilution with blood by a cell cycle kinetic method using cytofluorometry. In the entire group of patients, time to engraftment with donor marrow was found to be 16.6 × 4.4 days and more rapid when a nucleated donor cell dose of ≥4.0 × 10⁸ cells/kg was given. For lymphocytes in peripheral blood in patients in remission, the effective D₀ ranged from 373 rad in 10 children ≤10 y old, to 536 rad in the four patients between 11–17 y old, while n = 1.0 in all groups. There was no trend observed according to age. Granulocytes had a much higher effective D₀, approximately 1000 rad in vivo. Absolute nucleated cell concentration in marrow dropped slowly initially, due to an increased lymphocyte concentration in marrow during a concurrent drop in lymphocyte concentration in peripheral blood, but eventually fell on the last day of TBI ranging from 7–44 % of the initial marrow nucleated cell concentration. Marrow myeloid elements, however, dropped continuously throughout the course of TBI.     

Effect of irradiation on thermal sensitivity of bone marrow progenitors

The heat sensitivity of murine CFU-GM and CFU-E following 2.5 Gy of total body irradiation (TBI) was studied. C3H f/Sed female mice were treated with 2.5 Gy TBI and femoral bone marrow was heated in vitro at 43°C. CFU-GM show heat radio-sensitization when bone marrow was heated immediately following irradiation. There was a brief decline in heat and radiation interaction when cells were heated 3 hours following 2.5 Gy of TBI, but heat radiosensitization returned to its maximum from 1 to 2 days following irradiation and remained significantly different from the control on days 5 and 7 following irradiation. The heat and radiation interaction disappeared by 30 days. CFU-E shows significant heat radiosensitization only on day 2 following 2.5 Gy of TBI. Total nucleated cells per femur showed a decrease by 70 per cent in days 1 to 2 following TBI, recovered to control values by day 5, and did not correlate with the changes in heat radiosensitization. Cell cycle analysis of CFU-GM using hydroxyurea showed no significant changes in cell cycle parameters on days 1 and 2 following 2.5 Gy, when maximum heat sensitization was observed. It is concluded that bone marrow progenitors may respond in a different way from other normal tissues to heat and irradiation sequencing, and that these differences must be considered when designing clinical trials.     

Similar effects on murine haemopoietic compartment of low dose rate single dose and high dose rate fractionated total body irradiation. Preliminary results after a unique dose of 750 cGy

This study was designed to compare two different modalities of TBI which are currently used in clinical practice. The same dose of 750 cGy was given to CBA mice either in a single dose at a low dose rate (4 cGy min-1) (STBI) or in a fractionated regimen (six fractions of 125 cGy three times a day) at a higher dose rate (25 cGy min-1) (FTBI). After TBI completion we simultaneously studied the in vivo radiation response of bone marrow cells, two murine bone marrow clonogenic cells (CFU-S and GM-CFC) and peripheral blood lymphocytes and granulocytes for a period of 1 month. The percentage of spleen erythrocytic and granulocytic colonies was also determined. No significant differences were observed between the two groups in the first 48 hours after irradiation except in bone marrow cell numbers, probably due to differences in the overall treatment time between the two TBI schedules. After the first 48 hours the repopulation patterns of the different cells were very similar in both groups. These findings suggest that the different dose rates and fractionation used in this study caused similar radiation damage to the murine haemopoietic system. Moreover, no significant repopulation occurred during the longer overall treatment time of the fractionated regimen. These preliminary results must be corroborated with a larger range of doses before any firm conclusion can be drawn.     

Influence of dose on regeneration of murine hematopoietic stem cells after total body irradiation and 5-fluorouracil.

The regeneration of murine bone marrow stem cells after treatment with 5-fluorouracil (5-FU) and total body irradiation (TBI) was investigated by means of the spleen colony assay. 5-FU was given intraperitoneally in doses ranging from 75 to 225 mg/kg body weight. TBI was administrated as a single fraction in the dose range 0.75-3.50 Gy. The frequency of stem cells declined rapidly and reached a minimum (nadir) on day 1-2 after either of the 2 treatments. Their number returned to near-pretreatment values by day 7-10. Increasing the dose of either 5-FU or TBI resulted in a lower nadir in stem cell survival and a higher regeneration rate of the stem cells. The maximal regeneration rate corresponded to a doubling time of 19 h. The time to complete regeneration was constant and neither influenced by the treatment modality nor by the dose. Both this work and previous studies on cyclophosphamide and combined 5-FU and TBI may indicate that the regeneration rate of bone marrow stem cells after a cytotoxic assault is influenced primarily by the stem cell surviving the treatment rather than the treatment modality.     

冬虫夏草对小鼠骨髓和小肠上皮急性损伤防护效应的研究

观察冬虫夏草发酵菌丝（ＤＣＸＣ）对小鼠受Ｘ线照射后骨髓及小肠的防护作用。实验结果显示：（１）照射前服用ＤＣＸＣ的各照射剂量组，小鼠骨髓干细胞的存活数均高于单纯照射组，防护系数为１．６７；（２）昆明小鼠无论照射前给药或照前、照后连续给药，对小肠隐窝干细胞均有一定保护作用。表明，ＤＣＸＣ对辐射诱发的小鼠骨髓及小肠急性损伤具有较好防护效果，是一值得继续开发和探索的辐射损伤防护剂