大豆异黄酮对电离辐射小鼠淋巴细胞的防护作用

目的研究大豆异黄酮(SI)对小鼠淋巴细胞的辐射防护作用。方法24只雄性昆明小鼠,随机分为正常对照组、辐射对照组和辐射补充0.5%SI组,喂养2周后,4.0Gy照射。照射后24 h处死小鼠,取血、胸腺和脾脏分离淋巴细胞,进行血淋巴细胞计数、观察DNA损伤情况;培养胸腺和脾脏淋巴细胞,检测3H-dT掺入量,观察淋巴细胞的增殖能力,计算淋巴细胞的增殖指数。结果辐射使小鼠胸腺和脾脏淋巴细胞数明显减少、胸腺淋巴细胞增殖能力和脾脏淋巴细胞转化指数降低、血淋巴细胞DNA损伤增加,这些变化均具有统计学意义;补充SI可降低胸腺和脾脏淋巴细胞数的减少幅度,降低胸腺淋巴细胞增殖能力和脾脏淋巴细胞转化指数下降幅度,减少辐射对血淋巴细胞DNA损伤程度,其中SI对胸腺淋巴细胞增殖能力和对血淋巴细胞DNA损伤程度的防护作用与辐射对照组相比有统计学意义。结论大豆异黄酮可对小鼠的血、胸腺和脾脏淋巴细胞有一定的辐射防护作用。     

迷迭香酸对辐射损伤小鼠造血功能的保护作用

迷迭香酸是一种天然的水溶性多酚类化合物,最新研究发现具有抗紫外线辐射及细胞保护之效,实验中口服给予辐射损伤的ICR种小鼠不同剂量迷迭香酸(50、100、150、200 mg/kg),通过脏器系数、骨髓DNA含量、股骨有核细胞数(BMNC)、脾结节数(CFU-S)等指标检测其对小鼠造血系统的作用,并以外周血(WBC、HGB、PLT)及体重等指标,从整体水平上衡量迷迭香酸对小鼠造血功能的影响.研究发现不同剂量迷迭香酸对小鼠造血系统均有一定的保护作用,但呈倒“U”形剂量反应曲线,100 mg/kg作用尤为突出,各指标与单纯照射组相比均有显著性差异.     

IRM-2近交系小鼠对电离辐射抗性的研究

目的观察IRM-2小鼠对电离辐射的耐受性.方法分析测定了IRM-2小鼠对137Csγ射线的LD50及经4.0Gy137Csγ射线照射后不同时间外周血白细胞、骨髓有核细胞总数、骨髓细胞DNA含量和脾结节的变化,并与亲代小鼠ICR和615进行了比较.结果用不同剂量的137Csγ射线照射后,IRM-2小鼠对γ射线的LD50比ICR和615小鼠分别高1.73～1.57Gy和1.44Gy;外周血白细胞数和骨髓有核细胞总数、骨髓细胞DNA含量下降的幅度小且恢复得快;CFU-S的增加也较ICR和615小鼠明显.结论IRM-2小鼠比一般的纯系和杂交品系小鼠具有更强的辐射抗性.     

脐带间充质干细胞对辐射损伤小鼠造血功能的影响

目的:探讨脐带间充质干细胞(UCMSC)对辐射损伤模型小鼠造血功能的影响,为相关放射性造血功能损伤疾病的治疗提供理论依据。方法:选取清洁的雄性小鼠80只,随机分为正常组、放射对照组、普通组(注射1次UCMSC)和加强组(注射2次UCMSC),放射组、普通组和加强组的小鼠均经2 Gy射线照射剂量辐射1 h,建立辐射损伤模型。UCMSC悬浮液(每次注射剂量均为230 IU/g)经尾静脉注射方式分别在辐射12和24 h后给予小鼠。照射处理后定期在第1、2、7、14 d对小鼠进行心脏取血并做血象分析,测定血清内超氧化物歧化酶(SOD)活性和丙二醛(MDA)含量。结果:与辐射对照组比较,尾静脉注射UCMSC悬浮液组能改善外周血的数目,血清中SOD活性有所升高,MDA含量显著降低。结论:脐带间充质干细胞对辐射损伤小鼠造血功能具有一定的防护作用。     

湖南黑茶对小鼠辐射损伤的保护作用

研究湖南黑茶对辐射损伤小鼠的保护作用。本研究采用外周血液学指标、免疫系统指标、体内抗氧化指标超氧化物歧化酶(Superoxide dismutase,SOD)及存活时间考察黑茶对辐射损伤小鼠的保护作用。结果显示,小鼠受到137Cs-γ6.5 Gy的照射后,与单纯照射组相比,黑茶能明显提高小鼠的外周血白细胞数(white blood cell,WBC)、股骨有核细胞数(the number of bone narrow nucleated cells,BMNC)、骨髓DNA含量以及脾结节数(colony forming unit-spleen,CFU-S),差异具有显著性(P0.05);小鼠受到137Cs-γ6.0 Gy后,与单纯照射组相比,黑茶可显著提高小鼠肝组织和肺组织中SOD的活力,差异具有显著性(P0.05);黑茶可使受照(8.0Gy)小鼠存活天数由8.6±1.96 d延长到11.13±2.75 d。提示黑茶有较好辐射防护的作用。     

升血汤促进辐射损伤小鼠造血与免疫功能恢复的实验研究

本文研究了中药升血汤对辐射损伤小鼠造血与免疫功能恢复的促进作用,结果证明,升血汤能显著地增加辐射损伤小鼠外周血中各类血细胞数以及骨髓及脾脏中的有核细胞数,明显促进小鼠巨噬细胞,Ｔ及Ｂ淋巴细胞功能的恢复,表明升血汤可以明显促进辐射损伤小鼠造血与免疫功能的恢复。     

黑果枸杞对X射线辐射小鼠的保护作用研究

探讨黑果枸杞水提取液对辐射损伤小鼠的保护作用。除正常对照组外各组小鼠接受5Gy的X射线一次性全身均匀照射。检查各组小鼠外周血中红细胞数(RBC)、白细胞数(WBC)、血小板数(PLT)、血红蛋白数(HGB),计算胸腺和脾脏指数,测定抗氧化酶活力、DNA和Caspase-3、Caspase-6的含量。黑果枸杞水提取液对辐射后小鼠的血象有明显回升,SOD活力明显增强,DNA含量增多,CAT、GSH-PX、T-AOC活力下降,Caspase-3和Caspase-6的含量降低,脏器指数无统计学意义。研究结果显示,黑果枸杞水提取液对X射线所致小鼠辐射损伤有一定保护作用。     

淋巴细胞对造血干细胞增殖的放大作用

小鼠的胸腺细胞对多向性造血干细胞(CFU-S)的增殖有明显的“放大”。作用,而这个“放大”作用的实现,是胸腺细胞与骨髓细胞之比例达到500∶1时为最好。小鼠胸腺细胞对多向性造血干细胞的分化也有明显的影响;本实验表明首先出现红系分化的增强,然后再出现粒系分化的增强。被激活的胸腺及脾脏的淋巴细胞(T细胞)对CFU-S的增殖具有更强的“放大”作用。在去胸腺小鼠的实验中表明ConA对CFU-S没有直接的作用。ConA作用于胸腺等T淋巴细胞,被激活的T细胞再作用于造血干细胞。本实验初步表明,淋巴细胞对造血干细胞的激活作用,必须在完整的活的淋巴细胞与造血干细胞直接接触或接近下才能实现。     

造血干细胞的浓集及其功能研究

应用细胞毒剂和非连续密度梯度离必法可以提高小鼠骨髓CFU-S的相对浓度。实验研究表明，骨髓CFU-S是维持正常机体恒定造血或对照射小鼠促进造血恢复的主要细胞成分。在对750拉德γ线照射小鼠注射相同细胞数量的基础上，治疗效果随CFU-S浓集程度的提高而增强。小鼠注射细胞毒剂有影响骨髓CFU-S生理状态的作用，促进处于砌期的CFU-S转入增殖活动。因此，对照射小鼠移植经细胞毒剂或合并非连纹密度梯度离心后的造血干细胞，它将在CFU-S浓集的基础上迸一步提高其促进照射动物造血功能的恢复。     

17α-乙炔-雌三醇-3-环戊醚对小鼠造血干细胞的影响

小鼠灌胃给予辐射防护有效剂量17α-乙炔-雌三醇-3-环戊醚(CEE_3)后10天内,骨髓与脾脏CFU-S都出现一过性抑制,到15天时恢复正常。CEE_3对CFU-S的抑制程度和辐射防护效价与药量有一定关系。切除脾脏或切除肾上腺可减轻CEE_3对造血干细胞的抑制作用。照前切除脾脏可提高照射小鼠骨髓CFU-S含量,但不能提高CEE_3对造血干细胞的防护效果。切除肾上腺对照射小鼠骨髓CFU-S的含量无明显影响,但可明显减低CEE_3对照射小鼠CFU-S的防护效果。对CEE_3等雌激素的辐射防护作用机理进行简短的讨论。     

The opposing actions in vivo on murine myelopoiesis of purified preparations of lactoferrin and the colony stimulating factors

The actions of purified iron-saturated human lactoferrin (LF), purified preparations of human MiaPaCa colony stimulating factor-1 (CSF-1), and recombinant murine interleukin-3 (IL-3) were evaluated in vivo in mice. Studies in vitro were compared at lowered (5%), as well as at normal incubator (20%), oxygen (O2) tension because of the potentially greater physiologic relevance of in vitro studies performed at lowered O2 tension. The results demonstrate that 1) increased release of granulocyte-macrophage colony stimulating factor (GM-CSF) in vitro from pokeweed mitogen stimulated mouse spleen cells and from human mononuclear blood cells occurred at lowered O2 tension, and that human mononuclear blood leukocytes were more sensitive to the LF-induced suppression of GM-CSF release when cells were cultured at 5%, compared to 20%, O2 tension; 2) LF administered intravenously (IV) to mice pretreated with sublethal intraperitoneal dosages of Cytoxan decreased the cycling status of marrow and spleen granulocyte-macrophage (CFU-GM), erythroid (BFU-E-2 and BFU-E-1) and multipotential (CFU-GEMM) progenitor cells and the absolute numbers of these progenitors; these effects were most noticeable if care was taken to deplete endotoxin from the LF samples prior to testing LF in vivo and if the control medium was endotoxin free; 3) endotoxin-depleted LF decreased the cycling status of marrow and spleen CFU-GM, BFU-E, and CFU-GEMM and the numbers of these progenitors in the marrows of mice previously untreated with Cytoxan; these effects were most apparent when assessment of progenitor cells and their cycling rates were evaluated in vitro at lowered (5%) O2 tension; 4) purified natural human CSF-1 increased the absolute numbers of marrow CFU-GM and the cycling status of marrow CFU-GM and CFU-GEMM in mice pretreated with LF; and 5) purified recombinant murine IL-3 stimulated proliferation of day 8 and day 12 CFU-S (colony forming unit-spleen) in mice not previously treated with Cytoxan. These results substantiate the in vivo myelosuppressive effects of LF on CFU-GM and extend these effects to erythroid and multipotential progenitor cells, provide evidence that human CSF-1 has an in vivo action in mice, and confirm the studies of others showing that IL-3 stimulates the proliferation of CFU-S in vivo.     

人骨髓细胞培养液中干细胞刺激物的分析研究

人骨髓细胞体外培养液中含有高活力的 CSF,在长期培养过程中,CSF 活力的变化,与 CFU-C 数量的变化有大致平行的趋势。这种 CSF 对狗和小鼠也同样有效。人骨體条件液中的 CSF 对培养中的 CFU-S 也有明显的激发作用。这一结论可以从几个方面获得证据:第一,小鼠骨髓细胞与人骨髓条件液保温六小时后,再测定其中 CFU-S 数,结果是增加了。第二,经亚致死剂量照射的小鼠,腹腔注射适量的人骨髓条件液,其内源性脾结节也明显增多。第三,采用阿糖胞苷自杀的方法,测定小鼠骨髓经与人骨髓条件液保温后,其中 CFU-S 的自杀率也有增高的趋势。上述几方面的实验,说明人骨髓长期培养中存在着某种活性物质,调节体外造血。至于这种物质的来源,以及在体外造血中所起的作用,还需要做很多工作,逐步予以澄清。     

白介素3治疗小鼠造血功能低下的疗效及机理初探

通过整体实验观察国产重组白介素３（ＩＬ－３）对射线和环磷酰胺所致小鼠造血功能低下的疗效;以体外实验分析其疗效机理。实验结果表明：（１）ｒｈＩＬ－３腹腔或皮下连续５天注射能全面提高７Ｇｙ照射小鼠９天时股骨骨髓ＣＦＵ－Ｅ、ＢＦＵ－Ｅ、ＣＦＵ－Ｍｉｘ和ＣＦＵ－ＧＭ的产率和数量,其效果强弱与注射途径和用药剂量有关。ｒｈＩＬ－３对小鼠股骨骨髓有核细胞总数和内源性脾结节数的改善影响小。（２）ｒｈＩＬ－３对环磷酰胺所致小鼠造血功能低下亦有改善效果,并与起用时间和剂量有关。（３）ｒｈＩＬ－３对人骨髓细胞和ＣＦＵ－ＧＭ集落形成有明显的增强作用。小鼠骨髓细胞对ｒｈＩＬ－３缺乏反应;对ｒｍＩＬ－３有增殖分化加强的反应。ｒｍＩＬ－３体外共育能提高正常及照射２Ｇｙ小鼠骨髓细胞体外培养后ＣＦＵ－ＧＭ的产率和数量。文中讨论了ＩＬ－３的应用前景及合理方案问题。     

Smad3基因剔除对小鼠造血功能的影响

研究Smad3基因剔除对小鼠造血功能的影响。实验小鼠分为 5组 ,每组有Smad3基因剔除小鼠(Smad3 - - )和其同窝孪生的野生型小鼠 (Smad3 + + )各 1只。小鼠的造血功能用 14天形成的脾结节 (CFU S1 4 )、多系祖细胞 (CFU GEMM)、粒 单系祖细胞 (CFU GM)、红系祖细胞 (BFU E)测定及外周血象、骨髓象等实验血液学指标来确定。每组小鼠取尾血作白细胞、红细胞和血小板计数 ,涂片作白细胞分类计数。将一侧股骨的骨髓冲出 ,制成单细胞悬液 ,计数其中有核细胞数 ,测定CFU GM、BFU E、CFU GEMM值。将每只小鼠的 4× 10 4个骨髓有核细胞 ,经尾静脉注入 3只 8～ 10周经致死量射线照射的同系雌性小鼠体内 ,测定 14天的CFU S。取一部分胸骨、肝脏、脾脏固定做病理切片 ,其余胸骨冲出骨髓 ,涂片作分类计数。结果Smad3 - - 小鼠外周血白细胞和血小板计数明显高于Smad3 + + 小鼠 ,红细胞数无显著差异。外周血白细胞分类结果也表明粒细胞显著增高。骨髓有核细胞数无显著差异 ,CFU GM显著增高 ,BFU E无显著差异 ,CFU GEMM明显减少 ,CFU S显著减少。病理形态学观察发现骨髓增生极度活跃 ,以粒系为主 ,肝脾无显著差别。骨髓涂片分类表明粒系增多 ,粒系 :红系比例增高。因此得出结论Smad3基因剔除使小鼠造血干祖细胞数目     

小鼠胎肝基质细胞造血刺激因子体外生物活性研究

本实验对基质细胞造血刺激因子－１（ＳＨＦ－１）的体外生物活性进行了研究。结果表明,ＳＨＦ－１可刺激小鼠骨髓ＣＦＵ－Ｅ、ＢＦＵ－Ｅ、ＣＦＵ－ＧＭ、ＣＦＵ－Ｍｉｘ集落的形成,它产生的这些广泛造血刺激作用是其自身所具活性的直接影响。正常小鼠骨髓细胞与ＳＨＦ－１在体外孵育４ｈ,其中ＣＦＵ－Ｓ的自杀率可提高约１０％,显示它对造血干细胞也有诱导增殖作用。     

辐射对造血微环境损伤在放疗中的意义

本文观察了500～3000rad、局部照射后一年内骨髓中CFU-S数的变化动态,同时了解造血微环境支持造血的功能之演变过程。实验发现,500rad照射后局部骨髓中CFU-S含量明显减少,恢复不稳定,同时造血微环境支持造血的功能亦有相类似的波形起伏的损伤修复过程。1000rad局部照射的骨髓中CFU-S有更显著的降低,恢复缓慢而不稳定,造血微环境支持造血的功能早期明显受损,以后虽有修复但不能恢复到正常水平。2000rad以上的X线照射可导致局部骨髓长期再生不良,造血微环境亦见剧烈而持久的功能缺陷,这一结果表明:局部照射后,屏蔽区正常造血干细胞不能在照射部位骨髓中正常种植增殖,其原因与局部造血微环境的功能障碍密切相关。     

5-Fluorouracil treatment after irradiation impairs recovery of bone marrow functions

Summary In mice, persisting radiation-induced growth retardation of hematopoietic tissue suggested that at least part of the surviving stem cells are genetically injured. Additional mitotic stress some time after the radiation insult might remove injured stem cells, thus improving the overall recovery of the irradiated bone marrow.Mice were treated with 5 Gy whole-body gamma irradiation. Two weeks later half of the animals were injected i.v. with 150 mg/kg 5-fluorouracil (5-FU), the other half remained untreated (5 Gy-controls). 2 or 10 weeks later, femoral cellularity and CFU-S content, proliferation ability of transplanted bone marrow and the compartment ratio (CR; ratio of splenic IUdR incorporation at day 3 and number of CFU-S transfused) were determined.Four weeks after 5 Gy and 2 weeks after 5-FU treatment all parameters showed significant impairment of recovery. 12 weeks after 5 Gy and 10 weeks after 5-FU CFU-S and CR were still reduced compared to the 5 Gy-controls. 5-FU treatment of unirradiated mice did not produce permanent effects on the quality of stem cells or the hematopoietic microenvironment. It is concluded, therefore, that an increased proliferation stimulus does not aid in the removal of injured CFU-S and may even impair recovery of bone marrow functions by increasing the proportion of genetically injured stem cells which continue proliferation.Dedicated to Prof. L.E. Feinendegen on the occasion of his 60th birthday     

Radioprotective effects of ginsan,an immunomodulator

We previously reported that ginsan, a purified polysaccharide isolated from Panax ginseng, had a mitogenic activity, induced LAK cells, and increased levels of several cytokines. In an effort to identify other immunostimulatory effects, we evaluated the protective effects of ginsan injected in vivo against radiation by measuring its effects on the CFU-S bone marrow cells and spleen cells. Ginsan was found to significantly increase the number of bone marrow cells, spleen cells, granulocyte-macrophage colony-forming cells (GM-CFC), and circulating neutrophils, lymphocytes and platelets in irradiated mice. In addition, ginsan induced the endogenous production of cytokines such as Il1, Il6, Ifng and Il12, which are required for hematopoietic recovery, and was able to enhance Th1 function while interfering with the Th2 response in irradiated mice. We demonstrated that pretreatment with ginsan protected mice from the lethal effects of ionizing radiation more effectively than when it was given immediately after or at various times after irradiation. A significant increase in the LD(50/30) from 7.54 Gy for PBS injection to 10.93 Gy for mice pretreated with 100 mg/kg ginsan was observed. These findings indicate that ginsan may be a useful agent to reduce the time necessary for reconstituting hematopoietic cells after irradiation.     

The development of radiation late effects to the bone marrow after single and chronic exposure

The marrow is a tissue distributed in numerous skeletal parts and works as an organ which is composed of a haemopoietic cell parenchyma and a supporting stroma. The pathophysiological mechanisms involved in the radiation-induced late effects depend mainly on the damage produced to each of these elements. Parenchymal cell damage ends with a failure of the stem cell pool to supply an adequate number of highly differentiated functional blood cells and is clinically manifested as aplastic anaemia or leukaemia. The effects of radiation on the haemopoietic stem cell can be measured by means of spleen colony forming units (CFU-S) in rodents. The self-maintaining capacity of the CFU-S was found to be lower than normal 16 weeks after a dose of 0.64 Gy. In larger animals it is only possible to measure the activity of some of the progenitor cells, estimating the number of granulocyte-macrophage colonies in culture (CFU-GM) as an indicator of stem cell changes. Their number in the blood is about 50 per cent of normal even 160 days after about 0.78 Gy. The stromal cells are also radiosensitive if measured with respect to their capacity to support long-term cell replication in vitro. Marrow fibrosis develops after single, repeated and chronic radiation exposure, and a dose of 40 Gy impairs the capacity of the marrow to support haemopoiesis.     

Ex vivo expansion of primitive hematopoietic cells for cellular therapies: An overview

Sources of hematopoietic cells for bone marrow transplantation are limited by the supply of compatible donors, the possibility of viral infection, and autologous (patient) marrow that is depleted from prior chemo- or radiotherapy or has cancerous involvement. Anex vivo system to amplify hematopoietic progenitor cells could increase the number of patients eligible for autologous transplant, allow use of cord blood hematopoietic cells to repopulate an adult, reduce the amount of bone marrow and/or mobilized peripheral blood stem and progenitor cells required for transplantation, and reduce the time to white cell and platelet engraftment. The cloning of hematopoietic growth factors and the identification of appropriate conditions has enabled the development of successfulex vivo hematopoietic cell cultures. Purification systems based on the CD34 marker (which is expressed by the most primitive hematopoietic cells) have proven an essential tool for research and clinical applications. Present methods for hematopoietic cultures (HC) on stromal (i.e. accessory cells that support hematopoiesis) layers in flasks lack a well-controlled growth environment. Several bioreactor configurations have been investigated, and a first generation of reactors and cultures has reached the clinical trial stage. Our research suggests that perfusion conditions improve substantially the performance of hematopoietic reactors. We have designed and tested a perfusion bioreactor system which is suitable for the culture of non-adherent cells (without stromal cells) and readily scaleable for clinical therapies. Eliminating the stromal layer eliminates the need for a stromal cell donor, reduces culture time, and simplifies the culture system. In addition, we have compared the expansion characteristics of both mononuclear and CD34⁺ cells, since the latter are frequently assumed to give a superior performance for likely transplantation therapies.Abbreviations BFU0-E burst forming unit-erythroid - BM bone marrow - CB cord blood - CFU-C colony forming unit-culture - CFU-E colony forming unit-erythroid - CFU-F colony forming unit-fibroblast - CFU-GEMM colony forming unit-granulocyte, erythroid, macrophage, megakaryocyte - CFU-GM colony forming unit-granulocyte, macrophage - CFU-Mix colony forming unit-mixed (also known as CFU-GEMM) - CML chronic myeloid leukemia - CSF colony stimulating factor - DMSO dimethyl sulfoxide - ECM extracellular matrix - EPO erythropoietin - FL fetal liver - HC hematopoietic culture - LTBMC long-term bone marrow culture - LTC-IC long-term culture initiating cell - LTHC long-term hematopoietic culture - MNC mononuclear cells - PB peripheral blood