PI3K激活NF-κB信号通路保护中子辐射致IEC-6细胞损伤

中子属于高传能线密度电离辐射,能产生比κ射线更为严重的放射损伤,肠上皮对中子辐射高度敏感,迄今未见有关中子辐射致肠上皮细胞损伤中PI3K对NF-κB信号通路调控的研究报道.本研究旨在探讨中子照射后肠上皮细胞中PI3K对NF-κB信号通路的调控及其在中子辐射致肠上皮细胞损伤中的作用.选取肠上皮细胞系-6（intestinal epithelial cell No.6,IEC-6）进行传代培养,随机分为对照组、4Gy中子照射组和4Gy中子照射＋LY294002处理组,照射组和LY294002处理组细胞采用4Gy中子均匀照射,LY294002处理组细胞在照前24h给予终浓度为10κmol/L的LY294002,各组于照射后6和24h采用MTT比色法、流式细胞术和免疫印迹（Western blot）方法检测IEC-6细胞增殖活力、凋亡与坏死率以及NF-κB信号通路相关分子NF-κB（p65）,IKKκ和IκBκ的表达变化.研究发现,4Gy中子照射后6和24h,IEC-6细胞增殖活力下降,凋亡和坏死率增加;应用LY294002后IEC-6细胞增殖活力较照射组明显下降,IEC-6细胞凋亡和坏死率较照射组增加.4Gy中子照射后6和24h,IEC-6细胞NF-κB（p65）和IKKκ表达升高,IκBκ表达降低;应用LY294002后NF-κB（p65）和IKKκ表达降低,IκBκ表达升高,表明4Gy中子照射可引起IEC-6细胞增殖活力下降,凋亡和坏死率增加;PI3K可激活NF-κB信号通路,对中子辐射IEC-6细胞损伤发挥保护作用.     

γ射线对红细胞嘌呤核苷磷酸化酶的影响 一、酶活力的比较研究

用大鼠、小鼠及狗作为实验动物进行辐射对红细胞嘌呤核苷磷酸化酶活力影响的比较研究。在离体实验中,用9700或97000拉德照射大鼠及小鼠的肝素抗凝血,可观察到红细胞嘌呤核苷磷酸化酶活力轻微减低,而在整体实验中却得到以下的不同结果: 1.用400、600及700拉德分别照射大鼠,均可使酶活力下降。400或600拉德照射后酶活力下降至一定时日可恢复至照射前水平,但700拉德照射后酶活力一直下降且始终未恢复。照射后大鼠酶活力的抑制与恢复程度与辐射损伤的轻重有关。2.以800、900和1000拉德的剂量分别照射小鼠后,酶活力无明显变化。经700或1200拉德照射后第1天,狗的红细胞嘌呤核苷磷酸化酶活力较照射前显著下降,而且以后各日,一直处于低的水平。     

X射线全身辐射对乌龟存活率的影响

利用X射线全身辐射对乌龟进行了抗辐射作用的研究,并用小鼠作参照.根据前人的研究资料,照射剂量选用6.5 Gy和15 Gy两个水平. 结果显示,乌龟受照射后,6.5 Gy剂量组60 d内无死亡;15 Gy剂量组从第16 d开始出现死亡,30 d内死亡率为20%,60 d内死亡率为70%.乌龟体重照射前后变化不大.小鼠受照射后,6.5 Gy剂量组第14d开始出现死亡,30 d内死亡率为30%,60 d内死亡率为80%:15 Gy剂量组第4 d出现死亡,第5 d死亡率达100%.研究结果表明,乌龟的抗辐射能力明显高于小鼠.     

狗和猴离体血液丙线照射后淋巴细胞酸性磷酸酯酶活力改变与辐射剂量的关系

本文用细胞化学的方法证实了在不加培养基的狗和猴离体血液中,淋巴细胞形志及Acp活力在24小时内仍能保持正常。在上述条件的基础上得出了离体血液丙线200、300、400及600拉德照射后,淋巴细胞Acp活力“4”分型改变与辐射剂量相关的方程。 文中还就离体血液丙线照射后Acp活力测定应用于生物剂量估算、某些抗放药物效价的验证以及照射后淋巴细胞Acp活力改变的机理作了初步讨论。 作者曾在丙线外照射中小剂量诊断指标的研究过程中,观察到家免的外周血液淋巴细胞酸性磷酸酯酶(Acp)活力与受照剂量之间存在线性关系 (顾远锡,1980)。以后又发现具有抗放作用的胰岛素对丙线照射狗的外周血液淋巴细胞Acp活力有一定的保护作用 (顾远锡,1980)。这些结果提示,外周血液淋巴细胞Acp活力的测定有可能作为受照剂量的估算方法,并可用来判断某些抗放药物的效价。后经动物实验证实了上述设想 (顾远锡,1981)。剂量估算方法和药物效价判断的应用,其主要对象是人,但是在人身上是无法进行整体照射情况下淋巴细胞Acp活力的剂量—效应关系的研究。进一步设想如果动物及人的离体血液经照射后,其淋巴细胞Acp活力与照射剂量之间仍存在线性关系,而且在给抗放药后取血照射仍能看出药物的保获作用,则这一生物指标不仅有可能用于整体动物     

γ射线对小鼠调节性T细胞功能及相关细胞因子的影响及其意义

调节性T细胞（regulatory T cells,Tregs）及相关细胞因子在机体免疫平衡的调节中发挥重要作用,而其在放射免疫损伤中的作用尚不明确.本实验以6Gyγ射线照射C57BL/6小鼠,于照射后1～28d不同时间,检测外周血、胸腺和脾脏Treg细胞亚群及血清中细胞因子IL-2,IL-10及TGF-γ含量的变化,以探讨其在放射免疫损伤中的作用机制.结果显示,小鼠经6Gyγ射线照射后各组织CD4＋CD25＋Treg细胞比例明显增加（P〈0.05或P〈0.01）,胸腺CD4＋CD25＋Foxp3＋Treg细胞比例于照后1d即明显增高（P〈0.01）,而在照后7d明显低于未照射组（P〈0.01）;血清抑制性细胞因子IL-10（7d）,TGF-γ（3d）含量明显增高（P〈0.05）,而IL-2浓度持续降低.本文揭示了Treg细胞及其相关细胞因子与辐射所致免疫功能受抑和免疫调节功能失衡密切相关,为进一步的辐射损伤机制研究奠定基础.     

不同刺激因子对狗骨髓CFU-C的影响

集落刺激因子(CSF)是集落形成单位(CFU-C)体外培养中的一个必不可少的条件,对培养结果的好坏起着决定性的作用。在狗的骨髓细胞培养中,同种异体血清是目前普遍采用的一种CSF,但由于制备方法不同,其刺激活力可有显著差别。为了确定狗血清刺激活力的适宜条件,我们对几种不同的CSF进行了研究。     

谷氨酰胺对低剂量电离辐射损害的保护作用

观察小剂量电离辐射条件下大鼠补充谷氨酰胺(Gln)对体内谷胱甘肽(GSH)代谢的影响。SD雄性大鼠受照射后经饲料补充2%Gln。照射源为60Co;剂量率6×10-2Gy/h,1h/d,5d/周,累积剂量1.5Gy。与对照组相比,受照射大鼠睾丸重量降低,精子畸变率增高,肝脏GSH含量降低,外周血白细胞计数降低,血清Gln及Glu+Gln含量降低,差异具有显著性,补充Gln则与对照组无明显差异。表明小剂量电离辐射导致大鼠出现可逆性损害,机体的Gln需求有所增加,补充Gln对大鼠的GSH代谢有一定益处。     

SD大鼠急性放射性皮炎模型的建立

目的:建立安全的SD大鼠急性放射性皮炎模型。方法:将28只雄性SD大鼠随机分为7组,分别为空白对照组,电子线照射组(60 Gy,45 Gy,30 Gy),X线照射组(45 Gy,30 Gy,15 Gy),每组4只。选择臀背部皮肤,去毛后照射。放疗后第一天起开始观察动物皮肤表现,采用Douglas and Fowler评分方法记录每只动物皮炎情况,并定期测量动物体重,观察动物一般情况并记录死亡情况。于放疗后第28天处死动物,取照射区域皮肤行HE染色及免疫组化染色(CD3,CD11c,CD68,IV型胶原),以通过光镜分析射线照射后皮肤组织变化情况、真皮层内炎症细胞浸润类型及胶原形成情况。结果:至放疗后第28天X线照射组动物出现大量死亡,电子线照射组动物均存活,电子线各组均出现不同程度的放射性皮炎反应,镜下可见局部组织不同程度的表皮层坏死、炎症细胞浸润、毛囊及附属器减少等表现,以电子线照射60 Gy及45 Gy组表现明显。免疫组化结果显示放射线照射可使真皮层内以CD68为表面标志的巨噬细胞浸润增加,并促进以IV型胶原为标志胶原细胞形成。结论:电子线60 Gy及45 Gy照射SD大鼠臀背部皮肤可建立一种安全有效的急性放射性皮炎动物模型,其临床表现及病理表现可用于实验研究。     

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。     

γ射线对人淋巴细胞cx43和ANLN基因转录表达的影响

目的研究γ射线对人外周血淋巴细胞cx43和ANLN基因转录表达的影响。方法对数生长期的淋巴细胞,分别给予1、2、3、4、5、6 Gy的^60Coγ射线照射,照射后12h,以及2Gy照射后4、8、12、24、36、48、72h,分别提取总RNA,反转录成cDNA。利用实时荧光定量PCR技术,检测各组cx43和ANLN基因表达改变。结果人外周血淋巴细胞cx43 mRNA表达水平在2Gy照射后4、8、12h明显增高,分别为对照组（未照射组）的6．74、9．06、7．22倍（P〈0．05）;24～72h,其表达水平与对照组相比没有明显变化。1、2、3、4、5、6Gy剂量照射后12h,cx43 mRNA表达水平显著增高（P〈0．05）。ANLN mRoNA表达水平在2Gy不同时间点及1～5Gy照射后12h,表达降低（P〈0．05）,6Gy照射后12h其表达开始升高,为对照组的6．08倍（P〈0．05）。结论γ射线照射2Gy不同时间点及不同剂量照射后12h,cx43基因表达上调,ANLN基因表达下调。1～3Gy剂量照射后12h,cx43 mRNA表达在此范围内有时间和剂量的依赖性。cx43可能会发展为核事故受照射人员的分子生物学剂量标记物。     

Increase in the activity of nuclear DNA polymerases in the tissues of mice subjected to long-term gamma irradiation

Activity of nuclear DNA-polymerase in the liver, lung and spleen tissues of mice subjected to long-term chronic gamma-irradiation (1.3 mGy/h) has been investigated. Chronic gamma-irradiation with a cumulative dose of 1.7 Gy during 55 days raises DNA polymerase activity in the irradiated tissue nuclei. Analysis of DNA-polymerase activity in the liver nuclei have demonstrated that this increase is connected with activation of DNA-polymerase beta.     

Lithium-stimulated recovery of granulopoiesis after sublethal irradiation is not mediated via increased levels of colony stimulating factor (CSF)

We have previously demonstrated that lithium (Li) is an effective agent in accelerating the recovery of granulopoiesis following sublethal (2 Gy) whole body irradiation. In this report, studies are described that further define this Li-mediated recovery by measuring the levels of colony-stimulating factor (CSF) present in serum from mice administered 105 micrograms/mouse (total dose) of ultra-pure Li2CO3 for 3 days immediately following irradiation. On days 1-28 following the last lithium dose, the serum was tested for its CSF activity against both normal non-adherent derived bone marrow target cells and non-adherent marrow cells from mice administered cyclophosphamide (200 mg/kg body weight). Serum was assayed at 0.01, 0.1, 1 and 10 per cent final concentration. No significant difference in the total number of CFU-GM was observed from normal marrow using either serum from irradiated mice or lithium-treated and irradiated mice, although the irradiation did produce a 300 per cent rise in CFU-GM colonies compared to normal serum (days 4 and 10-15). From regenerating marrow, we observed a significant difference (P less than or equal to 0.01) in CFU-GM cultured with serum at 0.1 per cent concentration from irradiated and lithium-treated mice compared to irradiated mice without lithium. The presence of CSF was confirmed by its reduced activity in the presence of anti-(CSF). These results suggest (Li) may increase the sensitivity of CFU-GM to CSF, thereby producing more CFU-GM ultimately providing more circulating granulocytes.     

Insulin-like growth factor-1 and growth hormone (GH) levels in canine cerebrospinal fluid are unaffected by GH or GH secretagogue (MK-0677) administration.

Elevation in circulating GH levels results in a dose-related increase in serum insulin-like growth factor-1 (IGF-1) levels in dogs. However, it is not known whether elevations in systemic IGF-1 and GH levels contribute to the cerebrospinal fluid (CSF) levels of these hormones. Therefore, a study was designed in dogs to determine if elevated circulating GH levels was a result of a GH secretagogue (MK-0677) or if exogenous GH administration resulted in increased IGF-1 and GH levels in the CSF of dogs. A total of 12 normal, young adult male dogs were randomized to three treatment groups (4 dogs/group) based on body weight. There were 4 vehicle control dogs. A group of 4 dogs were dosed orally with MK-0677 (5 mg/kg/day) dissolved in deionized water. A third group of 4 dogs received subcutaneous injections of porcine GH (pGH) at a dose of 0.1 IU/kg/day. From all dogs, blood and CSF samples were collected prior to the initiation of treatment and on days 7 and 15 of treatment. All samples were assayed using a validated radioimmunoassay. Administration of MK-0677 or pGH resulted in a statistically significant (P < or = 0.05) increased body weight gain and increased serum IGF-1 and GH levels. In contrast, administration of MK-0677 resulted in no significant (P > 0.05) increase in CSF IGF-1 or GH levels on days 7 or 15 of the study. The CSF IGF-1 values ranged from 1.2 to 2.0 ng/ml with minimal variation among three separate samples taken during the course of the study from each dog. Similarly, the CSF GH levels were very low (< 0.98 ng/ml to 2.4 ng/ml) in all dogs irrespective of treatment group. This study has demonstrated that there is no correlation between the circulating levels of IGF-1 or GH and the levels of these hormones in the CSF of normal dogs. An approximately 100-fold difference between serum and CSF IGF-1 levels in vehicle control dogs suggest that there is a blood-brain barrier for the circulating IGF-1. Similarly, failure to see an elevation in CSF GH levels despite increases in serum GH levels shows that there is a blood-brain barrier for GH in normal dogs. These results suggest that the likely source of GH and IGF-1 in the CSF of dogs is from the CNS.     

Induction of external abnormalities in offspring of male mice irradiated with 252Cf neutron.

To assess the genetic effects of fission neutron, the induction of external malformations was studied in F1 fetuses after F0 male mice were irradiated. Male mice of the ICR:MCH strain were irradiated with 252Cf neutron at doses of 0.238, 0.475, 0.95 and 1.9 Gy. They were mated with non-irradiated female mice at 71-120 days after the irradiation. Pregnant females were autopsied on day 18 of gestation and their fetuses were examined for deaths and external abnormalities. No increases of pre- and post-implantation losses were noted at any dose. External abnormalities were observed at rates of 1.40% in the 0.238 Gy, 2.23% in the 0.475 Gy, 3.36% in the 0.95 Gy and 3.26% in the 1.9 Gy groups; the rate in the control group was 1.65%. The dose-response curve was linear up to 0.95 Gy, and then flattened out; the induction rate of external abnormalities was 2.7 x 10(-4)/gamete/cGy based on the linear regression. These results indicated that fission neutron effectively induces external abnormalities in F1 fetuses after spermatogonial irradiation.     

Two-phase response of rat pineal melatonin to lethal whole-body irradiation with gamma rays.

Male Wistar rats adapted to artificial light:dark (LD) regimen 12:12 h were whole-body irradiated with a single dose of 9.6 Gy of gamma rays and sham/irradiated in the night in darkness. The rats were examined 60 min, 1, 3 and 5 days after exposure between 22:00 and 01:30 h in the darkness. The results obtained indicate a two-phase reaction of pineal melatonin after the lethal irradiation of rats: the decline of melatonin concentration early after the exposure (at 60 min) with unchanged serotonin N-acetyltransferase (NAT) activity followed by an increase of melatonin synthesis, accompanied by an increase of pineal and serum melatonin on day 5 after the exposure. NAT activity was increased on day 3 after the exposure. Serum corticosterone concentrations in irradiated rats were increased 60 min and 3 days after exposure. With respect to the antioxidant, immunomodulating and stress-diminishing properties of melatonin, we consider the increase in melatonin synthesis during later periods after irradiation as part of adaptation of the organism to overcome radiation stress.     

Changes in lipoprotein lipase activity in the adipose tissue and heart of non-lethally X-irradiated rats

After 16 h nocturnal deprivation of food, male Wistar rats were irradiated by a single whole body dose of 2.40 Gy X-rays. Both the irradiated and sham-irradiated (control) rats were pair-fed for the first six days after irradiation, but for the rest of the time they were fed ad libitum. Lipoprotein lipase activity (LPLA) in the adipose tissue fell between 24 and 48 h; LPLA in the heart fell at 24 h and 21 days and rose on the 14th days. The serum triacylglycerol concentration rose between 24 and 72 h. Comparison with the fed control group showed LPLA in adipose tissue to be reduced at 6 and 72 h and on the 28th day and raised between the 7th and the 14th day. In the heart it was raised at 1 h and between 72 h and the 14th day, it was reduced on the 21st day and rose on the 35th day. The triacylglycerol concentration was raised between 48 and 72 h and on the 28th day. Pair-feeding after non-lethal X-irradiation allowed more exact differentiation of the specific effect of ionizing radiation on LPLA in the adipose tissue and heart at the early post-irradiation intervals.     

Effects of single-dose external gamma irradiation on rat thyroid status as observed during the year

We studied the rat thyroid status depending on the dose of external radiation and the time passed after the exposure. The experiments were carried out on female albino Wistar rats. The doses absorbed amounted to 0.25; 0.5; 1.0; 2.0 and 5.0 Gy. The animals were decapitated after 3, 6, 24 hours and 7, 30, 180 and 365 days following the radiation. The blood serum was assayed for the contents of thyroxin (T4) and triiodothyronine (T3) using a radioimmunological technique. The liver tissue was assayed spectrophotometrically for the activity of thyroid-induced NADP malate dehydrogenase (NADP-MDH). No changes were found in the blood thyroid hormone contents within short periods after the radiation effect. After 6 hours the T4 levels was 1.2-1.3-fold decreased in the blood of rats receiving the radiation doses of 1.0; 2.0; and 5.0 Gy. After a day the T4 concentration was diminished by 1.21-193-fold in all the experimental animals independently of the radiation dose and that of T3--in 2.0 Gy--and 5.0 Gy--irradiated group. After 7 days following the radiation the T4 and T3 contents remained to be decreased by 1.37-1.75 fold and those of NADP-MDH--by 1.3-1.8-fold in all the animal groups. In a month, the low dose-treated animals (0.25, 0.5, 1.0 Gy) showed the level of thyroid hormones reduced to the control values, whereas the 2.0 and 5.0 Gy--treated rats demonstrated this reduction only by 6 months. The decreased concentration of blood thyroid hormones was due not to the activation of their peripheral metabolism, but, probably, to inhibition of their biosynthesis in thyroid cells under conditions of radiation-induced activation of oxidative stress.     

Modulatory effect of Mentha piperita (Linn.) on serum phosphatases activity in Swiss albino mice against gamma irradiation

Mentha extract (ME; 1 g/kg body wt) given orally for three consecutive days prior to whole body irradiation (8 Gy) showed modulation of activity of serum phosphatases in albino mice. Values of acid phosphatase activities were significantly higher in untreated irradiated group throughout the experiment. Irradiated animals pretreated with ME showed significant decline in acid phosphatase activity as compared to untreated irradiated animals at all autopsy intervals and attained normalcy at day 5. A marked decrease in serum alkaline phosphatase activity was recorded in both irradiated groups. However, in ME pretreated irradiated group, values of alkaline phosphatase activity remained significantly higher than untreated irradiated animals at all intervals and attained normalcy from day 5 onwards.     

松茸多糖抗辐射功能的初步研究

预防给予小鼠松茸多糖(PTM),测定小鼠在受到2 Gy X线照射后,脾和胸腺重量、T淋巴细胞转化能力、外周血中白细胞数和肝组织中SOD、CAT、GSH-Px活性及MDA含量的变化,结果显示PTM处理组小鼠的各项指标与辐射对照组比较均有显著性差别,提示PTM可促进机体自由基的清除,增加机体抗氧化能力,对辐射所致的免疫损伤有明显的保护作用。     

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。