阿霉素肾病大鼠肾皮质糖皮质激素受体与核因子-κB 活性的变化

目的探讨阿霉素(ADM)肾病综合征(NS)大鼠肾组织转录因子核因子-кB(NF-κB)和糖皮质激素受体(GR)DNA结合活性的变化及地塞米松(Dex)、环孢霉素(CsA)对其活性的影响。方法应用凝胶电泳迁移率法(EMSA)和同位素放射自显影等方法检测:⑴阿霉素肾病大鼠模型形成过程的不同时间点NF-κB、GR的DNA结合活性、血液生化指标和尿蛋白的排泄量;⑵Dex和CsA治疗对上述指标的影响。结果大鼠经尾静脉注射阿霉素后7 d,24 h尿蛋白排泄量(UpV)(mg/24h)开始升高,21 d出现大量蛋白尿,低蛋白血症,高胆固醇血症。注射阿霉素后第7天,大鼠肾皮质组织NF-κB活性开始升高,28 d达高峰,明显高于正常对照组(P<0.01);正常对照组及肾病7 d组GR活性最高,14 d、21 d、28 d组GR活性明显降低(P<0.01)。Dex不能抑制ADM大鼠的肾皮质细胞中已经升高的NF-κB结合活性(P>0.05),同时,也不能使GR DNA结合活性升高(P<0.01);而CsA能抑制NF-κB结合活性(P<0.01)。Dex不能减少ADM大鼠UpV(P>0.05);而CsA可减少(P<0.05)。结论ADM肾病大鼠肾皮质中NF-κB DNA结合活性异常升高,GR活性降低。CsA可抑制NF-κB活性,减少尿蛋白排泄量。     

长期接触低剂量氯化甲基汞致脑发育损伤大鼠动物实验模型的研究

目的:观察大鼠长期接触低剂量氯化甲基汞(MMC)不同发育阶段仔鼠脑汞含量。方法:雌性Wistar大鼠体重(120±20)g,随机分为3个实验组和对照组各30只。实验组母鼠从妊娠前90天至仔鼠出生后30天连续喂饲含有不同剂量(0.75、1.50和3.00 mg/kg)MMC的普通饲料,对照组给予普通饲料。采用干烧法用F732-V冷原子吸收测汞仪分别测定仔鼠大脑、小脑和海马组织汞含量。结果:在建立MMC所致脑发育损伤动物实验模型过程中,发现实验组母鼠的产子率低于正常鼠,高剂量组尤为明显,生后仔鼠外观无异常表现,不同脑区(大脑、小脑和海马)脑汞含量差异无统计学意义(P>0.05)。各实验组生后不同时间仔鼠脑汞含量明显高于相应对照组(P<0.05),各实验组仔鼠脑汞含量随母鼠接触剂量的增高而有不同程度升高(P<0.05),同一实验组内仔鼠脑汞含量随生后时间的延长逐渐升高,在生后30天达峰值。结论:甲基汞可透过血脑屏障蓄积于仔鼠脑内,且仔鼠脑汞含量与母鼠接触剂量之间存在一定剂量反应关系。     

氯化甲基汞致脑发育损伤形态学研究

目的:探讨氯化甲基汞(MMC)致大鼠不同发育阶段脑组织形态结构改变。方法:雌性Wistar大鼠体重(120±20)g,随机分为3个实验组和对照组各30只。3个实验组母鼠从妊娠前90天至仔鼠出生后30天连续喂饲含有不同剂量(0.75、1.50和3.00 mg/kg)MMC的普通饲料,对照组给予普通饲料。脑切片经苏木精-伊红染色,在光镜下观察长期接触低剂量MMC仔鼠脑组织形态结构。采用荧光原位末端标记法,在激光共聚焦显微镜下计数两组生后不同时间点仔鼠大脑、小脑和海马发生凋亡的神经元,以PCD阳性率表示凋亡的多少。结果:在实验设计的接触剂量下,仔鼠大脑、小脑和海马神经元体积缩小、固缩,核染色质致密等凋亡形态特征。实验组和对照组生后不同时间点仔鼠脑神经元都存在凋亡。结论:大鼠长期低剂量接触不同浓度MMC可使其生后仔鼠大脑、小脑和海马部分神经元出现凋亡形态学改变,随仔鼠脑汞含量升高神经元凋亡率明显增加。     

实验性肺纤维化肺组织胶原形成与核因子κB活性变化

目的 观察实验性肺纤维化大鼠肺组织胶原形成与NF-κB活性变化,初步探讨NF-κB活性变化与肺组织胶原合成的关系.方法 用SD雄性大鼠一次性气管内注入博来霉素(BLM)(5 mg/kg体重)复制肺纤维化模型,对照组气管内注入生理盐水,于第1、7、14、21、28天处死,用羟脯氨酸含量测定法评估肺纤维化胶原形成程度.用HE染色结合NF-κB p65免疫组织化学方法观察肺组织NF-κB活性变化.结果 模型组大鼠肺组织自第1天开始即出现炎症细胞浸润,肺泡间隔增厚,自第7天开始肺组织出现胶原沉积,羟脯氨酸含量逐渐增高,于28 d达高峰.NF-κB p65蛋白表达自第1天增高,第7天达峰值,第14天回落,随着肺纤维化加重,第21天以后接近对照组水平.结论 肺组织NF-κB活化是肺纤维化过程中早期事件,可能在肺组织胶原沉积的形成中起重要作用.     

窒息新生鼠脑核因子-κB的表达及地塞米松的影响

【目的】观察核因子-κB（nuclear factor kappa B，NP-κB）在窒息新生大鼠窒息后不同时间脑组织内表达的动态变化及地塞米松（dexamethasone，DXM）早期干预的影响。【方法】出生7d新生大鼠随机分为：对照组，窒息组和DXM预处理组；制备新生大鼠窒息模型，并于窒息后1、3、5和7d取脑组织行石蜡切片。采用免疫组织化学方法检测窒息新生鼠脑组织NF-κB p65活性表达水平及应用DXM预处理后NF-κB p65活性变化情况，用苏木精-伊红染色观察窒息后脑组织形态学变化。【结果】窒息组新生鼠脑组织NF-κB的表达较对照组显著增强（P〈0．05），以5d时升高最明显；给予DXM预处理后，NF-κB的表达减弱（P〈0．05），窒息组神经元有凋亡现象，DXM预处理组凋亡神经元明显减少。【结论】窒息新生鼠脑组织NF—κB表达增加，凋亡神经元增多，窒息后5d时NF-κB的活性达高峰；DXM可抑制窒息新生鼠NF-κB的表达，减少神经元的凋亡。DXM可能通过抑制NF-κB活性减轻窒息脑损伤。     

甲基汞对幼年大鼠认知及甲状腺激素的干扰效应

目的探讨出生后早期暴露甲基汞(MMC)所致仔鼠神经行为的发育毒性及对甲状腺激素的干扰作用。方法将112只新生清洁级大鼠随机分为对照组(生理盐水)和低(0.5mg/kg)、中(1.0mg/kg)和高(2.0mg/kg)剂量MMC染毒组,每组28只。于出生后第2～8天,采用口服方式进行染毒,染毒容量为1.0ml/kg。评价仔鼠一般生理指标和反射行为发育,采用恋窝实验和水迷宫实验检测感官、认知和记忆,并检测血中游离甲状腺激素(T4)、三碘甲腺原氨酸(T3)含量和促甲状激素(TSH)活性。结果与对照组相比,MMC染毒仔鼠的生长发育及一般神经行为发育基本无变化。在恋窝实验中,随着MMC染毒剂量的升高,仔鼠达标率呈下降趋势,达标仔鼠潜伏时间呈延长趋势。在传统水迷宫实验中,MMC染毒仔鼠游泳时间缩短;在反向水迷宫实验中,MMC染毒仔鼠游泳时间延长。与对照组比较,高剂量MMC染毒组仔鼠全血中游离T4含量较高,T3含量较低,而MMC染毒组仔鼠全血中TSH活性均较高,差异有统计学意义(P0.05或P0.01)。结论出生后早期MMC暴露可明显影响仔鼠的感觉、学习和认知能力,其机制可能与甲状腺激素紊乱有关。     

宫内γ线照射对仔鼠脑发育影响的研究

目的：探讨宫内γ线照射对仔鼠脑发育的影响。方法：采用整体动物实验，于SD大鼠孕17天给予1．0Gyγ线照射，观察生后1，4，7，14及21日龄仔鼠脑重、脑DNA和蛋白质含量的动态变化。结果：照射组仔鼠生后1，4，7，14及21日龄脑重、脑／体比值及脑组织蛋白质含量，与对照组比较均显降低；对照组仔鼠出生后脑组织DNA含量逐天下降，至生后第4天后又逐渐增高，照射组仔鼠第1及21日龄脑组织DNA含量显低于对照组。结论：孕17天大鼠受1．0Gyγ线照射可影响仔鼠脑发育，表现为脑重、脑DNA和蛋白质含量的降低。     

白介素-18水平及核因子-κB活性变化与早产儿脑室周围白质软化相关性研究

目的:探讨白介素-18(IL-18)及核因子-κB(NF-κB)在早产儿脑室周围白质软化(PVL)发病机制中的作用,并为预防和治疗早产儿PVL提供可能的理论依据。方法:经头颅B超诊断为PVL的早产儿21例,并根据PVL病变程度分为Ⅰ级PVL组早产儿8例、Ⅱ级PVL组6例,Ⅲ、Ⅳ级PVL组7例;对照组早产儿20例。分别取出生6h内静脉血应用酶联免疫吸附试验测定血清IL-18水平及单核细胞NF-κB活性。结果:①Ⅰ级PVL组早产儿血清IL-18水平及外周血单核细胞NF-κBp65活性明显高于对照组(P<0.05);②Ⅱ级PVL组早产儿血清IL-18水平明显高于Ⅰ级PVL组(P<0.05);③Ⅲ、Ⅳ级PVL组早产儿血清IL-18水平及血单核细胞NF-κBp65活性明显高于Ⅱ级PVL组(P<0.05);④PVL组早产儿血清IL-18水平与单核细胞NF-κBp65活性呈正相关性(P<0.01)。结论:PVL早产儿血清IL-18水平及单核细胞NF-κB活性增高,且PVL病变越重,IL-18水平及NF-κB活性越高,提示IL-18与NF-κB在早产儿脑损伤机制中可能发挥了重要的作用,为进一步研究拮抗NF-κB活性,减少IL-18表达以预防和治疗早产儿PVL开辟新的思路。     

水杨酸钠对百草枯所致大鼠肺组织核转录因子和炎性因子的影响

[目的]探讨水杨酸钠（sodium salicylate,NaSAL）对抗百草枯（paraquat,PQ）致肺损伤的作用机制。[方法]将72只雄性SD大鼠随机分为溶剂对照组（生理盐水灌胃）、PQ染毒组和PQ＋NaSAL干预组。PQ经口染毒,剂量为80 mg/kg（按大鼠体重）,干预组给予染毒组相等剂量PQ后立即腹腔注射NaSAL（120 mg/kg）。在染毒后第1、3、7、14天取对照组、染毒组、干预组各6只动物进行检查,分别用凝胶电泳迁移率实验（EMSA）和酶联免疫吸附法（ELISA）检测肺组织中核转录因子（NF-κB）活性变增高;肺组织中TNF-α、TGF-β1蛋白表达增强,各时点蛋白水平与对照组相比差异有统计学意义（P〈0.05）。干预组NF-κB活性较染毒组降低;与染毒组相比,干预组肺组织TNF-α、TGF-β1蛋白水平表达明显下降,相应时点差异也有统计学意义（P〈0.05）。[结论]NaSAL干预后可以降低肺组织中NF-κB的活性,减少TNF-α、TGF-β1的蛋化及肿瘤坏死因子（TNF-α）、转化生长因子-β（1TGF-β1）蛋白含量的改变。[结果]染毒组大鼠肺组织中NF-κB活性白表达,提示NaSAL可以改善PQ中毒引起的肺损伤。     

磺胺二甲嘧啶对大鼠早期身体及智力发育的影响

目的研究受孕中晚期及哺乳期母体接触磺胺二甲嘧啶对子代大鼠早期身体及智力发育的影响。方法受孕母鼠在受孕第7天起至哺乳期结束分别给予磺胺二甲嘧啶0、50、100、200mg/(kg·d),观察F1代大鼠体重及开眼时间,测定各组出生后不同天龄大鼠的血清游离T4及TSH水平,并用水迷宫实验检测各组20～30天龄大鼠的学习和记忆功能。结果实验组F1代大鼠血清游离T4水平低于对照组,同时发现实验组F1代大鼠体重降低,开眼延后,以及学习记忆功能障碍,均表现出剂量-反应关系,尤其是100、200mg/kg剂量组与对照组相比,差异有统计学意义(P<0.05)。结论发育期接触磺胺二甲嘧啶,可以通过干扰体内甲状腺素水平从而影响大鼠身体及智力的发育。     

Effects of FK506 on acute allograft rejection in transplanted rat heart: the role of mitogen-activated protein kinases, AP-1 and NF-kappaB

To scrutinize the effect of the immunosuppressant on acute allograft rejection as related to the intracellular signal transduction, heterotopic cardiac transplantation was performed from DA rat to Lewis rat with/without FK506. In the experimental group, recipients were given FK506 intramuscularly for 5 days. The control group received placebo. Allograft survivals were compared between two groups. For the assay of mitogen-activated protein kinase (MAPKs) families, activator protein-1 (AP-1) and nuclear factor-kappaB (NF-kappaB) in the left ventricular free wall (LV) and septum (SEP) of the grafts, rats were sacrificed on POD 5 (n=5 in each group). Extracellular signal-regulated kinase (ERK) and p38MAPK were measured using Western blot analysis. AP-1 and NF-kappaB DNA binding activities were measured by electrophoretic mobility shift assay. FK506 prolonged allograft survival (6.5 vs 31 days), and suppressed activation of myocardial MAPKs (ERK: 66% in LV and 67% in SEP, p38MAPK: 62% in LV and 72% in SEP), AP-1 (24% in LV and 18% in SEP), and NF-kappaB (41% in LV and 20% in SEP) (the mean value of activities in the control group was represented as 100%). These results suggest that the signal transduction pathways may play important roles in acute allograft rejection in rat cardiac transplantation.     

Effect of vitamin A compounds on the covalent binding of benzo(a)pyrene to nuclear macromolecules

Effect of vitamin A compounds on the covalent binding of benzo(a)pyrene [B(a)P] to subnuclear components was studied using rat liver nuclei in vitro. The retinol-induced inhibition of the covalent binding of B(a)P depended both on the dose and the time of addition. Retinal and retinyl acetate were both potent inhibitors, but retinoic acid was less effective. Retinol induced marked inhibition of B(a)P binding to DNA and protein of the nuclear matrix but had little or no effect on the labeling of bulk DNA and protein in chromatin fractions.     

低强度2 450 MHz微波对丝裂霉素C遗传毒性作用影响的体外实验

目的研究低强度2 450 MHz微波是否增强丝裂霉素C(MMC)对人外周血淋巴细胞的遗传毒性.方法采用彗星试验和胞质分裂阻断微核试验(CBMN),在体外检测2 450 MHz微波(5.0mW/cm2)与MMC诱发的DNA单链断裂及染色体损伤的情况.结果微波辐射组外周血淋巴细胞的彗星长度[男、女分别为(29.1±8.1)、(25.9±7.5)μm]与对照组[男、女分别为(26.3±6.6)、(24.1±4.3)μm]比较,差异无显著性(P＞0.05);MMC各剂量组(0.012 5、0.025 0、0.050 0、0.100 0μg/ml)的彗星长度均长于对照组,差异有显著性(P＜0.01),而且随着MMC剂量的增加,彗星长度增长;微波联合MMC(MW+MMC)各剂量组的彗星长度也随着MMC剂量增加而增长,且明显长于对照组,差异亦有显著性(P＜0.01),当MMC≥0.025 0μg/ml时,微波与MMC可协同增加DNA单链断裂.微核试验结果表明,微波组的微核率与对照组的差异无显著性(P＞0.05).MMC组和MW+MMC组在MMC≥0.050 0μg/ml时,其微核率与对照组比较,差异有显著性(P＜0.05或P＜0.01).MW+MMC组的微核率高于相应的MMC组,但差异无显著性(P＞0.05).结论低强度2450 MHz微波辐射未能诱发人外周血淋巴细胞DNA和染色体损伤,但彗星试验显示,其可增强MMC诱发的DNA单链断裂效应.     

A telomere-binding protein (TRF2/MTBP) from mouse nuclear matrix with motives of an intermediate filament-type rod domain

In previous work, we identified a telomeric DNA-binding protein (termed telomere-membrane binding protein, MTBP) in the envelope of the frog oocyte nucleus and raised antibodies against it. Here we present immunological evidence which suggests strongly that MTBP is identical with the vertebrate telomeric DNA-binding protein TRF2 (telomere-repeat factor 2). MTBP/TRF2 possesses motif which resembles rod domain characteristic of intermediate filament (IF) proteins as shown by immunological cross-reactivity with characteristic antibodies, as well as amino acid sequence homology. Anti-MTBP antibodies recognised a protein of the same M, as TRF2 in extracts of mouse nuclei and nuclear matrix as shown by ion-exchange chromatography, gel shift assays, and Western blots. This mouse MTBP analogue forms more stable complexes with the vertebrate telomeric DNA fragment (T(2)AG(3))(135) than with the corresponding fragment from Tetrahymena (T(2)G(4))(130). Proteins in each of these complexes are recognised by anti-MTBP antibody. In situ hybridization with the vertebrate telomeric DNA sequence (T(2)AG(3))(135) and immunofluorescence with anti-MTBP antibody had shown earlier that these are co-localised in the nucleus of mouse cells, and here MTBP is shown to be associated with the residual membrane of hepatocyte nuclei using Western blotting and immunofluorescence. Some immunofluorescence signal from MTBP is localized at chromosome extremities on metaphase plates from mouse cell culture, but the main signal is seen in patches scattered around the chromosomes which were identified as remnants of the nuclear envelope by double labelling with antibodies against lamin B. These observations suggest that MTBP/TRF2 is a good candidate for the attachment of telomeres to the nuclear envelope in somatic cells.     

Comparative study of p38 MAPK signal transduction pathway of peripheral blood mononuclear cells from patients with coal-combustion-type fluorosis with and without high hair selenium levels

Coal-combustion-type fluorosis has only been reported in China and its pathogenesis has not been fully understood. Fluoride causes chronic toxic effects and selenium modulates cellular activities through signal transduction in human cells. The present study enrolled three groups of subjects with well-defined serum and urine fluoride and hair selenium: high fluoride + high selenium group, high fluoride group and normal control group. The expressions of p38, NF-kB p65, caspase-3 and p53 genes at both protein and mRNA levels in peripheral blood mononuclear cells (PBMCs) were determined by Western blotting and quantitative real-time RT-PCR, respectively. The results showed that the expressions of p38, NF-kB p65, and caspase-3 protein in high fluoride group were higher than those in the other two groups. The mRNA level of NF-kB p65 and caspase-3 was significantly higher in high fluoride + high selenium group than control and lower than high fluoride group. The mRNA and protein level of p53 was significantly higher in high fluoride + high selenium group than that in other two groups. These results suggest that selenium may influence the protein and gene expression associated with p38 signal transduction pathway and up-regulate p53 expression in PBMCs from patients with coal-combustion-type fluorosis.     

汽车尾气对大鼠肺脏的氧化应激和遗传毒性作用

[目的]探讨汽车尾气对大鼠肺脏生物大分子的氧化损伤作用。[方法]将汽车尾气的颗粒物、冷凝物和半挥发性有机物的二氯甲烷提取物（extracts of gasoline engine exhaust,EGE）减压挥干后用二甲亚砜（dimethyl sulfoxide,DMSO）定容到200L／mL。将40只SD大鼠分为5组,每组8只。以DMSO为溶剂对照,以汽车尾气0、5．6、16．7、50．0L／kg的剂量经气管滴注染毒,每周1次,共4次,末次染毒24h后处死,测定肺脏脏器系数以及肺组织内丙二醛（malondialdehyde,MDA）和羰基蛋白（carbonyl protein,CP）含量以及超氧化物岐化酶（superoxide dismutase,SOD）、谷胱甘肽过氧化物酶（glutathione peroxidase,GSH—Px）活力;并用彗星试验检测肺组织细胞中DNA的损伤程度。[结果]各组动物体重和肺脏脏器系数与对照组相比差异无统计学意义（P〉0．05）;16．7、50．0L／kg剂量组肺组织中MDA含量分别达到了4．57、4．48nmol／mg蛋白,故对照组明显升高（P〈0．05）;CP含量在50．0L／kg剂量组为8．91μmol／mg蛋白,较对照组明显增加（P〈0．05）;SOD和GSH—Px活力在50．0L／kg剂量组分别为1697．61NU／mg蛋白和14．80U／mg蛋白,与对照组比较均明显降低（P〈0．05）。在16．7L／kg和50．0L／kg组,肺组织细胞的拖尾率与对照组比较均明显增加（P〈0．05）,但各组尾长的增加无统计学意义。[结论]汽车尾气可诱导大鼠肺组织生物大分子的氧化损伤和DNA单链断裂。