母鼠孕前慢性应激对成年雄性子代行为学和脑区单胺递质的影响

Objective To examine the effects of maternal rats exposed to chronic unpredictable stress before pregnancy on the behaviors and brain monamine of their adult male offspring.Methods Sixteen SD rats were divided into chronic unpredictable stress (CUS) group and controls.CUS rats were exposed to 21 days chronic unpredictable stressors ,and the controls were stress-free.Ten days after the last stressor, all the female rats were caged with sexually experienced males of the same strain.Then we performed the following experiments on the two months male progeny, sucrose consumption measuring anhedonia, Morris water maze measuring cognitive function and high performance liquid chromatography detecting the contents of monoamine.Results The sucrose consumption showed that both sucrose intake and sucrose consumption percentage of the control progeny were higher than those of the CUS progeny ( sucrose consumption: ( 10.23 ± 4.12 ) g vs ( 6.48 ± 3.19 ) g; sucrose consumption percentage: ( 85.43 ± 20.15 ) % vs (60.98 ± 24.65 ) % ) (P ＜ 0.05 ).The number of times crossing the removed hidden platform in the CUS progeny ( 1.64 ± 1.69) was significantly fewer than that in the control progeny (4.17±2.29 ) in Morris water maze (P ＜ 0.05 ).The contents of serotonin in the hypothalamus of CUS progeny ( ( 500.17 ± 80.94 ) ng/g tissue) was lower than that of the control progeny ( ( 569.63 ± 50.91 ) ng/g tissue) (P ＜0.05) ,while the norepinephrine in the hippocampus of CUS progeny( (2315.01 ± 1397.12) ng/g tissue) was higher than that of the control progeny( (907.56 ± 207.27) ng/g tissue) (P＜0.05) by high performance liquid chromatography.Conclusions Depression or stressful events before pregnancy of dams result in anhedonia, decreased spatial memory and abnormalities in brain monoamine of their adult male progeny.     

抗人白介素-28A单克隆抗体的制备与鉴定

目的制备抗人白介素-28A（hIL-28A）单克隆抗体（mAb）。方法以hIL-28A融合蛋白为免疫原免疫BALB／c小鼠，运用杂交瘤技术制备抗hIL-28A mAb。采用Ig类与亚类鉴定试剂盒鉴定mAb的Ig亚类，通过ELISA、Western blot、免疫组化染色法、流式细胞分析、激光共聚焦显微镜技术鉴定mAb的特异性。结果获得2株稳定分泌抗hIL-28A mAb的杂交瘤细胞，但2株mAb均与hIL-29有弱交叉反应。2株mAb的类别均为IgM。Western blot分析表明，此mAb所识别的靶分子的相对分子质量（Mr）约为19000。免疫组化染色表明，mAb与包皮和肺组织中的上皮细胞、结肠组织中的疑似巨噬细胞呈阳性反应。流式细胞仪分析显示，2株mAb与人脐静脉内皮细胞系HUVEC细胞的反应均呈阳性。激光共聚焦扫描显微镜观察到荧光标记的阳性反应物主要位于HUVEC细胞胞浆内。结论成功地制备抗hIL-28A mAb，为病毒感染性疾病、肿瘤和其他疾病的研究提供新的手段。     

抗蛋白酶激活受体?鄄2单克隆抗体的制备与鉴定

目的:制备出一种具有多种用途的抗蛋白酶激活受体-2(PAR-2)单克隆抗体(mAb).方法:用人工合成的11肽PAR-2作为免疫原,采用皮下多点注射方法免疫BALB/c小鼠,取其脾细胞与NS-1骨髓瘤细胞融合,杂交瘤细胞采用间接ELISA法筛选.通过ELISA、Dot blot、免疫组化染色、流式细胞分析、激光共聚焦显微镜技术鉴定mAb的特异性.结果:获得1株可稳定分泌抗:PAR-2 mAb的杂交瘤细胞.其分泌的mAb为IgM型,ELJSA法检测杂交瘤细胞培养上清效价为1:16;点杂交分析表明此抗体可特异性结合PAR-2;免疫组化染色显示该单克隆抗体与人肺组织中的肺泡上皮细胞和平滑肌细胞,结肠组织中的腺上皮细胞和疑似淋巴细胞,扁桃体中的淋巴细胞,包皮组织中的鳞状上皮细胞呈阳性反应;流式细胞仪分析显示与人肺腺癌细胞系A549细胞中的PAR-2呈阳性反应;激光共聚焦扫描显微镜观察发现荧光标记的阳性反应物位于A549细胞的胞膜和胞浆.结论:成功地制备出可用于免疫组化和点杂交的抗PAR-2单克隆抗体,为PAR-2相关性疾病的研究提供了有用的工具.     

母鼠孕前慢性应激对成年雄性子代行为学和脑区单胺递质的影响

Objective To examine the effects of maternal rats exposed to chronic unpredictable stress before pregnancy on the behaviors and brain monamine of their adult male offspring.Methods Sixteen SD rats were divided into chronic unpredictable stress (CUS) group and controls.CUS rats were exposed to 21 days chronic unpredictable stressors ,and the controls were stress-free.Ten days after the last stressor, all the female rats were caged with sexually experienced males of the same strain.Then we performed the following experiments on the two months male progeny, sucrose consumption measuring anhedonia, Morris water maze measuring cognitive function and high performance liquid chromatography detecting the contents of monoamine.Results The sucrose consumption showed that both sucrose intake and sucrose consumption percentage of the control progeny were higher than those of the CUS progeny ( sucrose consumption: ( 10.23 ± 4.12 ) g vs ( 6.48 ± 3.19 ) g; sucrose consumption percentage: ( 85.43 ± 20.15 ) % vs (60.98 ± 24.65 ) % ) (P ＜ 0.05 ).The number of times crossing the removed hidden platform in the CUS progeny ( 1.64 ± 1.69) was significantly fewer than that in the control progeny (4.17±2.29 ) in Morris water maze (P ＜ 0.05 ).The contents of serotonin in the hypothalamus of CUS progeny ( ( 500.17 ± 80.94 ) ng/g tissue) was lower than that of the control progeny ( ( 569.63 ± 50.91 ) ng/g tissue) (P ＜0.05) ,while the norepinephrine in the hippocampus of CUS progeny( (2315.01 ± 1397.12) ng/g tissue) was higher than that of the control progeny( (907.56 ± 207.27) ng/g tissue) (P＜0.05) by high performance liquid chromatography.Conclusions Depression or stressful events before pregnancy of dams result in anhedonia, decreased spatial memory and abnormalities in brain monoamine of their adult male progeny.     

母鼠孕前慢性应激对成年雄性子代行为学和脑区单胺递质的影响

目的 研究母鼠孕前慢性应激对成年雄性子代行为学和脑内单胺类神经递质的影响.方法 16只成年雌性SD大鼠随机分为正常对照组和慢性不可预见性应激组(CUS),CUS组大鼠每天接受一种应激,共21d,正常对照组不接受任何应激.应激结束后10 d,2组大鼠均与同种SD雄性大鼠合笼,以成年2月龄雄性子代作为研究对象.用糖水消耗实验测量快感缺失,Morris水迷宫检测空间记忆能力,高效液相色谱法测量海马、下丘脑和前额皮质单胺递质的含量.结果 糖水消耗实验示正常对照组子代的糖水消耗量和糖水消耗百分比均显著高于CUS组子代[糖水消耗量:(10.23±4.12)g,(6.48±3.19)g;糖水消耗百分比:(85.43±20.15)%,(60.98±24.65)%](P＜0.05).Morris水迷宫检测示2组的逃避潜伏期无差异,但空间探索测试中正常对照组子代[(4.17±2.29)次]穿越平台的次数明显多CUS组子代[(1.64±1.69)次](P＜0.05).CUS组子代[(500.17±80.94)ng/g脑组织]下丘脑5-羟色胺的含量低于正常对照组子代[(569.63±50.91)ng/g脑组织](P＜0.05),而CUS组子代[(2315.01±1397.12)ng/g脑组织]海马去甲肾上腺素的含量却要高于正常对照组子代[(907.56±207.27)ng/g脑组织](P＜0.05).结论 母鼠孕前慢性应激或抑郁会造成成年雄性子代的快感缺失和空间记忆力下降,以及脑内单胺类神经递质异常.     

生命早期应激小鼠成年后免疫功能的改变

目的 观察生命早期应激对小鼠成年后免疫功能的影响.方法 生命早期应激采用新生鼠母爱剥夺和孤养两种方式,分别在小鼠1,2和3月龄检测外周血白细胞比例、刀豆蛋白A(Con A)和脂多糖(LPS)诱导的脾淋巴细胞转化以及外周血清对正常小鼠类别细胞增殖的影响.结果 生命早期母爱剥夺小鼠成年后外周血中性粒细胞比例由(17.94±1.24)%,(17.33±1.80)%和(21.25±2.82)%分别升高至1,2,3月末的(27.17±3.19)%,(29.75±2.76)%和(28.33±3.12)%;淋巴细胞比例降低.由(79.06±1.18)%,(77.83±3.05)%和(72.46±4.45)%分别降低至1,2,3月末的(69.72±4.97)%,(64.00±3.9)%,(66.74±2.67)%.Con A和LPS诱导的脾淋T和B巴细胞转化增强,外周血清对正常淋巴细胞增殖具有显著的抑制作用.结论 生命早期应激可导致小鼠成年后细胞性免疫功能亢进.     

慢性不可预见性应激和氟西汀治疗对雌鼠类抑郁样和焦虑样行为的影响

目的：研究慢性不可预见性应激（CUS）和氟西汀（Flu）治疗对雌鼠类抑郁样和焦虑样行为的影响。方法：成年雌性SD大鼠随机分为NS组、Flu组、CUS＋NS组和CUS＋Flu组,每组6只。CUS＋NS和CUS＋Flu组每日CUS前30 min分别注射2.5 mL/kg NS或10 mg/kg Flu;NS和Flu组仅给予相同剂量的NS或Flu。最后一次应激注射后第2 d用糖水消耗实验和强迫游泳评估类抑郁行为,开场试验评估类焦虑行为。结果：糖水消耗实验示糖水消耗百分比无组间差异,但CUS＋NS组糖水消耗量明显低于其它3组;强迫游泳示CUS＋NS组的不动时间显著长于其它各组,而NS、Flu和CUS＋Flu组间无统计学意义;开场实验示CUS和CUS＋Flu组中央/总路程低于NS和Flu组（P〈0.05）,CUS＋NS组的中央活动时间少于Flu组,CUS＋Flu组的中央活动时间少于NS和Flu组。结论：CUS大鼠具有类抑郁样行为,同时伴类焦虑样行为,Flu治疗可以逆转类抑郁样行为,但对类焦虑样行为无效;Flu对正常大鼠无情绪改变作用。     

双重免疫胶体金标记组织中肥大细胞内分泌颗粒的超微结构

目的：观察肥大细胞分泌颗粒内特异性酶的分布及其超微结构，并探讨肥大细胞的功能。方法：用双重免疫胶体金标记类胰蛋白酶抗体（AA5）及类糜蛋白酶抗体（CC4），在透射电子显微镜下观察肥大细胞内分泌颗粒的超微结构。结果：在肥大细胞内分泌颗粒中富集类胰蛋白酶和（或）类糜蛋白酶，多数酶蛋白呈细小球形结构；在细胞膜内侧分泌颗粒融合形成分泌通道，细胞的外侧可见肥大细胞脱出的颗粒；细胞内的另侧分泌颗粒呈球形小泡。结论：肥大细胞内酶蛋白的富集可为肥大细胞参与Ⅰ型变态反应提供物质基础。     

抗人蛋白酶激活受体3单克隆抗体的制备与鉴定

目的:制备并鉴定抗人蛋白酶激活受体3(PAR3)单克隆抗体(mAb).方法:以人PAR3为免疫原免疫BALB/c小鼠,运用杂交瘤技术制备抗PAR3 mAb.采用捕获ELISA法鉴定mAb的Ig亚类;通过ELISA、流式细胞分析、激光共聚焦显微镜技术、免疫组织化学染色法、斑点杂交技术鉴定mAb的特异性.结果:获得2株可稳定分泌抗人PAR3 mAb的杂交瘤细胞,其分泌的mAb均为IgM.斑点杂交技术分析表明,其mAb能与PAR3抗原有效结合.免疫组织化学染色表明,mAb与肺组织中的巨噬细胞、结肠组织中的淋巴细胞及疑似肥大细胞、扁桃体和包皮组织中的淋巴细胞的反应呈阳性.流式细胞仪分析及激光共聚焦显微镜观察显示,2株mAb与人肺腺癌A549细胞胞内及膜上的PAR3均呈阳性反应.结论:成功地制备抗PAR3 mAb,为变态反应性和炎症性疾病的研究提供了有用的试剂.     

白介素-1β对原代培养海马神经元毒性的研究

目的 探讨IL-β对体外培养的海马神经元活性及合成和分泌脑源性神经营养因子(BDNF)的影响.方法 采用SD新生鼠进行海马神经元原代培养.培养7d的细胞以细胞免疫化学法枪测神经元特异性烯醇化酶(NSE)的表达,计算阳性细胞率.实验分IL-β为0ng/ml的正常组和IL-β为5 ng/ml、10ng/ml、20ng/ml 3个实验组,进行海马神经元无血清培养48 h.采用甲基噻唑基四唑(MTT)试验检测细胞活性,ELISA法检测培养上清BDNF的含量.结果 NSE细胞免疫化学鉴定显示阳性细胞达到90%以上;IL-1β降低体外培养的海马神经元活性,各剂量组按浓度在590nm波长处检测OD值为(0.5585±0.2407; 0.4295±0.1401; 0.4191±0.1050),与正常组(0.9462±0.2972)比较,差异具有显著性(分别为P<0.05,P<0.01,P<0.01 ANOVA);IL-1β降低体外培养的海马神经元合成和分泌BDNF水平,各剂量组显著BDNF含量分别是[(8.65±0.71)pg/ml,(8.90±0.35)pg/ml,(7.90±0.35)pg/ml],与正常组(12.40±1.77)pg/ml比较差异具有显著性(均P<0.05 ANOVA).结论 IL-1β可导致体外培养海马原代神经元的损伤,其机制可能与海马神经元合成和分泌BDNF的减少有关.