菠萝蛋白酶诱导小鼠急性过敏性气道炎症的作用研究

探讨菠萝蛋白酶诱导小鼠急性过敏性气道炎症中2型固有淋巴细胞(group 2innate lymphoid cell,ILC2)及相关细胞因子的变化及意义。对未致敏BALB/c雌性小鼠在第1天和第3天给予菠萝蛋白酶滴鼻,ELISA检测12h内小鼠肺组织匀浆上清中IL-33变化,5d后处死小鼠,肺组织切片病理观察,瑞氏染色计数BALF中嗜酸性粒细胞含量,流式检测肺组织中ILC2数量变化,ELISA检测BALF中IL-5、IL-13水平。结果显示,菠萝蛋白酶诱导气道上皮细胞产生IL-33,3h时开始,6h时达高峰;菠萝蛋白酶滴鼻小鼠肺组织切片显示大量炎性细胞浸润和杯状细胞黏液分泌增加,BALF中嗜酸性粒细胞比例增加,IL-5和IL-13含量增加,流式检测肺组织中ILC2数量增多。研究表明,菠萝蛋白酶刺激小鼠气道上皮细胞释放IL-33,引起小鼠肺ILC2活化增殖并分泌大量Th2型细胞因子,引起嗜酸性粒细胞浸润等急性过敏性气道炎症表现,提示ILC2促进早期急性过敏性气道炎症的发生。     

Thl7淋巴细胞在哮喘小鼠气道炎症中的初步研究

目的:初步探索Th17细胞及其分泌的炎症介质在哮喘小鼠气道炎症中的作用机制.方法:20只小鼠随机均分为哮喘组和正常对照组.哮喘组用卵白蛋白(OVA)致敏与激发建立小鼠哮喘模型.正常对照组致敏与激发均以生理盐水代替.HE染色观察小鼠气道及肺组织病理变化;光学显微镜下观察小鼠支气管肺泡灌洗液(BALF)中细胞分类及计数;酶联免疫吸附试验(ELISA)检测小鼠BALF上清中IL-4、IL-5、IL-13、IFN-γ及IL-17的含量,流式细胞技术(FCM)检测小鼠外周血Th1、Th2及Th17淋巴细胞占CD4+T淋巴细胞百分率情况.结果:哮喘组小鼠BALF中细胞总数和中性粒细胞、嗜酸性粒细胞、淋巴细胞百分率均显著高于对照组(P<0.05),BALF上清中IL-4、IL-5、IL-13及IL-17的水平显著增高(P<0.05),而IFN-γ差异无统计学意义(P>0.05),外周血Th2、Th17细胞明显增高(P<0.05),而Th1细胞无明显变化.结论:Th17细胞及其分泌的炎症介质可促进中性粒细胞及嗜酸性粒细胞在气道内聚集,加重哮喘气道炎症,可能与哮喘气道重塑密切相关.     

Th17淋巴细胞在哮喘小鼠气道炎症中的初步研究

目的:初步探索Th17细胞及其分泌的炎症介质在哮喘小鼠气道炎症中的作用机制。方法:20只小鼠随机均分为哮喘组和正常对照组。哮喘组用卵白蛋白(OVA)致敏与激发建立小鼠哮喘模型。正常对照组致敏与激发均以生理盐水代替。HE染色观察小鼠气道及肺组织病理变化;光学显微镜下观察小鼠支气管肺泡灌洗液(BALF)中细胞分类及计数;酶联免疫吸附试验(ELISA)检测小鼠BALF上清中IL-4、IL-5、IL-13、IFN-γ及IL-17的含量,流式细胞技术(FCM)检测小鼠外周血Th1、Th2及Th17淋巴细胞占CD4+T淋巴细胞百分率情况。结果:哮喘组小鼠BALF中细胞总数和中性粒细胞、嗜酸性粒细胞、淋巴细胞百分率均显著高于对照组(P<0.05),BALF上清中IL-4、IL-5、IL-13及IL-17的水平显著增高(P<0.05),而IFN-γ差异无统计学意义(P>0.05),外周血Th2、Th17细胞明显增高(P<0.05),而Th1细胞无明显变化。结论:Th17细胞及其分泌的炎症介质可促进中性粒细胞及嗜酸性粒细胞在气道内聚集,加重哮喘气道炎症,可能与哮喘气道重塑密切相关。     

呼吸道合胞病毒感染后期OVA刺激对小鼠气道炎症及AHR的影响

目的研究呼吸道合胞病毒(RSV)感染后期过敏原刺激对小鼠气道炎症及AHR的影响。方法 6~8周龄雌性Balb/c小鼠随机分为Mock组、RSV组、OVA组及R+O组。检测各组小鼠支气管肺泡灌洗液(BALF)中细胞总数及分类计数;苏木精-伊红(HE)染色观察肺部病理损伤;全身体积描述记法检测小鼠气道反应性;ELISA检测BALF中IL-4、IL-5、CXCL10、CCL5、CCL20的水平。结果 R+O组小鼠BALF中炎症细胞总数、巨噬细胞、中性粒细胞及嗜酸性粒细胞分类计数较其他3组均显著增高(P0.05);R+O组小鼠肺组织病理评分及AHR较其他3组亦显著增高(P0.05);BALF中各处理组IL-4、IL-5较Mock组明显增高(P0.05),但处理组之间无显著性差异;R+O组CXCL10水平较其他3组显著升高(P0.05)。结论 RSV感染后期接受OVA刺激可致CXCL10表达增加,进而促使巨噬细胞、中性粒细胞及嗜酸性粒细胞募集,加重小鼠气道炎症及AHR。     

地塞米松减轻过敏性哮喘模型小鼠气道炎性反应

目的 观察地塞米松(DEX)对过敏性哮喘模型小鼠气道炎性的影响并探讨相关机制。方法 将小鼠随机分为对照组、哮喘组、地塞米松组。哮喘组小鼠于实验第0、7、14天皮下注射卵清蛋白(OVA)/氢氧化铝混合液,第21、22天1%OVA混悬液雾化吸入20 min;地塞米松组在哮喘组基础上于每次雾化吸入前3 h地塞米松(2 mg/kg)腹腔注射。末次雾化结束测定小鼠气道阻力,24 h后收集肺泡灌洗液(BALF),Wright-Giemsa染色后行细胞分类计数;ELISA测定BALF中IL-6、IL-18和IL-1β等浓度;免疫组织化学法(IHC)检测肺组织NLRP3、IL-1β蛋白表达,HE染色观察肺组织病理。结果 与对照组比较,哮喘组小鼠气道高反应性(AHR)明显增高;BALF中嗜酸性粒细胞、中性粒细胞等炎性细胞数量明显增加;IL-5、IL-6、IL-18和IL-1β表达均升高(P<0.01);肺组织NLRP3、IL-1β蛋白表达增强。与哮喘组比较,地塞米松组小鼠AHR显著降低,BALF中炎性细胞总数、嗜酸性粒细胞、中性粒细胞均明显减少(P<0.01),IL-5、IL-6、IL-18...     

磷脂酰肌醇3激酶对哮喘大鼠气道上皮细胞诱导型一氧化氮合酶表达的调控作用

目的： 探讨磷脂酰肌醇3激酶（PI3K）抑制剂wortmannin对哮喘大鼠支气管上皮细胞诱导型一氧化氮合酶（iNOS）表达的影响。方法: 24只成年哮喘大鼠随机分成对照组、哮喘组以及PI3K抑制剂wortmannin干预组。对支气管肺泡灌洗液（BALF）细胞总数及嗜酸性粒细胞进行计数,免疫组织化学检测大鼠支气管上皮细胞iNOS蛋白的表达,RT-PCR检测肺组织iNOS mRNA的表达,分光光度计检测肺组织PI3K活性、iNOS活性及NO含量。结果: 哮喘组大鼠BALF细胞总数计数及嗜酸性粒细胞分类均高于对照组;PI3K抑制剂wortmannin干预组BALF嗜酸性粒细胞计数及分类明显低于哮喘组,差异显著。哮喘组肺组织PI3K活性、iNOS活性及NO含量高于对照组,PI3K抑制剂wortmannin干预组肺组织PI3K活性、iNOS活性及NO含量低于哮喘组。哮喘组大鼠支气管上皮细胞iNOS蛋白及肺组织iNOS mRNA表达较对照组明显增强,但PI3K抑制剂wortmannin组iNOS蛋白及mRNA表达均明显弱于哮喘组。结论: PI3K可调节哮喘大鼠气道iNOS表达,影响哮喘气道炎症反应。     

转录因子GATA-3 mRNA在哮喘模型小鼠体内的表达

目的:探讨转录因子GATA-3 mRNA在哮喘模型小鼠体内的表达。 方法:建立卵白蛋白（OVA）致小鼠哮喘模型，计数支气管肺泡灌洗液（BALF）中炎症细胞总数和分类，评价肺组织炎症细胞浸润；ELISA检测BALF和脾细胞培养上清液中IL-4和IFN-γ浓度；RT-PCR检测脾细胞和肺组织GATA-3 mRNA表达水平。 结果:哮喘组BALF中炎症细胞总数及嗜酸粒细胞百分比明显高于对照组，支气管出现明显炎症细胞浸润、黏液分泌和支气管收缩，BALF和脾细胞培养上清液中IL-4明显高于对照组，肺组织和脾细胞GATA-3 mRNA表达水平均明显高于对照组。 结论:哮喘模型小鼠肺组织和脾细胞GATA-3 mRNA表达增加，可能在促进Th2细胞因子合成和介导气道炎症细胞浸润中具有重要作用。     

地塞米松可增强支气管哮喘小鼠肺组织RECK的表达

目的研究Kazal基序逆向诱导富含半胱氨酸蛋白(RECK)在支气管哮喘小鼠肺组织的表达及地塞米松处理后对其表达的影响。方法 BALB/c小鼠分为哮喘组、对照组、地塞米松组,每组10只。采用鸡卵清蛋白(OVA)腹腔注射致敏及雾化吸入的方法制作哮喘模型,进行支气管肺泡灌洗液(BALF)中嗜酸性粒细胞(EOS)及淋巴细胞(Lym)分类计数;HE染色观察气道炎症细胞浸润及气道重塑情况,实时荧光定量PCR检测RECK mRNA的表达,免疫组织化学法检测RECK蛋白的表达及定位。结果与对照组和地塞米松组相比,哮喘组BALF中细胞总数、EOS明显增多;RECK mRNA在哮喘组表达较对照组、地塞米松组明显降低。免疫组织化学结果显示RECK主要表达于气道上皮细胞、上皮下炎症细胞。哮喘组表达最低,对照组表达最高,地塞米松组表达较哮喘组有所增高。结论 RECK在哮喘小鼠肺组织中表达明显降低,而地塞米松可上调RECK的表达。     

原癌基因表达与哮喘气道炎症

目的:观察原癌基因表达与气道炎症细胞浸润的关系。方法:卵蛋白致敏豚鼠,复制哮喘模型。Dot-blot\,Northern-blot分子杂交及免疫组织化学技术观察哮喘发作前、后豚鼠气道上皮及肺组织中c-fos\,c-myc\,c-jun和c-sis的表达及其与炎症细胞浸润的关系。结果:正常豚鼠气道及肺组织中c-fos\,c-mycmRNA无或很少表达,极少炎症细胞浸润。哮喘发作后,豚鼠气道上皮及肺组织c-fos\,c-mycmRNA表达明显增加。免疫组织化学研究显示Fos、Myc、Jun及Sis在正常豚鼠气道及肺组织低水平表达,哮喘发作后即刻,4种原癌基因表达产物明显增加,气道内有少量淋巴细胞、嗜酸性粒细胞及中性粒细胞浸润;12-24h,上述细胞浸润加重,粘膜下层及粘膜上皮内以中性粒细胞和嗜酸性粒细胞浸润为主。结论:原癌基因表达与气道炎症在哮喘发病中有密切关系。     

Development in in vitro toxicology

There are three principal pressures driving the development of in vitro toxicology: (1) the need for more efficient testing systems to cope with the large number of xenobiotics currently being developed; (2) public pressure to reduce animal experimentation; and (3) a need for a better understanding of the mechanisms of toxicity. Within this, in vitro toxicology is focused on local, systemic, and target-organ toxicity. It is becoming increasingly apparent that a step or decision-tree approach using input of a variety of experimental data (physicochemical properties, biokinetics, cytotoxicity) provides the most efficient system for predicting toxicity. Examples of the use of in vitro toxicity systems for prediction of systemic toxicity and target-organ (liver) toxicity are presented.Originally presented at ECCP 93.     

Seasonal variations in acute toxoplasmosis in pregnant women in Slovenia

Between December 1999 and December 2004, 40 081 pregnant women were examined for toxoplasmosis with Toxo-IgG, Toxo-IgM enzyme immunoassay. Women with positive results were then retested with the Toxo-IgG avidity assay for recent toxoplasmosis. Recent acute toxoplasmosis in pregnant women was found to be significantly more frequent (p < 0.01) during winter than summer. The incidence of acute toxoplasmosis during winter-spring was also significantly more frequent (p < 0.025) than summer-autumn. This phenomenon should be taken into account when formulating preventive measures for toxoplasmosis, especially for pregnant women.     

Age-related changes in polyamines in memory-associated brain structures in rats

Polyamines putrescine, spermidine and spermine are positively charged aliphatic amines and have important roles in maintaining normal cellular function, regulating neurotransmitter receptors and modulating learning and memory. Recent evidence suggests a role of putrescine in hippocampal neurogenesis, that is significantly impaired during aging. The present study measured the polyamine levels in memory-related brain structures in 24- (aged), 12- (middle-aged) and 4- (young) month-old rats using liquid chromatography/mass spectrometry and high performance liquid chromatography. In the hippocampus, the putrescine levels were significantly decreased in the CA1 and dentate gyrus, and increased in the CA2/3 with age. Significant age-related increases in the spermidine levels were found in the CA1 and CA2/3. There was no difference between groups in spermine in any sub-regions examined. In the parahippocampal region, increased putrescine level with age was observed in the entorhinal cortex, and age did not alter the spermidine levels. The spermine level was significantly decreased in the perirhinal cortex and increased in the postrhinal cortex with age. In the prefrontal cortex, there was age-related decrease in putrescine, and the spermidine and spermine levels were significantly increased with age. This study, for the first time, demonstrates age-related region-specific changes in polyamines in memory-associated structures, suggesting that polyamine system dysfunction may potentially contribute to aged-related impairments in hippocampal neurogenesis and learning and memory.     

休克过程中肾上腺髓质素变化的研究进展

Adrenomedullin (AM) is a new peptidergic regulator of vascular function. AM serves as a hormone, which has many biological properties, plays an important role in the many pathophysiological processes, especially shock. This review will highlight the structure, biological properties of AM and the relationship between AM and shock.     

Secular change in menarche in women in Madrid

The age at menarche was estimated by recollection in 1617 women between the ages of 18 and 60 in Madrid and a nearby suburb, Pinto. The population of Pinto is working-class and the Madrid group, taken from residential neighbourhoods , belongs to the upper middle class. In both groups we found a diminution in average age at menarche, from 14.04 to 13.02 years in Madrid and from 14.55 to 13.16 years from about 1935 to about 1965 in Pinto. These changes have been more intense in the group which is less well-off economically, where living conditions have varied much more drastically.     

Pitfalls in TRAP assay in routine detection of malignancy in effusions

Telomerase has been found to be reactivated in a majority of cancers but is inactive in most somatic cells. Our principal goal was to determine the potential use of the telomeric repeat amplification protocol (TRAP) assay as marker for malignancy in cytological effusions. The simple selection criterion was the cytological diagnosis, and routine samples were classified into malignant (58 samples) and nonmalignant (233 samples). Of the malignant samples, 44/58 (76%) were positive by TRAP assay. Of the 14 telomerase-negative cytology-positive samples, RNA integrity was poor in 9, indicating suboptimal sample conservation for molecular analysis. In 3 of the remaining 5 samples with a negative TRAP assay, a high number of malignant cells was observed, and these cells might have been telomerase-negative. Thus, the sensitivity of TRAP assay for the presence of malignant cells was about 76%. In the cytologically nonmalignant effusions, the presence of telomerase activity was observed in 24% (55/233). Of these, 6% were highly suspicious for malignancy, 9% were doubtful, and 9% were cytologically nonmalignant effusions confirmed by a follow-up of 12 mo or more. According to these data, the specificity of the TRAP assay to detect tumor cells in effusions ranged only between 82-91%. Our results indicate that, although the TRAP assay is positive in 6-15% of putative malignant effusions, the relatively high number of TRAP false-negative and false-positive cases renders this test unsuitable for routine diagnostic purposes.