Differential in vivo cytokine mRNA expression in lesional skin of intrinsic vs. extrinsic atopic dermatitis patients using semiquantitative RT-PCR

BACKGROUND: A small subgroup of atopic dermatitis (AD) patients with normal serum IgE levels and without specific IgE sensitization has been termed 'intrinsic type of AD' (ADi) as a counterpart to the term 'extrinsic type of AD' (ADe). However, there are neither molecular markers nor clinically diagnostic tools for distinguishing between ADi and ADe. OBJECTIVE: The present studies were undertaken to clarify the pathogenesis and in vivo cytokine micromilieu of ADi patients in comparison with ADe patients. METHODS: We used semiquantitative RT-PCR to investigate the expression of various cytokines and assessed the tissue eosinophil counts in skin biopsies from both types of AD patients. RESULTS: Although there was no significant difference of cellular infiltrates in the lesional skin between ADe and ADi patients, ADe had significantly increased tissue eosinophilia than ADi. Based on our RT-PCR, the expression patterns of cytokines could be categorized into four groups. The first group includes IL-5, IL-13, and IL-1beta, whose levels of mRNA expression were higher in both types of AD patients than non-atopic (NA) subjects, while ADe patients had even higher levels than ADi patients. The second group includes interferon-gamma (IFN-gamma), IL-12, granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-4, and IL-10, whose levels of mRNA expression were elevated in both types of AD patients without differences between ADe and ADi patients. The third group includes tumour necrosis factor-alpha (TNF-alpha), whose mRNA expression was more decreased in both types of AD patients than NA, and the fourth group includes IL-6 and transforming growth factor-beta (TGF-beta), which did not show any differences among the three groups. CONCLUSION: These current data demonstrate that the expressions of cytokines IL-5, IL-13, and IL-1beta mRNA and the number of dermal infiltrating eosinophils are increased in ADe patients compared with ADi patients.     

发酵支原体的套式PCR检测研究及其临床应用

发酵支原体（Ｍｆ）已从孕妇的胎盘和羊水中分离检测到，这一发现揭示Ｍｆ存在宫内感染的可能性。Ｍｆ的人工培养更难于其它致病性支原体。为此，我们以Ｍｆ的１６ＳｒＲＮＡ基因为靶基因建立了Ｍｆ－ＤＮＡ的套式检测技术。结果表明Ｍｆ－ＤＮＡ的ＰＣＲ法检测是一种灵敏、特异、简便和有价值的手段。     

用多重及套式PCR同时检测血清中的HBV和HCV方法的建立

首次应用多重及套式ＰＣＲ技术同时检测人血清中的ＨＢＶ和ＨＣＶ。将抽提的ＨＢＶＤＮＡ和（或）ＨＣＶＲＮＡ在含ＡＭＶ逆转录酶、ＴａｑＤＮＡ多聚酶以及ＨＢＶ和ＨＣＶ外套引物的ＰＣＲ缓冲引物的ＰＣＲ缓冲液中进行逆转录后连续进行ＰＣＲ扩增，以第一轮扩增产物为模板，在含ＨＢＶ和ＨＣＶ内套引物的ＰＣＲ反应体系中进行第二轮扩增，产物经电泳后，以ＤＮＡ分子量标志物或已知片段做参考，出现５２３ｂｐ和（或）２６０ｂｐ产     

人前列腺干细胞抗原在前列腺组织中的表达

目的：克隆人前列腺干细胞抗原(PSCA)基因，并利用半定量逆转录聚合酶链反应(RT—PCR)对其组织表达谱进行分析。方法：从人前列腺癌组织提取总RNA，利用RT—PCR技术扩增出人PSCA基因编码区序列，并通过半定量RT—PCR方法检测6例人新鲜正常前列腺组织、16例前列腺增生组织及11例前列腺肿瘤组织中PSCA基因表达水平。结果：序列测定表明，克隆获得的369bp片段与文献报道的人PSCA基因编码区cDNA序列一致，PSCA为前列腺组织特异表达，在前列腺癌组织中的表达明显高于前列腺增生及前列腺正常组织。结论：PSCA是一种新的前列腺肿瘤标记物。     

胃癌患者外周血细胞角蛋白20mRNA的检测及临床意义

目的　采用逆转录聚合酶链反应检测进展期胃癌患者外周血中细胞角蛋白 2 0 (cytokeratin 2 0 ,CK2 0 )mRNA的表达 ,以检测胃癌外周血循环中微转移的情况 ,并比较CK2 0mRNA的阳性检出率与胃癌患者的性别、年龄、肿瘤部位和胃癌组织的分化程度的关系。方法　 6 0例胃癌患者和 2 0例慢性胃炎患者分别作为试验组和对照组。采用密度梯度离心法分离获取全部病例外周静脉血中的单个核细胞 ,随后应用逆转录聚合酶链反应的方法检测其中的CK2 0mRNA表达情况。结果　试验组中有 32例患者外周血循环中有阳性表达 ,阳性率为 5 3.3% ;而在对照组患者外周血中无 1例标本检出细胞角蛋白 2 0mRNA ;两组间的阳性率差异具有显著性 (P <0 .0 0 5 ) ;癌细胞的检出率与胃癌组织的分化程度呈正相关 (P <0 .0 5 ) ,而与胃癌患者的性别、年龄和肿瘤部位等因素无明显相关 (P >0 .0 5 )。结论　CK2 0mRNA可作为标志物用于检测胃癌的血行转移 ;CK2 0mRNA的阳性检出率与胃癌组织的分化程度具有相关性。     

逆转录PCR检测多源耐药基因在儿童白血病中的表达

多源耐药基因的过度表达是导致血病化学药物治疗失败的主要原因之一。本研究建立了应用逆转录ＰＣＲ技术对多源耐药基因的定量检测，其方法灵敏，快速，具有极大的临床应用价值。     

Mutation and expression analysis of LZTS1 in ovarian cancer

LZTS1 has been shown to have tumour suppressor activities against prostate and breast cancer and is located within a region of frequent loss of heterozygosity (LOH) at 8p22 in ovarian cancer. We have analysed the expression of LZTS1 in ovarian cancer and found no evidence of loss of expression relative to normal ovarian surface epithelial cells. We have also analysed the coding region of the LZTS1 gene in 87 primary ovarian adenocarcinomas by DHPLC and detected a single silent somatic mutation. These data indicate that LZTS1 is not the target of LOH at 8p22 in ovarian cancer.