应用原位聚合酶链反应检测淋病患者HSV-Ⅱ感染情况的报告

为了解淋病患者ＨＳＶ Ⅱ的感染情况，应用原位聚合酶链反应（ＩＳＰＣＲ）和ＰＣＲ技术对５８例淋病患者进行ＨＳＶ Ⅱ检测。结果两种方法ＨＳＶ ⅡＤＮＡ阳性检出率均为５３．４５％（３１／５８）。ＩＳＰＣＲ显示，１１例有皮损者中，９例阳性信号存在于上皮细胞核及胞浆，既往有疱疹皮损者６例，阳性信号均存在于细胞核，４１例无明显皮损者，１６例阳性，其中１２例阳性信号在细胞核，４例在胞核及胞浆。结果提示，淋病患者合并有高的ＨＳＶ Ⅱ感染率。应用ＩＳＰＣＲ不仅可以敏感特异地检测ＨＳＶ Ⅱ，而且可以定位，为临床诊断和治疗提供重要依据     

原位聚合酶链反应技术检测结节性红斑皮损中Ⅱ型单纯疱疹...

目的 探讨结节性红斑（ＥＮ）的发病与Ⅱ型单纯疱疹病毒（ＨＳＶ－２）的关系。方法 用原位聚合酶链反应法（ＩＳＰＣＲ）,对３５ＮＥ皮损组织及１４例正常皮肤组织切片中的ＨＳＶ－２ＤＮＡ进行扩增,ＳＰ免疫组化染色,ＡＥＣ显色,结果 ＨＳＶ－２ＤＮＡ在ＥＮ及正常皮肤中的检出率分别为５７．１％（２０／３５）、８．３％（２／１４）,Ｐ〈０．０１,阳性信号主要位于皮损部位,以小血管为著,细胞内的分布以核内型为主,     

gD基因在生殖器疱疹PCR诊断中的研究

依据国外已发表的疱疹病毒（ＨＳＶ）糖蛋白Ｄ基因（ｇＤ基因）的核苷酸保守区序列设计了一对引物，建立了用疱疹病毒ｇＤ基因做疱疹病毒诊断基因的ＰＣＲ方法。对４８例拟诊为疱疹病毒感染的临床标本进行病毒细胞培养和ＰＣＲ检测。结果表明，ＰＣＲ对ＨＳＶ２感染的敏感性为８４％（２０／２４），特异性为９１％（２２／２４）。经标准株及限制性内切酶的酶切证实和序列分析，表明该方法特异、敏感     

男性生殖器疱疹病毒的检测和治疗

目的：研究生殖器疱疹（ＧＨ）的治疗和探讨其病原学。方法：采用聚合酶链反应（ＰＣＲ）检测生殖器皮肤粘膜及精液中的单纯疱疹病毒（ＨＳＶ），并用盐酸万乃洛韦（ＶＣＶ）与阿昔洛韦（ＡＣＶ）对照治疗６０例ＧＨ患者。结果：发现生死器皮肤粘膜ＨＳＶ－Ⅱ阳性率高于精液中。ＶＣＶ疗效与ＡＣＶ相当，但ＶＣＶ起效时间明显快于ＡＣＶ（Ｐ〈０．０１）。结论：ＶＣＶ和ＡＣＶ适合治疗ＧＨ；泌尿生殖器标本ＨＳＶ－Ⅱ的检测，对了解ＧＨ的病原很重要。     

PCR检测164例STD患者5种常见病原体结果

ＰＣＲ检测１６４例ＳＴＤ患者５种常见病原体结果黄干军近年来我们用聚合酶链反应（ＰＣＲ）对１６４例性传播疾病（ＳＴＤ）患者进行了淋球菌（ＧＣ）、沙眼衣原体（ＣＴ）、解脲支原体（ＵＵ）、人类乳头瘤病毒（ＨＰＶ）和单纯疱疹病毒（ＨＳＶ）的ＤＮＡ检测，现报告...     

生殖器疱疹的诊断及感染特征探讨

细胞培养法对８０例临床诊断为典型生殖器疱疹的患者进行了人类单纯疱疹病毒（ＨＳＶ）分离，并结合链霉亲和素过氧化物酶免疫组化法（ＬＳＡＢ）及ＰＣＲ法对其中部分患者进行了ＨＳＶ及其型别检查。三种方法的ＨＳＶ检出率分别为５８．８％、２０％、９０％；培养法作为金标准特异性高，对早期患者检出率达８６．４％；ＰＣＲ法与其他两法比较敏感性高，并利于ＨＳＶ型别的诊断；ＬＳＡＢ法对临床标本的检出率虽很低，却可提高培养法的检出率。ＨＳＶ的感染型别ＨＳＶ２占９１．７％、ＨＳＶ１占８．３％。还讨论了ＰＣＲ引物的优点，并总结了本组患者的临床感染特征。     

人类乳头瘤病毒16全基因在体外培养的正常人角质形成细胞 …

目的 利用含有人类乳头瘤病毒（ＨＰＶ）１６全基因的质粒转染正常人角质形成细胞，观察ＨＰＶ１６ｍＲＮＡ在转染细胞的表达。方法 用ＦｕＧＥＮＥ＾ＴＭ６转染试剂，将携带ＨＰＶ１６全基因的质粒ｐＳＶ２－ｎｅｏ／１６转染体外培养的正常人角质形成细胞，在转染后２４ｈ提取细胞总ＲＮＡ和ＤＮＡ，进行ＲＴ－ＰＣＲ和Ｓｏｕｔｈｅｒｎ印迹分析，结果 ２４ｈ后，ＲＴ－ＰＣＲ成功地扩增出１１０ｂｐ的片段，转染细胞中已出现Ｈ     

原位聚合酶链反应技术检测结节性红斑皮损中Ⅱ型单纯疱疹病毒DNA

目的　探讨结节性红斑（ Ｅ Ｎ） 的发病与Ⅱ型单纯疱疹病毒（ Ｈ Ｓ Ｖ２） 的关系。方法　用原位聚合酶链反应法（ Ｉ Ｓ Ｐ Ｃ Ｒ） ,对３５ 例 Ｅ Ｎ 皮损组织及１４ 例正常皮肤组织切片中的 Ｈ Ｓ Ｖ２ Ｄ Ｎ Ａ 进行扩增, Ｓ Ｐ 免疫组化染色, Ａ Ｅ Ｃ 显色。结果　 Ｈ Ｓ Ｖ２ Ｄ Ｎ Ａ 在 Ｅ Ｎ 及正常皮肤中的检出率分别为５７ ．１ ％ （２０／３５） 、８ ．３ ％ （２／１４） , Ｐ＜ ０ ．０１ ,阳性信号主要位于皮损部位,以小血管为著,细胞内的分布以核内型为主,亦有核浆混合型。结论　 Ｅ Ｎ 的发病与 Ｈ Ｓ Ｖ２ 有关。     

性乱人群中2型单纯疱疹病毒感染的分析

目的 为了解性乱人群不典型皮损中及无皮损的生殖道分泌物中２型单纯疱疹病毒（ＨＳＶ２）的感染情况。方法 在ＨＳＶ２高度保守的基因区设计一对引物，利用聚合酶链反应，对合肥地区６２０例性乱者作ＨＳＶ２－ＤＮＡ检测。结果 ３７５例不典型皮损的恬乱人群中，１４１例检出ＨＳＶ２－ＤＮＡ，阳性率为３７．６％，其中男性占３３．６％，女性占４７．７％。     

PCR检测银屑病患者鳞屑中的乙型肝炎病毒

我们应用聚合酶链反应（ＰＣＲ）法。对３１例寻常性银屑病患者的鳞屑进行乙型肝炎病毒（ＨＢＶ）ＤＮＡ检测。结果报告如下。病例　住院病人皮损分布广泛,同形反应阳性的寻常性进行期银屑病患者３１例,男２３例,女８例,年龄１８～５９岁（平均４３．５岁）,病程３月～５０年。酶联免疫吸附试验和ＰＣＲ同时检测血清ＨＢｓＡｇ及ＨＢＶＤＮＡ。标本采集和处理　钝刀片刮下患者鳞屑约０．５ｇ,入蒸馏水中浸泡冲洗,再用无水乙醇浸洗,室温晾干,然后加０．５ｍｌＴＥＳ缓冲液和加１５ｍｌ蛋白酶Ｋ（２５ｇ／Ｌ）,３７℃保温１２ｈ～１…     

皮肤良恶性肿瘤中p16、p21ras和ki-67蛋白表达研究

目的:探讨细胞周期依赖性激酶抑制蛋白、癌基因蛋白和细胞增殖指数在皮肤基底细胞癌(BBC)、鳞状细胞癌(SCC)和脂溢性角化病(SK)中的表达,并就其结果进行分析。方法:采用DAKO En-Vision免疫组化法检测了18例BCC、10例SCC皮肤恶性肿瘤和17例SK皮肤良性肿瘤组织中p16、p21~(ras)及ki-67蛋白的表达。结果:BCC肿瘤组织中p16、p21~(ras)及ki-67阳性表达率分别为61.1％、77.8％、94％;SCC肿瘤组织中阳性表达率分别90％、50％、90％;SK组织中阳性表达率分别为70.5％、88.2％、94.1％。BCC中p16阳性率明显低于SCC(P<0.01);SCC中p21~(ras)阳性率明显低于SK(P<0.01);三组ki-67蛋白均呈高表达,它们之间无统计学差异。结论:p16、p21、ki-67基因与肿瘤的发生发展及增殖程度密切相关。p16、p21~(ras)基因蛋白的缺失程度或超表达在临床上可作为对良恶性肿瘤预后判断的参考值,它们的表达可能与肿瘤的分化程度有关。     

An evaluation of the efficacy of a single‐session 577 nm pro‐yellow laser treatment in patients with postacne erythema and scarring

Erythema and scarring are among the most common complications of severe inflammatory acne. In this study, we aimed to share our experience with pro‐yellow laser and document the efficacy and safety of this treatment in postacne erythema and scarring. The study included 40 patients, 24 (60%) females, and 16 (40%) males with a mean age of 29.5 ± 8.16 (min. 18 years, max. 57 years). The pro‐yellow laser was applied to all patients as a single session with irradiation of 22 J/cm². Improvement in postacne erythema and scars were evaluated after the treatment. The study included 40 patients, 24 patients (60%) were females and 16 patients (40%) were males with the mean age of 29.5 ± 8.16 (ranged between 18 and 57 years old). A total of 21 patients (52.5%) had good improvement (51%‐75% regression), 10 patients (25%) had excellent improvement (76%‐100% regression), and a moderate improvement (26%‐50%) was detected in 9 patients (22.5%). Also, there were mild improvement (1%‐25%) in 20 patients (76.9%) and a moderate improvement (26%‐50%) in 6 patients (23.1%). We found that pro‐yellow laser is highly effective in the treatment of postacne erythema, while its effectiveness was mild to moderate in atrophic acne scars. Also, it has been observed that the pro‐yellow laser system can be used safely immediately after cessation of systemic isotretinoin treatment.     

UV fingerprints predominate in the PTCH mutation spectra of basal cell carcinomas independent of clinical phenotype

Basal cell carcinoma (BCC) shows a wide interpatient variation in lesion accrual. To determine whether certain tumorigenic fingerprints and potentially predisposing patched (PTCH) tumor suppressor single-nucleotide polymorphisms (SNPs) are distributed differently among sporadic BCC patients, we compared the PTCH mutation spectra in early-onset BCC (first lesion at age < 35 years), regular BCC (first lesion at age > or = 35 years and < 10 lesions), and multiple BCC (> or = 10 lesions). The PTCH gene was mutated in 29 of 60 cases (48%). Most of the PTCH mutations bore the UV fingerprint (i.e., C --> T or tandem CC --> TT transitions at dipyrimidine sites). However, neither the proportion nor the spectra of exonic PTCH mutations differed significantly among the three groups. A large number of SNPs (IVS10+99C/T, IVS11-51G/C, 1665T/C, 1686C/T, IVS15+9G/C, IVS16-80G/C, IVS17+21G/A, and 3944C/T or its combinations) were also detected, but again their incidence did not differ significantly among the groups. Interestingly, expression of the IVS16-80G/C and the IVS17+21G/A genotype did not achieve the Hardy-Weinberg equilibrium in patients with regular and/or early-onset BCC. These data suggest that a (UV-) mutated PTCH gene is important for sporadic BCC formation independent of clinical phenotype and that the IVS16-80G/C and/or IVS17+21G/A SNP site might be important for tumorigenesis in certain BCC patients.     

Human papillomavirus (HPV) detection among human immunodeficiency virus-infected pregnant Thai women: implications for future HPV immunization

BACKGROUND: Human immunodeficiency virus (HIV)-infected women are at increased risk for developing cervical cancer and for infection with human papillomavirus (HPV). Prophylactic vaccines targeting HPV types 16 and 18 are being evaluated for efficacy among young women. GOAL: The goal was to assess the prevalence of HPV among HIV-infected pregnant women in Bangkok and to evaluate the need for prophylactic HPV vaccines studies in this population. STUDY DESIGN: The study population consisted of 256 HIV-infected pregnant women who participated in a mother-to-child HIV transmission trial. Stored cervicovaginal lavage samples were tested for the presence of HPV DNA by polymerase chain reaction with PGMY09/11 primers and reverse line-blot hybridization for determination of anogenital HPV types. RESULTS: HPV prevalence was 35.5% (91/256); high-risk HPV prevalence was 23.4% (60/256). HPV type 16 or 18 was present in 8.2% (21/256). Almost half of all infections were multiple. Furthermore, overall HPV detection was associated with abnormal cervical cytology (P<0.001) and higher HIV-plasma viral load (P=0.007). CONCLUSIONS: Only one-quarter of HIV-infected pregnant women in Bangkok had high-risk HPV types; less than 10% had HPV types 16 or 18. As the HPV prevalence is expected to increase during HIV disease, prophylactic vaccines targeting HPV types 16 and 18 should be studied among HIV-infected women not yet infected with these HPV types and not previously exposed.     

外阴恶性肿瘤中P21~(WAF1/CIP1) P53及HPV6/11 HPV16/18的检测和意义

目的　探讨P2 1WAF1/CIP1,P5 3及HPV6/11,16/18与外阴恶性肿瘤的关系。方法　P2 1WAF1/CIP1,P5 3用免疫组化SP法检测 ;HPV6/11,16/18用原位杂交法 (ISH)检测。结果　P2 1WAF1/CIP1,P5 3在外阴恶性肿瘤组、外阴上皮内瘤变 (VIN)组、正常对照组中的阳性检测率分别为 40 .0 0 % (12 /3 0 )、63 .3 3 % (19/3 0 ) ,5 2 .3 8% (11/2 1) ,47.62 % (10 /2 1) ,0 (0 /10 )和 0 (0 /10 ) ;外阴病变各组P2 1WAF1/CIP1,P5 3与正常组比较差异均有显著性 (P均 <0 .0 5 ) ;HPV6/11、16/18在外阴恶性肿瘤组、VIN组中的阳性检测率分别为 60 .0 0 % (18/3 0 )和 3 3 .3 3 % (10 /3 0 ) ,42 .86% (9/2 1)和 2 8.5 7% (6/2 1) ,正常对照组没有检测出 ;HPV 6/11阳性率外阴恶性肿瘤组、VIN组与正常组比较差异均有显著性 (P均 <0 .0 5 ) ;外阴恶性肿瘤组HPV16/18阳性率与正常组比较差异有显著性 (P <0 .0 5 )。结论　P2 1WAF1/CIP1P5 3及HPV感染在外阴恶性肿瘤的发生发展中可能起一定的作用     

P16、P21WAF1/CIP1、PCNA和cyclin E在脂溢性角化病中的表达及其意义

目的：研究脂溢性角化病皮损组织中P16、P21^WAF1/CIP1、PCNA、cyclinE四种细胞周期素相关因子的表达及意义。方法：应用免疫组化PV法（改进的SP法），对50例脂溢性角化病病人和10例正常人皮肤进行P16、P21^WAF1/CIP1、PCNA、cyclinE四种细胞周期素相关因子的测定。结果：脂溢性角化病患者中这四种因子的表达阳性率分别为：78％、68％、70％、46％；而正常对照组的阳性率仅为：40％、30％、50％、10％。结论：P16、P21^WAF1/CIP1、PCNA、cyclinE四种细胞周期素相关因子在脂溢性角化病的发病中可能起重要作用。     

p16INK4a expression is frequently decreased and associated with 9p21 loss of heterozygosity in sporadic melanoma

The product of the p16/INK4a/CDKN2/MTS1 tumor-suppressor gene acts as a negative cell cycle regulator by inhibiting G₁ cyclin-dependent kinases that phosphorylate the retinoblastoma protein. p16 is inactivated in a wide range of human malignancies, including familial melanoma. However, its expression and function in sporadic melanoma has not been extensively investigated. We studied p16 expression in 62 archival melanomas and 30 archival nevi and lentigines by immunohistochemistry. Eighteen of 26 (69%) superficial spreading melanomas, 17 of 28 (61%) nodular melanomas, all of three lentigo maligna melanomas, and all of five melanoma metastases were found to harbor less than 10% p16 -positive tumor cells. In contrast, only six of 24 (25%) nevi had less than 10% positive cells. No correlation between tumor thickness and loss of p16 expression was found. Using DNA from micro-dissected tumor and matched normal tissues, five of seven (71%) p16 -negative melanoma cases had 9p21 loss of heterozygosity (LOH), and one of these 9p21 LOH cases had promoter region hypermethylation of the remaining p16 allele. These data demonstrate that partial or complete loss of p16 expression is prevalent in sporadic melanoma and is frequently associated with 9p21 LOH.     

Molecular alterations at chromosome 9p21 in melanocytic naevi and melanoma

BACKGROUND: The chromosome 9p21 and its CDKN locus, with the p16 tumour suppressor gene (CDKN2A), are recognized as the genomic regions involved in the pathogenesis of melanoma. OBJECTIVES: To elucidate further the role of such regions during the different phases of melanocytic tumorigenesis. METHODS: Tissue sections from naevi, primary and metastatic melanomas were investigated by fluorescence in situ hybridization for allelic loss at the 9p21 chromosome and by immunochemistry for p16CDKN2A expression. RESULTS: Dysplastic naevi and primary or secondary melanomas were found to carry hemizygous deletions within the entire 9p21 region at similar frequencies (varying from 55% to 62%). Allelic deletion spanning the CDKN locus was observed at significantly increased rates moving from early (7%) to advanced (28%) primary melanomas and to secondary melanoma lesions (37%) (P=0.018). Also, inactivation of the p16 gene (CDKN2A) was absent in naevi and present at steadily increasing rates moving from primary melanomas (7% early lesions to 17% advanced lesions) to melanoma metastases (62%) (P=0.004). CONCLUSIONS: Our findings indicate that, in a model of sequential accumulation of genetic alterations, 9p21 deletions may play a role in melanocytic transformation and tumour initiation whereas rearrangements at the CDKN locus, and p16 gene (CDKN2A) inactivation may contribute to tumour progression.     

Analysis of gene probes and gene amplification techniques for diagnosis and monitoring of treatment in childhood leprosy

Nucleic acid sequences of Mycobacterium leprae were detected using gene probes hybridizing with targeting ribosomal RNA (16S rRNA), ribosomal DNA (16S rDNA) and gene amplification techniques (PCR) in skin lesion of paediatric leprosy patients and the effect of treatment on the by these methods. Eighty paediatric leprosy patients were included in the study. Most cases (79%) were between 9 and 16 years of age. Cases were divided into three groups according to treatment status, viz. untreated (30), undergoing treatment (30), and at the end of treatment (20). Clinical and slit smear examination for acid fast bacilli (AFB) was performed and nucleic acids were extracted and fractionated from skin biopsies. M. leprae specific 16S rRNA and 16S rDNA was detected by hybridization with gene probes whereas the 36 kDa gene sequence was detected by a gene amplification assay (PCR). The cases were classified as paucibacillary (PB) and multibacillary (MB) by the standard criteria of WHO (1988). Positivity of 16S rRNA in PB cases decreased from 60% in untreated to 10.5% after 4-8 months of treatment whereas for 16S r DNA, it decreased from 50% to 21%, for PCR from 70% to 36.8% for the same specimen, and all became negative at 1 year. Similar trends were seen in MB cases where positivity in smear positive untreated cases decreased from 100% to 56.2% with 16S rRNA and 42.8% with 16S rDNA and PCR, respectively, after 9-12 months of treatment and all became negative at 2 years, except one case which remained positive with PCR. Similar results were observed in smear negative MB cases, 100% positivity detected by 16S r RNA and PCR, 75% detected by 16S rDNA decreased to zero after 9-12 months of therapy. This study suggests the potential usefulness of gene probes targeting 16S rRNA and 16S rDNA and PCR as supportive molecular tools for diagnosis of smear negative evolving MB disease and also monitoring the response to treatment, these observations however, needs to be validated in prospective follow up studies.     

p16INK4a and p14ARF tumor suppressor genes are commonly inactivated in cutaneous squamous cell carcinoma

The p16(INK4a) and p14(ARF) tumor suppressor genes (TSGs) are encoded within the CDKN2A locus on chromosome 9p21 and function as cell cycle regulatory proteins in the p53 and RB pathways. Inactivation of these genes by genetic and epigenetic changes has been described in some human cancers, but their importance in cutaneous squamous cell carcinoma (SCC) has not been established. Our detailed examination of 40 cutaneous SCC revealed loss of heterozygosity of 9p21 markers in 32.5% of cases. Mutational analysis confirmed five point mutations in four of 40 SCCs. These mutations changed the amino acid sequence of p16(INK4a) in four tumors and p14(ARF) in three tumors. Promoter methylation of p16(INK4a) and p14(ARF) was detected in 13 of 36 (36%) and 16 of 38 (42%) cases, respectively. Absent protein expression was confirmed by immunohistochemistry in 13 of 16 (82%) of the tumors with biallelic inactivating events. Overall, the frequency of 9p21 alterations was 76% and for both p16(INK4a) and p14(ARF), promoter methylation is the commonest mechanism of gene inactivation. Alterations at this locus were significantly more common in tumors from immunocompetent compared with immunosuppressed individuals. These data confirm the importance of inactivation of p16(INK4a) and p14(ARF) TSGs in the pathogenesis of cutaneous SCCs.