喉乳头状瘤相关HPV—DNA序列检测及临床观察

对１６例喉和喉气管乳头状瘤的组织切片复习，其中１３例可见凹空细胞，应用同位素＾３２Ｐ标记ＨＰＶ６ｂ，１１，１６，以Ｓｏｌｔ ｂｌｏｔ杂交技术，对瘤组织ＤＮＡ进行检测。结果阳性率为ＨＰＶ６ｂ为８７．５％，ＨＰＶ１１为９３．７％、ＨＰＶ１６为８１．２％。对１２例阳性的瘤组织ＤＮＡ做Ｓｏｕｔｈｅｒｎ ｂｌｏｔ杂交，结果３例为典型的ＨＰＶ１１型，５例弱阳性反应，为未定型。全部病例采用间接喉镜或支撑喉显微镜     

喉乳头状瘤患儿血清人乳头状瘤病毒11b病毒样颗粒抗体检测及意义

目的　制备人乳头状瘤病毒 (humanpapillomavirus,HPV) 11b型病毒样颗粒 (viruslikeparticles ,VLP)检测儿童喉乳头状瘤 (juvenilelarynxpapilloma ,JLP)患者血清抗HPV11bVLP抗体 ,探讨与疾病发生、发展的关系。方法　采用重组杆状病毒昆虫细胞系统制备HPV11bVLP。通过酶联免疫吸附试验 (enzymelinkedimmunosorbentassay,ELISA)方法检测 4 6份复发性儿童喉乳头状瘤患儿不同发病时期的血清和 2 0份健康对照组的血清抗体。将标本分 5组 :A组 :健康对照 (2 0份 ) ;B组 :发病 1年(15份 ) ;C组 :发病 2年 (15份 ) ;D组 :未复发 1年 (8份 ) ;E组 :未复发 2年 (8份 )。结果A、B、C、D、E各组血清抗体吸光度 (A)值平均 ( x±s)分别为 0 0 73± 0 0 35 ;0 12 0± 0 0 4 9;0 137± 0 0 5 7;0 5 18±0 12 2 ;0 5 5 7± 0 14 4 ,B、C、D、E各组与A组比较差异均有显著性意义 (P <0 0 5 )。未复发组 (D +F组 )和复发组 (B +C组 )间A均值比较差异有高度显著性意义 (t=14 90 ,P <0 0 0 0 1)。两组年龄比较差异有显著性意义 (6 2 5 0± 1 6 93和 3 2 2 7± 1 4 5 3;t=6 76 7,P <0 0 0 0 1)。结论　HPV感染的JLP患者血清能产生抗HPV抗体 ,疾病发展和预后与血清抗病毒中和抗体水平密切相关。HPV11bVLP及血     

儿童喉乳头状瘤细胞的原代培养及生物学特性研究

目的 探讨人乳头状瘤病毒（human papilloma virus,HPV)感染阳性患儿喉乳头状瘤细胞在体外培养的生物学特性。方法 2000年3月－2001年4月采用组织块培养法培养10例12例次喉乳头状瘤患儿手术的标本－HPV感染阳性患儿喉乳头状瘤细胞，观察其生长情况，计数法绘制细胞生长曲线，运用共同引物聚合酶链反应（polymerase chain reaction,PCR)法及核酸分子斑点杂交方法对喉乳头状瘤细胞在培养前后的HPV DNA进行检测。结果 HPV感染阳性的喉乳头状瘤细胞在体外生长时间可长达6周，培养前后细胞内均含有HPV的DNA。细胞生长分为3期，延缓期、生长期及停滞期。培养前1－4d细胞从组织块中大量游出，5－7d为延缓期，此期间细胞渐渐开始贴壁，8－18d为生长期，细胞数目迅速增多，生长速度很快，继而进入停滞期，细胞数目增长缓慢，细胞空泡化明显，渐渐走向死亡。结论 HPV感染阳性的喉乳头状瘤细胞在体外生长情况良好，但要建立HPV感染阳性的喉乳头状瘤细胞的动物模型尚需进一步研究。     

人乳头状瘤病毒与喉癌,喉内翻性乳头状瘤发病的关系

应用聚合酶链反应（ＰＣＲ）法检测人乳头状瘤病毒（ＨＰＶ）抗原，并在透射电镜下查找ＨＰＶ颗粒，在１３例喉内翻性乳头状瘤的标本中，有７例找到ＨＰＶ颗粒，８例ＨＰＶ抗原阳性；其中３例喉内翻性乳头瘤恶变的标本中２例找到ＨＰＶ颗粒且抗原阳性，证明喉内翻性乳头状瘤发病及其癌变与ＨＰＶ密切相关。     

成人喉乳头状瘤

１９７５年７月－１９９１年１２月收治住院成人喉乳头状瘤４２例，单发３５例，多发７例。平均年龄５２．４岁，均获随访，４２例中１１例恶变（２６．２％）１９例伴上皮非典型增生，占４５．２％。成人单发喉乳头状瘤手术治疗应留有一定的安全缘，而多发性的应用电烧，术后辅助干扰素治疗取得了满意的效果。本文并对其病因，恶变、恶变与皮皮非典型增生的关系以及治疗复发问题的讨论。     

儿童喉乳头状瘤与人乳头状瘤病毒的相关性及其免疫综合治疗

对2002～2005年收治的60例儿童喉乳头状瘤（JLP）的病变组织进行人乳头状瘤病毒（HPV）分型及体液、细胞免疫检测，并对其临床资料进行回顾性分析，以探讨JLP患儿HPV感染类型、途径及细胞、体液免疫水平，提高对其临床特点的认识，探讨有效的治疗方法。     

小儿喉乳头状瘤31例临床小结

报告３１例小儿喉乳头状瘤的病例资料。本病特点：病情发展快，病变范围广，声门下发生率高，气管切开率高和绝少癌变。认为似不应以手术次数作为分型依据；术前要对肿瘤精确定位和明确范围；术中应用激光消除残存瘤组织和止血，可获较好疗效；术后密切随访观察，及时发现和处理复发病例。     

原位PCR技术检测喉乳头状瘤病毒DNA的体会

聚合酶链反应(PCR)技术的诞生,为分子生物学的发展开僻了新的途径。新近,我们应用原位PCR对喉部病变的陈旧蜡块标本进行了乳头状瘤病毒(HPV)DNA的检测,并就实验的一些体会作一总结。1　材料和方法1.1　材料选择21例经HPV6、11、18型特异引物PCR检测为阳性的喉良、恶性肿瘤蜡块标本(由301医院病理科提供)。每个标本作3个连续切片,片厚4μm,分别进行HE、ISH、PCR-ISH检测。阳性对照标本为HPV6、11、18阳性的尖锐湿疣和宫颈癌;阴性对照标本为HPV阴性的喉癌及乳头状瘤。HPV6、11、16、18型E6引物,合成于赛百盛公司;TaqDNA聚合…     

HPV11对小儿喉乳头状瘤预后的影响

目的 :研究人乳头状瘤病毒 (HPV)型别对小儿喉乳头状瘤 (JLP)预后的影响。方法 :应用聚合酶链反应结合斑点杂交技术对 2 5例JLP的石蜡标本进行HPV定型分析 ,并统计HPV11、HPV6 感染组的气管切开率和术后复发率。结果 :HPV总检出率为 96.0 % ,其中HPV11为 5 6.0 % ,HPV6 为 4 0 .0 % ,HPV16、18、33无一例阳性。HPV11感染组的气管切开率为 71.4 % ,术后复发率为 85 .7% ;HPV6 感染组的气管切开率为 3 0 .0 % ,术后复发率为4 0 .0 %。两组分别比较 ,其差异均有显著性意义 (P <0 .0 5 )。结论 :HPV6、11与JLP发生密切相关 ,HPV11感染与JLP的喉梗阻和术后复发率相关 ,HPV11感染可作为JLP预后评判的重要依据。     

喉乳头状瘤HPV_(16/18)感染与p53蛋白表达的相关性

目的 :了解喉乳头状瘤组织内 HPV1 6 / 1 8的感染与抑癌基因 p5 3变异的关系 ,以及 HPV感染在喉乳头状瘤发病中的作用。方法 :采用 PCR和免疫组化技术 ,检测 35例喉乳头状瘤组织中 HPV1 6 / 1 8DNA及 p5 3蛋白的表达。结果 :2 4例组织中检出 HPV1 6 / 1 8DNA(6 8.6 % ) ;19例 p5 3蛋白呈过度表达 (5 4.3% ) ;在 12例中同时检出HPV1 6 / 1 8DNA和 p5 3蛋白过度表达 (34 .3% )。结论 :提示 HPV1 6 / 1 8感染和 p5 3变异与喉乳头状瘤的发生明确相关 ,其内在分子机理及临床意义有待进一步阐明。     

Effect of human papilloma virus expression on clinical course of laryngeal papilloma

Conclusion. Our observations suggest that human papilloma virus (HPV) 6/11 is the main causative agent of laryngeal papilloma and that detection of active HPV DNA expression may be helpful in identifying patients with aggressive recurrent laryngeal papilloma. Objectives. HPV is assumed to be the main causative agent of this disease. We investigated the expression of the entire genotype of HPV in cases of laryngeal papilloma and correlated their expression with the clinical course of the disease. Subjects and methods. Seventy cases of laryngeal papilloma were evaluated for the presence of the HPV genome by in situ hybridization (ISH) using wide-spectrum HPV DNA probe. Specific types of HPV infection were determined by DNA ISH using type-specific HPV DNA probes (HPV 6, 11, 16, 18, 31, 33). Separate analyses were conducted comparing viral types, frequency of recurrences and duration of disease-free periods. Results. We detected HPV DNA in 40 of the 70 laryngeal papilloma cases (57%). In particular, HPV DNA was detected in 75% of the juvenile types. There were significant associations between HPV and laryngeal papilloma (p<0.01). Among the HPV-positive cases, major specific types were HPV 6/11 (97%). Significant associations were also noted between viral expression and clinical course.     

鼻、鼻窦恶性肿瘤中EB病毒和人乳头状瘤病毒的检测

目的：探讨ＥＢ病毒和人乳头状瘤病毒（ＨＰＶ）与鼻、鼻窦恶性肿瘤的关系。方法：用聚合酶链反应（ＰＣＲ）方法检测,３２例鼻、鼻窦恶性肿瘤组织蜡块的ＥＢ病毒和ＨＰＶ（ＨＰＶ６、１１、１６、１８、３３型）基因,分析其与病理分型及ＴＮＭ分期的关系。结果：３２例中检出ＥＢ病毒１２例（３７．５％）,ＨＰＶ２１例（６５．６％）,其中混合感染６例,５例均未检出。与之对照的１０例鼻息肉中未检出ＥＢＶ和ＨＰＶ。ＴＮＭ分     

High incidence of malignant transformation of laryngeal papilloma in Taiwan

OBJECTIVES: Papillomas of the larynx include solitary laryngeal papilloma and recurrent respiratory papillomatosis. This study investigated the incidence of malignant transformation and assessed possible risk factors for laryngeal papillomas. STUDY DESIGN: A prospective, longitudinal study. METHODS: Twenty-six consecutive laryngeal papilloma patients were prospectively studied for 5 or more years, and each patient was periodically examined at 3 to 6 month intervals. A detailed epidemiologic questionnaire was administered at the initial visit. After enrollment, tissue obtained during each laryngeal surgery was examined by polymerase chain reaction assay for human papilloma virus (HPV) and typing. RESULTS: During 237 person-years of follow-up, six new, pathologically confirmed cases of laryngeal carcinoma were ascertained (incidence 2.5/100 person-years), and all were associated with HPV-6 or HPV-11. Malignant transformation revealed no correlation with the following: age less than 3 years at diagnosis, sex, history of tobacco use, history of alcohol consumption, family history of laryngeal cancer, or type of laryngeal papilloma. Laryngeal papilloma without demonstrable HPV DNA was the only significant risk factor for malignant transformation (P < .05). The cumulative risk of malignant transformation in subjects without demonstrable HPV DNA was significantly higher than that in HPV-positive patients (relative risk, 8.0; 95% confidence interval, 1.1-60.3; P = .05). CONCLUSIONS: A relatively high incidence of malignant transformation of laryngeal papilloma was noted in Taiwanese patients. Patients without demonstrable HPV DNA require more frequent follow-up and may benefit from anti-HPV vaccinations.     

Absence of high-risk human papilloma virus in p16 positive inverted sinonasal papilloma

ObjectivesSinonasal inverted papilloma (SIP) is a relatively rare disease, and its etiology is not understood. It is characterized by locally aggressive growth and a strong tendency to recur despite its benign histology.AimsThe aim of this study was to identify the presence of human papilloma virus (HPV) and its surrogate marker p16 in SIP tissue samples from a regional cohort.Material and methodsSubjects were identified from our regional center cohort of 88 SIP patients treated between 1984–2014. From these subjects, 54 were included in this study. Of these, 53 biopsies were analyzed with PCR, and 54 samples were immunohistochemically stained for p16. DNA was extracted from histopathologically verified SIP. Genotype screening for 13 high risk-, 5 oncogenic and 6 low risk HPV types was performed using the PapilloCheck® HPV-screening test.ResultsHPV analysis was successful for 38 of 53 samples. Of the 38 successfully analyzed samples, only 2 samples were positive for HPV 11. Notably, p16 was present in the epithelia in all samples, and in the papilloma lesions in 37 samples.ConclusionSince only 2 out of 38 SIPs were positive for HPV (type 11), and at the same time p16 was positive in epithelia in all samples and in 37 of 38 papilloma lesions of the samples, it is concluded that p16 cannot be used as a surrogate marker for high-risk HPV-infection in SIP. We are currently planning a prospective, multicenter study in order to increase the study power and in order to be able to better evaluate the clinical implications of HPV-and p16 in SIP.     

复发性喉乳头状瘤中潜在生物学标志物的筛选及生物信息学分析

目的通过生物信息学技术筛选复发性喉乳头状瘤中表达的差异基因,为复发性喉乳头状瘤的治疗提供新的思路和靶点。方法从GEO数据库下载复发性喉乳头状瘤的芯片数据集GSE10935,使用在线分析工具GEO2R筛选复发性喉乳头状瘤及其相邻正常喉黏膜组织间的差异表达基因,使用网络分析工具Metascape进行差异基因的GO分析及KEGG通路富集,通过STRING在线软件进行差异表达基因的蛋白-蛋白相互作用分析,并应用Cytoscape对蛋白-蛋白相互作用网络进行模块分析。最后,使用转录因子富集分析数据库ChEA3预测复发性喉乳头状瘤组织中差异表达基因的关键转录因子。结果在10对复发性喉乳头状瘤组织及其相邻的正常喉黏膜组织间共筛选出53个差异表达基因(DEGs)(|logFC|>1,adj P<0.05),其中上调的DEGs共30个,下调的DEGs共23个。GO分析和KEGG通路富集显示,这些差异表达的基因主要富集在多个代谢、免疫调节及PPAR信号通路等。通过蛋白-蛋白相互作用网络的构建及模块分析,筛选出SLC27A2、SCD、ECI2及FADS2四个枢纽差异表达基因。进一步的分析发现,TP63为复发性喉乳头状瘤中最为重要的转录因子。结论为进一步深入理解复发性喉乳头状瘤的生物学机制、探索其治疗的有效方案提供了新思路。     

中西医结合治疗小儿喉尖锐湿疣,喉乳头状瘤疗效分析

1991年1月～1995年4月间对8例喉尖锐湿疣、6例喉乳头状瘤患儿施行手术、激光、干扰素等综合治疗,除1例治愈外,其余皆复发,治愈率7．1％。1995年6月开始,对上述复发患儿及另3例喉尖锐湿疣和喉乳头状瘤患儿进行中西医结合治疗,随访2～25年无复发,治愈率86．67％。治疗前后进行嗓音电脑分析,各项指标P值均小于0．01。因此认为：中西医结合治疗具有手术范围小、种植机会少、复发率低、手术出数少、费用低、音质恢复良好等优点,并指出治疗及方剂中的要点和注意事项。     

Detection of HHV-8 and HPV in laryngeal carcinoma

OBJECTIVE: In this study we aimed to investigate the presence of human herpesvirus 8 (HHV-8) and human papillomavirus (HPV) in laryngeal carcinoma. MATERIALS AND METHODS: Fifty patients operated on because of laryngeal carcinoma were included in the study. Forty-seven had squamous cell carcinoma (SCC) whereas three had verrucous carcinoma. Fresh tumoral tissues, or tumoral tissues obtained from archival paraffin-embedded blocks, were examined. HHV-8 DNA and HPV DNA were detected using polymerase chain reaction (PCR) and viral genotypes of HPV were determined via the hybrid capture method. The presence of HHV-8 DNA and HPV DNA were also investigated in normal appearing laryngeal tissue collected from 50 cadavers at autopsy. RESULTS: HPV DNA was detected in seven patients (7/50; 14%) (5 out of 47 patients with SCC (5/47; 10.6%) and two out of three patients with verrucous carcinoma). HHV-8 DNA was detected in five patients and they all had SCC (5/47; 10.6%). One case had both HHV-8 and HPV DNA. None of the control samples from cadavers harbored HHV-8 DNA, or HPV DNA. There was a statistically significant correlation between HHV-8 DNA and HPV DNA positivity and laryngeal SCC (Fisher exact test; p=0.023 for each). No statistically significant correlations were found between the presence of HHV-8 and/or HPV and age, gender, tumor stage, differentiation, the site of the tumor, smoking and alcohol use. CONCLUSIONS: The findings of the present study suggest that beside HPV, HHV-8 might have a role in laryngeal carcinogenesis. Further investigations are necessary to clarify the exact role of these viruses in laryngeal carcinoma.