原位杂交在成年和幼年型喉乳头状瘤HPV病因学的对比研究

探讨成年和幼年型喉乳头状瘤ＨＰＶ感染发病差异及其影响因素。方法：用地高辛配基（Ｄｉｇｏｘｉｇｅｎｉｎ）标记ＨＰＶ６和ＨＰＶ１１型作探针，原位核酸杂交方法在２９例成年型喉乳头状瘤（ＡＬＰ）和２１例幼年型喉乳头状瘤（ＪＬＰ）石蜡包埋标本检测ＨＰＶ同源序列。结果：ＡＬＰＨＰＶ６和ＨＰＶ１１阳性率分别为４１４％（１２／２９）和４８３％（１４／２９）；ＪＬＰＨＰＶ６及ＨＰＶ１１阳性率均为７６２％（１６／２１）。ｘ２统计示：两型喉乳头状瘤ＨＰＶ６及ＨＰＶ１１阳性率明显不同（ＨＰＶ６ｘ２＝５９９，ＨＰＶ１１ｘ２＝３９５，Ｐ均小于００５）。结论：１）ＡＬＰ和ＪＬＰＨＰＶ感染发病存在差异。２）ＡＬＰ除了ＨＰＶ感染外，其促发因素不可忽视，ＪＬＰ更倾向于依赖ＨＰＶ感染而发病。     

上呼吸道乳头状瘤HPVDNA的检测

应用多重引物ＰＣＲ技术对４７例复发性呼吸道乳头状瘤病（ＲＲＰ）和９例声带息肉石蜡包埋组织中ＨＰＶ６／１１、１６、１８ＤＮＡ进行了检测。结果发现：①ＲＲＰ组织中ＨＰＶ６／１１ＤＮＡ的阳性率为６８．１％（３２／４７），其中喉乳头状瘤、口咽乳头状瘤、鼻腔和鼻前庭乳头状瘤组织中ＨＰＶ６／１１ＤＮＡ的阳性率分别为７０．４％（１９／２７）、６６．７％（４／６）和６４．３％（９／１４）；②所有ＲＲＰ标本中均未发现特异性的ＨＰＶ１６和ＨＰＶ１８ＤＮＡ；③９例声带息肉标本中四种型号的ＨＰＶＤＮＡ全部阴性。实验结果表明，ＲＲＰ的发病与ＨＰＶ６／１１感染密切相关。多重引物ＰＣＲ检测ＨＰＶＤＮＡ具有敏感、特异、快速、简便等优点，适合于临床广泛开展和进行流行病学调查。     

PCR和PCR产物斑占杂交技术检测成人咽喉病变中人乳头状？…

目的：研究成人咽喉部良、恶性病变与人乳头状瘤病毒（ＨＰＶ）感染的关系。方法：应用聚合酶链反应（ＰＣＲ）和斑点杂交技术,对５５例咽喉不同病变的新鲜组织标本进行ＨＰＶ６,１１,１６,１８,３３共５型ＨＰＶ－ＤＮＡ感染的检测。结果：在咽乳状瘤组ＨＰＶ感染率为６０％（６／１０）,喉乳头状瘤组为７０％（７／１０）,喉鳞状上皮非典型增生组为２０％（１／５）,声带息肉组为２０％（１／５）,喉癌组为２０％（１／５     

国人喉癌与人类乳头状瘤病毒相关性的研究

本项研究对１２４例喉不同病变的新鲜组织标本，采用共同引物和多重引物ＰＣＲ的方法，进行ＨＰＶＤＮＡ检测。用共同引物ＰＣＲ检测ＨＰＶ＿（６，１１，１６，１８，３１，３３，３５，４２，５８）九个型别的感染，阳性病例再用多重引物ＰＣＲ进一步分型。结果，①喉癌组：ＨＰＶ感染的总阳性率为４９．１％，其中ＨＰＶ＿（１８）型阳性率１５．８％，ＨＰＶ＿（１６）型为１２．３％，ＨＰＶ＿（１６）和＿（１８）双重型感染为５．３％，ＨＰＶ＿（６／１１）和＿（１８）型混合感染为３．５％，其它型为１２．３％。②颈转移淋巴结组：总阳性率为２１．４％，其中ＨＰＶ＿（１６）、ＨＰＶ＿（１８）及ＨＰＶ＿（１６）和＿（１８）双重型感染各为７．１％。③癌前病变组：ＨＰＶ感染阳性率为１１．１％，为ＨＰＶ＿（６／１１）和＿（１８）型混合感染。④声带息肉组：阳性率７．１％，为ＨＰＶ＿（６／１１）型感染。⑤癌旁及癌周正常喉组织：均为ＨＰＶＤＮＡ阴性。本文对ＨＰＶ在喉癌中致病作用进行了讨论。     

国人喉癌与人类乳头状瘤病毒相关性的研究

本项研究对１２４例喉不同病变的新鲜组织标本，采用共同引物和多重引物ＰＣＲ的方法，进行ＨＰＶ ＤＮＡ检测。用共同引物ＰＣＲ检测ＨＰＶ６．，１６，１８，３１，３３，３５，４２，５８九个型的感染，阳性病例再用多重引物ＰＣＲ进一步分型。结果，（１）喉癌组；ＨＰＶ感染的总阳性率为４９．１％，其中ＨＰＶ１８型阳性率１５．８％，ＨＰＶ１６型为１２．３％，ＨＰＶ１６和１８双重感染为５．３％，ＨＰＶ６／１１和１８型     

喉乳头状瘤HPV16/18感染与p53蛋白表达的相关性

目的：了解喉乳头状瘤组织内ＨＰＶ１６／１８的感染与抑癌基因ｐ５３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ＨＰＶ１６／１８ＤＮＡ及ｐ５３蛋白的表达。结果：２４例组织中检出ＨＰＶ１６／１８ＤＮＡ（６８．６％）;１９例ｐ５３蛋白呈过度表达（５４．３％）;在１２例中同时检出ＨＰＶ１６／１８ＤＮＡ和ｐ５３蛋白过度表达（３４．３％）。结论：     

婴幼儿咽喉乳头瘤组织人乳头瘤病毒感染的探讨

目的 探讨温州地区人乳头瘤病毒感染和婴幼儿咽喉乳头状瘤的关系。方法 应用聚合酶链反应和核酸斑点杂交技术检测３５例婴幼儿咽喉乳头瘤组织和１０例对照组组织（小儿声带小结）ＨＰＶ６、１１、１６、１８、３３５个型别的ＤＮＡ。结果 乳头瘤组织ＨＰＶ感染率为９１．４％（３０／３５）,其中ＨＰＶ６型检出率为５４．２％（１９／３５）,ＨＰＶ１１型感染率为２５．７％（９／３５）,多重型别ＨＰＶ６＋１１感染率为１１．     

喉乳头状瘤临床行为与细胞增殖活性关系的研究

利用ＩＳＨ方法以及免疫病理手段对不同型的ＬＰ组织的标本进行ＰＣＮＡ、Ｐ５３检测，探讨不同型ＨＰＶ与ＬＰ的发生发展的相关机制，以寻求有效的检测方法帮助临床对ＬＰ的预后进行评估。标本选自１９９４年１月￣１９９５年１２月间我院收治的ＬＰ共３６例。ＩＳＨ法ＨＰＶ６ｂ／１１阳性率为７５％，明显高于ＨＰＶ１６和／或ＨＰＶ１８的表达。１０例喉鳞癌各有１例ＨＰＶ１６、１８阳性，无ＨＰＶ６ｂ／１１阳性。ＡＢＣ法行Ｐ     

原位免位PCR检测喉癌中人类乳头状瘤病毒E6蛋白

针对常规免疫组化检测石蜡包埋的喉癌组织中人类乳头状瘤病毒（ＨＰＶ）１６／１８型Ｅ６蛋白阳性率低、信号模糊等缺点，我们探索出一种敏感而有效的检测方法———原位免疫ＰＣＲ（ＩｎＳｉｔｕＩｍｍｕｎｏ－ＰＣＲ，ＩＳＩ－ＰＣＲ），报告如下：１　材料与方法１．１　材料：９例１６／１８型ＨＰＶＤＮＡ阳性的喉癌高分化鳞癌石蜡包埋组织。生物素标记ＰａＴＥＬ１４／ＥｃｏＲＩＤＮＡ片断。ＰＣＲ引物：５’－ＡＴＡＣＣＴＡＴＴＧＣＣＴＡＣＧＧＣＡＧ－３’；５’－ＣＧＴＴＡＧＴＡＡＡＴＧＡＡＴＴＴＴＣＴ－３’。１．２　方法…     

人乳头状瘤病毒与鼻内翻性乳头状瘤的关系

为探讨人乳头状瘤病毒（ＨＰＶ）感染与鼻内翻性乳头状瘤（ＩＰ）发病及其恶变的关系，研究选取组织病理确认为ＩＰ（３６例）和ＩＰ癌变（１６例）的石蜡包埋标本，采用多聚酶链反应（ＰＣＲ）方法对标本进行ＨＰＶ相关ＤＮＡ序列扩增。结果显示：３６例ＩＰ组织有２１例（５８．３％）为ＨＰＶ阳性；１６例ＩＰ癌变组织中有１１例（６８．８％）阳性。经统计学处理，二者间无显著性差异（Ｐ〉０．０５）．提示ＨＰＶ感染在ＩＰ的发     

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。     

Human papilloma virus infection and expression of p16 protein in laryngeal papilloma and laryngeal carcinoma]

OBJECTIVE: To evaluate the role of human papilloma virus (HPV) infection and inactivation of p16 gene in laryngeal papilloma (LP) and laryngeal squamous cell carcinoma (LC). METHODS: HPV consensus primers direct in situ polymerase chain reaction (ISPCR) and immunohistochemical method were applied to detect the presence of HPV genomes (1, 6, 8, 11, 13, 16, 18, 30, 31, 32, 33, 45, 51) and the expression of p16 protein respectively in 93 cases of formalin-fixed, paraffin-imbedded specimens, which contained 46 cases of LPs [adult-onset laryngeal papilloma (ALP) 21, juvenile-onset laryngeal papilloma (JLP)25], 26 cases of LCs, 6 cases of normal tissues adjacent to carcinoma, and 15 cases of vocal noduli. RESULTS: (1) The difference of positive rates of HPV-DNA in JLP group (84%, 21/25) and other groups were statistically significant (chi 2 test, P < 0.05). The difference of positive rates of HPV-DNA in ALPs(38.1%, 8/21), in LCs(19.2%, 5/26), in vocal noduli(0%, 0/15), and in normal tissues adjacent to carcinoma(0%, 0/6) were not significant statistically (chi 2 test or Fisher's exact probability test, P > 0.05). (2) The positive rates of expression of p16 protein in ALP group(57.1%, 12/21) and LC group(38.5%, 10/26) were significantly lower than that in vocal nodule group(93.3%, 14/15), in JLP group(88%, 22/25), and in normal tissues adjacent to carcinoma group (100%, 6/6) (chi 2 test or Fisher's exact probability test, P > 0.05). There were no significant differences of positive rates of expression of p16 protein between ALP group and LC group, and between JLP group and vocal nodule group (chi 2 test, P > 0.05). (3) In LPs, the difference of positive rates of p16 protein expression between HPV positive cases and HPV negative cases was significant statistically (chi 2 test, P < 0.05). In LCs, there was no difference in p16 protein expression rate between the two teams(Fisher exact probability test, P > 0.05). CONCLUSION: The pathogenesis of JLP is closely associated with HPV infection and not associated with the inactivation of p16 gene. Conversely, the pathogenesis of ALP and LC is associated with the inactivation of p16 gene and not associated with the HPV infection.     

The detection and significance of human papilloma virus 11b virus like particles and its serum antibody in juvenile larynx papilloma]

OBJECTIVE: To study the relationship between human papilloma virus 11b virus like particles (HPV11bVLPS) serum antibody and the development and prognosis of juvenile larynx papilloma (JLP). METHODS: Enzyme linked immunosorbent assay (ELISA) was used to detect the serum HPV11bVLP antibody (Ab) of 46 JLP's samples in different stage and 20 controls using HPV11bVLPS which was produced by recombinant bacilovirus in insect cells. Grouping: A: control group (n = 20); B: the time of onset was 1 years (n = 15); C: the time of onset was 2 years (n = 15); the patients were followed-up 1 year without recurrence (n = 8); E: The patients were followed-up 2 years without recurrence (n = 8). RESULTS: A value of HPV11bVLP Ab among A, B, C, D, E. group were: (0.073 +/- 0.035); (0.120 +/- 0.049); (0.137 +/- 0.057); (0.518 +/- 0.122); (0.557 +/- 0.144). There was a significant difference between JLP patients and the control group (P < 0.05). The level of HPV11bVLP Ab in (D + E) group (0.534 +/- 0.132) was higher than (B + C) group (0.128 +/- 0.053) (t = 14.90, P < 0.001). CONCLUSION: The results suggested that HPV serum antibody was produced in JLP with HPV infection. There is close relationship between the development and prognosis of the disease and the level of HPV11Ab in serum. The assay of serum HPV11bVLPAb and HPV-VLP could be used as immunological study of HPV11-infection associated disease.     

鼻腔及鼻窦内翻性乳头状瘤与人乳头状瘤病毒感染及病毒亚型的关系

目的 采用多聚酶链反应(polymerase chain reaction，PCR)检测鼻腔及鼻窦内翻性乳头状瘤(nasal inverted papilloma，NIP)中人乳头状瘤病毒(human papilloma virus，HPV)DNA及其亚型，旨在探讨HPV在NIP发病中的作用及其与预后的关系。方法将28例鼻腔及鼻窦NIP分为无复发组、复发组、癌变组，用聚合酶链反应检测其HPV通用型、6、11、16及18型的感染情况，同时以10例慢性炎症鼻黏膜或炎症性鼻息肉病例为对照。结果28例NIP患者HPV-DNA总阳性率为75％(21例)。无复发组病例HPV-DNA阳性率为42％(5／12)，均为单一的低危型HPV感染(4例为HPV6型，1例为HPV11型)；复发组的13例和癌变组的3例中，均可检出HPV-DNA。复发者以检出HPV6、HPV11型DNA为主，4例具有双重感染。癌变者以检出HPV16、HPV18型DNA为主，其中2例为双重感染。结论NIP的发生与HPV感染密切相关。应用PCR检测HPV及各亚型感染情况有助于预测肿瘤的临床行为及预后，指导临床对高危患者进行重点监测。     

喉癌和喉乳头状瘤组织中人乳头状瘤病毒和p16蛋白的检测

目的探讨人类乳头状瘤病毒（humanpapillomavirus,HPV）感染和抑癌基因p16的失活与喉癌和喉乳头状瘤（laryngealpapilloma,LP）发生的相关性,以进一步阐明喉癌和LP的病因和发病机理。方法收集LP46例,其中成人型喉乳头状瘤（adult-onsetLP,ALP）21例,青少年型喉乳头状瘤（juvenile-onsetLP,JLP）25例、喉癌26例、癌旁正常组织6例、声带小结15例,用标记的HPV1,6,8,11,13,16,18,30,31,32,33,45,51通用引物直接法原位聚合酶链反应（polymerasechainreaction,PCR）方法和免疫组化（SP法）方法分别检测HPV-DNA和p16蛋白。结果①HPV阳性率JLP组（84%,21/25）显著高于ALP组（38.1%,8/21）、喉癌组（19.2%,5/26）、声带小结组（0/15）和癌旁组织组（0/6）（χ     

HPV6b病毒样颗粒免疫治疗儿童喉乳头状瘤临床初步研究

目的 :研究HPV6bL1病毒样颗粒 (VLP)治疗儿童喉乳头状瘤 (JLP)的安全性和免疫原性。方法 :应用基因工程制备的HPV6bL1VLP 5、10、2 5 μg 3种剂量递增方法对 10例严重复发性JLP患儿进行免疫接种 ,记录不良反应及行血、尿常规和生化检测 ,ELISA法检测血清特异性HPV6bL1VLP抗体 ,对 7例患儿进行迟发性超敏反应 (DTH)试验 ,纤维喉镜随访观察喉部病变情况。结果 :接种后患儿无局部和全身不良反应 ,血清均能产生特异性的中和抗体 ,接种前 3天和 3种剂量完成后及开始治疗 1年后的血清抗体吸收度A均值分别为 0 .110± 0 .0 35 ,0 .310± 0 .0 12 ,0 .5 87± 0 .0 12 ,0 .75 2± 0 .0 19,0 .772± 0 .0 13。第 1剂量完成后与接种前 3天A均值比较 ,第 2剂量与第 1剂量完成后比较 ,第 3剂量与第 2剂量完成后比较 ,接种 1年后与对照组比较 ,各组间差异均有统计学意义 (均P <0 .0 1)。 7例行DTH试验的患儿均呈阳性反应。经免疫治疗后的 10例患儿未见复发。结论 :HPV6bL1VLP对JLP具有安全性和免疫原性 ,可成为防治JLP的有效疫苗。     

Short-fragment PCR assay for highly sensitive broad-spectrum detection of human papillomaviruses in laryngeal squamous cell carcinoma and normal mucosa: clinico-pathological evaluation

The aim of the study was to determine the prevalence and genotypes of HPV infection in laryngeal cancer specimens, normal mucosa obtained from the surgical margin and laryngeal nodules using a novel high sensitive and specific SPF₁₀ HPV DNA test, PCR/DEIA method and INNO-LiPA genotyping assay. The correlation between HPV presence and clinico-pathological features was analyzed. Tissue samples were collected from 93 primary laryngeal squamous cell carcinoma (LSCC), 49 specimens of normal mucosa and from 22 specimens of laryngeal nodules serving as control group. HPV DNA was amplified by the short PCR fragment (SPF₁₀) primer set using HPV DNA enzyme immunoassay (DNA/DEIA) method and INNO-LiPA HPV genotyping assay. Human papillomavirus was detected in 33 (35.5%) of the 93 samples from LSCC, in 4 (8.2%) of 49 samples of the normal mucosa and it was not detected in any of the sample from the control group. Twenty-eight of 33 (81.8%) were positive for HPV-16, 6 of 33 (18.2%) were positive for HPV-18 and 5 of 33 (15.1%) were positive for HPV-33. Multiple infection was found in 5 of 33 (15.1%); 3 samples were positive for HPV-16 and HPV-33, 2 samples for HPV-16 and HPV-18. There was a statistically significant correlation between the presence of HPV in LSCC tumors and in control group samples and between the presence of HPV in the tumors and normal mucosa from the free surgical margin. The presence of HPV infection in 35.5% of the cases suggests a possible role in the etiology of laryngeal cancer and supports the role of high-risk types of HPV (16, 18 and 33) in LSCC. HPV infection is not likely to influence survival rates as an independent prognostic factor in patients with laryngeal cancer.     

Human papillomavirus and risk of laryngeal cancer

We determined the relationship between human papillomavirus (HPV) infection and the HPV types detected in 44 patients with squamous cell carcinoma, 10 laryngeal leukoplakia patients, and 12 patients evaluated for benign laryngeal conditions (controls). The sources of HPV DNA were from brushings from the upper respiratory tract and lesion (benign or malignant), oral rinses, and biopsies of patient lesions. Polymerase chain reaction (PCR) and DNA sequencing were used to identify and type HPV. We detected HPV in 25.0% (11/44) of patients with laryngeal cancer, in 30.0% (3/10) of patients with laryngeal leukoplakia, and in 16.7% (2/12) of noncancer controls. Patients with cancer were not more likely to be identified with oncogenic HPV types ( 18.2%) than either the leukoplakia group (20%) or the control group (16.7%). An increased risk of disease was associated with current tobacco use and former alcohol drinking in cancer patients versus controls and in leukoplakia patients versus controls (all p < .05). After we controlled for tobacco and alcohol effects on the risk of disease, exposure to oncogenic HPV types was associated with an increased risk of laryngeal cancer (odds ratio = 3.0) and of laryngeal leukoplakia (odds ratio = 6.0) compared to controls, although the results were not statistically significant. This study suggests that although HPV infection and HPV oncogenic types are not found at a higher frequency in laryngeal cancer or laryngeal leukoplakia as compared to controls, infection is associated with an increased risk of disease after controlling for the effects of alcohol and tobacco use.