[Investigation on HPV DNA detection and genotyping practices used in French laboratories in 2009]. / Enquête sur les pratiques de détection et de génotypage des papillomavirus humains dans les laboratoires de microbiologie en France en 2009.

The French National reference Laboratory for Human papillomavirus (HPV) performed in 2009 a national study in order to review the methods used to detect and identify HR HPV genotypes in microbiology laboratories. Results from this study show a great diversity in volumes of samples treated in laboratories. Among clinical indications, the most frequent is a result of ASC-US at a Pap smear. This indication in the only one covered by the National Public Insurance System and is mostly performed in laboratories from private sector. Other indications mainly correspond to research programs and are performed in public Hospitals. This study allowed also to review the adequacy between the liquid based cytology samples and the assays used for direct detection of HR HPV or identification of the genotypes present in the sample. The right tests were not carried in the right solution storage according to the recommendations from different HPV testing assays. National recommendations should be elaborated in order to improve the performance of the test used.

[Detection and typification of human papilloma virus in pre cancerous cervical lesions]. / Detección y tipificación de virus papiloma humano en lesiones preneoplásicas del cuello uterino mediante PCR-RFLP.

BACKGROUND: The association of different genotypes of human papilloma virus (HPV) with cervical cancer is well known. However, there is little information about their association with pre-cancerous lesions. AIM: To assess the frequency of different HPV genotypes in pre cancerous cervical lesions. MATERIAL AND METHODS: A cervical sample was obtained by cytobrush in 15 women with low grade lesions and 40 women with high grade lesions, subjected to conization by loop electrical excision procedure (LEEP). Detection and typification of HPV was done by polymerase chain reaction and restriction fragment length polymorphism. RESULTS: All women were infected with HPV. Eighty five percent of samples were typified. A unique HPV subtype was found in 76% of women. Fourteen percent had an infection with multiple subtypes and in 10%, the viral genotype was not identified. The most common subtypes found were HPV 16, HPV 52 and HPV 53. CONCLUSIONS: There is a high rate of infection with HPV with a high oncogenic risk among these women.

PathogenMip assay: a multiplex pathogen detection assay.

The Molecular Inversion Probe (MIP) assay has been previously applied to a large-scale human SNP detection. Here we describe the PathogenMip Assay, a complete protocol for probe production and applied approaches to pathogen detection. We have demonstrated the utility of this assay with an initial set of 24 probes targeting the most clinically relevant HPV genotypes associated with cervical cancer progression. Probe construction was based on a novel, cost-effective, ligase-based protocol. The assay was validated by performing pyrosequencing and Microarray chip detection in parallel experiments. HPV plasmids were used to validate sensitivity and selectivity of the assay. In addition, 20 genomic DNA extracts from primary tumors were genotyped with the PathogenMip Assay results and were in 100% agreement with conventional sequencing using an L1-based HPV genotyping protocol. The PathogenMip Assay is a widely accessible protocol for producing and using highly discriminating probes, with experimentally validated results in pathogen genotyping, which could potentially be applied to the detection and characterization of any microbe.

Detección y tipificación de virus papiloma humano en lesiones preneoplásicas del cuello uterino mediante PCR-RFLP / Detection and typification of human papilloma virus in pre cancerous cervical lesions

Background: The association of different genotypes of human papilloma virus (HPV) with cervical cancer is well known. However, there is little information about their association with pre-cancerous lesions. Aim: To assess the frequency of different HPV genotypes in pre cancerous cervical lesions. Material and methods: A cervical sample was obtained by cytobrush in 15 women with low grade lesions and 40 women with high grade lesions, subjected to conization by loop electrical excision procedure (LEEP). Detection and typification of HPV was done by polymerase chain reaction and restriction fragment length polymorphism. Results: All women were infected with HPV. Eighty five percent of samples were typified. A unique HPV subtype was found in 76 percent of women. Fourteen percent had an infection with multiple subtypes and in 10 percent, the viral genotype was not identified. The most common subtypes found were HPV 16, HPV 52 and HPV 53. Conclusions: There is a high rate of infection with HPV with a high oncogenic risk among these women.

[Detection rate of human papillomavirus-16 in esophageal squamous cell carcinoma from different Chinese populations].

BACKGROUND & OBJECTIVE: Anyang in Henan Province of China is a hyperendemic area of esophageal cancer. The infection of human papillomavirus (HPV) is thought as an important pathogenesis of esophageal cancer in Anyang. This study was to detect infection rate and level of HPV-16 in esophageal squamous cell carcinoma (ESCC) patients from 3 different Chinese areas, and investigate its relationship with the pathogenesis of ESCC. METHODS: Infection status of HPV-16 in 119 ESCC specimens (43 collected from Anyang, 43 from Beijing, and the rest 33 from Mongolia nationality of Inner Mongolia) was detected by in situ hybridization (ISH) technique with digoxin-labeled HPV-16 E6 probe. RESULTS: HPV16 infection rates were 81.4%, 69.8%, and 63.6% in the specimens from Anyang, Beijing, and Inner Mongolia, respectively. Infection level of HPV-16 was significantly higher in Anyang group than in Beijing group (H=3.91, P<0.05) and Inner Mongolia group(H=4.22,P<0.05). There was no significant difference between the latter 2 groups. Furthermore, the proportion of strong expression of HPV16 (++ and +++) was significantly higher in Anyang group than in the other 2 groups(H=3.95, P<0.05). CONCLUSIONS: HPV-16 infection rate is high in the esophageal specimens from the 3 different areas. Infection status of HPV16 is serious in Anyang.

Inhibición del crecimiento de la línea de melanoma murino B16F10 por diferentes compuestos polifenólicos / Growth inhibition of the murine melanoma cell line B16F10 with poliphenols

Introducción: Estudiamos la respuesta de la línea B16F10 al tratamiento con seis polifenoles vegetales comparándola con la del melfalán. Material y métodos: Se han realizado pruebas de citoxicidad (24 horas) y antiproliferación (72 horas) de los polifenoles y la cuantificación de la viabilidad celular con el test de MTT. Resultados: Observamos una pequeña inhibición con tangeretina y luteonina (24 h) a la máxima concentración ensayada (25 y 50 pM), mientras que el melfalán mostró una curva dosis-respuesta. A las 72 horas, los cultivos presentaron una inhibición del crecimiento a la máxima concentración (50 pM con 7,3'-dimetilhesperetina, mientras que la tangeretina mostró una acción antiproliferante efectiva con una curva dosis-respuesta evidente. Conclusiones: Estos resultados sugieren que la presencia de al menos tres grupos metoxilados adyacentes, como ocurre en la tangeretina, potencia el efecto antiproliferante y la ausencia del doble enlace C2-C3 en los polifenoles hidroxilados conlleva una pérdida importante de actividad (AU)

[Clinical and molecular biology studies of laryngeal papillomatosis]. / Klinische und molekularbiologische Untersuchungen zur laryngealen Papillomatose.

BACKGROUND: Recurrent laryngeal papillomatosis is a benign neoplastic disease which is probably caused by but at least associated with the Human Papilloma Virus. It can be of significant importance for the affected patients because of its recurrent clinical course. A wide variety of therapeutic measures have been described including the surgical removal either with conventional instruments or laser. Malignancies developing from papillomas have been reported. PATIENTS: The clinical courses of all 95 patients who have been treated for laryngeal papillomatosis since 1960 were analysed retrospectively. The two most common forms of treatment, surgical removal either conventionally or with the use of the laser, were compared. "Hot-start" polymerase chain reaction and Southern blot hybridization were used to detect HPV-DNA. The case reports of all patients developing cancer of the larynx are included. RESULTS: Laryngeal papillomatosis is a disease of all ages, more often first diagnosed before age 10 or after age 30. Puberty had no effect on the clinical course. However, the rate of complications such as tracheostomy and glottic webs was significantly reduced after laser surgery. HPV-DNA was found in 10 of 11 samples. Squamous cell carcinoma subsequently developed in four cases, three of which occurred almost simultaneously and were therefore not included. CONCLUSION: The term juvenile laryngeal papillomatosis should be replaced by recurrent respiratory papillomatosis. The occurrence of squamous cell carcinomas in patients previously treated for papillomas underlines the need for repeated histological studies. The surgical treatment remains the mainstay in the management of laryngeal papillomatosis. The laser surgical technique is superior to conventional removal. Using the most sensitive and specific methods presently available, HPV-DNA can be detected in a large percentage of laryngeal papillomas.

Resultados de la biopsia por colposcopia en pacientes con citología PAP II y cambios por PVH

Se realizó una revisión de la s 245 pacientes enviadas a Colposcopia por citología PAP II más cambios virales, en un período de 3 años, se encontró en la biopsia dirigida un 68.16 por ciento de lesión por condiloma, un 31.42 por ciento asociado a NIC y un 0.4 por ciento con cáncer invasor. Esto confirma los informes de la relación de PVH y cáncer de cérvix y por lo tanto la necesidad de colposcopia y biopsia en las pacientes con citologías PAP II y cambios virales (coilocitos, disqueratocitos, binucleación)

Detección del papilomavirus humano en ADN de tejido normal y lesiones premalignas del cérvix por hibridación molecular / Identification of human papilloma virus in DNA of normal tissue and precursor lesions of uterine cervix by molecular hybridization

Se analizaron 64 muestras de ADN obtenido de tejido histológicamente normal del cérvix y de lesiones tempranas del cáncer cervical por hibridación dot-blot, para detectar la presencia de papilomavirus humano(PVH). Utilizando ADN de controles adecuados y sondas específicas, detectamos la infección por PVH en 39 por ciento(25 de 64) de los casos; mientras que 61 por ciento (39 de 64) resultó negativo a PVH. Se detectó PVH del grupo de ®bajo riesgo¼ (tipo 6/11) en un 80 por ciento (20 de 25) de las muestras de ADN; mientras que un 8 por ciento (2 de 25) fue positivo a PVH del grupo de ®alto riesgo¼ (tipo 16/18). Además, encontramos 12 por ciento (3 de 25) de las muestras positivas a ambos grupos de virus

Evaluation of two commercially available in situ hybridization kits for detection of human papillomavirus DNA in cervical biopsies: comparison to Southern blot hybridization.

In order to determine the distribution of human papillomavirus (HPV) types and to compare in situ hybridization (ISH) with Southern blot hybridization (SBH), paired colposcopically directed cervical biopsies and cervical-vaginal lavages were obtained from 92 women referred for abnormal Pap smears. The lavages and one of the biopsies were snap-frozen and subsequently tested by SBH for HPV DNA. The other biopsy was formalin-fixed and paraffin-embedded for conventional light microscopy and for ISH using two commercially available kits from Digene Diagnostics Inc.: ViraType In Situ containing probe mixtures to HPVs 6 and 11, 16 and 18, and 31, 33, and 35; and Omniprobe containing a single probe mixture to 14 different anogenital HPVs. In addition to the aforementioned types Omniprobe contains probes for types 42, 43, 44, 45, 51, 52, and 56. HPV DNA was detected by SBH in 27% of specimens showing no specific pathological change and in 92% of cases of cervical intraepithelial neoplasia (CIN). The HPVs by SBH in CIN were type 16 in 25%, types 31 or 35 in 23%, types 52 or 56 in 17%, uncharacterized types in 13%, mixed types in 8%, and type 18 in 6%. None of the specimens was positive for the low-risk HPV types 6, 11, 42, 43, or 44. The proportion of cases of CIN containing HPV 16 increased as the grade of CIN increased. Otherwise there were no striking differences detected in the HPV types in the different grades of CIN. ISH using ViraType In Situ was positive in only 50% of cases of CIN that were HPV positive by SBH.(ABSTRACT TRUNCATED AT 250 WORDS)

Human papillomavirus DNA in adenocarcinoma in situ, microinvasive adenocarcinoma of the uterine cervix, and coexisting cervical squamous intraepithelial neoplasia.

Previously, human papillomavirus (HPV) DNA, mainly HPV-18 DNA, was detected in more than 40% (17/40 cases) of invasive adenocarcinoma of the uterine cervix in our laboratory. In order to identify HPV DNA in the precursor lesions of adenocarcinoma of the cervix, 11 cases of adenocarcinoma in situ containing microinvasive adenocarcinoma and 10 cases of adenocarcinoma in situ were studied for the presence of HPV DNA by in situ hybridization using highly sensitive 3H-labeled HPV-16 and HPV-18 DNA probes. HPV types present in cervical squamous intraepithelial neoplasia (CIN) coexisting with adenocarcinoma in situ and microinvasive adenocarcinoma were also studied. Apart from the coexisting CIN II-III with glandular neoplasms, 48 cases of CIN III (severe dysplasia and squamous carcinoma in situ) removed by conization or hysterectomy and known to be free of adenocarcinoma were used for comparison. HPV DNA was detected in 64% of microinvasive adenocarcinoma, 70% of adenocarcinoma in situ, and 63% of the control CIN III. HPV-18 DNA was the preponderant type of HPV DNA found in adenocarcinoma in situ and microinvasive adenocarcinoma. All cases of HPV DNA-positive microinvasive adenocarcinoma contained the same type of HPV DNA as the lesions of coexisting adenocarcinoma in situ. CIN coexisting with microinvasive adenocarcinoma or adenocarcinoma in situ contained the same type of HPV as identified in the glandular lesions, whereas all of the HPV DNA-positive control CIN III cases contained HPV-16 DNA. These results suggest that adenocarcinoma in situ is a precursor lesion of adenocarcinoma of the cervix that contains HPV DNA, and that CIN coexisting with adenocarcinoma may be a result of a metaplastic process of adenocarcinoma or of bidirectional differentiation of the affected reserve cells.