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Sample records for hpv-16 early gene

  1. HPV16 early gene E5 specifically reduces miRNA-196a in cervical cancer cells

    Science.gov (United States)

    Liu, Chanzhen; Lin, Jianfei; Li, Lianqin; Zhang, Yonggang; Chen, Weiling; Cao, Zeyi; Zuo, Huancong; Chen, Chunling; Kee, Kehkooi

    2015-01-01

    High-risk human papillomavirus (HPV) type 16, which is responsible for greater than 50% of cervical cancer cases, is the most prevalent and lethal HPV type. However, the molecular mechanisms of cervical carcinogenesis remain elusive, particularly the early steps of HPV infection that may transform normal cervical epithelium into a pre-neoplastic state. Here, we report that a group of microRNAs (microRNAs) were aberrantly decreased in HPV16-positive normal cervical tissues, and these groups of microRNAs are further reduced in cervical carcinoma. Among these miRNAs, miR196a expression is the most reduced in HPV16-infected tissues. Interestingly, miR196a expression is low in HPV16-positive cervical cancer cell lines but high in HPV16-negative cervical cancer cell lines. Furthermore, we found that only HPV16 early gene E5 specifically down-regulated miRNA196a in the cervical cancer cell lines. In addition, HoxB8, a known miR196a target gene, is up-regulated in the HPV16 cervical carcinoma cell line but not in HPV18 cervical cancer cell lines. Various doses of miR196a affected cervical cancer cell proliferation and apoptosis. Altogether, these results suggested that HPV16 E5 specifically down-regulates miR196a upon infection of the human cervix and initiates the transformation of normal cervix cells to cervical carcinoma. PMID:25563170

  2. Depletion of HPV16 early genes induces autophagy and senescence in a cervical carcinogenesis model, regardless of viral physical state.

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    Hanning, Jennifer E; Saini, Harpreet K; Murray, Matthew J; Caffarel, Maria M; van Dongen, Stijn; Ward, Dawn; Barker, Emily M; Scarpini, Cinzia G; Groves, Ian J; Stanley, Margaret A; Enright, Anton J; Pett, Mark R; Coleman, Nicholas

    2013-11-01

    In cervical carcinomas, high-risk human papillomavirus (HR-HPV) may be integrated into host chromosomes or remain extra-chromosomal (episomal). We used the W12 cervical keratinocyte model to investigate the effects of HPV16 early gene depletion on in vitro cervical carcinogenesis pathways, particularly effects shared by cells with episomal versus integrated HPV16 DNA. Importantly, we were able to study the specific cellular consequences of viral gene depletion by using short interfering RNAs known not to cause phenotypic or transcriptional off-target effects in keratinocytes. We found that while cervical neoplastic progression in vitro was characterized by dynamic changes in HPV16 transcript levels, viral early gene expression was required for cell survival at all stages of carcinogenesis, regardless of viral physical state, levels of early gene expression or histology in organotypic tissue culture. Moreover, HPV16 early gene depletion induced changes in host gene expression that were common to both episome-containing and integrant-containing cells. In particular, we observed up-regulation of autophagy genes, associated with enrichment of senescence and innate immune-response pathways, including the senescence-associated secretory phenotype (SASP). In keeping with these observations, HPV16 early gene depletion induced autophagy in both episome-containing and integrant-containing W12 cells, as evidenced by the appearance of autophagosomes, punctate expression of the autophagy marker LC3, conversion of LC3B-I to LC3B-II, and reduced levels of the autophagy substrate p62. Consistent with the reported association between autophagy and senescence pathways, HPV16 early gene depletion induced expression of the senescence marker beta-galactosidase and increased secretion of the SASP-related protein IGFBP3. Together, these data indicate that depleting HR-HPV early genes would be of potential therapeutic benefit in all cervical carcinogenesis pathways, regardless of viral

  3. HPV16 early gene E5 specifically reduces miRNA-196a in cervical cancer cells

    OpenAIRE

    Liu, Chanzhen; Lin, Jianfei; Li, Lianqin; Zhang, Yonggang; Chen, Weiling; Cao, Zeyi; Zuo, Huancong; Chen, Chunling; Kee, Kehkooi

    2015-01-01

    High-risk human papillomavirus (HPV) type 16, which is responsible for greater than 50% of cervical cancer cases, is the most prevalent and lethal HPV type. However, the molecular mechanisms of cervical carcinogenesis remain elusive, particularly the early steps of HPV infection that may transform normal cervical epithelium into a pre-neoplastic state. Here, we report that a group of microRNAs (microRNAs) were aberrantly decreased in HPV16-positive normal cervical tissues, and these groups of...

  4. HPV-16 L1 genes with inactivated negative RNA elements induce potent immune responses

    International Nuclear Information System (INIS)

    Rollman, Erik; Arnheim, Lisen; Collier, Brian; Oeberg, Daniel; Hall, Haakan; Klingstroem, Jonas; Dillner, Joakim; Pastrana, Diana V.; Buck, Chris B.; Hinkula, Jorma; Wahren, Britta; Schwartz, Stefan

    2004-01-01

    Introduction of point mutations in the 5' end of the human papillomavirus type 16 (HPV-16) L1 gene specifically inactivates negative regulatory RNA processing elements. DNA vaccination of C57Bl/6 mice with the mutated L1 gene resulted in improved immunogenicity for both neutralizing antibodies as well as for broad cellular immune responses. Previous reports on the activation of L1 by codon optimization may be explained by inactivation of the regulatory RNA elements. The modified HPV-16 L1 DNA that induced anti-HPV-16 immunity may be seen as a complementary approach to protein subunit immunization against papillomavirus

  5. Six1 overexpression at early stages of HPV16-mediated transformation of human keratinocytes promotes differentiation resistance and EMT

    International Nuclear Information System (INIS)

    Xu, Hanwen; Pirisi, Lucia; Creek, Kim E.

    2015-01-01

    Previous studies in our laboratory discovered that SIX1 mRNA expression increased during in vitro progression of HPV16-immortalized human keratinocytes (HKc/HPV16) toward a differentiation-resistant (HKc/DR) phenotype. In this study, we explored the role of Six1 at early stages of HPV16-mediated transformation by overexpressing Six1 in HKc/HPV16. We found that Six1 overexpression in HKc/HPV16 increased cell proliferation and promoted cell migration and invasion by inducing epithelial–mesenchymal transition (EMT). Moreover, the overexpression of Six1 in HKc/HPV16 resulted in resistance to serum and calcium-induced differentiation, which is the hallmark of the HKc/DR phenotype. Activation of MAPK in HKc/HPV16 overexpressing Six1 is linked to resistance to calcium-induced differentiation. In conclusion, this study determined that Six1 overexpression resulted in differentiation resistance and promoted EMT at early stages of HPV16-mediated transformation of human keratinocytes. - Highlights: • Six1 expression increases during HPV16-mediated transformation. • Six1 overexpression causes differentiation resistance in HPV16-immortalized cells. • Six1 overexpression in HPV16-immortalized keratinocytes activates MAPK. • Activation of MAPK promotes EMT and differentiation resistance. • Six1 overexpression reduces Smad-dependent TGF-β signaling

  6. Multiple-integrations of HPV16 genome and altered transcription of viral oncogenes and cellular genes are associated with the development of cervical cancer.

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    Xulian Lu

    Full Text Available The constitutive expression of the high-risk HPV E6 and E7 viral oncogenes is the major cause of cervical cancer. To comprehensively explore the composition of HPV16 early transcripts and their genomic annotation, cervical squamous epithelial tissues from 40 HPV16-infected patients were collected for analysis of papillomavirus oncogene transcripts (APOT. We observed different transcription patterns of HPV16 oncogenes in progression of cervical lesions to cervical cancer and identified one novel transcript. Multiple-integration events in the tissues of cervical carcinoma (CxCa are significantly more often than those of low-grade squamous intraepithelial lesions (LSIL and high-grade squamous intraepithelial lesions (HSIL. Moreover, most cellular genes within or near these integration sites are cancer-associated genes. Taken together, this study suggests that the multiple-integrations of HPV genome during persistent viral infection, which thereby alters the expression patterns of viral oncogenes and integration-related cellular genes, play a crucial role in progression of cervical lesions to cervix cancer.

  7. Sites of disruption within E1 and E2 genes of HPV16 and association with cervical dysplasia.

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    Tsakogiannis, D; Gortsilas, P; Kyriakopoulou, Z; Ruether, I G A; Dimitriou, T G; Orfanoudakis, G; Markoulatos, P

    2015-11-01

    Integration of HPV16 DNA into the host chromosome usually disrupts the E1 and/or E2 genes. The present study investigated the disruption of E1, E2 genes in a total of eighty four HPV16-positive precancerous and cervical cancer specimens derived from Greek women (seventeen paraffin-embedded cervical biopsies and sixty seven Thin Prep samples). Complete E2 and E1 genes were amplified using three and nine overlapping primer sets respectively, in order to define the sites of disruption. Extensive mapping analysis revealed that disruption/deletion events within E2 gene occurred in high grade and cervical cancer samples (x(2) test, P disruption was documented among low grade cervical intraepithelial neoplasias. In addition, disruptions within the E1 gene occur both in high and low grade cervical intraepithelial neoplasia. This leads to the assumption that in low grade cervical intraepithelial neoplasias only E1 gene disruption was involved (Fisher's exact test, P disruption of E1 gene was located between nucleotides 1059 and 1323, while the most prevalent deleted region of the E2 gene was located between nucleotides 3172 and 3649 (E2 hinge region). Therefore, it is proposed that each population has its own profile of frequencies and sites of disruptions and extensive mapping analysis of E1 and E2 genes is mandatory in order to determine suitable markers for HPV16 DNA integration analysis in distinct populations. © 2015 Wiley Periodicals, Inc.

  8. Changes in global gene expression profiles induced by HPV 16 E6 oncoprotein variants in cervical carcinoma C33-A cells

    International Nuclear Information System (INIS)

    Zacapala-Gómez, Ana Elvira; Del Moral-Hernández, Oscar; Villegas-Sepúlveda, Nicolás; Hidalgo-Miranda, Alfredo; Romero-Córdoba, Sandra Lorena

    2016-01-01

    We analyzed the effects of the expression of HPV 16 E6 oncoprotein variants (AA-a, AA-c, E-A176/G350, E-C188/G350, E-G350), and the E-Prototype in global gene expression profiles in an in vitro model. E6 gene was cloned into an expression vector fused to GFP and was transfected in C33-A cells. Affymetrix GeneChip Human Transcriptome Array 2.0 platform was used to analyze the expression of over 245,000 coding transcripts. We found that HPV16 E6 variants altered the expression of 387 different genes in comparison with E-Prototype. The altered genes are involved in cellular processes related to the development of cervical carcinoma, such as adhesion, angiogenesis, apoptosis, differentiation, cell cycle, proliferation, transcription and protein translation. Our results show that polymorphic changes in HPV16 E6 natural variants are sufficient to alter the overall gene expression profile in C33-A cells, explaining in part the observed differences in oncogenic potential of HPV16 variants. - Highlights: • Amino acid changes in HPV16 E6 variants modulate the transciption of specific genes. • This is the first comparison of global gene expression profile of HPV 16 E6 variants. • Each HPV 16 E6 variant appears to have its own molecular signature.

  9. Changes in global gene expression profiles induced by HPV 16 E6 oncoprotein variants in cervical carcinoma C33-A cells

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    Zacapala-Gómez, Ana Elvira, E-mail: zak_ana@yahoo.com.mx [Laboratorio de Biomedicina Molecular, Unidad Académica de Ciencias Químico Biológicas, Universidad Autónoma de Guerrero, Chilpancingo, Gro., México (Mexico); Del Moral-Hernández, Oscar, E-mail: odelmoralh@gmail.com [Laboratorio de Biomedicina Molecular, Unidad Académica de Ciencias Químico Biológicas, Universidad Autónoma de Guerrero, Chilpancingo, Gro., México (Mexico); Villegas-Sepúlveda, Nicolás, E-mail: nvillega@cinvestav.mx [Departamento de Biomedicina Molecular, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional (CINVESTAV-IPN), México, D.F., México (Mexico); Hidalgo-Miranda, Alfredo, E-mail: ahidalgo@inmegen.gob.mx [Laboratorio de Genómica del Cáncer, Instituto Nacional de Medicina Genómica (INMEGEN), México, D.F., México (Mexico); Romero-Córdoba, Sandra Lorena, E-mail: sromero_cordoba@hotmail.com [Laboratorio de Genómica del Cáncer, Instituto Nacional de Medicina Genómica (INMEGEN), México, D.F., México (Mexico); and others

    2016-01-15

    We analyzed the effects of the expression of HPV 16 E6 oncoprotein variants (AA-a, AA-c, E-A176/G350, E-C188/G350, E-G350), and the E-Prototype in global gene expression profiles in an in vitro model. E6 gene was cloned into an expression vector fused to GFP and was transfected in C33-A cells. Affymetrix GeneChip Human Transcriptome Array 2.0 platform was used to analyze the expression of over 245,000 coding transcripts. We found that HPV16 E6 variants altered the expression of 387 different genes in comparison with E-Prototype. The altered genes are involved in cellular processes related to the development of cervical carcinoma, such as adhesion, angiogenesis, apoptosis, differentiation, cell cycle, proliferation, transcription and protein translation. Our results show that polymorphic changes in HPV16 E6 natural variants are sufficient to alter the overall gene expression profile in C33-A cells, explaining in part the observed differences in oncogenic potential of HPV16 variants. - Highlights: • Amino acid changes in HPV16 E6 variants modulate the transciption of specific genes. • This is the first comparison of global gene expression profile of HPV 16 E6 variants. • Each HPV 16 E6 variant appears to have its own molecular signature.

  10. Genetic variability in L1 and L2 genes of HPV-16 and HPV-58 in Southwest China.

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    Yaofei Yue

    Full Text Available HPV account for most of the incidence of cervical cancer. Approximately 90% of anal cancers and a smaller subset (<50% of other cancers (oropharyngeal, penile, vaginal, vulvar are also attributed to HPV. The L1 protein comprising HPV vaccine formulations elicits high-titre neutralizing antibodies and confers type restricted protection. The L2 protein is a promising candidate for a broadly protective HPV vaccine. In our previous study, we found the most prevalent high-risk HPV infectious serotypes were HPV-16 and HPV-58 among women of Southwest China. To explore gene polymorphisms and intratypic variations of HPV-16 and HPV-58 L1/L2 genes originating in Southwest China, HPV-16 (L1: n = 31, L2: n = 28 and HPV-58 (L1: n = 21, L2: n = 21 L1/L2 genes were sequenced and compared to others described and submitted to GenBank. Phylogenetic trees were then constructed by Neighbor-Joining and the Kimura 2-parameters methods (MEGA software, followed by an analysis of the diversity of secondary structure. Then selection pressures acting on the L1/L2 genes were estimated by PAML software. Twenty-nine single nucleotide changes were observed in HPV-16 L1 sequences with 16/29 non-synonymous mutations and 13/29 synonymous mutations (six in alpha helix and two in beta turns. Seventeen single nucleotide changes were observed in HPV-16 L2 sequences with 8/17 non-synonymous mutations (one in beta turn and 9/17 synonymous mutations. Twenty-four single nucleotide changes were observed in HPV-58 L1 sequences with 10/24 non-synonymous mutations and 14/24 synonymous mutations (eight in alpha helix and four in beta turn. Seven single nucleotide changes were observed in HPV-58 L2 sequences with 4/7 non-synonymous mutations and 3/7 synonymous mutations. The result of selective pressure analysis showed that most of these mutations were of positive selection. This study may help understand the intrinsic geographical relatedness and biological differences of HPV-16/HPV-58 and

  11. The transcriptional regulator gene E2 of the Human Papillomavirus (HPV) 16 influences the radiosensitivity of cervical keratinocytes

    International Nuclear Information System (INIS)

    Lindel, Katja; Rieken, Stefan; Daffinger, Sigrid; Weber, Klaus J; Villiers, Ethel-Michele de; Debus, Jürgen

    2012-01-01

    Clinical studies have demonstrated that HPV induced tumors constitute a specific subclass of cancer with a better response to radiation treatment. The purpose of this study was to investigate meaning of viral E2-gene for radiosensitivity. W12 cells contain episomal HPV 16 genomes, whereas S12 cells, which derive from the W12 line, contain HPV DNA as integrated copies. Clonogenic survival was analyzed using 96-well in vitro test. Using flow cytometry cell cycle analyses were performed. Expression of pRb and p53 were analyzed using intracellular staining. W12 cells (intact E2 gene) showed a lower survival fraction than S12 cells. W12 cells developed a G2/M block 24 h after irradiation with 2 Gy whereas S12 showed no G2/M bloc. After irradiation S12 cells developed polyploidy and pRb-positive cells decreased. W12 cells showed no change of pRb-positive cells. Depending on E2 gene status differences in cell cycle regulation might cause radioresistance. The E2/E7/pRb pathway seems to influence HPV-induced radiosensitivity. Our experiments demonstrated an effect of HPV on radiosensitivity of cervical keratinocytes via viral transcription regulator E2 pathway

  12. The HPV16 E7 oncoprotein increases the expression of Oct3/4 and stemness-related genes and augments cell self-renewal

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    Organista-Nava, Jorge; Gómez-Gómez, Yazmín [Programa de Doctorado en Ciencias Biomédicas, Instituto de Fisiología Celular (IFC), Universidad Nacional Autónoma de México (UNAM), Ciudad de México 04510, México (Mexico); Departamento de Genética y Biología Molecular, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional (CINVESTAV-IPN), Ciudad de México 07360, México (Mexico); Ocadiz-Delgado, Rodolfo; García-Villa, Enrique [Departamento de Genética y Biología Molecular, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional (CINVESTAV-IPN), Ciudad de México 07360, México (Mexico); Bonilla-Delgado, José [Unidad de Investigación, Hospital Juárez de México, Ciudad de México 07760, México (Mexico); Lagunas-Martínez, Alfredo [División de Biología Molecular de Patógenos, CISEI, Instituto Nacional de Salud Pública, Cuernavaca, Morelos, México (Mexico); and others

    2016-12-15

    Oct3/4 is a transcription factor involved in maintenance of the pluripotency and self-renewal of stem cells. The E7 oncoprotein and 17β-estradiol (E{sub 2}) are key factors in cervical carcinogenesis. In the present study, we aimed to investigate the effect of the HPV16 E7 oncoprotein and E{sub 2} on the expression pattern of Oct3/4, Sox2, Nanog and Fgf4. We also determined whether the E7 oncoprotein is associated with cell self-renewal. The results showed that Oct3/4, Sox2, Nanog and Fgf4 were upregulated by the E7 oncoprotein in vivo and in vitro and implicate E{sub 2} in the upregulation of these factors in vivo. We also demonstrated that E7 is involved in cell self-renewal, suggesting that the HPV16 E7 oncoprotein upregulates Oct3/4, Sox2, Nanog and Fgf4 expression to maintain the self-renewal capacity of cancer stem cells. -- Graphical abstract: The HPV16 E7 oncoprotein and 17β-estradiol are involved in the upregulation of Oct3/4, Sox2, Nanog and Fgf4 expression to maintain the self-renewal ability of cancer stem cells in cervical cancer. - Highlights: •The HPV16 E7 oncoprotein enhances cellular proliferation and dedifferentiation. •The E7 oncoprotein induces stemness-related genes expression in vivo and in vitro. •The 17β-estradiol induces stemness-related genes expression in vivo. •The HPV16 E7 oncoprotein is involved in the cell self-renewal of cancer cells.

  13. The HPV16 E7 oncoprotein increases the expression of Oct3/4 and stemness-related genes and augments cell self-renewal

    International Nuclear Information System (INIS)

    Organista-Nava, Jorge; Gómez-Gómez, Yazmín; Ocadiz-Delgado, Rodolfo; García-Villa, Enrique; Bonilla-Delgado, José; Lagunas-Martínez, Alfredo

    2016-01-01

    Oct3/4 is a transcription factor involved in maintenance of the pluripotency and self-renewal of stem cells. The E7 oncoprotein and 17β-estradiol (E 2 ) are key factors in cervical carcinogenesis. In the present study, we aimed to investigate the effect of the HPV16 E7 oncoprotein and E 2 on the expression pattern of Oct3/4, Sox2, Nanog and Fgf4. We also determined whether the E7 oncoprotein is associated with cell self-renewal. The results showed that Oct3/4, Sox2, Nanog and Fgf4 were upregulated by the E7 oncoprotein in vivo and in vitro and implicate E 2 in the upregulation of these factors in vivo. We also demonstrated that E7 is involved in cell self-renewal, suggesting that the HPV16 E7 oncoprotein upregulates Oct3/4, Sox2, Nanog and Fgf4 expression to maintain the self-renewal capacity of cancer stem cells. -- Graphical abstract: The HPV16 E7 oncoprotein and 17β-estradiol are involved in the upregulation of Oct3/4, Sox2, Nanog and Fgf4 expression to maintain the self-renewal ability of cancer stem cells in cervical cancer. - Highlights: •The HPV16 E7 oncoprotein enhances cellular proliferation and dedifferentiation. •The E7 oncoprotein induces stemness-related genes expression in vivo and in vitro. •The 17β-estradiol induces stemness-related genes expression in vivo. •The HPV16 E7 oncoprotein is involved in the cell self-renewal of cancer cells.

  14. Treatment of minimal residual disease after surgery or chemotherapy in mice carrying HPV16-associated tumours: cytokine and gene therapy with IL-2 and GM-CSF

    Czech Academy of Sciences Publication Activity Database

    Mikyšková, Romana; Indrová, Marie; Šímová, Jana; Jandlová, Táňa; Bieblová, Jana; Jinoch, P.; Bubeník, Jan; Vonka, V.

    2004-01-01

    Roč. 24, č. 1 (2004), s. 161-167 ISSN 1019-6439 R&D Projects: GA MZd NC7148; GA MZd NC7552; GA ČR GA301/01/0985 Institutional research plan: CEZ:AV0Z5052915 Keywords : HPV16 * gene therapy * minimal residual tumour disease Subject RIV: FD - Oncology ; Hematology Impact factor: 3.056, year: 2004

  15. Specific interaction between hnRNP H and HPV16 L1 proteins: Implications for late gene auto-regulation enabling rapid viral capsid protein production

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    Zheng, Zi-Zheng; Sun, Yuan-Yuan; Zhao, Min; Huang, Hui [National Institute of Diagnostics and Vaccine Development in Infectious Diseases, Xiamen University, Xiamen, Fujian 361005 (China); School of Life Sciences, Xiamen University, Xiamen, Fujian 361005 (China); Zhang, Jun; Xia, Ning-Shao [National Institute of Diagnostics and Vaccine Development in Infectious Diseases, Xiamen University, Xiamen, Fujian 361005 (China); School of Life Sciences, Xiamen University, Xiamen, Fujian 361005 (China); School of Public Health, Xiamen University, Xiamen, Fujian 361005 (China); Miao, Ji, E-mail: jmiao@xmu.edu.cn [National Institute of Diagnostics and Vaccine Development in Infectious Diseases, Xiamen University, Xiamen, Fujian 361005 (China); School of Life Sciences, Xiamen University, Xiamen, Fujian 361005 (China); Zhao, Qinjian, E-mail: qinjian_zhao@xmu.edu.cn [National Institute of Diagnostics and Vaccine Development in Infectious Diseases, Xiamen University, Xiamen, Fujian 361005 (China); School of Public Health, Xiamen University, Xiamen, Fujian 361005 (China)

    2013-01-18

    Highlights: ► The RNA-binding hnRNP H regulates late viral gene expression. ► hnRNP H activity was inhibited by a late viral protein. ► Specific interaction between HPV L1 and hnRNP H was demonstrated. ► Co-localization of HPV L1 and hnRNP H inside cells was observed. ► Viral capsid protein production, enabling rapid capsid assembly, was implicated. -- Abstract: Heterogeneous nuclear ribonucleoproteins (hnRNPs), including hnRNP H, are RNA-binding proteins that function as splicing factors and are involved in downstream gene regulation. hnRNP H, which binds to G triplet regions in RNA, has been shown to play an important role in regulating the staged expression of late proteins in viral systems. Here, we report that the specific association between hnRNP H and a late viral capsid protein, human papillomavirus (HPV) L1 protein, leads to the suppressed function of hnRNP H in the presence of the L1 protein. The direct interaction between the L1 protein and hnRNP H was demonstrated by complex formation in solution and intracellularly using a variety of biochemical and immunochemical methods, including peptide mapping, specific co-immunoprecipitation and confocal fluorescence microscopy. These results support a working hypothesis that a late viral protein HPV16 L1, which is down regulated by hnRNP H early in the viral life cycle may provide an auto-regulatory positive feedback loop that allows the rapid production of HPV capsid proteins through suppression of the function of hnRNP H at the late stage of the viral life cycle. In this positive feedback loop, the late viral gene products that were down regulated earlier themselves disable their suppressors, and this feedback mechanism could facilitate the rapid production of capsid proteins, allowing staged and efficient viral capsid assembly.

  16. Highly potent and specific siRNAs against E6 or E7 genes of HPV16- or HPV18-infected cervical cancers

    OpenAIRE

    Chang, J T-C; Kuo, T-F; Chen, Y-J; Chiu, C-C; Lu, Y-C; Li, H-F; Shen, C-R; Cheng, A-J

    2010-01-01

    Infection with high-risk types (type 16 or type 18) of human papillomaviruses (HPVs) increases a patient's risk of cervical cancer. Given the importance of the cervix and the severe side effects resulting from traditional cancer therapies, this study aimed to achieve targeted inhibition of viral oncogenes in tumor cells using small interfering RNAs (siRNA). To accomplish this, we developed nine siRNAs against either the E6 or E7 genes of HPV-16 or HPV-18 in several combinations, yielding siRN...

  17. Rapid identification of HPV 16 and 18 by multiplex nested PCR-immunochromatographic test.

    Science.gov (United States)

    Kuo, Yung-Bin; Li, Yi-Shuan; Chan, Err-Cheng

    2015-02-01

    Human papillomavirus (HPV) types 16 and 18 are known to be high-risk viruses that cause cervical cancer. An HPV rapid testing kit that could help physicians to make early and more informed decisions regarding patient care is needed urgently but not yet available. This study aimed to develop a multiplex nested polymerase chain reaction-immunochromatographic test (PCR-ICT) for the rapid identification of HPV 16 and 18. A multiplex nested PCR was constructed to amplify the HPV 16 and 18 genotype-specific L1 gene fragments and followed by ICT which coated with antibodies to identify rapidly the different PCR products. The type-specific gene regions of high-risk HPV 16 and 18 could be amplified successfully by multiplex nested PCR at molecular sizes of approximately 99 and 101bp, respectively. The capture antibodies raised specifically against the moleculars labeled on the PCR products could be detected simultaneously both HPV 16 and 18 in one strip. Under optimal conditions, this PCR-ICT assay had the capability to detect HPV in a sample with as low as 100 copies of HPV viral DNA. The PCR-ICT system has the advantage of direct and simultaneous detection of two high-risk HPV 16 and 18 DNA targets in one sample, which suggested a significant potential of this assay for clinical application. Copyright © 2014. Published by Elsevier B.V.

  18. HPV DNA methylation at the early promoter and E1/E2 integrity: A comparison between HPV16, HPV18 and HPV45 in cervical cancer.

    Science.gov (United States)

    Amaro-Filho, Sérgio Menezes; Pereira Chaves, Cláudia Bessa; Felix, Shayany Pinto; Basto, Diogo Lisbôa; de Almeida, Liz Maria; Moreira, Miguel Angelo Martins

    2018-04-09

    To compare and describe type-specific characteristics of HPV16, HPV18 and HPV45 in cervical cancer with respect to 3'LCR methylation and disruption of E1/E2. The methylation level of 137 cervical cancer samples (70 with HPV16, 37 with HPV18, and 30 with HPV45) of Brazilian patients was analyzed by pyrosequencing. PCR amplifications were performed to characterize E1 and E2 disruption as an episomal surrogate. The 3'LCR of HPV16 showed a higher methylation at all CpG sites (7%, 9%, 11%, 10% and 10%) than homologous HPV18 regions (4%, 5%. 6%, 9% and 5%) and HPV45 regions (7%, 7% and 5%). Presence of intact E1/E2 was associated with higher HPV16 and HPV18 methylation levels at all CpG sites (p < 0.05). Disruption of E1/E2 was more frequently found in HPV45 (97%) and HPV18 (84%) than in HPV16 DNA (30%). HPV16 disruption was more frequently found in E1 (48%) unlike HPV18, where it was found in E2 (61%). Concomitant disruption of E1/E2 was most frequent in HPV45 (72%). The findings showed a higher methylation associated with intact E1/E2 for HPV16 and HPV18. The closely phylogenetic related HPV18 and HPV45 share a similar methylation level and the frequency of viral genome disruption. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

  19. Disruption of HPV16-E7 by CRISPR/Cas System Induces Apoptosis and Growth Inhibition in HPV16 Positive Human Cervical Cancer Cells

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    Zheng Hu

    2014-01-01

    Full Text Available High-risk human papillomavirus (HR-HPV has been recognized as a major causative agent for cervical cancer. Upon HPV infection, early genes E6 and E7 play important roles in maintaining malignant phenotype of cervical cancer cells. By using clustered regularly interspaced short palindromic repeats- (CRISPR- associated protein system (CRISPR/Cas system, a widely used genome editing tool in many organisms, to target HPV16-E7 DNA in HPV positive cell lines, we showed for the first time that the HPV16-E7 single-guide RNA (sgRNA guided CRISPR/Cas system could disrupt HPV16-E7 DNA at specific sites, inducing apoptosis and growth inhibition in HPV positive SiHa and Caski cells, but not in HPV negative C33A and HEK293 cells. Moreover, disruption of E7 DNA directly leads to downregulation of E7 protein and upregulation of tumor suppressor protein pRb. Therefore, our results suggest that HPV16-E7 gRNA guided CRISPR/Cas system might be used as a therapeutic strategy for the treatment of cervical cancer.

  20. Risk of progression of early cervical lesions is associated with integration and persistence of HPV-16 and expression of E6, Ki-67, and telomerase

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    Arianna Vega-Peña

    2013-01-01

    Full Text Available Background: Low-grade squamous intraepithelial lesions (LSIL are the earliest lesions of the uterine cervix, the persistence and integration of high-risk human papillomavirus (HR-HPV as type 16, which promotes the development of more aggressive lesions. Aim: To select more aggressive lesions with tendency to progress to invasive cervical cancer. Materials and Methods: A total of 75 cytological specimens in liquid base (Liqui-PREP were analyzed: 25 specimens were with no signs of SIL (NSIL and without HPV; 25 NSIL with HPV-16, and 25 with both LSIL and HPV-16. The expression of Ki-67, telomerase, and viral E6 was evaluated by immunocytochemistry; and the detection of viral DNA was done by polymerase chain reaction (PCR and restriction fragment length polymorphism (RFLPs for genotyping or sequencing of HPV-16. The physical state of HPV-16 was evaluated by in situ hybridization with amplification with tyramide. Results: Of the total group, 58.6% had LSIL associated with persistence and of these 59.3% was associated with integrated state of HPV as intense expression of E6, Ki-67 (P = 0.013, P = 0.055 has except for the expression of telomerase present a non-significant association (P<0.341. Conclusions: Overexpression of E6 and Ki-67 is associated with the integration of HPV-16, favoring viral persistence, and increasing the risk of progression in women with NSIL and LSIL.

  1. AKT1 loss correlates with episomal HPV16 in vulval intraepithelial neoplasia.

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    Arucha L Ekeowa-Anderson

    Full Text Available Anogenital malignancy has a significant association with high-risk mucosal alpha-human papillomaviruses (alpha-PV, particularly HPV 16 and 18 whereas extragenital SCC has been linked to the presence of cutaneous beta and gamma-HPV types. Vulval skin may be colonised by both mucosal and cutaneous (beta-, mu-, nu- and gamma- PV types, but there are few systematic studies investigating their presence and their relative contributions to vulval malignancy. Dysregulation of AKT, a serine/threonine kinase, plays a significant role in several cancers. Mucosal HPV types can increase AKT phosphorylation and activity whereas cutaneous HPV types down-regulate AKT1 expression, probably to weaken the cornified envelope to promote viral release. We assessed the presence of mucosal and cutaneous HPV in vulval malignancy and its relationship to AKT1 expression in order to establish the corresponding HPV and AKT1 profile of normal vulval skin, vulval intraepithelial neoplasia (VIN and vulval squamous cell carcinoma (vSCC. We show that HPV16 is the principle HPV type present in VIN, there were few detectable beta types present and AKT1 loss was not associated with the presence of these cutaneous HPV. We show that HPV16 early gene expression reduced AKT1 expression in transgenic mouse epidermis. AKT1 loss in our VIN cohort correlated with presence of high copy number, episomal HPV16. Maintained AKT1 expression correlated with low copy number, an increased frequency of integration and increased HPV16E7 expression, a finding we replicated in another untyped cohort of vSCC. Since expression of E7 reflects tumour progression, these findings suggest that AKT1 loss associated with episomal HPV16 may have positive prognostic implications in vulval malignancy.

  2. Monitoring HPV-16 E7 phosphorylation events

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    Nogueira, Marcela O.; Hošek, Tomáš; Calçada, Eduardo O.; Castiglia, Francesca [Magnetic Resonance Center (CERM) and Department of Chemistry “Ugo Schiff”, University of Florence, via Luigi Sacconi 6, Sesto Fiorentino (Italy); Massimi, Paola; Banks, Lawrence [International Centre for Genetic Engineering and Biotechnology (ICGEB), Padriciano 99, Trieste (Italy); Felli, Isabella C., E-mail: felli@cerm.unifi.it [Magnetic Resonance Center (CERM) and Department of Chemistry “Ugo Schiff”, University of Florence, via Luigi Sacconi 6, Sesto Fiorentino (Italy); Pierattelli, Roberta, E-mail: pierattelli@cerm.unifi.it [Magnetic Resonance Center (CERM) and Department of Chemistry “Ugo Schiff”, University of Florence, via Luigi Sacconi 6, Sesto Fiorentino (Italy)

    2017-03-15

    HPV-16 E7 is one of the key proteins that, by interfering with the host metabolism through many protein-protein interactions, hijacks cell regulation and contributes to malignancy. Here we report the high resolution investigation of the CR3 region of HPV-16 E7, both as an isolated domain and in the full-length protein. This opens the way to the atomic level study of the many interactions in which HPV-16 E7 is involved. Along these lines we show here the effect of one of the key post-translational modifications of HPV-16 E7, the phosphorylation by casein kinase II.

  3. Chemotherapy, IL-12 gene therapy and combined adjuvant therapy of HPV 16-associated MHC class I-proficient and -deficient tumours

    Czech Academy of Sciences Publication Activity Database

    Indrová, Marie; Bieblová, Jana; Jandlová, Táňa; Vonka, V.; Pajtasz-Piasecka, E.; Reiniš, Milan

    2006-01-01

    Roč. 28, č. 1 (2006), s. 253-260 ISSN 1019-6439 R&D Projects: GA MZd(CZ) NR7807; GA MZd(CZ) NR8004 Grant - others:Ministry of Scientific Research Information Society Technologies(PL) PBZ-KBN-091/PO5/2003 Institutional research plan: CEZ:AV0Z50520514 Keywords : HPV 16 * MHC class I-deficient and MHC class I-proficient tumour cells * CMRTD Subject RIV: EC - Immunology Impact factor: 2.556, year: 2006

  4. The HPV16 E7 oncoprotein increases the expression of Oct3/4 and stemness-related genes and augments cell self-renewal.

    Science.gov (United States)

    Organista-Nava, Jorge; Gómez-Gómez, Yazmín; Ocadiz-Delgado, Rodolfo; García-Villa, Enrique; Bonilla-Delgado, José; Lagunas-Martínez, Alfredo; Tapia, Jesús Santa-Olalla; Lambert, Paul F; García-Carrancá, Alejandro; Gariglio, Patricio

    2016-12-01

    Oct3/4 is a transcription factor involved in maintenance of the pluripotency and self-renewal of stem cells. The E7 oncoprotein and 17β-estradiol (E 2 ) are key factors in cervical carcinogenesis. In the present study, we aimed to investigate the effect of the HPV16 E7 oncoprotein and E 2 on the expression pattern of Oct3/4, Sox2, Nanog and Fgf4. We also determined whether the E7 oncoprotein is associated with cell self-renewal. The results showed that Oct3/4, Sox2, Nanog and Fgf4 were upregulated by the E7 oncoprotein in vivo and in vitro and implicate E 2 in the upregulation of these factors in vivo. We also demonstrated that E7 is involved in cell self-renewal, suggesting that the HPV16 E7 oncoprotein upregulates Oct3/4, Sox2, Nanog and Fgf4 expression to maintain the self-renewal capacity of cancer stem cells. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. Expression of HPV16 E5 produces enlarged nuclei and polyploidy through endoreplication

    International Nuclear Information System (INIS)

    Hu Lulin; Potapova, Tamara A.; Li Shibo; Rankin, Susannah; Gorbsky, Gary J.; Angeletti, Peter C.; Ceresa, Brian P.

    2010-01-01

    Anogenital cancers and head and neck cancers are causally associated with infection by high-risk human papillomavirus (HPV). The mechanism by which high-risk HPVs contribute to oncogenesis is poorly understood. HPV16 encodes three genes (HPV16 E5, E6, and E7) that can transform cells when expressed independently. HPV16 E6 and E7 have well-described roles causing genomic instability and unregulated cell cycle progression. The role of HPV16 E5 in cell transformation remains to be elucidated. Expression of HPV16 E5 results in enlarged, polyploid nuclei that are dependent on the level and duration of HPV16 E5 expression. Live cell imaging data indicate that these changes do not arise from cell-cell fusion or failed cytokinesis. The increase in nuclear size is a continual process that requires DNA synthesis. We conclude that HPV16 E5 produces polyploid cells by endoreplication. These findings provide insight into how HPV16 E5 can contribute to cell transformation.

  6. Gene silencing of HPV16 E6/E7 induced by promoter-targeting siRNA in SiHa cells

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    Hong, D; Lu, W; Ye, F; Hu, Y; Xie, X

    2009-01-01

    Background: Recently, transcriptional gene silencing induced by small interfering RNA (siRNA) was found in mammalian and human cells. However, previous studies focused on endogenous genes. Methods: In this study, we designed siRNA targeting the promoter of human papillomavirus 16 E6/E7 and transfected it into the cervical cancer cell line, SiHa. E6 and E7 mRNA and protein expression were detected in cells treated by promoter-targeting siRNA. Futhermore, cellular growth, proliferation, apoptos...

  7. Risk allelic load in Th2 and Th3 cytokines genes as biomarker of susceptibility to HPV-16 positive cervical cancer: a case control study

    International Nuclear Information System (INIS)

    Torres-Poveda, K.; Burguete-García, A. I.; Bahena-Román, M.; Méndez-Martínez, R.; Zurita-Díaz, M. A.; López-Estrada, G.; Delgado-Romero, K.; Peralta-Zaragoza, O.; Bermúdez-Morales, V. H.; Cantú, D.; García-Carrancá, A.; Madrid-Marina, V.

    2016-01-01

    Alterations in the host cellular immune response allow persistent infections with High-Risk Human Papillomavirus (HR-HPV) and development of premalignant cervical lesions and cervical cancer (CC). Variations of immunosuppressive cytokine levels in cervix are associated with the natural history of CC. To assess the potential role of genetic host immunity and cytokines serum levels in the risk of developing CC, we conducted a case–control study paired by age. Peripheral blood samples from patients with CC (n = 200) and hospital controls (n = 200), were used to evaluate nine biallelic SNPs of six cytokine genes of the adaptive immune system by allelic discrimination and cytokines serum levels by ELISA. After analyzing the SNP association by multivariate logistic regression adjusted by age, CC history and smoking history, three Th2 cytokines (IL-4, IL-6 and IL-10) and one Th3 (TGFB1) cytokine were significantly associated with CC. Individuals with at least one copy of the following risk alleles: T of SNP (−590C > T IL-4), C of SNP (−573G > C IL-6), A of SNP (−592C > A IL-10), T of SNP (−819C > T IL-10) and T of SNP (−509C > T TGFB1), had an adjusted odds ratio (OR) of 2.08 (95 % CI 1.475–2.934, p = 0.0001), an OR of 1.70 (95 % CI 1.208–2.404, p = 0.002), an OR of 1.87 (95 % CI 1.332–2.630, p = 0.0001), an OR of 1.67 (95 % CI 1.192–2.353, p = 0.003) and an OR of 1.91 (95 % CI 1.354–2.701, p = 0.0001), respectively, for CC. The burden of carrying two or more of these risk alleles was found to have an additive effect on the risk of CC (p trend = 0.0001). Finally, the serum levels of Th2 and Th3 cytokines were higher in CC cases than the controls; whereas IFNG levels, a Th1 cytokine, were higher in controls than CC cases. The significant associations of five SNPs with CC indicate that these polymorphisms are potential candidates for predicting the risk of development of CC, representing a risk allelic load for CC and can be used as a biomarker of

  8. HPV16-E2 protein modifies self-renewal and differentiation rate in progenitor cells of human immortalized keratinocytes.

    Science.gov (United States)

    Domínguez-Catzín, Victoria; Reveles-Espinoza, Alicia-María; Sánchez-Ramos, Janet; Cruz-Cadena, Raúl; Lemus-Hernández, Diana; Garrido, Efraín

    2017-04-03

    Cervical cancer is the fourth cause of death worldwide by cancer in women and is a disease associated to persistent infection with human papillomavirus (HPV), particularly from two high-risk types HPV16 and 18. The virus initiates its replicative cycle infecting cells located in the basal layer of the epithelium, where a small population of epithelial stem cells is located performing important functions of renewal and maintenance of the tissue. Viral E2 gene is one of the first expressed after infection and plays relevant roles in the replicative cycle of the virus, modifying fundamental processes in the infected cells. Thus, the aim of the present study was to demonstrate the presence of hierarchic subpopulations in HaCaT cell line and evaluate the effect of HPV16-E2 expression, on their biological processes. HaCaT-HPV16-E2 cells were generated by transduction of HaCaT cell line with a lentiviral vector. The α6-integrin-CD71 expression profile was established by immunostaining and flow cytometric analysis. After sorting, cell subpopulations were analyzed in biological assays for self-renewal, clonogenicity and expression of stemness factors (RT-qPCR). We identified in HaCaT cell line three different subpopulations that correspond to early differentiated cells (α6-integrin dim ), transitory amplifying cells (α6-integrin bri /CD71 bri ) and progenitor cells (α6-integrin bri /CD71 dim ). The last subpopulation showed stem cell characteristics, such as self-renewal ability, clonogenicity and expression of the well-known stem cell factors SOX2, OCT4 and NANOG, suggesting they are stem-like cells. Interestingly, the expression of HPV16-E2 in HaCaT cells changed its α6-integrin-CD71 immunophenotype modifying the relative abundance of the cell subpopulations, reducing significantly the percentage of α6-integrin bri /CD71 dim cells. Moreover, the expression of the stem cell markers was also modified, increasing the expression of SOX2 and NANOG, but decreasing notably

  9. Sequence imputation of HPV16 genomes for genetic association studies.

    Directory of Open Access Journals (Sweden)

    Benjamin Smith

    Full Text Available Human Papillomavirus type 16 (HPV16 causes over half of all cervical cancer and some HPV16 variants are more oncogenic than others. The genetic basis for the extraordinary oncogenic properties of HPV16 compared to other HPVs is unknown. In addition, we neither know which nucleotides vary across and within HPV types and lineages, nor which of the single nucleotide polymorphisms (SNPs determine oncogenicity.A reference set of 62 HPV16 complete genome sequences was established and used to examine patterns of evolutionary relatedness amongst variants using a pairwise identity heatmap and HPV16 phylogeny. A BLAST-based algorithm was developed to impute complete genome data from partial sequence information using the reference database. To interrogate the oncogenic risk of determined and imputed HPV16 SNPs, odds-ratios for each SNP were calculated in a case-control viral genome-wide association study (VWAS using biopsy confirmed high-grade cervix neoplasia and self-limited HPV16 infections from Guanacaste, Costa Rica.HPV16 variants display evolutionarily stable lineages that contain conserved diagnostic SNPs. The imputation algorithm indicated that an average of 97.5±1.03% of SNPs could be accurately imputed. The VWAS revealed specific HPV16 viral SNPs associated with variant lineages and elevated odds ratios; however, individual causal SNPs could not be distinguished with certainty due to the nature of HPV evolution.Conserved and lineage-specific SNPs can be imputed with a high degree of accuracy from limited viral polymorphic data due to the lack of recombination and the stochastic mechanism of variation accumulation in the HPV genome. However, to determine the role of novel variants or non-lineage-specific SNPs by VWAS will require direct sequence analysis. The investigation of patterns of genetic variation and the identification of diagnostic SNPs for lineages of HPV16 variants provides a valuable resource for future studies of HPV16

  10. HPV16 genetic variation and the development of cervical cancer worldwide

    Science.gov (United States)

    Cornet, I; Gheit, T; Iannacone, M R; Vignat, J; Sylla, B S; Del Mistro, A; Franceschi, S; Tommasino, M; Clifford, G M

    2013-01-01

    Background: Factors that favour a small proportion of HPV16 infections to progress to cancer are still poorly understood, but several studies have implicated a role of HPV16 genetic variation. Methods: To evaluate the association between HPV16 genetic variants and cervical cancer risk, we designed a multicentre case–control study based on HPV16-positive cervical samples (1121 cervical cancer cases and 400 controls) from the International Agency for Research on Cancer biobank. By sequencing the E6 gene, HPV16 isolates were classified into variant lineages and the European (EUR)-lineage isolates were subclassified by the common polymorphism T350G. Results: Incidence of variant lineages differed between cases and controls in Europe/Central Asia (P=0.006, driven by an underrepresentation of African lineages in cases), and South/Central America (P=0.056, driven by an overrepresentation of Asian American/North American lineages in cases). EUR-350G isolates were significantly underrepresented in cervical cancer in East Asia (odds ratio (OR)=0.02 vs EUR-350T; 95% confidence interval (CI)=0.00–0.37) and Europe/Central Asia (OR=0.42; 95% CI=0.27–0.64), whereas the opposite was true in South/Central America (OR=4.69; 95% CI=2.07–10.66). Conclusion: We observed that the distribution of HPV16 variants worldwide, and their relative risks for cervical cancer appear to be population-dependent. PMID:23169278

  11. Viral load and genomic integration of HPV 16 in cervical samples from HIV-1-infected and uninfected women in Burkina Faso.

    Science.gov (United States)

    Rousseau, Marie-Noelle Didelot; Costes, Valérie; Konate, Issouf; Nagot, Nicolas; Foulongne, Vincent; Ouedraogo, Abdoulaye; Van de Perre, Philippe; Mayaud, Philippe; Segondy, Michel

    2007-06-01

    The relationships between human papillomavirus type 16 (HPV 16) viral load, HPV 16 integration status, human immunodeficiency virus type 1 (HIV-1) status, and cervical cytology were studied among women enrolled in a cohort of female sex workers in Burkina Faso. The study focused on 24 HPV 16-infected women. The HPV 16 viral load in cervical samples was determined by real-time PCR. Integration ratio was estimated as the ratio between E2 and E6 genes DNA copy numbers. Integrated HPV16 viral load was defined as the product of HPV 16 viral load by the integration ratio. High HPV 16 viral load and high integration ratio were more frequent among women with squamous intraepithelial lesions compared with women with normal cytology (33% vs. 11%, and 33% vs. 0%, respectively), and among women with high-grade squamous intraepithelial lesions compared with women without high-grade squamous intraepithelial lesions (50% vs. 17%, and 50% vs. 11%, respectively). High HPV 16 DNA load, but not high integration ratio, was also more frequent among HIV-1-positive women (39% vs. 9%; and 23% vs. 18%, respectively). The absence of statistical significance of these differences might be explained by the small study sample size. High-integrated HPV 16 DNA load was significantly associated with the presence of high-grade squamous intraepithelial lesions (50% vs. 5%, P = 0.03) in univariate and multivariate analysis (adjusted odds-ratio: 19.05; 95% confidence interval (CI), 1.11-328.3, P = 0.03), but not with HIV-1 or other high-risk HPV types (HR-HPV). Integrated HPV 16 DNA load may be considered as a useful marker of high-grade cervical lesions in HPV 16-infected women. (c) 2007 Wiley-Liss, Inc.

  12. Genome wide expression analysis in HPV16 Cervical Cancer: identification of altered metabolic pathways

    Directory of Open Access Journals (Sweden)

    Salcedo Mauricio

    2007-09-01

    Full Text Available Abstract Background Cervical carcinoma (CC is a leading cause of death among women worldwide. Human papilloma virus (HPV is a major etiological factor in CC and HPV 16 is the more frequent viral type present. Our aim was to characterize metabolic pathways altered in HPV 16 tumor samples by means of transcriptome wide analysis and bioinformatics tools for visualizing expression data in the context of KEGG biological pathways. Results We found 2,067 genes significantly up or down-modulated (at least 2-fold in tumor clinical samples compared to normal tissues, representing ~3.7% of analyzed genes. Cervical carcinoma was associated with an important up-regulation of Wnt signaling pathway, which was validated by in situ hybridization in clinical samples. Other up-regulated pathways were those of calcium signaling and MAPK signaling, as well as cell cycle-related genes. There was down-regulation of focal adhesion, TGF-β signaling, among other metabolic pathways. Conclusion This analysis of HPV 16 tumors transcriptome could be useful for the identification of genes and molecular pathways involved in the pathogenesis of cervical carcinoma. Understanding the possible role of these proteins in the pathogenesis of CC deserves further studies.

  13. Immunogenicity of an HPV-16 L2 DNA vaccine

    Science.gov (United States)

    Hitzeroth, Inga I.; Passmore, Jo-Ann S.; Shephard, Enid; Stewart, Debbie; Müller, Martin; Williamson, Anna-Lise; Rybicki, Edward P.; Kast, W. Martin

    2009-01-01

    The ability to elicit cross-neutralizing antibodies makes human papillomavirus (HPV) L2 capsid protein a possible HPV vaccine. We examined and compared the humoral response of mice immunised with a HPV-16 L2 DNA vaccine or with HPV-16 L2 protein. The L2 DNA vaccine elicited a non-neutralising antibody response unlike the L2 protein. L2 DNA vaccination suppressed the growth of L2-expressing C3 tumor cells, which is a T cell mediated effect, demonstrating that the lack of non-neutralizing antibody induction by L2 DNA was not caused by lack of T cell immunogenicity of the construct. PMID:19559114

  14. Photon-induced cell migration and integrin expression promoted by DNA integration of HPV16 genome

    Energy Technology Data Exchange (ETDEWEB)

    Rieken, Stefan; Simon, Florian; Habermehl, Daniel; Dittmar, Jan Oliver; Combs, Stephanie E.; Weber, Klaus; Debus, Juergen; Lindel, Katja [University Hospital of Heidelberg, Department of Radiation Therapy and Radiation Oncology, Heidelberg (Germany)

    2014-10-15

    Persistent human papilloma virus 16 (HPV16) infections are a major cause of cervical cancer. The integration of the viral DNA into the host genome causes E2 gene disruption which prevents apoptosis and increases host cell motility. In cervical cancer patients, survival is limited by local infiltration and systemic dissemination. Surgical control rates are poor in cases of parametrial infiltration. In these patients, radiotherapy (RT) is administered to enhance local control. However, photon irradiation itself has been reported to increase cell motility. In cases of E2-disrupted cervical cancers, this phenomenon would impose an additional risk of enhanced tumor cell motility. Here, we analyze mechanisms underlying photon-increased migration in keratinocytes with differential E2 gene status. Isogenic W12 (intact E2 gene status) and S12 (disrupted E2 gene status) keratinocytes were analyzed in fibronectin-based and serum-stimulated migration experiments following single photon doses of 0, 2, and 10 Gy. Quantitative FACS analyses of integrin expression were performed. Migration and adhesion are increased in E2 gene-disrupted keratinocytes. E2 gene disruption promotes attractability by serum components, therefore, effectuating the risk of local infiltration and systemic dissemination. In S12 cells, migration is further increased by photon RT which leads to enhanced expression of fibronectin receptor integrins. HPV16-associated E2 gene disruption is a main predictor of treatment-refractory cancer virulence. E2 gene disruption promotes cell motility. Following photon RT, E2-disrupted tumors bear the risk of integrin-related infiltration and dissemination. (orig.) [German] Persistierende Infektionen mit humanen Papillomaviren 16 (HPV16) sind ein Hauptausloeser des Zervixkarzinoms. Die Integration der viralen DNS in das Wirtszellgenom fuehrt zum Integritaetsverlust des E2-Gens, wodurch in der Wirtszelle Apoptose verhindert und Motilitaet gesteigert werden. In

  15. [Colorimetric detection of HPV6 and HPV16 by loop mediated isothermal amplification].

    Science.gov (United States)

    Lu, Chun-bin; Luo, Le; Yang, Meng-jie; Nie, Kai; Wang, Miao; Ma, Xue-Jun

    2011-01-01

    A simple, rapid and sensitive colorimetric loop mediated isothermal amplification (LAMP) method was established to detect HPV6 and HPV 16 respectively. The method employed a set of four specially designed primers that recognized six distinct sequences of HPV6-E6 or HPV16-E7 for amplification of nucleic acid under isothermal conditions at 63 degrees C for one hour. The amplification process of LAMP was monitored by the addition of HNB (hydroxy naphthol blue) dye prior to amplification. A positive reaction was indicated by a color change from violet to sky blue and confirmed by real-time turbidimeter and agarose electrophoresis. Thirteen cervical swab samples having single infection with 13 different HPV genotypes were examined to evaluate the specificity. A serial dilution of a cloned plasmid containing HPV-E6 or HPV-E7 gene was examined to evaluate the sensitivity. The results showed that no cross-reaction with other HPV genotypes was observed. The colorimetric LAMP assay could achieve a sensitivity of 1000 copies, 10-20 times lower than that of real-time PCR. The assay was further evaluated with 62 clinical specimens and consistent results were obtained compared with the detection using Kai Pu HPV Genotyping Kit. We concluded that this colorimetric LAMP assay had potential usefulness for the rapid screening of the HPV6 or HPV16 infection in the laboratories and hospitals of provincial and municipal region in China.

  16. Tumour-inhibitory and antimetastatic effects of IL-2 in mice carrying MHC class I- tumours of HPV16 origin

    Czech Academy of Sciences Publication Activity Database

    Indrová, Marie; Bubeník, Jan; Mikyšková, Romana; Vonka, V.; Šmahel, M.; Žák, R.; Šímová, Jana; Bieblová, Jana; Mendoza, Luis; Jandlová, Táňa

    2002-01-01

    Roč. 2002, č. 20 (2002), s. 643-646 ISSN 1019-6439 Institutional research plan: CEZ:AV0Z5052915 Keywords : HPV16, IL-2 * tumour vaccines * gene therapy Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.931, year: 2002

  17. Local IFN-gamma therapy of HPV16-associated tumours

    Czech Academy of Sciences Publication Activity Database

    Mikyšková, Romana; Bieblová, Jana; Šímová, Jana; Indrová, Marie; Jandlová, Táňa; Vonka, V.; Šmahel, M.; Bubeník, Jan; Mendoza, Luis

    2003-01-01

    Roč. 49, č. 1 (2003), s. 26-32 ISSN 0015-5500 R&D Projects: GA MZd NC7148; GA MZd NC5900; GA ČR GA301/01/0985 Grant - others:GA CZ(CZ) Liga proti rakovině Institutional research plan: CEZ:AV0Z5052915 Keywords : MHC class I * IFN-gamma * HPV16 Subject RIV: FD - Oncology ; Hematology Impact factor: 0.527, year: 2003

  18. Early Detection of Autism (ASD) by a Non-invasive Quick Measurement of Markedly Reduced Acetylcholine & DHEA and Increased β-Amyloid (1-42), Asbestos (Chrysotile), Titanium Dioxide, Al, Hg & often Coexisting Virus Infections (CMV, HPV 16 and 18), Bacterial Infections etc. in the Brain and Corresponding Safe Individualized Effective Treatment.

    Science.gov (United States)

    Omura, Yoshiaki; Lu, Dominic; Jones, Marilyn K; Nihrane, Ahdallah; Duvvi, Harsha; Shimotsuura, Yasuhiro; Ohki, Motomu

    2015-01-01

    of above abnormal substances & coexisting infections, if treatment is given early. When HPV-16 & HPV-18 coexist, at triangular central area of the top of head, in addition to inability to talk, severe neuromuscular problems of lower extremity were found to also exist. However, if treatment is given 3-4 years after onset of Autism symptoms, even when successful biochemical reduction of above abnormal substances occurs, clinical improvement is less significant, since permanent damage in brain tissue seems to already exist. Therefore, early diagnosis & early treatment is very important for both Autism & Alzheimer's disease. In addition the optimal doses of Vitamin D3 and Taurine may play an important role in the future treatment of Autism, Alzheimer's Disease and memory disturbances by significantly increasing Acetylcholine and DHEA levels, enhancing the excretion of toxic substances in the urine, as well as having an anticancer effect.

  19. [Human Papilloma virus in Quechua women from Jujuy with high frequency of cervical cancer: viral types and HPV-16 variants].

    Science.gov (United States)

    Picconi, Maria Alejandra; Gronda, Jorge; Alonio, Lidia V; Villa, Luisa L; Sichero, Laura; Miranda, Sergio; Barcena, Martin; Teyssie, Angelica

    2002-01-01

    Human Papillomaviruses (HPVs) are etiologically associated to cervical carcinoma. In order to evaluate HPV infection and its relationship with the high frequency of this neoplasia in Quechua women from Jujuy (Argentina), 271 cervical samples from preneoplastic and neoplastic lesions (biopsies) and normal controls (cytologies) were studied. Detection and typing were performed using PCR-RFLP or PCR-hybridization and the HPV-16 variability in L1 and E6 genes (by PCR-hybridization) was analysed. HPV was detected in 52% of controls, 91% of low-grade lesions, 97% of high-grade lesions and 100% of invasive carcinomas, corresponding 55% to HPV-16. HPV-16 European variants were predominant, most of them being non-prototypic strains. The high frequency of high risk infection types and the raised proportion of HPV-16 non-prototypic variants related to a greater oncogenic potential could explain, in part, the high cervical cancer frequency of this native population. These data may contribute to disease control and vaccinal formulation.

  20. Unexpected inverse correlation between Native American ancestry and Asian American variants of HPV16 in admixed Colombian cervical cancer cases.

    Science.gov (United States)

    Lopera, Esteban A; Baena, Armando; Florez, Victor; Montiel, Jehidys; Duque, Constanza; Ramirez, Tatiana; Borrero, Mauricio; Cordoba, Carlos M; Rojas, Fredy; Pareja, Rene; Bedoya, Astrid M; Bedoya, Gabriel; Sanchez, Gloria I

    2014-12-01

    European (E) variants of HPV 16 are evenly distributed among world regions, meanwhile Non-European variants such as European-Asian (EAs), Asian American (AA) and African (Af) are mostly confined to Eastern Asia, The Americas and African regions respectively. Several studies have shown that genetic variation of HPV 16 is associated with the risk of cervical cancer, which also seems to be dependent on the population. This relationship between ethnicity and variants have led to the suggestion that there is co-evolution of variants with humankind. Our aim was to evaluate the relationship between the individual ancestry proportion and infection with HPV 16 variants in cervical cancer. We examined the association between ancestry and HPV 16 variants in samples of 82 cervical cancer cases from different regions of Colombia. Individual ancestry proportions (European, African and Native American) were estimated by genotyping 106 ancestry informative markers. Variants were identified by PCR amplification of the E6 gene, followed by reverse line blot hybridization (RLB) with variants specific probes. Overall European (E) and Asian American (AA) variants frequency was 66.5% and 33.5% respectively. Similar distribution was observed in cases with higher proportions of European or African ancestry. A higher Native American ancestry was significantly associated with higher frequency of E variants (median ancestry>23.6%, Age and place of birth adjusted OR: 3.55, 95% CI: 1.26-10.03, p=0.01). Even further, an inverse geographic correlation between Native American ancestry and frequency of infections with AA variants was observed (ρ=-0.825, p=0.008). Regions with higher proportion of Native American ancestry had a lower frequency of AA variants of HPV 16. This study suggests replacement of AA variants by E variants of human papillomavirus 16 in cervical cancer cases with high Native American ancestry. Copyright © 2014 Elsevier B.V. All rights reserved.

  1. Detection of the E7 transform gene of human papilloma virus type 16 in human oral squamous cell carcinoma.

    Science.gov (United States)

    Wang, J; Li, J; Huang, H; Fu, Y

    1998-12-01

    To determine, with the use of polymerase chain reaction, the prevalence of human papillomavirus (HPV) 16 in 30 patients with primary oral squamous cell carcinoma (OSCC) and 30 healthy control patients. DNA was extracted from freshly frozen tumor tissues of 30 patients with primary oral squamous cell carcinoma and from the oral mucosa of 30 controls. A pair of specific primers of the E7 early gene of HPV 16 were designed. PCR products were run by 1.5% agarose gel and the results of electrophoresis were photographed. HPV 16 was detected in 36.7% (11/30) of oral squamous cell carcinoma patients and 11.1% (4/30) of controls. HPV 16 has a significant association with oral squamous cell carcinoma. However, the role HPV 16 plays in the tumorigenesis of oral cancer and its clinical significance remain to be investigated.

  2. Repression of MHC class I transcription by HPV16E7 through interaction with a putative RXRβ motif and NF-κB cytoplasmic sequestration

    International Nuclear Information System (INIS)

    Li, Hui; Zhan, TaiLan; Li, Chang; Liu, Mugen; Wang, Qing K.

    2009-01-01

    Down-regulation of transcription of the MHC class I genes in HPV16 tumorigenic cells is partly due to HPV16E7 associated with the MHC class I promoter and repressed chromatin activation. In this study, we further demonstrated that HPV16E7 is physically associated with a putative RXRβ binding motif (GGTCA) of the proximal promoter of the MHC class I genes by using reporter transcriptional assays and chromatin immunoprecipitation assays. Our data also provide evidence that HPV16E7 inhibits TNF-α-induced up-regulation of MHC class I transcription by impaired nuclear translocation of NF-κB. More importantly, CaSki tumor cells treated with TSA and transfected with the constitutively active mutant form of IKK-α (which can activate NF-κB directly) showed a maximal level of up-regulation of MHC-I expression. Taken together, our results suggest that HPV16E7 may employ two independent mechanisms to ensure that either the constitutive or inducible transcription of MHC class I genes is down-regulated.

  3. Photon-induced cell migration and integrin expression promoted by DNA integration of HPV16 genome

    International Nuclear Information System (INIS)

    Rieken, Stefan; Simon, Florian; Habermehl, Daniel; Dittmar, Jan Oliver; Combs, Stephanie E.; Weber, Klaus; Debus, Juergen; Lindel, Katja

    2014-01-01

    Persistent human papilloma virus 16 (HPV16) infections are a major cause of cervical cancer. The integration of the viral DNA into the host genome causes E2 gene disruption which prevents apoptosis and increases host cell motility. In cervical cancer patients, survival is limited by local infiltration and systemic dissemination. Surgical control rates are poor in cases of parametrial infiltration. In these patients, radiotherapy (RT) is administered to enhance local control. However, photon irradiation itself has been reported to increase cell motility. In cases of E2-disrupted cervical cancers, this phenomenon would impose an additional risk of enhanced tumor cell motility. Here, we analyze mechanisms underlying photon-increased migration in keratinocytes with differential E2 gene status. Isogenic W12 (intact E2 gene status) and S12 (disrupted E2 gene status) keratinocytes were analyzed in fibronectin-based and serum-stimulated migration experiments following single photon doses of 0, 2, and 10 Gy. Quantitative FACS analyses of integrin expression were performed. Migration and adhesion are increased in E2 gene-disrupted keratinocytes. E2 gene disruption promotes attractability by serum components, therefore, effectuating the risk of local infiltration and systemic dissemination. In S12 cells, migration is further increased by photon RT which leads to enhanced expression of fibronectin receptor integrins. HPV16-associated E2 gene disruption is a main predictor of treatment-refractory cancer virulence. E2 gene disruption promotes cell motility. Following photon RT, E2-disrupted tumors bear the risk of integrin-related infiltration and dissemination. (orig.) [de

  4. Interaction between polymorphisms of the Human Leukocyte Antigen and HPV-16 Variants on the risk of invasive cervical cancer

    International Nuclear Information System (INIS)

    Araujo Souza, Patricia S de; Maciag, Paulo C; Ribeiro, Karina B; Petzl-Erler, Maria Luiza; Franco, Eduardo L; Villa, Luisa L

    2008-01-01

    Persistent infection with oncogenic types of human papillomavirus (HPV) is the major risk factor for invasive cervical cancer (ICC), and non-European variants of HPV-16 are associated with an increased risk of persistence and ICC. HLA class II polymorphisms are also associated with genetic susceptibility to ICC. Our aim is to verify if these associations are influenced by HPV-16 variability. We characterized HPV-16 variants by PCR in 107 ICC cases, which were typed for HLA-DQA1, DRB1 and DQB1 genes and compared to 257 controls. We measured the magnitude of associations by logistic regression analysis. European (E), Asian-American (AA) and African (Af) variants were identified. Here we show that inverse association between DQB1*05 (adjusted odds ratio [OR] = 0.66; 95% confidence interval [CI]: 0.39–1.12]) and HPV-16 positive ICC in our previous report was mostly attributable to AA variant carriers (OR = 0.27; 95%CI: 0.10–0.75). We observed similar proportions of HLA DRB1*1302 carriers in E-P positive cases and controls, but interestingly, this allele was not found in AA cases (p = 0.03, Fisher exact test). A positive association with DRB1*15 was observed in both groups of women harboring either E (OR = 2.99; 95% CI: 1.13–7.86) or AA variants (OR = 2.34; 95% CI: 1.00–5.46). There was an inverse association between DRB1*04 and ICC among women with HPV-16 carrying the 350T [83L] single nucleotide polymorphism in the E6 gene (OR = 0.27; 95% CI: 0.08–0.96). An inverse association between DQB1*05 and cases carrying 350G (83V) variants was also found (OR = 0.37; 95% CI: 0.15–0.89). Our results suggest that the association between HLA polymorphism and risk of ICC might be influenced by the distribution of HPV-16 variants

  5. Modulation of basal cell fate during productive and transforming HPV-16 infection is mediated by progressive E6-driven depletion of Notch.

    Science.gov (United States)

    Kranjec, Christian; Holleywood, Christina; Libert, Diane; Griffin, Heather; Mahmood, Radma; Isaacson, Erin; Doorbar, John

    2017-08-01

    In stratified epithelia such as the epidermis, homeostasis is maintained by the proliferation of cells in the lower epithelial layers and the concomitant loss of differentiated cells from the epithelial surface. These differentiating keratinocytes progressively stratify and form a self-regenerating multi-layered barrier that protects the underlying dermis. In such tissue, the continual loss and replacement of differentiated cells also limits the accumulation of oncogenic mutations within the tissue. Inactivating mutations in key driver genes, such as TP53 and NOTCH1, reduce the proportion of differentiating cells allowing for the long-term persistence of expanding mutant clones in the tissue. Here we show that through the expression of E6, HPV-16 prevents the early fate commitment of human keratinocytes towards differentiation and confers a strong growth advantage to human keratinocytes. When E6 is expressed either alone or with E7, it promotes keratinocyte proliferation at high cell densities, through the combined inactivation of p53 and Notch1. In organotypic raft culture, the activity of E6 is restricted to the basal layer of the epithelium and is enhanced during the progression from productive to abortive or transforming HPV-16 infection. Consistent with this, the expression of p53 and cleaved Notch1 becomes progressively more disrupted, and is associated with increased basal cell density and reduced commitment to differentiation. The expression of cleaved Notch1 is similarly disrupted also in HPV-16-positive cervical lesions, depending on neoplastic grade. When taken together, these data depict an important role of high-risk E6 in promoting the persistence of infected keratinocytes in the basal and parabasal layers through the inactivation of gene products that are commonly mutated in non-HPV-associated neoplastic squamous epithelia. © 2017 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great

  6. Clinical and epidemiological correlations between the infection with HPV 16 and HPV 18 and female cervical lesions.

    Science.gov (United States)

    Stoian, M; Repanovici, R; Corniţescu, F

    1995-01-01

    A number of 66 specimens from female cervical lesions were examined for infection with human papillomavirus (HPV) types 6, 11, 16, and 18 by nucleic acid hybridization in dot-blot techniques and 35 sera were tested by the immunodot-blot technique, in order to detect the presence of anti E4 and E7 HPV protein antibodies. The findings were compared with the histologic diagnosis. Fifty-six per cent of specimens contained HPV DNA sequences. In 47% of specimens from cervical carcinoma, HPV 11 was detected in 4 cases, HPV 16 in 21 cases, and HPV 18 in 7 cases. Serum antibodies against HPV 16 E4 and HPV 16 E7 occurred in all the cases of uterine carcinoma, in 4 of 10 cases of CIN I-II, and in 3 of 5 sera obtained from apparently healthy women. The analysis of risk factors disclosed the early onset of sexual activity, a relatively high number of births and abortions before the age of 22 years, the use of oral oestroprogestative contraceptive agents, the presence in anamnesis of genital infections with bacterial flora--Candida albicans, Trichomonas vaginalis, Chlamydia trachomatis, Mycoplasma, etc. Our results showed that HPV typing by nucleic acid hybridization was useful for differentiating low- from high-risk cervical lesions and also tried to elucidate the risk factors associated with HPV infections and progression to malignancy.

  7. Gene therapy pf HPV-16 induced tumours in rodents

    Czech Academy of Sciences Publication Activity Database

    Vonka, V.; Sobotková, E.; Šmahel, M.; Žák, R.; Hamšíková, E.; Bubeník, Jan

    1999-01-01

    Roč. 19, - (1999), s. 2014 ISSN 0250-7005. [Symposium on Local Cytokine Therapy of Cancer: Interleukin-2, Interferon and Related Cytokines /1./. Hamburg, 29.04.1999-01.05.1999] Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.375, year: 1999

  8. Adjuvant cytokine treatment of minimal residual disease after surgical therapy in mice carrying HPV16-associated tumours: Cytolytic activity of spleen cells from tumour regressors

    Czech Academy of Sciences Publication Activity Database

    Indrová, Marie; Mikyšková, Romana; Jandlová, Táňa; Vonka, V.; Bubeník, Jan; Bieblová, Jana

    2003-01-01

    Roč. 2003, č. 49 (2003), s. 217-222 ISSN 0015-5500 Institutional research plan: CEZ:AV0Z5052915 Keywords : HPV16 * gene therapy * IL-2 Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.527, year: 2003

  9. Immunization with mutant HPV16 E7 protein inhibits the growth of TC-1 cells in tumor-bearing mice.

    Science.gov (United States)

    Li, Yan-Li; Ma, Zhong-Liang; Zhao, Yue; Zhang, Jing

    2015-04-01

    Two human papillomavirus (HPV) 16 oncogenic proteins, E6 and E7, are co-expressed in the majority of HPV16-induced cervical cancer cells. Thus, the E6 and E7 proteins are good targets for developing therapeutic vaccines for cervical cancer. In the present study, immunization with the mutant non-transforming HPV16 E7 (mE7) protein was demonstrated to inhibit the growth of TC-1 cells in the TC-1 mouse model. The HPV16 mE7 gene was amplified by splicing overlap extension polymerase chain reaction using pET-28a(+)-E7 as a template, and the gene was cloned into pET-28a(+) to form pET-28a(+)-mE7. Compared with the E7 protein, mE7 lacks amino acid residues 94-98, and at residue 24, there is a Cys to Gly substitution. pET-28a(+)-mE7 was then introduced into Escherichia coli Rosetta. The expression of mE7 was induced by isopropyl β-D-1-thiogalactopyranoside. The mE7 protein was purified using Ni-NTA agarose and detected by SDS-PAGE and western blot analysis. In the tumor prevention model, no tumor was detected in the mice vaccinated with the mE7 protein. After 40 days, the tumor-free mice and control mice were challenged with 2×10 5 TC-1 cells. All control mice developed tumors six days later, but mE7 immunized mice were tumor free until 90 days. In the tumor therapy model, the TC-1 cells were initially injected subcutaneously, and the mice were subsequently vaccinated. Vaccination against the mE7 protein may significantly inhibit TC-1 cell growth compared to the control. These results demonstrated that immunization with the HPV16 mE7 protein elicited a long-term protective immunity against TC-1 tumor growth and generated a significant inhibition of TC-1 growth in a TC-1 mouse model.

  10. EXPRESSION OF HPV 16 AND 18 IN CERVICAL INTRAEPITHELIAL NEOPLASIA

    Directory of Open Access Journals (Sweden)

    Kodali Venkataramana

    2017-03-01

    Full Text Available BACKGROUND Cervical cancer is by far the most common human papilloma virus related disease. Nearly, all cases of cervical cancer can be attributable to human papilloma virus infection. Infection with the human papilloma virus is the main risk factors for cervical intraepithelial neoplasia and cervical cancer especially the high-risk types. The aim of the study is to study the prevalence of high-risk human papilloma virus 16 and 18 in various grades of cervical intraepithelial neoplasia. MATERIALS AND METHODS It is a prospective study for a period of two years. 50 cases of cervical intraepithelial neoplasia of various grades on histopathology were included in the study. Polymerase chain reaction DNA sequencing was done in all the cases. The patients were followed up for 1 year with Pap smears and results tabulated. RESULTS 77.77% of cases were human papilloma virus 16 positive and 22.22% for human papilloma virus 18. High-risk human papilloma virus was positive in 66.66% of cases beyond 30 years of age. In cases with positive HPV 16 or 18, 62.5% of CIN 1 cases progressed to CIN 2 on follow up for one year,all the CIN2 cases progressed to CIN 3 and CIN 3 cases persisted in the same phase. CONCLUSION High-risk human papilloma virus testing beyond 30 years should be included in the screening test along with Pap smears.

  11. Investigating Reports of Complex Regional Pain Syndrome: An Analysis of HPV-16/18-Adjuvanted Vaccine Post-Licensure Data

    Directory of Open Access Journals (Sweden)

    Frank Huygen

    2015-09-01

    There is not sufficient evidence to suggest an increased risk of developing CRPS following vaccination with HPV-16/18-adjuvanted vaccine. Post-licensure safety surveillance confirms the acceptable benefit-risk of HPV-16/18 vaccination.

  12. Reactivity of human sera in a sensitive, high-throughput pseudovirus-based papillomavirus neutralization assay for HPV16 and HPV18

    International Nuclear Information System (INIS)

    Pastrana, Diana V.; Buck, Christopher B.; Pang, Y.-Y. S.; Thompson, Cynthia D.; Castle, Philip E.; FitzGerald, Peter C.; Krueger Kjaer, Susanne; Lowy, Douglas R.; Schiller, John T.

    2004-01-01

    Sensitive high-throughput neutralization assays, based upon pseudoviruses carrying a secreted alkaline phosphatase (SEAP) reporter gene, were developed and validated for human papillomavirus (HPV)16, HPV18, and bovine papillomavirus 1 (BPV1). SEAP pseudoviruses were produced by transient transfection of codon-modified papillomavirus structural genes into an SV40 T antigen expressing line derived from 293 cells, yielding sufficient pseudovirus from one flask for thousands of titrations. In a 96-well plate format, in this initial characterization, the assay was reproducible and appears to be as sensitive as, but more specific than, a standard papillomavirus-like particle (VLP)-based enzyme-linked immunosorbent assay (ELISA). The neutralization assay detected type-specific HPV16 or HPV18 neutralizing antibodies (titers of 160-10240) in sera of the majority of a group of women infected with the corresponding HPV type, but not in virgin women. Sera from HPV16 VLP vaccinees had high anti-HPV16 neutralizing titers (mean: 45000; range: 5120-163840), but no anti-HPV18 neutralizing activity. The SEAP pseudovirus-based neutralization assay should be a practical method for quantifying potentially protective antibody responses in HPV natural history and prophylactic vaccine studies

  13. HPV16 E6 regulates annexin 1 (ANXA1) protein expression in cervical carcinoma cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Calmon, Marilia Freitas [Department of Biology, Institute of Bioscience, Language and Exact Science, São Paulo State University, São Jose do Rio Preto (Brazil); Sichero, Laura [Molecular Biology Laboratory, Centre for Translational Research in Oncology, Instituto do Câncer do Estado de São Paulo (ICESP), São Paulo (Brazil); Boccardo, Enrique [Department of Microbiology, Institute of Biomedical Sciences, University of São Paulo., São Paulo (Brazil); Villa, Luisa Lina [Department of Radiology and Oncology, Faculdade de Medicina, Universidade de São Paulo, São Paulo (Brazil); Rahal, Paula, E-mail: rahalp@yahoo.com.br [Department of Biology, Institute of Bioscience, Language and Exact Science, São Paulo State University, São Jose do Rio Preto (Brazil)

    2016-09-15

    Annexin 1 (ANXA1) is a substrate for E6AP mediated ubiquitylation. It has been hypothesized that HPV 16 E6 protein redirects E6AP away from ANXA1, increasing its stability and possibly contributing to viral pathogenesis. We analyzed ANXA1 expression in HPV-positive and negative cervical carcinoma-derived cells, in cells expressing HPV-16 oncogenes and in cells transduced with shRNA targeting E6AP. We observed that ANXA1 protein expression increased in HPV-16-positive tumor cells, in keratinocytes expressing HPV-16 E6wt (wild-type) or E6/E7 and C33 cells expressing HPV-16 E6wt. ANXA1 protein expression decreased in cells transfected with E6 Dicer-substrate RNAs (DsiRNA) and C33 cells cotransduced with HPV-16 E6wt and E6AP shRNA. Moreover, colony number and proliferation rate decreased in HPV16-positive cells transduced with ANXA1 shRNA. We observed that in cells infected with HPV16, the E6 binds to E6AP to degrade p53 and upregulate ANXA1. We suggest that ANXA1 may play a role in HPV-mediated carcinogenesis. - Highlights: • ANXA1 upregulation requires the presence of E6 and E6AP and is dependent on E6 integrity. • E6 binds to E6AP to degrade p53 and upregulate ANXA1 in cells infected with HPV16. • ANXA1 plays a role in cell proliferation in HPV-positive cervical cells.

  14. HPV16 E6 regulates annexin 1 (ANXA1) protein expression in cervical carcinoma cell lines

    International Nuclear Information System (INIS)

    Calmon, Marilia Freitas; Sichero, Laura; Boccardo, Enrique; Villa, Luisa Lina; Rahal, Paula

    2016-01-01

    Annexin 1 (ANXA1) is a substrate for E6AP mediated ubiquitylation. It has been hypothesized that HPV 16 E6 protein redirects E6AP away from ANXA1, increasing its stability and possibly contributing to viral pathogenesis. We analyzed ANXA1 expression in HPV-positive and negative cervical carcinoma-derived cells, in cells expressing HPV-16 oncogenes and in cells transduced with shRNA targeting E6AP. We observed that ANXA1 protein expression increased in HPV-16-positive tumor cells, in keratinocytes expressing HPV-16 E6wt (wild-type) or E6/E7 and C33 cells expressing HPV-16 E6wt. ANXA1 protein expression decreased in cells transfected with E6 Dicer-substrate RNAs (DsiRNA) and C33 cells cotransduced with HPV-16 E6wt and E6AP shRNA. Moreover, colony number and proliferation rate decreased in HPV16-positive cells transduced with ANXA1 shRNA. We observed that in cells infected with HPV16, the E6 binds to E6AP to degrade p53 and upregulate ANXA1. We suggest that ANXA1 may play a role in HPV-mediated carcinogenesis. - Highlights: • ANXA1 upregulation requires the presence of E6 and E6AP and is dependent on E6 integrity. • E6 binds to E6AP to degrade p53 and upregulate ANXA1 in cells infected with HPV16. • ANXA1 plays a role in cell proliferation in HPV-positive cervical cells.

  15. HPV-16 viral load in oropharyngeal squamous cell carcinoma using digital PCR.

    Science.gov (United States)

    Antonsson, Annika; Knight, Lani; Panizza, Benedict J; Porceddu, Sandro V; Emmett, Sarah; Whiteman, David C

    2018-05-09

    We did not identify any strong associations between HPV-16 viral load and any of the clinical or lifestyle factors. The epidemiology of oropharyngeal SCC is changing, with an increasing proportion of HPV-positive cases seen in the last decade. It is known that a high viral load is linked to the development of cervical cancer, the relation between viral load and oropharyngeal SCC is less clear. We sought to determine HPV-16 viral load in HPV-positive oropharyngeal SCCs using highly sensitive digital PCR and to identify clinical and lifestyle factors associated with viral load. We analysed 45 HPV-16 positive oropharyngeal SCCs diagnosed between 2013 and 2015. All patients completed a lifestyle questionnaire and clinical data were extracted from medical charts. Viral load was determined using digital PCR assays for HPV-L1 and RNAseP. We found large variations in HPV-16 viral load from 1 to 930 copies per cell (median 34 copies per cell).

  16. Reduction in HPV 16/18 prevalence in sexually active young women following the introduction of HPV immunisation in England.

    Science.gov (United States)

    Mesher, D; Soldan, K; Howell-Jones, R; Panwar, K; Manyenga, P; Jit, M; Beddows, S; Gill, O N

    2013-12-17

    Reduction in the prevalence of vaccine type HPV infection in young women is an early indication of the impact of the HPV immunisation programme and a necessary outcome if the subsequent impact on cervical cancer is to be realised. Residual vulva-vaginal swab (VVS) specimens from young women aged 16-24 years undergoing chlamydia screening in community sexual health services (formerly known as family planning clinics), general practice (GP), and youth clinics in 2010-2012 were submitted from 10 laboratories in seven regions around England. These specimens were linked to demographic and sexual behaviour data reported with the chlamydia test, anonymised, and tested for type-specific HPV DNA using a multiplex PCR and Luminex-based genotyping test. Estimated immunisation coverage was calculated and findings were compared to a baseline survey conducted prior to the introduction of HPV immunisation in 2008. A total of 4664 eligible specimens were collected and 4178 had a valid test result. The post-immunisation prevalence of HPV 16/18 infection was lowest in this youngest age group (16-18 years) and increased with age. This increase with age was a reversal of the pattern seen prior to immunisation and was inversely associated with estimates of age-specific immunisation coverage (65% for 16-18 year olds). The prevalence of HPV 16/18 infection in the post-immunisation survey was 6.5% amongst 16-18 year olds, compared to 19.1% in the similar survey conducted prior to the introduction of HPV immunisation. These findings are the first indication that the national HPV immunisation programme is successfully preventing HPV 16/18 infection in sexually active young women in England. The reductions seen suggest, for the estimated coverage, high vaccine effectiveness and some herd-protection benefits. Continued surveillance is needed to determine the effects of immunisation on non-vaccine HPV types. Crown Copyright © 2013. Published by Elsevier Ltd. All rights reserved.

  17. Expressions of HPV 16-E6 in Esophageal Carcinoma and its Clinical Significance

    Directory of Open Access Journals (Sweden)

    Xing Zhao

    2015-07-01

    Full Text Available Background and Objective: The role of (Human Papilloma Virus HPV in cancer of certain anatomical location, such as cervix, has been widely recognized. The present study was conducted to explore the association between HPV 16-E6 protein and esophageal squamous cell carcinoma. Methods: SP immunohistochemical method was used to examine the expression of HPV 16-E6 in 50 cases of esophageal squamous cell carcinoma, 10 cases of normal esophageal squamous cell and 10 cases of adjacent tissue. Results: The expression of HPV 16-E6 was significantly higher in esophageal carcinoma than in normal esophageal mucosa and in adjacent tissue. The expressions of HPV 16-E6 had significant correlation with invasive depth (P<0.05, but not with patient age, lymph node metastasis, tumor size (P>0.05. Conclusion: HPV 16-E6 can promote the growth and metastasis of esophageal squamous cell carcinoma and can be a prognostic factor of esophageal squamous cell carcinoma.DOI: http://dx.doi.org/10.3126/jcmsn.v10i4.12970 JCMS Nepal 2014; 10(4:1-5 

  18. Karyopherin β3: A new cellular target for the HPV-16 E5 oncoprotein

    International Nuclear Information System (INIS)

    Krawczyk, Ewa; Hanover, John A.; Schlegel, Richard; Suprynowicz, Frank A.

    2008-01-01

    Epidemiological and experimental studies have shown that high-risk human papillomaviruses (HPVs) are the causative agents of cervical cancer worldwide, and that HPV-16 is associated with more than half of these cases. In addition to the well-characterized E6 and E7 oncoproteins of HPV-16, recent evidence increasingly has implicated the HPV-16 E5 protein (16E5) as an important mediator of oncogenic transformation. Since 16E5 has no known intrinsic enzymatic activity, its effects on infected cells are most likely mediated by interactions with various cellular proteins and/or its documented association with lipid rafts. In the present study, we describe a new cellular target that binds to 16E5 in COS cells and in stable human ectocervical cell lines. This target is karyopherin β3, a member of the nuclear import receptor family with critical roles in the nuclear import of ribosomal proteins and in the secretory pathway

  19. E6D25E, HPV16 Asian variant shows specific proteomic pattern correlating in cells transformation and suppressive innate immune response

    International Nuclear Information System (INIS)

    Chopjitt, Peechanika; Pientong, Chamsai; Sunthamala, Nuchsupha; Kongyingyoes, Bunkerd; Haonon, Ornuma; Boonmars, Thidarut; Kikawa, Satomi; Nakahara, Tomomi; Kiyono, Tohru; Ekalaksananan, Tipaya

    2016-01-01

    HPV16 Asian variant (HPV16As) containing E6D25E oncogene, is commonly associated with cervical cancers of Asian populations. To explore a mechanism of E6D25E oncoprotein in carcinogenesis, we compared protein profiles in human keratinocytes expressing E6D25E with E6 of HPV16 prototype (E6Pro). A human cervical keratinocyte cell line, HCK1T, was transduced with retroviruses containing E6D25E or E6Pro genes. Biological properties of E6D25E or E6Pro transduced HCK1T cells were characterized. Protein profiles of the transduced HCK1T cells were analyzed using 2D-PAGE and characterized by mass spectrometry and western blotting. Reactomes of modulated proteins were analyzed by using the Reactome Knowledgebase. The E6D25E and E6Pro oncoproteins were comparable for their abilities to degrade p53 and suppress the induction of p21, and induce cell proliferation. Interestingly, the protein profiles of the HCK1T cells transduced with E6D25E showed specific proteomic patterns different from those with E6Pro. Among altered proteins, more than 1.5-fold up- or down- regulation was observed in E6D25E-expressing cells for gp96 and keratin7 which involved in activation of TLR signaling and transformation of squamocolumnar junction cells, respectively. This report describes new cellular proteins specifically targeted by E6D25E oncoprotein that may contribute to impair immune response against viral infection and cell transformation associated with oncogenic property of HPV16As variant. - Highlights: • E6D25E HPV16 specifically modulates protein profile of human keratinocytes. • E6D25E HPV16 modulates protein profile which involves in TLR signalling and transformation of squamocolumnar junction cells. • E6D25E oncoprotein may correlate to impair of immune response against viral infection and cells transformation.

  20. E6D25E, HPV16 Asian variant shows specific proteomic pattern correlating in cells transformation and suppressive innate immune response

    Energy Technology Data Exchange (ETDEWEB)

    Chopjitt, Peechanika; Pientong, Chamsai; Sunthamala, Nuchsupha [Department of Microbiology, Faculty of Medicine, Khon Kaen University, Khon Kaen, 40002 (Thailand); HPV & EBV and Carcinogenesis Research Group, Khon Kaen University (Thailand); Kongyingyoes, Bunkerd [Department of Pharmacology, Faculty of Medicine, Khon Kaen University, Khon Kaen, 40002 (Thailand); Haonon, Ornuma; Boonmars, Thidarut [Department of Parasitology, Faculty of Medicine, Khon Kaen University, Khon Kaen, 40002 (Thailand); Kikawa, Satomi; Nakahara, Tomomi [Division of Carcinogenesis and Cancer Prevention, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo, 104-0045 (Japan); Kiyono, Tohru, E-mail: tkiyono@ncc.go.jp [Division of Carcinogenesis and Cancer Prevention, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo, 104-0045 (Japan); Ekalaksananan, Tipaya, E-mail: tipeka@kku.ac.th [Department of Microbiology, Faculty of Medicine, Khon Kaen University, Khon Kaen, 40002 (Thailand); HPV & EBV and Carcinogenesis Research Group, Khon Kaen University (Thailand)

    2016-09-09

    HPV16 Asian variant (HPV16As) containing E6D25E oncogene, is commonly associated with cervical cancers of Asian populations. To explore a mechanism of E6D25E oncoprotein in carcinogenesis, we compared protein profiles in human keratinocytes expressing E6D25E with E6 of HPV16 prototype (E6Pro). A human cervical keratinocyte cell line, HCK1T, was transduced with retroviruses containing E6D25E or E6Pro genes. Biological properties of E6D25E or E6Pro transduced HCK1T cells were characterized. Protein profiles of the transduced HCK1T cells were analyzed using 2D-PAGE and characterized by mass spectrometry and western blotting. Reactomes of modulated proteins were analyzed by using the Reactome Knowledgebase. The E6D25E and E6Pro oncoproteins were comparable for their abilities to degrade p53 and suppress the induction of p21, and induce cell proliferation. Interestingly, the protein profiles of the HCK1T cells transduced with E6D25E showed specific proteomic patterns different from those with E6Pro. Among altered proteins, more than 1.5-fold up- or down- regulation was observed in E6D25E-expressing cells for gp96 and keratin7 which involved in activation of TLR signaling and transformation of squamocolumnar junction cells, respectively. This report describes new cellular proteins specifically targeted by E6D25E oncoprotein that may contribute to impair immune response against viral infection and cell transformation associated with oncogenic property of HPV16As variant. - Highlights: • E6D25E HPV16 specifically modulates protein profile of human keratinocytes. • E6D25E HPV16 modulates protein profile which involves in TLR signalling and transformation of squamocolumnar junction cells. • E6D25E oncoprotein may correlate to impair of immune response against viral infection and cells transformation.

  1. Significant difference in p53 and p21 protein immunoreactivity in HPV 16 positive and HPV negative breast carcinomas

    International Nuclear Information System (INIS)

    Hennig, E.M.; Norwegian Radium Hospital, Oslo; Kvinnsland, S.; Holm, R.; Nesland, J.M.

    1999-01-01

    Human papillomavirus (HPV) 16 has previously been found in 19/41 breast carcinomas (46%) in women with a history of HPV 16 positive CIN III lesions. There was no significant difference in distribution of histological subtypes, mean or median tumour diameter or number of regional lymph node metastases in the HPV positive and HPV negative breast carcinoma groups. P53, p21 and c-erbB-2 proteins were analyzed by immunohistochemistry in the HPV 16 positive and HPV negative breast carcinomas. There was a significant difference in p53 and p21 protein immunoreactivity between HPV 16 positive and HPV negative breast carcinomas (p=0.0091 and p=0.0040), with a significant less detectable p53 and p21 protein immunoreactivity in the HPV 16 positive cases. There was also a significant difference in the coexpression of p53/p21 between the HPV 16 positive and HPV 16 negative breast carcinomas (p=0.002). No significant difference in immunostaining for c-erbB-2 protein in the two groups was found (p=0.15), or for the coexpression of p53/c-erbB-2 (p=0.19). The significantly lower expression of p53 and p21 proteins in HPV 16 positive than in HPV 16 negative breast carcinomas supports the hypothesis of inactivation and degradation of wild-type p53 proteins by HPV 16 E6 and that p53 mutation is not necessary for transformation in the HPV 16 positive cases. (orig.)

  2. The Subcellular Localisation of the Human Papillomavirus (HPV 16 E7 Protein in Cervical Cancer Cells and Its Perturbation by RNA Aptamers

    Directory of Open Access Journals (Sweden)

    Özlem Cesur

    2015-06-01

    Full Text Available Human papillomavirus (HPV is the most common viral infection of the reproductive tract, affecting both men and women. High-risk oncogenic types are responsible for almost 90% of anogenital and oropharyngeal cancers including cervical cancer. Some of the HPV “early” genes, particularly E6 and E7, are known to act as oncogenes that promote tumour growth and malignant transformation. Most notably, HPV-16 E7 interacts with the tumour suppressor protein pRb, promoting its degradation, leading to cell cycle dysregulation in infected cells. We have previously shown that an RNA aptamer (termed A2 selectively binds to HPV16 E7 and is able to induce apoptosis in HPV16-transformed cervical carcinoma cell lines (SiHa through reduction of E7 levels. In this study, we investigated the effects of the A2 aptamer on E7 localisation in order to define its effects on E7 activity. We demonstrate for the first time that E7 localised to the plasma membrane. In addition, we show that A2 enhanced E7 localisation in the ER and that the A2-mediated reduction of E7 was not associated with proteasomal degradation. These data suggest that A2 perturbs normal E7 trafficking through promoting E7 ER retention.

  3. A non-pathogenic live vector as an efficient delivery system in vaccine design for the prevention of HPV16 E7-overexpressing cancers.

    Science.gov (United States)

    Hosseinzadeh, Sahar; Bolhassani, Azam; Rafati, Sima; Taheri, Tahereh; Zahedifard, Farnaz; Daemi, Amin; Taslimi, Yasaman; Hashemi, Mehrdad; Memarnejadian, Arash

    2013-01-01

    The attenuated or non-pathogenic live vectors have been evolved specifically to deliver DNA into cells as efficient delivery tools in gene therapy. Recently, a non-pathogenic protozoan, Leishmania tarentolae (L.tar) has attracted a great attention. In current study, we used Leishmania expression system (LEXSY) for stable expression of HPV16 E7 linked to different mini-chaperones [N-/C-terminal of gp96] and compared their immunogenicity and protective effects in C57BL/6 mice against TC-1 challenge. TC-1 murine model is primary C57BL/6 mice lung epithelial cells co-transformed with HPV16 E6, HPV16 E7 and ras oncogenes. Our results showed that subcutaneous administration of mice with both the recombinant L.tar-E7-NT (gp96) and L.tar-E7-CT (gp96) led to enhance the levels of IFN-γ and also IgG2a before and after challenge with TC-1. Furthermore, L.tar-E7-CT (gp96) live vaccine indicated significant protective effects as compared to control groups as well as group vaccinated with L.tar-E7. Indeed, the recombinant live vector is capable of eliciting effective humoral and cellular immune responses in mice, but however, further studies are required to increase their efficacy.

  4. Therapy of HPV 16-induced tumours with IL-2, IL-12 and genetically modified tumour vaccines

    Czech Academy of Sciences Publication Activity Database

    Bubeník, Jan; Mikyšková, Romana; Indrová, Marie; Vonka, V.; Šmahel, M.

    2002-01-01

    Roč. 98, Suppl. 13 (2002), s. P 1001 ISSN 0020-7136. [UICC International Cancer Congress /18./. 30.06.2002-05.07.2002, Oslo] Institutional research plan: CEZ:AV0Z5052915 Keywords : HPV 16 * interleukins * vaccine therapy Subject RIV: EB - Genetics ; Molecular Biology

  5. Genetically modified cellular vaccines for therapy of human papilloma virus type 16 (HPV 16)-associated tumours

    Czech Academy of Sciences Publication Activity Database

    Bubeník, Jan

    2008-01-01

    Roč. 8, č. 3 (2008), s. 180-186 ISSN 1568-0096 Grant - others:EU-FP6-NoE Clinigene(XE) 018933 Institutional research plan: CEZ:AV0Z50520514 Keywords : HPV 16 * genetically modified vaccines Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.316, year: 2008

  6. Ski protein levels increase during in vitro progression of HPV16-immortalized human keratinocytes and in cervical cancer

    International Nuclear Information System (INIS)

    Chen, Yi; Pirisi, Lucia; Creek, Kim E.

    2013-01-01

    We compared the levels of the Ski oncoprotein, an inhibitor of transforming growth factor-beta (TGF-β) signaling, in normal human keratinocytes (HKc), HPV16 immortalized HKc (HKc/HPV16), and differentiation resistant HKc/HPV16 (HKc/DR) in the absence and presence of TGF-β. Steady-state Ski protein levels increased in HKc/HPV16 and even further in HKc/DR, compared to HKc. TGF-β treatment of HKc, HKc/HPV16, and HKc/DR dramatically decreased Ski. TGF-β-induced Ski degradation was delayed in HKc/DR. Ski and phospho-Ski protein levels are cell cycle dependent with maximal Ski expression and localization to centrosomes and mitotic spindles during G2/M. ShRNA knock down of Ski in HKc/DR inhibited cell proliferation. More intense nuclear and cytoplasmic Ski staining and altered Ski localization were found in cervical cancer samples compared to adjacent normal tissue in a cervical cancer tissue array. Overall, these studies demonstrate altered Ski protein levels, degradation and localization in HPV16-transformed human keratinocytes and in cervical cancer. - Highlights: • Ski oncoprotein levels increase during progression of HPV16-transformed cells. • Ski and phospho-Ski protein levels are cell cycle dependent. • Ski knock-down in HPV16-transformed keratinocytes inhibited cell proliferation. • Cervical cancer samples overexpress Ski

  7. Skin reactions to human papillomavirus (HPV) 16 specific antigens intradermally injected in healthy subjects and patients with cervical neoplasia

    NARCIS (Netherlands)

    van den Hende, Muriel; van Poelgeest, Mariëtte I. E.; van der Hulst, Jeanette M.; de Jong, Joan; Drijfhout, Jan W.; Fleuren, Gert Jan; Valentijn, A. Rob P. M.; Wafelman, Amon R.; Slappendel, Gijs M.; Melief, Cornelis J. M.; Offringa, Rienk; van der Burg, Sjoerd H.; Kenter, Gemma G.

    2008-01-01

    We have tested the safety and feasibility of a synthetic long peptide-based HPV16-specific skin test to detect cellular immune responses to HPV16 E2, E6 and E7 in vivo. Women with cervical neoplasia (n = 11) and healthy individuals (n = 19) were intradermally challenged with 8 different pools of

  8. Análisis de variantes de HPV-16 como marcador molecular antropólogico

    Directory of Open Access Journals (Sweden)

    Badano, Ines

    2007-01-01

    Full Text Available El Virus Papiloma Humano tipo 16 (HPV16 es el principal responsable del desarrollo del cáncer de cuello uterino. Además de su significado clínico, el estudio de la variación genética en las regiones E6, L1 y LCR de este virus ha permitido identificar variantes específicas de diferentes áreas geográficas. Este descubrimiento sugiere una antigua propagación del HPV16 y su coevolución con el género humano. En este contexto, las variantes podrían servir como marcador molecular antropológico, aportando nuevos datos al análisis de patrones migratorios humanos. El objetivo del trabajo fue determinar las variantes de HPV16 en las regiones genéticas E6 y L1 que infectan mujeres guaraníes de Misiones. Para ello se analizaron 39 muestras de cepillados cervicales de mujeres guaraníes infectadas con HPV16. Las variantes en E6 y L1 se identificaron por PCR e hibridación en dot blot. Los resultados obtenidos fueron: el 77% de las variantes Europeas, 20% Africanas y 3% Asiático Americanas. La baja prevalencia de variantes Asiático Americanas coincide con lo reportado por Picconi y col, 2002 para mujeres quechuas, y haría suponer una limitada diseminación del HPV16 durante la época prehispánica. El predominio de las variantes Europeas podría ser resultado de la colonización española y la inmigración europea, mientras que el tráfico de esclavos negros explicaría la presencia de variantes Africanas. Por otra parte, la hipótesis de competencia viral tampoco puede ser descartada.

  9. HPV16-E7-Specific Activated CD8 T Cells in E7 Transgenic Skin and Skin Grafts

    Directory of Open Access Journals (Sweden)

    Seyed Davoud Jazayeri

    2017-05-01

    Full Text Available Human papillomavirus (HPV 16 E7 (E7 protein expression in skin promotes epithelial hyperproliferation and transformation to malignancy. Grafts of murine skin expressing E7 protein as a transgene in keratinocytes are not rejected from immunocompetent recipients, whereas grafts expressing ovalbumin (OVA, with or without coexpression of E7 protein, are promptly rejected, demonstrating that E7-associated non-antigen-specific local immunosuppression is not a major determinant of lack of rejection of E7 transgenic skin. To determine whether failure of rejection of E7 skin grafts is due to failure to attract E7-specific effector T cells, E7- and OVA-specific effector CD8+ T cells, activated in vitro, were transferred to animals bearing E7 transgenic skin grafts. Three days after T cell transfer, E7-specific T cells were present in significantly greater numbers than OVA-specific T cells in the grafted skin on animals bearing recently placed or healed E7 grafts, without graft rejection, and also in the ear skin of E7 transgenic animals, without obvious pathology. E7 and OVA-specific T cells were present in lesser numbers in healed E7 grafts than in recently placed grafts and in lesser numbers in recently placed E7 transgenic epidermal grafts without E7-associated hyperproliferation, derived from E7 transgenic mice with a mutated retinoblastoma gene. These data demonstrate that effector T cells are to some extent attracted to E7 transgenic skin specifically by E7 expression, but in large measure non-specifically by the epithelial proliferation associated with E7 expression, and by the local inflammation produced by grafting. Failure of E7 graft rejection was observed despite trafficking of E7-specific effector T cells to E7-expressing epithelium, a finding of consequence for immunotherapy of HPV 16 E7-associated human cancers.

  10. The epithelial-mesenchymal transition induced by keratinocyte growth conditions is overcome by E6 and E7 from HPV16, but not HPV8 and HPV38: Characterization of global transcription profiles

    International Nuclear Information System (INIS)

    Azzimonti, Barbara; Dell'Oste, Valentina; Borgogna, Cinzia; Mondini, Michele; Gugliesi, Francesca; De Andrea, Marco; Chiorino, Giovanna; Scatolini, Maria; Ghimenti, Chiara; Landolfo, Santo; Gariglio, Marisa

    2009-01-01

    The aim of this study was to evaluate the growth properties of primary human keratinocytes expressing E6 and E7 proteins, which are from either the β- or α-genotypes, under different culture conditions. We demonstrated that keratinocytes expressing E6 and E7, from both HPV8 and 38, irreversibly underwent the epithelial-mesenchymal transition (EMT) when grown on plastic with FAD medium (F12/DMEM/5%FBS). Expression of E6/E7 from HPV16 was capable of fully overcoming the FAD-induced EMT. Immortalization was only observed in HPV16-transduced cell lines, while the more proliferating phenotype of both KerHPV8 and 38 was mainly related to FAD-induced EMT. Microarray analysis of exponentially growing cells identified 146 cellular genes that were differentially regulated in HPV16 compared to HPV8- and 38-transduced cells. A large accumulation of transcripts associated with epidermal development and differentiation was observed in HPV16-transduced cells, whereas transcripts of genes involved in the extracellular matrix, multicellular organismal processes, and inflammatory response were affected in HPV8 and 38-transduced cells.

  11. Minimal residual disease after surgery of HPV 16-associated tumours as target for immunotherapy

    Czech Academy of Sciences Publication Activity Database

    Bubeník, Jan; Reiniš, Milan; Šímová, Jana

    2006-01-01

    Roč. 18, Supplement 1 (2006), - ISSN 1107-3756. [World Congress on Advances in Oncology /11./ and International Symposium on Molecular Medicine /9./. 12.10.2006-14.10.2006, Hersonissos] R&D Projects: GA MZd(CZ) NR7807; GA ČR(CZ) GA301/04/0492; GA AV ČR(CZ) IAA500520605 Institutional research plan: CEZ:AV0Z50520514 Keywords : minimal residual disease * HPV16 * immunotherapy Subject RIV: EB - Genetics ; Molecular Biology

  12. Tumour-inhibitory effects of dendritic cells administered at the site of HPV 16-induced neoplasms

    Czech Academy of Sciences Publication Activity Database

    Mendoza, Luis; Bubeník, Jan; Šímová, Jana; Korb, Jan; Bieblová, Jana; Vonka, V.; Indrová, Marie; Mikyšková, Romana; Jandlová, Táňa

    2002-01-01

    Roč. 48, č. 3 (2002), s. 114-119 ISSN 0015-5500 R&D Projects: GA MZd NC7148; GA ČR GA301/00/0114; GA AV ČR IAA7052002; GA AV ČR IAA5052203 Institutional research plan: CEZ:AV0Z5052915 Keywords : HPV 16 * dendritic cells * adjuvant therapy Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.615, year: 2002

  13. Depletion of Treg cells inhibits minimal residual disease after surgery of HPV16-associated tumours

    Czech Academy of Sciences Publication Activity Database

    Šímová, Jana; Bubeník, Jan; Bieblová, Jana; Rosalia, Rodney Alexander; Frič, Jan; Reiniš, Milan

    2006-01-01

    Roč. 29, č. 6 (2006), s. 1567-1571 ISSN 1019-6439 R&D Projects: GA ČR(CZ) GA301/04/0492 EU Projects: European Commission(XE) 18933 - CLINIGENE Grant - others:Liga proti rakovině(CZ) XX Institutional research plan: CEZ:AV0Z50520514 Keywords : HPV 16 * residual tumour disease * Treg cells Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.556, year: 2006

  14. Cytokines and dendritic cells as adjuvants for therapy of HPV16-associated tumours

    Czech Academy of Sciences Publication Activity Database

    Bubeník, Jan; Mikyšková, Romana; Reiniš, Milan; Mendoza, Luis; Indrová, Marie; Šmahel, M.; Vonka, V.

    2003-01-01

    Roč. 12, Supplement 1 (2003), s. S7 ISSN 1107-3756. [The 8th World Congress on Advances in Oncology and 6th Internationl Symposium on Molecular Medicine . Hernissos, Crete, 16.10.2003-18.10.2003] Institutional research plan: CEZ:AV0Z5052915 Keywords : HPV16 * dendritic cells * cytokines Subject RIV: FD - Oncology ; Hematology Impact factor: 1.940, year: 2003

  15. HPV16-associated tumours: Therapy of surgical minimal residual disease with dendritic cell-based vaccines

    Czech Academy of Sciences Publication Activity Database

    Reiniš, Milan; Indrová, Marie; Mendoza, Luis; Mikyšková, Romana; Bieblová, Jana; Bubeník, Jan; Šímová, Jana

    2004-01-01

    Roč. 25, č. 4 (2004), s. 1165-1170 ISSN 1019-6439 R&D Projects: GA MZd NC7148; GA ČR GA301/04/0492; GA ČR GA301/01/0985; GA AV ČR IAA5052203 Institutional research plan: CEZ:AV0Z5052915 Keywords : HPV16 * minimal residual tumour disease * dendritic cells Subject RIV: EC - Immunology Impact factor: 3.056, year: 2004

  16. Immunotherapeutic efficacy of vaccines generated by fusion of dendritic cells and HPV16-associated tumour cells

    Czech Academy of Sciences Publication Activity Database

    Bubeník, Jan; Šímová, Jana; Bieblová, Jana; Reiniš, Milan; Indrová, Marie

    2005-01-01

    Roč. 16, Suppl. 1 (2005), s. 101 ISSN 1107-3756. [World Congress on Advances in Oncology /10./ and International Symposium on Molecular Medicine /8./. 05.10.13-05.10.15, Hersonissos] R&D Projects: GA ČR(CZ) GA301/04/0492; GA MZd(CZ) NR8004 Institutional research plan: CEZ:AV0Z50520514 Keywords : HPV16 * dendritic cells * vaccines Subject RIV: EC - Immunology

  17. Immunotherapeutic efficacy of vaccines generated by fusion of dendritic cells and HPV16-associated tumour cells

    Czech Academy of Sciences Publication Activity Database

    Šímová, Jana; Bubeník, Jan; Bieblová, Jana; Indrová, Marie; Jandlová, Táňa

    2005-01-01

    Roč. 51, č. 1 (2005), s. 19-24 ISSN 0015-5500 R&D Projects: GA AV ČR(CZ) IAA5052203; GA ČR(CZ) GA301/04/0492 Grant - others:Liga proti rakovině, Praha(CZ) - Institutional research plan: CEZ:AV0Z50520514 Keywords : HPV16 * DC-tumour hybrids * immunotherapy Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.719, year: 2005

  18. Cervical intraepithelial neoplasia grade 2 or worse in Galicia, Spain: HPV 16 prevalence and vaccination impact.

    Science.gov (United States)

    Pérez-Castro, Sonia; Lorenzo-Mahía, Yolanda; Iñarrea Fernández, Amparo; Lamas-González, María José; Sarán-Díez, María Teresa; Rubio-Alarcón, Joaquín; Reboredo-Reboredo, María Consuelo; Mosteiro-Lobato, Sonia; López-Miragaya, Isabel; Torres-Piñón, Julio; Melón-García, Santiago

    2014-10-01

    The etiology of cervical intraepithelial neoplasia grade 2 or worse (CIN2+) can influence the efficacy of Public Health preventive strategies. This study aimed to determine the high-risk papillomavirus (HR-HPV) prevalence in CIN2+ cases in unvaccinated women in Galicia (Spain), the expected impact of bivalent vaccination, and the distribution of HPV 16 in squamous lesions. Ninety-four histologically confirmed cases of CIN2+ (2009-2010) were retrospectively studied: 23 CIN2, 58 CIN3- squamous carcinoma in situ (CIN3-CIS), 5 adenocarcinoma in situ (AIS), and 8 invasive squamous cervical cancer (SCC). Linear Array HPV Genotyping Test (Roche Diagnostics, Mannheim, Germany) was performed on the cervical specimens. Bivalent vaccination impact was calculated, based on regional vaccination coverage data, local HR-HPV prevalence, and reported efficacy (direct and cross-protection) of the vaccine. HR-HPV prevalence was 96.8%. The most frequent genotypes were HPV 16 (48.8-58.2%) and HPV 31 (9.3%-12.1%), considering single infections or single-multiple infections, respectively (hierarchical attribution). In squamous lesions, HPV 16 prevalence in women younger than 45 years of age increased in severe lesions (CIN3-CIS/SCC, OR 4.2), and was higher than in older women (OR 5.5). The vaccine could reduce the cumulative incidence of CIN2+ by 50.6% (direct protection), or by 62.7% (direct and cross-protection). HPV vaccination could have a great impact in women younger than 45 years of age due to the high prevalence of HPV 16 in their lesions. Copyright © 2013 Elsevier España, S.L.U. y Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

  19. HPV16/18 genotyping for the triage of HPV positive women in primary cervical cancer screening in Chile.

    Science.gov (United States)

    Lagos, Marcela; Van De Wyngard, Vanessa; Poggi, Helena; Cook, Paz; Viviani, Paola; Barriga, María Isabel; Pruyas, Martha; Ferreccio, Catterina

    2015-01-01

    We previously conducted a population-based screening trial of high-risk human papillomavirus (hrHPV) testing and conventional cytology, demonstrating higher sensitivity (92.7 % vs 22.1 % for CIN2+) but lower positive predictive value (10.5 % vs 23.9 %) of hrHPV testing. Here we report the performance of HPV16/18 genotyping to triage the hrHPV positive participants. Women aged 25 years and older received hrHPV (Hybrid Capture 2) and Papanicolaou testing; positives by either test underwent colposcopy and directed biopsy, as did a sample of double-negatives. hrHPV positive women were reflex-tested with HPV16/18 genotyping (Digene HPV Genotyping PS Test). Among the 8,265 participants, 10.7 % were hrHPV positive, 1.7 % had ASCUS+ cytology, 1.2 % had CIN2+; 776 (88 %) hrHPV positive women had complete results, of whom 38.8 % were positive for HPV16 (24.0 %), HPV18 (9.7 %) or both (5.1 %). CIN2+ prevalence in HPV16/18 positive women (16.3 %, 95 % CI 12.3-20.9) was twice that of HPV16/18 negative women (8.0 %, 95 % CI 5.7-10.8). HPV16/18 genotyping identified 40.5 % of CIN2, 66.7 % of CIN3 and 75.0 % of cancers. Compared to hrHPV screening alone, HPV16/18 triage significantly reduced the referral rate (10.7 % vs 3.7 %) and the number of colposcopies required to detect one CIN2+ (9 vs 6). When HPV16/18 negative women with baseline ASCUS+ cytology were also colposcopied, an additional 14 % of CIN2+ was identified; referral increased slightly to 4.2 %. HPV16/18 triage effectively stratified hrHPV positive women by their risk of high-grade lesions. HPV16/18 positive women must be referred immediately; referral could be deferred in HPV16/18 negative women given the slower progression of non-HPV16/18 lesions, however, they will require active follow-up.

  20. An Examination of HPV16 Natural Immunity in Men Who Have Sex with Men (MSM) in the HPV in Men (HIM) Study.

    Science.gov (United States)

    Beachler, Daniel C; Pinto, Ligia A; Kemp, Troy J; Nyitray, Alan G; Hildesheim, Allan; Viscidi, Raphael; Schussler, John; Kreimer, Aimée R; Giuliano, Anna R

    2018-04-01

    Background: Evidence suggests that natural antibodies developed after HPV16 infection may protect some women but not men against subsequent HPV16 reacquisition. Less is known whether antibodies developed following HPV16 infection are protective among men who have sex with men (MSM). Methods: Four hundred seventy-five MSM from the Human Papillomavirus Infection in Men (HIM) study were tested for serum antibodies to HPV16 L1 using enzyme-linked immunosorbent assays, and for anal and genital HPV16 DNA using PCR consensus primer system (PGMY 09/11). Adjusted Cox regression was used to evaluate whether baseline HPV16 seropositivity impacts subsequent genital or anal HPV16 DNA. Results: The risk of subsequent genital HPV16 [aHR = 1.05, 95% confidence interval (CI) = 0.66-1.68] and anal HPV16 infections among MSM (aHR = 2.34, 95% CI = 0.92-5.98) was similar or nonsignificantly higher in HPV16-seropositive than HPV16-seronegative MSM. The risk of genital HPV16 was also similar between HPV16-seronegative and HPV16-seropositive MSM in the highest tertile of HPV16 antibody levels and when restricting to those with new sex partners during follow-up ( P > 0.20). Among the 118 MSM who were HPV16 seropositive, 90% remained HPV16 seropositive up to 4 years later. When tested together, MSM with the highest antibody titers (top tertile) had similar levels to females (mean = 130.3 vs. 134.5 EU/mL, P = 0.84). Conclusions: Despite years of HPV16 seropositivity persistence and antibody titers comparable with females, this study suggested no evidence of HPV16 natural antibodies protecting against subsequent genital or anal HPV16 infection in MSM. Impact: This could help partially explain the high incidence of genital and anal HPV16 infection and related anal cancer seen in middle-aged and older MSM. Cancer Epidemiol Biomarkers Prev; 27(4); 496-502. ©2018 AACR . ©2018 American Association for Cancer Research.

  1. Radioimmunotherapy with an antibody to HPV16 E6 oncoprotein is effective in experimental cervical tumor expressing low levels of E6

    Science.gov (United States)

    Jiang, Zewei; Wang, Xing Guo; Einstein, Mark H; Goldberg, Gary L; Casadevall, Arturo

    2010-01-01

    Purpose HPV16 is associated with ∼50% of all cervical cancers worldwide. The E6 and E7 genes of oncogenic HPV types, such as HPV16, are necessary for the HPV transforming function and tumorogenesis making them ideal targets for novel treatments. Radioimmunotherapy employs systemically administered radiolabeled monoclonal antibodies (mAbs) that bind to tumor-associated antigens. Previously we demonstrated in mice that radioimmunotherapy targeting viral antigens with mAb to HPV16 E6 suppressed CasKi cervical tumors expressing high levels of E6 (∼600 copies of HPV per cell). However, that study opened the question whether radioimmunotherapy can suppress the growth of cervical tumors with low E6 and E7 expression, such as may be seen in patients. Experimental Design We evaluated the expression of E6 in patients' tumors and in the SiHa cell line expressing low levels of E6 and E7 (1–2 copies of HPV per cell) and found them comparable. We initiated SiHa tumors in nude mice, radiolabeled C1P5 mAb to E6 with a beta-emitter 188-Rhenium (188Re) and treated tumor-bearing mice with: (1) 200 µCi 188Re-C1P5 alone; (2) proteasome inhibitor MG132 alone; (3) MG132 followed by 200 µCi 188Re-C1P5; (4) unlabeled C1P5; (5) 200 µCi 188Re-18B7 (isotype-matching control mAb); (6) no treatment. 188Re-C1P5 alone and in combination with MG-132 significantly retarded tumor growth compared to all control groups. Conclusions Our data demonstrate the possibility to suppress tumor growth by targeting viral antigens even in cervical tumors with low E6 expression and provide additional evidence for the potential usefulness of radioimmunotherapy targeting HPV-related antigens in the clinic. PMID:20861673

  2. Local cytokine treatment of HPV16-associated tumours results in inhibition of their lung metastases

    Czech Academy of Sciences Publication Activity Database

    Mikyšková, Romana; Bubeník, Jan; Mendoza, Luis; Vonka, V.; Šmahel, M.; Šímová, Jana; Jandlová, Táňa

    2001-01-01

    Roč. 18, č. 7 (2001), s. 581-587 ISSN 0262-0898 R&D Projects: GA MZd NC5526; GA MZd NC5900; GA ČR GA312/99/0542; GA ČR GA301/01/0985; GA ČR GA301/00/0114; GA AV ČR IAA7052002 Institutional research plan: CEZ:AV0Z5052915 Keywords : HPV 16 * cytokines * lung metastasis Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.966, year: 2001

  3. Dendritic cell-based vaccines for therapy of HPV16-induced tumours

    Czech Academy of Sciences Publication Activity Database

    Bubeník, Jan; Šímová, Jana; Vonka, V.; Šmahel, M.; Mikyšková, Romana; Mendoza, Luis

    2001-01-01

    Roč. 495, - (2001), s. 359-363 ISSN 0065-2598 R&D Projects: GA MZd NC5526; GA ČR GA312/98/0826; GA ČR GA312/99/0542; GA ČR GA301/00/0114; GA ČR GA301/01/0985; GA AV ČR IAA7052002 Institutional research plan: CEZ:AV0Z5052915 Keywords : HPV16 * dendritic cell s * tumour vaccines Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.513, year: 2000

  4. The HPV-16 E7 oncoprotein induces centriole multiplication through deregulation of Polo-like kinase 4 expression

    Directory of Open Access Journals (Sweden)

    Duensing Stefan

    2011-05-01

    Full Text Available Abstract Background Infection with high-risk human papillomaviruses (HPVs such as HPV-16 is intimately associated with squamous cell carcinomas (SCCs of the anogenital tract and a subset of oropharyngeal carcinomas. Such lesions, including pre-invasive precursors, frequently show multipolar mitoses and aneuploidy. The high-risk HPV-16-encoded E7 oncoprotein has been shown to rapidly induce centrosome abnormalities thereby causing the formation of supernumerary mitotic spindle poles and increasing the risk for chromosome missegregation. HPV-16 E7 has been found to rapidly induce centriole overduplication, in part, through the simultaneous formation of more than one daughter centriole at single maternal centrioles (centriole multiplication. The precise molecular mechanism that underlies HPV-16 E7-induced centriole multiplication, however, remains poorly understood. Findings Here, we show that human keratinocytes engineered to stably express the HPV-16 E7 oncoprotein exhibit aberrant Polo-like kinase 4 (PLK4 protein expression at maternal centrioles. Real-time quantitative reverse transcriptase (qRT-PCR analysis of these cells revealed an increase of PLK4 mRNA levels compared to control cells. Importantly, the ability of the HPV-16 E7 oncoprotein to induce centriole multiplication was found to correlate with its ability to activate the PLK4 promoter and to up-regulate PLK4 mRNA. Conclusions These results highlight the critical role of PLK4 transcriptional deregulation in centriole multiplication in HPV-16 E7-expressing cells. Our findings encourage further experiments to test transcriptional inhibitors or small molecules targeting PLK4 to prevent centriole abnormalities, mitotic infidelity and malignant progression in HPV-associated neoplasms and other tumors in which PLK4 regulation is disrupted.

  5. Human papillomavirus detection with genotyping by the cobas and Aptima assays: Significant differences in HPV 16 detection?

    Science.gov (United States)

    Chorny, Joseph A; Frye, Teresa C; Fisher, Beth L; Remmers, Carol L

    2018-03-23

    The primary high-risk human papillomavirus (hrHPV) assays in the United States are the cobas (Roche) and the Aptima (Hologic). The cobas assay detects hrHPV by DNA analysis while the Aptima detects messenger RNA (mRNA) oncogenic transcripts. As the Aptima assay identifies oncogenic expression, it should have a lower rate of hrHPV and genotype detection. The Kaiser Permanente Regional Reference Laboratory in Denver, Colorado changed its hrHPV assay from the cobas to the Aptima assay. The rates of hrHPV detection and genotyping were compared over successive six-month periods. The overall hrHPV detection rates by the two platforms were similar (9.5% versus 9.1%) and not statistically different. For genotyping, the HPV 16 rate by the cobas was 1.6% and by the Aptima it was 1.1%. These differences were statistically different with the Aptima detecting nearly one-third less HPV 16 infections. With the HPV 18 and HPV 18/45, there was a slightly higher detection rate of HPV 18/45 by the Aptima platform (0.5% versus 0.9%) and this was statistically significant. While HPV 16 represents a low percentage of hrHPV infections, it was detected significantly less by the Aptima assay compared to the cobas assay. This has been previously reported, although not highlighted. Given the test methodologies, one would expect the Aptima to detect less HPV 16. This difference appears to be mainly due to a significantly increased number of non-oncogenic HPV 16 infections detected by the cobas test as there were no differences in HPV 16 detection rates in the high-grade squamous intraepithelial lesions indicating that the two tests have similar sensitivities for oncogenic HPV 16. © 2018 Wiley Periodicals, Inc.

  6. Retinoblastoma-independent antiproliferative activity of novel intracellular antibodies against the E7 oncoprotein in HPV 16-positive cells

    International Nuclear Information System (INIS)

    Accardi, Luisa; Tommasino, Massimo; Banks, Lawrence; Chirullo, Barbara; Giorgi, Colomba; Donà, Maria Gabriella; Mileo, Anna M; Paggi, Marco G; Federico, Antonio; Torreri, Paola; Petrucci, Tamara C; Accardi, Rosita; Pim, David

    2011-01-01

    'High risk' Human Papillomavirus strains are the causative agents of the vast majority of carcinomas of the uterine cervix. In these tumors, the physical integration of the HPV genome is a frequent, though not invariable occurrence, but the constitutive expression of the E6 and E7 viral genes is always observed, suggesting key roles for the E6 and E7 oncoproteins in the process of malignant transformation. The 'intracellular antibody' technology using recombinant antibodies in single-chain format offers the possibility of targeting a protein in its intracellular environment even at the level of definite domains thus representing a valuable strategy to 'knock out' the function of specific proteins. In this study, we investigate the in vitro activity of two single-chain antibody fragments directed against the 'high-risk' HPV 16 E7 oncoprotein, scFv 43M2 and scFv 51. These scFvs were expressed by retroviral system in different cell compartments of the HPV16-positive SiHa cells, and cell proliferation was analyzed by Colony Formation Assay and EZ4U assay. The binding of these scFvs to E7, and their possible interference with the interaction between E7 and its main target, the tumor suppressor pRb protein, were then investigated by immunoassays, PepSet™technology and Surface Plasmon Resonance. The expression of the two scFvs in the nucleus and the endoplasmic reticulum of SiHa cells resulted in the selective growth inhibition of these cells. Analysis of binding showed that both scFvs bind E7 via distinct but overlapping epitopes not corresponding to the pRb binding site. Nevertheless, the binding of scFv 43M2 to E7 was inhibited by pRb in a non-competitive manner. Based on the overall results, the observed inhibition of HPV-positive SiHa cells proliferation could be ascribed to an interaction between scFv and E7, involving non-pRb targets. The study paves the way for the employment of specific scFvs in immunotherapeutic

  7. Direct identification of an HPV-16 tumor antigen from cervical cancer biopsy specimens

    Directory of Open Access Journals (Sweden)

    Derin B Keskin

    2011-12-01

    Full Text Available Persistent infection with high-risk human papilloma viruses (HPV is the worldwide cause of many cancers, including cervical, anal, vulval, vaginal, penile and oropharyngeal. Since T cells naturally eliminate the majority of chronic HPV infections by recognizing epitopes displayed on virally altered epithelium, we exploited Poisson detection mass spectrometry (MS3 to identify those epitopes and inform future T cell-based vaccine design. Nine cervical cancer biopsies from HPV-16 positive HLA-A*02 patients were obtained, histopathology determined, and E7 oncogene PCR-amplified from tumor DNA and sequenced. Conservation of E7 oncogene coding segments was found in all tumors. MS3 analysis of HLA-A*02 immunoprecipitates detected E711-19 peptide (YMLDLQPET in seven of the nine tumor biopsies. The remaining two samples were E711-19 negative and lacked the HLA-A*02 binding GILT thioreductase peptide despite possessing binding-competent HLA-A*02 alleles. Thus, the conserved E711-19 peptide is a dominant HLA-A*02 binding tumor antigen in HPV-16 transformed cervical squamous and adenocarcinomas. Findings that a minority of HLA-A*02:01 tumors lack expression of both E711-19 and a peptide from a thioreductase important in processing of cysteine-rich proteins like E7 underscore the value of physical detection, define a potential additional tumor escape mechanism and have implications for therapeutic cancer vaccine development.

  8. Characterization of IgA response among women with incident HPV 16 infection

    International Nuclear Information System (INIS)

    Onda, Takashi; Carter, Joseph J.; Koutsky, Laura A.; Hughes, James P.; Lee, Shu-Kuang; Kuypers, Jane; Kiviat, Nancy; Galloway, Denise A.

    2003-01-01

    Previous studies have characterized the prevalence and duration of serum IgG antibodies to human papillomavirus type 16 (HPV 16) in a well-studied cohort of college women, using viruslike particle- (VLP) based ELISAs. In this study IgA antibodies in cervical secretions and sera were examined using a newly developed capsomer-based ELISA and the patterns observed for serum IgG, serum IgA, and cervical IgA antibodies were compared. The median time to antibody detection from the first detection of HPV 16 DNA was 10.5 months for IgA in cervical secretions and 19.1 months for serum IgA. Serum IgA antibody conversion was observed less frequently and occurred later than IgA conversion in cervical secretions (P = 0.011) or serum IgG conversion (P 0.051). The median time to antibody reversion, following seroconversion, was 12.0 months for IgA in cervical secretions and 13.6 months for serum IgA, whereas approximately 20% of women with serum IgG antibodies reverted within 36 months. Thus, the duration of IgA in cervical secretions and sera was shorter than the duration of serum IgG (P = 0.007 and 0.001)

  9. Investigating Reports of Complex Regional Pain Syndrome: An Analysis of HPV-16/18-Adjuvanted Vaccine Post-Licensure Data

    Science.gov (United States)

    Huygen, Frank; Verschueren, Kristin; McCabe, Candida; Stegmann, Jens-Ulrich; Zima, Julia; Mahaux, Olivia; Van Holle, Lionel; Angelo, Maria-Genalin

    2015-01-01

    Complex regional pain syndrome (CRPS) is a chronic pain disorder that typically follows trauma or surgery. Suspected CRPS reported after vaccination with human papillomavirus (HPV) vaccines led to temporary suspension of proactive recommendation of HPV vaccination in Japan. We investigated the potential CRPS signal in relation to HPV-16/18-adjuvanted vaccine (Cervarix®) by database review of CRPS cases with independent expert confirmation; a disproportionality analysis and analyses of temporality; an observed versus expected analysis using published background incidence rates; systematic reviews of aggregate safety data, and a literature review. The analysis included 17 case reports of CRPS: 10 from Japan (0.14/100,000 doses distributed) and seven from the United Kingdom (0.08/100,000). Five cases were considered by independent experts to be confirmed CRPS. Quantitative analyses did not suggest an association between CRPS and HPV-16/18-adjuvanted vaccine. Observed CRPS incidence after HPV-16/18 vaccination was statistically significantly below expected rates. Systematic database reviews using search terms varying in specificity and sensitivity did not identify new cases. No CRPS was reported during clinical development and no unexpected results found in the literature. There is not sufficient evidence to suggest an increased risk of developing CRPS following vaccination with HPV-16/18-adjuvanted vaccine. Post-licensure safety surveillance confirms the acceptable benefit-risk of HPV-16/18 vaccination. PMID:26501109

  10. Augmentation of cellular and humoral immune responses to HPV16 and HPV18 E6 and E7 antigens by VGX-3100

    Directory of Open Access Journals (Sweden)

    Matthew P Morrow

    2016-01-01

    Full Text Available We have previously demonstrated the immunogenicity of VGX-3100, a multicomponent DNA immunotherapy for the treatment of Human Papillomavirus (HPV16/18-positive CIN2/3 in a phase 1 clinical trial. Here, we report on the ability to boost immune responses with an additional dose of VGX-3100. Patients completing our initial phase 1 trial were offered enrollment into a follow on trial consisting of a single boost dose of VGX-3100. Data show both cellular and humoral immune responses could be augmented above pre-boost levels, including the induction of interferon (IFNγ production, tumor necrosis factor (TNFα production, CD8+ T cell activation and the synthesis of lytic proteins. Moreover, observation of antigen-specific regulation of immune-related gene transcripts suggests the induction of a proinflammatory response following the boost. Analysis of T cell receptor (TCR sequencing suggests the localization of putative HPV-specific T cell clones to the cervical mucosa, which underscores the putative mechanism of action of lesion regression and HPV16/18 elimination noted in our double-blind placebo-controlled phase 2B trial. Taken together, these data indicate that VGX-3100 drives the induction of robust cellular and humoral immune responses that can be augmented by a fourth “booster” dose. These data could be important in the scope of increasing the clinical efficacy rate of VGX-3100.

  11. A novel trivalent HPV 16/18/58 vaccine with anti-HPV 16 and 18 neutralizing antibody responses comparable to those induced by the Gardasil quadrivalent vaccine in rhesus macaque model

    Directory of Open Access Journals (Sweden)

    Fei Yin

    2017-06-01

    Full Text Available Persistent infection with human papillomavirus (HPV is a key factor in the development of precancerous lesions and invasive cervical cancer. Prophylactic vaccines to immunize against HPV are an effective approach to reducing HPV related disease burden. In this study, we investigated the immunogenicity and dosage effect of a trivalent HPV 16/18/58 vaccine (3vHPV produced in Escherichia coli (E.coli, with Gardasil quadrivalent vaccine (4vHPV, Merck & Co. as a positive control. Sera collected from rhesus macaques vaccinated with three dosage formulations of 3vHPV (termed low-, mid-, and high-dosage formulations, respectively, and the 4vHPV vaccine were analyzed by both Pseudovirus-Based Neutralization Assay (PBNA and Enzyme-Linked Immunosorbent Assay (ELISA. Strong immune responses against HPV 16/18/58 were successfully elicited, and dosage-dependence was observed, with likely occurrence of immune interference between different L1-VLP antigens. HPV 16/18 specific neutralizing antibody (nAb and total immunoglobulin G (IgG antibody responses in rhesus macaques receiving 3vHPV at the three dosages tested were generally non-inferior to those observed in rhesus macaques receiving 4vHPV throughout the study period. Particularly, HPV 18 nAb titers induced by the mid-dosage formulation that contained the same amounts of HPV 16/18 L1-VLPs as Gardasil 4vHPV were between 7.3 to 12.7-fold higher compared to the positive control arm from weeks 24–64. The durability of antibody responses specific to HPV 16/18 elicited by 3vHPV vaccines was also shown to be non-inferior to that associated with Gardasil 4vHPV. Keywords: Human papillomavirus, HPV 16/18/58, GMTs, Trivalent, Immunogenicity

  12. A novel HPV 16 L1-based chimeric virus-like particle containing E6 and E7 seroreactive epitopes permits highly specific detection of antibodies in patients with CIN 1 and HPV-16 infection

    Directory of Open Access Journals (Sweden)

    Santiago-Osorio Edelmiro

    2011-02-01

    Full Text Available Abstract Background The presence of IgG antibodies to HPV-16 L1-virus like particles (VLPs in serum has been reported as a result of persistent exposure to the virus and as a marker of disease progression. However, detection of VLP-specific antibodies in sera does not always indicate a malignant lesion as positive results may also be due to a nonmalignant viral infection. Furthermore, malignant lesions are associated with an increased antibody titer for E6 and E7 proteins. The aim of this study was to develop an ELISA using a novel chimeric virus-like particle (cVLP encoding an L1 protein fused with a string of HPV-16 E6 and E7 seroreactive epitopes to its C-terminus to be used for detection of HPV-16 specific antibodies in patients with cervical intraepithelial lesion grade 1 (CIN 1. Results The sera of 30 patients with CIN 1 who also tested positive for HPV-16 DNA and of 30 age-matched normal donors negative for HPV infection were tested for the presence of IgG antibodies specific for either VLP-L1 (HPV-16 L1, gVLP (derived from Gardasil, or cVLP by ELISA. The cVLP-reactive sera yielded two distinct groups of results: (H reactivity levels that presented very strong cVLP-specific titers, and (L reactivity levels with significantly lower titers similar to those obtained with VLP-L1 and gVLP antigens. Additionally, the sera that presented the higher cVLP titers closely matched those that had significantly stronger reactivity to E6 and E7 epitopes. Interestingly, the samples with the highest titers corresponded to patients with the higher numbers of sexual partners and pregnancies. On the other hand only 4 out of the 12 sera that harbored antibodies with VLP neutralizing ability corresponded to the group with high cVLP antibody titers. Conclusion We report for the first time that chimeric particles containing HPV-16 L1 protein fused with E6 and E7 seroreactive epitopes enable much better detection of IgG antibodies in the sera of CIN 1 patients

  13. HPV16 RNA patterns defined by novel high-throughput RT-qPCR as triage marker in HPV-based cervical cancer precursor screening.

    Science.gov (United States)

    Höfler, Daniela; Böhmer, Gerd; von Wasielewski, Reinhard; Neumann, Heinrich; Halec, Gordana; Holzinger, Dana; Dondog, Bolormaa; Gissmann, Lutz; Pawlita, Michael; Schmitt, Markus

    2015-09-01

    Cervical cancer precursor screening by HPV testing has a low positive predictive value for advanced lesion. HPV16 RNA patterns characteristic for HPV16-transformed cells but based on laborious, cost-intensive singleplex NASBA reactions promised high value in triaging HPV16 DNA-positive women. We developed two high-throughput reverse transcriptase quantitative (RT-q) PCR assays for the HPV16 transcripts E6*I, E1^E4 and E1C and the cellular transcript ubiquitin C and analysed RNA of 158 singly HPV16 DNA-positive cervical cell samples archived in PreservCyt buffer for the presence of transformation-associated HPV16 RNA patterns, i.e., upregulation of E6*I relative to E1^E4 and/or presence of E1C. HPV16 RNA pattern analyses classified 85% of 58 samples diagnosed ≤CIN1 (no cytologically and histologically detectable cervical lesion or CIN grade 1) as negative and 90% of 59 samples diagnosed as ≥CIN3 (CIN grade 3 or invasive cancer) as positive. Among 41 CIN grade 2 samples representing an intermediate lesion group, 49% were HPV16 RNA patterns-positive. Interestingly, 3 of 4 HPV16 RNA patterns-positive lesions initially diagnosed as ≤CIN1 at follow-up 5-24 months later had progressed to ≥CIN2. We successfully developed and validated a second generation of HPV16 RNA patterns assay by rapid RT-qPCR as triage marker for HPV16 DNA-positive women offering clinical utility to distinguish between the need for immediate colposcopy and continued observation. Limited follow-up data suggests that HPV16 RNA patterns-positivity in ≤CIN1 lesions can predict disease progression. Copyright © 2015 Elsevier Inc. All rights reserved.

  14. Strategies for continuous evaluation of the benefit-risk profile of HPV-16/18-AS04-adjuvanted vaccine.

    Science.gov (United States)

    Angelo, Maria-Genalin; Taylor, Sylvia; Struyf, Frank; Tavares Da Silva, Fernanda; Arellano, Felix; David, Marie-Pierre; Dubin, Gary; Rosillon, Dominique; Baril, Laurence

    2014-11-01

    The HPV types 16/18-AS04-adjuvanted cervical cancer vaccine, Cervarix(®) (HPV-16/18-vaccine, GlaxoSmithKline, Belgium) was first approved in 2007 and is licensed in 134 countries for the prevention of persistent infection, premalignant cervical lesions and cervical cancer caused by oncogenic HPV. Benefit-risk status requires continual re-evaluation as vaccine uptake increases, as the epidemiology of the disease evolves and as new information becomes available. This paper provides an example of benefit-risk considerations and risk-management planning. Evaluation of the benefit-risk of HPV-16/18-vaccine post-licensure includes studies with a range of designs in many countries and in collaboration with national public agencies and regulatory authorities. The strategy to assess benefit versus risk will continue to evolve and adapt to the changing HPV-16/18-vaccine market.

  15. Health and economic outcomes of HPV 16,18 vaccination in 72 GAVI-eligible countries.

    Science.gov (United States)

    Goldie, Sue J; O'Shea, Meredith; Campos, Nicole Gastineau; Diaz, Mireia; Sweet, Steven; Kim, Sun-Young

    2008-07-29

    The risk of dying from cervical cancer is disproportionately borne by women in developing countries. Two new vaccines are highly effective in preventing HPV 16,18 infection, responsible for approximately 70% of cervical cancer, in girls not previously infected. The GAVI Alliance (GAVI) provides technical assistance and financial support for immunization in the world's poorest countries. Using population-based and epidemiologic data for 72 GAVI-eligible countries we estimate averted cervical cancer cases and deaths, disability-adjusted years of life (DALYs) averted and incremental cost-effectiveness ratios (I$/DALY averted) associated with HPV 16,18 vaccination of young adolescent girls. In addition to vaccine coverage and efficacy, relative and absolute cancer reduction depended on underlying incidence, proportion attributable to HPV types 16 and 18, population age-structure and competing mortality. With 70% coverage, mean reduction in the lifetime risk of cancer is below 40% in some countries (e.g., Nigeria, Ghana) and above 50% in others (e.g., India, Uganda, Kenya). At I$10 per vaccinated girl (approximately $2.00 per dose assuming three doses, plus wastage, administration, program support) vaccination was cost-effective in all countries using a per capita GDP threshold; for 49 of 72 countries, the cost per DALY averted was less than I$100 and for 59 countries, it was less than I$200. Taking into account country-specific assumptions (per capita GNI, DPT3 coverage, percentage of girls who are enrolled in fifth grade) for the year of introduction, percent coverage achieved in the first year, and years to maximum coverage, a 10-year modeled scenario prevented the future deaths of approximately 2 million women vaccinated as adolescents. Despite favorable cost-effectiveness, assessment of financial costs raised concerns about affordability; as the cost per vaccinated girl was increased from I$10 to I$25 (approximately $2 to $5 per dose), the financial costs for the

  16. Curcumin modulates cellular AP-1, NF-kB, and HPV16 E6 proteins in oral cancer.

    Science.gov (United States)

    Mishra, Alok; Kumar, Rakesh; Tyagi, Abhishek; Kohaar, Indu; Hedau, Suresh; Bharti, Alok C; Sarker, Subhodeep; Dey, Dipankar; Saluja, Daman; Das, Bhudev

    2015-01-01

    In this study, we investigated the effects of the natural antioxidant curcumin on the HPV16-positive oral carcinoma cell line 93VU147T and demonstrated that curcumin is not only a potent inhibitor for the activity of host nuclear transcription factors AP-1 and NF-kB but it also selectively suppresses transcription of the HPV16/E6 oncogene during the carcinogenic process in oral cancer cells. This study suggests a therapeutic potential of curcumin for high-risk human papilloma virus (HPV)-infected oral cancers.

  17. Correlation between levels of human papillomavirus (HPV)-16 and 18 antibodies in serum and cervicovaginal secretions in girls and women vaccinated with the HPV-16/18 AS04-adjuvanted vaccine

    DEFF Research Database (Denmark)

    Schwarz, Tino F; Kocken, Mariëlle; Petäjä, Tiina

    2010-01-01

    and CVS samples were collected from a subset of women aged 10-65 years (N=350) at pre-specified time-points from 7 to 36 months post-vaccination. Anti-HPV-16/18 antibody levels in serum and CVS were measured by enzyme-linked immunosorbent assay. Pearson correlation coefficients between serum and CVS...... in serum were substantially higher at all time-points than those in a control group of women who had cleared a natural HPV infection in another trial. In women with detectable antibodies in both serum and CVS, good correlation was seen between HPV-16/18 antibody levels at all time-points (Pearson......This pooled analysis of data from four Phase III clinical trials was undertaken to assess the correlation between levels of anti-human papillomavirus (HPV)-16/18 antibodies in serum and cervicovaginal secretions (CVS) in girls and women vaccinated with the HPV-16/18 AS04-adjuvanted vaccine. Serum...

  18. Human papillomavirus genotypes and phylogenetic analysis of HPV-16 variants in HIV-1 infected subjects in Italy.

    Science.gov (United States)

    Tanzi, Elisabetta; Amendola, Antonella; Bianchi, Silvia; Fasolo, M Michela; Beretta, Rosangela; Pariani, Elena; Zappa, Alessandra; Frati, Elena; Orlando, Giovanna

    2009-05-29

    A cross-sectional study was carried out to improve the state of evidence regarding the spectrum of HPV types and HPV-16 LCR variants circulating among men and women infected with HIV-1 in Italy. This study, conducted in 518 HIV-positive subjects (346 males and 172 females), showed a high prevalence of HPV anal infections (88.7%) in men and of cervical infections (65.1%) in women. A wide spectrum of HPV genotypes has been observed, as both single and multiple infections. Low-risk HPV types 6, 11 and 61 were frequently detected. HPV-16 was the prevalent high-risk type. Fourteen different HPV-16 LCR variants were found. Ten belonged to the European lineage (78.7% were detected in Italian subjects and 21.3% in foreign-born, all homo/bisexual men), two to the Asiatic lineage and two to the African-2 lineage. This study underlines the great genotypic heterogeneity characterizing anal and cervical HPV infections and the marked polymorphism of the predominant HPV-16 in this high-risk population in Italy.

  19. Inhibitory effects of unmethylated CpG oligodeoxynucleotides on MHC class I-deficient and -proficient HPV16-associated tumours

    Czech Academy of Sciences Publication Activity Database

    Reiniš, Milan; Šímová, Jana; Bubeník, Jan

    2006-01-01

    Roč. 118, č. 7 (2006), s. 1836-1842 ISSN 0020-7136 R&D Projects: GA ČR(CZ) GA301/04/0492 Institutional research plan: CEZ:AV0Z50520514 Keywords : HPV16 * immunotherapy * CpG oligodeoxynucleotides Subject RIV: EC - Immunology Impact factor: 4.693, year: 2006

  20. DNA methylation regulated microRNAs in HPV-16-induced head and neck squamous cell carcinoma (HNSCC).

    Science.gov (United States)

    Sannigrahi, M K; Sharma, Rajni; Singh, Varinder; Panda, Naresh K; Rattan, Vidya; Khullar, Madhu

    2018-02-17

    Epigenetic modifications have been reported to play an important role in regulating gene expression and these modifications become critical when they have a role in controlling another important layer of epigenetic regulation namely microRNAs. In the present study, we have identified the microRNAs that may be regulated by promoter DNA methylation and histone acetylation in Human papilloma virus-positive head and neck squamous cell carcinoma. HPV-negative cell line (UPCI:SCC-116) and HPV-16 +ve cell line (UPCI:SCC-090) were treated with methylation inhibitor (5-aza-2'-deoxycytidine, AZA) and acetylation inhibitor (Trichostatin-A, TSA), followed by micro-array analysis. The differentially expressed miRNAs were validated in control (n = 10), HPV-16 +ve (n = 30), and HPV -ve (n = 30) HNC, TCGA (n = 529) tissue samples, and two HPV -ve (SCC116 and Hacat) and two HPV +ve (SCC090 and SiHa) cell lines. Methylation-specific PCR (MSP) and chromatin immunoprecipitation assay (CHIP) were performed to validate their regulation. In silico and in vitro analyses of identified miRNAs were done to study putative pathways they target and their possible role in carcinogenesis. Among 10 miRNAs specifically up-regulated in microarray analysis of AZA-treated SCC090 cells, we observed significantly decreased expression of hsa-miR-181c-5p, hsa-miR-132-5p, hsa-miR-658 in HPV +ve HNC cohort, TCGA tissue samples, and cell lines as compared to their HPV -ve counterpart, and their promoter region also possesses CpG islands. MSP and analysis of TCGA data (MethHC) revealed increased frequency of methylation at the promoter of hsa-miR-132-5p that is negatively correlated with its expression. In TSA-treated SCC090 cells, out of 7 miRNAs, two namely Hsa-miR-129-2-3p and Hsa-miR-449a were found to be up-regulated as compared to HPV -ve cells. However, the levels of enrichment by anti-acetyl-H3 and anti-acetyl-H4 were significantly low in cell lines compared to respective controls

  1. Success or failure of vaccination for HPV16-positive vulvar lesions correlates with kinetics and phenotype of induced T-cell responses

    NARCIS (Netherlands)

    Welters, Marij J. P.; Kenter, Gemma G.; de Vos van Steenwijk, Peggy J.; Löwik, Margriet J. G.; Berends-van der Meer, Dorien M. A.; Essahsah, Farah; Stynenbosch, Linda F. M.; Vloon, Annelies P. G.; Ramwadhdoebe, Tamara H.; Piersma, Sytse J.; van der Hulst, Jeanette M.; Valentijn, A. Rob P. M.; Fathers, Lorraine M.; Drijfhout, Jan W.; Franken, Kees L. M. C.; Oostendorp, Jaap; Fleuren, Gert Jan; Melief, Cornelis J. M.; van der Burg, Sjoerd H.

    2010-01-01

    One half of a group of 20 patients with human papillomavirus type 16 (HPV16)-induced vulvar intraepithelial neoplasia grade 3 displayed a complete regression (CR) after therapeutic vaccination with HPV16 E6/E7 synthetic long peptides. Patients with relatively larger lesions generally did not display

  2. Integration of Oncogenes via Sleeping Beauty as a Mouse Model of HPV16+ Oral Tumors and Immunologic Control.

    Science.gov (United States)

    Lin, Yi-Hsin; Yang, Ming-Chieh; Tseng, Ssu-Hsueh; Jiang, Rosie; Yang, Andrew; Farmer, Emily; Peng, Shiwen; Henkle, Talia; Chang, Yung-Nien; Hung, Chien-Fu; Wu, T-C

    2018-01-23

    Human papillomavirus type 16 (HPV16) is the etiologic factor for cervical cancer and a subset of oropharyngeal cancers. Although several prophylactic HPV vaccines are available, no effective therapeutic strategies to control active HPV diseases exist. Tumor implantation models are traditionally used to study HPV-associated buccal tumors. However, they fail to address precancerous phases of disease progression and display tumor microenvironments distinct from those observed in patients. Previously, K14-E6/E7 transgenic mouse models have been used to generate spontaneous tumors. However, the rate of tumor formation is inconsistent, and the host often develops immune tolerance to the viral oncoproteins. We developed a preclinical, spontaneous, HPV16 + buccal tumor model using submucosal injection of oncogenic plasmids expressing HPV16-E6/E7, NRas G12V , luciferase, and sleeping beauty (SB) transposase, followed by electroporation in the buccal mucosa. We evaluated responses to immunization with a pNGVL4a-CRT/E7(detox) therapeutic HPV DNA vaccine and tumor cell migration to distant locations. Mice transfected with plasmids encoding HPV16-E6/E7, NRas G12V , luciferase, and SB transposase developed tumors within 3 weeks. We also found transient anti-CD3 administration is required to generate tumors in immunocompetent mice. Bioluminescence signals from luciferase correlated strongly with tumor growth, and tumors expressed HPV16-associated markers. We showed that pNGVL4a-CRT/E7(detox) administration resulted in antitumor immunity in tumor-bearing mice. Lastly, we demonstrated that the generated tumor could migrate to tumor-draining lymph nodes. Our model provides an efficient method to induce spontaneous HPV + tumor formation, which can be used to identify effective therapeutic interventions, analyze tumor migration, and conduct tumor biology research. Cancer Immunol Res; 6(3); 1-15. ©2018 AACR. ©2018 American Association for Cancer Research.

  3. Human papillomavirus (HPV vaccination for the prevention of HPV 16/18 induced cervical cancer and its precursors

    Directory of Open Access Journals (Sweden)

    Greiner, Wolfgang

    2009-03-01

    Full Text Available Introduction: Essential precondition for the development of cervical cancer is a persistent human papillomavirus (HPV infection. The majority - approximately 70% - of cervical carcinomas is caused by two high-risk HPV types (16 and 18. Recently, two vaccines have been approved to the German market with the potential to induce protection against HPV 16 and HPV 18 among additional low-risk virus types. Objectives: To analyse whether HPV vaccination is effective with regard to the reduction of cervical cancer and precursors of cervical carcinoma (CIN, respectively? Does HPV vaccination represent a cost-effective alternative or supplement to present screening practice? Are there any differences concerning cost-effectiveness between the two available vaccines? Should HPV vaccination be recommended from a health economic point of view? If so, which recommendations can be conveyed with respect to a (reorganization of the German vaccination strategy? Which ethical, social and legal implications have to be considered? Methods: Based on a systematic literature review, randomized controlled trials (RCT looking at the effectiveness of HPV vaccination for the prevention of cervical carcinoma and its precursors - cervical intraepithelial neoplasia - have been identified. In addition, health economic models were identified to address the health economic research questions. Quality assessment of medical and economic literature was assured by application of general assessment standards for the systematic and critical appraisal of scientific studies. Results: Vaccine efficacy in prevention of CIN 2 or higher lesions in HPV 16 or HPV 18 negative women, who received all vaccination doses, ranges between 98% and 100%. Side effects of the vaccination are mainly associated with injection site reactions (redness, turgor, pain. No significant differences concerning serious complications between the vaccination- and the placebo-groups were reported. Results of base case

  4. Prevalência dos HPV 16, 18, 45 e 31 em mulheres com lesão cervical Prevalence of HPV 16, 18, 45 and 31 in women with cervical lesions

    Directory of Open Access Journals (Sweden)

    Denise Rocha Pitta

    2010-07-01

    Full Text Available OBJETIVO: avaliar a prevalência dos HPV 16, 18, 31 e 45 em amostras de raspado cervical de mulheres com alterações celulares e/ou colposcopia sugestiva de lesão de alto grau ou lesão de baixo grau persistente submetidas à conização. MÉTODOS: Foram incluídas 120 mulheres. A análise histológica dos cones cervicais revelou 7 casos de cervicite, 22 de NIC1, 31 de NIC2, 54 de NIC3 e 6 carcinomas invasores. Foram analisadas as amostras de raspado cervical coletadas antes da conização para a presença do DNA-HPV por PCR com os primers de consenso, PGMY09/11. As amostras positivas para DNA de HPV foram testadas para presença do HPV16, 18, 31 e 45 utilizando-se primers tipo específico para esses HPV. RESULTADOS: O DNA-HPV foi detectado em 67,5% das mulheres. O HPV 16 (40% foi o tipo mais prevalente na maioria das lesões, seguido dos HPV 31 (13,3%, 45 (13,3% e 18 (4,1%. Infecções múltiplas ocorreram em 15% dos casos e as infecções por outros tipos de HPV foram detectadas em 14% da amostra. CONCLUSÕES: as infecções pelos HPV 16 e 18 nem sempre ocorrem de maneira solitária (infecção única, estando associadas a outros tipos de HPV em diversas ocasiões.PURPOSE: to determine the prevalence of HPV 16, 18, 31 and 45 in cervical screening samples of women with cellular changes and/or colposcopy suggestive of persistent high grade or low grade lesion who were submitted to conization. METHODS: a total of 120 women were included in the study. Histological analysis of the cervical cones revealed 7 cases of cervicitis, 22 of CIN1, 31 of CIN2, 54 of CIN3, and 6 invasive carcinomas. The cervical screening samples were analyzed before conization for the presence of HPV-DNA by PCR using the consensus primers PGMY09/11. HPV-DNA-positive samples were tested for the presence of HPV16, 18, 31 and 45 using type-specific primers for these HPV. RESULTS: HPV-DNA was detected in 67.5% of the studied women. HPV 16 (40% was the most prevalent type in

  5. Nuclear import of high risk HPV16 E7 oncoprotein is mediated by its zinc-binding domain via hydrophobic interactions with Nup62

    Energy Technology Data Exchange (ETDEWEB)

    Eberhard, Jeremy; Onder, Zeynep; Moroianu, Junona, E-mail: moroianu@bc.edu

    2013-11-15

    We previously discovered that nuclear import of high risk HPV16 E7 is mediated by a cNLS located within the zinc-binding domain via a pathway that is independent of karyopherins/importins (Angeline et al., 2003; Knapp et al., 2009). In this study we continued our characterization of the cNLS and nuclear import pathway of HPV16 E7. We find that an intact zinc-binding domain is essential for the cNLS function in mediating nuclear import of HPV16 E7. Mutagenesis of cysteine residues to alanine in each of the two CysXXCys motifs involved in zinc-binding changes the nuclear localization of the EGFP-16E7 and 2xEGFP-16E7 mutants. We further discover that a patch of hydrophobic residues, {sub 65}LRLCV{sub 69}, within the zinc-binding domain of HPV16 E7 mediates its nuclear import via hydrophobic interactions with the FG domain of the central channel nucleoporin Nup62. - Highlights: • An intact zinc-binding domain is essential for the nuclear localization of HPV16 E7. • Identification of a hydrophobic patch that is critical for the nuclear import of HPV16 E7. • HPV16 E7 interacts via its zinc-binding domain with the FG domain of Nup62.

  6. Epithelium Expressing the E7 Oncoprotein of HPV16 Attracts Immune-Modulatory Dendritic Cells to the Skin and Suppresses Their Antigen-Processing Capacity.

    Directory of Open Access Journals (Sweden)

    Janin Chandra

    Full Text Available Antigen presenting cells (APCs in skin can promote either antigen-specific effector functions or antigen tolerance, and thus determine clearance or persistence of cutaneous viral infections. Human papillomavirus (HPV infections can persist in squamous epithelium in immunocompetent individuals, and some persisting HPV infections, particularly with HPV16, promote malignant epithelial transformation. Here, we investigate whether local expression of the HPV16 protein most associated with malignant transformation, HPV16-E7, affects the phenotype and function of APC subsets in the skin. We demonstrate an expanded population of Langerhans cells in HPV16-E7 transgenic skin with distinct cell surface markers which express immune-modulatory enzymes and cytokines not expressed by cells from non transgenic skin. Furthermore, HPV16-E7 transgene expression in keratinocytes attracts new APC subsets to the epidermis. In vivo migration and transport of antigen to the draining lymph node by these APCs is markedly enhanced in HPV16-E7 expressing skin, whereas antigen-processing, as measured by proteolytic cleavage of DQ-OVA and activation of T cells in vivo by APCs, is significantly impaired. These data suggest that local expression of HPV16-E7 in keratinocytes can contribute to persisting infection with this oncogenic virus, by altering the phenotype and function of local APCs.

  7. The role of NK1.1+ cells in the protection against MHC class I+ HPV16-associated tumours

    Czech Academy of Sciences Publication Activity Database

    Šímová, Jana; Bubeník, Jan; Bieblová, Jana; Jandlová, Táňa

    2004-01-01

    Roč. 50, č. 6 (2004), s. 200-202 ISSN 0015-5500 R&D Projects: GA AV ČR IAA5052203; GA AV ČR KSK5011112; GA MZd NC7148; GA MZd NR7807 Institutional research plan: CEZ:AV0Z5052915 Keywords : HPV16 * MHC class I expression * NK1.1+ cells Subject RIV: EC - Immunology Impact factor: 0.507, year: 2004

  8. Prophylactic, therapeutic and anti-metastatic effects of BMDC and DC lines in mice carrying HPV16-associated tumours

    Czech Academy of Sciences Publication Activity Database

    Mendoza, Luis; Bubeník, Jan; Šímová, Jana; Jandlová, Táňa; Vonka, V.; Mikyšková, Romana

    2003-01-01

    Roč. 23, č. 1 (2003), s. 243-247 ISSN 1019-6439 R&D Projects: GA MZd NC7148; GA ČR GA301/01/0985; GA AV ČR IAA5052203 Grant - others:Liga proti rakovině(CZ) - Institutional research plan: CEZ:AV0Z5052915 Keywords : HPV 16 * dendritic cells * residual tumour disease Subject RIV: FD - Oncology ; Hematology Impact factor: 2.536, year: 2003

  9. Attenuated Recombinant Influenza A Virus Expressing HPV16 E6 and E7 as a Novel Therapeutic Vaccine Approach.

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    Christoph Jindra

    Full Text Available Persistent infection with high-risk human papillomavirus (HPV types, most often HPV16 and HPV18, causes all cervical and most anal cancers, and a subset of vulvar, vaginal, penile and oropharyngeal carcinomas. Two prophylactic virus-like particle (VLPs-based vaccines, are available that protect against vaccine type-associated persistent infection and associated disease, yet have no therapeutic effect on existing lesions or infections. We have generated recombinant live-attenuated influenza A viruses expressing the HPV16 oncogenes E6 and E7 as experimental immunotherapeutic vaccine candidates. The influenza A virus life cycle lacks DNA intermediates as important safety feature. Different serotypes were generated to ensure efficient prime and boost immunizations. The immune response to vaccination in C57BL/6 mice was characterized by peptide ELISA and IFN-γ ELISpot, demonstrating induction of cell-mediated immunity to HPV16 E6 and E7 oncoproteins. Prophylactic and therapeutic vaccine efficacy was analyzed in the murine HPV16-positive TC-1 tumor challenge model. Subcutaneous (s.c. prime and boost vaccinations of mice with recombinant influenza A serotypes H1N1 and H3N2, followed by challenge with TC-1 cells resulted in complete protection or significantly reduced tumor growth as compared to control animals. In a therapeutic setting, s.c. vaccination of mice with established TC-1 tumors decelerated tumor growth and significantly prolonged survival. Importantly, intralesional vaccine administration induced complete tumor regression in 25% of animals, and significantly reduced tumor growth in 50% of mice. These results suggest recombinant E6E7 influenza viruses as a promising new approach for the development of a therapeutic vaccine against HPV-induced disease.

  10. A Phase I Trial of a Human Papillomavirus (HPV) DNA Vaccine for HPV16+ Cervical Intraepithelial Neoplasia 2/3

    Science.gov (United States)

    Trimble, Cornelia L.; Peng, Shiwen; Kos, Ferdynand; Gravitt, Patti; Viscidi, Raphael; Sugar, Elizabeth; Pardoll, Drew; Wu, TC

    2010-01-01

    Purpose: To evaluate the safety and immunogenicity of a therapeutic HPV16 DNA vaccine administered to women with HPV16+CIN2/3. Experimental Design: This phase I trial incorporated the standard ‘3+3” dose escalation design with an additional 6 patients allocated to the maximally tolerated dose (MTD). Healthy adult women with colposcopically-directed biopsy-proven HPV16+ CIN2/3 received three intramuscular (IM) vaccinations (0.5 mg, 1 mg, or 3mg) of a plasmid expressing a Sig-E7(detox)-HSP70 fusion protein on days 0, 28 and 56, and underwent standard therapeutic resection of the cervical squamocolumnar junction at day 105 (week 15). Safety and immunogenicity of the vaccine and histologic outcome based on resection at week 15 were assessed. Results: Fifteen patients were evaluable (3 each at 0.5 mg and 1mg, 9 at 3mg). The vaccine was well tolerated: most adverse events were mild transient injection-site discomfort; no dose-limiting toxicities were observed. Although HPVE7-specific T-cell responses to E7 detected by enzyme-linked immunospot assays (IFNγ) were of low frequency and magnitude, detectable increases in response subsequent to vaccination were identified in subjects in the second and third cohorts. Complete histologic regression occurred in 3/9 (33%, CI: 7%-70%)) individuals in the highest dose cohort, Although the difference is not significant, it is slightly higher than would be expected in an unvaccinated cohort (25%). Conclusions: This HPV16 DNA vaccine was safe and well tolerated. While it appears possible to elicit HPV-specific T cell responses in patients with established dysplastic lesions, other factors are likely to play a role in lesion regression. PMID:19118066

  11. Interleukin 2 gene therapy of residual disease in mice carrying tumours induced by HPV 16

    Czech Academy of Sciences Publication Activity Database

    Bubeník, Jan; Šímová, Jana; Mikyšková, Romana; Sobota, Vesna; Jandlová, Táňa; Šmahel, M.; Sobotková, E.; Vonka, V.

    1999-01-01

    Roč. 14, č. 3 (1999), s. 593-597 ISSN 1019-6439 R&D Projects: GA MZd IZ3516; GA MZd IZ4069; GA MZd NC45011; GA ČR GA312/96/1211; GA ČR GA312/98/0826 Institutional research plan: CEZ:AV0Z5052915 Subject RIV: EC - Immunology Impact factor: 1.381, year: 1999

  12. The Prevalence and pattern of HPV-16 immunostaining in uterine cervical carcinomas in Ethiopian women: a pilot study

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    Mona M Rashed

    2011-03-01

    Full Text Available INTRODUCTION: Cancer of the cervix uteri is the second most common cancer among women worldwide. The association of human papillomavirus (HPV infection with cervical carcinogenesis is well documented. This is a pilot study aiming to studying the prevalence and the pattern of Human Papilloma Virus Type 16 (HPV16 by immunostaining in the tissues of cervical carcinomas of Ethiopian women. METHODS: 20 specimens of uterine cervical carcinomas were studied histopathologically and immunohistochemically for HPV16. RESULTS: Histologically the specimens were classified as: Ten cases were Non Keratinized Squamous cell carcinoma (NKSCC, six cases were Keratinized Squamous Cell Carcinoma (KSCC and four cases were Adenocarcinoma (ADC. Immunohistochemistry study showed positivity in eleven cases (55%; seven cases (35% were non-keratinized squamous cell carcinoma; three cases (15% were keratinized squamous cell carcinoma and one case (5% belonged to the adenocarcinomas. CONCLUSION: This study reveals a significant detection of HPV in Ethiopian women by the use of advanced techniques such as Immunohistochemistry (IHC. The data of this study suggested that the marked expression of the HPV 16 was in the less differentiated uterine cervix carcinomas

  13. Studies towards the potential of poliovirus as a vector for the expression of HPV 16 virus-like-particles.

    Science.gov (United States)

    van Kuppeveld, Frank J M; de Jong, Arjan; Dijkman, Henri B P M; Andino, Raul; Melchers, Willem J G

    2002-11-15

    Development of human cervical carcinomas is associated with infection by certain human papillomavirus (HPV) types. Thus, protection against HPV infection through vaccination may prevent development of cervical cancer. The purpose of this study was to investigate the possibility of using a poliovirus recombinant vector to induce immunity against HPV. A poliovirus recombinant was constructed which contained the complete coding sequence of the HPV 16 major capsid protein L1, between the P1 and P2 region of the poliovirus polyprotein. A replication-competent virus was obtained after transfection of the recombinant RNA into tissue culture cells. Electron microscopically examination of cells infected with the poliovirus-HPV L1 recombinant indicated that HPV 16 L1 self-assembles into virus-like particles. To investigate the immunological response in vivo, susceptible transgenic mice carrying the poliovirus receptor were infected with the recombinant poliovirus. In all mice a modest but consistent immune response against HPV 16 was observed. Based on these results, the potential for picornavirus-derived vectors in vaccine development against HPV infection is discussed.

  14. The high risk HPV16 L2 minor capsid protein has multiple transport signals that mediate its nucleocytoplasmic traffic

    International Nuclear Information System (INIS)

    Mamoor, Shahan; Onder, Zeynep; Karanam, Balasubramanyam; Kwak, Kihyuck; Bordeaux, Jennifer; Crosby, Lauren; Roden, Richard B.S.; Moroianu, Junona

    2012-01-01

    In this study we examined the transport signals contributing to HPV16 L2 nucleocytoplasmic traffic using confocal microscopy analysis of enhanced green fluorescent protein—L2 (EGFP-L2) fusions expressed in HeLa cells. We confirmed that both nuclear localization signals (NLSs), the nNLS (1MRHKRSAKRTKR12) and cNLS (456RKRRKR461), previously characterized in vitro (Darshan et al., 2004), function independently in vivo. We discovered that a middle region rich in arginine residues (296SRRTGIRYSRIGNKQTLRTRS316) functions as a nuclear retention sequence (NRS), as mutagenesis of critical arginine residues within this NRS reduced the fraction of L2 in the nucleus despite the presence of both NLSs. Significantly, the infectivity of HPV16 pseudoviruses containing either RR297AA or RR297EE within the L2 NRS was strongly reduced both in HaCaT cells and in a murine challenge model. Experiments using Ratjadone A nuclear export inhibitor and mutation-localization analysis lead to the discovery of a leucine-rich nuclear export signal ( 462 LPYFFSDVSL) mediating 16L2 nuclear export. These data indicate that HPV16 L2 nucleocytoplasmic traffic is dependent on multiple functional transport signals.

  15. The high risk HPV16 L2 minor capsid protein has multiple transport signals that mediate its nucleocytoplasmic traffic

    Energy Technology Data Exchange (ETDEWEB)

    Mamoor, Shahan; Onder, Zeynep [Biology Department, Boston College, Chestnut Hill, MA 02467 (United States); Karanam, Balasubramanyam; Kwak, Kihyuck [Department of Pathology, The Johns Hopkins University, Baltimore, MD 21231 (United States); Bordeaux, Jennifer; Crosby, Lauren [Biology Department, Boston College, Chestnut Hill, MA 02467 (United States); Roden, Richard B.S. [Department of Pathology, The Johns Hopkins University, Baltimore, MD 21231 (United States); Moroianu, Junona, E-mail: moroianu@bc.edu [Biology Department, Boston College, Chestnut Hill, MA 02467 (United States)

    2012-01-20

    In this study we examined the transport signals contributing to HPV16 L2 nucleocytoplasmic traffic using confocal microscopy analysis of enhanced green fluorescent protein-L2 (EGFP-L2) fusions expressed in HeLa cells. We confirmed that both nuclear localization signals (NLSs), the nNLS (1MRHKRSAKRTKR12) and cNLS (456RKRRKR461), previously characterized in vitro (Darshan et al., 2004), function independently in vivo. We discovered that a middle region rich in arginine residues (296SRRTGIRYSRIGNKQTLRTRS316) functions as a nuclear retention sequence (NRS), as mutagenesis of critical arginine residues within this NRS reduced the fraction of L2 in the nucleus despite the presence of both NLSs. Significantly, the infectivity of HPV16 pseudoviruses containing either RR297AA or RR297EE within the L2 NRS was strongly reduced both in HaCaT cells and in a murine challenge model. Experiments using Ratjadone A nuclear export inhibitor and mutation-localization analysis lead to the discovery of a leucine-rich nuclear export signal ({sub 462}LPYFFSDVSL) mediating 16L2 nuclear export. These data indicate that HPV16 L2 nucleocytoplasmic traffic is dependent on multiple functional transport signals.

  16. Clinicopathological Implications of Human Papilloma Virus (HPV) L1 Capsid Protein Immunoreactivity in HPV16-Positive Cervical Cytology

    Science.gov (United States)

    Lee, Sung-Jong; Lee, Ah-Won; Kang, Chang-Suk; Park, Jong-Sup; Park, Dong-Choon; Ki, Eun-Young; Lee, Keun-Ho; Yoon, Joo-Hee; Hur, Soo-Young; Kim, Tae-Jung

    2014-01-01

    Background: The objective of this study was to investigate the expression of human papilloma virus (HPV) L1 capsid protein in abnormal cervical cytology with HPV16 infection and analyze its association with cervical histopathology in Korean women. Material and Methods: We performed immunocytochemistry for HPV L1 in 475 abnormal cervical cytology samples from patients with HPV16 infections using the Cytoactiv® HPV L1 screening set. We investigated the expression of HPV L1 in cervical cytology samples and compared it with the results of histopathological examination of surgical specimens. Results: Of a total of 475 cases, 188 (39.6%) were immunocytochemically positive and 287 (60.4%) negative for HPV L1. The immunocytochemical expression rates of HPV L1 in atypical squamous cells of unknown significance (ASCUS), low-grade squamous intraepithelial lesions (LSIL), high-grade squamous intraepithelial lesions (HSIL), and cancer were 21.8%, 59.7%, 19.1%, and 0.0%, respectively. LSIL exhibited the highest rate of HPV L1 positivity. Of a total of 475 cases, the multiple-type HPV infection rate, including HPV16, in HPV L1-negative cytology samples was 27.5%, which was significantly higher than that in HPV L1-positive cytology samples (p = 0.037). The absence of HPV L1 expression in ASCUS and LSIL was significantly associated with high-grade (≥cervical intraepithelial neoplasia [CIN] 2) than low-grade (≤CIN1) histopathology diagnoses (p 0.05). On the other hand, among 188 HPV L1-positive cases, 30.6% of multiple-type HPV infections showed high-grade histopathology diagnoses (≥CIN3), significantly higher than the percentage of HPV16 single infections (8.6%) (p = 0.0004) Conclusions: Our study demonstrates that the expression of HPV L1 is low in advanced dysplasia. Furthermore, the absence of HPV L1 in HPV16-positive low-grade cytology (i.e., ASCUS and LSIL) is strongly associated with high-grade histopathology diagnoses. The multiplicity of HPV infections may have an

  17. The synergistic transactivation of the hepatitis B viral (HBV) pregenomic promoter by the E6 protein of human papillomavirus type 16 (HPV-16 E6) with HBV X protein was mediated through the AP1 site of E element in the enhancer I (EnI) in human liver cell.

    Science.gov (United States)

    Lee, D H; Choi, B H; Rho, H M

    1999-11-01

    Infection by HBV of a cell already infected with other viral species or vice versa has been suggested as being involved in hepatocellular carcinoma. Using the CAT assay method, we investigated the interactive roles of HBx and potentially oncogenic and transactivating viral early proteins such as Ad5 E1A, HPV-16 E6, and SV40 T ag. In the presence of HBx, only HPV-16 E6 showed significant synergistic transactivation of EnI. We further investigated the function of the HPV-16 E6 using deletion, heterologous promoter, and mutation analyses on the EnI promoter. The results showed that the synergistic effect was mediated through the AP1 site of the E element in EnI by the direct activation of AP1 and support the idea that the infection by HBV of the cell with other viral species such as HPV-16 could increase the transcription activity of the HBV and other oncogenes containing an AP1 site in the promoter. Copyright 1999 Academic Press.

  18. Evaluation of HPV-16 and HPV-18 specific antibody measurements in saliva collected in oral rinses and merocel® sponges.

    Science.gov (United States)

    Parker, Katherine H; Kemp, Troy J; Pan, Yuanji; Yang, Zhen; Giuliano, Anna R; Pinto, Ligia A

    2018-05-03

    Current Human papillomavirus (HPV) L1 VLP vaccines protect against HPV-16 and HPV-18-associated cancers, in females and males. Although correlates of protection have not been identified, HPV-specific antibodies at sites of infection are thought to be the main mechanism of protection afforded by vaccination. Oral sampling has gained increased attention as a potential alternative to serum in monitoring immunity to vaccination and understanding local immunity in oral cancers. Serum was collected via venipuncture, and saliva was collected via oral rinses and Merocel® sponges from healthy volunteers: 16 unvaccinated females, 6 females (ages 24-41) and 6 mid-adult aged males (ages 27-45) recipients of three doses of the HPV-16/18/6/11 vaccine (Gardasil®). Mid-adult male vaccine trial participants were compared to female participants. Samples were tested for anti-HPV-16 and anti-HPV-18 immunoglobulin G levels by an L1 virus-like particle-based enzyme-linked immunosorbent assay (ELISA). All vaccinated participants had detectable serum anti-HPV-16 and anti-HPV-18 antibodies. Optimal standard concentration range and sample serial dilutions for oral rinses were determined. The standard curve was not affected by the type of solution examined. Reproducibility of HPV-16 and HPV-18 antibody titers in mouthwash (overall CV  0.9) was observed for sera spiked controls in both solutions. HPV-16 and HPV-18 specific antibodies were detectable in saliva from vaccine recipients, both in mouthwash and in Merocel® sponges but levels were several logs lower than those in serum. This study confirms the application of HPV-16 and HPV-18 ELISAs currently used in sero-epidemiological studies of immunogenicity of HPV vaccines for use with oral samples. Oral samples may be a useful resource for the detection of HPV-16 and HPV-18-specific antibodies in saliva following vaccination. Copyright © 2018 Elsevier Ltd. All rights reserved.

  19. Analysis of peripheral blood immune cells after prophylactic immunization with HPV-16/18 ASO4-adjuvanted vaccine

    Directory of Open Access Journals (Sweden)

    Iwona Hus

    2015-04-01

    Full Text Available Persistent infection with oncogenic types of human papillomavirus (HPV is a causal factor for more than 99% of cervical cancers. Recently, prophylactic vaccines have been developed to prevent infections with cancer-associated HPV types (HPV16 and HPV18. The aim of this study was to analyze the changes in the immune system that occur within four weeks of the first dose of HPV-16/18 ASO4-adjuvanted vaccine. Assessment of the percentages of selected cell populations in peripheral blood of 20 healthy volunteers vaccinated with Cervarix was performed using flow cytometry. The analysis revealed an increase in the proportion of activated B and CD4+ T helper cells and an absence of significant differences in cytotoxic CD8+ T lymphocytes, indicating activation of the humoral response after vaccination, without a significant effect on cellular response. There were no significant changes in the NK cell population, and there was a reduction of the percentage of NKT-like cells, which may result from expiry of the primary response at the time of analysis. The presented results are preliminary, and in the context of the increasing use of the anti-HPV vaccine, it would be worth continuing the study in larger groups of patients and at earlier and later time points in combination with the measurement of specific anti-HPV16 and -HPV18 antibody levels. Such an assessment could therefore contribute not only to better understanding of the exact mechanism of action of the vaccine, but also to defining the immunological parameters that determine its effectiveness.

  20. Immune escape phenotype of HPV16-associated tumours: MHC class I expression changes during progression and therapy

    Czech Academy of Sciences Publication Activity Database

    Mikyšková, Romana; Bubeník, Jan; Vonka, V.; Šmahel, M.; Indrová, Marie; Bieblová, Jana; Šímová, Jana; Jandlová, Táňa

    2005-01-01

    Roč. 26, č. 2 (2005), s. 521-527 ISSN 1019-6439 R&D Projects: GA MZd(CZ) NC7148; GA MZd(CZ) NR8004; GA MZd(CZ) NR7807; GA ČR(CZ) GA301/04/0492; GA AV ČR(CZ) IAA5052203 Institutional research plan: CEZ:AV0Z5052915 Keywords : HPV16 * MHC class I- tumour cells * CTLs Subject RIV: EC - Immunology Impact factor: 2.681, year: 2005

  1. Epidemiology of HPV 16 and cervical cancer in Finland and the potential impact of vaccination: mathematical modelling analyses.

    Directory of Open Access Journals (Sweden)

    Ruanne V Barnabas

    2006-05-01

    Full Text Available BACKGROUND: Candidate human papillomavirus (HPV vaccines have demonstrated almost 90%-100% efficacy in preventing persistent, type-specific HPV infection over 18 mo in clinical trials. If these vaccines go on to demonstrate prevention of precancerous lesions in phase III clinical trials, they will be licensed for public use in the near future. How these vaccines will be used in countries with national cervical cancer screening programmes is an important question. METHODS AND FINDINGS: We developed a transmission model of HPV 16 infection and progression to cervical cancer and calibrated it to Finnish HPV 16 seroprevalence over time. The model was used to estimate the transmission probability of the virus, to look at the effect of changes in patterns of sexual behaviour and smoking on age-specific trends in cancer incidence, and to explore the impact of HPV 16 vaccination. We estimated a high per-partnership transmission probability of HPV 16, of 0.6. The modelling analyses showed that changes in sexual behaviour and smoking accounted, in part, for the increase seen in cervical cancer incidence in 35- to 39-y-old women from 1990 to 1999. At both low (10% in opportunistic immunisation and high (90% in a national immunisation programme coverage of the adolescent population, vaccinating women and men had little benefit over vaccinating women alone. We estimate that vaccinating 90% of young women before sexual debut has the potential to decrease HPV type-specific (e.g., type 16 cervical cancer incidence by 91%. If older women are more likely to have persistent infections and progress to cancer, then vaccination with a duration of protection of less than 15 y could result in an older susceptible cohort and no decrease in cancer incidence. While vaccination has the potential to significantly reduce type-specific cancer incidence, its combination with screening further improves cancer prevention. CONCLUSIONS: HPV vaccination has the potential to

  2. Immunogenicity of dendritic cell-based HPV16 E6/E7 peptide vaccines: CTL activation and protective effects

    Czech Academy of Sciences Publication Activity Database

    Indrová, Marie; Reiniš, Milan; Bubeník, Jan; Jandlová, Táňa; Bieblová, Jana; Vonka, V.; Velek, Jiří

    2004-01-01

    Roč. 50, č. 6 (2004), s. 184-193 ISSN 0015-5500 R&D Projects: GA MZd NC7148; GA MZd NR7807; GA ČR GA301/01/0985; GA ČR GA301/04/0492; GA AV ČR KSK5011112 Grant - others:Fogarty International Center(US) D43 TW000233 Institutional research plan: CEZ:AV0Z5052915 Keywords : HPV16 E6/E7 peptides * dendritic cell s * cytotoxic lymphocytes Subject RIV: EC - Immunology Impact factor: 0.507, year: 2004

  3. Búsquedas de epitopes conformacionales en la proteína HPV16-E7 del virus de papiloma humano

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    Julio C. Calvo

    2010-11-01

    Full Text Available El virus de papiloma humano tipo 16 (HPV-16 es la principal causa de cáncer entre las mujeres en Colombia. Es importante desarrollar pruebas de bajo costo y confiables para detectar la presencia de este virus en las primeras etapas de la enfermedad. Los péptidos pueden ser usados para detectar anticuerpos en sueros, pero a menudo no son activos. Sin embargo, péptidos restringidos que imiten epitopes conformacionales pueden ser más efectivos.

  4. The early noncoding region of human papillomavirus type 16 is regulated by cytoplasmic polyadenylation factors

    DEFF Research Database (Denmark)

    Glahder, Jacob-Andreas Harald; Kristiansen, Karen; Durand, Marjorie

    2010-01-01

    All human papillomavirus type 16 (HPV-16) early mRNAs are polyadenylated at the poly(A) signal within the early 3' untranslated region (3'UTR). The 3'end of the early E5 open reading frame and the 3'UTR of HPV-16 is very AU-rich, with five regions similar to cytoplasmic polyadenylation elements (...

  5. Vitamin K3 induces antiproliferative effect in cervical epithelial cells transformed by HPV 16 (SiHa cells) through the increase in reactive oxygen species production.

    Science.gov (United States)

    de Carvalho Scharf Santana, Natália; Lima, Natália Alves; Desoti, Vânia Cristina; Bidóia, Danielle Lazarin; de Souza Bonfim Mendonça, Patrícia; Ratti, Bianca Altrão; Nakamura, Tânia Ueda; Nakamura, Celso Vataru; Consolaro, Marcia Edilaine Lopes; Ximenes, Valdecir Farias; de Oliveira Silva, Sueli

    2016-10-01

    Cervical cancer is characterized as an important public health problem. According to latest estimates, cancer of the cervix is the fourth most common cancer among women. Due to its high prevalence, the search for new and efficient drugs to treat this infection is continuous. The progression of HPV-associated cervical cancer involves the expression of two viral proteins, E6 and E7, which are rapidly degraded by the ubiquitin-proteasome system through the increase in reactive oxygen species generation. Vitamins are essential to human substances, participate in the regulation of metabolism, and facilitate the process of energy transfer. Some early studies have indicated that vitamin K3 exerts antitumor activity by inducing cell death by apoptosis through an increase in the generation of reactive oxygen species. Thus, we evaluated the antiproliferative effect and a likely mechanism of action of vitamin K3 against cervical epithelial cells transformed by HPV 16 (SiHa cells) assessing the production of total ROS, the mitochondrial membrane potential, the cell morphology, the cell volume, and the cell membrane integrity. Our results show that vitamin K3 induces an increase in ROS production in SiHa cells, triggering biochemical and morphological events, such as depolarization of mitochondrial membrane potential and decreasing cell volume. Our data showed that vitamin K3 generates an oxidative imbalance in SiHa cells, leading to mechanisms that induce cell death by apoptosis.

  6. HPV-16 E7 expression up-regulates phospholipase D activity and promotes rapamycin resistance in a pRB-dependent manner.

    Science.gov (United States)

    Rabachini, Tatiana; Boccardo, Enrique; Andrade, Rubiana; Perez, Katia Regina; Nonogaki, Suely; Cuccovia, Iolanda Midea; Villa, Luisa Lina

    2018-04-27

    Human Papillomavirus (HPV) infection is the main risk factor for the development and progression of cervical cancer. HPV-16 E6 and E7 expression is essential for induction and maintenance of the transformed phenotype. These oncoproteins interfere with the function of several intracellular proteins, including those controlling the PI3K/AKT/mTOR pathway in which Phospolipase D (PLD) and Phosphatidic acid (PA) play a critical role. PLD activity was measured in primary human keratinocytes transduced with retroviruses expressing HPV-16 E6, E7 or E7 mutants. The cytostatic effect of rapamycin, a well-known mTOR inhibitor with potential clinical applications, was evaluated in monolayer and organotypic cultures. HPV-16 E7 expression in primary human keratinocytes leads to an increase in PLD expression and activity. Moreover, this activation is dependent on the ability of HPV-16 E7 to induce retinoblastoma protein (pRb) degradation. We also show that cells expressing HPV-16 E7 or silenced for pRb acquire resistance to the antiproliferative effect of rapamycin. This is the first indication that HPV oncoproteins can affect PLD activity. Since PA can interfere with the ability of rapamycin to bind mTOR, the use of combined strategies to target mTOR and PLD activity might be considered to treat HPV-related malignancies.

  7. Cellular immune responses to HPV-18, -31, and -53 in healthy volunteers immunized with recombinant HPV-16 L1 virus-like particles

    International Nuclear Information System (INIS)

    Pinto, Ligia A.; Viscidi, Raphael; Harro, Clayton D.; Kemp, Troy J.; Garcia-Pineres, Alfonso J.; Trivett, Matthew; Demuth, Franklin; Lowy, Douglas R.; Schiller, John T.; Berzofsky, Jay A.; Hildesheim, Allan

    2006-01-01

    Human papillomavirus-like particles (HPV VLP) are candidate vaccines that have shown to be efficacious in reducing infection and inducing robust antiviral immunity. Neutralizing antibodies generated by vaccination are largely type-specific, but little is known about the type-specificity of cellular immune responses to VLP vaccination. To determine whether vaccination with HPV-16 L1VLP induces cellular immunity to heterologous HPV types (HPV-18, HPV-31, and HPV-53), we examined proliferative and cytokine responses in vaccine (n = 11) and placebo (n = 5) recipients. Increased proliferative and cytokine responses to heterologous types were observed postvaccination in some individuals. The proportion of women responding to heterologous types postvaccination (36%-55%) was lower than that observed in response to HPV-16 (73%). Response to HPV-16 VLP predicted response to other types. The strongest correlations in response were observed between HPV-16 and HPV-31, consistent with their phylogenetic relatedness. In summary, PBMC from HPV-16 VLP vaccine recipients can respond to L1VLP from heterologous HPV types, suggesting the presence of conserved T cell epitopes

  8. Genetic variants of NOXA and MCL1 modify the risk of HPV16-associated squamous cell carcinoma of the head and neck

    International Nuclear Information System (INIS)

    Zhou, Ziyuan; Sturgis, Erich M; Liu, Zhensheng; Wang, Li-E; Wei, Qingyi; Li, Guojun

    2012-01-01

    The cooperation between phorbol 12-myristate 13-acetate induced protein 1 (NOXA) and myeloid cell leukemia 1 (MCL1) is critical in the intrinsic apoptotic pathway. Human papillomavirus 16 (HPV16), by inducing p53 and pRb-E2F degradation, may play an essential role in development of squamous cell carcinoma of the head and neck (SCCHN) through NOXA-MCL1 axis-mediated apoptosis. Therefore, genetic variants of NOXA and MCL1 may modify the SCCHN risk associated with HPV16 seropositivity. HPV16 serology was obtained by immunoadsorption assay. Four functional SNPs in the promoter of NOXA (rs9957673, rs4558496) and MCL1 (rs9803935, rs3738485) were genotyped for 380 cases and 335 frequency-matched cancer-free controls of non-Hispanic whites. Associations between the four polymorphisms and SCCHN risk were not significant, while we observed a significantly joint effect on SCCHN risk between the polymorphisms and HPV16 seropositivity. Notably, this effect modification was particularly pronounced for oropharyngeal cancer in subgroups including never smokers, never drinkers and younger subjects. Our results suggested that polymorphisms of NOXA and MCL1 may modify the risk of HPV16-associated oropharyngeal cancer. The further identification of population subgroups at higher risk provides evidence that HPV-targeting treatment may help benefit SCCHN. However, larger studies are needed to validate our findings

  9. Long-term persistence of systemic and mucosal immune response to HPV-16/18 AS04-adjuvanted vaccine in preteen/adolescent girls and young women

    DEFF Research Database (Denmark)

    Petäjä, T; Pedersen, Court; Poder, A

    2011-01-01

    Vaccination against oncogenic human papillomavirus (HPV) types is one key intervention for cervical cancer prevention. This follow-up study assessed the persistence of the systemic and mucosal immune responses together with the safety profile of the HPV-16/18 AS04-adjuvanted vaccine administered...... of transudation or exudation of serum IgG antibodies through the cervical epithelium. The HPV-16/18 AS04-adjuvanted vaccine had a clinically acceptable safety profile. In conclusion, this follow-up study shows that the HPV-16/18 AS04-adjuvanted vaccine administered to preteen/adolescents girls and young women...... induces long-term systemic and mucosal immune response and has a clinically acceptable safety profile up to four years after the first vaccine dose....

  10. Long-term persistence of systemic and mucosal immune response to HPV-16/18 AS04-adjuvanted vaccine in preteen/adolescent girls and young women

    DEFF Research Database (Denmark)

    Petäjä, T; Pedersen, C; Andersen, Anne Poder

    2010-01-01

    Vaccination against oncogenic human papillomavirus (HPV) types is one key intervention for cervical cancer prevention. This follow-up study assessed the persistence of the systemic and mucosal immune responses together with the safety profile of the HPV-16/18 AS04-adjuvanted vaccine administered...... of transudation or exudation of serum IgG antibodies through the cervical epithelium. The HPV-16/18 AS04-adjuvanted vaccine had a clinically acceptable safety profile. In conclusion, this follow-up study shows that the HPV-16/18 AS04-adjuvanted vaccine administered to preteen/adolescents girls and young women...... induces long-term systemic and mucosal immune response and has a clinically acceptable safety profile up to four years after the first vaccine dose....

  11. DNA of HPV and antibodies toward the protein E7 of HPV 16 as prediction factors in women with cervical cancer submitted to radiotherapy

    International Nuclear Information System (INIS)

    Bravo, Maria Mercedes; Combita R, Alba Lucia; Molano L, Monica; Gonzalez Florez, Hector; Orozco D, Oscar

    2002-01-01

    The effects of HPV infection on intrinsic tumor cell sensitivity to radiation therapy (RT) are not clear. Antibodies to HPV16-E7 protein are consistently detected in cervical cancer patients, the changes in the levels of these antibodies after RT thus may have prognostic implications. The aim of this study was to evaluate the antibodies to HPV16-E7 protein and the HPV status in cervical cancer patients before and after RT and to correlate these with clinic pathological parameters. Antibodies to peptide E7 and HPV DNA status before and after RT could have prognosis significance for patients with locally advanced uterine cervical carcinoma

  12. Immunohistochemical characteristic of expression levels of Kі-67, p16INK4a, HPV16 in cervical intraepithelial neoplasia and cervical cancer

    Directory of Open Access Journals (Sweden)

    V. A. Tumanskiy

    2017-08-01

    Full Text Available Squamous cervical cancer (SCC is a common tumor in women, which is preceded by the series of pathological processes, among which the key role is played by cervical intraepithelial neoplasia (CIN. Aim. To study the characteristics of immunohistochemical (IHC expression of Ki-67, p16INK4a, HPV16 in squamous cervical epithelium (SCE with dysplastic changes of varying degree (CIN I–III and also in the tumor cells of SCC. Materials and methods. Pathohistological and IHC studies of uterine cervix biopsies from 53 patients (the age ranged from 18 to 45 years were performed. Results. It was found that SCE with CIN I is characterized by the low Ki-67 expression level (Me = 17.87 % (13.76, 22.44 and the extremely low p16INK4a expression level (Me = 0.00 CUOD (0.00; 29.64. The proportion of HPV16-positive patients with CIN I is 27.27 %. CIN II is characterized by the average proliferation level in SCE (Me = 44.96 % (34.91, 55.41 and the moderate p16INK4a expression level (Me = 75.71 CUOD (51.24, 82, 41. The proportion of HPV16-positive patients with CIN II is 71.43 %. CIN III is characterized by the high proliferation level (Me = 74.62 % (68.50, 84.67 and by the high p16INK4a expression level of in SCE (Me = 117.47 CUOD (95.38, 123, 93; the proportion of HPV16-positive patients with CIN III is 77.78%. In all the patients with SСС, nuclear and cytoplasmic expression of HPV16 was detected in the tumor cells. High expression levels of Ki-67 and p16INK4a were detected in the tumor cells. There are direct correlations between the expression levels of Ki-67, p16INK4a, HPV16 and CIN degree. Conclusions. These data indicate that the expression levels of Ki-67, p16INK4a and HPV16 increase with the increasing of CIN grade. The absence of statistically significant differences between the expression levels of Ki-67, p16INK4a and HPV16 in CIN III and the same levels in the tumor cells of SCC indicates that these markers cannot be used for differential diagnosis

  13. Morphological correlates of genital HPV infection: Viral replication, transcription and gene expression

    International Nuclear Information System (INIS)

    Crum, C.P.; Friedman, D.; Nuovo, G.; Silverstein, S.J.

    1987-01-01

    Current studies indicate a strong correlation between specific morphological changes and the presence of certain HPV strains in precancerous squamous epithelium of the cervix, vulva and vagina. HPV type 16 is the most commonly detected HPV type in cervical lesions in our experience, and 85% of these lesions exhibit some morphological features associated with aneuploid epithelium (CIN). However, over 50% of these lesions containing HPV 16 DNA exhibit, in addition, foci of epithelium indistinguishable from condyloma, although in our experience, only one HPV type(16) is detected in the majority of these lesions. DNA-DNA in situ hybridization analysis of these lesions containing HPV 16 DNA has demonstrated nucleic acids in areas resembling both condyloma and CIN, with the greatest concentration in mature cells containing cytoplasmic maturation. Ten percent of lesions containing HPV 16 produce detectable capsid antigens, and we have confirmed the presence of these antigens in the same areas which hybridize in-situ for HPV DNA. Recent studies using biotin and S-35 labeled RNa probes constructed in GEM-1 vectors indicate that early HPV genes are expressed primarily in the upper (more mature) regions of the neoplastic epithelium. Thus maturation appears to exert a positive influence on a variety of HPV functions in neoplastic epithelium, including DNA replication, early and late gene expression. It is possible that patterns of gene expression may vary between lesions associated with different HPV types or different morphologies. This possibility is being explored

  14. Prevalence of HPV 16 and 18 and attitudes toward HPV vaccination trials in patients with cervical cancer in Mali

    Science.gov (United States)

    Téguété, Ibrahima; Dolo, Amadou; Sangare, Kotou; Sissoko, Abdoulaye; Rochas, Mali; Beseme, Sarah; Tounkara, Karamoko; Yekta, Shahla; De Groot, Anne S.; Koita, Ousmane A.

    2017-01-01

    Background Cervical cancer is one of the most common and lethal cancers in West Africa. Even though vaccines that protect against the most common Human papillomavirus (HPV) strains, 16 and 18, are currently in use in developed countries, the implementation of these vaccines in developing countries has been painfully slow, considering the pre-eminence of HPV-associated cervical cancer among women in those countries. Aim We performed serological and PCR-based assessment of blood and tissue specimens obtained from women undergoing cervical cancer-related surgery at a major urban hospital in Bamako. Since several therapeutic HPV vaccines are currently in clinical trials, we also assessed willingness to participate in HPV cancer vaccine trials. Methods Blood and biopsy samples of 240 women were evaluated for HPV types 16 and 18 by serology and PCR. Knowledge regarding the HPV vaccine and autonomy to decide to vaccinate their own child was assessed with a standardized questionnaire. Results HPV 16 and 18 were identified in 137/166 (82.5%) cervical cancer biopsy samples by PCR. Co-infection with both HPV 16 and 18 was significantly more frequent in women over 50 years of age than in younger women (63.0% vs. 37.0%). 44% of study participants said they would be willing to vaccinate their child with HPV vaccine. Only 39% of women participating in this study reported that they would be able to make an autonomous decision to receive HPV vaccination. Permission from a male spouse or head of household was identified as important for participation by 59% of the women. Conclusion This study provides strong support for the introduction of currently available HPV vaccines in Mali, and also provides key information about conditions for obtaining informed consent for HPV vaccine trials and HPV vaccination in Mali. PMID:28231334

  15. Development of a simple and quick immunochromatography method for detection of anti-HPV-16/-18 antibodies

    Science.gov (United States)

    Endo, Fumiko; Tabata, Tsutomu; Sadato, Daichi; Kawamura, Machiko; Ando, Noriyuki; Ukaji, Masako; Kobayashi, Kaoru; Kobayashi, Yukuharu; Ikeda, Tomoaki; Shibasaki, Futoshi

    2017-01-01

    Immunochromatography (IC) is widely used to detect target molecules in biological fluids. Since this method can be performed without a special technique or device, IC is a convenient way to assess the existence of antibodies or pathogens such as viruses and bacteria, simply and quickly. In this study, we established an IC method to detect serum antibodies against oncogenic human papillomavirus (HPV)-16 and HPV-18 L1 proteins using recombinant L1 proteins produced by silkworms as antigens. Infection of oncogenic HPVs is a major risk factor of cervical cancer, which is one of the most common cancers in women worldwide. We first measured blood sera of two groups by magnetic beads enzyme-linked immunosorbent assay (MB-ELISA). For the first group, sera were collected prospectively from young women who planned to receive HPV vaccination. The second group consisted of children under 20 years of age, non-vaccinated healthy women, vaccinated healthy women, dysplasia, cervical intraepithelial neoplasia III, and cervical cancer patients. We confirmed that standard vaccination doses significantly increased serum HPV antibody concentrations, and the level was sustained at least more than 30 months after vaccination. In contrast, an increase in antibody concentration was not observed in patients with precancerous cervical changes and cervical cancer. We next measured the samples in both groups using the IC method we originally developed, and found that the measurement values of IC highly correlated with those of MB-ELISA. The simple and quick IC method would be a useful tool for rapid monitoring of L1 specific antibody levels in a non-laboratory environment. With less than one drop of serum, our IC can easily detect serum HPV-16/-18 antibodies within 15 minutes, without the need for electronic devices or techniques. PMID:28158224

  16. Development of a simple and quick immunochromatography method for detection of anti-HPV-16/-18 antibodies.

    Directory of Open Access Journals (Sweden)

    Fumiko Endo

    Full Text Available Immunochromatography (IC is widely used to detect target molecules in biological fluids. Since this method can be performed without a special technique or device, IC is a convenient way to assess the existence of antibodies or pathogens such as viruses and bacteria, simply and quickly. In this study, we established an IC method to detect serum antibodies against oncogenic human papillomavirus (HPV-16 and HPV-18 L1 proteins using recombinant L1 proteins produced by silkworms as antigens. Infection of oncogenic HPVs is a major risk factor of cervical cancer, which is one of the most common cancers in women worldwide. We first measured blood sera of two groups by magnetic beads enzyme-linked immunosorbent assay (MB-ELISA. For the first group, sera were collected prospectively from young women who planned to receive HPV vaccination. The second group consisted of children under 20 years of age, non-vaccinated healthy women, vaccinated healthy women, dysplasia, cervical intraepithelial neoplasia III, and cervical cancer patients. We confirmed that standard vaccination doses significantly increased serum HPV antibody concentrations, and the level was sustained at least more than 30 months after vaccination. In contrast, an increase in antibody concentration was not observed in patients with precancerous cervical changes and cervical cancer. We next measured the samples in both groups using the IC method we originally developed, and found that the measurement values of IC highly correlated with those of MB-ELISA. The simple and quick IC method would be a useful tool for rapid monitoring of L1 specific antibody levels in a non-laboratory environment. With less than one drop of serum, our IC can easily detect serum HPV-16/-18 antibodies within 15 minutes, without the need for electronic devices or techniques.

  17. MHC class I+ and class I(-)HPV16-associated tumours expressing the E7 oncoprotein do not cross-react in immunization/challenge experiments

    Czech Academy of Sciences Publication Activity Database

    Šímová, Jana; Mikyšková, Romana; Vonka, V.; Bieblová, Jana; Bubeník, Jan; Jandlová, Táňa

    2003-01-01

    Roč. 2003, č. 49 (2003), s. 230-234 ISSN 0015-5500 Institutional research plan: CEZ:AV0Z5052915 Keywords : HPV 16 * MHC class I expression * tumour vaccines Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.527, year: 2003

  18. Immunotherapy of HPV-16-associated tumours with tumour cell line/dendritic cell line (TC-1/DC2.4) hybrid vaccines

    Czech Academy of Sciences Publication Activity Database

    Šímová, Jana; Bieblová, Jana; Jandlová, Táňa; Bubeník, Jan

    2003-01-01

    Roč. 2003, č. 49 (2003), s. 203-206 ISSN 0015-5500 Institutional research plan: CEZ:AV0Z5052915 Keywords : HPV16 * dendritic cells * fusion Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.527, year: 2003

  19. No evidence for cross-protection of the HPV-16/18 vaccine against HPV-6/11 positivity in female STI clinic visitors

    NARCIS (Netherlands)

    Woestenberg, Petra J.; King, Audrey J.; van der Sande, Marianne A. B.; Donken, Robine; Leussink, Suzan; van der Klis, Fiona R. M.; Hoebe, Christian J. P. A.; Bogaards, Johannes A.; van Benthem, Birgit H. B.; Adema, D.; Buist-Arkema, R.; Beerens, A.; Luijt, D.; Meijer, S.; Schirm, J.; Buiting, A.; Peeters, M.; Rossen, J.; Verbakel, H.; van Esch, P.; Verweij, J.; van der Eijk, A.; Huisman, R.; Kerkhof, C.; Korff, H.; Schutten, M.; Velzing, J.; Verduyn-Lunel, F.; Lakbiach, S.; van Rosmalen, P.; Schuurman, R.; Abma, D.; Adams, K.; Bruisten, S.; Linde, I.; Oostvogel, P.; Touwen, C.; Vermeulen, W.; Brink, A.; Nelissen, J.; Wolffs, P.; Duijvendijk, N.; Schneeberger, P.; Dinnissen-van Poppel, M.; Melchers, W.; Poort, Y.; Hooghiemstra, M.; Huisman, H.; Weel, J.; Stam, J.

    2017-01-01

    Data from a vaccine trial and from post-vaccine surveillance in the United Kingdom have suggested that the bivalent HPV-16/18 vaccine offers cross-protection against HPV-6/11 and protection against anogenital warts (AGW). We studied the effect of the bivalent vaccine on genital HPV-6/11 positivity

  20. No evidence for cross-protection of the HPV-16/18 vaccine against HPV-6/11 positivity in female STI clinic visitors

    NARCIS (Netherlands)

    Woestenberg, Petra J.; King, Audrey J.; van der Sande, Marianne A B; Donken, Robine; Leussink, Suzan; van der Klis, Fiona R M; Hoebe, Christian J P A; Bogaards, Johannes A.; van Benthem, Birgit H B

    OBJECTIVES: Data from a vaccine trial and from post-vaccine surveillance in the United Kingdom have suggested that the bivalent HPV-16/18 vaccine offers cross-protection against HPV-6/11 and protection against anogenital warts (AGW). We studied the effect of the bivalent vaccine on genital HPV-6/11

  1. Therapy of MHC Class I+ and Class I- HPV 16-Associated Tumours with Cytokines, Genetically Modified Tumour Vaccines and Dendritic Cells

    Czech Academy of Sciences Publication Activity Database

    Bubeník, Jan; Šímová, Jana; Mikyšková, Romana; Vonka, V.

    2004-01-01

    Roč. 27, č. 4 (2004), s. 161D ISSN 0147-958X. [International Congress of Immunology /12./ and Annual Conference of FOCIS /4./. Montreal, 18.07.2004-23.07.2004] Institutional research plan: CEZ:AV0Z5052915 Keywords : HPV 16 * tumour vaccines * dendritic cells Subject RIV: EC - Immunology Impact factor: 1.051, year: 2004

  2. Threshold cost-effectiveness analysis for a therapeutic vaccine against HPV-16/18-positive cervical intraepithelial neoplasia in the Netherlands

    NARCIS (Netherlands)

    Luttjeboer, Jos; Setiawan, Didik; Cao, Qi; Daemen, Toos CAHH; Postma, Maarten J.

    2016-01-01

    In this study, the potential price for a therapeutic vaccine against Human Papilloma Virus (HPV)-16 & 18 (pre)-malignant cervical lesions is examined. A decision tree model was built in the context of the new Dutch cervical cancer-screening program and includes a primary test for the presence of

  3. The nuclear retention signal of HPV16 L2 protein is essential for incoming viral genome to transverse the trans-Golgi network

    International Nuclear Information System (INIS)

    DiGiuseppe, Stephen; Bienkowska-Haba, Malgorzata; Hilbig, Lydia; Sapp, Martin

    2014-01-01

    The Human papillomavirus (HPV) capsid is composed of the major and minor capsid proteins, L1 and L2, respectively. Infectious entry requires a complex series of conformational changes in both proteins that lead to uptake and allow uncoating to occur. During entry, the capsid is disassembled and host cyclophilins dissociate L1 protein from the L2/DNA complex. Herein, we describe a mutant HPV16 L2 protein (HPV16 L2-R302/5A) that traffics pseudogenome to the trans-Golgi network (TGN) but fails to egress. Our data provide further evidence that HPV16 traffics through the TGN and demonstrates that L2 is essential for TGN egress. Furthermore, we show that cyclophilin activity is required for the L2/DNA complex to be transported to the TGN which is accompanied by a reduced L1 protein levels. - Highlights: • mNLS mutant HPV16 L2 protein traffics pseudogenome to the TGN but fails to egress. • Cyclophilin activity is required for trafficking of the L2/DNA complex to the TGN. • Majority of L1 protein is shed from the L2/DNA complex prior to reaching the TGN

  4. The nuclear retention signal of HPV16 L2 protein is essential for incoming viral genome to transverse the trans-Golgi network

    Energy Technology Data Exchange (ETDEWEB)

    DiGiuseppe, Stephen; Bienkowska-Haba, Malgorzata; Hilbig, Lydia; Sapp, Martin, E-mail: msapp1@lsuhsc.edu

    2014-06-15

    The Human papillomavirus (HPV) capsid is composed of the major and minor capsid proteins, L1 and L2, respectively. Infectious entry requires a complex series of conformational changes in both proteins that lead to uptake and allow uncoating to occur. During entry, the capsid is disassembled and host cyclophilins dissociate L1 protein from the L2/DNA complex. Herein, we describe a mutant HPV16 L2 protein (HPV16 L2-R302/5A) that traffics pseudogenome to the trans-Golgi network (TGN) but fails to egress. Our data provide further evidence that HPV16 traffics through the TGN and demonstrates that L2 is essential for TGN egress. Furthermore, we show that cyclophilin activity is required for the L2/DNA complex to be transported to the TGN which is accompanied by a reduced L1 protein levels. - Highlights: • mNLS mutant HPV16 L2 protein traffics pseudogenome to the TGN but fails to egress. • Cyclophilin activity is required for trafficking of the L2/DNA complex to the TGN. • Majority of L1 protein is shed from the L2/DNA complex prior to reaching the TGN.

  5. Human papillomavirus type 16 E6-specific antitumor immunity is induced by oral administration of HPV16 E6-expressing Lactobacillus casei in C57BL/6 mice.

    Science.gov (United States)

    Lee, Tae-Young; Kim, Yang-Hyun; Lee, Kyung-Soon; Kim, Jeong-Ki; Lee, Il-Han; Yang, Jai-Myung; Sung, Moon-Hee; Park, Jong-Sup; Poo, Haryoung

    2010-11-01

    Given that local cell-mediated immunity (CMI) against the human papillomavirus type 16 E6 (HPV16 E6) protein is important for eradication of HPV16 E6-expressing cancer cells in the cervical mucosa, the HPV16 E6 protein may be a target for the mucosal immunotherapy of cervical cancer. Here, we expressed the HPV16 E6 antigen on Lactobacillus casei (L. casei) and investigated E6-specific CMI following oral administration of the L. casei-PgsA-E6 to mice. Surface expression of HPV16 E6 antigens was confirmed and mice were orally inoculated with the L. casei-PgsA or the L. casei-PgsA-E6. Compared to the L. casei-PgsA-treated mice, significantly higher levels of serum IgG and mucosal IgA were observed in L. casei-PgsA-E6-immunized mice; these differences were significantly enhanced after boost. Consistent with this, systemic and local CMI were significantly increased after the boost, as shown by increased counts of IFN-gamma-secreting cells in splenocytes, mesenteric lymph nodes (MLN), and vaginal samples. Furthermore, in the TC-1 tumor model, animals receiving the orally administered L. casei-PgsA-E6 showed reduced tumor size and increased survival rate versus mice receiving control (L. casei-PgsA) immunization. We also found that L. casei-PgsA-E6-induced antitumor effect was decreased by in vivo depletion of CD4(+) or CD8(+) T cells. Collectively, these results indicate that the oral administration of lactobacilli bearing the surface-displayed E6 protein induces T cell-mediated cellular immunity and antitumor effects in mice.

  6. The differences in heparin binding for the C-terminal basic-sequence-rich peptides of HPV-16 and HPV-18 capsid protein L1

    International Nuclear Information System (INIS)

    Sun Jian; Yu Jisheng; Yu Zhiwu; Zha Xiao; Wu Yuqing

    2012-01-01

    Graphial abstract: The differences in heparin binding for the C-terminal basic-sequence-rich peptides of HPV-16 and HPV-18 capsid protein L1. Highlights: ► Several driving forces contribute to the interaction between heparin and peptides. ► C-terminal of HPV L1 is a potential candidate for the attachment to host cells. ► The C-terminal peptides of HPV-16 and -18 L1 have different heparin-binding. ► The different heparin-binding provides an explanation for the distinct prevalences. - Abstract: The high-risk types of human papillomaviruses (HPV) HPV-16 and -18 are the predominant types associated with cervical cancer. HPV-16 and -18 account for about 50% and 20%, respectively, of cervical cancers worldwide. While the reason and molecular mechanism of the distinct prevalence and distributions between them remain poorly understood, the binding affinity of cell surface receptor with capsid proteins, especially L1, may be involved. We examined heparin binding with two synthetic peptides corresponding to the 14 amino acid C-terminal peptides of HPV-16 and -18 L1 with the goal of comparing the equivalent residues in different HPV types. Using isothermal titration calorimetry (ITC) and static right-angle light scattering (SLS), we determined the binding constant K, reaction enthalpy ΔH, and other thermodynamic parameters in the interaction. Especially, we assessed the role of specific residues in binding with heparin by comparing the NMR spectra of free and heparin-bound peptides.

  7. Induction of MAGE-A3 and HPV-16 immunity by Trojan vaccines in patients with head and neck carcinoma.

    Science.gov (United States)

    Voskens, Caroline J; Sewell, Duane; Hertzano, Ronna; DeSanto, Jennifer; Rollins, Sandra; Lee, Myounghee; Taylor, Rodney; Wolf, Jeffrey; Suntharalingam, Mohan; Gastman, Brian; Papadimitriou, John C; Lu, Changwan; Tan, Ming; Morales, Robert; Cullen, Kevin; Celis, Esteban; Mann, Dean; Strome, Scott E

    2012-12-01

    We performed a pilot study using Trojan vaccines in patients with advanced squamous cell carcinoma of the head and neck (SCCHN). These vaccines are composed of HLA-I and HLA-II restricted melanoma antigen E (MAGE)-A3 or human papillomavirus (HPV)-16 derived peptides, joined by furin-cleavable linkers, and linked to a "penetrin" peptide sequence derived from HIV-TAT. Thirty-one patients with SCCHN were screened for the trial and 5 were enrolled. Enrolled patients were treated with 300 μg of Trojan peptide supplemented with Montanide and granulocyte-macrophage colony-stimulating factor (GM-CSF) at 4-week intervals for up to 4 injections. Following vaccination, peripheral blood mononuclear cells (PBMCs) from 4 of 5 patients recognized both the full Trojan constructs and constituent HLA-II peptides, whereas responses to HLA-I restricted peptides were less pronounced. This treatment regimen seems to have acceptable toxicity and elicits measurable systemic immune responses against HLA-II restricted epitopes in a subset of patients with advanced SCCHN. Copyright © 2012 Wiley Periodicals, Inc.

  8. Human papillomavirus (HPV types 16, 18, 31, 45 DNA loads and HPV-16 integration in persistent and transient infections in young women

    Directory of Open Access Journals (Sweden)

    Ferenczy Alex

    2010-11-01

    Full Text Available Abstract Background HPV burden is a predictor for high-grade cervical intraepithelial neoplasia and cancer. The natural history of HPV load in young women being recently exposed to HPV is described in this paper. Methods A total of 636 female university students were followed for 2 years. Cervical specimens with HPV-16, -18, -31, or -45 DNA by consensus PCR were further evaluated with type-specific and β-globin real-time PCR assays. Proportional hazards regression was used to estimate hazard ratios (HR of infection clearance. Generalized estimating equations assessed whether HPV loads was predictive of HPV infection at the subsequent visit. Results HPV loads were consistently higher among women Conclusions The association between HPV load and persistence is not uniform across high-risk genital genotypes. HPV-16 integration was only rarely demonstrated in young women.

  9. IL-12 immunotherapy of minimal residual disease in murine models of HPV16-associated tumours: induction of immune responses, cytokine production and kinetics of immune cell subsets

    Czech Academy of Sciences Publication Activity Database

    Indrová, Marie; Bieblová, Jana; Bubeník, Jan; Reiniš, Milan

    2008-01-01

    Roč. 32, č. 2 (2008), s. 499-507 ISSN 1019-6439 R&D Projects: GA ČR GA301/06/0774 EU Projects: European Commission(XE) 18933 - CLINIGENE Institutional research plan: CEZ:AV0Z50520514 Keywords : HPV 16 * MHC class I-positive and -deficient tumours * immature myeloid cells Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.234, year: 2008

  10. Genetically modified cellular vaccines against human papillomavirus type 16 (HPV16)-associated tumors: adjuvant treatment of minimal residual disease after surgery/chemotherapy

    Czech Academy of Sciences Publication Activity Database

    Bubeník, Jan; Šímová, Jana

    2009-01-01

    Roč. 14, č. 1 (2009), s. 169-173 ISSN 1107-0625 R&D Projects: GA ČR GA301/06/0774; GA ČR GA301/07/1410 EU Projects: European Commission(XE) 18933 - CLINIGENE Institutional research plan: CEZ:AV0Z50520514 Keywords : residual tumour disease * HPV16 * cellular vaccines Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.600, year: 2009

  11. HPV 16-associated tumours: IL-12 can repair the absence of cytotoxic and proliferative responses of tumour infiltrating cells after chemotherapy

    Czech Academy of Sciences Publication Activity Database

    Indrová, Marie; Bieblová, Jana; Rossowska, J.; Kuropka, P.; Pajtasz-Piasecka, E.; Bubeník, Jan; Reiniš, Milan

    2009-01-01

    Roč. 34, č. 1 (2009), s. 173-180 ISSN 1019-6439 R&D Projects: GA ČR GA301/06/0774 EU Projects: European Commission(XE) 18933 - CLINIGENE Grant - others:Polish Ministry of Science and Higher Education(PL) N401 235334 Institutional research plan: CEZ:AV0Z50520514 Keywords : HPV 16 * vaccines * tumour-infiltrating cells Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.447, year: 2009

  12. Immunization with MHC class I-negative but not -positive HPV16-associated tumour cells inhibits growth of MHC class I-negative tumours

    Czech Academy of Sciences Publication Activity Database

    Reiniš, Milan; Šímová, Jana; Indrová, Marie; Bieblová, Jana; Přibylová, Hana; Moravcová, Simona; Jandlová, Táňa; Bubeník, Jan

    2007-01-01

    Roč. 30, č. 4 (2007), s. 1011-1017 ISSN 1019-6439 R&D Projects: GA MZd NR7807 EU Projects: European Commission(XE) 18933 - CLINIGENE Grant - others:Liga proti rakovině, Praha(CZ) XX Institutional research plan: CEZ:AV0Z50520514 Keywords : HPV16 * MHC class I-deficient tumours * immunologic crossreaction Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.295, year: 2007

  13. Introduction and sustained high coverage of the HPV bivalent vaccine leads to a reduction in prevalence of HPV 16/18 and closely related HPV types.

    Science.gov (United States)

    Kavanagh, K; Pollock, K G J; Potts, A; Love, J; Cuschieri, K; Cubie, H; Robertson, C; Donaghy, M

    2014-05-27

    In 2008, a national human papillomavirus (HPV) immunisation programme began in Scotland for 12-13 year old females with a three-year catch-up campaign for those under the age of 18. Since 2008, three-dose uptake of bivalent vaccine in the routine cohort aged 12-13 has exceeded 90% annually, while in the catch-up cohort overall uptake is 66%. To monitor the impact of HPV immunisation, a programme of national surveillance was established (pre and post introduction) which included yearly sampling and HPV genotyping of women attending for cervical screening at age 20. By linking individual vaccination, screening and HPV testing records, we aim to determine the impact of the immunisation programme on circulating type-specific HPV infection particularly for four outcomes: (i) the vaccine types HPV 16 or 18 (ii) types considered to be associated with cross-protection: HPV 31, 33 or 45; (iii) all other high-risk types and (iv) any HPV. From a total of 4679 samples tested, we demonstrate that three doses (n=1100) of bivalent vaccine are associated with a significant reduction in prevalence of HPV 16 and 18 from 29.8% (95% confidence interval 28.3, 31.3%) to 13.6% (95% confidence interval 11.7, 15.8%). The data also suggest cross-protection against HPV 31, 33 and 45. HPV 51 and 56 emerged as the most prevalent (10.5% and 9.6%, respectively) non-vaccine high-risk types in those vaccinated, but at lower rates than HPV 16 (25.9%) in those unvaccinated. This data demonstrate the positive impact of bivalent vaccination on the prevalence of HPV 16, 18, 31, 33 and 45 in the target population and is encouraging for countries which have achieved high-vaccine uptake.

  14. Fowlpox virus recombinants expressing HPV-16 E6 and E7 oncogenes for the therapy of cervical carcinoma elicit humoral and cell-mediated responses in rabbits.

    Science.gov (United States)

    Radaelli, Antonia; Pozzi, Eleana; Pacchioni, Sole; Zanotto, Carlo; Morghen, Carlo De Giuli

    2010-04-21

    Around half million new cases of cervical cancer arise each year, making the development of an effective therapeutic vaccine against HPV a high priority. As the E6 and E7 oncoproteins are expressed in all HPV-16 tumour cells, vaccines expressing these proteins might clear an already established tumour and support the treatment of HPV-related precancerous lesions. Three different immunisation regimens were tested in a pre-clinical trial in rabbits to evaluate the humoral and cell-mediated responses of a putative HPV-16 vaccine. Fowlpoxvirus (FP) recombinants separately expressing the HPV-16 E6 (FPE6) and E7 (FPE7) transgenes were used for priming, followed by E7 protein boosting. All of the protocols were effective in eliciting a high antibody response. This was also confirmed by interleukin-4 production, which increased after simultaneous priming with both FPE6 and FPE7 and after E7 protein boost. A cell-mediated immune response was also detected in most of the animals. These results establish a preliminary profile for the therapy with the combined use of avipox recombinants, which may represent safer immunogens than vaccinia-based vectors in immuno-compromised individuals, as they express the transgenes in most mammalian cells in the absence of a productive replication.

  15. Fowlpox virus recombinants expressing HPV-16 E6 and E7 oncogenes for the therapy of cervical carcinoma elicit humoral and cell-mediated responses in rabbits

    Directory of Open Access Journals (Sweden)

    Pacchioni Sole

    2010-04-01

    Full Text Available Abstract Background Around half million new cases of cervical cancer arise each year, making the development of an effective therapeutic vaccine against HPV a high priority. As the E6 and E7 oncoproteins are expressed in all HPV-16 tumour cells, vaccines expressing these proteins might clear an already established tumour and support the treatment of HPV-related precancerous lesions. Methods Three different immunisation regimens were tested in a pre-clinical trial in rabbits to evaluate the humoral and cell-mediated responses of a putative HPV-16 vaccine. Fowlpoxvirus (FP recombinants separately expressing the HPV-16 E6 (FPE6 and E7 (FPE7 transgenes were used for priming, followed by E7 protein boosting. Results All of the protocols were effective in eliciting a high antibody response. This was also confirmed by interleukin-4 production, which increased after simultaneous priming with both FPE6 and FPE7 and after E7 protein boost. A cell-mediated immune response was also detected in most of the animals. Conclusion These results establish a preliminary profile for the therapy with the combined use of avipox recombinants, which may represent safer immunogens than vaccinia-based vectors in immuno-compromised individuals, as they express the transgenes in most mammalian cells in the absence of a productive replication.

  16. HPV16-E2 induces prophase arrest and activates the cellular DNA damage response in vitro and in precursor lesions of cervical carcinoma.

    Science.gov (United States)

    Xue, Yuezhen; Toh, Shen Yon; He, Pingping; Lim, Thimothy; Lim, Diana; Pang, Chai Ling; Abastado, Jean-Pierre; Thierry, Françoise

    2015-10-27

    Cervical intraepithelial neoplasia (CIN) is caused by human papillomavirus (HPV) infection and is the precursor to cervical carcinoma. The completion of the HPV productive life cycle depends on the expression of viral proteins which further determines the severity of the cervical neoplasia. Initiation of the viral productive replication requires expression of the E2 viral protein that cooperates with the E1 viral DNA helicase. A decrease in the viral DNA replication ability and increase in the severity of cervical neoplasia is accompanied by simultaneous elevated expression of E6 and E7 oncoproteins. Here we reveal a novel and important role for the HPV16-E2 protein in controlling host cell cycle during malignant transformation. We showed that cells expressing HPV16-E2 in vitro are arrested in prophase alongside activation of a sustained DDR signal. We uncovered evidence that HPV16-E2 protein is present in vivo in cells that express both mitotic and DDR signals specifically in CIN3 lesions, immediate precursors of cancer, suggesting that E2 may be one of the drivers of genomic instability and carcinogenesis in vivo.

  17. The levels of anti-HPV16/18 and anti-HPV31/33/35/45/52/58 antibodies among AS04-adjuvanted HPV16/18 vaccinated and non-vaccinated Ugandan girls aged 10-16 years.

    Science.gov (United States)

    Nakalembe, Miriam; Banura, Cecily; Namujju, Proscovia B; Mirembe, Florence M

    2014-01-01

    Data on Human Papilloma virus (HPV) vaccine immune response in sub-Saharan Africa is still sparse yet such knowledge is critical for optimal implementation and monitoring of HPV vaccines. Our primary objective was to evaluate levels of anti-HPV-16/18 antibodies and six other 'high risk' HPV (hrHPV) types among the vaccinated and unvaccinated Ugandan girls. We conducted a cross sectional study among AS04-adjuvanted HPV-16/18 vaccinated and unvaccinated school girls aged 10-16 years in Western Uganda using purposive sampling. The vaccinated girls were at 18 months post vaccination. After consenting and assenting, data was collected using interviewer administered questionnaires for demographics and sexual history. Blood was drawn from which serum samples were analysed by the multiplex HPV serology technology to determine anti-HPV antibody levels to HPV-16/18 and six other hrHPV types (31, 33, 35, 45, 52 and 58). The antibody levels were expressed as Median Fluorescent Intensity (MFI). A total of 207 vaccinated [mean age 13.1 years (SD 1.5); range 10-16 years] and 197 unvaccinated girls [mean age 13.6 years (SD 1.3); range 10-16 years] participated in the study. Sexual activity was self reported among 14/207 (6.8%) vaccinated and 5/197 (2.5%) unvaccinated girls. The MFI levels for HPV-16 and HPV-18 were 15 and 20 times higher respectively in the vaccinated girls than in the unvaccinated girls. HPV-16 mean MFI level was 4691(SD 1812; 95% CI: 4438-4958) among the vaccinated compared to 218 (SD 685; 95% CI: 190-252) among the unvaccinated girls. For HPV-18 the mean MFI level was 1615 (SD 1326; 95% CI: 1470-1776) among the vaccinated compared to MFI 103 (SD 506; 95% CI: 88 -121) among unvaccinated girls. In addition antibody levels to non vaccine hrHPV types (31, 33, 35, 45, 52 and 58) were all significantly higher in the vaccinated group than in the unvaccinated group (plevel of antibodies to HPV-16/18 and other non-vaccine hrHPV types compared to the unvaccinated girls

  18. Detecção sorológica de anti-HPV 16 e 18 e sua associação com os achados do papanicolaou em adolescentes e mulheres jovens Serological detection of anti HPV 16/18 and its association with pap smear in adolescents and young women

    Directory of Open Access Journals (Sweden)

    Cristina Helena Rama

    2006-02-01

    Full Text Available OBJETIVO: Verificar a taxa de anticorpos neutralizantes anti-HPV 16 e/ou 18, e a sua associação com os achados da citologia oncológica do colo uterino em adolescentes e mulheres jovens. MÉTODOS: Foram incluídas, neste estudo transversal, 541 mulheres de 15 a 25 anos de idade, saudáveis, sexualmente ativas, que apresentaram exame ginecológico normal, no período de setembro a novembro de 2000. Foi obtida uma amostra cervical para citologia em meio líquido e uma amostra de sangue para identificação dos anticorpos anti-HPV 16 e/ou 18, por meio do método ELISA. As amostras foram encaminhadas para um laboratório de referência na Bélgica. Para análise estatística, foram estimadas a prevalência e a razão de prevalência (RP, com intervalo de confiança de 95%. RESULTADOS: Entre as mulheres incluídas, 150 (27,7% apresentaram resultados positivos da sorologia sendo: 79 (14,6% por anticorpos anti-HPV 16, 35 (6,4% anti-HPV 18 e 36 (6,6% anti-HPV 16 e 18. Foram detectadas anormalidades citológicas em 107 casos (20,5%, sendo 63 classificadas como atipias celulares de significado indeteminado (ASCUS, 41 casos sugestivos de lesões de baixo (LSIL e três casos sugestivos de lesões de alto grau (HSIL. A prevalência de citologias anormais quando a sorologia foi positiva foi apenas 1,75 vez a prevalência de citologias positivas observadas com sorologia negativa. Nesta amostra não houve evidência de associação entre os resultados anormais da citologia e a positividade da sorologia. CONCLUSÃO: O resultado deste estudo indica uma alta prevalência de sorologia positiva para o HPV 16 e 18 em mulheres jovens sadias sem relação com os achados anormais da colpocitologia.OBJECTIVE: Verify the relation between HPV 16/18 antibodies and the association with cervical cytology findings in adolescents and young women. METHODS: A cross sectional study with 541 healthy and sexually active women from 15 to 25 years of age was carried on from

  19. Suppression of HPV-16 late L1 5′-splice site SD3632 by binding of hnRNP D proteins and hnRNP A2/B1 to upstream AUAGUA RNA motifs

    Science.gov (United States)

    Li, Xiaoze; Johansson, Cecilia; Glahder, Jacob; Mossberg, Ann-Kristin; Schwartz, Stefan

    2013-01-01

    Human papillomavirus type 16 (HPV-16) 5′-splice site SD3632 is used exclusively to produce late L1 mRNAs. We identified a 34-nt splicing inhibitory element located immediately upstream of HPV-16 late 5′-splice site SD3632. Two AUAGUA motifs located in these 34 nt inhibited SD3632. Two nucleotide substitutions in each of the HPV-16 specific AUAGUA motifs alleviated splicing inhibition and induced late L1 mRNA production from episomal forms of the HPV-16 genome in primary human keratinocytes. The AUAGUA motifs bind specifically not only to the heterogeneous nuclear RNP (hnRNP) D family of RNA-binding proteins including hnRNP D/AUF, hnRNP DL and hnRNP AB but also to hnRNP A2/B1. Knock-down of these proteins induced HPV-16 late L1 mRNA expression, and overexpression of hnRNP A2/B1, hnRNP AB, hnRNP DL and the two hnRNP D isoforms hnRNP D37 and hnRNP D40 further suppressed L1 mRNA expression. This inhibition may allow HPV-16 to hide from the immune system and establish long-term persistent infections with enhanced risk at progressing to cancer. There is an inverse correlation between expression of hnRNP D proteins and hnRNP A2/B1 and HPV-16 L1 production in the cervical epithelium, as well as in cervical cancer, supporting the conclusion that hnRNP D proteins and A2/B1 inhibit HPV-16 L1 mRNA production. PMID:24013563

  20. Estimating long-term clinical effectiveness and cost-effectiveness of HPV 16/18 vaccine in China.

    Science.gov (United States)

    Zhang, Qian; Liu, Yi-Jun; Hu, Shang-Ying; Zhao, Fang-Hui

    2016-11-04

    Human papillomavirus (HPV) 16 and 18 are the two most common HPV oncogenic types that can be prevented by vaccination. This study aimed at assessing the cost-effectiveness of 3 doses of the bivalent HPV vaccine in rural and urban settings in China. A Markov model was adapted to reflect the lifetime of a modelled 100,000 12-year-old girls cohort in rural and urban settings in China. Input parameters were obtained from published literature, official reports and a two-round expert review panel. Clinical and economic outcomes of vaccination at age 12 with screening was compared to screening only. In the base case analysis, a 3 % discount rate, the vaccine cost of 247 CNY (US$ 39, PAHO vaccine cost in 2013), two rounds of screening in a life time and 70 % coverage for both screening and vaccination were used. One-way, two-way and probabilistic sensitivity analyses were performed. We used different thresholds of cost-effectiveness to reflect the diversity of economic development in China. Vaccination in addition to screening could prevent 60 % more cervical cancer cases and deaths than screening only. The incremental cost effectiveness ratio varied largely when changing cost of vaccination and discount in one way analysis. Vaccination was very cost-effective when the vaccine cost ranged 87-630 CNY (US$ 13.8-100) in rural and 87-750 CNY (US$ 13.8-119) in urban; and remained cost-effective when the vaccine cost ranged 630-1,700 CNY (US$ 100-270) in rural and 750-1,900 CNY (US$ 119-302) in urban in two way analysis. Probabilistic sensitivity analyses showed that model results were robust. In both rural and urban, the vaccination cost and discounting are important factors determining the cost-effectiveness of HPV vaccination; policy makers in China should take these into account when making a decision on the introduction of HPV vaccine. In areas with a high burden of cervical cancer and limited screening activities, HPV vaccination should be prioritized. However, the vaccine

  1. Cost-Effectiveness of Cervical Cancer Screening With Human Papillomavirus DNA Testing and HPV-16,18 Vaccination

    Science.gov (United States)

    Goldhaber-Fiebert, Jeremy D.; Stout, Natasha K.; Salomon, Joshua A.; Kuntz, Karen M.; Goldie, Sue J.

    2011-01-01

    Background The availability of human papillomavirus (HPV) DNA testing and vaccination against HPV types 16 and 18 (HPV-16,18) motivates questions about the cost-effectiveness of cervical cancer prevention in the United States for unvaccinated older women and for girls eligible for vaccination. Methods An empirically calibrated model was used to assess the quality-adjusted life years (QALYs), lifetime costs, and incremental cost-effectiveness ratios (2004 US dollars per QALY) of screening, vaccination of preadolescent girls, and vaccination combined with screening. Screening varied by initiation age (18, 21, or 25 years), interval (every 1, 2, 3, or 5 years), and test (HPV DNA testing of cervical specimens or cytologic evaluation of cervical cells with a Pap test). Testing strategies included: 1) cytology followed by HPV DNA testing for equivocal cytologic results (cytology with HPV test triage); 2) HPV DNA testing followed by cytology for positive HPV DNA results (HPV test with cytology triage); and 3) combined HPV DNA testing and cytology. Strategies were permitted to switch once at age 25, 30, or 35 years. Results For unvaccinated women, triennial cytology with HPV test triage, beginning by age 21 years and switching to HPV testing with cytology triage at age 30 years, cost $78 000 per QALY compared with the next best strategy. For girls vaccinated before age 12 years, this same strategy, beginning at age 25 years and switching at age 35 years, cost $41 000 per QALY with screening every 5 years and $188 000 per QALY screening triennially, each compared with the next best strategy. These strategies were more effective and cost-effective than screening women of all ages with cytology alone or cytology with HPV triage annually or biennially. Conclusions For both vaccinated and unvaccinated women, age-based screening by use of HPV DNA testing as a triage test for equivocal results in younger women and as a primary screening test in older women is expected to be more

  2. Gene expression in early stage cervical cancer

    NARCIS (Netherlands)

    Biewenga, Petra; Buist, Marrije R.; Moerland, Perry D.; van Thernaat, Emiel Ver Loren; van Kampen, Antoine H. C.; ten Kate, Fiebo J. W.; Baas, Frank

    2008-01-01

    Objective. Pelvic lymph node metastases are the main prognostic factor for survival in early stage cervical cancer, yet accurate detection methods before surgery are lacking. In this study, we examined whether gene expression profiling can predict the presence of lymph node metastasis in early stage

  3. An evaluation of clinical performance of FTA cards for HPV 16/18 detection using cobas 4800 HPV Test compared to dry swab and liquid medium.

    Science.gov (United States)

    Dong, Li; Lin, Chunqing; Li, Li; Wang, Margaret; Cui, Jianfeng; Feng, Ruimei; Liu, Bin; Wu, Zeni; Lian, Jia; Liao, Guangdong; Chen, Wen; Qiao, Youlin

    2017-09-01

    Effective dry storage and transport media as an alternative to conventional liquid-based medium would facilitate the accessibility of women in the low-resource settings to human papillomavirus (HPV)- based cervical cancer screening. To evaluate analytical and clinical performance of indicating FTA™ Elute Cartridge (FTA card) for the detection of HPV16/18 and cervical precancerous lesions and cancer compared to dry swab and liquid medium. Ninety patients with abnormal cytology and/or HPV infection were included for analysis. Three specimens of cervical exfoliated cells from each woman were randomly collected by FTA card, dry swab or liquid-based medium prior to colposcopy examination. The subsequent HPV DNA tests were performed on cobas 4800 HPV platform. High-risk HPV (hrHPV) positivity rate was 63.3%, 62.2% and 65.6% for samples collected by FTA card, dry swab and liquid medium, respectively. The overall agreements and kappa values for the detection of hrHPV, HPV 16 and HPV 18 between FTA card and liquid-based medium were 88.9% (κ=0.76), 97.8% (κ=0.94) and 100% (κ=1.0),respectively; between FTA card and dry swab were 92.1% (κ=0.83), 94.5% (κ=0.87) and 100% (κ=1.0), respectively. The performances of hrHPV tested by FTA card, dry swab, and liquid-based medium for detecting CIN2+ were comparable in terms of the sensitivity and specificity. The specificity of detection of CIN2+ by HPV16/18 increased by approximately 40% compared to hrHPV for any medium albeit at cost of a moderate loss of sensitivity. Dry medium might offer an alternative to conventional liquid-based medium in the HPV-based cervical cancer screening program especially in low-resource settings but still needs further evaluation. Copyright © 2017. Published by Elsevier B.V.

  4. Expression profile of microRNA-146a along HPV-induced multistep carcinogenesis: a study in HPV16 transgenic mice.

    Science.gov (United States)

    Araújo, Rita; Santos, Joana M O; Fernandes, Mara; Dias, Francisca; Sousa, Hugo; Ribeiro, Joana; Bastos, Margarida M S M; Oliveira, Paula A; Carmo, Diogo; Casaca, Fátima; Silva, Sandra; Medeiros, Rui; Gil da Costa, Rui M

    2018-02-01

    Persistent human papillomavirus (HPV) infection is associated with the development of certain types of cancer and the dysregulation of microRNAs has been implicated in HPV-associated carcinogenesis. This is the case of microRNA-146a (miR-146a), which is thought to regulate tumor-associated inflammation. We sought to investigate the expression levels of miR-146a during HPV16-mediated carcinogenesis using skin samples from K14-HPV16 transgenic mice which develop the consecutive phases of the carcinogenesis process. Female transgenic (HPV +/- ) and wild-type (HPV -/- ) mice were sacrificed at 24-26 weeks-old or 28-30 weeks-old. Chest and ear skin samples from HPV +/- and HPV -/- mice were histologically classified and used for microRNA extraction and quantification by qPCR. Chest skin samples from 24 to 26 weeks-old HPV +/- mice presented diffuse epidermal hyperplasia and only 22.5% showed multifocal dysplasia, while at 28-30 weeks-old all (100.0%) HPV +/- animals showed epidermal dysplasia. All HPV +/- ear skin samples showed carcinoma in situ (CIS). MiR-146a expression levels were higher in HPV +/- compared to HPV -/- mice (p = 0.006). There was also an increase in miR-146a expression in dysplastic skin lesions compared with hyperplasic lesions (p = 0.011). Samples showing CIS had a significant decrease in miR-146a expression when compared to samples showing epidermal hyperplasia (p = 0.018) and epidermal dysplasia (p = 0.009). These results suggest that HPV16 induces the overexpression of miR-146a in the initial stages of carcinogenesis (hyperplasia and dysplasia), whereas decreases its expression at later stages (CIS). Taken together, these data implicate and suggest different roles of miR-146a in HPV-mediated carcinogenesis.

  5. A pilot study to compare the detection of HPV-16 biomarkers in salivary oral rinses with tumour p16INK4a expression in head and neck squamous cell carcinoma patients

    International Nuclear Information System (INIS)

    Chai, Ryan C.; Lim, Yenkai; Frazer, Ian H.; Wan, Yunxia; Perry, Christopher; Jones, Lee; Lambie, Duncan; Punyadeera, Chamindie

    2016-01-01

    Human papilloma virus-16 (HPV-16) infection is a major risk factor for a subset of head and neck squamous cell carcinoma (HNSCC), in particular oropharyngeal squamous cell carcinoma (OPSCC). Current techniques for assessing the HPV-16 status in HNSCC include the detection of HPV-16 DNA and p16 INK4a expression in tumor tissues. When tumors originate from hidden anatomical sites, this method can be challenging. A non-invasive and cost-effective alternative to biopsy is therefore desirable for HPV-16 detection especially within a community setting to screen at-risk individuals. The present study compared detection of HPV-16 DNA and RNA in salivary oral rinses with tumor p16 INK4a status, in 82 HNSCC patients using end-point and quantitative polymerase chain reaction (PCR). Of 42 patients with p16 INK4a -positive tumours, 39 (sensitivity = 92.9 %, PPV = 100 % and NPV = 93 %) had oral rinse samples with detectable HPV-16 DNA, using end-point and quantitative PCR. No HPV-16 DNA was detected in oral rinse samples from 40 patients with p16 INK4a negative tumours, yielding a test specificity of 100 %. For patients with p16 INK4a positive tumours, HPV-16 mRNA was detected using end-point reverse transcription PCR (RT-PCR) in 24/40 (sensitivity = 60 %, PPV = 100 % and NPV = 71 %), and using quantitative RT-PCR in 22/40 (sensitivity = 55 %, PPV = 100 % and NPV = 69 %). No HPV-16 mRNA was detected in oral rinse samples from the p16 INK4a -negative patients, yielding a specificity of 100 %. We demonstrate that the detection of HPV-16 DNA in salivary oral rinse is indicative of HPV status in HNSCC patients and can potentially be used as a diagnostic tool in addition to the current methods. The online version of this article (doi:10.1186/s12885-016-2217-1) contains supplementary material, which is available to authorized users

  6. Primary Screening for Cervical Cancer Based on High-Risk Human Papillomavirus (HPV) Detection and HPV 16 and HPV 18 Genotyping, in Comparison to Cytology

    Science.gov (United States)

    Constantinidis, Theocharis; Constantinidis, Theodoros C.

    2015-01-01

    Objectives The objective of the present study is to assess the performance of a high-risk human papillomavirus (HR-HPV) DNA test with individual HPV-16/HPV-18 genotyping as a method for primary cervical cancer screening compared with liquid-based cytology (LBC) in a population of Greek women taking part in routine cervical cancer screening. Methods The study, conducted by the “HEllenic Real life Multicentric cErvical Screening” (HERMES) study group, involved the recruitment of 4,009 women, aged 25–55, who took part in routine cervical screening at nine Gynecology Departments in Greece. At first visit cervical specimens were collected for LBC and HPV testing using the Roche Cobas 4800 system. Women found positive for either cytology or HPV were referred for colposcopy, whereas women negative for both tests will be retested after three years. The study is ongoing and the results of the first screening round are reported herein. Results Valid results for cytology and HPV testing were obtained for 3,993 women. The overall prevalence of HR-HPV was 12.7%, of HPV-16 2.7% and of HPV-18 1.4%. Of those referred for colposcopy, cervical intraepithelial neoplasia grade 2 or worse (CIN2+) was detected in 41 women (1.07%). At the threshold of CIN2+, cytology [atypical squamous cells of undetermined significance (ASC-US) or worse] and HPV testing showed a sensitivity of 53.7% and 100% respectively, without change between age groups. Cytology and HPV testing showed specificity of 96.8% and 90.3% respectively, which was increased in older women (≥30) in comparison to younger ones (25–29). Genotyping for HPV16/18 had similar accuracy to cytology for the detection of CIN2+ (sensitivity: 58.5%; specificity 97.5%) as well as for triage to colposcopy (sensitivity: 58.5% vs 53.7% for cytology). Conclusion HPV testing has much better sensitivity than cytology to identify high-grade cervical lesions with slightly lower specificity. HPV testing with individual HPV-16/HPV-18

  7. Procjena citomorfologije, HPV statusa i HPV 16 genotipa u predikciji ishoda bolesti kod pacijentica s citološkim nalazom ASCUS i LSIL

    OpenAIRE

    Vrdoljak-Mozetič, Danijela; Štemberger-Papić, Snježana; Verša Ostojić, Damjana; Rubeša-Mihaljević, Roberta; Dinter, Morana; Brnčić-Fischer, Alemka; Krašević, Maja

    2016-01-01

    Cilj: Istražiti međusobnu povezanost i prognostički značaj HPV statusa (humani papiloma-virus), HPV 16 genotipa i citomorfologije kod pacijentica s citološkim nalazom abnormalnih stanica pločastog epitela vrata maternice graničnog i blagog stupnja. Materijali i metode: U studiju su uključene pacijentice s inicijalnim citološkim nalazom vrata maternice atipičnih skvamoznih stanica neodređenog značenja (ASCUS, N = 160) i skvamozne intraepitelne lezije niskog stupnja (LSIL, N = 155). Analizirane...

  8. Focal epithelial hyperplasia by human papillomavirus (HPV)-32 misdiagnosed as HPV-16 and treated with combination of retinoids, imiquimod and quadrivalent HPV vaccine.

    Science.gov (United States)

    Gemigniani, Franco; Hernández-Losa, Javier; Ferrer, Berta; García-Patos, Vicente

    2015-12-01

    Focal epithelial hyperplasia (FEH) or Heck's disease is a rare, benign and asymptomatic mucosal proliferation associated with human papillomavirus (HPV) infection, mainly with genotypes 13 and 32. We report a florid case of FEH in an 11-year-old Haitian girl with systemic lupus erythematosus receiving immunosuppressive therapy. Cryotherapy was previously performed on numerous occasions with no results. We decided to prescribe a non-invasive and more comfortable treatment. A combination of topical retinoid and imiquimod cream was well tolerated and led to an important improvement. The evidence of infection by HPV-16 detected by polymerase chain reaction (PCR) technique, prompted us to prescribe the quadrivalent HPV vaccine (types 6, 11,16 and 18). Subsequent PCR sequencing with generic primers GP5-GP6 and further BLAST comparative analysis confirmed that genomic viral sequence in our case truly corresponded with HPV-32. This molecular misdiagnosis can be explained by the similarity between genomic sequences of both HPV-16 and -32 genotypes. At the 1-year follow up, we observed total clinical improvement and no recurrences of the disease. Complete healing in this case may correspond to a potential action of topical retinoid, imiquimod and the cross-protection mechanism of the quadrivalent HPV vaccine. © 2015 Japanese Dermatological Association.

  9. HPV16-E7 expression in squamous epithelium creates a local immune suppressive environment via CCL2- and CCL5- mediated recruitment of mast cells.

    Directory of Open Access Journals (Sweden)

    Anne-Sophie Bergot

    2014-10-01

    Full Text Available Human Papillomavirus (HPV 16 E7 protein promotes the transformation of HPV infected epithelium to malignancy. Here, we use a murine model in which the E7 protein of HPV16 is expressed as a transgene in epithelium to show that mast cells are recruited to the basal layer of E7-expressing epithelium, and that this recruitment is dependent on the epithelial hyperproliferation induced by E7 by inactivating Rb dependent cell cycle regulation. E7 induced epithelial hyperplasia is associated with increased epidermal secretion of CCL2 and CCL5 chemokines, which attract mast cells to the skin. Mast cells in E7 transgenic skin, in contrast to those in non-transgenic skin, exhibit degranulation. Notably, we found that resident mast cells in E7 transgenic skin cause local immune suppression as evidenced by tolerance of E7 transgenic skin grafts when mast cells are present compared to the rejection of mast cell-deficient E7 grafts in otherwise competent hosts. Thus, our findings suggest that mast cells, recruited towards CCL2 and CCL5 expressed by epithelium induced to proliferate by E7, may contribute to an immunosuppressive environment that enables the persistence of HPV E7 protein induced pre-cancerous lesions.

  10. A prime/boost strategy using DNA/fowlpox recombinants expressing the genetically attenuated E6 protein as a putative vaccine against HPV-16-associated cancers.

    Science.gov (United States)

    Bissa, Massimiliano; Illiano, Elena; Pacchioni, Sole; Paolini, Francesca; Zanotto, Carlo; De Giuli Morghen, Carlo; Massa, Silvia; Franconi, Rosella; Radaelli, Antonia; Venuti, Aldo

    2015-03-05

    Considering the high number of new cases of cervical cancer each year that are caused by human papilloma viruses (HPVs), the development of an effective vaccine for prevention and therapy of HPV-associated cancers, and in particular against the high-risk HPV-16 genotype, remains a priority. Vaccines expressing the E6 and E7 proteins that are detectable in all HPV-positive pre-cancerous and cancer cells might support the treatment of HPV-related lesions and clear already established tumors. In this study, DNA and fowlpox virus recombinants expressing the E6F47R mutant of the HPV-16 E6 oncoprotein were generated, and their correct expression verified by RT-PCR, Western blotting and immunofluorescence. Immunization protocols were tested in a preventive or therapeutic pre-clinical mouse model of HPV-16 tumorigenicity using heterologous (DNA/FP) or homologous (DNA/DNA and FP/FP) prime/boost regimens. The immune responses and therapeutic efficacy were evaluated by ELISA, ELISPOT assays, and challenge with TC-1* cells. In the preventive protocol, while an anti-E6-specific humoral response was just detectable, a specific CD8(+) cytotoxic T-cell response was elicited in immunized mice. After the challenge, there was a delay in cancer appearance and a significant reduction of tumor volume in the two groups of E6-immunized mice, thus confirming the pivotal role of the CD8(+) T-cell response in the control of tumor growth in the absence of E6-specific antibodies. In the therapeutic protocol, in-vivo experiments resulted in a higher number of tumor-free mice after the homologous DNA/DNA or heterologous DNA/FP immunization. These data establish a preliminary indication for the prevention and treatment of HPV-related tumors by the use of DNA and avipox constructs as safe and effective immunogens following a prime/boost strategy. The combined use of recombinants expressing both E6 and E7 proteins might improve the antitumor efficacy, and should represent an important approach to

  11. Association of the Plasma and Tissue Riboflavin Levels with C20orf54 Expression in Cervical Lesions and Its Relationship to HPV16 Infection

    Science.gov (United States)

    Kelimu, Alimujiang; Guo, Xia; Mamtimin, Batur; Abudula, Abuliz; Upur, Halmurat

    2013-01-01

    Riboflavin deficiency can cause a variety of metabolic problems that lead to skin and mucosal disorders. Limited evidence suggests that high intake of riboflavin may reduce overall risks of cancer. However, association of this deficiency with cervical cancer and precancerous lesions are still not definitively known. In this study, we characterized the relationship between plasma and tissue riboflavin levels and C20orf54 protein expression in patients with cervical intraepithelial neoplasia (CIN) and cervical squamous cell carcinoma (CSCC) as well as the relationship of these levels with human papillomavirus virus 16, 18 (HPV16/18) infections. High-performance liquid chromatography (HPLC) was used to measure blood riboflavin levels in patients with CIN and CSCC, and an enzyme-linked immunosorbent assay (ELISA) was used to determine tissue riboflavin levels in patients with CSCC and matched normal mucous epithelia. The expression of C20orf54 in fresh CSCC and matched tissues were detected by qRT-PCR and western blot, respectively. And it was further confirmed by immunohistochemistry (IHC) with formalin-fixed, paraffin-embedded CIN and CSCC. An HPV genotyping chip was used to analyze HPV infection and typing. The results showed that patients with CIN and CSCC had decreased plasma riboflavin levels as compared with normal controls. There was also significantly decreased riboflavin in tissues from CSCC patients, when compared with normal cervical epithelia. C20orf54 expression were significantly up-regulated in CSCC compared to matched control on both mRNA and protein level. Tissue riboflavin levels were significantly lower in HPV16/18 positive tissue compared with HPV16/18-negative tissue, and an inverse association was found between tissue riboflavin levels and C20orf54 mRNA and protein expression in CSCC. Additionally, C20orf54 was significantly correlated with tumor stages. In conclusion, C20orf54 tend to play a protective role in Uyghur cervical carcinogenesis of

  12. Safety and Immunogenicity of the HPV-16/18 AS04-adjuvanted Vaccine in 4-6-year-old Girls: Results to Month 12 From a Randomized Trial.

    Science.gov (United States)

    Lin, Lan; Parra, Mercedes Macias; Sierra, Victor Y; Cespedes, Albino Salas; Granados, Maria Angelica; Luque, Adriana; Damaso, Silvia; Castrejon Alba, Maria Mercedes; Romano-Mazzotti, Luis; Struyf, Frank

    2018-04-01

    The burden of cervical cancer caused by human papillomavirus (HPV) is high in Latin America. The suboptimal HPV vaccination coverage in adolescents could be improved by pediatric immunization. HPV vaccination has not yet been reported in girls HPV-16/18 vaccine in 4-6-year-old girls. Healthy girls (randomized 1:1) received either 2 doses of AS04-HPV-16/18 vaccine (HPV group, N=74) or 1 dose of each measles-mumps-rubella and diphtheria-tetanus-acellular-pertussis vaccines (control group, N=74) 6 months apart. We report the safety and serum anti-HPV-16 and anti-HPV-18 antibodies (measured by enzyme-linked immunosorbent assay) up to 6 months postvaccination, that is, month (M) 12. Injection site pain was the most frequently reported solicited local symptom in HPV vaccinees. The incidence of other solicited and unsolicited symptoms after each vaccination was similar between the HPV and control group. Until M12, 1 girl in the HPV group and 2 in the control group reported serious adverse events; all serious adverse events were assessed as unrelated to study vaccines. No potential immune-mediated diseases were identified. All girls seroconverted for both antigens after 2 doses of AS04-HPV-16/18. In initially seronegative girls, anti-HPV-16 geometric mean concentrations were 20080.0 enzyme-linked immunosorbent assay units (EU)/mL at M7 and 3246.5 EU/mL at M12; anti-HPV-18 geometric mean concentrations were 10621.8 EU/mL at M7 and 1216.6 EU/mL at M12. Two-dose vaccination with AS04-HPV-16/18 was well tolerated and induced adequate antibody responses in 4-6-year-old girls.

  13. Impact of an HPV6/11/16/18 L1 virus-like particle vaccine on progression to cervical intraepithelial neoplasia in seropositive women with HPV16/18 infection

    DEFF Research Database (Denmark)

    Haupt, Richard M; Wheeler, Cosette M; Brown, Darron R

    2011-01-01

    The impact of a human papillomavirus (HPV) vaccine on development of cervical intraepithelial neoplasia grade 2-3 or adenocarcinoma in situ (CIN2-3/AIS) in women with ongoing HPV16 or 18 infections prevaccination is reported. Seventeen thousand six-hundred and twenty-two women aged 16-26 were.......9). These data suggest HPV vaccination neither reduces nor enhances progression to HPV16/18-related high grade cervical lesions, and cervical cytology screening and corresponding management should continue as per local recommendations. Ultimately, population-based surveillance of vaccinated individuals beyond...

  14. HPV16 DNA status is a strong prognosticator of loco-regional control after postoperative radiochemotherapy of locally advanced oropharyngeal carcinoma: Results from a multicentre explorative study of the German Cancer Consortium Radiation Oncology Group (DKTK-ROG)

    International Nuclear Information System (INIS)

    Lohaus, Fabian; Linge, Annett; Tinhofer, Inge; Budach, Volker; Gkika, Eleni; Stuschke, Martin; Balermpas, Panagiotis; Rödel, Claus; Avlar, Melanie; Grosu, Anca-Ligia

    2014-01-01

    Objective: To investigate the impact of HPV status in patients with locally advanced head and neck squamous cell carcinoma (HNSCC), who received surgery and cisplatin-based postoperative radiochemotherapy. Materials and methods: For 221 patients with locally advanced squamous cell carcinoma of the hypopharynx, oropharynx or oral cavity treated at the 8 partner sites of the German Cancer Consortium, the impact of HPV DNA, p16 overexpression and p53 expression on outcome were retrospectively analysed. The primary endpoint was loco-regional tumour control; secondary endpoints were distant metastases and overall survival. Results: In the total patient population, univariate analyses revealed a significant impact of HPV16 DNA positivity, p16 overexpression, p53 positivity and tumour site on loco-regional tumour control. Multivariate analysis stratified for tumour site showed that positive HPV 16 DNA status correlated with loco-regional tumour control in patients with oropharyngeal carcinoma (p = 0.02) but not in the oral cavity carcinoma group. Multivariate evaluation of the secondary endpoints in the total population revealed a significant association of HPV16 DNA positivity with overall survival (p < 0.01) but not with distant metastases. Conclusions: HPV16 DNA status appears to be a strong prognosticator of loco-regional tumour control after postoperative cisplatin-based radiochemotherapy of locally advanced oropharyngeal carcinoma and is now being explored in a prospective validation trial

  15. The prevalence of the HPV 16 genome, integrated viral status and p53 genotype in cervical cancer population of north-eastern Hungary, the correlation with the established markers of tumour progression.

    Science.gov (United States)

    Hernádi, Zoltán; Sápy, Tamás; Krasznai, Zoárd T

    2004-03-15

    To evaluate the prevalence of the HPV 16 integrated status and the p53 genotype in cervical cancer in north-eastern Hungary and their correlation with the established prognostic factors. Parallel with the routine histological examination, Southern blot hybridisation and multiplex PCRs were used to detect type/physical state of HPV DNA in primary tumours and in regional lymph nodes combined with p53 genotyping of 83 patients. 46.9% (39/83) prevalence rate of HPV 16 genome was found. The frequency of viral integration (76.9% in primary tumours and 95.2% in regional lymph nodes) and that of the p53Arg homozygous genotype (64.1%) proved to be higher than reported from other parts of the world. The HPV 16 integration and the p53 genotype, failed to correlate with the FIGO stage and lymphatic spread. The prevalence of the integrated status of the HPV 16 genome combined with homozygous p53Arg genotype is relatively high in Hungary. These factors however failed to show a strong correlation with the established markers of tumour progression.

  16. Combination of intratumoral injections of vaccinia virus MVA expressing GM-CSF and immunization with DNA vaccine prolongs the survival of mice bearing HPV16 induced tumors with downregulated expression of MHC class I molecules

    Czech Academy of Sciences Publication Activity Database

    Němečková, Š.; Šmahel, M.; Hainz, P.; Macková, J.; Zurková, K.; Gabriel, P.; Indrová, Marie; Kutinová, L.

    2007-01-01

    Roč. 54, č. 4 (2007), s. 326-333 ISSN 0028-2685 R&D Projects: GA MZd NR8004 Institutional research plan: CEZ:AV0Z50520514 Keywords : vaccinia virus MVA expressing GM- CSF * DNA vaccine * HPV16 induced tumors Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.208, year: 2007

  17. Interferon-Tau has Antiproliferative effects, Represses the Expression of E6 and E7 Oncogenes, Induces Apoptosis in Cell Lines Transformed with HPV16 and Inhibits Tumor Growth In Vivo

    Science.gov (United States)

    Padilla-Quirarte, Herbey Oswaldo; Trejo-Moreno, Cesar; Fierros-Zarate, Geny; Castañeda, Jhoseline Carnalla; Palma-Irizarry, Marie; Hernández-Márquez, Eva; Burguete-Garcia, Ana Isabel; Peralta-Zaragoza, Oscar; Madrid-Marina, Vicente; Torres-Poveda, Kirvis; Bermúdez-Morales, Victor Hugo

    2016-01-01

    Interferon tau (IFN-τ) is a promising alternative antiviral and immunotherapeutic agent in a wide variety of diseases including infectious, neurodegenerative, autoimmune and cancer due to its low toxicity in comparison with other type I interferon´s. The objective of our study was established the effect of the bovine IFN-τ on human (SiHa) and murine (BMK-16/myc) cells transformed with HPV 16 and evaluates the antitumor effect in a murine tumor model HPV 16 positive. We determine that bovine IFN-τ has antiproliferative effects, pro-apoptotic activity and induces repression of viral E6 and E7 oncogenes (time- and dose-dependent) on human and murine cells transformed with HPV 16 similar to the effects of IFN-β. However, IFN-τ induces greater antiproliferative effect, apoptosis and repression of both oncogenes in BMK-16/myc cells compared to SiHa cells. The differences were explained by the presence and abundance of the type I interferon receptor (IFNAR) in each cell line. On the other hand, we treated groups of tumor-bearing mice (HPV16 positive) with IFN-τ and showed the inhibition tumor growth effect in vivo. Our finding indicates that bovine IFN-τ may be a good candidate for immunotherapy against cervical cancer. PMID:27994659

  18. Modeling the impact of the difference in cross-protection data between a human papillomavirus (HPV-16/18 AS04-adjuvanted vaccine and a human papillomavirus (HPV-6/11/16/18 vaccine in Canada

    Directory of Open Access Journals (Sweden)

    Kohli Michele

    2012-10-01

    Full Text Available Background In Canada, two vaccines that have demonstrated high efficacy against infection with human papillomavirus (HPV types −16 and −18 are available. The HPV-6/11/16/18 vaccine provides protection against genital warts (GW while the HPV-16/18 vaccine may provide better protection against other oncogenic HPV types. In this analysis, the estimated clinical and economic benefit of each of these vaccines was compared in the Canadian setting. Methods A Markov model of the natural history of HPV infection among women, cervical cancer (CC and GW was used to estimate the impact of vaccinating a cohort of 100,000 12-year-old females on lifetime outcomes and healthcare system costs (no indirect benefit in males included. A budget impact model was used to estimate the impact of each vaccine by province. Results In the base case, vaccination with the HPV-16/18 vaccine was predicted to prevent 48 additional CC cases, and 16 additional CC deaths, while vaccination with the HPV-6/11/16/18 vaccine was predicted to prevent 6,933 additional GW cases. Vaccination with the HPV-16/18 vaccine was estimated to save 1 additional discounted quality adjusted life year (QALY at an overall lower lifetime cost to the healthcare system compared to the HPV-6/11/16/18 vaccine (assuming vaccine price parity. In sensitivity analyses, the HPV-6/11/16/18 vaccine was associated with greater QALYs saved when the cross-protection efficacy of the HPV-16/18 vaccine was reduced, or the burden of GW due to HPV-6/11 was increased. In most scenarios with price parity, the lifetime healthcare cost of the strategy with the HPV-16/18 vaccine was predicted to be lower than the HPV-6/11/16/18 vaccine. In the probabilistic sensitivity analyses, the HPV-16/18 vaccine provided more QALY benefit than the HPV-6/11/16/18 vaccine in 49.2% of scenarios, with lower relative lifetime costs in 83.5% of scenarios. Conclusions Overall, the predicted lifetime healthcare costs and QALYs saved by

  19. MG132 plus apoptosis antigen-1 (APO-1) antibody cooperate to restore p53 activity inducing autophagy and p53-dependent apoptosis in HPV16 E6-expressing keratinocytes.

    Science.gov (United States)

    Lagunas-Martínez, Alfredo; García-Villa, Enrique; Arellano-Gaytán, Magaly; Contreras-Ochoa, Carla O; Dimas-González, Jisela; López-Arellano, María E; Madrid-Marina, Vicente; Gariglio, Patricio

    2017-01-01

    The E6 oncoprotein can interfere with the ability of infected cells to undergo programmed cell death through the proteolytic degradation of proapoptotic proteins such as p53, employing the proteasome pathway. Therefore, inactivation of the proteasome through MG132 should restore the activity of several proapoptotic proteins. We investigated whether in HPV16 E6-expressing keratinocytes (KE6 cells), the restoration of p53 levels mediated by MG132 and/or activation of the CD95 pathway through apoptosis antigen-1 (APO-1) antibody are responsible for the induction of apoptosis. We found that KE6 cells underwent apoptosis mainly after incubation for 24 h with MG132 alone or APO-1 plus MG132. Both treatments activated the extrinsic and intrinsic apoptosis pathways. Autophagy was also activated, principally by APO-1 plus MG132. Inhibition of E6-mediated p53 proteasomal degradation by MG132 resulted in the elevation of p53 protein levels and its phosphorylation in Ser46 and Ser20; the p53 protein was localized mainly at nucleus after treatment with MG132 or APO-1 plus MG132. In addition, induction of its transcriptional target genes such as p21, Bax and TP53INP was observed 3 and 6 h after treatment. Also, LC3 mRNA was induced after 3 and 6 h, which correlates with lipidation of LC3B protein and induction of autophagy. Finally, using pifithrin alpha we observed a decrease in apoptosis induced by MG132, and by APO-1 plus MG132, suggesting that restoration of APO-1 sensitivity occurs in part through an increase in both the levels and the activity of p53. The use of small molecules to inhibit the proteasome pathway might permit the activation of cell death, providing new opportunities for CC treatment.

  20. Threshold cost-effectiveness analysis for a therapeutic vaccine against HPV-16/18-positive cervical intraepithelial neoplasia in the Netherlands.

    Science.gov (United States)

    Luttjeboer, Jos; Setiawan, Didik; Cao, Qi; Cahh Daemen, Toos; Postma, Maarten J

    2016-12-07

    In this study, the potential price for a therapeutic vaccine against Human Papilloma Virus (HPV)-16 & 18 (pre)-malignant cervical lesions is examined. A decision tree model was built in the context of the new Dutch cervical cancer-screening program and includes a primary test for the presence of HPV. Based on data of cervical cancer screening and HPV prevalence in the Netherlands, cohorts were created with HPV-16 or 18 positive women with cervical intraepithelial neoplasia (CIN) 2 or 3 or cervical cancer stage 1A (FIGO 1A). In the base case, the vaccine price was based on equal numbers of effective treatments in the vaccine branch and the current treatments branch of the model, and parity in cost, i.e. total cost in both branches are the same. The vaccine price is calculated by subtracting the cost of the vaccine branch from cost in the standard treatment branch and divided by the total number of women in the cohort, thereby equalizing costs in both strategies. Scenario analyses were performed taking quality adjusted life years (QALYs) into account with €20,000/QALY, €50,000/QALY and €80,000/QALY as corresponding thresholds. Sensitivity analyses were specifically targeted at the characteristics of the type-specific HPV test in the screening practice and vaccine efficacy. A probabilistic sensitivity analysis (PSA) was performed to quantify the level of uncertainty of the results found in the base case. In the base case, break-even vaccine prices of €381, €568 and €1697 were found for CIN 2, CIN 3 and FIGO 1A, respectively. The PSA showed vaccine pricing below €310, €490 and €1660 will be cost saving with a likelihood of 95% for CIN 2, CIN 3 and FIGO 1A, respectively. The vaccine price proved to be very sensitive for inclusion of QALY gains, including the HPV-type specific test into the Dutch screening practice and vaccine efficacy. Copyright © 2016 Elsevier Ltd. All rights reserved.

  1. Immunization of early adolescent females with human papillomavirus type 16 and 18 L1 virus-like particle vaccine containing AS04 adjuvant

    DEFF Research Database (Denmark)

    Pedersen, Court; Petaja, Tiina; Strauss, Gitte

    2007-01-01

    will require prophylactic vaccination against oncogenic HPV 16 and 18 before the onset of sexual activity in early adolescent girls. To establish the feasibility of vaccination in girls 10-14 years of age, we compared the immunogenicity and safety in early adolescent female individuals to those 15-25 years...... measured by enzyme-linked immunosorbent assay. Vaccine safety was assessed at 7 or 30 days after each dose; serious adverse events were recorded during the entire study period. RESULTS: Both age groups achieved 100% seroconversion for HPV 16 and 18. Participants in the group aged 10-14 years were not only......PURPOSE: In female individuals 15-25-years of age, the AS04-containing human papillomavirus (HPV)-16/18 vaccine is highly immunogenic and provides up to 100% protection against HPV-16/18 persistent infection and associated cervical lesions up to 4.5 years. Optimal cervical cancer prevention...

  2. The Human Papillomavirus Type 16 E6 Gene Alone Is Sufficient To Induce Carcinomas in Transgenic Animals

    Science.gov (United States)

    Song, Shiyu; Pitot, Henry C.; Lambert, Paul F.

    1999-01-01

    High-risk human papillomaviruses (HPVs) are the causative agents of certain human cancers. HPV type 16 (HPV16) is the papillomavirus most frequently associated with cervical cancer in women. The E6 and E7 genes of HPV are expressed in cells derived from these cancers and can transform cells in tissue culture. Animal experiments have demonstrated that E6 and E7 together cause tumors. We showed previously that E6 and E7 together or E7 alone could induce skin tumors in mice when these genes were expressed in the basal epithelia of the skin. In this study, we investigated the role that the E6 gene plays in carcinogenesis. We generated K14E6 transgenic mice, in which the HPV16 E6 gene was directed in its expression by the human keratin 14 promoter (hK14) to the basal layer of the epidermis. We found that E6 induced cellular hyperproliferation and epidermal hyperplasia and caused skin tumors in adult mice. Interestingly, the tumors derived from E6 were mostly malignant, as opposed to the tumors from E7 mice, which were mostly benign. This result leads us to hypothesize that E6 may contribute differently than E7 to HPV-associated carcinogenesis; whereas E7 primarily contributes to the early stages of carcinogenesis that lead to the formation of benign tumors, E6 primarily contributes to the late stages of carcinogenesis that lead to malignancy. PMID:10364340

  3. Murine HPV16 E7-expressing transgenic skin effectively emulates the cellular and molecular features of human high-grade squamous intraepithelial lesions

    Directory of Open Access Journals (Sweden)

    Z.K. Tuong

    2018-06-01

    Full Text Available Currently available vaccines prevent HPV infection and development of HPV-associated malignancies, but do not cure existing HPV infections and dysplastic lesions. Persistence of infection(s in immunocompetent patients may reflect induction of local immunosuppressive mechanisms by HPV, providing a target for therapeutic intervention. We have proposed that a mouse, expressing HPV16 E7 oncoprotein under a Keratin 14 promoter (K14E7 mice, and which develops epithelial hyperplasia, may assist with understanding local immune suppression mechanisms that support persistence of HPV oncogene-induced epithelial hyperplasia. K14E7 skin grafts recruit immune cells from immunocompetent hosts, but consistently fail to be rejected. Here, we review the literature on HPV-associated local immunoregulation, and compare the findings with published observations on the K14E7 transgenic murine model, including comparison of the transcriptome of human HPV-infected pre-malignancies with that of murine K14E7 transgenic skin. We argue from the similarity of i the literature findings and ii the transcriptome profiles that murine K14E7 transgenic skin recapitulates the cellular and secreted protein profiles of high-grade HPV-associated lesions in human subjects. We propose that the K14E7 mouse may be an appropriate model to further study the immunoregulatory effects of HPV E7 expression, and can facilitate development and testing of therapeutic vaccines.

  4. Early gene regulation of osteogenesis in embryonic stem cells

    KAUST Repository

    Kirkham, Glen R.; Lovrics, Anna; Byrne, Helen M.; Jensen, Oliver E.; King, John R.; Shakesheff, Kevin M.; Buttery, Lee D. K.

    2012-01-01

    The early gene regulatory networks (GRNs) that mediate stem cell differentiation are complex, and the underlying regulatory associations can be difficult to map accurately. In this study, the expression profiles of the genes Dlx5, Msx2 and Runx2

  5. Comparison of the sensitivity and specificity of real-time PCR and in situ hybridization in HPV16 and 18 detection in archival cervical cancer specimens

    Directory of Open Access Journals (Sweden)

    Beata Biesaga

    2012-07-01

    Full Text Available The aim of this study was to analyze the correlation between real-time PCR (RT-PCR treated as a reference method and in situ hybridization with tyramide amplification system (ISH-TSA in the detection of HPV16 and 18 infection and the assessment of viral genome status. The study was performed on cervical cancer biopsies fixed in 10% neutral buffered formalin and embedded in paraffin obtained from 85 women. TaqMan-based 5’exonuclease RT-PCR with type-specific primers was used to assess HPV16 and 18 infections and genome status. Viral infection and genome status was also assessed by ISH-TSA. RT-PCR revealed 76 (89.4%, and ISH-TSA 81 (95.3% cancers with HPV16 and 18 infections. The ISH-TSA sensitivity and specificity were: 96.1% and 11.1% compared to RT-PCR. The difference between these techniques in HPV detection was significant (p = 0.000. Among 76 HPV16/18 positive cancers in RT-PCR, there were 30 (39.5% with integrated and 46 (60.5% with mixed viral genome form. According to ISH-TSA, there were 39 (51.3% samples with integrated and 37 with mixed form (48.7%. The sensitivity and specificity of ISH-TSA in genome status assessment were 70.0% and 60.9%, respectively. The difference between RT-PCR and ISH-TSA in genome state detection was not statistically significant (p = 0.391. These results suggest that ISH-TSA shows insufficient specificity in HPV detection for use in clinical practice. However, this assay could be applied for viral genome status assessment.

  6. Comparative immunogenicity and safety of human papillomavirus (HPV)-16/18 AS04-adjuvanted vaccine and 4vHPV vaccine administered according to two- or three-dose schedules in girls aged 9-14 years: Results to month 36 from a randomized trial.

    Science.gov (United States)

    Leung, Ting Fan; Liu, Anthony Pak-Yin; Lim, Fong Seng; Thollot, Franck; Oh, Helen May Lin; Lee, Bee Wah; Rombo, Lars; Tan, Ngiap Chuan; Rouzier, Roman; De Simoni, Stéphanie; Suryakiran, Pemmaraju; Hezareh, Marjan; Thomas, Florence; Folschweiller, Nicolas; Struyf, Frank

    2018-01-02

    This observer-blind study (clinicaltrials.gov NCT01462357) compared the immunogenicity and safety of two doses (2D) of the HPV-16/18 AS04-adjuvanted vaccine (2D of AS04-HPV-16/18) vs. two or three doses of the 4vHPV vaccine [2D or 3D of 4vHPV] in 1075 healthy girls aged 9-14 years. Girls were randomized (1:1:1) to receive 2D of AS04-HPV-16/18 at months (M) 0, 6 (N = 359), 2D of 4vHPV at M0, 6 (N = 358) or 3D of 4vHPV at M0, 2, 6 (N = 358). 351, 339 and 346 girls, respectively, returned for the concluding visit at M36. Superiority was demonstrated at M7 and M12; comparison of the immune response to both vaccine antigens was made between 2D of AS04-HPV-16/18 and 2D or 3D of 4vHPV at subsequent time points in the according-to-protocol immunogenicity cohort (ATP-I; N = 958 at M36) and the total vaccinated cohort (TVC: N = 1036 at M36). HPV-16/18-specific T-cell- and B-cell-mediated immune responses and safety were also investigated. At M36, anti-HPV-16/18 ELISA responses in the 2D AS04-HPV-16/18 group remained superior to those of the 2D and 3D 4vHPV groups. In the M36 TVC, geometric mean titers were 2.78-fold (HPV-16) and 6.84-fold (HPV-18) higher for 2D of AS04-HPV-16/18 vs. 2D of 4vHPV and 2.3-fold (HPV-16) and 4.14-fold (HPV-18) higher vs. 3D of 4vHPV. Results were confirmed by vaccine pseudovirion-based neutralisation assay. Numbers of circulating CD4 + T cells and B cells appeared similar across groups. Safety was in line with the known safety profiles of both vaccines. In conclusion, superior HPV-16/18 antibody responses were elicited by 2D of the AS04-HPV-16/18 compared with 2D or 3D of the 4vHPV vaccine in girls aged 9-14 years. NCT0146235. Copyright © 2017. Published by Elsevier Ltd.

  7. The combinational effect of E6/E7 siRNA and anti-miR-182 on apoptosis induction in HPV16-positive cervical cells.

    Science.gov (United States)

    Javadi, Hamidreza; Lotfi, Abbas Sahebghadam; Hosseinkhani, Saman; Mehrani, Hossein; Amani, Jafar; Soheili, Zahra Soheila; Hojati, Zahra; Kamali, Mehdi

    2018-06-06

    In the present research, we assumed that reducing the amounts of E6 and E7 oncoproteins by a specific siRNA sequence and recovering p53 and RB proteins, along with the recovery of the FOXO1 protein by applying anti-miR-182, would increase apoptosis and reduce proliferation rate in cancer cells. The HPV16-positive CaSki cervical cancer cell line was used. 48 hours after transfection of siRNA for targeting E6 and E7 oncoproteins and anti-miR-182, expression of its cellular targets p53, p21 and FOXO1 was assessed by real-time PCR, western blot analysis and immunocytofluorescence staining. In all treatments, apoptosis rate and viability were evaluated using Annexin-V-FITC apoptosis detection kits and MTT assays, respectively. Among the designed siRNAs, E6-1 and E7-2 proved the most effective in reducing E6 and E7 expressions by increasing the apoptotic rates to 12.4% and 16%, respectively, after 48 hours. Also, using anti-miR-182 increased apoptotic rate to 12.7% 48 hours after transfection of cervical cancer cells. The combinational use of either E6-1 or E7-2 siRNAs with anti-miR-182 resulted in a rise in apoptosis to 19.3% and 26%, respectively, higher than those obtained from the individual application of either without anti-miR-182. The simultaneous use of siRNA E6-1 and siRNA E7-2 with cisplatin increased sensitivity to cisplatin and reduced the viability of the cancer cells as compared to the use of cisplatin alone. The simultaneous use of cisplatin and anti-miR-182 had no considerable effect on viability or apoptosis rate compared to cisplatin alone.

  8. In situ hybridization detection methods for HPV16 E6/E7 mRNA in identifying transcriptionally active HPV infection of oropharyngeal carcinoma: an updating.

    Science.gov (United States)

    Volpi, Chiara C; Ciniselli, Chiara M; Gualeni, Ambra V; Plebani, Maddalena; Alfieri, Salvatore; Verderio, Paolo; Locati, Laura; Perrone, Federica; Quattrone, Pasquale; Carbone, Antonino; Pilotti, Silvana; Gloghini, Annunziata

    2018-04-01

    The aim of this study is to compare 2 in situ hybridization (ISH) detection methods for human papilloma virus (HPV) 16 E6/E7 mRNA, that is, the RNAscope 2.0 High Definition (HD) and the upgraded RNAscope 2.5 HD version. The RNAscope 2.5 HD has recently replaced the RNAscope 2.0 HD detection kit. Therefore, this investigation starts from the need to analytically validate the new mRNA ISH assay and, possibly, to refine the current algorithm for HPV detection in oropharyngeal squamous cell carcinoma with the final goal of applying it to daily laboratory practice. The study was based on HPV status and on generated data, interpreted by a scoring algorithm. The results highlighted that the compared RNAscope HPV tests had a good level of interchangeability and enabled to identify oropharyngeal squamous cell carcinoma that are truly driven by high-risk HPV infection. This was also supported by the comparison of the RNAscope HPV test with HPV E6/E7 mRNA real-time reverse-transcription polymerase chain reaction in a fraction of cases where material for HPV E6/E7 mRNA real-time reverse-transcription polymerase chain reaction was available. Furthermore, the algorithm that associates p16 immunohistochemistry with the identification of HPV mRNA by RNAscope was more effective than the one that associated p16 immunohistochemistry with the identification of HPV DNA by ISH. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. Rejection of large HPV-16 expressing tumors in aged mice by a single immunization of VacciMax® encapsulated CTL/T helper peptides

    Directory of Open Access Journals (Sweden)

    MacDonald Lisa

    2007-06-01

    Full Text Available Abstract The incidence of cancer increases significantly in later life, yet few pre-clinical studies of cancer immunotherapy use mice of advanced age. A novel vaccine delivery platform (VacciMax®,VM is described that encapsulates antigens and adjuvants in multilamellar liposomes in a water-in-oil emulsion. The therapeutic potential of VM-based vaccines administered as a single dose was tested in HLA-A2 transgenic mice of advanced age (48–58 weeks old bearing large palpable TC1/A2 tumors. The VM-based vaccines contained one or more peptides having human CTL epitopes derived from HPV 16 E6 and E7. VM formulations contained a single peptide, a mixture of four peptides or the same four peptides linked together in a single long peptide. All VM formulations contained PADRE and CpG as adjuvants and ISA51 as the hydrophobic component of the water-in-oil emulsion. VM-formulated vaccines containing the four peptides as a mixture or linked together in one long peptide eradicated 19-day old established tumors within 21 days of immunization. Peptide-specific cytotoxic cellular responses were confirmed by ELISPOT and intracellular staining for IFN-γ producing CD8+ T cells. Mice rendered tumor-free by vaccination were re-challenged in the opposite flank with 10 million HLF-16 tumor cells, another HLA-A2/E6/E7 expressing tumor cell line. None of these mice developed tumors following the re-challenge. In summary, this report describes a VM-formulated therapeutic vaccine with the following unprecedented outcome: a eradication of large tumors (> 700 mm3 b in mice of advanced age c in less than three weeks post-immunization d following a single vaccination.

  10. LOW MOLECULAR MASS POLYPEPTIDE AND TRANSPORTER ANTIGEN PEPTIDE GENES POLYMORPHISM AS THE RISK FACTORS OF CERVICAL CANCER WHICH CAUSED BY HUMAN PAPILLOMAVIRUS TYPE-16 INFECTION IN BALI

    Directory of Open Access Journals (Sweden)

    I N. B. Mahendra

    2015-12-01

    Full Text Available Background: Until recently, cervical cancer is one of the major problem in women’s health issue related to its high incidence and mortality rate. The etiology of cervical cancer is the high risk oncogenic group of Human Papillomavirus (HPV, especially HPV-16 and 18 and its phylogenies. Meanwhile in Bali, more than 50% of infection are caused by HPV-16 infection. The main objective of this study was to investigate the role of LMP-2, LMP-7, TAP-1 and TAP-2 gene polymorphism as the risk factor in the cervical cancer carcinogenesis that is caused by HPV-16 infection. Method: A nested non-paired case-control study was conducted at Obstetric and Gynecologic Department Sanglah General Hospital, Bali-Indonesia from March 1 until August 31, 2013. Laboratory testing was carried out at Laboratory of Histopathology Leiden University Medical Centre, Netherlands,. Results: A total of 40 samples were collected, consist of 20epithelial cervical cancer patients with positive HPV-16 infection as the case group and 20 non-cervical cancer patients with positive HPV-16 infection as the control group. Women infected by HPV-16 with LMP-7 gene polymorphism had a higher risk (OR=7.36, CI 95%=1.38-40.55, p=0.013 to be diagnosed with cervical cancer. Balinese women who were infected by HPV-16 with TAP-2 gene polymorphism had a higher risk (OR= 9.33, CI 95%=2.18-39.96, p=0.001 to be diagnosed with cervical cancer. Meanwhile, Balinese women who were infected by HPV-16 with LMP-7 and TAP-2 genes polymorphism had a higher risk (OR=12.67, CI 95%=1.40-114.42, p=0.020 to be diagnosed with cervical cancer. As the result, it was shown that both of this gene polymorphism was working synergistically. Conclusion: TAP-2 and LMP-7 genes polymorphism play a role in the carcinogenesis mechanism of cervical cancer that is caused by HPV-16 infection in Bali. Meanwhile, LMP-2 and TAP-1 genes polymorphism were not found to play a role in the immunology pathway of cervical cancer that is

  11. Human Papillomavirus 16 (HPV-16), HPV-18, and HPV-31 E6 Override the Normal Phosphoregulation of E6AP Enzymatic Activity.

    Science.gov (United States)

    Thatte, Jayashree; Banks, Lawrence

    2017-11-15

    The human papillomavirus (HPV) E6 oncoproteins recruit the cellular ubiquitin ligase E6AP/UBE3A to target cellular substrates for proteasome-mediated degradation, and one consequence of this activity is the E6 stimulation of E6AP autoubiquitination and degradation. Recent studies identified an autism-linked mutation within E6AP at T485, which was identified as a protein kinase A phosphoacceptor site and which could directly regulate E6AP ubiquitin ligase activity. In this study, we have analyzed how T485-mediated regulation of E6AP might affect E6 targeting of some of its known substrates. We show that modulation of T485 has no effect on the ability of E6 to direct either p53 or Dlg for degradation. Furthermore, T485 regulation has no effect on HPV-16 or HPV-31 E6-induced autodegradation of E6AP but does affect HPV-18 E6-induced autodegradation of E6AP. In cells derived from cervical cancers, we find low levels of both phosphorylated and nonphosphorylated E6AP in the nucleus. However, ablation of E6 results in a dramatic accumulation of phospho-E6AP in the cytoplasm, whereas nonphosphorylated E6AP accumulates primarily in the nucleus. Interestingly, E6AP phosphorylation at T485 confers association with 14-3-3 proteins, and this interaction seems to be important, in part, for the ability of E6 to recruit phospho-E6AP into the nucleus. These results demonstrate that HPV E6 overrides the normal phosphoregulation of E6AP, both in terms of its enzymatic activity and its subcellular distribution. IMPORTANCE Recent reports demonstrate the importance of phosphoregulation of E6AP for its normal enzymatic activity. Here, we show that HPV E6 is capable of overriding this regulation and can promote degradation of p53 and Dlg regardless of the phosphorylation status of E6AP. Furthermore, E6 interaction with E6AP also significantly alters how E6AP is subject to autodegradation and suggests that this is not a simple stimulation of an already-existing activity but rather a

  12. A pilot study of pNGVL4a-CRT/E7(detox) for the treatment of patients with HPV16+ cervical intraepithelial neoplasia 2/3 (CIN2/3).

    Science.gov (United States)

    Alvarez, Ronald D; Huh, Warner K; Bae, Sejong; Lamb, Lawrence S; Conner, Michael G; Boyer, Jean; Wang, Chenguang; Hung, Chien-Fu; Sauter, Elizabeth; Paradis, Mihaela; Adams, Emily A; Hester, Shirley; Jackson, Bradford E; Wu, T C; Trimble, Cornelia L

    2016-02-01

    The purpose of this study was to evaluate the safety, efficacy, and immunogenicity of a plasmid vaccine, pNGVL4a-CRT-E7(detox), administered either intradermally, intramuscularly, or directly into the cervical lesion, in patients with HPV16-associated CIN2/3. Eligible patients with HPV16(+) CIN2/3 were enrolled in treatment cohorts evaluating pNGVL4a-CRT-E7(detox), administered by either particle-mediated epidermal delivery (PMED), intramuscular injection (IM), or cervical intralesional injection, at study weeks 0, 4, and 8. Patients were monitored for local injection site and systemic toxicity. A standard therapeutic resection was performed at week 15. The primary endpoints were safety and tolerability. Secondary endpoints included histologic regression and change in cervical HPV viral load. Exploratory endpoints included immune responses in the blood and in the target tissue. Thirty-two patients with HPV16(+) CIN2/3 were enrolled onto the treatment phase of the study, and were vaccinated. Twenty-two of 32 patients (69%) experienced vaccine-specific related adverse events. The most frequent vaccine-related events were constitutional and local injection site in nature, and were grade 1 or less in severity. Histologic regression to CIN 1 or less occurred in 8 of 27 (30%) patients who received all vaccinations and underwent LEEP. In subject-matched comparisons, intraepithelial CD8+ T cell infiltrates increased after vaccination in subjects in the intralesional administration cohort. pNGVL4a-CRT-E7(detox) was well-tolerated, elicited the most robust immune response when administered intralesionally, and demonstrated preliminary evidence of potential clinical efficacy. Copyright © 2015 Elsevier Inc. All rights reserved.

  13. HPV16型E6/E7遺伝子発現誘導が正常ヒト線維芽細胞に及ぼす影響の検討

    OpenAIRE

    黄田, 光博; 清水, 久美子; 白澤, 浩; 清水, 文七; 磯野, 可一

    1997-01-01

    HPV16型E6/E7遺伝子発現を亜鉛により誘導可能な正常ヒト線維芽細胞を樹立し, HPV-16型E6/E7がヒト線維芽細胞に及ぼす影響を細胞学的及び分子生物学的に検討した。HPV-16型E6/E7の発現誘導による正常ヒト線維芽細胞の形態変化は観察されなかったが, E6/E7の発現誘導により, 細胞飽和密度の上昇が観察された。またマウス10T1/2細胞で観察されるE6のHMG-I(Y)発現への影響は, ヒト線維芽細胞では見いだされなかったが, E6/E7によりc-mycの発現が誘導されうることが明らかになった。この結果はE6/E7の細胞不死化に及ぼす過程を究明する上で有用であると考えられる。...

  14. Therapy of HPV 16-associated carcinoma with dendritic cell-based vaccines: In vitro priming of the effector cell responses by DC pulsed with tumour lysates and synthetic RAHYNIVTF peptide

    Czech Academy of Sciences Publication Activity Database

    Indrová, Marie; Bubeník, Jan; Šímová, Jana; Vonka, V.; Němečková, Š.; Mendoza, Luis; Reiniš, Milan

    2001-01-01

    Roč. 7, č. 1 (2001), s. 97-100 ISSN 1107-3756 R&D Projects: GA MZd NC5526; GA MZd NC45011; GA ČR GA312/98/0826; GA ČR GA312/99/0542; GA ČR GA301/00/0114; GA AV ČR IAA7052002 Institutional research plan: CEZ:AV0Z5052915 Keywords : tumour vaccines * HPV 16 * dendritic cells Subject RIV: FD - Oncology ; Hematology Impact factor: 1.689, year: 2001

  15. CpG oligodeoxynucleotides are effective in therapy of minimal residual tumour disease after chemotherapy or surgery in a murine model of MHC class I-deficient, HPV16-associated tumours

    Czech Academy of Sciences Publication Activity Database

    Reiniš, Milan; Šímová, Jana; Indrová, Marie; Bieblová, Jana; Bubeník, Jan

    2007-01-01

    Roč. 30, č. 5 (2007), s. 1247-1251 ISSN 1019-6439 R&D Projects: GA ČR GA301/04/0492; GA ČR GA301/06/0774 EU Projects: European Commission(XE) 18933 - CLINIGENE Grant - others:Liga proti rakovině, Praha(CZ) XX Institutional research plan: CEZ:AV0Z50520514 Keywords : HPV 16 * minimal residual tumour disease * CpG oligonucleotides Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.295, year: 2007

  16. Relationship between Humoral Immune Responses against HPV16, HPV18, HPV31 and HPV45 in 12-15 Year Old Girls Receiving Cervarix® or Gardasil® Vaccine.

    Directory of Open Access Journals (Sweden)

    Anna Godi

    Full Text Available Human papillomavirus (HPV vaccines confer protection against the oncogenic genotypes HPV16 and HPV18 through the generation of type-specific neutralizing antibodies raised against virus-like particles (VLP representing these genotypes. The vaccines also confer a degree of cross-protection against HPV31 and HPV45, which are genetically-related to the vaccine types HPV16 and HPV18, respectively, although the mechanism is less certain. There are a number of humoral immune measures that have been examined in relation to the HPV vaccines, including VLP binding, pseudovirus neutralization and the enumeration of memory B cells. While the specificity of responses generated against the vaccine genotypes are fairly well studied, the relationship between these measures in relation to non-vaccine genotypes is less certain.We carried out a comparative study of these immune measures against vaccine and non-vaccine genotypes using samples collected from 12-15 year old girls following immunization with three doses of either Cervarix® or Gardasil® HPV vaccine.The relationship between neutralizing and binding antibody titers and HPV-specific memory B cell levels for the vaccine genotypes, HPV16 and HPV18, were very good. The proportion of responders approached 100% for both vaccines while the magnitude of these responses induced by Cervarix® were generally higher than those following Gardasil® immunization. A similar pattern was found for the non-vaccine genotype HPV31, albeit at a lower magnitude compared to its genetically-related vaccine genotype, HPV16. However, both the enumeration of memory B cells and VLP binding responses against HPV45 were poorly related to its neutralizing antibody responses. Purified IgG derived from memory B cells demonstrated specificities similar to those found in the serum, including the capacity to neutralize HPV pseudoviruses.These data suggest that pseudovirus neutralization should be used as the preferred humoral immune

  17. Chemoimmunotherapy in mice carrying HPV16-associated, MHC class I+ and class I- tumours: effects of CBM-4A potentiated with IL-2, IL-12, GM-CSF and genetically modified tumour vaccines

    Czech Academy of Sciences Publication Activity Database

    Indrová, Marie; Bubeník, Jan; Mikyšková, Romana; Mendoza, Luis; Šímová, Jana; Bieblová, Jana; Jandlová, Táňa; Jinoch, P.; Šmahel, M.; Vonka, V.; Pajtasz-Piasecka, E.

    2003-01-01

    Roč. 22, č. 3 (2003), s. 691-695 ISSN 1019-6439 R&D Projects: GA MZd NC7148; GA ČR GA301/00/0114; GA ČR GA301/01/0985; GA AV ČR IAA5052203; GA AV ČR IAA7052002 Grant - others:KBN(PL) P04A12314; Liga proti rakovině(CZ) - Institutional research plan: CEZ:AV0Z5052915 Keywords : chemotherapy -induced residual tumour disease * immunotherapy * HPV16-associated tumours Subject RIV: FD - Oncology ; Hematology Impact factor: 2.536, year: 2003

  18. T-cell activation and early gene response in dogs.

    Directory of Open Access Journals (Sweden)

    Sally-Anne Mortlock

    Full Text Available T-cells play a crucial role in canine immunoregulation and defence against invading pathogens. Proliferation is fundamental to T-cell differentiation, homeostasis and immune response. Initiation of proliferation following receptor mediated stimuli requires a temporally programmed gene response that can be identified as immediate-early, mid- and late phases. The immediate-early response genes in T-cell activation engage the cell cycle machinery and promote subsequent gene activation events. Genes involved in this immediate-early response in dogs are yet to be identified. The present study was undertaken to characterise the early T-cell gene response in dogs to improve understanding of the genetic mechanisms regulating immune function. Gene expression profiles were characterised using canine gene expression microarrays and quantitative reverse transcription PCR (qRT-PCR, and paired samples from eleven dogs. Significant functional annotation clusters were identified following stimulation with phytohemagluttinin (PHA (5μg/ml, including the Toll-like receptor signaling pathway and phosphorylation pathways. Using strict statistical criteria, 13 individual genes were found to be differentially expressed, nine of which have ontologies that relate to proliferation and cell cycle control. These included, prostaglandin-endoperoxide synthase 2 (PTGS2/COX2, early growth response 1 (EGR1, growth arrest and DNA damage-inducible gene (GADD45B, phorbol-12-myristate-13-acetate-induced protein 1 (PMAIP1, V-FOS FBJ murine osteosarcoma viral oncogene homolog (FOS, early growth response 2 (EGR2, hemogen (HEMGN, polo-like kinase 2 (PLK2 and polo-like kinase 3 (PLK3. Differential gene expression was re-examined using qRT-PCR, which confirmed that EGR1, EGR2, PMAIP1, PTGS2, FOS and GADD45B were significantly upregulated in stimulated cells and ALAS2 downregulated. PTGS2 and EGR1 showed the highest levels of response in these dogs. Both of these genes are involved in

  19. The 3' region of Human Papillomavirus type 16 early mRNAs decrease expression

    DEFF Research Database (Denmark)

    Vinther, J.; Rosenstierne, M.W.; Kristiansen, Karen

    2005-01-01

    Background: High risk human papillomavirus (HR-HPV) infects mucosal surfaces and HR-HPV infection is required for development of cervical cancer. Accordingly, enforced expression of the early HR-HPV proteins can induce immortalisation of human cells. In most cervical cancers and cervical cancer...... cell lines the HR-HPV double stranded DNA genome has been integrated into the host cell genome. Methods: We have used a retroviral GUS reporter system to generate pools of stably transfected HaCaT and SiHa cells. The HPV-16 early sequences that are deleted upon integration of the HPV-16 genome...

  20. Alteration of gene expression by alcohol exposure at early neurulation.

    Science.gov (United States)

    Zhou, Feng C; Zhao, Qianqian; Liu, Yunlong; Goodlett, Charles R; Liang, Tiebing; McClintick, Jeanette N; Edenberg, Howard J; Li, Lang

    2011-02-21

    We have previously demonstrated that alcohol exposure at early neurulation induces growth retardation, neural tube abnormalities, and alteration of DNA methylation. To explore the global gene expression changes which may underline these developmental defects, microarray analyses were performed in a whole embryo mouse culture model that allows control over alcohol and embryonic variables. Alcohol caused teratogenesis in brain, heart, forelimb, and optic vesicle; a subset of the embryos also showed cranial neural tube defects. In microarray analysis (accession number GSM9545), adopting hypothesis-driven Gene Set Enrichment Analysis (GSEA) informatics and intersection analysis of two independent experiments, we found that there was a collective reduction in expression of neural specification genes (neurogenin, Sox5, Bhlhe22), neural growth factor genes [Igf1, Efemp1, Klf10 (Tieg), and Edil3], and alteration of genes involved in cell growth, apoptosis, histone variants, eye and heart development. There was also a reduction of retinol binding protein 1 (Rbp1), and de novo expression of aldehyde dehydrogenase 1B1 (Aldh1B1). Remarkably, four key hematopoiesis genes (glycophorin A, adducin 2, beta-2 microglobulin, and ceruloplasmin) were absent after alcohol treatment, and histone variant genes were reduced. The down-regulation of the neurospecification and the neurotrophic genes were further confirmed by quantitative RT-PCR. Furthermore, the gene expression profile demonstrated distinct subgroups which corresponded with two distinct alcohol-related neural tube phenotypes: an open (ALC-NTO) and a closed neural tube (ALC-NTC). Further, the epidermal growth factor signaling pathway and histone variants were specifically altered in ALC-NTO, and a greater number of neurotrophic/growth factor genes were down-regulated in the ALC-NTO than in the ALC-NTC embryos. This study revealed a set of genes vulnerable to alcohol exposure and genes that were associated with neural tube

  1. Using immediate-early genes to map hippocampal subregional functions

    Czech Academy of Sciences Publication Activity Database

    Kubík, Štěpán; Miyashita, T.; Guzowski, J. F.

    2007-01-01

    Roč. 14, č. 11 (2007), s. 758-770 ISSN 1072-0502 Grant - others:NIH(US) MH060123 Institutional research plan: CEZ:AV0Z50110509 Keywords : immediate-early genes * hippocampus * CA3 Subject RIV: FH - Neurology Impact factor: 4.037, year: 2007

  2. Telomerase activation by the E6 gene product of human papillomavirus type 16.

    Science.gov (United States)

    Klingelhutz, A J; Foster, S A; McDougall, J K

    1996-03-07

    Activation of telomerase, a ribonucleoprotein complex that synthesizes telomere repeat sequences, is linked to cell immortalization and is characteristic of most cell lines and tumours. Here we show that expression of the human papillomavirus type 16 (HPV-16) E6 protein activates telomerase in early-passage human keratinocytes and mammary epithelial cells. This activation was observed in cells pre-crisis, that is, before they became immortal, and occurred within one passage of retroviral infection with vectors expressing HPV-16 E6. Studies using HPV-16 E6 mutants showed that there was no correlation between the ability of the mutants to activate telomerase and their ability to target p53 for degradation, suggesting that telomerase activation by HPV-16 E6 is p53 independent. Keratinocytes expressing wild-type HPV-16 E6 have an extended lifespan, but do not become immortal, indicating that telomerase activation and E6-mediate degradation of p53 are insufficient for their immortalization. These results show that telomerase activation is an intrinsic, but insufficient, component of transformation by HPV.

  3. Distribution of human papilloma virus type 16 E6/E7 gene mutation in cervical precancer or cancer: A case control study in Guizhou Province, China.

    Science.gov (United States)

    Yang, Yingjie; Ren, Jie; Zhang, Qizhu

    2016-02-01

    HPV-16 varies geographically and is correlated with cervical cancer genesis and progression. This study aimed to determine the distribution of HPV-16 E6/E7 genetic variation in patients with invasive cervical cancer or precancer in Guizhou Province, China. A case-control study was designed, and the distribution of HPV-16 E6/E7 genetic variation was compared among women with cervical cancer, precancer, and sexually active without cervical lesion. HPV infection was detected through flow-through hybridization and gene chip techniques to determine the prevalence of HPV 16 E6/E7 genetic variation. Among 90 specimens (30 cervical cancer, 30 precancer, 30 controls), 81 were subjected to HPV-16 E6/E7 gene sequencing. The rates of DNA sequence mutation and amino acid mutation were 76.5% (62/81) and 66.7% (54/81), respectively. Both E6 and E7 genes showed higher mutation rate than their prototypes. The prevalence of E6/E7 mutation significantly differed between the cervical cancer and the controls (P prevalent in cervical cancer or precancer than those in the controls. The possible correlation between genetic variation and cancerigenesis may be used to design an HPV vaccine for cervical carcinoma. © 2015 Wiley Periodicals, Inc.

  4. Gene function in early mouse embryonic stem cell differentiation

    Directory of Open Access Journals (Sweden)

    Campbell Pearl A

    2007-03-01

    Full Text Available Abstract Background Little is known about the genes that drive embryonic stem cell differentiation. However, such knowledge is necessary if we are to exploit the therapeutic potential of stem cells. To uncover the genetic determinants of mouse embryonic stem cell (mESC differentiation, we have generated and analyzed 11-point time-series of DNA microarray data for three biologically equivalent but genetically distinct mESC lines (R1, J1, and V6.5 undergoing undirected differentiation into embryoid bodies (EBs over a period of two weeks. Results We identified the initial 12 hour period as reflecting the early stages of mESC differentiation and studied probe sets showing consistent changes of gene expression in that period. Gene function analysis indicated significant up-regulation of genes related to regulation of transcription and mRNA splicing, and down-regulation of genes related to intracellular signaling. Phylogenetic analysis indicated that the genes showing the largest expression changes were more likely to have originated in metazoans. The probe sets with the most consistent gene changes in the three cell lines represented 24 down-regulated and 12 up-regulated genes, all with closely related human homologues. Whereas some of these genes are known to be involved in embryonic developmental processes (e.g. Klf4, Otx2, Smn1, Socs3, Tagln, Tdgf1, our analysis points to others (such as transcription factor Phf21a, extracellular matrix related Lama1 and Cyr61, or endoplasmic reticulum related Sc4mol and Scd2 that have not been previously related to mESC function. The majority of identified functions were related to transcriptional regulation, intracellular signaling, and cytoskeleton. Genes involved in other cellular functions important in ESC differentiation such as chromatin remodeling and transmembrane receptors were not observed in this set. Conclusion Our analysis profiles for the first time gene expression at a very early stage of m

  5. Rat primary embryo fibroblast cells suppress transformation by the E6 and E7 genes of human papillomavirus type 16 in somatic hybrid cells.

    OpenAIRE

    Miyasaka, M; Takami, Y; Inoue, H; Hakura, A

    1991-01-01

    The E6 and E7 genes of human papillomavirus type 16 (HPV-16) transform established lines of rat cells but not rat cells in primary culture irrespective of the expression of the two genes. The reason for this difference between the susceptibilities of cell lines and primary cells was examined by using hybrid cells obtained by somatic cell fusion of rat cell lines transformed by the E6 and E7 genes of HPV-16 and freshly isolated rat embryo fibroblast cells. In these hybrid cells, transformed ph...

  6. The PCDH1 gene and asthma in early childhood

    DEFF Research Database (Denmark)

    Mortensen, Li J; Kreiner-Møller, Eskil; Hakonarson, Hakon

    2014-01-01

    Previous studies have suggested that variants in the protocadherin-1 (PCDH1) gene, which is important for cell-cell adhesion, are associated with asthma, bronchial, hyperresponsiveness and atopic dermatitis in school children. Our aim was to associate common variants of the PCDH1 gene with longit......Previous studies have suggested that variants in the protocadherin-1 (PCDH1) gene, which is important for cell-cell adhesion, are associated with asthma, bronchial, hyperresponsiveness and atopic dermatitis in school children. Our aim was to associate common variants of the PCDH1 gene...... with longitudinally assessed asthma phenotypes and atopic dermatitis in early childhood. We analysed eight single-nucleotide polymorphisms in PCDH1 from 411 children born to asthmatic mothers from the Copenhagen Prospective Studies on Asthma in Childhood birth cohort. Asthma and atopic dermatitis were diagnosed...

  7. Risk of HPV-16/18 Infections and Associated Cervical Abnormalities in Women Seropositive for Naturally Acquired Antibodies: Pooled Analysis Based on Control Arms of Two Large Clinical Trials.

    Science.gov (United States)

    Safaeian, Mahboobeh; Castellsagué, Xavier; Hildesheim, Allan; Wacholder, Sholom; Schiffman, Mark H; Bozonnat, Marie-Cécile; Baril, Laurence; Rosillon, Dominique

    2018-06-05

    Studies on the role of antibodies produced after infection with human papillomavirus 18 (HPV-18) and subsequent protection from HPV-18 infection have been conflicting, mainly due to inadequate sample size. We pooled data from the control arms of the Costa Rica Vaccine Trial and the PATRICIA trial. Using Poisson regression we compared the risk of newly detected 1-time HPV-18 infection, HPV-18 1-year persistent infection (12MPI), and HPV-18-associated atypical squamous cells of undetermined significance or greater (ASC-US+) lesions between HPV-18 seropositive and seronegative women. High HPV-18 antibodies at enrollment was associated with reduced subsequent HPV-18 detection (P trend = 0.001; relative rate [RR] = 0.69; 95% confidence interval [CI], 0.47-1.01 for the third quartile; RR = 0.63; 95% CI, 0.43-0.94 for the fourth quartile, compared to seronegative). The risk of 12MPI showed a decreasing trend with increasing antibodies (P trend = 0.06; RR = 0.72; 95% CI, 0.29-1.77; RR = 0.42; 95% CI, 0.13-1.32 for the third and fourth quartiles, respectively). Lastly, we observed a significant decreased risk of HPV-18 ASC-US+ with increasing antibody (P trend = 0.01; RR = 0.46; 95% CI, 0.21-0.97 for the fourth quartile). We also observed a significant decreased risk of HPV-16 infection, 12MPI, and ASC-US+ with increasing HPV-16 antibody level. High HPV-18 naturally acquired antibodies were associated with partial protection from future HPV-18 infections and associated lesions. NCT00128661 and NCT001226810.

  8. Gene expression profiles reveal key genes for early diagnosis and treatment of adamantinomatous craniopharyngioma.

    Science.gov (United States)

    Yang, Jun; Hou, Ziming; Wang, Changjiang; Wang, Hao; Zhang, Hongbing

    2018-04-23

    Adamantinomatous craniopharyngioma (ACP) is an aggressive brain tumor that occurs predominantly in the pediatric population. Conventional diagnosis method and standard therapy cannot treat ACPs effectively. In this paper, we aimed to identify key genes for ACP early diagnosis and treatment. Datasets GSE94349 and GSE68015 were obtained from Gene Expression Omnibus database. Consensus clustering was applied to discover the gene clusters in the expression data of GSE94349 and functional enrichment analysis was performed on gene set in each cluster. The protein-protein interaction (PPI) network was built by the Search Tool for the Retrieval of Interacting Genes, and hubs were selected. Support vector machine (SVM) model was built based on the signature genes identified from enrichment analysis and PPI network. Dataset GSE94349 was used for training and testing, and GSE68015 was used for validation. Besides, RT-qPCR analysis was performed to analyze the expression of signature genes in ACP samples compared with normal controls. Seven gene clusters were discovered in the differentially expressed genes identified from GSE94349 dataset. Enrichment analysis of each cluster identified 25 pathways that highly associated with ACP. PPI network was built and 46 hubs were determined. Twenty-five pathway-related genes that overlapped with the hubs in PPI network were used as signatures to establish the SVM diagnosis model for ACP. The prediction accuracy of SVM model for training, testing, and validation data were 94, 85, and 74%, respectively. The expression of CDH1, CCL2, ITGA2, COL8A1, COL6A2, and COL6A3 were significantly upregulated in ACP tumor samples, while CAMK2A, RIMS1, NEFL, SYT1, and STX1A were significantly downregulated, which were consistent with the differentially expressed gene analysis. SVM model is a promising classification tool for screening and early diagnosis of ACP. The ACP-related pathways and signature genes will advance our knowledge of ACP pathogenesis

  9. Early gene regulation of osteogenesis in embryonic stem cells

    KAUST Repository

    Kirkham, Glen R.

    2012-01-01

    The early gene regulatory networks (GRNs) that mediate stem cell differentiation are complex, and the underlying regulatory associations can be difficult to map accurately. In this study, the expression profiles of the genes Dlx5, Msx2 and Runx2 in mouse embryonic stem cells were monitored over a 48 hour period after exposure to the growth factors BMP2 and TGFβ1. Candidate GRNs of early osteogenesis were constructed based on published experimental findings and simulation results of Boolean and ordinary differential equation models were compared with our experimental data in order to test the validity of these models. Three gene regulatory networks were found to be consistent with the data, one of these networks exhibited sustained oscillation, a behaviour which is consistent with the general view of embryonic stem cell plasticity. The work cycle presented in this paper illustrates how mathematical modelling can be used to elucidate from gene expression profiles GRNs that are consistent with experimental data. © 2012 The Royal Society of Chemistry.

  10. Therapeutic misconception in early phase gene transfer trials.

    Science.gov (United States)

    Henderson, Gail E; Easter, Michele M; Zimmer, Catherine; King, Nancy M P; Davis, Arlene M; Rothschild, Barbra Bluestone; Churchill, Larry R; Wilfond, Benjamin S; Nelson, Daniel K

    2006-01-01

    Many subjects in early phase clinical trials expect to benefit in some way from the research intervention. It is understandable that people hope for improvement in their condition, no matter what the evidence. Yet unreasonable expectation of medical benefit may reflect problems with informed consent: Investigators may not disclose clearly that direct medical benefit from an early phase experimental intervention is unlikely or impossible, or subjects may not appreciate the differences between treatment and research. This paper presents findings from recent interviews with researchers and subjects and analysis of consent forms in early phase gene transfer research, a cutting-edge technology often called 'gene therapy'. We use three variables to construct a composite measure of therapeutic misconception TM, tapping misconceptions about the purposes of early phase research and the potential for direct medical benefit in these trials. Our multivariate model demonstrates the importance of both subject- and study-level factors as predictors of this TM index: education, disease type, and communication by study personnel about the likelihood of benefit. We hope that this work will deepen the discussion of how to define and measure TM, and refine the specification of factors that are related to subjects' TM.

  11. Early evolution of the LIM homeobox gene family

    Energy Technology Data Exchange (ETDEWEB)

    Srivastava, Mansi; Larroux, Claire; Lu, Daniel R; Mohanty, Kareshma; Chapman, Jarrod; Degnan, Bernard M; Rokhsar, Daniel S

    2010-01-01

    LIM homeobox (Lhx) transcription factors are unique to the animal lineage and have patterning roles during embryonic development in flies, nematodes and vertebrates, with a conserved role in specifying neuronal identity. Though genes of this family have been reported in a sponge and a cnidarian, the expression patterns and functions of the Lhx family during development in non-bilaterian phyla are not known. We identified Lhx genes in two cnidarians and a placozoan and report the expression of Lhx genes during embryonic development in Nematostella and the demosponge Amphimedon. Members of the six major LIM homeobox subfamilies are represented in the genomes of the starlet sea anemone, Nematostella vectensis, and the placozoan Trichoplax adhaerens. The hydrozoan cnidarian, Hydra magnipapillata, has retained four of the six Lhx subfamilies, but apparently lost two others. Only three subfamilies are represented in the haplosclerid demosponge Amphimedon queenslandica. A tandem cluster of three Lhx genes of different subfamilies and a gene containing two LIM domains in the genome of T. adhaerens (an animal without any neurons) indicates that Lhx subfamilies were generated by tandem duplication. This tandem cluster in Trichoplax is likely a remnant of the original chromosomal context in which Lhx subfamilies first appeared. Three of the six Trichoplax Lhx genes are expressed in animals in laboratory culture, as are all Lhx genes in Hydra. Expression patterns of Nematostella Lhx genes correlate with neural territories in larval and juvenile polyp stages. In the aneural demosponge, A. queenslandica, the three Lhx genes are expressed widely during development, including in cells that are associated with the larval photosensory ring. The Lhx family expanded and diversified early in animal evolution, with all six subfamilies already diverged prior to the cnidarian-placozoan-bilaterian last common ancestor. In Nematostella, Lhx gene expression is correlated with neural

  12. Early evolution of the LIM homeobox gene family

    Directory of Open Access Journals (Sweden)

    Degnan Bernard M

    2010-01-01

    Full Text Available Abstract Background LIM homeobox (Lhx transcription factors are unique to the animal lineage and have patterning roles during embryonic development in flies, nematodes and vertebrates, with a conserved role in specifying neuronal identity. Though genes of this family have been reported in a sponge and a cnidarian, the expression patterns and functions of the Lhx family during development in non-bilaterian phyla are not known. Results We identified Lhx genes in two cnidarians and a placozoan and report the expression of Lhx genes during embryonic development in Nematostella and the demosponge Amphimedon. Members of the six major LIM homeobox subfamilies are represented in the genomes of the starlet sea anemone, Nematostella vectensis, and the placozoan Trichoplax adhaerens. The hydrozoan cnidarian, Hydra magnipapillata, has retained four of the six Lhx subfamilies, but apparently lost two others. Only three subfamilies are represented in the haplosclerid demosponge Amphimedon queenslandica. A tandem cluster of three Lhx genes of different subfamilies and a gene containing two LIM domains in the genome of T. adhaerens (an animal without any neurons indicates that Lhx subfamilies were generated by tandem duplication. This tandem cluster in Trichoplax is likely a remnant of the original chromosomal context in which Lhx subfamilies first appeared. Three of the six Trichoplax Lhx genes are expressed in animals in laboratory culture, as are all Lhx genes in Hydra. Expression patterns of Nematostella Lhx genes correlate with neural territories in larval and juvenile polyp stages. In the aneural demosponge, A. queenslandica, the three Lhx genes are expressed widely during development, including in cells that are associated with the larval photosensory ring. Conclusions The Lhx family expanded and diversified early in animal evolution, with all six subfamilies already diverged prior to the cnidarian-placozoan-bilaterian last common ancestor. In

  13. Early signatures of regime shifts in gene expression dynamics

    Science.gov (United States)

    Pal, Mainak; Pal, Amit Kumar; Ghosh, Sayantari; Bose, Indrani

    2013-06-01

    Recently, a large number of studies have been carried out on the early signatures of sudden regime shifts in systems as diverse as ecosystems, financial markets, population biology and complex diseases. The signatures of regime shifts in gene expression dynamics are less systematically investigated. In this paper, we consider sudden regime shifts in the gene expression dynamics described by a fold-bifurcation model involving bistability and hysteresis. We consider two alternative models, models 1 and 2, of competence development in the bacterial population B. subtilis and determine some early signatures of the regime shifts between competence and noncompetence. We use both deterministic and stochastic formalisms for the purpose of our study. The early signatures studied include the critical slowing down as a transition point is approached, rising variance and the lag-1 autocorrelation function, skewness and a ratio of two mean first passage times. Some of the signatures could provide the experimental basis for distinguishing between bistability and excitability as the correct mechanism for the development of competence.

  14. Early signatures of regime shifts in gene expression dynamics

    International Nuclear Information System (INIS)

    Pal, Mainak; Pal, Amit Kumar; Ghosh, Sayantari; Bose, Indrani

    2013-01-01

    Recently, a large number of studies have been carried out on the early signatures of sudden regime shifts in systems as diverse as ecosystems, financial markets, population biology and complex diseases. The signatures of regime shifts in gene expression dynamics are less systematically investigated. In this paper, we consider sudden regime shifts in the gene expression dynamics described by a fold-bifurcation model involving bistability and hysteresis. We consider two alternative models, models 1 and 2, of competence development in the bacterial population B. subtilis and determine some early signatures of the regime shifts between competence and noncompetence. We use both deterministic and stochastic formalisms for the purpose of our study. The early signatures studied include the critical slowing down as a transition point is approached, rising variance and the lag-1 autocorrelation function, skewness and a ratio of two mean first passage times. Some of the signatures could provide the experimental basis for distinguishing between bistability and excitability as the correct mechanism for the development of competence. (paper)

  15. Persistent HPV16/18 infection in Indian women with the A-allele (rs6457617) of HLA-DQB1 and T-allele (rs16944) of IL-1β -511 is associated with development of cervical carcinoma.

    Science.gov (United States)

    Dutta, Sankhadeep; Chakraborty, Chandraditya; Mandal, Ranajit Kumar; Basu, Partha; Biswas, Jaydip; Roychoudhury, Susanta; Panda, Chinmay Kumar

    2015-07-01

    The aim of this study was to understand the association of human papillomavirus (HPV) type 16/18 infection and polymorphisms in the HLA-DQB1 (rs6457617) and IL-1β -511 (rs16944) loci with the development of uterine cervical cancer (CaCx). The distribution of HLA-DQB1 G > A and IL-1β -511 C/T polymorphisms was determined in HPV-negative cervical swabs from normal women (N = 111) and compared with cervical swabs of HPV-cleared normal women (once HPV infected followed by natural clearance of the infection, N = 86), HPV16/18-positive cervical intraepithelial neoplasia (CIN, N = 41) and CaCx biopsies (N = 107). The A-allele containing genotypes (i.e. G/A and A/A) of HLA-DQB1 was significantly associated with CaCx compared with HPV-negative [OR = 2.56(1.42-4.62), p = 0.001] or HPV-cleared [OR = 2.07(1.12-3.87), p = 0.01] normal women, whereas the T-allele containing genotypes (i.e. C/T and T/T) of IL-1β showed increased risk of CIN [OR = 3.68(0.97-16.35), p = 0.03; OR = 3.59(0.92-16.38), p = 0.03] and CaCx development [OR = 2.03(1.03-5.2), p = 0.02; OR = 2.25(0.96-5.31), p = 0.04] compared with HPV-negative or HPV-cleared normal women. Considering these two loci together, it was evident that the T- and A-alleles rendered significantly increased susceptibility for development of CIN and CaCx compared with HPV-negative and HPV-cleared normal women. Moreover, the T-allele of IL-1β showed increased susceptibility for CIN [OR = 3.62(0.85-17.95), p = 0.04] and CaCx [OR = 2.39(0.91-6.37), p = 0.05] development compared with the HPV-cleared women, even in the presence of the HLA-DQB1 G-allele. Thus, our data suggest that persistent HPV16/18 infection in the cervix due to the presence of the HLA-DQB1 A-allele and chronic inflammation due to the presence of the IL-1β -511 T-allele might predispose women to CaCx development.

  16. CpG Methylation Analysis of HPV16 in Laser Capture Microdissected Archival Tissue and Whole Tissue Sections from High Grade Anal Squamous Intraepithelial Lesions: A Potential Disease Biomarker.

    Directory of Open Access Journals (Sweden)

    Monica Molano

    Full Text Available Incidence and mortality rates of anal cancer are increasing globally. More than 90% of anal squamous cell carcinomas (ASCC are associated with human papillomavirus (HPV. Studies on HPV-related anogenital lesions have shown that patterns of methylation of viral and cellular DNA targets could potentially be developed as disease biomarkers. Lesion-specific DNA isolated from formalin-fixed paraffin-embedded (FFPE tissues from existing or prospective patient cohorts may constitute a valuable resource for methylation analysis. However, low concentrations of DNA make these samples technically challenging to analyse using existing methods. We therefore set out to develop a sensitive and reproducible nested PCR-pyrosequencing based method to accurately quantify methylation at 10 CpG sites within the E2BS1, E2BS2,3,4 and Sp1 binding sites in the viral upstream regulatory region of HPV16 genome. Methylation analyses using primary and nested PCR-pyrosequencing on 52 FFPE tissue [26 paired whole tissue sections (WTS and laser capture microdissected (LCM tissues] from patients with anal squamous intraepithelial lesions was performed. Using nested PCR, methylation results were obtained for the E2BS1, E2BS2,3,4 and Sp1 binding sites in 86.4% of the WTS and 81.8% of the LCM samples. Methylation patterns were strongly correlated within median values of matched pairs of WTS and LCM sections, but overall methylation was higher in LCM samples at different CpG sites. High grade lesions showed low methylation levels in the E2BS1 and E2BS2 regions, with increased methylation detected in the E2BS,3,4/Sp1 regions, showing the highest methylation at CpG site 37. The method developed is highly sensitive in samples with low amounts of DNA and demonstrated to be suitable for archival samples. Our data shows a possible role of specific methylation in the HPV16 URR for detection of HSIL.

  17. Learning abilities and disabilities: generalist genes in early adolescence.

    Science.gov (United States)

    Davis, Oliver S P; Haworth, Claire M A; Plomin, Robert

    2009-01-01

    The new view of cognitive neuropsychology that considers not just case studies of rare severe disorders but also common disorders, as well as normal variation and quantitative traits, is more amenable to recent advances in molecular genetics, such as genome-wide association studies, and advances in quantitative genetics, such as multivariate genetic analysis. A surprising finding emerging from multivariate quantitative genetic studies across diverse learning abilities is that most genetic influences are shared: they are "generalist", rather than "specialist". We exploited widespread access to inexpensive and fast Internet connections in the United Kingdom to assess over 5000 pairs of 12-year-old twins from the Twins Early Development Study (TEDS) on four distinct batteries: reading, mathematics, general cognitive ability (g) and, for the first time, language. Genetic correlations remain high among all of the measured abilities, with language as highly correlated genetically with g as reading and mathematics. Despite developmental upheaval, generalist genes remain important into early adolescence, suggesting optimal strategies for molecular genetic studies seeking to identify the genes of small effect that influence learning abilities and disabilities.

  18. Establishment and characterization of murine small cell lung carcinoma cell lines derived from HPV-16 E6/E7 transgenic mice.

    Science.gov (United States)

    Carraresi, Laura; Martinelli, Rosanna; Vannoni, Alessandro; Riccio, Massimo; Dembic, Maja; Tripodi, Sergio; Cintorino, Marcella; Santi, Spartaco; Bigliardi, Elisa; Carmellini, Mario; Rossini, Mara

    2006-01-08

    We have established two murine cell lines derived from Small Cell Lung Carcinomas (SCLCs) developed by HPV-E6/E7 transgenic mice. These cells named PPAP-9 and PPAP-10 were isolated from mice bearing tumors, 9 and 10 months old, respectively. The cells, 5 microm in diameter, express HPV oncoproteins and sustain tumor formation after subcutaneous injection in syngenic mice. A detailed analysis indicated the epithelial origin and the neuroendocrine differentiation of these cells. We showed by confocal immunofluorescence the expression of the epithelial marker cytokeratin 5, whose gene promoter was used to direct the expression of HPV E6/E. Cells express several neuroendocrine markers such as CGRP, MAP-2, Ash1, CgrA, Scg2. The neuroendocrine differentiation of these cells was further confirmed by electron microscopy demonstrating neuropeptides secreting granules in their cytoplasm. Furthermore, in agreement with the altered expression observed in the majority of human SCLC we showed in these cells the absence of both p53 and pRB and a dramatic reduction in the expression of Caveolin-1.

  19. Induction of anchorage-independent growth of human embryonic fibroblasts with a deletion in the short arm of chromosome 11 by human papillomavirus type 16 DNA

    International Nuclear Information System (INIS)

    Smits, H.L.; Raadsheer, E.; Rood, I.; Mehendale, S.; Slater, R.M.; van der Noordaa, J.; Ter Schegget, J.

    1988-01-01

    Human embryonic fibroblasts with a large deletion (11p11.11p15.1) in the short arm of one chromosome 11 (del-11 cells) appeared to be susceptible to transformation by early human papillomavirus type 16 (HPV-16) DNA, whereas diploid human embryonic fibroblasts were not. This difference in susceptibility might be explained by the absence of a tumor suppressor gene located within the deleted part on the short arm of chromosome 11. The presence of abundant viral early-gene transcripts in transformed cells suggests that transformation was induced by an elevated level of an HPV-16 early-gene product(s). The low transcriptional activity of HPV-16 in diploid cells may indicate that cellular genes affect viral transcription. Interruption of the HPV-16 E2 early open reading frame is probably required for high-level HPV-16 early-gene expression driven from the homologous enhancer-promoter region

  20. The Curcumin Analogue 1,5-Bis(2-hydroxyphenyl-1,4-pentadiene-3-one Induces Apoptosis and Downregulates E6 and E7 Oncogene Expression in HPV16 and HPV18-Infected Cervical Cancer Cells

    Directory of Open Access Journals (Sweden)

    Felicia Paulraj

    2015-06-01

    Full Text Available In an effort to study curcumin analogues as an alternative to improve the therapeutic efficacy of curcumin, we screened the cytotoxic potential of four diarylpentanoids using the HeLa and CaSki cervical cancer cell lines. Determination of their EC50 values indicated relatively higher potency of 1,5-bis(2-hydroxyphenyl-1,4-pentadiene-3-one (MS17, 1.03 ± 0.5 μM; 2.6 ± 0.9 μM and 1,5-bis(4-hydroxy-3-methoxyphenyl-1,4-pentadiene-3-one (MS13, 2.8 ± 0.4; 6.7 ± 2.4 μM in CaSki and HeLa, respectively, with significantly greater growth inhibition at 48 and 72 h of treatment compared to the other analogues or curcumin. Based on cytotoxic and anti-proliferative activity, MS17 was selected for comprehensive apoptotic studies. At 24 h of treatment, fluorescence microscopy detected that MS17-exposed cells exhibited significant morphological changes consistent with apoptosis, corroborated by an increase in nucleosomal enrichment due to DNA fragmentation in HeLa and CaSki cells and activation of caspase-3 activity in CaSki cells. Quantitative real-time PCR also detected significant down-regulation of HPV18- and HPV16-associated E6 and E7 oncogene expression following treatment. The overall data suggests that MS17 treatment has cytotoxic, anti-proliferative and apoptosis-inducing potential in HPV-positive cervical cancer cells. Furthermore, its role in down-regulation of HPV-associated oncogenes responsible for cancer progression merits further investigation into its chemotherapeutic role for cervical cancer.

  1. Comparison of HPV detection technologies: Hybrid capture 2, PreTect HPV-Proofer and analysis of HPV DNA viral load in HPV16, HPV18 and HPV33 E6/E7 mRNA positive specimens.

    LENUS (Irish Health Repository)

    Keegan, Helen

    2012-02-01

    Human papillomavirus (HPV) testing using molecular methods in liquid based cytology (LBC) specimens may be useful as an adjunct to cervical screening by cytology. We compared the positivity rate of the commercially available HPV DNA method hybrid capture 2 (hc2) and the commercially available E6\\/E7 mRNA method PreTect HPV-Proofer in cytological specimens (n=299). LBC specimens collected (n=299) represented the following cervical cytological disease categories: Normal (n=60), borderline nuclear abnormalities (BNA) (n=34), CIN1 (n=121), CIN2 (n=60), CIN3 (n=24). Overall, 69% (205\\/299) of the cases were positive by hc2 and 38% (112\\/299) of the cases were positive by PreTect HPV-Proofer. Concordance rates between the two tests were highest in the high-grade cytology cases (CIN2: 67% and CIN3: 83%) and the normal cytology cases (88%) and lowest in the BNA and CIN1 categories (56% and 52%). HPV DNA viral load analyses were carried out on HPV16 (n=55), HPV18 (n=9) and HPV33 (n=13) samples that were positive by PreTect HPV-Proofer. The sensitivity and specificity of PreTect HPV-Proofer and the hc2 DNA test for the detection of high-grade cytology (i.e. CIN2+) were 71.4% and 75.8% vs 100% and 43.7%, respectively. The relatively low detection rate observed by PreTect HPV-Proofer in the whole range of cytological positive cases, combined with a relatively higher specificity and PPV, suggests that PreTect HPV-Proofer may be more useful than hc2 for triage and in predicting high-grade disease.

  2. Highly preserved consensus gene modules in human papilloma virus 16 positive cervical cancer and head and neck cancers.

    Science.gov (United States)

    Zhang, Xianglan; Cha, In-Ho; Kim, Ki-Yeol

    2017-12-26

    In this study, we investigated the consensus gene modules in head and neck cancer (HNC) and cervical cancer (CC). We used a publicly available gene expression dataset, GSE6791, which included 42 HNC, 14 normal head and neck, 20 CC and 8 normal cervical tissue samples. To exclude bias because of different human papilloma virus (HPV) types, we analyzed HPV16-positive samples only. We identified 3824 genes common to HNC and CC samples. Among these, 977 genes showed high connectivity and were used to construct consensus modules. We demonstrated eight consensus gene modules for HNC and CC using the dissimilarity measure and average linkage hierarchical clustering methods. These consensus modules included genes with significant biological functions, including ATP binding and extracellular exosome. Eigengen network analysis revealed the consensus modules were highly preserved with high connectivity. These findings demonstrate that HPV16-positive head and neck and cervical cancers share highly preserved consensus gene modules with common potentially therapeutic targets.

  3. Integrative analyses reveal novel strategies in HPV11,-16 and-45 early infection

    DEFF Research Database (Denmark)

    Kaczkowski, Bogumil; Rossing, Maria; Andersen, Ditte

    2012-01-01

    of genes not previously implicated in HPV biology, such as the PSG family and ANKRD1, and of genes implicated in the biology of other viruses, e. g. MX1, IFI44 and DDX60. Carcinogenesis-related genes, e. g. ABL2, MGLL and CYR61, were upregulated by high-risk HPV16 and -45. The integrative analysis revealed...... the suppression of DNA repair by HPV11 and -16, and downregulation of cytoskeleton genes by all HPV types. Various signalling pathways were affected by the HPVs: IL-2 by HPV11; JAK-STAT by HPV16; and TGF-beta, NOTCH and tyrosine kinase signalling by HPV45. This study uncovered novel strategies employed by HPV...... to establish infection and promote uncontrolled growth....

  4. Effects of Transcranial Direct Current Stimulation on Expression of Immediate Early Genes (IEG’s)

    Science.gov (United States)

    2015-12-01

    TRANSCRANIAL DIRECT CURRENT STIMULATION OF EXPRESSION OF IMMEDIATE EARLY GENES (IEG’S) Jessica...AND SUBTITLE Effects of Transcranial Direct Current Stimulation on Expression of Immediate Early Genes (IEG’s) 5a. CONTRACT NUMBER In-House 5b...community in better understanding what is occurring biologically during tDCS. 15. SUBJECT TERMS Transcranial direct current stimulation

  5. Dynamic changes in the interchromosomal interaction of early histone gene loci during development of sea urchin.

    Science.gov (United States)

    Matsushita, Masaya; Ochiai, Hiroshi; Suzuki, Ken-Ichi T; Hayashi, Sayaka; Yamamoto, Takashi; Awazu, Akinori; Sakamoto, Naoaki

    2017-12-15

    The nuclear positioning and chromatin dynamics of eukaryotic genes are closely related to the regulation of gene expression, but they have not been well examined during early development, which is accompanied by rapid cell cycle progression and dynamic changes in nuclear organization, such as nuclear size and chromatin constitution. In this study, we focused on the early development of the sea urchin Hemicentrotus pulcherrimus and performed three-dimensional fluorescence in situ hybridization of gene loci encoding early histones (one of the types of histone in sea urchin). There are two non-allelic early histone gene loci per sea urchin genome. We found that during the morula stage, when the early histone gene expression levels are at their maximum, interchromosomal interactions were often formed between the early histone gene loci on separate chromosomes and that the gene loci were directed to locate to more interior positions. Furthermore, these interactions were associated with the active transcription of the early histone genes. Thus, such dynamic interchromosomal interactions may contribute to the efficient synthesis of early histone mRNA during the morula stage of sea urchin development. © 2017. Published by The Company of Biologists Ltd.

  6. Influence of Isoflurane on Immediate-Early Gene Expression.

    Science.gov (United States)

    Bunting, Kristopher M; Nalloor, Rebecca I; Vazdarjanova, Almira

    2015-01-01

    Anterograde amnesia is a hallmark effect of volatile anesthetics. Isoflurane is known to affect both the translation and transcription of plasticity-associated genes required for normal memory formation in many brain regions. What is not known is whether isoflurane anesthesia prevents the initiation of transcription or whether it halts transcription already in progress. We tested the hypothesis that general anesthesia with isoflurane prevents learning-induced initiation of transcription of several memory-associated immediate-early genes (IEGs) correlated with amnesia; we also assessed whether it stops transcription initiated prior to anesthetic administration. Using a Tone Fear Conditioning paradigm, rats were trained to associate a tone with foot-shock. Animals received either no anesthesia, anesthesia immediately after training, or anesthesia before, during, and after training. Animals were either sacrificed after training or tested 24 h later for long-term memory. Using Cellular Compartment Analysis of Temporal Activity by Fluorescence in situ Hybridization (catFISH), we examined the percentage of neurons expressing the IEGs Arc/Arg3.1 and Zif268/Egr1/Ngfi-A/Krox-24 in the dorsal hippocampus, primary somatosensory cortex, and primary auditory cortex. On a cellular level, isoflurane administered at high doses (general anesthesia) prevented initiation of transcription, but did not stop transcription of Arc and Zif268 mRNA initiated prior to anesthesia. On a behavioral level, the same level of isoflurane anesthesia produced anterograde amnesia for fear conditioning when administered before and during training, but did not produce retrograde amnesia when administered immediately after training. General anesthesia with isoflurane prevents initiation of learning-related transcription but does not stop ongoing transcription of two plasticity-related IEGs, Arc and Zif268, a pattern of disruption that parallels the effects of isoflurane on memory formation. Combined with

  7. Influence of isoflurane on Immediate-Early Gene expression

    Directory of Open Access Journals (Sweden)

    Kristopher M Bunting

    2016-01-01

    Full Text Available Background: Anterograde amnesia is a hallmark effect of volatile anesthetics. Isoflurane is known to affect both the translation and transcription of plasticity-associated genes required for normal memory formation in many brain regions. What is not known is whether isoflurane anesthesia prevents the initiation of transcription or whether it halts transcription already in progress. We tested the hypothesis that general anesthesia with isoflurane prevents learning-induced initiation of transcription of several memory-associated immediate-early genes (IEGs correlated with amnesia; we also assessed whether it stops transcription initiated prior to anesthetic administration.Methods: Using a Tone Fear Conditioning paradigm, rats were trained to associate a tone with foot-shock. Animals received either no anesthesia, anesthesia immediately after training, or anesthesia before, during, and after training. Animals were either sacrificed after training or tested 24 hours later for memory. Using Cellular Compartment Analysis of Temporal Activity by Fluorescence in situ Hybridization (catFISH, we examined the percentage of neurons expressing the IEGs Arc/Arg3.1 and Zif268/Egr1/Ngfi-A/Krox-24 in the dorsal hippocampus, primary somatosensory cortex, and primary auditory cortex.Results: On a cellular level, isoflurane administered at high doses (general anesthesia prevented initiation of transcription, but did not stop transcription of Arc and Zif268 mRNA initiated prior to anesthesia. On a behavioral level, the same level of isoflurane anesthesia produced anterograde amnesia for fear conditioning when administered before and during training, but did not produce retrograde amnesia when administered immediately after training. Conclusions: General anesthesia with isoflurane prevents initiation of learning-related transcription but does not stop ongoing transcription of two plasticity-related IEGs, Arc and Zif268, a pattern of disruption that parallels the

  8. Early pregnancy peripheral blood gene expression and risk of preterm delivery: a nested case control study

    Directory of Open Access Journals (Sweden)

    Muhie Seid Y

    2009-12-01

    Full Text Available Abstract Background Preterm delivery (PTD is a significant public health problem associated with greater risk of mortality and morbidity in infants and mothers. Pathophysiologic processes that may lead to PTD start early in pregnancy. We investigated early pregnancy peripheral blood global gene expression and PTD risk. Methods As part of a prospective study, ribonucleic acid was extracted from blood samples (collected at 16 weeks gestational age from 14 women who had PTD (cases and 16 women who delivered at term (controls. Gene expressions were measured using the GeneChip® Human Genome U133 Plus 2.0 Array. Student's T-test and fold change analysis were used to identify differentially expressed genes. We used hierarchical clustering and principle components analysis to characterize signature gene expression patterns among cases and controls. Pathway and promoter sequence analyses were used to investigate functions and functional relationships as well as regulatory regions of differentially expressed genes. Results A total of 209 genes, including potential candidate genes (e.g. PTGDS, prostaglandin D2 synthase 21 kDa, were differentially expressed. A set of these genes achieved accurate pre-diagnostic separation of cases and controls. These genes participate in functions related to immune system and inflammation, organ development, metabolism (lipid, carbohydrate and amino acid and cell signaling. Binding sites of putative transcription factors such as EGR1 (early growth response 1, TFAP2A (transcription factor AP2A, Sp1 (specificity protein 1 and Sp3 (specificity protein 3 were over represented in promoter regions of differentially expressed genes. Real-time PCR confirmed microarray expression measurements of selected genes. Conclusions PTD is associated with maternal early pregnancy peripheral blood gene expression changes. Maternal early pregnancy peripheral blood gene expression patterns may be useful for better understanding of PTD

  9. Gene therapy, early promises, subsequent problems, and recent breakthroughs.

    Science.gov (United States)

    Razi Soofiyani, Saeideh; Baradaran, Behzad; Lotfipour, Farzaneh; Kazemi, Tohid; Mohammadnejad, Leila

    2013-01-01

    Gene therapy is one of the most attractive fields in medicine. The concept of gene delivery to tissues for clinical applications has been discussed around half a century, but scientist's ability to manipulate genetic material via recombinant DNA technology made this purpose to reality. Various approaches, such as viral and non-viral vectors and physical methods, have been developed to make gene delivery safer and more efficient. While gene therapy initially conceived as a way to treat life-threatening disorders (inborn errors, cancers) refractory to conventional treatment, to date gene therapy is considered for many non-life-threatening conditions including those adversely influence on a patient's quality of life. Gene therapy has made significant progress, including tangible success, although much slower than was initially predicted. Although, gene therapies still at a fairly primitive stage, it is firmly science based. There is justifiable hope that with enhanced pathobiological understanding and biotechnological improvements, gene therapy will be a standard part of clinical practice within 20 years.

  10. Gene Therapy, Early Promises, Subsequent Problems, and Recent Breakthroughs

    Directory of Open Access Journals (Sweden)

    Saeideh Razi Soofiyani

    2013-08-01

    Full Text Available Gene therapy is one of the most attractive fields in medicine. The concept of gene delivery to tissues for clinical applications has been discussed around half a century, but scientist’s ability to manipulate genetic material via recombinant DNA technology made this purpose to reality. Various approaches, such as viral and non-viral vectors and physical methods, have been developed to make gene delivery safer and more efficient. While gene therapy initially conceived as a way to treat life-threatening disorders (inborn errors, cancers refractory to conventional treatment, to date gene therapy is considered for many non–life-threatening conditions including those adversely influence on a patient’s quality of life. Gene therapy has made significant progress, including tangible success, although much slower than was initially predicted. Although, gene therapies still at a fairly primitive stage, it is firmly science based. There is justifiable hope that with enhanced pathobiological understanding and biotechnological improvements, gene therapy will be a standard part of clinical practice within 20 years.

  11. Early events in the evolution of spider silk genes.

    Directory of Open Access Journals (Sweden)

    James Starrett

    Full Text Available Silk spinning is essential to spider ecology and has had a key role in the expansive diversification of spiders. Silk is composed primarily of proteins called spidroins, which are encoded by a multi-gene family. Spidroins have been studied extensively in the derived clade, Orbiculariae (orb-weavers, from the suborder Araneomorphae ('true spiders'. Orbicularians produce a suite of different silks, and underlying this repertoire is a history of duplication and spidroin gene divergence. A second class of silk proteins, Egg Case Proteins (ECPs, is known only from the orbicularian species, Lactrodectus hesperus (Western black widow. In L. hesperus, ECPs bond with tubuliform spidroins to form egg case silk fibers. Because most of the phylogenetic diversity of spiders has not been sampled for their silk genes, there is limited understanding of spidroin gene family history and the prevalence of ECPs. Silk genes have not been reported from the suborder Mesothelae (segmented spiders, which diverged from all other spiders >380 million years ago, and sampling from Mygalomorphae (tarantulas, trapdoor spiders and basal araneomorph lineages is sparse. In comparison to orbicularians, mesotheles and mygalomorphs have a simpler silk biology and thus are hypothesized to have less diversity of silk genes. Here, we present cDNAs synthesized from the silk glands of six mygalomorph species, a mesothele, and a non-orbicularian araneomorph, and uncover a surprisingly rich silk gene diversity. In particular, we find ECP homologs in the mesothele, suggesting that ECPs were present in the common ancestor of extant spiders, and originally were not specialized to complex with tubuliform spidroins. Furthermore, gene-tree/species-tree reconciliation analysis reveals that numerous spidroin gene duplications occurred after the split between Mesothelae and Opisthothelae (Mygalomorphae plus Araneomorphae. We use the spidroin gene tree to reconstruct the evolution of amino acid

  12. Diffuse optical measurements of head and neck tumor hemodynamics for early prediction of radiation therapy (Conference Presentation)

    Science.gov (United States)

    Dong, Lixin; Kudrimoti, Mahesh; Irwin, Daniel; Chen, Li; Shang, Yu; Li, Xingzhe; Stevens, Scott D.; Shelton, Brent J.; Yu, Guoqiang

    2016-03-01

    Radiation therapy is a principal modality for head and neck cancers and its efficacy depends on tumor hemodynamics. Our laboratory developed a hybrid diffuse optical instrument allowing for simultaneous measurements of tumor blood flow and oxygenation. In this study, the clinically involved cervical lymph node was monitored by the hybrid instrument once a week over the treatment period of seven weeks. Based on treatment outcomes within one year, patients were classified into a complete response group (CR) and an incomplete response group (IR) with remote metastasis and/or local recurrence. A linear mixed models was used to compare tumor hemodynamic responses to the treatment between the two groups. Interestingly, we found that human papilloma virus (HPV-16) status largely affected tumor hemodynamic responses. For HPV-16 negative tumors, significant differences in blood flow index (BFI, p = 0.007) and reduced scattering coefficient (μs', p = 0.0005) were observed between the two groups; IR tumors exhibited higher μs' values and a continuous increase in BFI over the treatment period. For HPV-16 positive tumors, oxygenated hemoglobin concentration ([HbO2]) and blood oxygen saturation (StO2) were significant different (p = 0.003 and 0.01, respectively); IR group showed lower [HbO2] and StO2. Our results imply HPV-16 negative tumors with higher density of vasculature (μs') and higher blood flow show poor responses to radiotherapy and HPV-16 positive tumors with lower tissue oxygenation level (lower StO2 and [HbO2]) exhibit poor treatment outcomes. Our diffuse optical measurements show the great potential for early prediction of radiotherapy in head and neck cancers.

  13. Evolutionary analysis of the kinesin light chain genes in the yellow fever mosquito Aedes aegypti: gene duplication as a source for novel early zygotic genes.

    Science.gov (United States)

    Biedler, James K; Tu, Zhijian

    2010-07-08

    The maternal zygotic transition marks the time at which transcription from the zygotic genome is initiated and a subset of maternal RNAs are progressively degraded in the developing embryo. A number of early zygotic genes have been identified in Drosophila melanogaster and comparisons to sequenced mosquito genomes suggest that some of these early zygotic genes such as bottleneck are fast-evolving or subject to turnover in dipteran insects. One objective of this study is to identify early zygotic genes from the yellow fever mosquito Aedes aegypti to study their evolution. We are also interested in obtaining early zygotic promoters that will direct transgene expression in the early embryo as part of a Medea gene drive system. Two novel early zygotic kinesin light chain genes we call AaKLC2.1 and AaKLC2.2 were identified by transcriptome sequencing of Aedes aegypti embryos at various time points. These two genes have 98% nucleotide and amino acid identity in their coding regions and show transcription confined to the early zygotic stage according to gene-specific RT-PCR analysis. These AaKLC2 genes have a paralogous gene (AaKLC1) in Ae. aegypti. Phylogenetic inference shows that an ortholog to the AaKLC2 genes is only found in the sequenced genome of Culex quinquefasciatus. In contrast, AaKLC1 gene orthologs are found in all three sequenced mosquito species including Anopheles gambiae. There is only one KLC gene in D. melanogaster and other sequenced holometabolous insects that appears to be similar to AaKLC1. Unlike AaKLC2, AaKLC1 is expressed in all life stages and tissues tested, which is consistent with the expression pattern of the An. gambiae and D. melanogaster KLC genes. Phylogenetic inference also suggests that AaKLC2 genes and their likely C. quinquefasciatus ortholog are fast-evolving genes relative to the highly conserved AaKLC1-like paralogs. Embryonic injection of a luciferase reporter under the control of a 1 kb fragment upstream of the AaKLC2.1 start

  14. Evolutionary analysis of the kinesin light chain genes in the yellow fever mosquito Aedes aegypti: gene duplication as a source for novel early zygotic genes

    Directory of Open Access Journals (Sweden)

    Tu Zhijian

    2010-07-01

    Full Text Available Abstract Background The maternal zygotic transition marks the time at which transcription from the zygotic genome is initiated and a subset of maternal RNAs are progressively degraded in the developing embryo. A number of early zygotic genes have been identified in Drosophila melanogaster and comparisons to sequenced mosquito genomes suggest that some of these early zygotic genes such as bottleneck are fast-evolving or subject to turnover in dipteran insects. One objective of this study is to identify early zygotic genes from the yellow fever mosquito Aedes aegypti to study their evolution. We are also interested in obtaining early zygotic promoters that will direct transgene expression in the early embryo as part of a Medea gene drive system. Results Two novel early zygotic kinesin light chain genes we call AaKLC2.1 and AaKLC2.2 were identified by transcriptome sequencing of Aedes aegypti embryos at various time points. These two genes have 98% nucleotide and amino acid identity in their coding regions and show transcription confined to the early zygotic stage according to gene-specific RT-PCR analysis. These AaKLC2 genes have a paralogous gene (AaKLC1 in Ae. aegypti. Phylogenetic inference shows that an ortholog to the AaKLC2 genes is only found in the sequenced genome of Culex quinquefasciatus. In contrast, AaKLC1 gene orthologs are found in all three sequenced mosquito species including Anopheles gambiae. There is only one KLC gene in D. melanogaster and other sequenced holometabolous insects that appears to be similar to AaKLC1. Unlike AaKLC2, AaKLC1 is expressed in all life stages and tissues tested, which is consistent with the expression pattern of the An. gambiae and D. melanogaster KLC genes. Phylogenetic inference also suggests that AaKLC2 genes and their likely C. quinquefasciatus ortholog are fast-evolving genes relative to the highly conserved AaKLC1-like paralogs. Embryonic injection of a luciferase reporter under the control of a

  15. Development of a Combination Cell and Gene Therapy Approach for Early-Stage Breast Cancer

    National Research Council Canada - National Science Library

    Lewis, Michael T

    2005-01-01

    The unique biology of the breast presents the opportunity to these cell and gene therapy techniques in a way that circumvents many of these technical limitations for the treatment of early stage breast cancer...

  16. Differential neutrophil gene expression in early bovine pregnancy

    Directory of Open Access Journals (Sweden)

    Kizaki Keiichiro

    2013-02-01

    Full Text Available Abstract Background In food production animals, especially cattle, the diagnosis of gestation is important because the timing of gestation directly affects the running of farms. Various methods have been used to detect gestation, but none of them are ideal because of problems with the timing of detection or the accuracy, simplicity, or cost of the method. A new method for detecting gestation, which involves assessing interferon-tau (IFNT-stimulated gene expression in peripheral blood leukocytes (PBL, was recently proposed. PBL fractionation methods were used to examine whether the expression profiles of various PBL populations could be used as reliable diagnostic markers of bovine gestation. Methods PBL were collected on days 0 (just before artificial insemination, 7, 14, 17, 21, and 28 of gestation. The gene expression levels of the PBL were assessed with microarray analysis and/or quantitative real-time reverse transcription (q PCR. PBL fractions were collected by flow cytometry or density gradient cell separation using Histopaque 1083 or Ficoll-Conray solutions. The expression levels of four IFNT-stimulated genes, interferon-stimulated protein 15 kDa (ISG15, myxovirus-resistance (MX 1 and 2, and 2′-5′-oligoadenylate synthetase (OAS1, were then analyzed in each fraction through day 28 of gestation using qPCR. Results Microarray analysis detected 72 and 28 genes in whole PBL that were significantly higher on days 14 and 21 of gestation, respectively, than on day 0. The upregulated genes included IFNT-stimulated genes. The expression levels of these genes increased with the progression of gestation until day 21. In flow cytometry experiments, on day 14 the expression levels of all of the genes were significantly higher in the granulocyte fraction than in the other fractions. Their expression gradually decreased through day 28 of gestation. Strong correlations were observed between the expression levels of the four genes in the granulocyte

  17. Weighted gene co-expression network analysis reveals potential genes involved in early metamorphosis process in sea cucumber Apostichopus japonicus.

    Science.gov (United States)

    Li, Yongxin; Kikuchi, Mani; Li, Xueyan; Gao, Qionghua; Xiong, Zijun; Ren, Yandong; Zhao, Ruoping; Mao, Bingyu; Kondo, Mariko; Irie, Naoki; Wang, Wen

    2018-01-01

    Sea cucumbers, one main class of Echinoderms, have a very fast and drastic metamorphosis process during their development. However, the molecular basis under this process remains largely unknown. Here we systematically examined the gene expression profiles of Japanese common sea cucumber (Apostichopus japonicus) for the first time by RNA sequencing across 16 developmental time points from fertilized egg to juvenile stage. Based on the weighted gene co-expression network analysis (WGCNA), we identified 21 modules. Among them, MEdarkmagenta was highly expressed and correlated with the early metamorphosis process from late auricularia to doliolaria larva. Furthermore, gene enrichment and differentially expressed gene analysis identified several genes in the module that may play key roles in the metamorphosis process. Our results not only provide a molecular basis for experimentally studying the development and morphological complexity of sea cucumber, but also lay a foundation for improving its emergence rate. Copyright © 2017 Elsevier Inc. All rights reserved.

  18. GCN5 Regulates FGF Signaling and Activates Selective MYC Target Genes during Early Embryoid Body Differentiation

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    Li Wang

    2018-01-01

    Full Text Available Precise control of gene expression during development is orchestrated by transcription factors and co-regulators including chromatin modifiers. How particular chromatin-modifying enzymes affect specific developmental processes is not well defined. Here, we report that GCN5, a histone acetyltransferase essential for embryonic development, is required for proper expression of multiple genes encoding components of the fibroblast growth factor (FGF signaling pathway in early embryoid bodies (EBs. Gcn5−/− EBs display deficient activation of ERK and p38, mislocalization of cytoskeletal components, and compromised capacity to differentiate toward mesodermal lineage. Genomic analyses identified seven genes as putative direct targets of GCN5 during early differentiation, four of which are cMYC targets. These findings established a link between GCN5 and the FGF signaling pathway and highlighted specific GCN5-MYC partnerships in gene regulation during early differentiation.

  19. Influence of Isoflurane on Immediate-Early Gene Expression

    OpenAIRE

    Bunting, Kristopher M.; Nalloor, Rebecca I.; Vazdarjanova, Almira

    2016-01-01

    Background: Anterograde amnesia is a hallmark effect of volatile anesthetics. Isoflurane is known to affect both the translation and transcription of plasticity-associated genes required for normal memory formation in many brain regions. What is not known is whether isoflurane anesthesia prevents the initiation of transcription or whether it halts transcription already in progress. We tested the hypothesis that general anesthesia with isoflurane prevents learning-induced initiation of transcr...

  20. Transient expression of fusion gene coding for the HPV-16 epitopes fused to the sequence of potyvirus coat protein using different means of inoculation of Nicotiana benthamiana and Brassica rapa, cv. Rapa plants

    Czech Academy of Sciences Publication Activity Database

    Hoffmeisterová, Hana; Čeřovská, Noemi; Moravec, Tomáš; Plchová, Helena; Kmoníčková, Jitka; Velemínský, Jiří

    2008-01-01

    Roč. 94, č. 3 (2008), s. 261-267 ISSN 0167-6857 R&D Projects: GA ČR GA521/06/0973 Institutional research plan: CEZ:AV0Z50380511 Keywords : Edible vaccine * epitope expression * E7 oncogen Subject RIV: EE - Microbiology, Virology Impact factor: 1.017, year: 2008

  1. Early maternal alcohol consumption alters hippocampal DNA methylation, gene expression and volume in a mouse model.

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    Heidi Marjonen

    Full Text Available The adverse effects of alcohol consumption during pregnancy are known, but the molecular events that lead to the phenotypic characteristics are unclear. To unravel the molecular mechanisms, we have used a mouse model of gestational ethanol exposure, which is based on maternal ad libitum ingestion of 10% (v/v ethanol for the first 8 days of gestation (GD 0.5-8.5. Early neurulation takes place by the end of this period, which is equivalent to the developmental stage early in the fourth week post-fertilization in human. During this exposure period, dynamic epigenetic reprogramming takes place and the embryo is vulnerable to the effects of environmental factors. Thus, we hypothesize that early ethanol exposure disrupts the epigenetic reprogramming of the embryo, which leads to alterations in gene regulation and life-long changes in brain structure and function. Genome-wide analysis of gene expression in the mouse hippocampus revealed altered expression of 23 genes and three miRNAs in ethanol-exposed, adolescent offspring at postnatal day (P 28. We confirmed this result by using two other tissues, where three candidate genes are known to express actively. Interestingly, we found a similar trend of upregulated gene expression in bone marrow and main olfactory epithelium. In addition, we observed altered DNA methylation in the CpG islands upstream of the candidate genes in the hippocampus. Our MRI study revealed asymmetry of brain structures in ethanol-exposed adult offspring (P60: we detected ethanol-induced enlargement of the left hippocampus and decreased volume of the left olfactory bulb. Our study indicates that ethanol exposure in early gestation can cause changes in DNA methylation, gene expression, and brain structure of offspring. Furthermore, the results support our hypothesis of early epigenetic origin of alcohol-induced disorders: changes in gene regulation may have already taken place in embryonic stem cells and therefore can be seen in

  2. Advances in gene therapy and early imaging monitoring for avascular necrosis of the femoral head

    International Nuclear Information System (INIS)

    Wang Peng; Lan Xiaoli; Zhang Yongxue; Qi Hongyan

    2012-01-01

    Gene therapy is a method that transfers foreign gene to target cells, so as to correct or compensate the disease which is caused by the gene defects and abnormalities. As a new technology, gene therapy has been used in many fields, such as cancer, cardiovascular and nervous system disease, and it brings some hope for patients with difficult and complicated disease. Avascular necrosis of femoral head is a refractory and common disease in clinical, but the traditional surgery therapy and conservative treatment both have many shortcomings,and the effect is unsatisfactory. As a new technology,gene therapy showed bright future in orthopedics ischemic disease, and its potential feasibility has been confirmed by many animal experiments. This article focuses on the research progress of gene therapy and early monitoring in the avascular necrosis of the femoral head. (authors)

  3. Regulation of radiation-induced apoptosis by early growth response-1 gene in solid tumors

    International Nuclear Information System (INIS)

    Ahmed, M.

    2003-01-01

    Ionizing radiation exposure is associated with activation of certain immediate-early genes that function as transcription factors. These include members of jun or fos and early growth response (EGR) gene families. In particular, the functional role of EGR-1 in radiation-induced signaling is pivotal since the promoter of EGR-1 contains radiation-inducible CArG DNA sequences. The Egr-1 gene belongs to a family of Egr genes that includes EGR-2, EGR-3, EGR-4, EGR-α and the tumor suppressor, Wilms' tumor gene product, WT1. The Egr-1 gene product, EGR-1, is a nuclear protein that contains three zinc fingers of the C 2 H 2 subtype. The EGR-1 GC-rich consensus target sequence, 5'-GCGT/GGGGCG-3' or 5'-TCCT/ACCTCCTCC-3', has been identified in the promoter regions of transcription factors, growth factors, receptors, cell cycle regulators and pro-apoptotic genes. The gene targets mediated by Egr-1 in response to ionizing radiation include TNF-α , p53, Rb and Bax, all these are effectors of apoptosis. Based on these targets, Egr-1 is a pivotal gene that initiates early signal transduction events in response to ionizing radiation leading to either growth arrest or cell death in tumor cells. There are two potential application of Egr-1 gene in therapy of cancer. First, the Egr-1 promoter contains information for appropriate spatial and temporal expression in-vivo that can be regulated by ionizing radiation to control transcription of genes that have pro-apoptotic and suicidal function. Secondly, EGR-1 protein can eliminate 'induced-radiation resistance' by inhibiting the functions of radiation-induced pro-survival genes (NFκB activity and bcl-2 expression) and activate pro-apoptotic genes (such as bax) to confer a significant radio-sensitizing effect. Together, the reported findings from my laboratory demonstrate clearly that EGR-1 is an early central gene that confers radiation sensitivity and its pro-apoptotic functions are synergized by abrogation of induced radiation

  4. Early Infantile Leigh-like Gene Defects Have a Poor Prognosis: Report and Review

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    Majid Alfadhel

    2017-10-01

    Full Text Available Solute carrier family 19 (thiamine transporter, member 3 ( SCL19A3 gene defect produces an autosomal recessive neurodegenerative disorder associated with different phenotypes and acronyms. One of the common presentations is early infantile lethal Leigh-like syndrome. We report a case of early infantile Leigh-like SLC19A3 gene defects of patients who died at 4 months of age with no response to a high dose of biotin and thiamine. In addition, we report a novel mutation that was not reported previously. Finally, we review the literature regarding early infantile Leigh-like SLC19A3 gene defects and compare the literature with our patient.

  5. Impact of viral E2-gene status on outcome after radiotherapy for patients with human papillomavirus 16-positive cancer of the uterine cervix

    International Nuclear Information System (INIS)

    Lindel, Katja; Villiers, Ethel-Michele de; Burri, Philipp; Studer, Ueli; Altermatt, Hans J.; Greiner, Richard H.; Gruber, Guenther

    2006-01-01

    Purpose: Integration of high-risk papillomavirus DNA has been considered an important step in oncogenic progression to cervical carcinoma. Disruption of the human papillomavirus (HPV) genome within the E2 gene is frequently a consequence. This study investigated the influence of episomal viral DNA on outcome in patients with advanced cervical cancer treated with primary radiotherapy. Methods and Materials: Paraffin-embedded biopsies of 82 women with locally advanced cervical cancer could be analyzed for HPV infection by multiplex polymerase chain reaction (PCR) by use of SPF1/2 primers. E2-gene intactness of HPV-16-positive samples was analyzed in 3 separate amplification reactions by use of the E2A, E2B, E2C primers. Statistical analyses (Kaplan-Meier method; log-rank test) were performed for overall survival (OS), disease-free survival (DFS), local progression-free survival (LPFS), and distant metastases-free survival (DMFS). Results: Sixty-one (75%) of 82 carcinomas were HPV positive, 44 of them for HPV-16 (72%). Seventeen of the 44 HPV-16-positive tumors (39%) had an intact E2 gene. Patients with a HPV-16-positive tumor and an intact E2 gene showed a trend for a better DFS (58% vs. 38%, p = 0.06) compared with those with a disrupted E2 gene. A nonsignificant difference occurred regarding OS (87% vs. 66%, p = 0.16) and DMFS (57% vs. 48%, p = 0.15). Conclusion: E2-gene status may be a promising new target, but more studies are required to elucidate the effect of the viral E2 gene on outcome after radiotherapy in HPV-positive tumors

  6. Early gene expression divergence between allopatric populations of the house mouse (Mus musculus domesticus).

    Science.gov (United States)

    Bryk, Jarosław; Somel, Mehmet; Lorenc, Anna; Teschke, Meike

    2013-03-01

    Divergence of gene expression is known to contribute to the differentiation and separation of populations and species, although the dynamics of this process in early stages of population divergence remains unclear. We analyzed gene expression differences in three organs (brain, liver, and testis) between two natural populations of Mus musculus domesticus that have been separated for at most 3000 years. We used two different microarray platforms to corroborate the results at a large scale and identified hundreds of genes with significant expression differences between the populations. We find that although the three tissues have similar number of differentially expressed genes, brain and liver have more tissue-specific genes than testis. Most genes show changes in a single tissue only, even when expressed in all tissues, supporting the notion that tissue-specific enhancers act as separable targets of evolution. In terms of functional categories, in brain and to a smaller extent in liver, we find transcription factors and their targets to be particularly variable between populations, similar to previous findings in primates. Testis, however, has a different set of differently expressed genes, both with respect to functional categories and overall correlation with the other tissues, the latter indicating that gene expression divergence of potential importance might be present in other datasets where no differences in fraction of differentially expressed genes were reported. Our results show that a significant amount of gene expression divergence quickly accumulates between allopatric populations.

  7. The BRCA1 and BRCA2 Genes in Early-Onset Breast Cancer Patients.

    Science.gov (United States)

    Saleem, Mohamed; Ghazali, Mohd Bazli; Wahab, Md Azlan Mohamed Abdul; Yusoff, Narazah Mohd; Mahsin, Hakimah; Seng, Ch'ng Ewe; Khalid, Imran Abdul; Rahman, Mohd Nor Gohar; Yahaya, Badrul Hisham

    2018-04-24

    Approximately 5-10% of breast cancers are attributable to genetic susceptibility. Mutations in the BRCA1 and BRCA2 genes are the best known genetic factors to date. The goal of this study was to determine the structure and distribution of haplotypes of the BRCA1 and BRCA2 genes in early-onset breast cancer patients. We enrolled 70 patients diagnosed with early-onset breast cancer. A total of 21 SNPs (11 on BRCA1 and 10 on BRCA2) and 1 dinucleotide deletion on BRCA1 were genotyped using nested allele-specific PCR methods. Linkage disequilibrium (LD) analysis was conducted, and haplotypes were deduced from the genotype data. Two tightly linked LD blocks were observed on each of the BRCA1 and BRCA2 genes. Variant-free haplotypes (TAT-AG for BRCA1 and ATA-AAT for BRCA2) were observed at a frequency of more than 50% on each gene along with variable frequencies of derived haplotypes. The variant 3'-subhaplotype CGC displayed strong LD with 5'-subhaplotypes GA, AA, and GG on BRCA1 gene. Haplotypes ATA-AGT, ATC-AAT, and ATA-AAC were the variant haplotypes frequent on BRCA2 gene. Although the clinical significance of these derived haplotypes has not yet been established, it is expected that some of these haplotypes, especially the less frequent subhaplotypes, eventually will be shown to be indicative of a predisposition to early-onset breast cancer.

  8. Genes as early responders regulate quorum-sensing and control bacterial cooperation in Pseudomonas aeruginosa.

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    Kelei Zhao

    Full Text Available Quorum-sensing (QS allows bacterial communication to coordinate the production of extracellular products essential for population fitness at higher cell densities. It has been generally accepted that a significant time duration is required to reach appropriate cell density to activate the relevant quiescent genes encoding these costly but beneficial public goods. Which regulatory genes are involved and how these genes control bacterial communication at the early phases are largely un-explored. By determining time-dependent expression of QS-related genes of the opportunistic pathogen Pseudomonas aerugionsa, we show that the induction of social cooperation could be critically influenced by environmental factors to optimize the density of population. In particular, small regulatory RNAs (RsmY and RsmZ serving as early responders, can promote the expression of dependent genes (e.g. lasR to boost the synthesis of intracellular enzymes and coordinate instant cooperative behavior in bacterial cells. These early responders, acting as a rheostat to finely modulate bacterial cooperation, which may be quickly activated under environment threats, but peter off when critical QS dependent genes are fully functional for cooperation. Our findings suggest that RsmY and RsmZ critically control the timing and levels of public goods production, which may have implications in sociomicrobiology and infection control.

  9. Pyrosequencing of Haliotis diversicolor transcriptomes: insights into early developmental molluscan gene expression.

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    Zi-Xia Huang

    Full Text Available BACKGROUND: The abalone Haliotis diversicolor is a good model for study of the settlement and metamorphosis, which are widespread marine ecological phenomena. However, information on the global gene backgrounds and gene expression profiles for the early development of abalones is lacking. METHODOLOGY/PRINCIPAL FINDINGS: In this study, eight non-normalized and multiplex barcode-labeled transcriptomes were sequenced using a 454 GS system to cover the early developmental stages of the abalone H. diversicolor. The assembly generated 35,415 unigenes, of which 7,566 were assigned GO terms. A global gene expression profile containing 636 scaffolds/contigs was constructed and was proven reliable using qPCR evaluation. It indicated that there may be existing dramatic phase transitions. Bioprocesses were proposed, including the 'lock system' in mature eggs, the collagen shells of the trochophore larvae and the development of chambered extracellular matrix (ECM structures within the earliest postlarvae. CONCLUSION: This study globally details the first 454 sequencing data for larval stages of H. diversicolor. A basic analysis of the larval transcriptomes and cluster of the gene expression profile indicates that each stage possesses a batch of specific genes that are indispensable during embryonic development, especially during the two-cell, trochophore and early postlarval stages. These data will provide a fundamental resource for future physiological works on abalones, revealing the mechanisms of settlement and metamorphosis at the molecular level.

  10. The pnk/pnl gene (ORF 86) of Autographa californica nucleopolyhedrovirus is a non-essential, immediate early gene.

    Science.gov (United States)

    Durantel, D; Croizier, L; Ayres, M D; Croizier, G; Possee, R D; López-Ferber, M

    1998-03-01

    Autographa californica nucleopolyhedrovirus (AcMNPV) ORF 86, located within the HindIII C fragment, potentially encodes a protein which shares sequence similarity with two T4 bacteriophage gene products, RNA ligase and polynucleotide kinase. This AcMNPV gene has been designated pnk/pnl but has yet to be assigned a function in virus replication. It has been classified as an immediate early virus gene, since the promoter was active in uninfected insect cells and mRNA transcripts were detectable from 4 to 48 h post-infection and in the presence of cycloheximide or aphidicolin in virus-infected cells. The extremities of the transcript have been mapped by primer extension and 3' RACE-PCR to positions -18 from the translational start codon and +15 downstream of the stop codon. The function of pnk/pnl was investigated by producing a recombinant virus (Acdel86lacZ) with the coding region replaced with that of lacZ. This virus replicated normally in Spodoptera frugiperda (Sf 21) cells, indicating that pnk/pnl is not essential for propagation in these cells. Virus protein production in Acdel86lacZ-infected Sf 21 cells also appeared to be unaffected, with normal synthesis of the IE-1, GP64, VP39 and polyhedrin proteins. Shut-down of host protein synthesis was not abolished in recombinant infection. When other baculovirus genomes were examined for the presence of pnk/pnl by restriction enzyme digestion and PCR, a deletion was found in AcMNPV 1.2, Galleria mellonella NPV (GmMNPV) and Bombyx mori NPV (BmNPV), suggesting that in many isolates this gene has either never been acquired or has been lost during genome evolution. This is one of the first baculovirus immediate early genes that appears to be nonessential for virus survival.

  11. Reverse-engineering of gene networks for regulating early blood development from single-cell measurements.

    Science.gov (United States)

    Wei, Jiangyong; Hu, Xiaohua; Zou, Xiufen; Tian, Tianhai

    2017-12-28

    Recent advances in omics technologies have raised great opportunities to study large-scale regulatory networks inside the cell. In addition, single-cell experiments have measured the gene and protein activities in a large number of cells under the same experimental conditions. However, a significant challenge in computational biology and bioinformatics is how to derive quantitative information from the single-cell observations and how to develop sophisticated mathematical models to describe the dynamic properties of regulatory networks using the derived quantitative information. This work designs an integrated approach to reverse-engineer gene networks for regulating early blood development based on singel-cell experimental observations. The wanderlust algorithm is initially used to develop the pseudo-trajectory for the activities of a number of genes. Since the gene expression data in the developed pseudo-trajectory show large fluctuations, we then use Gaussian process regression methods to smooth the gene express data in order to obtain pseudo-trajectories with much less fluctuations. The proposed integrated framework consists of both bioinformatics algorithms to reconstruct the regulatory network and mathematical models using differential equations to describe the dynamics of gene expression. The developed approach is applied to study the network regulating early blood cell development. A graphic model is constructed for a regulatory network with forty genes and a dynamic model using differential equations is developed for a network of nine genes. Numerical results suggests that the proposed model is able to match experimental data very well. We also examine the networks with more regulatory relations and numerical results show that more regulations may exist. We test the possibility of auto-regulation but numerical simulations do not support the positive auto-regulation. In addition, robustness is used as an importantly additional criterion to select candidate

  12. Transcriptome Analysis of Early Responsive Genes in Rice during Magnaporthe oryzae Infection

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    Yiming Wang

    2014-12-01

    Full Text Available Rice blast disease caused by Magnaporthe oryzae is one of the most serious diseases of cultivated rice (Oryza sativa L. in most rice-growing regions of the world. In order to investigate early response genes in rice, we utilized the transcriptome analysis approach using a 300 K tilling microarray to rice leaves infected with compatible and incompatible M. oryzae strains. Prior to the microarray experiment, total RNA was validated by measuring the differential expression of rice defense-related marker genes (chitinase 2, barwin, PBZ1, and PR-10 by RT-PCR, and phytoalexins (sakuranetin and momilactone A with HPLC. Microarray analysis revealed that 231 genes were up-regulated (>2 fold change, p < 0.05 in the incompatible interaction compared to the compatible one. Highly expressed genes were functionally characterized into metabolic processes and oxidation-reduction categories. The oxidative stress response was induced in both early and later infection stages. Biotic stress overview from MapMan analysis revealed that the phytohormone ethylene as well as signaling molecules jasmonic acid and salicylic acid is important for defense gene regulation. WRKY and Myb transcription factors were also involved in signal transduction processes. Additionally, receptor-like kinases were more likely associated with the defense response, and their expression patterns were validated by RT-PCR. Our results suggest that candidate genes, including receptor-like protein kinases, may play a key role in disease resistance against M. oryzae attack.

  13. Comparative gene expression analysis of Dtg, a novel target gene of Dpp signaling pathway in the early Drosophila melanogaster embryo.

    Science.gov (United States)

    Hodar, Christian; Zuñiga, Alejandro; Pulgar, Rodrigo; Travisany, Dante; Chacon, Carlos; Pino, Michael; Maass, Alejandro; Cambiazo, Verónica

    2014-02-10

    In the early Drosophila melanogaster embryo, Dpp, a secreted molecule that belongs to the TGF-β superfamily of growth factors, activates a set of downstream genes to subdivide the dorsal region into amnioserosa and dorsal epidermis. Here, we examined the expression pattern and transcriptional regulation of Dtg, a new target gene of Dpp signaling pathway that is required for proper amnioserosa differentiation. We showed that the expression of Dtg was controlled by Dpp and characterized a 524-bp enhancer that mediated expression in the dorsal midline, as well as, in the differentiated amnioserosa in transgenic reporter embryos. This enhancer contained a highly conserved region of 48-bp in which bioinformatic predictions and in vitro assays identified three Mad binding motifs. Mutational analysis revealed that these three motifs were necessary for proper expression of a reporter gene in transgenic embryos, suggesting that short and highly conserved genomic sequences may be indicative of functional regulatory regions in D. melanogaster genes. Dtg orthologs were not detected in basal lineages of Dipterans, which unlike D. melanogaster develop two extra-embryonic membranes, amnion and serosa, nevertheless Dtg orthologs were identified in the transcriptome of Musca domestica, in which dorsal ectoderm patterning leads to the formation of a single extra-embryonic membrane. These results suggest that Dtg was recruited as a new component of the network that controls dorsal ectoderm patterning in the lineage leading to higher Cyclorrhaphan flies, such as D. melanogaster and M. domestica. Copyright © 2013 Elsevier B.V. All rights reserved.

  14. Differential maturation of rhythmic clock gene expression during early development in medaka (Oryzias latipes).

    Science.gov (United States)

    Cuesta, Ines H; Lahiri, Kajori; Lopez-Olmeda, Jose Fernando; Loosli, Felix; Foulkes, Nicholas S; Vallone, Daniela

    2014-05-01

    One key challenge for the field of chronobiology is to identify how circadian clock function emerges during early embryonic development. Teleosts such as the zebrafish are ideal models for studying circadian clock ontogeny since the entire process of development occurs ex utero in an optically transparent chorion. Medaka (Oryzias latipes) represents another powerful fish model for exploring early clock function with, like the zebrafish, many tools available for detailed genetic analysis. However, to date there have been no reports documenting circadian clock gene expression during medaka development. Here we have characterized the expression of key clock genes in various developmental stages and in adult tissues of medaka. As previously reported for other fish, light dark cycles are required for the emergence of clock gene expression rhythms in this species. While rhythmic expression of per and cry genes is detected very early during development and seems to be light driven, rhythmic clock and bmal expression appears much later around hatching time. Furthermore, the maturation of clock function seems to correlate with the appearance of rhythmic expression of these positive elements of the clock feedback loop. By accelerating development through elevated temperatures or by artificially removing the chorion, we show an earlier onset of rhythmicity in clock and bmal expression. Thus, differential maturation of key elements of the medaka clock mechanism depends on the developmental stage and the presence of the chorion.

  15. Methylation in the promoter regions of WT1, NKX6-1 and DBC1 genes in cervical cancer tissues of Uygur women in Xinjiang

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    Dan Wu

    Full Text Available Abstract This study aimed to explore: 1 DNA methylation in the promoter regions of Wilms tumor gene 1 (WT1, NK6 transcription factor related locus 1 gene (NKX6-1 and Deleted in bladder cancer 1 (DBC1 gene in cervical cancer tissues of Uygur women in Xinjiang, and 2 the correlation of gene methylation with the infection of HPV16/18 viruses. We detected HPV16/18 infection in 43 normal cervical tissues, 30 cervical intraepithelial neoplasia lesions (CIN and 48 cervical cancer tissues with polymerase chain reaction (PCR method. Methylation in the promoter regions of the WT1, NKX6-1 and DBC1 genes in the above-mentioned tissues was measured by methylation-specific PCR (MSP and cloning sequencing. The expression level of these three genes was measured by real-time PCR (qPCR in 10 methylation-positive cervical cancer tissues and 10 methylation-negative normal cervical tissues. We found that the infection of HPV16 in normal cervical tissues, CIN and cervical cancer tissues was 14.0, 36.7 and 66.7%, respectively. The infection of HPV18 was 0, 6.7 and 10.4%, respectively. The methylation rates of WT1, NKX6-1 and DBC1 genes were 7.0, 11.6 and 23.3% in normal cervical tissues, 36.7, 46.7 and 30.0% in CIN tissues, and 89.6, 77.1 and 85.4% in cervical cancer tissues. Furthermore, WT1, NKX6-1 and DBC1 genes were hypermethylated in the high-grade squamous intraepithelial lesion (CIN2, CIN3 and in the cervical cancer tissues with infection of HPV16/18 (both P< 0.05. The expression of WT1, NKX6-1 and DBC1 was significantly lower in the methylation-positive cervical cancer tissues than in methylation-negative normal cervical tissues. Our findings indicated that methylation in the promoter regions of WT1, NKX6-1 and DBC1 is correlated with cervical cancer tumorigenesis in Uygur women. The infection of HPV16/18 might be correlated with methylation in these genes. Gene inactivation caused by methylation might be related to the incidence and development of cervical

  16. Early vertebrate chromosome duplications and the evolution of the neuropeptide Y receptor gene regions

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    Brenner Sydney

    2008-06-01

    Full Text Available Abstract Background One of the many gene families that expanded in early vertebrate evolution is the neuropeptide (NPY receptor family of G-protein coupled receptors. Earlier work by our lab suggested that several of the NPY receptor genes found in extant vertebrates resulted from two genome duplications before the origin of jawed vertebrates (gnathostomes and one additional genome duplication in the actinopterygian lineage, based on their location on chromosomes sharing several gene families. In this study we have investigated, in five vertebrate genomes, 45 gene families with members close to the NPY receptor genes in the compact genomes of the teleost fishes Tetraodon nigroviridis and Takifugu rubripes. These correspond to Homo sapiens chromosomes 4, 5, 8 and 10. Results Chromosome regions with conserved synteny were identified and confirmed by phylogenetic analyses in H. sapiens, M. musculus, D. rerio, T. rubripes and T. nigroviridis. 26 gene families, including the NPY receptor genes, (plus 3 described recently by other labs showed a tree topology consistent with duplications in early vertebrate evolution and in the actinopterygian lineage, thereby supporting expansion through block duplications. Eight gene families had complications that precluded analysis (such as short sequence length or variable number of repeated domains and another eight families did not support block duplications (because the paralogs in these families seem to have originated in another time window than the proposed genome duplication events. RT-PCR carried out with several tissues in T. rubripes revealed that all five NPY receptors were expressed in the brain and subtypes Y2, Y4 and Y8 were also expressed in peripheral organs. Conclusion We conclude that the phylogenetic analyses and chromosomal locations of these gene families support duplications of large blocks of genes or even entire chromosomes. Thus, these results are consistent with two early vertebrate

  17. Three neuropeptide Y receptor genes in the spiny dogfish, Squalus acanthias, support en bloc duplications in early vertebrate evolution.

    Science.gov (United States)

    Salaneck, Erik; Ardell, David H; Larson, Earl T; Larhammar, Dan

    2003-08-01

    It has been debated whether the increase in gene number during early vertebrate evolution was due to multiple independent gene duplications or synchronous duplications of many genes. We describe here the cloning of three neuropeptide Y (NPY) receptor genes belonging to the Y1 subfamily in the spiny dogfish, Squalus acanthias, a cartilaginous fish. The three genes are orthologs of the mammalian subtypes Y1, Y4, and Y6, which are located in paralogous gene regions on different chromosomes in mammals. Thus, these genes arose by duplications of a chromosome region before the radiation of gnathostomes (jawed vertebrates). Estimates of duplication times from linearized trees together with evidence from other gene families supports two rounds of chromosome duplications or tetraploidizations early in vertebrate evolution. The anatomical distribution of mRNA was determined by reverse-transcriptase PCR and was found to differ from mammals, suggesting differential functional diversification of the new gene copies during the radiation of the vertebrate classes.

  18. Differential effects of simple repeating DNA sequences on gene expression from the SV40 early promoter.

    Science.gov (United States)

    Amirhaeri, S; Wohlrab, F; Wells, R D

    1995-02-17

    The influence of simple repeat sequences, cloned into different positions relative to the SV40 early promoter/enhancer, on the transient expression of the chloramphenicol acetyltransferase (CAT) gene was investigated. Insertion of (G)29.(C)29 in either orientation into the 5'-untranslated region of the CAT gene reduced expression in CV-1 cells 50-100 fold when compared with controls with random sequence inserts. Analysis of CAT-specific mRNA levels demonstrated that the effect was due to a reduction of CAT mRNA production rather than to posttranscriptional events. In contrast, insertion of the same insert in either orientation upstream of the promoter-enhancer or downstream of the gene stimulated gene expression 2-3-fold. These effects could be reversed by cotransfection of a competitor plasmid carrying (G)25.(C)25 sequences. The results suggest that a G.C-binding transcription factor modulates gene expression in this system and that promoter strength can be regulated by providing protein-binding sites in trans. Although constructs containing longer tracts of alternating (C-G), (T-G), or (A-T) sequences inhibited CAT expression when inserted in the 5'-untranslated region of the CAT gene, the amount of CAT mRNA was unaffected. Hence, these inhibitions must be due to posttranscriptional events, presumably at the level of translation. These effects of microsatellite sequences on gene expression are discussed with respect to recent data on related simple repeat sequences which cause several human genetic diseases.

  19. Early embryonic failure: Expression and imprinted status of candidate genes on human chromosome 21

    Energy Technology Data Exchange (ETDEWEB)

    Sherman, L.S.; Bennett, P.R.; Moore, G.E. [Queen Charlotte`s and Chelsea Hospital, London (United Kingdom)

    1994-09-01

    Two cases of maternal uniparental (hetero)disomy for human chromosome 21 (mUPD21) have been identified in a systematic search for UPD in 23 cases of early embryonic failure (EEF). Bi-parental origin of the other chromosome pairs was confirmed using specific VNTR probes or dinucleotide repeat analysis. Both maternally and paternally derived isochromosomes 21q have previously been identified in two individuals with normal phenotypes. Full UPD21 has a different mechanism of origin than uniparental isochromosome 21q and its effect on imprinted genes and phenotypic outcome will therefore not necessarily be the same. EEF associated with mUPD21 suggests that developmentally important genes on HSA 21 may be imprinted such that they are only expressed from either the maternally or paternally derived alleles. We have searched for monoallelic expression of candidate genes on HSA 21 in human pregnancy (CBS, IFNAR, COL6A1) using intragenic DNA polymorphisms. These genes were chosen either because their murine homologues lie in imprinted regions or because they are potentially important in embryogenesis. Once imprinted candidate genes have been identified, their methylation status and expression in normal, early embryonic failure and uniparental disomy 21 pregnancies will be studied. At the same time, a larger number of cases of EEF are being examined to further investigate the incidence of UPD21 in this group.

  20. Prevalence of anal infection due to high-risk human papillomavirus and analysis of E2 gene integrity among women with cervical abnormalities.

    Science.gov (United States)

    Gonzalez-Losa, María Del Refugio; Puerto-Solís, Marylin; Ayora-Talavera, Guadalupe; Gómez-Carvallo, Jesús; Euán-López, Alejandra; Cisneros-Cutz, José I; Rosado-López, Ariel; Echeverría Salazar, Jesúa; Conde-Ferráez, Laura

    2018-04-01

    High-risk human papillomaviruses (HR-HPV) infection has been associated with 90% of anal cancer cases. Women with abnormal cytology are a high-risk group to develop anal neoplasia. The aim of this study is to describe the prevalence and epidemiology of HR-HPV 16, 18, 45, and 58 anal infections in women with cervical abnormalities, as well as to assess E2 gene integrity. A cross-sectional study was performed on 311 cervical and 311 anal samples from patients with abnormal cytology in two colposcopy clinics in Yucatan, Mexico. A specific PCR for oncogenes was performed in order to identify HVP 16, 18, 45 and 58. Real time PCR was used to amplify the whole HPV 16, 18, and 58 E2 gene to verify its integrity in anal samples. High risk HPV 16, 18, 58, and/or 45 were found in 41.47% (129/311) of cervical samples, and in 30.8% (96/331) of anal samples, with 18% (57/311) of the patients being positive in both samples. The same genotypes in both anatomical sites were observed in 11.25% (35/311). The E2 gene was disrupted in 82% of all tested samples. The frequency of genome disruption viral integration in anal samples by genotype was: HPV 58 (97.2%); HPV 16 (72.4%), and HPV 18 (0%). Women with cervical disease have HR-HPV anal infections, and most of them have the E2 gene disrupted, which represents a risk to develop anal cancer. Copyright © 2016 Elsevier España, S.L.U. and Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

  1. Transcriptomic profiling of bovine IVF embryos revealed candidate genes and pathways involved in early embryonic development

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    Yandell Brian S

    2010-01-01

    Full Text Available Abstract Background Early embryonic loss is a large contributor to infertility in cattle. Although genetic factors are known to affect early embryonic development, the discovery of such factors has been a serious challenge. The objective of this study was to identify genes differentially expressed between blastocysts and degenerative embryos at early stages of development. Results Using microarrays, genome-wide RNA expression was profiled and compared for in vitro fertilization (IVF - derived blastocysts and embryos undergoing degenerative development up to the same time point. Surprisingly similar transcriptomic profiles were found in degenerative embryos and blastocysts. Nonetheless, we identified 67 transcripts that significantly differed between these two groups of embryos at a 15% false discovery rate, including 33 transcripts showing at least a two-fold difference. Several signaling and metabolic pathways were found to be associated with the developmental status of embryos, among which were previously known important steroid biosynthesis and cell communication pathways in early embryonic development. Conclusions This study presents the first direct and comprehensive comparison of transcriptomes between IVF blastocysts and degenerative embryos, providing important information for potential genes and pathways associated with early embryonic development.

  2. A robust prognostic gene expression signature for early stage lung adenocarcinoma

    DEFF Research Database (Denmark)

    Krzystanek, Marcin; Moldvay, Judit; Szüts, David

    2016-01-01

    Stage I lung adenocarcinoma is usually not treated with adjuvant chemotherapy; however, around half of these patients do not survive 5 years. Therefore, a reliable prognostic biomarker for early stage patients would be critical to identify those most likely to benefit from early additional treatm...... not given adjuvant therapy. Seven genes consistently obtained statistical significance in Cox regression for overall survival. The combined signature has a weighted mean hazard ratio of 3.2 in all cohorts and 3.0 (C.I. 1.3-7.4, p ...

  3. Intrauterine growth restriction and placental gene expression in severe preeclampsia, comparing early-onset and late-onset forms.

    Science.gov (United States)

    Nevalainen, Jaana; Skarp, Sini; Savolainen, Eeva-Riitta; Ryynänen, Markku; Järvenpää, Jouko

    2017-10-26

    To evaluate placental gene expression in severe early- or late-onset preeclampsia with intrauterine growth restriction compared to controls. Chorionic villus sampling was conducted after cesarean section from the placentas of five women with early- or late-onset severe preeclampsia and five controls for each preeclampsia group. Microarray analysis was performed to identify gene expression differences between the groups. Pathway analysis showed over-representation of gene ontology (GO) biological process terms related to inflammatory and immune response pathways, platelet development, vascular development, female pregnancy and reproduction in early-onset preeclampsia. Pathways related to immunity, complement and coagulation cascade were overrepresented in the hypergeometric test for the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. Ten genes (ABI3BP, C7, HLA-G, IL2RB, KRBOX1, LRRC15, METTL7B, MPP5, RFLNB and SLC20A) had a ≥±1 fold expression difference in severe early-onset preeclampsia group compared to early controls. There were 362 genes that had a ≥±1 fold expression difference in severe early-onset preeclampsia group compared to late-onset preeclampsia group including ABI3BP, C7, HLA-G and IL2RB. There are significant differences in placental gene expression between severe early- and late-onset preeclampsia when both are associated with intrauterine growth restriction. ABI3BP, C7, HLA-G and IL2RB might contribute to the development of early form of severe preeclampsia.

  4. Global gene expression analysis of early response to chemotherapy treatment in ovarian cancer spheroids

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    Tetu Bernard

    2008-02-01

    Full Text Available Abstract Background Chemotherapy (CT resistance in ovarian cancer (OC is broad and encompasses diverse unrelated drugs, suggesting more than one mechanism of resistance. To better understand the molecular mechanisms controlling the immediate response of OC cells to CT exposure, we have performed gene expression profiling in spheroid cultures derived from six OC cell lines (OVCAR3, SKOV3, TOV-112, TOV-21, OV-90 and TOV-155, following treatment with 10,0 μM cisplatin, 2,5 μM paclitaxel or 5,0 μM topotecan for 72 hours. Results Exposure of OC spheroids to these CT drugs resulted in differential expression of genes associated with cell growth and proliferation, cellular assembly and organization, cell death, cell cycle control and cell signaling. Genes, functionally involved in DNA repair, DNA replication and cell cycle arrest were mostly overexpressed, while genes implicated in metabolism (especially lipid metabolism, signal transduction, immune and inflammatory response, transport, transcription regulation and protein biosynthesis, were commonly suppressed following all treatments. Cisplatin and topotecan treatments triggered similar alterations in gene and pathway expression patterns, while paclitaxel action was mainly associated with induction of genes and pathways linked to cellular assembly and organization (including numerous tubulin genes, cell death and protein synthesis. The microarray data were further confirmed by pathway and network analyses. Conclusion Most alterations in gene expression were directly related to mechanisms of the cytotoxics actions in OC spheroids. However, the induction of genes linked to mechanisms of DNA replication and repair in cisplatin- and topotecan-treated OC spheroids could be associated with immediate adaptive response to treatment. Similarly, overexpression of different tubulin genes upon exposure to paclitaxel could represent an early compensatory effect to this drug action. Finally, multicellular

  5. Copper induces expression and methylation changes of early development genes in Crassostrea gigas embryos.

    Science.gov (United States)

    Sussarellu, Rossana; Lebreton, Morgane; Rouxel, Julien; Akcha, Farida; Rivière, Guillaume

    2018-03-01

    Copper contamination is widespread along coastal areas and exerts adverse effects on marine organisms such as mollusks. In the Pacific oyster, copper induces severe developmental abnormalities during early life stages; however, the underlying molecular mechanisms are largely unknown. This study aims to better understand whether the embryotoxic effects of copper in Crassostrea gigas could be mediated by alterations in gene expression, and the putative role of DNA methylation, which is known to contribute to gene regulation in early embryo development. For that purpose, oyster embryos were exposed to 4 nominal copper concentrations (0.1, 1, 10 and 20 μg L -1 Cu 2+ ) during early development assays. Embryotoxicity was monitored through the oyster embryo-larval bioassay at the D-larva stage 24 h post fertilization (hpf) and genotoxicity at gastrulation 7 hpf. In parallel, the relative expression of 15 genes encoding putative homeotic, biomineralization and DNA methylation proteins was measured at three developmental stages (3 hpf morula stage, 7 hpf gastrula stage, 24 hpf D-larvae stage) using RT-qPCR. Global DNA content in methylcytosine and hydroxymethylcytosine were measured by HPLC and gene-specific DNA methylation levels were monitored using MeDIP-qPCR. A significant increase in larval abnormalities was observed from copper concentrations of 10 μg L -1 , while significant genotoxic effects were detected at 1 μg L -1 and above. All the selected genes presented a stage-dependent expression pattern, which was impaired for some homeobox and DNA methylation genes (Notochord, HOXA1, HOX2, Lox5, DNMT3b and CXXC-1) after copper exposure. While global DNA methylation (5-methylcytosine) at gastrula stage didn't show significant changes between experimental conditions, 5-hydroxymethylcytosine, its degradation product, decreased upon copper treatment. The DNA methylation of exons and the transcript levels were correlated in control samples for HOXA1 but such

  6. Lethal influenza virus infection in macaques is associated with early dysregulation of inflammatory related genes.

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    Cristian Cillóniz

    2009-10-01

    Full Text Available The enormous toll on human life during the 1918-1919 Spanish influenza pandemic is a constant reminder of the potential lethality of influenza viruses. With the declaration by the World Health Organization of a new H1N1 influenza virus pandemic, and with continued human cases of highly pathogenic H5N1 avian influenza virus infection, a better understanding of the host response to highly pathogenic influenza viruses is essential. To this end, we compared pathology and global gene expression profiles in bronchial tissue from macaques infected with either the reconstructed 1918 pandemic virus or the highly pathogenic avian H5N1 virus A/Vietnam/1203/04. Severe pathology was observed in respiratory tissues from 1918 virus-infected animals as early as 12 hours after infection, and pathology steadily increased at later time points. Although tissues from animals infected with A/Vietnam/1203/04 also showed clear signs of pathology early on, less pathology was observed at later time points, and there was evidence of tissue repair. Global transcriptional profiles revealed that specific groups of genes associated with inflammation and cell death were up-regulated in bronchial tissues from animals infected with the 1918 virus but down-regulated in animals infected with A/Vietnam/1203/04. Importantly, the 1918 virus up-regulated key components of the inflammasome, NLRP3 and IL-1beta, whereas these genes were down-regulated by A/Vietnam/1203/04 early after infection. TUNEL assays revealed that both viruses elicited an apoptotic response in lungs and bronchi, although the response occurred earlier during 1918 virus infection. Our findings suggest that the severity of disease in 1918 virus-infected macaques is a consequence of the early up-regulation of cell death and inflammatory related genes, in which additive or synergistic effects likely dictate the severity of tissue damage.

  7. Induction of the early-late Ddc gene during Drosophila metamorphosis by the ecdysone receptor.

    Science.gov (United States)

    Chen, Li; Reece, Christian; O'Keefe, Sandra L; Hawryluk, Gregory W L; Engstrom, Monica M; Hodgetts, Ross B

    2002-06-01

    During Drosophila metamorphosis, the 'early-late' genes constitute a unique class regulated by the steroid hormone 20-hydroxyecdysone. Their induction is comprised of both a primary and a secondary response to ecdysone. Previous work has suggested that the epidermal expression of the dopa decarboxylase gene (Ddc) is likely that of a typical early-late gene. Accumulation of the Ddc transcript is rapidly initiated in the absence of protein synthesis, which implies that the ecdysone receptor plays a direct role in induction. However, full Ddc expression requires the participation of one of the transcription factors encoded by the Broad-Complex. In this paper, we characterize an ecdysone response element (EcRE) that contributes to the primary response. Using gel mobility shift assays and transgenic assays, we identified a single functional EcRE, located at position -97 to -83 bp relative to the transcription initiation site. This is the first report of an EcRE associated with an early-late gene in Drosophila. Competition experiments indicated that the affinity of the Ddc EcRE for the ecdysone receptor complex was at least four-fold less than that of the canonical EcRE of the hsp27 gene. Using in vitro mutagenesis, we determined that the reduced affinity of the EcRE resided at two positions where the nucleotides differed from those found in the canonical sequence. The ecdysone receptor, acting through this EcRE, releases Ddc from a silencing mechanism, whose cis-acting domain we have mapped to the 5'-upstream region between -2067 and -1427 bp. Deletion of this repressive element resulted in precocious expression of Ddc in both epidermis and imaginal discs. Thus, epidermal Ddc induction at pupariation is under the control of an extended genomic region that contains both positive and negative regulatory elements. Copyright 2002 Elsevier Science Ireland Ltd.

  8. Expression of putative immune response genes during early ontogeny in the coral Acropora millepora.

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    Eneour Puill-Stephan

    Full Text Available Corals, like many other marine invertebrates, lack a mature allorecognition system in early life history stages. Indeed, in early ontogeny, when corals acquire and establish associations with various surface microbiota and dinoflagellate endosymbionts, they do not efficiently distinguish between closely and distantly related individuals from the same population. However, very little is known about the molecular components that underpin allorecognition and immunity responses or how they change through early ontogeny in corals.Patterns in the expression of four putative immune response genes (apextrin, complement C3, and two CELIII type lectin genes were examined in juvenile colonies of Acropora millepora throughout a six-month post-settlement period using quantitative real-time PCR (qPCR. Expression of a CELIII type lectin gene peaked in the fourth month for most of the coral juveniles sampled and was significantly higher at this time than at any other sampling time during the six months following settlement. The timing of this increase in expression levels of putative immune response genes may be linked to allorecognition maturation which occurs around this time in A. millepora. Alternatively, the increase may represent a response to immune challenges, such as would be involved in the recognition of symbionts (such as Symbiodinium spp. or bacteria during winnowing processes as symbioses are fine-tuned.Our data, although preliminary, are consistent with the hypothesis that lectins may play an important role in the maturation of allorecognition responses in corals. The co-expression of lectins with apextrin during development of coral juveniles also raises the possibility that these proteins, which are components of innate immunity in other invertebrates, may influence the innate immune systems of corals through a common pathway or system. However, further studies investigating the expression of these genes in alloimmune-challenged corals are

  9. Expression of putative immune response genes during early ontogeny in the coral Acropora millepora.

    Science.gov (United States)

    Puill-Stephan, Eneour; Seneca, François O; Miller, David J; van Oppen, Madeleine J H; Willis, Bette L

    2012-01-01

    Corals, like many other marine invertebrates, lack a mature allorecognition system in early life history stages. Indeed, in early ontogeny, when corals acquire and establish associations with various surface microbiota and dinoflagellate endosymbionts, they do not efficiently distinguish between closely and distantly related individuals from the same population. However, very little is known about the molecular components that underpin allorecognition and immunity responses or how they change through early ontogeny in corals. Patterns in the expression of four putative immune response genes (apextrin, complement C3, and two CELIII type lectin genes) were examined in juvenile colonies of Acropora millepora throughout a six-month post-settlement period using quantitative real-time PCR (qPCR). Expression of a CELIII type lectin gene peaked in the fourth month for most of the coral juveniles sampled and was significantly higher at this time than at any other sampling time during the six months following settlement. The timing of this increase in expression levels of putative immune response genes may be linked to allorecognition maturation which occurs around this time in A. millepora. Alternatively, the increase may represent a response to immune challenges, such as would be involved in the recognition of symbionts (such as Symbiodinium spp. or bacteria) during winnowing processes as symbioses are fine-tuned. Our data, although preliminary, are consistent with the hypothesis that lectins may play an important role in the maturation of allorecognition responses in corals. The co-expression of lectins with apextrin during development of coral juveniles also raises the possibility that these proteins, which are components of innate immunity in other invertebrates, may influence the innate immune systems of corals through a common pathway or system. However, further studies investigating the expression of these genes in alloimmune-challenged corals are needed to further

  10. Global gene expression profiling of individual human oocytes and embryos demonstrates heterogeneity in early development.

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    Lisa Shaw

    Full Text Available Early development in humans is characterised by low and variable embryonic viability, reflected in low fecundity and high rates of miscarriage, relative to other mammals. Data from assisted reproduction programmes provides additional evidence that this is largely mediated at the level of embryonic competence and is highly heterogeneous among embryos. Understanding the basis of this heterogeneity has important implications in a number of areas including: the regulation of early human development, disorders of pregnancy, assisted reproduction programmes, the long term health of children which may be programmed in early development, and the molecular basis of pluripotency in human stem cell populations. We have therefore investigated global gene expression profiles using polyAPCR amplification and microarray technology applied to individual human oocytes and 4-cell and blastocyst stage embryos. In order to explore the basis of any variability in detail, each developmental stage is replicated in triplicate. Our data show that although transcript profiles are highly stage-specific, within each stage they are relatively variable. We describe expression of a number of gene families and pathways including apoptosis, cell cycle and amino acid metabolism, which are variably expressed and may be reflective of embryonic developmental competence. Overall, our data suggest that heterogeneity in human embryo developmental competence is reflected in global transcript profiles, and that the vast majority of existing human embryo gene expression data based on pooled oocytes and embryos need to be reinterpreted.

  11. EBV tegument protein BNRF1 disrupts DAXX-ATRX to activate viral early gene transcription.

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    Kevin Tsai

    2011-11-01

    Full Text Available Productive infection by herpesviruses involve the disabling of host-cell intrinsic defenses by viral encoded tegument proteins. Epstein-Barr Virus (EBV typically establishes a non-productive, latent infection and it remains unclear how it confronts the host-cell intrinsic defenses that restrict viral gene expression. Here, we show that the EBV major tegument protein BNRF1 targets host-cell intrinsic defense proteins and promotes viral early gene activation. Specifically, we demonstrate that BNRF1 interacts with the host nuclear protein Daxx at PML nuclear bodies (PML-NBs and disrupts the formation of the Daxx-ATRX chromatin remodeling complex. We mapped the Daxx interaction domain on BNRF1, and show that this domain is important for supporting EBV primary infection. Through reverse transcription PCR and infection assays, we show that BNRF1 supports viral gene expression upon early infection, and that this function is dependent on the Daxx-interaction domain. Lastly, we show that knockdown of Daxx and ATRX induces reactivation of EBV from latently infected lymphoblastoid cell lines (LCLs, suggesting that Daxx and ATRX play a role in the regulation of viral chromatin. Taken together, our data demonstrate an important role of BNRF1 in supporting EBV early infection by interacting with Daxx and ATRX; and suggest that tegument disruption of PML-NB-associated antiviral resistances is a universal requirement for herpesvirus infection in the nucleus.

  12. EBV Tegument Protein BNRF1 Disrupts DAXX-ATRX to Activate Viral Early Gene Transcription

    Science.gov (United States)

    Tsai, Kevin; Thikmyanova, Nadezhda; Wojcechowskyj, Jason A.; Delecluse, Henri-Jacques; Lieberman, Paul M.

    2011-01-01

    Productive infection by herpesviruses involve the disabling of host-cell intrinsic defenses by viral encoded tegument proteins. Epstein-Barr Virus (EBV) typically establishes a non-productive, latent infection and it remains unclear how it confronts the host-cell intrinsic defenses that restrict viral gene expression. Here, we show that the EBV major tegument protein BNRF1 targets host-cell intrinsic defense proteins and promotes viral early gene activation. Specifically, we demonstrate that BNRF1 interacts with the host nuclear protein Daxx at PML nuclear bodies (PML-NBs) and disrupts the formation of the Daxx-ATRX chromatin remodeling complex. We mapped the Daxx interaction domain on BNRF1, and show that this domain is important for supporting EBV primary infection. Through reverse transcription PCR and infection assays, we show that BNRF1 supports viral gene expression upon early infection, and that this function is dependent on the Daxx-interaction domain. Lastly, we show that knockdown of Daxx and ATRX induces reactivation of EBV from latently infected lymphoblastoid cell lines (LCLs), suggesting that Daxx and ATRX play a role in the regulation of viral chromatin. Taken together, our data demonstrate an important role of BNRF1 in supporting EBV early infection by interacting with Daxx and ATRX; and suggest that tegument disruption of PML-NB-associated antiviral resistances is a universal requirement for herpesvirus infection in the nucleus. PMID:22102817

  13. Characterization of Betula platyphylla gene transcripts associated with early development of male inflorescence.

    Science.gov (United States)

    Xing, Lei; Liu, Xue-Mei

    2012-02-01

    Birch (Betula platyphylla), an eminent tree species in Northeast and Inner Mongolia of China, has been widely used in architecture, furniture, and paper making in recent years. In order to retrieve genes involved in early development of B. platyphylla male inflorescence, RNA populations extracted from early and late developmental stage were analyzed by cDNA-Amplified Fragment Length Polymorphism (cDNA-AFLP) technique. Following amplification of 256 pairs of primer combinations, ~7000 fragments were generated, of which 350 transcripts expressing more in early stage than late. Of 350 specific transcripts, 198 clear and reproducible electrophoresis bands were retrieved and sequenced successfully, 74 of them (37%) showing significant homologies to known genes after GO annotation. Majority of the predicted gene products were involved in metabolism (24.56%), cellular process (27.19%), response to stimulus (11.4%) and cell growth (8.7%). Transcripts ME56, ME108, ME206 and ME310, representing metabolism, cellular process, response to stimulus and cell growth, respectively, were selected for further study to validate cDNA-AFLP expression patterns via RT-PCR and qRT-PCR analysis. RT-PCR and qRT-PCR expression pattern results were consistent with cDNA-AFLP analysis results.

  14. Salicylate-induced changes in immediate-early genes in the hippocampal CA1 area.

    Science.gov (United States)

    Wu, Hao; Xu, Feng-Lei; Yin, Yong; Da, Peng; You, Xiao-Dong; Xu, Hui-Min; Tang, Yan

    2015-08-01

    Studies have suggested that salicylate affects neuronal function via interactions with specific membrane channels/receptors. However, the effect of salicylate on activity and synaptic morphology of the hippocampal Cornu Ammonis (CA) 1 area remains to be elucidated. The activation of immediate-early genes (IEGs) was reported to correlate with neuronal activity, in particular activity-regulated cytoskeleton-associated protein and early growth response gene 1. The aim of the present study was to evaluate the expression of these IEGs, as well that of N-methyl D-aspartate (NMDA) receptor subunit 2B in rats following acute and chronic salicylate treatment. Protein and messenger RNA levels of all three genes were increased in rats following chronic administration of salicylate (300 mg/kg for 10 days), returning to baseline levels 14 days post-cessation of treatment. The transient upregulation of gene expression following treatment was accompanied by ultrastructural alterations in hippocampal CA1 area synapses. An increase in synaptic interface curvature was observed as well as an increased number of presynaptic vesicles; in addition, postsynaptic densities thickened and lengthened. In conclusion, the results of the present study indicated that chronic exposure to salicylate may lead to structural alteration of hippocampal CA1 neurons, and it was suggested that this process occurs through induced expression of IEGs via NMDA receptor activation.

  15. Early experiences mediate distinct adult gene expression and reproductive programs in Caenorhabditis elegans

    Science.gov (United States)

    Ow, Maria C.; Nichitean, Alexandra M.; Dorus, Steve; Hall, Sarah E.

    2018-01-01

    Environmental stress during early development in animals can have profound effects on adult phenotypes via programmed changes in gene expression. Using the nematode C. elegans, we demonstrated previously that adults retain a cellular memory of their developmental experience that is manifested by differences in gene expression and life history traits; however, the sophistication of this system in response to different environmental stresses, and how it dictates phenotypic plasticity in adults that contribute to increased fitness in response to distinct environmental challenges, was unknown. Using transcriptional profiling, we show here that C. elegans adults indeed retain distinct cellular memories of different environmental conditions. We identified approximately 500 genes in adults that entered dauer due to starvation that exhibit significant opposite (“seesaw”) transcriptional phenotypes compared to adults that entered dauer due to crowding, and are distinct from animals that bypassed dauer. Moreover, we show that two-thirds of the genes in the genome experience a 2-fold or greater seesaw trend in gene expression, and based upon the direction of change, are enriched in large, tightly linked regions on different chromosomes. Importantly, these transcriptional programs correspond to significant changes in brood size depending on the experienced stress. In addition, we demonstrate that while the observed seesaw gene expression changes occur in both somatic and germline tissue, only starvation-induced changes require a functional GLP-4 protein necessary for germline development, and both programs require the Argonaute CSR-1. Thus, our results suggest that signaling between the soma and the germ line can generate phenotypic plasticity as a result of early environmental experience, and likely contribute to increased fitness in adverse conditions and the evolution of the C. elegans genome. PMID:29447162

  16. Early experiences mediate distinct adult gene expression and reproductive programs in Caenorhabditis elegans.

    Directory of Open Access Journals (Sweden)

    Maria C Ow

    2018-02-01

    Full Text Available Environmental stress during early development in animals can have profound effects on adult phenotypes via programmed changes in gene expression. Using the nematode C. elegans, we demonstrated previously that adults retain a cellular memory of their developmental experience that is manifested by differences in gene expression and life history traits; however, the sophistication of this system in response to different environmental stresses, and how it dictates phenotypic plasticity in adults that contribute to increased fitness in response to distinct environmental challenges, was unknown. Using transcriptional profiling, we show here that C. elegans adults indeed retain distinct cellular memories of different environmental conditions. We identified approximately 500 genes in adults that entered dauer due to starvation that exhibit significant opposite ("seesaw" transcriptional phenotypes compared to adults that entered dauer due to crowding, and are distinct from animals that bypassed dauer. Moreover, we show that two-thirds of the genes in the genome experience a 2-fold or greater seesaw trend in gene expression, and based upon the direction of change, are enriched in large, tightly linked regions on different chromosomes. Importantly, these transcriptional programs correspond to significant changes in brood size depending on the experienced stress. In addition, we demonstrate that while the observed seesaw gene expression changes occur in both somatic and germline tissue, only starvation-induced changes require a functional GLP-4 protein necessary for germline development, and both programs require the Argonaute CSR-1. Thus, our results suggest that signaling between the soma and the germ line can generate phenotypic plasticity as a result of early environmental experience, and likely contribute to increased fitness in adverse conditions and the evolution of the C. elegans genome.

  17. EGR3 Immediate Early Gene and the Brain-Derived Neurotrophic Factor in Bipolar Disorder

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    Bianca Pfaffenseller

    2018-02-01

    Full Text Available Bipolar disorder (BD is a severe psychiatric illness with a consistent genetic influence, involving complex interactions between numerous genes and environmental factors. Immediate early genes (IEGs are activated in the brain in response to environmental stimuli, such as stress. The potential to translate environmental stimuli into long-term changes in brain has led to increased interest in a potential role for these genes influencing risk for psychiatric disorders. Our recent finding using network-based approach has shown that the regulatory unit of early growth response gene 3 (EGR3 of IEGs family was robustly repressed in postmortem prefrontal cortex of BD patients. As a central transcription factor, EGR3 regulates an array of target genes that mediate critical neurobiological processes such as synaptic plasticity, memory and cognition. Considering that EGR3 expression is induced by brain-derived neurotrophic factor (BDNF that has been consistently related to BD pathophysiology, we suggest a link between BDNF and EGR3 and their potential role in BD. A growing body of data from our group and others has shown that peripheral BDNF levels are reduced during mood episodes and also with illness progression. In this same vein, BDNF has been proposed as an important growth factor in the impaired cellular resilience related to BD. Taken together with the fact that EGR3 regulates the expression of the neurotrophin receptor p75NTR and may also indirectly induce BDNF expression, here we propose a feed-forward gene regulatory network involving EGR3 and BDNF and its potential role in BD.

  18. Social defeat during adolescence and adulthood differentially induce BDNF-regulated immediate early genes

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    Caroline M. Coppens

    2011-11-01

    Full Text Available Stressful life events generally enhance the vulnerability for the development of human psychopathologies such as anxiety disorders and depression. The incidence rates of adult mental disorders steeply rises during adolescence in parallel with a structural and functional reorganization of the neural circuitry underlying stress reactivity. However, the mechanisms underlying susceptibility to stress and manifestation of mental disorders during adolescence are little understood. We hypothesized that heightened sensitivity to stress during adolescence reflects age-dependent differences in the expression of activity-dependent genes involved in synaptic plasticity. Therefore, we compared the effect of social stress during adolescence with social stress in adulthood on the expression of a panel of genes linked to induction of long-term potentiation (LTP and brain-derived neurotrophic factor (BDNF signaling. We show that social defeat during adolescence and adulthood differentially regulates expression of the immediate early genes BDNF, Arc, Carp, and Tieg1, as measured by qPCR in tissue lysates from prefrontal cortex, nucleus accumbens, and hippocampus. In the hippocampus, mRNA levels for all four genes were robustly elevated following social defeat in adolescence, whereas none were induced by defeat in adulthood. The relationship to coping style was also examined using adult reactive and proactive coping rats. Gene expression levels of reactive and proactive animals were similar in the prefrontal cortex and hippocampus. However, a trend toward a differential expression of BDNF and Arc mRNA in the nucleus accumbens was detected. BDNF mRNA was increased in the nucleus accumbens of proactive defeated animals, whereas the expression level in reactive defeated animals was comparable to control animals. The results demonstrate striking differences in immediate early gene expression in response to social defeat in adolescent and adult rats.

  19. Early Developmental and Evolutionary Origins of Gene Body DNA Methylation Patterns in Mammalian Placentas.

    Directory of Open Access Journals (Sweden)

    Diane I Schroeder

    2015-08-01

    Full Text Available Over the last 20-80 million years the mammalian placenta has taken on a variety of morphologies through both divergent and convergent evolution. Recently we have shown that the human placenta genome has a unique epigenetic pattern of large partially methylated domains (PMDs and highly methylated domains (HMDs with gene body DNA methylation positively correlating with level of gene expression. In order to determine the evolutionary conservation of DNA methylation patterns and transcriptional regulatory programs in the placenta, we performed a genome-wide methylome (MethylC-seq analysis of human, rhesus macaque, squirrel monkey, mouse, dog, horse, and cow placentas as well as opossum extraembryonic membrane. We found that, similar to human placenta, mammalian placentas and opossum extraembryonic membrane have globally lower levels of methylation compared to somatic tissues. Higher relative gene body methylation was the conserved feature across all mammalian placentas, despite differences in PMD/HMDs and absolute methylation levels. Specifically, higher methylation over the bodies of genes involved in mitosis, vesicle-mediated transport, protein phosphorylation, and chromatin modification was observed compared with the rest of the genome. As in human placenta, higher methylation is associated with higher gene expression and is predictive of genic location across species. Analysis of DNA methylation in oocytes and preimplantation embryos shows a conserved pattern of gene body methylation similar to the placenta. Intriguingly, mouse and cow oocytes and mouse early embryos have PMD/HMDs but their placentas do not, suggesting that PMD/HMDs are a feature of early preimplantation methylation patterns that become lost during placental development in some species and following implantation of the embryo.

  20. Microarray gene expression during early healing of GBR-treated calvarial critical size defects.

    Science.gov (United States)

    Al-Kattan, R; Retzepi, M; Calciolari, E; Donos, N

    2017-10-01

    To investigate the gene expression and molecular pathways implicated in the regulation of the osseous healing process following guided bone regeneration (GBR). Six 6-month-old Wistar male rats were used. Standardized 5-mm critical size defects were created in the parietal bones of each animal and treated with an extracranial and intracranial ePTFE membrane, according to the GBR principle. Three animals were randomly sacrificed after 7 and 15 days of healing. Total RNA was extracted from each sample and prepared for gene expression analysis. RNA quality and quantity were assessed, followed by hybridization of the cRNA to Affymetrix GeneChip Rat Genome 230 2.0 Arrays. The Affymetrix data were processed, and first-order analysis, quality control and statistical analysis were performed. Biological interpretation was performed via pathway and Gene Ontology (GO) analysis. Between the 7- and 15-day samples, 538 genes were differently regulated. At day 7, inflammatory and immune responses were clearly upregulated. In addition, GO terms related to angiogenesis and cell cycle regulation were overexpressed. At day 15, a more complex cellular activity and cell metabolism were evident. The bone formation processes were significantly overexpressed, with several genes encoding growth factors, enzyme activity, and extracellular matrix formation found as upregulated. Remarkably, a negative regulation of Wnt signalling pathway was observed at 15 days. The gene expression profile of the cells participating in osseous formation varied depending on the healing stage. A number of candidate genes that seem differentially expressed during early stages of intramembranous bone regeneration was suggested. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  1. [Analysis of gene mutation of early onset epileptic spasm with unknown reason].

    Science.gov (United States)

    Yang, X; Pan, G; Li, W H; Zhang, L M; Wu, B B; Wang, H J; Zhang, P; Zhou, S Z

    2017-11-02

    Objective: To summarize the gene mutation of early onset epileptic spasm with unknown reason. Method: In this prospective study, data of patients with early onset epileptic spasm with unknown reason were collected from neurological department of Children's Hospital of Fudan University between March 2016 and December 2016. Patients with known disorders such as infection, metabolic, structural, immunological problems and known genetic mutations were excluded. Patients with genetic disease that can be diagnosed by clinical manifestations and phenotypic characteristics were also excluded. Genetic research methods included nervous system panel containing 1 427 epilepsy genes, whole exome sequencing (WES), analysis of copy number variation (CNV) and karyotype analysis of chromosome. The basic information, phenotypes, genetic results and the antiepileptic treatment of patients were analyzed. Result: Nine of the 17 cases with early onset epileptic spasm were boys and eight were girls. Patients' age at first seizure onset ranged from 1 day after birth to 8 months (median age of 3 months). The first hospital visit age ranged from 1 month to 2 years (median age of 4.5 months). The time of following-up ranged from 8 months to 3 years and 10 months. All the 17 patients had early onset epileptic spasm. Video electroencephalogram was used to monitor the spasm seizure. Five patients had Ohtahara syndrome, 10 had West syndrome, two had unclear classification. In 17 cases, 10 of them had detected pathogenic genes. Nine cases had point mutations, involving SCN2A, ARX, UNC80, KCNQ2, and GABRB3. Except one case of mutations in GABRB3 gene have been reported, all the other cases had new mutations. One patient had deletion mutation in CDKL5 gene. One CNV case had 6q 22.31 5.5MB repeats. Ten cases out of 17 were using 2-3 antiepileptic drugs (AEDs) and the drugs had no effect. Seven cases used adrenocorticotropic hormone (ACTH) and prednisone besides AEDs (a total course for 8 weeks

  2. Early community context, genes, and youth body mass index trajectories: an investigation of gene-community interplay over early life course.

    Science.gov (United States)

    Wickrama, Kandauda K A S; O'Neal, Catherine Walker; Lee, Tae Kyoung

    2013-09-01

    To investigate additive and interactive influences of community adversity and cumulative genetic sensitivity on youth body mass index (BMI) trajectories over adolescence and young adulthood. We used latent growth curve modeling to examine BMI trajectories over three waves (1995, 2001, and 2008) of the National Longitudinal Study of Adolescent Health (n = 14,563). We measured genetic sensitivity by a cumulative index of genes associated with serotonin and dopamine functions. Community adversity was positively associated with the initial level and rate of change in BMI trajectories over time. Adolescents experiencing community adversity had a higher BMI at Wave 1 and gained weight more quickly than those who did not live in adverse communities. Community adversity interacted with cumulative genetic sensitivity to explain variation in the rate of change in BMI trajectories. The influence of community adversity was greater for those with more sensitivity alleles than those with fewer sensitivity alleles. Gender, race/ethnicity, and family contexts were also associated with youth BMI trajectories. Community adversity in early adolescence, and its interaction with genes, has far-reaching consequences, including the rate of change in BMI trajectories extending into adulthood. This work has practical implications for future intervention/prevention programs. Published by Elsevier Inc.

  3. Natural variation in gene expression in the early development of dauer larvae of Caenorhabditis elegans

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    Barker Gary LA

    2009-07-01

    Full Text Available Abstract Background The free-living nematode Caenorhabditis elegans makes a developmental decision based on environmental conditions: larvae either arrest as dauer larva, or continue development into reproductive adults. There is natural variation among C. elegans lines in the sensitivity of this decision to environmental conditions; that is, there is variation in the phenotypic plasticity of dauer larva development. We hypothesised that these differences may be transcriptionally controlled in early stage larvae. We investigated this by microarray analysis of different C. elegans lines under different environmental conditions, specifically the presence and absence of dauer larva-inducing pheromone. Results There were substantial transcriptional differences between four C. elegans lines under the same environmental conditions. The expression of approximately 2,000 genes differed between genetically different lines, with each line showing a largely line-specific transcriptional profile. The expression of genes that are markers of larval moulting suggested that the lines may be developing at different rates. The expression of a total of 89 genes was putatively affected by dauer larva or non-dauer larva-inducing conditions. Among the upstream regions of these genes there was an over-representation of DAF-16-binding motifs. Conclusion Under the same environmental conditions genetically different lines of C. elegans had substantial transcriptional differences. This variation may be due to differences in the developmental rates of the lines. Different environmental conditions had a rather smaller effect on transcription. The preponderance of DAF-16-binding motifs upstream of these genes was consistent with these genes playing a key role in the decision between development into dauer or into non-dauer larvae. There was little overlap between the genes whose expression was affected by environmental conditions and previously identified loci involved in

  4. Alternative haplotypes of antigen processing genes in zebrafish diverged early in vertebrate evolution

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    McConnell, Sean C.; Hernandez, Kyle M.; Wcisel, Dustin J.; Kettleborough, Ross N.; Stemple, Derek L.; Andrade, Jorge; de Jong, Jill L. O.

    2016-01-01

    Antigen processing and presentation genes found within the MHC are among the most highly polymorphic genes of vertebrate genomes, providing populations with diverse immune responses to a wide array of pathogens. Here, we describe transcriptome, exome, and whole-genome sequencing of clonal zebrafish, uncovering the most extensive diversity within the antigen processing and presentation genes of any species yet examined. Our CG2 clonal zebrafish assembly provides genomic context within a remarkably divergent haplotype of the core MHC region on chromosome 19 for six expressed genes not found in the zebrafish reference genome: mhc1uga, proteasome-β 9b (psmb9b), psmb8f, and previously unknown genes psmb13b, tap2d, and tap2e. We identify ancient lineages for Psmb13 within a proteasome branch previously thought to be monomorphic and provide evidence of substantial lineage diversity within each of three major trifurcations of catalytic-type proteasome subunits in vertebrates: Psmb5/Psmb8/Psmb11, Psmb6/Psmb9/Psmb12, and Psmb7/Psmb10/Psmb13. Strikingly, nearby tap2 and MHC class I genes also retain ancient sequence lineages, indicating that alternative lineages may have been preserved throughout the entire MHC pathway since early diversification of the adaptive immune system ∼500 Mya. Furthermore, polymorphisms within the three MHC pathway steps (antigen cleavage, transport, and presentation) are each predicted to alter peptide specificity. Lastly, comparative analysis shows that antigen processing gene diversity is far more extensive than previously realized (with ancient coelacanth psmb8 lineages, shark psmb13, and tap2t and psmb10 outside the teleost MHC), implying distinct immune functions and conserved roles in shaping MHC pathway evolution throughout vertebrates. PMID:27493218

  5. Early bovine embryos regulate oviduct epithelial cell gene expression during in vitro co-culture.

    Science.gov (United States)

    Schmaltz-Panneau, Barbara; Cordova, Amanda; Dhorne-Pollet, Sophie; Hennequet-Antier, Christelle; Uzbekova, Sveltlana; Martinot, Emmanuelle; Doret, Sarah; Martin, Patrice; Mermillod, Pascal; Locatelli, Yann

    2014-10-01

    In mammals, the oviduct may participate to the regulation of early embryo development. In vitro co-culture of early bovine embryos with bovine oviduct epithelial cells (BOEC) has been largely used to mimic the maternal environment. However, the mechanisms of BOEC action have not been clearly elucidated yet. The aim of this study was to determine the response of BOEC cultures to the presence of developing bovine embryos. A 21,581-element bovine oligonucleotide array was used compare the gene expression profiles of confluent BOEC cultured for 8 days with or without embryos. This study revealed 34 differentially expressed genes (DEG). Of these 34 genes, IFI6, ISG15, MX1, IFI27, IFI44, RSAD2, IFITM1, EPSTI1, USP18, IFIT5, and STAT1 expression increased to the greatest extent due to the presence of embryos with a major impact on antiviral and immune response. Among the mRNAs at least 25 are already described as induced by interferons. In addition, transcript levels of new candidate genes involved in the regulation of transcription, modulation of the maternal immune system and endometrial remodeling were found to be increased. We selected 7 genes and confirmed their differential expression by quantitative RT-PCR. The immunofluorescence imaging of cellular localization of STAT1 protein in BOEC showed a nuclear translocation in the presence of embryos, suggesting the activation of interferon signaling pathway. This first systematic study of BOEC transcriptome changes in response to the presence of embryos in cattle provides some evidences that these cells are able to adapt their transcriptomic profile in response to embryo signaling. Copyright © 2014 Elsevier B.V. All rights reserved.

  6. Comparative Genetic Variability in HIV-1 Subtype C vpu Gene in Early Age Groups of Infants.

    Science.gov (United States)

    Sharma, Uma; Gupta, Poonam; Gupta, Sunil; Venkatesh, S; Husain, Mohammad

    2018-01-01

    Identifying the genetic variability in vertically transmitted viruses in early infancy is important to understand the disease progression. Being important in HIV-1 disease pathogenesis, vpu gene, isolated from young infants was investigated to understand the viral characteristics. Blood samples were obtained from 80 HIV-1 positive infants, categorized in two age groups; acute (6-18 months). A total of 77 PCR positive samples, amplified for vpu gene, were sequenced and analyzed. 73 isolates belonged to subtype C. Analysis of heterogeneity of amino acid sequences in infant groups showed that in the sequences of acute age group both insertions and deletions were present while in the early age group only deletions were present. In the acute age group, a deletion of 3 residues (RAE) in the first alfa helix in one sequence and insertions of 1-2 residues (DM, GH, G and H) in the second alfa helix in 4 sequences were observed. In the early age group, deletion of 2 residues (VN) in the cytoplasmic tail region in 2 sequences was observed. Length of the amino terminal was observed to be gradually increasing with the increasing age of the infants. Protein Variation Effect Analyzer software showed that deleterious mutations were more in the acute than the early age group. Entropy analysis revealed that heterogeneity of the residues was comparatively higher in the sequences of acute than the early age group. Mutations observed in the helixes may affect the conformation and lose the ability to degrade CD4 receptors. Heterogeneity was decreasing with the increasing ages of the infants, indicating positive selection for robust virion survival. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  7. In vitro modeling of human pancreatic duct epithelial cell transformation defines gene expression changes induced by K-ras oncogenic activation in pancreatic carcinogenesis.

    Science.gov (United States)

    Qian, Jiaying; Niu, Jiangong; Li, Ming; Chiao, Paul J; Tsao, Ming-Sound

    2005-06-15

    Genetic analysis of pancreatic ductal adenocarcinomas and their putative precursor lesions, pancreatic intraepithelial neoplasias (PanIN), has shown a multistep molecular paradigm for duct cell carcinogenesis. Mutational activation or inactivation of the K-ras, p16(INK4A), Smad4, and p53 genes occur at progressive and high frequencies in these lesions. Oncogenic activation of the K-ras gene occurs in >90% of pancreatic ductal carcinoma and is found early in the PanIN-carcinoma sequence, but its functional roles remain poorly understood. We show here that the expression of K-ras(G12V) oncogene in a near diploid HPV16-E6E7 gene immortalized human pancreatic duct epithelial cell line originally derived from normal pancreas induced the formation of carcinoma in 50% of severe combined immunodeficient mice implanted with these cells. A tumor cell line established from one of these tumors formed ductal cancer when implanted orthotopically. These cells also showed increased activation of the mitogen-activated protein kinase, AKT, and nuclear factor-kappaB pathways. Microarray expression profiling studies identified 584 genes whose expression seemed specifically up-regulated by the K-ras oncogene expression. Forty-two of these genes have been reported previously as differentially overexpressed in pancreatic cancer cell lines or primary tumors. Real-time PCR confirmed the overexpression of a large number of these genes. Immunohistochemistry done on tissue microarrays constructed from PanIN and pancreatic cancer samples showed laminin beta3 overexpression starting in high-grade PanINs and occurring in >90% of pancreatic ductal carcinoma. The in vitro modeling of human pancreatic duct epithelial cell transformation may provide mechanistic insights on gene expression changes that occur during multistage pancreatic duct cell carcinogenesis.

  8. The interaction of corticotropin-releasing hormone receptor gene and early life stress on emotional empathy.

    Science.gov (United States)

    Grimm, Simone; Wirth, Katharina; Fan, Yan; Weigand, Anne; Gärtner, Matti; Feeser, Melanie; Dziobek, Isabel; Bajbouj, Malek; Aust, Sabine

    2017-06-30

    Early life stress (ELS) is associated with increased vulnerability for depression, changes to the corticotropin-releasing hormone (CRH) system and structural and functional changes in hippocampus. Single nucleotide polymorphisms in the CRH receptor 1 (CRHR1) gene interact with ELS to predict depression, cognitive functions and hippocampal activity. Social cognition has been related to hippocampal function and might be crucial for maintaining mental health. However, the interaction of CRHR1 gene variation and ELS on social cognition has not been investigated yet. We assessed social cognition in 502 healthy subjects to test effects of ELS and the CRHR1 gene. Participants were genotyped for rs110402 and rs242924. ELS was assessed by Childhood Trauma Questionnaire, social cognition was measured via Multifaceted Empathy Test and Empathy Quotient. Severity of ELS was associated with decreased emotional, but not cognitive empathy. Subjects with the common homozygous GG GG genotype showed decreased implicit emotional empathy after ELS exposure regardless of its severity. The results reveal that specific CRHR1 polymorphisms moderate the effect of ELS on emotional empathy. Exposure to ELS in combination with a vulnerable genotype results in impaired emotional empathy in adulthood, which might represent an early marker of increased vulnerability after ELS. Copyright © 2017 Elsevier B.V. All rights reserved.

  9. Gene expression in the mouse brain following early pregnancy exposure to ethanol

    Directory of Open Access Journals (Sweden)

    Christine R. Zhang

    2016-12-01

    Full Text Available Exposure to alcohol during early embryonic or fetal development has been linked with a variety of adverse outcomes, the most common of which are structural and functional abnormalities of the central nervous system [1]. Behavioural and cognitive deficits reported in individuals exposed to alcohol in utero include intellectual impairment, learning and memory difficulties, diminished executive functioning, attention problems, poor motor function and hyperactivity [2]. The economic and social costs of these outcomes are substantial and profound [3,4]. Improvement of neurobehavioural outcomes following prenatal alcohol exposure requires greater understanding of the mechanisms of alcohol-induced damage to the brain. Here we use a mouse model of relatively moderate ethanol exposure early in pregnancy and profile gene expression in the hippocampus and caudate putamen of adult male offspring. The effects of offspring sex and age on ethanol-sensitive hippocampal gene expression were also examined. All array data are available at the Gene Expression Omnibus (GEO repository under accession number GSE87736.

  10. Oxytocin receptor gene polymorphisms (rs53576) and early paternal care sensitize males to distressing female vocalizations.

    Science.gov (United States)

    Truzzi, Anna; Poquérusse, Jessie; Setoh, Peipei; Shinohara, Kazuyuki; Bornstein, Marc H; Esposito, Gianluca

    2018-04-01

    The oxytocinergic system is highly involved in social bonding and early caregiver-infant interactions. Here, we hypothesize that oxytocin receptor (OXTR) gene genotype and parental bonding history interact in influencing social development. To address this question, we assessed adult males' arousal (heart rate changes) in response to different distress vocalizations (human female, human infant and bonobo). Region rs53576 of the OXTR gene was genotyped from buccal mucosa cell samples, and a self-report Parental Bonding Instrument was used (which provide information about parental care or parental overprotection). A significant gene-environment interaction between OXTR genotype and parenting style was found to influence participants' social responsivity to female cry vocalizations. Specifically, a history of appropriate paternal care in participants accentuated the heightened social sensitivity determined by G/G homozygosity, while higher versus lower paternal overprotection lead to distinct levels of physiological arousal particularly in A carriers individuals. These results add to our understanding of the dynamic interplay between genetic susceptibility and early environmental experience in shaping the development of appropriate social sensitivity in males. © 2018 Wiley Periodicals, Inc.

  11. Expression of immediate-early genes in the dorsal cochlear nucleus in salicylate-induced tinnitus.

    Science.gov (United States)

    Hu, Shou-Sen; Mei, Ling; Chen, Jian-Yong; Huang, Zhi-Wu; Wu, Hao

    2016-02-01

    Spontaneous neuronal activity in dorsal cochlear nucleus (DCN) may be involved in the physiological processes underlying salicylate-induced tinnitus. As a neuronal activity marker, immediate-early gene (IEG) expression, especially activity-dependent cytoskeletal protein (Arc/Arg3.1) and the early growth response gene-1 (Egr-1), appears to be highly correlated with sensory-evoked neuronal activity. However, their relationships with tinnitus induced by salicylate have rarely been reported in the DCN. In this study, we assessed the effect of acute and chronic salicylate treatment on the expression of N-methyl D-aspartate receptor subunit 2B (NR2B), Arg3.1, and Egr-1. We also observed ultrastructural alterations in the DCN synapses in an animal model of tinnitus. Levels of mRNA and protein expression of NR2B and Arg3.1 were increased in rats that were chronically administered salicylate (200 mg/kg, twice daily for 3, 7, or 14 days). These levels returned to baseline 14 days after cessation of treatment. However, no significant changes were observed in Egr-1 gene expression in any groups. Furthermore, rats subjected to long-term salicylate administration showed more presynaptic vesicles, thicker and longer postsynaptic densities, and increased synaptic interface curvature. Alterations of Arg3.1 and NR2B may be responsible for the changes in the synaptic ultrastructure. These changes confirm that salicylate can cause neural plasticity changes at the DCN level.

  12. Type II cytokeratin gene expression is indicative of early cell differentiation in the chick embryo

    International Nuclear Information System (INIS)

    Charlebois, T.S.

    1988-01-01

    Embryonic development in vertebrates appears to involve a series of inductive tissue interactions that lead to regional specializations, which eventually become elaborated in the basic body plan of the embryo. The inductive interactions leading to early regionalization of the embryo are often particularly difficult to evaluate because of the absence of available morphological or biochemical evidence that such events have occurred. In the 36 hour chick embryo, the regional subdivision of the early ectoderm is evidence by a marked lens-forming bias in the head ectoderm, which is absent in the presumptive dorsal epidermis of the trunk region. As a strategy for isolating genes whose differential expression might reflect this regional subdivision, a cDNA library from 36 hour embryos was prepared and screened for differential hybridization to [ 32 P]cDNA probes synthesized using template RNA isolated from 36 hour head ectoderm and trunk ectoderm. A cDNA clone (T4) was isolated which hybridizes to transcripts present at much higher levels in trunk ectoderm than in head ectoderm. Partial nucleotide and deduced amino acid sequences of this clone indicate that it represents a gene encoding a type II cytokeratin. The distribution of transcripts complementary to the T4 probe was evaluated in early embryos using RNA gel blot analysis and in situ hybridization to tissue sections

  13. Inositol polyphosphate multikinase is a coactivator for serum response factor-dependent induction of immediate early genes

    Science.gov (United States)

    Kim, Eunha; Tyagi, Richa; Lee, Joo-Young; Park, Jina; Kim, Young-ran; Beon, Jiyoon; Chen, Po Yu; Cha, Jiyoung Y.; Snyder, Solomon H.; Kim, Seyun

    2013-01-01

    Inositol polyphosphate multikinase (IPMK) is a notably pleiotropic protein. It displays both inositol phosphate kinase and phosphatidylinositol kinase catalytic activities. Noncatalytically, IPMK stabilizes the mammalian target of rapamycin complex 1 and acts as a transcriptional coactivator for CREB-binding protein/E1A binding protein p300 and tumor suppressor protein p53. Serum response factor (SRF) is a major transcription factor for a wide range of immediate early genes. We report that IPMK, in a noncatalytic role, is a transcriptional coactivator for SRF mediating the transcription of immediate early genes. Stimulation by serum of many immediate early genes is greatly reduced by IPMK deletion. IPMK stimulates expression of these genes, an influence also displayed by catalytically inactive IPMK. IPMK acts by binding directly to SRF and thereby enhancing interactions of SRF with the serum response element of diverse genes. PMID:24248338

  14. The Immediate Early Gene Egr3 Is Required for Hippocampal Induction of Bdnf by Electroconvulsive Stimulation

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    Kimberly T. Meyers

    2018-05-01

    Full Text Available Early growth response 3 (Egr3 is an immediate early gene (IEG that is regulated downstream of a cascade of genes associated with risk for psychiatric disorders, and dysfunction of Egr3 itself has been implicated in schizophrenia, bipolar disorder, and depression. As an activity-dependent transcription factor, EGR3 is poised to regulate the neuronal expression of target genes in response to environmental events. In the current study, we sought to identify a downstream target of EGR3 with the goal of further elucidating genes in this biological pathway relevant for psychiatric illness risk. We used electroconvulsive stimulation (ECS to induce high-level expression of IEGs in the brain, and conducted expression microarray to identify genes differentially regulated in the hippocampus of Egr3-deficient (-/- mice compared to their wildtype (WT littermates. Our results replicated previous work showing that ECS induces high-level expression of the brain-derived neurotrophic factor (Bdnf in the hippocampus of WT mice. However, we found that this induction is absent in Egr3-/- mice. Quantitative real-time PCR (qRT-PCR validated the microarray results (performed in males and replicated the findings in two separate cohorts of female mice. Follow-up studies of activity-dependent Bdnf exons demonstrated that ECS-induced expression of both exons IV and VI requires Egr3. In situ hybridization demonstrated high-level cellular expression of Bdnf in the hippocampal dentate gyrus following ECS in WT, but not Egr3-/-, mice. Bdnf promoter analysis revealed eight putative EGR3 binding sites in the Bdnf promoter, suggesting a mechanism through which EGR3 may directly regulate Bdnf gene expression. These findings do not appear to result from a defect in the development of hippocampal neurons in Egr3-/- mice, as cell counts in tissue sections stained with anti-NeuN antibodies, a neuron-specific marker, did not differ between Egr3-/- and WT mice. In addition, Sholl

  15. The presence of p53 influences the expression of multiple human cytomegalovirus genes at early times postinfection.

    Science.gov (United States)

    Hannemann, Holger; Rosenke, Kyle; O'Dowd, John M; Fortunato, Elizabeth A

    2009-05-01

    Human cytomegalovirus (HCMV) is a common cause of morbidity and mortality in immunocompromised and immunosuppressed individuals. During infection, HCMV is known to employ host transcription factors to facilitate viral gene expression. To further understand the previously observed delay in viral replication and protein expression in p53 knockout cells, we conducted microarray analyses of p53(+/+) and p53(-/-) immortalized fibroblast cell lines. At a multiplicity of infection (MOI) of 1 at 24 h postinfection (p.i.), the expression of 22 viral genes was affected by the absence of p53. Eleven of these 22 genes (group 1) were examined by real-time reverse transcriptase, or quantitative, PCR (q-PCR). Additionally, five genes previously determined to have p53 bound to their nearest p53-responsive elements (group 2) and three control genes without p53 binding sites in their upstream sequences (group 3) were also examined. At an MOI of 1, >3-fold regulation was found for five group 1 genes. The expression of group 2 and 3 genes was not changed. At an MOI of 5, all genes from group 1 and four of five genes from group 2 were found to be regulated. The expression of control genes from group 3 remained unchanged. A q-PCR time course of four genes revealed that p53 influences viral gene expression most at immediate-early and early times p.i., suggesting a mechanism for the reduced and delayed production of virions in p53(-/-) cells.

  16. Early life trauma, depression and the glucocorticoid receptor gene--an epigenetic perspective.

    Science.gov (United States)

    Smart, C; Strathdee, G; Watson, S; Murgatroyd, C; McAllister-Williams, R H

    2015-12-01

    Hopes to identify genetic susceptibility loci accounting for the heritability seen in unipolar depression have not been fully realized. Family history remains the 'gold standard' for both risk stratification and prognosis in complex phenotypes such as depression. Meanwhile, the physiological mechanisms underlying life-event triggers for depression remain opaque. Epigenetics, comprising heritable changes in gene expression other than alterations of the nucleotide sequence, may offer a way to deepen our understanding of the aetiology and pathophysiology of unipolar depression and optimize treatments. A heuristic target for exploring the relevance of epigenetic changes in unipolar depression is the hypothalamic-pituitary-adrenal (HPA) axis. The glucocorticoid receptor (GR) gene (NR3C1) has been found to be susceptible to epigenetic modification, specifically DNA methylation, in the context of environmental stress such as early life trauma, which is an established risk for depression later in life. In this paper we discuss the progress that has been made by studies that have investigated the relationship between depression, early trauma, the HPA axis and the NR3C1 gene. Difficulties with the design of these studies are also explored. Future efforts will need to comprehensively address epigenetic natural histories at the population, tissue, cell and gene levels. The complex interactions between the epigenome, genome and environment, as well as ongoing nosological difficulties, also pose significant challenges. The work that has been done so far is nevertheless encouraging and suggests potential mechanistic and biomarker roles for differential DNA methylation patterns in NR3C1 as well as novel therapeutic targets.

  17. Interactions of early adversity with stress-related gene polymorphisms impact regional brain structure in females.

    Science.gov (United States)

    Gupta, Arpana; Labus, Jennifer; Kilpatrick, Lisa A; Bonyadi, Mariam; Ashe-McNalley, Cody; Heendeniya, Nuwanthi; Bradesi, Sylvie; Chang, Lin; Mayer, Emeran A

    2016-04-01

    Early adverse life events (EALs) have been associated with regional thinning of the subgenual cingulate cortex (sgACC), a brain region implicated in the development of disorders of mood and affect, and often comorbid functional pain disorders, such as irritable bowel syndrome (IBS). Regional neuroinflammation related to chronic stress system activation has been suggested as a possible mechanism underlying these neuroplastic changes. However, the interaction of genetic and environmental factors in these changes is poorly understood. The current study aimed to evaluate the interactions of EALs and candidate gene polymorphisms in influencing thickness of the sgACC. 210 female subjects (137 healthy controls; 73 IBS) were genotyped for stress and inflammation-related gene polymorphisms. Genetic variation with EALs, and diagnosis on sgACC thickness was examined, while controlling for race, age, and total brain volume. Compared to HCs, IBS had significantly reduced sgACC thickness (p = 0.03). Regardless of disease group (IBS vs. HC), thinning of the left sgACC was associated with a significant gene-gene environment interaction between the IL-1β genotype, the NR3C1 haplotype, and a history of EALs (p = 0.05). Reduced sgACC thickness in women with the minor IL-1β allele, was associated with EAL total scores regardless of NR3C1 haplotype status (p = 0.02). In subjects homozygous for the major IL-1β allele, reduced sgACC with increasing levels of EALs was seen only with the less common NR3C1 haplotype (p = 0.02). These findings support an interaction between polymorphisms related to stress and inflammation and early adverse life events in modulating a key region of the emotion arousal circuit.

  18. A gene expression profile indicative of early stage HER2 targeted therapy response.

    Science.gov (United States)

    O'Neill, Fiona; Madden, Stephen F; Clynes, Martin; Crown, John; Doolan, Padraig; Aherne, Sinéad T; O'Connor, Robert

    2013-07-01

    Efficacious application of HER2-targetting agents requires the identification of novel predictive biomarkers. Lapatinib, afatinib and neratinib are tyrosine kinase inhibitors (TKIs) of HER2 and EGFR growth factor receptors. A panel of breast cancer cell lines was treated with these agents, trastuzumab, gefitinib and cytotoxic therapies and the expression pattern of a specific panel of genes using RT-PCR was investigated as a potential marker of early drug response to HER2-targeting therapies. Treatment of HER2 TKI-sensitive SKBR3 and BT474 cell lines with lapatinib, afatinib and neratinib induced an increase in the expression of RB1CC1, ERBB3, FOXO3a and NR3C1. The response directly correlated with the degree of sensitivity. This expression pattern switched from up-regulated to down-regulated in the HER2 expressing, HER2-TKI insensitive cell line MDAMB453. Expression of the CCND1 gene demonstrated an inversely proportional response to drug exposure. A similar expression pattern was observed following the treatment with both neratinib and afatinib. These patterns were retained following exposure to traztuzumab and lapatinib plus capecitabine. In contrast, gefitinib, dasatinib and epirubicin treatment resulted in a completely different expression pattern change. In these HER2-expressing cell line models, lapatinib, neratinib, afatinib and trastuzumab treatment generated a characteristic and specific gene expression response, proportionate to the sensitivity of the cell lines to the HER2 inhibitor.Characterisation of the induced changes in expression levels of these genes may therefore give a valuable, very early predictor of the likely extent and specificity of tumour HER2 inhibitor response in patients, potentially guiding more specific use of these agents.

  19. Early Gene Expression in Wounded Human Keratinocytes Revealed by DNA Microarray Analysis

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    Pascal Barbry

    2006-04-01

    Full Text Available Wound healing involves several steps: spreading of the cells, migration and proliferation. We have profiled gene expression during the early events of wound healing in normal human keratinocytes with a home-made DNA microarray containing about 1000 relevant human probes. An original wounding machine was used, that allows the wounding of up to 40% of the surface of a confluent monolayer of cultured cells grown on a Petri dish (compared with 5% with a classical ‘scratch’ method. The two aims of the present study were: (a to validate a limited number of genes by comparing the expression levels obtained with this technique with those found in the literature; (b to combine the use of the wounding machine with DNA microarray analysis for large-scale detection of the molecular events triggered during the early stages of the wound-healing process. The time-courses of RNA expression observed at 0.5, 1.5, 3, 6 and 15 h after wounding for genes such as c-Fos, c-Jun, Egr1, the plasminogen activator PLAU (uPA and the signal transducer and transcription activator STAT3, were consistent with previously published data. This suggests that our methodologies are able to perform quantitative measurement of gene expression. Transcripts encoding two zinc finger proteins, ZFP36 and ZNF161, and the tumour necrosis factor α-induced protein TNFAIP3, were also overexpressed after wounding. The role of the p38 mitogen-activated protein kinase (p38MAPK in wound healing was shown after the inhibition of p38 by SB203580, but our results also suggest the existence of surrogate activating pathways.

  20. Early Involvement of Immune/Inflammatory Response Genes in Retinal Degeneration in DBA/2J Mice

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    W. Fan

    2010-01-01

    , representative of different functions/pathways, were validated with RT-PCR, and changes in glial responses were visualized in the retina with immunocytochemistry. Conclusions The results showed that the expression of genes related to the immune response and acute stress were altered independently of the development of elevated IOP, and indicated early involvement of the immune system in the onset of the disease. The later development of elevated IOP, observed in this animal model, was coincident with continued changes in expression of genes observed at earlier time points. Further studies are warranted to identify the roles of specific genes identified here with respect to the death of the RGCs.

  1. Early Involvement of Immune/Inflammatory Response Genes in Retinal Degeneration in DBA/2J Mice

    Directory of Open Access Journals (Sweden)

    W. Fan

    2010-03-01

    , representative of different functions/pathways, were validated with RT-PCR, and changes in glial responses were visualized in the retina with immunocytochemistry. Conclusions: The results showed that the expression of genes related to the immune response and acute stress were altered independently of the development of elevated IOP, and indicated early involvement of the immune system in the onset of the disease. The later development of elevated IOP, observed in this animal model, was coincident with continued changes in expression of genes observed at earlier time points. Further studies are warranted to identify the roles of specific genes identified here with respect to the death of the RGCs.

  2. Transcription mapping and expression patterns of genes in the major immediate-early region of Kaposi's sarcoma-associated herpesvirus.

    Science.gov (United States)

    Saveliev, Alexei; Zhu, Fan; Yuan, Yan

    2002-08-01

    Viral immediate-early (IE) genes are the first class of viral genes expressed during primary infection or reactivation from latency. They usually encode regulatory proteins that play crucial roles in viral life cycle. In a previous study, four regions in the KSHV genome were found to be actively transcribed in the immediate-early stage of viral reactivation in primary effusion lymphoma cells. Three immediate-early transcripts were characterized in these regions, as follows: mRNAs for ORF50 (KIE-1), ORF-45 (KIE-2), and ORF K4.2 (KIE-3) (F. X. Zhu, T. Cusano, and Y. Yuan, 1999, J. Virol. 73, 5556-5567). In the present study, we further analyzed the expression of genes in these IE regions in BC-1 and BCBL-1 cells. One of the immediate-early regions (KIE-1) that encompasses ORF50 and other genes was intensively studied to establish a detailed transcription map and expression patterns of genes in this region. This study led to identification of several novel IE transcripts in this region. They include a 2.6-kb mRNA which encodes ORF48/ORF29b, a family of transcripts that are complementary to ORF50 mRNA and a novel K8 IE mRNA of 1.5 kb. Together with the IE mRNA for ORF50 which was identified previously, four immediate-early genes have been mapped to KIE-1 region. Therefore, we would designate KIE-1 the major immediate-early region of KSHV. In addition, we showed that transcription of K8 gene is controlled by two promoters, yielding two transcripts, an immediate-early mRNA of 1.5 kb and a delayed-early mRNA of 1.3 kb.

  3. Novel Pectate Lyase Genes of Heterodera glycines Play Key Roles in the Early Stage of Parasitism.

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    Huan Peng

    Full Text Available Pectate lyases are known to play a key role in pectin degradation by catalyzing the random cleavage of internal polymer linkages (endo-pectinases. In this paper, four novel cDNAs, designated Hg-pel-3, Hg-pel-4, Hg-pel-6 and Hg-pel-7, that encode pectate lyases were cloned and characterized from the soybean cyst nematode, Heterodera glycines. The predicted protein sequences of HG-PEL-3, HG-PEL-4 and HG-PEL-6 differed significantly in both their amino acid sequences and their genomic structures from other pectate lyases of H. glycines (HG-PEL-1, HG-PEL-2 and HG-PEL-7. A phylogenetic study revealed that the pectate lyase proteins of H. glycines are clustered into distinct clades and have distinct numbers and positioning of introns, which suggests that the pectate lyase genes of H. glycines may have evolved from at least two ancestral genes. A Southern blot analysis revealed that multiple Hg-pel-6-like genes were present in the H. glycines genome. In situ hybridization showed that four novel pectate lyases (Hg-pel-3, Hg-pel-4, Hg-pel-6 and Hg-pel-7 were actively transcribed in the subventral esophageal gland cells. A semi-quantitative RT-PCR assay supported the finding that the expression of these genes was strong in the egg, pre-parasitic second-stage juvenile (J2 and early parasitic J2 stages and that it declined in further developmental stages of the nematode. This expression pattern suggests that these proteins play a role in the migratory phase of the nematode life cycle. Knocking down Hg-pel-6 using in vitro RNA interference resulted in a 46.9% reduction of the number of nematodes that invaded the plants and a 61.5% suppression of the development of H. glycines females within roots compared to the GFP-dsRNA control. Plant host-derived RNAi induced the silencing of the Hg-pel-6gene, which significantly reduced the nematode infection levels at 7 Days post inoculation (dpi. Similarly, this procedure reduced the number of female adults at 40 dpi

  4. Gene methylation profiles of normal mucosa, and benign and malignant colorectal tumors identify early onset markers

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    Vatn Morten

    2008-12-01

    Full Text Available Abstract Background Multiple epigenetic and genetic changes have been reported in colorectal tumors, but few of these have clinical impact. This study aims to pinpoint epigenetic markers that can discriminate between non-malignant and malignant tissue from the large bowel, i.e. markers with diagnostic potential. The methylation status of eleven genes (ADAMTS1, CDKN2A, CRABP1, HOXA9, MAL, MGMT, MLH1, NR3C1, PTEN, RUNX3, and SCGB3A1 was determined in 154 tissue samples including normal mucosa, adenomas, and carcinomas of the colorectum. The gene-specific and widespread methylation status among the carcinomas was related to patient gender and age, and microsatellite instability status. Possible CIMP tumors were identified by comparing the methylation profile with microsatellite instability (MSI, BRAF-, KRAS-, and TP53 mutation status. Results The mean number of methylated genes per sample was 0.4 in normal colon mucosa from tumor-free individuals, 1.2 in mucosa from cancerous bowels, 2.2 in adenomas, and 3.9 in carcinomas. Widespread methylation was found in both adenomas and carcinomas. The promoters of ADAMTS1, MAL, and MGMT were frequently methylated in benign samples as well as in malignant tumors, independent of microsatellite instability. In contrast, normal mucosa samples taken from bowels without tumor were rarely methylated for the same genes. Hypermethylated CRABP1, MLH1, NR3C1, RUNX3, and SCGB3A1 were shown to be identifiers of carcinomas with microsatellite instability. In agreement with the CIMP concept, MSI and mutated BRAF were associated with samples harboring hypermethylation of several target genes. Conclusion Methylated ADAMTS1, MGMT, and MAL are suitable as markers for early tumor detection.

  5. Influence of Dopamine-Related Genes on Neurobehavioral Recovery after Traumatic Brain Injury during Early Childhood.

    Science.gov (United States)

    Treble-Barna, Amery; Wade, Shari L; Martin, Lisa J; Pilipenko, Valentina; Yeates, Keith Owen; Taylor, H Gerry; Kurowski, Brad G

    2017-06-01

    The present study examined the association of dopamine-related genes with short- and long-term neurobehavioral recovery, as well as neurobehavioral recovery trajectories over time, in children who had sustained early childhood traumatic brain injuries (TBI) relative to children who had sustained orthopedic injuries (OI). Participants were recruited from a prospective, longitudinal study evaluating outcomes of children who sustained a TBI (n = 68) or OI (n = 72) between the ages of 3 and 7 years. Parents completed ratings of child executive function and behavior at the immediate post-acute period (0-3 months after injury); 6, 12, and 18 months after injury; and an average of 3.5 and 7 years after injury. Thirty-two single nucleotide polymorphisms (SNPs) in dopamine-related genes (dopamine receptor D2 [DRD2], solute carrier family 6 member 3 [SLC6A3], solute carrier family 18 member A2 [SLC18A2], catechol-o-methyltransferase [COMT], and ankyrin repeat and kinase domain containing 1 [ANKK1]) were examined in association with short- and long-term executive function and behavioral adjustment, as well as their trajectories over time. After controlling for premorbid child functioning, genetic variation within the SLC6A3 (rs464049 and rs460000) gene was differentially associated with neurobehavioral recovery trajectories over time following TBI relative to OI, with rs464049 surviving multiple testing corrections. In addition, genetic variation within the ANKK1 (rs1800497 and rs2734849) and SLC6A3 (rs464049, rs460000, and rs1042098) genes was differentially associated with short- and long-term neurobehavioral recovery following TBI, with rs460000 and rs464049 surviving multiple testing corrections. The findings provide preliminary evidence that genetic variation in genes involved in DRD2 expression and density (ANKK1) and dopamine transport (SLC6A3) plays a role in neurobehavioral recovery following pediatric TBI.

  6. The full-length E1-circumflexE4 protein of human papillomavirus type 18 modulates differentiation-dependent viral DNA amplification and late gene expression

    International Nuclear Information System (INIS)

    Wilson, Regina; Ryan, Gordon B.; Knight, Gillian L.; Laimins, Laimonis A.; Roberts, Sally

    2007-01-01

    Activation of the productive phase of the human papillomavirus (HPV) life cycle in differentiated keratinocytes is coincident with high-level expression of E1-circumflexE4 protein. To determine the role of E1-circumflexE4 in the HPV replication cycle, we constructed HPV18 mutant genomes in which expression of the full-length E1-circumflexE4 protein was abrogated. Undifferentiated keratinocytes containing mutant genomes showed enhanced proliferation when compared to cells containing wildtype genomes, but there were no differences in maintenance of viral episomes. Following differentiation, cells with mutant genomes exhibited reduced levels of viral DNA amplification and late gene expression, compared to wildtype genome-containing cells. This indicates that HPV18 E1-circumflexE4 plays an important role in regulating HPV late functions, and it may also function in the early phase of the replication cycle. Our finding that full-length HPV18 E1-circumflexE4 protein plays a significant role in promoting viral genome amplification concurs with a similar report with HPV31, but is in contrast to an HPV11 study where viral DNA amplification was not dependent on full-length E1-circumflexE4 expression, and to HPV16 where only C-terminal truncations in E1-circumflexE4 abrogated vegetative genome replication. This suggests that type-specific differences exist between various E1-circumflexE4 proteins

  7. Gene Expression Associated with Early and Late Chronotypes in Drosophila melanogaster

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    Mirko ePegoraro

    2015-05-01

    Full Text Available The circadian clock provides the temporal framework for rhythmic behavioural and metabolic functions. In the modern era of industrialization, work and social pressures, the clock function is often jeopardized, resulting in adverse and chronic effects on health. Understanding circadian clock function, particularly individual variation in diurnal phase preference (chronotype, and the molecular mechanisms underlying such chronotypes may lead to interventions that could abrogate clock dysfunction and improve human (and animal health and welfare. Our preliminary studies suggested that fruitflies, like humans, can be classified as early rising ‘larks’ or late rising ‘owls’, providing a convenient model system for these types of studies. We have identified strains of flies showing increased preference for morning emergence (Early or E from the pupal case, or more pronounced preference for evening emergence (Late or L. We have sampled pupae the day before eclosion (4th day after pupariation at 4 h intervals in the E and L strains, and examined differences in gene expression by RNAseq. We have identified differentially expressed transcripts between the E and L strains which provide candidate genes for studies of Drosophila chronotypes and their human orthologues.

  8. Association between polymorphisms in cancer-related genes and early onset of esophageal adenocarcinoma.

    Science.gov (United States)

    Wu, I-Chen; Zhao, Yang; Zhai, Rihong; Liu, Geoffrey; Ter-Minassian, Monica; Asomaning, Kofi; Su, Li; Liu, Chen-Yu; Chen, Feng; Kulke, Matthew H; Heist, Rebecca S; Christiani, David C

    2011-04-01

    There is an increasing incidence of esophageal adenocarcinoma (EA) among younger people in the western populations. However, the association between genetic polymorphisms and the age of EA onset is unclear. In this study, 1330 functional/tagging single-nucleotide polymorphisms (SNPs) from 354 cancer-related genes were genotyped in 335 white EA patients. Twenty important SNPs that have the highest importance scores and lowest classification error rate were identified by the random forest algorithm to be associated with early onset of EA (age ≤ 55 years). Subsequent logistic regression analysis indicated that 10 SNPs (rs2070744 of NOS3, rs720321 of BCL2, rs17757541 of BCL2, rs11775256 of TNFRSF10A, rs1035142 of CASP8, rs2236302 of MMP14, rs4740363 of ABL1, rs696217 of GHRL, rs2445762 of CYP19A1, and rs11941492 of VEGFR2/KDR) were significantly associated with early onset of EA (≤55 vs >55 years, all P polymorphisms in cancer-related genes, especially those in the apoptotic pathway, play an important role in the development of younger-aged EA in a dose-response manner.

  9. Filaggrin gene variants and atopic diseases in early childhood assessed longitudinally from birth

    DEFF Research Database (Denmark)

    Bønnelykke, Klaus; Pipper, Christian Bressen; Tavendale, Roger

    2010-01-01

    Copenhagen Prospective Study on Asthma in Childhood (COPSAC) was one of the discovery cohorts of the association between eczema and variants in the filaggrin coding gene (FLG). Here, we study the FLG-associated risk of asthma symptoms in early life and describe the temporal relationship in the de......Copenhagen Prospective Study on Asthma in Childhood (COPSAC) was one of the discovery cohorts of the association between eczema and variants in the filaggrin coding gene (FLG). Here, we study the FLG-associated risk of asthma symptoms in early life and describe the temporal relationship...... diagnosed prospectively by the investigators. FLG variants R501X and Del4 were determined in 382 Caucasians. Filaggrin variants increased risk of developing recurrent wheeze, asthma and asthma exacerbations (hazard ratio 1.82 [1.06-3.12], p = 0.03), which was expressed within the first 1.5 yr of life...... fully in the first year of life (point prevalence ratio for age 0-5 was 1.75 [1.29-2.37]; p-value = 0.0003) contrasting the increased risk of specific sensitization by age 4 (odds ratio 3.52 [1.72-7.25], p = 0.0007) but not age 1.5. This study describes a FLG-associated pattern of atopic diseases...

  10. Suicide attempts in children and adolescents: The place of clock genes and early rhythm dysfunction.

    Science.gov (United States)

    Olliac, Bertrand; Ouss, Lisa; Charrier, Annaëlle

    2016-11-01

    Suicide remains one of the leading causes of death among young people, and suicidal ideation and behavior are relatively common in healthy and clinical populations. Suicide risk in childhood and adolescence is often approached from the perspective of nosographic categories to which predictive variables for suicidal acts are often linked. The cascading effects resulting from altered clock genes in a pediatric population could participate in biological rhythm abnormalities and the emergence of suicide attempts through impaired regulation of circadian rhythms and emotional states with neurodevelopmental effects. Also, early trauma and stressful life events can alter the expression of clock genes and contribute to the emergence of suicide attempts. Alteration of clock genes might lead to desynchronized and abnormal circadian rhythms impairing in turn the synchronization between external and internal rhythms and therefore the adaptation of the individual to his/her internal and external environment with the development of psychiatric disorders associated with increased risk for suicide attempts. Copyright © 2017 Elsevier Ltd. All rights reserved.

  11. Advance in plasma SEPT9 gene methylation assay for colorectal cancer early detection.

    Science.gov (United States)

    Wang, Yu; Chen, Pei-Min; Liu, Rong-Bin

    2018-01-15

    This review article summarizes the research advances of the plasma-based SEPT9 gene methylation assay for the clinical detection of colorectal cancer and its limitations. Colorectal cancer is a common malignancy with a poor prognosis and a high mortality, for which early detection and diagnosis are particularly crucial for the high-risk groups. Increasing evidence supported that SEPT9 gene methylation is associated with the pathogenesis of colorectal cancer and that detecting the level of methylation of SEPT9 in the peripheral blood can be used for screening of colorectal cancer in susceptible populations. In recent years, the data obtained in clinical studies demonstrated that the SEPT9 gene methylation assay has a good diagnostic performance with regard to both sensitivity and specificity with the advantage of better acceptability, convenience and compliance with serological testing compared with fecal occult blood tests and carcinoembryonic antigen for colorectal cancer (CRC). Furthermore, the combination of multiple methods or markers has become a growing trend for CRC detection and screening. Nevertheless, the clinical availability of the methylated SEPT9 assay is still limited because of the large degree of sample heterogeneity caused by demographic characteristics, pathological features, comorbidities and/or technique selection. Another factor is the cost-effectiveness of colorectal cancer screening strategies that hinders its large-scale application. In addition, improvements in its accuracy in detecting adenomas and premalignant polyps are required.

  12. A peroxidase gene expressed during early developmental stages of the parasitic plant Orobanche ramosa.

    Science.gov (United States)

    González-Verdejo, Clara Isabel; Barandiaran, Xabier; Moreno, Maria Teresa; Cubero, José Ignacio; Di Pietro, Antonio

    2006-01-01

    Broomrapes (Orobanche spp.) are holoparasitic weeds that cause devastating losses in many economically important crops. The molecular mechanisms that control the early stages of host infection in Orobanche are poorly understood. In the present study, the role of peroxidase has been examined during pre-infection growth and development of O. ramosa, using an in vitro model system. Peroxidase activity was histochemically localized at the tips of actively growing radicles and nascent attachment organs. Addition of exogenous catalase resulted in a significant reduction in the apical growth rate of the radicle. The prx1 gene encoding a putative class III peroxidase was cloned from a cDNA library of O. ramosa and was found to be expressed specifically during the early stages of the parasitic life cycle. The exogenous addition of sucrose resulted in significantly reduced prx1 transcript levels and in a dramatic change in radicle development from polarized apical growth to isotropic growth and the formation of tubercle-like structures. The results indicate an important role of peroxidases during the early parasitic stages of Orobanche.

  13. Roles of Polypyrimidine Tract Binding Proteins in Major Immediate-Early Gene Expression and Viral Replication of Human Cytomegalovirus▿

    Science.gov (United States)

    Cosme, Ruth S. Cruz; Yamamura, Yasuhiro; Tang, Qiyi

    2009-01-01

    Human cytomegalovirus (HCMV), a member of the β subgroup of the family Herpesviridae, causes serious health problems worldwide. HCMV gene expression in host cells is a well-defined sequential process: immediate-early (IE) gene expression, early-gene expression, DNA replication, and late-gene expression. The most abundant IE gene, major IE (MIE) gene pre-mRNA, needs to be spliced before being exported to the cytoplasm for translation. In this study, the regulation of MIE gene splicing was investigated; in so doing, we found that polypyrimidine tract binding proteins (PTBs) strongly repressed MIE gene production in cotransfection assays. In addition, we discovered that the repressive effects of PTB could be rescued by splicing factor U2AF. Taken together, the results suggest that PTBs inhibit MIE gene splicing by competing with U2AF65 for binding to the polypyrimidine tract in pre-mRNA. In intron deletion mutation assays and RNA detection experiments (reverse transcription [RT]-PCR and real-time RT-PCR), we further observed that PTBs target all the introns of the MIE gene, especially intron 2, and affect gene splicing, which was reflected in the variation in the ratio of pre-mRNA to mRNA. Using transfection assays, we demonstrated that PTB knockdown cells induce a higher degree of MIE gene splicing/expression. Consistently, HCMV can produce more viral proteins and viral particles in PTB knockdown cells after infection. We conclude that PTB inhibits HCMV replication by interfering with MIE gene splicing through competition with U2AF for binding to the polypyrimidine tract in MIE gene introns. PMID:19144709

  14. Roles of polypyrimidine tract binding proteins in major immediate-early gene expression and viral replication of human cytomegalovirus.

    Science.gov (United States)

    Cosme, Ruth S Cruz; Yamamura, Yasuhiro; Tang, Qiyi

    2009-04-01

    Human cytomegalovirus (HCMV), a member of the beta subgroup of the family Herpesviridae, causes serious health problems worldwide. HCMV gene expression in host cells is a well-defined sequential process: immediate-early (IE) gene expression, early-gene expression, DNA replication, and late-gene expression. The most abundant IE gene, major IE (MIE) gene pre-mRNA, needs to be spliced before being exported to the cytoplasm for translation. In this study, the regulation of MIE gene splicing was investigated; in so doing, we found that polypyrimidine tract binding proteins (PTBs) strongly repressed MIE gene production in cotransfection assays. In addition, we discovered that the repressive effects of PTB could be rescued by splicing factor U2AF. Taken together, the results suggest that PTBs inhibit MIE gene splicing by competing with U2AF65 for binding to the polypyrimidine tract in pre-mRNA. In intron deletion mutation assays and RNA detection experiments (reverse transcription [RT]-PCR and real-time RT-PCR), we further observed that PTBs target all the introns of the MIE gene, especially intron 2, and affect gene splicing, which was reflected in the variation in the ratio of pre-mRNA to mRNA. Using transfection assays, we demonstrated that PTB knockdown cells induce a higher degree of MIE gene splicing/expression. Consistently, HCMV can produce more viral proteins and viral particles in PTB knockdown cells after infection. We conclude that PTB inhibits HCMV replication by interfering with MIE gene splicing through competition with U2AF for binding to the polypyrimidine tract in MIE gene introns.

  15. The immediate early gene product EGR1 and polycomb group proteins interact in epigenetic programming during chondrogenesis.

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    Frank Spaapen

    Full Text Available Initiation of and progression through chondrogenesis is driven by changes in the cellular microenvironment. At the onset of chondrogenesis, resting mesenchymal stem cells are mobilized in vivo and a complex, step-wise chondrogenic differentiation program is initiated. Differentiation requires coordinated transcriptomic reprogramming and increased progenitor proliferation; both processes require chromatin remodeling. The nature of early molecular responses that relay differentiation signals to chromatin is poorly understood. We here show that immediate early genes are rapidly and transiently induced in response to differentiation stimuli in vitro. Functional ablation of the immediate early factor EGR1 severely deregulates expression of key chondrogenic control genes at the onset of differentiation. In addition, differentiating cells accumulate DNA damage, activate a DNA damage response and undergo a cell cycle arrest and prevent differentiation associated hyper-proliferation. Failed differentiation in the absence of EGR1 affects global acetylation and terminates in overall histone hypermethylation. We report novel molecular connections between EGR1 and Polycomb Group function: Polycomb associated histone H3 lysine27 trimethylation (H3K27me3 blocks chromatin access of EGR1. In addition, EGR1 ablation results in abnormal Ezh2 and Bmi1 expression. Consistent with this functional interaction, we identify a number of co-regulated targets genes in a chondrogenic gene network. We here describe an important role for EGR1 in early chondrogenic epigenetic programming to accommodate early gene-environment interactions in chondrogenesis.

  16. Epstein-Barr virus immediate-early gene product trans-activates gene expression from the human immunodeficiency virus long terminal repeat

    International Nuclear Information System (INIS)

    Kenney, S.; Kamine, J.; Markovitz, D.; Fenrick, R.; Pagano, J.

    1988-01-01

    Acquired immunodeficiency syndrome patients are frequently coinfected with Epstein-Barr virus (EBV). In this report, the authors demonstrate that an EBV immediate-early gene product, BamHI MLF1, stimulates expression of the bacterial chloramphenicol acetyltransferase (CAT) gene linked to the human immunodeficiency virus (HIV) promoter. The HIV promoter sequences necessary for trans-activation by EBV do not include the tat-responsive sequences. In addition, in contrast to the other herpesvirus trans-activators previously studied, the EBV BamHI MLF1 gene product appears to function in part by a posttranscriptional mechanism, since it increases pHIV-CAT protein activity more than it increases HIV-CAT mRNA. This ability of an EBV gene product to activate HIV gene expression may have biologic consequences in persons coinfected with both viruses

  17. Gene Expression Patterns during the Early Stages of Chemically Induced Larval Metamorphosis and Settlement of the Coral Acropora millepora

    Science.gov (United States)

    Siboni, Nachshon; Abrego, David; Motti, Cherie A.; Tebben, Jan; Harder, Tilmann

    2014-01-01

    The morphogenetic transition of motile coral larvae into sessile primary polyps is triggered and genetically programmed upon exposure to environmental biomaterials, such as crustose coralline algae (CCA) and bacterial biofilms. Although the specific chemical cues that trigger coral larval morphogenesis are poorly understood there is much more information available on the genes that play a role in this early life phase. Putative chemical cues from natural biomaterials yielded defined chemical samples that triggered different morphogenetic outcomes: an extract derived from a CCA-associated Pseudoalteromonas bacterium that induced metamorphosis, characterized by non-attached metamorphosed juveniles; and two fractions of the CCA Hydrolithon onkodes (Heydrich) that induced settlement, characterized by attached metamorphosed juveniles. In an effort to distinguish the genes involved in these two morphogenetic transitions, competent larvae of the coral Acropora millepora were exposed to these predictable cues and the expression profiles of 47 coral genes of interest (GOI) were investigated after only 1 hour of exposure using multiplex RT–qPCR. Thirty-two GOI were differentially expressed, indicating a putative role during the early regulation of morphogenesis. The most striking differences were observed for immunity-related genes, hypothesized to be involved in cell recognition and adhesion, and for fluorescent protein genes. Principal component analysis of gene expression profiles resulted in separation between the different morphogenetic cues and exposure times, and not only identified those genes involved in the early response but also those which influenced downstream biological changes leading to larval metamorphosis or settlement. PMID:24632854

  18. Spatio-Temporal Gene Expression Profiling during In Vivo Early Ovarian Folliculogenesis: Integrated Transcriptomic Study and Molecular Signature of Early Follicular Growth.

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    Agnes Bonnet

    Full Text Available The successful achievement of early ovarian folliculogenesis is important for fertility and reproductive life span. This complex biological process requires the appropriate expression of numerous genes at each developmental stage, in each follicular compartment. Relatively little is known at present about the molecular mechanisms that drive this process, and most gene expression studies have been performed in rodents and without considering the different follicular compartments.We used RNA-seq technology to explore the sheep transcriptome during early ovarian follicular development in the two main compartments: oocytes and granulosa cells. We documented the differential expression of 3,015 genes during this phase and described the gene expression dynamic specific to these compartments. We showed that important steps occurred during primary/secondary transition in sheep. We also described the in vivo molecular course of a number of pathways. In oocytes, these pathways documented the chronology of the acquisition of meiotic competence, migration and cellular organization, while in granulosa cells they concerned adhesion, the formation of cytoplasmic projections and steroid synthesis. This study proposes the involvement in this process of several members of the integrin and BMP families. The expression of genes such as Kruppel-like factor 9 (KLF9 and BMP binding endothelial regulator (BMPER was highlighted for the first time during early follicular development, and their proteins were also predicted to be involved in gene regulation. Finally, we selected a data set of 24 biomarkers that enabled the discrimination of early follicular stages and thus offer a molecular signature of early follicular growth. This set of biomarkers includes known genes such as SPO11 meiotic protein covalently bound to DSB (SPO11, bone morphogenetic protein 15 (BMP15 and WEE1 homolog 2 (S. pombe(WEE2 which play critical roles in follicular development but other biomarkers

  19. Changes in nucleosome repeat lengths precede replication in the early replicating metallothionein II gene region of cells synchronized in early S phase

    International Nuclear Information System (INIS)

    D'Anna, J.A.; Tobey, R.A.

    1989-01-01

    Previous investigations showed that inhibition of DNA synthesis by hydroxyurea, aphidicolin, or 5-fluorodeoxyuridine produced large changes in the composition and nucleosome repeat lengths of bulk chromatin. There the authors report results of investigations to determine whether the changes in nucleosome repeat lengths might be localized in the initiated replicons, as postulated. In most experiments, Chinese hamster (line CHO) cells were synchronized in G1, or they were synchronized in early S phase by allowing G1 cells to enter S phase in medium containing 1 mM hydroxyurea or 5 μg mL -1 aphidicolin, a procedure believed to produce an accumulation of initiated replicons that arise from normally early replicating DNA. Measurements of nucleosome repeat lengths of bulk chromatin, the early replicating unexpressed metallothionein II (MTII) gene region, and a later replicating repeated sequence indicate that the changes in repeat lengths occur preferentially in the early replicating MTII gene region as G1 cells enter and become synchronized in early S phase. During that time, the MTII gene region is not replicated nor is there any evidence for induction of MTII messenger RNA. Thus, the results are consistent with the hypothesis that changes in chromatin structure occur preferentially in the early replicating (presumably initiated) replicons at initiation or that changes in chromatin structure can precede replication during inhibition of DNA synthesis. The shortened repeat lengths that precede MTII replication are, potentially, reversible, because they become elongated when the synchronized early S-phase cells are released to resume cell cycle progression

  20. Mapping of brain activity by automated volume analysis of immediate early genes

    Science.gov (United States)

    Renier, Nicolas; Adams, Eliza L.; Kirst, Christoph; Wu, Zhuhao; Azevedo, Ricardo; Kohl, Johannes; Autry, Anita E.; Kadiri, Lolahon; Venkataraju, Kannan Umadevi; Zhou, Yu; Wang, Victoria X.; Tang, Cheuk Y.; Olsen, Olav; Dulac, Catherine; Osten, Pavel; Tessier-Lavigne, Marc

    2016-01-01

    Summary Understanding how neural information is processed in physiological and pathological states would benefit from precise detection, localization and quantification of the activity of all neurons across the entire brain, which has not to date been achieved in the mammalian brain. We introduce a pipeline for high speed acquisition of brain activity at cellular resolution through profiling immediate early gene expression using immunostaining and light-sheet fluorescence imaging, followed by automated mapping and analysis of activity by an open-source software program we term ClearMap. We validate the pipeline first by analysis of brain regions activated in response to Haloperidol. Next, we report new cortical regions downstream of whisker-evoked sensory processing during active exploration. Lastly, we combine activity mapping with axon tracing to uncover new brain regions differentially activated during parenting behavior. This pipeline is widely applicable to different experimental paradigms, including animal species for which transgenic activity reporters are not readily available. PMID:27238021

  1. REST mediates androgen receptor actions on gene repression and predicts early recurrence of prostate cancer

    DEFF Research Database (Denmark)

    Svensson, Charlotte; Ceder, Jens; Iglesias Gato, Diego

    2014-01-01

    The androgen receptor (AR) is a key regulator of prostate tumorgenesis through actions that are not fully understood. We identified the repressor element (RE)-1 silencing transcription factor (REST) as a mediator of AR actions on gene repression. Chromatin immunoprecipitation showed that AR binds...... in cell cycle progression, including Aurora Kinase A, that has previously been implicated in the growth of NE-like castration-resistant tumors. The analysis of prostate cancer tissue microarrays revealed that tumors with reduced expression of REST have higher probability of early recurrence, independently...... of their Gleason score. The demonstration that REST modulates AR actions in prostate epithelia and that REST expression is negatively correlated with disease recurrence after prostatectomy, invite a deeper characterization of its role in prostate carcinogenesis....

  2. Early and long-standing rheumatoid arthritis: distinct molecular signatures identified by gene-expression profiling in synovia

    Science.gov (United States)

    Lequerré, Thierry; Bansard, Carine; Vittecoq, Olivier; Derambure, Céline; Hiron, Martine; Daveau, Maryvonne; Tron, François; Ayral, Xavier; Biga, Norman; Auquit-Auckbur, Isabelle; Chiocchia, Gilles; Le Loët, Xavier; Salier, Jean-Philippe

    2009-01-01

    Introduction Rheumatoid arthritis (RA) is a heterogeneous disease and its underlying molecular mechanisms are still poorly understood. Because previous microarray studies have only focused on long-standing (LS) RA compared to osteoarthritis, we aimed to compare the molecular profiles of early and LS RA versus control synovia. Methods Synovial biopsies were obtained by arthroscopy from 15 patients (4 early untreated RA, 4 treated LS RA and 7 controls, who had traumatic or mechanical lesions). Extracted mRNAs were used for large-scale gene-expression profiling. The different gene-expression combinations identified by comparison of profiles of early, LS RA and healthy synovia were linked to the biological processes involved in each situation. Results Three combinations of 719, 116 and 52 transcripts discriminated, respectively, early from LS RA, and early or LS RA from healthy synovia. We identified several gene clusters and distinct molecular signatures specifically expressed during early or LS RA, thereby suggesting the involvement of different pathophysiological mechanisms during the course of RA. Conclusions Early and LS RA have distinct molecular signatures with different biological processes participating at different times during the course of the disease. These results suggest that better knowledge of the main biological processes involved at a given RA stage might help to choose the most appropriate treatment. PMID:19563633

  3. Inflammatory Gene Expression in Whole Peripheral Blood at Early Stages of Sporadic Amyotrophic Lateral Sclerosis

    Directory of Open Access Journals (Sweden)

    Pol Andrés-Benito

    2017-10-01

    Full Text Available ObjectiveCharacterization of altered expression of selected transcripts linked to inflammation in the peripheral blood of sporadic amyotrophic lateral sclerosis (sALS patients at early stage of disease to increase knowledge about peripheral inflammatory response in sALS.MethodsRNA expression levels of 45 genes were assessed by RT-qPCR in 22 sALS cases in parallel with 13 age-matched controls. Clinical and serum parameters were assessed at the same time.ResultsUpregulation of genes coding for factors involved in leukocyte extravasation (ITGB2, INPP5D, SELL, and ICAM1 and extracellular matrix remodeling (MMP9 and TIMP2, as well as downregulation of certain chemokines (CCL5 and CXC5R, anti-inflammatory cytokines (IL10, TGFB2, and IL10RA, pro-inflammatory cytokines (IL-6, and T-cell regulators (CD2 and TRBC1 was found in sALS cases independently of gender, clinical symptoms at onset (spinal, respiratory, or bulbar, progression, peripheral leukocyte number, and integrity of RNA. MMP9 levels positively correlated with age, whereas CCR5, CCL5, and TRBC1 negatively correlated with age in sALS but not in controls. Relatively higher TNFA expression levels correlate with higher creatinine kinase protein levels in plasma.ConclusionPresent findings show early inflammatory responses characterized by upregulation of factors enabling extravasation of leukocytes and extracellular matrix remodeling in blood in sALS cases, in addition to increased TNFA levels paralleling skeletal muscle damage.

  4. Ribosomal genes as early targets of cadmium-induced toxicity in Chironomus riparius larvae

    International Nuclear Information System (INIS)

    Planello, R.; Martinez-Guitarte, J.L.; Morcillo, G.

    2007-01-01

    Cadmium is a widespread environmental pollutant that causes severe impacts in organisms. Although the effects of cadmium on aquatic insects have been studied in terms of their toxicity and changes in developmental parameters, little is known about its molecular and genetic effects. We have investigated the alterations in the pattern of gene expression provoked by acute exposure to cadmium in Chironomus riparius Mg. (Diptera, Chironomidae), a sentinel organism widely used in aquatic toxicity testing. The early cytotoxic effects were evaluated using immunocytochemistry and specific fluorescent probes in fourth instar larvae after 12 h of 10 mM cadmium treatments; under these conditions no significant effect on larvae mortality was detected until after 36 h of exposure. The changes in the pattern of gene expression were analysed by means of DNA/RNA hybrid antibodies in the polytene chromosomes from salivary gland cells. A decrease in the activity of the nucleolus is especially remarkable, accompanied by a significant reduction in size and the modification in nucleolar architecture, as shown by FISH. The inhibition of rDNA transcription was further evaluated by Northern blot analysis, which showed a marked decrease in the level of preribosomal rRNA (54% 45S 12 h). However, the BR genes, whose products are the giant polypeptides that constitute the silk-like secretion for constructing housing tubes, remain active. Simultaneously, decondensation and activation take place at some chromosomal regions, especially at the centromeres. The changes observed in the pattern of gene expression do not resemble those found after heat shock or other cell stressors. These data provide the first evidence that cadmium interacts with ribosomal genes and results in a drastic impairment of the functional activity of the nucleolus, an essential organelle for cellular survival. Therefore, the depletion of ribosomes would be a long-term effect of Cd-induced cellular damage. These findings may

  5. Ribosomal genes as early targets of cadmium-induced toxicity in Chironomus riparius larvae

    Energy Technology Data Exchange (ETDEWEB)

    Planello, R. [Biologia Ambiental, Facultad de Ciencias, Universidad Nacional de Educacion a Distancia, Senda del Rey 9, 28040, Madrid (Spain); Martinez-Guitarte, J.L. [Biologia Ambiental, Facultad de Ciencias, Universidad Nacional de Educacion a Distancia, Senda del Rey 9, 28040, Madrid (Spain); Morcillo, G. [Biologia Ambiental, Facultad de Ciencias, Universidad Nacional de Educacion a Distancia, Senda del Rey 9, 28040, Madrid (Spain)]. E-mail: gmorcillo@ccia.uned.es

    2007-02-01

    Cadmium is a widespread environmental pollutant that causes severe impacts in organisms. Although the effects of cadmium on aquatic insects have been studied in terms of their toxicity and changes in developmental parameters, little is known about its molecular and genetic effects. We have investigated the alterations in the pattern of gene expression provoked by acute exposure to cadmium in Chironomus riparius Mg. (Diptera, Chironomidae), a sentinel organism widely used in aquatic toxicity testing. The early cytotoxic effects were evaluated using immunocytochemistry and specific fluorescent probes in fourth instar larvae after 12 h of 10 mM cadmium treatments; under these conditions no significant effect on larvae mortality was detected until after 36 h of exposure. The changes in the pattern of gene expression were analysed by means of DNA/RNA hybrid antibodies in the polytene chromosomes from salivary gland cells. A decrease in the activity of the nucleolus is especially remarkable, accompanied by a significant reduction in size and the modification in nucleolar architecture, as shown by FISH. The inhibition of rDNA transcription was further evaluated by Northern blot analysis, which showed a marked decrease in the level of preribosomal rRNA (54% 45S 12 h). However, the BR genes, whose products are the giant polypeptides that constitute the silk-like secretion for constructing housing tubes, remain active. Simultaneously, decondensation and activation take place at some chromosomal regions, especially at the centromeres. The changes observed in the pattern of gene expression do not resemble those found after heat shock or other cell stressors. These data provide the first evidence that cadmium interacts with ribosomal genes and results in a drastic impairment of the functional activity of the nucleolus, an essential organelle for cellular survival. Therefore, the depletion of ribosomes would be a long-term effect of Cd-induced cellular damage. These findings may

  6. Hydroxymethylation at Gene Regulatory Regions Directs Stem/Early Progenitor Cell Commitment during Erythropoiesis

    Directory of Open Access Journals (Sweden)

    Jozef Madzo

    2014-01-01

    Full Text Available Hematopoietic stem cell differentiation involves the silencing of self-renewal genes and induction of a specific transcriptional program. Identification of multiple covalent cytosine modifications raises the question of how these derivatized bases influence stem cell commitment. Using a replicative primary human hematopoietic stem/progenitor cell differentiation system, we demonstrate dynamic changes of 5-hydroxymethylcytosine (5-hmC during stem cell commitment and differentiation to the erythroid lineage. Genomic loci that maintain or gain 5-hmC density throughout erythroid differentiation contain binding sites for erythroid transcription factors and several factors not previously recognized as erythroid-specific factors. The functional importance of 5-hmC was demonstrated by impaired erythroid differentiation, with augmentation of myeloid potential, and disrupted 5-hmC patterning in leukemia patient-derived CD34+ stem/early progenitor cells with TET methylcytosine dioxygenase 2 (TET2 mutations. Thus, chemical conjugation and affinity purification of 5-hmC-enriched sequences followed by sequencing serve as resources for deciphering functional implications for gene expression during stem cell commitment and differentiation along a particular lineage.

  7. Genetic evidence for the association between the early growth response 3 (EGR3 gene and schizophrenia.

    Directory of Open Access Journals (Sweden)

    Rui Zhang

    Full Text Available Recently, two genome scan meta-analysis studies have found strong evidence for the association of loci on chromosome 8p with schizophrenia. The early growth response 3 (EGR3 gene located in chromosome 8p21.3 was also found to be involved in the etiology of schizophrenia. However, subsequent studies failed to replicate this finding. To investigate the genetic role of EGR3 in Chinese patients, we genotyped four SNPs (average interval ∼2.3 kb in the chromosome region of EGR3 in 470 Chinese schizophrenia patients and 480 healthy control subjects. The SNP rs35201266 (located in intron 1 of EGR3 showed significant differences between cases and controls in both genotype frequency distribution (P = 0.016 and allele frequency distribution (P = 0.009. Analysis of the haplotype rs35201266-rs3750192 provided significant evidence for association with schizophrenia (P = 0.0012; a significant difference was found for the common haplotype AG (P = 0.0005. Furthermore, significant associations were also found in several other two-, and three-SNP tests of haplotype analyses. The meta-analysis revealed a statistically significant association between rs35201266 and schizophrenia (P = 0.0001. In summary, our study supports the association of EGR3 with schizophrenia in our Han Chinese sample, and further functional exploration of the EGR3 gene will contribute to the molecular basis for the complex network underlying schizophrenia pathogenesis.

  8. The Polynesian gene pool: an early contribution by Amerindians to Easter Island.

    Science.gov (United States)

    Thorsby, Erik

    2012-03-19

    It is now generally accepted that Polynesia was first settled by peoples from southeast Asia. An alternative that eastern parts of Polynesia were first inhabited by Amerindians has found little support. There are, however, many indications of a 'prehistoric' (i.e. before Polynesia was discovered by Europeans) contact between Polynesia and the Americas, but genetic evidence of a prehistoric Amerindian contribution to the Polynesian gene pool has been lacking. We recently carried out genomic HLA (human leucocyte antigen) typing as well as typing for mitochondrial DNA (mtDNA) and Y chromosome markers of blood samples collected in 1971 and 2008 from reputedly non-admixed Easter Islanders. All individuals carried HLA alleles and mtDNA types previously found in Polynesia, and most of the males carried Y chromosome markers of Polynesian origin (a few had European Y chromosome markers), further supporting an initial Polynesian population on Easter Island. The HLA investigations revealed, however, that some individuals also carried HLA alleles which have previously almost only been found in Amerindians. We could trace the introduction of these Amerindian alleles to before the Peruvian slave trades, i.e. before the 1860s, and provide suggestive evidence that they were introduced already in prehistoric time. Our results demonstrate an early Amerindian contribution to the Polynesian gene pool on Easter Island, and illustrate the usefulness of typing for immunogenetic markers such as HLA to complement mtDNA and Y chromosome analyses in anthropological investigations.

  9. Early-Onset Alzheimer Disease and Candidate Risk Genes Involved in Endolysosomal Transport.

    Science.gov (United States)

    Kunkle, Brian W; Vardarajan, Badri N; Naj, Adam C; Whitehead, Patrice L; Rolati, Sophie; Slifer, Susan; Carney, Regina M; Cuccaro, Michael L; Vance, Jeffery M; Gilbert, John R; Wang, Li-San; Farrer, Lindsay A; Reitz, Christiane; Haines, Jonathan L; Beecham, Gary W; Martin, Eden R; Schellenberg, Gerard D; Mayeux, Richard P; Pericak-Vance, Margaret A

    2017-09-01

    Mutations in APP, PSEN1, and PSEN2 lead to early-onset Alzheimer disease (EOAD) but account for only approximately 11% of EOAD overall, leaving most of the genetic risk for the most severe form of Alzheimer disease unexplained. This extreme phenotype likely harbors highly penetrant risk variants, making it primed for discovery of novel risk genes and pathways for AD. To search for rare variants contributing to the risk for EOAD. In this case-control study, whole-exome sequencing (WES) was performed in 51 non-Hispanic white (NHW) patients with EOAD (age at onset 65 years) from the Alzheimer's Disease Genetics Consortium. The study was conducted from January 21, 2013, to October 13, 2016. Alzheimer disease diagnosed according to standard National Institute of Neurological and Communicative Disorders and Stroke and the Alzheimer Disease and Related Disorders Association criteria. Association between Alzheimer disease and genetic variants and genes was measured using logistic regression and sequence kernel association test-optimal gene tests, respectively. Of the 1524 NHW patients with EOAD, 765 (50.2%) were women and mean (SD) age was 60.0 (4.9) years; of the 7046 NHW patients with LOAD, 4171 (59.2%) were women and mean (SD) age was 77.4 (8.6) years; and of the 7001 NHW controls, 4215 (60.2%) were women and mean (SD) age was 77.4 (8.6) years. The gene PSD2, for which multiple unrelated NHW cases had rare missense variants, was significantly associated with EOAD (P = 2.05 × 10-6; Bonferroni-corrected P value [BP] = 1.3 × 10-3) and LOAD (P = 6.22 × 10-6; BP = 4.1 × 10-3). A missense variant in TCIRG1, present in a NHW patient and segregating in 3 cases of a Hispanic family, was more frequent in EOAD cases (odds ratio [OR], 2.13; 95% CI, 0.99-4.55; P = .06; BP = 0.413), and significantly associated with LOAD (OR, 2.23; 95% CI, 1.37-3.62; P = 7.2 × 10-4; BP = 5.0 × 10-3). A missense variant in the LOAD risk

  10. Changes in protein synthetic activity in early Drosophila embryos mutant for the segmentation gene Krueppel

    International Nuclear Information System (INIS)

    Bedian, V.; Summers, M.C.; Kauffman, S.A.

    1988-01-01

    We have identified early embryo proteins related to the segmentation gene Krueppel by [35S]methionine pulse labelling and two-dimensional gel electrophoresis. Protein synthesis differences shared by homozygous embryos of two Krueppel alleles when compared to heterozygous and wild-type embryos are reported. The study was extended to syncytial blastoderm stages by pulse labelling and gel analysis of single embryos, using Krueppel-specific proteins from gastrula stages as molecular markers for identifying homozygous Krueppel embryos. Localized expression of interesting proteins was examined in embryo fragments. The earliest differences detected at nuclear migration stages showed unregulated synthesis in mutant embryos of two proteins that have stage specific synthesis in normal embryos. At the cellular blastoderm stage one protein was not synthesized and two proteins showed apparent shifts in isoelectric point in mutant embryos. Differences observed in older embryos included additional proteins with shifted isoelectric points and a number of qualitative and quantitative changes in protein synthesis. Five of the proteins with altered rates of synthesis in mutant embryos showed localized synthesis in normal embryos. The early effects observed are consistent with the hypothesis that the Krueppel product can be a negative or positive regulator of expression of other loci, while blastoderm and gastrula stage shifts in isoelectric point indicate that a secondary effect of Krueppel function may involve post-translational modification of proteins

  11. Post-transcriptional regulation of macrophage ABCA1, an early response gene to IFN-γ

    International Nuclear Information System (INIS)

    Alfaro Leon, Martha Leticia; Evans, Glenn F.; Farmen, Mark W.; Zuckerman, Steven H.

    2005-01-01

    Interferon-γ (IFN-γ) down-regulates receptors associated with reverse cholesterol transport including ABCA1. In the present study, the kinetics and mechanism of ABCA1 down-regulation were determined in mouse peritoneal macrophages. IFN-γ decreased ABCA1 mRNA 1 h following IFN-γ addition and was maximally reduced by 3 h. Down-regulation was protein synthesis dependent and involved post-transcriptional processes. ABCA1 message had a T 1/2 of 115 min in actinomycin treated cells that was reduced to a T 1/2 of 37 min by IFN-γ. The decrease in message stability was also associated with a rapid loss of ABCA1 protein, significant 3 h following IFN-γ addition. The kinetics of ABCA1 message and protein decrease was consistent with the early IFN-γ-induced changes in Stat1 phosphorylation and nuclear translocation observed in these cells. Therefore, ABCA1 can be considered as an early response gene to macrophage activation by IFN-γ with down-regulation occurring by message destabilization

  12. Beyond Neuronal Activity Markers: Select Immediate Early Genes in Striatal Neuron Subtypes Functionally Mediate Psychostimulant Addiction

    Directory of Open Access Journals (Sweden)

    Ramesh Chandra

    2017-06-01

    Full Text Available Immediate early genes (IEGs were traditionally used as markers of neuronal activity in striatum in response to stimuli including drugs of abuse such as psychostimulants. Early studies using these neuronal activity markers led to important insights in striatal neuron subtype responsiveness to psychostimulants. Such studies have helped identify striatum as a critical brain center for motivational, reinforcement and habitual behaviors in psychostimulant addiction. While the use of IEGs as neuronal activity markers in response to psychostimulants and other stimuli persists today, the functional role and implications of these IEGs has often been neglected. Nonetheless, there is a subset of research that investigates the functional role of IEGs in molecular, cellular and behavioral alterations by psychostimulants through striatal medium spiny neuron (MSN subtypes, the two projection neuron subtypes in striatum. This review article will address and highlight the studies that provide a functional mechanism by which IEGs mediate psychostimulant molecular, cellular and behavioral plasticity through MSN subtypes. Insight into the functional role of IEGs in striatal MSN subtypes could provide improved understanding into addiction and neuropsychiatric diseases affecting striatum, such as affective disorders and compulsive disorders characterized by dysfunctional motivation and habitual behavior.

  13. Early diagnosis of typhoid fever by nested PCR for flagellin gene of Salmonella enterica serotype Typhi.

    Science.gov (United States)

    Khan, S; Harish, B N; Menezes, G A; Acharya, N S; Parija, S C

    2012-11-01

    Typhoid fever caused by Salmonella Typhi continues to be a major health problem in spite of the use of antibiotics and the development of newer antibacterial drugs. Inability to make an early laboratory diagnosis and resort to empirical therapy, often lead to increased morbidity and mortality in cases of typhoid fever. This study was aimed to optimize a nested PCR for early diagnosis of typhoid fever and using it as a diagnostic tool in culture negative cases of suspected typhoid fever. Eighty patients with clinical diagnosis of typhoid fever and 40 controls were included in the study. The blood samples collected were subjected to culture, Widal and nested PCR targeting the flagellin gene of S. Typhi. The sensitivity of PCR on blood was found to be 100 per cent whereas the specificity was 76.9 per cent. The positive predictive value (PPV) of PCR was calculated to be 76.9 per cent with an accuracy of 86 per cent. None of the 40 control samples gave a positive PCR. Due to its high sensitivity and specificity nested PCR can be used as a useful tool to diagnose clinically suspected, culture negative cases of typhoid fever.

  14. Time-Series Interactions of Gene Expression, Vascular Growth and Hemodynamics during Early Embryonic Arterial Development.

    Directory of Open Access Journals (Sweden)

    Selda Goktas

    Full Text Available The role of hemodynamic forces within the embryo as biomechanical regulators for cardiovascular morphogenesis, growth, and remodeling is well supported through the experimental studies. Furthermore, clinical experience suggests that perturbed flow disrupts the normal vascular growth process as one etiology for congenital heart diseases (CHD and for fetal adaptation to CHD. However, the relationships between hemodynamics, gene expression and embryonic vascular growth are poorly defined due to the lack of concurrent, sequential in vivo data. In this study, a long-term, time-lapse optical coherence tomography (OCT imaging campaign was conducted to acquire simultaneous blood velocity, pulsatile micro-pressure and morphometric data for 3 consecutive early embryonic stages in the chick embryo. In conjunction with the in vivo growth and hemodynamics data, in vitro reverse transcription polymerase chain reaction (RT-PCR analysis was performed to track changes in transcript expression relevant to histogenesis and remodeling of the embryonic arterial wall. Our non-invasive extended OCT imaging technique for the microstructural data showed continuous vessel growth. OCT data coupled with the PIV technique revealed significant but intermitted increases in wall shear stress (WSS between first and second assigned stages and a noticeable decrease afterwards. Growth rate, however, did not vary significantly throughout the embryonic period. Among all the genes studied, only the MMP-2 and CASP-3 expression levels remained unchanged during the time course. Concurrent relationships were obtained among the transcriptional modulation of the genes, vascular growth and hemodynamics-related changes. Further studies are indicated to determine cause and effect relationships and reversibility between mechanical and molecular regulation of vasculogenesis.

  15. Specific interactions between transcription factors and the promoter-regulatory region of the human cytomegalovirus major immediate-early gene

    International Nuclear Information System (INIS)

    Ghazal, P.; Lubon, H.; Hennighausen, L.

    1988-01-01

    Repeat sequence motifs as well as unique sequences between nucleotides -150 and -22 of the human cytomegalovirus immediate-early 1 gene interact in vitro with nuclear proteins. The authors show that a transcriptional element between nucleotides -91 and -65 stimulated promoter activity in vivo and in vitro by binding specific cellular transcription factors. Finally, a common sequence motif, (T)TGG/AC, present in 15 of the determined binding sites suggests a particular class of nuclear factors associated with the immediate-early 1 gene

  16. Selecting postoperative adjuvant systemic therapy for early stage breast cancer: A critical assessment of commercially available gene expression assays

    Science.gov (United States)

    Schuur, Eric; Angel Aristizabal, Javier; Bargallo Rocha, Juan Enrique; Cabello, Cesar; Elizalde, Roberto; García‐Estévez, Laura; Gomez, Henry L.; Katz, Artur; Nuñez De Pierro, Aníbal

    2017-01-01

    Risk stratification of patients with early stage breast cancer may support adjuvant chemotherapy decision‐making. This review details the development and validation of six multi‐gene classifiers, each of which claims to provide useful prognostic and possibly predictive information for early stage breast cancer patients. A careful assessment is presented of each test's analytical validity, clinical validity, and clinical utility, as well as the quality of evidence supporting its use. PMID:28211064

  17. Animal models for development of therapeutic HPV16 vaccines

    Czech Academy of Sciences Publication Activity Database

    Bubeník, Jan

    2002-01-01

    Roč. 20, č. 1 (2002), s. 207-212 ISSN 1019-6439 Institutional research plan: CEZ:AV0Z5052915 Keywords : human papilloma viruses * tumour vaccines Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.931, year: 2002

  18. The HPV16 E5 oncogene inhibits endocytic trafficking

    DEFF Research Database (Denmark)

    Thomsen, P; van Deurs, B; Norrild, B

    2000-01-01

    fibroblasts to study the effects of E5. Various endocytic markers including the pH-sensitive probe DM-NERF coupled to dextran, TransFluoSpheres and TRITC-concanavalin A, were applied. In E5-transfected cells, none of these markers colocalized with the membrane permeable probe LysoTracker Red, which...

  19. Dominant Role of HPV16 E7 in Anal Carcinogenesis

    Science.gov (United States)

    Thomas, Marie K.; Pitot, Henry C.; Liem, Amy; Lambert, Paul F.

    2011-01-01

    Ninety percent of anal cancer is associated with human papilloma viruses (HPVs). Using our previously established HPV transgenic mouse model for anal cancer, we tested the role of the individual oncogenes E6 and E7. K14E6 and K14E7 transgenic mice were treated with dimethylbenz[a]anthracene (DMBA) to the anal canal and compared to matched nontransgenic and doubly transgenic K14E6/E7 mice. K14E7 and K14E6/E7 transgenic mice developed anal tumors (papillomas, atypias and carcinomas combined) at significantly higher rates (88% and 100%, respectively) than either K14E6 or NTG mice (18% and 19%, respectively). Likewise, K14E7 and K14E6/E7 transgenic mice developed frank cancer (carcinomas) at significantly higher rates (85% and 85%, respectively) than either K14E6 or NTG mice (18% and 10%, respectively). These findings indicate that E7 is the more potent oncogene in anal cancer caused by HPVs. PMID:21999991

  20. High prevalence of oncogenic HPV-16 in cervical smears of ...

    Indian Academy of Sciences (India)

    2012-01-27

    Jan 27, 2012 ... demographic, e.g., lower age of conception, high parity, .... and purified using Gel extraction kit (Fermentas, USA) .... (11.9%) and women belonging to lower income group ..... Epidemiologic classification of human papillo-.

  1. EXPRESSION OF HPV 16 AND 18 IN CERVICAL INTRAEPITHELIAL NEOPLASIA

    OpenAIRE

    Kodali Venkataramana; Prasad Usha

    2017-01-01

    BACKGROUND Cervical cancer is by far the most common human papilloma virus related disease. Nearly, all cases of cervical cancer can be attributable to human papilloma virus infection. Infection with the human papilloma virus is the main risk factors for cervical intraepithelial neoplasia and cervical cancer especially the high-risk types. The aim of the study is to study the prevalence of high-risk human papilloma virus 16 and 18 in various grades of cervical intraepithelia...

  2. Identification of early zygotic genes in the yellow fever mosquito Aedes aegypti and discovery of a motif involved in early zygotic genome activation.

    Science.gov (United States)

    Biedler, James K; Hu, Wanqi; Tae, Hongseok; Tu, Zhijian

    2012-01-01

    During early embryogenesis the zygotic genome is transcriptionally silent and all mRNAs present are of maternal origin. The maternal-zygotic transition marks the time over which embryogenesis changes its dependence from maternal RNAs to zygotically transcribed RNAs. Here we present the first systematic investigation of early zygotic genes (EZGs) in a mosquito species and focus on genes involved in the onset of transcription during 2-4 hr. We used transcriptome sequencing to identify the "pure" (without maternal expression) EZGs by analyzing transcripts from four embryonic time ranges of 0-2, 2-4, 4-8, and 8-12 hr, which includes the time of cellular blastoderm formation and up to the start of gastrulation. Blast of 16,789 annotated transcripts vs. the transcriptome reads revealed evidence for 63 (P<0.001) and 143 (P<0.05) nonmaternally derived transcripts having a significant increase in expression at 2-4 hr. One third of the 63 EZG transcripts do not have predicted introns compared to 10% of all Ae. aegypti genes. We have confirmed by RT-PCR that zygotic transcription starts as early as 2-3 hours. A degenerate motif VBRGGTA was found to be overrepresented in the upstream sequences of the identified EZGs using a motif identification software called SCOPE. We find evidence for homology between this motif and the TAGteam motif found in Drosophila that has been implicated in EZG activation. A 38 bp sequence in the proximal upstream sequence of a kinesin light chain EZG (KLC2.1) contains two copies of the mosquito motif. This sequence was shown to support EZG transcription by luciferase reporter assays performed on injected early embryos, and confers early zygotic activity to a heterologous promoter from a divergent mosquito species. The results of these studies are consistent with the model of early zygotic genome activation via transcriptional activators, similar to what has been found recently in Drosophila.

  3. Identification of early zygotic genes in the yellow fever mosquito Aedes aegypti and discovery of a motif involved in early zygotic genome activation.

    Directory of Open Access Journals (Sweden)

    James K Biedler

    Full Text Available During early embryogenesis the zygotic genome is transcriptionally silent and all mRNAs present are of maternal origin. The maternal-zygotic transition marks the time over which embryogenesis changes its dependence from maternal RNAs to zygotically transcribed RNAs. Here we present the first systematic investigation of early zygotic genes (EZGs in a mosquito species and focus on genes involved in the onset of transcription during 2-4 hr. We used transcriptome sequencing to identify the "pure" (without maternal expression EZGs by analyzing transcripts from four embryonic time ranges of 0-2, 2-4, 4-8, and 8-12 hr, which includes the time of cellular blastoderm formation and up to the start of gastrulation. Blast of 16,789 annotated transcripts vs. the transcriptome reads revealed evidence for 63 (P<0.001 and 143 (P<0.05 nonmaternally derived transcripts having a significant increase in expression at 2-4 hr. One third of the 63 EZG transcripts do not have predicted introns compared to 10% of all Ae. aegypti genes. We have confirmed by RT-PCR that zygotic transcription starts as early as 2-3 hours. A degenerate motif VBRGGTA was found to be overrepresented in the upstream sequences of the identified EZGs using a motif identification software called SCOPE. We find evidence for homology between this motif and the TAGteam motif found in Drosophila that has been implicated in EZG activation. A 38 bp sequence in the proximal upstream sequence of a kinesin light chain EZG (KLC2.1 contains two copies of the mosquito motif. This sequence was shown to support EZG transcription by luciferase reporter assays performed on injected early embryos, and confers early zygotic activity to a heterologous promoter from a divergent mosquito species. The results of these studies are consistent with the model of early zygotic genome activation via transcriptional activators, similar to what has been found recently in Drosophila.

  4. Prediction of lymphatic metastasis based on gene expression profile analysis after brachytherapy for early-stage oral tongue carcinoma

    International Nuclear Information System (INIS)

    Watanabe, Hiroshi; Mogushi, Kaoru; Miura, Masahiko; Yoshimura, Ryo-ichi; Kurabayashi, Tohru; Shibuya, Hitoshi; Tanaka, Hiroshi; Noda, Shuhei; Iwakawa, Mayumi; Imai, Takashi

    2008-01-01

    Background and purpose: The management of lymphatic metastasis of early-stage oral tongue carcinoma patients is crucial for its prognosis. The purpose of this study was to evaluate the predictive ability of lymphatic metastasis after brachytherapy (BRT) for early-stage tongue carcinoma based on gene expression profiling. Patients and methods: Pre-therapeutic biopsies from 39 patients with T1 or T2 tongue cancer were analyzed for gene expression signatures using Codelink Uniset Human 20K Bioarray. All patients were treated with low dose-rate BRT for their primary lesions and underwent strict follow-up under a wait-and-see policy for cervical lymphatic metastasis. Candidate genes were selected for predicting lymph-node status in the reference group by the permutation test. Predictive accuracy was further evaluated by the prediction strength (PS) scoring system using an independent validation group. Results: We selected a set of 19 genes whose expression differed significantly between classes with or without lymphatic metastasis in the reference group. The lymph-node status in the validation group was predicted by the PS scoring system with an accuracy of 76%. Conclusions: Gene expression profiling using 19 genes in primary tumor tissues may allow prediction of lymphatic metastasis after BRT for early-stage oral tongue carcinoma

  5. Domestication-driven Gossypium profilin 1 (GhPRF1) gene transduces early flowering phenotype in tobacco by spatial alteration of apical/floral-meristem related gene expression.

    Science.gov (United States)

    Pandey, Dhananjay K; Chaudhary, Bhupendra

    2016-05-13

    Plant profilin genes encode core cell-wall structural proteins and are evidenced for their up-regulation under cotton domestication. Notwithstanding striking discoveries in the genetics of cell-wall organization in plants, little is explicit about the manner in which profilin-mediated molecular interplay and corresponding networks are altered, especially during cellular signalling of apical meristem determinacy and flower development. Here we show that the ectopic expression of GhPRF1 gene in tobacco resulted in the hyperactivation of apical meristem and early flowering phenotype with increased flower number in comparison to the control plants. Spatial expression alteration in CLV1, a key meristem-determinacy gene, is induced by the GhPRF1 overexpression in a WUS-dependent manner and mediates cell signalling to promote flowering. But no such expression alterations are recorded in the GhPRF1-RNAi lines. The GhPRF1 transduces key positive flowering regulator AP1 gene via coordinated expression of FT4, SOC1, FLC1 and FT1 genes involved in the apical-to-floral meristem signalling cascade which is consistent with our in silico profilin interaction data. Remarkably, these positive and negative flowering regulators are spatially controlled by the Actin-Related Protein (ARP) genes, specifically ARP4 and ARP6 in proximate association with profilins. This study provides a novel and systematic link between GhPRF1 gene expression and the flower primordium initiation via up-regulation of the ARP genes, and an insight into the functional characterization of GhPRF1 gene acting upstream to the flowering mechanism. Also, the transgenic plants expressing GhPRF1 gene show an increase in the plant height, internode length, leaf size and plant vigor. Overexpression of GhPRF1 gene induced early and increased flowering in tobacco with enhanced plant vigor. During apical meristem determinacy and flower development, the GhPRF1 gene directly influences key flowering regulators through ARP-genes

  6. Unstable Expression of Commonly Used Reference Genes in Rat Pancreatic Islets Early after Isolation Affects Results of Gene Expression Studies.

    Directory of Open Access Journals (Sweden)

    Lucie Kosinová

    Full Text Available The use of RT-qPCR provides a powerful tool for gene expression studies; however, the proper interpretation of the obtained data is crucially dependent on accurate normalization based on stable reference genes. Recently, strong evidence has been shown indicating that the expression of many commonly used reference genes may vary significantly due to diverse experimental conditions. The isolation of pancreatic islets is a complicated procedure which creates severe mechanical and metabolic stress leading possibly to cellular damage and alteration of gene expression. Despite of this, freshly isolated islets frequently serve as a control in various gene expression and intervention studies. The aim of our study was to determine expression of 16 candidate reference genes and one gene of interest (F3 in isolated rat pancreatic islets during short-term cultivation in order to find a suitable endogenous control for gene expression studies. We compared the expression stability of the most commonly used reference genes and evaluated the reliability of relative and absolute quantification using RT-qPCR during 0-120 hrs after isolation. In freshly isolated islets, the expression of all tested genes was markedly depressed and it increased several times throughout the first 48 hrs of cultivation. We observed significant variability among samples at 0 and 24 hrs but substantial stabilization from 48 hrs onwards. During the first 48 hrs, relative quantification failed to reflect the real changes in respective mRNA concentrations while in the interval 48-120 hrs, the relative expression generally paralleled the results determined by absolute quantification. Thus, our data call into question the suitability of relative quantification for gene expression analysis in pancreatic islets during the first 48 hrs of cultivation, as the results may be significantly affected by unstable expression of reference genes. However, this method could provide reliable information

  7. Early gene expression profiles of patients with chronic hepatitis C treated with pegylated interferon-alfa and ribavirin.

    Science.gov (United States)

    Younossi, Zobair M; Baranova, Ancha; Afendy, Arian; Collantes, Rochelle; Stepanova, Maria; Manyam, Ganiraju; Bakshi, Anita; Sigua, Christopher L; Chan, Joanne P; Iverson, Ayuko A; Santini, Christopher D; Chang, Sheng-Yung P

    2009-03-01

    Responsiveness to hepatitis C virus (HCV) therapy depends on viral and host factors. Our aim was to assess sustained virologic response (SVR)-associated early gene expression in patients with HCV receiving pegylated interferon-alpha2a (PEG-IFN-alpha2a) or PEG-IFN-alpha2b and ribavirin with the duration based on genotypes. Blood samples were collected into PAXgene tubes prior to treatment as well as 1, 7, 28, and 56 days after treatment. From the peripheral blood cells, total RNA was extracted, quantified, and used for one-step reverse transcription polymerase chain reaction to profile 154 messenger RNAs. Expression levels of messenger RNAs were normalized with six "housekeeping" genes and a reference RNA. Multiple regression and stepwise selection were performed to assess differences in gene expression at different time points, and predictive performance was evaluated for each model. A total of 68 patients were enrolled in the study and treated with combination therapy. The results of gene expression showed that SVR could be predicted by the gene expression of signal transducer and activator of transcription-6 (STAT-6) and suppressor of cytokine signaling-1 in the pretreatment samples. After 24 hours, SVR was predicted by the expression of interferon-dependent genes, and this dependence continued to be prominent throughout the treatment. Early gene expression during anti-HCV therapy may elucidate important molecular pathways that may be influencing the probability of achieving virologic response.

  8. Transcriptome profiling and digital gene expression by deep sequencing in early somatic embryogenesis of endangered medicinal Eleutherococcus senticosus Maxim.

    Science.gov (United States)

    Tao, Lei; Zhao, Yue; Wu, Ying; Wang, Qiuyu; Yuan, Hongmei; Zhao, Lijuan; Guo, Wendong; You, Xiangling

    2016-03-01

    Somatic embryogenesis (SE) has been studied as a model system to understand molecular events in physiology, biochemistry, and cytology during plant embryo development. In particular, it is exceedingly difficult to access the morphological and early regulatory events in zygotic embryos. To understand the molecular mechanisms regulating early SE in Eleutherococcus senticosus Maxim., we used high-throughput RNA-Seq technology to investigate its transcriptome. We obtained 58,327,688 reads, which were assembled into 75,803 unique unigenes. To better understand their functions, the unigenes were annotated using the Clusters of Orthologous Groups, Gene Ontology, and Kyoto Encyclopedia of Genes and Genomes databases. Digital gene expression libraries revealed differences in gene expression profiles at different developmental stages (embryogenic callus, yellow embryogenic callus, global embryo). We obtained a sequencing depth of >5.6 million tags per sample and identified many differentially expressed genes at various stages of SE. The initiation of SE affected gene expression in many KEGG pathways, but predominantly that in metabolic pathways, biosynthesis of secondary metabolites, and plant hormone signal transduction. This information on the changes in the multiple pathways related to SE induction in E. senticosus Maxim. embryogenic tissue will contribute to a more comprehensive understanding of the mechanisms involved in early SE. Additionally, the differentially expressed genes may act as molecular markers and could play very important roles in the early stage of SE. The results are a comprehensive molecular biology resource for investigating SE of E. senticosus Maxim. Copyright © 2015 Elsevier B.V. All rights reserved.

  9. Cpt1a gene expression in peripheral blood mononuclear cells as an early biomarker of diet-related metabolic alterations

    KAUST Repository

    Diaz-Rua, Ruben

    2016-11-23

    Background: Research on biomarkers that provide early information about the development of future metabolic alterations is an emerging discipline. Gene expression analysis in peripheral blood mononuclear cells (PBMC) is a promising tool to identify subjects at risk of developing diet-related diseases.

  10. Xp22.3 genomic deletions involving the CDKL5 gene in girls with early onset epileptic encephalopathy.

    Science.gov (United States)

    Mei, Davide; Marini, Carla; Novara, Francesca; Bernardina, Bernardo D; Granata, Tiziana; Fontana, Elena; Parrini, Elena; Ferrari, Anna R; Murgia, Alessandra; Zuffardi, Orsetta; Guerrini, Renzo

    2010-04-01

    Mutations of the X-linked gene cyclin-dependent kinase-like 5 (CDKL5) cause an X-linked encephalopathy with early onset intractable epilepsy, including infantile spasms and other seizure types, and a Rett syndrome (RTT)-like phenotype. Very limited information is available on the frequency and phenotypic spectrum associated with CDKL5 deletions/duplications. We investigated the role of CDKL5 deletions/duplications in causing early onset intractable epilepsy of unknown etiology in girls. We studied 49 girls with early onset intractable epilepsy, with or without infantile spasms, and developmental impairment, for whom no etiologic factors were obvious after clinical examination, brain magnetic resonance imaging (MRI) and expanded screening for inborn errors of metabolism. We performed CDKL5 gene mutation analysis in all and multiplex ligation dependent probe amplification assay (MLPA) in those who were mutation negative. Custom Array-comparative genomic hybridization (CGH), breakpoint polymerase chain reaction (PCR) analysis, and X-inactivation studies were performed in patients in whom MLPA uncovered a genomic alteration. We found CDKL5 mutations in 8.2% (4 of 49) of patients and genomic deletions in 8.2% (4 of 49). Overall, abnormalities of the CDKL5 gene accounted for 16.3% (8 of 49) of patients. CDKL5 gene deletions are an under-ascertained cause of early onset intractable epilepsy in girls. Genetic testing of CDKL5, including both mutation and deletion/duplication analysis, should be considered in this clinical subgroup.

  11. Early-Onset Severe Encephalopathy with Epilepsy: The BRAT1 Gene Should Be Added to the List of Causes

    NARCIS (Netherlands)

    van de Pol, L.A.; Wolf, N.I.; van Weissenbruch, M.M.; Stam, C.J.; Weiss, M.M.; Waisfisz, Q.; Kevelam, S.H.; Bugiani, M.; van de Kamp, J.M.; Knaap, M.

    2015-01-01

    A variety of pathologies can underlie early-onset severe encephalopathy with epilepsy. To aid the diagnostic process in such patients we present an overview of causes, including the rapidly expanding list of genes involved. When no explanation is found, whole-exome sequencing (WES) can be used in an

  12. Immediate-early gene region of human cytomegalovirus trans-activates the promoter of human immunodeficiency virus

    International Nuclear Information System (INIS)

    Davis, M.G.; Kenney, S.C.; Kamine, J.; Pagano, J.S.; Huang, E.S.

    1987-01-01

    Almost all homosexual patients with acquired immunodeficiency syndrome are also actively infected with human cytomegalovirus (HCMV). The authors have hypothesized that an interaction between HCMV and human immunodeficiency virus (HIV), the agent that causes acquired immunodeficiency syndrome, may exist at a molecular level and contribute to the manifestations of HIV infection. In this report, they demonstrate that the immediate-early gene region of HCMV, in particular immediate-early region 2, trans-activates the expression of the bacterial gene chloramphenicol acetyltransferase that is fused to the HIV long terminal repeat and carried by plasmid pHIV-CAT. The HCMV immediate-early trans-activator increases the level of mRNA from the plamid pHIV-CAT. The sequences of HIV that are responsive to trans-activation by the HDMV immediate-early region are distinct from HIV sequences that are required for response to the HIV tat. The stimulation of HIV gene expression by HDMV gene functions could enhance the consequences of HIV infection in persons with previous or concurrent HCMV infection

  13. Bm59 is an early gene, but is unessential for the propagation and assembly of Bombyx mori nucleopolyhedrovirus.

    Science.gov (United States)

    Hu, Xiaolong; Shen, Yunwang; Zheng, Qin; Wang, Guobao; Wu, Xiaofeng; Gong, Chengliang

    2016-02-01

    Bombyx mori nucleopolyhedrovirus (BmNPV) is a major pathogen that specifically infects the domestic silkworm and causes serious economic loss to sericulture around the world. The function of BmNPV Bm59 gene in the viral life cycle is inconclusive. To investigate the role of Bm59 during viral infection, the transcription initiation site and temporal expression of Bm59 were analyzed, and Bm59-knockout virus was generated through homologous recombination in Escherichia coli. The results showed that Bm59 is an early transcription gene with an atypia early transcriptional start motif. Budded virion (BV) production and DNA replication in the BmN cells transfected with the Bm59-knockout virus bacmid were similar to those in the cells transfected with the wild-type virus. Electron microscopy revealed that the occlusion-derived virus can be produced in cells infected with the Bm59-knockout virus. These results indicated that Bm59 is an early gene and is not essential for viral replication or assembly of BmNPV. These findings suggested that non-essential gene (Bm59) remained in the viral genome, which may interact with other viral/host genes in a certain situation.

  14. Early-Onset Severe Encephalopathy with Epilepsy: The BRAT1 Gene Should Be Added to the List of Causes.

    Science.gov (United States)

    van de Pol, Laura A; Wolf, Nicole I; van Weissenbruch, Mirjam M; Stam, Cornelie J; Weiss, Janneke M; Waisfisz, Quinten; Kevelam, Sietske H; Bugiani, Mariana; van de Kamp, Jiddeke M; van der Knaap, Marjo S

    2015-12-01

    A variety of pathologies can underlie early-onset severe encephalopathy with epilepsy. To aid the diagnostic process in such patients we present an overview of causes, including the rapidly expanding list of genes involved. When no explanation is found, whole-exome sequencing (WES) can be used in an attempt to identify gene defects in patients suspected to suffer from a genetic form. We describe three siblings, born to consanguineous parents, with a lethal severe epileptic encephalopathy with early-infantile onset, including their magnetic resonance imaging, electroencephalography and, in one case, neuropathological findings. Using WES a homozygous frameshift mutation in the BRAT1 gene, c.638dup p.(Val214Glyfs*189), was identified. We present our cases in the context of all published cases with mutations in the BRAT1 gene and conclude that BRAT1 should be added to the growing list of genes related to early-onset severe encephalopathy with epilepsy. Georg Thieme Verlag KG Stuttgart · New York.

  15. Dietary tomato and lycopene impact androgen signaling- and carcinogenesis-related gene expression during early TRAMP prostate carcinogenesis

    Science.gov (United States)

    Wan, Lei; Tan, Hsueh-Li; Thomas-Ahner, Jennifer M.; Pearl, Dennis K.; Erdman, John W.; Moran, Nancy E.; Clinton, Steven K.

    2014-01-01

    Consumption of tomato products containing the carotenoid lycopene is associated with a reduced risk of prostate cancer. To identify gene expression patterns associated with early testosterone-driven prostate carcinogenesis, which are impacted by dietary tomato and lycopene, wild type (WT) and transgenic adenocarcinoma of the mouse prostate (TRAMP) mice were fed control or tomato- or lycopene-containing diets from 4-10 wk-of-age. Eight-week-old mice underwent sham surgery, castration, or castration followed by testosterone-repletion (2.5 mg/kg/d initiated 1 wk after castration). Ten-wk-old intact TRAMP mice exhibit early multifocal prostatic intraepithelial neoplasia (PIN). Of the 200 prostate cancer-related genes measured by quantitative NanoString®, 189 are detectable, 164 significantly differ by genotype, 179 by testosterone status, and 30 by diet type (Plycopene feeding (Srd5a1) and by tomato-feeding (Srd5a2, Pxn, and Srebf1). Additionally, tomato-feeding significantly reduced expression of genes associated with stem cell features, Aldh1a and Ly6a, while lycopene-feeding significantly reduced expression of neuroendocrine differentiation-related genes, Ngfr and Syp. Collectively, these studies demonstrate a profile of testosterone-regulated genes associated with early stages of prostate carcinogenesis that are potential mechanistic targets of dietary tomato components. Future studies on androgen signaling/metabolism, stem cell features, and neuroendocrine differentiation pathways may elucidate the mechanisms by which dietary tomato and lycopene impact prostate cancer risk. PMID:25315431

  16. Neuropeptide Y gene polymorphisms confer risk of early-onset atherosclerosis.

    Directory of Open Access Journals (Sweden)

    Svati H Shah

    2009-01-01

    Full Text Available Neuropeptide Y (NPY is a strong candidate gene for coronary artery disease (CAD. We have previously identified genetic linkage to familial CAD in the genomic region of NPY. We performed follow-up genetic, biostatistical, and functional analysis of NPY in early-onset CAD. In familial CAD (GENECARD, N = 420 families, we found increased microsatellite linkage to chromosome 7p14 (OSA LOD = 4.2, p = 0.004 in 97 earliest age-of-onset families. Tagged NPY SNPs demonstrated linkage to CAD of a 6-SNP block (LOD = 1.58-2.72, family-based association of this block with CAD (p = 0.02, and stronger linkage to CAD in the earliest age-of-onset families. Association of this 6-SNP block with CAD was validated in: (a 556 non-familial early-onset CAD cases and 256 controls (OR 1.46-1.65, p = 0.01-0.05, showing stronger association in youngest cases (OR 1.84-2.20, p = 0.0004-0.09; and (b GENECARD probands versus non-familial controls (OR 1.79-2.06, p = 0.003-0.02. A promoter SNP (rs16147 within this 6-SNP block was associated with higher plasma NPY levels (p = 0.04. To assess a causal role of NPY in atherosclerosis, we applied the NPY1-receptor-antagonist BIBP-3226 adventitially to endothelium-denuded carotid arteries of apolipoprotein E-deficient mice; treatment reduced atherosclerotic neointimal area by 50% (p = 0.03. Thus, NPY variants associate with atherosclerosis in two independent datasets (with strong age-of-onset effects and show allele-specific expression with NPY levels, while NPY receptor antagonism reduces atherosclerosis in mice. We conclude that NPY contributes to atherosclerosis pathogenesis.

  17. Cord blood gene expression supports that prenatal exposure to perfluoroalkyl substances causes depressed immune functionality in early childhood.

    Science.gov (United States)

    Pennings, Jeroen L A; Jennen, Danyel G J; Nygaard, Unni C; Namork, Ellen; Haug, Line S; van Loveren, Henk; Granum, Berit

    2016-01-01

    Perfluoroalkyl and polyfluoroalkyl substances (PFAS) are a class of synthetic compounds that have widespread use in consumer and industrial applications. PFAS are considered environmental pollutants that have various toxic properties, including effects on the immune system. Recent human studies indicate that prenatal exposure to PFAS leads to suppressed immune responses in early childhood. In this study, data from the Norwegian BraMat cohort was used to investigate transcriptomics profiles in neonatal cord blood and their association with maternal PFAS exposure, anti-rubella antibody levels at 3 years of age and the number of common cold episodes until 3 years. Genes associated with PFAS exposure showed enrichment for immunological and developmental functions. The analyses identified a toxicogenomics profile of 52 PFAS exposure-associated genes that were in common with genes associated with rubella titers and/or common cold episodes. This gene set contains several immunomodulatory genes (CYTL1, IL27) as well as other immune-associated genes (e.g. EMR4P, SHC4, ADORA2A). In addition, this study identified PPARD as a PFAS toxicogenomics marker. These markers can serve as the basis for further mechanistic or epidemiological studies. This study provides a transcriptomics connection between prenatal PFAS exposure and impaired immune function in early childhood and supports current views on PPAR- and NF-κB-mediated modes of action. The findings add to the available evidence that PFAS exposure is immunotoxic in humans and support regulatory policies to phase out these substances.

  18. E6-associated transcription patterns in human papilloma virus 16-positive cervical tissues.

    Science.gov (United States)

    Lin, Kezhi; Lu, Xulian; Chen, Jun; Zou, Ruanmin; Zhang, Lifang; Xue, Xiangyang

    2015-01-01

    The change in transcription pattern induced by post-transcriptional RNA splicing is an important mechanism in the regulation of the early gene expression of human papilloma virus (HPV). The present study was conducted to establish a method to specifically amplify HPV-16 E6-associated transcripts. The E6-related transcripts from 63 HPV-16-positive cervical tumor tissue samples were amplified, consisting of eight cases of low-risk intraepithelial lesions, 38 cases of high-risk intraepithelial lesions and 17 cases of cervical cancer (CxCa). The appropriate amplified segments were recovered following agarose gel electrophoresis, and subjected to further sequencing and sequence alignment analysis. Six groups of E6 transcription patterns were identified from HPV-16-positive cervical tumor tissue, including five newly-discovered transcripts. Different HPV-16 E6-associated transcription patterns were detected during the development of CxCa. Over the course of the progression of the low-grade squamous intraepithelial lesions to CxCa, the specific HPV-16 E6-associated transcription patterns and the dominant transcripts were all different. As indicated by this study, the transcription pattern of the E6 early gene of HPV-16 was closely associated with the stages of cervical carcinogenesis, and may also be involved in the development of CxCa.

  19. Meta-analysis of differentiating mouse embryonic stem cell gene expression kinetics reveals early change of a small gene set.

    Directory of Open Access Journals (Sweden)

    Clive H Glover

    2006-11-01

    Full Text Available Stem cell differentiation involves critical changes in gene expression. Identification of these should provide endpoints useful for optimizing stem cell propagation as well as potential clues about mechanisms governing stem cell maintenance. Here we describe the results of a new meta-analysis methodology applied to multiple gene expression datasets from three mouse embryonic stem cell (ESC lines obtained at specific time points during the course of their differentiation into various lineages. We developed methods to identify genes with expression changes that correlated with the altered frequency of functionally defined, undifferentiated ESC in culture. In each dataset, we computed a novel statistical confidence measure for every gene which captured the certainty that a particular gene exhibited an expression pattern of interest within that dataset. This permitted a joint analysis of the datasets, despite the different experimental designs. Using a ranking scheme that favored genes exhibiting patterns of interest, we focused on the top 88 genes whose expression was consistently changed when ESC were induced to differentiate. Seven of these (103728_at, 8430410A17Rik, Klf2, Nr0b1, Sox2, Tcl1, and Zfp42 showed a rapid decrease in expression concurrent with a decrease in frequency of undifferentiated cells and remained predictive when evaluated in additional maintenance and differentiating protocols. Through a novel meta-analysis, this study identifies a small set of genes whose expression is useful for identifying changes in stem cell frequencies in cultures of mouse ESC. The methods and findings have broader applicability to understanding the regulation of self-renewal of other stem cell types.

  20. Expression stability and selection of optimal reference genes for gene expression normalization in early life stage rainbow trout exposed to cadmium and copper.

    Science.gov (United States)

    Shekh, Kamran; Tang, Song; Niyogi, Som; Hecker, Markus

    2017-09-01

    Gene expression analysis represents a powerful approach to characterize the specific mechanisms by which contaminants interact with organisms. One of the key considerations when conducting gene expression analyses using quantitative real-time reverse transcription-polymerase chain reaction (qPCR) is the selection of appropriate reference genes, which is often overlooked. Specifically, to reach meaningful conclusions when using relative quantification approaches, expression levels of reference genes must be highly stable and cannot vary as a function of experimental conditions. However, to date, information on the stability of commonly used reference genes across developmental stages, tissues and after exposure to contaminants such as metals is lacking for many vertebrate species including teleost fish. Therefore, in this study, we assessed the stability of expression of 8 reference gene candidates in the gills and skin of three different early life-stages of rainbow trout after acute exposure (24h) to two metals, cadmium (Cd) and copper (Cu) using qPCR. Candidate housekeeping genes were: beta actin (b-actin), DNA directed RNA polymerase II subunit I (DRP2), elongation factor-1 alpha (EF1a), glyceraldehyde 3-phosphate dehydrogenase (GAPDH), glucose-6-phosphate dehydrogenase (G6PD), hypoxanthine phosphoribosyltransferase (HPRT), ribosomal protein L8 (RPL8), and 18S ribosomal RNA (18S). Four algorithms, geNorm, NormFinder, BestKeeper, and the comparative ΔCt method were employed to systematically evaluate the expression stability of these candidate genes under control and exposed conditions as well as across three different life-stages. Finally, stability of genes was ranked by taking geometric means of the ranks established by the different methods. Stability of reference genes was ranked in the following order (from lower to higher stability): HPRT

  1. Differential impact of Met receptor gene interaction with early-life stress on neuronal morphology and behavior in mice.

    Science.gov (United States)

    Heun-Johnson, Hanke; Levitt, Pat

    2018-02-01

    Early adversity in childhood increases the risk of anxiety, mood, and post-traumatic stress disorders in adulthood, and specific gene-by-environment interactions may increase risk further. A common functional variant in the promoter region of the gene encoding the human MET receptor tyrosine kinase (rs1858830 ' C' allele) reduces expression of MET and is associated with altered cortical circuit function and structural connectivity. Mice with reduced Met expression exhibit changes in anxiety-like and conditioned fear behavior, precocious synaptic maturation in the hippocampus, and reduced neuronal arbor complexity and synaptogenesis. These phenotypes also can be produced independently by early adversity in wild-type mice. The present study addresses the outcome of combining early-life stress and genetic influences that alter timing of maturation on enduring functional and structural phenotypes. Using a model of reduced Met expression ( Met +/- ) and early-life stress from postnatal day 2-9, social, anxiety-like, and contextual fear behaviors in later life were measured. Mice that experienced early-life stress exhibited impairments in social interaction, whereas alterations in anxiety-like behavior and fear learning were driven by Met haploinsufficiency, independent of rearing condition. Early-life stress or reduced Met expression decreased arbor complexity of ventral hippocampal CA1 pyramidal neurons projecting to basolateral amygdala. Paradoxically, arbor complexity in Met +/- mice was increased following early-life stress, and thus not different from arbors in wild-type mice raised in control conditions. The changes in dendritic morphology are consistent with the hypothesis that the physiological state of maturation of CA1 neurons in Met +/- mice influences their responsiveness to early-life stress. The dissociation of behavioral and structural changes suggests that there may be phenotype-specific sensitivities to early-life stress.

  2. Transcriptional activation of immediate-early gene ETR101 by human T-cell leukaemia virus type I Tax

    DEFF Research Database (Denmark)

    Chen, Li; Ma, Shiliang; Li, Bo

    2003-01-01

    Human T-cell leukaemia virus type I (HTLV-I) Tax regulates viral and cellular gene expression through interactions with multiple cellular transcription pathways. This study describes the finding of immediate-early gene ETR101 expression in HTLV-I-infected cells and its regulation by Tax. ETR101...... was persistently expressed in HTLV-I-infected cells but not in HTLV-I uninfected cells. Expression of ETR101 was dependent upon Tax expression in the inducible Tax-expressing cell line JPX-9 and also in Jurkat cells transiently transfected with Tax-expressing vectors. Tax transactivated the ETR101 gene promoter......-DNA complex in HTLV-I-infected cell lines. EMSA with specific antibodies confirmed that the CREB transcription factor was responsible for formation of this specific protein-DNA complex. These results suggested that Tax directly transactivated ETR101 gene expression, mainly through a CRE sequence via the CREB...

  3. Genetic Analysis of PARK2 and PINK1 Genes in Brazilian Patients with Early-Onset Parkinson's Disease

    Directory of Open Access Journals (Sweden)

    Karla Cristina Vasconcelos Moura

    2013-01-01

    Full Text Available Parkinson's disease is the second most frequent neurodegenerative disorder in the world, affecting 1-2% of individuals over the age of 65. The etiology of Parkinson's disease is complex, with the involvement of gene-environment interactions. Although it is considered a disease of late manifestation, early-onset forms of parkinsonism contribute to 5–10% of all cases. In the present study, we screened mutations in coding regions of PARK2 and PINK1 genes in 136 unrelated Brazilian patients with early-onset Parkinson's disease through automatic sequencing. We identified six missense variants in PARK2 gene: one known pathogenic mutation, two variants of uncertain role, and three nonpathogenic changes. No pathogenic mutation was identified in PINK1 gene, only benign polymorphisms. All putative pathogenic variants found in this study were in heterozygous state. Our data show that PARK2 point mutations are more common in Brazilian early-onset Parkinson's disease patients (2.9% than PINK1 missense variants (0%, corroborating other studies worldwide.

  4. Hypoxia-activated genes from early placenta are elevated in Preeclampsia, but not in Intra-Uterine Growth Retardation

    Directory of Open Access Journals (Sweden)

    Danan Jean-Louis

    2005-08-01

    Full Text Available Abstract Background As a first step to explore the possible relationships existing between the effects of low oxygen pressure in the first trimester placenta and placental pathologies developing from mid-gestation, two subtracted libraries totaling 2304 cDNA clones were constructed. For achieving this, two reciprocal suppressive/subtractive hybridization procedures (SSH were applied to early (11 weeks human placental villi after incubation either in normoxic or in hypoxic conditions. The clones from both libraries (1440 hypoxia-specific and 864 normoxia-specific were spotted on nylon macroarrays. Complex cDNAs probes prepared from placental villi (either from early pregnancy, after hypoxic or normoxic culture conditions, or near term for controls or pathological placentas were hybridized to the membranes. Results Three hundred and fifty nine clones presenting a hybridization signal above the background were sequenced and shown to correspond to 276 different genes. Nine of these genes are mitochondrial, while 267 are nuclear. Specific expression profiles characteristic of preeclampsia (PE could be identified, as well as profiles specific of intra-uterine growth retardation (IUGR. Focusing on the chromosomal distribution of the fraction of genes that responded in at least one hybridization experiment, we could observe a highly significant chromosomal clustering of 54 genes into 8 chromosomal regions, four of which containing imprinted genes. Comparative mapping data indicate that these imprinted clusters are maintained in synteny in mice, and apparently in cattle and pigs, suggesting that the maintenance of such syntenies is requested for achieving a normal placental physiology in eutherian mammals. Conclusion We could demonstrate that genes induced in PE were also genes highly expressed under hypoxic conditions (P = 5.10-5, which was not the case for isolated IUGR. Highly expressed placental genes may be in syntenies conserved interspecifically

  5. Farm exposure and time trends in early childhood may influence DNA methylation in genes related to asthma and allergy.

    Science.gov (United States)

    Michel, S; Busato, F; Genuneit, J; Pekkanen, J; Dalphin, J-C; Riedler, J; Mazaleyrat, N; Weber, J; Karvonen, A M; Hirvonen, M-R; Braun-Fahrländer, C; Lauener, R; von Mutius, E; Kabesch, M; Tost, J

    2013-03-01

    Genetic susceptibility and environmental influences are important contributors to the development of asthma and atopic diseases. Epigenetic mechanisms may facilitate gene by environment interactions in these diseases. We studied the rural birth cohort PASTURE (Protection against allergy: study in rural environments) to investigate (a) whether epigenetic patterns in asthma candidate genes are influenced by farm exposure in general, (b) change over the first years of life, and (c) whether these changes may contribute to the development of asthma. DNA was extracted from cord blood and whole blood collected at the age of 4.5 years in 46 samples per time point. DNA methylation in 23 regions in ten candidate genes (ORMDL1, ORMDL2, ORMDL3, CHI3L1, RAD50, IL13, IL4, STAT6, FOXP3, and RUNX3) was assessed by pyrosequencing, and differences between strata were analyzed by nonparametric Wilcoxon-Mann-Whitney tests. In cord blood, regions in ORMDL1 and STAT6 were hypomethylated in DNA from farmers' as compared to nonfarmers' children, while regions in RAD50 and IL13 were hypermethylated (lowest P-value (STAT6) = 0.001). Changes in methylation over time occurred in 15 gene regions (lowest P-value (IL13) = 1.57*10(-8)). Interestingly, these differences clustered in the genes highly associated with asthma (ORMDL family) and IgE regulation (RAD50, IL13, and IL4), but not in the T-regulatory genes (FOXP3, RUNX3). In this first pilot study, DNA methylation patterns change significantly in early childhood in specific asthma- and allergy-related genes in peripheral blood cells, and early exposure to farm environment seems to influence methylation patterns in distinct genes. © 2013 John Wiley & Sons A/S. Published by Blackwell Publishing Ltd.

  6. Hierarchical clustering of gene expression patterns in the Eomes + lineage of excitatory neurons during early neocortical development

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    Cameron David A

    2012-08-01

    Full Text Available Abstract Background Cortical neurons display dynamic patterns of gene expression during the coincident processes of differentiation and migration through the developing cerebrum. To identify genes selectively expressed by the Eomes + (Tbr2 lineage of excitatory cortical neurons, GFP-expressing cells from Tg(Eomes::eGFP Gsat embryos were isolated to > 99% purity and profiled. Results We report the identification, validation and spatial grouping of genes selectively expressed within the Eomes + cortical excitatory neuron lineage during early cortical development. In these neurons 475 genes were expressed ≥ 3-fold, and 534 genes ≤ 3-fold, compared to the reference population of neuronal precursors. Of the up-regulated genes, 328 were represented at the Genepaint in situ hybridization database and 317 (97% were validated as having spatial expression patterns consistent with the lineage of differentiating excitatory neurons. A novel approach for quantifying in situ hybridization patterns (QISP across the cerebral wall was developed that allowed the hierarchical clustering of genes into putative co-regulated groups. Forty four candidate genes were identified that show spatial expression with Intermediate Precursor Cells, 49 candidate genes show spatial expression with Multipolar Neurons, while the remaining 224 genes achieved peak expression in the developing cortical plate. Conclusions This analysis of differentiating excitatory neurons revealed the expression patterns of 37 transcription factors, many chemotropic signaling molecules (including the Semaphorin, Netrin and Slit signaling pathways, and unexpected evidence for non-canonical neurotransmitter signaling and changes in mechanisms of glucose metabolism. Over half of the 317 identified genes are associated with neuronal disease making these findings a valuable resource for studies of neurological development and disease.

  7. The Tyrosyl-DNA Phosphodiesterase 1β (Tdp1β Gene Discloses an Early Response to Abiotic Stresses

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    Maria Elisa Sabatini

    2017-11-01

    Full Text Available Tyrosyl-DNA phosphodiesterase 1 (Tdp1 is involved in DNA repair pathways as it mends the topoisomerase I—DNA covalent complexes. In plants, a small Tdp1 gene family, composed by Tdp1α and Tdp1β genes, was identified, but the roles of these genes in abiotic stress responses are not fully understood. To investigate their specific stress response patterns, the present study made use of bioinformatic and molecular tools to look into the Tdp1β gene function, so far described only in the plant kingdom, and compare it with Tdp1α gene coding for the canonical, highly conserved α isoform. The expression profiles of Tdp1α and Tdp1β genes were examined under abiotic stress conditions (cold, heat, high osmolarity, salt, and UV-B in two model species, Arabidopsis thaliana and Medicago truncatula. The two isoforms of topoisomerase I (TOP1α and TOP1β were also taken into consideration in view of their known roles in DNA metabolism and cell proliferation. Data relative to gene expression in Arabidopsis were retrieved from the AtGenExpress microarray dataset, while quantitative Real-Time PCR was carried out to evaluate the stress response in M. truncatula cell cultures. These analyses revealed that Tdp1β gene expression was enhanced during the first hour of treatment, whereas Tdp1α enhanced expression succeeded at subsequent timepoints. In agreement with the gene-specific responses to abiotic stress conditions, the promoter regions of Tdp1α and Tdp1β genes are well equipped with stress-related cis-elements. An in-depth bioinformatic characterization of the HIRAN motif, a distinctive feature of the Tdp1β protein, showed its wide distribution in chromatin remodeling and DNA repair proteins. The reported data suggests that Tdp1β functions in the early response to abiotic stresses.

  8. Targeted gene panel sequencing in children with very early onset inflammatory bowel disease--evaluation and prospective analysis.

    Science.gov (United States)

    Kammermeier, Jochen; Drury, Suzanne; James, Chela T; Dziubak, Robert; Ocaka, Louise; Elawad, Mamoun; Beales, Philip; Lench, Nicholas; Uhlig, Holm H; Bacchelli, Chiara; Shah, Neil

    2014-11-01

    Multiple monogenetic conditions with partially overlapping phenotypes can present with inflammatory bowel disease (IBD)-like intestinal inflammation. With novel genotype-specific therapies emerging, establishing a molecular diagnosis is becoming increasingly important. We have introduced targeted next-generation sequencing (NGS) technology as a prospective screening tool in children with very early onset IBD (VEOIBD). We evaluated the coverage of 40 VEOIBD genes in two separate cohorts undergoing targeted gene panel sequencing (TGPS) (n=25) and whole exome sequencing (WES) (n=20). TGPS revealed causative mutations in four genes (IL10RA, EPCAM, TTC37 and SKIV2L) discovered unexpected phenotypes and directly influenced clinical decision making by supporting as well as avoiding haematopoietic stem cell transplantation. TGPS resulted in significantly higher median coverage when compared with WES, fewer coverage deficiencies and improved variant detection across established VEOIBD genes. Excluding or confirming known VEOIBD genotypes should be considered early in the disease course in all cases of therapy-refractory VEOIBD, as it can have a direct impact on patient management. To combine both described NGS technologies would compensate for the limitations of WES for disease-specific application while offering the opportunity for novel gene discovery in the research setting. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

  9. Targeted next generation sequencing identifies functionally deleterious germline mutations in novel genes in early-onset/familial prostate cancer.

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    Paula Paulo

    2018-04-01

    Full Text Available Considering that mutations in known prostate cancer (PrCa predisposition genes, including those responsible for hereditary breast/ovarian cancer and Lynch syndromes, explain less than 5% of early-onset/familial PrCa, we have sequenced 94 genes associated with cancer predisposition using next generation sequencing (NGS in a series of 121 PrCa patients. We found monoallelic truncating/functionally deleterious mutations in seven genes, including ATM and CHEK2, which have previously been associated with PrCa predisposition, and five new candidate PrCa associated genes involved in cancer predisposing recessive disorders, namely RAD51C, FANCD2, FANCI, CEP57 and RECQL4. Furthermore, using in silico pathogenicity prediction of missense variants among 18 genes associated with breast/ovarian cancer and/or Lynch syndrome, followed by KASP genotyping in 710 healthy controls, we identified "likely pathogenic" missense variants in ATM, BRIP1, CHEK2 and TP53. In conclusion, this study has identified putative PrCa predisposing germline mutations in 14.9% of early-onset/familial PrCa patients. Further data will be necessary to confirm the genetic heterogeneity of inherited PrCa predisposition hinted in this study.

  10. Early developmental gene enhancers affect subcortical volumes in the adult human brain.

    Science.gov (United States)

    Becker, Martin; Guadalupe, Tulio; Franke, Barbara; Hibar, Derrek P; Renteria, Miguel E; Stein, Jason L; Thompson, Paul M; Francks, Clyde; Vernes, Sonja C; Fisher, Simon E

    2016-05-01

    Genome-wide association screens aim to identify common genetic variants contributing to the phenotypic variability of complex traits, such as human height or brain morphology. The identified genetic variants are mostly within noncoding genomic regions and the biology of the genotype-phenotype association typically remains unclear. In this article, we propose a complementary targeted strategy to reveal the genetic underpinnings of variability in subcortical brain volumes, by specifically selecting genomic loci that are experimentally validated forebrain enhancers, active in early embryonic development. We hypothesized that genetic variation within these enhancers may affect the development and ultimately the structure of subcortical brain regions in adults. We tested whether variants in forebrain enhancer regions showed an overall enrichment of association with volumetric variation in subcortical structures of >13,000 healthy adults. We observed significant enrichment of genomic loci that affect the volume of the hippocampus within forebrain enhancers (empirical P = 0.0015), a finding which robustly passed the adjusted threshold for testing of multiple brain phenotypes (cutoff of P < 0.0083 at an alpha of 0.05). In analyses of individual single nucleotide polymorphisms (SNPs), we identified an association upstream of the ID2 gene with rs7588305 and variation in hippocampal volume. This SNP-based association survived multiple-testing correction for the number of SNPs analyzed but not for the number of subcortical structures. Targeting known regulatory regions offers a way to understand the underlying biology that connects genotypes to phenotypes, particularly in the context of neuroimaging genetics. This biology-driven approach generates testable hypotheses regarding the functional biology of identified associations. Hum Brain Mapp 37:1788-1800, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  11. The Transcriptomes of Xiphinema index and Longidorus elongatus Suggest Independent Acquisition of Some Plant Parasitism Genes by Horizontal Gene Transfer in Early-Branching Nematodes.

    Science.gov (United States)

    Danchin, Etienne G J; Perfus-Barbeoch, Laetitia; Rancurel, Corinne; Thorpe, Peter; Da Rocha, Martine; Bajew, Simon; Neilson, Roy; Guzeeva, Elena Sokolova; Da Silva, Corinne; Guy, Julie; Labadie, Karine; Esmenjaud, Daniel; Helder, Johannes; Jones, John T; den Akker, Sebastian Eves-van

    2017-10-23

    Nematodes have evolved the ability to parasitize plants on at least four independent occasions, with plant parasites present in Clades 1, 2, 10 and 12 of the phylum. In the case of Clades 10 and 12, horizontal gene transfer of plant cell wall degrading enzymes from bacteria and fungi has been implicated in the evolution of plant parasitism. We have used ribonucleic acid sequencing (RNAseq) to generate reference transcriptomes for two economically important nematode species, Xiphinema index and Longidorus elongatus , representative of two genera within the early-branching Clade 2 of the phylum Nematoda. We used a transcriptome-wide analysis to identify putative horizontal gene transfer events. This represents the first in-depth transcriptome analysis from any plant-parasitic nematode of this clade. For each species, we assembled ~30 million Illumina reads into a reference transcriptome. We identified 62 and 104 transcripts, from X. index and L. elongatus , respectively, that were putatively acquired via horizontal gene transfer. By cross-referencing horizontal gene transfer prediction with a phylum-wide analysis of Pfam domains, we identified Clade 2-specific events. Of these, a GH12 cellulase from X. index was analysed phylogenetically and biochemically, revealing a likely bacterial origin and canonical enzymatic function. Horizontal gene transfer was previously shown to be a phenomenon that has contributed to the evolution of plant parasitism among nematodes. Our findings underline the importance and the extensiveness of this phenomenon in the evolution of plant-parasitic life styles in this speciose and widespread animal phylum.

  12. A Single Dose of LSD Does Not Alter Gene Expression of the Serotonin 2A Receptor Gene (HTR2A) or Early Growth Response Genes (EGR1-3) in Healthy Subjects

    Science.gov (United States)

    Dolder, Patrick C.; Grünblatt, Edna; Müller, Felix; Borgwardt, Stefan J.; Liechti, Matthias E.

    2017-01-01

    Rationale: Renewed interest has been seen in the use of lysergic acid diethylamide (LSD) in psychiatric research and practice. The repeated use of LSD leads to tolerance that is believed to result from serotonin (5-HT) 5-HT2A receptor downregulation. In rats, daily LSD administration for 4 days decreased frontal cortex 5-HT2A receptor binding. Additionally, a single dose of LSD acutely increased expression of the early growth response genes EGR1 and EGR2 in rat and mouse brains through 5-HT2A receptor stimulation. No human data on the effects of LSD on gene expression has been reported. Therefore, we investigated the effects of single-dose LSD administration on the expression of the 5-HT2A receptor gene (HTR2A) and EGR1-3 genes. Methods: mRNA expression levels were analyzed in whole blood as a peripheral biomarker in 15 healthy subjects before and 1.5 and 24 h after the administration of LSD (100 μg) and placebo in a randomized, double-blind, placebo-controlled, cross-over study. Results: LSD did not alter the expression of the HTR2A or EGR1-3 genes 1.5 and 24 h after administration compared with placebo. Conclusion: No changes were observed in the gene expression of LSD’s primary target receptor gene or genes that are implicated in its downstream effects. Remaining unclear is whether chronic LSD administration alters gene expression in humans. PMID:28701958

  13. Combined chromatin and expression analysis reveals specific regulatory mechanisms within cytokine genes in the macrophage early immune response.

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    Maria Jesus Iglesias

    Full Text Available Macrophages play a critical role in innate immunity, and the expression of early response genes orchestrate much of the initial response of the immune system. Macrophages undergo extensive transcriptional reprogramming in response to inflammatory stimuli such as Lipopolysaccharide (LPS.To identify gene transcription regulation patterns involved in early innate immune responses, we used two genome-wide approaches--gene expression profiling and chromatin immunoprecipitation-sequencing (ChIP-seq analysis. We examined the effect of 2 hrs LPS stimulation on early gene expression and its relation to chromatin remodeling (H3 acetylation; H3Ac and promoter binding of Sp1 and RNA polymerase II phosphorylated at serine 5 (S5P RNAPII, which is a marker for transcriptional initiation. Our results indicate novel and alternative gene regulatory mechanisms for certain proinflammatory genes. We identified two groups of up-regulated inflammatory genes with respect to chromatin modification and promoter features. One group, including highly up-regulated genes such as tumor necrosis factor (TNF, was characterized by H3Ac, high CpG content and lack of TATA boxes. The second group, containing inflammatory mediators (interleukins and CCL chemokines, was up-regulated upon LPS stimulation despite lacking H3Ac in their annotated promoters, which were low in CpG content but did contain TATA boxes. Genome-wide analysis showed that few H3Ac peaks were unique to either +/-LPS condition. However, within these, an unpacking/expansion of already existing H3Ac peaks was observed upon LPS stimulation. In contrast, a significant proportion of S5P RNAPII peaks (approx 40% was unique to either condition. Furthermore, data indicated a large portion of previously unannotated TSSs, particularly in LPS-stimulated macrophages, where only 28% of unique S5P RNAPII peaks overlap annotated promoters. The regulation of the inflammatory response appears to occur in a very specific manner at

  14. Molecular pathways of early CD105-positive erythroid cells as compared with CD34-positive common precursor cells by flow cytometric cell-sorting and gene expression profiling

    International Nuclear Information System (INIS)

    Machherndl-Spandl, S; Suessner, S; Danzer, M; Proell, J; Gabriel, C; Lauf, J; Sylie, R; Klein, H-U; Béné, M C; Weltermann, A; Bettelheim, P

    2013-01-01

    Special attention has recently been drawn to the molecular network of different genes that are responsible for the development of erythroid cells. The aim of the present study was to establish in detail the immunophenotype of early erythroid cells and to compare the gene expression profile of freshly isolated early erythroid precursors with that of the CD34-positive (CD34 + ) compartment. Multiparameter flow cytometric analyses of human bone marrow mononuclear cell fractions (n=20) defined three distinct early erythroid stages. The gene expression profile of sorted early erythroid cells was analyzed by Affymetrix array technology. For 4524 genes, a differential regulation was found in CD105-positive erythroid cells as compared with the CD34 + progenitor compartment (2362 upregulated genes). A highly significant difference was observed in the expression level of genes involved in transcription, heme synthesis, iron and mitochondrial metabolism and transforming growth factor-β signaling. A comparison with recently published data showed over 1000 genes that as yet have not been reported to be upregulated in the early erythroid lineage. The gene expression level within distinct pathways could be illustrated directly by applying the Ingenuity software program. The results of gene expression analyses can be seen at the Gene Expression Omnibus repository

  15. Downregulation of immediate-early genes linking to suppression of neuronal plasticity in rats after 28-day exposure to glycidol

    International Nuclear Information System (INIS)

    Akane, Hirotoshi; Saito, Fumiyo; Shiraki, Ayako; Takeyoshi, Masahiro; Imatanaka, Nobuya; Itahashi, Megu; Murakami, Tomoaki; Shibutani, Makoto

    2014-01-01

    We previously found that the 28-day oral toxicity study of glycidol at 200 mg/kg/day in rats resulted in axonopathy in both the central and peripheral nervous systems and aberrations in the late-stage of hippocampal neurogenesis targeting the process of neurite extension. To capture the neuronal parameters in response to glycidol toxicity, these animals were subjected to region-specific global gene expression profiling in four regions of cerebral and cerebellar architectures, followed by immunohistochemical analysis of selected gene products. Expression changes of genes related to axonogenesis and synaptic transmission were observed in the hippocampal dentate gyrus, cingulate cortex and cerebellar vermis at 200 mg/kg showing downregulation in most genes. In the corpus callosum, genes related to growth, survival and functions of glial cells fluctuated their expression. Immunohistochemically, neurons expressing gene products of immediate-early genes, i.e., Arc, Fos and Jun, decreased in their number in the dentate granule cell layer, cingulate cortex and cerebellar vermis. We also applied immunohistochemical analysis in rat offspring after developmental exposure to glycidol through maternal drinking water. The results revealed increases of Arc + neurons at 1000 ppm and Fos + neurons at ≥ 300 ppm in the dentate granule cell layer of offspring only at the adult stage. These results suggest that glycidol suppressed neuronal plasticity in the brain after 28-day exposure to young adult animals, in contrast to the operation of restoration mechanism to increase neuronal plasticity at the adult stage in response to aberrations in neurogenesis after developmental exposure. - Highlights: • Neuronal toxicity parameters after 28-day glycidol treatment were examined in rats. • Region-specific global gene expression profiling was conducted in brain regions. • Cortical tissues downregulated genes on axonogenesis and synaptic transmission. • Cortical tissues decreased

  16. Whole transcriptome profiling of maize during early somatic embryogenesis reveals altered expression of stress factors and embryogenesis-related genes.

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    Stella A G D Salvo

    Full Text Available Embryogenic tissue culture systems are utilized in propagation and genetic engineering of crop plants, but applications are limited by genotype-dependent culture response. To date, few genes necessary for embryogenic callus formation have been identified or characterized. The goal of this research was to enhance our understanding of gene expression during maize embryogenic tissue culture initiation. In this study, we highlight the expression of candidate genes that have been previously regarded in the literature as having important roles in somatic embryogenesis. We utilized RNA based sequencing (RNA-seq to characterize the transcriptome of immature embryo explants of the highly embryogenic and regenerable maize genotype A188 at 0, 24, 36, 48, and 72 hours after placement of explants on tissue culture initiation medium. Genes annotated as functioning in stress response, such as glutathione-S-transferases and germin-like proteins, and genes involved with hormone transport, such as PINFORMED, increased in expression over 8-fold in the study. Maize genes with high sequence similarity to genes previously described in the initiation of embryogenic cultures, such as transcription factors BABY BOOM, LEAFY COTYLEDON, and AGAMOUS, and important receptor-like kinases such as SOMATIC EMBRYOGENESIS RECEPTOR LIKE KINASES and CLAVATA, were also expressed in this time course study. By combining results from whole genome transcriptome analysis with an in depth review of key genes that play a role in the onset of embryogenesis, we propose a model of coordinated expression of somatic embryogenesis-related genes, providing an improved understanding of genomic factors involved in the early steps of embryogenic culture initiation in maize and other plant species.

  17. Gene Expression Profiling of Peripheral Blood From Kidney Transplant Recipients for the Early Detection of Digestive System Cancer.

    Science.gov (United States)

    Kusaka, M; Okamoto, M; Takenaka, M; Sasaki, H; Fukami, N; Kataoka, K; Ito, T; Kenmochi, T; Hoshinaga, K; Shiroki, R

    2017-06-01

    Kidney transplant recipients are at increased risk of developing cancer in comparison with the general population. To effectively manage post-transplantation malignancies, it is essential to proactively monitor patients. A long-term intensive screening program was associated with a reduced incidence of cancer after transplantation. This study evaluated the usefulness of the gene expression profiling of peripheral blood samples obtained from kidney transplant patients and adopted a screening test for detecting cancer of the digestive system (gastric, colon, pancreas, and biliary tract). Nineteen patients were included in this study and a total of 53 gene expression screening tests were performed. The gene expression profiles of blood-delivered total RNA and whole genome human gene expression profiles were obtained. We investigated the expression levels of 2665 genes associated with digestive cancers and counted the number of genes in which expression was altered. A hierarchical clustering analysis was also performed. The final prediction of the cancer possibility was determined according to an algorithm. The number of genes in which expression was altered was significantly increased in the kidney transplant recipients in comparison with the general population (1091 ± 63 vs 823 ± 94; P = .0024). The number of genes with altered expression decreased after the induction of mechanistic target of rapamycin (mTOR) inhibitor (1484 ± 227 vs 883 ± 154; P = .0439). No cases of possible digestive cancer were detected in this study period. The gene expression profiling of peripheral blood samples may be a useful and noninvasive diagnostic tool that allows for the early detection of cancer of the digestive system. Copyright © 2017 Elsevier Inc. All rights reserved.

  18. Downregulation of immediate-early genes linking to suppression of neuronal plasticity in rats after 28-day exposure to glycidol

    Energy Technology Data Exchange (ETDEWEB)

    Akane, Hirotoshi [Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo 183-8509 (Japan); Saito, Fumiyo [Chemicals Evaluation and Research Institute, Japan, 1-4-25 Koraku, Bunkyo-ku, Tokyo 112-0004 (Japan); Shiraki, Ayako [Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo 183-8509 (Japan); Pathogenetic Veterinary Science, United Graduate School of Veterinary Sciences, Gifu University, 1-1 Yanagido, Gifu-shi, Gifu 501-1193 (Japan); Takeyoshi, Masahiro; Imatanaka, Nobuya [Chemicals Evaluation and Research Institute, Japan, 1-4-25 Koraku, Bunkyo-ku, Tokyo 112-0004 (Japan); Itahashi, Megu [Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo 183-8509 (Japan); Pathogenetic Veterinary Science, United Graduate School of Veterinary Sciences, Gifu University, 1-1 Yanagido, Gifu-shi, Gifu 501-1193 (Japan); Murakami, Tomoaki [Laboratory of Veterinary Toxicology, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo 183-8509 (Japan); Shibutani, Makoto, E-mail: mshibuta@cc.tuat.ac.jp [Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo 183-8509 (Japan)

    2014-09-01

    We previously found that the 28-day oral toxicity study of glycidol at 200 mg/kg/day in rats resulted in axonopathy in both the central and peripheral nervous systems and aberrations in the late-stage of hippocampal neurogenesis targeting the process of neurite extension. To capture the neuronal parameters in response to glycidol toxicity, these animals were subjected to region-specific global gene expression profiling in four regions of cerebral and cerebellar architectures, followed by immunohistochemical analysis of selected gene products. Expression changes of genes related to axonogenesis and synaptic transmission were observed in the hippocampal dentate gyrus, cingulate cortex and cerebellar vermis at 200 mg/kg showing downregulation in most genes. In the corpus callosum, genes related to growth, survival and functions of glial cells fluctuated their expression. Immunohistochemically, neurons expressing gene products of immediate-early genes, i.e., Arc, Fos and Jun, decreased in their number in the dentate granule cell layer, cingulate cortex and cerebellar vermis. We also applied immunohistochemical analysis in rat offspring after developmental exposure to glycidol through maternal drinking water. The results revealed increases of Arc{sup +} neurons at 1000 ppm and Fos{sup +} neurons at ≥ 300 ppm in the dentate granule cell layer of offspring only at the adult stage. These results suggest that glycidol suppressed neuronal plasticity in the brain after 28-day exposure to young adult animals, in contrast to the operation of restoration mechanism to increase neuronal plasticity at the adult stage in response to aberrations in neurogenesis after developmental exposure. - Highlights: • Neuronal toxicity parameters after 28-day glycidol treatment were examined in rats. • Region-specific global gene expression profiling was conducted in brain regions. • Cortical tissues downregulated genes on axonogenesis and synaptic transmission. • Cortical tissues

  19. Gene expression profiling reveals underlying molecular mechanisms of the early stages of tamoxifen-induced rat hepatocarcinogenesis

    International Nuclear Information System (INIS)

    Pogribny, Igor P.; Bagnyukova, Tetyana V.; Tryndyak, Volodymyr P.; Muskhelishvili, Levan; Rodriguez-Juarez, Rocio; Kovalchuk, Olga; Han Tao; Fuscoe, James C.; Ross, Sharon A.; Beland, Frederick A.

    2007-01-01

    Tamoxifen is a widely used anti-estrogenic drug for chemotherapy and, more recently, for the chemoprevention of breast cancer. Despite the indisputable benefits of tamoxifen in preventing the occurrence and re-occurrence of breast cancer, the use of tamoxifen has been shown to induce non-alcoholic steatohepatitis, which is a life-threatening fatty liver disease with a risk of progression to cirrhosis and hepatocellular carcinoma. In recent years, the high-throughput microarray technology for large-scale analysis of gene expression has become a powerful tool for increasing the understanding of the molecular mechanisms of carcinogenesis and for identifying new biomarkers with diagnostic and predictive values. In the present study, we used the high-throughput microarray technology to determine the gene expression profiles in the liver during early stages of tamoxifen-induced rat hepatocarcinogenesis. Female Fisher 344 rats were fed a 420 ppm tamoxifen containing diet for 12 or 24 weeks, and gene expression profiles were determined in liver of control and tamoxifen-exposed rats. The results indicate that early stages of tamoxifen-induced liver carcinogenesis are characterized by alterations in several major cellular pathways, specifically those involved in the tamoxifen metabolism, lipid metabolism, cell cycle signaling, and apoptosis/cell proliferation control. One of the most prominent changes during early stages of tamoxifen-induced hepatocarcinogenesis is dysregulation of signaling pathways in cell cycle progression from the G 1 to S phase, evidenced by the progressive and sustained increase in expression of the Pdgfc, Calb3, Ets1, and Ccnd1 genes accompanied by the elevated level of the PI3K, p-PI3K, Akt1/2, Akt3, and cyclin B, D1, and D3 proteins. The early appearance of these alterations suggests their importance in the mechanism of neoplastic cell transformation induced by tamoxifen

  20. Early diffusion of gene expression profiling in breast cancer patients associated with areas of high income inequality.

    Science.gov (United States)

    Ponce, Ninez A; Ko, Michelle; Liang, Su-Ying; Armstrong, Joanne; Toscano, Michele; Chanfreau-Coffinier, Catherine; Haas, Jennifer S

    2015-04-01

    With the Affordable Care Act reducing coverage disparities, social factors could prominently determine where and for whom innovations first diffuse in health care markets. Gene expression profiling is a potentially cost-effective innovation that guides chemotherapy decisions in early-stage breast cancer, but adoption has been uneven across the United States. Using a sample of commercially insured women, we evaluated whether income inequality in metropolitan areas was associated with receipt of gene expression profiling during its initial diffusion in 2006-07. In areas with high income inequality, gene expression profiling receipt was higher than elsewhere, but it was associated with a 10.6-percentage-point gap between high- and low-income women. In areas with low rates of income inequality, gene expression profiling receipt was lower, with no significant differences by income. Even among insured women, income inequality may indirectly shape diffusion of gene expression profiling, with benefits accruing to the highest-income patients in the most unequal places. Policies reducing gene expression profiling disparities should address low-inequality areas and, in unequal places, practice settings serving low-income patients. Project HOPE—The People-to-People Health Foundation, Inc.

  1. Identification of nickel response genes in abnormal early developments of sea urchin by differential display polymerase chain reaction.

    Science.gov (United States)

    Ryu, Tae Kwon; Lee, Gunsup; Rhee, Yong; Park, Heung-Sik; Chang, Man; Lee, Sukchan; Lee, Jaean; Lee, Taek-Kyun

    2012-10-01

    Bioassays and biomarkers have been previously developed to assess the effects of heavy metal contaminants on the early life stages of the sea urchin. In this study, malformation in the early developmental processes was observed in sea urchin (Strongylocentrotus intermedius) larvae exposed to 10 ppm Ni for over 30 h. The most critical stage at which the triggering of nickel effects takes place is thought to be the blastula stage, which occurs after fertilization in larval development. To investigate the molecular-level responses of sea urchin exposed to heavy metal stress and to explore the differentially expressed genes that are induced or repressed by nickel, differential display polymerase chain reaction (DD-PCR) was used with sea urchin mRNAs. The malformation-related genes expressed in the early life stages of the sea urchin were cloned from larvae exposed to 10 ppm of nickel for 15 h, and accessed via DD-PCR. Sequence analysis results revealed that each of the genes evidenced high homology with EGF2, PCSK9, serine/threonine protein kinase, apolipophorin precursor protein, and MGC80921 protein/transcript variant 2. This result may prove useful in the development of novel biomarkers for the assessment of heavy metal stresses on sea urchin embryos. Copyright © 2012 Elsevier Inc. All rights reserved.

  2. The p. N103K mutation of leptin (LEP gene and severe early onset obesity in Pakistan

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    Shabana

    Full Text Available BACKGROUND: Obesity is a complex disorder and has been increasing globally at alarming rates including Pakistan. However, there is scarce research on understanding obesity genetics in Pakistan. Leptin is a hormone secreted by adipocytes in response to satiety and correlates with body weight. Any mutations in the LEP gene have an adverse effect on energy regulation pathway and lead to severe, early onset obesity. To date, only eight mutations have been described in the LEP gene of which p. N103K is one. METHODS: We aimed to analyze the prevalence of this mutation in Pakistani subjects. A total of 475 subjects were genotyped by PCR-RFLP analysis and their serum profiling was done. RESULTS: Results showed that this mutation was present only in one male child with early onset obesity (10 year. He had very low serum leptin levels suggestive of functional impact of the mutation. The prevalence of such mutations is, however, low due to the drastic effects on the energy regulation. CONCLUSION: In conclusion, LEP gene mutations contribute significantly to the monogenic forms of obesity and are important due to the availability of treatment options. Such mutations may exert their effect by directly affecting energy regulation pathway and are more prominent in the early stages of life only.

  3. Expectations and beliefs in science communication: Learning from three European gene therapy discussions of the early 1990s.

    Science.gov (United States)

    Meyer, Gitte

    2016-04-01

    There is widespread agreement that the potential of gene therapy was oversold in the early 1990s. This study, however, comparing written material from the British, Danish and German gene therapy discourses of the period finds significant differences: Over-optimism was not equally strong everywhere; gene therapy was not universally hyped. Against that background, attention is directed towards another area of variation in the material: different basic assumptions about science and scientists. Exploring such culturally rooted assumptions and beliefs and their possible significance to science communication practices, it is argued that deep beliefs may constitute drivers of hype that are particularly difficult to deal with. To participants in science communication, the discouragement of hype, viewed as a practical-ethical challenge, can be seen as a learning exercise that includes critical attention to internalised beliefs. © The Author(s) 2014.

  4. Transcriptome Sequencing, De Novo Assembly and Differential Gene Expression Analysis of the Early Development of Acipenser baeri.

    Directory of Open Access Journals (Sweden)

    Wei Song

    Full Text Available The molecular mechanisms that drive the development of the endangered fossil fish species Acipenser baeri are difficult to study due to the lack of genomic data. Recent advances in sequencing technologies and the reducing cost of sequencing offer exclusive opportunities for exploring important molecular mechanisms underlying specific biological processes. This manuscript describes the large scale sequencing and analyses of mRNA from Acipenser baeri collected at five development time points using the Illumina Hiseq2000 platform. The sequencing reads were de novo assembled and clustered into 278167 unigenes, of which 57346 (20.62% had 45837 known homologues proteins in Uniprot protein databases while 11509 proteins matched with at least one sequence of assembled unigenes. The remaining 79.38% of unigenes could stand for non-coding unigenes or unigenes specific to A. baeri. A number of 43062 unigenes were annotated into functional categories via Gene Ontology (GO annotation whereas 29526 unigenes were associated with 329 pathways by mapping to KEGG database. Subsequently, 3479 differentially expressed genes were scanned within developmental stages and clustered into 50 gene expression profiles. Genes preferentially expressed at each stage were also identified. Through GO and KEGG pathway enrichment analysis, relevant physiological variations during the early development of A. baeri could be better cognized. Accordingly, the present study gives insights into the transcriptome profile of the early development of A. baeri, and the information contained in this large scale transcriptome will provide substantial references for A. baeri developmental biology and promote its aquaculture research.

  5. Expression of immediate-early genes in the inferior colliculus and auditory cortex in salicylate-induced tinnitus in rat.

    Science.gov (United States)

    Hu, S S; Mei, L; Chen, J Y; Huang, Z W; Wu, H

    2014-03-12

    Tinnitus could be associated with neuronal hyperactivity in the auditory center. As a neuronal activity marker, immediate-early gene (IEG) expression is considered part of a general neuronal response to natural stimuli. Some IEGs, especially the activity-dependent cytoskeletal protein (Arc) and the early growth response gene-1 (Egr-1), appear to be highly correlated with sensory-evoked neuronal activity. We hypothesize, therefore, an increase of Arc and Egr-1 will be observed in a tinnitus model. In our study, we used the gap prepulse inhibition of acoustic startle (GPIAS) paradigm to confirm that salicylate induces tinnitus-like behavior in rats. However, expression of the Arc gene and Egr-1 gene were decreased in the inferior colliculus (IC) and auditory cortex (AC), in contradiction of our hypothesis. Expression of N-methyl d-aspartate receptor subunit 2B (NR2B) was increased and all of these changes returned to normal 14 days after treatment with salicylate ceased. These data revealed long-time administration of salicylate induced tinnitus markedly but reversibly and caused neural plasticity changes in the IC and the AC. Decreased expression of Arc and Egr-1 might be involved with instability of synaptic plasticity in tinnitus.

  6. Expression of immediate-early genes in the inferior colliculus and auditory cortex in salicylate-induced tinnitus in rat

    Directory of Open Access Journals (Sweden)

    S.S. Hu

    2014-03-01

    Full Text Available Tinnitus could be associated with neuronal hyperactivity in the auditory center. As a neuronal activity marker, immediate-early gene (IEG expression is considered part of a general neuronal response to natural stimuli. Some IEGs, especially the activity-dependent cytoskeletal protein (Arc and the early growth response gene-1 (Egr-1, appear to be highly correlated with sensory-evoked neuronal activity. We hypothesize, therefore, an increase of Arc and Egr-1 will be observed in a tinnitus model. In our study, we used the gap prepulse inhibition of acoustic startle (GPIAS paradigm to confirm that salicylate induces tinnitus-like behavior in rats. However, expression of the Arc gene and Egr-1 gene were decreased in the inferior colliculus (IC and auditory cortex (AC, in contradiction of our hypothesis. Expression of N-methyl d-aspartate receptor subunit 2B (NR2B was increased and all of these changes returned to normal 14 days after treatment with salicylate ceased. These data revealed long-time administration of salicylate induced tinnitus markedly but reversibly and caused neural plasticity changes in the IC and the AC. Decreased expression of Arc and Egr-1 might be involved with instability of synaptic plasticity in tinnitus.

  7. Levetiracetam attenuates hippocampal expression of synaptic plasticity-related immediate early and late response genes in amygdala-kindled rats

    Directory of Open Access Journals (Sweden)

    Watson William P

    2010-01-01

    Full Text Available Abstract Background The amygdala-kindled rat is a model for human temporal lobe epilepsy and activity-dependent synaptic plasticity. Hippocampal RNA isolated from amygdala-kindled rats at different kindling stages was analyzed to identify kindling-induced genes. Furthermore, effects of the anti-epileptic drug levetiracetam on kindling-induced gene expression were examined. Results Cyclooxygenase-2 (Cox-2, Protocadherin-8 (Pcdh8 and TGF-beta-inducible early response gene-1 (TIEG1 were identified and verified as differentially expressed transcripts in the hippocampus of kindled rats by in situ hybridization and quantitative RT-PCR. In addition, we identified a panel of 16 additional transcripts which included Arc, Egr3/Pilot, Homer1a, Ania-3, MMP9, Narp, c-fos, NGF, BDNF, NT-3, Synaptopodin, Pim1 kinase, TNF-α, RGS2, Egr2/krox-20 and β-A activin that were differentially expressed in the hippocampus of amygdala-kindled rats. The list consists of many synaptic plasticity-related immediate early genes (IEGs as well as some late response genes encoding transcription factors, neurotrophic factors and proteins that are known to regulate synaptic remodelling. In the hippocampus, induction of IEG expression was dependent on the afterdischarge (AD duration. Levetiracetam, 40 mg/kg, suppressed the development of kindling measured as severity of seizures and AD duration. In addition, single animal profiling also showed that levetiracetam attenuated the observed kindling-induced IEG expression; an effect that paralleled the anti-epileptic effect of the drug on AD duration. Conclusions The present study provides mRNA expression data that suggest that levetiracetam attenuates expression of genes known to regulate synaptic remodelling. In the kindled rat, levetiracetam does so by shortening the AD duration thereby reducing the seizure-induced changes in mRNA expression in the hippocampus.

  8. Shear stress upregulates regeneration-related immediate early genes in liver progenitors in 3D ECM-like microenvironments.

    Science.gov (United States)

    Nishii, Kenichiro; Brodin, Erik; Renshaw, Taylor; Weesner, Rachael; Moran, Emma; Soker, Shay; Sparks, Jessica L

    2018-05-01

    The role of fluid stresses in activating the hepatic stem/progenitor cell regenerative response is not well understood. This study hypothesized that immediate early genes (IEGs) with known links to liver regeneration will be upregulated in liver progenitor cells (LPCs) exposed to in vitro shear stresses on the order of those produced from elevated interstitial flow after partial hepatectomy. The objectives were: (1) to develop a shear flow chamber for application of fluid stress to LPCs in 3D culture; and (2) to determine the effects of fluid stress on IEG expression in LPCs. Two hours of shear stress exposure at ∼4 dyn/cm 2 was applied to LPCs embedded individually or as 3D spheroids within a hyaluronic acid/collagen I hydrogel. Results were compared against static controls. Quantitative reverse transcriptase polymerase chain reaction was used to evaluate the effect of experimental treatments on gene expression. Twenty-nine genes were analyzed, including IEGs and other genes linked to liver regeneration. Four IEGs (CFOS, IP10, MKP1, ALB) and three other regeneration-related genes (WNT, VEGF, EpCAM) were significantly upregulated in LPCs in response to fluid mechanical stress. LPCs maintained an early to intermediate stage of differentiation in spheroid culture in the absence of the hydrogel, and addition of the gel initiated cholangiocyte differentiation programs which were abrogated by the onset of flow. Collectively the flow-upregulated genes fit the pattern of an LPC-mediated proliferative/regenerative response. These results suggest that fluid stresses are potentially important regulators of the LPC-mediated regeneration response in liver. © 2017 Wiley Periodicals, Inc.

  9. Evolution of GHF5 endoglucanase gene structure in plant-parasitic nematodes: no evidence for an early domain shuffling event.

    Science.gov (United States)

    Kyndt, Tina; Haegeman, Annelies; Gheysen, Godelieve

    2008-11-03

    the whole gene cassette, i.e. the GHF5 catalytic domain and the CBM2, rather than that it evolved by domain shuffling. Our evolutionary model for the gene structure in PPN GHF5 endoglucanases implies the occurrence of an early duplication event, and more recent gene duplications at genus or species level.

  10. Evolution of GHF5 endoglucanase gene structure in plant-parasitic nematodes: no evidence for an early domain shuffling event

    Directory of Open Access Journals (Sweden)

    Gheysen Godelieve

    2008-11-01

    PPN GHF5 endoglucanase gene most probably consisted of the whole gene cassette, i.e. the GHF5 catalytic domain and the CBM2, rather than that it evolved by domain shuffling. Our evolutionary model for the gene structure in PPN GHF5 endoglucanases implies the occurrence of an early duplication event, and more recent gene duplications at genus or species level.

  11. Regulation of early signaling and gene expression in the α-particle and bystander response of IMR-90 human fibroblasts

    Directory of Open Access Journals (Sweden)

    Hei Tom K

    2010-07-01

    Full Text Available Abstract Background The existence of a radiation bystander effect, in which non-irradiated cells respond to signals from irradiated cells, is well established. To understand early signaling and gene regulation in bystander cells, we used a bio-informatics approach, measuring global gene expression at 30 minutes and signaling pathways between 30 minutes and 4 hours after exposure to α-particles in IMR-90 fibroblasts. Methods We used whole human genome microarrays and real time quantitative PCR to measure and validate gene expression. Microarray analysis was done using BRB-Array Tools; pathway and ontology analyses were done using Ingenuity Pathway Analysis and PANTHER, respectively. We studied signaling in irradiated and bystander cells using immunoblotting and semi-quantitative image analysis. Results Gene ontology suggested signal transduction and transcriptional regulation responding 30 minutes after treatment affected cell structure, motility and adhesion, and interleukin synthesis. We measured time-dependent expression of genes controlled by the NF-κB pathway; matrix metalloproteinases 1 and 3; chemokine ligands 2, 3 and 5 and interleukins 1β, 6 and 33. There was an increased response of this set of genes 30 minutes after treatment and another wave of induction at 4 hours. We investigated AKT-GSK3β signaling and found both AKT and GSK3β are hyper-phosphorylated 30 minutes after irradiation and this effect is maintained through 4 hours. In bystander cells, a similar response was seen with a delay of 30 minutes. We proposed a network model where the observed decrease in phosphorylation of β-catenin protein after GSK3β dependent inactivation can trigger target gene expression at later times after radiation exposure Conclusions These results are the first to show that the radiation induced bystander signal induces a widespread gene expression response at 30 minutes after treatment and these changes are accompanied by modification of

  12. Dynamics of immediate early gene and neuropeptide gene response to prolonged immobilization stress: evidence against a critical role of the termination of exposure to the stressor.

    Science.gov (United States)

    Trnecková, Lenka; Rotllant, David; Klenerová, Vera; Hynie, Sixtus; Armario, Antonio

    2007-02-01

    Stress-induced expression of immediate early genes (IEGs) appears to be transient even if the exposure to the stressor persists. However, there are some exceptions which suggest that particular characteristics of stressors can affect the dynamics of IEG expression. We studied in selected telencephalic, diencephalic and brainstem regions the mRNA levels of two clearly distinct IEGs (c-fos and arc) during prolonged exposure to a severe stressor such as immobilization (IMO) and after releasing the rats from the situation. Although regional differences were observed with the two IEGs, overall, c-fos mRNA levels progressively declined over the course of 4 h of continuous exposure to IMO, whereas arc mRNA levels were maintained at high levels in the brain regions that express this gene under stress (telencephalon). Levels of CRF hnRNA in the hypothalamus paraventricular nucleus only slightly declined during prolonged exposure to IMO. Surprisingly, termination of exposure to IMO did not modify CRF gene expression in the paraventricular nucleus or the pattern of IEGs expression, with the exception of c-fos in the lateral septum. Thus, putative signals associated to the termination of exposure to IMO were unable to modify either IEG expression in most brain areas or CRF gene expression in the paraventricular nucleus.

  13. Visualization of odor-induced neuronal activity by immediate early gene expression

    Directory of Open Access Journals (Sweden)

    Bepari Asim K

    2012-11-01

    Full Text Available Abstract Background Sensitive detection of sensory-evoked neuronal activation is a key to mechanistic understanding of brain functions. Since immediate early genes (IEGs are readily induced in the brain by environmental changes, tracing IEG expression provides a convenient tool to identify brain activity. In this study we used in situ hybridization to detect odor-evoked induction of ten IEGs in the mouse olfactory system. We then analyzed IEG induction in the cyclic nucleotide-gated channel subunit A2 (Cnga2-null mice to visualize residual neuronal activity following odorant exposure since CNGA2 is a key component of the olfactory signal transduction pathway in the main olfactory system. Results We observed rapid induction of as many as ten IEGs in the mouse olfactory bulb (OB after olfactory stimulation by a non-biological odorant amyl acetate. A robust increase in expression of several IEGs like c-fos and Egr1 was evident in the glomerular layer, the mitral/tufted cell layer and the granule cell layer. Additionally, the neuronal IEG Npas4 showed steep induction from a very low basal expression level predominantly in the granule cell layer. In Cnga2-null mice, which are usually anosmic and sexually unresponsive, glomerular activation was insignificant in response to either ambient odorants or female stimuli. However, a subtle induction of c-fos took place in the OB of a few Cnga2-mutants which exhibited sexual arousal. Interestingly, very strong glomerular activation was observed in the OB of Cnga2-null male mice after stimulation with either the neutral odor amyl acetate or the predator odor 2, 3, 5-trimethyl-3-thiazoline (TMT. Conclusions This study shows for the first time that in vivo olfactory stimulation can robustly induce the neuronal IEG Npas4 in the mouse OB and confirms the odor-evoked induction of a number of IEGs. As shown in previous studies, our results indicate that a CNGA2-independent signaling pathway(s may activate the

  14. High-throughput sequencing of the T cell receptor β gene identifies aggressive early-stage mycosis fungoides.

    Science.gov (United States)

    de Masson, Adele; O'Malley, John T; Elco, Christopher P; Garcia, Sarah S; Divito, Sherrie J; Lowry, Elizabeth L; Tawa, Marianne; Fisher, David C; Devlin, Phillip M; Teague, Jessica E; Leboeuf, Nicole R; Kirsch, Ilan R; Robins, Harlan; Clark, Rachael A; Kupper, Thomas S

    2018-05-09

    Mycosis fungoides (MF), the most common cutaneous T cell lymphoma (CTCL) is a malignancy of skin-tropic memory T cells. Most MF cases present as early stage (stage I A/B, limited to the skin), and these patients typically have a chronic, indolent clinical course. However, a small subset of early-stage cases develop progressive and fatal disease. Because outcomes can be so different, early identification of this high-risk population is an urgent unmet clinical need. We evaluated the use of next-generation high-throughput DNA sequencing of the T cell receptor β gene ( TCRB ) in lesional skin biopsies to predict progression and survival in a discovery cohort of 208 patients with CTCL (177 with MF) from a 15-year longitudinal observational clinical study. We compared these data to the results in an independent validation cohort of 101 CTCL patients (87 with MF). The tumor clone frequency (TCF) in lesional skin, measured by high-throughput sequencing of the TCRB gene, was an independent prognostic factor of both progression-free and overall survival in patients with CTCL and MF in particular. In early-stage patients, a TCF of >25% in the skin was a stronger predictor of progression than any other established prognostic factor (stage IB versus IA, presence of plaques, high blood lactate dehydrogenase concentration, large-cell transformation, or age). The TCF therefore may accurately predict disease progression in early-stage MF. Early identification of patients at high risk for progression could help identify candidates who may benefit from allogeneic hematopoietic stem cell transplantation before their disease becomes treatment-refractory. Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

  15. Gene expression changes as markers of early lapatinib response in a panel of breast cancer cell lines

    LENUS (Irish Health Repository)

    O’Neill, Fiona

    2012-06-18

    expressed in response to lapatinib at the 12 hour time point examined. The expression of these 5 genes correlated directly with lapatinib sensitivity. We propose that the gene expression profile may represent both an early measure of the likelihood of sensitivity and the level of response to lapatinib and may therefore have application in early response detection.

  16. A 7-Gene Signature Depicts the Biochemical Profile of Early Prefibrotic Myelofibrosis

    DEFF Research Database (Denmark)

    Skov, Vibe; Burton, Mark; Thomassen, Mads

    2016-01-01

    was performed in 17 and 9 patients diagnosed with ET and PMF, respectively. Using elevated LDH obtained at the time of diagnosis as a marker of prePMF, a 7-gene signature was identified which correctly predicted the prePMF group with a sensitivity of 100% and a specificity of 89%. The 7 genes included MPO......, CEACAM8, CRISP3, MS4A3, CEACAM6, HEMGN, and MMP8, which are genes known to be involved in inflammation, cell adhesion, differentiation and proliferation. Evaluation of bone marrow biopsies and the 7-gene signature showed a concordance rate of 71%, 79%, 62%, and 38%. Our 7-gene signature may be a useful...

  17. Differentially expressed androgen-regulated genes in androgen-sensitive tissues reveal potential biomarkers of early prostate cancer.

    Directory of Open Access Journals (Sweden)

    Dogus Murat Altintas

    Full Text Available BACKGROUND: Several data favor androgen receptor implication in prostate cancer initiation through the induction of several gene activation programs. The aim of the study is to identify potential biomarkers for early diagnosis of prostate cancer (PCa among androgen-regulated genes (ARG and to evaluate comparative expression of these genes in normal prostate and normal prostate-related androgen-sensitive tissues that do not (or rarely give rise to cancer. METHODS: ARG were selected in non-neoplastic adult human prostatic epithelial RWPE-1 cells stably expressing an exogenous human androgen receptor, using RNA-microarrays and validation by qRT-PCR. Expression of 48 preselected genes was quantified in tissue samples (seminal vesicles, prostate transitional zones and prostate cancers, benign prostatic hypertrophy obtained from surgical specimens using TaqMan® low-density arrays. The diagnostic performances of these potential biomarkers were compared to that of genes known to be associated with PCa (i.e. PCA3 and DLX1. RESULTS AND DISCUSSION: By crossing expression studies in 26 matched PCa and normal prostate transitional zone samples, and 35 matched seminal vesicle and PCa samples, 14 genes were identified. Similarly, 9 genes were overexpressed in 15 benign prostatic hypertrophy samples, as compared to PCa samples. Overall, we selected 8 genes of interest to evaluate their diagnostic performances in comparison with that of PCA3 and DLX1. Among them, 3 genes: CRYAB, KCNMA1 and SDPR, were overexpressed in all 3 reference non-cancerous tissues. The areas under ROC curves of these genes reached those of PCA3 (0.91 and DLX1 (0.94. CONCLUSIONS: We identified ARG with reduced expression in PCa and with significant diagnostic values for discriminating between cancerous and non-cancerous prostatic tissues, similar that of PCA3. Given their expression pattern, they could be considered as potentially protective against prostate cancer. Moreover, they could

  18. Caffeine induces high expression of cyp-35A family genes and inhibits the early larval development in Caenorhabditis elegans.

    Science.gov (United States)

    Min, Hyemin; Kawasaki, Ichiro; Gong, Joomi; Shim, Yhong-Hee

    2015-03-01

    Intake of caffeine during pregnancy can cause retardation of fetal development. Although the significant influence of caffeine on animal development is widely recognized, much remains unknown about its mode of action because of its pleiotropic effects on living organisms. In the present study, by using Caenorhabditis elegans as a model organism, the effects of caffeine on development were examined. Brood size, embryonic lethality, and percent larval development were investigated, and caffeine was found to inhibit the development of C. elegans at most of the stages in a dosage-dependent fashion. Upon treatment with 30 mM caffeine, the majority (86.1 ± 3.4%) of the L1 larvae were irreversibly arrested without further development. In contrast, many of the late-stage larvae survived and grew to adults when exposed to the same 30 mM caffeine. These results suggest that early-stage larvae are more susceptible to caffeine than later-stage larvae. To understand the metabolic responses to caffeine treatment, the levels of expression of cytochrome P450 (cyp) genes were examined with or without caffeine treatment using comparative micro-array, and it was found that the expression of 24 cyp genes was increased by more than 2-fold (p family was the most prominent. Interestingly, depletion of the cyp-35A family genes one-by-one or in combination through RNA interference resulted in partial rescue from early larval developmental arrest caused by caffeine treatment, suggesting that the high-level induction of cyp-35A family genes can be fatal to the development of early-stage larvae.

  19. MicroRNAs and Target Genes As Biomarkers for the Diagnosis of Early Onset of Parkinson Disease

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    Ahmad R. Arshad

    2017-10-01

    Full Text Available Among the neurodegenerative disorders, Parkinson's disease (PD ranks as the second most common disorder with a higher prevalence in individuals aged over 60 years old. Younger individuals may also be affected with PD which is known as early onset PD (EOPD. Despite similarities between the characteristics of EOPD and late onset PD (LODP, EOPD patients experience much longer disease manifestations and poorer quality of life. Although some individuals are more prone to have EOPD due to certain genetic alterations, the molecular mechanisms that differentiate between EOPD and LOPD remains unclear. Recent findings in PD patients revealed that there were differences in the genetic profiles of PD patients compared to healthy controls, as well as between EOPD and LOPD patients. There were variants identified that correlated with the decline of cognitive and motor symptoms as well as non-motor symptoms in PD. There were also specific microRNAs that correlated with PD progression, and since microRNAs have been shown to be involved in the maintenance of neuronal development, mitochondrial dysfunction and oxidative stress, there is a strong possibility that these microRNAs can be potentially used to differentiate between subsets of PD patients. PD is mainly diagnosed at the late stage, when almost majority of the dopaminergic neurons are lost. Therefore, identification of molecular biomarkers for early detection of PD is important. Given that miRNAs are crucial in controlling the gene expression, these regulatory microRNAs and their target genes could be used as biomarkers for early diagnosis of PD. In this article, we discussed the genes involved and their regulatory miRNAs, regarding their roles in PD progression, based on the findings of significantly altered microRNAs in EOPD studies. We also discussed the potential of these miRNAs as molecular biomarkers for early diagnosis.

  20. The NRF2-KEAP1 Pathway Is an Early Responsive Gene Network in Arsenic Exposed Lymphoblastoid Cells

    Science.gov (United States)

    Córdova, Emilio J.; Martínez-Hernández, Angélica; Uribe-Figueroa, Laura; Centeno, Federico; Morales-Marín, Mirna; Koneru, Harsha; Coleman, Matthew A.; Orozco, Lorena

    2014-01-01

    Inorganic arsenic (iAs), a major environmental contaminant, has risen as an important health problem worldwide. More detailed identification of the molecular mechanisms associated with iAs exposure would help to establish better strategies for prevention and treatment. Although chronic iAs exposures have been previously studied there is little to no information regarding the early events of exposure to iAs. To better characterize the early mechanisms of iAs exposure we conducted gene expression studies using sublethal doses of iAs at two different time-points. The major transcripts differentially regulated at 2 hrs of iAs exposure included antioxidants, detoxificants and chaperones. Moreover, after 12 hrs of exposure many of the down-regulated genes were associated with DNA replication and S phase cell cycle progression. Interestingly, the most affected biological pathway by both 2 or 12 hrs of iAs exposure were the Nrf2-Keap1 pathway, represented by the highly up-regulated HMOX1 transcript, which is transcriptionally regulated by the transcription factor Nrf2. Additional Nrf2 targets included SQSTM1 and ABCB6, which were not previously associated with acute iAs exposure. Signalling pathways such as interferon, B cell receptor and AhR route were also responsive to acute iAs exposure. Since HMOX1 expression increased early (20 min) and was responsive to low iAs concentrations (0.1 µM), this gene could be a suitable early biomarker for iAs exposure. In addition, the novel Nrf2 targets SQSTM1 and ABCB6 could play an important and previously unrecognized role in cellular protection against iAs. PMID:24516582

  1. Novel mutations in the BRCA1 and BRCA2 genes in Iranian women with early-onset breast cancer

    International Nuclear Information System (INIS)

    Yassaee, Vahid R; Zeinali, Sirous; Harirchi, Iraj; Jarvandi, Soghra; Mohagheghi, Mohammad A; Hornby, David P; Dalton, Ann

    2002-01-01

    Breast cancer is the most common female malignancy and a major cause of death in middle-aged women. So far, germline mutations in the BRCA1 and BRCA2 genes in patients with early-onset breast and/or ovarian cancer have not been identified within the Iranian population. With the collaboration of two main centres for cancer in Iran, we obtained clinical information, family history and peripheral blood from 83 women under the age of 45 with early-onset breast cancer for scanning of germline mutations in the BRCA1 and BRCA2 genes. We analysed BRCA1 exons 11 and BRCA2 exons 10 and 11 by the protein truncation test, and BRCA1 exons 2, 3, 5, 13 and 20 and BRCA2 exons 9, 17, 18 and 23 with the single-strand conformation polymorphism assay on genomic DNA amplified by polymerase chain reaction. Ten sequence variants were identified: five frameshifts (putative mutations – four novel); three missense changes of unknown significance and two polymorphisms, one seen commonly in both Iranian and British populations. Identification of these novel mutations suggests that any given population should develop a mutation database for its programme of breast cancer screening. The pattern of mutations seen in the BRCA genes seems not to differ from other populations studied. Early-onset breast cancer (less than 45 years) and a limited family history is sufficient to justify mutation screening with a detection rate of over 25% in this group, whereas sporadic early-onset breast cancer (detection rate less than 5%) is unlikely to be cost-effective

  2. Global gene analysis of oocytes from early stages in human folliculogenesis shows high expression of novel genes in reproduction

    DEFF Research Database (Denmark)

    Markholt, Sara; Grøndahl, M L; Ernst, Erik

    2012-01-01

    The pool of primordial follicles in humans is laid down during embryonic development and follicles can remain dormant for prolonged intervals, often decades, until individual follicles resume growth. The mechanisms that induce growth and maturation of primordial follicles are poorly understood...... but follicles once activated either continue growth or undergo atresia. We have isolated pure populations of oocytes from human primordial, intermediate and primary follicles using laser capture micro-dissection microscopy and evaluated the global gene expression profiles by whole-genome microarray analysis......) and the mitochondrial-encoded ATPase6 (ATP6). Thus, the present study provides not only a technique to capture and perform transcriptome analysis of the sparse material of human oocytes from the earliest follicle stages but further includes a comprehensive basis for our understanding of the regulatory factors...

  3. Early life stress affects mortality rate more than social behavior, gene expression or oxidative damage in honey bee workers.

    Science.gov (United States)

    Rueppell, Olav; Yousefi, Babak; Collazo, Juan; Smith, Daniel

    2017-04-01

    Early life stressors can affect aging and life expectancy in positive or negative ways. Individuals can adjust their behavior and molecular physiology based on early life experiences but relatively few studies have connected such mechanisms to demographic patterns in social organisms. Sociality buffers individuals from environmental influences and it is unclear how much early life stress affects later life history. Workers of the honey bee (Apis mellifera L.) were exposed to two stressors, Varroa parasitism and Paraquat exposure, early in life. Consequences were measured at the molecular, behavioral, and demographic level. While treatments did not significantly affect levels of oxidative damage, expression of select genes, and titers of the common deformed wing virus, most of these measures were affected by age. Some of the age effects, such as declining levels of deformed wing virus and oxidative damage, were opposite to our predictions but may be explained by demographic selection. Further analyses suggested some influences of worker behavior on mortality and indicated weak treatment effects on behavior. The latter effects were inconsistent among the two experiments. However, mortality rate was consistently reduced by Varroa mite stress during development. Thus, mortality was more responsive to early life stress than our other response variables. The lack of treatment effects on these measures may be due to the social organization of honey bees that buffers the individual from the impact of stressful developmental conditions. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. Upregulation of inflammatory genes and downregulation of sclerostin gene expression are key elements in the early phase of fragility fracture healing.

    Directory of Open Access Journals (Sweden)

    Joana Caetano-Lopes

    Full Text Available BACKGROUND: Fracture healing is orchestrated by a specific set of events that culminates in the repair of bone and reachievement of its biomechanical properties. The aim of our work was to study the sequence of gene expression events involved in inflammation and bone remodeling occurring in the early phases of callus formation in osteoporotic patients. METHODOLOGY/PRINCIPAL FINDINGS: Fifty-six patients submitted to hip replacement surgery after a low-energy hip fracture were enrolled in this study. The patients were grouped according to the time interval between fracture and surgery: bone collected within 3 days after fracture (n = 13; between the 4(th and 7(th day (n = 33; and after one week from the fracture (n = 10. Inflammation- and bone metabolism-related genes were assessed at the fracture site. The expression of pro-inflammatory cytokines was increased in the first days after fracture. The genes responsible for bone formation and resorption were upregulated one week after fracture. The increase in RANKL expression occurred just before that, between the 4(th-7(th days after fracture. Sclerostin expression diminished during the first days after fracture. CONCLUSIONS: The expression of inflammation-related genes, especially IL-6, is highest at the very first days after fracture but from day 4 onwards there is a shift towards bone remodeling genes, suggesting that the inflammatory phase triggers bone healing. We propose that an initial inflammatory stimulus and a decrease in sclerostin-related effects are the key components in fracture healing. In osteoporotic patients, cellular machinery seems to adequately react to the inflammatory stimulus, therefore local promotion of these events might constitute a promising medical intervention to accelerate fracture healing.

  5. The Transcriptomes of Xiphinema index and Longidorus elongatus Suggest Independent Acquisition of Some Plant Parasitism Genes by Horizontal Gene Transfer in Early-Branching Nematodes

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    Etienne G.J. Danchin

    2017-10-01

    Full Text Available Nematodes have evolved the ability to parasitize plants on at least four independent occasions, with plant parasites present in Clades 1, 2, 10 and 12 of the phylum. In the case of Clades 10 and 12, horizontal gene transfer of plant cell wall degrading enzymes from bacteria and fungi has been implicated in the evolution of plant parasitism. We have used ribonucleic acid sequencing (RNAseq to generate reference transcriptomes for two economically important nematode species, Xiphinema index and Longidorus elongatus, representative of two genera within the early-branching Clade 2 of the phylum Nematoda. We used a transcriptome-wide analysis to identify putative horizontal gene transfer events. This represents the first in-depth transcriptome analysis from any plant-parasitic nematode of this clade. For each species, we assembled ~30 million Illumina reads into a reference transcriptome. We identified 62 and 104 transcripts, from X. index and L. elongatus, respectively, that were putatively acquired via horizontal gene transfer. By cross-referencing horizontal gene transfer prediction with a phylum-wide analysis of Pfam domains, we identified Clade 2-specific events. Of these, a GH12 cellulase from X. index was analysed phylogenetically and biochemically, revealing a likely bacterial origin and canonical enzymatic function. Horizontal gene transfer was previously shown to be a phenomenon that has contributed to the evolution of plant parasitism among nematodes. Our findings underline the importance and the extensiveness of this phenomenon in the evolution of plant-parasitic life styles in this speciose and widespread animal phylum.

  6. The Transcriptomes of Xiphinema index and Longidorus elongatus Suggest Independent Acquisition of Some Plant Parasitism Genes by Horizontal Gene Transfer in Early-Branching Nematodes

    Science.gov (United States)

    Danchin, Etienne G.J.; Perfus-Barbeoch, Laetitia; Rancurel, Corinne; Thorpe, Peter; Da Rocha, Martine; Bajew, Simon; Neilson, Roy; Sokolova (Guzeeva), Elena; Da Silva, Corinne; Guy, Julie; Labadie, Karine; Esmenjaud, Daniel; Helder, Johannes; Jones, John T.

    2017-01-01

    Nematodes have evolved the ability to parasitize plants on at least four independent occasions, with plant parasites present in Clades 1, 2, 10 and 12 of the phylum. In the case of Clades 10 and 12, horizontal gene transfer of plant cell wall degrading enzymes from bacteria and fungi has been implicated in the evolution of plant parasitism. We have used ribonucleic acid sequencing (RNAseq) to generate reference transcriptomes for two economically important nematode species, Xiphinema index and Longidorus elongatus, representative of two genera within the early-branching Clade 2 of the phylum Nematoda. We used a transcriptome-wide analysis to identify putative horizontal gene transfer events. This represents the first in-depth transcriptome analysis from any plant-parasitic nematode of this clade. For each species, we assembled ~30 million Illumina reads into a reference transcriptome. We identified 62 and 104 transcripts, from X. index and L. elongatus, respectively, that were putatively acquired via horizontal gene transfer. By cross-referencing horizontal gene transfer prediction with a phylum-wide analysis of Pfam domains, we identified Clade 2-specific events. Of these, a GH12 cellulase from X. index was analysed phylogenetically and biochemically, revealing a likely bacterial origin and canonical enzymatic function. Horizontal gene transfer was previously shown to be a phenomenon that has contributed to the evolution of plant parasitism among nematodes. Our findings underline the importance and the extensiveness of this phenomenon in the evolution of plant-parasitic life styles in this speciose and widespread animal phylum. PMID:29065523

  7. Post-Transcriptional Control of Gene Expression in Mouse Early Embryo Development: A View from the Tip of the Iceberg

    Directory of Open Access Journals (Sweden)

    Claudio Sette

    2011-04-01

    Full Text Available Fertilization is a very complex biological process that requires the perfect cooperation between two highly specialized cells: the male and female gametes. The oocyte provides the physical space where this process takes place, most of the energetic need, and half of the genetic contribution. The spermatozoon mostly contributes the other half of the chromosomes and it is specialized to reach and to penetrate the oocyte. Notably, the mouse oocyte and early embryo are transcriptionally inactive. Hence, they fully depend on the maternal mRNAs and proteins stored during oocyte maturation to drive the onset of development. The new embryo develops autonomously around the four-cell stage, when maternal supplies are exhausted and the zygotic genome is activated in mice. This oocyte-to-embryo transition needs an efficient and tightly regulated translation of the maternally-inherited mRNAs, which likely contributes to embryonic genome activation. Full understanding of post-transcriptional regulation of gene expression in early embryos is crucial to understand the reprogramming of the embryonic genome, it might help driving reprogramming of stem cells in vitro and will likely improve in vitro culturing of mammalian embryos for assisted reproduction. Nevertheless, the knowledge of the mechanism(s underlying this fundamental step in embryogenesis is still scarce, especially if compared to other model organisms. We will review here the current knowledge on the post-transcriptional control of gene expression in mouse early embryos and discuss some of the unanswered questions concerning this fascinating field of biology.

  8. A New Nonsense Mutation in CDKL5 Gene in a Male Patient with Early Onset Refractory Epilepsy: a Case Report

    Directory of Open Access Journals (Sweden)

    Soudeh Ghafouri-Fard

    2016-02-01

    Full Text Available Background The X-linked cyclin-dependent kinase like 5 (CDKL5/STK9 gene has been shown to be responsible for a severe encephalopathy condition characterized by early onset of epilepsy and severe developmental delay. CDKL5 mutations have been shown to be more frequent among female patients. Results Here we report a 6- month male patient, second child of a healthy non consanguineous in the Iranian population. He has been affected by early onset epileptic refractory seizures and developmental delay. Whole-exome sequencing (WES has revealed a base substitution c.173T>A in CDKL5 gene, resulting in the formation of stop codon p.L58X. This mutation resides in the catalytic domain of the corresponding protein and is expected to result in premature RNA break down with no CDKL5 resulting protein. Conclusion   The present report highlights the importance of CDKL5 mutation analysis in male patients affected with early onset refractory epilepsy.

  9. Causality analysis detects the regulatory role of maternal effect genes in the early Drosophila embryo

    Directory of Open Access Journals (Sweden)

    Zara Ghodsi

    2017-03-01

    Full Text Available In developmental studies, inferring regulatory interactions of segmentation genetic network play a vital role in unveiling the mechanism of pattern formation. As such, there exists an opportune demand for theoretical developments and new mathematical models which can result in a more accurate illustration of this genetic network. Accordingly, this paper seeks to extract the meaningful regulatory role of the maternal effect genes using a variety of causality detection techniques and to explore whether these methods can suggest a new analytical view to the gene regulatory networks. We evaluate the use of three different powerful and widely-used models representing time and frequency domain Granger causality and convergent cross mapping technique with the results being thoroughly evaluated for statistical significance. Our findings show that the regulatory role of maternal effect genes is detectable in different time classes and thereby the method is applicable to infer the possible regulatory interactions present among the other genes of this network.

  10. RNA-seq analysis of clinical-grade osteochondral allografts reveals activation of early response genes

    NARCIS (Netherlands)

    Lin, Yang; Lewallen, Eric A.; Camilleri, Emily T.; Bonin, Carolina A.; Jones, Dakota L.; Dudakovic, Amel; Galeano-Garces, Catalina; Wang, Wei; Karperien, Marcel J.; Larson, Annalise N.; Dahm, Diane L.; Stuart, Michael J.; Levy, Bruce A.; Smith, Jay; Ryssman, Daniel B.; Westendorf, Jennifer J.; Im, Hee-Jeong; van Wijnen, Andre J.; Riester, Scott M.; Krych, Aaron J.

    2016-01-01

    Preservation of osteochondral allografts used for transplantation is critical to ensure favorable outcomes for patients after surgical treatment of cartilage defects. To study the biological effects of protocols currently used for cartilage storage, we investigated differences in gene expression

  11. Transcription and replication result in distinct epigenetic marks following repression of early gene expression

    OpenAIRE

    Kallestad, Les; Woods, Emily; Christensen, Kendra; Gefroh, Amanda; Balakrishnan, Lata; Milavetz, Barry

    2013-01-01

    Simian Virus 40 (SV40) early transcription is repressed when the product of early transcription, T-antigen, binds to its cognate regulatory sequence, Site I, in the promoter of the SV40 minichromosome. Because SV40 minichromosomes undergo replication and transcription potentially repression could occur during active transcription or during DNA replication. Since repression is frequently epigenetically marked by the introduction of specific forms of methylated histone H3, we characterized th...

  12. Early stages of functional diversification in the Rab GTPase gene family revealed by genomic and localization studies in Paramecium species.

    Science.gov (United States)

    Bright, Lydia J; Gout, Jean-Francois; Lynch, Michael

    2017-04-15

    New gene functions arise within existing gene families as a result of gene duplication and subsequent diversification. To gain insight into the steps that led to the functional diversification of paralogues, we tracked duplicate retention patterns, expression-level divergence, and subcellular markers of functional diversification in the Rab GTPase gene family in three Paramecium aurelia species. After whole-genome duplication, Rab GTPase duplicates are more highly retained than other genes in the genome but appear to be diverging more rapidly in expression levels, consistent with early steps in functional diversification. However, by localizing specific Rab proteins in Paramecium cells, we found that paralogues from the two most recent whole-genome duplications had virtually identical localization patterns, and that less closely related paralogues showed evidence of both conservation and diversification. The functionally conserved paralogues appear to target to compartments associated with both endocytic and phagocytic recycling functions, confirming evolutionary and functional links between the two pathways in a divergent eukaryotic lineage. Because the functionally diversifying paralogues are still closely related to and derived from a clade of functionally conserved Rab11 genes, we were able to pinpoint three specific amino acid residues that may be driving the change in the localization and thus the function in these proteins. © 2017 Bright et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

  13. Properties of a herpes simplex virus multiple immediate-early gene-deleted recombinant as a vaccine vector

    International Nuclear Information System (INIS)

    Watanabe, Daisuke; Brockman, Mark A.; Ndung'u, Thumbi; Mathews, Lydia; Lucas, William T.; Murphy, Cynthia G.; Felber, Barbara K.; Pavlakis, George N.; Deluca, Neal A.; Knipe, David M.

    2007-01-01

    Herpes simplex virus (HSV) recombinants induce durable immune responses in rhesus macaques and mice and have induced partial protection in rhesus macaques against mucosal challenge with virulent simian immunodeficiency virus (SIV). In this study, we evaluated the properties of a new generation HSV vaccine vector, an HSV-1 multiple immediate-early (IE) gene deletion mutant virus, d106, which contains deletions in the ICP4, ICP27, ICP22, and ICP47 genes. Because several of the HSV IE genes have been implicated in immune evasion, inactivation of the genes encoding these proteins was expected to result in enhanced immunogenicity. The d106 virus expresses few HSV gene products and shows minimal cytopathic effect in cultured cells. When d106 was inoculated into mice, viral DNA accumulated at high levels in draining lymph nodes, consistent with an ability to transduce dendritic cells and activate their maturation and movement to lymph nodes. A d106 recombinant expressing Escherichia coli β-galactosidase induced durable β-gal-specific IgG and CD8 + T cell responses in naive and HSV-immune mice. Finally, d106-based recombinants have been constructed that express simian immunodeficiency virus (SIV) gag, env, or a rev-tat-nef fusion protein for several days in cultured cells. Thus, d106 shows many of the properties desirable in a vaccine vector: limited expression of HSV gene products and cytopathogenicity, high level expression of transgenes, ability to induce durable immune responses, and an ability to transduce dendritic cells and induce their maturation and migration to lymph nodes

  14. Copy number variations in "classical" obesity candidate genes are not frequently associated with severe early-onset obesity in children.

    Science.gov (United States)

    Windholz, Jan; Kovacs, Peter; Schlicke, Marina; Franke, Christin; Mahajan, Anubha; Morris, Andrew P; Lemke, Johannes R; Klammt, Jürgen; Kiess, Wieland; Schöneberg, Torsten; Pfäffle, Roland; Körner, Antje

    2017-05-01

    Obesity is genetically heterogeneous and highly heritable, although polymorphisms explain the phenotype in only a small proportion of obese children. We investigated the presence of copy number variations (CNVs) in "classical" genes known to be associated with (monogenic) early-onset obesity in children. In 194 obese Caucasian children selected for early-onset and severe obesity from our obesity cohort we screened for deletions and/or duplications by multiplex ligation-dependent probe amplification reaction (MLPA). As we found one MLPA probe to interfere with a polymorphism in SIM1 we investigated its association with obesity and other phenotypic traits in our extended cohort of 2305 children. In the selected subset of most severely obese children, we did not find CNV with MLPA in POMC, LEP, LEPR, MC4R, MC3R or MC2R genes. However, one SIM1 probe located at exon 9 gave signals suggestive for SIM1 insufficiency in 52 patients. Polymerase chain reaction (PCR) analysis identified this as a false positive result due to interference with single nucleotide polymorphism (SNP) rs3734354/rs3734355. We, therefore, investigated for associations of this polymorphism with obesity and metabolic traits in our extended cohort. We found rs3734354/rs3734355 to be associated with body mass index-standard deviation score (BMI-SDS) (p = 0.003), but not with parameters of insulin metabolism, blood pressure or food intake. In our modest sample of severely obese children, we were unable to find CNVs in well-established monogenic obesity genes. Nevertheless, we found an association of rs3734354 in SIM1 with obesity of early-onset type in children, although not with obesity-related traits.

  15. Microarray and RT-PCR screening for white spot syndrome virus immediate-early genes in cycloheximide-treated shrimp

    International Nuclear Information System (INIS)

    Liu Wangjing; Chang Yunshiang; Wang Chunghsiung; Kou, Guang-Hsiung; Lo Chufang

    2005-01-01

    Here, we report for the first time the successful use of cycloheximide (CHX) as an inhibitor to block de novo viral protein synthesis during WSSV (white spot syndrome virus) infection. Sixty candidate IE (immediate-early) genes were identified using a global analysis microarray technique. RT-PCR showed that the genes corresponding to ORF126, ORF242 and ORF418 in the Taiwan isolate were consistently CHX-insensitive, and these genes were designated ie1, ie2 and ie3, respectively. The sequences for these IE genes also appear in the two other WSSV isolates that have been sequenced. Three corresponding ORFs were identified in the China WSSV isolate, but only an ORF corresponding to ie1 was predicted in the Thailand isolate. In a promoter activity assay in Sf9 insect cells using EGFP (enhanced green fluorescence protein) as a reporter, ie1 showed very strong promoter activity, producing higher EGFP signals than the insect Orgyia pseudotsugata multicapsid nuclear polyhedrosis virus (OpMNPV) ie2 promoter

  16. Cpt1a gene expression in peripheral blood mononuclear cells as an early biomarker of diet-related metabolic alterations

    KAUST Repository

    Diaz Rua, Ruben; Palou, Andreu; Oliver, Paula

    2016-01-01

    subjects at risk of developing diet-related diseases.Objective: We analysed PBMC expression of key energy homeostasis-related genes in a time-course analysis in order to find out early markers of metabolic alterations due to sustained intake of high-fat (HF) and highprotein (HP) diets.Design: We administered HF and HP diets (4 months) to adult Wistar rats in isocaloric conditions to a control diet, mainly to avoid overweight associated with the intake of hyperlipidic diets and, thus, to be able to characterise markers of metabolically obese normal-weight (MONW) syndrome. PBMC samples were collected at different time points of dietary treatment and expression of relevant energy homeostatic genes analysed by real-time reverse transcription-polymerase chain reaction. Serum parameters related with metabolic syndrome, as well as fat deposition in liver, were also analysed.Results: The most outstanding results were those obtained for the expression of the lipolytic gene carnitine palmitoyltransferase 1a (Cpt1a). Cpt1a expression in PBMC increased after only 1 month of exposure to both unbalanced diets, and this increased expression was maintained thereafter. Interestingly, in the case of the HF diet, Cpt1a expression was altered even in the absence of increased body weight but correlated with alterations such as higher insulin resistance, alteration of serum lipid profile and, particularly, increased fat deposition in liver, a feature characteristic of metabolic syndrome, which was even observed in animals fed with HP diet.Conclusions: We propose Cpt1a gene expression analysis in PBMC as an early biomarker of metabolic alterations associated with MONW phenotype due to the intake of isocaloric HF diets, as well as a marker of increased risk of metabolic diseases

  17. Differential gene expression profile reveals deregulation of pregnancy specific β1 glycoprotein 9 early during colorectal carcinogenesis

    Directory of Open Access Journals (Sweden)

    Gallinger Steven

    2005-06-01

    Full Text Available Abstract Background APC (Adenomatous polyposis coli plays an important role in the pathogenesis of both familial and sporadic colorectal cancer. Patients carrying germline APC mutations develop multiple colonic adenomas at younger age and higher frequency than non-carrier cases which indicates that silencing of one APC allele may be sufficient to initiate the transformation process. Methods To elucidate the biological dysregulation underlying adenoma formation we examined global gene expression profiles of adenomas and corresponding normal mucosa from an FAP patient. Differential expression of the most significant gene identified in this study was further validated by mRNA in situ hybridization, reverse transcriptase PCR and Northern blotting in different sets of adenomas, tumours and cancer cell lines. Results Eighty four genes were differentially expressed between all adenomas and corresponding normal mucosa, while only seven genes showed differential expression within the adenomas. The first group included pregnancy specific β-1 glycoprotein 9 (PSG9 (p PSG9 is a member of the carcinoembryonic antigen (CEA/PSG family and is produced at high levels during pregnancy, mainly by syncytiotrophoblasts. Further analysis of sporadic and familial colorectal cancer confirmed that PSG9 is ectopically upregulated in vivo by cancer cells. In total, deregulation of PSG9 mRNA was detected in 78% (14/18 of FAP adenomas and 75% (45/60 of sporadic colorectal cancer cases tested. Conclusion Detection of PSG9 expression in adenomas, and at higher levels in FAP cases, indicates that germline APC mutations and defects in Wnt signalling modulate PSG9 expression. Since PSG9 is not found in the non-pregnant adult except in association with cancer, and it appears to be an early molecular event associated with colorectal cancer monitoring of its expression may be useful as a biomarker for the early detection of this disease.

  18. Cpt1a gene expression in peripheral blood mononuclear cells as an early biomarker of diet-related metabolic alterations

    Directory of Open Access Journals (Sweden)

    Rubén Díaz-Rúa

    2016-11-01

    Full Text Available Background: Research on biomarkers that provide early information about the development of future metabolic alterations is an emerging discipline. Gene expression analysis in peripheral blood mononuclear cells (PBMC is a promising tool to identify subjects at risk of developing diet-related diseases. Objective: We analysed PBMC expression of key energy homeostasis-related genes in a time-course analysis in order to find out early markers of metabolic alterations due to sustained intake of high-fat (HF and high-protein (HP diets. Design: We administered HF and HP diets (4 months to adult Wistar rats in isocaloric conditions to a control diet, mainly to avoid overweight associated with the intake of hyperlipidic diets and, thus, to be able to characterise markers of metabolically obese normal-weight (MONW syndrome. PBMC samples were collected at different time points of dietary treatment and expression of relevant energy homeostatic genes analysed by real-time reverse transcription-polymerase chain reaction. Serum parameters related with metabolic syndrome, as well as fat deposition in liver, were also analysed. Results: The most outstanding results were those obtained for the expression of the lipolytic gene carnitine palmitoyltransferase 1a (Cpt1a. Cpt1a expression in PBMC increased after only 1 month of exposure to both unbalanced diets, and this increased expression was maintained thereafter. Interestingly, in the case of the HF diet, Cpt1a expression was altered even in the absence of increased body weight but correlated with alterations such as higher insulin resistance, alteration of serum lipid profile and, particularly, increased fat deposition in liver, a feature characteristic of metabolic syndrome, which was even observed in animals fed with HP diet. Conclusions: We propose Cpt1a gene expression analysis in PBMC as an early biomarker of metabolic alterations associated with MONW phenotype due to the intake of isocaloric HF diets, as

  19. Depletion of Key Meiotic Genes and Transcriptome-Wide Abiotic Stress Reprogramming Mark Early Preparatory Events Ahead of Apomeiotic Transition

    Directory of Open Access Journals (Sweden)

    Jubin N Shah

    2016-10-01

    Full Text Available Molecular dissection of apomixis - an asexual reproductive mode - is anticipated to solve the enigma of loss of meiotic sex, and to help fixing elite agronomic traits. The Brassicaceae genus Boechera comprises of both sexual and apomictic species, permitting comparative analyses of meiotic circumvention (apomeiosis and parthenogenesis. Whereas previous studies reported local transcriptome changes during these events, it remained unclear whether global changes associated with hybridization, polyploidy and environmental adaptation that arose during evolution of Boechera might hint as (epigenetic regulators of early development prior apomictic initiation. To identify these signatures during vegetative stages, we compared seedling RNA-seq transcriptomes of an obligate triploid apomict and a diploid sexual, both isolated from a drought-prone habitat. Uncovered were several genes differentially expressed between sexual and apomictic seedlings, including homologues of meiotic genes ASYNAPTIC 1 (ASY1 and MULTIPOLAR SPINDLE 1 (MPS1 that were down-regulated in apomicts. An intriguing class of apomict-specific deregulated genes included several NAC transcription factors, homologues of which are known to be transcriptionally reprogrammed during abiotic stress in other plants. Deregulation of both meiotic and stress-response genes during seedling stages might possibly be important in preparation for meiotic circumvention, as similar transcriptional alteration was discernible in apomeiotic floral buds too. Furthermore, we noted that the apomict showed better tolerance to osmotic stress in vitro than the sexual, in conjunction with significant upregulation of a subset of NAC genes. In support of the current model that DNA methylation epigenetically regulates stress, ploidy, hybridization and apomixis, we noted that ASY1, MPS1 and NAC019 homologues were deregulated in Boechera seedlings upon DNA demethylation, and ASY1 in particular seems to be repressed by

  20. Gene Expression Profiling of Early Stage Non-Small Cell Lung Cancer

    NARCIS (Netherlands)

    J. Hou (Jun)

    2010-01-01

    textabstractNSCLC is a highly heterogeneous malignancy with a poor prognosis. Treatment for NSCLC is currently based on a combination of pathological staging and histological classification. Recently, gene expression-based NSCLC profiling is proven a superior approach to stratify cancer cases with

  1. Early results of sarcomeric gene screening from the Egyptian National BA-HCM Program.

    Science.gov (United States)

    Kassem, Heba Sh; Azer, Remon S; Saber-Ayad, Maha; Ayad, Maha S; Moharem-Elgamal, Sarah; Magdy, Gehan; Elguindy, Ahmed; Cecchi, Franco; Olivotto, Iacopo; Yacoub, Magdi H

    2013-02-01

    The present study comprised sarcomeric genotyping of the three most commonly involved sarcomeric genes: MYBPC3, MYH7, and TNNT2 in 192 unrelated Egyptian hypertrophic cardiomyopathy (HCM) index patients. Mutations were detected in 40 % of cases. Presence of positive family history was significantly (p=0.002) associated with a higher genetic positive yield (49/78, 62.8 %). The majority of the detected mutations in the three sarcomeric genes were novel (40/62, 65 %) and mostly private (47/62, 77 %). Single nucleotide substitution was the most frequently detected mutation type (51/62, 82 %). Over three quarters of these substitutions (21/27, 78 %) involved CpG dinucleotide sites and resulted from C>T or G>A transition in the three analyzed genes, highlighting the significance of CpG high mutability within the sarcomeric genes examined. This study could aid in global comparative studies in different ethnic populations and constitutes an important step in the evolution of the integrated clinical, translational, and basic science HCM program.

  2. The predictive value of the 70-gene signature for adjuvant chemotherapy in early breast cancer

    NARCIS (Netherlands)

    Knauer, Michael; Mook, Stella; Rutgers, Emiel J. T.; Bender, Richard A.; Hauptmann, Michael; van de Vijver, Marc J.; Koornstra, Rutger H. T.; Bueno-de-Mesquita, Jolien M.; Linn, Sabine C.; van 't Veer, Laura J.

    2010-01-01

    Multigene assays have been developed and validated to determine the prognosis of breast cancer. In this study, we assessed the additional predictive value of the 70-gene MammaPrint signature for chemotherapy (CT) benefit in addition to endocrine therapy (ET) from pooled study series. For 541

  3. A novel whole exon deletion in WWOX gene causes early epilepsy, intellectual disability and optic atrophy.

    Science.gov (United States)

    Ben-Salem, Salma; Al-Shamsi, Aisha M; John, Anne; Ali, Bassam R; Al-Gazali, Lihadh

    2015-05-01

    Recent studies have implicated the WW domain-containing oxidoreductase encoding gene (WWOX) in a severe form of autosomal recessive neurological disorder. This condition showed an overlapping spectrum of clinical features including spinocerebellar ataxia associated with generalized seizures and delayed psychomotor development to growth retardation, spasticity, and microcephaly. We evaluated a child from a consanguineous Emirati family that presented at birth with growth retardation, microcephaly, epileptic seizures, and later developed spasticity and delayed psychomotor development. Screening for deletions and duplications using whole-chromosomal microarray analysis identified a novel homozygous microdeletion encompassing exon 5 of the WWOX gene. Analysis of parental DNA indicated that this deletion was inherited from both parents and lies within a large region of homozygosity. Sanger sequencing of the cDNA showed that the deletion resulted in exon 5 skipping leading to a frame-shift and creating a premature stop codon at amino acid position 212. Quantification of mRNA revealed striking low level of WWOX expression in the child and moderate level of expression in the mother compared to a healthy control. To the best of our knowledge, this is the first homozygous germline structural variation in WWOX gene resulting in truncated transcripts that were presumably subject to NMD pathway. Our findings extend the clinical and genetic spectrum of WWOX mutations and support a crucial role of this gene in neurological development.

  4. Learning Motivation Mediates Gene-by-Socioeconomic Status Interaction on Mathematics Achievement in Early Childhood

    Science.gov (United States)

    Tucker-Drob, Elliot M.; Harden, K. Paige

    2012-01-01

    There is accumulating evidence that genetic influences on achievement are more pronounced among children living in higher socioeconomic status homes, and that these gene-by-environment interactions occur prior to children's entry into formal schooling. We hypothesized that one pathway through which socioeconomic status promotes genetic influences…

  5. High frequencies of antibiotic resistance genes in infants’ meconium and early fecal samples

    DEFF Research Database (Denmark)

    Gosalbes, M. J.; Vallès, Y.; Jiménez-Hernández, N.

    2016-01-01

    The gastrointestinal tract (GIT) microbiota has been identified as an important reservoir of antibiotic resistance genes (ARGs) that can be horizontally transferred to pathogenic species. Maternal GIT microbes can be transmitted to the offspring, and recent work indicates that such transfer start...

  6. Liver alpha-amylase gene expression as an early obesity biomarker.

    Science.gov (United States)

    Mojbafan, Marzieh; Afsartala, Zohreh; Amoli, Mahsa M; Mahmoudi, Mahdi; Yaghmaei, Parichehreh; Larijani, Bagher; Ebrahim-Habibi, Azadeh

    2017-04-01

    Obesity is a major health problem worldwide, for which preventive and therapeutic means are still needed. Alpha-amylase is a digestive enzyme whose inhibition has been targeted as a potential anti-obesity strategy. However, alpha-amylase gene expression has not been particularly attended to, and in contrast with pancreatic and salivary amylases, fewer studies have focused on liver alpha-amylase. The present study aimed at investigating the expression of alpha-amylase gene in obese and normal mice at RNA and protein level as well as acarbose effect on this gene expression in hepatocyte cell culture. Control and case groups were fed by normal mouse pellet and high-fat diet respectively, during 8 weeks. After this period, serum biochemical parameters including glucose, cholesterol, triglycerides, AST, ALT and alpha-amylase were assayed. Liver alpha-amylase gene was analyzed by real time PCR, and liver enzyme was assayed with Bernfeld and ELISA methods Hepatocyte cell culture derived from both group were also treated by acarbose and alpha-amylase activity and gene expression was analyzed by above mentioned methods. All biochemical factors showed an increase in obese mice, but the increase in ALT and AST were not statistically significant. Alpha-amylase levels were also increased in obese mice, both at RNA and protein level, while a decrease was seen in obese mice derived hepatocytes after acarbose treatment. Elevated liver alpha-amylase levels may be indicative of initial stages of obesity and the use of acarbose could be considered as a treatment of obesity which could be potentially effective at multiple levels. Copyright © 2016. Published by Elsevier Urban & Partner Sp. z o.o.

  7. Thalidomide induced early gene expression perturbations indicative of human embryopathy in mouse embryonic stem cells

    International Nuclear Information System (INIS)

    Gao, Xiugong; Sprando, Robert L.; Yourick, Jeffrey J.

    2015-01-01

    Developmental toxicity testing has traditionally relied on animal models which are costly, time consuming, and require the sacrifice of large numbers of animals. In addition, there are significant disparities between human beings and animals in their responses to chemicals. Thalidomide is a species-specific developmental toxicant that causes severe limb malformations in humans but not in mice. Here, we used microarrays to study transcriptomic changes induced by thalidomide in an in vitro model based on differentiation of mouse embryonic stem cells (mESCs). C57BL/6 mESCs were allowed to differentiate spontaneously and RNA was collected at 24, 48, and 72 h after exposure to 0.25 mM thalidomide. Global gene expression analysis using microarrays revealed hundreds of differentially expressed genes upon thalidomide exposure that were enriched in gene ontology (GO) terms and canonical pathways associated with embryonic development and differentiation. In addition, many genes were found to be involved in small GTPases-mediated signal transduction, heart development, and inflammatory responses, which coincide with clinical evidences and may represent critical embryotoxicities of thalidomide. These results demonstrate that transcriptomics in combination with mouse embryonic stem cell differentiation is a promising alternative model for developmental toxicity assessment. - Highlights: • Studied genomic changes in mouse embryonic stem cells upon thalidomide exposure • Identified gene expression changes that may represent thalidomide embryotoxicity • The toxicogenomic changes coincide well with known thalidomide clinical outcomes. • The mouse embryonic stem cell model is suitable for developmental toxicity testing. • The model has the potential for high-throughput screening of a multitude of compounds

  8. Thalidomide induced early gene expression perturbations indicative of human embryopathy in mouse embryonic stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Gao, Xiugong, E-mail: xiugong.gao@fda.hhs.gov; Sprando, Robert L.; Yourick, Jeffrey J.

    2015-08-15

    Developmental toxicity testing has traditionally relied on animal models which are costly, time consuming, and require the sacrifice of large numbers of animals. In addition, there are significant disparities between human beings and animals in their responses to chemicals. Thalidomide is a species-specific developmental toxicant that causes severe limb malformations in humans but not in mice. Here, we used microarrays to study transcriptomic changes induced by thalidomide in an in vitro model based on differentiation of mouse embryonic stem cells (mESCs). C57BL/6 mESCs were allowed to differentiate spontaneously and RNA was collected at 24, 48, and 72 h after exposure to 0.25 mM thalidomide. Global gene expression analysis using microarrays revealed hundreds of differentially expressed genes upon thalidomide exposure that were enriched in gene ontology (GO) terms and canonical pathways associated with embryonic development and differentiation. In addition, many genes were found to be involved in small GTPases-mediated signal transduction, heart development, and inflammatory responses, which coincide with clinical evidences and may represent critical embryotoxicities of thalidomide. These results demonstrate that transcriptomics in combination with mouse embryonic stem cell differentiation is a promising alternative model for developmental toxicity assessment. - Highlights: • Studied genomic changes in mouse embryonic stem cells upon thalidomide exposure • Identified gene expression changes that may represent thalidomide embryotoxicity • The toxicogenomic changes coincide well with known thalidomide clinical outcomes. • The mouse embryonic stem cell model is suitable for developmental toxicity testing. • The model has the potential for high-throughput screening of a multitude of compounds.

  9. Methylation of Breast Cancer Predisposition Genes in Early-Onset Breast Cancer: Australian Breast Cancer Family Registry.

    Directory of Open Access Journals (Sweden)

    Cameron M Scott

    Full Text Available DNA methylation can mimic the effects of both germline and somatic mutations for cancer predisposition genes such as BRCA1 and p16INK4a. Constitutional DNA methylation of the BRCA1 promoter has been well described and is associated with an increased risk of early-onset breast cancers that have BRCA1-mutation associated histological features. The role of methylation in the context of other breast cancer predisposition genes has been less well studied and often with conflicting or ambiguous outcomes. We examined the role of methylation in known breast cancer susceptibility genes in breast cancer predisposition and tumor development. We applied the Infinium HumanMethylation450 Beadchip (HM450K array to blood and tumor-derived DNA from 43 women diagnosed with breast cancer before the age of 40 years and measured the methylation profiles across promoter regions of BRCA1, BRCA2, ATM, PALB2, CDH1, TP53, FANCM, CHEK2, MLH1, MSH2, MSH6 and PMS2. Prior genetic testing had demonstrated that these women did not carry a germline mutation in BRCA1, ATM, CHEK2, PALB2, TP53, BRCA2, CDH1 or FANCM. In addition to the BRCA1 promoter region, this work identified regions with variable methylation at multiple breast cancer susceptibility genes including PALB2 and MLH1. Methylation at the region of MLH1 in these breast cancers was not associated with microsatellite instability. This work informs future studies of the role of methylation in breast cancer susceptibility gene silencing.

  10. Maternal nutrient restriction in early gestation upregulates myogenic genes in cattle fetal muscle tissue

    Science.gov (United States)

    Prenatal myogenesis is a critical factor in determining the muscle growth potential of cattle. We hypothesized that maternal nutrient restriction during early gestation would alter the transcriptome of fetal primordial muscle tissue in cattle. A total of 14 Angus-cross heifers were estrus synchroniz...

  11. Reconstructing the early evolution of the fungi using a six gene phylogeny

    NARCIS (Netherlands)

    James, T.Y.; Kauff, F.; Schoch, C.L.; Matheny, P.B.; Hofstetter, V.; Cox, C.J.; Celio, G.; Gueidan, C.; Fraker, E.; Miadlikowska, J.; Lumbsch, H.T.; Rauhut, A.; Reeb, V.; Arnold, A.E.; Amtoft, A.; Stajich, J.E.; Hosaka, K.; Sung, G.H.; Johnson, D.; O'Rourke, B.; Binder, M.; Curtis, J.M.; Slot, J.C.; Wang, Z.; Wilson, A.W.; Schüßler, A.; Longcore, J.E.; O'Donnell, K.; Mozley-Standridge, S.; Porter, D.; Letcher, P.M.; Powell, M.J.; Taylor, J.W.; White, M.M.; Griffith, G.W.; Davies, D.R.; Sugiyama, J.; Rossman, A.Y.; Rogers, J.D.; Pfister, D.H.; Hewitt, D.; Hansen, K.; Hambleton, S.; Shoemaker, R.A.; Kohlmeyer, J.; Volkmann-Kohlmeyer, B.; Spotts, R.A.; Serdani, M.; Crous, P.W.; Hughes, K.W.; Matsuura, K.; Langer, E.; Langer, G.; Untereiner, W.A.; Lücking, R.; Büdel, B.; Geiser, D.M.; Aptroot, A.; Diederich, P.; Schmitt, I.; Schultz, M.; Yahr, R.; Hibbett, D.S.; Lutzoni, F.; McLaughlin, D.J.; Spatafora, J.W.; Vilgalys, R.

    2006-01-01

    The ancestors of fungi are believed to be simple aquatic forms with flagellated spores, similar to members of the extant phylum Chytridiomycota (chytrids). Current classifications assume that chytrids form an early-diverging clade within the kingdom Fungi and imply a single loss of the spore

  12. Gene expression profiling of porcine skeletal muscle in the early recovery phase following acute physical activity

    DEFF Research Database (Denmark)

    Hansen, Jeanette; Conley, Lene; Hedegaard, Jakob

    2012-01-01

    Acute physical activity elicits changes in gene expression in skeletal muscles to promote metabolic changes and to repair exercise-induced muscle injuries. In the present time-course study, pigs were submitted to an acute bout of treadmill running until near exhaustion to determine the impact...... associated with proteolytic events, such as the muscle-specific E3 ubiquitin ligase atrogin-1, were significantly upregulated, suggesting that protein breakdown, prevention of protein aggregation and stabilization of unfolded proteins are important processes for restoration of cellular homeostasis. We also...... detected an upregulation of genes that are associated with muscle cell proliferation and differentiation, including MUSTN1, ASB5 and CSRP3, possibly reflecting activation, differentiation and fusion of satellite cells to facilitate repair of muscle damage. In addition, exercise increased expression...

  13. The GCR2 gene family is not required for ABA control of seed germination and early seedling development in Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Jianjun Guo

    Full Text Available BACKGROUND: The plant hormone abscisic acid (ABA regulates diverse processes of plant growth and development. It has recently been proposed that GCR2 functions as a G-protein-coupled receptor (GPCR for ABA. However, the structural relationships and functionality of GCR2 have been challenged by several independent studies. A central question in this controversy is whether gcr2 mutants are insensitive to ABA, because gcr2 mutants were shown to display reduced sensitivity to ABA under one experimental condition (e.g. 22 degrees C, continuous white light with 150 micromol m(-2 s(-1 but were shown to display wild-type sensitivity under another slightly different condition (e.g. 23 degrees C, 14/10 hr photoperiod with 120 micromol m(-2 s(-1. It has been hypothesized that gcr2 appears only weakly insensitive to ABA because two other GCR2-like genes in Arabidopsis, GCL1 and GCL2, compensate for the loss of function of GCR2. PRINCIPAL FINDINGS: In order to test this hypothesis, we isolated a putative loss-of-function allele of GCL2, and then generated all possible combinations of mutations in each member of the GCR2 gene family. We found that all double mutants, including gcr2 gcl1, gcr2 gcl2, gcl1 gcl2, as well as the gcr2 gcl1 gcl2 triple mutant displayed wild-type sensitivity to ABA in seed germination and early seedling development assays, demonstrating that the GCR2 gene family is not required for ABA responses in these processes. CONCLUSION: These results provide compelling genetic evidence that GCR2 is unlikely to act as a receptor for ABA in the context of either seed germination or early seedling development.

  14. The role of early life growth development, the FTO gene and exclusive breastfeeding on child BMI trajectories.

    Science.gov (United States)

    Wu, Yan Yan; Lye, Stephen; Briollais, Laurent

    2017-10-01

    Recent studies have implicated the FTO gene in child and adult obesity. A longer duration of exclusive breastfeeding (EXBF) has been shown to reduce body mass index (BMI) and the risk of being overweight in the general population and among FTO gene carriers. However, it remains unclear whether the preventive effect of EXBF could be explained by its impact on early life growth development, e.g. ages at adiposity peak (AP) and adiposity rebound (AR) and BMI velocities in the first years of life, which are major determinants of overweight and obesity later in life. We studied 5590 children from the British Avon Longitudinal Study of Parents and Children (ALSPAC) cohort and modelled their longitudinal BMI profiles with mixed effects models from birth to 16 years of age, as well as their ages at AP, AR and BMI velocities in relation to the FTO gene variant and EXBF. A longer duration of EXBF (i.e. at least 5 months) has substantial impact on BMI growth trajectories among children carrying the FTO adverse variant by modulating the age at AP, age at AR and BMI velocities. EXBF acts antagonistically to the FTO rs9939609 risk allele and by the age of 15, the predicted reduction in BMI after 5 months of EXBF is 0.56 kg/m2 [95% confidence interval (CI) 0.11-1.01; P = 0.003] and 1.14 kg/m2 (95% CI 0.67-1.62; P < 0.0001) in boys and girls, respectively. EXBF influences early life growth development and thus plays a critical role in preventing the risks of overweight and obesity even when those are exacerbated by genetic factors. © The Author 2017; all rights reserved. Published by Oxford University Press on behalf of the International Epidemiological Association

  15. Early Phenylpropanoid Biosynthetic Steps in Cannabis sativa: Link between Genes and Metabolites

    Directory of Open Access Journals (Sweden)

    Immacolata Coraggio

    2013-06-01

    Full Text Available Phenylalanine ammonia-lyase (PAL, Cinnamic acid 4-hydroxylase (C4H and 4-Coumarate: CoA ligase (4CL catalyze the first three steps of the general phenylpropanoid pathway whereas chalcone synthase (CHS catalyzes the first specific step towards flavonoids production. This class of specialized metabolites has a wide range of biological functions in plant development and defence and a broad spectrum of therapeutic activities for human health. In this study, we report the isolation of hemp PAL and 4CL cDNA and genomic clones. Through in silico analysis of their deduced amino acid sequences, more than an 80% identity with homologues genes of other plants was shown and phylogenetic relationships were highlighted. Quantitative expression analysis of the four above mentioned genes, PAL and 4CL enzymatic activities, lignin content and NMR metabolite fingerprinting in different Cannabis sativa tissues were evaluated. Furthermore, the use of different substrates to assay PAL and 4CL enzymatic activities indicated that different isoforms were active in different tissues. The diversity in secondary metabolites content observed in leaves (mainly flavonoids and roots (mainly lignin was discussed in relation to gene expression and enzymatic activities data.

  16. Early phenylpropanoid biosynthetic steps in Cannabis sativa: link between genes and metabolites.

    Science.gov (United States)

    Docimo, Teresa; Consonni, Roberto; Coraggio, Immacolata; Mattana, Monica

    2013-06-28

    Phenylalanine ammonia-lyase (PAL), Cinnamic acid 4-hydroxylase (C4H) and 4-Coumarate: CoA ligase (4CL) catalyze the first three steps of the general phenylpropanoid pathway whereas chalcone synthase (CHS) catalyzes the first specific step towards flavonoids production. This class of specialized metabolites has a wide range of biological functions in plant development and defence and a broad spectrum of therapeutic activities for human health. In this study, we report the isolation of hemp PAL and 4CL cDNA and genomic clones. Through in silico analysis of their deduced amino acid sequences, more than an 80% identity with homologues genes of other plants was shown and phylogenetic relationships were highlighted. Quantitative expression analysis of the four above mentioned genes, PAL and 4CL enzymatic activities, lignin content and NMR metabolite fingerprinting in different Cannabis sativa tissues were evaluated. Furthermore, the use of different substrates to assay PAL and 4CL enzymatic activities indicated that different isoforms were active in different tissues. The diversity in secondary metabolites content observed in leaves (mainly flavonoids) and roots (mainly lignin) was discussed in relation to gene expression and enzymatic activities data.

  17. Cryptic Transcription and Early Termination in the Control of Gene Expression

    Directory of Open Access Journals (Sweden)

    Jessie Colin

    2011-01-01

    Full Text Available Recent studies on yeast transcriptome have revealed the presence of a large set of RNA polymerase II transcripts mapping to intergenic and antisense regions or overlapping canonical genes. Most of these ncRNAs (ncRNAs are subject to termination by the Nrd1-dependent pathway and rapid degradation by the nuclear exosome and have been dubbed cryptic unstable transcripts (CUTs. CUTs are often considered as by-products of transcriptional noise, but in an increasing number of cases they play a central role in the control of gene expression. Regulatory mechanisms involving expression of a CUT are diverse and include attenuation, transcriptional interference, and alternative transcription start site choice. This review focuses on the impact of cryptic transcription on gene expression, describes the role of the Nrd1-complex as the main actor in preventing nonfunctional and potentially harmful transcription, and details a few systems where expression of a CUT has an essential regulatory function. We also summarize the most recent studies concerning other types of ncRNAs and their possible role in regulation.

  18. Early Right Ventricular Apical Pacing-Induced Gene Expression Alterations Are Associated with Deterioration of Left Ventricular Systolic Function

    Directory of Open Access Journals (Sweden)

    Haiyan Xu

    2017-01-01

    Full Text Available The chronic high-dose right ventricular apical (RVA pacing may have deleterious effects on left ventricular (LV systolic function.