Soft tissue sarcoma: biology and therapeutic correlates

Soft tissue sarcomas (STS) comprise a heterogenenous group of greater than 50 malignancies of putative mesenchymal cell origin and as such they may arise in diverse tissue types in various anatomical locations throughout the whole body. Collectively they account for approximately 1% of all human malignancies yet have a spectrum of aggressive behaviours amongst their subtypes. They thus pose a particular challenge to manage and remain an under investigated group of cancers with no generally applicable new therapies in the past 40 years and an overall 5-year survival rate that remains stagnant at around 50%. From September 2000 to July 2006 I undertook a full time post-doctoral level research fellowship at the MD Anderson Cancer Center, Houston, Texas, USA in the department of Surgical Oncology to investigate the biology of soft tissue sarcoma and test novel anti- sarcoma adenovirus-based therapy in the preclinical nude rat model of isolated limb perfusion against human sarcoma xenografts. This work, in collaboration with colleagues as indicated herein, led to a number of publications in the scientific literature furthering our understanding of the malignant phenotype of sarcoma and reported preclinical studies with wild-type p53, in a replication deficient adenovirus vector, and oncolytic adenoviruses administered by isolated limb perfusion. Additional collaborative and pioneering preclinical studies reported the molecular imaging of sarcoma response to systemically delivered therapeutic phage RGD-4c AAVP. Doxorubicin chemotherapy is the single most active broadly applicable anti-sarcoma chemotherapeutic yet only has an approximate 30% overall response rate with additional breakthrough tumour progression and recurrence after initial chemo-responsiveness further problematic features in STS management. Doxorubicin is a substrate for the multi- drug resistance (mdr) gene product p-glycoprotein drug efflux pump and exerts its main mode of action by induction of DNA double-strand breaks during the S-phase of the cell cycle. Two papers in my thesis characterise different aspects of chemoresistance in sarcoma. The first shows that wild-type p53 suppresses Protein Kinase Calpha (PKCα) phosphorylation (and activation) of p-glycoprotein by transcriptional repression of PKCα through a Sp-1 transcription factor binding site in its -244/-234 promoter region. The second paper demonstrates that Rad51 (a central mediator of homologous recombination repair of double strand breaks) has elevated levels in sarcoma and particularly in the S- G2 phase of the cell cycle. Suppression of Rad51 with small interfering RNA in sarcoma cell culture led to doxorubicin chemosensitisation. Reintroduction of wild-type p53 into STS cell lines resulted in decreased Rad51 protein and mRNA expression via transcriptional repression of the Rad51 promoter through increased AP-2 binding. In light of poor response rates to chemotherapy, escape from local control portends a poor prognosis for patients with sarcoma. Two papers in my thesis characterise aspects of sarcoma angiogenesis, invasion and metastasis. Human sarcoma samples were found to have high levels of matrix metalloproteinase-9 (MMP-9) with expression levels that correlated with p53 mutational status. MMP-9 is known to degrade extracellular collagen, contribute to the control of the angiogenic switch necessary in primary tumour progression and facilitate invasion and metastasis. Reconstitution of wild-type p53 function led to decreased levels of MMP-9 protein and mRNA as well as zymography-assessed MMP-9 proteolytic activity and decreased tumour cell invasiveness. Reintroduction of wild-type p53 into human sarcoma xenografts in-vivo decreased tumour growth and MMP-9 protein expression. Wild-type p53 was found to suppress mmp-9 transcription via decreased binding of NF-κB to its -607/-595 mmp-9 promoter element. Studies on the role of the VEGF165 in sarcoma found that sarcoma cells stably transfected with VEGF165 formed more aggressive xenografted tumours with increased vascularity, growth rate, metastasis, and resistance to chemotherapy. Use of the anti-VEGFR2 antibody DC101 enhanced doxorubicin sensitivity at sub-conventional dosing, inhibited tumour growth, decreased development of metastases, and reduced tumour micro-vessel density while increasing the vessel maturation index. These effects were explained primarily through effects on endothelial cells (e.c.s), rather than the tumour cells per se, where DC101 induced e.c. sensitivity to doxorubicin and suppressed e.c. production of MMPs. The p53 tumour suppressor pathway is the most frequently mutated pathway in sarcoma. Recapitulation of wild-type p53 function in sarcoma exerts a number of anti-cancer outcomes such as growth arrest, resensitisation to chemotherapy, suppression of invasion, and attenuation of angiogenesis. Using a modified nude rat-human sarcoma xenograft model for isolated limb perfusion (ILP) delivery of wild-type p53 in a replication deficient adenovirus vector I showed that functionally competent wild-type p53 could be delivered to and detected in human leiomyosarcoma xenografts confirming preclinical feasibility - although not efficacious due to low transgene expression. Viral fibre modification to express the RGD tripeptide motif led to greater viral uptake by sarcoma cells in vitro (transductional targeting) and changing the transgene promoter to a response element active in cells with active telomerase expression restricted the transgene expression to the tumour intracellular environment (transcriptional targeting). Delivery of the fibre-modified, selectively replication proficient oncolytic adenovirus Ad.hTC.GFP/ E1a.RGD by ILP demonstrated a more robust, and tumour-restricted, transgene expression with evidence of anti-sarcoma effect confirmed microscopically. Collaborative studies using the fibre modified phage RGD-4C AAVP confirmed that systemic delivery specifically, efficiently, and repeatedly targets human sarcoma xenografts, binds to αv integrins in tumours, and demonstrates a durable, though heterogeneous, transgene expression of 1-4 weeks. Incorporation of the Herpes Simplex Virus thymidine kinase (HSVtk) transgene into RGD-4C AAVP permitted CT-PET spatial and temporal molecular imaging in vivo of transgene expression and allowed quantification of tumour metabolic activity both before and after interval administration of a systemic cytotoxic with predictable and measurable response to treatment before becoming apparent clinically. These papers further the medical and scientific community’s understanding of the biology of soft tissue sarcoma and report preclinical studies with novel and promising anti- sarcoma therapeutics.

Association of human papillomavirus and Chlamydia trachomatis with intraepithelial alterations in cervix samples

The influence of different infectious agents and their association with human papillomavirus (HPV) in cervical carcinogenesis have not been completely elucidated. This study describes the association between cytological changes in cervical epithelium and the detection of the most relevant aetiological agents of sexually transmitted diseases. Samples collected from 169 patients were evaluated by conventional cytology followed by molecular analysis to detect HPV DNA, Chlamydia trachomatis , herpes simplex virus 1 and 2, Neisseria gonorrhoeae , Mycoplasma genitalium , Trichomonas vaginalis, and Treponema pallidum , besides genotyping for most common high-risk HPV. An association between cytological lesions and different behavioural habits such as smoking and sedentariness was observed. Intraepithelial lesions were also associated with HPV and C. trachomatis detection. An association was also found between both simple and multiple genotype infection and cytological changes. The investigation of HPV and C. trachomatis proved its importance and may be considered in the future for including in screening programs, since these factors are linked to the early diagnosis of patients with precursor lesions of cervical cancer.

Cytomegalovirus and Langerhans Cell Histiocytosis: Is There a Link?

Background: Langerhans cell histiocytosis is a rare proliferative histiocytic disease of unknown etiology. Histologically, it is characterized by granuloma-like proliferation of Langerhans-type dendritic cells derived from bone marrow. Many investigators have suggested the possible role of viruses such as Epstein-Barr virus, human herpesvirus-6 (HHV-6), herpes simplex virus (HSV) types 1 and 2, and Cytomegalovirus in the pathogenesis of Langerhans cell histiocytosis. Objectives: In this study, we have investigated the presence of Cytomegalovirus in Langerhans cell histiocytosis in Iranian children. Patients and Methods: In this retrospective study, we have investigated the presence of Cytomegalovirus DNA expression, using paraffin-embedded tissue samples of 30 patients with Langerhans cell histiocytosis and 30 age and site-matched controls by qualitative Polymerase Chain Reaction (PCR) method. Results: No significant difference in prevalence of Cytomegalovirus presence between patients and controls was found. Cytomegalovirus was found by qualitative PCR in only 2 (6.66%) out of 30 patients and in 1 (3.3%) of 30 control samples with a P value of 1 (1.00 > 0.05) using chi-square test with OR: 2.07; 95% CI of OR: 0.18 - 24.15. Conclusions: Our findings do not support the hypothesis of a possible role for Cytomegalovirus in the pathogenesis of Langerhans cell histiocytosis.

Determination of antioxidant activity, phenolic contents and antiviral potential of methanol extract of Euphorbia spinidens (Euphorbiaceae)

Purpose: This study was aimed to evaluate the antioxidant activity of the methanol extract of Euphorbia spinidens (Euphorbiaceae) and its effect on Herpes simplex virus type-1 (HSV-1) replication. Methods: The methanol extract of aerial parts of E. spinidens collected from Khorasan State in North- Eastern part of Iran was used in this study. Total phenolic, flavonoid contents and the antioxidant activity were evaluated using Folin-Ciocalteu method, aluminum chloride colorimetric method and β- carotene-linoleate model system, respectively. Both the cytotoxic and antiviral effects of the crude extract on Vero cell line were determined by quantifying the viability of Vero cells using 3-(4,5- dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)2H-tetrazolium (MTS) assay. Results: Total phenolic and flavonoid contents of E.spinidens were 70 ± 1 mg of gallic acid equivalent/g of dry extract (mg GAE/g extract) and 49.66 ± 1.00 mg rutin equivalent/g of dry extract (mg RTN/g extract), respectively. Antioxidant activity was 44 ± 1 % compared with the standard, buthylated hydroxytuloene (BHT). The 50 % cytotoxic concentration (CC50) of the extract on Vero cells was 5.072 ± 0.063 mg/ml and its antiviral concentration of 50 % effectiveness (EC50) value was 0.34 ± 0.003 mg/ml. Conclusion: The findings of this study show that the methanol extract of E. spinidens has high content of phenolic and flavonoid compounds with good antioxidant activity. Furthermore, this extract has significant antiviral effect on HSV-1 probably due to the inhibition of viral replication.

The Herpesvirus Nuclear Egress Complex Component, UL31, is Recruited to Sites of DNA Damage Through Poly(ADP-RIBOSE) Binding

Thesis (Master, Biomedical & Molecular Sciences) -- Queen's University, 2016-08-23 15:03:30.807

Étude de la fonction anti-apoptotique de la sous-unité R1 de la ribonucléotide réductase des virus de l’herpès simplex

L’élimination des cellules infectées par apoptose constitue un mécanisme de défense antivirale. Les virus de l’herpès simplex (HSV) de type 1 et 2 encodent des facteurs qui inhibent l’apoptose induite par la réponse antivirale. La sous-unité R1 de la ribonucléotide réductase d’HSV-2 (ICP10) possède une fonction anti-apoptotique qui protège les cellules épithéliales de l’apoptose induite par les récepteurs de mort en agissant en amont ou au niveau de l’activation de la procaspase-8. Puisqu’une infection avec un mutant HSV-1 déficient pour la R1 diminue la résistance des cellules infectées vis à vis du TNFα, il a été suggéré que la R1 d’HSV-1 (ICP6) pourrait posséder une fonction anti-apoptotique. Le but principal de cette thèse est d’étudier le mécanisme et le potentiel de la fonction anti-apoptotique de la R1 d’HSV-1 et de la R1 d'HSV-2. Dans une première étude, nous avons investigué le mécanisme de la fonction anti-apoptotique de la R1 d’HSV en utilisant le TNFα et le FasL, deux inducteurs des récepteurs de mort impliqués dans la réponse immune anti-HSV. Cette étude a permis d’obtenir trois principaux résultats concernant la fonction anti-apoptotique de la R1 d’HSV. Premièrement, la R1 d’HSV-1 inhibe l’apoptose induite par le TNFα et par le FasL aussi efficacement que la R1 d’HSV-2. Deuxièmement, la R1 d’HSV-1 est essentielle à l’inhibition de l’apoptose induite par le FasL. Troisièmement, la R1 d’HSV interagit constitutivement avec la procaspase-8 d’une manière qui inhibe la dimérisation et donc l’activation de la caspase-8. Ces résultats suggèrent qu’en plus d’inhiber l’apoptose induite par les récepteurs de mort la R1 d’HSV peut prévenir l’activation de la caspase-8 induite par d’autres stimuli pro-apoptotiques. Les ARN double-brins (ARNdb) constituant un intermédiaire de la transcription du génome des HSV et activant l’apoptose par une voie dépendante de la caspase-8, nous avons testé dans une seconde étude l’impact de la R1 d’HSV sur l’apoptose induite par l’acide polyriboinosinique : polyribocytidylique (poly(I:C)), un analogue synthétique des ARNdb. Ces travaux ont montré qu’une infection avec les HSV protège les cellules épithéliales de l’apoptose induite par le poly(I:C). La R1 d’HSV-1 joue un rôle majeur dans l’inhibition de l’activation de la caspase-8 induite par le poly(I:C). La R1 d’HSV interagit non seulement avec la procaspase-8 mais aussi avec RIP1 (receptor interacting protein 1). En interagissant avec RIP1, la R1 d’HSV-2 inhibe l’interaction entre RIP1 et TRIF (Toll/interleukine-1 receptor-domain-containing adapter-inducing interferon β), l’adaptateur du Toll-like receptor 3 qui est un détecteur d’ARNdb , laquelle est essentielle pour signaler l’apoptose induite par le poly(I:C) extracellulaire et la surexpression de TRIF. Ces travaux démontrent la capacité de la R1 d’HSV à inhiber l’apoptose induite par divers stimuli et ils ont permis de déterminer le mécanisme de l’activité anti-apoptotique de la R1 d’HSV. Très tôt durant l’infection, cFLIP, un inhibiteur cellulaire de la caspase-8, est dégradé alors que la R1 d’HSV s’accumule de manière concomitante. En interagissant avec la procapsase-8 et RIP1, la R1 d’HSV se comporte comme un inhibiteur viral de l’activation de la procaspase-8 inhibant l’apoptose induite par les récepteurs de mort et les détecteurs aux ARNdb.

Prevention of recurrent herpes labialis outbreaks through low-intensity laser therapy: a clinical protocol with 3-year follow-up

Conselho Nacional de Pesquisa e Desnsenvolvimento Tecnologico (CNPq)

Epstein–Barr virus nuclear protein 2 interacts with p300, CBP, and PCAF histone acetyltransferases in activation of the LMP1 promoter

The Epstein–Barr virus (EBV) nuclear protein 2 (EBNA2) and herpes simplex virion protein 16 (VP16) acidic domains that mediate transcriptional activation now are found to have affinity for p300, CBP, and PCAF histone acetyltransferases (HATs). Transcriptionally inactive point mutations in these domains lack affinity for p300, CBP, or PCAF. P300 and CBP copurify with the principal HAT activities that bind to EBNA2 or VP16 acidic domains through velocity sedimentation and anion-exchange chromatography. EBNA2 binds to both the N- and C-terminal domains of p300 and coimmune-precipitates from transfected 293T cells with p300. In EBV-infected Akata Burkitt's tumor cells that do not express the EBV encoded oncoproteins EBNA2 or LMP1, p300 expression enhances the ability of EBNA2 to up-regulate LMP1 expression. Through its intrinsic HAT activity, PCAF can further potentiate the p300 effect. In 293 T cells, P300 and CBP (but not PCAF) can also coactivate transcription mediated by the EBNA2 or VP16 acidic domains and HAT-negative mutants of p300 have partial activity. Thus, the EBNA2 and VP16 acidic domains can utilize the intrinsic HAT or scaffolding properties of p300 to activate transcription.

Study of knowledge of genital herpes infection and attitudes to testing for genital herpes among antenatal clinic attendees

A descriptive survey of knowledge of genital herpes and attitudes to testing was conducted among antenatal clinic attendees at the Gold Coast Hospital, Australia. The study subjects showed a good knowledge of genital herpes, to a level that appears sufficient for an informed choice regarding herpes serology testing to be made. A preference for testing for genital herpes was suggested. Although serological testing is not routinely required, the results of the study indicate that discussion of genital herpes should be considered in the antenatal clinic, setting.

Complement factor C4 and immunoglobulins in recurrent or chronic mucosal infections

The study assessed whether plasma concentrations of complement factors C3, C4, or immunoglobulins, serum classical pathway hemolytyic activity, or polymorphisms in the class I and II HLA genes, isotypes and gene numbers of C4, or allotypes of IgG1 and IgG3 heavy chain genes were associated with severe frequently recurring or chronic mucosal infections. According to strict clinical criteria, 188 consecutive voluntary patients without a known immunodeficiency and 198 control subjects were recruited. Frequencies of low levels in IgG1, IgG2, IgG3 and IgG4 were for the first time tested from adult general population and patients with acute rhinosinusitis. Frequently recurring intraoral herpes simplex type 1 infections, a rare form of the disease, was associated with homozygosity in HLA -A*, -B*, -C*, and -DR* genes. Frequently recurrent genital HSV-2 infections were associated with low levels of IgG1 and IgG3, present in 54% of the recruited patients. This association was partly allotype-dependent. The G3mg,G1ma/ax haplotype, together with low IgG3, was more common in patients than in control subjects who lacked antibodies against herpes simplex viruses. This is the first found immunogenetic deficiency in otherwise healthy adults that predisposes to highly frequent mucosal herpes recurrences. According to previous studies, HSV effectively evades the allotype G1ma/ax of IgG1, whereas G3mg is associated with low IgG3. Certain HLA genes were more common in patients than in control subjects. Having more than one C4A or C4B gene was associated with neuralgias caused by the virus. Low levels of IgA, IgG1, IgG2, IgG3, and IgG4 were common in the general adult population, but even more frequent in patients with chronic sinusitis. Only low IgG1 was more common chronic than in acute rhinosinusitis. Clinically, nasal polyposis and bronchial asthma were associated with complicated disease forms. The best differentiating immunologic parameters were C4A deficiency and the combination of low plasma IgG4 together with low IgG1 or IgG2, performing almost equally. The lack of C4A, IgA, and IgG4, all known to possess anti-inflammatory activity, together with a concurrently impaired immunity caused by low subclass levels, may predispose to chronic disease forms. In severe chronic adult periodontitis, any C4A or C4B deficiency combined was associated with the disease. The new quantitative analysis of C4 genes and the conventional C4 allotyping method complemented each other. Lowered levels of plasma C3 or C4 or both, and serum CH50 were found in herpes and periodontitis patients. In rhinosinusitis, there was a linear trend with the highest levels found in the order: acute > chronic rhinosinusitis > general population > blood donors with no self-reported history of rhinosinusitis. Complement is involved in the defense against the tested mucosal infections. Seemingly immunocompetent patients with chronic or recurrent mucosal infections frequently have subtle weaknesses in different arms of immunity. Their susceptibility to chronic disease forms may be caused by these. Host s subtly impaired immunity often coincides with effective immune evasion from the same arms of immunity by the disease-causing pathogens. The interpretation of low subclass levels, if no additional predisposing immunologic factors are tested, is difficult and of limited value in early diagnosis and treatment.

Syndrome d'immunodéficience acquise (AIDS) révélé par un herpes génital sévère. A propos de deux cas.

The authors present 2 cases of AIDS revealed by severe recurrent genital herpes simplex. The patients are 2 young, previously healthy, African women without histories of homosexuality or drug abuse. The first patient died after 5 months of follow-up (post mortem findings: viral bronchopneumonia with positive cultures for herpes and cytomegalovirus (CMV), viral colitis due to CMV). The second patient survived. She has been treated, during the last 11 months, for filariasis, buccal and vaginal candidiasis and cerebral toxoplasmosis.

Étude des voies d’apprêtement des antigènes viraux menant à la présentation antigénique par les CMH de classe I

Le contrôle immunitaire des infections virales est effectué, en grande partie, par les lymphocytes T CD8+ cytotoxiques. Pour y parvenir, les lymphocytes T CD8+ doivent être en mesure de reconnaître les cellules infectées et de les éliminer. Cette reconnaissance des cellules infectées s’effectue par l’interaction du récepteur T (TCR) des lymphocytes T CD8+ et des peptides viraux associés au complexe majeur d’histocompatibilité (CMH) de classe I à la surface des cellules hôtes. Cette interaction constitue l’élément déclencheur permettant l’élimination de la cellule infectée. On comprend donc toute l’importance des mécanismes cellulaires menant à la génération des peptides antigéniques à partir des protéines virales produites au cours d’une infection. La vision traditionnelle de cet apprêtement protéique menant à la présentation d’antigènes par les molécules du CMH propose deux voies cataboliques distinctes. En effet, il est largement admis que les antigènes endogènes sont apprêtés par la voie dite ‘‘classique’’ de présentation antigénique par les CMH de classe I. Cette voie implique la dégradation des antigènes intracellulaires par le protéasome dans le cytoplasme, le transport des peptides résultant de cette dégradation à l’intérieur du réticulum endoplasmique, leur chargement sur les molécules du CMH de classe I et finalement le transport des complexes peptide-CMH à la surface de la cellule où ils pourront activer les lymphocytes T CD8+. Dans la seconde voie impliquant des antigènes exogènes, le dogme veut que ceux-ci soient apprêtés par les protéases du compartiment endovacuolaire. Les peptides ainsi générés sont directement chargés sur les molécules de CMH de classe II à l’intérieur de ce compartiment. Par la suite, des mécanismes de recyclage vésiculaire assurent le transport des complexes peptide-CMH de classe II à la surface de la cellule afin de stimuler les lymphocytes T CD4+. Cependant, cette stricte ségrégation des voies d’apprêtement antigénique a été durement éprouvée par la capacité des cellules présentatrices d’antigènes à effectuer l’apprêtement d’antigènes exogènes et permettre leur présentation sur des molécules de CMH de classe I. De plus, l’identification récente de peptides d’origine intracellulaire associés à des molécules de CMH de classe II a clairement indiqué la présence d’interactions entre les deux voies d’apprêtement antigénique permettant de transgresser le dogme préalablement établi. L’objectif du travail présenté ici était de caractériser les voies d’apprêtement antigénique menant à la présentation d’antigènes viraux par les molécules du CMH de classe I lors d’une infection par le virus de l’Herpès simplex de type I (HSV-1). Dans les résultats rapportés ici, nous décrivons une nouvelle voie d’apprêtement antigénique résultant de la formation d’autophagosomes dans les cellules infectées. Cette nouvelle voie permet le transfert d’antigènes viraux vers un compartiment vacuolaire dégradatif dans la phase tardive de l’infection par le virus HSV-1. Cette mise en branle d’une seconde voie d’apprêtement antigénique permet d’augmenter le niveau de présentation de la glycoprotéine B (gB) virale utilisée comme modèle dans cette étude. De plus, nos résultats décrivent la formation d’une nouvelle forme d’autophagosomes dérivés de l’enveloppe nucléaire en réponse à l’infection par le virus HSV-1. Ces nouveaux autophagosomes permettent le transfert d’antigènes viraux vers un compartiment vacuolaire lytique, action également assurée par les autophagosomes dits classiques. Dans la deuxième partie du travail présenté ici, nous utilisons l’infection par le virus HSV-1 et la production de la gB qui en résulte pour étudier le trafic membranaire permettant le transfert de la gB vers un compartiment vacuolaire dégradatif. Nos résultats mettent en valeur l’importance du réticulum endoplasmique, et des compartiments autophagiques qui en dérivent, dans ces mécanismes de transfert antigénique permettant d’amplifier la présentation antigénique de la protéine virale gB sur des CMH de classe I via une voie vacuolaire. L’ensemble de nos résultats démontrent également une étroite collaboration entre la voie classique de présentation antigénique par les CMH de classe I et la voie vacuolaire soulignant, encore une fois, la présence d’interaction entre les deux voies.

Prevalência da infecção pelo Papilomavírus humano, Chlamydia Trachomatis e Herpes Simples do tipo 2 em adolescentes atendidas em unidades de saúde pública de Natal

This study assessed the level of knowledge, attitude and practice of Pap smear and human papillomavirus (HPV), in addition to analyzing the prevalence of genital HPV infection, Herpes Simplex Type 2 (HSV-2) and Chlamydia trachomatis in teenagers. The study consisted of two approaches, one based only on interviews conducted with adolescents enrolled in public schools or in public health facilities in the city of Natal. The other approach involved only a group of 132 adolescents enrolled among those admitted to two health units in Natal-RN. This second group of participants two specimens were collected for laboratory analysis: one was directed to prepare the blade for the Pap test, and other processed for DNA extraction for molecular analysis, focusing on the detection of HPV, HSV-2 and C . trachomatis. The presence of DNA of the three pathogens was investigated by the technique of polymerase chain reaction (PCR). The presence of each of the three pathogens was analyzed in terms of socio-demographic characteristics, as well as sexual and reproductive activity to identify risk factors for infection and development of lesions of the uterine cervix. The results show that the adolescents in this study had levels of knowledge and attitude very low, both in relation to cytology to HPV as though they have made a reasonable percentage of adequate practice exam and prevention of HPV infection. The overall prevalence of HPV infection was 54.5% and 48.2% in adolescents with normal cytology and 86.4% in those with abnormal cytology. We observed a higher proportion of cases of infection in the age group of 18 to 21. The prevalence of HPV infection was slightly higher among pregnant teenagers. The overall prevalence of HSV-2 infection was 13.6% and 11.8% in women with normal cytology and 22.7% in those with abnormal cytology. A higher proportion of cases of infection was found in the age group from 14 to 17, with a slightly higher prevalence among pregnant women. The C. trachomatis was found with an overall prevalence of 19.7% and 21.8% in adolescents with normal cytology and 9.1% in those with abnormal cytology. The prevailing rate was highest in the age group 18 to 21 years and in nonpregnant

Antiviral activities of plants occurring in the state of Minas Gerais, Brazil: Part 2. Screening Bignoniaceae species

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Immunohistochemical approach to the pathogenesis of clinical cases of bovine Herpesvirus type 5 infections

Meningoencephalitis by Herpesvirus type 5 (BoHV-5) in cattle has some features that are similar to those of herpetic encephalitis in humans and other animal species. Human Herpesvirus 3 (commonly known as Varicella-zoster virus 1), herpes simplex viruses (HSV), and equid Herpesvirus 1 (EHV-1) induce an intense inflammatory, vascular and cellular response. In spite of the many reports describing the histological lesions associated with natural and experimental infections, the immunopathological mechanisms for the development of neurological disorder have not been established. A total of twenty calf brains were selected from the Veterinary School, University of São Paulo State, Araçatuba, Brazil, after confirmation of BoHV-5 infection by virus isolation as well as by a molecular approach. The first part of the study characterized the microscopic lesions associated with the brain areas in the central nervous system (CNS) that tested positive in a viral US9 gene hybridization assay. The frontal cortex (Fc), parietal cortex (Pc), thalamus (T) and mesencephalon (M) were studied. Secondly, distinct pathogenesis mechanisms that take place in acute cases were investigated by an immunohistochemistry assay. This study found the frontal cortex to be the main region where intense oxidative stress phenomena (AOP-1) and synaptic protein expression (SNAP-25) were closely related to inflammatory cuffs, satellitosis and gliosis, which represent the most frequently observed neurological lesions. Moreover, MMP-9 expression was shown to be localized in the leptomeninges, in the parenchyma and around mononuclear infiltrates (p < 0.0001). These data open a new perspective in understanding the role of the AOP-1, MMP-9 and SNAP-25 proteins in mediating BoHV-5 pathogenesis and the strategies of host-virus interaction in order to invade the CNS.