Influence of IL-6, IL-8, and TGF-β 1 gene polymorphisms on the risk of human papillomavirus-infection in women from Pernambuco, Brazil

Human papillomavirus (HPV) infections are strongly associated with the development of cervical intraepithelial neoplasias and invasive cervical cancer. Polymorphisms in cytokine-encoding genes and behavioural cofactors could play an important role in protecting an individual against viral infections and cancer. Here, we investigated whether IL-6 -174 G>C, IL-8 +396 G>T, and TGF-β1 +869 G>C and +915 G>C polymorphisms were associated with susceptibility to HPV infection in women from north-east (Pernambuco) Brazil. We analysed 108 healthy uninfected women (HC) and 108 HPV-positive women with cervical lesions. Genetic polymorphisms were assessed using Sanger sequencing and polymerase chain reaction-restriction fragment length polymorphism. Comparison of the distribution of the genotypic and allelic frequencies of the IL-18 +396 T>G polymorphism between HPV infected woman an uninfected controls showed that the GG genotype and G allele were both more frequent in the HC group, and were associated with protection from HPV infection (p = 0.0015; OR = 0.29 CI95% = 0.13-0.61; p = 0.0005; OR = 0.45 CI95% 0.29-0.7, respectively). Individuals from the control group could have previously had HPV infection that was spontaneously eliminated; however, it was undetectable at the time of sample collection. Based on our findings, we hypothesize that the IL-8 +396 G>T polymorphism could interfere with susceptibility to HPV infection, by modulating the ability of immune system to fight the virus.

CCR2 and CCR5 genes polymorphisms in women with cervical lesions from Pernambuco, Northeast Region of Brazil: a case-control study

Polymorphisms in chemokine receptors play an important role in the progression of cervical intraepithelial neoplasia (CIN) to cervical cancer (CC). Our study examined the association of CCR2-64I (rs1799864) andCCR5-Δ32 (rs333) polymorphisms with susceptibility to develop cervical lesion (CIN and CC) in a Brazilian population. The genotyping of 139 women with cervical lesions and 151 women without cervical lesions for the CCR2-64I and CCR5-Δ32 polymorphisms were performed using polymerase chain reaction-restriction fragment length polymorphism. The individuals carrying heterozygous or homozygous genotypes (GA+AA) for CCR2-64I polymorphisms seem to be at lower risk for cervical lesion [odds ratio (OR) = 0.37, p = 0.0008)]. The same was observed for the A allele (OR = 0.39, p = 0.0002), while no association was detected (p > 0.05) with CCR5-Δ32 polymorphism. Regarding the human papillomavirus (HPV) type, patients carrying the CCR2-64Ipolymorphism were protected against infection by HPV type 16 (OR = 0.35, p = 0.0184). In summary, our study showed a protective effect ofCCR2-64I rs1799864 polymorphism against the development of cervical lesions (CIN and CC) and in the susceptibility of HPV 16 infection.

Prevalência dos genótipos do papilomavírus humano: comparação entre três métodos de detecção em pacientes de Pernambuco, Brasil / Prevalence of human papillomavirus genotypes: comparison between three detection methods in patients of Pernambuco, Brazil

OBJETIVO: comparar três métodos para detecção do HPV e determinar a prevalência dos genótipos encontrados. MÉTODOS: um total de 120 amostras de raspagem da região cervical de mulheres portadoras de neoplasia intraepitelial cervical foram analisadas pela reação em cadeia da polimerase convencional, usando os sistemas de primers MY09/11, GP05+/06+ e pela Nested-PCR. As amostras foram submetidas à extração de DNA e, logo após, amplificadas com os primers GH20 e PC04 (β-globina) para verificação da qualidade do DNA obtido e pela reação em cadeia da polimerase convencional e Nested-PCR. Os fragmentos amplificados foram visualizados em gel de agarose a 1,2 por cento, corados com Blue Green Loading Dye I. As amostras positivas foram sequenciadas usando o sequenciador automático de DNA "MegaBACE 1000". Para análise estatística foram utilizados os teste do Χ2 e o de Fisher com nível de significância de 5 por cento. RESULTADOS: quinze amostras não se amplificaram para os primers de β-globina, sendo eliminadas do estudo. Das amostras restantes, 40 por cento (42/105) foram positivas para os primers MY09/11, 98 por cento (103/105) para os primers GP05+/06+ e 92 por cento (97/105) para Nested-PCR. Considerado as técnicas MY09/11 e GP05+/06+, foi possível observar 100 por cento de amostras positivas para o HPV. Neste estudo, a prevalência dos genótipos foi de 58, 23, 5, 4 e 3 por cento para HPVs 16, 18, 31, 33 e 56, respectivamente. Os HPV 67 e 83 apresentaram 2 por cento e os HPV 6, 11, 58 e candHPV85, 1 por cento cada. A prevalência dos genótipos neste estudo está de acordo com o reportado em todo o mundo (IC95 por cento=0,4657-0,8976). CONCLUSÕES: para obter resultados mais confiáveis, é necessário o uso de mais que um sistema de primers para detecção do HPV. Acredita-se que as três técnicas estudadas são importantes e adequadas para o diagnóstico clínico do HPV quando apropriadamente combinadas.

PURPOSE: to compare three methods for the detection of HPV infection and to determine the prevalence of the genotypes found. METHODS: a total of 120 cervical scrape samples from patients with cervical intraepithelial neoplasia were analyzed by the conventional polymerase chain reaction using the MY09/11 and GP05+/06+ primers, and by the Nested polymerase chain reaction. The samples were subjected to DNA amplification with the GH20 and PC04 primers (β-globin) to verify DNA quality and also by polymerase chain reaction and Nested polymerase chain reaction. The amplicons were visualized in 1.2 percent agarose gel stained with Blue Green Loading Dye I. Positive samples also were sequenced using the automatic DNA sequencer "MegaBACE 1000". The Χ2 and Fisher tests were used for statistical analysis with the level of significance set at 5 percent. RESULTS: fifteen samples were eliminated from the study because they failed to amplify the β-globin gene. Of the remaining samples, 40 percent (42/105) were positive using primers MY09/11, 98 percent (103/105) using primers GP05+/06+, and 92 percent (97/105) using Nested-PCR. With the MY09/11 and GP05+/06+ techniques, it was possible to obtain 100 percent HPV-positive samples. In this study, the prevalence of the genotypes found was 57, 23, 5, 4 and 3 percent for HPV genotypes 16, 18, 31, 33 and 56, respectively. HPVs 67 and 83 were present in 2 percent, and genotypes 6, 11, 58 and candHPV85 were present in 1 percent each. The prevalence of the more common genotypes (HPV 16 and 18) in this study agrees with that reported worldwide (IC95 percent=0.4657-0.8976). CONCLUSIONS: to obtain more reliable results, it is necessary the use of more than one primer system to detect HPV infections. We believe that the three techniques studied are important and suitable for the clinical diagnosis of HPV, when they are appropriately combined.

Immobilization of lipase from "Fusarium solani" FS1

Lipase from "Fusarium solani" FS1 was immobilized by covalent attachment to polyacrylamide beads and onto magnetized Dacron, retaining 12(per cent) and 97(per cent) of activity, respectively. Lipase was also entrapped within polyacrylamide beads, retaining 53(per cent) of activity. Investigations of the kinetic characteristics of the immobilized derivates using triolein as substrate showed that lipase immobilized onto polyacrilamide beads and Dracon did not follow Michaelis-Menten kinetics.

Production of extracellular lipase by the phytopathogenic fungus "Fusarium solani" FS1

A Brazilian strain of "Fusarium solani" was tested for extracellular lipase production in peptone-olive oil medium. The fungus produced 10,500 U.L(E-1) of lipase after 72 hours of cultivation at 25ºC in shake-flask at 120rpm in a medium containing 3(per cent) (w/v) peptone plus 0.5(per cent)(v/v) olive oil. Glucose (1(per cent) w/v) was found to inhibit the inductive effect of olive oil. Peptone concentrations below 3(per cent)(w/v) resulted in a reduced lipase production while increased olive oil concentration (above o.5(per cent)) did not further stimulate lipase production. The optimum lipase activity was achieved at pH 8.6 and 30ºC and a good enzyme stability (80(per cent) activity retention) was observed at pH ranging from 7.6 to 8.6, and the activity rapidly dropped at temperatures above 50ºC. Lipase activity was stimulated by addition of n-hexane to the culture medium supernatatnts, in contrast to incubation with water-soluble solvents