RESUMO
In human tumor cells, experimental and clinical evidence indicates that some factors involved in signal transduction and cell growth can also modulate the response to chemotherapeutic treatment. The aim of the present study was to investigate the role of folic acid (FA) as a modulator of carboplatin (CBDCA) activity. Genotoxicity and cytotoxicity induced by CBDCA alone and in combination with FA were assessed in cultured HeLa cells. We used comet assay, mitotic index analysis, MTT and NR assays, cytokinesis-block micronucleus cytome assay and annexin V-IP as different cytotoxicity and genotoxicity approaches for human cervical carcinoma cell line studies. The results showed that addition of 900nM FA together with 40.4mM CBDCA enhanced the activity of the platinum compound, increasing its effect on cell death by nearly 20%, as evidenced by the MTT and NR assays. Moreover, not only higher levels of DNA and chromosomal damage were reached but also the number of necrotic and apoptotic cells were significantly increased when cell cultures were treated with the combined procedure. This situation opens the possibility to explore the use of FA in platinum-based chemotherapy protocols to reduce the platinum doses for patient treatment and decrease the chance of developing the known side effects without losing biological activity.
Assuntos
Antineoplásicos/toxicidade , Carboplatina/toxicidade , Ácido Fólico/farmacologia , Mutagênicos/toxicidade , Apoptose/efeitos dos fármacos , Ensaio Cometa , Dano ao DNA , Sinergismo Farmacológico , Células HeLa , Humanos , Testes para Micronúcleos , Mitose/efeitos dos fármacosRESUMO
BACKGROUND: XRCC1 (X-ray repair cross-complementing group 1) plays a central role in the DNA base excision repair mechanism. Single nucleotide polymorphisms (SNPs) in the XRCC1 gene are thought to modulate DNA repair capacity and have been linked to cancer risk in several studies. MATERIALS AND METHODS: We conducted a case-control study comprising 217 cervical samples, including 103 cervical carcinomas and 114 normal tissue samples. Cervical samples were genotyped for two XRCC1 SNPs (Arg194Trp and Arg399Gln) by PCR-RFLPs. RESULTS: Subjects carrying heterozygous Arg399Gln or the combined Gln399Gln + Arg399Gln variant genotypes had a significantly reduced risk for cervical cancer development. In addition, the 194Arg-399Gln haplotype was also found to be associated with a decreased risk for cervical carcinoma. CONCLUSION: Our findings suggest that XRCC1 genotypes and haplotypes contribute in reducing the risk for cervical cancer development. Furthermore, genetic susceptibility conferred by Arg399Gln polymorphism operates independently of human papillomavirus infection of cervical tissue.
Assuntos
Proteínas de Ligação a DNA/genética , Predisposição Genética para Doença , Polimorfismo de Nucleotídeo Único , Neoplasias do Colo do Útero/genética , Adulto , Alelos , Argentina , Estudos de Casos e Controles , Feminino , Frequência do Gene , Genótipo , Haplótipos , Humanos , Pessoa de Meia-Idade , Papillomaviridae/genética , Infecções por Papillomavirus/genética , Risco , Neoplasias do Colo do Útero/virologia , Proteína 1 Complementadora Cruzada de Reparo de Raio-XRESUMO
Demodectic mange is a well-known parasitic skin disease characterized by the presence of a larger than normal number of Demodex mites (Demodex canis) in the skin of dogs. Recent research has suggested that major histocompatibility complex (MHC) class II expression is higher in the skin of dogs suffering from demodicosis than in normal ones. We have investigated whether canine Dog Leukocyte Antigen (DLA) class II alleles are associated with canine juvenile generalized demodicosis (JGD). In the present study, the analysis of microsatellite markers (FH2202, FH2975 and FH2054) linked to DLA was made in Boxer, Argentinean Mastiff and mixed breed dogs. DNA samples from 56 dogs affected with the disease and 60 breed-matched controls collected in Argentina were analysed. A highly significant association, in some of the analysed markers, in all breeds with the presence of demodicosis was observed with P < 0.05 and odds ratio (OR) > or =5. The results of this study suggest that an underlying DLA association exists with demodicosis in dogs and that this may represent an important immunological risk factor in the aetiology of this condition. This information could be used in the future to develop diagnostic tests to prevent canine JGD.
Assuntos
Doenças do Cão/genética , Predisposição Genética para Doença , Antígenos de Histocompatibilidade Classe I/genética , Repetições de Microssatélites/genética , Infestações por Ácaros/veterinária , Animais , Cães , Infestações por Ácaros/genéticaRESUMO
OBJECTIVE: The aim of the present work was to evaluate the presence of human papillomavirus genotypes in malignant and normal mucosa of the colon and rectum in order to determine if a relationship exists between HPV infection and colon neoplasms. MATERIALS AND METHODS: Thirty normal colon tissues and 54 sporadic adenocarcinomas were screened for HPV positivity using nested-PCR. Detection of viral types 6, 11, 16, 18, 33, 34 and 51 was performed by the LIS-SSCP (Low Ionic Strength-Single Strand Conformational Polymorphism) procedure. RESULTS: Significant differences in high risk HPV infection were found between normal samples and adenocarcinomas (P < 0.001). Among the cases, an inverse association between HPV infection and Dukes staging was also found (P = 0.020). Finally, there was no significant association between HPV and some classical clinicopathological features, although a gradient of infection form rectum to cecum was evident. CONCLUSION: The present study demonstrates that HPV may infect the glandular mucosa of the colon and suggests a possible association between HPV and colorectal cancer.
Assuntos
Adenocarcinoma/virologia , Neoplasias do Colo/virologia , Papillomaviridae/isolamento & purificação , Infecções por Papillomavirus/diagnóstico , Neoplasias Retais/virologia , Adenocarcinoma/genética , Idoso , Idoso de 80 Anos ou mais , Distribuição de Qui-Quadrado , Neoplasias do Colo/genética , DNA Viral/análise , Feminino , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Papillomaviridae/genética , Infecções por Papillomavirus/genética , Reação em Cadeia da Polimerase , Polimorfismo Conformacional de Fita Simples , Neoplasias Retais/genéticaRESUMO
Genital infection with human papillomavirus (HPV) is one of the most common sexually transmitted viral diseases. High risk HPV are now considered the main etiologic agent of cancer of the uterine cervix and their high-grade precursor lesions. The aim of the present study was to investigate the endemic HPV-genotype spectrum in a population of women from the city of La Plata, Argentina. With this purpose, 718 cervical scrapes or biopsies corresponding to 152 normal samples (Pap I/II), 84 samples classified as atypical squamous cells of undetermined significance (ASCUS), 100 condyloma, 279 low-grade squamous intraepithelial lesions (LGSIL), 82 high-grade squamous intraepithelial lesions (HGSIL), and 21 squamous cell carcinomas (SCC) were studied. The detection of HPV-DNA was performed by nested polymerase chain reaction, using My 09/11 and Gp 05/06. The viral genotypes were analyzed by single-stranded conformation polymorphisms, employing low ionic strength solution (LIS-SSCP). The overall prevalence of HPV infection was 75% in the analyzed population, with a frequency of 46% for normal cervix, 69% for ASCUS, 86% for condyloma, 80% for LGSIL, 98% for HGSIL and 100% for SCC. The most prevalent viral types were HPV 16 (35%), followed by HPV 6/11 (27% each one), HPV 33 (6%) and HPV 18 (5%). HPV 16 was the most prevalent viral type among women with LGSIL, HGSIL and SCC, representing 33%, 50% and 67% of the genital infections, respectively. HPV 6 and 11 were the most frequent viral types among samples classified as Pap I/II, ASCUS and condyloma. Women between 21 and 30 year old showed the highest prevalence of HPV positivity, compraising the 32.2% of total infections.
Assuntos
Carcinoma de Células Escamosas/virologia , Papillomaviridae/classificação , Infecções por Papillomavirus/virologia , Infecções Tumorais por Vírus/virologia , Neoplasias do Colo do Útero/virologia , Cervicite Uterina/virologia , Adolescente , Adulto , Idoso , Argentina/epidemiologia , Carcinoma de Células Escamosas/epidemiologia , Condiloma Acuminado/epidemiologia , Condiloma Acuminado/virologia , DNA Viral/genética , DNA Viral/isolamento & purificação , Feminino , Genótipo , Humanos , Pessoa de Meia-Idade , Papillomaviridae/genética , Papillomaviridae/isolamento & purificação , Infecções por Papillomavirus/epidemiologia , Prevalência , Infecções Tumorais por Vírus/epidemiologia , População Urbana , Neoplasias do Colo do Útero/epidemiologia , Cervicite Uterina/epidemiologia , Displasia do Colo do Útero/epidemiologia , Displasia do Colo do Útero/virologiaRESUMO
La infección genital por el virus papiloma humano (VPH) es la enfermedad de transmisión sexual de tipo viral más común en el mundo. Loas tipos virales de alto riesgo son considerados los agentes etiológicos del cáncer de cuello uterino. El objetivo de este estudio fue analizar los genotipos del VPH en un grupo de mujeres de la ciudad de La Plata, Argentina. Se estudiaron 718 hisopados y/o biopsias cervicales correspondientes a 152 muestras normales (Pap I/II), 84 muestras clasificadas como con atipías de significado incierto (ASCUS), 100 condilomas, 279 lesiones intraepiteliales de bajo grado (LGSIL), 82 lesiones intraepiteliales de alto grado (HGSIL) y 21 carcinomas de células escamosas (SCC). La detección del genoma viral se realizó por medio de la reacción en cadena de la polimerasa (PCR), utilizando un protocolo anidado con los cebadores My 09/11 y Gp 05/06. La genotipificación del VPH se realizó por medio de la técnica de análisis de polimorfismos en la conformación de cadenas simples, utilizando solución de siembra de baja fuerza iónica (LIS-SSCP). La prevalencia general de la infección fue 75 por ciento, con 46 por ciento de muestras ADN-VPH positivas para el grupo Pap I/II, 69 por ciento para las clasificadas como ASCUS, 86 por ciento para los condilomas, 80 por ciento para los LGSIL, 98 por ciento para los HGSIL y 100 por ciento para los carcinomas de células escamosas. Los tipos virales más prevalentes fueron VPH 16 (35 por ciento), VPH 6/11 (27 por ciento cada uno), VPH 33 (6 por ciento) y VPH 18 (5 por ciento). En el grupo de mujeres con LGSIL, HGSIL y SCC el tipo viral más prevalente fue VPH 16, representando el 33 por ciento, 50 por ciento y 67 por ciento de las infecciones, respectivamente. En las mujeres con Pap I/II, ASCUS y condilomas los tipos virales más frecuentes fueron VPH 6 y 11. El grupo etario con mayor prevalencia de VPH fue el comprendido por mujeres de 21 a 30 años, acumulando el 32,2 por ciento de las infecciones totales.
Assuntos
Humanos , Argentina , Genoma Viral , Papiloma , Reação em Cadeia da PolimeraseRESUMO
La infección genital por el virus papiloma humano (VPH) es la enfermedad de transmisión sexual de tipo viral más común en el mundo. Loas tipos virales de alto riesgo son considerados los agentes etiológicos del cáncer de cuello uterino. El objetivo de este estudio fue analizar los genotipos del VPH en un grupo de mujeres de la ciudad de La Plata, Argentina. Se estudiaron 718 hisopados y/o biopsias cervicales correspondientes a 152 muestras normales (Pap I/II), 84 muestras clasificadas como con atipías de significado incierto (ASCUS), 100 condilomas, 279 lesiones intraepiteliales de bajo grado (LGSIL), 82 lesiones intraepiteliales de alto grado (HGSIL) y 21 carcinomas de células escamosas (SCC). La detección del genoma viral se realizó por medio de la reacción en cadena de la polimerasa (PCR), utilizando un protocolo anidado con los cebadores My 09/11 y Gp 05/06. La genotipificación del VPH se realizó por medio de la técnica de análisis de polimorfismos en la conformación de cadenas simples, utilizando solución de siembra de baja fuerza iónica (LIS-SSCP). La prevalencia general de la infección fue 75 por ciento, con 46 por ciento de muestras ADN-VPH positivas para el grupo Pap I/II, 69 por ciento para las clasificadas como ASCUS, 86 por ciento para los condilomas, 80 por ciento para los LGSIL, 98 por ciento para los HGSIL y 100 por ciento para los carcinomas de células escamosas. Los tipos virales más prevalentes fueron VPH 16 (35 por ciento), VPH 6/11 (27 por ciento cada uno), VPH 33 (6 por ciento) y VPH 18 (5 por ciento). En el grupo de mujeres con LGSIL, HGSIL y SCC el tipo viral más prevalente fue VPH 16, representando el 33 por ciento, 50 por ciento y 67 por ciento de las infecciones, respectivamente. En las mujeres con Pap I/II, ASCUS y condilomas los tipos virales más frecuentes fueron VPH 6 y 11. El grupo etario con mayor prevalencia de VPH fue el comprendido por mujeres de 21 a 30 años, acumulando el 32,2 por ciento de las infecciones totales. (AU)
Assuntos
Humanos , Papiloma , Reação em Cadeia da Polimerase , Genoma Viral , ArgentinaRESUMO
The aim of the present study was to evaluate the relationship between viral type and copy number of human papillomavirus (HPV) with respect to the grade of cervical disease, and also to identify the existence of an HPV type-dependent viral load effect. DNA from 275 exocervical specimens, previously evaluated for histologic diagnosis, were evaluated for HPV presence, HPV type, and viral load. Viral load determination was performed using the low stringency PCR method (LS-PCR). Significant differences were found between the samples infected with HPV16 with respect to the samples infected with other 'high-risk' viral types (HPV -18, -31, -33 or -51) and 'low-risk' types (P < 0.05). However, highly significant differences were found between the viral loads observed in the high-grade squamous intraepithelial lesions group and normal epithelium (OR = 8.53) or the low grade ones (OR = 3.10). Moreover, a high viral load was detected in the condyloma acuminatum group compared to the normal epithelia samples (p< 0.05). This work confirms the genotype-specific association of viral load to the presence of HPV16. Also, a trend to higher viral loads could be seen in the more compromised cervical lesions. An unexpected level of viral particles appeared associated to the condylomas. This fact could be explained by a productive infection with high levels of viral replication.
Assuntos
Papillomaviridae/isolamento & purificação , Displasia do Colo do Útero/virologia , Neoplasias do Colo do Útero/virologia , Adulto , DNA Viral/análise , Feminino , Humanos , Papillomaviridae/classificação , Infecções por Papillomavirus/patologia , Infecções por Papillomavirus/virologia , Reação em Cadeia da Polimerase , Infecções Tumorais por Vírus/patologia , Infecções Tumorais por Vírus/virologia , Neoplasias do Colo do Útero/patologia , Carga Viral , Displasia do Colo do Útero/patologiaRESUMO
BACKGROUND: Helicobacter pylori infection is presumed to be the major causal agent of chronic active gastritis in humans. The persistent infection with this pathogen would be an important factor in the pathogenesis of peptic ulcer and also gastric cancer. METHODS: We investigated relationship between H. pylori characteristics in 42 patients with normal mucosa or gastritis with minor changes and 40 patients with mild and severe gastritis. Detection and typing of vacA and cagA genes were performed using a polymerase chain reaction method. RESULTS: The analysis of vacA prevalence and the type (S1 or S2) showed non-significant differences between the two groups studied (p > 0.05). However, cagA analysis showed highly significant differences between the groups classified as normal tissue-weak gastritis and mild-severe gastritis (p < 0.0001; OR = 8.4; CI = 3.1-22.8). CONCLUSIONS: cagA status is associated to the grade of gastritis, finding higher frequencies of H. pylori cagA+ in the moderate-severe gastritis group. These highly significant differences could make cagA status a genetic marker for disease progress.
Assuntos
Antígenos de Bactérias/genética , Proteínas de Bactérias/genética , Gastrite/microbiologia , Infecções por Helicobacter/microbiologia , Helicobacter pylori/genética , Reação em Cadeia da Polimerase , Adolescente , Adulto , Idoso , Antígenos de Bactérias/análise , Proteínas de Bactérias/análise , Proteínas de Bactérias/isolamento & purificação , Dispepsia/microbiologia , Dispepsia/patologia , Feminino , Gastrite/patologia , Marcadores Genéticos , Genótipo , Humanos , Mucosa Intestinal/microbiologia , Masculino , Pessoa de Meia-Idade , Estudos RetrospectivosRESUMO
BACKGROUND: Helicobacter pylori infection is presumed to be the major causal agent of chronic active gastritis in humans. The persistent infection with this pathogen would be an important factor in the pathogenesis of peptic ulcer and also gastric cancer. METHODS: We investigated relationship between H. pylori characteristics in 42 patients with normal mucosa or gastritis with minor changes and 40 patients with mild and severe gastritis. Detection and typing of vacA and cagA genes were performed using a polymerase chain reaction method. RESULTS: The analysis of vacA prevalence and the type (S1 or S2) showed non-significant differences between the two groups studied (p > 0.05). However, cagA analysis showed highly significant differences between the groups classified as normal tissue-weak gastritis and mild-severe gastritis (p < 0.0001; OR = 8.4; CI = 3.1-22.8). CONCLUSIONS: cagA status is associated to the grade of gastritis, finding higher frequencies of H. pylori cagA+ in the moderate-severe gastritis group. These highly significant differences could make cagA status a genetic marker for disease progress.
Assuntos
Humanos , Masculino , Feminino , Adolescente , Adulto , Pessoa de Meia-Idade , Antígenos de Bactérias/genética , Gastrite/microbiologia , Infecções por Helicobacter/microbiologia , Helicobacter pylori/genética , Reação em Cadeia da Polimerase , Antígenos de Bactérias/análise , Proteínas de Bactérias/genética , Proteínas de Bactérias/isolamento & purificação , Técnicas de Tipagem Bacteriana/métodos , DNA Bacteriano/análise , Dispepsia/microbiologia , Dispepsia/patologia , Mucosa Gástrica/microbiologia , Gastrite/patologia , Marcadores Genéticos , Genótipo , Helicobacter pylori/classificação , Helicobacter pylori/patogenicidade , Técnicas de Amplificação de Ácido Nucleico , Estudos Retrospectivos , VirulênciaRESUMO
BACKGROUND: Helicobacter pylori infection is presumed to be the major causal agent of chronic active gastritis in humans. The persistent infection with this pathogen would be an important factor in the pathogenesis of peptic ulcer and also gastric cancer. METHODS: We investigated relationship between H. pylori characteristics in 42 patients with normal mucosa or gastritis with minor changes and 40 patients with mild and severe gastritis. Detection and typing of vacA and cagA genes were performed using a polymerase chain reaction method. RESULTS: The analysis of vacA prevalence and the type (S1 or S2) showed non-significant differences between the two groups studied (p > 0.05). However, cagA analysis showed highly significant differences between the groups classified as normal tissue-weak gastritis and mild-severe gastritis (p < 0.0001; OR = 8.4; CI = 3.1-22.8). CONCLUSIONS: cagA status is associated to the grade of gastritis, finding higher frequencies of H. pylori cagA+ in the moderate-severe gastritis group. These highly significant differences could make cagA status a genetic marker for disease progress.(AU)
Assuntos
Humanos , Masculino , Feminino , Adolescente , Adulto , Pessoa de Meia-Idade , Idoso , Reação em Cadeia da Polimerase , Infecções por Helicobacter/microbiologia , Helicobacter pylori/genética , Gastrite/microbiologia , Antígenos de Bactérias/genética , Helicobacter pylori/classificação , Helicobacter pylori/patogenicidade , Técnicas de Tipagem Bacteriana/métodos , DNA Bacteriano/análise , Marcadores Genéticos , Virulência , Gastrite/patologia , Dispepsia/microbiologia , Dispepsia/patologia , Genótipo , Estudos Retrospectivos , Antígenos de Bactérias/análise , Mucosa Gástrica/microbiologia , Técnicas de Amplificação de Ácido Nucleico , Proteínas de Bactérias/genética , Proteínas de Bactérias/isolamento & purificaçãoRESUMO
Genital infection with human papillomavirus (HPV) is one of the most common sexually transmitted viral diseases. High risk HPV are now considered the main etiologic agent of cancer of the uterine cervix and their high-grade precursor lesions. The aim of the present study was to investigate the endemic HPV-genotype spectrum in a population of women from the city of La Plata, Argentina. With this purpose, 718 cervical scrapes or biopsies corresponding to 152 normal samples (Pap I/II), 84 samples classified as atypical squamous cells of undetermined significance (ASCUS), 100 condyloma, 279 low-grade squamous intraepithelial lesions (LGSIL), 82 high-grade squamous intraepithelial lesions (HGSIL), and 21 squamous cell carcinomas (SCC) were studied. The detection of HPV-DNA was performed by nested polymerase chain reaction, using My 09/11 and Gp 05/06. The viral genotypes were analyzed by single-stranded conformation polymorphisms, employing low ionic strength solution (LIS-SSCP). The overall prevalence of HPV infection was 75
in the analyzed population, with a frequency of 46
for normal cervix, 69
for ASCUS, 86
for condyloma, 80
for LGSIL, 98
for HGSIL and 100
for SCC. The most prevalent viral types were HPV 16 (35
), followed by HPV 6/11 (27
each one), HPV 33 (6
) and HPV 18 (5
). HPV 16 was the most prevalent viral type among women with LGSIL, HGSIL and SCC, representing 33
, 50
and 67
of the genital infections, respectively. HPV 6 and 11 were the most frequent viral types among samples classified as Pap I/II, ASCUS and condyloma. Women between 21 and 30 year old showed the highest prevalence of HPV positivity, compraising the 32.2
of total infections.
RESUMO
BACKGROUND: Helicobacter pylori infection is presumed to be the major causal agent of chronic active gastritis in humans. The persistent infection with this pathogen would be an important factor in the pathogenesis of peptic ulcer and also gastric cancer. METHODS: We investigated relationship between H. pylori characteristics in 42 patients with normal mucosa or gastritis with minor changes and 40 patients with mild and severe gastritis. Detection and typing of vacA and cagA genes were performed using a polymerase chain reaction method. RESULTS: The analysis of vacA prevalence and the type (S1 or S2) showed non-significant differences between the two groups studied (p > 0.05). However, cagA analysis showed highly significant differences between the groups classified as normal tissue-weak gastritis and mild-severe gastritis (p < 0.0001; OR = 8.4; CI = 3.1-22.8). CONCLUSIONS: cagA status is associated to the grade of gastritis, finding higher frequencies of H. pylori cagA+ in the moderate-severe gastritis group. These highly significant differences could make cagA status a genetic marker for disease progress.
RESUMO
c-erbB-2 gene amplification has been described in a variety of human cancers, but it has been poorly studied in noncancerous cytological samples from genital specimens positive for human papillomavirus (HPV). Furthermore, the relationship between this genetic event and the presence of high-risk and low-risk HPV types is poorly studied. Eighty-four noncancerous cytological samples from exocervical specimens that were positive for HPV types 6, 16, and 18 were analyzed for c-erbB-2 gene amplification using the genomic differential polymerase chain reaction with the single copy reference gene. An association between c-erbB-2 gene amplification and the group corresponding to HPV type 6 was found. Within the low-risk HPV group, c-erbB-2 amplification was associated to cervical intraepithelial neoplasia of grade I (CIN I). Because in the samples analyzed, most of the CIN I stage was characterized by a koilocytotic pattern, c-erbB-2 amplification could be related to this kind of cellular alteration. It would be important to study c-erbB-2 gene amplification and also gene expression in different CIN stages in order to determine its role and significance in cervical cancer.
Assuntos
Colo do Útero/virologia , Amplificação de Genes , Genes erbB-2/genética , Papillomaviridae/patogenicidade , Infecções por Papillomavirus/genética , Infecções Tumorais por Vírus/genética , Neoplasias do Colo do Útero/genética , Adulto , Transformação Celular Neoplásica , Feminino , Regulação da Expressão Gênica , Humanos , Papillomaviridae/isolamento & purificação , Infecções por Papillomavirus/patologia , Reação em Cadeia da Polimerase , Infecções Tumorais por Vírus/patologia , Neoplasias do Colo do Útero/virologia , Esfregaço Vaginal , Displasia do Colo do ÚteroRESUMO
Human exposure to metals is frequent due to their ubiquity, wide use in industry, and environmental persistence. Direct and indirect genotoxic effects of cadmium (Cd) and arsenic (As) were reported. However, the mechanisms of induction of genetic damage are not well known. The aim of the present work was to evaluate the degree of damage induced by Cd and As salts in a human lung fibroblasts cell line using the single cell gel electrophoresis assay (SCGE). MRC-5 cells were treated with cadmium chloride (CdCl(2)), cadmium sulfate (CdSO(4)), sodium arsenite (NaAsO(2)) and cacodylic acid (C(2)H(7)AsO(2)). A significant dose-dependent increment in the extent of DNA migration as well as in the percentage of cells with tails was observed (P<0.001) after treatment with CdSO(4) and NaAsO(2). Treatment with CdCl(2) induced a relatively low level of DNA strand breaks in comparison with that induced by CdSO(4). The increase migration observed with the three compounds could be originated either by the direct induction of DNA lesions or by the inhibition of excision repair mechanisms. On the other hand, cells treated with C(2)H(7)AsO(2) showed a decrease in the migration length with the three doses employed (P<0.001). The decrease in the rate of DNA migration could be a consequence of the induction of DNA cross-links by organic arsenicals.Cd and As salts induced DNA damage in fibroblast cells, detected as DNA migration in the single cell gel (SCG) assay. The distribution of DNA migration among cells as a function of dose revealed that the majority of exposed cells showed more DNA damage than cells obtained from control cultures. The potential for human exposure to both metals has been increased over the years due to the increment in their use. For this reason, elucidation of carcinogenic mechanisms is very important.
Assuntos
Arsênio/toxicidade , Compostos de Cádmio/toxicidade , Dano ao DNA , DNA/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Arsenitos/toxicidade , Ácido Cacodílico/toxicidade , Cloreto de Cádmio/toxicidade , Linhagem Celular , Ensaio Cometa , DNA/metabolismo , Relação Dose-Resposta a Droga , Fibroblastos/citologia , Fibroblastos/metabolismo , Humanos , Compostos de Sódio/toxicidade , Sulfatos/toxicidadeRESUMO
In the present study we used a simple and reliable method for HLA-DQA1 allele typing based on the single-stranded conformation polymorphism (SSCP) properties of DNA molecules obtained by PCR. The technique consists of PCR amplification of a DNA fragment comprising the second exon of the HLA-DQA1 gene, amplicon denaturation using a low ionic strength solution (LIS), and electrophoresis on a small native polyacrylamide gel, followed by a rapid silver staining procedure. In order to validate the technique and to obtain the allele patterns for the DQA1 gene, 50 cervical samples were typed using this methodology and the commercial Amplitype HLA DQA1 Amplification and Typing kit. All the alleles detected with the kit were characterized by the LIS-SSCP approach. This procedure proved to be useful for population screening and typing of the DQA1 gene as well as for detecting new alleles or mutations in the donor-recipient molecular matching of HLA class II genes.
Assuntos
Alelos , Antígenos HLA-DQ/genética , Reação em Cadeia da Polimerase/métodos , Eletroforese em Gel de Ágar , Genótipo , Cadeias alfa de HLA-DQ , Humanos , Concentração Osmolar , Polimorfismo Conformacional de Fita SimplesRESUMO
Although data on genetic alterations leading to development of colorectal cancer are abundant, no specific genetic alteration has been demonstrated for each class of tumor. The colorectal cancer phenotype is originated from an accumulation of different genetic alterations. The nature of these alterations, their order of appearance, and their associations vary greatly from one tumor to another, suggesting that the concept of a unique model of carcinogenesis is not applicable to these tumors. The aim of the present work was to study the association between K-ras and c-erbB-2 mutations and different clinicopathological variables in fifty-four samples from adenocarcinomas of the colon. The detection of K-ras activation was performed by specific enriched PCR. The genomic differential polymerase chain reaction with the single copy reference gene was employed for the detection of c-erbB-2 gene amplification. K-ras mutations were detected in 16 cases (29.63%) and c-erbB-2 amplifications in 1 sample (1.85%). Statistical analysis showed a significant association between K-ras codon 12 mutation frequency and Duke's stage B (p < 0.05). On the other hand, there was no association in relation to the other studied parameters. These results could indicate the occurrence of K-ras activation in early stages of the disease.
Assuntos
Adenocarcinoma/genética , Neoplasias do Colo/genética , Regulação Neoplásica da Expressão Gênica , Genes erbB-2/genética , Genes ras/genética , Adenocarcinoma/patologia , Neoplasias do Colo/patologia , Feminino , Amplificação de Genes , Humanos , Masculino , Mutação , Reação em Cadeia da PolimeraseRESUMO
In the present study we used a simple and reliable method for HLA-DQA1 allele typing based on the single-stranded conformation polymorphism (SSCP) properties of DNA molecules obtained by PCR. The technique consists of PCR amplification of a DNA fragment comprising the second exon of the HLA-DQA1 gene, amplicon denaturation using a low ionic strength solution (LIS), and electrophoresis on a small native polyacrylamide gel, followed by a rapid silver staining procedure. In order to validate the technique and to obtain the allele patterns for the DQA1 gene, 50 cervical samples were typed using this methodology and the commercial Amplitype® HLA DQA1 Amplification and Typing kit. All the alleles detected with the kit were characterized by the LIS-SSCP approach. This procedure proved to be useful for population screening and typing of the DQA1 gene as well as for detecting new alleles or mutations in the donor-recipient molecular matching of HLA class II genes
Assuntos
Humanos , Alelos , Antígenos HLA-DQ/genética , Eletroforese em Gel de Ágar , Genótipo , Concentração Osmolar , Reação em Cadeia da Polimerase , Polimorfismo Conformacional de Fita SimplesRESUMO
This study describes a fast and simple method for human papillomavirus (HPV) typing based on the polymerase chain reaction (PCR) amplification of a portion of the viral genome and single strand conformation polymorphism using low ionic strength solutions (LIS-SSCP). PCR products were obtained using My09/My11 and Gp5/Gp6 primers in a nested reaction. The band patterns corresponded to the plasmid HPV clones from HPV-6, -11, -16, -18, -31, -33 and -34. The SSCP minigels were stained with SYBR-Green II. In order to determine diagnostic applicability, 100 cervical samples were studied comprising liquid cytology and paraffin embedded biopsies from patients showing squamous intraepithelial lesions (SILs). The SSCP patterns obtained from the clinical samples and the HPV clones were similar when the same type was present. Therefore, the methodology proved to be efficient and with high reproducibility for the detection and typing of HPV in clinical samples.
Assuntos
Colo do Útero/virologia , Genoma Viral , Compostos Orgânicos , Papillomaviridae/isolamento & purificação , Infecções por Papillomavirus/virologia , Reação em Cadeia da Polimerase/métodos , Polimorfismo Conformacional de Fita Simples , Infecções Tumorais por Vírus/virologia , Biópsia , Carcinoma de Células Escamosas/virologia , Corantes , DNA de Neoplasias/isolamento & purificação , DNA Viral/isolamento & purificação , Feminino , Corantes Fluorescentes , Genótipo , Humanos , Soluções Hipotônicas , Desnaturação de Ácido Nucleico , Concentração Osmolar , Papillomaviridae/classificação , Papillomaviridae/genética , Inclusão em Parafina , Sensibilidade e Especificidade , Manejo de Espécimes , Neoplasias do Colo do Útero/virologia , Cervicite Uterina/virologia , Esfregaço Vaginal , Displasia do Colo do Útero/virologiaRESUMO
This study describes a fast and simple method for human papillomavirus (HPV) typing based on the polymerase chain reaction (PCR) amplification of a portion of the viral genome and single strand conformation polymorphism using low ionic strength solutions (LIS-SSCP). PCR products were obtained using My09/My11 and Gp5/Gp6 primers in a nested reaction. The band patterns corresponded to the plasmid HPV clones from HPV-6, -11, -16, -18, -31, -33 and -34. The SSCP minigels were stained with SYBR-Green II. In order to determine diagnostic applicability, 100 cervical samples were studied comprising liquid cytology and paraffin embedded biopsies from patients showing squamous intraepithelial lesions (SILs). The SSCP patterns obtained from the clinical samples and the HPV clones were similar when the same type was present. Therefore, the methodology proved to be efficient and with high reproducibility for the detection and typing of HPV in clinical samples.
