In vitro antiproliferative and apoptotic effects of thiosemicarbazones based on (-)-camphene and R-(+)-limonene in human melanoma cells.

A series of 38 thiosemicarbazone derivatives based on camphene and limonene were evaluated for their antiproliferative activity. Among them, 19 were synthesized and characterized using proton and carbon-13 nuclear magnetic resonance (1H and 13C NMR). For initial compound selection, human melanoma cells (SK-MEL-37) were exposed to a single concentration of a compound (100 µM) for 24, 48, and 72 hours, and cell detachment was visually observed. Cell viability was determined using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method. Nineteen compounds (4, 6, 8, 11, 13, 14, 15, 16, 17, 18, 20, 22, 25, 26, 31, 3', 4', 6', and 9') yielded cell viability below 20%. Subsequently, IC50 values for these compounds were determined, ranging from 11.56 to 55.38 µM, after 72 hours of treatment. Compound 17 (o-hydroxybenzaldehyde (-)-camphene-based thiosemicarbazone) demonstrated the lowest IC50 value, followed by compound 4 (benzaldehyde (-) camphene-based thiosemicarbazone) at 12.84 µM. Regarding compound 4, we observed the induction of a characteristic ladder pattern of DNA fragmentation through gel electrophoresis. Furthermore, fluorescence, flow cytometry and scanning microscopy assays revealed morphological changes consistent with apoptosis induction. Additionally, the measurement of caspase 6 and 8 activity in cellular extracts after treatment for 2, 4, 6, and 24 hours suggested the potential involvement of the extrinsic apoptosis pathway in the mechanism of action of compound 4. Further investigations, including molecular docking studies, are required to fully explore the potential of compound 4 and the other selected compounds, highlighting their promising role in future melanoma therapy research.

Circuit training reduces cardiometabolic risk factors in women / El entrenamiento en circuito reduce los factores de riesgo cardiometabólico en mujeres / Treinamento em circuito reduz os fatores de risco cardiometabólicos em mulheres

ABSTRACT Introduction Combined training is more effective than an isolated modality in reducing cardiometabolic risk indicators. Objective To evaluate the effect of circuit training volume on anthropometric and biochemical risk indicators for cardiometabolic diseases in overweight women. Methods Thirty-two participants underwent 24 weeks of circuit training with free weights combined with aerobic exercise. The training volume during the 24 weeks was used to distribute the women into moderate-volume physical activity (MVA), low-volume physical activity (LVA) and control (CON) groups. Anthropometric indices (body mass, body mass index (BMI), waist circumference (WC), waist-hip ratio (WHR)), blood glucose, insulin, insulin resistance (HOMA-IR), total cholesterol (TC), triglycerides, HDL-c, and LDL-c were evaluated at the beginning of the program and after 12 and 24 weeks. Results There was no interaction between training volume and time for any of the variables studied, but the intervention time influenced body mass (p=0.013) and BMI (p=0.012), and there was a tendency for participation time to reduce body mass (p=0.063) and BMI (p=0.062) after six months of intervention. The volume of the physical activity affected HDL-c (p=0.037), being significant (p=0.030) in the comparison between the MVA and CON groups. Additionally, there was a downward trend in HDL-c after six months of intervention (p=0.073), with a smaller reduction observed in the MVA group, indicating a protective role of moderate physical activity in the reduction of this lipid fraction. The association between physical activity volume and participation time resulted in a clinical improvement in total cholesterol (χ2 = 5.453, p = 0.02), with a higher probability of reaching clinically adequate values in the MVA group (OR = 0.126; 95%CI 0.019 - 0.827). Conclusion Training volume improved cardiometabolic risk factors in overweight women. Level of evidence II; Therapeutic Studies - Investigating the Results of Treatment.

RESUMEN Introducción El entrenamiento combinado es más eficiente que la modalidad aislada en indicadores de riesgo cardiometabólico. Objetivo Evaluar el efecto del volumen de entrenamiento en circuito sobre indicadores antropométricos y bioquímicos con riesgo de enfermedades cardiometabólicas en mujeres con sobrepeso. Métodos Treinta y dos participantes se sometieron a 24 semanas de entrenamiento en circuito con pesos libres combinados con ejercicio aeróbico. El volumen de entrenamiento durante las 24 semanas se utilizó para distribuir a las mujeres en los grupos: actividad física de volumen moderado (AVM), actividad física de volumen bajo (AVB) y control (CON). Se evaluaron los índices antropométricos masa corporal, índice de masa corporal (IMC), circunferencia de la cintura (CC), relación cintura-cadera (RCC), glucemia, insulina, resistencia a la insulina (HOMA-IR), colesterol total (CT), triglicéridos, HDL-c y LDL-c al inicio del programa y después de las semanas 12 y 24. Resultados No hubo interacción entre el volumen y el tiempo de entrenamiento para ninguna de las variables estudiadas, pero el tiempo de intervención influyó en la masa corporal (p=0,013) y en el IMC (p=0,012), y el tiempo de participación tendió a reducir la masa corporal (p=0,063) y el IMC (p=0,062), después de seis meses de intervención. El volumen de actividad física afectó al HDL-c (p =0,037), siendo significativo (p=0,030) en la comparación entre AVM y CON. Además, hubo una tendencia a la reducción del HDL-c después de seis meses de intervención (p=0,073), observándose la menor reducción en AVM, lo que indica el papel protector de la actividad física de volumen moderado en la reducción de esta fracción lipídica. La actividad física y el tiempo de participación mostraron una mejora clínica en colesterol total (χ2 = 5,453, p = 0,02), con mayor probabilidad de alcanzar valores clínicamente adecuados de AVM (OR = 0,126; IC95% 0,019 - 0,827). Conclusión El volumen de entrenamiento atenuó los factores de riesgo cardiometabólico en mujeres con sobrepeso. Nivel de Evidencia II; Estudios terapéuticos: investigación de los resultados del tratamiento.

ABSTRACT Introdução O treinamento combinado é mais eficiente do que a modalidade isolada com relação aos indicadores de risco cardiometabólico. Objetivos Avaliar o efeito do volume de treinamento em circuito sobre indicadores antropométricos e bioquímicos com risco de doenças cardiometabólicas em mulheres com excesso de peso. Métodos Trinta e duas participantes foram submetidas a 24 semanas de treinamento em circuito, com pesos livres combinados com exercício aeróbico. O volume de treinamento durante as 24 semanas foi utilizado para distribuir as mulheres nos grupos: atividade física de volume moderado (AVM), atividade física de baixo volume (AVB) e controle (CON). Os índices antropométricos massa corporal, índice de massa corporal (IMC), circunferência de cintura (CC), relação cintura-quadril (RCQ), glicemia, insulina, resistência à insulina (HOMA-IR), colesterol total (CT), triglicerídeos, HDL-c e LDL-c, foram avaliados no início do programa e depois de 12 e 24 semanas. Resultados Não houve interação entre o volume de treinamento e o tempo para nenhuma das variáveis estudadas, mas o tempo de intervenção influenciou a massa corporal (p = 0,013) e o IMC (p = 0,012), e o tempo de participação tendeu a reduzir a massa corporal (p = 0,063) e o IMC (p = 0,062), depois de seis meses de intervenção. O volume de atividade física afetou o HDL-c (p = 0,037), sendo significativo (p = 0,030) na comparação entre AVM e CON. Adicionalmente, verificou-se tendência de redução HDL-c depois seis meses de intervenção (p = 0,073), sendo a menor redução observada no AVM, que indica o papel protetor de atividade física de volume moderado na redução dessa fração lipídica. A associação entre o volume de atividade física e o tempo de participação mostrou melhora clínica do colesterol total (χ2= 5,453, p = 0,02), com maior probabilidade de atingir valores clinicamente adequados de AVM (OR = 0,126; IC de 95% 0,019 - 0,827). Conclusão O volume de treinamento atenuou os fatores de risco cardiometabólico em mulheres com excesso de peso. Nível de evidência II; Estudos terapêuticos - Investigação dos resultados do tratamento.

Curcumin supplementation reduces blood glucose and serum lipids of Brazilian women with high waist circumference: a randomized clinical trial

ABSTRACT Objective: To evaluate the effect of curcumin supplementation on the body compositions and biochemical parameters of Brazilian women with high waist circumferences. Materials and methods: This is a blind, randomized, placebo-controlled clinical trial conducted in 2016 with 35 Brazilian women with high waist circumference (WC). In total, 80 participants were randomized [38 in the placebo group (PG) and 42 in the supplemented group (SG)], but at the end of the protocol, 20 individuals in the PG and 15 in the SG were evaluated. The sample consumed one capsule of curcumin (500 mg/day) (Curcumin C3 Complex®) or a placebo for 90 days. Body weight, height, body mass index, WC, body fat, fat free mass, fasting glucose (FG), lipid profile [triglycerides (TGs), total cholesterol (TC), HDL-c and LDL-c], physical activity level and food intake (energy, carbohydrate, total fat and protein) data were evaluated before and after the intervention. Results: Curcumin supplementation reduced body mass (p < 0.040) but did not alter other anthropometric parameters or body composition (p ≥ 0.050). In relation to the biochemical profile, the SG saw reductions in FG (p < 0.001), TGs (p < 0.001) and TC (p = 0.001) compared with the PG. At the baseline and during the intervention, the practice of physical activity and food intake did not differ between the SG and PG (p ≥ 0.050). Conclusion: Curcumin supplementation improved the blood glucose and lipid profile of Brazilian women with high WC, without altering body composition. New studies with larger sample sizes and longer durations are important for identifying more robust data regarding the proposal of this work.

Expression of the cancer stem cell marker OCT4 is associated with worse prognosis and survival in cutaneous melanoma.

Cutaneous melanoma has an aggressive clinical presentation, showing rapid rate of growth and metastatic dissemination due to the permanence of cancer stem cells. The present study was to evaluate the expression of the self-renewal regulatory factor and the clinical significance of the transcription factor OCT4 in melanoma. Melanoma tissues were stained by immunohistochemistry and the correlation between the expression of this marker was determined through clinical-pathological variables and survival outcomes. Positive expression of nuclear and cytoplasmic OCT4 was observed in 49% and 41.2% of cases, respectively. The positive expression of nuclear OCT4 in melanoma was significantly associated with prognostic factors, such as Breslow depth, Clark's level, ulceration and metastasis. Survival of patients was 56% compared to positive nuclear OCT4 expression and 94.2% when compared to the low expression of the gene. Nuclear OCT4 positive genotype indicated aggressive tumor behavior with a worse clinical outcome, which indicates OCT4 as a useful biomarker in the prognosis of melanoma.

Endothelial nitric oxide concentrations in the saliva of jiu-jitsu athletes / Concentrações de óxido nítrico endotelial na saliva de atletas de jiu-jitsu / Concentraciones de óxido nítrico endotelial en la saliva de atletas de jiu-jitsu

ABSTRACT Introduction This article discusses the production of nitric oxide under the influence of sport-specific physical training, measured by the salivary nitrite of Jiu-Jitsu athletes. Objectives To verify the potential of the sport to produce optimal levels of nitric oxide stimulated by exertion, and to quantify training-related nitric oxide concentrations. Method The study participants were 14 volunteer athletes from the Tatame project (extension project), who were monitored for nine months in their training routine, providing samples of unstimulated saliva. Samples were collected each month, in three periods of the day: in the morning upon waking, immediately before training, and immediately after training. Salivary nitrite was quantified by the colorimetric Griess assay. Training heart rates were also monitored in order to establish training intensity. Results Mean monthly salivary nitrite levels showed a significant correlation with mean monthly heart rates, suggesting that salivary nitrite responds to training. However, salivary nitrite concentrations measured immediately after training were always lower than in the pre-training period. Conclusion The post-training reduction in concentrations was due to the nature of the sport studied, since because it involves a fight, the intense sympathetic stimulation inhibited salivary gland activity and irrigation, preventing salivary nitrite from producing an increase in circulating nitric oxide. Level of evidence IV; Case series.

RESUMO Introdução Este artigo discute a produção de óxido nítrico sob interferência do treinamento físico esportivo, medida pelo nitrito salivar em atletas de Jiu-Jitsu. Objetivos Verificar as potencialidades da modalidade na produção de níveis ideais de óxido nítrico estimulado pelo esforço e quantificar as concentrações de óxido nítrico relacionadas com o treinamento. Métodos O estudo teve a participação de 14 atletas voluntários do Projeto Tatame (projeto de extensão), que foram acompanhados por nove meses em sua rotina de treinamento e forneceram amostras de saliva não estimulada. Foram coletadas amostras a cada mês, em três períodos do dia: pela manhã ao acordar, imediatamente antes do treino e imediatamente após o treino. O nitrito salivar foi quantificado pelo método colorimétrico de Griess. Também foram monitoradas as frequências cardíacas de treinamento a fim de se estabelecer a intensidade do treinamento realizado. Resultados As médias mensais de nitrito salivar demonstraram correlação significativa com as médias mensais de frequência cardíaca, dando indícios de que o nitrito salivar responde ao treinamento. Contudo, as concentrações de nitrito salivar medidas imediatamente após o treinamento sempre eram menores com relação ao período anterior do treino. Conclusão A redução da concentração depois do treinamento deveu-se à natureza da modalidade estudada, visto que, por ser uma luta, a forte carga de estimulação simpática inibiu a atividade das glândulas salivares, bem como sua irrigação, impedindo que o nitrito salivar produzisse aumento do óxido nítrico circulante. Nível de evidência IV; Série de casos.

RESUMEN Introducción Este artículo analiza la producción de óxido nítrico bajo la influencia del entrenamiento físico deportivo, medida por el nitrito salival de atletas de Jiu-Jitsu. Objetivos Verificar el potencial de la modalidad en la producción de niveles ideales de óxido nítrico estimulado por el esfuerzo y cuantificar las concentraciones de óxido nítrico relacionadas con el entrenamiento. Métodos El estudio contó con la participación de 14 atletas voluntarios participantes del Proyecto Tatame (proyecto de extensión) a quienes se les dio seguimiento durante nueve meses en su rutina de entrenamiento y proporcionaron muestras de saliva no estimulada. Se recolectaron muestras cada mes, en tres períodos del día: en la mañana al despertar, inmediatamente antes del entrenamiento e inmediatamente después del entrenamiento. El nitrito salival se cuantific ó por el método colorimétrico de Griess. Las frecuencias cardíacas de entrenamiento se monitorearon para establecer la intensidad del entrenamiento realizado. Resultados Los promedios mensuales de nitrito salival mostraron una correlación significativa con los promedios mensuales de frecuencia cardíaca, lo que indica que el nitrito salival responde al entrenamiento. Sin embargo, las concentraciones de nitrito salival medidas inmediatamente después del entrenamiento siempre fueron más bajas en comparación con el período anterior al entrenamiento. Conclusión La reducción de la concentración después del entrenamiento se debió a la naturaleza de la modalidad estudiada, ya que, por ser una lucha, la fuerte carga de estimulación simpática inhibió la actividad de las glándulas salivales, así como su irrigación, evitando que el nitrito salival produjera aumento de óxido nítrico circulante. Nivel de evidencia IV; Serie de casos.

Waist Circumference Above 80 cm Predicts Increased Systolic Blood Pressure in Healthy Young Adult Women

Abstract Background: Among anthropometric measures for assessing adiposity-related risk, waist circumference (WC) is simple and fast to perform. Cut-off values for WC proposed by the International Diabetes Federation (IDF), and the Adult Treatment Panel III of the National Cholesterol Education Program (NCEP-ATP III) are categorized by gender and are not age-specific. Objective: To analyze the association between WC and cardiometabolic risk factors in adult women. Methods: A total of 164 healthy adult women were grouped by WC according to IDF and NCEP-ATP III cutoff values. Continuous variables were described as mean ± standard deviation or median (interquartile range). The Shapiro-Wilk test was used to assess the normality of data. Variables were analyzed by unpaired Student's t-test, Mann-Whitney U and Kruskal-Wallis tests. The correlation of WC categories with systolic (SBP) and diastolic (DBP) blood pressure, fasting blood glucose, high-density lipoprotein cholesterol (HDL-c), and triglycerides were examined by Spearman's rho correlation coefficient and linear regression analysis. A p value < 0.05 was considered statistically significant. Results: Increased WC showed a significant correlation with SBP, DBP, glucose, HDL-c, and triglycerides. In bivariate linear regression, approximately 63.0 % of the variability of SBP (≥ 130 mmHg) among the age group 20-40 years was predicted by increased WC according to both criteria. Conclusion: A WC above 80 cm in women aged 20-40 years strongly predicted variability in SBP, calling attention to the importance of measuring WC for the monitoring and prevention of cardiovascular and metabolic diseases in women in this age group.

Estrogen signaling in the proliferative endometrium: implications in endometriosis.

Even though the physiological role of estrogen in the female reproductive cycle and endometrial proliferative phase is well established, the signaling pathways by which estrogen exerts its action in the endometrial tissue are still little known. In this regard, advancements in cell culture techniques and maintenance of endometrial cells in cultures enabled the discovery of new signaling mechanisms activated by estrogen in the normal endometrium and in endometriosis. This review aims to present the recent findings in the genomic and non-genomic estrogen signaling pathways in the proliferative human endometrium specifically associated with the pathogenesis and development of endometriosis.

Estrogen signaling in the proliferative endometrium: implications in endometriosis / Sinalização pelo estrogênio no endométrio proliferativo: implicações na endometriose

SUMMARY Even though the physiological role of estrogen in the female reproductive cycle and endometrial proliferative phase is well established, the signaling pathways by which estrogen exerts its action in the endometrial tissue are still little known. In this regard, advancements in cell culture techniques and maintenance of endometrial cells in cultures enabled the discovery of new signaling mechanisms activated by estrogen in the normal endometrium and in endometriosis. This review aims to present the recent findings in the genomic and non-genomic estrogen signaling pathways in the proliferative human endometrium specifically associated with the pathogenesis and development of endometriosis.

RESUMO Embora esteja bem estabelecido o papel fisiológico do estrogênio no ciclo reprodutivo feminino e na fase proliferativa do endométrio, as vias de sinalização por meio das quais a ação do estrogênio é exercida no tecido endometrial são ainda pouco conhecidas. Nesse sentido, o avanço nas técnicas de cultura celular e a manutenção de células endometriais em cultivo possibilitaram a descoberta de novos mecanismos sinalizadores ativados pelo estrogênio no endométrio normal e na endometriose. Esta revisão tem o objetivo de apresentar as descobertas recentes envolvendo as vias de sinalização genômica e não genômica do estrogênio no endométrio proliferativo humano, especificamente associadas à patogênese e ao desenvolvimento da endometriose.

Citopatogenicidade de Trichomonas vaginalis: uma via de sinalização carboidrato-dependente / Trichomonas vaginalis' pathogenic effect and cell signaling in a carbohydrate-dependent manner

Trichomonas vaginalis é um agente infectante da microbiota vaginal que vem sendo correlacionado ao câncer cervical. Um receptor denominado alectina-1 (Gal 1) pode ser expresso em células epiteliais cervicais humanas se ligando à glicofosfolipídica (LPG) de T. vaginalis. A interação de T. vaginalis com as células epiteliais é mediada por cadeias galactose e N-acetilglucosamina (LacNac). Gal 1 se liga aos sítios poly-LacNAC e está relacionada com a aderência de T. vaginalis à célula humana. A sinalização ocorre por intermédio de sítios da proteína Src (SH2) que se associam, ocorrendo sob os domínios de PI3K que fosforilam a membrana de lípides fosfatidilinositol (PIP e PIP2). Aderindo-se às membranas citoplasmáticas e secretando enzimas, T. vaginalis pode ocasionar a ruptura do envoltório celular podendo fagocitar células epiteliais em meio vaginal. O núcleo N-acetilactosamina de Gal 1 pode mediar a regulação do crescimento celular com a ajuda da proteína GRB2; entretanto, Gal 1 pode contribuir para a supressão da inflamação por meio da indução de apoptose pelas células T ativadas. (AU)

Trichomonas vaginalis is an infectious agent of the vaginal flora which has been associated with cervical cancer. Galectin-1 (Gal 1) is a cell receptor expressed in cervical epithelial cells binding T. vaginalis? lipophosphoglican (LPG). Interaction between T. vaginalis and the epithelial cell is mediated by poly-LacNac domains (galactoside and acetil-lactosamin) and is related to cell adherence as well. Cell signaling occurs by the time Src (SH2) domains are correlated with this interaction and PI3K phosphorilation brings up phosphatidil inositol lipid membranes (PIP and PIP2). T. vaginalis adheres to cytoplasm membrane and secrets specific enzymes that probably lead to membrane rupture. Moreover this parasite may phagocyte epithelial cells in vaginal discharge. Gal 1 nucleus called N-acetil-lactosamin can mediate growth development through GRB2 protein and may contribute to inflammation suppression owing to apoptosis induction of activated T cells.(AU)

Arginine homozygosity in codon 72 of p53 correlates with failure to imatinib response in chronic myeloid leukemia.

PURPOSE: In this study, the role of the polymorphism at codon 72 of tumor protein p53 gene (TP53) was investigated regarding the response to treatment with imatinib in chronic myeloid leukemia (CML). METHODS: A total of 85 patients with CML were treated according to the Brazilian National Cancer Institute (INCA) guidelines and at the end of the 18th month a blood sample were collected for genotyping. Genomic DNA was extracted and TP53 codon 72 genotyping was performed by allele-specific polymerase chain reaction (AS-PCR), which detects argine or proline alleles. RESULT: Of the 85 CML samples, 27 samples were homozygous for arginine (Arg/Arg), 12 homozygous for proline (Pro/Pro) and 46 samples heterozygous (Arg/Pro). TP53 codon 72 polymorphism was in Hardy-Weinberg equilibrium (χ(2)=1.17, P=0.37). We did not find significant association between codon 72 polymorphism and age at diagnosis and sex (P=0.76 and P=0.33, respectively). High Sokal score are significantly associated with Arg/Arg genotype carriers (Odds ratio, OR=4.09; 95% confidence interval, CI 1.01=15.89; P=0.036). The arginine allele in homozygosis also have an increased risk of failure response to imatinib when compared with both, the heterozygous (Arg/Pro) and proline homozygous patients (P=0.021; OR=2.99, 95% CI=1.16-7.67). Additionally, interaction analysis with age at diagnosis revealed that among patients over 40-yr old, Arg/Arg genotype was significantly associated with non-responder patients (P=0.007; OR=5.13, 95% CI=1.5-17.55). CONCLUSION: The findings suggest that in CML patients, TP53 codon 72 polymorphism may contribute to a high Sokal score and failure to imatinib treatment.

In vitro mutagenicity and blood compatibility of paclitaxel and curcumin in poly (DL-lactide-co-glicolide) films.

Curcumin is considered to be a potential component for drug-eluting stents due to its anti-inflammatory properties. In this study we compared the mutagenicity and blood compatibility of curcumin to first generation drug eluting stent components: paclitaxel and sirolimus. The Ames test was used to assess mutagenicity. Blood compatibility was tested by measuring platelet activation and fibrinogen adsorption on poly (DL-lactide-co-glycolide, PLGA) films. We discovered that there was no significant increase in the number of revertants/plate following treatment with curcumin (up to 0.5mg/plate) or sirolimus (up to 0.5 µg/plate). However, a significant induction in the frequency of bacterial his(+) revertant colonies by paclitaxel at concentrations of 0.02, 0.05, 0.1, 0.2 and 0.5 µg/plate was observed. We also discovered a significant reduction in platelet activation by PLGA films containing 30% and 50% by weight curcumin. A similar reduction in platelet activation was also observed for PLGA films containing 1% by weight paclitaxel. In addition, we observed an increase of fibrinogen adsorption to PLGA-films containing curcumin. This would compromise the potential use of curcumin as a component of drug-eluting stents. Moreover, our data challenges the current view that paclitaxel does not significantly induce mutagenesis.

In vitro antiproliferative effects of the indole alkaloid vallesiachotamine on human melanoma cells.

In course of a screening for small molecules presenting potential anticancer properties, a known monoterpene indole alkaloid named vallesiachotamine was isolated from the leaves of Palicourea rigida (Rubiaceae) collected in the Brazilian Cerrado. The structure was determined by spectroscopic methods, mainly 1D- and 2D-NMR and its biological activities were investigated on cultured human (SK-MEL-37) melanoma cells. In vitro cytotoxicity was evaluated by the 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The inhibitory concentration (IC(50)) was 14.7 ± 1.2 µM for 24 h of drug exposure. Flow cytometry analysis revealed that vallesiachotamine induced G0/G1 arrest and increased the proportion of sub-G1 hypodiploid cells (at 11 µM and 22 µM) and this effect was not dependent on time of incubation. At these concentrations, a typical ladder was observed by agarose gel electrophoresis of the extracted DNA. Treatment of cells with 50 µM vallesiachotamine for 24 h caused extensive cytotoxicity and necrosis. Our results demonstrated that the indole alkaloid vallesiachotamine exhibited important cytotoxicity toward human melanoma cells and that apoptosis and necrosis might be responsible for the observed events.

TP53 codon 72 polymorphism in pigmentary phenotypes.

The p53 protein exerts different cellular functions, and recent findings have demonstrated its influence on the cascade of skin pigmentation during UV exposure. Among TP53 gene polymorphisms, the most studied is the G to C transversion in exon 4 at codon 72, which results in three distinct genotypes, Arg/Arg, Pro/Pro and Arg/Pro, each one encoding different p53 isoforms. Therefore, this study aimed to determine the relationship between TP53 codon 72 polymorphism and skin protection against sunburn. Genomic DNA was extracted from peripheral blood samples and genotyping was performed by PCR and confirmed by restriction enzyme digestion. The genotype frequency was 50% for Arg/Arg and 14.6% for Pro/Pro genotype. The frequency of heterozygous subjects was 35.4%. In our population, p53 genotypes were in Hardy-Weinberg (HW) equilibrium (X2 HM less than 3.84), showing a predominance of arginine allele (total Arg allele frequency of 68%). No significant association between p53 genotype and skin colour, hair or eye colour and susceptibility to sun exposure was found. However, further analysis demonstrated a significant association between the genotype Pro/Pro and blue/green eyes among participants who presented redness (P=0.016). Our findings indicate susceptibility to sun exposure when this phenotype (eye colour) occurs simultaneously with Pro/Pro genotype.

Curcumin associated magnetite nanoparticles inhibit in vitro melanoma cell growth.

Curcumin is a natural product possessing therapeutic properties but the low water solubility of this compound limits its use. We have successfully incorporated curcumin into a bilayer of dodecanoic acid attached to magnetite nanoparticles in an effort to maximize solubility and delivery efficiency. Curcumin/magnetite nanoparticles were characterized using diffused reflectance infra-red fourier transform spectroscopy (DRIFTS) and X-ray powder diffraction (XRD). Moreover curcumin associated magnetite nanoparticles inhibited in vitro melanoma cell growth. An inhibitory concentration (IC50) of 66.0 +/- 3.0 microM (48 +/- 2.2 microg-iron/mL) was observed for the curcumin/magnetite nanoparticles. Fluorescent microscopy revealed that curcumin associated magnetite nanoparticles were internalized by the melanoma cells and remained in the cytoplasm. The curcumin/magnetic nanoparticles synthesized in this study possess magnetic and water solubility properties making this a novel curcumin formulation with therapeutic potential.

Embryonic stem-like cells derived from in vitro produced bovine blastocysts

The aim of this work was to study the derivation of bovine embryonic stem-like (ES-like) cells from the inner cell mass (ICM) of in vitro produced blastocysts. The ICMs were mechanically isolated and six out of seventeen (35 percent) ICMs could attach to a monolayer of murine embryonic fibroblasts (MEF). Ten days after, primary outgrowths were mechanically dissected into several small clumps and transferred to a new MEF layer. Cells were further propagated and passaged by physical dissociation over a 60 days period. The pluripotency of the bovine ES-like cells was confirmed by RT-PCR of Oct-4 and STAT-3 gene markers. The colonies were weakly stained for alkaline phosphatase and the mesoderm and endoderm differentiation gene markers such as GATA-4 and Flk-1, respectively, were not expressed. Embryoid bodies were spontaneously formed at the seventh passage. Results showed that bovine ES-like cells could be obtained and passaged by mechanical procedures from the fresh in vitro produced blastocysts.

Morphological alterations and G0/G1 cell cycle arrest induced by curcumin in human SK-MEL-37 melanoma cells

The aim of this work was to study the effect of curcumin on cell cycle in the human SK-MEL-37 melanoma cell line. In addition, morphological and structural analyses were also performed. Flow cytometric analysis showed a G0/G1 arrest at 5 µM after 24 h exposure and a concentration-dependent increase in the proportion of sub-G0 hypodiploid cells. Typical apoptotic events were also observed by the fluorescence microscopy, transmission and scanning electronic microscopy. Loss of mitochondrial membrane potential was not detected. Results suggested that curcumin could arrest human melanoma cells at G0/G1 phase and induce a mitochondrial-independent apoptotic pathway.

O melanoma é um tipo agressivo de câncer cujo tratamento culmina com o estabelecimento de resistência aos quimioterápicos empregados. Portanto, é importante o desenvolvimento de novos agentes farmacológicos que sejam menos tóxicos e que não provoquem quimiorresistência. As inúmeras propriedades terapêuticas da curcumina vêm sendo confirmadas através de estudos sobre o seu mecanismo de ação em células cultivadas. No presente estudo, empregamos células de melanoma humano da linhagem SK-MEL-37, que desenvolveram resistência in vitro à doxorubicina e cisplatina, drogas normalmente utilizadas na clínica. Investigamos o efeito da curcumina sobre o ciclo celular através de citometria de fluxo. Além disso, análises morfológicas e estruturais também foram realizadas. Os resultados demonstraram que o tratamento com uma concentração de 5 ?M de curcumina provocou uma parada na subfase G0/G1. Além disso, observou-se um aumento dose-dependente na proporção de células hipodiplóides em sub-G0. Eventos apoptóticos típicos foram observados por microscopia de fluorescência, microscopia eletrônica de transmissão e microscopia eletrônica de varredura. Não foi detectada alteração no potencial de membrana mitocondrial. Os resultados indicam que futuros estudos poderão tornar possível a utilização da curcumina como um modulador para agentes quimioterápicos empregados na clínica no tratamento do melanoma.

Avaliação da produção de óxido nítrico em ratos, submetidos aos exercícios aeróbio e anaeróbio / Evaluation of the production of nitric oxide in mice, submitted to aerobic and anaerobic exercises

Óxido nítrico (NO) exerce influências muito importantes em vários processos fisiológicos. Neste trabalho avaliamos a produção de NO sanguíneo em ratos Wistar, submetidos ao nado aeróbio e anaeróbio agudos. A formação do óxido nítrico foi verificada através da dosagem dos produtos de oxidação estáveis do metabolismo do óxido nítrico (nitratos). Para isso utilizamos o método colorimétrico de Griess. Verificamos a existência de uma diferença significativa (p = 0,000261) na produção de óxido nítrico entre a realização do nado aeróbio e o anaeróbio, na qual o aeróbio mostrou-se mais eficiente na promoção de níveis mais elevados. O exercício aeróbio agudo com duração de no mínimo 10 minutos mostrou-se mais eficaz no quesito produção de NO em relação ao exercício de 5 minutos. A positiva relação observada entre o exercício aeróbio e a formação de NO pode ajudar a explicar os efeitos benéficos do exercício na saúde cardiovascular. Sabemos que a prática de exercício aeróbio e sua duração aumentam a biodisponibilidade de NO, o qual é considerado importante regulador fisiológico da pressão arterial.

Nitric Oxide (NO) exerts important influences in several physiological processes. In this work we evaluated the production of sanguine NO in Wistar rats, submitted to the acute aerobic and anaerobic exercises. The formation of nitric oxide was verified through the dosage of the end products of oxidation of the metabolism of nitric oxide (nitrates). For this we used the colorimetric Griess method. We verified the existence of a significant difference (p = 0.000261) in the production of NO among the accomplishment of the aerobic swimming and the anaerobic, where the aerobic was shown more efficient in the promotion of higher levels. The acute aerobic exercise with duration of at least 10 minutes was shown more effective in the requirement production of NO in relation to the 5 minutes exercise. The positive relationship observed between the aerobic exercise and the formation of NO can help to explain the beneficial effects of the exercise in the cardiovascular health. We know that the practice of aerobic exercise and your duration increases the biodisponibility of NO, which is an important physiologic regulator of the blood pressure.

In vitro biological activities of anionic gamma-Fe2O3 nanoparticles on human melanoma cells.

Three magnetic fluid (MF) samples containing gamma-Fe2O3 (maghemite) nanoparticles surface-coated with either meso-2,3-dimercaptosuccinic acid (DMSA), citric acid or lauric acid were prepared, characterized, and assessed for their cytotoxic potential on the human SK-MEL-37 melanoma cell line. Ultra-structural analysis was also performed using transmission electron microscopy (TEM). In vitro cytotoxicity was evaluated by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The inhibitory concentration (IC50) derived from the sigmoidal dose response curve was 254 microg-iron/mL (95% confidence interval 239-270 microg-iron/mL) for lauric acid-coated nanoparticles. DMSA-coated nanoparticles did not exhibit a clear trend toward toxicity (IC50 value is more than 2260 +/- 50 microg-iron/mL) and the IC50 value was about 433 +/- 14 microg-iron/mL for citric-acid coated nanoparticles. The cytotoxic response correlated with both the hydrodynamic diameter and the zeta potential suggests that the chain length of the carboxylic acid of the coating species may influence metabolic cellular process. Also the assayed nanoparticles can be considered non-cytotoxic to human melanoma cells since IC50 values are higher than plasma concentration usually observed in clinical use of contrast agents. Using TEM we verified that all assayed nanoparticles were internalized by cells through endocytic vesicles. Additionally, cells treated with lauric acid-coated nanoparticles at high concentration (588 or 840 microg-iron/mL) displayed morphological features of apoptosis (surface blebbing, intense vacuolization and chromatin condensation) or a typical DNA ladder pattern when analyzed by TEM or agarose gel electrophoresis, respectively. Apoptotic events may be operative, suggesting a promising therapeutic application for the lauric acid-coated nanoparticle in the treatment of cancer cells.

Role of G protein-coupled receptor kinases in the homologous desensitization of the human and mouse melanocortin 1 receptors.

The melanocortin 1 receptor, a G protein-coupled receptor positively coupled to adenylyl cyclase, is a key regulator of epidermal melanocyte proliferation and differentiation and a determinant of human skin phototype and skin cancer risk. Despite its potential importance for regulation of pigmentation, no information is available on homologous desensitization of this receptor. We found that the human melanocortin 1 receptor (MC1R) and its mouse ortholog (Mc1r) undergo homologous desensitization in melanoma cells. Desensitization is not dependent on protein kinase A, protein kinase C, calcium mobilization, or MAPKs, but is agonist dose-dependent. Both melanoma cells and normal melanocytes express two members of the G protein-coupled receptor kinase (GRK) family, GRK2 and GRK6. Cotransfection of the receptor and GRK2 or GRK6 genes in heterologous cells demonstrated that GRK2 and GRK6 impair agonist-dependent signaling by MC1R or Mc1r. However, GRK6, but not GRK2, was able to inhibit MC1R agonist-independent constitutive signaling. Expression of a dominant negative GRK2 mutant in melanoma cells increased their cAMP response to agonists. Agonist-stimulated cAMP production decreased in melanoma cells enriched with GRK6 after stable transfection. Therefore, GRK2 and GRK6 seem to be key regulators of melanocortin 1 receptor signaling and may be important determinants of skin pigmentation.

Psoralen derivatives and longwave ultraviolet irradiation are active in vitro against human melanoma cell line.

Cutaneous malignant melanoma is a very serious form of skin cancer that arises from melanocytes. Currently there is no effective treatment for metastatic melanoma so intense clinical trials are evaluating new drugs for this human malignancy. Psoralens are a group of compounds that bind to DNA in rapidly dividing cells and with ultraviolet light in the A band (UVA) cause DNA crosslinking, thereby preventing cellular division. They are used in the treatment of psoriasis and cutaneous T-cell lymphoma among other skin and blood diseases. We have investigated the cytotoxic potential of three psoralen derivatives plus UVA exposure (PUVA) on a established cell line of human melanoma. Cells were treated with different concentrations of 8-methoxypsoralen (8-MOP), 4,5',8-trimethylpsoralen (TMP) and 7-methylpyridopsoralen (MPP), for 1 h and after exposure to UVA light (0.3 J/cm(2)) were allowed to recover over a 24-72 h period. Viability was assessed by the microculture 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) colorimetric assay. Cisplatin, one of the most important drugs in the chemotherapy of melanoma, was included for comparative studies. All the psoralen derivatives tested were markedly cytotoxic in a dose and post-exposure-time dependent manner. The IC(50) values for 72 h of post-exposure time were as follows: MPP=0.05+/-0.01, TMP=0.13+/-0.003 and 8-MOP=10.79+/-1.85 micromol/L. Regardless of the limitations of the in vitro model, our results suggested that the lower IC(50) values of TMP and MPP might be of clinical importance.