Expression and distribution of kallikrein-related peptidases 5, 7, 8 and 10 in normal apocrine gland of canine skin / Expresión y distribución de péptidos relacionados con la calicreína 5, 7, 8 y 10 en glándula apocrina normal de piel canina

SUMMARY: Apocrine glands are sweat glands that are located in the skin of the dog. Anal sac apocrine, circunanal apocrine, and mammary glands are considered modified apocrine structures, and there are about nine possible types of neoplasms and other tumors in the apocrine glands of the dog and cat, including cysts, adenoma, carcinoma, and adenocarcinoma. Thus, it is important to provide new markers to characterize these glands to improve the histopathological diagnosis. In this article, we describe the distribution of kallikrein- related peptidases 5, 7, 8, and 10 in the normal apocrine glands of the dog's skin. These proteases have been shown to play a fundamental role in the homeostasis of the human skin barrier but have been scarcely studied in canine skin.

Las glándulas apocrinas son glándulas sudoríparas que se encuentran en la piel del perro. Las glándulas apocrinas del saco anal, apocrinas circunanales y mamarias se consideran estructuras apocrinas modificadas, y existen alrededor de nueve tipos posibles de neoplasias y otros tumores en las glándulas apocrinas del perro y el gato, incluidos quistes, adenoma, carcinoma y adenocarcinoma. Por lo tanto, es importante proporcionar nuevos marcadores para caracterizar estas glándulas para mejorar el diagnóstico histopatológico. En este artículo, describimos la distribución de las peptidasas 5, 7, 8 y 10 relacionadas con la calicreína en las glándulas apocrinas normales de la piel del perro. Se ha demostrado que estas proteasas desempeñan un papel fundamental en la homeostasis de la barrera de la piel humana, pero apenas se han estudiado en la piel canina.

Diretrizes brasileiras de angioedema hereditário 2022 ­ Parte 2: terapêutica / 2022 Brazilian guidelines for hereditary angioedema ­ Part 2: therapy

O tratamento do angioedema hereditário tem início com a educação dos pacientes e familiares sobre a doença, pois é fundamental o conhecimento da imprevisibilidade das crises, assim como os seus fatores desencadeantes. O tratamento medicamentoso se divide em terapia das crises e profilaxia das manifestações clínicas. As crises devem ser tratadas o mais precocemente possível com o uso do antagonista do receptor de bradicinina, o icatibanto ou o concentrado de C1-inibidor. É necessário estabeler um plano de ação em caso de crises para todos os pacientes. A profilaxia de longo prazo dos sintomas deve ser realizada preferencialmente com medicamentos de primeira linha, como concentrado do C1-inibidor ou o anticorpo monoclonal anti-calicreína, lanadelumabe. Como segunda linha de tratamento temos os andrógenos atenuados. Na profilaxia de curto prazo, antes de procedimentos que podem desencadear crises, o uso do concentrado de C1-inibidor é preconizado. Existem algumas restrições para uso desses tratamentos em crianças e gestantes que devem ser consideradas. Novos medicamentos baseados nos avanços do conhecimento da fisiopatologia do angioedema hereditário estão em desenvolvimento, devendo melhorar a qualidade de vida dos pacientes. O uso de ferramentas padronizadas para monitorização da qualidade de vida, do controle e da atividade da doença são fundamentais no acompanhamento destes pacientes. A criação de associações de pacientes e familiares de pacientes com angioedema hereditário tem desempenhado um papel muito importante no cuidado destes pacientes no nosso país.

The treatment of hereditary angioedema begins with the education of patients and their families about the disease, as it is essential to know the unpredictability of attacks as well as their triggering factors. Drug treatment is divided into attack therapy and prophylaxis of clinical manifestations. Attacks should be treated as early as possible with the bradykinin receptor antagonist icatibant or C1-inhibitor concentrate. An action plan needs to be established for all patients with attacks. Long-term prophylaxis of symptoms should preferably be performed with first-line drugs such as C1-inhibitor concentrate or the anti-kallikrein monoclonal antibody lanadelumab. Attenuated androgens are the second line of treatment. In short-term prophylaxis, before procedures that can trigger attacks, the use of C1-inhibitor concentrate is recommended. There are some restrictions for the use of these treatments in children and pregnant women that should be considered. New drugs based on advances in knowledge of the pathophysiology of hereditary angioedema are under development and are expected to improve patient quality of life. The use of standardized tools for monitoring quality of life and controlling disease activity is essential in the follow-up of these patients. The creation of associations of patients and families of patients with hereditary angioedema has played a very important role in the care of these patients in Brazil.

Polymorphisms in DSSP (rs36094464) and RUNX2 (rs566712) genes contribute to the susceptibility of dental caries in childhood / Los polimorfismos en los genes DSSP (rs36094464) y RUNX2 (rs566712) contribuyen a la susceptibilidad de la caries dental en la infancia

SUMMARY: Dental caries corresponds to an ecological and non-contagious, dynamic and chronic disease of multifactorial origin; currently there is evidence of how genetic factors could be included as predisposing agents to suffer it, however this evidence is diverse and incipient. a cross-sectional study was p erformed to investigate the possible associations of DSPP (rs36094464), RUNX2 (rs566712) and KLK4 (rs198968) polymorphisms in early childhood caries. Saliva samples of children (2-11years old) were collected and genotyped for DSPP (rs36094464), RUNX2 (rs566712) and KLK4 (rs198968) polymorphisms. Through the ceft index their caries history was determined and the gene variants were students through molecular biology techniques. polymorphisms of the DSSP (rs36094464) and RUNX2 (rs566712) are associated and contribute to the susceptibility of dental caries disease in early childhood, as they are related to their history of caries. KLK4 (rs198968) polymorphisms are not associated. In conclusions, the studied polymorphisms on DSSP and RUNX2 genes are associated with changes in the tooth microarchitecture, favoring the appearance of microlesions that would contribute to dental caries disease susceptibility in early childhood. Also, no association was found for the studied polymorphism of the KLK4 gene with dental caries disease susceptibility.

RESUMEN: La caries dental corresponde a una enfermedad crónica, no contagiosa, dinámica y de origen multifactorial. Actualmente existe evidencia de cómo los factores genéticos podrían incluirse como agentes predisponentes, sin embargo, esta evidencia es diversa e incipiente. Se realizó un estudio transversal para investigar las posibles asociaciones entre los polimorfismos DSPP (rs36094464), RUNX2 (rs566712) y KLK4 (rs198968) y la caries en la infancia. Se colectaron muestras de saliva de niños (de 2 a 11 años de edad) y se genotipificaron para los polimorfismos DSPP (rs36094464), RUNX2 (rs566712) y KLK4 (rs198968). Mediante el índice ceft se determinó su historial de caries y se estudiaron las variantes genéticas mediante técnicas de biología molecular. Los datos obtenidos indican que los polimorfismos del DSSP (rs36094464) y RUNX2 (rs566712) están asociados y contribuyen a la susceptibilidad de la enfermedad de caries dental en la infancia, ya que están - además - relacionados con el historial de caries. En conclusión, los polimorfismos estudiados en los genes DSSP y RUNX2 se asocian a la aparición de microlesiones que contribuirían a la susceptibilidad a la enfermedad de caries dental en la infancia. Creemos que este estudio es importante para la odontopediatría porque destaca el papel de DSSP (rs36094464) y RUNX2 (rs566712) y la susceptibilidad a la caries dental durante la infancia, además resalta la utilidad de la evaluación genética para la predicción y prevención de la caries dental y porque aporta evidencia que indica que los factores genéticos están implicados en la etiología de la caries.

Interactions amongst inflammation, renin-angiotensin-aldosterone and kallikrein-kinin systems: suggestive approaches for COVID-19 therapy

Coronavirus disease 2019 (COVID-19) is a rapid-spread infectious disease caused by the SARS-CoV-2 virus, which can culminate in the renin-angiotensin-aldosterone (RAAS) and kallikrein-kinin (KKS) systems imbalance, and in serious consequences for infected patients. This scoping review of published research exploring the RAAS and KKS was undertaken in order to trace the history of the discovery of both systems and their multiple interactions, discuss some aspects of the viral-cell interaction, including inflammation and the system imbalance triggered by SARS-CoV-2 infection, and their consequent disorders. Furthermore, we correlate the effects of continued use of the RAAS blockers in chronic diseases therapies with the virulence and physiopathology of COVID-19. We also approach the RAAS and KKS-related proposed potential therapies for treatment of COVID-19. In this way, we reinforce the importance of exploring both systems and the application of their components or their blockers in the treatment of coronavirus disease.(AU)

Subproteome of Lachesis muta rhombeata venom and preliminary studies on LmrSP-4, a novel snake venom serine proteinase

Lachesis muta rhombeata is one of the venomous snakes of medical importance in Brazil whose envenoming is characterized by local and systemic effects which may produce even shock and death. Its venom is mainly comprised of serine and metalloproteinases, phospholipases A2 and bradykinin-potentiating peptides. Based on a previously reported fractionation of L. m. rhombeata venom (LmrV), we decided to perform a subproteome analysis of its major fraction and investigated a novel component present in this venom. Methods: LmrV was fractionated through molecular exclusion chromatography and the main fraction (S5) was submitted to fibrinogenolytic activity assay and fractionated by reversed-phase chromatography. The N-terminal sequences of the subfractions eluted from reversed-phase chromatography were determined by automated Edman degradation. Enzyme activity of LmrSP-4 was evaluated upon chromogenic substrates for thrombin (S-2238), plasma kallikrein (S-2302), plasmin and streptokinase-activated plasminogen (S-2251) and Factor Xa (S-2222) and upon fibrinogen. All assays were carried out in the presence or absence of possible inhibitors. The fluorescence resonance energy transfer substrate Abz-KLRSSKQ-EDDnp was used to determine the optimal conditions for LmrSP-4 activity. Molecular mass of LmrSP-4 was determined by MALDI-TOF and digested peptides after trypsin and Glu-C treatments were analyzed by high resolution MS/MS using different fragmentation modes. Results: Fraction S5 showed strong proteolytic activity upon fibrinogen. Its fractionation by reversed-phase chromatography gave rise to 6 main fractions (S5C1-S5C6). S5C1-S5C5 fractions correspond to serine proteinases whereas S5C6 represents a C-type lectin. S5C4 (named LmrSP-4) had its N-terminal determined by Edman degradation up to the 53rd amino acid residue and was chosen for characterization studies. LmrSP-4 is a fibrinogenolytic serine proteinase with high activity against S-2302, being inhibited by PMSF and benzamidine, but not by 1,10-phenantroline. In addition, this enzyme exhibited maximum activity within the pH range from neutral to basic and between 40 and 50 °C. About 68% of the LmrSP-4 primary structure was covered, and its molecular mass is 28,190 Da. Conclusions: Novel serine proteinase isoforms and a lectin were identified in LmrV. Additionally, a kallikrein-like serine proteinase that might be useful as molecular tool for investigating bradykinin-involving process was isolated and partially characterized.(AU)

Preconditioning with L-alanyl-glutamine upon cerebral edema and hypocampus red neurons counting in rats subjected to brain ischemia /reperfusion injury

PURPOSE: To evaluate the effects of the dipeptide L-alanyl-glutamine (L-Ala-Gln) as a preconditioning agent to potentially promote reduction in the intensity of lesion or induction of resilience in rats subjected to global cerebral ischemia/reperfusion (I/R) injury. METHODS: Thirty-six male Wistar rats weighing 280-300g were randomly assigned to six groups (n=6). Groups Sham 1h and 24h were treated with saline and spared of further interventions. The remaining groups were submitted to clamping of the common carotid arteries for 30 minutes (ischemia) and treated with saline (SS) or L-Ala-Gln. Brain reperfusion was allowed for 1or 24 h. L-Ala-Gln was administered intravenously (0.75g/kg) 30 minutes before sham procedure or induction of global brain I/R injury. Brain edema and red neuron counting were determined. Results were expressed as Mean±SD for normal results and Median±Percentile for non parametric data. Significance was established at p<0.05. RESULTS: Global I/R injury promoted an increase in brain edema at 24 h after reperfusion, whereas preconditioning with L-Ala-Gln induced no change in edema. On the other hand, L-Ala-Gln preconditioning decreased significantly red neurons counting both at 1h and 24h post reperfusion (p<0.05). CONCLUSION: There was a significant preconditioning effect with L-Ala-Gln decreasing cell death (red neurons counting) at early (1h) and late reperfusion (24h) in the cerebral tissue. .

Napping during the night shift and recovery after work among hospital nurses / Cochilo durante o plantão noturno e a recuperação após o trabalho entre enfermeiros de hospitais / Siesta durante la guardia nocturna y la recuperación tras el trabajo entre enfermeros de hospitales

OBJECTIVE: To analyze the association between the length of napping during the night shift and the recovery after work among nurses. METHOD: Cross-sectional epidemiological study involving 1940 nurses from 18 public hospitals in the City of Rio de Janeiro. A multidimensional and self-applied questionnaire was used with information about health, sociodemographic and occupational characteristics, health-related behaviors and housework. Multiple logistic regression was applied to identify the association, adjusted for confounding variables. RESULTS: The gross analyses showed 44%, 127% and 66% higher chances of a high level of recovery after work for nurses who sleep up to two hours, between 2.1 and 3 hours and 3.1 hours or more, respectively, when compared to the nurses who do not sleep. After adjusting for confounding variables, the association only continues significant for the group that sleeps 2.1 to 3 hours during the night shift (OR=1.79; 95%CI=1.33-2.41). CONCLUSION: The association between the length of napping and the high level of recovery after work, confirmed in the present results, can be included in the studies that aim to support more appropriate policies aimed at improving the workers' work, life and health conditions, not only in nursing, but night-shift workers in general. .

OBJETIVO: analisar a associação entre duração do cochilo durante o plantão noturno e recuperação após o trabalho, entre enfermeiros. MÉTODO: estudo epidemiológico seccional com 1940 enfermeiros, de 18 hospitais públicos, do Município do Rio de Janeiro. Utilizou-se questionário multidimensional e autopreenchível com informações sobre saúde, características sociodemográficas, ocupacionais, comportamentos relacionados à saúde e trabalho doméstico. Utilizou-se a regressão logística múltipla, buscando identificar a associação ajustada por variáveis de confundimento. RESULTADOS: as análises brutas mostraram chances 44%, 127% e 66% mais elevadas de alta recuperação após o trabalho, para aqueles que dormem até 2 horas, de 2,1 a 3 horas e de 3,1 horas ou mais, respectivamente, comparados aos que não dormem. Após o ajuste por variáveis de confundimento, a associação permanece significativa apenas para o grupo que dorme de 2,1 a 3 horas durante o plantão noturno (OR=1,79; IC95%=1,33-2,41). CONCLUSÃO: a associação entre tempo de cochilo e alta recuperação após o trabalho, confirmada nos resultados, pode compor os estudos que buscam subsidiar políticas mais adequadas voltadas à melhoria das condições de trabalho, de vida e saúde dos trabalhadores, não apenas da enfermagem, mas trabalhadores noturnos de forma geral. .

OBJETIVO: Analizar la asociación entre la duración de la siesta durante la guardia nocturna y la recuperación tras el trabajo entre enfermeros. MÉTODO: Estudio epidemiológico seccional con 1940 enfermeros de dieciocho hospitales públicos del Municipio de Rio de Janeiro. Fue utilizado cuestionario tipo multidimensional y autollenado con informaciones sobre salud, características sociodemográficas, ocupacionales, comportamientos relacionados a la salud y trabajo doméstico. Fue utilizada la regresión logística múltipla, buscando identificar la asociación ajustada por variables de confusión. RESULTADOS: Los análisis brutos mostraron posibilidades 44%, 127% y 66% más elevadas de alta recuperación tras el trabajo, para aquellos que duermen hasta 2 horas, de 2,1 a 3 horas y de 3,1 horas o más, respectivamente, comparados a aquellos que no duermen. Tras el ajuste por variables de confusión, la asociación sigue significativa solamente para el grupo que duerme de 2,1 a 3 horas durante la guardia nocturna (OR=1,79; IC95%=1,33-2,41). CONCLUSIÓN: La asociación entre el tiempo de siesta y la alta recuperación tras el trabajo, confirmada en nuestros resultados, puede componer los estudios con objeto de subsidiar políticas más adecuadas dirigidas a la mejora de las condiciones de trabajo, de vida y salud de los trabajadores, no solamente enfermeros, pero trabajadores nocturnos de manera general. .

Sexual hormones modulate compensatory renal growth and function / Las hormonas sexuales modulan el crecimiento renal compensador (CRC) y la función renal en la uninefrectomía (uNx)

The role played by sexual hormones and vasoactive substances in the compensatory renal growth (CRG) that follows uninephrectomy (uNx) is still controversial. Intact and gonadectomized adult Wistar rats of both sexes, with and without uNx, performed at 90 days age, were studied at age 150 days. Daily urine volume, electrolyte excretion and kallikrein activity (UKa) were determined. Afterwards, glomerular filtration rate and blood pressure were measured, the kidneys weighed and DNA, protein and RNA studied to determine nuclei content and cell size. When the remnant kidney weight at age 150 days was compared with the weight of the kidney removed at the time of uNx, male uNx rats showed the greatest CRG (50%) while growth in the other uNx groups was 25%, 15% and 19% in orchidectomized, female and ovariectomized rats, respectively. The small CRG observed in the uNx female rats was accompanied by the lowest glomerular filtration value, 0.56 ± 0.02 ml/min/g kwt compared, with the other uNx groups, p < 0.05. Cell size (protein or RNA/DNA) was similar for all the groups except for uNx orchidectomized rats. In this group the cytoplasmatic protein or RNA content was lower than in the other groups while DNA (nuclei content) was similar. Some degree of hyperplasia was determined by DNA content in the uNx groups. Male sexual hormones positively influenced CRG and its absence modulated cell size. Female sexual hormones, instead, did not appear to stimulate CRG. The kallikrein kinin system may not be involved in CRG.

La importancia que pueden tener las hormonas sexuales y sustancias vasoactivas sobre el crecimiento renal compensador (CRC) que sigue a la uninefrectomía es aún materia de debate. Se estudiaron ratas Wistar de ambos sexos, a los 150 días de vida, intactas y gonadectomizadas con y sin uNx, realizada a los 90 días de vida. Se midió volumen urinario diario y excreción de electrolitos y actividad de kalikreína urinaria. Se midió filtrado glomerular y presión arterial media extrayéndose luego los riñones que fueron pesados y preparados para estudios histológicos y determinación de ADN, ARN y proteínas para estimar contenido nuclear y tamaño celular. El CRC fue calculado comparando el peso del riñón al momento de las uNx (90 dias de vida) con aquel obtenido a los 150 días de vida. En las ratas macho uNx se observó el mayor CRC (50%) mientras que, en los otros grupos uNx solo alcanzó un 25%, 15% y 19%. El filtrado glomerular acompañó los cambios morfológicos observándose el menor filtrado en las ratas hembras uNx respecto al resto de los grupos 0.56 ± 0.02, p < 0.05. El tamaño celular (proteína o ARN/ ADN) fue similar para todos los grupos excepto para los orquidectomizados uNx, cuyo contenido citoplasmático fue menor. El contenido nuclear (ADN) fue semejante en todos los grupos. Se observó que el CRC está influenciado positivamente por las hormonas sexuales masculinas y su ausencia modula el tamaño celular. La falta de hormonas sexuales femeninas, en cambio, afecta negativamente el CRC. El sistema kalikreína kinina no parecería estar involucrado en el CRC.

Cyclooxygenase-2 and hypoxia-regulated proteins are modulated by basic fibroblast growth factor in acute renal failure

Acute renal failure (ARF) can be caused by injuries that induce tissue hypoxia, which in turn can trigger adaptive or inflammatory responses. We previously showed the participation of basic fibroblast growth factor (FGF-2) in renal repair. Based on this, the aim of this study was to analyze the effect of FGF-2 signaling pathway manipulation at hypoxia-induced protein levels, as well as in key proteins from the vasoactive systems of the kidney. We injected rat kidneys with FGF-2 recombinant protein (r-FGF) or FGF-2 receptor antisense oligonucleotide (FGFR2-ASO) after bilateral ischemia, and evaluated the presence of iNOS, EPO and HO-1, in representation of hypoxia-induced proteins, as well as COX-2, renin, kallikrein, and B2KR, in representation of the vasoactive systems of the kidney. A reduction in iNOS, HO-1, EPO, renin, kallikrein, B2KR, and in renal damage was observed in animals treated with r-FGF. The opposite effect was found with FGF-2 receptor down-regulation. In contrast, COX-2 protein levels were higher in kidneys treated with r-FGF and lower in those that received FGFR2-ASO, as compared to saline treated kidneys. These results suggest that the protective role of FGF-2 in the pathogenesis of ARF induced by I/R is a complex process, through which a differential regulation of metabolic pathways takes place.

La diabetes mellitus experimental y las alteraciones de los tejidos blandos en periodoncia: efectos de la insulina y de un antagonista del receptor de cininas / Experimental diabetes mellitus and soft tissue alterations in periodontics: effects of insulin and a kinin receptor antagonist

La gingivopatía diabética ha sido reconocida no como una enfermedad infecciosa loca, sino como una enfermedad crónica inflamatoria para los tejidos gingivales. La diabetes parece contribuir a la mayor presencia de bacterias en la encía; sin embargo, es totalmente discutible que existen cambios cualitativos o cuantitativos entre la flora periodontopática diabética y no diabética. La enfermedad periodontal es una afección que tiene como respuesta un cuadro crónico inflamatorio. La disminución de insulina a consecuancia de alteraciones metabólicas induce hiperglucemia progresiva, lo que determina importantes factores de riesgo para las complicaciones orgánicas. Este trabajo muestra los efectos de dos factores críticos en la gingivopatía diabética: el efecto prolongado de hiperglucemia diabética y el estado inflamatorio y los componentes que determinan la destrucción del tejido gingival. El tratamiento experimental de acuerdo con los resultados obtejidos muestra que no sólo el control de la hiperglucemia por insulina es importante, sino también el ataque a los factores inflamatorios por drogas específicas.

El efecto de nefroprotección de los calcio-antagonistas: una contribución al conocimiento de su probable mecanismo / Nephroprotection from calcium channels blocking agents: a contribution to their probable mechanism of action

La prevalencia e incidencia de la Insuficiencia Renal Crónica (IRC) han ido aumentando progresivamente en los últimos 20 años, y tanto la Hipertensión Arterial (HTA) como la Diabetes Mellitus (DM) son las dos causas más importantes de ambas situaciones. Los cambios hemodinámicos y metabólicos contribuyen en el desarrollo y progresión de la enfermedad renal en ambas HTA y DM, que son factores claves en la nefropatía. La reducción de la presión arterial sola no es suficiente para una máxima protección renal, de allí la necesidad del conocimiento de un mecanismo adicional para la nefroprotección para establecer medidas preventivas. A manera de contribuir al conocimiento de tal mecanismo nefroprotector de las drogas calcio-antagonistas (CA) del grupo dihidropiridinas se realizó este estudio prospectivo, experimental y longitudinal controlado con placebo en 55 adultos normotensos e hipertensos no-diabéticos que recibieron 3 drogas antihipertensivas CA a monodosis matutinas. Se dividieron en 3 grupos. Grupo A (n = 25): Control, normotensos, que recibieron dosis única de 20 mg de Nitrendipina (NIT). Grupo B (n = 15): Hipertensos tipos I y II que recibieron dosis de 20 mg de Nifedipina (NIF) por 12 semanas y Grupo C (n = 15): Hipertensos tipo I y II que recibieron monodosis de Amlodipina (AML) de 5 mg/d durante 1 semana. En conjunto con las respuestas clínica y hemodinámica, se analizaron parámetros bioquímicos como hormonas vasoactivas renales renina, prostaglandinas, calicreínas y óxido nítrico y también cGMP. El efecto antihipertensivo de los CA se confirmó, y hubo un aumento significativo de la excreción urinaria de calicreínas relacionado con el aumento del óxido nítrico.

Análise de aspectos moleculares no modelo de progressão do melanoma cutânea por meio do estudo imunoistoquímico(Tissue Microarray-TMA) / Analysis of the molecular aspects on the progress melanoma model using the immunohistochemical study(tissue microrray-TMA

Introdução: O melanoma cutâneo é uma neoplasia de comportamento biológico muito agressivo e se origina dos melanócitos localizados na camada basal da epiderme. A sua incidência vem aumentando mundialmente na população branca. ... Novas tecnologias, como o TMA estão sendo utilizadas para o estudo de proteínas no modelo de progressão tumoral utilizando-se de nevos comuns, nevos atípicos, melanomas primários e metastáticos. ... Objetivos: Estudar as proteínas conexina 43, desmocolina 3, citoceratina 5, calicreina 6 e calicreinas 7, que estão envolvidas em adesão e comunicação celular no modelo de progressão do melanoma cutâneo. Material e métodos: Por meio da técnica de TMA foram analisados 59 amostras de nevos comuns, 29 de nevos atípicos 133 melanomas invasivos e 29 metástases. Foi realizada imunoistoquímica com utilização do permanent red para as proteínas conexina 43, desmocolina 3 e citoceratina 5. ... Resultados: A expressão de conexina 43 foi maior em melanoma em relação aos nevos atípicos e comuns. A comparação da expressão de conexina 43 entre nevos comuns e atípicos não se mostrou estatisticamente significativa. ... Conclusão final: as proteínas de adesão e comunicação celular poderiam contribuir no desenvolvimento tumoral descrito no modelo de progressão do melanoma.

Cutaneous melanoma is an agressive cancer, derived from the epidermal basal layer melanocytes. The incidence is growing around the world in caucasians. Based on experimental, clinical and histophatological observations Elder et al., proposed a development and progression melanoma model. New high throughput methods such as tissue microarray (TMA) are being used to evaluate the role of numerous proteins on the melanoma progression model using common nevus, atypical nevus, primary melanomas and metastatic melanomas. These studies allow to establish their participation in the melanoma model and to characterize the molecular and cellular events and help to better understand the pathways involved in melanoma pathogenesis. Adhesion molecules and related proteins have been reported to have a role in this pathway. The purpose of this study was to evaluate the immunohistochemical participation of connexin 43 (Cx43), desmocollin 3 (DSC3), citokeratin 5 (CK5), kalikrein 6 (KLK6) and kalikrein 7 (KLK7), cellular adhesion and communication proteins, in the progression melanoma model. We analyzed 59 samples of common nevus, 29 atypical nevus, 133 invasive and 29 metastatic melanomas by TMA. The immunostaining was evaluated with digital microscopy (ACIS III). Connexin 43 and DSC3 expression was higher in melanoma compared to atypical and common nevi (p<0,001). CK5 expression was higher in atypical nevus than in melanomas and common nevus (p<0,001). The CK5 expression on melanomas had higher expression than common nevus (p<0,001). Thin melanomas had higher CK5 expression than the intermediate and thick melanomas (>1mm) (P<0,001). Stage I had higher CK5 expression than stage II and III/IV (p=0,006 and p=0,004). CK5 expression was also higher in patients with lymph node metastasis (p<0,001). Higher KLK6 expression was found in melanomas compared to common nevus (p<0,001). A higher KLK7 expression was found in atypical nevus compared to common nevus and melanoma (p<0,001). Its expression was higher in melanomas than the common nevus (p<0,001). Higher KLK7 expression was found in thick melanomas (>4mm) compared to thin (<1mm) (p=0,014). Stage I melanomas had higher KLK7 expression in relation to the stage II melanomas (p=0,045). A higher expression was found in patients with no metastatic lymph node (p=0,005). The correlation was positive between Cx43 and DSC3 and between KLK6 and KLK7. The digital microscopy allowed us to study the expression of the biomarkers in melanoma. The expression of DSC3 and Cx43 were higher in melanomas than common and atypical nevus. The expression of CK5 and KLK7 were higher on atypical nevus than common nevus and melanomas and higher on the melanomas than the common nevus. The KLK6 expression was higher on melanomas than the common nevus. There was a positive and moderate correlation between Cx43 and DCS3 and KLK6 and KLK7. In the final conclusion: the cellular adhesion and communication proteins may contribute to melanoma pathogenesis in the context of the melanoma progression model.

El sistema formador de quinina y su relación con los trastornos cardiovasculares / The kinin-forming system and cardiovascular disorders

Cardiovascular diseases are the most common cause of death worldwide. In Kuwait, death related to cardio vascular diseases may account for about 40%. Thus, it will be the greatest health care burden of the twenty-first century. Hypertension and diabetes are the two major risk factors in the development of cardiac hypertrophy, ischemic heart disease, cardiac failure, cardiac arrhythmias and myocardial infarction. The kallikrein-kinin components (plasma and urinary kininogens, kallikreins, kininases and kinins) are able to regulate BP and blood glucose levels via promoting; vasodilator, natriuretic effects and glucose metabolism. These components are located in the cardiac tissue, kidney and vascular smooth muscle. The kinin system is found to be abnormally depressed in various experimental animal models of hypertension and diabetes which might be responsible for inducing cardiac complications. It has been pointed out that the development of a compound having renal kallikrein-like activity may serve the purpose of excreting excessive sodium from the kidney in the treatment of hypertension. Also it has been demonstrated that transgenic mice over-expressing renal tissue kallikrein were hypotensive and that administration of aprotinin, a tissue kallikrein inhibitor, restored the BP of the transgenic mice. These findings highlight a role of tissue kallikrein in the regulation of BP. It has been proposed that tissue kallikrein gene delivery into various hypertensive models exhibits protection, such as reduction in high BP, attenuation of cardiac hypertrophy, inhibition of renal damage and stenosis. This may indicate the prospect of this kallikrein gene therapy for cardiovascular pathology. Several reports indicate that urinary kallikrein excretion is decreased in essential hypertension in humans and in models of experimental hypertension. Thus, reduced urinary kallikrein may reflect a deficiency in the endogenous kallikrein/kinin vasodilatory system that contributes to the pathogenesis of hypertension. Previous studies conducted in white and black population in the USA demonstrated that urinary kallikrein excretion is diminished in family members at risk for hereditary (essential) hypertension and that urinary kallikrein is one of the major genetic markers associated with family history of hypertension. Also evidence for genetic linkage between the kallikrein locus and blood pressure has been reported in the rat. Previous studies have suggested that diminished urinary kallikrein excretion is associated with salt sensitivity of blood pressure. Kallikrein excretion is also diminished in African-Americans and deficiency of the kallikreinkinin renal vasodilatory system may explain many of the unique features of essential hypertension and heart diseases in some black subjects.

Las enfermedades cardiovasculares son la causa más frecuente de muerte en todo el mundo. En Kuwait, la muerte relacionada a enfermedades cardiovasculares puede llegar al 40%. De este modo, será la mayor carga para el sistema de salud del siglo XXI. La hipertensión y la diabetes son los dos factores de riesgo mayores en la producción de hipertrofia cardíaca, cardiopatía isquémica, insuficiencia cardíaca, arritmias cardíacas e infarto de miocardio. Los componentes del sistema calicreína-quinina (quininógenos plasmáticos y urinarios, calicreínas, quininasas y quininas) regulan la PA y los niveles de glucosa sanguínea mediante estimulación de la vasodilatación, efectos natriuréticos y metabolismo de la glucosa. Esos componentes se localizan en el tejido cardíaco, riñón y células musculares lisas. El sistema quinina se encuentra deprimido anormalmente en varios modelos animales experimentales de hipertensión y diabetes que podrían ser responsables de la aparición de complicaciones cardíacas. Se ha señalado que el hallar un compuesto con actividad renal similar a la calicreína puede ser útil al propósito de excretar el exceso de sodio por el riñón en el tratamiento de la hipertensión. También se demostró que los ratones transgénicos que sobreexpresan calicreína por el tejido renal eran hipotensos y que la administración de aprotinina, un inhibidor de la calicreína tisular, restaura la PA de los ratones transgénicos. Esos hallazgos realzaron el papel de la calicreína tisular en la regulación de la PA. Se ha propuesto que el gen de la calicreína tisular entregado en varios modelos de hipertensión ejerce protección, como reducción de la PA aumentada, atenuación de la hipertrofia cardíaca, inhibición de la estenosis y del daño renal. Esto puede indicar la posibilidad de este tratamiento con gen de calicreína para trastronos cardiovasculares. Varios informes indican que la excreción urinaria de calicreína está disminuida en la hipertensión esencial en humanos y en modelos de hipertensión experimental. De este modo, la reducción de la calicreína urinaria puede reflejar una deficiencia en el sistema vasodilatador endógeno calicreína/quinina que contribuye a la patogénesis de la hipertensión. Algunos estudios previos realizados en la población blanca y negra en los EE.UU. demostraron que la excreción urinaria de calicreína está disminuida en miembros familiares con riesgo de hipertensión hereditaria (esencial) y que la calicreína urinaria es uno de los mayores marcadores genéticos asociado con historia familiar de hipertensión. También se informó la existencia de unión genética entre el locus de la calicreína y la presión sanguínea, en ratas. Algunos estudios sugirieron que la excreción urinaria disminuida de calicreína se asocia con sensibilidad de la presión sanguínea a la sal. La excreción de calicreína también está disminuida en afroamericanos y la deficiencia del sistema renal vasodilatador de calicreína-quinina puede explicar muchas de las características singulares de la hipertensión y de las enfermedades cardíacas en algunos sujetos de raza negra.

¿Qué hay de nuevo sobre el PSA? / What's new about the PSA?

La determinación del PSA total forma parte integrante del diagnóstico del cáncer de próstata. Es importante conocer sus variaciones fisiopatológicas, una de las cuales es el aumento con la edad, la hipertrofia benigna, las intervenciones y las diversas afecciones de la próstata. La dificultad principal radica en el umbral del PSA a partir del cual la biopsia está indicada, y que actualmente es de 4 µg/L. Aún no hay consenso, pero estudios recientemente publicados preconizan bajarlo a 2,5 µg/L y añadirle la velocidad y la densidad del PSA. Para valores normales encontrados, la ANAES (Agence Nationale d'Accréditation et d'Evaluation en Santé, actual Haute Autorité de Santé) preconiza desde septiembre de 2004 los plazos pertinentes a respetar para repetir la cuantificación del PSA.

Substratos e inibidores para a calicreína tissular humana / Human tissue kallikrein substrates and inhibitors

Este trabalho focaliza o sistema calicreína cinina, do ponto de vista da - interaçao da calicreína tissular humana com seu inibidor proteico do tipo serpina, denominado calistatina. Para tanto, o mapeamento dos determinantes de especificidade da calicreína tissular humana foram explorados através do uso de substratos peptídicos sintéticos com supressao intramolecular de fluorescência. Este estudo, associado a informaçoes anteriores a cerca da especificidade desta enzima sobre seqüências derivadas do cininogênio humano foram capazes de gerar inibidores competitivos com alta afinidade, contendo inclusive aminoácidos nao naturais. Além disso, o estudo da calistatina também foi abordado, procurando-se determinar alguns dos fatores que podem regular sua atividade fisiológica, como a interaçao com heparina e a sua destruiçao por outras proteases. Os resultados apresentados indicam que o centro reativo da calistatina foi capaz de servir de molde para a síntese de inibidores eficazes para a calicreína tissular humana e que pequenos peptídeos, contendo em sua seqüência aminoácido nao naturais apresentam um caminho potencial para o desenvolvimento de inibidores peptídicos específicos. Estudamos ainda a interaçao da heparina com calistatina e apresentamos evidencias do mecanismo de sua perda de funçao inibitória do complexo. Finalmente, abordamos a da calistatina e da antiquimotripsina pela catepsina D, que além evidenciar um possível papel desta enzima no controle destas serpinas, coletamos informaçoes relevantes sobre a especificidade da catepsina D

Kallikrein-like amidase activity in renal ischemia and reperfusion

We assessed a kallikrein-like amidase activity probably related to the kallikrein-kinin system, as well as the participation of leukocyte infiltration in renal ischemia and reperfusion. Male C57BL/KSJmdb mice were subjected to 20 or 60 min of ischemia and to different periods of reperfusion. A control group consisted of sham-operated mice, under similar conditions, except for ischemia induction. Kallikrein-like amidase activity, Evans blue extravasation and myeloperoxidase activity were measured in kidney homogenates, previously perfused with 0.9 percent NaCl. Plasma creatinine concentration increased only in the 60-min ischemic group. After 20 min of ischemia and 1 or 24 h of reperfusion, no change in kallikrein-like amidase activity or Evans blue extravasation was observed. In the mice subjected to 20 min of ischemia, edema was evident at 1 h of reperfusion, but kidney water content returned to basal levels after 24 h of reperfusion. In the 60-min ischemic group, kallikrein-like amidase activity and Evans blue extravasation showed a similar significant increase along reperfusion time. Kallikrein-like amidase activity increased from 4 nmol PNA mg protein-1 min-1 in the basal condition to 15 nmol PNA mg protein-1 min-1 at 10 h of reperfusion. For dye extravasation the concentration measured was near 200 µg of Evans blue/g dry tissue in the basal condition and 1750 µg of Evans blue/g dry tissue at 10 h of reperfusion. No variation could be detected in the control group. A significant increase from 5 to 40 units of DAbs 655 nm g wet tissue-1 min-1 in the activity of the enzyme myeloperoxidase was observed in the 60-min ischemic group, when it was evaluated after 24 h of reperfusion. Histological analysis of the kidneys showed migration of polymorphonuclear leukocytes from the vascular bed to the interstitial tissue in the 60-min ischemic group after 24 h of reperfusion. We conclude that the duration of ischemia is critical for the development of damage during reperfusion and that the increase in renal cortex kallikrein-like amidase activity probably released from both the kidney and leukocytes may be responsible, at least in part, for the observed effects, probably through direct induction of increased vascular permeability.

Calicreína plasmática de rato e ativador tecidual recombinate de plasmogênio competem na ligaçäo a células hepáticas: estudo em ratos normais e submetidos a diferentes modelos de lesäo hepática / Plasma-kallikrein and recombinant tissue plasminogen activator in ligante hepatic cells

Nesta tese comparamos características de depuração da calicreína plasmática de rato (RPK) e do ativador tecidual do plasminogênio (tPA) em modelo experimental de perfusão de ligado isolado de rato e em células hepáticas isoladas. Especificamente testamos a hipótese destas duas proteínas competirem pelo mesmo receptor. Como controle para a perfusão de fígado, realizamos perfusão de veia cava inferior e verificamos que o tPA é removido eficientemente tanto veia cava como pelo ligado perfundido. Em contraste, a depuração da RPK pela veia cava é desprezível, comparada a depuração pelo fígado perfundido, O tPA é depurado da CIRCULAÇÃO de ratos em fase aguda e de ratos submetidos ao modelo de cirrose experimental com velocidade menor quando comparado aos ratos normais. A flbrose experimental não altera a depuração do tpA da circulação. A depuração do tPA pelo fígado perfundido de ratos submetidos ao modelo experimental de fase aguda e/ou inativação das células de Kupffer é menor que em ratos normais. Por outro lado, a depuração hepática da RPK aumenta nos animais submetidos ao modelo de fase aguda ou inativação das células de Kupffer. A fibrose ou a cirrose experimental diminui a depuração do tPA pelo fígado perfundido de rato. Por outro lado, a fibrose não altera a depuração hepática da RPK. A depuração hepática do tPA é inibida pela adição de b-galactosídeos (b-lactose octacetato ou metil b-galactosídeo) ou a-metil manosídeo ao líquido de perfusão, sugerindo a participação de lectinas S (inibição por b-galactosídeos) e receptor manosil-específico (inibição por manosídeo). A depuração hepática da RPK diminui somente na presença de metil b-galactosídeo. Entretanto, na depuração das duas proteínas não houve completa inibição pelos carboidratos utilizados sugerindo a presença de outro sistema de reconhecimento para estas proteínas, possivelmente via parte protéica. A ligação do 125I-tPA aos hepatócitos isolados é inibida por metil b-galactosídeo (50 e 100 mM), tiodigalactosídeo (50 mM), tio-b-galactosídeo (O,8 e 3,2 MM). Dextran T-40 não inibe esta ligação. Estes dados reafirmam a participação de lectinas S no reconhecimento do tPA. A marcação indireta da RPK com inibidores marcados radioativamente foi desenvolvida com sucesso. A 125I-a5CK-RPK ou a 3H-DFP-RPK são depuradas eficientemente pelo fígado perfundido de rato, em contraste com a 125I-RPK. O tPA e)...(au)

Effect of glandular kallikrein on distal bicarbonate transport. Role of basolateral Cl-/HCO3- exchanger and vacuolar H(+)-ATPase

The luminal membrane of collecting duct cells, specially the intercalated cells, is normally exposed to active kallikrein. This is due to the specific localization of renal kallikrein in the connecting tubule cells. We have previously reported inhibition of distal bicarbonate secretion by renal kallikrein. The present study was performed to evaluate the participation of basolateral Cl-/HCO3- exchanger and luminal H(+)-ATPase activity of cortical collecting duct segments (CCD) in the mechanism involved in the inhibition of bicarbonate secretion induced by the enzyme. The effect of orthograde injections of 1 microgram/ml (250 U/6.3 mg) pig pancreatic kallikrein, in the absence and presence of 1 mM DIDS (stilbene-disulfonic acid) in the renal tubule system, was evaluated. Urine fractions were collected after two-minutes stop-flow. Changes in the urine fraction (Fr) related to those in free-flow urine samples (Ff) were related to the respective polyfructosan (Inutest) ratio. Renal kallikrein activity (Fr:Ff kallikrein/Fr:Ff polyfructosan) increased significantly in the first 120 microliters urine fraction collected after glandular 1 microgram/ml kallikrein, P < 0.05, (first stop-flow) and after glandular 1 microgram/ml kallikrein plus 1 mM. DIDS P < 0.05 (second stop flow). Bicarbonate secretion rate (Fr:Ff HCO3-/Fr:Ff polyfructosan) of collecting ducts was significantly reduced in the first 120 microliters urine fraction collected, related to control, during the first and second stop-flow periods. No difference was shown in bicarbonate excretion between the first 120 microliters urine fractions collected after administration of glandular kallikrein and glandular kallikrein plus DIDS. To measure H(+)-ATPase activity, rat microdissected cortical collector tubules (CCD) were incubated in the presence of increasing glandular kallikrein doses (A: 93, B: 187 and C: 375 mU/200 microL) in the presence of ouabain (4 microM) and omeprazole (100 microM) to inhibit Na(+)-K(+)-ATPase and H(+)-K(+)-ATPase, respectively. In CCD, bafilomycin-sensitive H(+)-ATPase activity (pmol/mm/min) after increasing kallikrein doses did not differ significantly from control...

Renal function in the diabetic pregnant rats

Diabetic nephropathy is associated with increased urinary albumin and reduce kallikrein excretion. Increased activity of the renal kallikrein-kinin system has been suggested as one of the possible mechanisms underlying diabetic hyperfiltration. The present study shown that kallikrein-kinin system is progressively increased in the diabetic-pregnant rats at 7, 14, 21 days; 48 and 7 days after pregnancy (P<0.05 vs Control). However, this increase during diabetic pregnancy did not reached the levels of control pregnancy. On the other hand albumin excretion shown a significant and progressive renal damage in the diabetic state. These findings suggest that the diabetic pregnancy could impair the renal hemodynamic, but, on the other side could modulate the vasodilatador system at pregnancy in the attempt protect the fetus.