RESUMEN
Bioactive substances can be found in wine lees, a waste from the winemaking industry. This work developed two formulations, a nanoemulsion with coconut oil (NE-OC) and a nanoemulsion with coconut oil and 0.5% of wine lees extract (NE-OC-Ext), to investigate their effect on untreated, bleached, and bleached-colored hair. The oil-in-water (O/W) nanoemulsions were prepared with coconut oil, TweenTM 80, SpanTM 80, AristoflexTM AVC, Conserve NovaMit MFTM, wine lees extract, and deionized water. The hydration measurements were carried out using a Corneometer® CM 825 with the capacitance method. Scanning electron microscopy (SEM) was used to characterize the effect of formulations on hair fibers. Differential Thermal Analysis (DTA) was to assess the thermal stability and compatibility of wine lees and coconut oil in formulations. Compared to NE-OC, NE-OC-Ext showed a greater hydration effect on bleached-colored hair. DTA showed that NE-OC-Ext presented a smaller number of exothermic degradation events than those of NE-OC, suggesting good interaction and compatibility of the wine lees extract in this formulation. This study highlights the value of wine lees, a residue from the winemaking process, and its possibility of use as raw material for the cosmetic hair industry since it shows a greater moisturizing potential in colored hair.
Asunto(s)
Aceite de Coco , Emulsiones , Vino , Vino/análisis , Aceite de Coco/química , Microscopía Electrónica de Rastreo , Cabello/química , Cabello/efectos de los fármacos , Humanos , Tecnología Química Verde/métodosRESUMEN
Neisseria meningitidis, Haemophilus influenzae and Streptococcus pneumoniae are leading causes of meningitis and acute invasive infections. PCR-based methods are widely used for the diagnosis and surveillance of bacterial pathogens because of their high sensitivity, specificity and high-throughput capabilities compared with conventional laboratory methods. This study evaluated a high-resolution melting qualitative PCR analysis method for the simultaneous detection of these three pathogens. The assay has been optimized to detect three species-specific genes of each organism isolated from clinical samples, enabling accurate identification of the etiological agent. The method proved to be highly sensitive and cheaper than the real-time PCR TaqMan® system because it is probe-free; it could be used for the diagnosis of invasive diseases in public health laboratories of developing countries.
Asunto(s)
Meningitis Bacterianas , Neisseria meningitidis , Humanos , Análisis Costo-Beneficio , Meningitis Bacterianas/diagnóstico , Meningitis Bacterianas/microbiología , Neisseria meningitidis/genética , Streptococcus pneumoniae/genética , Reacción en Cadena en Tiempo Real de la Polimerasa/métodos , Sensibilidad y EspecificidadRESUMEN
Atherogenic events promote changes in vessel walls, with alteration of the redox state, and increased activity of matrix metalloproteinases (MMPs). Thus, this study aims to evaluate aortic remodeling, MMP activity, and reactive oxygen species (ROS) levels after treatment with doxycycline in ApoE-/- and ovariectomized mice (OVX). Female ApoE-/--knockout mice (5 weeks) were submitted to ovariectomy surgery to induce experimental menopause. They then received chow enriched with 1% cholesterol to induce hypercholesterolemia. The animals were divided into two experimental groups: ApoE-/-/OVX vehicle and ApoE-/-/OVX doxycycline (30 mg/kg) administered by gavage once a day for 28 days (15th to the 18th week of life). Blood samples were collected to measure total cholesterol and fractions. The aorta was used for morphometry and to measure the activity and expression of MMP-2 and ROS levels. The ApoE-/-/OVX doxycycline group showed no change in total and fraction cholesterol levels. However, there was a reduction in ROS levels, MMP-2 expression, and activity that correlated with a decrease in atherosclerotic lesions relative to the ApoE-/-/OVX vehicle (p > 0.05). Therefore, we conclude that doxycycline in ApoE-/-/OVX animals promotes a reduction in atherosclerotic lesions by reducing ROS and MMP-2 activity and expression.
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Aterosclerosis , Doxiciclina , Animales , Aorta/metabolismo , Apolipoproteínas E/genética , Apolipoproteínas E/metabolismo , Aterosclerosis/metabolismo , Colesterol/metabolismo , Doxiciclina/farmacología , Femenino , Metaloproteinasa 2 de la Matriz/genética , Metaloproteinasa 2 de la Matriz/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Noqueados para ApoE , Especies Reactivas de Oxígeno/metabolismoRESUMEN
A parasitological survey searching monogeneans infesting marine fish was carried out during June 2018 and January 2020 from the coastal zone of Puerto Pizarro, Tumbes (northern Peru) and from the coastal zone of Chorrillos, Lima (central Peru). The gills, skin, nasal cavities, or branchial gill-cover of seven species were sampled. Ten monogenean species assigned to six families and nine genera were identified. The monogeneans Callorhynchocotyle callorhynchi (Manter, 1955); Capsala biparasiticum (Goto, 1894) Price, 1938; Euryhaliotrema sagmatum Kritsky & Boeger, 2002; Listrocephalos kearni Bullard, Payne &Braswell, 2004; Magniexcipula lamothei Bravo-Hollis, 1981; Nasicola klawei (Stunkard, 1962) Yamaguti, 1968; and Pseudorhabdosynochus anulus Violante-Gonzalez & Rojas-Herrera, 2011 are registered for the first time in Peru. While Capsala gregalis (Wagner & Carter, 1967) Chisholm & Whittington, 2007; Heterocotyle margaritae Chero, Cruces, Sáez, Santos & Luque, 2020; and Monocotyle luquei Chero, Cruces, Iannacone, Sanchez, Minaya, Sáez & Alvariño, 2016 have been previously registered in Peruvian waters, however, the region of Tumbes (northern Peru) represent a new locality record for these species.
Durante junio de 2018 y enero de 2020 se realizó un levamiento parasitológico para buscar monogeneos que infectan peces marinos en la zona costera de Puerto Pizarro, departamento de Tumbes (norte de Perú) y en la zona costera de Chorrillos, departamento de Lima (centro de Perú). Se analizaron las branquias, la piel, las cavidades nasales o el opérculo de siete especies. Se identificaron diez especies de monogeneos, asignadas a seis familias y nueve géneros. Los monogeneos Callorhynchocotyle callorhynchi (Manter, 1955); Capsala biparasitica (Goto, 1894) Price, 1938; Euryhaliotrema sagmatum Kritsky y Boeger, 2002; Listrocephalos kearni Bullard, Payne y Braswell, 2004; Magniexcipula lamothei Bravo-Hollis, 1981; Nasicola klawei (Stunkard, 1962) Yamaguti, 1968; y Pseudorhabdosynochus anulus Violante-Gonzalez & Rojas-Herrera, 2011 son registrados por primera vez en Perú. Mientras que, Capsala gregalis (Wagner y Carter, 1967) Chisholm y Whittington, 2007; Heterocotyle margaritae Chero, Cruces, Sáez, Santos & Luque, 2020; y Monocotyle luquei Chero, Cruces, Iannacone, Sanchez, Minaya, Sáez & Alvariño, 2016 han sido previamente registrados en aguas peruanas, sin embargo, la región de Tumbes (norte de Perú) representa un nuevo registro de localidad para estas especies.
RESUMEN
Currently, the world has been devastated by an unprecedented pandemic in this century. The Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the agent of coronavirus disease 2019 (COVID-19), has been causing disorders, dysfunction and morphophysiological alterations in multiple organs as the disease evolves. There is a great scientific community effort to obtain a therapy capable of reaching the multiple affected organs in order to contribute for tissue repair and regeneration. In this regard, mesenchymal stem cells (MSCs) have emerged as potential candidates concerning the promotion of beneficial actions at different stages of COVID-19. MSCs are promising due to the observed therapeutic effects in respiratory preclinical models, as well as in cardiac, vascular, renal and nervous system models. Their immunomodulatory properties and secretion of paracrine mediators, such as cytokines, chemokines, growth factors and extracellular vesicles allow for long range tissue modulation and, particularly, blood-brain barrier crossing. This review focuses on SARS-CoV-2 impact to lungs, kidneys, heart, vasculature and central nervous system while discussing promising MSC's therapeutic mechanisms in each tissue. In addition, MSC's therapeutic effects in high-risk groups for COVID-19, such as obese, diabetic and hypertensive patients are also explored.
Asunto(s)
COVID-19/terapia , Trasplante de Células Madre Mesenquimatosas/métodos , Animales , COVID-19/inmunología , COVID-19/patología , Humanos , Inmunomodulación , Células Madre Mesenquimatosas/inmunología , SARS-CoV-2/inmunología , SARS-CoV-2/aislamiento & purificaciónRESUMEN
Meningococcal disease is a devastating infection caused by Neisseria meningitidis (meningococcus), and it is classified into serogroups according to its polysaccharide capsule composition. In Brazil, serogroup C is the most frequently responsible for the majority of cases, representing a serious public health challenge. In 2010, the meningococcal serogroup C conjugate vaccine was included in the calendar of the National Immunization Program. We have evaluated 163 meningococcal isolates collected during the pre (2006-2010) and post (2011-2016) vaccination periods. Epidemiological data were determined through Multilocus Sequence Typing (MLST) analysis, vaccine antigens and Bexsero Antigen Sequence Typing (BAST) variant. Clonal complex 103 remains the most prevalent in the country with a high number of serogroup C strains to which CC103 is directly associated. A total of 42 different ST were found. The two most prevalent ST were ST-3780 (CC103) with 38 strains and ST-10781, which was not associated with a CC with nine strains. Allele abcZ-276 was reported among 98% of the strains analyzed and it was not found among other CC103 strains worldwide, makes this allele an important genetic marker for a specific new clone only assigned to Brazilian serogroup C strains, ST-3780. FHbp-25 and NHBA-42 peptides were the most prevalent among isolates in both periods studied. BAST-824 and BAST-3073 have been expressed only in CC103 over the studied years, however, it was not possible to associate a BAST variant to a specific CC. Serogroup C phenotype [P1.22,14-6,36-2: F3-9: ST-3780 (CC103)] was the most prevalent according to the antigenic profiles of circulating strains in Brazil (2007-2016). Our study suggests that CC103 is still a major hypervirulent CC circulating in Brazil and ST-3780 is currently spreading all over the country even after the introduction of MenC in 2010.
Asunto(s)
Antígenos Bacterianos/genética , Infecciones Meningocócicas/prevención & control , Vacunas Meningococicas/administración & dosificación , Tipificación de Secuencias Multilocus/métodos , Neisseria meningitidis Serogrupo C/clasificación , Anticuerpos Antibacterianos/metabolismo , Antígenos Bacterianos/inmunología , Brasil , Variación Genética , Humanos , Infecciones Meningocócicas/inmunología , Vacunas Meningococicas/genética , Vacunas Meningococicas/inmunología , Neisseria meningitidis Serogrupo C/genética , Neisseria meningitidis Serogrupo C/inmunología , Neisseria meningitidis Serogrupo C/aislamiento & purificación , Filogenia , Vigilancia de la Población , SerogrupoRESUMEN
Increased reactive oxygen species (ROS) formation may enhance matrix metalloproteinase (MMP)-2 activity and promote cardiovascular dysfunction. We show for the first time that MMP-2 is upstream of increased ROS formation and activates signaling mechanisms impairing redox balance. Incubation of vascular smooth muscle cells (VSMC) with recombinant MMP-2 increased ROS formation assessed with dihydroethidium (DHE) by flow cytometry. This effect was blocked by the antioxidant apocynin or by polyethylene glycol-catalase (PEG-catalase), and by MMP inhibitors (doxycycline or GM6001). Next, we showed in HEK293 cells that MMP-2 transactivates heparin-binding epidermal growth factor (HB-EGF) leading to EGF receptor (EGFR) activation and increased ROS concentrations. This effect was prevented by the EGFR kinase inhibitor Ag1478, and by phospholipase C (PLC) or protein kinase C (PKC) inhibitors (A778 or chelerythrine, respectively), confirming the involvement of EGFR pathway in MMP-2-induce responses. Next, we showed that intraluminal exposure of aortas to MMP-2 increased vascular MMP-2 levels detected by immunofluorescence and gelatinolytic activity (by in situ zimography) in association with increased ROS formation. This effect was inhibited by MMP inhibitors (phenanthroline or doxycycline) and by apocynin or PEG-catalase. MMP-2 also increased aortic contractility to phenylephrine and this effect was prevented by MMP inhibitor GM6001 and by apocynin or PEG-catalase, showing again that increased ROS formation mediates functional effects of MMP-2. These results show that MMP-2 activates the EGFR and triggers downstream signaling pathways increasing ROS formation and promoting vasoconstriction. These findings may have various implications for cardiovascular diseases.
Asunto(s)
Aorta/fisiología , Receptores ErbB/genética , Metaloproteinasa 2 de la Matriz/metabolismo , Músculo Liso Vascular/fisiología , Activación Transcripcional , Vasoconstricción , Animales , Aorta/citología , Línea Celular , Receptores ErbB/metabolismo , Masculino , Músculo Liso Vascular/citología , Oxidación-Reducción , Conejos , Ratas , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Osteoporosis and cardiovascular diseases are common causes of morbidity and mortality worldwide, especially in people aged over 60 years. Osteoporosis is characterized by low bone mineral density, which deteriorates the microarchitecture of bones and increases the risk of bone fractures. Other pathologies also constitute risk factors for the development of osteoporosis, mainly cardiovascular diseases. In fact, a growing number of reports have shown a positive correlation between cardiovascular diseases and low bone mineral density. MMPs are proteases that participate in the organized degradation of the extracellular matrix (ECM) and which play essential physiological roles, such as cardiovascular and bone tissue remodeling. Overexpression of MMPs underlies pathological processes like osteoporosis and cardiovascular diseases. MMP-1, -2, -9, -13, and -14 are expressed in bone tissue and are key players in the digestion of bone matrix by osteoblasts. Considering this relationship between osteometabolic and cardiovascular pathologies and MMPs, this review focuses on the involvement of MMPs in osteoporosis and on their participation in cardiovascular diseases; it also deals with the positive correlation between osteoporosis and cardiovascular diseases. Although there are many drugs to treat osteoporosis, controversies exist. Here, we will describe these controversies and will discuss how inhibition of MMPs could be an alternative strategy to or an adjuvant therapy in the current treatment of osteoporosis.
Asunto(s)
Enfermedades Cardiovasculares/metabolismo , Metaloproteinasas de la Matriz/metabolismo , Osteoporosis/metabolismo , Animales , Enfermedades Cardiovasculares/tratamiento farmacológico , Humanos , Osteoporosis/tratamiento farmacológico , Inhibidores de Proteasas/química , Inhibidores de Proteasas/farmacologíaRESUMEN
A terapia de nutrição enteral é uma opção para pacientes que possuem limitações principalmente no sistemagastrointestinal e sistema neurológico. Para que ocorra a ingestão adequada de substâncias nutricionais, utiliza-se a sonda denutrição enteral. Uma vez que a maioria dos fármacos não está disponível em soluções, elixires, suspensões, é adotada a práticade macerar esses fármacos e os diluir em água. Objetivo: observar a técnica utilizada para triturar, diluir e administrar formulaçõessólidas via sonda. Método: estudo observacional com abordagem qualitativa e quantitativa, utilizando o Diagrama de Ishikawa,o checklist qualitativo e a fórmula quantitativa. Resultado: foi observado um percentual de 21,42% de obstrução das sondas denutrição enteral. Conclusão: a prática de trituração e diluição de fármacos sólidos influencia na obstrução das sondas de nutriçãoenteral, logo, é oportuna a sugestão da integração da equipe de saúde para maior conhecimento sobre formulações sólidas a serem trituradas.
Enteral nutrition therapy is an option for patients who have limitations primarily in the gastrointestinal system andneurological system. For the proper intake of nutritional substances, the enteral nutrition probe is used. Since most drugs are notavailable in solutions, elixirs, suspensions, the practice is to macerate these drugs and dilute them in water. Objective: to observethe technique used to crush, dilute and administer solid formulations via probe. Method: observational study with qualitative andquantitative approach, using the Ishikawa Diagram, the qualitative checklist and the quantitative formula. Result: a percentageof 21.42% of obstruction of the enteral nutrition probes was observed. Conclusion: the practice of grinding and dilution of soliddrugs influences the obstruction of the enteral nutrition probes, so it is timely to suggest the integration of the health team fora better knowledge about solid formulations to be crushed.
La terapia de nutrición enteral es una opción para pacientes que tienen limitaciones principalmente en el sistemagastrointestinal y el sistema neurológico. Para que ocurra la ingestión adecuada de sustancias nutricionales, se utiliza la sonda denutrición enteral. Una vez que la mayoría de los fármacos no están disponibles en soluciones, elixires, suspensiones, se adoptala práctica de macerar estos fármacos y diluirlos en agua. Objetivo: observar la técnica utilizada para triturar, diluir y administrarformulaciones sólidas vía sonda. Método: estudio observacional con enfoque cualitativo y cuantitativo, utilizando el Diagramade Ishikawa, el checklist cualitativo y la fórmula cuantitativa. Resultado: se observó un porcentaje de 21,42% de obstrucción delas sondas de nutrición enteral. Conclusión: la práctica de trituración y dilución de fármacos sólidos influye en la obstrucción delas sondas de nutrición enteral, luego es oportuno la sugerencia de la integración del equipo de salud para mayor conocimientosobre formulaciones sólidas a ser trituradas.
Asunto(s)
Humanos , Nutrición Enteral , Obstrucción del Catéter , Preparaciones Farmacéuticas/administración & dosificación , Stents , Unidades de Cuidados IntensivosRESUMEN
ABSTRACT This study aimed to determine the histological features of the endometrium of bitches, as well as the cell proliferation at specific moments of diestrus, 10, 20, 30, 40, 50 and 60 days post ovulation, correlating the endometrial thickness with the uterine cell proliferation and the metabolic state (weight, blood glucose and plasma cholesterol) of the animals. Therefore, the right and left uterine horns of 26 clinically healthy bitches submitted to ovariohysterectomy were histologically analyzed 10, 20, 30, 40, 50 and 60 days post ovulation. The hematoxylin-eosin and AgNOR staining techniques were performed. All parameters were evaluated by ANOVA and post-hoc Tukey test (p<0.05). The correlation between endometrial thickness and uterine cell proliferation, weight, blood glucose and plasma cholesterol of animals was observed using the Pearson method (p<0.05). In the present study, it is concluded that endometrial thickness does not differ at any of the moments analyzed in diestrus. The endometrial thickness is not influenced by hormones, weight, blood glucose or serum cholesterol of bitches in this phase of the estrous cycle. However, there is greater cell proliferation in the endometrium at day 40 compared to day 60 post ovulation under the influence of the endocrine profile.
Asunto(s)
Animales , Femenino , Perros , Diestro/fisiología , Colesterol/sangre , Proliferación Celular/fisiología , Endometrio/citología , Glucosa/análisis , Factores de Tiempo , Diestro/metabolismo , Endometrio/fisiologíaRESUMEN
This study aimed to determine the histological features of the endometrium of bitches, as well as the cell proliferation at specific moments of diestrus, 10, 20, 30, 40, 50 and 60 days post ovulation, correlating the endometrial thickness with the uterine cell proliferation and the metabolic state (weight, blood glucose and plasma cholesterol) of the animals. Therefore, the right and left uterine horns of 26 clinically healthy bitches submitted to ovariohysterectomy were histologically analyzed 10, 20, 30, 40, 50 and 60 days post ovulation. The hematoxylin-eosin and AgNOR staining techniques were performed. All parameters were evaluated by ANOVA and post-hoc Tukey test (p<0.05). The correlation between endometrial thickness and uterine cell proliferation, weight, blood glucose and plasma cholesterol of animals was observed using the Pearson method (p<0.05). In the present study, it is concluded that endometrial thickness does not differ at any of the moments analyzed in diestrus. The endometrial thickness is not influenced by hormones, weight, blood glucose or serum cholesterol of bitches in this phase of the estrous cycle. However, there is greater cell proliferation in the endometrium at day 40 compared to day 60 post ovulation under the influence of the endocrine profile.
Asunto(s)
Proliferación Celular/fisiología , Colesterol/sangre , Diestro/fisiología , Endometrio/citología , Glucosa/análisis , Animales , Diestro/metabolismo , Perros , Endometrio/fisiología , Femenino , Factores de TiempoRESUMEN
Abstract Background Several in-house PCR-based assays have been described for the detection of bacterial meningitis caused by Neisseria meningitidis, Streptococcus pneumoniae, and Haemophilus influenzae from clinical samples. PCR-based methods targeting different bacterial genes are frequently used by different laboratories worldwide, but no standard method has ever been established. The aim of our study was to compare different in-house and a commercial PCR-based tests for the detection of bacterial pathogens causing meningitis and invasive disease in humans. Methods A total of 110 isolates and 134 clinical samples (99 cerebrospinal fluid and 35 blood samples) collected from suspected cases of invasive disease were analyzed. Specific sets of primers frequently used for PCR-diagnosis of the three pathogens were used and compared with the results achieved using the multiplex approach described here. Several different gene targets were used for each microorganism, namely ctrA, crgA and nspA for N. meningitidis, ply for S. pneumoniae, P6 and bexA for H. influenzae. Results All used methods were fast, specific and sensitive, while some of the targets used for the in-house PCR assay detected lower concentrations of genomic DNA than the commercial method. An additional PCR reaction is described for the differentiation of capsulated and non-capsulated H. influenzae strains, the while commercial method only detects capsulated strains. Conclusions The in-house PCR methods here compared showed to be rapid, sensitive, highly specific, and cheaper than commercial methods. The in-house PCR methods could be easily adopted by public laboratories of developing countries for diagnostic purposes. The best results were achieved using primers targeting the genes nspA, ply, and P6 which were able to detect the lowest DNA concentrations for each specific target.
Asunto(s)
Humanos , Haemophilus influenzae/aislamiento & purificación , Reacción en Cadena de la Polimerasa/métodos , Meningitis por Haemophilus/diagnóstico , Meningitis Meningocócica/diagnóstico , Meningitis Neumocócica/diagnóstico , Neisseria meningitidis/aislamiento & purificación , Streptococcus pneumoniae/aislamiento & purificación , Streptococcus pneumoniae/genética , ADN Bacteriano/genética , Haemophilus influenzae/genética , Sensibilidad y Especificidad , Cartilla de ADN , Meningitis por Haemophilus/microbiología , Meningitis Meningocócica/microbiología , Meningitis Neumocócica/microbiología , Neisseria meningitidis/genéticaRESUMEN
AIMS: To evaluate the role of nitric oxide, reactive oxygen species (ROS), and natriuretic peptide receptor-B activation in C-type natriuretic peptide-anti-contractile effect on Phenylephrine-induced contraction in aorta isolated from septic rats. METHODS AND RESULTS: Cecal ligation and puncture (CLP) surgery was used to induce sepsis in male rats. Vascular reactivity was conducted in rat aorta and resistance mesenteric artery (RMA). Measurement of survival rate, mean arterial pressure (MAP), plasma nitric oxide, specific protein expression, and localization were evaluated. Septic rats had a survival rate about 37% at 4 h after the surgery, and these rats presented hypotension compared to control-operated (Sham) rats. Phenylephrine-induced contraction was decreased in sepsis. C-type natriuretic peptide (CNP) induced anti-contractile effect in aortas. Plasma nitric oxide was increased in sepsis. Nitric oxide-synthase but not natriuretic peptide receptor-B expression was increased in septic rat aortas. C-type natriuretic peptide-anti-contractile effect was dependent on nitric oxide-synthase, ROS, and natriuretic peptide receptor-B activation. Natriuretic peptide receptor-C, protein kinase-Cα mRNA, and basal nicotinamide adenine dinucleotide phosphate (NADPH)-dependent ROS production were lower in septic rats. Phenylephrine and CNP enhanced ROS production. However, stimulated ROS production was low in sepsis. CONCLUSION: CNP induced anti-contractile effect on Phenylephrine contraction in aortas from Sham and septic rats that was dependent on nitric oxide-synthase, ROS, and natriuretic peptide receptor-B activation.
RESUMEN
BACKGROUND: Several in-house PCR-based assays have been described for the detection of bacterial meningitis caused by Neisseria meningitidis, Streptococcus pneumoniae, and Haemophilus influenzae from clinical samples. PCR-based methods targeting different bacterial genes are frequently used by different laboratories worldwide, but no standard method has ever been established. The aim of our study was to compare different in-house and a commercial PCR-based tests for the detection of bacterial pathogens causing meningitis and invasive disease in humans. METHODS: A total of 110 isolates and 134 clinical samples (99 cerebrospinal fluid and 35 blood samples) collected from suspected cases of invasive disease were analyzed. Specific sets of primers frequently used for PCR-diagnosis of the three pathogens were used and compared with the results achieved using the multiplex approach described here. Several different gene targets were used for each microorganism, namely ctrA, crgA and nspA for N. meningitidis, ply for S. pneumoniae, P6 and bexA for H. influenzae. RESULTS: All used methods were fast, specific and sensitive, while some of the targets used for the in-house PCR assay detected lower concentrations of genomic DNA than the commercial method. An additional PCR reaction is described for the differentiation of capsulated and non-capsulated H. influenzae strains, the while commercial method only detects capsulated strains. CONCLUSIONS: The in-house PCR methods here compared showed to be rapid, sensitive, highly specific, and cheaper than commercial methods. The in-house PCR methods could be easily adopted by public laboratories of developing countries for diagnostic purposes. The best results were achieved using primers targeting the genes nspA, ply, and P6 which were able to detect the lowest DNA concentrations for each specific target.
Asunto(s)
Haemophilus influenzae/aislamiento & purificación , Meningitis por Haemophilus/diagnóstico , Meningitis Meningocócica/diagnóstico , Meningitis Neumocócica/diagnóstico , Neisseria meningitidis/aislamiento & purificación , Reacción en Cadena de la Polimerasa/métodos , Streptococcus pneumoniae/aislamiento & purificación , Cartilla de ADN , ADN Bacteriano/genética , Haemophilus influenzae/genética , Humanos , Meningitis por Haemophilus/microbiología , Meningitis Meningocócica/microbiología , Meningitis Neumocócica/microbiología , Neisseria meningitidis/genética , Sensibilidad y Especificidad , Streptococcus pneumoniae/genéticaRESUMEN
Leishmania contains two phosphoglycerate kinase (PGK) genes, PGKB and PGKC, which code for the cytosolic and glycosomal isoforms of the enzyme, respectively. Although differences in PGKB and PGKC transcript and protein levels and isoform activities have been well documented, the mechanisms of control of both transcript and protein abundance have not been described to date. To better understand the regulation of Leishmania PGK expression, we investigated the stabilities of both PGK transcripts using reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) in combination with transcription and trans-splicing inhibitors. Cells were treated with sinefungin and actinomycin D, and RNA decay kinetics were assessed. In addition, immunoblotting and protein synthesis inhibition by cycloheximide were employed to evaluate protein steady states and degradation. We observed increased stabilities of both PGKB mRNA and protein compared with the glycosomal isoform (PGKC). Our results indicate that both post-transcriptional and post-translational events contribute to the distinct expression levels of the PGKB and PGKC isoforms in Leishmania major.
Asunto(s)
Leishmania major/enzimología , Fosfoglicerato Quinasa/genética , Adenosina/análogos & derivados , Adenosina/farmacología , Antiprotozoarios/farmacología , Cicloheximida/farmacología , Citosol/enzimología , Dactinomicina/farmacología , Regulación de la Expresión Génica , Semivida , Immunoblotting , Isoenzimas/química , Isoenzimas/genética , Isoenzimas/metabolismo , Leishmania major/efectos de los fármacos , Leishmania major/genética , Microcuerpos/enzimología , Peso Molecular , Fosfoglicerato Quinasa/química , Fosfoglicerato Quinasa/metabolismo , Inhibidores de la Síntesis de la Proteína/farmacología , ARN Mensajero/metabolismo , ARN Protozoario/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transcripción GenéticaRESUMEN
OBJECTIVE: The present study evaluated the use of a reagent to stabilize the DNA extracted from human dental tissues stored under different temperature conditions and time intervals. MATERIAL AND METHODS: A total of 161 teeth were divided into two distinct groups: intact teeth and isolated dental pulp tissue. The samples were stored with or without the product at different time intervals and temperature. After storage, DNA extraction and genomic DNA quantification were performed using real-time PCR; the fragments of the 32 samples that represented each possible condition were analyzed to find the four pre-selected markers in STR analysis. RESULTS: The results of the quantification showed values ranging from 0.01 to 10,246.88 ng/µL of DNA. The statistical difference in the quantity of DNA was observed when the factors related to the time and temperature of storage were analyzed. In relation to the use of the specific reagent, its use was relevant in the group of intact teeth when they were at room temperature for 30 and 180 days. The analysis of the fragments in the 32 selected samples was possible irrespective of the amount of DNA, confirming that the STR analysis using an automated method yields good results. CONCLUSIONS: The use of a specific reagent showed a significant difference in stabilizing DNA in samples of intact human teeth stored at room temperature for 30 and 180 days, while the results showed no justification for using the product under the other conditions tested.
Asunto(s)
ADN/química , Pulpa Dental , Temperatura , Diente , Fragmentación del ADN , Odontología Forense/métodos , Humanos , Repeticiones de Microsatélite , Reacción en Cadena en Tiempo Real de la Polimerasa , Valores de Referencia , Manejo de Especímenes/métodos , Estadísticas no Paramétricas , Factores de TiempoRESUMEN
This MiniReview describes the essential biochemical and molecular aspects of matrix metalloproteinases (MMPs) and briefly discusses how they engage in different diseases, with particular emphasis on cardiovascular diseases. There is compelling scientific evidence that many MMPs, especially MMP-2, play important roles in the development of cardiovascular diseases; inhibition of these enzymes is beneficial to many cardiovascular conditions, sometimes precluding or postponing end-organ damage and fatal outcomes. Conducting comprehensive discussions and further studies on how MMPs participate in cardiovascular diseases is important, because inhibition of these enzymes may be an alternative or an adjuvant for current cardiovascular disease therapy.
Asunto(s)
Enfermedades Cardiovasculares/enzimología , Metaloproteinasas de la Matriz/metabolismo , Animales , Enfermedades Cardiovasculares/tratamiento farmacológico , Enfermedades Cardiovasculares/patología , Modelos Animales de Enfermedad , Humanos , Inhibidores de la Metaloproteinasa de la Matriz/uso terapéuticoRESUMEN
Objective: The present study evaluated the use of a reagent to stabilize the DNA extracted from human dental tissues stored under different temperature conditions and time intervals. Material and Methods: A total of 161 teeth were divided into two distinct groups: intact teeth and isolated dental pulp tissue. The samples were stored with or without the product at different time intervals and temperature. After storage, DNA extraction and genomic DNA quantification were performed using real-time PCR; the fragments of the 32 samples that represented each possible condition were analyzed to find the four pre-selected markers in STR analysis. Results: The results of the quantification showed values ranging from 0.01 to 10,246.88 ng/μL of DNA. The statistical difference in the quantity of DNA was observed when the factors related to the time and temperature of storage were analyzed. In relation to the use of the specific reagent, its use was relevant in the group of intact teeth when they were at room temperature for 30 and 180 days. The analysis of the fragments in the 32 selected samples was possible irrespective of the amount of DNA, confirming that the STR analysis using an automated method yields good results. Conclusions: The use of a specific reagent showed a significant difference in stabilizing DNA in samples of intact human teeth stored at room temperature for 30 and 180 days, while the results showed no justification for using the product under the other conditions tested. .
Asunto(s)
Humanos , ADN , Pulpa Dental , Temperatura , Diente , Fragmentación del ADN , Odontología Forense/métodos , Repeticiones de Microsatélite , Reacción en Cadena en Tiempo Real de la Polimerasa , Valores de Referencia , Manejo de Especímenes/métodos , Estadísticas no Paramétricas , Factores de TiempoRESUMEN
Increased levels of hydrogen peroxide (H2O2) can initiate protective responses to limit or repair oxidative damage. However, H2O2 signals also fine-tune responses to growth factors and cytokines controlling cell division, differentiation, and proliferation. Because 17ß-estradiol (E2) also plays important roles in these processes, and is considered a major risk factor in the development and progression of endometriosis, this study evaluated whether E2 has an antiapoptotic effect on oxidative stress in endometrial cells in combination with steady-state H2O2 levels ([H2O2]ss). Endometrial stromal cells were prepared from the eutopic endometrium of 18 women with and without endometriosis to produce primary cells. These cells were stimulated with E2 for 20h, exposed to [H2O2]ss, and examined for cell viability, proliferation, and apoptosis. The endometrial cells from women with endometriosis maintained the steady state for 120min at high H2O2 concentrations. When they were pretreated with E2 and exposed to [H2O2]ss, a decrease in apoptosis level was observed compared to the control cells (p<0.01). The endometrial cells from patients with endometriosis subjected to both E2 and [H2O2]ss showed increased ERK phosphorylation. These findings suggested that H2O2 is a signaling molecule that downregulates apoptosis in endometrial cells, supporting the fact that endometriosis, albeit a benign disease, shares some features with cancer such as decreased catalase levels. These results link the E2 effects on [H2O2]ss to resistance to apoptosis and progression of endometriosis.
Asunto(s)
Apoptosis/efectos de los fármacos , Endometriosis/tratamiento farmacológico , Estradiol/farmacología , Peróxido de Hidrógeno/farmacología , Adulto , División Celular/efectos de los fármacos , Células Cultivadas , Endometriosis/patología , Endometrio/citología , Endometrio/efectos de los fármacos , Femenino , Humanos , Persona de Mediana Edad , Estrés Oxidativo/efectos de los fármacosRESUMEN
O receptor B1 de cininas é uma proteína induzível, que mede a vasodilatação em diversos estados patológicos, tais como inflamação, dor e choque séptico. Considerando que sua expressão tem sido apontada como um indicador da gravidade de doenças, por mecanismos que envolvem espécies oxidarivos e nitrosativos, perguntamos se a indução de receptor B1 poderia ser desencadeada em condições pré-oxidantes. Assim, células endoteliais isoladas de um nódulo linfático axilar de camundongo, SVEC4-10E2, foram tratadas com 0,1 a 1.5 mm de H2O2 ou LPS (10g/ml) e mantidas por 3 h em cultura. Análises quantitativas de RT-PCR mostraram que a expressão do receptor B1 de cininas é dependente da dose de H2O2 com um aumento maximo no pulso 0,05mm. A funcionalidade do receptor B1 nas células tratadas com 1mMH2O2 foi similar àquela induzida por LPS, conforme determinado por medidas da taxa de acidificação extracelular em resposta ao agonista de B1, denominado Des-Arg9-bradicinina (DBK). A expressão das enzimas NADPH oxidase e prote´na do choque térmico 90 (hsp90) também foram analisadas, tendo em conta seus papés já bem conhecidos na disfunção endotelial associada com o estresse oxidativo, de moodo que ambas poderiam estar atuando como contribuintes para a indução do receptor B1 em nosso sistema de estudo. O aumento nas concentrações de H2O2 não causou mudança nos níveis de expressão da NADPH oxidase, mas diminui a expressão de hsp90. Esta mitigação nos níveis da hsp90 possivelmente potencializou a indução do receptor B1 por H2O2, já que níveis baixos ou mau funcionamento do hsp90 estão associados com a geração de superóxido por eNOS desacoplado. Portanto, nossos resultados mostram uma indução específica por H2O2 dos receptores de cininas B1 em céluas endoteliais de camundongo. Tal fenômeno pode significar uma resposta adaptativa das céluas às elevadas condições de estresse oxidativo.
The inducible kinin B, receptor mediates vasodilation in several pathological states such as inflammation, pain and septic shock. Since it has been considered and indicator of disease severity by mechanisms involving both oxidative and nitrosative species, we asked whether B1, receptor induction could be triggered by pro-oxidant conditions. Thus, endothelial cells from mouse lymph nodes, SVEC4-10E2, were treated with 0.1-1.5mM H2O2 or 10 ug/mL LPS and kept for 3h in culture. Quantitative RT-PCR analyses show H2O2 dose-dependent expression of the kinin B1 receptor, with the maximum increase at 0.05mM pulse. The functionality of the B1 receptor in 1mM H2O2-treated cells was similar to that induced by LPS, as determined by extracellular acidication rate measurements in response to B1 agonis Des-Arg³-Bradykinin (DBK). The expression of the enzymes NADPH oxidase and heat shock protein 90 (hsp90) was also examined, taking into account their well known roles in endothelial dysfunction associated with oxidative stress, so that both could be acting as contributors to rhe B1 receptor induction in our system. increasing H1O2 concentrations caused no changes in NADPH oxidase, but led to down regulation expression of hsp90. This mitigation of hsp90 levels possibly potentiates the B1 receptor induction by H2O2, since low levels or malfunction of hsp90 are associated with the generation of superoxide by eNOS. These results show a specific upregulation of the kinin B1 recepetor expression induced by H1O2 in mouse endothelial cells. This phenomenon could mean an adaptive response to elevated oxidative stress conditions.