Methotrexate reduces keratinocyte proliferation, migration and induces apoptosis in HaCaT keratinocytes in vitro and reduces wound closure in Skh1 mice in vivo.

The aim of the present work was to evaluate MTX treatment (0.1, 1 and 10 µg mL-1) in vitro in order to characterize its effects on cell proliferation alterations in cell cycle of HaCaT keratinocytes and wound healing in a Skh1 mice treated with MTX (low doses 30 mg kg-1, high doses 200 mg kg-1 and repeated doses at 1.5 mg kg-1). We analyzed the cytotoxic effect of methotrexate by a resazurin assay. The effects in the proliferation, cell cycle and apoptosis of HaCaT cells were analyzed by flow cytometry. The effects of MTX on wound healing in vivo were also analyzed. A trend toward reduction in the resazurin assay was found (p > 0.05). Reduced proliferation was also identified in a clonogenic assay and a CFSE assay (p < 0.05) due to the MTX treatment. A reduction in the G2/M and S phases was observed accompanied by apoptosis induction with increased sub G0 phase and annexin V FITC staining. Effect of MTX was evidenced in vivo on the wound closure process after day 10 (p < 0.05) with alterations in tissue architecture and remodeling. There is a marked effect of MTX on wound healing in vivo in Skh1 mice with implications for long-term therapy and surgical interventions.

Hidrotórax como complicación de la derivación ventriculoperitoneal: caso clínico / Hydrothorax as a complication of a ventricle peritoneal shunt. A case report

La derivación ventriculoperitoneal (DVP) es el tratamiento habitual en caso de hidrocefalia. Se trata de una técnica sencilla y eficaz, pero hasta un 70% de los pacientes presentan algún tipo de complicación a lo largo de su vida. La mayoría de ellas son debidas a infección o disfunción mecánica, siendo las complicaciones torácicas poco frecuentes. El presente caso es uno de los pocos encontrados en la literatura en el que se objetiva hidrotórax como complicación de la DVP sin migración de la punta de catéter y sin ascitis. Describimos el caso de una niña de 2años con DVP. La paciente es diagnosticada de derrame pleural compatible con hidrotórax. Tras análisis de β2-transferrina en líquido pleural se comprobó que se trataba de líquido cefalorraquídeo. El TAC craneal mostró un catéter de DVP normoposicionado. La radiografía y la ecografía de abdomen mostraron punta de catéter bien situada en peritoneo

The ventricle peritoneal (VP) shunt is commonly used in the treatment of hydrocephalus. It is a relatively simple and effective technique, but around 70% of the patients with a VP shunt have a complication in their lifetime. Most of these complications are due to infection or mechanical dysfunction. The thoracic complications are rare. The present case is one of the small number of them found in the literature, describing hydrothorax as a complication of a VP shunt without catheter migration and without ascites. The case is presented of a 2 year-old girl with VP shunt. The patient was diagnosed with pleural effusion compatible with hydrothorax. After finding beta-2-transferrin in the pleural fluid, it was it was shown to be from cerebrospinal fluid. Cranial CT showed the catheter in a proper position, and the Xray and ultrasound showed the catheter correctly positioned in the peritoneum

[Hydrothorax as a complication of a ventricle peritoneal shunt. A case report]. / Hidrotórax como complicación de la derivación ventriculoperitoneal: caso clínico.

The ventricle peritoneal (VP) shunt is commonly used in the treatment of hydrocephalus. It is a relatively simple and effective technique, but around 70% of the patients with a VP shunt have a complication in their lifetime. Most of these complications are due to infection or mechanical dysfunction. The thoracic complications are rare. The present case is one of the small number of them found in the literature, describing hydrothorax as a complication of a VP shunt without catheter migration and without ascites. The case is presented of a 2 year-old girl with VP shunt. The patient was diagnosed with pleural effusion compatible with hydrothorax. After finding beta-2-transferrin in the pleural fluid, it was it was shown to be from cerebrospinal fluid. Cranial CT showed the catheter in a proper position, and the Xray and ultrasound showed the catheter correctly positioned in the peritoneum.

Fallopian metaplastic papillary tumour: an atypical transdifferentiation of the tubal epithelium?

A metaplastic papillary tumor of the Fallopian tube is an extremely uncommon condition, with odd and confusing features that make it difficult to categorize as benign or borderline. Here, we summarize all the published cases to date and document the case of a 41-year-old woman diagnosed with this alteration after her last childbirth and ensuing tubal ligation. One of the tubes was bulky and filled with a caramel-like substance encircling a blurry spot. Light microscopy detailed a slender stalk covered by eosinophilic, columnar plump cells, showing atypical nuclei and focal budding. Mitotic figures were absent. The immunohistochemistry panel was positive for pan-cytokeratin, epithelial membrane antigen, cyclin D1, and hormone receptors. Additionally, a proliferation index of less than 5% was rated using Ki-67. The true nature of this tumor (reactive vs neoplastic) is uncertain. Nonetheless, its association with pregnancy suggests an adaptive change, likely similar to the atypical transdifferentiation proposed for Arias-Stella reaction.

Tumor de células granulares de la vulva / Granular cell tumor of the vulva

Antecedentes. El mioblastoma de células granulares o tumor de Abrikossoff, es una neoplasia poco común de la piel y tejido subcutáneo. Actualmente se cree que su origen es neurogénico, posiblemente de las células de Swann, pero el diagnóstico continúa siendo un enigma. Presentación del caso. Se trata de una mujer de 55 años con tumoración vulvar de 3 cm. La biopsia revela un tumor de células granulares. El tratamiento consiste en la extirpación completa de la lesión. Conclusiones. El tumor de células granulares es una neoplasia infrecuente pero debería ser incluida siempre en el diagnóstico diferencial de las lesiones vulvares, dado que pueden recidivar sin un tratamiento óptimo (AU)

Background. Granular cell myoblastoma or Abrikossoff's tumor is an uncommon neoplasm that can involve the skin and subcutaneous tissue. Currently, its origin is believed to be neurogenic, possibly from Swann cells, but diagnosis remains an enigma. Case presentation. The patient was a 55-year-old woman with a 3-cm vulvar tumor. Biopsy revealed a granular cell tumor. Treatment consisted of complete surgical excision. Conclusions. Granular cell tumor is an uncommon neoplasm but should always be included in the differential diagnosis of vulvar lesions, since then can recur without optimal treatment (AU)

Central Nervous System Strongyloidiasis and Cryptococcosis in an HIV-Infected Patient Starting Antiretroviral Therapy.

We report a case of Strongyloides stercoralis hyperinfection syndrome with central nervous system involvement, in a patient with late human immunodeficiency virus (HIV) infection starting antiretroviral therapy, in whom Strongyloides stercoralis larvae and Cryptococcus neoformans were isolated antemortem from cerebrospinal fluid. Our patient was not from an endemic region for the parasite, so strongyloidiasis was not originally suspected. For this reason, we conclude that Strongyloides stercoralis infection should be suspected in HIV-infected patients starting antiretroviral therapy in order to avoid potential fatal outcomes.

Influence of accumulation of heaps of steel slag on the environment: determination of heavy metals content in the soils.

The presence of high level of heavy metals involves a human healthy risk that could induce chronic diseases. This work reports on the metal contamination due to heaps of steel-slag accumulated during more than 40 years in allotments and industrial areas in the southern part of Madrid (Spain). Several slag and soil samples were collected in an area of 10 km(2) and characterized by different conventional (XRD and XRF) and no so common methods (ESEM, thermoluminescence and EDS-WDS). The analysis reveal the presence of: (i) important amounts of Fe (43%), Mg (26%), Cr (1.1%), Mn (4.6%), S (6.5%) in the form of Fe-rich slag phases (wustite, magnetite...), Si and Ca-rich phases (larnite, ghelenite...), Cr (chromite), Mn (bustamite) and graphite, (ii) traces of some other contaminants such as Cr (7700 ppm), Zn (3500 ppm), Ba (3000 ppm), Pb (700 ppm) or Cu (500 ppm) on pathway soil samples that come from the steel slag, and (iii) Co (13 ppm), Pb (78 ppm) and V (54 ppm) in farmland soil samples. Although the existing heavy metals content is not appropriate for the current use, the extremely high metal contamination of the surrounding areas is more worrying. The properties of the soil farmlands (pH circa 7, 13% of clay, mainly illite, and 1-4% of organic matter content) show suitable conditions for the retention of cationic metals, but further studies on the movilization of these elements have to be performed to determine the possibility of severe human health risks. This sort of study can provide useful information for the politicians regarding the appropriate use of the territory to prevent possible health hazard for the population.

Influence of accumulation of heaps of steel slag on the environment: determination of heavy metals content in the soils

The presence of high level of heavy metals involves a human healthy risk that could induce chronic diseases. This work reports on the metal contamination due to heaps of steel-slag accumulated during more than 40 years in allotments and industrial areas in the southern part of Madrid (Spain). Several slag and soil samples were collected in an area of 10 km² and characterized by different conventional (XRD and XRF) and no so common methods (ESEM, thermoluminescence and EDS-WDS). The analysis reveal the presence of: (i) important amounts of Fe (43 percent), Mg (26 percent), Cr (1.1 percent), Mn (4.6 percent), S (6.5 percent) in the form of Fe-rich slag phases (wustite, magnetite...), Si and Ca-rich phases (larnite, ghelenite...), Cr (chromite), Mn (bustamite) and graphite, (ii) traces of some other contaminants such as Cr (7700 ppm), Zn (3500 ppm), Ba (3000 ppm), Pb (700 ppm) or Cu (500 ppm) on pathway soil samples that come from the steel slag, and (iii) Co (13 ppm), Pb (78 ppm) and V (54 ppm) in farmland soil samples. Although the existing heavy metals content is not appropriate for the current use, the extremely high metal contamination of the surrounding areas is more worrying. The properties of the soil farmlands (pH circa 7, 13 percent of clay, mainly illite, and 1-4 percent of organic matter content) show suitable conditions for the retention of cationic metals, but further studies on the movilization of these elements have to be performed to determine the possibility of severe human health risks. This sort of study can provide useful information for the politicians regarding the appropriate use of the territory to prevent possible health hazard for the population.

A presença de altos niveis de metais pesados envolve riscos à saúde humana e pode induzir doenças crônicas. Este trabalho relata a contaminação metálica causada por pilhas de escória siderúrgica acumulada durante mais de 40 anos em áreas industriais na parte sul de Madrid (Espanha). Amostras de escória e solo foram coletadas em uma área de 10 km² e caracteri-zada por diferentes métodos, convencionais (XRD, XRF) ou não (ESEM, termoluminescência e EDS-WDS). A análise revela a presença de: i) quantidades importantes de Fe (43 por cento), Mg (26 por cento), Cr (1,1 por cento), Mn (4,6 por cento), S (6,5 por cento) formando várias fases ricas em Fe (wüstita, magnetita), Si e Ca (larnita, guelenita), Cr (cromita), Mn (bustamita) e grafite; (ii) tiajos de outros contaminantes, como Cr (7700 ppm), Zn (3500 ppm), Ba (3000 ppm), Pb (700 ppm) e Cu (500 ppm), no solo dos caminhos para as pilhas de resíduos e (iii) Co (13 ppm), Pb (78 ppm) e V (54 ppm) em amostras de solo agrícola. Embora os teores de metais pesados não sejam apropriados para uso corrente, a elevada contaminação de areas adjacentes é mais preocupante. Os solos adjacentes (pH ca. 7, 13 por cento de argila - principalmente ilita) e 1-4 por cento de matéria orgânica mostram condições adequadas para a retenção de cátions, mas outros estudos deverão ser realizados para determinar a possibilidade de riscos à saúde humana. Este tipo de trabalho pode fornecer informaçãoo útil para gestores públicos, com relação ao uso do território e prevenção de riscos à saúde da população.

The use of monoclonal antibodies anti-lectin from freshwater prawn Macrobrachium rosenbergii (DeMan, 1879) in the recognition of protein with lectin activity in Decapod's hemolymph.

We determined the cross-reactivity of a monoclonal antibody against the Macrobrachium rosenbergii lectin with proteins in the hemolymph from Procambarus clarkii (Pc), Procambarus americanus (Pa), Litopenaeus setiferus (Ls), and Pseudothelphusa americana (Psa). Crustaceans' hemolymph agglutinated erythrocytes from rat, mouse, guinea pig, and rabbit. Decapods' hemolymph hemagglutinating activity was inhibited by N-acetylated carbohydrates as well as by antibodies. Western blot assays indicated that the antibodies recognized two main proteins of 97.5 and 80.9 kDa in all hemolymphs studied; moreover, ELISA assays indicated that, in PSa, 7.2% of total proteins showed crossreactivity with antibodies in Pa, Pc, and Lc hemolymphs represented 4.2, 3.1%, and 2.5%, respectively. Our results suggested that antibodies recognize the lectin active site in the crustacean species tested; we propose the use of antibodies as an immunological marker for lectin identification and quantification among crustaceans.

Review: Immunity mechanisms in crustaceans.

Crustacean aquaculture represents a major industry in tropical developing countries. As a result of high culture densities and increasing extension of aquaculture farms, the presence of diseases has also increased, inducing economic losses. Invertebrates, which lack adaptive immune systems, have developed defense systems that respond against antigens on the surface of potential pathogens. The defense mechanisms of crustaceans depend completely on the innate immune system that is activated when pathogen-associated molecular patterns are recognized by soluble or by cell surface host proteins, such as lectins, antimicrobial, clotting, and pattern recognition proteins, which, in turn, activate cellular or humoral effector mechanisms to destroy invading pathogens. This work is aimed at presenting the main characteristics of the crustacean proteins that participate in immune defense by specific recognition of carbohydrate containing molecules, i.e. glycans, glycolipids, glycoproteins, peptidoglycans, or lipopolysaccharides from Gram-negative and Gram-positive bacteria, viruses, or fungi. We review some basic aspects of crustacean effector defense processes, like agglutination, encapsulation, phagocytosis, clottable proteins, and bactericidal activity, induced by these carbohydrate-driven recognition patterns.

The effect of sugars and free amino acids from the freshwater prawn Macrobrachium rosenbergii hemolymph on lectin activity and on oxidative burst.

We determined the effect of low molecular weight components (LMWC) from healthy juvenile and adult Macrobrachium rosenbergii hemolymph on lectin activity and oxidative burst (OB) in hemocytes. In an attempt to identify the LMWC that affect the lectin's hemagglutinating activity or oxidative burst, we determined the hemolymph carbohydrates and free amino acids (FAA) concentration. The LMWC (<2000 Da) were obtained after dialysis of the hemolymph. Our results showed that LMWC from juveniles exerted a greater inhibition on lectin than LMWC from adult hemolymph. Production of superoxide radicals by hemocytes was lower in the presence of juvenile (p<0.05) as compared to adult LMWC. FAA composition of the hemolymph and of LMWC from adults showed higher proportion of alanine (which corresponded to 25% of total FAA) and proline (>20%); whereas, in juveniles, the main FAA identified were glycine (>40%) and alanine (26%). N-Acetyl-D-glucosamine (GlcNAc) was the main sugar residue in the hemolymph and LMWC from juveniles; its concentration was 2.4 times higher than glucose (Glc), whereas, in adults, Glc was the main free sugar residue. Our results suggest that the proportion of FAA and carbohydrates in the hemolymph of M. rosenbergii seems to be correlated with the maturation process; furthermore, the high proportion of free GlcNAc and glycine regulate, in the juvenile stage, lectin activity and cellular oxidative mechanisms, respectively.

Myo-inositol-derived glycolipids with anti-inflammatory activity from Solanum lanceolatum.

Lanceolitols A1-A7 (1-7) and B1-B7 (9-15), two series of new myo-inositol-derived glycolipid analogues, in which a sugar moiety is replaced by a fatty acid esterified myo-inositol moiety, were isolated from the leaves of Solanum lanceolatum. Their structures were elucidated on the basis of spectroscopic analysis (1H NMR, 13C NMR, 1H-1H COSY, HMQC, HMBC, and HRFABMS), as well as chemical analysis. All the compounds showed in vivo anti-inflammatory activity against ear edema in mice produced by 12-O-tetradecanoylphorbol-13-acetate (TPA). In vitro enzyme inhibition studies showed that the mixture of lanceolitols A1-A7 inhibited by 58.56% phospholipase A2 from bee venom, while the mixture of lanceolitols B1-B7 was cyclooxygenase-2 (COX-2) inhibitors (IC50 = 237 microM).

Purification and characterization of a lectin from the white shrimp Litopenaeus setiferus (Crustacea decapoda) hemolymph.

A 291-kDa lectin (LsL) was purified from the hemolymph of the white shrimp Litopenaeus setiferus by affinity chromatography on glutaraldehyde-fixed stroma from rabbit erythrocytes. LsL is a heterotetramer of two 80-kDa and two 52-kDa subunits, with no covalently-liked carbohydrate, and mainly composed by aspartic and glutamic acids, glycine and alanine, with relatively lower methionine and cysteine contents. Edman degradation indicated that the NH2-terminal of the 80-kDa subunit is composed DASNAQKQHDVNFLL, whereas the NH2-terminal of the 52-kDa subunit is blocked. The peptide mass fingerprint of LsL was predicted from tryptic peptides from each subunit by MALDI-TOF, and revealed that each subunit showed 23 and 22%, respectively, homology with the hemocyanin precursor from Litopenaeus vannamei. Circular dichroism analysis revealed beta sheet and alpha helix contents of 52.7 and 6.1%, respectively. LsL agglutinate at higher titers guinea pig, murine, and rabbit erythrocytes its activity is divalent cation-dependent. N-acetylated sugars, such as GlcNAc, GalNAc, and NeuAc, were the most effective inhibitors of the LsL hemagglutinating activity. Sialylated O-glycosylated proteins, such as bovine submaxillary gland mucin, human IgA, and fetuin, showed stronger inhibitory activity than sialylated N-glycosylated proteins, such as human orosomucoid, IgG, transferrin, and lactoferrin. Desialylation of erythrocytes or inhibitory glycoproteins abolished their capacity to bind LsL, confirming the relevance of sialic acid in LsL-ligand interactions.

Participation of serum and membrane lectins on the oxidative burst regulation in Macrobrachium rosenbergii hemocytes.

Using a spectrophotometric NBT reduction assay and phagocytosis, we identified that production of superoxide anions and phagocytic activity of hemocytes from Macrobrachium rosenbergii were significantly higher in the presence of rat, rabbit, and chicken erythrocytes than with human, pig, or horse erythrocytes. Hemocytes stimulated with MrL, MrLMab, or PMA increased 4.7, 5.1, and 6.1 fold, respectively, the oxidative response as compared to non-stimulated hemocytes. MrLMab together with MrL increased 5.7 fold the oxidative capacity of hemocytes as compared to non-stimulated cells. These effects were inhibited with 100 mM GalNAc, GlcNAc, or Neu5Ac and 0.2 microM of sialylated submaxillary gland mucin and fetuin. Piroxicam inhibited (P < 0.05) the production of O(2)(-) induced by MrL, whereas iodoacetamide inhibited the effect of MrLMAb (P < 0.05) in a dose-dependent manner. Our results suggest that MrLMab might activate the oxidative burst through the metabolism of glucose as opposed to MrL which utilizes NADPH-independent mechanisms, very probably through pro-inflammatory metabolites.

Characterization of lectin aggregates in the hemolymph of freshwater prawn Macrobrachium rosenbergii.

In invertebrates, lectins play relevant roles in innate immunity; however, their regulatory mechanisms have not been identified yet. In this work, we purified, by gel filtration and affinity chromatography, lectin aggregates circulating in the hemolymph of the freshwater prawn Macrobrachium rosenbergii and compared their physicochemical properties with a previously described lectin (MrL). High-molecular weight MrL aggregates (MrL-I) lack hemagglutinating activity and showed bands of 62.1, 67.1 and 81.4 kDa, whereas MrL-III, which corresponds to MrL, showed hemagglutinating activity and is constituted by a single 9.6-kDa band as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis. MrL-I and MrL-III showed similar amino acid composition but different carbohydrates concentration. Edman degradation indicated NH2-terminal sequence of five amino acids for the 9.6-kDa MrL-III (DVPLL/A) and eleven for the main 81.4-kDa band identified in MrL-I (DVPLL/AXKQQQD); analysis by MALDI-TOF indicated a different tryptic pattern for MrL-I and MrL-III. MrL-I was recognized by monoclonal antibodies against MrL-III. Circular dichroism indicated that the secondary structure in both proteins is similar and contains 23% of beta-sheet and 24% of alpha-helix. Our results suggest that differential posttranslational processes that favor aggregation are involved in regulating the activity of the lectin.

Morphology of hemocytes from the freshwater prawn Macrobrachium rosenbergii.

Using morphological criteria, we identified three types of blood cells in the freshwater prawn Macrobrachium rosenbergii. Hyaline hemocytes, the most abundant type, have few large cytoplasmic granules, a large nucleocytoplasmic ratio, and lyse spontaneously in the absence of anticoagulant. Granular hemocytes are heterogeneous in size and in density of their granules. They are phagocytic and readily spread on substrates. The third type of hemocytes, identified as undifferentiated hemocytes, are the least abundant. The hemocytes of this economically relevant crustacean are compared with blood cells of other decapods. J. Morphol. 234:147-153, 1997. © 1997 Wiley-Liss, Inc.

Actitudes y prácticas de los cirujanos dentistas con respecto al control de infecciones en la práctica dental / Attitudes and practices of dental surgeons regarding infection control in the dental practice

El virus de la inmunodeficiencia humana (VIH) ha sido un problema clínico y epidemiológico de gran interés. En la práctica odontológica ha causado gran impacto, caracterizándose porque la percepción del público en general y de los profesionales acerca del contagio por VIH-SIDA es conflictiva, tal vez por factores como el miedo y la ignorancia. Debido a que el uso de barreras de protección al contagio no ha sido estudiado adecuadamente en nuestro país y con el fin de conocer las actitudes y prácticas de los dentistas con respecto al control de infecciones en la práctica dental, la presente investigación exploró estos aspectos de la práctica clínica. Un cuestionario se aplicó a 74 dentistas (21-35 años, 62 por ciento mujeres), alumnos de especialidades clínicas en la Divisón de Estudios de Posgrado, Facultad de Odontología, UNAM, en mayo de 1992, prestando particular atención al riesgo de contagio por VIH-SIDA y hepatitis B. Los datos fueron analizados con las pruebas t de Student, chi cuadrado y análisis de varianza de una vía (prueba de Scheffé). Los resultados mostraron que existe una actitud positiva ya que los encuestados aparentemente siguen normas apropiadas para el control de infecciones, utilizan barreras de protección personal y obtienen información actualizada de revistas médicas y pláticas. Un factor que puede influir en estas características es que la mayoría son jóvenes. Por otro lado, es notable que sólo una pequeña parte de los encuestados están vacunados contra la hepatitis B; esto hace pensar que la mayoría de los respondentes desconocen que el riesgo de contagio por hepatitis B es tres veces mayor para los dentistas que para el común de la población. En contraste, El riesgo de contagio por VIH-SIDA es mínimo. A pesar de los resultados obtenidos, en general positivos, es necesario seguir implementando nuevas estrategias clínicas y actualizando los conocimientos de los profesionales. Aunque este estudio fue hecho en una muestra pequeña y no podemos generalizar a toda la comunidad odontológica del país, esperamos que estudios como el presente muestren el camino para que en un futuro los individuos contagiados reciban la atención adecuada y los profesionales puedan prestarla con el mínimo de riesgo

Análisis estructural de las mucinas y su relevancia como receptor de agentes infecciosos / Structural analysis of mucins and their relevance as infectious agents receptor

La mucina es una glicoproteína de elevado peso molecular que se caracteriza por poseer complejas cadenas de oligosacáridos unidos a la proteína por enlaces O-glicosídicos. Se ha observado que estas glicoproteínas, que sirven como receptor de diversos agentes infecciosos, que sirven como receptor de diversos agentes infecciosos, se modifican estructural y funcionalmente como respuesta a procesos inflamatorios o de degeneración cancerosa. Su estudio está limitado por la polidispersión de la mucina, sin embargo en este trabajo se presentan diversos marcadores moleculares específicos para carbohidratos que permiten evidenciar tales transformaciones.