[Importance of the forensic autopsy in the diagnosis of septic shock: a case report]. / Importancia de la autopsia forense en el diagnóstico del shock séptico: a propósito de un caso.

Septic shock sometimes starts with unspecific symptoms that hamper the clinical diagnosis and, therefore an appropriate treatment. When the septic shock follows a fulminating course with a fatal outcome, the etiological diagnosis has to be conducted post-mortem. Sudden unexpected deaths in children and young adults are frequently the object of medico-legal autopsies. Some sudden unexpected deaths have an infectious origin, which requires further analyses, including microbiology, to establish the cause of death. Here, the case of a fatal septic shock in a 19-month old male infant is presented. After a mild foot injury, an infection by Streptococcus pyogenes progressed to septic shock with a fatal outcome as post-mortem studies demonstrated.

Producción de fosfolipasa y proteinasa en cepas de Malassezia pachydermatis aisladas de perros con otitis y sin otitis / Phospholipase and proteinase production by Malassezia pachydermatis isolated in dogs with and without otitis

Antecedentes. Malassezia pachydermatis forma parte de la microbiota cutánea de perros y gatos. M. pachydermatis se ha asociado frecuentemente a otitis externa y dermatitis seborreicas, sobre todo en el perro, y con menor frecuencia en el gato. M. pachydermatis podría actuar como patógeno cuando existen alteraciones en los mecanismos físicos, químicos o inmunológicos de la piel. Se han identificado diversos factores de virulencia como la capacidad de producir estearasas, lipasas, lipooxigenasas, proteinasas, condroitinsulfatasas e hialuronidasas. Objetivos. Se ha estudiado la actividad fosfolipasa medida a pH 6,3 y la actividad proteinasa medida a pH 6,3 y pH 6,8 (pH de oídos de perros con otitis) de cepas de M. pachydermatis aisladas de perros con otitis y sin otitis. Métodos. Se ha estudiado la actividad fosfolipasa mediante un método semicuantitativo con yema de huevo y la actividad proteinasa mediante un método semicuantitativo con agar albúmina sérica bovina. Se ha realizado el estudio en 96 aislamientos de M. pachydermatis, 43 de ellos aislados de perros sin sintomatología clínica de otitis y 52 aislados de perros con otitis. Resultados. Se observó que el 75,8% de los aislamientos presentaron actividad fosfolipasa a pH 6,3 y el 81% presentaron actividad proteinasa medida a pH 6,3, y el 97,9% a pH 6,8. Se detectó una mayor actividad fosfolipasa en cepas aisladas de perros con otitis. Con respecto a la actividad proteinasa, esta fue mayor a pH 6,8. Conclusiones. Estos hallazgos sugieren que la actividad fosfolipasa podría jugar un papel importante en la invasión de los tejidos del hospedador, por lo menos en la otitis crónica canina. Con respecto a la actividad proteinasa, estos hallazgos podrían ayudar a mejorar la terapéutica de la otitis cuando está implicada M. pachydermatis en el proceso, ajustando a pH bajos los tratamientos aplicados (AU)

[Phospholipase and proteinase production by Malassezia pachydermatis isolated in dogs with and without otitis]. / Producción de fosfolipasa y proteinasa en cepas de Malassezia pachydermatis aisladas de perros con otitis y sin otitis.

BACKGROUND: Malassezia pachydermatis is part of the skin microbiota of dogs and cats. M. pachydermatis has been associated with external otitis and seborrhoeic dermatitis, reported more often in dogs than in cats. When the physical, chemical or immunological mechanisms of the skin are altered, M. pachydermatis could act as a pathogen. Thus, several virulence factors, such as the ability to produce esterase, lipase, lipoxygenase, protease, chondroitin sulphatase, and hyaluronidase, have been studied. AIMS: In the present study, we aim to identify the phospholipase activity measured at pH 6.3, and the proteinase activity measured at pH 6.3 and pH 6.8 (pH from ears of dogs with external otitis) of M. pachydermatis strains isolated from dogs with and without external otitis. METHODS: The phospholipase activity was measured using a semi-quantitative method with egg yolk, and the proteinase activity with a semi-quantitative method using bovine serum albumin agar. The study was performed on 96 isolates of M. pachydermatis, 43 isolated from dogs without clinical symptoms of otitis, and 52 isolated from dogs with otitis. RESULTS: In our study, 75.8% of the isolates showed phospholipase activity at pH 6.3, and 81 and 97.9% of them showed proteinase activity measured at pH 6.3 and 6.8, respectively. A higher phospholipase activity was detected in strains isolated from dogs with otitis. The proteinase activity was increased at a pH of 6.8 (97.9%) in comparison to a pH of 6.3 (81%). CONCLUSIONS: Our results suggest that the phospholipase activity may play an important role in the invasion of host tissues in chronic canine otitis cases. The proteinase activity results obtained in this study suggest that a reduction in the pH of the treatment may improve its efficacy in the resolution of M. pachydermatis otitis.

[Methods to evaluate cognitive disorders in animal models]. / Métodos de evaluación de trastornos cognitivos en modelos animales.

INTRODUCTION: The increasing prevalence of cognitive dysfunction and dementia associated, among others, to population aging in developed countries has grown a great interest in the study of the etiopathogenesis of cognitive deficit and the likely pharmacological targets which improve intellectual function or alter the neurodegeneration underlying these symptoms. DEVELOPMENT AND CONCLUSIONS: An essential tool for that purpose is the use of animal models of human-related pathologies which clinically develop with cognitive impairment and dementia. In this review we will analyse the animal models of these disorders and, specially, the main tests that, by means of the observational evolution of the experimental animal, allow assessing its cognitive functions and its modification by experimental treatments that are wanted to investigate for its eventual introduction into clinics.

Antipsoriatic effects of avarol-3'-thiosalicylate are mediated by inhibition of TNF-alpha generation and NF-kappaB activation in mouse skin.

BACKGROUND AND PURPOSE: Avarol is a marine sesquiterpenoid hydroquinone with anti-inflammatory and antipsoriatic properties. The aim of this study was to evaluate the in vitro and in vivo pharmacological behaviour of the derivative avarol-3'-thiosalicylate (TA) on some inflammatory parameters related to the pathogenesis of psoriasis. EXPERIMENTAL APPROACH: Human neutrophils and monocytes as well as the human keratinocyte cell line HaCaT were used to study the effect of TA on oxidative stress, the arachidonic acid pathway, tumour necrosis factor-alpha (TNF-alpha) release and nuclear factor-kappaB (NF-kappaB) activation. All these parameters were also determined in vivo using the zymosan induced mouse air pouch model and the 12-O-tetradecanoylphorbol-13-acetate (TPA) induced mouse epidermal hyperplasia model. KEY RESULTS: TA showed antioxidant properties in human neutrophils and in the hypoxanthine/xanthine oxidase assay. This compound reduced, in a concentration-dependent manner, leukotriene B(4), prostaglandin E(2) and TNF-alpha production in activated leukocytes. Oral and intrapouch administration of TA in the mouse air pouch model produced a dose-dependent reduction of all these inflammatory mediators. TA also inhibited secretory phospholipase A(2) activity and NF-kappaB DNA-binding in HaCaT keratinocytes. In TPA-induced mouse epidermal hyperplasia, topical administration of TA reduced oedema, leukocyte infiltration, eicosanoid levels and TNF-alpha in skin. In addition, interleukin (IL)-1beta and IL-2 production were also inhibited. Finally, TA was also capable of suppressing NF-kappaB nuclear translocation in vivo. CONCLUSIONS AND IMPLICATIONS: TA inhibited several key biomarkers up-regulated in the inflammatory response of psoriatic skin and this compound could be a promising antipsoriatic agent.

Manifestaciones clínicas de la hipersensibilidad a alimentos mediada por IgE / Clinical manifestations of IgE mediated food hipersensitivity

Las reacciones alérgicas a los alimentos dan lugar a respuestas clínicas que afectan al tracto gastrointestinal, la piel y el tracto respiratorio y dependen en su expresión del mecanismo inmunológico implicado en la patogénesis de la reacción. La piel es el órgano diana en las reacciones de hipersensibilidad a alimentos. Los síntomas cutáneos son las manifestacioes más frecuentes en la patología alérgica alimentaria, junto con la clínica digestiva. La manifestación cutánea más frecuente es la urticaria aguada, que puede ir acompañada o no de angioedema. La urticaria se presenta desde el 30% hasta el 60% de pacientes alérgicos a alimentos en diferentes series, siendo en alrededor del 44% de los pacientes manifestación clínica aislada sin acompañarse de afectación de otros órganos. A su vez, en a población infantil la primera causa de urticaria/angioedema de mecanismo igE mediado son los alimentos. Dentro de las manifestaciones digestivas mediadas por IgE se incluyen el síndrome de alergia oral y la anafilaxia gastrointestinal. La sintomatología respiratoria puede producirse por ingestión o por inhalación del alimento, y es mucho menos frecuente que la clínica dermatológica, asociándose usualmente a cuadro generalizado de anafilaxia. Aunue cualquier alimento puede provocar cualquier manifestación clínica, ciertos alimentos han sido citados más frecuentemente como una causa de anafilaxia grave o mortal, como cacahuetes, nueces y mariscos, ocupando un segundo lugar, leche, huevo, pescado y otros. La sintomatología clínica incluye participación variable de la piel (prurito, urticaria, angioedema), gastrointestinal 8naúseas, vómitos, dolor, diarrea), respiratoria (nasal, laríngea, pulmonar) y cardiovascular (hipotensión, síncope, arritmias)

Food allergic reactions include gastrointestinal, cutaneous and respiratory manifestations, depending on the immunological mechanism implicated. Skin and digestive manifestations are the most frequent symptoms in food allergic reactions. Acute urticaria, with or without angioedema, is the most frequent cutaneous symptom. 30 to 60% of food allergic patients have urticaria, without any other clinical manifestation in 44%. Food allergy is the first cause of igE mediated urticaria/angioedema in children. Specific-IgE mediated gastrointestinal manifestations include the oral allergy syndrome and the gastrointestinal anaphylaxis. Respiratory symptoms can be produced eating or breathing the food, and are less frequent than skin manifestations. They usually appear taking part of an anaphylaxis. Any food allergen can produce any clinical manifestations, but some foods had been described several times as responsible of severe anaphylaxis (peanuts, nuts and seafood, and after them milk, egg, fish and other), including skin manifestations (pruritus, urticaria, angioedema), digestive manifstations (nausea, vomiting, abdominal pain, diarrhea), respiratory symptoms(rhinitis, laryngeal or bronquial symptoms) and cardiovascular symptoms (hypotension, arrhythmia)

VIP down-regulates TLR4 expression and TLR4-mediated chemokine production in human rheumatoid synovial fibroblasts.

OBJECTIVES: Vasoactive intestinal peptide (VIP) has demonstrated therapeutic effects in arthritis by inhibiting both innate and acquired immune responses. We investigated the potential effects of VIP in the regulation of Toll-like receptor (TLR) expression and function in synovial fibroblasts from patients with rheumatoid arthritis (RA) and osteoarthritis (OA). METHODS: Cultured fibroblast-like synoviocytes (FLS) were obtained from patients with RA and OA. The effects of VIP on basal or TNF-alpha or lipopolysaccharide (LPS)-induced TLR2, TLR4 and MyD88 expression and its effects on TLR4-mediated CCL2 and CXCL8 chemokine production were studied by reverse transcription-polymerase chain reaction, western blotting and enzyme-linked immunosorbent assay. RESULTS: TLR2, TLR4 and MyD88 mRNA expression was increased in RA FLS compared with OA FLS. The largest increase was observed for TLR4 and there was also overexpression at the protein level in RA FLS. TLR4 and MyD88 mRNA and proteins were induced by LPS and TNF-alpha in RA FLS. VIP down-regulated the induced but not the constitutive expression of TLR4 and MyD88 in RA FLS. VIP treatment decreased CCL2 and CXCL8 chemokine production in response to TLR4 activation with LPS in RA FLS. CONCLUSIONS: We demonstrate that VIP down-regulates LPS and TNF-alpha activation of TLR4 expression and the TLR4 functional response in terms of proinflammatory chemokine production. These studies suggest that the pleiotropic anti-inflammatory actions of VIP involve inhibitory effects on TLR4 expression and signalling.

Anemia hemolítica autoinmune refractaria: tratamiento con Rituximab, anticuerpo monoclonal anti-CD20 / Refractory autoimmune hemolytic anemia: treatment with Rituximab, monoclonal anti-CD20 antibody

La anemia hemolítica autoinmune en la infancia se caracteriza, en ocasiones por un curso severo y refractario al tratamiento. En estos casos a veces es necesario realizar esplenectomía, lo que supone graves riesgos. Rituximab es un nuevo anticuerpo monoclonal anti CD20 que ya se ha utilizado con éxito en esta enfermedad. Presentamos un caso de un niño diagnosticado de AHAI que fue tratado con esteroides, gammaglobulina y al que se le practicó una es-, plenectomia sin éxito. Se le administró Rituximab a dosis de 375 mg/m2/ semanal durante 4 semanas y, simultaneamente, se inició pauta de descenso corticoidea. A los tres meses del tratamiento con Rituximab presentó un ascenso de la cifra de Hb de 3,5 g/dl y un descenso de la cifra de reticulocito del 67 por ciento. En el seguimiento posterior, durante 14 meses, el paciente se ha mantenido asintomático sin tratamiento ninguno. El tratamiento con Rituximab es una alternativa para esos pacientes refractarios al tratamiento antes de contemplar la esplenectomía (AU)

Inhibition of acute and chronic inflammatory responses by the hydroxybenzoquinonic derivative rapanone.

Rapanone (2,5-dihydroxy-3-tridecyl-1,4-benzoquinone), a natural compound isolated from Myrsine guianensis growing in the Andean highlands of Colombia, was studied in different in vitro and in vivo models as a potential antioxidant and anti-inflammatory drug. Rapanone showed a mild anti-lipoperoxidative profile in rat liver microsomes and inhibited potently degranulation (IC(50) of 9.8 microM) and superoxide chemiluminescence (IC(50) of 3.0 microM) in human neutrophils. In addition, rapanone is a selective and potent human synovial PLA(2) inhibitor (IC(50) of 2.6 microM). In vivo experiments using the carrageenan paw oedema and the zymosan air pouch model in mice as well as the adjuvant arthritis model in rats have proved that rapanone is very efficient in controlling the inflammatory process by different administration routes.

New sesquiterpene derivatives from the sponge Dysidea species with a selective inhibitor profile against human phospholipase A2 and other leukocyte functions.

Two new sesquiterpene cyclopentenones, dysidenones A and B (2, 3), and a new sesquiterpene aminoquinone, dysidine (4), all containing the same rearranged drimane skeleton, have been isolated from a Dysidea sp. sponge, along with bolinaquinone (1). The structures were established from 2D NMR data. Bolinaquinone (1), dysidine (4), and a 1:1 mixture of dysidenones A and B (2, 3) significantly inhibited human synovial phospholipase A2 (PLA2) at 10 microM. Compound 4, which shows an IC50 value of 2.0 microM, exerts a higher potency and selectivity toward this enzyme than the reference inhibitor manoalide. In addition, all of these compounds modulated at 10 microM other human leukocyte functions such as the degranulation process measured as elastase release and the superoxide production measured by chemiluminescence.

Dysidotronic acid, a new sesquiterpenoid, inhibits cytokine production and the expression of nitric oxide synthase.

In a previous study, we reported a new bioactive sesquiterpenoid, named dysidotronic acid, to be a potent, selective human synovial phospholipase A(2) inhibitor. Dysidotronic acid is a novel, non-complex manoalide analogue lacking the pyranofuranone ring. We now investigate the effect of this compound on cytokine, nitric oxide and prostanoid generation on the mouse macrophage cell line RAW 264.7, where it showed a dose-dependent inhibition with inhibitory concentration 50% values in the micromolar range. This effect was also confirmed in the mouse air pouch injected with zymosan. Dysidotronic acid inhibited the production of tumor necrosis factor alpha and interleukin-1 beta as well as the production of nitric oxide, prostaglandin E(2) and leukotriene B(4). Decreased nitric oxide generation was the consequence of inhibition of the expression of nitric oxide synthase, whereas PGE(2) and LTB(4) reduction was due to inhibition of arachidonic acid bioavailability through a direct inhibitory effect of dysodotronic acid on secretory phospholipase A(2).

New pyridinium alkaloids from a marine sponge of the genus Spongia with a human phospholipase A(2) inhibitor profile.

Four new bioactive pyridinium alkaloids, named spongidines A-D (5-8), have been isolated from a Vanuatu sponge of the genus Spongia, together with known petrosaspongiolides D (1) and G (2). Compounds 3 and 4 are 21-hydroxy derivatives of petrosaspongiolides K and P. Structure elucidation was accomplished through extensive 2D NMR experiments (COSY, ROESY, HMBC, HMQC) and IR, UV, and FABMS data. All compounds significantly inhibited human synovial phospholipase A(2) (PLA(2)) at 10 microM, with an IC(50) value of 5.8 microM for compound 4, which is the most potent inhibitor, with a higher selectivity toward this enzyme than the reference inhibitor manoalide. Pyridinium alkaloids (5-8) mainly inhibited human synovial PLA(2). Compound 8, which contains a sulfonic acid group, is the most interesting inhibitor.

Synthesis and enzyme inhibitory activities of a series of lipidic diamine and aminoalcohol derivatives on cytosolic and secretory phospholipases A2.

We have synthesised some lipidic diamines and aminoalcohols and examined their behaviour as inhibitors of secretory and cytosolic PLA2. Some structure-activity relationships considerations have been deduced. Compound 14 was a potent and selective inhibitor of cPLA2 and compound 4 showed a dual inhibitory profile against both types of PLA2 while no cytotoxicity at 10 microM on human neutrophils or on murine macrophage line was observed for both.

Co-regulation between cyclo-oxygenase-2 and inducible nitric oxide synthase expression in the time-course of murine inflammation.

Many in vitro studies have used cell cultures to focus on the relationships between cyclo-oxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) isoforms. We have investigated the time-course of regulation and the role of COX-2 and iNOS in a model of experimental inflammation in mice, the air pouch injected with zymosan. This study demonstrates that there is an early acute phase (4 h) mediated mainly by eicosanoids, with high levels of prostaglandin E2 (PGE2) produced by cyclo-oxygenase-1. In addition, in the later phase (from 12 h) there is a participation of nitric oxide (NO) and PGE2 accompanied by co-induction of both iNOS and COX-2. These enzymes were detected in migrating leukocytes as well as in macrophages lining the air pouch. Administration of NS398 or indomethacin inhibited PGE2 levels and COX activity, but also nitrite levels and iNOS activity, which was accompanied by a reduction in iNOS expression. Aminoguanidine inhibited nitrite levels and iNOS activity in addition to exerting inhibitory effects on the COX pathway. Treatment of animals with dexamethasone reduced nitrite and PGE2 concentrations in air pouch exudates, as well as iNOS and COX-2 expression in migrating cells. Our results indicate that PGE2 and NO may play in vivo mutual modulatory roles in the inflammatory response caused by zymosan injection into the mouse air pouch, a suitable model to study drugs acting on those pathways.

In vitro infection of cells of the monocytic/macrophage lineage with bovine leukaemia virus.

The oncogenic retrovirus bovine leukaemia virus (BLV) primarily infects B cells. Most infected animals remain asymptomatic for long periods of time before an increase in circulating B cells or localized tumours can be observed. This long clinical latency period may be explained by cells of the monocyte/macrophage lineage (M/M) becoming infected and acting as a reservoir for the virus, as shown for other retroviruses (human immunodeficiency virus-1, feline immunodeficiency virus). M/M cells in different stages of differentiation (HL-60, THP-1, U-937, J774, BGM, PM2, primary macrophages of sheep and cows) were cultured with BLV produced by permanently infected donor cells (FLKBLV and BLV-bat(2)). Donor cells were inhibited from multiplying by either irradiation or treatment with mitomycin C. In other experiments, supernatant from donor cells containing virus was used. In co-culture with the donor cells, the less differentiated monocytic cells showed severe cellular changes such as differentiation, vacuolization, cell lysis and membrane blebbing; apoptosis was a frequent phenomenon. Budding and extracellular viruses were also observed. The more differentiated macrophage cells, although they showed less signs of infection by microscopy, had a complete BLV protein profile, as seen by Western blotting; bands corresponding to p24CA (Gag) and its precursors were clearly seen. In addition, gp51SU was identified by syncytia formation assays. It is concluded that M/M cells may be infected by BLV, the consequences of the infection differing according to the type of cell.

Cavernolide: a new inhibitor of human sPLA2 sharing unusual chemical features.

The inhibitory effect of cavernolide, a novel C2, terpene lactone isolated from the sponge Fasciospongia cavernosa, on PLA2 and other enzyme activities involved in the inflammatory process was studied. Cavernolide inhibited human synovial sPLA2 in a concentration-dependent manner with an IC50 value of 8.8 microM. Besides, this compound decreased in the nanomolar range the myeloperoxidase degranulation process using different stimuli. Cavernolide also inhibited TNFalpha, NO and PGE2 production in intact cell experiments. NO and PGE2 reduction was the consequence of the inhibition on iNOS and COX-2 expression because it did not affect inducible nitric oxide synthase and cyclooxygenase-2 activities in intact cells.

[Non-compliance of the treatment with antibiotics in non-severe acute infections]. / Incumplimiento del tratamiento con antibióticos en infecciones agudas no graves.

BACKGROUND: To determine the nonfulfillment of antiinfectious therapy in clinical practice. MATERIAL AND METHODS: Fulfillment was quantified by tablet counting (TC) in the homes of 366 patients undergoing antibiotic treatment and the motives and predictive factors were identified. RESULTS: Nonfulfillment was of 61% (95% confidence interval [CI] 55.4-66.6%). Patient improvement was the main reason for discontinuation (54.5%). The predictive factors were greater length of treatment (p = 0.000004), dose (p = 0.0019) and number of tablets (p = 0.0000). CONCLUSIONS: Nonfulfillment of antiinfectious treatment in clinical practice is high, mainly due to clinical improvement and to the greater complexity and length of treatment.