4-Deoxyphorbol inhibits HIV-1 infection in synergism with antiretroviral drugs and reactivates viral reservoirs through PKC/MEK activation synergizing with vorinostat.

Latent HIV reservoirs are the main obstacle to eradicate HIV infection. One strategy proposes to eliminate these viral reservoirs by pharmacologically reactivating the latently infected T cells. We show here that a 4-deoxyphorbol ester derivative isolated from Euphorbia amygdaloides ssp. semiperfoliata, 4ß-dPE A, reactivates HIV-1 from latency and could potentially contribute to decrease the viral reservoir. 4ß-dPE A shows two effects in the HIV replication cycle, infection inhibition and HIV transactivation, similarly to other phorboids PKC agonists such PMA and prostratin and to other diterpene esters such SJ23B. Our data suggest 4ß-dPE A is non-tumorigenic, unlike the related compound PMA. As the compounds are highly similar, the lack of tumorigenicity by 4ß-dPE A could be due to the lack of a long side lipophilic chain that is present in PMA. 4ß-dPE activates HIV transcription at nanomolar concentrations, lower than the concentration needed by other latency reversing agents (LRAs) such as prostratin and similar to bryostatin. PKCθ/MEK activation is required for the transcriptional activity, and thus, anti-latency activity of 4ß-dPE A. However, CD4, CXCR4 and CCR5 receptors down-regulation effect seems to be independent of PCK/MEK, suggesting the existence of at least two different targets for 4ß-dPE A. Furthermore, NF-κb transcription factor is involved in 4ß-dPE HIV reactivation, as previously shown for other PKCs agonists. We also studied the effects of 4ß-dPE A in combination with other LRAs. When 4ß-dPE A was combined with another PKC agonists such as prostratin an antagonic effect was achieved, while, when combined with an HDAC inhibitor such as vorinostat, a strong synergistic effect was obtained. Interestingly, the latency reversing effect of the combination was synergistically diminishing the EC50 value but also increasing the efficacy showed by the drugs alone. In addition, combinations of 4ß-dPE A with antiretroviral drugs as CCR5 antagonist, NRTIs, NNRTIs and PIs, showed a consistent synergistic effect, suggesting that the combination would not interefer with antiretroviral therapy (ART). Finally, 4ß-dPE A induced latent HIV reactivation in CD4 + T cells of infected patients under ART at similar levels than the tumorigenic phorbol derivative PMA, showing a clear reactivation effect. In summary, we describe here the mechanism of action of a new potent deoxyphorbol derivative as a latency reversing agent candidate to decrease the size of HIV reservoirs.

Disponibilidad de técnicas moleculares para niños con tumores de difícil caracterización. Un problema en la Oncopatología pediátrica. Aplicación y difusión de técnicas de diagnóstico complementarias en el ámbito público / Availability of molecular techniques for children with difficult-to-characterize tumors. A problem in pediatric oncopathology. Application and diffusion of complementary diagnostic techniques in the public setting

La revolución de la biología molecular y el desarrollo de la investigación biomédica básica para el diagnóstico y posterior manejo del cáncer infantil han llevado a la necesidad de organización de grupos interdisciplinarios de profesionales, los cuales se encargan de afrontar los nuevos desafíos diagnósticos y terapéuticos. Los sarcomas indiferenciados pediátricos constituyen un grupo heterogéneo de neoplasias malignas de aspecto primitivo y polifenotípico. La categorización de gran parte de este tipo de tumores es posible gracias a la aplicación de técnicas moleculares complementarias al estudio histopatológico. El objetivo del presente estudio fue recategorizar sarcomas indiferenciados mediante la implementación de una nueva metodología diagnóstica. Se efectuaron técnicas de inmunohistoquimica (IHQ), FISH de interfase y RT-PCR a partir de tejido fijado en formol e incluido en parafina en 144 casos de sarcomas indiferenciados. Se logró la recategorización del 95.1% de los casos, arribando a 24 diagnósticos diferentes. Sólo un 4.9% permanece aún como sarcoma indiferenciado o inclasificable. Los resultados alcanzados por este estudio demuestran la importancia de contar con nuevas herramientas diagnósticas a nivel molecular y recursos humanos especializados que posibiliten su correcta implementación para el diagnóstico de neoplasias de difícil caracterización (AU)

The revolution of molecular biology and the development of basic medical research for the diagnosis and subsequent management of childhood cancer have led to a need to organize interdisciplinary groups of professionals in charge of facing new diagnostic and treatment challenges. Childhood undifferentiated sarcomas are a heterogeneous group of malignant neoplasms that are primitive in appearance and have polyphenotypic features. Categorization of a large part of this type of tumor has become possible with molecular techniques as a complement to histopathological studies. The aim of this study was to categorize undifferentiated sarcomas using new diagnostic tools. Immunohistochemistry (IHC), interfase FISH, and RT-PCR techniques were used on formalin-fixed and paraffin-embedded tissues of 144 cases of undifferentiated sarcomas. Overall, 95.1% of the cases could be recategorized resulting in 24 different diagnoses. In only 4.9% the diagnosis of undifferentiated or unclassifiable sarcoma was maintained. These results emphasize the importance of the availability of new diagnostic tools at the molecular level and specialized human resources enabling adequate implementation for the diagnosis of difficult-to-characterize neoplasms (AU)

Pseudoaneurismas arteriales secundarios a fractura de fémur: opciones terapéuticas / Arterial pseudoaneurysms secondary to femur fracture: treatment options

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Long-term fiscal implications of funding assisted reproduction: a generational accounting model for Spain.

The aim of this study was to assess the lifetime economic benefits of assisted reproduction in Spain by calculating the return on this investment. We developed a generational accounting model that simulates the flow of taxes paid by the individual, minus direct government transfers received over the individual's lifetime. The difference between discounted transfers and taxes minus the cost of either IVF or artificial insemination (AI) equals the net fiscal contribution (NFC) of a child conceived through assisted reproduction. We conducted sensitivity analysis to test the robustness of our results under various macroeconomic scenarios. A child conceived through assisted reproduction would contribute €370,482 in net taxes to the Spanish Treasury and would receive €275,972 in transfers over their lifetime. Taking into account that only 75% of assisted reproduction pregnancies are successful, the NFC was estimated at €66,709 for IVF-conceived children and €67,253 for AI-conceived children. The return on investment for each euro invested was €15.98 for IVF and €18.53 for AI. The long-term NFC of a child conceived through assisted reproduction could range from €466,379 to €-9,529 (IVF) and from €466,923 to €-8,985 (AI). The return on investment would vary between €-2.28 and €111.75 (IVF), and €-2.48 and €128.66 (AI) for each euro invested. The break-even point at which the financial position would begin to favour the Spanish Treasury ranges between 29 and 41 years of age. Investment in assisted reproductive techniques may lead to positive discounted future fiscal revenue, notwithstanding its beneficial psychological effect for infertile couples in Spain.

Reflux patterns and risk factors of primary varicose veins' clinical severity.

OBJECTIVES: Primarily, to determine the association between the clinical severity of primary varicose veins and different reflux patterns in an anatomic and haemodynamic clinical study using duplex ultrasonography (DU). Secondly, to analyse the association of clinical severity with other aspects, such as risk factors for chronic venous insufficiency (CVI) and other concurrent diseases. METHOD: A total of 2036 limbs were evaluated using DU. Clinical status was characterized by the CEAP (clinical, aetiological, anatomical and pathological elements) classification. The degree of clinical severity was grouped into two categories, mild to moderate CVI (C1­C3)and severe CVI, characterized by the presence of skin changes (C4­C6). We analysed the association of the different reflux patterns with CEAP status. RESULTS: Saphenofemoral junction (SFJ) reflux of the great saphenous vein (GSV) was associated with the most severe form of the disease (odds ratio [OR] » 2.96; confidence interval [CI] 95%: 2.2­3.8), whereas competent SFJ of the GSV with reflux from proximal veins (OR » 2; CI 95%: 1.4­2.7) and the pure non-saphenous reflux (OR » 4.1; CI 95%:1.8­9.0) were associated with mild to moderate CVI. Obesity increased the frequency of severe CVI 2.7 times (OR » 2.7; CI 95%: 1.6­4.6); being a woman also increased the frequency of more severe disease 1.3 times (OR » 1.3; CI 95%: 1.0­1.7). CONCLUSION: Anatomical and haemodynamic studies by DU are postulated as a useful diagnostic tool that allow, by identifying the pattern of venous reflux of varicose pathology, characterization of the probable association to CVI clinical severity.

Immune response to racotumomab in a child with relapsed neuroblastoma.

Immunotherapy targeting ganglioside antigens is a powerful tool for the treatment of high risk neuroblastoma. However, only treatment with anti-GD2 antibodies has been used in clinical practice and other options may be pursued. We report the use of racotumomab, an anti-idiotype vaccine against N-glycolyl neuraminic acid (NeuGc)- containing gangliosides, eliciting an immune response in a child with relapsed neuroblastoma expressing the NeuGcGM3 ganglioside.

Resultados del tratamiento endovascular de los aneurismas de arteria poplítea / Results of endovascular treatment of popliteal artery aneurysms

Objetivo: Evaluar los resultados a largo plazo del tratamiento endovascular de los aneurismas de arteria poplítea (AAP). Pacientes y métodos: Análisis retrospectivo de una serie de 27 AAP intervenidos en 24 pacientes entre enero de 2004 y diciembre de 2010. Parámetros a valorar: complicaciones, salvamento de extremidad y permeabilidad según el método de Kaplan-Meier. Resultados: La edad media fue de 69±12 años. El diámetro medio de los AAP era de 29,5±13mm, con salida por 3 troncos distales en la mitad. Seis casos eran sintomáticos. Se utilizó técnica híbrida en 25 casos, mediante abordaje de la arteria femoral superficial proximal, y percutánea en 2. Se emplearon endoprótesis Fluency en 21 casos y Viabahn/Hemobahn en 6. Promedio de endoprótesis por caso: 2. Éxito técnico inicial: 100%. Morbimortalidad postoperatoria nula. Estancia media: 2,6±2 días. Control periódico clínico y ecográfico en todos los casos. Con una media de seguimiento de 51±24 meses se produjeron 9 oclusiones, 7 el primer año. Otras complicaciones: 2 migraciones con endoleak tipo I, 2 fracturas y una rotura contenida. Hubo 6 conversiones a cirugía abierta, la mayoría por trombólisis fallida, estando todos los bypass funcionantes. Permeabilidad primaria y secundaria a 4 años: 66 y 72%, respectivamente. Salvamento de la extremidad: 100%. No hubo pérdidas de seguimiento, durante el cual se produjeron 2 exitus por neoplasia. Conclusiones: El tratamiento endovascular de los AAP presenta una baja morbimortalidad y puede ser una alternativa terapéutica a la cirugía convencional(AU)

Objective: To evaluate the long term results of the endovascular treatment of popliteal aneurysms. Patients and methods: A retrospective analysis of 27 popliteal aneurysms in 24 patients operated on between January 2004 and December 2010. Endpoints: Complications, limb salvage and patency using the Kaplan-Meier method. Results: The mean age of the patients was 69±12 years. The mean diameter of aneurysms: 29.5±13mm. There were three run-off vessels patency in 52%. Six aneurysms were symptomatic. The technique with dissection of the proximal superficial femoral artery was used in 25 cases, and percutaneous in two. Fluency endografts were used in 21 cases, and Viabahn/Hemobahn in 6. The average number of stent-grafts per case was two. Initial tecnical success was obtained in 100%. There was no postoperative morbidity or mortality. The average in-hospital stay was 2.6±2 days. Clinical and duplex follow-up was possible in all patients. Nine occlusions (seven during the first year) were documented, with an average follow-up of 51±24 months. Other complications: two migrations with endoleak type I, two fractures, and one contained rupture. There were six conversions to bypass surgery, most of them due to failed thrombolysis, all remaining patent. Primary and secondary patency at 4 years was 66 and 72%, and limb salvage was 100%. No patient was lost in the follow-up, and two died of neoplasia. Conclusions: Endovascular treatment of popliteal aneurysms is a feasible and safe therapeutic alternative to conventional bypass surgery, with low morbidity and mortality(AU)

Engineering platforms for directed evolution of Laccase from Pycnoporus cinnabarinus.

While the Pycnoporus cinnabarinus laccase (PcL) is one of the most promising high-redox-potential enzymes for environmental biocatalysis, its practical use has to date remained limited due to the lack of directed evolution platforms with which to improve its features. Here, we describe the construction of a PcL fusion gene and the optimization of conditions to induce its functional expression in Saccharomyces cerevisiae, facilitating its directed evolution and semirational engineering. The native PcL signal peptide was replaced by the α-factor preproleader, and this construct was subjected to six rounds of evolution coupled to a multiscreening assay based on the oxidation of natural and synthetic redox mediators at more neutral pHs. The laccase total activity was enhanced 8,000-fold: the evolved α-factor preproleader improved secretion levels 40-fold, and several mutations in mature laccase provided a 13.7-fold increase in k(cat). While the pH activity profile was shifted to more neutral values, the thermostability and the broad substrate specificity of PcL were retained. Evolved variants were highly secreted by Aspergillus niger (∼23 mg/liter), which addresses the potential use of this combined-expression system for protein engineering. The mapping of mutations onto the PcL crystal structure shed new light on the oxidation of phenolic and nonphenolic substrates. Furthermore, some mutations arising in the evolved preproleader highlighted its potential for heterologous expression of fungal laccases in yeast (S. cerevisiae).

Tratamiento endovascular de los aneurismas poplíteos / No disponible

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Reflux or not reflux? Reflexiones sobre la publicación anglosajona del término en las varices primarias de los miembros inferiores en relación con nuestro entorno / Reflux or not reflux? Reflections on the term used in English language publications in relation to primary varicose veins of the lower limbs compared to that in Spanish literature

El significado práctico del término "reflux" en el estudio hemodinámico de las varices primarias publicado en la literatura varía en función del marco conceptual que se tenga de las distintas alteraciones hemodinámicas del sistema venoso. En este trabajo se exponen los fundamentos hemodinámicos y las discrepancias más interesantes entre la literatura anglosajona y "nuestro entorno"(AU)

The meaning of "reflux" in the haemodynamic study of primary varicose veins varies in the literature, according to different concepts of haemodynamic disorders of venous system. In this paper we present the haemodynamic basis and the main differences between English and Spanish literature(AU)

Tratamiento endovascular de una fístula aortocava complicado con una endofuga tipo II / Endovascular treatment of an aortocaval fistula complicated by a type II endoleak

Introducción. La fístula aortocava secundaria a rotura de aneurisma de aorta abdominal es una complicaciónmuy poco frecuente con elevada morbimortalidad. Caso clínico. Varón de 71 años de edad que presentaba aneurisma deaorta abdominal infrarrenal roto y fistulizado a cava inferior. Se realizó una intervención urgente mediante endoprótesisaortouniilíaca derecha e injerto femorofemoral. Como complicación, endofuga tipo II por arteria mesentérica inferiorque presurizaba el saco aneurismático y mantenía permeable la fístula. En un segundo tiempo se implantó una endoprótesisen la cava inferior y se embolizó el saco con Onyx ® mediante punción translumbar ecoguiada. En el postoperatorio,hubo mejoría hemodinámica, pero la endofuga y la fístula seguían permeables. Al mes se realizó una nueva intervenciónpor persistencia de la fístula aortocava con débito elevado y pseudoaneurisma femoral derecho de gran tamaño, conexplantación de la endoprótesis aórtica, cierre de la comunicación aortocava, reparación del seudoaneurisma femoral einterposición de prótesis aortobifemoral. La endoprótesis cava ayudó a atenuar la hemorragia intraoperatoria. Controlevolutivo a los dos años: paciente asintomático con endoprótesis cava permeable. Conclusión. Las complicaciones de lacirugía endovascular de la fístula aortocava parecen frecuentes y pueden requerir reconversión abierta si ésta fracasa

Introduction. An aortocaval fistula secondary to rupture of an abdominal aortic aneurysm is a very rarecomplication with high morbidity and mortality rates. Case report. A 71-year-old male who presented an infrarenalabdominal aorta aneurysm that was ruptured and fistulised to the inferior cava. Urgent surgery was performedinvolving a right aortouniiliac stent and a femorofemoral graft. The situation was complicated by a type II endoleak viathe inferior mesenteric artery that put pressure on the aneurysmal sac and kept the fistula patent. Later, a stent wasplaced in the inferior cava and the sac was embolised with Onyx ® by means of ultrasound-guided translumbar puncture.In the post-operative period a haemodynamic improvement was observed, but the endoleak and the fistula remainedpatent. At one month, a second operation was performed due to persistence of the aortocaval fistula with a high debitand a large right femoral pseudoaneurysm, with explantation of the aortic stent, closure of the aortocaval communication,repair of the femoral pseudo-aneurysm and placement of an aortobifemoral stent. The caval stent helped to reduceintraoperative bleeding. A control visit was carried out at two years and the patient was found to be asymptomatic and thecaval stent was patent. Conclusions. Complications of endovascular surgery carried out to treat aortocaval fistulas seemto be frequent and may require open restructuring if endovascular techniques fail

Effect of pH on the stability of Pleurotus eryngii versatile peroxidase during heterologous production in Emericella nidulans.

Complementary DNA (cDNA) encoding the new versatile peroxidase from the ligninolytic basidiomycete Pleurotus eryngii has been expressed in the ascomycete Emericella nidulans. In recombinant E. nidulans cultures, the pH reached values as high as 8.3, correlating with a sharp decrease in peroxidase activity. Peroxidase was rapidly inactivated at alkaline pH, but was comparatively stable at acidic pH. The peroxidase inactivation in alkaline buffer could be reversed by adding Ca(2+) and lowering the pH. However, reactivation did not result after incubating the enzyme in non-buffered E. nidulans cultures that reached pH 7.5. To optimize recombinant peroxidase production, the effect of controlling the pH in E. nidulans bioreactor cultures was studied. An extended growth period, and a significant increase in the recombinant peroxidase level (5.3-fold higher activity than in the bioreactor without pH control) was obtained when the pH was maintained at 6.8, showing that culture pH is an important parameter for recombinant peroxidase production.

Cirugía de revascularización tras angioplastia-stent renal en la nefropatía isquémica: a propósito de un caso y revisión de la literatura / Revascularisation surgery following renal stent-angioplasty in ischemic nephropathy: a case report and review of the literature

Introducción. La angioplastia y/o stent o la cirugía de revascularización de la arteria renal puede disminuir la hipertensión arterial y la necesidad de fármacos antihipertensivos, pero esta circunstancia puede desempeñar, actualmente, un papel se cundario en la estrategia terapéutica de la nefropatía isquémica, comparado con la posible prevención del fracaso renal. Existe un cambio sustancial en el objetivo principal del tratamiento de las estenosis arterioscleróticas de las arterias renales que van encaminadas a la prevención del fracaso de la función renal. Caso clínico. Presentamos un caso de cirugía de revascularización renal por reestenosis tras angioplastia-stent renal en un riñón único normofuncionante y se revisa la literatura relacionada con la nefropatía isquémica así como los resultados de los distintos procedimientos terapéuticos. Conclusiones. La cirugía de revascularización renal se presenta como una buena opción, avalada por sus aceptables resultados de permeabilidad y supervivencia. Se necesita un estudio clínico aleatorizado y prospectivo que compare el tratamiento médico, la angioplastia con y sin stent, y la cirugía de la arteria renal para definir el método más óptimo de intervención de los pacientes con arteriosclerosis de la arteria renal (AU)

Aneurisma de carótida extracraneal con estenosis crítica contralateral / Extracranial carotid aneurysm with contralateral critical stenosis

Introducción. Los aneurismas de carótida extracraneal son una patología poco frecuente y de etiología variable, aunque con una gran relevancia clínica. Sus manifestaciones son principalmente neurológicas y de origen tromboembólico. El tratamiento de elección es quirúrgico en la mayoría de casos, mediante la resección y revascularización. Caso clínico. Presentamos el caso de un paciente con un aneurisma de bulbo carotídeo, y una estenosis crítica contralateral tratado en nuestro servicio. El diagnóstico se realizó mediante ecografía Doppler asociada a una tomografía axial computarizada y una angiografía. El tratamiento consistió en una endarterectomía carotídea contralateral, seguida ocho semanas después de la resección del aneurisma y la reconstrucción mediante un injerto de Dacron impregnado en rifampicina, y el reimplante de la carótida externa. La evolución posterior ha sido favorable, sin complicaciones neurológicas. Discusión. Los aneurismas de carótida extracraneal son raros, pero tienen una gran relevancia clínica debido al alto porcentaje de complicaciones neurológicas que presentan. Además, debe tenerse en cuenta su posible etiología infecciosa. El tratamiento conservador sólo se justifica en una minoría de los casos, ya que el resto requiere únicamente un tratamiento quirúrgico. Los resultados de la reconstrucción justifican esta actitud; la ligadura carotídea se reserva en escasas indicaciones. Existen pocas publicaciones sobre la terapéutica endoluminal aplicada a esta patología, por lo que aún no puede considerarse una clara alternativa (AU)

Pycnoporus cinnabarinus laccases: an interesting tool for food or non-food applications.

The effects of the addition of ferulic acid and ethanol in P. cinnabarinus ss3 culture medium in fermentor were compared in 15-L fermentor. In the presence of 30 g l(-1) ethanol, laccase activity (270,000 U/L1) was 3-fold higher as compared with ferulic acid-induced cultures, and 150-fold higher as compared with non-induced cultures, respectively. High-quality flax pulp was bleached in a totally-chlorine free (TCF) sequence using a laccase-mediator system constituted by laccase from Pycnoporus cinnabarinus and 1-hydroxybenzotriazole (HBT) as mediator. Up to 90% delignification and strong brightness increase were attained after the laccase-mediator treatment followed by H2O2 bleaching. This TCF sequence was further improved by applying H2O2 under pressurized O2. In this way, up to 82% ISO brightness was obtained (compared with 37% in the initial pulp and 60% in the peroxide-bleached control) as well as very low kappa number. A positive evaluation of the laccase has been also performed in a food application. The colour of a tea-based beverage was significantly improved by incubating an infusion of green tea with the Pycnoporus laccase.

A new versatile peroxidase from Pleurotus.

Lignin peroxidase (LiP) and manganese peroxidase (MnP) have been investigated in Phanerochaete chrysosporium. A third ligninolytic peroxidase has been described in Pleurotus and Bjerkandera. Two of these versatile peroxidases (VPs) have been cloned, sequenced and characterized. They have high affinity for Mn(2+), hydroquinones and dyes, and also oxidize veratryl alcohol, dimethoxybenzene and lignin dimers. The deduced sequences show higher identity with Ph. chrysosporium LiP than MnP, but the molecular models obtained include a Mn(2+)-binding site. Concerning aromatic substrate oxidation, Pl. eryngii VP shows a putative long-range electron transfer pathway from an exposed trytophan to haem. Mutagenesis and chemical modification of this tryptophan and the acidic residues forming the Mn(2+)-binding site confirmed their role in catalysis. The existence of several substrate oxidation sites is supported further by biochemical evidence. Residue conservation in other fungal peroxidases is discussed.

The cloning of a new peroxidase found in lignocellulose cultures of Pleurotus eryngii and sequence comparison with other fungal peroxidases.

We report cloning and sequencing of gene ps1 encoding a versatile peroxidase combining catalytic properties of lignin peroxidase (LiP) and manganese peroxidase (MnP) isolated from lignocellulose cultures of the white-rot fungus Pleurotus eryngii. The gene contains 15 putative introns, and the deduced amino acid sequence consists of a 339-residue mature protein with a 31-residue signal peptide. Several putative response elements were identified in the promoter region. Amino acid residues involved in oxidation of Mn(2+) and aromatic substrates by direct electron transfer to heme and long-range electron transfer from superficial residues as predicted by analogy with Phanerochaete chrysosporium MnP and LiP, respectively. A dendrogram is presented illustrating sequence relationships between 29 fungal peroxidases.