Resistance to ethanol sensitization is associated with a loss of synaptic plasticity in the hippocampus.

Behavioral sensitization to repeated ethanol (EtOH) exposure induces an increase in locomotor activity in mice. However, not all animals express such sensitization. Although the literature indicated that the hippocampus may play a role in EtOH sensitization, it is not known whether behavioral sensitization to EtOH is associated with preferential changes in bidirectional synaptic plasticity, i.e., LTP and LTD, two markers of learning capabilities that have also been shown to be involved in addictive behavior. In the present study, we examined whether the vulnerability to develop and express behavioral sensitization to EtOH is associated with altered bidirectional synaptic plasticity in the CA1 area of the dorsal hippocampus. For this purpose, we analyzed both LTP and LTD in resistant and sensitized mice during the expression phase, i.e., 7 days after 10 days of repeated EtOH i.p. administration. We found that resistant mice showed a lack of LTD without changes in LTP. The lack of LTD was associated with an increase in GluN2A protein level and was not due to an altered level of neuronal activity, since no difference was observed between the number of c-FOS positive neurons in sensitized and resistant mice. Given that both types of synaptic plasticity signals may have distinct roles in specific learning and behaviors, our results suggest that resistant mice could exhibit different phenotypes in terms of learning/memory and addictive behaviors compared to sensitized ones. Synapse 71:e21899, 2017. © 2016 Wiley Periodicals, Inc.

[Gluteal artery based perforator flaps for sacral pressure sore reconstruction in children]. / Colgajos basados en arterias perforantes glúteas para la reconstrucción de úlceras sacras por presión en niños.

INTRODUCTION: Children have much lower incidence of pressure sores (PS) than adults and furthermore, they are diagnosed in earlier stages. Therefore, the reported experience with surgical treatment of advanced pediatric PS is scarce. MATERIAL AND METHODS: We present the surgical treatment of 2 chronic PS stage IV in children aged 11 and 14 years, by means of perforator flaps based on the gluteal arteries: in the first case we used a free-style flap based on a left medial gluteal perforator and in the second a large reusable rotation-advancement flap based on both right superior and inferior gluteal artery perforators. RESULTS: In both patients we achieved a rapid cure with 100% survival of the flaps and a stable cover over a 6 month and 1 year follow-up respectively. CONCLUSIONS: Gluteal artery perforator flaps can produce excellent and durable results in the reconstructive treatment of sacral pressure sores in children. These flaps carry lower morbidity than musculocutaneous flaps and are more reliable than traditional fasciocutaneous flaps. Furthermore they preserve more reconstructive options in case of recurrence during the children's lifetime.

A common polymorphism acts as an intragenic modifier of mutant p53 behaviour.

The p73 protein, a homologue of the tumour-suppressor protein p53, can activate p53-responsive promoters and induce apoptosis in p53-deficient cells. Here we report that some tumour-derived p53 mutants can bind to and inactivate p73. The binding of such mutants is influenced by whether TP53 (encoding p53) codon 72, by virtue of a common polymorphism in the human population, encodes Arg or Pro. The ability of mutant p53 to bind p73, neutralize p73-induced apoptosis and transform cells in cooperation with EJ-Ras was enhanced when codon 72 encoded Arg. We found that the Arg-containing allele was preferentially mutated and retained in squamous cell tumours arising in Arg/Pro germline heterozygotes. Thus, inactivation of p53 family members may contribute to the biological properties of a subset of p53 mutants, and a polymorphic residue within p53 affects mutant behaviour.

Regulatory and metabolic adaptations in the nitrogen assimilation of marine picocyanobacteria.

Prochlorococcus and Synechococcus are the two most abundant photosynthetic organisms on Earth, with a strong influence on the biogeochemical carbon and nitrogen cycles. Early reports demonstrated the streamlining of regulatory mechanisms in nitrogen metabolism and the removal of genes not strictly essential. The availability of a large series of genomes, and the utilization of latest generation molecular techniques have allowed elucidating the main mechanisms developed by marine picocyanobacteria to adapt to the environments where they thrive, with a particular interest in the strains inhabiting oligotrophic oceans. Given that nitrogen is often limited in those environments, a series of studies have explored the strategies utilized by Prochlorococcus and Synechococcus to exploit the low concentrations of nitrogen-containing molecules available in large areas of the oceans. These strategies include the reduction in the GC and the cellular protein contents; the utilization of truncated proteins; a reduced average amount of N in the proteome; the development of metabolic mechanisms to perceive and utilize nanomolar nitrate concentrations; and the reduced responsiveness of key molecular regulatory systems such as NtcA to 2-oxoglutarate. These findings are in sharp contrast with the large body of knowledge obtained in freshwater cyanobacteria. We will outline the main discoveries, stressing their relevance to the ecological success of these important microorganisms.

Relationship of oxidative stress and endothelial dysfunction in sleep apnoea.

The aim of the present study was to evaluate ischaemic reactive hyperaemia (IRH) in obstructive sleep apnoea (OSA) and its relationship with oxidative stress. We studied 69 consecutive patients referred to our Sleep Unit (Reina Sofia University Hospital, Cordoba, Spain). Patients with chronic diseases or those taking medication were excluded. IRH was assessed before and after polysomnography. Morning IRH and oxidative stress markers were compared between patients with (apnoea-hypopnoea index (AHI) ≥ 5) and without (AHI < 5) OSA. Measurements were repeated in 25 severe OSA patients after continuous positive airway pressure (CPAP) therapy. We included 46 OSA patients (mean ± sd AHI 49 ± 32.1) and 23 non-OSA subjects (AHI 3 ± 0.9). The OSA patients showed a significant worsening of morning IRH, and a significant increase in malondialdehyde and 8-hydroxydeoxyguanosine levels. Only the oxygen desaturation index independently explained morning IRH, while malondialdehyde levels showed a weak effect on IRH. In severe OSA patients, IRH improved significantly after CPAP treatment, as did malondialdehyde, 8-hydroxydeoxyguanosine and protein carbonyl levels. In OSA patients, endothelial dysfunction and oxidative stress were observed, and IRH worsened after sleep. The increase in oxidative stress was not associated with IRH, while intermittent hypoxia was strongly associated with IRH. In severe OSA patients, CPAP treatment improved oxidative stress and endothelial function.

[Phimosis: dorsal slit or circumcision?]. / Fimosis: prepucioplastia o circuncisión?

INTRODUCTION: Phimosis is perhaps one of the most frequent consultation on pediatric surgery clinics throught the world. The aim of this study is to compare the two procedures more frequently performed in our hospital: dorsal slit and circumcision. PATIENTS Y METHODS: Retrospective study of 1698 patients who were admitted for elective surgical treatment of phimosis between 2003 and 2009. We analyzed age, surgical and anesthethic times, surgical technique and complications. We also did transversal descriptive study through telephonic survey on parents and patients older than 16 years old. RESULTS: There was 76.6% of dorsal slit (n = 1300) and 23.4% (n= 398) of circumcisions. Mean age was 7.15 years y mean time of follow up was 42.3 months. Surgical time was significantly higher in circumcision (p < 0.0001). There were 3% (n = 51) of reoperations, no differences between groups. We didn't find differences in postoperative stenosis, but bleeding was more frequent in circumcision group (1.7%; p = 0.03). There were no differences on parental appreciation of postoperative pain, or functional and esthetic satisfaction between groups. CONCLUSIONS: We didn't find differences on subjective satisfaction between groups. Even if there are differences n postoperative bleeding, global incidence is very low. In our experience both techniques are valid and safe, so surgeon and parents must jointly make the decision.

Have there been changes in the application of mechanical ventilation in relation to scientific evidence? A multicenter observational study in Mexico. / ¿Se han producido cambios en la aplicación de la ventilación mecánica en relación con la evidencia científica? Estudio multicéntrico en México.

OBJECTIVE: The main study objectives were to describe the practice of mechanical ventilation over an 18-year period in Mexico, and estimate changes in mortality among critical patients subjected to invasive mechanical ventilation (IMV). DESIGN: A retrospective subanalysis of a prospective observational study conducted in 1998, 2004, 2010 and 2016 was carried out. SETTING: Intensive Care Units (ICUs) in Mexico. PARTICIPANTS: Adult patients consecutively enrolled in the ICU during one month and who underwent IMV for more than 12hours or noninvasive mechanical ventilation for more than one hour. Follow-up was performed up to a maximum of 28 days after inclusion. INTERVENTIONS: None. PRINCIPAL VARIABLES OF INTEREST: Age, sex, severity upon admission as estimated by SAPS II, parameters of daily arterial blood gases, treatment and complication variables, date and status at discharge from the ICU and from hospital. RESULTS: A total of 959 patients were included in 81 ICUs. Tidal volume (vt) decreased significantly both in patients with acute respiratory distress syndrome (ARDS) criteria (estimated 8.5ml/kg b.w. in 1998 to 6ml/kg in 2016; P<0.001) and in patients without ARDS (estimated 9ml/kg b.w. in 1998 to 6ml/kg in 2016; P<0.001). The ventilatory protective strategy (defined as vt < 6ml/kg or < 8ml/kg and a plateau pressure < 30cmH2O) was: 19% in 1998, 44% in 2004, 58% in 2010 and 75% in 2016 (P<0.001). The adjusted mortality rate in ICU over the 4 periods was: in 2004, odds ratio (OR) 1.05 (95% confidence interval, 95%CI: 0.73-1.72; P=0.764); in 2010, OR 1.68 (95%CI: 1.13-2.48; P=0.009); in 2016, OR 0.85 (95%CI: 0.60-1.20; P=0.368). CONCLUSIONS: The clinical practice of IMV in Mexican ICUs has been modified over a period of 18 years. The most significant change is the ventilatory strategy based on low vt. These changes have not been associated with significant changes in mortality.

Mixotrophy in marine picocyanobacteria: use of organic compounds by Prochlorococcus and Synechococcus.

Marine picocyanobacteria of the Prochlorococcus and Synechococcus genera have been longtime considered as autotrophic organisms. However, compelling evidence published over the last 15 years shows that these organisms can use different organic compounds containing key elements to survive in oligotrophic oceans, such as N (amino acids, amino sugars), S (dimethylsulfoniopropionate, DMSP), or P (ATP). Furthermore, marine picocyanobacteria can also take up glucose and use it as a source of carbon and energy, despite the fact that this compound is devoid of limiting elements and can also be synthesized by using standard metabolic pathways. This review will outline the main findings suggesting mixotrophy in the marine picocyanobacteria Prochlorococcus and Synechococcus, and its ecological relevance for these important primary producers.

n-3 Fatty acid supplementation in burned paediatric patients.

AIM: To determine the effect of dietary supplementation with n-3 fatty acids (FA) in paediatric burned patients who had less than 20% of total body surface affected. METHODS: Burned patients were randomly assigned into two groups, one of them received a supplement of n-3 FA during 5 weeks; the other group was considered as not n-3 supplemented burned group. A third group of no burned patients was selected as control. Blood samples were collected at admission and in burned groups at the final of the study. Plasma and erythrocyte phospholipid FA composition and some biochemical parameters related to the clinical evolution: total plasma proteins and C3 and C4 complement proteins were determined. RESULTS: In the early post-burn patients, there is an increase in saturated and monounsaturated FAs in plasma phospholipids, and a decrease in polyunsaturated FAs compared with control. These alterations are in favour of proinflammatory response to burn injury. In n-3 FA supplemented group, these changes were further reverted, and a favourable response in the amount of total plasma proteins and in C3 and C4 proteins of the complement system was demonstrated. CONCLUSION: Dietary n-3 FA supplementation might be beneficial for patients suffering thermal injury.

p73 cooperates with Ras in the activation of MAP kinase signaling cascade.

The p73 gene is capable of inducing cell cycle arrest, apoptosis, senescence, differentiation and to cooperate with oncogenic Ras in cellular transformation. Ras can be considered as a branch point in signal transduction, where diverse extracellular stimuli converge. The intensity of the mitogen-activated protein kinase (MAPK) cascade activation influences the cellular response to Ras. Despite the fundamental role of p53 in Ras-induced growth arrest and senescence, it remains unclear how the Ras/MEK/ERK pathway induces growth arrest in the absence of p53. We report here that oncogenic Ras stabilizes p73 resulting in p73 accumulation and enhancement of its activity. p73, in turn, induces a sustained activation of the MAP kinase cascade synergizing with oncogenic Ras. We also found that inhibition of p73 function modifies the cellular outcome to Ras activation inhibiting Ras-dependent differentiation. Here, we show for the first time that there is a signaling loop between Ras-dependent MAPK cascade activation and p73 function.

Calcium-activated potassium currents differentially modulate respiratory rhythm generation.

The pre-Bötzinger complex (PBC) generates eupnea and sighs in normoxia and gasping during hypoxia through particular mixtures of intrinsic and synaptic properties. Among intrinsic properties, little is known about the role of Ca(2+)-activated potassium channels in respiratory rhythms generation. To examine this role, we tested the effects of openers and blockers of the large-conductance (BK) and small-conductance (SK) Ca(2+)-activated potassium channels on the respiratory rhythms recorded both in vitro and in vivo, as well as on the discharge pattern of respiratory neurons in the PBC. Activation of SK channels with 1-ethyl-2-benzimidazolinone (1-EBIO) abolished sigh-like activity and inhibited eupneic-like activity, whereas blockade of SK channels with apamine (APA) increased frequency in both rhythms. In hypoxia, APA did not affect the transition to gasping-like activity. At the cellular level, activation of SK channels abolished pacemaker activity and decreased non-pacemaker neurons discharge; opposite effects were observed with SK blockade. In contrast to SK channel modulation, either activation or blockade of BK channels with NS 1619 or iberiotoxin and paxilline, respectively, produced mild effects on eupneic-like and sigh-like bursts during normoxia in vitro. However, BK blockers prevented the changes associated with the transition to gasping-like activity in vitro and perturbed gasping generation and autoresuscitation in vivo. At the cellular level BK channel modulation did not affect respiratory neurons discharge. We conclude that K(Ca) participate in rhythm generation in a state-dependent manner; SK channels are preferentially involved in rhythm generation in normoxia whereas BK channels participate in the transition to gasping generation in hypoxia.

Molecular heterogeneity in chronic lymphocytic leukemia is dependent on BCR signaling: clinical correlation.

Chronic lymphocytic leukemia (CLL), the most frequent form of adult leukemia in Western countries, is characterized by a highly variable clinical course. Expression profiling of a series of 160 CLL patients allowed interrogating the genes presumably playing a role in pathogenesis, relating the expression of functionally relevant signatures with the time to treatment. First, we identified genes relevant to the biology and prognosis of CLL to build a CLL disease-specific oligonucleotide microarray. Second, we hybridized a training series on the CLL-specific chip, generating a biology-based predictive model. Finally, this model was validated in a new CLL series. Clinical variability in CLL is related with the expression of two gene clusters, associated with B-cell receptor (BCR) signaling and mitogen-activated protein kinase (MAPK) activation, including nuclear factor-kappaB1 (NF-kappaB1). The expression of these clusters identifies three risk-score groups with treatment-free survival probabilities at 5 years of 83, 50 and 17%. This molecular predictor can be applied to early clinical stages of CLL. This signature is related to immunoglobulin variable region somatic hypermutation and surrogate markers. There is a molecular heterogeneity in CLL, dependent on the expression of genes defining BCR and MAPK/NF-kappaB clusters, which can be used to predict time to treatment in early clinical stages.

Viral oncoproteins discriminate between p53 and the p53 homolog p73.

p73 is a recently identified member of the p53 family. Previously it was shown that p73 can, when overproduced in p53-defective tumor cells, activate p53-responsive promoters and induce apoptosis. In this report we describe the generation of anti-p73 monoclonal antibodies and confirm that two previously described p73 isoforms are produced in mammalian cells. Furthermore, we show that these two isoforms can bind to canonical p53 DNA-binding sites in electrophoretic mobility shift assays. Despite the high degree of similarity between p53 and p73, we found that adenovirus E1B 55K, simian virus 40 T, and human papillomavirus E6 do not physically interact with p73. The observation that viral oncoproteins discriminate between p53 and p73 suggests that the functions of these two proteins may differ under physiological conditions. Furthermore, they suggest that inactivation of p73 may not be required for transformation.

Preferential retention of codon 72 arginine p53 in squamous cell carcinomas of the vulva occurs in cancers positive and negative for human papillomavirus.

We have sought to determine the basis for preferential loss of the codon 72 proline (72P) rather than the arginine (72R) allele in squamous cell carcinoma of the vulva with loss of heterozygosity (LOH) in p53. The proportion of cases containing human papillomavirus (HPV) 16 was not statistically different among individuals with either 72RR or 72RP in the germ line (P > 0.99), but p53 LOH was significantly more common in individuals heterozygous 72RP than in 72RR individuals (P = 0.04). LOH more commonly involved the 72P allele in both HPV-positive and HPV-negative cancers. Our results imply that preferential loss of the 72P allele in vulval squamous cell carcinoma occurs by HPV-dependent and -independent mechanisms.

Differential sensitivity of normal and Ha-ras-transformed C3H mouse embryo fibroblasts to tumor necrosis factor: induction of bcl-2, c-myc, and manganese superoxide dismutase in resistant cells.

In this study, we investigated the role of activated Ha-ras oncogene on the growth-regulatory properties of tumor necrosis factor (TNF) in C3H mouse embryo fibroblasts. TNF-resistant 10T1/2 cells transfected with an activated Ha-ras oncogene not only produced tumors in nude mice but also exhibited extreme sensitivity to cytolysis by TNF. TNF-induced cell death was mediated through apoptosis. The differential sensitivity of normal and Ha-ras transformed cells to TNF was not due to differences in the number of TNF receptors on their cell surface. However, TNF-resistant cells, but not sensitive cells, overexpressed bcl-2, c-myc, and manganese superoxide dismutase (MnSOD) mRNA following exposure to TNF. In addition, TNF treatment resulted in a marginal induction of p53 mRNA in both TNF-sensitive and resistant cells. These results suggest that TNF-induced cytotoxicity involves apoptosis and that TNF-induced over-expression of bcl-2, c-myc, and MnSOD genes is associated with TNF resistance in C3H mouse embryo fibroblasts.

Evidence that c-myc mediated apoptosis does not require wild-type p53 during lymphomagenesis.

Deregulation of c-myc, frequently implicated in oncogenesis, is associated with increased cell proliferation and also cell death. Similarly, the p53 tumor suppressor gene commonly mutated in human tumors, is known to induce apoptosis or cell cycle arrest in its wild-type conformation. Genetically altered mice simultaneously overexpressing c-myc and possessing a disrupted p53 gene were used to investigate whether c-myc mediated apoptosis requires wild-type p53. The accelerated development of malignant lymphomas in these mice was found to be a consequence of enhanced proliferation and not reduced apoptosis resulting from the synergistic effect of c-myc overexpression and p53 inactivation.

Differential regulation of endogenous endonuclease activation in isolated murine fibroblast nuclei by ras and bcl-2.

Transfection of a murine fibroblast cell line with an activated form of the Harvey ras oncogene conferred sensitivity to apoptosis induced by various agents. This intrinsic sensitivity to apoptosis correlated with the expression of endogenous endonuclease activity in isolated nuclei that was undetectable in the untransfected parental cell line. Subsequent transfection with the human bcl-2 oncogene prevented the morphological and biochemical features of apoptosis in whole cells, although it failed to confer complete protection against cell death. Furthermore, transfection of the bcl-2 oncogene also inhibited the enhanced endonuclease activity in isolated nuclei. Our results indicate that some of the effects of Ha-ras and bcl-2 and potentially other oncogenes, are exerted on the biochemical machinery of apoptosis at the level of the nucleus.

Evidence that p53 and bcl-2 are regulators of a common cell death pathway important for in vivo lymphomagenesis.

Multistep lymphomagenesis involves the deregulation of oncogenes and inactivation of tumor suppressor genes resulting in altered rates of proliferation as well as apoptotic cell death in tumor cells. The contribution of bcl-2 and p53 to the regulation of cell death during lymphomagenesis is assessed using bcl-2-1g, p53 'knock-out' (p53 KO), and p53 KO/bcl-2 hybrid mice. PCR-SSCP and DNA sequence analysis demonstrated that p53 somatic mutations are uncommon in lymphomas arising in bcl-2-Ig transgenic mice. Reduction in tumor latency was not observed in p53 KO/bcl-2 hybrid mice compared to p53 KO mice. Furthermore, overexpressed bcl-2 suppressed wild-type p53 associated apoptosis following gamma-radiation. These findings indicate that bcl-2 and p53 serve a suppressor and effector function, respectively, of a common cell death pathway. These findings also suggest that p53 somatic mutations provide no selective advantage during in vivo multistep lymphomagenesis in the context of bcl-2 gene deregulation.

Expression of human immunodeficiency virus type I tat results in down-regulation of bcl-2 and induction of apoptosis in hematopoietic cells.

Infection by human immunodeficiency virus type 1 (HIV-1) is characterized by progressive loss of various cell types, mainly CD4+ T lymphocytes. While a passive role for the virus in cell destruction is recognized, it does not account for the vast amount of cell death including those of uninfected "bystander' cells. Since in the past we and others have demonstrated the capacity of the Tat protein of HIV-1 to modulate the expression of various cellular genes and that Tat secreted by HIV-infected cells can be readily taken up by various cell types, we have investigated the role of Tat on inducing apoptosis. Our results indicate that T lymphocytes transfected to constitutively express HIV-1 tat, when grown under serum-free conditions results in rapid apoptosis characterized by typical ultrastructural features and DNA fragmentation. Additionally, we observed that in several hematopoietic cell types, including T and B lymphoid cells and monocytoid cells, the expression of HIV-1 tat results in down-regulation of bcl-2, an oncogene with known potential for inhibition of apoptosis. The tat-mediated down-regulation of bcl-2 was observed at both the transcriptional and translational levels. Also, tat-transfected cells expressed increased amounts of bax, a bcl-2 family protein known to induce apoptosis. While these results support reports in the literature of an active role for tat in inducing cell death in HIV-infected individuals, they point to a new mechanism involving Tat-mediated modulation of bcl-2 and bax.

Bcl-2 inhibits p53 nuclear import following DNA damage.

Bcl-2 is an integral membrane oncoprotein that localizes to membranes of the mitochondria, endoplasmic reticulum, and nuclear envelope. Bcl-2 is a member of a family of cell death regulators and functions to inhibit apoptosis. Using confocal microscopy and immunoblotting we show that the ability of bcl-2 to suppress cell death following genotoxic damage can be a consequence of inhibiting nuclear import of induced wild-type p53 protein. Our data suggests that the ability of bcl-2 to modulate trafficking events is not cell type specific. These data support a 'gatekeeper' mechanism for cell death suppression by bcl-2.