Sarcopenia in Children With Wilms Tumor: A Marker of Undernutrition Which May Impact Adversely on Clinical Outcomes.

The therapeutic approach to Wilms tumor (WT) is multidisciplinary and leads to significant patient impairment, increasing the risk of nutritional compromise and malnutrition. Children with cancer are vulnerable to sarcopenia which has been recognized as a negative impact of anticancer therapy. Recent studies have highlighted the reduction in the total psoas muscle area (TPMA) to be associated with a poor prognosis in many pediatric diseases, including cancer. This study aims to evaluate changes in the TPMA compartment during the treatment of children with WT. An observational, longitudinal, and retrospective study was undertaken in a single institution evaluating children (1 to 14 y, n=38) with WT between 2014 and 2020. TPMA was assessed by the analysis of previously collected, electronically stored computed tomography images of the abdomen obtained at 3 time points: diagnosis, preoperatively, and 1 year after surgery. For all patients, TPMA/age were calculated with a specific online calculator. Our data show a high incidence of sarcopenia (55.3%) at diagnosis which increased after 4 to 6 weeks of neoadjuvant chemotherapy (73.7%) and remained high (78.9%) 1 year after the surgical procedure. Using TPMA/age Z-score curves we have found significant and rapid muscle loss in children with WT, with little or no recovery in the study period.

Late clinical outcomes of myocardial hybrid revascularization versus coronary artery bypass grafting for complex triple-vessel disease: Long-term follow-up of the randomized MERGING clinical trial.

OBJECTIVES: This article aimed to compare the outcomes after hybrid revascularization with conventional coronary artery bypass grafting (CABG) surgery. BACKGROUND: The concept of hybrid coronary revascularization combines the advantages of CABG and percutaneous coronary intervention to improve the treatment of patients with complex multivessel disease. METHODS: The Myocardial hybrid revascularization versus coronary artERy bypass GraftING for complex triple-vessel disease-MERGING study is a pilot randomized trial that allocated 60 patients with complex triple-vessel disease to treatment with hybrid revascularization or conventional CABG (2:1 ratio). The primary outcome was the composite of all-cause death, myocardial infarction, stroke, or unplanned repeat revascularization at 2 years. RESULTS: Clinical and anatomical characteristics were similar between groups. After a mean follow-up of 802 ± 500 days, the primary endpoint rate was 19.3% in the hybrid arm and 5.9% in the CABG arm (p = NS). The incidence of unplanned revascularization increased over time in both groups, reaching 14.5 versus 5.9% in the hybrid and in the CABG groups, respectively (p = .4). Of note, in the hybrid group, there were no reinterventions driven by the occurrence of stent restenosis. CONCLUSIONS: Hybrid myocardial was feasible but associated with increasing rates of major adverse cardiovascular events during 2 years of clinical follow-up, while the control group treated with conventional surgery presented with low rates of complications during the same period. In conclusion, before more definitive data arise, hybrid revascularization should be applied with careful attention in practice, following a selective case-by-case indication.

The arrival of COVID-19 in Brazil and the impact on coronary artery bypass surgery.

BACKGROUND AND AIM OF THE STUDY: This study analyzed the arrival of coronavirus disease 2019 (COVID-19) in Brazil and its impact on coronary artery bypass grafting (CABG) surgery. METHODS: Patients undergoing isolated CABG in six hospitals in Brazil were divided into two periods: pre-COVID-19 (March-May 2019, N = 468) and COVID-19 era (March-May 2020, N = 182). Perioperative data were included on a dedicated REDCap platform. Patients with clinical and tomographic criteria and/or PCR (+) for severe acute respiratory syndrome coronavirus 2 infection were considered COVID-19 (+). Logistic regression analysis was performed to create a multiple predictive model for mortality after CABG in COVID-19 era. RESULTS: Compared to 2019, in 2020, CABG surgeries had a 2.8-fold increased mortality risk (95% confidence interval [CI]: 1-7.6, p = .041), patients who evolved with COVID-19 had a 11-fold increased mortality risk (95% CI: 2.2-54.9, p < .003), rates of morbidities and readmission to the intensive care unit. The surgical volume was decreased by 60%. The model to predict mortality after CABG in the COVID-19 era was validated with good calibration (Hosmer-Lemeshow = 1.43) and discrimination (receiver operating characteristic = 0.78). CONCLUSION: The COVID-19 pandemic had an adverse impact on mortality, morbidity and volume of patients undergoing CABG.

The glucoamylase from Aspergillus wentii: Purification and characterization.

This study describes for the first time the purification and characterization of a glucoamylase from Aspergillus wentii (strain PG18), a species of the Aspergillus genus Cremei section. Maximum enzyme production (∼3.5 U/ml) was obtained in submerged culture (72 h) with starch as the carbon source, at 25°C, and with orbital agitation (100 rpm). The enzyme was purified with one-step molecular exclusion chromatography. The 86 kDa purified enzyme hydrolyzed starch in a zymogram and had activity against p-nitrophenyl α- d-glucopyranoside. The optimal enzyme pH and temperature were 5.0 and 60°C (at pH 5.0), respectively. The Tm of the purified enzyme was 60°C, at pH 7.0. The purified glucoamylase had a KM for starch of 1.4 mg/ml and a Vmax of 0.057 mg/min of hydrolyzed starch. Molybdenum activated the purified enzyme, and sodium dodecyl sulfate inhibited it. A thin layer chromatography analysis revealed glucose as the enzyme's main starch hydrolysis product. An enzyme's peptide sequence was obtained by mass spectrometry and used to retrieve a glucoamylase within the annotated genome of A. wentii v1.0. An in silico structural model revealed a N-terminal glycosyl hydrolases family 15 (GH15) domain, which is ligated by a linker to a C-terminal carbohydrate-binding module (CBM) from the CBM20 family.

Protease inhibitor effects on prevalence of bacterial vaginosis and human papillomavirus-related lesions.

AIM: Antiretrovirals of the protease inhibitor (PI) class tend to achieve low concentrations in biological fluids. This study aimed to analyze possible changes in the vaginal microbiome and frequency of cervical human papillomavirus (HPV)-DNA and HPV-related lesions associated with the use of PI in antiretroviral therapy (ART). METHODS: Eighty-eight women with human immunodeficiency virus infection were divided in two groups: ART with PI and without PI. All the participants underwent anamnesis with demographic data collection. The total DNA, used as the template in the polymerase chain reaction-based assays for the detection of HPV-DNA, was extracted from cervical samples during cervical cytopathology. RESULTS: There were no differences between the groups with respect to HPV-related lesions. Despite the higher prevalence of bacterial vaginosis (BV) in the PI group (33.96% vs 17.14%), the difference was insignificant when considering all women (P = 0.066). When women with a detectable viral load and a CD4+ T-cell count <200 were excluded in both groups, BV was found to be more prevalent in the PI group (odds ratio, 3.349; 95% confidence interval, 1.113-11.41, P = 0.049). No associations were found between BV and age, condom use, cervical HPV, time with current ART regimen, unprotected receptive anal intercourse and cervical HPV-related lesions. CONCLUSION: The use of PI did not alter the frequencies of HPV-DNA and HPV-related lesions. However, an increased frequency of BV was found in women using PI after excluding women with a detectable viral load and a CD4+ T-cell count of <200.

Reproducibility of the NanoString 22-gene molecular subgroup assay for improved prognostic prediction of medulloblastoma.

Medulloblastoma is the most frequent malignant brain tumor in children. Four medulloblastoma molecular subgroups, MBSHH , MBWNT , MBGRP3 and MBGRP4 , have been identified by integrated high-throughput platforms. Recently, a 22-gene panel NanoString-based assay was developed for medulloblastoma molecular subgrouping, but the robustness of this assay has not been widely evaluated. Mutations in the gene for human telomerase reverse transcriptase (hTERT) have been found in medulloblastomas and are associated with distinct molecular subtypes. This study aimed to implement the 22-gene panel in a Brazilian context, and to associate the molecular profile with patients' clinical-pathological features. Formalin-fixed, paraffin-embedded (FFPE) medulloblastoma samples (n = 104) from three Brazilian centers were evaluated. Expression profiling of the 22-gene panel was performed by NanoString and a Canadian series (n = 240) was applied for training phase. hTERT mutations were analyzed by PCR followed by direct Sanger sequencing and the molecular profile was associated with patients' clinicopathological features. Overall, 65% of the patients were male, average age at diagnosis was 18 years and 7% of the patients presented metastasis at diagnosis. The molecular classification was attained in 100% of the cases, with the following frequencies: MBSHH (n = 51), MBWNT (n = 19), MBGRP4 (n = 19) and MBGRP3 (n = 15). The MBSHH and MBGRP3 subgroups were associated with older and younger patients, respectively. The MBGRP4 subgroup exhibited the lowest 5-year cancer-specific overall survival (OS), yet in the multivariate analysis, only metastasis at diagnosis and surgical resection were associated with OS. hTERT mutations were detected in 29% of the cases and were associated with older patients, increased hTERT expression and MBSHH subgroup. The 22-gene panel provides a reproducible assay for molecular subgrouping of medulloblastoma FFPE samples in a routine setting and is well-suited for future clinical trials.

Production, purification, and characterization of a novel serine-esterase from Aspergillus westerdijkiae.

Esterases hydrolyze water soluble short chain fatty acids esters and are biotechnologically important. A strain of Aspergillus westerdijkiae isolated from cooking oil for recycling was found to secrete an esterase. The best enzyme production (19-24 U/ml of filtrate) culture conditions were stablished. The protein was purified using ammonium sulphate precipitation, dialysis, and a chromatographic step in Sephacryl S-200 HR. The 32 kDa purified protein presented an optimal temperature of 40°C, with a T50 of 48.95°C, and an optimal pH of 8.0. KM and Vmax were 638.11 µM for p-NPB and 5.47 µmol of released p-NP · min-1 · µg-1 of protein, respectively. The purified enzyme was partially active in the presence of 25% acetone. PMSF inhibited the enzyme, indicating that it is a serine hydrolase. MS enzyme peptides sequences were used to find the protein in the A. westerdijkiae sequenced genome. A structure model demonstrated that the protein is a member of the a/ß -hydrolase fold superfamily.

Search for novel targets of the PII signal transduction protein in Bacteria identifies the BCCP component of acetyl-CoA carboxylase as a PII binding partner.

The PII family comprises a group of widely distributed signal transduction proteins. The archetypal function of PII is to regulate nitrogen metabolism in bacteria. As PII can sense a range of metabolic signals, it has been suggested that the number of metabolic pathways regulated by PII may be much greater than described in the literature. In order to provide experimental evidence for this hypothesis a PII protein affinity column was used to identify PII targets in Azospirillum brasilense. One of the PII partners identified was the biotin carboxyl carrier protein (BCCP), a component of the acetyl-CoA carboxylase which catalyses the committed step in fatty acid biosynthesis. As BCCP had been previously identified as a PII target in Arabidopsis thaliana we hypothesized that the PII -BCCP interaction would be conserved throughout Bacteria. In vitro experiments using purified proteins confirmed that the PII -BCCP interaction is conserved in Escherichia coli. The BCCP-PII interaction required MgATP and was dissociated by increasing 2-oxoglutarate. The interaction was modestly affected by the post-translational uridylylation status of PII ; however, it was completely dependent on the post-translational biotinylation of BCCP.

Purification of the Campylobacter jejuni Dps protein assisted by its high melting temperature.

Dps proteins (DNA binding protein from starved cell) form a distinct group within the ferritin superfamily. All Dps members are composed of 12 identical subunits that assemble into a conserved spherical protein shell. Dps oxidize Fe(2+) in a conserved ferroxidase center located at the interface between monomers, the product of the reaction Fe(3+), is then stored inside the protein shell in the form of non-reactive insoluble Fe2O3. The Campylobacter jejuni Dps (CjDps) has been reported to play a plethora of functions, such as DNA binding and protection, iron storage, survival in response to hydrogen peroxide and sulfatide binding. CjDps is also important during biofilm formation and caecal colonization in poultry. In order to facilitate in vitro characterisation of CjDps, it is important to have a simple and reproducible protocol for protein purification. Here we report an observation that CjDps has an unusual high melting temperature. We exploited this property for protein purification by introducing a thermal treatment step which allowed achieving homogeneity by using only two chromatographic steps. Gel filtration chromatography, circular dichroism, mass spectrometry, DNA-binding and iron oxidation analysis confirmed that the CjDps structure and function were unaffected.

Pharmacokinetic and pharmacodynamic evaluation of a nanotechnological topical formulation of lidocaine/prilocaine (nanorap) in healthy volunteers.

BACKGROUND: Nanorap is a new nanotechnological formulation for topical anesthesia composed of lidocaine (2.5%) and prilocaine (2.5%). This study evaluated the pharmacokinetics of Nanorap. For the determination of lidocaine and prilocaine in human plasma, a new method using high-performance liquid chromatography coupled with tandem mass spectrometry was developed. Nanorap pharmacodynamic (PD) and its physical proprieties were also evaluated. METHODS: Nanorap was administered by topical application of 2 g to healthy volunteers, and blood samples were collected for the pharmacokinetics analysis. The drugs were extracted from plasma by liquid-liquid extraction with ether/hexane (80/20, vol/vol). The chromatography separation was performed on a Genesis C18 analytical column 4 µm (100 × 2.1 mm i.d.) with a mobile phase of methanol/acetonitrile/water (40/30/30, for lidocaine, and 50/30/20, for prilocaine, vol/vol/vol) + 2 mM of ammonium acetate and ropivacaine as internal standard. The drugs were quantified using a mass spectrometer with an electrospray source in the electrospray ionization positive mode configured for multiple reaction monitoring. The PD of Nanorap was evaluated with the use of a visual analog scale. Nanorap was characterized by cryofracture. RESULTS: The chromatography run-time was 5.5 minutes for lidocaine and 3.3 minutes for prilocaine, and the lower limit of quantification was 0.05 ng/mL for both drugs. Mean Cmax was 6.62 and 1.72 ng/mL for lidocaine and prilocaine, respectively. Median Tmax was 6.5 hours for both drugs. Nanocapsules had a mean size of 88 nm and mean drug association of 92.5% and 89% for lidocaine and prilocaine, respectively. The PD study showed that Nanorap has a sufficient analgesic effect (>30% reduction in pain) after 10 minutes of application. CONCLUSIONS: A new simple, selective, and sensitive method for determination of lidocaine and prilocaine in human plasma was developed. Nanorap generated safe plasma levels of the drugs and satisfactory analgesic effect.

Sexual function in patients with deep infiltrating endometriosis.

INTRODUCTION: Endometriosis is a benign condition that causes pain and infertility. Sexual dysfunction, particularly deep dyspareunia, is common in patients with endometriosis and interferes with quality of life and conjugal satisfaction. AIM: The study aims to assess sexual function in women with deep infiltrating endometriosis. METHOD: Fifty-seven women diagnosed with deep infiltrating endometriosis were recruited from Hospital Universitário Pedro Ernesto (HUPE) between July and December 2011. The control group comprised 38 healthy women recruited at the HUPE family planning clinic. MAIN OUTCOME MEASURES: The main outcomes are full-scale and individual domain scores on the Female Sexual Function Index (FSFI), a validated questionnaire for functional assessment of sexual function in women. RESULTS: Patients with endometriosis had more pain in intercourse than controls, which correlates with lower scores in the FSFI pain domain. However, there were no statistically significant between-group differences in overall (full-scale) FSFI scores. CONCLUSION: Women with endometriosis exhibit significant dysfunction in the pain domain of the FSFI questionnaire, but this finding was not sufficient to affect the overall sexual function.

Mindfulness-Based Intervention Effect on the Psychophysiological Marker of Self-Regulation in Women With Endometriosis-Related Chronic Pain.

Endometriosis is a gynecological disease that involves a broad biopsychosocial compromise with the potential to create a negative vicious cycle. Despite the complexity of factors influencing women's improvement, most interventions investigated target just the peripheral nociceptive sources of endometriosis-related pain. An alternative is intervening in self-regulation, which can potentially influence multiple domains of the illness experience. The present study examines the effect of a brief Mindfulness-Based Intervention (bMBI) on attention and autonomic nervous system regulation in women with endometriosis-related pain. Also, explore the interaction between these self-regulation domains and the affective pain dimension. An exploratory analysis of the secondary outcomes of a pilot randomized controlled trial was performed. The vagally-mediated Heart Rate Variability (vmHRV) at rest, cognitive stress, and recovery was employed to measure autonomic regulation. The Flanker and Stroop tasks were used to estimate the attention domains. Results showed that bMBI (n = 26) significantly improved Flanker accuracy and Flanker and Stroop reaction time compared to the control group (n = 28). bMBI significantly increased vmHRV at rest and recovery after cognitive stress. Attention mediated the bMBI effect on affective pain improvement. Results suggest that bMBI improves self-regulation domains with the potential to develop a broad biopsychosocial benefit in the endometriosis context. PERSPECTIVE: This article demonstrates the positive impact of a brief Mindfulness-Based Intervention on attention and parasympathetic regulation in women suffering from endometriosis-related pain. This mindfulness-induced self-regulation improvement can benefit affective pain and potentially multiple psychophysiological processes relevant to endometriosis.

Data-driven coaching to improve statewide outcomes in CABG: before and after interventional study.

BACKGROUND: The impact of quality improvement initiatives program (QIP) on coronary artery bypass grafting surgery (CABG) remains scarce, despite improved outcomes in other surgical areas. This study aims to evaluate the impact of a package of QIP on mortality rates among patients undergoing CABG. MATERIALS AND METHODS: This prospective cohort study utilized data from the multicenter database Registro Paulista de Cirurgia Cardiovascular II (REPLICCAR II), spanning from July 2017 to June 2019. Data from 4018 isolated CABG adult patients were collected and analyzed in three phases: before-implementation, implementation, and after-implementation of the intervention (which comprised QIP training for the hospital team). Propensity Score Matching was used to balance the groups of 2170 patients each for a comparative analysis of the following outcomes: reoperation, deep sternal wound infection/mediastinitis ≤30 days, cerebrovascular accident, acute kidney injury, ventilation time >24 h, length of stay <6 days, length of stay >14 days, morbidity and mortality, and operative mortality. A multiple regression model was constructed to predict mortality outcomes. RESULTS: Following implementation, there was a significant reduction of operative mortality (61.7%, P =0.046), as well as deep sternal wound infection/mediastinitis ( P <0.001), sepsis ( P =0.002), ventilation time in hours ( P <0.001), prolonged ventilation time ( P =0.009), postoperative peak blood glucose ( P <0.001), total length of hospital stay ( P <0.001). Additionally, there was a greater use of arterial grafts, including internal thoracic ( P <0.001) and radial ( P =0.038), along with a higher rate of skeletonized dissection of the internal thoracic artery. CONCLUSIONS: QIP was associated with a 61.7% reduction in operative mortality following CABG. Although not all complications exhibited a decline, the reduction in mortality suggests a possible decrease in failure to rescue during the after-implementation period.

Protocol for the establishment of a serine integrase-based platform for functional validation of genetic switch controllers in eukaryotic cells.

Serine integrases (Ints) are a family of site-specific recombinases (SSRs) encoded by some bacteriophages to integrate their genetic material into the genome of a host. Their ability to rearrange DNA sequences in different ways including inversion, excision, or insertion with no help from endogenous molecular machinery, confers important biotechnological value as genetic editing tools with high host plasticity. Despite advances in their use in prokaryotic cells, only a few Ints are currently used as gene editors in eukaryotes, partly due to the functional loss and cytotoxicity presented by some candidates in more complex organisms. To help expand the number of Ints available for the assembly of more complex multifunctional circuits in eukaryotic cells, this protocol describes a platform for the assembly and functional screening of serine-integrase-based genetic switches designed to control gene expression by directional inversions of DNA sequence orientation. The system consists of two sets of plasmids, an effector module and a reporter module, both sets assembled with regulatory components (as promoter and terminator regions) appropriate for expression in mammals, including humans, and plants. The complete method involves plasmid design, DNA delivery, testing and both molecular and phenotypical assessment of results. This platform presents a suitable workflow for the identification and functional validation of new tools for the genetic regulation and reprogramming of organisms with importance in different fields, from medical applications to crop enhancement, as shown by the initial results obtained. This protocol can be completed in 4 weeks for mammalian cells or up to 8 weeks for plant cells, considering cell culture or plant growth time.

Uridylylation of Herbaspirillum seropedicae GlnB and GlnK proteins is differentially affected by ATP, ADP and 2-oxoglutarate in vitro.

PII are signal-transducing proteins that integrate metabolic signals and transmit this information to a large number of proteins. In proteobacteria, PII are modified by GlnD (uridylyltransferase/uridylyl-removing enzyme) in response to the nitrogen status. The uridylylation/deuridylylation cycle of PII is also regulated by carbon and energy signals such as ATP, ADP and 2-oxoglutarate (2-OG). These molecules bind to PII proteins and alter their tridimensional structure/conformation and activity. In this work, we determined the effects of ATP, ADP and 2-OG levels on the in vitro uridylylation of Herbaspirillum seropedicae PII proteins, GlnB and GlnK. Both proteins were uridylylated by GlnD in the presence of ATP or ADP, although the uridylylation levels were higher in the presence of ATP and under high 2-OG levels. Under excess of 2-OG, the GlnB uridylylation level was higher in the presence of ATP than with ADP, while GlnK uridylylation was similar with ATP or ADP. Moreover, in the presence of ADP/ATP molar ratios varying from 10/1 to 1/10, GlnB uridylylation level decreased as ADP concentration increased, whereas GlnK uridylylation remained constant. The results suggest that uridylylation of both GlnB and GlnK responds to 2-OG levels, but only GlnB responds effectively to variation on ADP/ATP ratio.

Heat stability of Proteobacterial PII protein facilitate purification using a single chromatography step.

The P(II) proteins comprise a family of widely distributed signal transduction proteins that integrate the signals of cellular nitrogen, carbon and energy status, and then regulate, by protein-protein interaction, the activity of a variety of target proteins including enzymes, transcriptional regulators and membrane transporters. We have previously shown that the P(II) proteins from Azospirillum brasilense, GlnB and GlnZ, do not alter their migration behavior under native gel electrophoresis following incubated for a few minutes at 95°C. This data suggested that P(II) proteins were either resistant to high temperatures and/or that they could return to their native state after having been unfolded by heat. Here we used (1)H NMR to show that the A. brasilense GlnB is stable up to 70°C. The melting temperature (Tm) of GlnB was determined to be 84°C using the fluorescent dye Sypro-Orange. P(II) proteins from other Proteobacteria also showed a high Tm. We exploited the thermo stability of P(II) by introducing a thermal treatment step in the P(II) purification protocol, this step significantly improved the homogeneity of A. brasilense GlnB and GlnZ, Herbaspirillum seropedicae GlnB and GlnK, and of Escherichia coli GlnK. Only a single chromatography step was necessary to obtain homogeneities higher than 95%. NMR(1) and in vitro uridylylation analysis showed that A. brasilense GlnB purified using the thermal treatment maintained its folding and activity. The purification protocol described here can facilitate the study of P(II) protein family members.

A protective effect of morning radiotherapy on acute skin toxicity in patients with breast cancer: A prospective cohort study.

ABSTRACT: The focus of this prospective cohort study was to evaluate the risk factors of severe acute skin toxicity (grade ≥2) in 100 patients with breast cancer (BC) during radiotherapy (RT).The patients were evaluated weekly during RT and 3 months after treatment. The endpoint included the occurrence of skin toxicity grade ≥2, according to Radiation Therapy Oncology Group (RTOG). Survival analysis was conducted by univariate and multivariate Cox regression analysis.In the multivariate analysis, RT in the afternoon (0-3 pm) (hazard ratios [HR] = 1.566, P = .042) was significantly associated with the early occurrence of skin toxicity, indicating a potential effect of chronotherapy related to this adverse event. In the univariate and multivariate analysis, skin phototype moderate brown (HR = 1.586, P = .042; HR = 1.706, P = .022, respectively) and dark brown or black (HR = 4.517, P < .001; HR = 5.336, P < 0.001, respectively) was significantly associated with the skin toxicity. Tangential field separation >21 cm (HR = 2.550, P = .009, HR = 2.923, P = .003), in women that were submitted to conservative surgery indicates indirectly that large breast size was also significantly associated with skin toxicity.Women with large breasts and dark brown or black skin should be followed more carefully during RT, which should be undergone in the morning, especially when submitted to conventional RT techniques, common in developing countries.

Polyelectrolytic gelatin nanoparticles as a drug delivery system for the promastigote form of Leishmania amazonensis treatment.

In this study, phthalocianato[bis(dimethylaminoethanoxy)] silicon (NzPC) was loaded onto gelatin nanoparticles functionalized with polyelectrolytes (polystyrene sulfonate/polyallylamine hydrochloride) by layer-by-layer (LbL) assembly for photodynamic therapy (PDT) application in promastigote form of Leishmania amazonensis treatment. The process yield, and encapsulation efficiency were 80.0% ± 1.8 and EE = 87.0% ± 1.1, respectively. The polyelectrolytic gelatin nanoparticles (PGN) had a mean diameter of 437.4 ± 72.85 nm, narrow distribution size with a polydispersity index of 0.086. The obvious switching of zeta potential indicates successful alternating deposition of the polyanion PSS and polycation PAH directly on the gelatin nanoparticles. Photosensitizer photophysical properties were shown to be preserved after gelatin nanoparticle encapsulation. The impact of the PDT in the viability and morphology of Leishmania amazonensis promastigote in culture medium was evaluated. The PGN-NzPc presented low toxicity at the dark and the PDT was capable of decreasing the viability in more than 80% in 0.1 µmol.L-1 concentration tested. The PDT also triggered significant morphological alterations in the Leishmania promastigotes. These results reinforce the idea that the use of PGN as photosensitizers carriers is useful for PDT of Leishmania promastigotes.

Feruloyl esterase activity and its role in regulating the feruloylation of maize cell walls.

Cell walls of grasses have ferulic acid (FA) ester-linked to the arabinosyl substitutions of arabinoxylan (AX). Feruloyl esterases (FAE) are carboxylic acid esterases that release FA from cell walls and synthetic substrates. Despite the importance of FA for cell wall recalcitrance and in response to biotic and abiotic stresses, the physiological function of plant FAEs remains unclear. Here, we developed a simple method for the determination of FAE activity (ZmFAE) in maize using the total protein extract and investigated its role in regulating the feruloylation of cell wall. The method includes a single protein extraction and enzymatic reaction with protein concentration as low as 65 µg at 35 °C for 30 min, using methyl ferulate as the substrate. The methodology allowed the determination of the apparent Km (392.82 µM) and Vmax (79.15 pkat mg-1 protein). We also found that ZmFAE activity was correlated (r = 0.829) with the levels of FA in seedling roots, plant roots and leaves of maize. Furthermore, the exposure to osmotic stress resulted in a 50% increase in ZmFAE activity in seedling roots. These data suggest that FAE-catalyzed reaction is important for cell wall feruloylation during plant development and in response to abiotic stress. We conclude proposing a model for the feruloylation and deferuloylation of AX, which explains the role of FAE in regulating the levels of ester-linked FA. Our model might orient further studies investigating the role of plant FAEs and assist strategies for genetic engineering of grasses to obtain plants with reduced biomass recalcitrance.

Glycemic homeostasis and hepatic metabolism are modified in rats with global cerebral ischemia.

Cerebral ischemia-induced hyperglycemia has been reported to accentuate neurological damage following focal or global cerebral ischemia. Hyperglycemia found in rats following focal brain ischemia occurs in the first 24 h and has been claimed to be caused by increased liver gluconeogenesis and insulin resistance. However, liver gluconeogenesis and the mechanisms leading to hyperglycemia after global cerebral ischemia remain uncertain. This study investigated the glycemic homeostasis and hepatic metabolism in rats after transient four-vessel occlusion (4-VO)-induced global cerebral ischemia, an event that mimics to a certain degree the situation during cardiac arrest. Several metabolic fluxes were measured in perfused livers. Activities and mRNA expressions of hepatic glycolysis and glyconeogenesis rate-limiting enzymes were assessed as well as respiratory activity of hepatic isolated mitochondria. Global cerebral ischemia was associated with hyperglycemia and hyperinsulinemia 24 h after ischemia. Insulin resistance developed later and was prominent after the 5th day. Hepatic anabolism and catabolism were both modified in a complex and time-dependent way. Gluconeogenesis, ß-oxidation, ketogenesis and glycolysis were diminished at 24 h after ischemia. At 5 days after ischemia glycolysis had normalized, but gluconeogenesis, ketogenesis and ß-oxidation were accelerated. The overall metabolic modifications suggest that a condition of depressed metabolism was established in response to the new conditions generated by the cerebral global ischemia. Whether the modifications in the liver metabolism found in rats after the ischemic insult can be translated to individuals following global brain ischemia remains uncertain, but the results of this study are hoped to encourage further investigations.