High-CO2 Levels Rather than Acidification Restrict Emiliania huxleyi Growth and Performance.

The coccolithophore Emiliania huxleyi shows a variety of responses to ocean acidification (OA) and to high-CO2 concentrations, but there is still controversy on differentiating between these two factors when using different strains and culture methods. A heavily calcified type A strain isolated from the Norwegian Sea was selected and batch cultured in order to understand whether acclimation to OA was mediated mainly by CO2 or H+, and how it impacted cell growth performance, calcification, and physiological stress management. Emiliania huxleyi responded differently to each acidification method. CO2-enriched aeration (1200 µatm, pH 7.62) induced a negative effect on the cells when compared to acidification caused by decreasing pH alone (pH 7.60). The growth rates of the coccolithophore were more negatively affected by high pCO2 than by low pH without CO2 enrichment with respect to the control (400 µatm, pH 8.1). High CO2 also affected cell viability and promoted the accumulation of reactive oxygen species (ROS), which was not observed under low pH. This suggests a possible metabolic imbalance induced by high CO2 alone. In contrast, the affinity for carbon uptake was negatively affected by both low pH and high CO2. Photochemistry was only marginally affected by either acidification method when analysed by PAM fluorometry. The POC and PIC cellular quotas and the PIC:POC ratio shifted along the different phases of the cultures; consequently, calcification did not follow the same pattern observed in cell stress and growth performance. Specifically, acidification by HCl addition caused a higher proportion of severely deformed coccoliths, than CO2 enrichment. These results highlight the capacity of CO2 rather than acidification itself to generate metabolic stress, not reducing calcification.

An Expedient Method for the Umpolung Coupling of Enols with Heteronucleophiles.

In this paper, we present an unprecedented and general umpolung protocol that allows the functionalization of silyl enol ethers and of 1,3-dicarbonyl compounds with a large range of heteroatom nucleophiles, including carboxylic acids, alcohols, primary and secondary amines, azide, thiols, and also anionic carbamates derived from CO2 . The scope of the reaction also extends to carbon-based nucleophiles. The reaction relies on the use of 1-bromo-3,3-dimethyl-1,3-dihydro-1λ3 [d][1,2]iodaoxole, which provides a key α-brominated carbonyl intermediate. The reaction mechanism has been studied experimentally and by DFT, and we propose formation of an unusual enolonium intermediate with a halogen-bonded bromide.

Unraveling the Mechanism of the IrIII -Catalyzed Regiospecific Synthesis of α-Chlorocarbonyl Compounds from Allylic Alcohols.

We have used experimental studies and DFT calculations to investigate the IrIII -catalyzed isomerization of allylic alcohols into carbonyl compounds, and the regiospecific isomerization-chlorination of allylic alcohols into α-chlorinated carbonyl compounds. The mechanism involves a hydride elimination followed by a migratory insertion step that may take place at Cß but also at Cα with a small energy-barrier difference of 1.8 kcal mol-1 . After a protonation step, calculations show that the final tautomerization can take place both at the Ir center and outside the catalytic cycle. For the isomerization-chlorination reaction, calculations show that the chlorination step takes place outside the cycle with an energy barrier much lower than that for the tautomerization to yield the saturated ketone. All the energies in the proposed mechanism are plausible, and the cycle accounts for the experimental observations.

Sarco-Endoplasmic Reticulum Calcium Release Model Based on Changes in the Luminal Calcium Content.

The sarcoplasmic/endoplasmic reticulum (SR/ER) is the main intracellular calcium (Ca2+) pool in muscle and non-muscle eukaryotic cells, respectively. The reticulum accumulates Ca2+ against its electrochemical gradient by the action of sarco/endoplasmic reticulum calcium ATPases (SERCA pumps), and the capacity of this Ca2+ store is increased by the presence of Ca2+ binding proteins in the lumen of the reticulum. A diversity of physical and chemical signals, activate the main Ca2+ release channels, i.e. ryanodine receptors (RyRs) and inositol (1, 4, 5) trisphosphate receptors (IP3Rs), to produce transient elevations of the cytoplasmic calcium concentration ([Ca2+]i) while the reticulum is being depleted of Ca2+. This picture is incomplete because it implies that the elements involved in the Ca2+ release process are acting alone and independently of each other. However, it appears that the Ca2+ released by RyRs and IP3Rs is trapped in luminal Ca2+ binding proteins (Ca2+ lattice), which are associated with these release channels, and the activation of these channels appears to facilitate that the trapped Ca2+ ions become available for release. This situation makes the initial stage of the Ca2+ release process a highly efficient one; accordingly, there is a large increase in the [Ca2+]i with minimal reductions in the bulk of the free luminal SR/ER [Ca2+] ([Ca2+]SR/ER). Additionally, it has been shown that active SERCA pumps are required for attaining this highly efficient Ca2+ release process. All these data indicate that Ca2+ release by the SR/ER is a highly regulated event and not just Ca2+ coming down its electrochemical gradient via the open release channels. One obvious advantage of this sophisticated Ca2+ release process is to avoid depletion of the ER Ca2+ store and accordingly, to prevent the activation of ER stress during each Ca2+ release event.

Effects of elevated CO2 on growth, calcification, and spectral dependence of photoinhibition in the coccolithophore Emiliania huxleyi (Prymnesiophyceae)1.

We studied the effects of elevated CO2 concentrations on cell growth, calcification, and spectral variation in the sensitivity of photosynthesis to inhibition by solar radiation in the globally important coccolithophore Emiliania huxleyi. Growth rates and chlorophyll a content per cell showed no significant differences between elevated (800 ppmv) and ambient (400 ppmv) CO2 conditions. However, the production of organic carbon and the cell quotas for both carbon and nitrogen, increased under elevated CO2 conditions, whilst particulate inorganic carbon production rates decreased under the same conditions. Biometric analyses of cells showed that coccoliths only presented significant differences due to treatments in the central area width. Most importantly, the size of the coccosphere decreased under elevated CO2 conditions. The susceptibility of photosynthesis to inhibition by ultraviolet radiation (UVR) was estimated using biological weighting functions (BWFs) and a model that predicts photosynthesis under photosynthetically active radiation and UVR exposures. BWF results demonstrated that the sensitivity of photosynthesis to UVR was not significantly different between E. huxleyi cells grown under elevated and present CO2 concentrations. We propose that the acclimation to elevated CO2 conditions involves a physiological mechanism of regulation and allocation of energy and metabolites in the cell, which is also responsible for altering the sensitivity to UVR. In coccolithophores, this mechanism might be affected by the decrease in the calcification rates.

Dural arteriovenous fistula at the foramen magnum: Report of a case and clinical-anatomical review.

Arterial supply and venous drainage at the foramen magnum is variable. Two main forms of clinical presentation, intracranial and spinal, can be differentiated when a dural arteriovenous fistula (DAVF) is found at this level. We describe a case of a 68-year-old patient with a progressive paraparesis, diagnosed of dural arteriovenous fistula located at the posterior lip of foramen magnum. We review, in this setting, the vascular radiological anatomy of those fistulas and its important correlation with neurologic clinical symptoms.

Influence of water electrolysis on hydrolysis and methanogenesis stages of anaerobic digestion at room temperature: Kinetic and metabolic analysis.

Batch cultures were performed to study hydrolysis and methanogenesis in the presence of an electric field at room temperature (i.e., 23 °C). Kinetic and metabolic analyses using RuO2/Ti electrodes were carried out in short reaction times to avoid biofilm formation, allowing the evaluation of the effect of O2 and H2 produced on anaerobic digestion during the imposition of three electric fields: 1.21, 1.45, and 1.64 V/cm. Results highlighted that at 1.21 V/cm, the electrolysis produced 0.0753 mg O2/L·min, where facultative microorganisms consumed 21 % oxygen, enhancing the hydrolysis phase by 52 %. Additionally, methane production was noticeably improved with an activity of 0.89 ± 0.02 g COD-CH4/g VSS·d, meaning 39 % higher than the control. The imposition of an electric field showed promising results since the methanogenic activity at room temperature was very close to the activities observed in conventional reactors at 35 °C.

Late clinical and radiological complications of stereotactical radiosurgery of arteriovenous malformations of the brain.

INTRODUCTION: Post-radiation injury of patients with brain arteriovenous malformations (AVM) include blood-brain barrier breakdown (BBBB), edema, and necrosis. Prevalence, clinical relevance, and response to treatment are poorly known. We present a series of consecutive brain AVM treated with stereotactic radiosurgery describing the appearance of radiation injury and clinical complications. METHODS: Consecutive patients with annual clinical and radiological follow-up (median length 63 months). Edema and BBBB were classified in four groups (minimal, perilesional, moderate, or severe), and noted together with necrosis. Clinical symptoms of interest were intracranial hypertension, new neurological deficits, new seizures, and brain hemorrhages. RESULTS: One hundred two cases, median age 34 years, 52% male. Median irradiated volume 3.8 cc, dose to the margin of the nidus 18.5 Gy. Nineteen patients underwent a second radiosurgery. Only 42.2% patients remained free from radiation injury. Edema was found in 43.1%, blood-brain barrier breakdown in 20.6%, necrosis in 6.9%. Major injury (moderate or severe edema, moderate or severe BBBB, or necrosis) was found in 20 of 102 patients (19.6%). AVM diameter >3 cm and second radiosurgery were independent predictors. Time to the worst imaging was 60 months. Patients with major radiation injury had a hazard ratio for appearance of focal deficits of 7.042 (p = 0.04), of intracranial hypertension 2.857 (p = 0.025), hemorrhage into occluded nidus 9.009 (p = 0.079), appearance of new seizures not significant. CONCLUSIONS: Major radiation injury is frequent and increases the risk of neurological complications. Its late appearance implies that current follow-up protocols need to be extended in time.

Heterogeneity of the endoplasmic reticulum Ca2+ store determines colocalization with mitochondria.

Contact sites between the endoplasmic reticulum (ER) and mitochondria play a pivotal role in cell signaling, and the interaction between these organelles is dynamic and finely regulated. We have studied the role of ER Ca2+ concentration ([Ca2+]ER) in modulating this association in HeLa and HEK293 cells and human fibroblasts. According to Manders' coefficient, ER-mitochondria colocalization varied depending on the ER marker; it was the highest with ER-Tracker and the lowest with ER Ca2+ indicators (Mag-Fluo-4, erGAP3, and G-CEPIA1er) in both HeLa cells and human fibroblasts. Only GEM-CEPIA1er displayed a high colocalization with elongated mitochondria in HeLa cells, this ER Ca2+ indicator reveals low Ca2+ regions because this ion quenches its fluorescence. On the contrary, the typical rounded and fragmented mitochondria of HEK293 cells colocalized with Mag-Fluo-4 and, to a lesser extent, with GEM-CEPIA1er. The ablation of the three IP3R isoforms in HEK293 cells increased mitochondria-GEM-CEPIA1er colocalization. This pattern of colocalization was inversely correlated with the rate of ER Ca2+ leak evoked by thapsigargin (Tg). Moreover, Tg and Histamine in the absence of external Ca2+ increased mitochondria-ER colocalization. On the contrary, in the presence of external Ca2+, both Bafilomycin A1 and Tg reduced the mitochondria-ER interaction. Notably, knocking down MCU decreased mitochondria-ER colocalization. Overall, our data suggest that the [Ca2+] is not homogenous within the ER lumen and that mitochondria-ER interaction is modulated by the ER Ca2+ leak and the [Ca2+]i.

How does a low-magnitude electric field influence anaerobic digestion in wastewater treatment? A review.

Anaerobic digestion (AD) is a physio-biochemical process widely used for treating industrial or municipal wastewater with concomitant methane production. Several technologies have been tested to improve AD's efficiency, like pretreatments and co-digestion, among others. Recently the imposition of a low-magnitude electric field (LMEF) has been applied at the AD to improve methane yield. Despite the positive results of imputing an electric field, many gaps are not understood yet. Therefore, this review focuses on the biochemical aspects of AD and electric field for a better understanding of the effect of the LMEF on the metabolisms of the AD during wastewater treatment and its application in methane production enhancement.

Enantioselective Catalysis with Pyrrolidinyl Gold(I) Complexes: DFT and NEST Analysis of the Chiral Binding Pocket.

A new generation of chiral gold(I) catalysts based on variations of complexes with JohnPhos-type ligands with a remote C2-symmetric 2,5-diarylpyrrolidine have been synthesized with different substitutions at the top and bottom aryl rings: from replacing the phosphine by a N-heterocyclic carbene (NHC) to increasing the steric hindrance with bis- or tris-biphenylphosphine scaffolds, or by directly attaching the C2-chiral pyrrolidine in the ortho-position of the dialkylphenyl phosphine. The new chiral gold(I) catalysts have been tested in the intramolecular [4+2] cycloaddition of arylalkynes with alkenes and in the atroposelective synthesis of 2-arylindoles. Interestingly, simpler catalysts with the C2-chiral pyrrolidine in the ortho-position of the dialkylphenyl phosphine led to the formation of opposite enantiomers. The chiral binding pockets of the new catalysts have been analyzed by DFT calculations. As revealed by non-covalent interaction plots, attractive non-covalent interactions between substrates and catalysts direct specific enantioselective folding. Furthermore, we have introduced the open-source tool NEST, specifically designed to account for steric effects in cylindrical-shaped complexes, which allows predicting experimental enantioselectivities in our systems.

Postradiosurgery hemorrhage rates of arteriovenous malformations of the brain: influencing factors and evolution with time.

BACKGROUND AND PURPOSE: The long-term benefit of radiosurgery of brain arteriovenous malformations (AVM), especially nonhemorrhagic cases, is controversial. We calculated hemorrhage rates pre- and posttreatment and analyzed the risk factors for bleeding based on cases followed at our site. METHODS: One hundred eight patients, age 36 ± 17 years, 56 men. The mean follow-up was 65 ± 44 months (median, 54; interquartile range, 33-94). Most AVMs were small (74.1% <3 cm in diameter); 48.1% were located in an eloquent area, 27.8% had deep drainage, and 39.8% presented with hemorrhage. RESULTS: The annual hemorrhage rate for any undiagnosed AVM was 1.2%, and 3.3% for AVMs with hemorrhagic presentation. Older patients, cortical or subcortical AVMs, and cases with multiple draining veins were less likely to present with bleeding. During the first 36 months postradiosurgery, hemorrhagic AVMs had a rebleeding rate of 2.1%, and a rate of 1.1% from 3 years onwards. Nonhemorrhagic AVMs had a hemorrhage rate of 1.4% during the first 3 years and 0.3% afterward. Arterial hypertension and nidus volume were independent predictors of bleeding after treatment. Mean nidus obliteration time was 37 ± 18 months (median, 32; interquartile range, 25-40), with hemorrhage rate of 1.3% before and 0.6% after obliteration, and 1.9% for AVMs that were not closed at the end of follow-up. CONCLUSIONS: Both hemorrhagic and nonhemorrhagic AVMs benefit from radiosurgical therapy, with gradual decrease in their bleeding rates over the years. Albeit small, the risk of hemorrhage persists during the entirety of follow-up, being higher for cases with hemorrhagic presentation and nonobliterated AVM.

Synthesis of α,γ-Chiral Trifluoromethylated Amines through the Stereospecific Isomerization of α-Chiral Allylic Amines.

Chiral γ-branched aliphatic amines are present in a large number of pharmaceuticals and natural products. However, enantioselective methods to access these compounds are scarce and mainly rely on the use of designed chiral transition-metal complexes. Herein, we combined an organocatalytic method for the stereospecific isomerization of chiral allylic amines with a diastereoselective reduction of the chiral imine/enamine intermediates, leading to γ-trifluoromethylated aliphatic amines with two noncontiguous stereogenic centers, in excellent yields and high diastereo- and enantioselectivities. This approach has been used with primary amine substrates. This approach also provides a new synthetic pathway to chiral trifluoromethylated scaffolds, of importance in medicinal chemistry. Additionally, a gram-scale reaction demonstrates the applicability of this synthetic procedure.

Corrosion Behavior of Al Modified with Zn in Chloride Solution.

Aluminum-based alloys have been considered candidate materials for cathodic protection anodes. However, the Al-based alloys can form a layer of alumina, which is a drawback in a sacrificial anode. The anodes must exhibit uniform corrosion to achieve better performance. Aluminum can be alloyed with Zn to improve their performance. In this sense, in the present research, the electrochemical corrosion performance of Al-xZn alloys (x = 1.5, 3.5, and 5 at.% Zn) exposed to 3.5 wt.% NaCl for 24 h was evaluated. Polarization curves, linear polarization resistance (LPR), and electrochemical impedance spectroscopy (EIS) were used to identify the electrochemical behavior. The microstructure of the samples before the corrosion assessment was characterized by means of X-ray diffraction analyses (XRD) and scanning electron microscopy (SEM). In addition, microstructures of the corroded surfaces were characterized using X-ray mappings via SEM. Polarization curves indicated that Zn additions changed the pseudo-passivation behavior from what pure Al exhibited in a uniform dissolution regime. Furthermore, the addition of Zn shifted the corrosion potential to the active side and increased the corrosion rate. This behavior was consistent with the proportional decrease in polarization resistance (Rp) and charge transfer resistance (Rct) in the EIS. The analysis of EIS was done using a mathematical model related to an adsorption electrochemical mechanism. The adsorption of chloride at the Al-Zn alloy surface formed aluminum chloride intermediates, which controlled the rate of the process. The rate constants of the reactions of a proposed chemical mechanism were evaluated.

Titanium Increases the Antioxidant Activity and Macronutrient Concentration in Tomato Seedlings Exposed to Salinity in Hydroponics.

Global climate change affects agriculture and tends to aggravate the effect of various environmental stress factors including soil salinity. Beneficial elements such as titanium (Ti) may improve the performance of plants facing restrictive environments such as saline soils. This research work evaluated the individual effect of sodium chloride (0, 50, and 100 mM NaCl) in solution, that of leaf-applied Ti (0, 500, and 1000 mg L-1 Ti), and their interactions on physiological, biochemical, and nutritional variables of tomato (Solanum lycopersicum L.) seedlings cv. Rio Grande in a factorial design in greenhouse hydroponics. NaCl reduced seedling height, stem diameter, leaf area, SPAD units, and sugar and K concentrations, and increased antioxidant activity in stems and roots, photosynthetic pigments, sugars. Titanium increased the N, P, K, Ca, Mg, and Ti concentrations in leaves, but the concentration of total sugars in leaves was reduced when applying 500 mg Ti L-1. Under moderate salinity conditions (50 mM NaCl) the application of Ti increased the antioxidant activity in roots, while, at all salinity levels tested, Ti increased the concentrations of macro-nutrients and Ti in leaves. Titanium is concluded to have a positive effect on the antioxidant activity and nutrition of seedlings under saline stress conditions.

PKC-Mediated Orai1 Channel Phosphorylation Modulates Ca2+ Signaling in HeLa Cells.

The overexpression of the Orai1 channel inhibits SOCE when using the Ca2+ readdition protocol. However, we found that HeLa cells overexpressing the Orai1 channel displayed enhanced Ca2+ entry and a limited ER depletion in response to the combination of ATP and thapsigargin (TG) in the presence of external Ca2+. As these effects require the combination of an agonist and TG, we decided to study whether the phosphorylation of Orai1 S27/S30 residues had any role using two different mutants: Orai1-S27/30A (O1-AA, phosphorylation-resistant) and Orai1-S27/30D (O1-DD, phosphomimetic). Both O1-wt and O1-AA supported enhanced Ca2+ entry, but this was not the case with O1-E106A (dead-pore mutant), O1-DD, and O1-AA-E106A, while O1-wt, O1-E106A, and O1-DD inhibited the ATP and TG-induced reduction of ER [Ca2+], suggesting that the phosphorylation of O1 S27/30 interferes with the IP3R activity. O1-wt and O1-DD displayed an increased interaction with IP3R in response to ATP and TG; however, the O1-AA channel decreased this interaction. The expression of mCherry-O1-AA increased the frequency of ATP-induced sinusoidal [Ca2+]i oscillations, while mCherry-O1-wt and mCherry-O1-DD decreased this frequency. These data suggest that the combination of ATP and TG stimulates Ca2+ entry, and the phosphorylation of Orai1 S27/30 residues by PKC reduces IP3R-mediated Ca2+ release.

A Pd-Catalyzed [4 + 2] Annulation Approach to Fluorinated N-Heterocycles.

3-Fluoro- and trifluoromethylthio-piperidines represent important building blocks for discovery chemistry. We report a simple and efficient method to access analogs of these compounds that are armed with rich functionality allowing them to be chemoselectively derivatized with high diastereocontrol.

Lessons from the COVID-19 air pollution decrease in Spain: Now what?

We offer an overview of the COVID-19 -driven air quality changes across 11 metropolises in Spain with the focus on lessons learned on how continuing abating pollution. Traffic flow decreased by up to 80% during the lockdown and remained relatively low during the full relaxation (June and July). After the lockdown a significant shift from public transport to private vehicles (+21% in Barcelona) persisted due to the pervasive fear that using public transport might increase the risk of SARS-CoV-2 infection, which need to be reverted as soon as possible. NO2 levels fell below 50% of the WHO annual air quality guidelines (WHOAQGs), but those of PM2.5 were reduced less than expected due to the lower contributions from traffic, increased contributions from agricultural and domestic biomass burning, or meteorological conditions favoring high secondary aerosol formation yields. Even during the lockdown, the annual PM2.5 WHOAQG was exceeded in cities within the NE and E regions with high NH3 emissions from farming and agriculture. Decreases in PM10 levels were greater than in PM2.5 due to reduced emissions from road dust, vehicle wear, and construction/demolition. Averaged O3 daily maximum 8-h (8hDM) experienced a generalized decrease in the rural receptor sites in the relaxation (June-July) with -20% reduced mobility. For urban areas O3 8hDM responses were heterogeneous, with increases or decreases depending on the period and location. Thus, after canceling out the effect of meteorology, 5 out of 11 cities experienced O3 decreases during the lockdown, while the remaining 6 either did not experience relevant reductions or increased. During the relaxation period and coinciding with the growing O3 season (June-July), most cities experienced decreases. However, the O3 WHOAQG was still exceeded during the lockdown and full relaxation periods in several cities. For secondary pollutants, such as O3 and PM2.5, further chemical and dispersion modeling along with source apportionment techniques to identify major precursor reduction targets are required to evaluate their abatement potential.

High-throughput and high-efficiency sample preparation for single-cell proteomics using a nested nanowell chip.

Global quantification of protein abundances in single cells could provide direct information on cellular phenotypes and complement transcriptomics measurements. However, single-cell proteomics is still immature and confronts many technical challenges. Herein we describe a nested nanoPOTS (N2) chip to improve protein recovery, operation robustness, and processing throughput for isobaric-labeling-based scProteomics workflow. The N2 chip reduces reaction volume to <30 nL and increases capacity to >240 single cells on a single microchip. The tandem mass tag (TMT) pooling step is simplified by adding a microliter droplet on the nested nanowells to combine labeled single-cell samples. In the analysis of ~100 individual cells from three different cell lines, we demonstrate that the N2 chip-based scProteomics platform can robustly quantify ~1500 proteins and reveal membrane protein markers. Our analyses also reveal low protein abundance variations, suggesting the single-cell proteome profiles are highly stable for the cells cultured under identical conditions.

[Breast cancer and subsequent pregnancy. Infertility, death risk and survival]. / Cáncer de mama y embarazo subsiguiente. Infertilidad, riesgo de muerte y supervivencia.

BACKGROUND: The chemotherapy commonly used for the treatment of breast cancer affect the fertility and could cause premature ovarian failure. The subsequent pregnancy to the breast cancer therefore it is not habitual. OBJECTIVE: The purpose of this study was to identify the effects of the chemotherapy in the women reproductive life; to evaluate the frequency of the women that experienced a later pregnancy after treatment of cancer, as well as the effects of the subsequent pregnancy on the breast cancer. We report our experience in the 14 year period. MATERIAL AND METHOD: As retrospective design, of 14 cases with breast cancer and subsequent pregnancy, from March 1994 to June 2008; demographic variables, clinical presentation, histopathological data, diagnostic procedures, treatments and results of the pregnancy were identified. RESULTS: The mean (M +/- SD) age of gestational women was of 31.5 +/- 5.2 years; the 83.3% women received adyuvant chemotherapy with antraciclines; the patients with regional illness (> 4 N+) and advanced illness had an adverse presage; the systemic relapse and progression showed in 42.8% of the cases. The pregnancy to term was presented in half of the cases in the first two years and in a third part, later to the 2 years of having concluded the oncological treatment; of the fourteen patients with breast cancer presented a total of 16 pregnancies: 9 were of term, 3 of preterm and 4 abortions. CONCLUSIONS: The study founds are based on a series of cases, which do not suggest that the pregnancy after the diagnosis and treatment of breast cancer has some adverse effect in the patients survival, for what the patients can conceive later to the oncological treatment. However, in this study it was observed that the effect of the advanced stage and positive axillary ganglion (> 4) influence in the relapses.