Role of HLA DRB1*15 and HLA DRB1*16alleles in the genetic susceptibility to develop systemic lupus erythematosus (SLE) after Chikungunya and Zika viruses infection in México

Systemic lupus erythematosus (SLE) is a clinically and genetically heterogeneous disease particularly prevalent in Mexico. Althoughits etiology is unknown, genetic factors strongly influence its presenceas well as triggering factors, such as viral infections, including Cytomegalovirus and Epstein-Barr virus. Here,the study presents the appearance of de novoSLE (patients who did not present SLE before de virus infection, corroborated by serological analysis and negative for antinuclear antibodies) cases in Mexicans who live near the southern border of Mexico, who presented clinical symptoms of arthritic, hematological, mucocutaneous and renal SLE, after Zika and/ or Chikungunya virus infection. Low resolution class Ⅱ HLA typing was performed, which found a significantly increased frequency of HLA DRB1*02 (15 and 16)when compared to a group of 99 healthy individuals (P =0.001, OR=4.5, IC95% 1.8~11.0). All the patients were diagnosed with SLE 1 to 3 years after being confirmed with the Zika, and/or Chikungunya infection. At the point of acute viral infection, none of the patients presented clinical signs or symptoms of autoimmunity or were negative for antinuclear antibodies. In genetically susceptible individuals, Zika and Chikungunya viral infection can trigger SLE.     

Diminished paracrine regulation of the epithelial Na+ channel by purinergic signaling in mice lacking connexin 30

We tested whether ATP release through Connexin 30 (Cx30) is part of a local purinergic regulatory system intrinsic to the aldosterone-sensitive distal nephron (ASDN) important for proper control of sodium excretion; if changes in sodium intake influence ATP release via Cx30; and if this allows a normal ENaC response to changes in systemic sodium levels. In addition, we define the consequences of disrupting ATP regulation of ENaC in Cx30(-/-) mice. Urinary ATP levels in wild-type mice increase with sodium intake, being lower and less dependent on sodium intake in Cx30(-/-) mice. Loss of inhibitory ATP regulation causes ENaC activity to be greater in Cx30(-/-) versus wild-type mice, particularly with high sodium intake. This results from compromised ATP release rather than end-organ resistance: ENaC in Cx30(-/-) mice responds to exogenous ATP. Thus, loss of paracrine ATP feedback regulation of ENaC in Cx30(-/-) mice disrupts normal responses to changes in sodium intake. Consequently, ENaC is hyperactive in Cx30(-/-) mice lowering sodium excretion particularly during increases in sodium intake. Clamping mineralocorticoids high in Cx30(-/-) mice fed a high sodium diet causes a marked decline in renal sodium excretion. This is not the case in wild-type mice, which are capable of undergoing aldosterone-escape. This loss of the ability of ENaC to respond to changes in sodium levels contributes to salt-sensitive hypertension in Cx30(-/-) mice.     

Molecular determinants of PI(4,5)P2 and PI(3,4,5)P3 regulation of the epithelial Na+ channel

Phosphatidylinositol 4,5-bisphosphate (PI(4,5)P(2)) and phosphatidylinositol 3,4,5-trisphosphate (PI(3,4,5)P(3)) are physiologically important second messengers. These molecules bind effector proteins to modulate activity. Several types of ion channels, including the epithelial Na(+) channel (ENaC), are phosphoinositide effectors capable of directly interacting with these signaling molecules. Little, however, is known of the regions within ENaC and other ion channels important to phosphoinositide binding and modulation. Moreover, the molecular mechanism of this regulation, in many instances, remains obscure. Here, we investigate modulation of ENaC by PI(3,4,5)P(3) and PI(4,5)P(2) to begin identifying the molecular determinants of this regulation. We identify intracellular regions near the inner membrane interface just following the second transmembrane domains in beta- and gamma- but not alpha-ENaC as necessary for PI(3,4,5)P(2) but not PI(4,5)P(2) modulation. Charge neutralization of conserved basic amino acids within these regions demonstrated that these polar residues are critical to phosphoinositide regulation. Single channel analysis, moreover, reveals that the regions just following the second transmembrane domains in beta- and gamma-ENaC are critical to PI(3,4,5)P(3) augmentation of ENaC open probability, thus, defining mechanism. Unexpectedly, intracellular domains within the extreme N terminus of beta- and gamma-ENaC were identified as being critical to down-regulation of ENaC activity and P(o) in response to depletion of membrane PI(4,5)P(2). These regions of the channel played no identifiable role in a PI(3,4,5)P(3) response. Again, conserved positive-charged residues within these domains were particularly important, being necessary for exogenous PI(4,5)P(2) to increase open probability. We conclude that beta and gamma subunits bestow phosphoinositide sensitivity to ENaC with distinct regions of the channel being critical to regulation by PI(3,4,5)P(3) and PI(4,5)P(2). This argues that these phosphoinositides occupy distinct ligand-binding sites within ENaC to modulate open probability.     

The chronoinotropic effects of new regulatory input to the heart of land pulmonates

The postjunctional potentials and chronoinotropic reactions of the heart evoked by activation of multimodal neurons and/or left pallial nerve were investigated in three species of land pulmonates: Achatina fulica Ferrussac, Helix lucorum L., Arianta arbustorum L. Both spikes of giant homologous neurons (d-VLN, d-RPLN by A. fulica, command neurons of pneumostoma LPa3, RPa3 by H. lucorum) and stimulation of the peripheral end of the left pallial nerve evoked the similar biphasic inhibitory-excitatory junction potentials in the heart, in mantle muscles and in different parts of visceral complex. The positive chronoinotropic effects of this input in the hearts of whole-mount preparations were modified due to interaction with well-known neural cardioregulating network of the system of intestinal nerve.     

On the empirical choice of the time window for restricted mean survival time

The t-year mean survival or restricted mean survival time (RMST) has been used as an appealing summary of the survival distribution within a time window [0, t]. RMST is the patient's life expectancy until time t and can be estimated nonparametrically by the area under the Kaplan-Meier curve up to t. In a comparative study, the difference or ratio of two RMSTs has been utilized to quantify the between-group-difference as a clinically interpretable alternative summary to the hazard ratio. The choice of the time window [0, t] may be prespecified at the design stage of the study based on clinical considerations. On the other hand, after the survival data have been collected, the choice of time point t could be data-dependent. The standard inferential procedures for the corresponding RMST, which is also data-dependent, ignore this subtle yet important issue. In this paper, we clarify how to make inference about a random “parameter.” Moreover, we demonstrate that under a rather mild condition on the censoring distribution, one can make inference about the RMST up to t, where t is less than or even equal to the largest follow-up time (either observed or censored) in the study. This finding reduces the subjectivity of the choice of t empirically. The proposal is illustrated with the survival data from a primary biliary cirrhosis study, and its finite sample properties are investigated via an extensive simulation study.     

Preservation of fresh edible cactus stems (<i>Opuntia ficus indica</i> Mill.) by modified atmosphere packaging

Cactus stems, the cladodes of Opuntia spp. cacti, are consumed in Mexico and other countries due to their fresh and herbaceous flavor, and because of their widely known nutraceutical benefits. In order to extend the postharvest life of this vegetable, the effect of a modified atmosphere packaging (MAP) was studied in cactus stems of the cultivar Atlixco stored at 4 ± 1 °C for 20 days under three types of atmospheres: (1) air (passive atmosphere), (2) 5 kPa O₂ + 4 kPa CO₂, and (3) N₂. During storage, the titratable acidity decreased and the color of cladodes became darker and less green; however, the 5 kPa O₂ + 4 kPa CO₂ atmosphere was able to preserve both quality characteristics. All modified atmospheres reduced weight loss (from 8 to <2%) and the symptoms of chilling injury, and this physiological disorder appeared earlier in controls than in MAP-stored cladodes. The levels of fermentation metabolites were low in all three evaluated atmospheres. Because of this, only cladodes stored under the N₂ atmosphere were selected for furthersensory analysis of the MAP effect on odor perception as evaluated by a trained panel. Results indicated that there was no detrimental effect (atypical odors) of MAP on this sensory characteristic. We conclude that cultivar Atlixco is suitable for preservation using MAP technology.     

The control of ionized calcium in squid axons

Measurements of the Ca content, [Ca](T), of freshly isolated squid axons show a value of 60 μmol/kg axoplasm. Axons in 3 mM Ca(Na) seawater show little change in Ca content over 4 h, while axons in 3 mM Ca(Na) seawater show little change in Ca content over 4 h, while axons in 10 mM Ca(Na) seawater show gains of 18 μmol/Ca/kgxh. In 10 Ca (Choline) seawater the gain is 2,400 μmol/kgxh. Using aequorin confined to a dialysis capillary in the center of an axon, one finds that [Ca](i) is in a steady state with 3 Ca (Na) seawater, and that both 10 Ca (Na) and 3 Ca (choline) seawater cause increases in [Ca](i). In 3 Ca (Na) seawater-3 Ca (choline) seawater mixtures, 180 mM [Na](0) (40 perecent Na) is as effective as 450 mM [Na](0) (100 percent Na) in maintaining a normal [Ca](1); lower [Na] causes an increase in [Ca](i). If axons are injected with the ATP-splitting enzyme apyrase, the resulting [Ca](1) is not loading with high [Ca](0) or low [Na](0) solutions. Depolarization of an axon with 100 mM K (Na) seawater leads to an increase in the steady-state level of [Ca](1) that is reversed upon returning the axon to normal seawater. Freshly isolated axons treated with either CN or FCCP to inhibit mitochondrial Ca buffering can still maintain a normal [Ca](i) in 1 Ca (Na) seawater.     

Synthesis of amide and sulfonamide substituted N-aryl 6-aminoquinoxalines as PFKFB3 inhibitors with improved physicochemical properties

In oncology, the “Warburg effect” describes the elevated production of energy by glycolysis in cancer cells. The ubiquitous and hypoxia-induced 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3) plays a noteworthy role in the regulation of glycolysis by producing fructose-2,6-biphosphate (F-2,6-BP), a potent activator of the glycolysis rate-limiting phosphofructokinase PFK-1. Series of amides and sulfonamides derivatives based on a N-aryl 6-aminoquinoxaline scaffold were synthesized and tested for their inhibition of PFKFB3 in vitro in a biochemical assay as well as in HCT116 cells. The carboxamide series displayed satisfactory kinetic solubility and metabolic stability, and within this class, potent lead compounds with low nanomolar activity have been identified with a suitable profile for further in vivo evaluation.     

Paracrine regulation of the epithelial Na+ channel in the mammalian collecting duct by purinergic P2Y2 receptor tone

Growing evidence implicates a key role for extracellular nucleotides in cellular regulation, including of ion channels and renal function, but the mechanisms for such actions are inadequately defined. We investigated purinergic regulation of the epithelial Na+ channel (ENaC) in mammalian collecting duct. We find that ATP decreases ENaC activity in both mouse and rat collecting duct principal cells. ATP and other nucleotides, including UTP, decrease ENaC activity via apical P2Y2 receptors. ENaC in collecting ducts isolated from mice lacking this receptor have blunted responses to ATP. P2Y2 couples to ENaC via PLC; direct activation of PLC mimics ATP action. Tonic regulation of ENaC in the collecting duct occurs via locally released ATP; scavenging endogenous ATP and inhibiting P2 receptors, in the absence of other stimuli, rapidly increases ENaC activity. Moreover, ENaC has greater resting activity in collecting ducts from P2Y2-/- mice. Loss of collecting duct P2Y2 receptors in the knock-out mouse is the primary defect leading to increased ENaC activity based on the ability of direct PLC stimulation to decrease ENaC activity in collecting ducts from P2Y2-/- mice in a manner similar to ATP in collecting ducts from wild-type mice. These findings demonstrate that locally released ATP acts in an autocrine/paracrine manner to tonically regulate ENaC in mammalian collecting duct. Loss of this intrinsic regulation leads to ENaC hyperactivity and contributes to hypertension that occurs in P2Y2 receptor-/- mice. P2Y2 receptor activation by nucleotides thus provides physiologically important regulation of ENaC and electrolyte handling in mammalian kidney.     

Visualization of elusive structures using intracardiac echocardiography: Insights from electrophysiology

Echocardiography has the ability to noninvasively explore hemodynamic variables during pharmacologic or exercise stress test in patients with heart failure. In this review, we detail some important potential applications of stress echocardiography in patients with heart failure. In patients with coronary artery disease and chronic LV dysfunction, dobutamine stress echocardiography is able to distinguish between viable and fibrotic tissue to make adequate clinical decisions. Exercise testing, in combination with echocardiographic monitoring, is a method of obtaining accurate information in the assessment of functional capacity and prognosis. Functional mitral regurgitation is a common finding in patients with dilated and ischaemic cardiomyopathy and stress echocardiography in the form of exercise or pharmacologic protocols can be useful to evaluate the behaviour of mitral regurgitation. It is clinical useful to search the presence of contractile reserve in non ischemic dilated cardiomyopathy such as to screen or monitor the presence of latent myocardial dysfunction in patients who had exposure to cardiotoxic agents. Moreover, in patients with suspected diastolic heart failure and normal systolic function, exercise echocardiography could be able to demonstrate the existence of such dysfunction and determine that it is sufficient to limit exercise tolerance. Finally, in the aortic stenosis dobutamine echocardiography can distinguish severe from non-severe stenosis in patients with low transvalvular gradients and depressed left ventricular function.