Sublithospheric diamond ages and the supercontinent cycle.

Subduction related to the ancient supercontinent cycle is poorly constrained by mantle samples. Sublithospheric diamond crystallization records the release of melts from subducting oceanic lithosphere at 300-700 km depths1,2 and is especially suited to tracking the timing and effects of deep mantle processes on supercontinents. Here we show that four isotope systems (Rb-Sr, Sm-Nd, U-Pb and Re-Os) applied to Fe-sulfide and CaSiO3 inclusions within 13 sublithospheric diamonds from Juína (Brazil) and Kankan (Guinea) give broadly overlapping crystallization ages from around 450 to 650 million years ago. The intracratonic location of the diamond deposits on Gondwana and the ages, initial isotopic ratios, and trace element content of the inclusions indicate formation from a peri-Gondwanan subduction system. Preservation of these Neoproterozoic-Palaeozoic sublithospheric diamonds beneath Gondwana until its Cretaceous breakup, coupled with majorite geobarometry3,4, suggests that they accreted to and were retained in the lithospheric keel for more than 300 Myr during supercontinent migration. We propose that this process of lithosphere growth-with diamonds attached to the supercontinent keel by the diapiric uprise of depleted buoyant material and pieces of slab crust-could have enhanced supercontinent stability.

A new family of glutamate-gated chloride channels in parasitic sea louse Caligus rogercresseyi: A subunit refractory to activation by ivermectin is dominant in heteromeric assemblies.

Sea louse ectoparasitosis is a major threat to fish aquaculture. Avermectins such as ivermectin and emamectin have been effectively used against sea louse infestation, but the emergence of resistance has limited their use. A better understanding of the molecular targets of avermectins is essential to the development of novel treatment strategies or new, more effective drugs. Avermectins are known to act by inhibiting neurotransmission through allosteric activation of glutamate-gated chloride channels (GluCls). We have investigated the GluCl subunit present in Caligus rogercresseyi, a sea louse affecting aquaculture in the Southern hemisphere. We identify four new subunits, CrGluCl-B to CrGluCl-E, and characterise them functionally. CrGluCl-A (previously reported as CrGluClα), CrGluCl-B and CrGluCl-C all function as glutamate channel receptors with different sensitivities to the agonist, but in contrast to subunit -A and -C, CrGluCl-B is not activated by ivermectin but is rather antagonised by the drug. CrGluCl-D channel appears active in the absence of any stimulation by glutamate or ivermectin and CrGluCl-E does not exhibit any activity. Notably, the expression of CrGluCl-B with either -A or -C subunits gives rise to receptors unresponsive to ivermectin and showing altered response to glutamate, suggesting that coexpression has led to the preferential formation of heteromers to which the presence of CrGluCl-B confers the property of ivermectin-activation refractoriness. Furthermore, there was evidence for heteromer formation with novel properties only when coexpressing pairs E/C and D/B CrGluCl subtypes. Site-directed mutagenesis shows that three transmembrane domain residues contribute to the lack of activation by ivermectin, most crucially Gln 15' in M2, with mutation Q15'T (the residue present in ivermectin-activated subunits A and C) conferring ivermectin activation to CrGluCl-B. The differential response to avermectin of these Caligus rogercresseyi GluClsubunits, which are highly conserved in the Northern hemisphere sea louse Lepeophtheirus salmonis, could have an influence on the response of these parasites to treatment with macrocyclic lactones. They could serve as molecular markers to assess susceptibility to existing treatments and might be useful molecular targets in the search for novel antiparasitic drugs.

Normal vision and development in mice with low functional expression of Kir7.1 in heterozygosis for a blindness-producing mutation inactivating the channel.

K+ channel Kir7.1 expressed at the apical membrane of the retinal pigment epithelium (RPE) plays an essential role in retinal function. An isoleucine-to-threonine mutation at position 120 of the protein is responsible for blindness-causing vitreo-retinal dystrophy. We have studied the molecular mechanism of action of Kir7.1-I120T in vitro by heterologous expression and in vivo in CRISPR-generated knockin mice. Full-size Kir7.1-I120T reaches the plasma membrane but lacks any activity. Analysis of Kir7.1 and the I120T mutant in mixed transfection experiments, and that of tandem tetrameric constructs made by combining wild type (WT) and mutant protomers, leads us to conclude that they do not form heterotetramers in vitro. Homozygous I120T/I120T mice show cleft palate and tracheomalacia and do not survive beyond P0, whereas heterozygous WT/I120T develop normally. Membrane conductance of RPE cells isolated from WT/WT and heterozygous WT/I120T mice is dominated by Kir7.1 current. Using Rb+ as a charge carrier, we demonstrate that the Kir7.1 current of WT/I120T RPE cells corresponds to approximately 50% of that in cells from WT/WT animals, in direct proportion to WT gene dosage. This suggests a lack of compensatory effects or interference from the mutated allele product, an interpretation consistent with results obtained using WT/- hemizygous mouse. Electroretinography and behavioral tests also show normal vision in WT/I120T animals. The hypomorphic ion channel phenotype of heterozygous Kir7.1-I120T mutants is therefore compatible with normal development and retinal function. The lack of detrimental effect of this degree of functional deficit might explain the recessive nature of Kir7.1 mutations causing human eye disease.NEW & NOTEWORTHY Human retinal pigment epithelium K+ channel Kir7.1 is affected by generally recessive mutations leading to blindness. We investigate one such mutation, isoleucine-to-threonine at position 120, both in vitro and in vivo in knockin mice. The mutated channel is inactive and in heterozygosis gives a hypomorphic phenotype with normal retinal function. Mutant channels do not interfere with wild-type Kir7.1 channels which are expressed concomitantly without hindrance, providing an explanation for the recessive nature of the disease.

Effect of essential oil of Alpinia zerumbet on cardiovascular and autonomic function in rats with isoproterenol induced acute myocardial infarction.

Alpinia zerumbet is a plant popularly used to treat hypertension and anxiety. Studies with Alpinia zerumbet demonstrate antihypertensive and vasodilator effects, among others. The objective of this study was to analyze the effect of essential oil of Alpinia zerumbet (EOAz) on cardiovascular and autonomic function in rats with isoproterenol-induced myocardial infarction. Male Wistar rats (n=32) were equally allocated into four groups: Control, ISO (150mg/kg, subcutaneous), EOAz (100mg/kg by gavage), ISO+EOAz. The rats were evaluated for cardiovascular and, autonomic parameters, electrocardiogram, and infarct size. EOAz was not able to reduce the electrocardiographic variations induced by ISO. Heart rate variability showed a decrease in sympathetic modulation on the heart in the groups treated with EOAz. The cardiopulmonary reflex induced by serotonin invoked a superior blood pressure variation at the 2 µg/kg dose in the EOAz treated groups, while the heart rate variation was significantly higher at the 16 µg/kg dose, when compared to other doses, in all groups, except EOAz+ISO. The sympathetic vagal index was higher in ISO group than in control. EOAz did not reduce the infarct size. We conclude that pretreatment with EOAz does not reverse the hemodynamic and electrocardiographic damage caused by isoproterenol but does reduce sympathetic modulation.

Molecular aspects of structure, gating, and physiology of pH-sensitive background K2P and Kir K+-transport channels.

K(+) channels fulfill roles spanning from the control of excitability to the regulation of transepithelial transport. Here we review two groups of K(+) channels, pH-regulated K2P channels and the transport group of Kir channels. After considering advances in the molecular aspects of their gating based on structural and functional studies, we examine their participation in certain chosen physiological and pathophysiological scenarios. Crystal structures of K2P and Kir channels reveal rather unique features with important consequences for the gating mechanisms. Important tasks of these channels are discussed in kidney physiology and disease, K(+) homeostasis in the brain by Kir channel-equipped glia, and central functions in the hearing mechanism in the inner ear and in acid secretion by parietal cells in the stomach. K2P channels fulfill a crucial part in central chemoreception probably by virtue of their pH sensitivity and are central to adrenal secretion of aldosterone. Finally, some unorthodox behaviors of the selectivity filters of K2P channels might explain their normal and pathological functions. Although a great deal has been learned about structure, molecular details of gating, and physiological functions of K2P and Kir K(+)-transport channels, this has been only scratching at the surface. More molecular and animal studies are clearly needed to deepen our knowledge.

Access to Contraceptive Services in Puerto Rico: An Analysis of Policy and Practice Change Strategies, 2015-2018.

CONTEXT: During the 2016-2017 Zika virus outbreak in Puerto Rico, preventing unintended pregnancy was a primary strategy to reduce Zika-related adverse birth outcomes. The Zika Contraception Access Network (Z-CAN) was a short-term emergency response intervention that used contraception to prevent unintended pregnancy among women who chose to delay or avoid pregnancy. OBJECTIVE: This analysis reports on the identified policy and practice change strategies to increase access to or provision of contraceptive services in Puerto Rico between 2015 and 2018. METHODS: A policy review was conducted to document federal- and territorial-level programs with contraceptive coverage and payment policies in Puerto Rico and to identify policy and practice change. Semistructured interviews with key stakeholders in Puerto Rico were also conducted to understand perceptions of policy and practice change efforts following the Zika virus outbreak, including emergency response, local, and policy efforts to improve contraception access in Puerto Rico. RESULTS: Publicly available information on federal and territorial programs with policies that facilitate access, delivery, and utilization of contraceptive coverage and family planning services in Puerto Rico to support contraceptive access was documented; however, interview results indicated that the implementation of the policies was often limited by barriers and that policy and practice changes as the result of the Zika virus outbreak were short-term. CONCLUSION: Consideration of long-term policy and practice changes related to contraceptive access is warranted. Similar analyses can be used to identify policies, practices, and perceptions in other settings in which the goal is to increase access to contraception or reduce unintended pregnancy.

Effects of Nb Additions and Heat Treatments on the Microstructure, Hardness and Wear Resistance of CuNiCrSiCoTiNbx High-Entropy Alloys.

In this research, a set of CuNiCrSiCoTi (H-0Nb), CuNiCrSiCoTiNb0.5 (H-0.5Nb) and CuNiCrSiCoTiNb1 (H-1Nb) high-entropy alloys (HEAs) were melted in a vacuum induction furnace. The effects of Nb additions on the microstructure, hardness, and wear behavior of these HEAs (compared with a CuBe commercial alloy) in the as-cast (AC) condition, and after solution (SHT) and aging (AT) heat treatments, were investigated using X-ray diffraction, optical microscopy, and electron microscopy. A ball-on-disc configuration tribometer was used to study wear behavior. XRD and SEM results showed that an increase in Nb additions and modification by heat treatment (HT) favored the formation of BCC and FCC crystal structures (CS), dendritic regions, and the precipitation of phases that promoted microstructure refinement during solidification. Increases in hardness of HEA systems were recorded after heat treatment and Nb additions. Maximum hardness values were recorded for the H-1Nb alloy with measured increases from 107.53 HRB (AC) to 112.98 HRB, and from 1104 HV to 1230 HV (aged for 60 min). However, the increase in hardness caused by Nb additions did not contribute to wear resistance response. This can be attributed to a high distribution of precipitated phases rich in high-hardness NiSiTi and CrSi. Finally, the H-0Nb alloy exhibited the best wear resistance behavior in the aged condition of 30 min, with a material loss of 0.92 mm3.

Altered phosphatidylinositol regulation of mutant inwardly rectifying K+ Kir7.1 channels associated with inherited retinal degeneration disease.

KEY POINTS: Kir7.1 K+ channel expressed in retinal pigment epithelium is mutated in inherited retinal degeneration diseases. We study Kir7.1 in heterologous expression to test the hypothesis that pathological R162 mutation to neutral amino acids results in loss of a crucial site that binds PI(4,5)P2 . Although R162W mutation inactivates Kir7.1, changes to smaller volume (e.g. Gln) amino acids are tolerated or even enhance function (Ala or Cys). Chemical modification of Kir7.1-R162C confirms that large residues of the size of Trp are incompatible with normal channel function even if positively charged. In addition to R162, K164 (and possibly K159) forms a binding site for the phosphoinositide and is essential for channel activity. R162 substitution with a large, neutral side chain like Trp exerts a dominant negative effect on Kir7.1 activity such that less than one fifth of the full activity is expected in a cell expressing the same amount of mutant and wild-type channels. ABSTRACT: Mutations in the Kir7.1 K+ channel, highly expressed in retinal pigment epithelium, have been linked to inherited retinal degeneration diseases. Examples are mutations changing Arg 162 to Trp in snowflake vitreoretinal degeneration (SVD) and Gln in retinitis pigmentosa. R162 is believed to be part of a site that binds PI(4,5)P2 and stabilises the open state. We have tested the hypothesis that R162 mutation to neutral amino acids will result in the loss of this crucial interaction to the detriment of channel function. Our findings indicate that although R612W mutation inactivates Kir7.1, changes to smaller volume (e.g. Gln) amino acids are tolerated or even enhance function (Ala or Cys). Cys chemical modification of Kir7.1-R162C confirms that large residues of the size of Trp are incompatible with normal channel function even if positively charged. Experiments titrating the levels of plasma membrane PI(4,5)P2 with voltage-dependent phosphatase DrVSP reveal that, in addition to R162, K164 (and possibly K159) forms a binding site for the phosphoinositide and ensures channel activity. Finally, the use of a concatemeric approach shows that substitution of R162 with a large, neutral side chain mimicking a Trp residue exerts a dominant negative effect on Kir7.1 activity such that less than one fifth of the full activity is expected in heterozygous cells carrying the SVD mutation. Our results suggest that if mutations in the human KCNJ13 gene resulting in the neutralisation of R162 and Kir7.1 malfunction led to retinal degeneration diseases, their severity might depend on the nature of the side chain of the replacing amino acid.

Delayed Lubricant Depletion of Slippery Liquid Infused Porous Surfaces Using Precision Nanostructures.

Slippery liquid infused porous surfaces (SLIPS) are an important class of repellent materials, comprising micro/nanotextures infused with a lubricating liquid. Unlike superhydrophobic surfaces, SLIPS do not rely on a stable air-liquid interface and thus can better manage low surface tension fluids, are less susceptible to damage under physical stress, and are able to self-heal. However, these collective properties are only efficient as long as the lubricant remains infused, which has proved challenging. We hypothesized that, in comparison to a nanohole and nanopillar morphology, the "hybrid" morphology of a hole within a nanopillar, namely a nanotube, would be able to retain and redistribute lubricant more effectively, owing to capillary forces trapping a reservoir of lubricant within the tube, while lubricant between tubes can facilitate redistribution to depleted areas. By virtue of recent fabrication advances in spacer defined intrinsic multiple patterning (SDIMP), we fabricated an array of silicon nanotubes and equivalent arrays of nanoholes and nanopillars (pitch, 560 nm; height, 2 µm). After infusing the nanostructures (prerendered hydrophobic) with lubricant Krytox 1525, we probed the lubricant stability under dynamic conditions and correlated the degree of the lubricant film discontinuity to changes in the contact angle hysteresis. As a proof of concept, the durability test, which involved consecutive deposition of droplets onto the surface amounting to 0.5 L, revealed 2-fold and 1.5-fold enhancements of lubricant retention in nanotubes in comparison to nanopillars and nanoholes, respectively, showing a clear trajectory for prolonging the lifetime of a slippery surface.

NT-proBNP levels in preeclampsia, intrauterine growth restriction as well as in the prediction on an imminent delivery.

OBJECTIVES: Studies of cardiovascular function in pregnancy have shown inconsistent and, in some cases, contradictory results, particularly regarding cardiac output. While some studies report preeclampsia associated with high cardiac output, other studies suggest that preeclampsia should be further subdivided into women with high or low cardiac output. This study was conducted to examine the NT-proBNP levels in preeclampsia, intrauterine growth restriction, and hypertensive pregnancies without preeclampsia. We also examined N-terminal pro-B natriuretic peptide (NT-proBNP) levels three to four months after delivery, in preeclamptic women as well as the prediction of delivery within 10 days. In a reduced number of preeclamptic women and controls we performed echocardiograms to study their diastolic function. METHODS: We investigated the NT-proBNP levels in 213 subjects with preeclampsia only, 73 with intrauterine growth restriction, 44 with preeclampsia and intrauterine growth restriction, 211 who were hypertensive and 662 unaffected pregnancies (controls). We also performed echocardiograms on 36 preeclampsia and 19 controls before delivery and three to five months after delivery. RESULTS: NT-proBNP levels are higher in early onset preeclampsia than in late onset preeclampsia. Intrauterine growth restriction pregnancies showed a NT-proBNP levels similar to hypertensive and unaffected pregnancies. Compared with healthy pregnancies, women with preterm preeclampsia (<37 gestational weeks) had altered left atrial segments. CONCLUSIONS: We observed that NT-proBNP levels are higher in early onset preeclampsia than in late onset. Moreover, diastolic dysfunction is higher in early onset than in late-onset term preeclampsia. An NT-proBNP value >136 pg/mL has a high positive predictive value for an imminent delivery within 10 days.

New 5-carba-pterocarpans: Synthesis and preliminary antiproliferative activity on a panel of human cancer cells.

Natural pterocarpans and synthetic 5-carba-pterocarpans are isosteres in which the oxygen atom at position 5 in the pyran-ring of pterocarpans is replaced by a methylene group. These 5-carba-analogues were obtained in good yields through the palladium-catalyzed oxyarylation of alcoxy-1,2-dihydronaphthalens with o-iodophenols in PEG-400. They were evaluated on human cancer cell lineages derived respectively from prostate tumor (PC3, IC50 = 11.84 µmol L-1, SI > 12)) and acute myeloid leukemia (HL-60, IC50 = 8.81 µmol L-1, SI > 16), highly incident cancer types presenting resistance against traditional chemotherapeutics. Compound 6c (LQB-492) was the most potent (IC50 = 3.85 µmol L-1, SI > 37) in SF-295 cell lineage (glioblastoma). Such findings suggest that 5-carba-pterocarpan can potentially be new hit compounds for further development of novel antiproliferative agents.

Household plastic waste habits and attitudes: A pilot study in the city of Valencia.

Bearing in mind that only 42% of plastic packaging post-consumer waste is recycled in Europe, the European Directive 2018/852 established the key target of a 55% plastic packaging waste recycling rate by 2030. For this reason, PlastiCircle, funded by the European Union's Horizon 2020 research and innovation program project, aims to foster the recycling of packaging, improve all stages of the waste collection, and promote responsible consumption. Three European cities have been selected as locations for pilot implementation: Valencia (Spain), Utrecht (The Netherlands) and Alba Iulia (Romania). The main objective of the present study has been to evaluate the participants' opinion and attitudes on plastic recycling. This paper presents the results from the district of San Marcelino in the city of Valencia, the first PlastiCircle pilot to face the challenges of encouraging households to participate more in plastic waste sorting and recycling.

The role of mitochondrial oxidative stress in the metabolic alterations in diet-induced obesity in rats.

The impact of the mitochondria-targeted antioxidant MitoQ was evaluated in the metabolic alterations and the adipose tissue remodeling associated with obesity. Male Wistar rats were fed either a high-fat diet (HFD; 35% fat) or a standard diet (3.5% fat) for 7 wk and treated with MitoQ (200 µM). A proteomic analysis of visceral adipose tissue from patients with obesity and patients without obesity was performed. MitoQ partially prevented the increase in body weight, adiposity, homeostasis model assessment index, and adipose tissue remodeling in HFD rats. It also ameliorated protein level changes of factors involved in insulin signaling observed in adipose tissue of obese rats: reductions in adiponectin and glucose transporter 4 (GLUT 4) and increases in dipeptidylpeptidase 4, suppressor of cytokine signaling 3 (SOCS3), and insulin receptor substrate 1 phosphorylation. MitoQ prevented down-regulation of adiponectin and GLUT 4 and increases in SOCS3 levels in a TNF-α-induced insulin-resistant 3T3-L1 adipocyte model. MitoQ also ameliorated alterations in mitochondrial proteins observed in obese rats: increases in cyclophylin F and carnitine palmitoyl transferase 1A and reductions in mitofusin1, peroxiredoxin 4, and fumarate hydratase. The proteomic analysis of the visceral adipose tissue from patients with obesity show alterations in mitochondrial proteins similar to those observed in obese rats. Therefore, the data show the beneficial effect of MitoQ in the metabolic dysfunction induced by obesity.-Marín-Royo, G., Rodríguez, C., Le Pape, A., Jurado-López, R., Luaces, M., Antequera, A., Martínez-González, J., Souza-Neto, F. V., Nieto, M. L., Martínez-Martínez, E., Cachofeiro, V. The role of mitochondrial oxidative stress in the metabolic alterations in diet-induced obesity in rats.

A more accurate prediction to rule in and rule out pre-eclampsia using the sFlt-1/PlGF ratio and NT-proBNP as biomarkers.

Background The management of potential pre-eclamptic patients using the soluble FMS-like tyrosine kinase 1 (sFlt-1)/ placental growth factor (PlGF) ratio is characterised by frequent false-positive results. Methods A retrospective cohort study was conducted to identify and validate cut-off values, obtained using a machine learning model, for the sFlt-1/PlGF ratio and NT-proBNP that would be predictive of the absence or presence of early-onset pre-eclampsia (PE) in singleton pregnancies presenting at 24 to 33 + 6 weeks of gestation. Results For the development cohort, we defined two sFlt-1/PlGF ratio cut-off values of 23 and 45 to rule out and rule in early-onset PE at any time between 24 and 33 + 6 weeks of gestation. Using an sFlt-1/PlGF ratio cut-off value of 23, the negative predictive value (NPV) for the development of early-onset PE was 100% (95% confidence interval [CI]: 99.5-100). The positive predictive value (PPV) of an sFlt-1/PlGF ratio >45 for a diagnosis of early-onset PE was 49.5% (95% CI: 45.8-55.6). When an NT-proBNP value >174 was combined with an sFlt-1/PlGF ratio >45, the PPV was 86% (95% CI: 79.2-92.6). In the validation cohort, the negative and positive values were very similar to those found for the development cohort. Conclusions An sFlt-1/PlGF ratio <23 rules out early-onset PE between 24 and 33 + 6 weeks of gestation at any time, with an NPV of 100%. An sFlt-1/PlGF ratio >45 with an NT-proBNP value >174 significantly enhances the probability of developing early-onset PE.

Chaotic Image Encryption Using Hopfield and Hindmarsh-Rose Neurons Implemented on FPGA.

Chaotic systems implemented by artificial neural networks are good candidates for data encryption. In this manner, this paper introduces the cryptographic application of the Hopfield and the Hindmarsh-Rose neurons. The contribution is focused on finding suitable coefficient values of the neurons to generate robust random binary sequences that can be used in image encryption. This task is performed by evaluating the bifurcation diagrams from which one chooses appropriate coefficient values of the mathematical models that produce high positive Lyapunov exponent and Kaplan-Yorke dimension values, which are computed using TISEAN. The randomness of both the Hopfield and the Hindmarsh-Rose neurons is evaluated from chaotic time series data by performing National Institute of Standard and Technology (NIST) tests. The implementation of both neurons is done using field-programmable gate arrays whose architectures are used to develop an encryption system for RGB images. The success of the encryption system is confirmed by performing correlation, histogram, variance, entropy, and Number of Pixel Change Rate (NPCR) tests.

The Effect of Nanosystems on ATP-Binding Cassette Transporters: Understanding the Influence of Nanosystems on Multidrug Resistance Protein-1 and P-glycoprotein.

The cancer multidrug resistance is involved in the failure of several treatments during cancer treatment. It is a phenomenon that has been receiving great attention in the last years due to the sheer amount of mechanisms discovered and involved in the process of resistance which hinders the effectiveness of many anti-cancer drugs. Among the mechanisms involved in the multidrug resistance, the participation of ATP-binding cassette (ABC) transporters is the main one. The ABC transporters are a group of plasma membrane and intracellular organelle proteins involved in the process of externalization of substrates from cells, which are expressed in cancer. They are involved in the clearance of intracellular metabolites as ions, hormones, lipids and other small molecules from the cell, affecting directly and indirectly drug absorption, distribution, metabolism and excretion. Other mechanisms responsible for resistance are the signaling pathways and the anti- and pro-apoptotic proteins involved in cell death by apoptosis. In this study we evaluated the influence of three nanosystem (Graphene Quantum Dots (GQDs), mesoporous silica (MSN) and poly-lactic nanoparticles (PLA)) in the main mechanism related to the cancer multidrug resistance such as the Multidrug Resistance Protein-1 and P-glycoprotein. We also evaluated this influence in a group of proteins involved in the apoptosis-related resistance including cIAP-1, XIAP, Bcl-2, BAK and Survivin proteins. Last, colonogenic and MTT (3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenyltetrazolium bromide) assays have also been performed. The results showed, regardless of the concentration used, GQDs, MSN and PLA were not cytotoxic to MDA-MB-231 cells and showed no impairment in the colony formation capacity. In addition, it has been observed that P-gp membrane expression was not significantly altered by any of the three nanomaterials. The results suggest that GQDs nanoparticles would be suitable for the delivery of other multidrug resistance protein 1 (MRP1) substrate drugs that bind to the transporter at the same binding pocket, while MSN can strongly inhibit doxorubicin efflux by MRP1. On the other hand, PLA showed moderate inhibition of doxorubicin efflux by MRP1 suggesting that this nanomaterial can also be useful to treat MDR (Multidrug resistance) due to MRP1 overexpression.

Elucidating the Structural Basis of the Intracellular pH Sensing Mechanism of TASK-2 K2P Channels.

Two-pore domain potassium (K2P) channels maintain the cell's background conductance by stabilizing the resting membrane potential. They assemble as dimers possessing four transmembrane helices in each subunit. K2P channels were crystallized in "up" and "down" states. The movements of the pore-lining transmembrane TM4 helix produce the aperture or closure of side fenestrations that connect the lipid membrane with the central cavity. When the TM4 helix is in the up-state, the fenestrations are closed, while they are open in the down-state. It is thought that the fenestration states are related to the activity of K2P channels and the opening of the channels preferentially occurs from the up-state. TASK-2, a member of the TALK subfamily of K2P channels, is opened by intracellular alkalization leading the deprotonation of the K245 residue at the end of the TM4 helix. This charge neutralization of K245 could be sensitive or coupled to the fenestration state. Here, we describe the relationship between the states of the intramembrane fenestrations and K245 residue in TASK-2 channel. By using molecular modeling and simulations, we show that the protonated state of K245 (K245+) favors the open fenestration state and, symmetrically, that the open fenestration state favors the protonated state of the lysine residue. We show that the channel can be completely blocked by Prozac, which is known to induce fenestration opening in TREK-2. K245 protonation and fenestration aperture have an additive effect on the conductance of the channel. The opening of the fenestrations with K245+ increases the entrance of lipids into the selectivity filter, blocking the channel. At the same time, the protonation of K245 introduces electrostatic potential energy barriers to ion entrance. We computed the free energy profiles of ion penetration into the channel in different fenestration and K245 protonation states, to show that the effects of the two transformations are summed up, leading to maximum channel blocking. Estimated rates of ion transport are in qualitative agreement with experimental results and support the hypothesis that the most important barrier for ion transport under K245+ and open fenestration conditions is the entrance of the ions into the channel.

A novel Kir7.1 splice variant expressed in various mouse tissues shares organisational and functional properties with human Leber amaurosis-causing mutations of this K+ channel.

Kir7.1 is an inwardly rectifying K+ channel present in epithelia where it shares membrane localization with the Na+/K+-pump. In the present communication we report the presence of a novel splice variant of Kir7.1 in mouse tissues including kidney, lung, choroid plexus and retinal pigment epithelium (RPE). The variant named mKir7.1-SV2 lacks most of the C-terminus domain but is predicted to have the two transmembrane domains and permeation pathway unaffected. Similarly truncated predicted proteins, Kir7.1-R166X and Kir7.1-Q219X, would arise from mutations associated with Leber Congenital Amaurosis, a rare recessive hereditary retinal disease that results in vision loss at early age. We found that mKir7.1-SV2 and the pathological variants do not produce any channel activity when expressed alone in HEK-293 cells due to their scarce presence in the plasma membrane. Simultaneous expression with the full length Kir7.1 however leads to a reduction in activity of the wild-type channel that might be due to partial proteasome degradation of WT-mutant channel heteromers.

Kir7.1 inwardly rectifying K+ channel is expressed in ciliary body non pigment epithelial cells and might contribute to intraocular pressure regulation.

Inwardly rectifying K+ channel Kir7.1 is expressed in epithelia where it shares membrane localisation with the Na+/K+-pump. The ciliary body epithelium (CBE) of the eye is a determinant of intraocular pressure (IOP) through NaCl-driven fluid secretion of aqueous humour. In the present study we explored the presence Kir7.1 in this epithelium in the mouse and its possible functional role in the generation of IOP. Use heterozygous animals for total Kir7.1 knockout expressing ß-galactosidase under the control of Kir7.1 promoter, identified the expression of Kir7.1 in non-pigmented epithelial cells of CBE. Using conditional, floxed knockout Kir7.1 mice as negative controls, we found Kir7.1 at the basolateral membrane of the same CBE cell layer. This was confirmed using a knockin mouse expressing the Kir7.1 protein tagged with a haemagglutinin epitope. Measurements using the conditional knockout mouse show only a minor effect of Kir7.1 inactivation on steady-state IOP. Transient increases in IOP in response to general anaesthetics, or to water injection, are absent or markedly curtailed in Kir7.1-deficient mice. These results suggest a role for Kir7.1 in IOP regulation through a possible modulation of aqueous humour production by the CBE non-pigmented epithelial cells. The location of Kir7.1 in the CBE, together with the effect of its removal on dynamic changes in IOP, point to a possible role of the channel as a leak pathway preventing cellular overload of K+ during the secretion process. Kir7.1 could be used as a potential therapeutic target in pathological conditions leading to elevated intraocular pressure.

Structure/Activity Analysis of TASK-3 Channel Antagonists Based on a 5,6,7,8 tetrahydropyrido[4,3-d]pyrimidine.

TASK-3 potassium (K+) channels are highly expressed in the central nervous system, regulating the membrane potential of excitable cells. TASK-3 is involved in neurotransmitter action and has been identified as an oncogenic K+ channel. For this reason, the understanding of the action mechanism of pharmacological modulators of these channels is essential to obtain new therapeutic strategies. In this study we describe the binding mode of the potent antagonist PK-THPP into the TASK-3 channel. PK-THPP blocks TASK-1, the closest relative channel of TASK-3, with almost nine-times less potency. Our results confirm that the binding is influenced by the fenestrations state of TASK-3 channels and occurs when they are open. The binding is mainly governed by hydrophobic contacts between the blocker and the residues of the binding site. These interactions occur not only for PK-THPP, but also for the antagonist series based on 5,6,7,8 tetrahydropyrido[4,3-d]pyrimidine scaffold (THPP series). However, the marked difference in the potency of THPP series compounds such as 20b, 21, 22 and 23 (PK-THPP) respect to compounds such as 17b, inhibiting TASK-3 channels in the micromolar range is due to the presence of a hydrogen bond acceptor group that can establish interactions with the threonines of the selectivity filter.