ResearchPad - basic-amino-acids https://www.researchpad.co Default RSS Feed en-us © 2020 Newgen KnowledgeWorks <![CDATA[The impact of rumen-protected amino acids on the expression of key- genes involved in the innate immunity of dairy sheep]]> https://www.researchpad.co/article/elastic_article_14615 Rumen protected amino acids inclusion in ewes’ diets has been proposed to enhance their innate immunity. The objective of this work was to determine the impact of dietary supplementation with rumen-protected methionine or lysine, as well as with a combination of these amino acids in two different ratios, on the expression of selected key-genes (NLRs, MyD88, TRIF, MAPK-1, IRF-3, JunD, TRAF-3, IRF-5, IL-1α, IL-10, IKK-α, STAT-3 and HO-1). Thus, sixty Chios dairy ewes (Ovis aries) were assigned to one of the following five dietary treatments (12 animals/ treatment): A: basal diet consist of concentrates, wheat straw and alfalfa hay (control group); B: basal diet +6.0 g/head rumen-protected methionine; C: basal diet + 5.0 g/head rumen-protected lysine; D: basal diet +6.0 g/head rumen-protected methionine + 5.0 g/head rumen-protected lysine and E: basal diet +12.0 g/head rumen-protected methionine + 5.0 g/head rumen-protected lysine. The results revealed a significant downregulation of relative transcript level of the IL-1α gene in the neutrophils of C and in monocytes of D ewes compared with the control. Significantly lower mRNA transcript accumulation was also observed for the MyD88 gene in the neutrophils of ewes fed with lysine only (C). The mRNA relative expression levels of JunD gene were highly induced in the monocytes, while those of IL-10 and HO-1 genes were declined in the neutrophils of ewes fed with the C and D diets compared with the control. Lower transcript levels of STAT-3 gene were observed in the neutrophils of ewes fed with either C or with E diets in comparison with the control. In conclusion, our results suggest that the dietary supplementation of ewes with rumen-protected amino acids, down regulate the expression of some genes involved in the pro-inflammatory signalling.

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<![CDATA[A modified arginine-depleting enzyme NEI-01 inhibits growth of pancreatic cancer cells]]> https://www.researchpad.co/article/elastic_article_11227 Arginine deprivation cancer therapy targets certain types of malignancies with positive result in many studies and clinical trials. NEI-01 was designed as a novel arginine-depleting enzyme comprising an albumin binding domain capable of binding to human serum albumin to lengthen its half-life. In the present work, NEI-01 is shown to bind to serum albumin from various species, including mice, rat and human. Single intraperitoneal administration of NEI-01 to mice reduced plasma arginine to undetectable level for at least 9 days. Treatment of NEI-01 specifically inhibited cell viability of MIA PaCa-2 and PANC-1 cancer cell lines, which were ASS1 negative. Using a human pancreatic mouse xenograft model, NEI-01 treatment significantly reduced tumor volume and weight. Our data provides proof of principle for a cancer treatment strategy using NEI-01.

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<![CDATA[Uptake and speciation of zinc in edible plants grown in smelter contaminated soils]]> https://www.researchpad.co/article/N21b4cc8f-fdc5-4198-8cfa-2868c1971919

Heavy metal accumulation in edible plants grown in contaminated soils poses a major environmental risk to humans and grazing animals. This study focused on the concentration and speciation of Zn in different edible plants grown in soils contaminated with smelter wastes (Spelter, WV, USA) containing high levels of the metals Zn, Cu, Pb, Cd. Their accumulation was examined in different parts (roots, stem, and leaves) of plants and as a function of growth stage (dry seed, sprouting seed, cotyledon, and leaves) in the root vegetables radish, the leafy vegetable spinach and the legume clover. Although the accumulation of metals varied significantly with plant species, the average metal concentrations were [Zn] > [Pb] > [Cu] > [Cd]. Metal uptake studies were complemented with bulk and micro X-ray absorption spectroscopy (XAS) at Zn K-edge and micro X-ray fluorescence (μXRF) measurements to evaluate the speciation and distribution of Zn in these plant species. Dynamic interplay between the histidine and malate complexation of Zn was observed in all plant species. XRF mapping of spinach leaves at micron spatial resolution demonstrated the accumulation of Zn in vacuoles and leaf tips. Radish root showed accumulation of Zn in root hairs, likely as ZnS nanoparticles. At locations of high Zn concentration in spinach leaves, μXANES suggests Zn complexation with histidine, as opposed to malate in the bulk leaf. These findings shed new light on the dynamic nature of Zn speciation in plants.

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<![CDATA[Citrulline protects mice from experimental cerebral malaria by ameliorating hypoargininemia, urea cycle changes and vascular leak]]> https://www.researchpad.co/article/5c8c194bd5eed0c484b4d370

Clinical and model studies indicate that low nitric oxide (NO) bioavailability due in part to profound hypoargininemia contributes to cerebral malaria (CM) pathogenesis. Protection against CM pathogenesis may be achieved by altering the diet before infection with Plasmodium falciparum infection (nutraceutical) or by administering adjunctive therapy that decreases CM mortality (adjunctive therapy). This hypothesis was tested by administering citrulline or arginine in experimental CM (eCM). We report that citrulline injected as prophylaxis immediately post infection (PI) protected virtually all mice by ameliorating (i) hypoargininemia, (ii) urea cycle impairment, and (iii) disruption of blood brain barrier. Citrulline prophylaxis inhibited plasma arginase activity. Parasitemia was similar in citrulline- and vehicle control-groups, indicating that protection from pathogenesis was not due to decreased parasitemia. Both citrulline and arginine administered from day 1 PI in the drinking water significantly protected mice from eCM. These observations collectively indicate that increasing dietary citrulline or arginine decreases eCM mortality. Citrulline injected ip on day 4 PI with quinine-injected ip on day 6 PI partially protected mice from eCM; citrulline plus scavenging of superoxide with pegylated superoxide dismutase and pegylated catalase protected all recipients from eCM. These findings indicate that ameliorating hypoargininemia with citrulline plus superoxide scavenging decreases eCM mortality.

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<![CDATA[Uncovering and resolving challenges of quantitative modeling in a simplified community of interacting cells]]> https://www.researchpad.co/article/5c99020dd5eed0c484b97589

Quantitative modeling is useful for predicting behaviors of a system and for rationally constructing or modifying the system. The predictive power of a model relies on accurate quantification of model parameters. Here, we illustrate challenges in parameter quantification and offer means to overcome these challenges, using a case example in which we quantitatively predict the growth rate of a cooperative community. Specifically, the community consists of two Saccharomyces cerevisiae strains, each engineered to release a metabolite required and consumed by its partner. The initial model, employing parameters measured in batch monocultures with zero or excess metabolite, failed to quantitatively predict experimental results. To resolve the model–experiment discrepancy, we chemically identified the correct exchanged metabolites, but this did not improve model performance. We then remeasured strain phenotypes in chemostats mimicking the metabolite-limited community environments, while mitigating or incorporating effects of rapid evolution. Almost all phenotypes we measured, including death rate, metabolite release rate, and the amount of metabolite consumed per cell birth, varied significantly with the metabolite environment. Once we used parameters measured in a range of community-like chemostat environments, prediction quantitatively agreed with experimental results. In summary, using a simplified community, we uncovered and devised means to resolve modeling challenges that are likely general to living systems.

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<![CDATA[Ser96Ala genetic variant of the human histidine-rich calcium-binding protein is a genetic predictor of recurrence after catheter ablation in patients with paroxysmal atrial fibrillation]]> https://www.researchpad.co/article/5c897742d5eed0c4847d2858

Background

Atrial fibrillation (AF) recurrence after radiofrequency catheter ablation (RFCA) still remains a serious issue. Ca2+ handling has a considerable effect on AF recurrence. The histidine-rich calcium-binding protein (HRC) genetic single nucleotide polymorphism (SNP), rs3745297 (T>G, Ser96Ala), is known to cause a sarcoplasmic reticulum Ca2+ leak. We investigated the association between HRC Ser96Ala and AF recurrence after RFCA in paroxysmal AF (PAF) patients.

Methods and results

We enrolled PAF patients who underwent RFCA (N = 334 for screening and N = 245 for replication) and were genotyped for HRC SNP (rs3745297). The patient age was younger and rate of diabetes and hypertension lower in the PAF patients with Ser96Ala than in those without (TT/TG/GG, 179/120/35; 64±10/60±12/59±13 y, P = 0.001; 18.5/ 9.2/8.6%, P = 0.04 and 66.1/50.0/37.1%, P = 0.001, respectively). During a mean 19 month follow-up, 57 (17.1%) patients suffered from AF recurrences. The rate of an Ser96Ala was significantly higher in patients with AF recurrence than in those without in the screening set (allele frequency model: odds ratio [OR], 1.80; P = 0.006). We also confirmed this significant association in the replication set (OR 1.74; P = 0.03) and combination (P = 0.0008). A multivariate analysis revealed that the AF duration, sinus node dysfunction, and HRC Ser96Ala were independent predictors of an AF recurrence (hazard ratio [HR], 1.04, P = 0.037; HR 2.42, P = 0.018; and HR 2.66, P = 0.007, respectively).

Conclusion

HRC SNP Ser96Ala is important as a new genetic marker of AF recurrence after RFCA.

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<![CDATA[Novel site-specific PEGylated L-asparaginase]]> https://www.researchpad.co/article/5c6c7587d5eed0c4843cfe5d

L-asparaginase (ASNase) from Escherichia coli is currently used in some countries in its PEGylated form (ONCASPAR, pegaspargase) to treat acute lymphoblastic leukemia (ALL). PEGylation refers to the covalent attachment of poly(ethylene) glycol to the protein drug and it not only reduces the immune system activation but also decreases degradation by plasmatic proteases. However, pegaspargase is randomly PEGylated and, consequently, with a high degree of polydispersity in its final formulation. In this work we developed a site-specific N-terminus PEGylation protocol for ASNase. The monoPEG-ASNase was purified by anionic followed by size exclusion chromatography to a final purity of 99%. The highest yield of monoPEG-ASNase of 42% was obtained by the protein reaction with methoxy polyethylene glycol-carboxymethyl N-hydroxysuccinimidyl ester (10kDa) in 100 mM PBS at pH 7.5 and PEG:ASNase ratio of 25:1. The monoPEG-ASNase was found to maintain enzymatic stability for more days than ASNase, also was resistant to the plasma proteases like asparaginyl endopeptidase and cathepsin B. Additionally, monoPEG-ASNase was found to be potent against leukemic cell lines (MOLT-4 and REH) in vitro like polyPEG-ASNase. monoPEG-ASNase demonstrates its potential as a novel option for ALL treatment, being an inventive novelty that maintains the benefits of the current enzyme and solves challenges.

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<![CDATA[SIRT5 deficiency suppresses mitochondrial ATP production and promotes AMPK activation in response to energy stress]]> https://www.researchpad.co/article/5c6dc99ed5eed0c484529ee3

Sirtuin 5 (SIRT5) is a member of the NAD+-dependent sirtuin family of protein deacylase that catalyzes removal of post-translational modifications, such as succinylation, malonylation, and glutarylation on lysine residues. In light of the SIRT5's roles in regulating mitochondrion function, we show here that SIRT5 deficiency leads to suppression of mitochondrial NADH oxidation and inhibition of ATP synthase activity. As a result, SIRT5 deficiency decreases mitochondrial ATP production, increases AMP/ATP ratio, and subsequently activates AMP-activated protein kinase (AMPK) in cultured cells and mouse hearts under energy stress conditions. Moreover, Sirt5 knockout attenuates transverse aortic constriction (TAC)-induced cardiac hypertrophy and cardiac dysfunction in mice, which is associated with decreased ATP level, increased AMP/ATP ratio and enhanced AMPK activation. Our study thus uncovers an important role of SIRT5 in regulating cellular energy metabolism and AMPK activation in response to energy stress.

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<![CDATA[Lysine 27 of replication-independent histone H3.3 is required for Polycomb target gene silencing but not for gene activation]]> https://www.researchpad.co/article/5c5b52c1d5eed0c4842bcf85

Proper determination of cell fates depends on epigenetic information that is used to preserve memory of decisions made earlier in development. Post-translational modification of histone residues is thought to be a central means by which epigenetic information is propagated. In particular, modifications of histone H3 lysine 27 (H3K27) are strongly correlated with both gene activation and gene repression. H3K27 acetylation is found at sites of active transcription, whereas H3K27 methylation is found at loci silenced by Polycomb group proteins. The histones bearing these modifications are encoded by the replication-dependent H3 genes as well as the replication-independent H3.3 genes. Owing to differential rates of nucleosome turnover, H3K27 acetylation is enriched on replication-independent H3.3 histones at active gene loci, and H3K27 methylation is enriched on replication-dependent H3 histones across silenced gene loci. Previously, we found that modification of replication-dependent H3K27 is required for Polycomb target gene silencing, but it is not required for gene activation. However, the contribution of replication-independent H3.3K27 to these functions is unknown. Here, we used CRISPR/Cas9 to mutate the endogenous replication-independent H3.3K27 to a non-modifiable residue. Surprisingly, we find that H3.3K27 is also required for Polycomb target gene silencing despite the association of H3.3 with active transcription. However, the requirement for H3.3K27 comes at a later stage of development than that found for replication-dependent H3K27, suggesting a greater reliance on replication-independent H3.3K27 in post-mitotic cells. Notably, we find no evidence of global transcriptional defects in H3.3K27 mutants, despite the strong correlation between H3.3K27 acetylation and active transcription.

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<![CDATA[Characterisation of a type II functionally-deficient variant of alpha-1-antitrypsin discovered in the general population]]> https://www.researchpad.co/article/5c424390d5eed0c4845e05dc

Lung disease in alpha-1-antitrypsin deficiency (AATD) results from dysregulated proteolytic activity, mainly by neutrophil elastase (HNE), in the lung parenchyma. This is the result of a substantial reduction of circulating alpha-1-antitrypsin (AAT) and the presence in the plasma of inactive polymers of AAT. Moreover, some AAT mutants have reduced intrinsic activity toward HNE, as demonstrated for the common Z mutant, as well as for other rarer variants. Here we report the identification and characterisation of the novel AAT reactive centre loop variant Gly349Arg (p.G373R) present in the ExAC database. This AAT variant is secreted at normal levels in cellular models of AATD but shows a severe reduction in anti-HNE activity. Biochemical and molecular dynamics studies suggest it exhibits unfavourable RCL presentation to cognate proteases and compromised insertion of the RCL into β-sheet A. Identification of a fully dysfunctional AAT mutant that does not show a secretory defect underlines the importance of accurate genotyping of patients with pulmonary AATD manifestations regardless of the presence of normal levels of AAT in the circulation. This subtype of disease is reminiscent of dysfunctional phenotypes in anti-thrombin and C1-inibitor deficiencies so, accordingly, we classify this variant as the first pure functionally-deficient (type II) AATD mutant.

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<![CDATA[Potent, multi-target serine protease inhibition achieved by a simplified β-sheet motif]]> https://www.researchpad.co/article/5c50c48fd5eed0c4845e88ff

Engagement of an extended β-sheet is a common substrate/inhibitor interaction at the active site of serine proteases and is an important feature of Laskowski mechanism inhibitors that present a substrate-like loop to a target protease. This loop is cleaved but subsequently relegated forming a stable inhibitor/protease complex. Laskowski inhibitors are ubiquitous in nature and are used extensively in serine protease inhibitor design. However, most studies concentrate on introducing new sidechain interactions rather than the direct contributions of the substrate-like β-sheet to enzyme inhibition. Here we report the crystal structure of an simplified β-sheet inhibitory motif within the Sunflower Trypsin Inhibitor (SFTI) in complex with trypsin. We show that the intramolecular hydrogen bond network of this SFTI variant (SFTI-TCTR) engages the inhibitor sidechains that would normally interact with a target protease, giving mainchain interactions a more prominent role in complex formation. Despite having reduced sidechain interactions, this SFTI variant is remarkably potent and inhibits a diverse range of serine proteases. Crystal structural analysis and molecular modelling of SFTI-TCTR complexes again indicates an interface dominated by β–sheet interactions, highlighting the importance of this motif and the adaptability of SFTI as a scaffold for inhibitor design.

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<![CDATA[Urease is an essential component of the acid response network of Staphylococcus aureus and is required for a persistent murine kidney infection]]> https://www.researchpad.co/article/5c390bf2d5eed0c48491f100

Staphylococcus aureus causes acute and chronic infections resulting in significant morbidity. Urease, an enzyme that generates NH3 and CO2 from urea, is key to pH homeostasis in bacterial pathogens under acidic stress and nitrogen limitation. However, the function of urease in S. aureus niche colonization and nitrogen metabolism has not been extensively studied. We discovered that urease is essential for pH homeostasis and viability in urea-rich environments under weak acid stress. The regulation of urease transcription by CcpA, Agr, and CodY was identified in this study, implying a complex network that controls urease expression in response to changes in metabolic flux. In addition, it was determined that the endogenous urea derived from arginine is not a significant contributor to the intracellular nitrogen pool in non-acidic conditions. Furthermore, we found that during a murine chronic renal infection, urease facilitates S. aureus persistence by promoting bacterial fitness in the low-pH, urea-rich kidney. Overall, our study establishes that urease in S. aureus is not only a primary component of the acid response network but also an important factor required for persistent murine renal infections.

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<![CDATA[Partitioning the efficiency of utilization of amino acids in growing broilers: Multiple linear regression and multivariate approaches]]> https://www.researchpad.co/article/5c1ab828d5eed0c484026eed

Determining the efficiency of amino acid (AA) utilization in growing animals is crucial to estimate their requirement accurately. In broiler chickens, the composition of AA in feather is different from feather-free body and the proportion of feathers will change along broiler’s growth, which may impact the efficiency of utilization on AA consumed. Therefore, in order to establish a method that predicts the efficiency of utilization for feather-free body and feather, two approaches were evaluated: a multiple linear regression and a multivariate analysis. Additionally, a new factorial model was proposed to predict AA requirements in broiler chickens. Data from 13 trials that evaluated the requirements for lysine (Lys), sulphur AA (SAA), threonine (Thr), and valine (Val) in male broilers were used for the analyses. Both methods of analysis were consistent in showing that the efficiency of utilization in feather-free body and feather were different. Using multiple linear regression, the values of efficiency of utilization estimated in feather-free body were 0.68, 0.72, 0.81, 0.79 (mg of amino acid deposited / mg of amino acid consumed above maintenance) and in feather were 0.58, 0.77, 0.78, and 1.57 (mg/mg) for Lys, SAA, Thr, and Val, respectively. Applying the multivariate approach, the corresponding predicted values were 0.68, 0.67, 4.23, 0.27 (mg/mg) in feather-free body and 1.16, 0.86, 0.16, and 1.10 (mg/mg) in feather, respectively. According to the results, efficiency of utilization may be related, to some extent, on the concentration determined in each tissue. The uncertainty around the amount of AA consumed for gain directed to feather-free body or feather deposition could be a limitation for multivariate analyses. The results indicated that multiple linear regression predictions may be better estimates of utilization efficiency. However, more studies are needed to elucidate the effect of age on deposition and partitioning of dietary AA in different parts of the broiler.

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<![CDATA[Towards a molecular basis of ubiquitin signaling: A dual-scale simulation study of ubiquitin dimers]]> https://www.researchpad.co/article/5bf86f60d5eed0c48405a957

Covalent modification of proteins by ubiquitin or ubiquitin chains is one of the most prevalent post-translational modifications in eukaryotes. Different types of ubiquitin chains are assumed to selectively signal respectively modified proteins for different fates. In support of this hypothesis, structural studies have shown that the eight possible ubiquitin dimers adopt different conformations. However, at least in some cases, these structures cannot sufficiently explain the molecular basis of the selective signaling mechanisms. This indicates that the available structures represent only a few distinct conformations within the entire conformational space adopted by a ubiquitin dimer. Here, molecular simulations on different levels of resolution can complement the structural information. We have combined exhaustive coarse grained and atomistic simulations of all eight possible ubiquitin dimers with a suitable dimensionality reduction technique and a new method to characterize protein-protein interfaces and the conformational landscape of protein conjugates. We found that ubiquitin dimers exhibit characteristic linkage type-dependent properties in solution, such as interface stability and the character of contacts between the subunits, which can be directly correlated with experimentally observed linkage-specific properties.

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<![CDATA[Conformational sampling of CpxA: Connecting HAMP motions to the histidine kinase function]]> https://www.researchpad.co/article/5c2400b7d5eed0c484098be2

In the histidine kinase family, the HAMP and DHp domains are considered to play an important role into the transmission of signal arising from environmental conditions to the auto-phosphorylation site and to the binding site of response regulator. Several conformational motions inside HAMP have been proposed to transmit this signal: (i) the gearbox model, (ii) α helices rotations, pistons and scissoring, (iii) transition between ordered and disordered states. In the present work, we explore by temperature-accelerated molecular dynamics (TAMD), an enhanced sampling technique, the conformational space of the cytoplasmic region of histidine kinase CpxA. Several HAMP motions, corresponding to α helices rotations, pistoning and scissoring have been detected and correlated to the segmental motions of HAMP and DHp domains of CpxA.

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<![CDATA[Selection of appropriate reference genes for RT-qPCR analysis in Propylea japonica (Coleoptera: Coccinellidae)]]> https://www.researchpad.co/article/5c06f054d5eed0c484c6d705

Reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) is a reliable technique commonly used in molecular biology to analyze RNA expression. The selection of suitable reference genes for data normalization is a precondition for credible measurements of gene expression levels using RT-qPCR. Propylea japonica is one of the most common pests of many crop systems throughout East Asia, and has often been used in the testing of non-target impacts during environmental risk assessments of genetically engineered plants. The present study assessed the suitability of nine frequently used reference genes for comparisons of P. japonica gene expression. Expression stability was compared across developmental stages, sex, a range of tissues, and following exposure to different temperatures. Data were analyzed using RefFinder, which integrated the results obtained using NormFinder, geNorm, BestKeeper, and the ΔCt method. This led to the identification of unique sets of reference genes for each experimental condition: ribosomal protein S18 (RPS18) and elongation factor 1 α (EF1A) for developmental stage comparisons, RPS18 and EF1A for sex comparisons, EF1A and ribosomal protein L4 for tissue comparisons, and RPS18 and EF1A for analyses of temperature-mediated effects. These reference genes will help to enhance the accuracy of RT-qPCR analyses of P. japonica gene expression. This work represents an initial move towards building a standardized system for RT-qPCR analysis of P. japonica, providing a basis for the ecological risk assessment of RNAi-based insect control products.

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<![CDATA[Biophysical and Pharmacological Characterization of Nav1.9 Voltage Dependent Sodium Channels Stably Expressed in HEK-293 Cells]]> https://www.researchpad.co/article/5989da16ab0ee8fa60b7b622

The voltage dependent sodium channel Nav1.9, is expressed preferentially in peripheral sensory neurons and has been linked to human genetic pain disorders, which makes it target of interest for the development of new pain therapeutics. However, characterization of Nav1.9 pharmacology has been limited due in part to the historical difficulty of functionally expressing recombinant channels. Here we report the successful generation and characterization of human, mouse and rat Nav1.9 stably expressed in human HEK-293 cells. These cells exhibit slowly activating and inactivating inward sodium channel currents that have characteristics of native Nav1.9. Optimal functional expression was achieved by coexpression of Nav1.9 with β1/β2 subunits. While recombinantly expressed Nav1.9 was found to be sensitive to sodium channel inhibitors TC-N 1752 and tetracaine, potency was up to 100-fold less than reported for other Nav channel subtypes despite evidence to support an interaction with the canonical local anesthetic (LA) binding region on Domain 4 S6. Nav1.9 Domain 2 S6 pore domain contains a unique lysine residue (K799) which is predicted to be spatially near the local anesthetic interaction site. Mutation of this residue to the consensus asparagine (K799N) resulted in an increase in potency for tetracaine, but a decrease for TC-N 1752, suggesting that this residue can influence interaction of inhibitors with the Nav1.9 pore. In summary, we have shown that stable functional expression of Nav1.9 in the widely used HEK-293 cells is possible, which opens up opportunities to better understand channel properties and may potentially aid identification of novel Nav1.9 based pharmacotherapies.

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<![CDATA[Controlling caspase activity in life and death]]> https://www.researchpad.co/article/5989db53ab0ee8fa60bdcee4 ]]> <![CDATA[68Ga and 188Re Starch-Based Microparticles as Theranostic Tool for the Hepatocellular Carcinoma: Radiolabeling and Preliminary In Vivo Rat Studies]]> https://www.researchpad.co/article/5989da35ab0ee8fa60b86084

Purpose

This work aims to develop, validate and optimize the radiolabeling of Starch-Based Microparticles (SBMP) by 188Re and 68Ga in the form of ready-to-use radiolabeling kits, the ultimate goal being to obtain a unique theranostic vector for the treatment of Hepatocellular Carcinoma.

Methods

Optimal labeling conditions and composition of freeze-dried kits were defined by monitoring the radiochemical purity while varying several parameters. In vitro stability studies were carried out, as well as an in vivo biodistribution as a preliminary approach with the intra-arterial injection of 68Ga radiolabeled SBMP into the hepatic artery of DENA-induced rats followed by PET/CT imaging.

Results

Kits were optimized for 188Re and 68Ga with high and stable radiochemical purity (>95% and >98% respectively). The in vivo preliminary study was successful with more than 95% of activity found in the liver and mostly in the tumorous part.

Conclusion

SBMP are a promising theranostic agent for the Selective Internal Radiation Therapy of Hepatocellular carcinoma.

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<![CDATA[Improved Survival and Initiation of Differentiation of Human Induced Pluripotent Stem Cells to Hepatocyte-Like Cells upon Culture in William’s E Medium followed by Hepatocyte Differentiation Inducer Treatment]]> https://www.researchpad.co/article/5989daefab0ee8fa60bc085c

Background

Hepatocyte differentiation inducer (HDI) lacks both glucose and arginine, but is supplemented with galactose and ornithine, and is added together with other reagents such as apoptosis inhibitor and oncostatin M. Although human induced pluripotent stem (iPS) cells initiate hepatocyte differentiation, most die within 7 days. In this study, we investigated both HDI and conventional media for their potential to improve cell survival.

Materials and Methods

201B7 iPS cells were cultured in conventional media. This consisted of three cycles of 5-day culture in William’s E (WE) medium, followed by a 2-day culture in HDI.

Results

Expression levels of α-feto protein (AFP) were higher in cells cultured in WE and in Dulbecco’s Modified Eagle’s Medium/Nutrient F-12 Ham (DF12). 201B7 cells expressed the highest AFP and albumin (ALB) when cultured in HDI for 2 days following 7-day culture in WE. After three cycles of 5-day culture in WE followed by 2 days in HDI, 201B7 cells expressed AFP and ALB 54 ± 2.3 (average ± standard deviation) and 73 ± 15.1 times higher, respectively, than those cultured in ReproFF (feeder-free condition).

Conclusion

201B7 cells survived culture in WE for 7 days followed HDI for 2 days. After three cycles of culture under these conditions, hepatocyte differentiation was enhanced, as evidenced by increased AFP and ALB expression.

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