Cell Membrane Components Research Articles

Regulatory Roles of Phospholipase A2 Enzymes and Bioactive Lipids in Mast Cell Biology.

Lipids play fundamental roles in life as an essential component of cell membranes, as a major source of energy, as a body surface barrier, and as signaling molecules that transmit intracellular and intercellular signals. Lipid mediators, a group of bioactive lipids that mediates intercellular signals, are produced via specific biosynthetic enzymes and transmit signals via specific receptors. Mast cells, a tissue-resident immune cell population, produce several lipid mediators that contribute to exacerbation or amelioration of allergic responses and also non-allergic inflammation, host defense, cancer and fibrosis by controlling the functions of microenvironmental cells as well as mast cell themselves in paracrine and autocrine fashions. Additionally, several bioactive lipids produced by stromal cells regulate the differentiation, maturation and activation of neighboring mast cells. Many of the bioactive lipids are stored in membrane phospholipids as precursor forms and released spatiotemporally by phospholipase A2 (PLA2) enzymes. Through a series of studies employing gene targeting and lipidomics, several enzymes belonging to the PLA2 superfamily have been demonstrated to participate in mast cell-related diseases by mobilizing unique bioactive lipids in multiple ways. In this review, we provide an overview of our current understanding of the regulatory roles of several PLA2-driven lipid pathways in mast cell biology.

Effects of Non-Opioid Analgesics on the Cell Membrane of Skin and Gastrointestinal Cancers.

Skin and gastrointestinal cancer cells are the target of research by many scientists due to the increasing morbidity and mortality rates around the world. New indications for drugs used in various conditions are being discovered. Non-opioid analgesics are worth noting as very popular, widely available, relatively cheap medications. They also have the ability to modulate the membrane components of tumor cells. The aim of this review is to analyze the impact of diclofenac, ibuprofen, naproxen, acetylsalicylic acid and paracetamol on skin and gastrointestinal cancers cell membrane. These drugs may affect the membrane through topical application, at the in vitro and in vivo level after oral or parenteral administration. They can lead to up- or downregulated expression of receptors, transporters and other molecules associated with plasma membrane. Medications may also alter the lipid bilayer composition of membrane, resulting in changes in its integrity and fluidity. Described modulations can cause the visualization of cancer cells, enhanced response of the immune system and the initiation of cell death. The outcome of this is inhibition of progression or reduction of tumor mass and supports chemotherapy. In conclusion, non-opioid analgesics may be used in the future as adjunctive therapy for the treatment of these cancers.

Evaluation of quinoxaline derivatives as potential ergosterol biosynthesis inhibitors: design, synthesis, ADMET, molecular docking studies, and antifungal activities

Because of the increased infection incidence and the advent of medication resistance, fungi have become a severe medical problem. Ergosterol is a structural component of the fungal cell membrane, and its synthetases (14α-demethylase (CYP51)) are thought to be the key to inhibiting ergosterol synthesis. Based on the active site (CYP51) analysis, we created a series of quinoxaline derivatives in this study. Following that, these target compounds were produced and tested for antifungal efficacy. The majority of the compounds show significant antifungal activity against a variety of Candida species. Compounds 9, 11, 17, 20, and 21 have antifungal properties that are similar to the positive control medicines and have considerable inhibitory effects, with MIC50 values ranging from 0.78 to 3.12 μg/mL. Compound 9 has the strongest antifungal activity against all Candida species, outperforming Ketoconazole and Fluconazole, and was thought to be the series' most active compound. As a result, the action mechanism for the most powerful blows was investigated further. They can alter ergosterol synthesis by blocking the activity of the 14α-demethylase (CYP51) target, according to preliminary mechanisms. According to this study, compounds 9, 11, 17, 20, and 21 have an efficient effect on the ergosterol production pathway. Moreover, the cytotoxicity of the most promising compounds was investigated, revealing the importance of 9, 11, 17, 20, and 21 hits as prospective anticandidal therapeutics. Furthermore, the ADMET characteristics, as well as the created binding models, will be valuable for further lead optimization.

Integrated bioinformatics analysis reveals marker genes and immune infiltration for pulmonary arterial hypertension

Pulmonary arterial hypertension (PAH) is a chronic cardiopulmonary syndrome with high pulmonary vascular load and eventually causing RV heart failure even death. However, the mechanism of pulmonary hypertension remains unclear. The purpose of this research is to detect the underlying key genes and potential mechanism of PAH using several bioinformatic methods. The microarrays GSE22356, GSE131793 and GSE168905 were acquired from the GEO. Subsequently, a host of bioinformatics techniques such as DAVID, STRING, R language and Cytoscape were utilized to investigate DEGs between PAH and healthy controls and conduct GO annotation, KEGG enrichment analysis and PPI network construction etc. Additionally, we predicted the transcription factors regulating DEGs through iRegulon plugin of Cytoscape and CIBERSORT was used to conduct immune infiltration analysis. One thousand two hundred and seventy-seven DEGs (403 up-regulated and 874 down-regulated) were identified from peripheral blood samples of 32 PAH patients and 29 controls, among which SLC4A1, AHSP, ALAS2, CA1, HBD, SNCA, HBM, SELENBP1, SERPINE1 and ITGA2B were detected as hub genes. The functional enrichment changes of DEGs were mainly enriched in protein binding, extracellular exosome, extracellular space, extracellular region and integral component of plasma membrane. The hub genes are chiefly enriched at extracellular exosome, hemoglobin complex, blood microparticle, oxygen transporter activity. Among TF-DEGs network, 42 target DEGs and 6 TFs were predicted with an NES > 4 (TEAD4, TGIF2LY, GATA5, GATA1, GATA2, FOS). Immune infiltration analysis showed that monocytes occupied the largest proportion of immune cells. The trend analysis results of infiltration immune cells illustrated that PAH patients had higher infiltration of NK cell activation, monocyte, T cell CD4 memory activation, and mast cell than healthy controls and lower infiltration of T cell CD4 naive. We detected SLC4A1, AHSP, ALAS2, CA1, HBD, SNCA, HBM, SELENBP1, SERPINE1 and ITGA2B as the most significant markers of PAH. The PAH patients had higher infiltration of NK cell activation, monocyte, T cell CD4 memory activation, and mast cell than healthy controls and lower infiltration of T cell CD4 naive. These identified genes and these immune cells probably have precise regulatory relationships in the development of PAH.

RNA-Seq Analysis of Magnaporthe grisea Transcriptome Reveals the High Potential of ZnO Nanoparticles as a Nanofungicide.

Magnaporthe grisea is one of the most destructive pathogen that encounters a challenge to rice production around the worldwide. The unique properties of ZnO nanoparticles (NPs), have high attractiveness as nanofungicide. In the present study, the response of fungi to ZnO NPs was evaluated using RNA sequencing (RNA-seq). Two different aligners (STAR and Hisat2) were used for aligning the clean reads, and the DEseq2 package was used to identify the differentially expressed genes (DEGs). In total, 1,438 and 761 fungal genes were significantly up- and down-regulated in response to ZnO NPs, respectively. The DEGs were subjected to functional enrichment analysis to identify significantly enriched biological pathways. Functional enrichment analysis revealed that “cell membrane components,” “ion (calcium) transmembrane transporter activity,” “steroid biosynthesis pathway” and “catalytic activity” were the contributed terms to fungal response mechanisms. The genes involved in aflatoxin efflux pumps and ribosome maturation were among the genes showing significant up- and down-regulation after ZnO NPs application. To confirm the obtained RNA-seq results, the expression of six randomly selected genes were evaluated using q-RT-PCR. Overall, the RNA-seq results suggest that ZnO NPs primarily act on the fungal cell membrane, but accumulation of ROS inside the cell induces oxidative stress, the fungal catalytic system is disrupted, resulting into the inhibition of ROS scavenging and eventually, to the death of fungal cells. Our findings provide novel insights into the effect of ZnO NPs as a promising nanofungicide for effective control of rice blast disease.

Functions and applications of glycolipid-hydrolyzing microbial glycosidases.

Glycolipids are important components of cell membranes in several organisms. The major glycolipids in mammals are glycosphingolipids (GSLs), which are composed of ceramides. In mammals, GSLs are degraded stepwise from the non-reducing end of the oligosaccharides via exo-type glycosidases. However, endoglycoceramidase (EGCase), an endo-type glycosidase found in actinomycetes, is a unique enzyme that directly acts on the glycosidic linkage between oligosaccharides and ceramides to generate intact oligosaccharides and ceramides. Three molecular species of EGCase, namely EGCase I, EGCase II, and endogalactosylceramidase, have been identified based on their substrate specificity. EGCrP1 and EGCrP2, which are homologs of EGCase in pathogenic fungi, were identified as the first fungal glucosylceramide- and sterylglucoside-hydrolyzing glycosidases, respectively. These enzymes are promising targets for antifungal drugs against pathogenic fungi. This review describes the functions and properties of these microbial glycolipid-degrading enzymes, the molecular basis of their differential substrate specificity, and their applications.

Novel Plasma Metabolomic Markers Associated with Diabetes Progression in Older Puerto Ricans.

We assessed longitudinal associations between plasma metabolites, their network-derived clusters, and type 2 diabetes (T2D) progression in Puerto Rican adults, a high-risk Hispanic subgroup with established health disparities. We used data from 1221 participants free of T2D and aged 40–75 years at baseline in the Boston Puerto Rican Health and San Juan Overweight Adult Longitudinal Studies. We used multivariable Poisson regression models to examine associations between baseline concentrations of metabolites and incident T2D and prediabetes. Cohort-specific estimates were combined using inverse-variance weighted fixed-effects meta-analyses. A cluster of 13 metabolites of branched chain amino acids (BCAA), and aromatic amino acid metabolism (pooled IRR = 1.87, 95% CI: 1.28; 2.73), and a cell membrane component metabolite cluster (pooled IRR = 1.54, 95% CI: 1.04; 2.27) were associated with a higher risk of incident T2D. When the metabolites were tested individually, in combined analysis, 5 metabolites involved in BCAA metabolism were associated with incident T2D. These findings highlight potential prognostic biomarkers to identify Puerto Rican adults who may be at high risk for diabetes. Future studies should examine whether diet and lifestyle can modify the associations between these metabolites and progression to T2D.

Screening of serum oxysterol biomarkers for colon cancer by liquid chromatography-tandem mass spectrometry

结肠癌(CC)是全球常见恶性肿瘤之一,发病率呈逐年上升趋势,目前没有有效的标志物用于疾病早期诊断和干预跟踪。胆固醇及其氧化衍生物氧固醇在众多恶性肿瘤发生发展中发挥关键作用。该研究采用液相色谱-串联质谱(LC-MS/MS)技术,对CC临床血清样本中胆固醇及相关10种氧固醇代谢物进行了定性定量分析,并采用偏最小二乘判别分析(PLS-DA)和正交偏最小二乘判别分析(OPLS-DA)进行多元统计分析,发现上述目标代谢物能够较好地区分CC组与健康对照组。为防止数据过拟合,该研究在PLS-DA模型各代谢物变量投影重要性(VIP)基础上,结合最优组分数及K-均值聚类结果,筛选得到3种代谢标志物。通过受试者操作特征曲线(ROC)的曲线下面积(AUC)分析,发现筛选得到的3种潜在标志物联合预测CC达到0.998,说明模型性能优良。GO(基因本体论)富集分析显示3种潜在标志物主要分布在内质网和包被囊泡上,参与胆固醇代谢、运输、低密度脂蛋白重塑等生物进程,发挥胆固醇运输活性和低密度脂蛋白颗粒受体结合的分子功能。KEGG(京都基因与基因组百科全书)通路分析显示3种潜在标志物富集于类固醇生物合成、PPAR(过氧化物酶体增殖物激活受体)信号通路及ABC(ATP结合盒)转运等通路上。该研究为寻找CC标志物及进一步阐明胆固醇及氧固醇在CC发病过程中的作用奠定了一定的基础。

25-Hydroxycholesterol as a Signaling Molecule of the Nervous System.

Cholesterol is an essential component of plasma membrane and precursor of biological active compounds, including hydroxycholesterols (HCs). HCs regulate cellular homeostasis of cholesterol; they can pass across the membrane and vascular barriers and act distantly as para- and endocrine agents. A small amount of 25-hydroxycholesterol (25-HC) is produced in the endoplasmic reticulum of most cells, where it serves as a potent regulator of the synthesis, intracellular transport, and storage of cholesterol. Production of 25-HC is strongly increased in the macrophages, dendrite cells, and microglia at the inflammatory response. The synthesis of 25-HC can be also upregulated in some neurological disorders, such as Alzheimer’s disease, amyotrophic lateral sclerosis, spastic paraplegia type 5, and X-linked adrenoleukodystrophy. However, it is unclear whether 25-HC aggravates these pathologies or has the protective properties. The molecular targets for 25-HC are transcriptional factors (LX receptors, SREBP2, ROR), G protein-coupled receptor (GPR183), ion channels (NMDA receptors, SLO1), adhesive molecules (α5β1 and ανβ3 integrins), and oxysterol-binding proteins. The diversity of 25-HC-binding proteins points to the ability of HC to affect many physiological and pathological processes. In this review, we focused on the regulation of 25-HC production and its universal role in the control of cellular cholesterol homeostasis, as well as the effects of 25-HC as a signaling molecule mediating the influence of inflammation on the processes in the neuromuscular system and brain. Based on the evidence collected, it can be suggested that 25-HC prevents accumulation of cellular cholesterol and serves as a potent modulator of neuroinflammation, synaptic transmission, and myelinization. An increased production of 25-HC in response to a various type of damage can have a protective role and reduce neuronal loss. At the same time, an excess of 25-HC may exert the neurotoxic effects.

Lipoprotein capture ELISA method for the sensitive detection of amphiphilic biomarkers

Enzyme-linked immunosorbent assays (ELISAs) are widely employed for the detection of protein targets due to their ease of use, sensitivity, and potential for high-throughput analyses. However, the use of ELISAs to detect non-protein targets such as lipids and amphiphiles is complicated by the physical properties of these molecules, which affects their association with functional surfaces and recognition ligands. Here, we developed a unique lipoprotein capture ELISA in which the natural association between lipoproteins and amphiphilic molecules facilitates detection of the target biomarker in a physiologically relevant conformation. An assay to detect the glycolipid lipoarabinomannan (LAM), a cell membrane component and virulence factor associated with Mycobacterial infections, was developed as a proof of concept.

Gene Expression Profiles of the Aging Rat Hippocampus Imply Altered Immunoglobulin Dynamics.

Aging is a process that leads to the deterioration in physiological functioning of the brain. Prior research has proposed that hippocampal aging is accompanied by genetic alterations in neural, synaptic, and immune functions. Nevertheless, interactome-based interrogations of gene alterations in hippocampal aging, remain scarce. Our study integrated gene expression profiles of the hippocampus from young and aged rats and functionally classified network-mapped genes based on their interactome. Hippocampal differentially expressed genes (DEGs) between young (5–8 months) and aged (21–26 months) male rats (Rattus norvegicus) were retrieved from five publicly available datasets (GSE14505, GSE20219, GSE14723, GSE14724, and GSE14725; 38 young and 29 aged samples). Encoded hippocampal proteins of age-related DEGs and their interactome were predicted. Clustered network DEGs were identified and the highest-ranked was functionally annotated. A single cluster of 19 age-related hippocampal DEGs was revealed, which was linked with immune response (biological process, P = 1.71E-17), immunoglobulin G binding (molecular function, P = 1.92E-08), and intrinsic component of plasma membrane (cellular component, P = 1.25E-06). Our findings revealed dysregulated hippocampal immunoglobulin dynamics in the aging rat brain. Whether a consequence of neurovascular perturbations and dysregulated blood-brain barrier permeability, the role of hippocampal immunoregulation in the pathobiology of aging warrants further investigation.

Pathogenetic and clinical significance of the gut-liver microbiota axis

Today, nonalcoholic fatty liver disease (NAFLD) and alcoholic liver disease (ALD) account for more than 50% of chronic liver diseases and cirrhosis in developed countries. The clinical significance of these pathologies lies in the formation of liver fibrosis and, therefore, in the increased risk of liver cirrhosis and hepatocellular carcinoma, which are life-threatening conditions. NAFLD is based on obesity and insulin resistance, whereas alcohol consumption is central to the etiopathogenesis of ALD. Recently, the role of the gut-liver microbiota axis in the genesis of NAFLD and ALD has also received increasing attention. Changes in qualitative and quantitative composition of intestinal microbiota and alterations of barrier function of intestinal mucosa can lead to entry of intraluminal antigens through portal vein system into the liver, inducing damage to hepatocytes and activation of proinflammatory processes. It is shown that in patients with CLD (NAFLD and ALD) there is an increase in the permeability of the intestinal mucosa and dysbiotic changes of the intestinal microbiome. The role of the gut-liver microbiota axis is well described in the genesis of NAFLD and ALD. Chronic alcohol consumption leads to increased colonies of Gram-negative bacteria in the intestinal mucosa and, therefore, to the accumulation of endotoxins (lipopolysaccharide components of bacterial cell membranes). Acetaldehyde produced by ADH of the intestinal epithelium stimulates tyrosine phosphorylation of tight cell contacts, increasing intestinal mucosal permeability, which leads to translocation of endotoxins into the portal bloodstream. Obesity is a major risk factor for NAFLD, which is also associated with dysbiotic changes in the intestinal microbiome. Obese individuals have increased Firmicutes, decreased Bacteroidetes, mediated reduction of short-chain fatty acid synthesis and increased intestinal wall permeability due to disruption of intercellular dense contacts, which leads to increased translocation of bacteria and endotoxins into the systemic bloodstream.

Green synthesis of core - Shell Te–Se bimetallic nanoparticles using Cinnamomum Camphora leaf extract and their in-vitro cholesterol degradation

Cholesterol is a ceraceous, fat-like organic molecule found in organisms and plasma lipoprotein. Its anabolism takes place from all animal germs, and essential structural component of the animal cell membrane. It is known that the accumulation of too much low-density cholesterol leads to the development of plaque inside the arteries known as atherosclerosis which leads to cerebrovascular, peripheral, and coronary vascular diseases. To prevent this, a wide assortment of pathogens could be helpful for cholesterol degradation. In this study, we have synthesized Te–Se BNps (Tellurium–Selenium bimetallic nanoparticles) from Cinnamomum Camphora leaf extract and tried to degrade low-density cholesterol. The biosynthesized Te–Se BNps have been characterized by DRS UV visible spectroscopy, XRD, HR-TEM, AFM, FT-IR, SEM and EDX spectroscopy. In addition, qualitative and quantitative analysis has been carried out by various techniques like UV spectrophotometer, thin layer chromatography, and HPLC studies. The estimation of cholesterol degradation showed that the percentage of degradation was 35% in acidic, basic as well as neutral medium. The mechanism of cholesterol degradation is also explained. The result revealed that camphor-mediated Te–Se BNps have great potential for cholesterol degradation; it can be used as a source for cholesterol oxidases products. This is the first report on cholesterol oxidation by Cinnamomum Camphora leaf extract mediated Te–Se BNps using the photochemical method.

Potent Virustatic Polymer-Lipid Nanomimics Block Viral Entry and Inhibit Malaria Parasites In Vivo.

Infectious diseases continue to pose a substantial burden on global populations, requiring innovative broad-spectrum prophylactic and treatment alternatives. Here, we have designed modular synthetic polymer nanoparticles that mimic functional components of host cell membranes, yielding multivalent nanomimics that act by directly binding to varied pathogens. Nanomimic blood circulation time was prolonged by reformulating polymer–lipid hybrids. Femtomolar concentrations of the polymer nanomimics were sufficient to inhibit herpes simplex virus type 2 (HSV-2) entry into epithelial cells, while higher doses were needed against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Given their observed virustatic mode of action, the nanomimics were also tested with malaria parasite blood-stage merozoites, which lose their invasive capacity after a few minutes. Efficient inhibition of merozoite invasion of red blood cells was demonstrated both in vitro and in vivo using a preclinical rodent malaria model. We envision these nanomimics forming an adaptable platform for developing pathogen entry inhibitors and as immunomodulators, wherein nanomimic-inhibited pathogens can be secondarily targeted to sites of immune recognition.

Phospholipid and menthol based nanovesicle impregnated transdermal patch for nutraceutical delivery to diminish folate and iron deficiency

Adequate micronutrient availability is particularly important in women, children and infants. Micronutrient deficiencies are the major cause of maternal and neonatal morbidity. To overcome this, WHO recommends the use of folic acid and iron supplements for reducing anaemia and improving the health of the mother and infants. Oral intake of supplements for nutritional deficiencies are associated with gastric irritation, nausea, constipation and non-patient compliance due to associated taste. In case of absorption deficiency nutrients administered orally pass-through digestive tract unabsorbed. In the present study, we propose transdermal delivery of nutraceuticals to avoid the limitations associated with oral intake. Transdermal delivery has limited use because of the closely packed barrier of the stratum corneum that limits the permeability of molecules across skin. Here, we have used biomimetic nanovesicles impregnated in transdermal patches for delivery of folic acid and iron. Nanovesicles are prepared using an abundant component of cell membrane, phosphatidyl choline and a permeation enhancer. Further these nanovesicles are impregnated onto polyacrylate based transdermal patch. In vitro studies have shown the ability of nanovesicles to fluidise skin lipids and penetrate into deeper skin. In vivo application of transdermal patches gradually increased the systemic concentration of nutraceuticals. Post application of the patch, five-fold increase in plasma folic acid concentration and 1.5-fold increase in plasma iron concertation was achieved in 6 h. Developed nanovesicles were compatible with keratinocytes and fibroblasts as tested in vitro and have the potential to enhance the cellular uptake of molecules. Skin irritation studies on human volunteers have confirmed the safety of nutraceutical loaded nanovesicles. Thus, the developed nutraceutical loaded transdermal patches provide a potential, easy to use platform for micronutrient delivery in infants and mothers.

Cytotoxic activity of Nep1-like proteins on monocots.

Summary Necrosis‐ and ethylene‐inducing peptide 1 (Nep1)‐like proteins (NLPs) are found throughout several plant‐associated microbial taxa and are typically considered to possess cytolytic activity exclusively on dicot plant species. However, cytolytic NLPs are also produced by pathogens of monocot plants such as the onion (Allium cepa) pathogen Botrytis squamosa.We determined the cytotoxic activity of B. squamosa BsNep1, as well as other previously characterized NLPs, on various monocot plant species and assessed the plant plasma membrane components required for NLP sensitivity.Leaf infiltration of NLPs showed that onion cultivars are differentially sensitive to NLPs, and analysis of their sphingolipid content revealed that the GIPC series A : series B ratio did not correlate to NLP sensitivity. A tri‐hybrid population derived from a cross between onion and two wild relatives showed variation in NLP sensitivity within the population. We identified a quantitative trait locus (QTL) for NLP insensitivity that colocalized with a previously identified QTL for B. squamosa resistance and the segregating trait of NLP insensitivity correlated with the sphingolipid content.Our results demonstrate the cytotoxic activity of NLPs on several monocot plant species and legitimize their presence in monocot‐specific plant pathogens.

Membrane phosphoinositides and renal epithelial cell polarity determination in the Xenopus pronephros in vivo

BackgroundThough they are only minor components of cell membranes, phosphoinositide lipids (PIs) participate in numerous signaling processes and in membrane identity determination. Many studies of cultured MDCK renal epithelial cells found that PIs are distributed with polarity among plasma membrane (PM) domains, and that their polarized distributions are required for the delivery of distinct populations of apical and basolateral membrane proteins. The extent to which PIs drive these processes in actual renal epithelial cells in vivo has never been examined. We investigate the distribution of PIs in vivo in the pronephros of Xenopus laevis tadpoles using MCherry‐tagged Pleckstrin Homology (PH) domains that selectively bind different PIs (PH‐AKT, PH‐PLCD1, which bind to (PI(3,4,5,)P3 and PI(4,5)P2 respectively) with the goal of assessing whether and how PI localizations affect cell polarity determination and the trafficking of proteins to their sites of ultimate functional residence in renal epithelial cells in situ.MethodsmRNA encoding MCherry‐PH‐AKT or MCherry‐PH‐PLCD1 was injected into fertilized oocytes and their distributions were assessed in the developed pronephros at stage NF45 via fluorescence microscopy. Knockdown (KD) of PTEN, a lipid 3’ phosphatase that regulates membrane PI composition, was achieved via injection of targeted morpholinos and confirmed by western blotting. The effects of PTEN KD on MCherry‐PH‐AKT distribution were assessed by fluorescence microscopy.ResultsIn MDCK cells PH‐AKT and PH‐PLCD1 localize to the basolateral and apical PMs, respectively. Their distributions are quite different in the pronephros, with both sensors localizing apically in the proximal portion of the tubule and basolaterally in the distal part. Interestingly, PTEN localization, at least in proximal tubules, is apical and not different than previously reported in cell models. Finally, PH‐AKT staining dramatically re‐distributes to the lateral domain of renal cells upon PTEN KD but this treatment does not alter the localization of protein markers of epithelial PM polarity.ConclusionsThese studies constitute the first effort to assess the role of PIs in establishing PM polarity in renal epithelial cells in vivo. Our data reveal important differences from observations previously made with in vitro systems. These findings highlight the need to explore the processes that produce renal epithelial cell polarity in vivo and in the context of intact renal tubules.

Snapshots of lipid synthesis and fat storage

During the past thirty years, the perceived role of lipids has shifted from simple structural components of cell membranes to bioactive molecules that regulate critical cellular and pathological processes. The enzymes that generate and breakdown these bioactive lipids have emerged as novel therapeutic targets for treating the leading causes of diseases in the United States, including cancer, heart disease, and diabetes. This talk will present new insight into how key enzymes in triglyceride and phospholipid metabolism work at the molecular and structural level.

Atorvastatin Modulates the Function and Trafficking of Dopamine Transporters

IntroductionDopaminergic signaling is involved in learning, memory, reward, movement, and emotion. The dopamine transporter (DAT) is the primary mechanism for terminating this signaling via DA uptake. A few lines of evidence indicate that DAT function is regulated by membrane cholesterol. Cholesterol is a critical component of plasma membranes and plays an essential role in regulating membrane fluidity, protein compartmentalization, and function. Statins are cholesterol‐lowering medications that have been linked to mild cognitive impairment. The goal of this project was to examine the effects of atorvastatin‐mediated reductions in cholesterol content on DAT function and trafficking.MethodsCultured cells & cholesterol measurement: HA‐tagged human DAT (hDAT) was stably expressed in neuroblastoma 2a (N2A) cells using G418 as selection. Cells were treated with atorvastatin (0.1, 0.3, 1.0, or 10 μM) and membrane cholesterol content was measured using Amplex Red.[3H]DA Uptake: After 24hr atorvastatin or vehicle treatment, cells were incubated with 15 nM [3H]DA in the presence of various concentrations of non‐labeled DA and [3H]DA uptake was measured to determine Vmax and Km.Cocaine inhibition of DA uptake & cholesterol replenishment: Cells were treated for 24hr with atorvastatin (1 μM) or vehicle and incubated with 15 nM [3H]DA and increasing concentrations of cocaine (100 nM‐10 μM). To determine if there was a causal effect of reduced cholesterol content and cocaine inhibition of DA uptake, atorvastatin‐treated cells were replenished with water soluble cholesterol (~40 mg per gram solid; MβCD) prior to cocaine inhibition.Amphetamine‐induced, DAT‐mediated DA efflux: Cells treated for 24hrs with either atorvastatin (1 μM) or vehicle were preloaded with 30 nM of [3H]DA. Then, cells were incubated with amphetamine (1 or 10 μM). AMPH‐induced DA efflux was determined by measuring DA content in the medium across time.Surface DAT Biotinylation: Basal surface levels of DAT in atorvastatin and vehicle treated cells, as well as DA‐induced DAT internalization, were measured by surface biotinylation using membrane non‐permeable sulfo‐NHS‐SS biotin.ImmunocytochemistryWe measured levels of F‐actin and G‐actin to determine the effects of atorvastatin treatment on actin dynamics using immunocytochemistry.ResultsWe found that atorvastatin dose dependently reduced membrane cholesterol content. This reduction was accompanied by decreases in the maximal rate of DA uptake and by decreases in the potency of cocaine to inhibit [3H]DA uptake. In contrast, atorvastatin treatment did not significantly alter AMPH‐induced DA efflux. Importantly, the reduced ability of cocaine to inhibit DA uptake was reversed by replenishing cholesterol in atorvastatin‐treated cells, suggesting a direct causal effect of reduced cholesterol content and cocaine inhibition. Lastly, we found that atorvastatin disrupted actin dynamics and abolished DA‐induced DAT internalization. The present study provides evidence for the first time that atorvastatin dysregulates DAT function and trafficking. Furthermore, this dysregulation may be associated with the cognitive deficits observed in patients receiving atorvastatin treatment.

Absorption of the [bmim][Cl] Ionic Liquid in DMPC Lipid Bilayers across Their Gel, Ripple, and Fluid Phases.

Lipid bilayers are a key component of cell membranes and play a crucial role in life and in bio-nanotechnology. As a result, controlling their physicochemical properties holds the promise of effective therapeutic strategies. Ionic liquids (ILs)—a vast class of complex organic electrolytes—have shown a high degree of affinity with lipid bilayers and can be exploited in this context. However, the chemical physics of IL absorption and partitioning into lipid bilayers is yet to be fully understood. This work focuses on the absorption of the model IL [bmim][Cl] into 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) lipid bilayers across their gel, ripple, and fluid phases. Here, by small-angle neutron scattering, we show that (i) the IL cations are absorbed in the lipid bilayer in all its thermodynamic phases and (ii) the amount of IL inserted into the lipid phase increased with increasing temperature, changing from three to four IL cations per 10 lipids with increasing temperature from 10 °C in the gel phase to 40 °C in the liquid phase, respectively. An explicative hypothesis, based on the entropy gain coming from the IL hydration water, is presented to explain the observed temperature trend. The ability to control IL absorption with temperature can be used as a handle to tune the effect of ILs on biomembranes and can be exploited in bio-nanotechnological applications.