Membrane Level Research Articles

Familial Steep Corneas in Posterior Polymorphous Corneal Dystrophy 3 Due to a Novel ZEB1 Gene Mutation.

To present 4 family members with posterior polymorphous corneal dystrophy (PPCD), nonkeratoconic steep corneas, and myopia caused by a previously unknown genetic alteration in the ZEB1 gene. Ophthalmic examinations and corneal curvature analyses were performed for all patients. Whole-exome targeted gene panel sequencing was performed for 1 patient. Pathogenic variant confirmation and segregation of the variant were performed for all 4 patients using Sanger sequencing. A mother and her 3 children presented with bilateral diffuse corneal opacities, vesicular aggregates at the Descemet membrane level, and endothelial stippling, compatible with the diagnosis of PPCD. All patients had steeper than average keratometry readings with normal globe axial lengths and refractive moderate to extremely high myopia, ranging from -2 to -16.5 diopters (spherical equivalent). The younger patients had more severe findings, with the youngest, an 11-month-old girl, presenting with keratometry readings of 66.72 × 69.48 @ 132 degrees and 66.10 × 67.32 @ 34 degrees in the right and left eyes, respectively. The ZEB1:c.794-1G>A; chr10-31809053G>A; NM_001174096.2 mutation was detected in all patients. We describe a novel ZEB1 mutation associated with PPCD, nonkeratoconic steep corneas, and myopia. PPCD3 should be considered not only as an endothelial pathology but also as an ectatic disorder and should be ruled out in young children presenting with high myopia.

Curvature Memory in Electrically Stimulated Lipid Membranes.

We demonstrate, using non-equilibrium molecular dynamics simulations, that lipid membrane capacitance varies with surface charge accumulation linked to membrane shape and curvature changes. Specifically, we show that lipid membranes exhibit a hysteretic response when exposed to oscillatory electric fields. The electromechanical coupling in these membranes leads to hysteretic buckling, in which the membrane can spontaneously buckle in one of two distinct directions along the electric field, even for the same ionic charge accumulation at the water-membrane interface. In this regard, these binary buckled membrane states suggest potential applications in neuromorphic computing. Their bistable nature, characterized by two distinct and stable configurations, could serve as a foundation for implementing memory storage systems and logic operations. Furthermore, we introduce a circuit model that captures these dynamic effects, offering insights into emergent memory effects in electrically stimulated lipid membranes. Finally, this work presents lipid bilayers as dynamic, adaptable elements and suggests a new platform for exploring energy storage, information processing, and memory encoding at the lipid membrane level.

Synergistically effects of n-3 PUFA and B vitamins prevent diabetic cognitive dysfunction through promoting TET2-mediated active DNA demethylation.

Diabetic cognitive dysfunction (DCD) refers to the cognitive impairment observed in individuals with diabetes. Epidemiological studies have suggested that supplementation with n-3 polyunsaturated fatty acid (PUFA) or B vitamins may prevent the development of diabetic complications. Post hoc studies indicate a potential synergistic effect of n-3 PUFA and B vitamins in preventing cognitive impairment. However, the precise effect and underlying mechanism of this combination on DCD remain unclear. In case-control study, we compared fatty acid composition of erythrocyte membrane and serum homocysteine levels between diabetic individuals with and without DCD. We found that insufficient levels of n-3 PUFA, along with elevated serum homocysteine, significantly increase the risk of developing DCD. Treatment with a combination of fish oil, folate, and vitamin B12 improved cognitive impairment and aberrant neuronal morphology in streptozotocin-induced DCD mice. Folic acid and vitamin B12 enhanced the efficiency of exogenous docosahexaenoic acid (DHA) transportation to the brain by preventing the accumulation of homocysteine and S-adenosylhomocysteine, thereby inhibiting neuronal apoptosis in diabetic brains. Furthermore, folic acid and vitamin B12 supplementation can provide sufficient 5-methylcytosine for diabetic brains by promoting DNA methylation, while increased DHA levels maintain TET-mediated active DNA demethylation in diabetic brains through enhancing TET2 function. Overall, our study provides novel insights into molecular mechanisms underlying the synergistic preventive effects of the combined supplementation with fish oil, folic acid and vitamin B12 on DCD, suggests that combining n-3 PUFA and B vitamins could be a promising strategy for preventing DCD among individuals with diabetes.

The Cardiomyocyte in Cirrhosis: Pathogenic Mechanisms Underlying Cirrhotic Cardiomyopathy

Cirrhotic cardiomyopathy is defined as systolic and diastolic dysfunction in patients with cirrhosis, in the absence of any primary heart disease. These changes are mainly due to the malfunction or abnormalities of cardiomyocytes. Similar to non-cirrhotic heart failure, cardiomyocytes in cirrhotic cardiomyopathy demonstrate a variety of abnormalities: from the cell membrane to the cytosol and nucleus. At the cell membrane level, biophysical plasma membrane fluidity, and membrane-bound receptors such as the beta-adrenergic, muscarinic and cannabinoid receptors are abnormal either functionally or structurally. Other changes include ion channels such as L-type calcium channels, potassium channels, and sodium transporters. In the cytosol, calcium release and uptake processes are dysfunctional and the myofilaments such as myosin heavy chain and titin, are either functionally abnormal or have structural alterations. Like the fibrotic liver, the heart in cirrhosis also shows fibrotic changes such as a collagen isoform switch from more compliant collagen III to stiffer collagen I which also impacts diastolic function. Other abnormalities include the secondary messenger cyclic adenosine monophosphate, cyclic guanosine monophosphate, and their downstream effectors such as protein kinase A and G-proteins. Finally, other changes such as excessive apoptosis of cardiomyocytes also play a critical role in the pathogenesis of cirrhotic cardiomyopathy. The present review aims to summarize these changes and review their critical role in the pathogenesis of cirrhotic cardiomyopathy.

Neuronal growth regulator 1 (NEGR1) promotes the synaptic targeting of glutamic acid decarboxylase 65 (GAD65).

Neuronal growth regulator 1 (NEGR1) is a synaptic plasma membrane localized cell adhesion molecule implicated in a wide spectrum of psychiatric disorders. By RNAseq analysis of the transcriptomic changes in the brain of NEGR1-deficient mice, we found that NEGR1 deficiency affects the expression of the Gad2 gene. We show that glutamic acid decarboxylase 65 (GAD65), the Gad2 - encoded enzyme synthesizing the inhibitory neurotransmitter GABA on synaptic vesicles, accumulates non-synaptically in brains of NEGR1-deficient mice. The density of non-synaptic GAD65 accumulations is also increased in NEGR1 deficient cultured hypothalamic neurons, and this effect is rescued by re-expression of NEGR1. By using a novel biosensor of the plasma membrane attachment of GAD65, we demonstrate that GAD65 attaches to the plasma membrane. NEGR1 promotes palmitoylation-dependent clearance of GAD65 from the plasma membrane and targeting of GAD65 to plasma membrane-derived endocytic vesicles. In NEGR1 deficient cultured hypothalamic neurons, the synaptic and extrasynaptic levels of the plasma membrane attached GAD65 are increased, and the synaptic levels of GABA are reduced. NEGR1-deficient mice are characterized by reduced body weight, lower GABAergic synapse densities in the arcuate nucleus, and blunted responsiveness to the reinforcing effects of food rewards. Our results indicate that abnormalities in synaptic GABA synthesis can contribute to brain disorders associated with abnormal expression of NEGR1 in humans.

Unveiling Interactions between Self-Assembling Peptides and Neuronal Membranes.

The use of self-assembling peptide hydrogels in the treatment of spinal cord and brain injuries, especially when combined with adult neural stem cells, has shown great potential. To advance tissue engineering, it is essential to understand the effect of mechanochemical signaling on cellular differentiation. The elucidation of the molecular interactions at the level of the neuronal membrane still represents a promising area of investigation for many drug delivery and tissue engineering applications. An innovative molecular dynamics framework has been introduced to investigate the effect of SAP fibrils with different charges on neural membrane lipid domain dynamics. Such advance enables the in silico exploration of the biomimetic properties of SAP hydrogels and other polymeric biomaterials for tissue engineering applications.

Oxysterol sulfates in fluids, cells and tissues: how much do we know about their clinical significance, biological relevance and biophysical implications?

Oxysterol sulfates are emerging as key players in lipid homeostasis, inflammation and immunity. Despite this, knowledge on their basal levels in fluids, cells and tissues and any changes associated with age, gender and diet in health and disease; as well as their spatio-temporal distribution in cell membranes and organelles have been greatly hampered by the lack of commercially available pure synthetic standards. Expansion of the panel of pure oxysterol sulfates standards is pivotal to improve our understanding on the impact of oxysterol sulfates at the membrane level and their role in cellular events. While the clinical significance, biophysical implications and biological relevance of oxysterol sulfates in fluids, cells and tissues remains largely unknown, knowledge already gathered on the precursors of oxysterol sulfates (e.g. oxysterols and cholesterol sulfate) can be used to guide researchers on the most relevant aspects to search for when screening for oxysterol sulfates bioavailability in (patho)physiological conditions which are crucial in the design of biophysical and of cell-based assays. Herein, we provide a review on the brief knowledge involving oxysterol sulfate and an overview on the biophysical implications and biological relevance of oxysterols and cholesterol sulfate useful to redirect further investigations on the role of oxysterol sulfates in health and disease.

Circ_0000284 Is Involved in Arsenite-Induced Hepatic Insulin Resistance Through Blocking the Plasma Membrane Translocation of GLUT4 in Hepatocytes via IGF2BP2/PPAR-γ.

Arsenic exposure can induce liver insulin resistance (IR) and diabetes (DM), but the underlying mechanisms are not yet clear. Circular RNAs (circRNAs) are involved in the regulation of the onset of diabetes, especially in the progression of IR. This study aimed to investigate the role of circRNAs in arsenic-induced hepatic IR and its underlying mechanism. Male C57BL/6J mice were given drinking water containing sodium arsenite (0, 0.5, 5, or 50 ppm) for 12 months. The results show that sodium arsenite increased circ_0000284 expression, decreased insulin-like growth factor 2 mRNA binding protein 2 (IGF2BP2) and peroxisome proliferator-activated receptor-γ (PPAR-γ), and inhibited cell membrane protein levels of insulin-responsive glucose transporter protein 4 (GLUT4) in the mouse livers, indicating that arsenic exposure causes liver damage and disruptions to glucose metabolism. Furthermore, sodium arsenite reduced glucose consumption and glycogen levels, increased the expression of circ_0000284, reduced the protein levels of IGF2BP2 and PPAR-γ, and inhibited GLUT4 protein levels in the cell membranes of insulin-treated HepG2 cells. However, a circ_0000284 inhibitor reversed arsenic exposure-induced reductions in IGF2BP2, PPAR-γ, and GLUT4 levels in the plasma membrane. These results indicate that circ_0000284 is involved in arsenite-induced hepatic insulin resistance through blocking the plasma membrane translocation of GLUT4 in hepatocytes via IGF2BP2/PPAR-γ. This study provides a scientific basis for finding early biomarkers for the control of arsenic exposure and type 2 diabetes mellitus (T2DM), and discovering new prevention and control measures.

Meniere's disease: Structural considerations in early cochlea hydrops.

Structural features of the human cochlea may control early lesion formation in endolymphatic hydrops. This process may hinge on three structural features: the flattened spiral shape of the human cochlea, the toroidal configuration of the distended cochlea duct, and the distensibility characteristics of Reissner's membrane. An analytical method is presented to assess the variation in hydropic distention that may occur in the several turns of the cochlea due to these structural features. A normal human cochlea is used to illustrate the method of analysis. Structural dimensions were taken from a mid-modiolar section. Reissner's membrane was projected to assume a spiral toroid shape as it distends. Peak membrane stress proclivities in each cochlea turn were calculated analytically. Membrane strain was assessed from a collagen model of Reissner's membrane. Sagittal membrane displacements were quantified geometrically. Stress levels in Reissner's membrane were projected to be the lowest in the lower basal turn and to increase progressively to a peak value in the apex. Strain in Reissner's membrane in the apical turn was projected to be substantially higher than in the lower turns. Sagittal displacement of Reissner's membrane was projected to be most pronounced in the apical turn in all the stages of early cochlea hydrops. Structural features appear to underlie a differential susceptibility to hydrops in the human cochlea. The flattened spiral shape of the human cochlea coupled with the anticlastic configuration and the distensile characteristics of Reissner's membrane are projected to result in distinct histological stages as hydropic disease in the cochlea progresses.

YAP/TAZ Signalling Controls Epidermal Keratinocyte Fate.

The paralogues Yes-associated protein (YAP) and transcriptional coactivator with PDZ-binding motif (TAZ) control cell proliferation and cell fate determination from embryogenesis to ageing. In the skin epidermis, these proteins are involved in both homeostatic cell renewal and injury-induced regeneration and also drive carcinogenesis and other pathologies. YAP and TAZ are usually considered downstream of the Hippo pathway. However, they are the central integrating link for the signalling microenvironment since they are involved in the interplay with signalling cascades induced by growth factors, cytokines, and physical parameters of the extracellular matrix. In this review, we summarise the evidence on how YAP and TAZ are activated in epidermal keratinocytes; how YAP/TAZ-mediated signalling cooperates with other signalling molecules at the plasma membrane, cytoplasmic, and nuclear levels; and how YAP/TAZ ultimately controls transcription programmes, defining epidermal cell fate.

Nano-laminated clay-essential oil composite formulations: Key mechanistic antibacterial processes and in vitro antibiofilm activity

This study involves fabrication of nano-clay-essential oil hybrid nanostructures using acoustic cavitation-based probe ultrasonication for controlled release antibacterial effect. Both the pristine and composites were characterized and showed no structural (shape and size) changes in clay nanoparticles (NPs) after adsorption of essential oil as examined by Transmission electron microscope. Adsorption of essential oil in the interlayer space of nano-clay particles was confirmed by increase in d-spacing value for both nano-bentonite (3.31–3.44 nm) and nano-MMT (3.36–3.45 nm) composites. The FTIR (Fourier transform infrared) spectroscopy showed disappearance or weakening of bands at 3500-3600 cm−1 (hydroxyl group) wavenumber and appearance of new bands at 1700 cm−1 (C-H bending) suggesting interlayer adsorption of essential oil within the nano-clay. Thermogravimetric analysis (TGA) revealed three step mass loss in nano-clay-essential oil composites with the lowest mass loss (81 %) reported in Nano-bentonite-Basil essential oil composite. The GC-MS analysis confirmed the presence of similar components in both pure and composite formulations but with varied compositions. The developed nanocomposites (nano-clay-essential oil) exhibited enhanced antibacterial and anti-biofilm activity against Staphylococcus aureus and Escherichia coli as compared to pure EOs. This work elucidates the antimicrobial mechanism of nano-clay-essential oil composite as identified by determining cell membrane damaging effect and level of reactive oxygen species (ROS). The present study provides insights on the potential role of nano-clay as adsorbent/nanocarrier and nano-clay-essential oil formulation for effective drug delivery.

Detecting and repairing micro defects in perfluorinated ion exchange membranes for redox flow batteries

Ion exchange membranes play a vital role in redox flow batteries. However, polymer membranes with a microscopic thickness of approximately 20–50 μm are susceptible to micro defects, which substantially reduces the battery's energy efficiency and cycling stability. Hence, there is a need for an effective strategy to identify and resolve membrane imperfections, which is currently missing in the literature. In this work, a pressure-retention setup and hot-pressing method are proposed and show that defective membranes can be effectively identified and resolved. For instance, a membrane with around 100-μm pinholes exhibits a low coulombic efficiency of 77.5 % at the current density of 100 mA cm−2. However, the coulombic efficiency can be raised to 96.3 % by removing the defects, thus attaining the level of the undamaged pristine membrane (96.4 %). The capacity retention rate of the vanadium redox flow batteries with the repaired membrane is 71.1 % over 100 cycles at the current density of 200 mA cm−2, close to that of the pristine membrane (72.2 %). In addition, the repaired membrane exhibits quite similar physicochemical properties to the pristine membrane from various characterizations. The proposed method represents a convenient, economical, and non-destructive membrane detecting and repairing strategy, demonstrating great potential for redox flow batteries.

Role of Rab10 in cocaine-induced behavioral effects is associated with GABAB receptor membrane expression in the nucleus accumbens.

Previous studies have demonstrated that Ras-related GTP-binding protein Rab10 (Rab10) plays a role in psychostimulant-induced behavioral effects. In this study, we showed that Rab10 in the nucleus accumbens (NAc) of male animals affects the development of cocaine-induced behavioral effects, which are associated with the plasma membrane expression of the GABAB heteroreceptor (GABABR). We performed flow cytometry, immunoendocytosis, pHluorin activity analysis, electrophysiology analysis, and open-field testing to explore the role of Rab10 in modulating the membrane expression and function of GABABR and its regulatory effect on cocaine-induced behavioral effects. Transcriptomics analysis showed that Rab10 was elevated following acute cocaine treatment. Membrane levels of Rab10 increased within day 1 of the cocaine treatment, subsequently decreasing at later time points. Rab10 deficiency in NAc regions significantly increased cocaine-inhibited membrane GABABR levels and inhibited cocaine-induced hyperlocomotion and behavioral sensitization. In addition, GAD 67 + -expressing neurons from NAc regions treated with cocaine revealed a significant decrease in Rab10 membrane expression. Furthermore, NAc neuron-specific Rab10 knockout resulted in a significant increase in the cocaine-inhibited membrane expression of GABABR, along with increased miniature inhibitory postsynaptic current (mIPSC) amplitude and attenuation of baclofen-amplified Ca2+ influx. These results uncover a new mechanism in which Rab10-GABABR signaling may serve as a potential pathway for regulating cocaine-induced behavioral effects.

Staphylococcus aureus can use an alternative pathway to be internalized by osteoblasts in absence of β1 integrins

Staphylococcus aureus main internalization mechanism in osteoblasts relies on a tripartite interaction between bacterial fibronectin-binding proteins, extracellular matrix soluble fibronectin, and osteoblasts’ β1 integrins. Caveolins, and particularly caveolin-1, have been shown to limit the plasma membrane microdomain mobility, and consequently reduce the uptake of S. aureus in keratinocytes. In this study, we aimed to deepen our understanding of the molecular mechanisms underlying S. aureus internalization in osteoblasts. Mechanistically, S. aureus internalization requires endosomal recycling of β1 integrins as well as downstream effectors such as Src, Rac1, and PAK1. Surprisingly, in β1 integrin deficient osteoblasts, S. aureus internalization is restored when Caveolin-1 is absent and requires αvβ3/5 integrins as backup fibronectin receptors. Altogether, our data support that β1 integrins regulate the level of detergent-resistant membrane at the plasma membrane in a an endosomal and Caveolin-1 dependent manner.

Guanidinium-Functionalized Carbon Dots: An Efficient Antibacterial Agent against Multidrug-Resistant ESKAPE Pathogens.

The rise of multidrug-resistant (MDR) bacteria poses a substantial challenge in clinical settings, particularly with the increasing prevalence of ESKAPE pathogens (E. faecium, S. aureus, K. pneumoniae, A. baumannii, P. aeruginosa, and E. coli) as critical MDR bacteria. These ESKAPE pathogens have the capability to undermine antibiotic treatments, leading to a high incidence of drug resistance. However, the development of efficient antibacterial agents against ESKAPE pathogens is still in the bottleneck. Herein, the first example of antibacterial carbon dots against ESKAPE pathogens was reported. Onion powder-based carbon dots were melted with poly(hexamethylene biguanide) hydrochloride (PHMB) to obtain guanidinium-functionalized carbon dots (GCDs), which exhibited satisfactory antibacterial activity against all the tested bacteria, including both Gram-positive and Gram-negative bacteria, and even ESKAPE pathogens. The efficient antibacterial ability of GCDs derives from the rupture of the bacterial cell membrane and elevated ROS levels. Safety assessments revealed that GCDs neither trigger detectable drug resistance nor exhibit any cytotoxic effects. Furthermore, GCDs effectively promoted wound healing without observable adverse reactions of mixed MDR bacteria-infected wounds in rats. The GCDs also showed excellent long-term stability. These findings indicate that GCDs hold promise as an efficient antibacterial agent for the treatment of MDR strain-caused clinical infected-wound healing.

CD73 restrains mutant β-catenin oncogenic activity in endometrial carcinomas.

Missense mutations in exon 3 of CTNNB1, the gene encoding β-catenin, are associated with poor outcomes in endometrial carcinomas (EC). Clinically, CTNNB1 mutation status has been difficult to use as a predictive biomarker as β-catenin oncogenic activity is modified by other factors, and these determinants are unknown. Here we reveal that CD73 restrains the oncogenic activity of exon 3 β-catenin mutants, and its loss associates with recurrence. Using 7 patient-specific mutants, with genetic deletion or ectopic expression of CD73, we show that CD73 loss increases β-catenin-TCF/LEF transcriptional activity. In cells lacking CD73, membrane levels of mutant β-catenin decreased which corresponded with increased levels of nuclear and chromatin-bound mutant β-catenin. These results suggest CD73 sequesters mutant β-catenin to the membrane to limit its oncogenic activity. Adenosine A1 receptor deletion phenocopied increased β-catenin-TCF/LEF activity seen with NT5E deletion, suggesting that the effect of CD73 loss on mutant β-catenin is mediated via attenuation of adenosine receptor signaling. RNA-seq analyses revealed that NT5E deletion alone drives pro-tumor Wnt/β-catenin gene expression and, with CD73 loss, β-catenin mutants dysregulate zinc-finger and non-coding RNA gene expression. We identify CD73 as a novel regulator of oncogenic β-catenin and help explain variability in patient outcomes in CTNNB1 mutant EC.

Bioactivity of Grape Pomace Extract and Sodium Selenite, Key Components of the OenoGrape Advanced Complex, on Target Human Cells: Intracellular ROS Scavenging and Nrf2/ARE Induction Following In Vitro Intestinal Absorption.

Oenobiol Sun Expert, a food formulation designed to enhance skin health prior to sun exposure, has been optimized by incorporating the OenoGrape Advanced Complex, which includes grape pomace extract, increased selenium content and 10% lycopene-rich tomato extract, with these constituents exhibiting high antioxidant potential. To evaluate the effects of these individual ingredients and the overall formulation at the cellular level, the AOP1 cell antioxidant efficacy assay was employed to measure the intracellular free radical scavenging activity, while the Cell Antioxidant Assay (CAA or DCFH-DA) assay was used to assess peroxidation scavenging at the plasma membrane level. The indirect antioxidant activity was examined using stably transfected cell lines containing a luciferase reporter gene controlled by the Antioxidant Response Element (ARE), which activates the endogenous antioxidant system via the Nrf2/Keap1-ARE pathway. Our results indicate that among the individual components, grape pomace extract and sodium selenite possess high and complementary antioxidant properties. Grape pomace extract was particularly effective in inhibiting free radicals (AOP1 EC50 = 6.80 μg/mL) and activating the ARE pathway (ARE EC50 = 231.1 μg/mL), whereas sodium selenite exerted its effects through potent ARE activation at sub-microgram levels (EC50 = 0.367 μg/mL). In contrast, the lycopene-rich tomato extract did not show a notable contribution to the antioxidant effects. The antiradical activity of the OenoGrape Advanced Complex, comprising these three ingredients, was very efficient and consistent with the results obtained for the individual components (AOP1 EC50 = 15.78 µg/mL and ARE EC50 of 707.7 μg/mL). Similarly, the free radical scavenging activity still persisted in the Oenobiol Sun Expert formulation (AOP1 EC50 = 36.63 µg/mL). Next, in vitro intestinal transepithelial transfer experiments were performed. The basolateral compartments of cells exposed to the ingredients were collected and assessed using the same antioxidant cell assays. The direct and indirect antioxidant activities were measured on both hepatocytes and keratinocytes, demonstrating the bioavailability and bioactivity of grape pomace extract and sodium selenite. These finding suggest that the ingredients of this food supplement contribute to enhanced cytoprotection following ingestion.

The contribution of mitochondria-associated ER membranes to cholesterol homeostasis.

Cellular demands for cholesterol are met by a balance between its biosynthesis in the endoplasmic reticulum (ER) and its uptake from lipoproteins. Cholesterol levels in intracellular membranes form a gradient maintained by a complex network of mechanisms including the control of the expression, compartmentalization and allosteric modulation of the enzymes that balance endogenous and exogenous sources of cholesterol. Low-density lipoproteins (LDLs) are internalized and delivered to lysosomal compartments to release their cholesterol content, which is then distributed within cellular membranes. High-density lipoproteins (HDLs), on the other hand, can transfer their cholesterol content directly into cellular membranes through the action of receptors such as the scavenger receptor B type 1 (SR-B1; gene SCARB1). We show here that SR-B1-mediated exogenous cholesterol internalization from HDL stimulates the formation of lipid-raft subdomains in the ER known as mitochondria-associated ER membranes (MAM), that, in turn, suppress de novo cholesterol biosynthesis machinery. We propose that MAM is a regulatory hub for cholesterol homeostasis that offers a novel dimension for understanding the intracellular regulation of this important lipid.

Comparison of Routine Preoperative Ultrasound-Guided Airway Examination Versus Clinical Airway Examination in Predicting Difficult Laryngoscopy in Patients Coming for Elective Surgery: An Observational Study.

Background As part of preoperative patient evaluations, it is customary to estimate the likelihood of difficulties during laryngoscopy and intubation. A diverse array of predictors is frequently employed by anesthesiologists to anticipate difficult laryngoscopy (DL); however, no single predictor has been established as the gold standard. In the present study, we compared routine preoperative ultrasound-guided airway examination and clinical airway examination in predicting DL in patients coming for elective surgery. Methodology The present study is a single-blinded observational study that spanned 12 months, beginning in March 2018 and concluding in February 2019. The study took place at Kovai Medical Center and Hospital (KMCH), which is a tertiary-care facility in Coimbatore, India. The data were obtained through face-to-face interviews, with a sample size of 135 research participants. The questionnaire contained comprehensive data regarding the patient's age, weight, gender, comorbidities, planned surgery, and preoperative assessment, which encompassed a detailed examination of the airway. The clinical airway examination comprised assessing mouth opening, performing the modified Mallampati test, measuring thyromental height (TMH) and thyromental distance (TMD), evaluating neck circumference, and observing neck extension. Ultrasound-guided airway examination was performed during the immediate pre-operative period, focusing on specific parameters such as the pre-epiglottic space, anterior neck soft tissue at the level of vocal cords, thyrohyoid membrane, suprasternal notch, hyoid bone (single parameter measured at different levels), hyo-mental ratio, and tongue volume (TV). Results Among the ultrasound parameters, TV (72.15 cm3) is the best ultrasound parameter to predict DL (95%CI: 0.65 to 0.82), with a high sensitivity of 82% and specificity of 45.88%. Other parameters that are useful are anterior neck soft tissue at the level of the thyrohyoid membrane (0.67 cm), suprasternal notch (1.01 cm), and, lastly, pre-epiglottic space (0.57 cm). Among clinical parameters, modified Mallampati grade (grade 3 or more) is the best parameter to assess the airway clinically and predict DL, followed by neck circumference (>42 cm) and TMH (>5 cm). Based on the findings, we observed that both clinical and ultrasound parameters are comparable to predict DL. Conclusion We observed that routine clinical airway examination and ultrasound examination yielded comparable results in predicting DL. Therefore, the routine use of clinical airway examination still holds good in predicting DL.

Noncanonical Amino Acid Tools and Their Application to Membrane Protein Studies.

Methods rooted in chemical biology have contributed significantly to studies of integral membrane proteins. One recent key approach has been the application of genetic code expansion (GCE), which enables the site-specific incorporation of noncanonical amino acids (ncAAs) with defined chemical properties into proteins. Efficient GCE is challenging, especially for membrane proteins, which have specialized biogenesis and cell trafficking machinery and tend to be expressed at low levels in cell membranes. Many eukaryotic membrane proteins cannot be expressed functionally in E. coli and are most effectively studied in mammalian cell culture systems. Recent advances have facilitated broader applications of GCE for studies of membrane proteins. First, AARS/tRNA pairs have been engineered to function efficiently in mammalian cells. Second, bioorthogonal chemical reactions, including cell-friendly copper-free "click" chemistry, have enabled linkage of small-molecule probes such as fluorophores to membrane proteins in live cells. Finally, in concert with advances in GCE methodology, the variety of available ncAAs has increased dramatically, thus enabling the investigation of protein structure and dynamics by multidisciplinary biochemical and biophysical approaches. These developments are reviewed in the historical framework of the development of GCE technology with a focus on applications to studies of membrane proteins.