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Recent advances in genetic diagnosis identified variants in genes encoding GABAA receptors as causative for genetic epilepsy. Here, we selected eight disease-associated variants in the α1 subunit of GABAA receptors causing mild to severe clinical phenotypes and showed that they are loss of function, mainly by reducing the folding and surface trafficking of the α1 protein. Furthermore, we sought client protein-specific pharmacological chaperones to restore the function of pathogenic receptors. Applications of positive allosteric modulators, including Hispidulin and TP003, increase the functional surface expression of the α1 variants. Mechanism of action study demonstrated that they enhance the folding, assembly, and trafficking and reduce the degradation of GABAA variants without activating the unfolded protein response in HEK293T cells and human iPSC-derived neurons. Since these compounds cross the blood-brain barrier, such a pharmacological chaperoning strategy holds great promise to treat genetic epilepsy in a GABAA receptor-specific manner.

Protein homeostasis (proteostasis) deficiency is an important contributing factor to neurological and metabolic diseases. However, how the proteostasis network orchestrates the folding and assembly of multi-subunit membrane proteins is poorly understood. Previous proteomics studies identified Hsp47 (Gene: SERPINH1), a heat shock protein in the endoplasmic reticulum lumen, as the most enriched interacting chaperone for gamma-aminobutyric acid type A (GABAA) receptors. Here, we show that Hsp47 enhances the functional surface expression of GABAA receptors in rat neurons and human HEK293T cells. Furthermore, molecular mechanism study demonstrates that Hsp47 acts after BiP (Gene: HSPA5) and preferentially binds the folded conformation of GABAA receptors without inducing the unfolded protein response in HEK293T cells. Therefore, Hsp47 promotes the subunit-subunit interaction, the receptor assembly process, and the anterograde trafficking of GABAA receptors. Overexpressing Hsp47 is sufficient to correct the surface expression and function of epilepsy-associated GABAA receptor variants in HEK293T cells. Hsp47 also promotes the surface trafficking of other Cys-loop receptors, including nicotinic acetylcholine receptors and serotonin type 3 receptors in HEK293T cells. Therefore, in addition to its known function as a collagen chaperone, this work establishes that Hsp47 plays a critical and general role in the maturation of multi-subunit Cys-loop neuroreceptors.

Abstract Background Newly graduated registered nurses leaving the nursing profession in the early stages of their career have enormous financial and time implications for nursing organizations and affect the quality of nursing care. Objective To identify the factors influencing newly graduated registered nurses’ intention to leave the nursing profession over the past 10 years. Methods The framework developed by Whittemore and Knafl was used to conduct this integrative review. An electronic search was conducted for English articles to identify research studies published between 2011-2022 using the following databases of PubMed, MEDLINE, CINAHL, PsycINFO, and Scopus. Eligible publications were critically reviewed and scored using the Critical Appraisal Skills Program Checklist and the Center for Evidence-Based Management appraisal. Results Twenty-one studies were analyzed. The main factors affecting newly graduated registered nurses’ intention to leave the nursing profession included demographic factors (age, educational level, year of experience, professional title, employment status, health status, shift, hospital location and size), supervisor and peer support, challenges in the workplace, cognitive and affective response to work, work environment (collegial nurse-physician relations, insufficient staffing level, person-work environment fit), gender stereotypes, autonomous motivation, role models, and resilience. Conclusions The factors affecting newly graduated registered nurses’ intention to leave the nursing profession are multifaceted and should receive continuous attention from nurse managers. The findings provide more comprehensive for nurse administrators to develop intervention strategies to mitigate newly graduated registered nurses’ turnover intention.

Background: The pollution of organophosphate flame retardants (OPFRs) is of global concern, but the site-specific data of OPFR concentrations in drinking water are scarce for many areas of the world outside of Europe and the US. This study aimed to investigate the occurrence and profiles of OPFRs in the tap water treatment and delivery process in Shanghai. Methods: In total, 106 samples were analyzed for 10 OPFRs, which were collected periodically from monitoring points of drinking water treatment plants and piped water between November 2021 and July 2023. The average daily doses of OPFRs through the ingestion of tap water were calculated by multiplying nominal volumes of water ingestion rates with the measured concentrations of OPFRs. Hazard quotients, the hazard index, and the carcinogenic risks of OPFRs via drinking water were used to estimate the health risks. Results: Tributyl phosphate (TBP), tris(2-chloroethyl) phosphate (TCEP), and tris (1-chloro-2-propyl) phosphate (TCIPP) were found in >90% of the tap water samples, whereas triethyl phosphate (TEP) and tris (2,3-dibromopropyl) phosphate (TDBPP) were not found in any samples. The concentrations of Σ10OPFRs were found at part-per-trillion ranges, with average concentrations that ranged from 86.0 ng/L in February 2023 (dry season) to 218 ng/L in July 2022 (wet season). TCIPP was the most abundant compound among the investigated OPFRs. The average daily dose of Σ10OPFRs via the ingestion of tap water was up to 20.4 ng/kg body weight/day. The hazard quotients of OPFRs through drinking water were in the range of 10−5–10−4, indicating low risk levels. Moreover, the hazard index of OPFRs indicated that the risk for children (2 × 10−4) was higher than adults (7 × 10−5). Conclusion: Tap water intake may be an important source of OPFRs exposure. But the risk of OPFRs for local residents is at a low level through drinking water.

IntroductionSweet sorghum juice is a typical production feedstock for natural, eco-friendly sweeteners and beverages. Clostridium tyrobutyricum is one of the widely used microorganisms in the food industry, and its principal product, bio-butyric acid is an important food additive. There are no published reports of Clostridium tyrobutyricum producing butyric acid using SSJ as the sole substrate without adding exogenous substances, which could reach a food-additive grade. This study focuses on tailoring a cost-effective, safe, and sustainable process and strategy for their production and application.MethodsThis study modeled the enzymolysis of non-reducing sugars via the first/second-order kinetics and added food-grade diatomite to the hydrolysate. Qualitative and quantitative analysis were performed using high-performance liquid chromatography, gas chromatography-mass spectrometer, full-scale laser diffraction method, ultra-performance liquid chromatography–tandem mass spectrometry, the cell double-staining assay, transmission electron microscopy, and Oxford nanopore technology sequencing. Quantitative real-time polymerase chain reaction, pathway and process enrichment analysis, and homology modeling were conducted for mutant genes.ResultsThe treated sweet sorghum juice showed promising results, containing 70.60 g/L glucose and 63.09 g/L fructose, with a sucrose hydrolysis rate of 98.29% and a minimal sucrose loss rate of 0.87%. Furthermore, 99.62% of the colloidal particles and 82.13% of the starch particles were removed, and the concentrations of hazardous substances were effectively reduced. A food microorganism Clostridium tyrobutyricum TGL-A236 with deep utilization value was developed, which showed superior performance by converting 30.65% glucose and 37.22% fructose to 24.1364 g/L bio-butyric acid in a treated sweet sorghum juice (1:1 dilution) fermentation broth. This titer was 2.12 times higher than that of the original strain, with a butyric acid selectivity of 86.36%. Finally, the Genome atlas view, Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and evolutionary genealogy of genes: Non-supervised Orthologous (eggNOG) functional annotations, three-dimensional structure and protein cavity prediction of five non-synonymous variant genes were obtained.ConclusionThis study not only includes a systematic process flow and in-depth elucidation of relevant mechanisms but also provides a new strategy for green processing of food raw materials, improving food microbial performance, and ensuring the safe production of food additives.

Abstract Background Genetic variants in the subunits of the gamma-aminobutyric acid type A (GABAA) receptors are implicated in the onset of multiple pathologic conditions including genetic epilepsy. Previous work showed that pathogenic GABAA subunits promote misfolding and inefficient assembly of the GABAA receptors, limiting receptor expression and activity at the plasma membrane. However, GABAA receptors containing variant subunits can retain activity, indicating that enhancing the folding, assembly, and trafficking of these variant receptors offers a potential opportunity to mitigate pathology associated with genetic epilepsy. Results Here, we demonstrate that pharmacologically enhancing endoplasmic reticulum (ER) proteostasis using small molecule activators of the ATF6 (Activating Transcription Factor 6) signaling arm of the unfolded protein response (UPR) increases the assembly, trafficking, and surface expression of variant GABAA receptors. These improvements are attributed to ATF6-dependent remodeling of the ER proteostasis environment, which increases protein levels of pro-folding ER proteostasis factors including the ER chaperone BiP (Immunoglobulin Binding Protein) and trafficking receptors, such as LMAN1 (Lectin Mannose-Binding 1) and enhances their interactions with GABAA receptors. Importantly, we further show that pharmacologic ATF6 activators increase the activity of GABAA receptors at the cell surface, revealing the potential for this strategy to restore receptor activity to levels that could mitigate disease pathogenesis. Conclusions These results indicate that pharmacologic ATF6 activators offer an opportunity to restore GABAA receptor activity in diseases including genetic epilepsy and point to the potential for similar pharmacologic enhancement of ER proteostasis to improve trafficking of other disease-associated variant ion channels implicated in etiologically-diverse diseases.

IntroductionClostridium tyrobutyricum has considerable prospect in the production of organic acids. Globally, refinery final molasses is rich in sugar and reported to have high levels of accumulation and high emission costs, recognized as an excellent substrate for C. tyrobutyricum fermentation, but there is no suitable method available at present.MethodsIn this study, an acid-base treatment combined with a new green membrane treatment technology – a dynamic ion-exchange membrane -was used to pretreat refinery final molasses, so that it could be used for C. tyrobutyricum to produce butyric acid. A high-performance liquid chromatography method was established to determine the conversion of a large amount of sucrose into fermentable sugars (71.88 g/L glucose and 38.06 g/L fructose) in the treated refinery final molasses. The process of sequential filtration with 3, 1, and 0.45 μm-pore diameter dynamic ion-exchange membranes could remove impurities, pigments, and harmful substances from the refinery final molasses, and retain the fermentable sugar.Results and discussionThis means that refinery final molasses from the sugar industry could be utilized as a high-value by-product and used for the growth of C. tyrobutyricum, with industrial feasibility and economic competitiveness. Using the treated refinery final molasses as a carbon source, C. tyrobutyricum was screened by the method of adaptive evolution. The strain with butyric acid yielded 52.54 g/L, and the yield of the six carbon sugar was increased from 0.240 to 0.478 g/g. The results showed that combination of C. tyrobutyricum and ionic membrane technology broke through the bottleneck of its utilization of refinery final molasses. This study provided an innovative idea for the C. tyrobutyricum fermentation to produce butyric acid.

Soil type is an important environmental factor that affects plant growth and development, especially during the early growth stage. In this study, P. chinense (Phellodendron chinense) seedlings were cultivated on red soil (RS), yellow soil (YS), acidic purple soil (ACPS), and alkaline purple soil (ALPS), and the influence of soil types on the growth parameters and physiological responses at three growth stages were investigated. The results show that soil types and growth stages can significantly influence seedling height, base diameter, and biomass in P. chinense seedlings. Moreover, the significant variations in chlorophyll, total polyphenols, soluble protein, soluble sugar, and malondialdehyde (MDA) content, as well as superoxide dismutase (SOD), peroxidase (POD), phenylalanine ammonia-lyase (PAL), and polyphenol oxidase (PPO) activity, were recorded at three growth stages among four soil types. These results show that P. chinense seedlings can grow normally in four soil types, and ACPS may be more beneficial for the artificial cultivation of P. chinense seedlings than those of RS, ALPS, and YS. Principal component analysis (PCA) reveals a strong correlation and clear differences in the test parameters between growth stage and organs in four soil types, and the cumulative contribution percentages of the total biplot range from 74.44% to 81.97%. This present information will be helpful for farmers in selecting soil type for the large-scale cultivation of P. chinense seedlings.

Summary: The endoplasmic reticulum membrane complex (EMC) plays a critical role in the biogenesis of tail-anchored proteins and a subset of multi-pass membrane proteins in the endoplasmic reticulum (ER). However, because of nearly exclusive expression of neurotransmitter-gated ion channels in the central nervous system (CNS), the role of the EMC in their biogenesis is not well understood. In this study, we demonstrated that the EMC positively regulates the surface trafficking and thus function of endogenous γ-aminobutyric acid type A (GABAA) receptors, the primary inhibitory ion channels in the mammalian brain. Moreover, among ten EMC subunits, EMC3 and EMC6 have the most prominent effect, and overexpression of EMC3 or EMC6 is sufficient to restore the function of epilepsy-associated GABAA receptor variants. In addition, EMC3 and EMC6 demonstrate endogenous interactions with major neuroreceptors, which depends on their transmembrane domains, suggesting a general role of the EMC in the biogenesis of neuroreceptors.

N-methyl-D-aspartate receptors (NMDARs) are glutamate-gated cation channels that mediate excitatory neurotransmission and are critical for synaptic development and plasticity in the mammalian central nervous system (CNS). Functional NMDARs typically form via the heterotetrameric assembly of GluN1 and GluN2 subunits. Variants within GRIN genes are implicated in various neurodevelopmental and neuropsychiatric disorders. Due to the significance of NMDAR subunit composition for regional and developmental signaling at synapses, properly folded receptors must reach the plasma membrane for their function. This review focuses on the protein quality control of NMDARs. Specifically, we review the quality control mechanisms that ensure receptors are correctly folded and assembled within the endoplasmic reticulum (ER) and trafficked to the plasma membrane. Further, we discuss disease-associated variants that have shown disrupted NMDAR surface expression and function. Finally, we discuss potential targeted pharmacological and therapeutic approaches to ameliorate disease phenotypes by enhancing the expression and surface trafficking of subunits harboring disease-associated variants, thereby increasing their incorporation into functional receptors.

In this paper, an aerostatic bearing model in the form of microorifice annular array is designed to explore the static performance of aerostatic bearing when the orifice diameter and working air film thickness are of the same order of magnitude. Firstly, the simulation model of the new bearing is established, the pressure distribution of the bearing gas film under the influence of the orifice diameter is analyzed by the CFD method, and the performance of the aerostatic bearing is compared with the traditional toroidal throttling method with the orifice diameter of 0.1 mm; at the same time, the effects of air supply pressure, orifice diameter, and orifice number on the static performance of the bearing under different air film thickness are analyzed; finally, a new bearing is developed, and the simulation results are verified by the experimental test. The results show that when the orifice diameter and the air film thickness are of the same order of magnitude, the maximum stiffness of the aerostatic bearing is significantly improved and the microporous throttled aerostatic bearing is more suitable for the field of high stiffness. The greater the air supply pressure, the greater the bearing capacity and stiffness of the bearing. The number of orifices has an effect on improving the bearing capacity of the bearing, but when the number of orifices is large, the effect will become less obvious. This study can provide a new theoretical reference for the design of aerostatic bearing.

Early-onset neonatal sepsis (EONS), a bacterial infection that occurs within 72 h after birth, is associated with high likelihood of neonatal mortality. Latamoxef, a semi-synthetic oxacephem antibiotic developed in 1980s, has been brought back into empirical EONS treatment in recent years. In the preliminary work, we established a population pharmacokinetics (PPK) model for latamoxef in Chinese neonates. Moreover, in order to better guide clinical treatment, we conducted dose simulation and found that ascending administration frequency could improve the target rate of 70% of patients having a free antimicrobial drug concentration exceeding the MIC during 70% of the dosing interval (70% fT > MIC). Accordingly, this study is aimed to compare the 70% fT > MIC, efficacy and safety between conventional regimen and PPK model regimen for rational use of latamoxef in EONS treatment. A single-blind, multicenter randomized controlled trial (RCT) for latamoxef will be conducted in Chinese EONS patients. Neonates (≤3 days of age, expected number = 114) admitted to the hospital with the diagnosis of EONS and fulfilling inclusion and exclusion criteria will be randomized (ratio of 1:1) to either a conventional regimen (30 mg/kg q12h) or model regimen (20 mg/kg q8h) latamoxef treatment group for at least 3 days. Primary outcome measure will be 70% fT > MIC and secondary outcome indicators will be the latamoxef treatment failure, duration of antibiotic therapy, changes of white blood cell count (WBC), C-reactive protein (CRP) and procalcitonin (PCT), blood culture results during administration and incidence of adverse event (AE)s. Assessments will be made at baseline, initial stage of latamoxef treatment (18–72 h) and before the end of latamoxef treatment. Ethical approval of our clinical trial has been granted by the ethics committee of the Beijing Children’s Hospital (ID: 2020-13-1). Written informed consent will be obtained from the parents of the participants. This trial is registered in the Chinese Clinical Trial Registry (ChiCTR 2000040064).It is hoped that our study will provide a clinical basis for the rational clinical use of latamoxef in EONS treatment.

Lipopolysaccharide (LPS) can induce bone loss by stimulating osteoclast formation. Colony-stimulating factor 1 receptor (CSF 1R) inhibitors have great potential for the treatment of rheumatoid arthritis and tumor-related bone erosion. However, its role in LPS-induced bone loss is still not clarified. In this study, we observed the effects of CSF 1R inhibitor, PLX3397, on LPS-induced bone damage in an animal model. The models were established by LPS administration in male Sprague-Dawley rats. PLX3397 (30 mg/kg body weight) was given by oral gavage. MicroCT analysis, biomechanical properties, biomarker assay, histological examination, and mRNA expression of osteoclast differentiation-related genes (Traf6, Fra1, c-fos and NFATc1) were performed on the 8th week. LPS induced bone loss was shown as the decrease in bone volume fraction and trabecular number and increase in trabecular separation (p

Mitochondria defects caused by mutant huntingtin (mtHtt) have been implicated in Huntington's disease. Here authors show that VCP binds to mtHtt on the mitochondria, and that treatment with a peptide that disrupts this interaction reduces the cellular and behavioural deficits in mouse models of HD.

Biogenesis of membrane proteins is controlled by the protein homeostasis (proteostasis) network. We have been focusing on protein quality control of γ-aminobutyric acid type A (GABAA) receptors, the major inhibitory neurotransmitter-gated ion channels in mammalian central nervous system. Proteostasis deficiency in GABAA receptors causes loss of their surface expression and thus function on the plasma membrane, leading to epilepsy and other neurological diseases. One well-characterized example is the A322D mutation in the α1 subunit that causes its extensive misfolding and expedited degradation in the endoplasmic reticulum (ER), resulting in autosomal dominant juvenile myoclonic epilepsy. We aimed to correct misfolding of the α1(A322D) subunits in the ER as an approach to restore their functional surface expression. Here, we showed that application of BIX, a specific, potent ER resident HSP70 family protein BiP activator, significantly increases the surface expression of the mutant receptors in human HEK293T cells and neuronal SH-SY5Y cells. BIX attenuates the degradation of α1(A322D) and enhances their forward trafficking and function. Furthermore, because BiP is one major target of the two unfolded protein response (UPR) pathways: ATF6 and IRE1, we continued to demonstrate that modest activations of the ATF6 pathway and IRE1 pathway genetically enhance the plasma membrane trafficking of the α1(A322D) protein in HEK293T cells. Our results underlie the potential of regulating the ER proteostasis network to correct loss-of-function protein conformational diseases.

High blood pressure (BP) is the most common cardiovascular risk factor worldwide and a major contributor to heart disease and stroke. We previously discovered a BP-associated missense SNP (single nucleotide polymorphism)-rs2272996-in the gene encoding vanin-1, a glycosylphosphatidylinositol (GPI)-anchored membrane pantetheinase. In the present study, we first replicated the association of rs2272996 and BP traits with a total sample size of nearly 30,000 individuals from the Continental Origins and Genetic Epidemiology Network (COGENT) of African Americans (P=0.01). This association was further validated using patient plasma samples; we observed that the N131S mutation is associated with significantly lower plasma vanin-1 protein levels. We observed that the N131S vanin-1 is subjected to rapid endoplasmic reticulum-associated degradation (ERAD) as the underlying mechanism for its reduction. Using HEK293 cells stably expressing vanin-1 variants, we showed that N131S vanin-1 was degraded significantly faster than wild type (WT) vanin-1. Consequently, there were only minimal quantities of variant vanin-1 present on the plasma membrane and greatly reduced pantetheinase activity. Application of MG-132, a proteasome inhibitor, resulted in accumulation of ubiquitinated variant protein. A further experiment demonstrated that atenolol and diltiazem, two current drugs for treating hypertension, reduce the vanin-1 protein level. Our study provides strong biological evidence for the association of the identified SNP with BP and suggests that vanin-1 misfolding and degradation are the underlying molecular mechanism.

This paper indicated that inactivated Bordetella pertussis (iBp) can enhance the lung airway hyperreactivity of the rats sensitized and challenged with OVA. The mechanisms were involved in the upregulation of cAMP-PDE activity and PDE4A, PDE4D, and PDE3 gene expression in the lungs. But only PDE4 activity was different between the OVA and OVA+iBp groups, and PDE4D expression was significantly increased in iBp rats alone. So, our data suggested that cosensitization with OVA and iBp affects lung airway reactivity by modulating the lung cAMP-PDE activity and PDE4D gene expression.

Gamma-aminobutyric acid type A (GABAA) receptors are the primary inhibitory neurotransmitter-gated ion channels in the mammalian central nervous system. Maintenance of GABAA receptor protein homeostasis (proteostasis) in cells utilizing its interacting proteins is essential for the function of GABAA receptors. However, how the proteostasis network orchestrates GABAA receptor biogenesis in the endoplasmic reticulum is not well understood. Here, we employed a proteomics-based approach to systematically identify the interactomes of GABAA receptors. We carried out a quantitative immunoprecipitation-tandem mass spectrometry analysis utilizing stable isotope labeling by amino acids in cell culture. Furthermore, we performed comparative proteomics by using both WT α1 subunit and a misfolding-prone a1 subunit carrying the A322D variant as the bait proteins. We identified 125 interactors for WT α1-containing receptors, 105 proteins for a1(A322D)-containing receptors, and 54 overlapping proteins within these two interactomes. Our bioinformatics analysis identified potential GABAA receptor proteostasis network components, including chaperones, folding enzymes, trafficking factors, and degradation factors, and we assembled a model of their potential involvement in the cellular folding, degradation, and trafficking pathways for GABAA receptors. In addition, we verified endogenous interactions between a1 subunits and selected interactors by using coimmunoprecipitation in mouse brain homogenates. Moreover, we showed that TRIM21 (tripartite motif containing-21), an E3 ubiquitin ligase, positively regulated the degradation of misfolding-prone a1(A322D) subunits selectively. This study paves the way for understanding the molecular mechanisms as well as fine-tuning of GABAA receptor proteostasis to ameliorate related neurological diseases such as epilepsy. [ABSTRACT FROM AUTHOR]

N-methyl-D-aspartate receptors (NMDARs) are glutamate-gated cation channels that mediate excitatory neurotransmission and are critical for synaptic development and plasticity in the mammalian central nervous system (CNS). Functional NMDARs typically form via the heterotetrameric assembly of GluN1 and GluN2 subunits. Variants within GRIN genes are implicated in various neurodevelopmental and neuropsychiatric disorders. Due to the significance of NMDAR subunit composition for regional and developmental signaling at synapses, properly folded receptors must reach the plasma membrane for their function. This review focuses on the protein quality control of NMDARs. Specifically, we review the quality control mechanisms that ensure receptors are correctly folded and assembled within the endoplasmic reticulum (ER) and trafficked to the plasma membrane. Further, we discuss disease-associated variants that have shown disrupted NMDAR surface expression and function. Finally, we discuss potential targeted pharmacological and therapeutic approaches to ameliorate disease phenotypes by enhancing the expression and surface trafficking of subunits harboring disease-associated variants, thereby increasing their incorporation into functional receptors. [ABSTRACT FROM AUTHOR]

The radiolabled acyl-homoserine lactones (acyl-HSL) are synthesized to examine their fate when amended at low concentrations, relevant to quorum sensing, to buffered bulk soil slurries and other samples. The biological signal decay might either promote or complicate cellular communications and the accuracy of population density-based controls on gene expression in species-rich ecosystems.