Efficacious human metapneumovirus vaccine based on AI-guided engineering of a closed prefusion trimer.

The prefusion conformation of human metapneumovirus fusion protein (hMPV Pre-F) is critical for eliciting the most potent neutralizing antibodies and is the preferred immunogen for an efficacious vaccine against hMPV respiratory infections. Here we show that an additional cleavage event in the F protein allows closure and correct folding of the trimer. We therefore engineered the F protein to undergo double cleavage, which enabled screening for Pre-F stabilizing substitutions at the natively folded protomer interfaces. To identify these substitutions, we developed an AI convolutional classifier that successfully predicts complex polar interactions often overlooked by physics-based methods and visual inspection. The combination of additional processing, stabilization of interface regions and stabilization of the membrane-proximal stem, resulted in a Pre-F protein vaccine candidate without the need for a heterologous trimerization domain that exhibited high expression yields and thermostability. Cryo-EM analysis shows the complete ectodomain structure, including the stem, and a specific interaction of the newly identified cleaved C-terminus with the adjacent protomer. Importantly, the protein induces high and cross-neutralizing antibody responses resulting in near complete protection against hMPV challenge in cotton rats, making the highly stable, double-cleaved hMPV Pre-F trimer an attractive vaccine candidate.

Structural basis for the oligomerization-facilitated NLRP3 activation.

The NACHT-, leucine-rich-repeat-, and pyrin domain-containing protein 3 (NLRP3) is a critical intracellular inflammasome sensor and an important clinical target against inflammation-driven human diseases. Recent studies have elucidated its transition from a closed cage to an activated disk-like inflammasome, but the intermediate activation mechanism remains elusive. Here we report the cryo-electron microscopy structure of NLRP3, which forms an open octamer and undergoes a ~ 90° hinge rotation at the NACHT domain. Mutations on open octamer's interfaces reduce IL-1ß signaling, highlighting its essential role in NLRP3 activation/inflammasome assembly. The centrosomal NIMA-related kinase 7 (NEK7) disrupts large NLRP3 oligomers and forms NEK7/NLRP3 monomers/dimers which is a critical step preceding the assembly of the disk-like inflammasome. These data demonstrate an oligomeric cooperative activation of NLRP3 and provide insight into its inflammasome assembly mechanism.

Structural basis of the human NAIP/NLRC4 inflammasome assembly and pathogen sensing.

The NLR family caspase activation and recruitment domain-containing 4 (NLRC4) inflammasome is a critical cytosolic innate immune machine formed upon the direct sensing of bacterial infection and in response to cell stress during sterile chronic inflammation. Despite its major role in instigating the subsequent host immune response, a more complete understanding of the molecular events in the formation of the NLRC4 inflammasome in humans is lacking. Here we identify Bacillus thailandensis type III secretion system needle protein (Needle) as a potent trigger of the human NLR family apoptosis inhibitory protein (NAIP)/NLRC4 inflammasome complex formation and determine its structural features by cryogenic electron microscopy. We also provide a detailed understanding of how type III secretion system pathogen components are sensed by human NAIP to form a cascade of NLRC4 protomer through a critical lasso-like motif, a 'lock-key' activation model and large structural rearrangement, ultimately forming the full human NLRC4 inflammasome. These results shed light on key regulatory mechanisms specific to the NLRC4 inflammasome assembly, and the innate immune modalities of pathogen sensing in humans.

Structural and mechanistic insights into the inhibition of respiratory syncytial virus polymerase by a non-nucleoside inhibitor.

The respiratory syncytial virus polymerase complex, consisting of the polymerase (L) and phosphoprotein (P), catalyzes nucleotide polymerization, cap addition, and cap methylation via the RNA dependent RNA polymerase, capping, and Methyltransferase domains on L. Several nucleoside and non-nucleoside inhibitors have been reported to inhibit this polymerase complex, but the structural details of the exact inhibitor-polymerase interactions have been lacking. Here, we report a non-nucleoside inhibitor JNJ-8003 with sub-nanomolar inhibition potency in both antiviral and polymerase assays. Our 2.9 Å resolution cryo-EM structure revealed that JNJ-8003 binds to an induced-fit pocket on the capping domain, with multiple interactions consistent with its tight binding and resistance mutation profile. The minigenome and gel-based de novo RNA synthesis and primer extension assays demonstrated that JNJ-8003 inhibited nucleotide polymerization at the early stages of RNA transcription and replication. Our results support that JNJ-8003 binding modulates a functional interplay between the capping and RdRp domains, and this molecular insight could accelerate the design of broad-spectrum antiviral drugs.

Does acupuncture have advantages in the rehabilitation of vascular mild cognitive impairment? A systematic review and meta-analysis.

Background: Vascular mild cognitive impairment (VMCI) is a common impairment caused by vascular factors. VMCI often occurs after stroke, and it is the main clinical manifestation of long-term disability. Many patients are treated with acupuncture in combination with other therapies. However, evidence regarding the effectiveness of this treatment regimen is lacking. Aims: This meta-analysis aimed to evaluate the efficacy of acupuncture therapy for treating VMCI. Methods: This systematic review was conducted in accordance with the preferred reporting and meta-analysis guidelines. The CNKI, Wanfang, VIP, CBM, Cochrane Library, PubMed and Embase databases were searched from inception to August 20, 2022. After two researchers independently screened the literature, they extracted the data and evaluated the risk of bias in the included studies. Revman 5.3 software was used for the meta-analysis. Summary of review: Thirty-two randomized controlled trials (RCTs) were included. The overall effective rate of acupuncture for treating VMCI was 3.06, 95% CI [2.39, 3.91], (P < 0.05). Montreal Cognitive Assessment (MoCA), Mini-Mental State Examination (MMSE), Barthel Index and Activities of Daily Living (ADLs) scores significantly differed between the treatment and control groups, with weighted mean differences (WMDs) [95% CI] (P value) of 1.97 [1.44, 2.49] (P < 0.05), 2.02 [1.50, 2.54] (P < 0.05), 5.54 [3.81, 7.28] (P < 0.05), and 3.43 [2.53, 4.33] (P < 0.05), respectively. The overall effective rate of electroacupuncture (EA) for treating VMCI was better than that of the control group (RR = 2.25, 95% CI, [1.13, 4.50], P < 0.05). MoCA, MMSE, Barthel index and ADL scores differed significantly between the treatment and control groups, with WMDs [95% CI] (P value) of 1.79 [1.20, 2.38] (P < 0.05), 1.45 [0.87, 2.03] (P < 0.05), 5.78 [2.38, 9.18] (P < 0.05), and 3.15 [2.15, 4.15] (P < 0.05), respectively. Acupuncture alone and combined with drug therapy were thus superior to drug therapy alone for improving cognitive function. EA also has potential advantages. Conclusions: Acupuncture combined with another therapy is better than other therapies alone, such as simple drug therapy, for treating VMCI. However, variations in study duration (4-12 weeks) limit us from drawing any definitive conclusions about long-term effects. Therefore, more RCTs with rigorous designs and reasonable treatment and follow-up durations are needed.

Convergence of immune escape strategies highlights plasticity of SARS-CoV-2 spike.

The global spread of the SARS-CoV-2 virus has resulted in emergence of lineages which impact the effectiveness of immunotherapies and vaccines that are based on the early Wuhan isolate. All currently approved vaccines employ the spike protein S, as it is the target for neutralizing antibodies. Here we describe two SARS-CoV-2 isolates with unusually large deletions in the N-terminal domain (NTD) of the spike. Cryo-EM structural analysis shows that the deletions result in complete reshaping of the NTD supersite, an antigenically important region of the NTD. For both spike variants the remodeling of the NTD negatively affects binding of all tested NTD-specific antibodies in and outside of the NTD supersite. For one of the variants, we observed a P9L mediated shift of the signal peptide cleavage site resulting in the loss of a disulfide-bridge; a unique escape mechanism with high antigenic impact. Although the observed deletions and disulfide mutations are rare, similar modifications have become independently established in several other lineages, indicating a possibility to become more dominant in the future. The observed plasticity of the NTD foreshadows its broad potential for immune escape with the continued spread of SARS-CoV-2.

Rational design of a highly immunogenic prefusion-stabilized F glycoprotein antigen for a respiratory syncytial virus vaccine.

Respiratory syncytial virus (RSV) is the leading, global cause of serious respiratory disease in infants and is an important cause of respiratory illness in older adults. No RSV vaccine is currently available. The RSV fusion (F) glycoprotein is a key antigen for vaccine development, and its prefusion conformation is the target of the most potent neutralizing antibodies. Here, we describe a computational and experimental strategy for designing immunogens that enhance the conformational stability and immunogenicity of RSV prefusion F. We obtained an optimized vaccine antigen after screening nearly 400 engineered F constructs. Through in vitro and in vivo characterization studies, we identified F constructs that are more stable in the prefusion conformation and elicit ~10-fold higher serum-neutralizing titers in cotton rats than DS-Cav1. The stabilizing mutations of the lead construct (847) were introduced onto F glycoprotein backbones of strains representing the dominant circulating genotypes of the two major RSV subgroups, A and B. Immunization of cotton rats with a bivalent vaccine formulation of these antigens conferred complete protection against RSV challenge, with no evidence of disease enhancement. The resulting bivalent RSV prefusion F investigational vaccine has recently been shown to be efficacious against RSV disease in two pivotal phase 3 efficacy trials, one for passive protection of infants by immunization of pregnant women and the second for active protection of older adults by direct immunization.

Formation of Core-Shell AuCu@Ag Nanocrystals through the Nanoscale Kirkendall Effect.

Polymetallic nanocrystals (NCs) consist of multiple metal elements. A powerful platform to achieve the flexible construction of polymetallic NCs is highly desired but challenging. Herein, we devise a model system that realizes metal atom diffusion between different NCs, resulting in the formation of polymetallic NCs. The differential bond strength between different metal atoms is proposed to initiate such metal atom diffusion, and the specific high surface-to-volume ratio of the NCs can expedite the diffusion process. Taking the Au-Cu-Ag trimetallic system as an example, core-shell AuCu@Ag NCs were successfully formed by combining AgCu NCs with Au NCs. The evolution process was explored, and the gradual fusion of simple NCs into AuCu@Ag NCs was unambiguously observed, which could be attributed to the larger bond strength of Au-Cu than that of Ag-Cu. This work offers an opportunity/platform in theory and experiment to expand the synthesis framework as well as the polymetallic NC list.

Mix-and-Detection Assay with Multiple Cyclic Enzymatic Repairing Amplification for Rapid and Ultrasensitive Detection of Long Noncoding RNAs in Breast Tissues.

Long noncoding RNAs (lncRNAs) are valuable biomarkers and therapeutic targets, and they play essential roles in various pathological and biological processes. So far, the reported lncRNA assays usually suffer from unsatisfactory sensitivity and time-consuming procedures. Herein, we develop a mix-and-read assay based on multiple cyclic enzymatic repairing amplification (ERA) for sensitive and rapid detection of mammalian metastasis-associated lung adenocarcinoma transcript 1 (lncRNA MALAT1). In this assay, we design two three-way junction (3WJ) probes including a 3WJ template and a 3WJ primer to specifically recognize lncRNA MALAT1, and the formation of a stable 3WJ structure induces cyclic ERA to generate triggers. The resulting triggers subsequently hybridize with a free 3WJ template and act as primers to initiate new rounds of cyclic ERA, generating abundant triggers. The hybridization of triggers with signal probes forms stable double-stranded DNA duplexes that can be specifically cleaved by apurinic/apyrimidinic endonuclease 1 to produce a high fluorescence signal. This assay can be carried out in a mix-and-read manner within 10 min under an isothermal condition (50 °C), which is the rapidest and simplest method reported so far for the lncRNA MALAT1 assay. This method can sensitively detect lncRNA MALAT1 with a limit of detection of 0.87 aM, and it can accurately measure endogenous lncRNA MALAT1 at the single-cell level. Moreover, this method can distinguish lncRNA MALAT1 expression in breast cancer patient tissues and their corresponding healthy adjacent tissues. Importantly, the extension of this assay to different RNAs detection can be achieved by simply replacing the corresponding target recognition sequences.

Molecular-Shape-Controlled Nonfused Ring Electron Acceptors for High-Performance Organic Solar Cells with Tunable Phase Morphology.

Two nonfused ring electron acceptors (NFREAs), BTh-OC8-2F and DTh-OC8-2F, with different molecular shapes are designed and synthesized. Both acceptors can form planar molecular shapes by the assistance of S···O intramolecular interactions. Differently, BTh-OC8-2F, with a linear molecular backbone and two trans-arranged side chains at the core unit, exhibits much stronger crystallinity than DTh-OC8-2, with a C-shape molecular shape and two cis-arranged steric side chains at the core unit. Thus, the DTh-OC8-2F based blend film displays a better nanoscale phase separation, more suppressed charge recombination, more efficient exciton dissociation, and lower nonradiative energy loss. Organic solar cells based on DTh-OC8-2F can deliver a power conversion efficiency of 14.13%, which is much higher than BTh-OC8-2F based ones (11.95%) and is also one of the highest values reported for organic solar cells based on NFREAs.

A sensitive and simple competitive nanozyme-linked apta-sorbent assay for the dual-mode detection of ochratoxin A.

The enzyme-linked apta-sorbent assay (ELASA) is widely used for the detection of small-molecule compounds as a result of low cost and reagent stability of aptamers. However, enzyme labels used in ELASA still suffer from some drawbacks, such as high production cost and limited stability. To overcome the drawbacks, we reported a nanozyme-linked apta-sorbent assay (NLASA) coupled with surface-enhanced Raman scattering (SERS)-colorimetric dual-mode detection. For nanozyme labels, Pd-Pt bimetallic nanocrystals (Pd-Pt NRs) could catalyze 3,3',5,5'-tetramethylbenzidine (TMB) to blue TMB2+, whose color variation could not only be distinguished by naked eyes but also had a strong SERS signal. The NLASA method was employed to detect ochratoxin A (OTA) with a limit of detection values of 0.097 nM (0.039 ppb) and 0.042 nM (0.017 ppb) via the colorimetric and SERS methods, respectively. This method was applied for the determination of OTA in wine and grape samples, and the detection results were in a satisfied agreement with those determined by the high performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) method. The proposed NLASA method provided a rapid and sensitive detection for OTA and could also be broadened for other small-molecules.

Refined Capsid Structure of Human Adenovirus D26 at 3.4 Å Resolution.

Various adenoviruses are being used as viral vectors for the generation of vaccines against chronic and emerging diseases (e.g., AIDS, COVID-19). Here, we report the improved capsid structure for one of these vectors, human adenovirus D26 (HAdV-D26), at 3.4 Å resolution, by reprocessing the previous cryo-electron microscopy dataset and obtaining a refined model. In addition to overall improvements in the model, the highlights of the structure include (1) locating a segment of the processed peptide of VIII that was previously believed to be released from the mature virions, (2) reorientation of the helical appendage domain (APD) of IIIa situated underneath the vertex region relative to its counterpart observed in the cleavage defective (ts1) mutant of HAdV-C5 that resulted in the loss of interactions between the APD and hexon bases, and (3) the revised conformation of the cleaved N-terminal segments of pre-protein VI (pVIn), located in the hexon cavities, is highly conserved, with notable stacking interactions between the conserved His13 and Phe18 residues. Taken together, the improved model of HAdV-D26 capsid provides a better understanding of protein-protein interactions in HAdV capsids and facilitates the efforts to modify and/or design adenoviral vectors with altered properties. Last but not least, we provide some insights into clotting factors (e.g., FX and PF4) binding to AdV vectors.

The anti-infective outcomes of the distal femoral replacement coated with antibiotic cement in limb salvage surgery: A randomized clinical trial.

BACKGROUND: The aim of this study was to observe the anti-infective effect of the distal femoral tumor prosthesis coated with antibiotic cement during limb salvage treatment, and evaluate its potential prospect in clinic. METHODS: In this randomized controlled trial, the en bloc resection and reconstruction were performed in 36 patients with distal femoral primary bone tumor. Patients were divided into 2 groups randomly according to the application of antibiotic bone cement coating, which included antibiotic cement coating group (16 cases) and control group (18 cases). There were 10 men and 6 women in anti-infection group, aged from 18 to 54 years (23.47 ±â€Š3.53), and there were 12 men and 6 women in control group, aged from 19 to 56 years (24.16 ±â€Š4.32). The tumor type, age, sex, and Enneking stage were enrolled with well-matched of the 2 groups of patients. There was no difference between bundles and routine standard care for each group. The antibiotic cement was coated on the surface of polyethylene jacket with punched holes during operation. The peri-prosthetic infection, local recurrence and distant metastasis were followed up and limb functions were evaluated by Musculoskeletal Tumor Society 93 (MSTS93) scoring system. RESULTS: Patients were followed up till 34.7 months (range 18∼62 months). There was no periprosthetic infection in anti-infection group. Four cases in control group showed deep infection. Infection rate had significant differences between the 2 groups (P < .05). Infection-related prosthesis mortality was 0% (0/16) in anti-infection group and 16.67% (3/18) in control group. Local recurrence and distant metastasis occurred in 7 of 34 patients with primary malignant bone tumor, wherein 2 cases of local recurrence and 1 cases of distant metastasis occurred in anti-infective group; 2 cases of local recurrence and 2 cases of distant metastasis occurred in the control group. During a latest follow-up, MSTS93 function scoring revealed a mean of 25.6 ±â€Š4.2 in anti-infection group and 18.5 ±â€Š3.3 in control group. The survival rate of anti-infective group is 75%, and the survival rate of control group is 61.11%. CONCLUSION: The antibiotic cement-coated technique on the surface of the polyethylene jacket of custom-made distal femoral prosthesis is simple and effective in controlling the periprosthetic infection after tumor prosthesis reconstruction.

Structure of a Cell Entry Defective Human Adenovirus Provides Insights into Precursor Proteins and Capsid Maturation.

Maturation of adenoviruses is distinguished by proteolytic processing of several interior minor capsid proteins and core proteins by the adenoviral protease and subsequent reorganization of adenovirus core. We report the results derived from the icosahedrally averaged cryo-EM structure of a cell entry defective form of adenovirus, designated ts1, at a resolution of 3.7 Å as well as of the localized reconstructions of unique hexons and penton base. The virion structure revealed the structures and organization of precursors of minor capsid proteins, pIIIa, pVI and pVIII, which are closely associated with the hexons on the capsid interior. In addition to a well-ordered helical domain (a.a. 310-397) of pIIIa, highlights of the structure include the precursors of VIII display significantly different structures near the cleavage sites. Moreover, we traced residues 4-96 of the membrane lytic protein (pVI) that includes an amphipathic helix occluded deep in the hexon cavity suggesting the possibility of co-assembly of hexons with the precursors of VI. In addition, we observe a second copy of pVI ordered up to residue L40 in the peripentonal hexons and a few fragments of density corresponding to 2nd and 3rd copies of pVI in other hexons. However, we see no evidence of precursors of VII binding in the hexon cavity. These findings suggest the possibility that differently bound pVI molecules undergo processing at the N-terminal cleavage sites at varying efficiencies, subsequently creating competition between the cleaved and uncleaved forms of VI, followed by reorganization, processing, and release of VI molecules from the hexon cavities.

Integrative structural and functional analysis of human malic enzyme 3: A potential therapeutic target for pancreatic cancer.

Malic enzymes (ME1, ME2, and ME3) are involved in cellular energy regulation, redox homeostasis, and biosynthetic processes, through the production of pyruvate and reducing agent NAD(P)H. Recent studies have implicated the third and least well-characterized isoform, mitochondrial NADP+-dependent malic enzyme 3 (ME3), as a therapeutic target for pancreatic cancers. Here, we utilized an integrated structure approach to determine the structures of ME3 in various ligand-binding states at near-atomic resolutions. ME3 is captured in the open form existing as a stable tetramer and its dynamic Domain C is critical for activity. Catalytic assay results reveal that ME3 is a non-allosteric enzyme and does not require modulators for activity while structural analysis suggests that the inner stability of ME3 Domain A relative to ME2 disables allostery in ME3. With structural information available for all three malic enzymes, the foundation has been laid to understand the structural and biochemical differences of these enzymes and could aid in the development of specific malic enzyme small molecule drugs.

Synthesis of hybrid Au-Ag2S-Cu2-xS nanocrystals with disparate interfacial features.

Hybrid nanocrystals (NCs) with multiple components and junctions have attracted considerable attention due to their promising synergistic properties. In particular, great attention has been paid to the manipulation of buried multijunction heterointerfaces because they are closely related to the surface energy and carrier transfer of NCs. However, heterointerfaced NCs are usually constructed by sequential step-by-step pathways, and buried interfaces can only be formed along a given direction, resulting in the one and only spatial orientation of multiple interfaces. In this work, we demonstrate two types of Au-Ag2S-Cu2-xS NCs with disparate interfacial features. Specifically, the first type (Type I) is prepared through a routine two-step method and shows that Au domain close to the Ag2S-Cu2-xS interface; another type (Type II) is achieved by a facile one-pot synthesis procedure and contains Au domain with an interphase only with the Ag2S domain, far from the Cu2-xS domain. More importantly, type II NCs could not be formed through other traditional strategies and an underlying mechanism of formation is developed by monitoring the evolution process. Au@Ag core-shell NCs, metastable Au@Ag2S NCs and Janus Au-Ag2S NCs are formed successively before the Cu2-xS domain appears. We speculate that the Au@Ag2S intermediate plays an essential role in building the final complex nanostructure. We expect that such a simple and facile one-pot method will be used to fabricate additional asymmetric multicomponent NCs with distinctive interfacial features and promising potential applications.

Boosting the anode performance of microbial fuel cells with a bacteria-derived biological iron oxide/carbon nanocomposite catalyst.

Modifying the electrodes of microbial fuel cells (MFCs) with iron oxides can improve the bacterial attachment performances and electrocatalytic activities for energy conversion, which is of significance in the fabrication of MFCs. However, the conventional modification methods usually result in the aggregation of iron sites, producing the electrodes of poor qualities. Herein, we report a novel method for the modification of electrochemical electrodes to boost the anode performance of MFC. The Shewanella precursor adhered on carbon felt electrode was directly carbonized to form a bacteria-derived biological iron oxide/carbon (Bio-FeOx/C) nanocomposite catalyst. The large spatial separation between the bacteria, as well as those between the iron containing proteins in the bacteria, deliver a highly dispersed Bio-FeOx/C nanocomposite with good electrocatalytic activities. The excellent microbial attachment performance and electron transfer rate of the Bio-FeOx/C modified electrode significantly promote the transfer of produced electrons between bacteria and electrode. Accordingly, the MFC with the Bio-FeOx/C electrode exhibits the maximum power density of 797.0 mW m-2, much higher than that obtained with the conventional carbon felt anode (226.1 mW m-2). Our works have paved a new avenue to the conversion of the natural bacterial precursors into active iron oxide nanoparticles as the anode catalyst of MFCs. The high catalytic activity of the prepared Bio-FeOx endows it great application potentials in the construction of high-performance electrodes.

Improving the power generation performances of Gram-positive electricigens by regulating the peptidoglycan layer with lysozyme.

The power generation performance of a microbial fuel cell (MFC) greatly depends on the relative amount of electricigens in the anodic microbial community. Running the MFC multiple times can practically enrich the electricigens, and thus improve its power generation efficiency. However, Gram-positive electricigens cannot be enriched well because of their thick non-conductive peptidoglycan layer. Herein, we report a new Gram-positive electricigen enrichment method by regulating the peptidoglycan layer of the bacteria using lysozyme. Lysozyme can partially hydrolyze the peptidoglycans layer of Gram-positive Firmicutes to improve the permeability of cell wall, and thus enhance its electricity generation activity. The stimulation of Gram-positive electricigen endows MFCs a high power generation community structure, which results in the power density 42% higher than that of the control sample. Our work has provided a new and simple method for optimizing the anode community structure by regulating weak electricigens in the community with lysozyme.

Cryo-EM structures of the human glutamine transporter SLC1A5 (ASCT2) in the outward-facing conformation.

Alanine-serine-cysteine transporter 2 (ASCT2, SLC1A5) is the primary transporter of glutamine in cancer cells and regulates the mTORC1 signaling pathway. The SLC1A5 function involves finely tuned orchestration of two domain movements that include the substrate-binding transport domain and the scaffold domain. Here, we present cryo-EM structures of human SLC1A5 and its complex with the substrate, L-glutamine in an outward-facing conformation. These structures reveal insights into the conformation of the critical ECL2a loop which connects the two domains, thus allowing rigid body movement of the transport domain throughout the transport cycle. Furthermore, the structures provide new insights into substrate recognition, which involves conformational changes in the HP2 loop. A putative cholesterol binding site was observed near the domain interface in the outward-facing state. Comparison with the previously determined inward-facing structure of SCL1A5 provides a basis for a more integrated understanding of substrate recognition and transport mechanism in the SLC1 family.

Lovastatin attenuates hypertension induced by renal tubule-specific knockout of ATP-binding cassette transporter A1, by inhibiting epithelial sodium channels.

BACKGROUND AND PURPOSE: We have shown that cholesterol is synthesized in the principal cells of renal cortical collecting ducts (CCD) and stimulates the epithelial sodium channels (ENaC). Here we have determined whether lovastatin, a cholesterol synthesis inhibitor, can antagonize the hypertension induced by activated ENaC, following deletion of the cholesterol transporter (ATP-binding cassette transporter A1; ABCA1). EXPERIMENTAL APPROACH: We selectively deleted ABCA1 in the principal cells of mouse CCD and used the cell-attached patch-clamp technique to record ENaC activity. Western blot and immunofluorescence staining were used to evaluate protein expression levels. Systolic BP was measured with the tail-cuff method. KEY RESULTS: Specific deletion of ABCA1 elevated BP and ENaC single-channel activity in the principal cells of CCD in mice. These effects were antagonized by lovastatin. ABCA1 deletion elevated intracellular cholesterol levels, which was accompanied by elevated ROS, increased expression of serum/glucocorticoid regulated kinase 1 (Sgk1), phosphorylated neural precursor cell-expressed developmentally down-regulated protein 4-2 (Nedd4-2) and furin, along with shorten the primary cilium, and reduced ATP levels in urine. CONCLUSIONS AND IMPLICATIONS: These data suggest that specific deletion of ABCA1 in principal cells increases BP by stimulating ENaC channels via a cholesterol-dependent pathway which induces several secondary responses associated with oxidative stress, activated Sgk1/Nedd4-2, increased furin expression, and reduced cilium-mediated release of ATP. As ABCA1 can be blocked by cyclosporine A, these results suggest further investigation of the possible use of statins to treat CsA-induced hypertension.