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BACKGROUND AND AIM: Postoperative complications are important clinical outcomes for colon cancer patients. This study aimed to investigate the predictive value of inflammatory-nutritional indicators combined with computed tomography body composition on postoperative complications in patients with stage II-III colon cancer. METHODS: We retrospectively collected data from patients with stage II-III colon cancer admitted to our hospital from 2017 to 2021, including 198 patients in the training cohort and 50 patients in the validation cohort. Inflammatory-nutritional indicators and body composition were included in the univariate and multivariate analyses. Binary regression was used to develop a nomogram and evaluate its predictive value. RESULTS: In the multivariate analysis, the monocyte-lymphocyte ratio (MLR), systemic immune-inflammation index (SII), nutritional risk score (NRS), skeletal muscle index (SMI), and visceral fat index (VFI) were independent risk factors for postoperative complications of stage II-III colon cancer. In the training cohort, the area under the receiver operating characteristic curve of the predictive model was 0.825 (95% confidence interval [CI] 0.764-0.886). In the validation cohort, it was 0.901 (95% CI 0.816-0.986). The calibration curve showed that the prediction results were in good agreement with the observational results. Decision curve analysis showed that colon cancer patients could benefit from the predictive model. CONCLUSIONS: A nomogram combining MLR, SII, NRS, SMI, and VFI with good accuracy and reliability in predicting postoperative complications in patients with stage II-III colon cancer was established, which can help guide treatment decisions.
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Neoplasias do Colo , Humanos , Reprodutibilidade dos Testes , Estudos Retrospectivos , Neoplasias do Colo/diagnóstico por imagem , Neoplasias do Colo/cirurgia , Composição Corporal , Inflamação/diagnóstico por imagem , Inflamação/etiologia , Nomogramas , Complicações Pós-Operatórias/diagnóstico por imagem , Complicações Pós-Operatórias/etiologia , TomografiaRESUMO
BACKGROUND: Infectious endocarditis (IE) is a disease caused by the colonization of toxic microorganisms on the endocardium of heart valves [1]. Although much progress has been made in the diagnosis and treatment of IE, its complications, such as annular abscesses [2], still have a high mortality rate. In this case, we describe a patient with infective endocarditis complicated by occult deteriorated aortic annular abscess. CASE PRESENTATION: A 44-year-old man was admitted due to weakness of his right limbs and unclear speech for 10 h. He had recurrent fevers for 1 month before admission. Transthoracic echocardiography showed a mix-echoic vegetation attached to the bicuspid aortic valve, moderate aortic regurgitation and a possible aortic annular abscess. Blood cultures were negative and empiric antibiotic therapy was begun. The patient did not have fever again and seem to be clinically improved. However, follow-up transesophageal echocardiography revealed a large periaortic abscess led to aortic sinus pseudoaneurysm. The patient underwent mechanical prosthetic valve replacement and annulus reconstruction successfully. Perivalvular abscess may be insidious deterioration in patients who seem to be clinically improved, which requires us to pay more attention. DISCUSSION: Occult deterioration of an aortic annular abscess is rare and more attention should be paid. Re-evaluation of echocardiography is required even if the patient's symptoms improve.
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Falso Aneurisma , Doença da Válvula Aórtica Bicúspide , Masculino , Humanos , Adulto , Abscesso/diagnóstico por imagem , Abscesso/etiologia , Abscesso/terapia , Falso Aneurisma/diagnóstico por imagem , Falso Aneurisma/etiologia , Falso Aneurisma/terapia , Catéteres , AortaRESUMO
Complement component C3, mainly synthesized by hepatocytes, acts as the convergence point of three different pathways in activating the complement cascade. Besides its well-established roles in the extracellular milieu, C3 performs various intracellular functions such as immunomodulation and pathogen recognition. Although C3 is present at extremely high concentrations in hepatocytes, little is known about its intrahepatic function. In this study, we found that C3 knockout (C3-/- ) mice displayed accelerated hepatic triglyceride (TG) accumulation compared with C57BL/6J wild type mice. Mechanistically, C3 deficiency impaired lipophagy in hepatocytes, owing to the disrupted interaction between C3 and autophagy-related 16 like 1, which is essential for autolysosome assembly. Furthermore, lipophagy deficiency affected the function of the endoplasmic reticulum in C3-/- mice, subsequently affecting the expression of protein disulfide isomerase and activity of microsomal TG transfer protein, and ultimately impairing the production of hepatic very-low-density lipoproteins (VLDLs). Rapamycin and thapsigargin treatment accelerated VLDL secretion and alleviated hepatic lipid accumulation in C3-/- mice. Our study demonstrates that C3 promotes lipophagy to facilitate VLDL secretion in hepatocytes, thus playing an essential role in balancing TG levels in the liver.
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Proteínas Relacionadas à Autofagia/metabolismo , Autofagia , Complemento C3/fisiologia , Lipoproteínas VLDL/metabolismo , Hepatopatia Gordurosa não Alcoólica/prevenção & controle , Sirolimo/farmacologia , Animais , Proteínas Relacionadas à Autofagia/genética , Imunossupressores/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Hepatopatia Gordurosa não Alcoólica/etiologia , Hepatopatia Gordurosa não Alcoólica/metabolismo , Hepatopatia Gordurosa não Alcoólica/patologiaRESUMO
Chronic exposure to high concentrations of stearic acid (C18:0) can result in ß-cell dysfunction, leading to development of type 2 diabetes. However, the molecular mechanisms underlying the destructive effects of stearic acid on ß-cells remain largely unknown. In this study, we aimed to investigate the role of miR-297b-5p on stearic acid-induced ß-cell apoptosis. Differential expression of microRNAs (miRNAs) was assessed in a ß-TC6 cell line exposed to stearic acid, palmitic acid, or a normal culture medium by high-throughput sequencing. The apoptosis rate was measured by flow cytometry after miR-297b-5p mimic/inhibitor transfection, and large-tumor suppressor kinase 2 (LATS2) was identified as a target of miR-297b-5p using a luciferase activity assay. In vivo, C57BL/6 mice were fed with normal and high-stearic-acid diet, respectively. Mouse islets were used for similar identification of miR-297b-5p and Lats2 in ß-TC6 cell. We selected two differentially expressed miRNAs in stearic acid compared with those in the palmitic acid and control groups. miR-297b-5p expression was significantly lower in ß-TC6 cells and mouse islets in stearic acid than in control group. Upregulation of miR-297b-5p alleviated the stearic acid-induced cell apoptosis and reduction in insulin secretion by inhibiting Lats2 expression in vitro. Meanwhile, silencing Lats2 significantly reversed the stearic acid-stimulated ß-cell dysfunction in both ß-TC6 cells and islets. Our findings indicate a suppressive role for miR-297b-5p in stearic acid-induced ß-cell apoptosis, which may reveal a potential target for the treatment of ß-cell dysfunction in the pathogenesis of type 2 diabetes.
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Apoptose/efeitos dos fármacos , Células Secretoras de Insulina/efeitos dos fármacos , MicroRNAs/metabolismo , Proteínas Serina-Treonina Quinases/genética , Ácidos Esteáricos/farmacologia , Proteínas Supressoras de Tumor/genética , Animais , Linhagem Celular , Células Cultivadas , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/metabolismo , Ácidos Graxos não Esterificados/sangue , Citometria de Fluxo , Humanos , Secreção de Insulina/efeitos dos fármacos , Secreção de Insulina/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ácido Palmítico/metabolismo , Regulação para Cima/efeitos dos fármacosRESUMO
PURPOSE: Sepsis continues to be a critical issue in intensive care, characterized by significant morbidity and mortality. This study explores the association between Vasoactive Inotrope Score (VIS) trajectories and 28-day mortality in ICU patients with sepsis, employing VIS trajectories as a marker for assessing severity and guiding therapy. METHODS: We conducted a retrospective analysis of the MIMIC-IV database, which included sepsis patients admitted to the ICU between 2008 and 2019. VIS calculations were performed bi-hourly during the first 72 hours of ICU admission. Using latent growth mixture modeling, we identified distinct VIS trajectory patterns, and multivariate Cox proportional hazards models were employed to evaluate their association with 28-day mortality. FINDINGS: Among 6,802 sepsis patients who met the inclusion criteria, four distinct VIS trajectory patterns were identified: "Low-Decreasing" (52.1%), "Mild-Ascending" (13.2%), "Moderate-Decreasing" (23.0%), and "High-Stable" (11.6%). The 28-day survival analysis demonstrated that, compared to the "Low-Decreasing" group, the "Mild-Ascending" group had a hazard ratio (HR) for mortality of 2.55 (95% CI: 2.19-2.97, P < 0.001), the "Moderate-Decreasing" group had an HR of 1.20 (95% CI: 1.03-1.41, P = 0.021), and the "High-Stable" group presented the highest risk with an HR of 4.19 (95% CI: 3.43-5.12, P < 0.001). IMPLICATIONS: This study offers significant insights into the prognostic value of VIS trajectories in sepsis patients. The identification of distinct trajectory patterns not only underscores the heterogeneity in sepsis but also emphasizes the importance of personalized management strategies. The findings underscore the potential of VIS trajectory monitoring in predicting 28-day outcomes and in guiding clinical decision-making in ICU settings.
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Native functionality directed the C-H activation cascade to enable rapid construction of molecular complexity, featuring step-economy and synthetic efficiency. Herein, by exploiting bifunctional α-alcohol haloalkynes, we developed Ru(II)-catalyzed carboxylic acid, amine, and amide assisted divergent C-H alkynylation and annulation cascade, affording polyfunctional heterocycles. Significantly, a bilateral aryl C-H polycyclization cascade of azobenzenes was achieved using the versatile haloalkynes.
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PURPOSE: The purpose of this study is to evaluate the impact of pre- and postoperative pulmonary-related training, including respiratory and aerobic training, on postoperative pulmonary complications (PPCs) after radical resection of gastric cancer (GC). METHODS: We conducted a randomized controlled trial of 121 participants who received a systematic, comprehensive, high-intensity pulmonary exercise regimen for 5 days preoperatively and at least 5 days postoperatively. The control group received standard preoperative care. We analyzed the occurrence of PPCs in both groups as the primary outcome RESULTS: 43 out of 121 participants (35.5%) were diagnosed with PPCs, and the incidence of PPCs was markedly lower in the training group than in the control group (26.2%, 16/61 vs. 45.0%, 27/60, P= 0.031). Moreover, the duration of the postoperative hospital stay was shorter in the training group (8.69 ± 1.92 days) compared to the control group (9.57 ± 2.16 days, P=0.020) as well as reduced hospitalization costs, amounting to 9605.1 ± 2556.26 in the training group and 10594.6 ± 2560.7 in the control group (P=0.035). CONCLUSION: Our study established that a perioperative pulmonary-related training notably diminishes the incidence of PPCs, curtails the duration of hospitalization, and mitigates hospitalization expenses for patients undergoing gastric cancer surgery. CLINICAL TRIAL REGISTRATION: ChiCTR2400081853.
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BACKGROUND: Venetoclax (Ven) combined with intensive chemotherapy was proven effective in the management of acute myeloid leukemia (AML). However, the severe and prolonged myelosuppression remains a concern to worry about. To explore more appropriate combination regimens, we designed Ven combining daunorubicin and cytarabine (DA 2 + 6) regimen as induction therapy, aimed to evaluate the effectiveness and safety in adults de novo AML. METHODS: A phase 2 clinical trial was performed in 10 Chinese hospitals to investigate Ven combined with daunorubicin and cytarabine (DA 2 + 6) in patients with AML. The primary endpoints were overall response rate (ORR), comprising of complete remission (CR), complete remission with incomplete blood cell count recovery (CRi), and partial response (PR). Secondary endpoints included measurable residual disease (MRD) of bone marrow assessed by flow cytometry, overall survival (OS), event-free survival (EFS), disease-free survival (DFS), and the safety of regimens. This study is a currently ongoing trial listed on the Chinese Clinical Trial Registry as ChiCTR2200061524. RESULTS: Overall, 42 patients were enrolled from January 2022 to November 2022; 54.8% (23/42) were male, and the median age was 40 (range, 16-60) years. The ORR after one cycle of induction was 92.9% (95% confidence interval [CI], 91.6-94.1; 39/42) with a composite complete response rate (CR + CRi) 90.5% (95% CI, 89.3-91.6, CR 37/42, CRi 1/42). Moreover, 87.9% (29/33) of the CR patients with undetectable MRD (95% CI, 84.9-90.8). Grade 3 or worse adverse effects included neutropenia (100%), thrombocytopenia (100%), febrile neutropenia (90.5%), and one mortality. The median neutrophil and platelet recovery times were 13 (5-26) and 12 (8-26) days, respectively. Until Jan 30, 2023, the estimated 12-month OS, EFS, and DFS rates were 83.1% (95% CI, 78.8-87.4), 82.7% (95% CI, 79.4-86.1), and 92.0% (95% CI, 89.8-94.3), respectively. CONCLUSION: Ven with DA (2 + 6) is a highly effective and safe induction therapy for adults with newly diagnosed AML. To the best of our knowledge, this induction therapy has the shortest myelosuppressive period but has similar efficacy to previous studies.
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Rationale: Invariant natural killer T (iNKT) cells and Kupffer cells represent major hepatic populations of innate immune cells. However, their roles in steatohepatitis remain poorly understood. To elucidate their functions in steatohepatitis development, real-time, in vivo analysis is necessary to understand the pathophysiological events in the dynamic interactions between them during diet-induced steatohepatitis. Methods: We used a steatohepatitis animal model induced by a methionine-choline-deficient (MCD) diet. Multi-photon confocal live imaging and conventional experimental techniques were employed to investigate the hepatic pathological microenvironment of iNKT and Kupffer cells, interactions between them, and the biological effects of these interactions in steatohepatitis. Results: We found that iNKT cells were recruited and aggregated into small clusters and interacted dynamically with Kupffer cells in the early stage of steatohepatitis. Most significantly, the iNKT cells in the cluster cleared free lipids released by necrotic hepatocytes and presented a non-classical activation state with high IFN-γ expression. Furthermore, the Kupffer cells in the cell cluster were polarized to type M1. The transcriptome sequencing of iNKT cells showed upregulation of genes related to phagocytosis and lipid processing. Adoptive transfer of iNKT cells to Jα18-/- mice showed that iNKT and Kupffer cell clusters were essential for balancing the liver and peripheral lipid levels and inhibiting liver fibrosis development. Conclusions: Our study identified an essential role for dynamic interactions between iNKT cells and Kupffer cells in promoting lipid phagocytosis and clearance by iNKT cells during early liver steatohepatitis. Therefore, modulating iNKT cells is a potential therapeutic strategy for early steatohepatitis.
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Modelos Animais de Doenças , Microscopia Intravital/métodos , Células de Kupffer/patologia , Lipídeos/química , Células T Matadoras Naturais/patologia , Hepatopatia Gordurosa não Alcoólica/patologia , Animais , Células de Kupffer/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Células T Matadoras Naturais/metabolismo , Hepatopatia Gordurosa não Alcoólica/etiologia , Hepatopatia Gordurosa não Alcoólica/metabolismoRESUMO
Iron is one of the most important trace elements in the body, and its homeostasis is essential to the normal function of the immune system. Complement component C3, which is the converging of three main pathways of complement system activation, plays a key role in the innate immunity. However, the relationship between iron homeostasis and complement C3 remains unknown. The aim of our study was to analyze the relationship between serum iron and ferritin level and complement C3 and C4. A total of 590 healthy individuals were recruited in our study. Higher serum complement C3 level (p < 0.001) was found in individuals with higher serum ferritin level (> 104.0 µg/L). Moreover, serum iron level and serum ferritin level were positively correlated with complement C3 (r = 0.133, p = 0.001; r = 0.221, p < 0.001) and complement C4 (r = 0.117, p = 0.004; r = 0.123, p = 0.003). The linear regression analysis displayed that both serum iron level and serum ferritin level were linearly correlated with serum complement C3 level (adjusted beta: 2.382, 95% CI: 0.841-3.923; adjusted beta: 42.911, 95% CI: 29.070-56.751). To explore the relationship between iron homeostasis and complement C3 further, the serum samples from C3-/- mice and the wild-type (WT) control mice were obtained. Significantly lower serum iron level and higher ferritin level were found in C3-/- mice than those in WT mice (p < 0.001; p < 0.001), indicating that complement C3 might influence iron distribution and utilization. Overall, these data suggested that serum iron and ferritin levels were correlated with complement C3. The deficiency of complement C3 may disrupt the regular iron metabolism in the body.
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Complemento C3 , Complemento C4 , Animais , Complemento C3/metabolismo , Complemento C4/metabolismo , Ferritinas , Ferro , CamundongosRESUMO
The health effect of dietary fat has been one of the most vexing issues in the field of nutrition. Few animal studies have examined the impact of high-fat diets on lifespan by controlling energy intake. In this study, we found that compared to a normal diet, an isocaloric moderately high-fat diet (IHF) significantly prolonged lifespan by decreasing the profiles of free fatty acids (FFAs) in serum and multiple tissues via downregulating FFA anabolism and upregulating catabolism pathways in rats and flies. Proteomics analysis in rats identified PPRC1 as a key protein that was significantly upregulated by nearly 2-fold by IHF, and among the FFAs, only palmitic acid (PA) was robustly and negatively associated with the expression of PPRC1. Using PPRC1 transgenic RNAi/overexpression flies and in vitro experiments, we demonstrated that IHF significantly reduced PA, which could upregulate PPRC1 through PPARG, resulting in improvements in oxidative stress and inflammation and prolonging the lifespan.
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Gorduras na Dieta/farmacologia , Longevidade/efeitos dos fármacos , Ácido 3-Hidroxibutírico/farmacologia , Animais , Drosophila , Proteínas de Drosophila/antagonistas & inibidores , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Ácidos Graxos não Esterificados/sangue , Ácidos Graxos não Esterificados/metabolismo , Fígado/metabolismo , Masculino , Obesidade/patologia , Estresse Oxidativo/efeitos dos fármacos , PPAR gama/antagonistas & inibidores , PPAR gama/genética , PPAR gama/metabolismo , Ácido Palmítico/metabolismo , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Ratos , Ratos Wistar , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Regulação para Cima/efeitos dos fármacosRESUMO
With the aggravation of the energy crisis, increasing attention has been paid to electrocatalytic technology for renewable energy devices. In particular, the research on catalysts towards the oxygen evolution reaction (OER), oxygen reduction reaction (ORR), and hydrogen evolution reaction (HER) has become more urgent, and the development of multifunctional electrocatalysts has become a research trend. Here we report the synthesis of waxberry-like cobalt-nickel oxide/S,N-codoped carbon hollow nanocomposites as trifunctional catalysts. Uniform cobalt-nickel glycerate solid spheres are first synthesized as the precursor and subsequently chemically transformed into cobalt-nickel oxide/S,N-codoped carbon hollow nanospheres. Benefiting from the synergistic coupling of cobalt-nickel oxide and S,N-codoped carbon nanocomposites, hierarchical porosity and hollow structure, the cobalt-nickel oxide/S,N-codoped carbon nanohybrids exhibit superior trifunctional electrocatalytic activity and durability towards OER, ORR, and HER in alkaline media.
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BACKGROUND/OBJECTIVES: The purpose of our study was to explore the relation of prenatal exposure to the Chinese famine and the risk of metabolic syndrome in adulthood in consecutive generations. SUBJECTS/METHODS: A total of 960 families, including 1920 parents (F1) who were born near the time of the Great Chinese famine and 1145 of their offspring (F2), were selected from the Suihua rural area. Parental participants were defined as nonexposed (born between 1 October 1956 and 30 September 1958 or between 1 October 1962 and 30 September 1964) and famine exposed (born between 1 October 1959 and 30 September 1961). In F2, participants were divided into having no parents exposed to famine, only a mother exposed to famine, only a father exposed to famine or both parents exposed to famine. Metabolic syndrome was defined by the International Diabetes Federation criteria. RESULTS: Exposure to famine during gestation was associated with an increased risk of metabolic syndrome (odds ratio: 2.79, 95% confidence interval: 2.16, 3.60) in F1 adults. However, similar association was not observed (maternal: odds ratio: 1.33, 95% confidence interval: 0.69, 2.52; paternal: odds ratio: 1.67, 95% confidence interval: 0.87, 3.21; parental: odds ratio: 1.25, 95% confidence interval: 0.68, 2.31) in F2 adults. CONCLUSIONS: Exposure to the Chinese famine during foetal life was associated with an increased risk of metabolic syndrome in F1 participants. In F2 adults, there might be an important relationship between exposure to famine and the risk of metabolic syndrome with increasing age.
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Síndrome Metabólica , Efeitos Tardios da Exposição Pré-Natal , Inanição , Adulto , China/epidemiologia , Fome Epidêmica , Feminino , Humanos , Síndrome Metabólica/epidemiologia , Síndrome Metabólica/etiologia , Gravidez , Efeitos Tardios da Exposição Pré-Natal/epidemiologia , Fatores de RiscoRESUMO
Methods are described to determine the structures of viral membrane fusion domains in detergent micelles by NMR and in lipid bilayers by site-directed spin labeling and EPR spectroscopy. Since in favorable cases, the lower-resolution spin label data obtained in lipid bilayers fully support the higher-resolution structures obtained by solution NMR, it is possible to graft the NMR structural coordinates into membranes using the EPR-derived distance restraints to the lipid bilayer. Electron paramagnetic dynamics and distance measurements in bilayers support conclusions drawn from NMR in detergent micelles. When these methods are applied to a structure determination of the influenza virus fusion domain and four point mutations with different functional phenotypes, it is evident that a fixed-angle boomerang structure with a glycine edge on the outside of the N-terminal arm is both necessary and sufficient to support membrane fusion. The human immunodeficiency virus fusion domain forms a straight helix with a flexible C-terminus. While EPR data for this fusion domain are not yet available, it is tentatively speculated that, because of its higher hydrophobicity, a critically tilted insertion may occur even in the absence of a kinked boomerang structure in this case.
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Espectroscopia de Ressonância de Spin Eletrônica/métodos , Espectroscopia de Ressonância Magnética/métodos , Proteína gp41 do Envelope de HIV/química , Glicoproteínas de Hemaglutininação de Vírus da Influenza/química , Humanos , Bicamadas Lipídicas/química , Micelas , Estrutura Terciária de ProteínaRESUMO
Near infrared diffuse reflectance spectra of 50 tobacco samples were pretreated with PCA. The calibration models of determination of the main components in tobacco were developed with support v ector regression (SVR). The models weretested with leave-one-out (LOOCV) method and optimized with parameters of kernel function, penalty coefficient C and insensitive loss function. The root mean square errors (RMSE) with leave-one-out cross validation of the optimal models of nicotine, and total sugars, reductive sugar, and total nitrogen were 0.313, 1.581, 1.412 and 0.117 respectively. Based on the comparison of RMSE of the SVM model with those of the partial least square (PLS), multiplicative linear regression (MLR) and back propagation artificial neuron network (BP-ANN) models, it was found that the SVR model was the most robust one. This study suggested that it is feasible to rapidly determine the main components concentrations by near infrared spectroscopy method based on SVR.
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Nicotiana/química , Extratos Vegetais/análise , Espectroscopia de Luz Próxima ao Infravermelho/métodos , Computação Matemática , Folhas de Planta/química , Análise de Componente PrincipalRESUMO
The human immunodeficiency virus (HIV) gp41 fusion domain plays a critical role in membrane fusion during viral entry. A thorough understanding of the relationship between the structure and the activity of the fusion domain in different lipid environments helps to formulate mechanistic models on how it might function in mediating membrane fusion. The secondary structure of the fusion domain in small liposomes composed of different lipid mixtures was investigated by circular dichroism spectroscopy. The fusion domain formed an α-helix in membranes containing less than 30 mol% cholesterol and formed ß-sheet secondary structure in membranes containing ≥30 mol% cholesterol. EPR spectra of spin-labeled fusion domains also indicated different conformations in membranes with and without cholesterol. Power saturation EPR data were further used to determine the orientation and depth of α-helical fusion domains in lipid bilayers. Fusion and membrane perturbation activities of the gp41 fusion domain were measured by lipid mixing and contents leakage. The fusion domain fused membranes in both its helical form and its ß-sheet form. High cholesterol, which induced ß-sheets, promoted fusion; however, acidic lipids, which promoted relatively deep membrane insertion as an α-helix, also induced fusion. The results indicate that the structure of the HIV gp41 fusion domain is plastic and depends critically on the lipid environment. Provided that their membrane insertion is deep, α-helical and ß-sheet conformations contribute to membrane fusion.
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Colesterol/química , Proteína gp41 do Envelope de HIV/química , Sequência de Aminoácidos , Dicroísmo Circular , Proteína gp41 do Envelope de HIV/síntese química , Humanos , Bicamadas Lipídicas/química , Lipídeos/química , Lipossomos/química , Fusão de Membrana , Dados de Sequência Molecular , Estrutura Secundária de Proteína , Análise de SequênciaRESUMO
A thorough understanding of the structure of fusion domains of enveloped viruses in changing lipid environments helps us to formulate mechanistic models on how they might function in mediating viral entry by membrane fusion. We have expressed the N-terminal fusion domain of HIV-1 gp41 as a construct that is water-soluble in the absence of membranes, but that also binds with high affinity to lipid micelles and bilayers in their presence. We have solved the structure and studied the dynamics of this domain bound to dodecylphosphocholine micelles by homo- and heteronuclear NMR spectroscopy. The fusion peptide forms a stable hydrophobic helix from Ile(4) to Ala(14), but is increasingly more disordered and dynamic in a segment of intermediate polarity that stretches from Ala(15) to Ser(23). When bound to lipid bilayers at low concentration, the HIV fusion domain is also largely alpha-helical, as determined by CD and FTIR spectroscopy. However, at higher protein/lipid ratios, the domain is partially converted to form beta-structures in lipid bilayers. Controlled lipid mixing occurs at concentrations that support the alpha-helical, but not the beta-strand conformation.
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Proteína gp41 do Envelope de HIV/química , Bicamadas Lipídicas , Sítios de Ligação , Dicroísmo Circular , HIV/química , HIV/isolamento & purificação , Lipossomos , Espectroscopia de Ressonância Magnética , Micelas , Ligação Proteica , Espectroscopia de Infravermelho com Transformada de Fourier , Proteínas Virais de Fusão/químicaRESUMO
Influenza virus hemagglutinin (HA)-mediated membrane fusion is initiated by a conformational change that releases a V-shaped hydrophobic fusion domain, the fusion peptide, into the lipid bilayer of the target membrane. The most N-terminal residue of this domain, a glycine, is highly conserved and is particularly critical for HA function; G1S and G1V mutant HAs cause hemifusion and abolish fusion, respectively. We have determined the atomic resolution structures of the G1S and G1V mutant fusion domains in membrane environments. G1S forms a V with a disrupted "glycine edge" on its N-terminal arm and G1V adopts a slightly tilted linear helical structure in membranes. Abolishment of the kink in G1V results in reduced hydrophobic penetration of the lipid bilayer and an increased propensity to form beta-structures at the membrane surface. These results underline the functional importance of the kink in the fusion peptide and suggest a structural role for the N-terminal glycine ridge in viral membrane fusion.
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Glicoproteínas de Hemaglutininação de Vírus da Influenza/química , Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Vírus da Influenza A/genética , Vírus da Influenza A/fisiologia , Fusão de Membrana/genética , Fusão de Membrana/fisiologia , Proteínas Virais de Fusão/química , Proteínas Virais de Fusão/genética , Glicina/química , Glicoproteínas de Hemaglutininação de Vírus da Influenza/fisiologia , Técnicas In Vitro , Vírus da Influenza A/patogenicidade , Bicamadas Lipídicas , Espectroscopia de Ressonância Magnética , Modelos Biológicos , Modelos Moleculares , Conformação Proteica , Estrutura Terciária de Proteína , Espectroscopia de Infravermelho com Transformada de Fourier , Proteínas Virais de Fusão/fisiologiaRESUMO
The fusion peptides of viral membrane fusion proteins play a key role in the mechanism of viral spike glycoprotein mediated membrane fusion. These peptides insert into the lipid bilayers of cellular target membranes where they adopt mostly helical secondary structures. To better understand how membranes may be converted to high-energy intermediates during fusion, it is of interest to know how much energy, enthalpy and entropy, is provided by the insertion of fusion peptides into lipid bilayers. Here, we describe a detailed thermodynamic analysis of the binding of analogues of the influenza hemagglutinin fusion peptide of different lengths and amino acid compositions. In small unilamellar vesicles, the interaction of these peptides with lipid bilayers is driven by enthalpy (-16.5 kcal/mol) and opposed by entropy (-30 cal mol(-1) K(-1)). Most of the driving force (deltaG = -7.6 kcal/mol) comes from the enthalpy of peptide insertion deep into the lipid bilayer. Enthalpic gains and entropic losses of peptide folding in the lipid bilayer cancel to a large extent and account for only about 40% of the total binding free energy. The major folding event occurs in the N-terminal segment of the fusion peptide. The C-terminal segment mainly serves to drive the N-terminus deep into the membrane. The fusion-defective mutations G1S, which causes hemifusion, and particularly G1V, which blocks fusion, have major structural and thermodynamic consequences on the insertion of fusion peptides into lipid bilayers. The magnitudes of the enthalpies and entropies of binding of these mutant peptides are reduced, their helix contents are reduced, but their energies of self-association at the membrane surface are increased compared to the wild-type fusion peptide.