COPD symptoms in the morning: impact,evaluation and management

Chronic obstructive pulmonary disease (COPD) symptoms in the morning, including dyspnea and sputum production, affect patients’ quality of life and limit their ability to carry out even simple morning activities. It is now emerging that these symptoms are associated with increased risk of exacerbations and work absenteeism, suggesting that they have a more profound impact on patients than previously thought. The development of validated patient-reported outcome (PRO) questionnaires to capture patients’ experience of COPD symptoms in the morning is, therefore, vital for establishing effective and comprehensive management strategies. Although it is well established that long-acting bronchodilators are effective in improving COPD symptoms, the limited available data on their impact on morning symptoms and activities have been obtained with non-validated PRO questionnaires. In this review, we discuss the impact of COPD symptoms in the morning and available tools used to evaluate them, and highlight specific gaps that need to be addressed to develop standardized instruments able to meet regulatory requirement. We also present available evidence on the effect of pharmacological therapies on morning symptoms.     

Safety and tolerability of once-daily umeclidinium/vilanterol 125/25 mcg and umeclidinium 125 mcg in patients with chronic obstructive pulmonary disease: results from a 52-week,randomized, double-blind,placebo-controlled study

Background

The long-acting muscarinic antagonist (LAMA) umeclidinium (UMEC) and the combination of UMEC with the long-acting β₂-agonist (LABA) vilanterol (UMEC/VI) are approved maintenance treatments for chronic obstructive pulmonary disease (COPD) in the US and EU. They are not indicated for the treatment of asthma.

Methods

In this 52-week, double-blind, placebo-controlled, parallel-group safety study (GSK study DB2113359; {"type":"clinical-trial","attrs":{"text":"NCT01316887","term_id":"NCT01316887"}}NCT01316887), patients were randomized 2:2:1 to UMEC/VI 125/25 mcg, UMEC 125 mcg, or placebo. Study endpoints included adverse events (AEs), clinical chemistry and hematology parameters, vital signs, 12-lead, and 24-hour Holter electrocardiograms. COPD exacerbations and rescue medication use were assessed as safety parameters; lung function was also evaluated.

Results

The incidence of on-treatment AEs, serious AEs (SAEs), and drug-related AEs was similar between treatment groups (AEs: 52–58%; SAEs: 6–7%; drug-related AEs: 12–13%). Headache was the most common AE in each treatment group (8–11%). AEs associated with the LAMA and LABA pharmacologic classes occurred at a low incidence across treatment groups. No clinically meaningful effects on vital signs or laboratory assessments were reported for active treatments versus placebo. The incidences of atrial arrhythmias with UMEC/VI 125/25 mcg were similar to placebo; for UMEC 125 mcg, the incidences of ectopic supraventricular beats, sustained supraventricular tachycardia, and ectopic supraventricular rhythm were ≥2% greater than placebo. With active treatments, COPD exacerbations were fewer (13–15% of patients reporting ≥1 exacerbation) and on average less rescue medication was required (1.6–2.2 puffs/day) versus placebo (24% reporting ≥1 exacerbation, 2.6 puffs/day). Both active treatments improved lung function versus placebo.

Conclusion

UMEC/VI 125/25 mcg and UMEC 125 mcg were well tolerated over 12 months in patients with COPD.     

Dose-finding evaluation of once-daily treatment with olodaterol,a novel long-acting β2-agonist,in patients with asthma: results of a parallel-group study and a crossover study

Background

Olodaterol is a novel, inhaled long-acting β₂-agonist (LABA) with >24-hour duration of action investigated in asthma and chronic obstructive pulmonary disease.

Methods

Two multicentre studies examined the efficacy and safety of 4 weeks’ once-daily (QD) olodaterol (2, 5, 10 and 20 μg, with background inhaled corticosteroids) in patients with asthma. One randomised, double-blind, parallel-group study (1222.6; 296 patients) administered treatment in the morning. Pulmonary function tests (PFTs) were performed pre-dose (trough) and ≤3 hours post-dose (weeks 1 and 2), and ≤6 hours post-dose after 4 weeks; primary end point was trough forced expiratory volume in 1 second (FEV₁) response (change from baseline mean FEV₁) after 4 weeks. A second randomised, double-blind, placebo- and active-controlled (formoterol 12 μg twice-daily) incomplete-block crossover study (1222.27; 198 patients) administered QD treatments in the evening. PFTs were performed over a 24-hour dosing interval after 4 weeks; primary end point was FEV₁ area under the curve from 0–24 hours (AUC_0–24) response (change from study baseline [mean FEV₁] after 4 weeks).

Results

Study 1222.6 showed a statistically significant increase in trough FEV₁ response with olodaterol 20 μg (0.147 L; 95 % confidence interval [CI]: 0.059, 0.234; p = 0.001) versus placebo, with more limited efficacy and no evidence of dose response compared to placebo across the other olodaterol doses (2, 5 and 10 μg). Study 1222.27 demonstrated increases in FEV₁ AUC_0–24 responses at 4 weeks with all active treatments (p < 0.0001); adjusted mean (95 % CI) differences from placebo were 0.140 (0.097, 0.182), 0.182 (0.140, 0.224), 0.205 (0.163, 0.248) and 0.229 (0.186, 0.272) L for olodaterol 2, 5, 10 and 20 μg, respectively, and 0.169 (0.126, 0.211) for formoterol, providing evidence of increased efficacy with higher olodaterol dose. Olodaterol was generally well tolerated, with a few events associated with known sympathomimetic effects, mainly with 20 μg.

Conclusions

The LABA olodaterol has >24-hour duration of action. In patients with asthma, evidence of bronchodilator efficacy was demonstrated with statistically and clinically significant improvements in the primary end point of trough FEV₁ response measured in clinics over placebo for the highest administered dose of 20 μg in Study 1222.6, and statistically and clinically significant improvements versus placebo in FEV₁ AUC_0–24 responses at 4 weeks for all doses tested in Study 1222.27, which also exhibited a dose response. Bronchodilator efficacy was seen over placebo for all olodaterol doses for morning and evening peak expiratory flow in both studies. All doses were well tolerated.

Trial registrations

{"type":"clinical-trial","attrs":{"text":"NCT00467740","term_id":"NCT00467740"}}NCT00467740 (1222.6) and {"type":"clinical-trial","attrs":{"text":"NCT01013753","term_id":"NCT01013753"}}NCT01013753 (1222.27).

Electronic supplementary material

The online version of this article (doi:10.1186/s12931-015-0249-8) contains supplementary material, which is available to authorized users.     

Effect of β2-adrenergic receptor gene (ADRB2) 3′ untranslated region polymorphisms on inhaled corticosteroid/long-acting β2-adrenergic agonist response

Background

Evidence suggests that variation in the length of the poly-C repeat in the 3′ untranslated region (3′UTR) of the β₂-adrenergic receptor gene (ADRB2) may contribute to interindividual variation in β-agonist response. However, methodology in previous studies limited the assessment of the effect of sequence variation in the context of poly-C repeat length. The objectives of this study were to design a novel genotyping method to fully characterize sequence variation in the ADRB2 3′UTR poly-C repeat in asthma patients treated with inhaled corticosteroid and long-acting β₂-adrenergic agonist (ICS/LABA) combination therapy, and to analyze the effect of the poly-C repeat polymorphism on clinical response.

Methods

In 2,250 asthma patients randomized to treatment with budesonide/formoterol or fluticasone/salmeterol in a six-month study (AstraZeneca study code: SD-039-0735), sequence diversity in the ADRB2 poly-C repeat region was determined using a novel sequencing-based genotyping method. The relationship between the poly-C repeat polymorphism and the incidence of severe asthma exacerbations, and changes in pulmonary function and asthma symptoms from baseline to the average during the treatment period, were analyzed.

Results

Poly-C repeat genotypes were assigned in 97% (2,192/2,250) of patients. Of the 13 different poly-C repeat alleles identified, six alleles occurred at a frequency of >5% in one or more population in this study. The repeat length of these six common alleles ranged from 10 to 14 nucleotides. Twelve poly-C repeat genotypes were observed at a frequency of >1%. No evidence of an association between poly-C repeat genotype and the incidence of severe asthma exacerbations was observed. Patients’ pulmonary function measurements improved and asthma symptoms declined when treated with ICS/LABA combination therapy regardless of poly-C repeat genotype.

Conclusions

The extensive sequence diversity present in the poly-C repeat region of the ADRB2 3′UTR did not predict therapeutic response to ICS/LABA therapy.     

Efficacy and safety of fixed-dose combinations of aclidinium bromide/formoterol fumarate: the 24-week,randomized, placebo-controlled AUGMENT COPD study

Background

Combining two long-acting bronchodilators with complementary mechanisms of action may provide treatment benefits to patients with chronic obstructive pulmonary disease (COPD) that are greater than those derived from either treatment alone. The efficacy and safety of a fixed-dose combination (FDC) of aclidinium bromide, a long-acting muscarinic antagonist, and formoterol fumarate, a long-acting β₂-agonist, in patients with moderate to severe COPD are presented.

Methods

In this 24-week double-blind study, 1692 patients with stable COPD were equally randomized to twice-daily treatment with FDC aclidinium 400 μg/formoterol 12 μg (ACL400/FOR12 FDC), FDC aclidinium 400 μg/formoterol 6 μg (ACL400/FOR6 FDC), aclidinium 400 μg, formoterol 12 μg, or placebo administered by a multidose dry powder inhaler (Genuair^®/Pressair^®)*. Coprimary endpoints were change from baseline to week 24 in 1-hour morning postdose FEV₁ (FDCs versus aclidinium) and change from baseline to week 24 in morning predose (trough) FEV₁ (FDCs versus formoterol). Secondary endpoints were change from baseline in St. George’s Respiratory Questionnaire (SGRQ) total score and improvement in Transition Dyspnea Index (TDI) focal score at week 24. Safety and tolerability were also assessed.

Results

At study end, improvements from baseline in 1-hour postdose FEV₁ were significantly greater in patients treated with ACL400/FOR12 FDC or ACL400/FOR6 FDC compared with aclidinium (108 mL and 87 mL, respectively; p < 0.0001). Improvements in trough FEV₁ were significantly greater in patients treated with ACL400/FOR12 FDC versus formoterol (45 mL; p = 0.0102), a numerical improvement of 26 mL in trough FEV₁ over formoterol was observed with ACL400/FOR6 FDC. Significant improvements in both SGRQ total and TDI focal scores were observed in the ACL400/FOR12 FDC group at study end (p < 0.0001), with differences over placebo exceeding the minimal clinically important difference of ≥4 points and ≥1 unit, respectively. All treatments were well tolerated, with safety profiles of the FDCs similar to those of the monotherapies.

Conclusions

Treatment with twice-daily aclidinium 400 μg/formoterol 12 μg FDC provided rapid and sustained bronchodilation that was greater than either monotherapy; clinically significant improvements in dyspnea and health status were evident compared with placebo. Aclidinium/formoterol FDC may be an effective and well tolerated new treatment option for patients with COPD.

Trial registration

Clinicaltrials.gov {"type":"clinical-trial","attrs":{"text":"NCT01437397","term_id":"NCT01437397"}}NCT01437397.*Registered trademarks of Almirall S.A., Barcelona, Spain; for use within the US as Pressair^® and Genuair^® within all other licensed territories.

Electronic supplementary material

The online version of this article (doi:10.1186/s12931-014-0123-0) contains supplementary material, which is available to authorized users.     

Add-on therapy options in asthma not adequately controlled by inhaled corticosteroids: a comprehensive review

Many patients with persistent asthma can be controlled with inhaled corticosteroids (ICS). However, a considerable proportion of patients remain symptomatic, despite the use of ICS. We present systematically evidence that supports the different treatment options. A literature search was made of Medline/PubMed to identify randomised and blinded trials. To demonstrate the benefit that can be obtained by increasing the dose of ICS, dose-response studies with at least three different ICS doses were identified. To demonstrate whether more benefit can be obtained by adding long-acting β₂-agonist (LABA), leukotriene antagonist (LTRA) or theophylline than by increasing the dose of ICS, studies comparing these options were identified. Thirdly, studies comparing the different "add-on" options were identified. The addition of a LABA is more effective than increasing the dose of ICS in improving asthma control. By increasing the dose of ICS, clinical improvement is likely to be of small magnitude. Addition of a LTRA or theophylline to the treatment regimen appears to be equivalent to doubling the dose of ICS. Addition of a LABA seems to be superior to an LTRA in improving lung function. However, addition of LABA and LTRA may be equal with respect to asthma exacerbations. However, more and longer studies are needed to better clarify the role of LTRAs and theophylline as add-on therapies.     

The relationship between osteoclastogenic and anti-osteoclastogenic pro-inflammatory cytokines differs in human osteoporotic and osteoarthritic bone tissues

Background

Pro-inflammatory cytokines possess osteoclastogenic or anti-osteoclastogenic activities. They influence osteoclasts directly or via the receptor activator of nuclear factor κB (RANK), RANK ligand (RANKL) and osteoprotegerin (OPG) system. Recent evidence suggests that inflammation may play a role in osteoporosis (OP) and osteoarthritis (OA). We aimed therefore to determine whether there is a difference between both groups: first, in the expression of the osteoclastogenic and anti-osteoclastogenic cytokines, second, in correlation of these cytokines with bone mineral density (BMD) and levels of bone turnover markers (BTM) and third, in correlation between the expression of these cytokines and osteoclast specific genes and RANK/RANKL/OPG genes.

Methods

Human bone samples from 54 age and sex matched patients with OP or OA were collected during hip arthroplasty surgery. The expression of 25 genes encoding pro-inflammatory cytokines, their receptors, osteoclast specific genes and RANK/RANKL/OPG genes was measured using quantitative real-time PCR. Total hip, femoral neck and lumbar spine BMD and BTM in blood samples were measured. The comparison between OP and OA was assessed using Student''s t-test or Mann-Whitney U test and correlations between gene expression, BMD and BTM were determined using nonparametric correlation.

Results

The results demonstrated a higher expression of interleukin (IL)-6 and IL-1α in OP, and interferon (IFN)-γ in OA (p < 0.0005). Negative correlations of total hip BMD with tumor necrosis factor-α (TNF-α) in OA and with RANKL/RANK in OP were found (p < 0.05). Significant correlations with BTM were shown for IL-1α and IFN-γ in OP (rho = 0.608 and -0.634) and for TNF-α, IL-6 and transforming growth factor-β1 (TGF-β1) in OA (rho = 0.591, -0.521 and 0.636). Results showed OP specific negative correlations (IFN-γ with ITGB3, IFN-β1 with CTSK, tartrate resistant acid phosphatase (TRAP), CALCR, RANK, RANKL, IL-1α with CTSK, OPG, IL-17A with CALCR) and positive (TGF-β1 with CTSK, TRAP, RANK), and OA specific negative (IL-1α with osteoclast associated immunoglobulin-like receptor (OSCAR), TNF-α with RANK, RANKL, OPG) and positive (IL-6 with RANK, RANKL, OPG) correlations.

Conclusions

Our results demonstrate that the relationship between osteoclastogenic and anti-osteoclastogenic pro-inflammatory cytokines differs in human OP and OA bone and could present an important factor for characteristics of OP and OA bone phenotypes.     

The Structure of a Full-length Membrane-embedded Integrin Bound to a Physiological Ligand

Increased ligand binding to integrin (“activation”) underpins many biological processes, such as leukocyte trafficking, cell migration, host-pathogen interaction, and hemostasis. Integrins exist in several conformations, ranging from compact and bent to extended and open. However, the exact conformation of membrane-embedded, full-length integrin bound to its physiological macromolecular ligand is still unclear. Integrin α_IIbβ₃, the most abundant integrin in platelets, has been a prototype for integrin activation studies. Using negative stain electron microscopy and nanodisc-embedding to provide a membrane-like environment, we visualized the conformation of full-length α_IIbβ₃ in both a Mn²⁺-activated, ligand-free state and a Mn²⁺-activated, fibrin-bound state. Activated but ligand-free integrins exist mainly in the compact conformation, whereas fibrin-bound α_IIbβ₃ predominantly exists in a fully extended, headpiece open conformation. Our results show that membrane-embedded, full-length integrin adopts an extended and open conformation when bound to its physiological macromolecular ligand.     

Modularity in Protein Evolution: Modular Organization and De Novo Domain Evolution in Mollusk Metallothioneins

Metallothioneins (MTs) are proteins devoted to the control of metal homeostasis and detoxification, and therefore, MTs have been crucial for the adaptation of the living beings to variable situations of metal bioavailability. The evolution of MTs is, however, not yet fully understood, and to provide new insights into it, we have investigated the MTs in the diverse classes of Mollusks. We have shown that most molluskan MTs are bimodular proteins that combine six domains—α, β1, β2, β3, γ, and δ—in a lineage-specific manner. We have functionally characterized the Neritimorpha β₃β₁ and the Patellogastropoda γβ₁ MTs, demonstrating the metal-binding capacity of the new γ domain. Our results have revealed a modular organization of mollusk MT, whose evolution has been impacted by duplication, loss, and de novo emergence of domains. MTs represent a paradigmatic example of modular evolution probably driven by the structural and functional requirements of metal binding.     

Extension of a de novo TIM barrel with a rationally designed secondary structure element

The ability to construct novel enzymes is a major aim in de novo protein design. A popular enzyme fold for design attempts is the TIM barrel. This fold is a common topology for enzymes and can harbor many diverse reactions. The recent de novo design of a four‐fold symmetric TIM barrel provides a well understood minimal scaffold for potential enzyme designs. Here we explore opportunities to extend and diversify this scaffold by adding a short de novo helix on top of the barrel. Due to the size of the protein, we developed a design pipeline based on computational ab initio folding that solves a less complex sub‐problem focused around the helix and its vicinity and adapt it to the entire protein. We provide biochemical characterization and a high‐resolution X‐ray structure for one variant and compare it to our design model. The successful extension of this robust TIM‐barrel scaffold opens opportunities to diversify it towards more pocket like arrangements and as such can be considered a building block for future design of binding or catalytic sites.     

Karrikins force a rethink of strigolactone mode of action

Strigolactones (SL) and karrikins (KAR) both contain essential butenolide moieties, and both require the F-box protein MAX2 to control seed germination and photomorphogenesis in Arabidopsis thaliana. A new discovery that SL and KAR also require related α/β-hydrolase proteins for such activity suggests that they operate through a similar molecular mechanism. Based on structural similarity, a previously proposed mode of action for SL was also considered for KAR, but recent structure-activity studies suggest that this mechanism may not apply. Here we rationalise these observations into a hypothesis whereby different α/β-hydrolases distinguish SL and KAR by virtue of their non-butenolide moieties and catalyze nucleophilic attack on the butenolide. The products would be different for SL and KAR, and in the case of SL they have no biological activity. The inference is that nucleophilic attack on SL and KAR by α/β-hydrolases is required for their bioactivity, but the hydrolysis products are not.     

Characterization and structural analyses of a novel glycosyltransferase acting on the β-1,2-glucosidic linkages

The IALB_1185 protein, which is encoded in the gene cluster for endo-β-1,2-glucanase homologs in the genome of Ignavibacterium album, is a glycoside hydrolase family (GH) 35 protein. However, most known GH35 enzymes are β-galactosidases, which is inconsistent with the components of this gene cluster. Thus, IALB_1185 is expected to possess novel enzymatic properties. Here, we showed using recombinant IALB_1185 that this protein has glycosyltransferase activity toward β-1,2-glucooligosaccharides, and that the kinetic parameters for β-1,2-glucooligosaccharides are not within the ranges for general GH enzymes. When various aryl- and alkyl-glucosides were used as acceptors, glycosyltransfer products derived from these acceptors were subsequently detected. Kinetic analysis further revealed that the enzyme has wide aglycone specificity regardless of the anomer, and that the β-1,2-linked glucose dimer sophorose is an appropriate donor. In the complex of wild-type IALB_1185 with sophorose, the electron density of sophorose was clearly observed at subsites −1 and +1, whereas in the E343Q mutant–sophorose complex, the electron density of sophorose was clearly observed at subsites +1 and +2. This observation suggests that binding at subsites −1 and +2 competes through Glu102, which is consistent with the preference for sophorose as a donor and unsuitability of β-1,2-glucooligosaccharides as acceptors. A pliable hydrophobic pocket that can accommodate various aglycone moieties was also observed in the complex structures with various glucosides. Overall, our biochemical and structural data are indicative of a novel enzymatic reaction. We propose that IALB_1185 be redefined β-1,2-glucooligosaccharide:d-glucoside β-d-glucosyltransferase as a systematic name and β-1,2-glucosyltransferase as an accepted name.     

Association of plasma sRAGE,but not esRAGE with lung function impairment in COPD

Rationale

Plasma soluble Receptor for Advanced Glycation End Product (sRAGE) is considered as a biomarker in COPD. The contribution of endogenous sRAGE (esRAGE) to the pool of plasma sRAGE and the implication of both markers in COPD pathogenesis is however not clear yet. The aim of the current study was therefore to measure plasma levels of esRAGE comparative to total sRAGE in patients with COPD and a control group. Further, we established the relations of esRAGE and total sRAGE with disease specific characteristics such as lung function and D_LCO, and with different circulating AGEs.

Methods

Plasma levels of esRAGE and sRAGE were measured in an 88 patients with COPD and in 55 healthy controls. FEV₁ (%predicted) and FEV₁/VC (%) were measured in both groups; D_LCO (%predicted) was measured in patients only. In this study population we previously reported that the AGE N^ϵ-(carboxymethyl) lysine (CML) was decreased, N^ϵ-(carboxyethyl) lysine (CEL) increased and pentosidine was not different in plasma of COPD patients compared to controls.

Results

Plasma esRAGE (COPD: 533.9 ± 412.4, Controls: 848.7 ± 690.3 pg/ml; p = 0.000) was decreased in COPD compared to controls. No significant correlations were observed between plasma esRAGE levels and lung function parameters or plasma AGEs. A positive correlation was present between esRAGE and total sRAGE levels in the circulation. Confirming previous findings, total sRAGE (COPD: 512.6 ± 403.8, Controls: 1834 ± 804.2 pg/ml; p < 0.001) was lower in patients compared to controls and was positively correlated FEV₁ (r = 0.235, p = 0.032), FEV₁/VC (r = 0.218, p = 0.047), and D_LCO (r = 0.308, p = 0.006). sRAGE furthermore did show a significant positive association with CML (r = 0.321, p = 0.003).

Conclusion

Although plasma esRAGE is decreased in COPD patients compared to controls, only total sRAGE showed a significant and independent association with FEV₁, FEV₁/VC and D_LCO, indicating that total sRAGE but not esRAGE may serve as marker of COPD disease state and severity.     

Carotenoid metabolism in mammals,including man: formation,occurrence, and function of apocarotenoids: Thematic Review Series: Fat-Soluble Vitamins: Vitamin A

Vitamin A was recognized as an essential nutrient 100 years ago. In the 1930s, it became clear that dietary β-carotene was cleaved at its central double to yield vitamin A (retinal or β-apo-15′-carotenal). Thus a great deal of research has focused on the central cleavage of provitamin A carotenoids to form vitamin A (retinoids). The mechanisms of formation and the physiological role(s) of noncentral (eccentric) cleavage of both provitamin A carotenoids and nonprovitamin A carotenoids has been less clear. It is becoming apparent that the apocarotenoids exert unique biological activities themselves. These compounds are found in the diet and thus may be absorbed in the intestine, or they may form from enzymatic or nonenzymatic cleavage of the parent carotenoids. The mechanism of action of apocarotenoids in mammals is not fully worked out. However, as detailed in this review, they have profound effects on gene expression and work, at least in part, through the modulation of ligand-activated nuclear receptors. Understanding the interactions of apocarotenoids with other lipid-binding proteins, chaperones, and metabolizing enzymes will undoubtedly increase our understanding of the biological roles of these carotenoid metabolites.     

A newly introduced salt bridge cluster improves structural and biophysical properties of de novo TIM barrels

Protein stability can be fine‐tuned by modifying different structural features such as hydrogen‐bond networks, salt bridges, hydrophobic cores, or disulfide bridges. Among these, stabilization by salt bridges is a major challenge in protein design and engineering since their stabilizing effects show a high dependence on the structural environment in the protein, and therefore are difficult to predict and model. In this work, we explore the effects on structure and stability of an introduced salt bridge cluster in the context of three different de novo TIM barrels. The salt bridge variants exhibit similar thermostability in comparison with their parental designs but important differences in the conformational stability at 25°C can be observed such as a highly stabilizing effect for two of the proteins but a destabilizing effect to the third. Analysis of the formed geometries of the salt bridge cluster in the crystal structures show either highly ordered salt bridge clusters or only single salt bridges. Rosetta modeling of the salt bridge clusters results in a good prediction of the tendency on stability changes but not the geometries observed in the three‐dimensional structures. The results show that despite the similarities in protein fold, the salt bridge clusters differently influence the structural and stability properties of the de novo TIM barrel variants depending on the structural background where they are introduced.     

The vitamin E isoforms α-tocopherol and γ-tocopherol have opposite associations with spirometric parameters: the CARDIA study

Background

Clinical studies of the associations of vitamin E with lung function have reported conflicting results. However, these reports primarily examine the α-tocopherol isoform of vitamin E and have not included the isoform γ-tocopherol which we recently demonstrated in vitro opposes the function of α-tocopherol. We previously demonstrated, in vitro and in animal studies, that the vitamin E isoform α-tocopherol protects, but the isoform γ-tocopherol promotes lung inflammation and airway hyperresponsiveness.

Methods

To translate these findings to humans, we conducted analysis of 4526 adults in the Coronary Artery Risk Development in Young Adults (CARDIA) multi-center cohort with available spirometry and tocopherol data in blacks and whites. Spirometry was obtained at years 0, 5, 10, and 20 and serum tocopherol was from years 0, 7 and 15 of CARDIA.

Results

In cross-sectional regression analysis at year 0, higher γ-tocopherol associated with lower FEV₁ (p = 0.03 in blacks and p = 0.01 in all participants) and FVC (p = 0.01 in blacks, p = 0.05 in whites, and p = 0.005 in all participants), whereas higher α-tocopherol associated with higher FVC (p = 0.04 in blacks and whites and p = 0.01 in all participants). In the lowest quartile of α-tocopherol, higher γ-tocopherol associated with a lower FEV₁ (p = 0.05 in blacks and p = 0.02 in all participants). In contrast, in the lowest quartile of γ-tocopherol, higher α-tocopherol associated with a higher FEV₁ (p = 0.03) in blacks. Serum γ-tocopherol >10 μM was associated with a 175–545 ml lower FEV₁ and FVC at ages 21–55 years.

Conclusion

Increasing serum concentrations of γ-tocopherol were associated with lower FEV1 or FVC, whereas increasing serum concentrations of α-tocopherol was associated with higher FEV1 or FVC. Based on the prevalence of serum γ-tocopherol >10 μM in adults in CARDIA and the adult U.S. population in the 2011 census, we expect that the lower FEV₁ and FVC at these concentrations of serum γ-tocopherol occur in up to 4.5 million adults in the population.     

Plasmin-sensitive dibasic sequences in the third fibronectin-like domain of L1-cell adhesion molecule (CAM) facilitate homomultimerization and concomitant integrin recruitment

L1 is a multidomain transmembrane neural recognition molecule essential for neurohistogenesis. While moieties in the immunoglobulin-like domains of L1 have been implicated in both heterophilic and homophilic binding, the function of the fibronectin (FN)-like repeats remains largely unresolved. Here, we demonstrate that the third FN-like repeat of L1 (FN3) spontaneously homomultimerizes to form trimeric and higher order complexes. Remarkably, these complexes support direct RGD-independent interactions with several integrins, including alpha(v)beta(3) and alpha(5)beta(1). A pep- tide derived from the putative C-C' loop of FN3 (GSQRKHSKRHIHKDHV(852)) also forms trimeric complexes and supports alpha(v)beta(3) and alpha(5)beta(1) binding. Substitution of the dibasic RK(841) and KR(845) sequences within this peptide or the FN3 domain limited multimerization and abrogated integrin binding. Evidence is presented that the multimerization of, and integrin binding to, the FN3 domain is regulated both by conformational constraints imposed by other domains and by plasmin- mediated cleavage within the sequence RK( downward arrow)HSK( downward arrow)RH(846). The integrin alpha(9)beta(1), which also recognizes the FN3 domain, colocalizes with L1 in a manner restricted to sites of cell-cell contact. We propose that distal receptor ligation events at the cell-cell interface may induce a conformational change within the L1 ectodomain that culminates in receptor multimerization and integrin recruitment via interaction with the FN3 domain.     

Strong acids induce amyloid fibril formation of β2-microglobulin via an anion-binding mechanism

Amyloid fibrils, crystal-like fibrillar aggregates of proteins associated with various amyloidoses, have the potential to propagate via a prion-like mechanism. Among known methodologies to dissolve preformed amyloid fibrils, acid treatment has been used with the expectation that the acids will degrade amyloid fibrils similar to acid inactivation of protein functions. Contrary to our expectation, treatment with strong acids, such as HCl or H₂SO₄, of β₂-microglobulin (β2m) or insulin actually promoted amyloid fibril formation, proportionally to the concentration of acid used. A similar promotion was observed at pH 2.0 upon the addition of salts, such as NaCl or Na₂SO₄. Although trichloroacetic acid, another strong acid, promoted amyloid fibril formation of β2m, formic acid, a weak acid, did not, suggesting the dominant role of anions in promoting fibril formation of this protein. Comparison of the effects of acids and salts confirmed the critical role of anions, indicating that strong acids likely induce amyloid fibril formation via an anion-binding mechanism. The results suggest that although the addition of strong acids decreases pH, it is not useful for degrading amyloid fibrils, but rather induces or stabilizes amyloid fibrils via an anion-binding mechanism.