Overexpression of Nmnat3 efficiently increases NAD and NGD levels and ameliorates age‐associated insulin resistance

Nicotinamide adenine dinucleotide (NAD) is an important cofactor that regulates various biological processes, including metabolism and gene expression. As a coenzyme, NAD controls mitochondrial respiration through enzymes of the tricarboxylic acid (TCA) cycle, β‐oxidation, and oxidative phosphorylation and also serves as a substrate for posttranslational protein modifications, such as deacetylation and ADP‐ribosylation by sirtuins and poly(ADP‐ribose) polymerase (PARP), respectively. Many studies have demonstrated that NAD levels decrease with aging and that these declines cause various aging‐associated diseases. In contrast, activation of NAD metabolism prevents declines in NAD levels during aging. In particular, dietary supplementation with NAD precursors has been associated with protection against age‐associated insulin resistance. However, it remains unclear which NAD synthesis pathway is important and/or efficient at increasing NAD levels in vivo. In this study, Nmnat3 overexpression in mice efficiently increased NAD levels in various tissues and prevented aging‐related declines in NAD levels. We also demonstrated that Nmnat3‐overexpressing (Nmnat3 Tg) mice were protected against diet‐induced and aging‐associated insulin resistance. Moreover, in skeletal muscles of Nmnat3 Tg mice, TCA cycle activity was significantly enhanced, and the energy source for oxidative phosphorylation was shifted toward fatty acid oxidation. Furthermore, reactive oxygen species (ROS) generation was significantly suppressed in aged Nmnat3 Tg mice. Interestingly, we also found that concentrations of the NAD analog nicotinamide guanine dinucleotide (NGD) were dramatically increased in Nmnat3 Tg mice. These results suggest that Nmnat3 overexpression improves metabolic health and that Nmnat3 is an attractive therapeutic target for metabolic disorders that are caused by aging.     

Oxygen Therapy Use in Older Adults with Chronic Obstructive Pulmonary Disease

Rationale

Oxygen therapy improves survival and function in severely hypoxemic chronic obstructive pulmonary disease (COPD) patients based on two landmark studies conducted over 40 years ago. We hypothesize that oxygen users in the current era may be very different. We examined trends and subject characteristics associated with oxygen therapy use from 2001–2010 in the United States.

Methods

We examined Medicare beneficiaries with COPD who received oxygen from 2001 to 2010. COPD subjects were identified by: 1) ≥2 outpatient visits >30 days apart within one year with an encounter diagnosis of COPD; or 2) an acute care hospitalization with COPD as the primary or secondary discharge diagnosis. Oxygen therapy and sustained oxygen therapy were defined as ≥1 and ≥11 claims for oxygen, respectively, in the durable medical equipment file in a calendar year. Primary outcome measures were factors associated with oxygen therapy and sustained oxygen therapy over the study period.

Results

Oxygen therapy increased from 33.7% in 2001 to 40.5% in 2010 (p-value of trend <0.001). Sustained oxygen therapy use increased from 19.5% in 2001, peaked in 2008 to 26.9% and declined to 18.5% in 2010. The majority of subjects receiving oxygen therapy and sustained oxygen therapy were female. Besides gender, factors associated with any oxygen use or sustained oxygen therapy were non-Hispanic white race, low socioeconomic status and ≥2 comorbidities.

Conclusions

Any oxygen use among fee-for service Medicare beneficiaries with COPD is high. Current users of oxygen are older females with multiple comorbidities. Decline in sustained oxygen therapy use after 2008 may be related to reimbursement policy change.     

Interaction pattern for the complex of B-DNA Fullerene compounds with a set of known replication proteins using docking study

Fullerenes have attracted considerable attention due to their unique chemical structure and potential applications which has opened wide venues for possible human exposure to various fullerene types. Therefore, in depth knowledge of how fullerene may interfere with various cellular processes becomes quite imperative. The present study was designed to investigate how the presence of fullerene affect the binding of DNA with different enzymes involved in replication process. Different fullerenes were first docked with DNA and then binding scores of different enzymes was analyzed with fullerene docked DNA. C30, C40 & C50 once docked with DNA, reduced the binding score of primase, whereas no significant change in the binding score was observed with the helicase, ssb protein, dna pol δ, dna pol ε, ligase, DNA clamp, and topoisomerases. On the contrast, the binding score of RPA14 decreases in fluctuating manner while interacting with increasing molecular weight of fullerene bound single-stranded DNA complex. The study revealed the affect of fullerene family interacting with DNA on the binding pattern of enzymes involved in replication process. Study suggests that the presence of most of fullerenes may not affect the activity of these enzymes necessary for replication process whereas C30, C40 & C50 may disrupt the activity of primase, (strating point for DNA polymerase) its docking score decreases from 13820 to 10702.     

Compartment-specific Synthesis of Phosphatidylethanolamine Is Required for Normal Heavy Metal Resistance

Control of lipid composition of membranes is crucial to ensure normal cellular functions. Saccharomyces cerevisiae has two different phosphatidylserine decarboxylase enzymes (Psd1 and Psd2) that catalyze formation of phosphatidylethanolamine. The mitochondrial Psd1 provides roughly 70% of the phosphatidylethanolamine (PE) biosynthesis in the cell with Psd2 carrying out the remainder. Here, we demonstrate that loss of Psd2 causes cells to acquire sensitivity to cadmium even though Psd1 remains intact. This cadmium sensitivity results from loss of normal activity of a vacuolar ATP-binding cassette transporter protein called Ycf1. Measurement of phospholipid levels indicates that loss of Psd2 causes a specific reduction in vacuolar membrane PE levels, whereas total PE levels are not significantly affected. The presence of a phosphatidylinositol transfer protein called Pdr17 is required for Psd2 function and normal cadmium tolerance. We demonstrate that Pdr17 and Psd2 form a complex in vivo that seems essential for maintenance of vacuolar PE levels. Finally, we refine the localization of Psd2 to the endosome arguing that this enzyme controls vacuolar membrane phospholipid content by regulating phospholipids in compartments that will eventually give rise to the vacuole. Disturbance of this regulation of intracellular phospholipid balance leads to selective loss of membrane protein function in the vacuole.     

Process and Outcome Measures among COPD Patients with a Hospitalization Cared for by an Advance Practice Provider or Primary Care Physician

Objectives

To examine the process and outcomes of care of COPD patients by Advanced Practice Providers (APPs) and primary care physicians.

Methods

We conducted a cross sectional retrospective cohort study of Medicare beneficiaries with COPD who had at least one hospitalization in 2010. We examined the process measures of receipt of spirometry evaluation, influenza and pneumococcal vaccine, use of COPD medications, and referral to a pulmonary specialist visit. Outcome measures were emergency department (ER) visit, number of hospitalizations and 30-day readmission in 2010.

Results

A total of 7,257 Medicare beneficiaries with COPD were included. Of these, 1,999 and 5,258 received primary care from APPs and primary care physicians, respectively. Patients in the APP group were more likely to be white, younger, male, residing in non-metropolitan areas and have fewer comorbidities. In terms of process of care measures, APPs were more likely to prescribe short acting bronchodilators (adjusted odds ratio [aOR] = 1.18, 95%Confidence Interval [CI] 1.05–1.32), oxygen therapy (aOR = 1.25, 95% CI 1.12–1.40) and consult a pulmonary specialist (aOR = 1.39, 95% CI 1.23–1.56), but less likely to give influenza and pneumococcal vaccinations. Patients receiving care from APPs had lower rates of ER visits for COPD (aOR = 0.84, 95%CI 0.71–0.98) and had a higher follow-up rate with pulmonary specialist within 30 days of hospitalization for COPD (aOR = 1.25, 95%CI 1.07–1.48) than those cared for by physicians.

Conclusions

Compared to patients cared for by physicians, patients cared for by APPs were more likely to receive short acting bronchodilator, oxygen therapy and been referred to pulmonologist, however they had lower rates of vaccination probably due to lower age group. Patients cared for by APPs were less like to visit an ER for COPD compared to patients care for by physicians, conversely there was no differences in hospitalization or readmission for COPD between MDs and APPs.     

Deficiency of Nicotinamide Mononucleotide Adenylyltransferase 3 (Nmnat3) Causes Hemolytic Anemia by Altering the Glycolytic Flow in Mature Erythrocytes

NAD biosynthesis is of substantial interest because of its important roles in regulating various biological processes. Nicotinamide mononucleotide adenylyltransferase 3 (Nmnat3) is considered a mitochondria-localized NAD synthesis enzyme involved in de novo and salvage pathways. Although the biochemical properties of Nmnat3 are well documented, its physiological function in vivo remains unclear. In this study, we demonstrated that Nmnat3 was localized in the cytoplasm of mature erythrocytes and critically regulated their NAD pool. Deficiency of Nmnat3 in mice caused splenomegaly and hemolytic anemia, which was associated with the findings that Nmnat3-deficient erythrocytes had markedly lower ATP levels and shortened lifespans. However, the NAD level in other tissues were not apparently affected by the deficiency of Nmnat3. LC-MS/MS-based metabolomics revealed that the glycolysis pathway in Nmnat3-deficient erythrocytes was blocked at a glyceraldehyde 3-phosphate dehydrogenase (GAPDH) step because of the shortage of the coenzyme NAD. Stable isotope tracer analysis further demonstrated that deficiency of Nmnat3 resulted in glycolysis stall and a shift to the pentose phosphate pathway. Our findings indicate the critical roles of Nmnat3 in maintenance of the NAD pool in mature erythrocytes and the physiological impacts at its absence in mice.     

Molecular docking analysis of new generation cephalosporins interactions with recently known SHV-variants

Extended-spectrum-β-lactamases (ESBLs), constitutes the growing class of betalactamses, these are enzymes produced by bacteria which impart resistance against advanced-generation-cephalosporins. SHV enzymes are among the most prevalent ESBLs. The mode of molecular interactions of recent SHV-variants to advanced generation cephalosporins has not been reported yet. This is the first time we are reporting the insilico study of these recent variants with new generation cephaosporins. Homology models for SHV-105, SHV-95, SHV-89, SHV-61 and SHV-48 were generated using MODELLER9v3. New generation Cephalosporins were selected to target the active site amino acid residues of these modeled SHV enzymes for predicting comparative efficacies of these inhibitors against the said enzymes on the basis of interaction energies of docking. The docked complexes were analyzed by using DISCOVERY STUDIO 2.5. In this study A237, S70, K234, R275, N132, R244 and S130 were found crucial to the correct positioning of drugs within the binding site of SHV enzymes in 11, 6, 6, 6, 5, 5 and 5 instances, respectively. On the basis of interaction energy and Ki calculations cefatoxime emerged as the most efficient among the other advanced cephalosporins against all the studied SHV variants, excluding SHV-48 where ceftazidime was found to be most effective drug. Furthermore, this study identified amino acid residues crucial to 'SHV-Cephalosporins' interactions and this information will be useful in designing effective and versatile drug candidates.