Aminopeptidases do not directly degrade tau protein

BACKGROUND: Tau hyperphosphorylation and aggregation to form intracellular neurofibrillar tangles is prevalent in a number of tauopathies. Thus there is current interest in the mechanisms involved in Tau clearance. It was recently reported that Tau can be degraded by an aminopeptidase known as the puromycin sensitive aminopeptidase (PSA). Until now PSA has been reported to only cleave peptides, with the largest reported substrates having 30-50 amino acids. We have studied this unique PSA cleavage reaction using a number of different PSA preparations. RESULTS: An N-terminally His tagged-PSA was expressed and purified from Sf9 insect cells. Although this PSA preparation cleaved Tau, product analysis with N and C terminal Tau antibodies coupled with mass spectrometry showed an endoproteolytic cleavage atypical for an aminopeptidase. Furthermore, the reaction was not blocked by the general aminopeptidase inhibitor bestatin or the specific PSA inhibitor puromycin. In order to test whether Tau hydrolysis might be caused by a protease contaminant the enzyme was expressed in E. coli as glutathione S-transferase and maltose binding protein fusion proteins or in Sf9 cells as a C-terminally His-tagged protein. After purification to near homogeneity none of these other recombinant forms of PSA cleaved Tau. Further, Tau-cleaving activity and aminopeptidase activities derived from the Sf9 cell expression system were separable by molecular sieve chromatography. When tested in a cellular context we again failed to see a PSA dependent cleavage of Tau. A commercial preparation of a related aminopeptidase, aminopeptidase N, also exhibited Tau cleaving activity, but this activity could also be separated from aminopeptidase activity. CONCLUSION: It is concluded that PSA does not directly cleave Tau

Aminopeptidases do not directly degrade tau protein

BACKGROUND: Tau hyperphosphorylation and aggregation to form intracellular neurofibrillar tangles is prevalent in a number of tauopathies. Thus there is current interest in the mechanisms involved in Tau clearance. It was recently reported that Tau can be degraded by an aminopeptidase known as the puromycin sensitive aminopeptidase (PSA). Until now PSA has been reported to only cleave peptides, with the largest reported substrates having 30-50 amino acids. We have studied this unique PSA cleavage reaction using a number of different PSA preparations. RESULTS: An N-terminally His tagged-PSA was expressed and purified from Sf9 insect cells. Although this PSA preparation cleaved Tau, product analysis with N and C terminal Tau antibodies coupled with mass spectrometry showed an endoproteolytic cleavage atypical for an aminopeptidase. Furthermore, the reaction was not blocked by the general aminopeptidase inhibitor bestatin or the specific PSA inhibitor puromycin. In order to test whether Tau hydrolysis might be caused by a protease contaminant the enzyme was expressed in E. coli as glutathione S-transferase and maltose binding protein fusion proteins or in Sf9 cells as a C-terminally His-tagged protein. After purification to near homogeneity none of these other recombinant forms of PSA cleaved Tau. Further, Tau-cleaving activity and aminopeptidase activities derived from the Sf9 cell expression system were separable by molecular sieve chromatography. When tested in a cellular context we again failed to see a PSA dependent cleavage of Tau. A commercial preparation of a related aminopeptidase, aminopeptidase N, also exhibited Tau cleaving activity, but this activity could also be separated from aminopeptidase activity. CONCLUSION: It is concluded that PSA does not directly cleave Tau

Neutrophilia and NETopathy as Key Pathologic Drivers of Progressive Lung Impairment in Patients With COVID-19

There is an urgent need for new therapeutic strategies to contain the spread of the novel coronavirus disease 2019 (COVID-19) and to curtail its most severe complications. Severely ill patients experience pathologic manifestations of acute respiratory distress syndrome (ARDS), and clinical reports demonstrate striking neutrophilia, elevated levels of multiple cytokines, and an exaggerated inflammatory response in fatal COVID-19. Mechanical respirator devices are the most widely applied therapy for ARDS in COVID-19, yet mechanical ventilation achieves strikingly poor survival. Many patients, who recover, experience impaired cognition or physical disability. In this review, we argue the need to develop therapies aimed at inhibiting neutrophil recruitment, activation, degranulation, and neutrophil extracellular trap (NET) release. Moreover, we suggest that currently available pharmacologic approaches should be tested as treatments for ARDS in COVID-19. In our view, targeting host-mediated immunopathology holds promise to alleviate progressive pathologic complications of ARDS and reduce morbidities and mortalities in severely ill patients with COVID-19

Stakeholder preferences for attributes of digital health technologies to consider in health service funding

Objectives: Health service providers are currently making decisions on the public funding of digital health technologies (DHTs) for managing chronic diseases with limited understanding of stakeholder preferences for DHT attributes. This study aims to understand the community, patient/carer, and health professionals' preferences to help inform a prioritized list of evaluation criteria. Methods: An online best-worst scaling survey was conducted in Australia, New Zealand, Canada, and the United Kingdom to ascertain the relative importance of twenty-four DHT attributes among stakeholder groups using an efficient incomplete block design. The attributes were identified from a systematic review of DHT evaluation frameworks for consideration in a health technology assessment. Results were analyzed with multinomial models by stakeholder group and latent class. Results: A total of 1,251 participants completed the survey (576 general community members, 543 patients/carers, and 132 health professionals). Twelve attributes achieved a preference score above 50 percent in the stakeholder group model, predominantly related to safety but also covering technical features, effectiveness, ethics, and economics. Results from the latent class model supported this prioritization. Overall, connectedness with the patient's healthcare team seemed the most important; with Helps health professionals respond quickly when changes in patient care are needed as the most highly prioritized of all attributes. Conclusions: It is proposed that these prioritized twelve attributes be considered in all evaluations of DHTs that manage chronic disease, supplemented with a limited number of attributes that reflect the specific perspective of funders, such as equity of access, cost, and system-level implementation considerations

Manifolds with almost nonnegative curvature operator and principal bundles

We study manifolds with almost nonnegative curvature operator (ANCO) and provide first examples of closed simply connected ANCO mannifolds that do not admit nonnegative curvature operator

Quantum Maxwell-Bloch equations for spatially inhomogeneous semiconductor lasers

We present quantum Maxwell-Bloch equations (QMBE) for spatially inhomogeneous semiconductor laser devices. The QMBE are derived from fully quantum mechanical operator dynamics describing the interaction of the light field with the quantum states of the electrons and the holes near the band gap. By taking into account field-field correlations and field-dipole correlations, the QMBE include quantum noise effects which cause spontaneous emission and amplified spontaneous emission. In particular, the source of spontaneous emission is obtained by factorizing the dipole-dipole correlations into a product of electron and hole densities. The QMBE are formulated for general devices, for edge emitting lasers and for vertical cavity surface emitting lasers, providing a starting point for the detailed analysis of spatial coherence in the near field and far field patterns of such laser diodes. Analytical expressions are given for the spectra of gain and spontaneous emission described by the QMBE. These results are applied to the case of a broad area laser, for which the frequency and carrier density dependent spontaneous emission factor beta and the evolution of the far field pattern near threshold are derived.Comment: 22 pages RevTex and 7 figures, submitted to Phys.Rev.A, revisions in abstract and in the discussion of temporal coherenc

Immunization of Alpacas (\u3cem\u3eLama pacos\u3c/em\u3e) with Protein Antigens and Production of Antigen-Specific Single Domain Antibodies

In this manuscript, a method for the immunization of alpaca and the use of molecular biology methods to produce antigen-specific single domain antibodies is described and demonstrated. Camelids, such as alpacas and llamas, have become a valuable resource for biomedical research since they produce a novel type of heavy chain-only antibody which can be used to produce single domain antibodies. Because the immune system is highly flexible, single domain antibodies can be made to many different protein antigens, and even different conformations of the antigen, with a very high degree of specificity. These features, among others, make single domain antibodies an invaluable tool for biomedical research. A method for the production of single domain antibodies from alpacas is reported. A protocol for immunization, blood collection, and B-cell isolation is described. The B-cells are used for the construction of an immunized library, which is used in the selection of specific single domain antibodies via panning. Putative specific single domain antibodies obtained via panning are confirmed by pull-down, ELISA, or gel-shift assays. The resulting single domain antibodies can then be used either directly or as a part of an engineered reagent. The uses of single domain antibody and single domain antibody-based regents include structural, biochemical, cellular, in vivo, and therapeutic applications. Single domain antibodies can be produced in large quantities as recombinant proteins in prokaryotic expression systems, purified, and used directly or can be engineered to contain specific markers or tags that can be used as reporters in cellular studies or in diagnostics

Environmental Profile of a Community’s Health (EPOCH): An Ecometric Assessment of Measures of the Community Environment Based on Individual Perception

Background: Public health research has turned towards examining upstream, community-level determinants of cardiovascular disease risk factors. Objective measures of the environment, such as those derived from direct observation, and perception-based measures by residents have both been associated with health behaviours. However, current methods are generally limited to objective measures, often derived from administrative data, and few instruments have been evaluated for use in rural areas or in low-income countries. We evaluate the reliability of a quantitative tool designed to capture perceptions of community tobacco, nutrition, and social environments obtained from interviews with residents in communities in 5 countries. Methodology/Principal Findings: Thirteen measures of the community environment were developed from responses to questionnaire items from 2,360 individuals residing in 84 urban and rural communities in 5 countries (China, India, Brazil, Colombia, and Canada) in the Environmental Profile of a Community’s Health (EPOCH) study. Reliability and other properties of the community-level measures were assessed using multilevel models. High reliability (>0.80) was demonstrated for all community-level measures at the mean number of survey respondents per community (n = 28 respondents). Questionnaire items included in each scale were found to represent a common latent factor at the community level in multilevel factor analysis models. Conclusions/ Significance: Reliable measures which represent aspects of communities potentially related to cardiovascular disease (CVD)/risk factors can be obtained using feasible sample sizes. The EPOCH instrument is suitable for use in different settings to explore upstream determinants of CVD/risk factors

Renormalization Group Analysis of a Quivering String Model of Posture Control

Scaling concepts and renormalization group (RG) methods are applied to a simple linear model of human posture control consisting of a trembling or quivering string subject to damping and restoring forces. The string is driven by uncorrelated white Gaussian noise intended to model the corrections of the physiological control system. We find that adding a weak quadratic nonlinearity to the posture control model opens up a rich and complicated phase space (representing the dynamics) with various non-trivial fixed points and basins of attraction. The transition from diffusive to saturated regimes of the linear model is understood as a crossover phenomenon, and the robustness of the linear model with respect to weak non-linearities is confirmed. Correlations in posture fluctuations are obtained in both the time and space domain. There is an attractive fixed point identified with falling. The scaling of the correlations in the front-back displacement, which can be measured in the laboratory, is predicted for both the large-separation (along the string) and long-time regimes of posture control.Comment: 20 pages, 13 figures, RevTeX, accepted for publication in PR

Theory of coherent acoustic phonons in InGaN/GaN multi-quantum wells

A microscopic theory for the generation and propagation of coherent LA phonons in pseudomorphically strained wurzite (0001) InGaN/GaN multi-quantum well (MQW) p-i-n diodes is presented. The generation of coherent LA phonons is driven by photoexcitation of electron-hole pairs by an ultrafast Gaussian pump laser and is treated theoretically using the density matrix formalism. We use realistic wurzite bandstructures taking valence-band mixing and strain-induced piezo- electric fields into account. In addition, the many-body Coulomb ineraction is treated in the screened time-dependent Hartree-Fock approximation. We find that under typical experimental conditions, our microscopic theory can be simplified and mapped onto a loaded string problem which can be easily solved.Comment: 20 pages, 17 figure