Targeting Calcium Sensitization Mechanisms In Human Airway Smooth Muscle For Novel Bronchodilator Therapy In Asthma

Asthma is characterized by airway hyperresponsiveness (AHR) and inflammation. Management of asthma involves controlling airway inflammation with inhaled corticosteroids, and using bronchodilators such as β-2 agonists that inhibit airway smooth muscle (ASM) shortening and prevents or reverses airway narrowing. Despite β-2 agonists’ ability to mitigate asthma exacerbations, they have limitations, especially concerning efficacy and safety. Studies have shown that β-2 agonist use can result in adverse patient outcomes, β-2 adrenergic receptor tachyphylaxis, deterioration of asthma control, and death. New therapeutics are needed to overcome these limitations. Rho kinase (ROCK) inhibitors have generated excitement as potential bronchodilators. Since ROCK activation is necessary to maintain ASM tone by inhibiting MLC phosphatase, ROCK inhibitors allow the constitutively active MLC phosphatase to de-phosphorylate MLC and promote relaxation of ASM. ROCK inhibitors blunt ASM contraction and induce bronchodilation in vitro, ex vivo, and in vivo. Unfortunately, ROCK is ubiquitously expressed and adverse effects such as vasodilation preclude any clinical trials. We present an alternate strategy to induce bronchodilation in human airways by inhibiting the p110δ isoform of phosphoinositide 3-kinase (PI3Kδ) that is not ubiquitously expressed. In primary human ASM cells (HASMCs), we demonstrate that PI3Kδ modulates ROCK activation, and that inhibition of PI3Kδ using a FDA approved drug Cal101 (Idelalisib) attenuates ROCK activation. Cal101 induces relaxation of cultured HASMCs by inhibiting PI3Kδ-mediated activation of ROCK. Using human precision-cut lung slices, we also demonstrate that Cal101 induces bronchodilation of human small airways as effectively as formoterol, an industry standard β-2 agonist. In our model of β-2 agonist tachyphylaxis, the effectiveness of β-2 agonists was significantly attenuated while the effectiveness of Cal101 was not. Thus, our results suggest that PI3Kδ inhibitors represent a novel class of bronchodilators. Furthermore, we demonstrate that Gα12 mediates agonist-induced activation of PI3K and ROCK. These results unveil a novel pathway regulating HASMC signaling that is necessary to elicit shortening. Importantly, our work has uncovered several new targets for bronchodilation and offers new avenues for the treatment of asthma

Are We Over-Lawyering International Affairs

This panel will discuss the role of lawyers — particularly government lawyers — in addressing questions of legal policy. We will discuss fundamental questions such as: Should lawyers decide legal policy? Or, is that best left to the policymakers? Should lawyers give advice as to legal policy, or should they stick to providing answers as to what the law is? How should lawyers respond to what a policymaker thinks is the legal question, but is really a question of legal policy? If lawyers find the law vague or lacking, should they fill in the gaps, advising as to what the law should be? Was Secretary of State Rice right when she warned the American Society of International Law that lawyers should not stretch laws, such as the Geneva Conventions, to apply to circumstances they were not designed for? Did the Office of the Justice Department’s opinions on interrogation techniques stretch in the other direction when they held that laws did not restrict the President’s authority? Should lawyers indicate the quality of the response to a question? For example, should they say how a court would, or should, decide, or is it just enough to say that this is a reasonable answer and others may differ? What should a government lawyer do after losing an intra-governmental policy argument on a legal issue? Is the answer different if the argument was over a legal policy issue

The Brescia Internationally Validated European Guidelines on Minimally Invasive Pancreatic Surgery (EGUMIPS)

Objective: To develop and update evidence-based and consensus-based guidelines on laparoscopic and robotic pancreatic surgery. Summary Background Data: Minimally invasive pancreatic surgery (MIPS), including laparoscopic and robotic surgery, is complex and technically demanding. Minimizing the risk for patients requires stringent, evidence-based guidelines. Since the International Miami Guidelines on MIPS in 2019, new developments and key publications have been reported, necessitating an update. Methods: Evidence-based guidelines on 22 topics in 8 domains were proposed: terminology, indications, patients, procedures, surgical techniques and instrumentation, assessment tools, implementation and training, and artificial intelligence. The Brescia Internationally Validated European Guidelines on Minimally Invasive Pancreatic Surgery (EGUMIPS, September 2022) used the Scottish Intercollegiate Guidelines Network (SIGN) methodology to assess the evidence and develop guideline recommendations, the Delphi method to establish consensus on the recommendations among the Expert Committee, and the AGREE II-GRS tool for guideline quality assessment and external validation by a Validation Committee. Results: Overall, 27 European experts, 6 international experts, 22 international Validation Committee members, 11 Jury Committee members, 18 Research Committee members, and 121 registered attendees of the 2-day meeting were involved in the development and validation of the guidelines. In total, 98 recommendations were developed, including 33 on laparoscopic, 34 on robotic, and 31 on general MIPS, covering 22 topics in 8 domains. Out of 98 recommendations, 97 reached at least 80% consensus among the experts and congress attendees, and all recommendations were externally validated by the Validation Committee. Conclusions: The EGUMIPS evidence-based guidelines on laparoscopic and robotic MIPS can be applied in current clinical practice to provide guidance to patients, surgeons, policy-makers, and medical societies.</p

Non-Standard Errors

In statistics, samples are drawn from a population in a data-generating process (DGP). Standard errors measure the uncertainty in estimates of population parameters. In science, evidence is generated to test hypotheses in an evidence-generating process (EGP). We claim that EGP variation across researchers adds uncertainty: Non-standard errors (NSEs). We study NSEs by letting 164 teams test the same hypotheses on the same data. NSEs turn out to be sizable, but smaller for better reproducible or higher rated research. Adding peer-review stages reduces NSEs. We further find that this type of uncertainty is underestimated by participants

Robust estimation of bacterial cell count from optical density

Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals &lt;1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data

Discutindo a educação ambiental no cotidiano escolar: desenvolvimento de projetos na escola formação inicial e continuada de professores

A presente pesquisa buscou discutir como a Educação Ambiental (EA) vem sendo trabalhada, no Ensino Fundamental e como os docentes desta escola compreendem e vem inserindo a EA no cotidiano escolar., em uma escola estadual do município de Tangará da Serra/MT, Brasil. Para tanto, realizou-se entrevistas com os professores que fazem parte de um projeto interdisciplinar de EA na escola pesquisada. Verificou-se que o projeto da escola não vem conseguindo alcançar os objetivos propostos por: desconhecimento do mesmo, pelos professores; formação deficiente dos professores, não entendimento da EA como processo de ensino-aprendizagem, falta de recursos didáticos, planejamento inadequado das atividades. A partir dessa constatação, procurou-se debater a impossibilidade de tratar do tema fora do trabalho interdisciplinar, bem como, e principalmente, a importância de um estudo mais aprofundado de EA, vinculando teoria e prática, tanto na formação docente, como em projetos escolares, a fim de fugir do tradicional vínculo “EA e ecologia, lixo e horta”.Facultad de Humanidades y Ciencias de la Educació

Inflammatory cytokines induce human bronchial smooth muscle cell proliferation via an NCX-1 dependent mechanism

Airway smooth muscle hyperplasia is a characteristic of airway remodeling in asthma and this is thought to be, at least in part, cytokine mediated. Because cytosolic free calcium ([Ca²⁺]cyt) plays an important role in smooth muscle proliferation, we chose to examine the role of [Ca²⁺]cyt, focusing on the expression of the Na⁺/Ca²⁺ exchanger 1 (NCX1) protein and its link to human airway smooth muscle proliferation. In vitro studies were done to examine the function and expression of NCX1 protein in human bronchial smooth muscle cells (HBSMC). Cells were grown in the presence/absence of the inflammatory cytokines TNF[alpha], IL-13, and IL-33 and assessed for proliferation using a colorimetric assay. Proliferation was induced in the presence of inflammatory cytokines and blocked in the presence of SN-6, a selective NCX1 inhibitor. Immunoblotting, immunocytochemistry, and quantitative PCR revealed that inflammatory cytokines upregulate NCX1 protein and mRNA expression. In vivo studies were done in mice using an ovalbumin model of asthma. NCX1 expression in asthmatic mice airway was compared with control mice. Immunoblotting revealed a substantial increase in NCX1 protein expression in asthmatic mice. We have demonstrated that NCX1 is expressed in HBSMC and that this expression is increased by cytokines associated with asthma. Moreover, cells proliferate with these cytokines, which is blocked by NCX1 inhibition. NCX1 is also expressed and upregulated in asthmatic mice airway. These data suggest that NCX1 may play an important role in airway remodeling associated with asthm