Human Cys-loop receptors are important therapeutic targets. High-resolution structures are essential for rational drug design, but only a few are available due to difficulties in obtaining sufficient quantities of protein suitable for structural studies. Although expression of proteins in E. coli offers advantages of high yield, low cost, and fast turnover, this approach has not been thoroughly explored for full-length human Cys-loop receptors because of the conventional wisdom that E. coli lacks the specific chaperones and post-translational modifications potentially required for expression of human Cys-loop receptors. Here we report the successful production of full-length wild type human ␣7nAChR from E. coli. Chemically induced chaperones promote high expression levels of well-folded proteins. The choice of detergents, lipids, and ligands during purification determines the final protein quality. The purified ␣7nAChR not only forms pentamers as imaged by negativestain electron microscopy, but also retains pharmacological characteristics of native ␣7nAChR, including binding to bungarotoxin and positive allosteric modulators specific to ␣7nAChR. Moreover, the purified ␣7nAChR injected into Xenopus oocytes can be activated by acetylcholine, choline, and nicotine, inhibited by the channel blockers QX-222 and phencyclidine, and potentiated by the ␣7nAChR specific modulators PNU-120596 and TQS. The successful generation of functional human ␣7nAChR from E. coli opens a new avenue for producing mammalian Cys-loop receptors to facilitate structure-based rational drug design.Human Cys-loop receptors are promising therapeutic targets for various neurological disorders and diseases (1-4). Structure-based drug design for these receptors requires their high-resolution structures (5). Although Cys-loop receptors contain only four major receptor types, including nicotinic acetylcholine receptors (nAChRs), 2 serotonin 5-HT3 receptors, glycine receptors, and GABA A receptors, each receptor type often has multiple subtypes that form numerous functional distinct receptors. Among the human Cys-loop receptors, highresolution structures have been obtained for only the ␤3 GABA A and ␣3 glycine receptors (6, 7). Structures for other eukaryotic Cys-loop receptors include the mouse serotonin 5-HT3A receptor (8), the zebrafish ␣1 glycine receptor (9), the Caenorhabditis elegans GluCl (10, 11) and the muscle-type nicotinic acetylcholine receptor (nAChR) from Torpedo marmorota (12). The dichotomy between the small number of available structures and the relatively large receptor population in the superfamily indicates the technical difficulties for structural determination of these receptors. One of the greatest challenges for structural determination of Cys-loop receptors and similarly complex human membrane proteins is the production of a large quantity of well-folded functional proteins.The ␣7 nAChR (␣7nAChR) is one of the most abundant nAChR subtypes found in the brain (13,14). It is also expressed in a wide variety of non-neuronal tiss...

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Background Perioperative stroke-related mortality in the non-cardiovascular, non-neurological surgery population is an uncommon, yet devastating outcome. A combination of emboli and hypoperfusion may cause the large vessel occlusions leading to perioperative strokes and mortality. Identifying independent risk factors for perioperative stroke-related mortality may enhance risk-stratification algorithms and preventative therapies. Objectives This study utilised cause-of-death data to determine independent risk scores for common surgical comorbidities that may lead to perioperative stroke-related mortality, including atrial fibrillation and asymptomatic carotid stenosis. Methods This retrospective, IRB-exempt, case–control study evaluated non-cardiovascular, non-neurological surgical patients in a claims-based database. ICD-10-CM and ICD-9-CM codes identified cause of death and comorbidity incidences, respectively. A multivariate regression analysis then established adjusted independent risk scores of each comorbidity in relation to perioperative stroke-related mortality. Results Patients with atrial fibrillation were more likely (1.7 aOR, 95% CI (1.1, 2.8) p = 0.02) to die from perioperative stroke-related mortality than from other causes. No association was found with asymptomatic carotid stenosis. Further, in-hospital strokes (25.9 aOR, 95% CI (16.0, 41.8) p < 0.001) or diabetes (1.8 aOR, 95% CI (1.1, 2.9) p = 0.02) may increase perioperative stroke-related mortality risk. Conclusions Atrial fibrillation, diabetes and in-hospital strokes may be independent risk factors for perioperative stroke-related mortality in the non-cardiovascular, non-neurological surgery population.

Coronavirus Disease 2019 (COVID-19) has had a devastating impact on the ability of highly trained healthcare providers to render sufficient care, due to both the significant demand on resources and the unique nature of this disease that make it resistant to traditional therapies. This review sought to determine the potential role of phosphodiesterase-5 inhibitors (PDE-5) in the management of COVID-19 by extrapolating relevant data and clinical studies from other related disease states, including acute respiratory distress syndrome, acute lung injury, and high altitude pulmonary edema. Following a literature search, 4 reports were analyzed and included in this review. While the heterogenicity of data and the small number of trials included limit the interpretation and applicability, it was consistently demonstrated that PDE-5 inhibitors lowered pulmonary arterial pressures. The overall benefit of these agents is seemingly dependent upon the etiology of the respiratory failure, which warrants expanded clinical investigation for COVID-19.