Cysteamine-mediated blockade of the glycine cleavage system modulates epithelial cell inflammatory and innate immune responses to viral infection.

Transient blockade of glycine decarboxylase (GLDC) can restrict de novo pyrimidine synthesis, which is a well-described strategy for enhancing the host interferon response to viral infection and a target pathway for some licenced anti-inflammatory therapies. The aminothiol, cysteamine, is produced endogenously during the metabolism of coenzyme A, and is currently being investigated in a clinical trial as an intervention in community acquired pneumonia resulting from viral (influenza and SARS-CoV-2) and bacterial respiratory infection. Cysteamine is known to inhibit both bacterial and the eukaryotic host glycine cleavage systems via competitive inhibition of GLDC at concentrations, lower than those required for direct antimicrobial or antiviral activity. Here, we demonstrate for the first time that therapeutically achievable concentrations of cysteamine can inhibit glycine utilisation by epithelial cells and improve cell-mediated responses to infection with respiratory viruses, including human coronavirus 229E and Influenza A. Cysteamine reduces interleukin-6 (IL-6) and increases the interferon-λ (IFN-λ) response to viral challenge and in response to liposomal polyinosinic:polycytidylic acid (poly I:C) simulant of RNA viral infection.

Chiral Indolizinium Salts Derived from 2-Pyridinecarbaldehyde-First Diastereoselective Syntheses of (-)-1-epi-lentiginosine.

The first diastereoselective synthesis of (-)-1-epi-lentiginosine from a common chiral trans-epoxyamide derived from 2-pyridincarbaldehyde is reported. This methodology involves a sequential oxirane ring opening and intramolecular 5-exo-tet cyclization of tosylate trans-epoxyalcohol to afford a diastereomeric mixture of indolizinium salts in a one-pot fashion, followed by regio- and diastereospecific pyridinium ring reduction.

Piperidine-Iodine as Efficient Dual Catalyst for the One-Pot, Three-Component Synthesis of Coumarin-3-Carboxamides.

A simple and efficient one-pot, three-component synthetic method for the preparation of coumarin-3-carboxamides was carried out by the reaction of salicylaldehyde, aliphatic primary/secondary amines, and diethylmalonate. The protocol employs piperidine-iodine as a dual system catalyst and ethanol, a green solvent. The main advantages of this approach are that it is a metal-free and clean reaction, has low catalyst loading, and requires no tedious workup.

Single-Locus-Sequence-Based Typing of the mgpB Gene Reveals Transmission Dynamics in Mycoplasma genitalium.

Sexually transmitted infections (STIs) by Mycoplasma genitalium are a major problem worldwide, especially given their marked and rapid propensity for developing antimicrobial resistance. Since very few treatment options exist, clinicians face an important challenge in the management of the infection. In this scenario, little is known regarding the transmission dynamics of M. genitalium and the epidemiology of antimicrobial resistance. This mgpB-based molecular typing study, conducted among 54 asymptomatically infected individuals prospectively recruited from an STI screening service, reveals two distinct epidemiological clusters that significantly correlate with sexual conduct in heterosexuals and men who have sex with men (MSM), respectively. This well-defined structuration suggests the presence of two independent sexual networks with little connectivity between them. On the other hand, the study demonstrates the multiclonal feature of the emergence of antibiotic resistance in M. genitalium to both macrolides and fluoroquinolones. The high prevalence of macrolide resistance in M. genitalium among MSM, influenced by dense network connectivity and strong antibiotic selective pressure, may correspond to allodemics affecting other STIs such as gonorrhea, syphilis and enteric pathogens. Collaterally, the structural and functional impact of mutations in the mgpB gene, encoding the major adhesin P140 (MgpB), may require further investigation.

Temporal network alignment via GoT-WAVE.

MOTIVATION: Network alignment (NA) finds conserved regions between two networks. NA methods optimize node conservation (NC) and edge conservation. Dynamic graphlet degree vectors are a state-of-the-art dynamic NC measure, used within the fastest and most accurate NA method for temporal networks: DynaWAVE. Here, we use graphlet-orbit transitions (GoTs), a different graphlet-based measure of temporal node similarity, as a new dynamic NC measure within DynaWAVE, resulting in GoT-WAVE. RESULTS: On synthetic networks, GoT-WAVE improves DynaWAVE's accuracy by 30% and speed by 64%. On real networks, when optimizing only dynamic NC, the methods are complementary. Furthermore, only GoT-WAVE supports directed edges. Hence, GoT-WAVE is a promising new temporal NA algorithm, which efficiently optimizes dynamic NC. We provide a user-friendly user interface and source code for GoT-WAVE. AVAILABILITY AND IMPLEMENTATION: http://www.dcc.fc.up.pt/got-wave/. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Structural asymmetry and disulfide bridges among subunits modulate the activity of human malonyl-CoA decarboxylase.

Decarboxylation of malonyl-CoA to acetyl-CoA by malonyl-CoA decarboxylase (MCD; EC 4.1.1.9) is an essential facet in the regulation of fatty acid metabolism. The structure of human peroxisomal MCD reveals a molecular tetramer that is best described as a dimer of structural heterodimers, in which the two subunits present markedly different conformations. This molecular organization is consistent with half-of-the-sites reactivity. Each subunit has an all-helix N-terminal domain and a catalytic C-terminal domain with an acetyltransferase fold (GNAT superfamily). Intersubunit disulfide bridges, Cys-206-Cys-206 and Cys-243-Cys-243, can link the four subunits of the tetramer, imparting positive cooperativity to the catalytic process. The combination of a half-of-the-sites mechanism within each structural heterodimer and positive cooperativity in the tetramer produces a complex regulatory picture that is further complicated by the multiple intracellular locations of the enzyme. Transport into the peroxisome has been investigated by docking human MCD onto the peroxisomal import protein peroxin 5, which revealed interactions that extend beyond the C-terminal targeting motif.

Transdiaphragmatic intercostal hernia induced by sternutation: A case report.

INTRODUCTION AND IMPORTANCE: Sternutation is, by definition, a situation that increases abdominal pressure. However, it has not been clearly linked to protrusion of abdominal content through weaknesses in the abdominal boundaries. CASE PRESENTATION: Here we present a case report in which the only trigger factor found for an abdominal content protrusion was a sternutation episode. The patient arrived in our institution with the diagnosis of a transdiaphragmatic intercostal hernia, proven in CT-scan. He was, then, submitted to emergent surgery, where through thoracotomy and subcostal laparotomy, hernia content was reduced. The patient had a favorable evolution. CLINICAL DISCUSSION: TDIH is a rare entity, for which there are still no consensus regarding its management. This makes clinical practice more challenging, leaving to the surgeon the therapeutic decision tailored to each patient. CONCLUSION: This entity should be further studied, and consensus reached regarding its management.

Synchronous mixed medullary-papillary carcinoma and papillary multifocal carcinoma of the thyroid with cervical lymph node metastases.

Key Clinical Message: The occurrence of simultaneous multifocal papillary thyroid carcinoma and mixed medullary-papillary carcinoma, as far as we know, has not been previously described. We suggest the surgical approach to be driven by the medullary component. Abstract: Patient underwent total thyroidectomy with central compartment lymph node dissection. Histological examination revealed a simultaneous multifocal papillary thyroid carcinoma and mixed medullary-papillary carcinoma. He was disease-free at 1-year-follow-up.

Role of choledochotomy after mechanical lithotripsy-related adverse event in endoscopic retrograde cholangiopancreatography.

In case of rupture of the mechanical lithotripter's traction wires during an ERCP, we suggest performing a choledochotomy to remove the stone, and remove the closed Dormia basket through the mouth.

Discharge protocol in acute pancreatitis: an international survey and cohort analysis.

There are several overlapping clinical practice guidelines in acute pancreatitis (AP), however, none of them contains suggestions on patient discharge. The Hungarian Pancreatic Study Group (HPSG) has recently developed a laboratory data and symptom-based discharge protocol which needs to be validated. (1) A survey was conducted involving all members of the International Association of Pancreatology (IAP) to understand the characteristics of international discharge protocols. (2) We investigated the safety and effectiveness of the HPSG-discharge protocol. According to our international survey, 87.5% (49/56) of the centres had no discharge protocol. Patients discharged based on protocols have a significantly shorter median length of hospitalization (LOH) (7 (5;10) days vs. 8 (5;12) days) p < 0.001), and a lower rate of readmission due to recurrent AP episodes (p = 0.005). There was no difference in median discharge CRP level among the international cohorts (p = 0.586). HPSG-protocol resulted in the shortest LOH (6 (5;9) days) and highest median CRP (35.40 (13.78; 68.40) mg/l). Safety was confirmed by the low rate of readmittance (n = 35; 5%). Discharge protocol is necessary in AP. The discharge protocol used in this study is the first clinically proven protocol. Developing and testifying further protocols are needed to better standardize patients' care.

Spontaneous internal hernia through a defect in the hepatogastric ligament.

Transomental internal hernias are a rare cause of intestinal obstruction and most commonly iatrogenic, resulting from previous surgical interventions, abdominal trauma or inflammation. Occasionally, they may occur spontaneously. We report the case of a 44-year-old healthy male admitted to the emergency room with acute abdominal pain and vomiting, consistent with intestinal obstruction. An internal hernia of small bowel in the lesser sac was suspected after performing a computed tomography (CT) scan and emergent laparotomy confirmed herniation of a jejunal loop through a defect in the hepatogastric ligament, resulting in strangulation and requiring enterectomy. The patient had a favourable outcome and was discharged a few days after surgery. Both radiologists and surgeons must be aware of rare internal hernia subtypes, to avoid delays in diagnosis and treatment. Abdominal CT is the first-line imaging of choice, providing useful diagnostic hallmarks. Nevertheless, surgical exploration is typically essential to confirm the diagnosis, identify the defect and assess bowel viability.

Diastereoselective synthesis of new zwitterionic bicyclic lactams, scaffolds for construction of 2-substituted-4-hydroxy piperidine and its pipecolic acid derivatives.

The synthesis of new chiral highly functionalized zwitterionic bicyclic lactams starting from acyclic ß-enaminoesters derived from (R)-(-)-2-phenylglycinol is described. The key step involved an intramolecular non-classical Corey-Chaykovsky ring-closing reaction of the corresponding sulfonium salts derived from ß-enaminoesters. This methodology permits the generation of two or three new stereogenic centers with high diastereoselectivity. The utility of these intermediates was demonstrated by the stereocontrolled total synthesis of cis-4-hydroxy-2-methyl piperidine and its corresponding pipecolic acid derivative.

Predicting bacterial promoter function and evolution from random sequences.

Predicting function from sequence is a central problem of biology. Currently, this is possible only locally in a narrow mutational neighborhood around a wildtype sequence rather than globally from any sequence. Using random mutant libraries, we developed a biophysical model that accounts for multiple features of σ70 binding bacterial promoters to predict constitutive gene expression levels from any sequence. We experimentally and theoretically estimated that 10-20% of random sequences lead to expression and ~80% of non-expressing sequences are one mutation away from a functional promoter. The potential for generating expression from random sequences is so pervasive that selection acts against σ70-RNA polymerase binding sites even within inter-genic, promoter-containing regions. This pervasiveness of σ70-binding sites implies that emergence of promoters is not the limiting step in gene regulatory evolution. Ultimately, the inclusion of novel features of promoter function into a mechanistic model enabled not only more accurate predictions of gene expression levels, but also identified that promoters evolve more rapidly than previously thought.

The Sialoglycan Binding Adhesins of Mycoplasma genitalium and Mycoplasma pneumoniae.

Mycoplasma genitalium (Mge) and Mycoplasma pneumoniae (Mpn) are two human pathogens associated with urogenital and respiratory tract infections, respectively. The recent elucidation of the tridimensional structure of their major cytoadhesins by X-ray crystallography and cryo-electron microscopy/tomography, has provided important insights regarding the mechanics of infection and evasion of immune surveillance.

Robotic-assisted parathyroidectomy via transaxillary approach: feasibility and learning curves.

BACKGROUND: There have been few reports of robotic-assisted transaxillary parathyroidectomy in the literature. We aim to report our experience with robotic-assisted transaxillary parathyroidectomy for primary hyperparathyroidism (PHPT) in the Western population. METHODS: A retrospective study was performed from July 2010 through July 2019 at two institutions, one in the United States and one in France. Demographic characteristics and perioperative data were collected for all patients undergoing robotic-assisted transaxillary parathyroidectomy by a single surgeon at each institution. A linear regression model was developed to describe the learning curve for this procedure at each institution. RESULTS: One-hundred and two patients with PHPT were included with a median age of 55.6±12.4 years and median body mass index (BMI) of 25.5±6.1 kg/m2. The majority of patients were female (80.4%). Median total operative time was 116±53 minutes. Minor complications were reported in 2 patients (1.96%), and one case was converted to a trans-cervical approach (TCA) for four-gland exploration. Median patient follow-up time was 6.5±12.2 months, and disease recurrence was reported in one patient. Calculated learning curves showed that one surgeon achieved proficiency by the eighth case, and the other achieved proficiency by the fourteenth case. CONCLUSIONS: This is the largest reported experience of robotic-assisted transaxillary parathyroidectomy for PHPT in the Asian and Western population. Analysis of the procedural learning curve demonstrates that proficiency in this technique was achieved after performance of less than 15 surgeries. This procedure is safe and feasible in the hands of experienced surgeons for select patients with localized disease.

Cysteamine Inhibits Glycine Utilisation and Disrupts Virulence in Pseudomonas aeruginosa.

Pseudomonas aeruginosa is a major opportunistic human pathogen which employs a myriad of virulence factors. In people with cystic fibrosis (CF) P. aeruginosa frequently colonises the lungs and becomes a chronic infection that evolves to become less virulent over time, but often adapts to favour persistence in the host with alginate-producing mucoid, slow-growing, and antibiotic resistant phenotypes emerging. Cysteamine is an endogenous aminothiol which has been shown to prevent biofilm formation, reduce phenazine production, and potentiate antibiotic activity against P. aeruginosa, and has been investigated in clinical trials as an adjunct therapy for pulmonary exacerbations of CF. Here we demonstrate (for the first time in a prokaryote) that cysteamine prevents glycine utilisation by P. aeruginosa in common with previously reported activity blocking the glycine cleavage system in human cells. Despite the clear inhibition of glycine metabolism, cysteamine also inhibits hydrogen cyanide (HCN) production by P. aeruginosa, suggesting a direct interference in the regulation of virulence factor synthesis. Cysteamine impaired chemotaxis, lowered pyocyanin, pyoverdine and exopolysaccharide production, and reduced the toxicity of P. aeruginosa secreted factors in a Galleria mellonella infection model. Thus, cysteamine has additional potent anti-virulence properties targeting P. aeruginosa, further supporting its therapeutic potential in CF and other infections.

P2X7 receptors mediate ischemic damage to oligodendrocytes.

Brain ischemia leading to stroke is a major cause of disability in developed countries. Therapeutic strategies have most commonly focused on protecting neurons from ischemic damage. However, ischemic damage to white matter causes oligodendrocyte death, myelin disruption, and axon dysfunction, and it is partially mediated by glutamate excitotoxicity. We have previously demonstrated that oligodendrocytes express ionotropic purinergic receptors. The objective of this study was to investigate the role of purinergic signaling in white matter ischemia. We show that, in addition to glutamate, enhanced ATP signaling during ischemia is also deleterious to oligodendrocytes and myelin, and impairs white matter function. Thus, ischemic oligodendrocytes in culture display an inward current and cytosolic Ca(2+) overload, which is partially mediated by P2X7 receptors. Indeed, oligodendrocytes release ATP after oxygen and glucose deprivation through the opening of pannexin hemichannels. Consistently, ischemia-induced mitochondrial depolarization as well as oxidative stress culminating in cell death are partially reversed by P2X7 receptor antagonists, by the ATP degrading enzyme apyrase and by blockers of pannexin hemichannels. In turn, ischemic damage in isolated optic nerves, which share the properties of brain white matter, is greatly attenuated by all these drugs. Ultrastructural analysis and electrophysiological recordings demonstrated that P2X7 antagonists prevent ischemic damage to oligodendrocytes and myelin, and improved action potential recovery after ischemia. These data indicate that ATP released during ischemia and the subsequent activation of P2X7 receptor is critical to white matter demise during stroke and point to this receptor type as a therapeutic target to limit tissue damage in cerebrovascular diseases.

Alternative conformation of the C-domain of the P140 protein from Mycoplasma genitalium.

The human pathogen Mycoplasma genitalium is responsible for urethritis in men, and for cervicitis and pelvic inflammatory disease in women. The adherence of M. genitalium to host target epithelial cells is mediated through an adhesion complex called Nap, which is essential for infectivity. Nap is a transmembrane dimer of heterodimers of the immunodominant proteins P110 and P140. The M. genitalium genome contains multiple copies of portions that share homology with the extracellular regions of P140 and P110 encoded by the genes mg191 and mg192, respectively. Homologous recombination between the genes and the copies allows the generation of a large diversity of P140 and P110 variants to overcome surveillance by the host immune system. Interestingly, the C-terminal domain (C-domain) of the extracellular region of P140, which is essential for the function of Nap by acting as a flexible stalk anchoring the protein to the mycoplasma membrane, presents a low degree of sequence variability. In the present work, the X-ray crystal structures of two crystal forms of a construct of the P140 C-domain are reported. In both crystal forms, the construct forms a compact octamer with D4 point-group symmetry. The structure of the C-domain determined in this work presents significant differences with respect to the structure of the C-domain found recently in intact P140. The structural plasticity of the C-domain appears to be a possible mechanism that may help in the functioning of the mycoplasma adhesion complex.

Structure and mechanism of the Nap adhesion complex from the human pathogen Mycoplasma genitalium.

Mycoplasma genitalium is a human pathogen adhering to host target epithelial cells and causing urethritis, cervicitis and pelvic inflammatory disease. Essential for infectivity is a transmembrane adhesion complex called Nap comprising proteins P110 and P140. Here we report the crystal structure of P140 both alone and in complex with the N-terminal domain of P110. By cryo-electron microscopy (cryo-EM) and tomography (cryo-ET) we find closed and open Nap conformations, determined at 9.8 and 15 Å, respectively. Both crystal structures and the cryo-EM structure are found in a closed conformation, where the sialic acid binding site in P110 is occluded. By contrast, the cryo-ET structure shows an open conformation, where the binding site is accessible. Structural information, in combination with functional studies, suggests a mechanism for attachment and release of M. genitalium to and from the host cell receptor, in which Nap conformations alternate to sustain motility and guarantee infectivity.

Immunodominant proteins P1 and P40/P90 from human pathogen Mycoplasma pneumoniae.

Mycoplasma pneumoniae is a bacterial human pathogen that causes primary atypical pneumonia. M. pneumoniae motility and infectivity are mediated by the immunodominant proteins P1 and P40/P90, which form a transmembrane adhesion complex. Here we report the structure of P1, determined by X-ray crystallography and cryo-electron microscopy, and the X-ray structure of P40/P90. Contrary to what had been suggested, the binding site for sialic acid was found in P40/P90 and not in P1. Genetic and clinical variability concentrates on the N-terminal domain surfaces of P1 and P40/P90. Polyclonal antibodies generated against the mostly conserved C-terminal domain of P1 inhibited adhesion of M. pneumoniae, and serology assays with sera from infected patients were positive when tested against this C-terminal domain. P40/P90 also showed strong reactivity against human infected sera. The architectural elements determined for P1 and P40/P90 open new possibilities in vaccine development against M. pneumoniae infections.