NGF Enhances CGRP Release Evoked by Capsaicin from Rat Trigeminal Neurons: Differential Inhibition by SNAP-25-Cleaving Proteases

Nerve growth factor (NGF) is known to intensify pain in various ways, so perturbing pertinent effects without negating its essential influences on neuronal functions could help the search for much-needed analgesics. Towards this goal, cultured neurons from neonatal rat trigeminal ganglia—a locus for craniofacial sensory nerves—were used to examine how NGF affects the Ca2+-dependent release of a pain mediator, calcitonin gene-related peptide (CGRP), that is triggered by activating a key signal transducer, transient receptor potential vanilloid 1 (TRPV1) with capsaicin (CAP). Measurements utilised neurons fed with or deprived of NGF for 2 days. Acute re-introduction of NGF induced Ca2+-dependent CGRP exocytosis that was inhibited by botulinum neurotoxin type A (BoNT/A) or a chimera of/E and/A (/EA), which truncated SNAP-25 (synaptosomal-associated protein with Mr = 25 k) at distinct sites. NGF additionally caused a Ca2+-independent enhancement of the neuropeptide release evoked by low concentrations (<100 nM) of CAP, but only marginally increased the peak response to ≥100 nM. Notably, BoNT/A inhibited CGRP exocytosis evoked by low but not high CAP concentrations, whereas/EA effectively reduced responses up to 1 µM CAP and inhibited to a greater extent its enhancement by NGF. In addition to establishing that sensitisation of sensory neurons to CAP by NGF is dependent on SNARE-mediated membrane fusion, insights were gleaned into the differential ability of two regions in the C-terminus of SNAP-25 (181–197 and 198–206) to support CAP-evoked Ca2+-dependent exocytosis at different intensities of stimulation.

Genotypic and phenotypic portrait of Candida albicans clinical isolates colonizing the airways of patients with cystic fibrosis

Candida albicans colonizes the respiratory tract of patients with Cystic Fibrosis (CF). It competes with CF-associated pathogens, such as Pseudomonas aeruginosa and Staphylococcus aureus, and contributes to disease severity. We serially recovered 160 C. albicans clinical isolates over a period of 30 months from the sputum of 23 pediatric and 2 adult antifungal-naive CF patients at Children’s Hospital Tunis and characterized the genotype and phenotype of a subset of strains using multilocus sequence typing (MLST) and growth assays on multiple stress-, filamentous growth- and biofilm-inducing media. Out of 16 patients regularly sampled for at least 9 months, 8 and 4 were chronically and transiently colonized with C. albicans, respectively. MLST analyses of 56 strains originating from 15 patients indicated that each patient was colonized with a single strain, while 8 patients (53%) carried isolates from clade 4 known to be enriched with strains from Middle East-Africa. A subset of these isolates with the same sequence type and colonizing 3 unrelated patients displayed altered susceptibility to cell wall-perturbing agents, suggesting changes in cell wall structure/function during growth in the CF lung. We also observed differential ability to filament and/or form biofilms in a set of identical isolates from clade 10 sampled over a period of 9 months in a pediatric CF patient, suggesting alterations in phenotypes associated with virulence. Our findings will rely on future whole-genome sequencing analyses to identify polymorphisms that could explain the emergence of new traits in C. albicans strains thriving in the CF host environment.

Radiomics Nomogram Based on Spectral CT Imaging to Diagnose Osteoporosis

Background: Osteoporosis is associated with a decrease of bone mineralized component as well as a increase of bone marrow fat. At present, there are few studies using radiomics nomogram based fat-water material decomposition (MD) images of spectral CT as an evaluation method of osteoporosis. This study aims to establish and validate a radiomics nomogram based the fat-water imaging of spectral CT in diagnosing osteoporosis.Methods: 95 patients who underwent spectral CT included T11-L2 and dual x-ray absorptiometry (DXA) were collected. The patients were divided into two groups according to T-score, normal bone mineral density (BMD) (T≥-1) and abnormally low BMD (T<-1). Radiomic features were selected from fat-water imaging of the spectral CT. Radscore was calculated by summing the selected features weighted by their coefficients. A nomogram combining the radiomics signature and significant clinical variables was built. The ROC curve was performed to evaluate the performance of the model. Finally, we used decision curve analysis (DCA) to evaluate the clinical usefulness of the model.Results: Five radiomic features based on fat-water imaging of spectral CT were constructed to distinguish abnormally low BMD from normal BMD, and its differential performance was high with an area under the curve (AUC) of 0.95 (95% CI, 0.89-1.00) in the training cohort and 0.97 (95% CI, 0.91-1.00) in the test cohort. The radiomics nomogram showed excellent differential ability with AUC of 0.96 (95%CI, 0.91-1.00) in the training cohort and 0.98 (95%CI, 0.93-1.00) in the test cohort, which performed better than the radiomics model and clinics model only. The DCA showed that the radiomics nomogram had a higher benefit in differentiating abnormally low BMD from normal BMD than the clinical model alone.Conclusion: The radiomics nomogram incorporated radiomics features and clinical factor based the fat-water imaging of spectral CT may serve as an efficient tool to identify abnormally low BMD from normal BMD well.

Neutrophil Phenotypes Result in Differential Responses to Pathogens

Introduction: We have previously reported that neutrophils from healthy subjects vary in their susceptibility to degranulate in response to immune complexes and bacterial ligands (Duarte 2020 [abstract]). In longitudinal testing, neutrophil responses are specific to each individual, resulting in "high" or "low" amounts of degranulation and represent a neutrophil phenotype (Duarte 2019). It is unknown if the phenotype extends to other neutrophil effector functions or if the phenotype is relevant to host-pathogen interactions. Methods/Results: To determine if the neutrophil phenotype extends to other effector functions relevant to pathogen responses, we performed a series of functional assays using whole blood and isolated neutrophils from previously phenotyped "high" and "low" subjects. Our first objective was to determine if the neutrophil degranulation phenotype is conserved in response to a broad range of pathogens. To do this, we chose Staphylococcus aureus (MRSA) strain USA300 (UAMS 1182), Escherichia coli (strain DH5alpha), and Candida albicans (strain SC5314) as model pathogens for gram-positive bacteria, gram-negative bacteria, and fungus, respectively. As shown in Figure 1, degranulation responses were preserved in response to supernatant secreted from all organisms (Fig 1A, p=0.001, p=0.001, and p=0.01 for S. aureus, E. coli, and C. albicans, respectively) and in response to the organism itself (Fig 1B, p=0.01, p=0.005, p=0.005 for respective pathogens). For all organisms, "high" subjects degranulated and released more MMP9 (representative of tertiary granules) when compared to "low" subjects. Besides exocytosis of granules, phagocytosis of pathogens is critical for host defense. To determine if the neutrophil phenotype results in differential ability to phagocytose, neutrophils were isolated from "high" and "low" subjects and uptake of fluorescently-labeled S. aureus bioparticles was measured (Invitrogen, Waltham, MA). As shown in Figure 2A, neutrophils from "high" subjects were less efficient at phagocytosis when compared to neutrophils from "low" subjects (p&lt;0.001). These findings were confirmed by direct visualization using immunofluorescent microscopy. As shown in Figure 2B/2C, neutrophils from "high" subjects had a lower phagocytic index compared to neutrophils from "low" subjects, defined as percent of S. aureus-engulfed neutrophils (p=0.002). Finally, to determine if the neutrophil phenotype results in differential ability to kill, we performed bacterial kill assays using S. aureus. As shown in Figure 3, "high" subjects were less efficient at bacterial kill when compared to "low" subjects resulting in higher bacterial survival at 60 minutes (p&lt;0.001). Conclusions: Taken together, these studies continue to build on our prior observations that neutrophils from healthy subjects vary in their susceptibility to activation, resulting in differential ability to degranulate, phagocytose, and kill pathogens. We demonstrate that excessive exocytosis of granules is correlated with less efficient ability to phagocytose and kill. These differences in effector function are likely relevant to host defense and the innate immune response. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

Reason-giving for resistance: obfuscation, justification and earmarking in resisting informal financial assistance

Monetary assistance is a common request from social ties and can be both an economic lifeline and a financial burden. This study examines the relational work at the heart of such exchanges, examining when and how attempts at mobilizing informal financial assistance are resisted. Using qualitative data from West African factory workers, we demonstrate that individuals who wish to resist the provision of assistance without causing relational damage employ rhetorical strategies designed to justify or obfuscate their refusal, relying heavily on socially legitimate reason-giving. The findings reveal that subjective calculations of resource availability are central to the mobilization of assistance, resistant givers differentiate between justification and obfuscation of refusals, and earmarks can play a role in protecting resources from social demands. More broadly, these findings suggest ways that a differential ability to resist social capital mobilization may generate inequalities within social groups.

Patents in the History of the Semiconductor Industry

Chips can be easily copied and semiconductor firms are not monopolies. Nevertheless, in the semiconductor industry patents protect Ricardian rents against free riding. Ricardian rents—rents wrought by a firm’s differential ability to produce more output or value per unit of input—remunerate the investments in R&D that semiconductor firms make in the expectation of profit. In addition, patents enlarge the set of business models, strategies, and contracts that firms can use to trade. Many practices that emerged over time—for example technical marketing, second sourcing, licensing, trade in intellectual property—and the observed evolution of horizontal and vertical specialization would not have been feasible without patents. Last, patents and Ricardian rents in the semiconductor industry conciliate protracted investments in R&D with exceptionally fast growth of multifactor productivity and falling prices over almost 70 years.

Changes in Executive Function over time in Pediatric Cancer Survivors

Objective. Pediatric oncology survivors are at risk for executive function (EF) and working memory (WM) deficits, which can be measured via performance-based measures or rating scales. Previous studies have shown these measurement methods to be weakly correlated. This study aimed to describe parent rated EF and performance-based working memory (PBWM) in pediatric cancer survivors; examine change in EF and PBWM across time; and investigate the relationship between parent rated WM and PBWM. Method. The sample included 59 patients (50 brain tumor, 9 Leukemia) diagnosed in childhood (Mage=6.92 years; SD=4.12) seen twice for clinical neuropsychological evaluation. PBWM was examined via the auditory working memory scale from a Wechsler intelligence measure or Differential Ability Scales-II. Parents completed the BRIEF/2 as a measure of global EF (GEC), metacognitive skills (MI/CRI), and behavioral regulation (BRI). Results. MI/CRI and GEC at Time 1 were significantly above the mean (p<.01), while PBWM did not differ from the normative mean. GEC, MI/CRI, and BRI were significantly higher than the normative mean at Time 2 (p<.05). PBWM was both clinically and statistically elevated (p<.001). There was a significant change across time in GEC, MI/CRI, and PBWM (p<.05), but not BRI. PBWM was only weakly correlated with the BRIEF WM subscale at Time 1 and Time 2 (all p>.05). Conclusions. Multiple measures of EF should be considered when providing diagnoses and recommendations for pediatric cancer survivors. Furthermore, given declines across time, findings document need for continued monitoring and re-assessment of pediatric survivors as they get further out from treatment.

SARS-CoV-2 Isolates Show Impaired Replication in Human Immune Cells but Differential Ability to Replicate and Induce Innate Immunity in Lung Epithelial Cells

With the rapid spread of the coronavirus disease 2019 (COVID-19) pandemic, information on the replication of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and regulation of innate immunity in human immune cells and lung epithelial cells is needed. In the present study, we show that SARS-CoV-2 failed to productively infect human immune cells, but different isolates of SARS-CoV-2 showed differential ability to replicate and regulate innate interferon responses in human lung epithelial Calu-3 cells.

Determination of IgG1 and IgG3 SARS-CoV-2 spike protein and nucleocapsid binding. Who is binding who and why?

The involvement of IgG3 within the humoral immune response to SARS-CoV2 infection has been implicated in the pathogenesis of ARDS in COVID-19. The exact molecular mechanism is unknown but is thought to involve this IgG subtypes differential ability to fix complement and stimulate cytokine release. We examined convalescent patients antibodies binding to immobilised nucleocapsid and spike protein by MALDI-ToF mass spectrometry. IgG3 was a major immunoglobulin found in all samples. Differential analysis of the spectral signatures found for nucleocapsid versus spike protein demonstrated that the predominant humoral immune response to nucleocapsid was IgG3, whilst against spike it was IgG1. However, the spike protein displayed a strong affinity for IgG3 itself which it would bind from control plasma samples as well as from those previously infected with SARS-CoV2, much in the way Protein-G binds IgG1. Furthermore, detailed spectral analysis indicated a mass shift consistent with hyper-glycosylation or glycation was a characteristic of the IgG3 captured by the spike protein.