Human amylin-derived oligomers and aggregates are believed to play an important role in the pathogenesis of type II diabetes mellitus (T2DM). In addition to amylin-evoked cell attrition, T2DM is often accompanied by elevated serum copper levels. Although previous studies have shown that human amylin, in the course of its aggregation, produces hydrogen peroxide (H2O2) in solution, and that this process is exacerbated in the presence of copper(II) ions (Cu2+), very little is known about the mechanism of interaction between Cu2+ and amylin in pancreatic β-cells, including its pathological significance. Hence, in this study we investigated the mechanism by which Cu2+ and human amylin catalyze formation of reactive oxygen species (ROS) in cells and in vitro, and examined the modulatory effect of Cu2+ on amylin aggregation and toxicity in pancreatic rat insulinoma (RIN-m5F) β-cells. Our results indicate that Cu2+ interacts with human and rat amylin to form metalo-peptide complexes with low aggregative and oxidative properties. Human and non-amyloidogenic rat amylin produced minute (nM) amounts of H2O2, the accumulation of which was slightly enhanced in the presence of Cu2+. In a marked contrast to human and rat amylin, and in the presence of the reducing agents glutathione and ascorbate, Cu2+ produced μM concentrations of H2O2 surpassing the amylin effect by several fold. The current study shows that human and rat amylin not only produce but also quench H2O2, and that human but not rat amylin significantly decreases the amount of H2O2 in solution produced by Cu2+ and glutathione. Similarly, human amylin was found to also decrease hydroxyl radical formation elicited by Cu2+ and glutathione. Furthermore, Cu2+ mitigated the toxic effect of human amylin by inhibiting activation of pro-apoptotic caspase-3 and stress-kinase signaling pathways in rat pancreatic insulinoma cells in part by stabilizing human amylin in its native conformational state. This sacrificial quenching of metal-catalyzed ROS by human amylin and copper’s anti-aggregative and anti-apoptotic properties suggest a novel and protective role for the copper–amylin complex.

Autism spectrum disorder (ASD) describes a group of neurodevelopmental disorders with core deficits in social communication and manifestation of restricted, repetitive, and stereotyped behaviors. Despite the core symptomatology, ASD is extremely heterogeneous with respect to the severity of symptoms and behaviors. This heterogeneity presents an inherent challenge to all large-scale genome-wide omics analyses. In the present study, we address this heterogeneity by stratifying ASD probands from simplex families according to the severity of behavioral scores on the Autism Diagnostic Interview-Revised diagnostic instrument, followed by re-analysis of existing DNA methylation data from individuals in three ASD subphenotypes in comparison to that of their respective unaffected siblings. We demonstrate that subphenotyping of cases enables the identification of over 1.6 times the number of statistically significant differentially methylated regions (DMR) and DMR-associated genes (DAGs) between cases and controls, compared to that identified when all cases are combined. Our analyses also reveal ASD-related neurological functions and comorbidities that are enriched among DAGs in each phenotypic subgroup but not in the combined case group. Moreover, relational gene networks constructed with the DAGs reveal signaling pathways associated with specific functions and comorbidities. In addition, a network comprised of DAGs shared among all ASD subgroups and the combined case group is enriched in genes involved in inflammatory responses, suggesting that neuroinflammation may be a common theme underlying core features of ASD. These findings demonstrate the value of phenotype definition in methylomic analyses of ASD and may aid in the development of subtype-directed diagnostics and therapeutics.

The endosomal innate receptor CD158d (KIR2DL4) induces cellular senescence in human NK cells in response to soluble ligand (HLA-G or agonist antibody). These senescent NK cells display a senescence-associated secretory phenotype (SASP) and their secretome promotes vascular remodeling and angiogenesis. To understand how CD158d initiates signaling for a senescence response, we mapped the region in its cytoplasmic tail that controls signaling. We identified a conserved TRAF6 binding motif, which was required for CD158d-induced NF-κB activation and IL-8 secretion, for TRAF6 association with CD158d, and for TRAF6 recruitment to CD158d+ endosomes in transfected cells. The adaptor TRAF6 is known to couple proximal signals from receptors such as endosomal TLRs and CD40 through the kinase TAK1 for NF-κB-dependent pro-inflammatory responses. siRNA-mediated silencing of TRAF6 and TAK1, and inhibition of TAK1 blocked CD158d-dependent IL-8 secretion. Stimulation of primary, resting NK cells with soluble Ab to CD158d induced TRAF6 association with CD158d, induced TAK1 phosphorylation, and inhibition of TAK1 blocked the CD158d-dependent reprogramming of NK cells that produces the SASP signature. Our results reveal that a prototypic TLR and TNF-receptor signaling pathway is used by a killer cell immunoglobulin-like receptor that promotes secretion of pro-inflammatory and pro-angiogenic mediators as part of a unique senescence phenotype in NK cells.

Background: Cholera surveillance relies on clinical diagnosis of acute watery diarrhea. Suspected cholera case definitions have high sensitivity but low specificity, challenging our ability to characterize cholera burden and epidemiology. Our objective was to estimate the proportion of clinically suspected cholera that are true Vibrio cholerae infections and identify factors that explain variation in positivity. Methods: We conducted a systematic review of studies from 2000-2021 that tested ≥10 suspected cholera cases for V. cholerae O1/O139 using culture, PCR and/or a rapid diagnostic test. We estimated diagnostic test sensitivity and specificity using a latent class meta-analysis. We estimated positivity using a random-effects meta-analysis, adjusting for test performance and study methodology. Results: We included 113 studies from 28 countries. V. cholerae positivity was lower in studies with representative sampling and lower minimum ages in suspected case definitions. After adjusting for sampling methods, case definitions, and tests, on average half (49%, 95% Credible Interval: 43%-54%) of suspected cases represented true V. cholerae infections, although variation across studies was high. Odds of a suspected case having a true infection were 1.64 (95% Credible Interval: 1.06-2.52) times higher when surveillance was initiated in response to an outbreak than in non-outbreak settings. Conclusions: Burden estimates based on suspected cases alone may overestimate the incidence of medically attended cholera about twofold. However, accounting for cases missed by traditional clinical surveillance is key to unbiased overall cholera burden estimates. Given variability between settings, assumptions about positivity, which are necessary without exhaustive testing, should be based on local data.

Essential physiological functions in eukaryotic cells, such as release of hormones and digestive enzymes, neurotransmission, and intercellular signaling, are all achieved by cell secretion. In regulated (calcium-dependent) secretion, membrane-bound secretory vesicles dock and transiently fuse with specialized, permanent, plasma membrane structures, called porosomes or fusion pores. Porosomes are supramolecular, cup-shaped lipoprotein structures at the cell plasma membrane that mediate and control the release of vesicle cargo to the outside of the cell. The sizes of porosomes range from 150nm in diameter in acinar cells of the exocrine pancreas to 12nm in neurons. In recent years, significant progress has been made in our understanding of the porosome and the cellular activities required for cell secretion, such as membrane fusion and swelling of secretory vesicles. The discovery of the porosome complex and the molecular mechanism of cell secretion are summarized in this article.