A phosphatidyl-inositol-3-kinase-dependent anti-inflammatory pathway activated by Salmonella in epithelial cells

Salmonella activates the phosphatidyl-inositol-3-kinase (PI3K)/Akt pathway in epithelial cells, but its role in inflammation has not been previously elucidated. We show here that inhibition of PI3K in T84 intestinal epithelial cells results in augmentation of Salmonella-induced interleukin-8 (IL-8) production at the level of both protein and mRNA. The mechanism of this effect appears to involve altered activation of the extracellular growth factor-regulated kinase (ERK), a molecule that is implicated in the regulation of IL-8 expression. These results identify activation of the PI3K/Akt pathway as an anti-inflammatory signal that may contribute to the establishment of Salmonella in the intestine.     

Developmentally regulated intestinal expression of IFN-gamma and its target genes and the age-specific response to enteric Salmonella infection

Young infants are highly susceptible to systemic dissemination of enteric pathogens such as Salmonella typhimurium when compared with older individuals. The mechanisms underlying this differential susceptibility have not been defined clearly. To better understand this phenomenon, we examined the responses of adult mice and preweaned pups to oral infection by S. typhimurium. We found clear age-specific differences, namely, an attenuated intestinal inflammatory response and a higher systemic bacterial burden in the pups compared with the adults. To elucidate the molecular basis for these differences, we obtained a microarray-based profile of gene expression in the small intestines of uninfected adult and preweaned animals. The results indicated a striking age-dependent increase in the intestinal expression of a number of IFN-gamma-regulated genes involved in antimicrobial defense. This finding was confirmed by real-time quantitative PCR, which also demonstrated an age-dependent increase in intestinal expression of IFN-gamma. The developmental up-regulation of the IFN-gamma-regulated genes was dependent on both IFN-gamma and a normal commensal microflora, as indicated by experiments in IFN-gamma-knockout mice and germfree mice, respectively. However, the increase in expression of IFN-gamma itself was independent of the commensal flora. The functional importance of IFN-gamma in the immunological maturation of the intestine was confirmed by the observation that the response of adult IFN-gamma-knockout animals to S. typhimurium infection resembled that of the wild-type pups. Our findings thus reveal a novel role for IFN-gamma in the developmental regulation of antimicrobial responses in the intestine.     

Increased alpha3-fucosylation of alpha1-acid glycoprotein in Type I diabetic patients is related to vascular function

Diabetic mellitus is attended by the development of endothelial dysfunction which is suggested to be accompanied with a chronic low-degree of inflammation. During a chronic hepatic inflammatory response, specific changes in glycosylation of the acute phase protein ₁-acid glycoprotein (AGP) can be detected. In this report we studied the changes in glycosylation of AGP in more detail and evaluated the relation between a change in glycosylation of AGP and urinary albumin secretion in Type I diabetic patients. The glycosylation of AGP, studied by crossed affinity immunoelectrophoresis (CAIE) and high pH anion exchange chromatography with pulse amperometric detection (HPAEC-PAD), showed an increase in 3-fucosylation. Staining with an antibody against sialyl Lewis^x (sLe^x) implied that part of the 3-fucosylation was present in a sLe^x-conformation. In the group of Type I diabetic patients with increased urinary albumin excretion, a significant increase in 3-fucosylation of AGP (p[emsp4 ]<[emsp4 ]0.0005) could be detected. Therefore, the increased 3-fucosylation of AGP can be used as an additional marker for the development of vascular complications in Type I diabetic patients.     

Radiation-hormesis phenotypes,the related mechanisms and implications for disease prevention and therapy

Humans are continuously exposed to ionizing radiation throughout life from natural sources that include cosmic, solar, and terrestrial. Much harsher natural radiation and chemical environments existed during our planet’s early years. Mammals survived the harsher environments via evolutionarily-conserved gifts a continuously evolving system of stress-induced natural protective measures (i.e., activated natural protection [ANP]). The current protective system is differentially activated by stochastic (i.e., variable) low-radiation-dose thresholds and when optimally activated in mammals includes antioxidants, DNA damage repair, p53-related apoptosis of severely-damaged cells, reactive-oxygen-species (ROS)/reactive-nitrogen-species (RNS)- and cytokine-regulated auxiliary apoptosis that selectively removes aberrant cells (e.g., precancerous cells), suppression of disease promoting inflammation, and immunity against cancer cells. The intercellular-signaling-based protective system is regulated at least in part via epigenetic reprogramming of adaptive-response genes. When the system is optimally activated, it protects against cancer and some other diseases, thereby leading to hormetic phenotypes (e.g., reduced disease incidence to below the baseline level; reduced pain from inflammation-related problems). Here, some expressed radiation hormesis phenotypes and related mechanisms are discussed along with their implications for disease prevention and therapy.     

Assessing Population Viability of Black Bears using Spatial Capture-Recapture Models

The Central Georgia Bear Population (CGP) is the least abundant and most isolated of Georgia's 3 American black bear (Ursus americanus) populations. Beginning in 2011, changes to regulations governing harvest of the CGP resulted in an increase in female bear harvest, creating concern that future harvest could be an important influence on population viability. Hence, our objective was to assess viability of the CGP under various levels of female mortality. During 2012–2016, we used barbed-wire hair snares to collect bear hair samples from within the range of the CGP in Georgia, USA. We used microsatellite genotyping to identify individual bears and created robust-design, spatial detection histories for all female bears detected. We fit open population spatial capture-recapture (SCR) models to the detection histories in a Bayesian framework. We used the Widely Applicable Information Criterion (WAIC) to rank models that varied with respect to sources of variation in detection probability, survival, and per capita recruitment, and used the model with the lowest WAIC to forecast dynamics of the CGP 50 years into the future under various levels of female mortality. We assessed the 50-year extinction probability under a continuation of mortality levels documented during 2012–2016, and under incremental increases in female mortality above this baseline. The top model included density-dependent per capita recruitment, annual variation in detection probability, and a trap-level behavioral response. Abundance increased from 106 (95% CI = 86–132) females in 2012 to 136 (95% CI = 113–161) females in 2013 and remained relatively stable thereafter. Annual female survival was 0.75 (95% CI = 0.69–0.82) and did not vary among years. The per capita recruitment rate decreased over time as density increased, and was 0.49 (95% CI = 0.33–0.66) during the first time interval and 0.29 (95% CI = 0.20–0.38) during the final time interval. Annual growth rate () was 1.28 (95% CI = 1.07–1.52) between 2012 and 2013 but decreased throughout the study, ending at 1.04 (95% CI = 0.93–1.17). Forecasts indicated continuation of the female mortality levels experienced from 2012–2016 were sustainable over 50 years, with the estimated extinction risk being <0.001%. Increasing annual harvest by 5 females introduced a negligible increase in the 50-year probability of extinction, but harvesting an additional 10 females/year caused extinction risk to rise to 1.15%. We recommend that harvest regulations are structured such that mortality rates remain at current levels or do not increase by more than an annual average of 5 females above levels observed during our study. Furthermore, we recommend that managers continue to monitor the population so that harvest regulations and population models can be refined over time. © 2020 The Wildlife Society.     

Structure and dynamics of human Nedd4-1 WW3 in complex with the αENaC PY motif

Nedd4-1 (neuronal precursor cell expressed developmentally downregulated gene 4-1) is an E3 ubiquitin ligase that interacts with and negatively regulates the epithelial Na⁺ channel (ENaC). The WW domains of Nedd4-1 bind to the ENaC subunits via recognition of PY motifs. Human Nedd4-1 (hNedd4-1) contains four WW domains with the third domain (WW3*) showing the strongest affinity to the PY motif. To understand the mechanism underlying this binding affinity, we have carried out NMR structural and dynamics analyses of the hNedd4-1 WW3* domain in complex with a peptide comprising the C-terminal tail of the human ENaC α-subunit. The structure reveals that the peptide interacts in a similar manner to other WW domain–ENaC peptide structures. Crucial interactions that likely provide binding affinity are the broad XP groove facilitating additional contacts between the WW3* domain and the peptide, compared to similar complexes, and the large surface area buried (83 Å²) between R430 (WW3*) and L647′ (αENaC). This corroborates the model-free analysis of the ¹⁵N backbone relaxation data, which showed that R430 is the most rigid residue in the domain (S² = 0.90 ± 0.01). Carr–Purcell–Meiboom–Gill relaxation dispersion analysis identified two different conformational exchange processes on the μs–ms time-scale. One of these processes involves residues located at the peptide binding interface, suggesting conformational exchange may play a role in peptide recognition. Thus, both structural and dynamic features of the complex appear to define the high binding affinity. The results should aid interpretation of biochemical data and modeling interfaces between Nedd4-1 and other interacting proteins.     

Evaluation of APOE polymorphisms and the risk for age-related macular degeneration in a Southeastern Brazilian population

This study aimed to evaluate the role of APOE polymorphisms (rs429358 and rs7412) in the risk of age-related macular degeneration in a sample of the Southeastern Brazilian population. Seven hundred and five unrelated individuals were analyzed, 334 with age-related macular degeneration (case group), and 371 without the disease (control group). In the case group, patients were further stratified according to disease phenotypes, divided into dry and wet age-related macular degeneration, and non-advanced and advanced age-related macular degeneration. APOE polymorphisms (rs429358 and rs7412) were evaluated through polymerase chain reaction and direct sequencing. In the comparison of cases vs. controls, none of the associations reached statistical significance, considering the Bonferroni-adjusted P-value, although there was a suggestive protection for the E3/E4 genotype (OR = 0.626; P-value = 0.037) and E4 carriers (OR = 0.6515; P-value = 0.047). Statistically significant protection for both the E3/E4 genotype and E4 carriers was observed in the comparisons: advanced age-related macular degeneration vs. controls (OR = 0.3665, P-value = 0.491 × 10⁻³ and OR = 0.4031, P-value = 0.814 × 10⁻³, respectively), advanced age-related macular degeneration vs. non-advanced age-related macular degeneration (OR = 0.2529, P-value = 0.659 × 10⁻⁴ and OR = 0.2692, P-value = 0.631 × 10⁻⁴, respectively). In the comparison of wet age-related macular degeneration vs. control, protection was statistically significant only for E3/E4 (OR = 0.4052, P-value = 0.001). None of the comparisons demonstrated any significant association for E2 genotypes or E2 carriers in age-related macular degeneration risk in this study. Findings suggest a protective role of the E4 haplotype in the APOE gene in the risk for advanced and wet forms of age-related macular degeneration, in a sample of the Brazilian population. To our knowledge, this is the first Brazilian study to show the association between APOE polymorphisms and age-related macular degeneration.     

Molecular pathogenesis of EBV susceptibility in XLP as revealed by analysis of female carriers with heterozygous expression of SAP

X-linked lymphoproliferative disease (XLP) is a primary immunodeficiency caused by mutations in SH2D1A which encodes SAP. SAP functions in signalling pathways elicited by the SLAM family of leukocyte receptors. A defining feature of XLP is exquisite sensitivity to infection with EBV, a B-lymphotropic virus, but not other viruses. Although previous studies have identified defects in lymphocytes from XLP patients, the unique role of SAP in controlling EBV infection remains unresolved. We describe a novel approach to this question using female XLP carriers who, due to random X-inactivation, contain both SAP(+) and SAP(-) cells. This represents the human equivalent of a mixed bone marrow chimera in mice. While memory CD8(+) T cells specific for CMV and influenza were distributed across SAP(+) and SAP(-) populations, EBV-specific cells were exclusively SAP(+). The preferential recruitment of SAP(+) cells by EBV reflected the tropism of EBV for B cells, and the requirement for SAP expression in CD8(+) T cells for them to respond to Ag-presentation by B cells, but not other cell types. The inability of SAP(-) clones to respond to Ag-presenting B cells was overcome by blocking the SLAM receptors NTB-A and 2B4, while ectopic expression of NTB-A on fibroblasts inhibited cytotoxicity of SAP(-) CD8(+) T cells, thereby demonstrating that SLAM receptors acquire inhibitory function in the absence of SAP. The innovative XLP carrier model allowed us to unravel the mechanisms underlying the unique susceptibility of XLP patients to EBV infection in the absence of a relevant animal model. We found that this reflected the nature of the Ag-presenting cell, rather than EBV itself. Our data also identified a pathological signalling pathway that could be targeted to treat patients with severe EBV infection. This system may allow the study of other human diseases where heterozygous gene expression from random X-chromosome inactivation can be exploited.