RSV Vaccine-Enhanced Disease Is Orchestrated by the Combined Actions of Distinct CD4 T Cell Subsets

There is no currently licensed vaccine for respiratory syncytial virus (RSV) despite being the leading cause of lower respiratory tract infections in children. Children previously immunized with a formalin-inactivated RSV (FI-RSV) vaccine exhibited enhanced respiratory disease following natural RSV infection. Subsequent studies in animal models have implicated roles for CD4 T cells, eosinophils and non-neutralizing antibodies in mediating enhanced respiratory disease. However, the underlying immunological mechanisms responsible for the enhanced respiratory disease and other disease manifestations associated with FI-RSV vaccine-enhanced disease remain unclear. We demonstrate for the first time that while CD4 T cells mediate all aspects of vaccine-enhanced disease, distinct CD4 T cell subsets orchestrate discrete and specific disease parameters. A Th2-biased immune response, but not eosinophils specifically, was required for airway hyperreactivity and mucus hypersecretion. In contrast, the Th1-associated cytokine TNF-α was necessary to mediate airway obstruction and weight loss. Our data demonstrate that individual disease manifestations associated with FI-RSV vaccine-enhanced disease are mediated by distinct subsets of CD4 T cells.     

Differences in the Metabolic Rates of Exploited and Unexploited Fish Populations: A Signature of Recreational Fisheries Induced Evolution?

Non-random mortality associated with commercial and recreational fisheries have the potential to cause evolutionary changes in fish populations. Inland recreational fisheries offer unique opportunities for the study of fisheries induced evolution due to the ability to replicate study systems, limited gene flow among populations, and the existence of unexploited reference populations. Experimental research has demonstrated that angling vulnerability is heritable in Largemouth Bass Micropterus salmoides, and is correlated with elevated resting metabolic rates (RMR) and higher fitness. However, whether such differences are present in wild populations is unclear. This study sought to quantify differences in RMR among replicated exploited and unexploited populations of Largemouth Bass. We collected age-0 Largemouth Bass from two Connecticut drinking water reservoirs unexploited by anglers for almost a century, and two exploited lakes, then transported and reared them in the same pond. Field RMR of individuals from each population was quantified using intermittent-flow respirometry. Individuals from unexploited reservoirs had a significantly higher mean RMR (6%) than individuals from exploited populations. These findings are consistent with expectations derived from artificial selection by angling on Largemouth Bass, suggesting that recreational angling may act as an evolutionary force influencing the metabolic rates of fishes in the wild. Reduced RMR as a result of fisheries induced evolution may have ecosystem level effects on energy demand, and be common in exploited recreational populations globally.     

Meteorologically Driven Simulations of Dengue Epidemics in San Juan,PR

Meteorological factors influence dengue virus ecology by modulating vector mosquito population dynamics, viral replication, and transmission. Dynamic modeling techniques can be used to examine how interactions among meteorological variables, vectors and the dengue virus influence transmission. We developed a dengue fever simulation model by coupling a dynamic simulation model for Aedes aegypti, the primary mosquito vector for dengue, with a basic epidemiological Susceptible-Exposed-Infectious-Recovered (SEIR) model. Employing a Monte Carlo approach, we simulated dengue transmission during the period of 2010–2013 in San Juan, PR, where dengue fever is endemic. The results of 9600 simulations using varied model parameters were evaluated by statistical comparison (r²) with surveillance data of dengue cases reported to the Centers for Disease Control and Prevention. To identify the most influential parameters associated with dengue virus transmission for each period the top 1% of best-fit model simulations were retained and compared. Using the top simulations, dengue cases were simulated well for 2010 (r² = 0.90, p = 0.03), 2011 (r² = 0.83, p = 0.05), and 2012 (r² = 0.94, p = 0.01); however, simulations were weaker for 2013 (r² = 0.25, p = 0.25) and the entire four-year period (r² = 0.44, p = 0.002). Analysis of parameter values from retained simulations revealed that rain dependent container habitats were more prevalent in best-fitting simulations during the wetter 2010 and 2011 years, while human managed (i.e. manually filled) container habitats were more prevalent in best-fitting simulations during the drier 2012 and 2013 years. The simulations further indicate that rainfall strongly modulates the timing of dengue (e.g., epidemics occurred earlier during rainy years) while temperature modulates the annual number of dengue fever cases. Our results suggest that meteorological factors have a time-variable influence on dengue transmission relative to other important environmental and human factors.     

Influence of local-scale abiotic and biotic factors on stress and nutrition in invasive silver carp

Community structure and dynamics in aquatic ecosystems are influenced by a variety of abiotic and biotic factors including productivity, species composition, and temperature. These factors may also affect local-scale community resilience to nonnative species invasions. We used habitat characteristics, zooplankton concentrations, fish abundances, and species composition and richness data collected by two fish population monitoring programs to define relationships that influence stress and nutrition in invasive silver carp (Hypophthalmichthys molitrix). We collected blood samples and quantified nutritional (alkaline phosphatase, cholesterol, protein, and triglycerides) and stress metrics (cortisol and glucose) from individuals across three distinct time periods. Nutritional patterns in silver carp were explained by temperature and food resources, indicating elevated feeding in warmer months. Patterns in blood-based stress parameters were most strongly driven by environmental characteristics, elevating with high water temperatures and increased turbidity. Nutrient levels and community richness parameters did not influence the stress or condition of silver carp, likely due to the absence of limiting resources or competition for this species. Together, our results provide insights into the factors that may influence the spread and distribution of silver carp, as well as the characteristics of habitats that could be vulnerable to future silver carp invasion.     

Lack of REDD1 reduces whole body glucose and insulin tolerance,and impairs skeletal muscle insulin signaling

A lack of the REDD1 promotes dysregulated growth signaling, though little has been established with respect to the metabolic role of REDD1. Therefore, the goal of this study was to determine the role of REDD1 on glucose and insulin tolerance, as well as insulin stimulated growth signaling pathway activation in skeletal muscle. First, intraperitoneal (IP) injection of glucose or insulin were administered to REDD1 wildtype (WT) versus knockout (KO) mice to examine changes in blood glucose over time. Next, alterations in skeletal muscle insulin (IRS-1, Akt, ERK 1/2) and growth (4E-BP1, S6K1, REDD1) signaling intermediates were determined before and after IP insulin treatment (10 min). REDD1 KO mice were both glucose and insulin intolerant when compared to WT mice, evident by higher circulating blood glucose concentrations and a greater area under the curve following IP injections of glucose or insulin. While the REDD1 KO exhibited significant though blunted insulin-stimulated increases (p < 0.05) in Akt S473 and T308 phosphorylation versus the WT mice, acute insulin treatment has no effect (p < 0.05) on REDD1 KO skeletal muscle 4E-BP1 T37/46, S6K1 T389, IRS-1 Y1222, and ERK 1/2 T202/Y204 phosphorylation versus the WT mice. Collectively, these novel data suggest that REDD1 has a more distinct role in whole body and skeletal muscle metabolism and insulin action than previously thought.     

Ablation of LKB1 in the heart leads to energy deprivation and impaired cardiac function

Energy deprivation in the myocardium is associated with impaired heart function and increased morbidity. LKB1 is a kinase that is required for activation of AMP-activated protein kinase (AMPK) as well as 13 AMPK-related protein kinases. AMPK stimulates ATP production during ischemia and prevents post-ischemic dysfunction. We used the Cre–Lox system to generate mice where LKB1 was selectively knocked out in cardiomyocytes and muscle cells (LKB1-KO) to assess the role of LKB1 on cardiac function in these mice.Heart rates of LKB1-KO mice were reduced and ventricle diameter was increased. Ex vivo, cardiac function was impaired during aerobic perfusion of isolated working hearts, and recovery of function after ischemia was reduced. Although oxidative metabolism and mitochondrial function were normal, the AMP/ATP ratio was increased in LKB1-KO hearts. This was associated with a complete ablation of AMPKα2 activity, and a stimulation of signaling through the mammalian target of rapamycin. Our results establish a critical role for LKB1 for normal cardiac function under both aerobic conditions and during recovery after ischemia. Ablation of LKB1 leads to a decreased cardiac efficiency despite normal mitochondrial oxidative metabolism.     

Cannabinoid-mediated modulation of neuropathic pain and microglial accumulation in a model of murine type I diabetic peripheral neuropathic pain

Background

Despite the frequency of diabetes mellitus and its relationship to diabetic peripheral neuropathy (DPN) and neuropathic pain (NeP), our understanding of underlying mechanisms leading to chronic pain in diabetes remains poor. Recent evidence has demonstated a prominent role of microglial cells in neuropathic pain states. One potential therapeutic option gaining clinical acceptance is the cannabinoids, for which cannabinoid receptors (CB) are expressed on neurons and microglia. We studied the accumulation and activation of spinal and thalamic microglia in streptozotocin (STZ)-diabetic CD1 mice and the impact of cannabinoid receptor agonism/antagonism during the development of a chronic NeP state. We provided either intranasal or intraperitoneal cannabinoid agonists/antagonists at multiple doses both at the initiation of diabetes as well as after establishment of diabetes and its related NeP state.

Results

Tactile allodynia and thermal hypersensitivity were observed over 8 months in diabetic mice without intervention. Microglial density increases were seen in the dorsal spinal cord and in thalamic nuclei and were accompanied by elevation of phosphorylated p38 MAPK, a marker of microglial activation. When initiated coincidentally with diabetes, moderate-high doses of intranasal cannabidiol (cannaboid receptor 2 agonist) and intraperitoneal cannabidiol attenuated the development of an NeP state, even after their discontinuation and without modification of the diabetic state. Cannabidiol was also associated with restriction in elevation of microglial density in the dorsal spinal cord and elevation in phosphorylated p38 MAPK. When initiated in an established DPN NeP state, both CB1 and CB2 agonists demonstrated an antinociceptive effect until their discontinuation. There were no pronociceptive effects demonstated for either CB1 or CB2 antagonists.

Conclusions

The prevention of microglial accumulation and activation in the dorsal spinal cord was associated with limited development of a neuropathic pain state. Cannabinoids demonstrated antinociceptive effects in this mouse model of DPN. These results suggest that such interventions may also benefit humans with DPN, and their early introduction may also modify the development of the NeP state.     

Microbial community shifts influence patterns in tropical forest nitrogen fixation

The role of biodiversity in ecosystem function receives substantial attention, yet despite the diversity and functional relevance of microorganisms, relationships between microbial community structure and ecosystem processes remain largely unknown. We used tropical rain forest fertilization plots to directly compare the relative abundance, composition and diversity of free-living nitrogen (N)-fixer communities to in situ leaf litter N fixation rates. N fixation rates varied greatly within the landscape, and ‘hotspots’ of high N fixation activity were observed in both control and phosphorus (P)-fertilized plots. Compared with zones of average activity, the N fixation ‘hotspots’ in unfertilized plots were characterized by marked differences in N-fixer community composition and had substantially higher overall diversity. P additions increased the efficiency of N-fixer communities, resulting in elevated rates of fixation per nifH gene. Furthermore, P fertilization increased N fixation rates and N-fixer abundance, eliminated a highly novel group of N-fixers, and increased N-fixer diversity. Yet the relationships between diversity and function were not simple, and coupling rate measurements to indicators of community structure revealed a biological dynamism not apparent from process measurements alone. Taken together, these data suggest that the rain forest litter layer maintains high N fixation rates and unique N-fixing organisms and that, as observed in plant community ecology, structural shifts in N-fixing communities may partially explain significant differences in system-scale N fixation rates.     

Therapeutic Efficacy of Cintredekin Besudotox (IL13-PE38QQR) in Murine Lung Fibrosis Is Unaffected by Immunity to Pseudomonas aeruginosa Exotoxin A

Background

We have previously explored a therapeutic strategy for specifically targeting the profibrotic activity of IL-13 during experimental pulmonary fibrosis using a fusion protein comprised of human IL-13 and a mutated form of Pseudomonas aeruginosa exotoxin A (IL13-PE) and observed that the intranasal delivery of IL13-PE reduced bleomycin-induced pulmonary fibrosis through its elimination of IL-13-responsive cells in the lung. The aim of the present study was to determine whether the presence of an immune response to P. aeruginosa and/or its exotoxin A (PE) would diminish the anti-fibrotic properties of IL13-PE.

Methodology/Principal Findings

Fourteen days after P. aeruginosa infection, C57BL/6 mice were injected with bleomycin via the intratracheal route. Other groups of mice received 4 doses of saline or IL13-PE by either intranasal or intraperitoneal application, and were challenged i.t. with bleomycin 28 days later. At day 21 after bleomycin, all mice received either saline vehicle or IL13-PE by the intranasal route and histopatological analyses of whole lung samples were performed at day 28 after bleomycin. Intrapulmonary P. aeruginosa infection promoted a neutralizing IgG2A and IgA antibody response in BALF and serum. Surprisingly, histological analysis showed that a prior P. aeruginosa infection attenuated the development of bleomycin-induced pulmonary fibrosis, which was modestly further attenuated by the intranasal administration of IL13-PE. Although prior intranasal administration of IL13-PE failed to elicit an antibody response, the systemic administration of IL13-PE induced a strong neutralizing antibody response. However, the prior systemic sensitization of mice with IL13-PE did not inhibit the anti-fibrotic effect of IL13-PE in fibrotic mice.

Conclusions

Thus, IL13-PE therapy in pulmonary fibrosis works regardless of the presence of a humoral immune response to Pseudomonas exotoxin A. Interestingly, a prior infection with P. aeruginosa markedly attenuated the pulmonary fibrotic response suggesting that the immune elicitation by this pathogen exerts anti-fibrotic effects.     

Effect of active site and surface mutations on the reduction potential of yeast cytochrome c peroxidase and spectroscopic properties of the oxidized and reduced enzyme

The reduction potentials of 22 yeast cytochrome c peroxidase (CcP) mutants were determined at pH 7.0 in order to determine the effect of both heme pocket and surface mutations on the Fe(III)/Fe(II) redox couple of CcP, as well as to determine the range in redox potentials that could be obtained through point mutations in the enzyme. Spectroscopic properties of the Fe(III) and Fe(II) forms of the mutant enzymes are also reported. The mutations include variants in the distal and proximal heme pockets as well as on the enzyme surface and involve single, double, and triple point mutations. A spectrochemical redox titration technique used in this study gave an E(0') value of -189 mV for yeast CcP compared to a previously reported value of -194 mV determined by potentiometry [C.W. Conroy, P. Tyma, P.H. Daum, J.E. Erman, Biochim. Biophys. Acta 537 (1978) 62-69]. Both positive and negative shifts in the reduction potential from that of the wild-type enzyme were observed, spanning a range of 113 mV. The His-52-->Asn mutation gave the most negative potential, -259 mV, while a triple mutant in which the three distal pocket residues, Arg-48, Trp-51, and His-52, were all converted to leucine residues gave the most positive potential, -146 mV.