18F-EF5 PET Is Predictive of Response to Fractionated Radiotherapy in Preclinical Tumor Models

We evaluated the relationship between pre-treatment positron emission tomography (PET) using the hypoxic tracer ¹⁸F-[2-(2-nitro-1-H-imidazol-1-yl)-N-(2,2,3,3,3- pentafluoropropyl) acetamide] (¹⁸F-EF5) and the response of preclinical tumor models to a range of fractionated radiotherapies. Subcutaneous HT29, A549 and RKO tumors grown in nude mice were imaged using ¹⁸F-EF5 positron emission tomography (PET) in order to characterize the extent and heterogeneity of hypoxia in these systems. Based on these results, 80 A549 tumors were subsequently grown and imaged using ¹⁸F-EF5 PET, and then treated with one, two, or four fraction radiation treatments to a total dose of 10–40 Gy. Response was monitored by serial caliper measurements of tumor volume. Longitudinal post-treatment ¹⁸F-EF5 PET imaging was performed on a subset of tumors. Terminal histologic analysis was performed to validate ¹⁸F-EF5 PET measures of hypoxia. EF5-positive tumors responded more poorly to low dose single fraction irradiation relative to EF5-negative tumors, however both groups responded similarly to larger single fraction doses. Irradiated tumors exhibited reduced ¹⁸F-EF5 uptake one month after treatment compared to control tumors. These findings indicate that pre- treatment ¹⁸F-EF5 PET can predict the response of tumors to single fraction radiation treatment. However, increasing the number of fractions delivered abrogates the difference in response between tumors with high and low EF5 uptake pre-treatment, in agreement with traditional radiobiology.     

Retinal Development and Ommin Pigment in the Cranchiid Squid Teuthowenia pellucida (Cephalopoda: Oegopsida)

The cranchiid Teuthowenia pellucida, like many deep-sea squid species, possesses large eyes that maximise light sensitivity in a nearly aphotic environment. To assess ontogenetic changes in the visual system, we conducted morphometric and histological analyses of the eyes using specimens from New Zealand collections. While the ratio between eye diameter and mantle length maintained a linear relationship throughout development, histological sections of the retina revealed that the outer photoreceptor layer became proportionally longer as the animal aged, coincident with a habitat shift into deeper, darker ocean strata. Other retinal layers maintained the same absolute thickness as was observed in paralarvae. Granules of the pigment ommin, normally located in the screening layer positioned at the base of the photoreceptors, were also observed at the outer end of the photoreceptor segments throughout the retina in young and mid-sized specimens. Early developmental stages of this species, dwelling in shallow waters, may therefore rely on migratory ommin to help shield photoreceptors from excess light and prevent over-stimulation. The oldest, deeper-dwelling specimens of T. pellucida examined had longer photoreceptors, and little or no migrated ommin was observed; we suggest therefore that short-term adaptive mechanisms for bright light conditions may be used primarily during epipelagic, early life stages in this species.     

Shading and Watering as a Tool to Mitigate the Impacts of Climate Change in Sea Turtle Nests

Increasing sand temperatures resulting from climate change may negatively impact sea turtle nests by altering sex ratios and decreasing reproductive output. We analyzed the effect of nest shading and watering on sand temperatures as climate mitigation strategies in a beach hatchery at Playa Grande, Costa Rica. We set up plots and placed thermocouples at depths of 45cm and 75cm. Half of the plots were shaded and half were exposed to the sun. Within these exposure treatments, we applied three watering treatments over one month, replicating local climatic conditions experienced in this area. We also examined gravimetric water content of sand by collecting sand samples the day before watering began, the day after watering was complete, and one month after completion. Shading had the largest impact on sand temperature, followed by watering and depth. All watering treatments lowered sand temperature, but the effect varied with depth. Temperatures in plots that received water returned to control levels within 10 days after watering stopped. Water content increased at both depths in the two highest water treatments, and 30 days after the end of water application remained higher than plots with low water. While the impacts of watering on sand temperature dissipate rapidly after the end of application, the impacts on water content are much more lasting. Although less effective at lowering sand temperatures than shading, watering may benefit sea turtle clutches by offsetting negative impacts of low levels of rain in particularly dry areas. Prior to implementing such strategies, the natural conditions at the location of interest (e.g. clutch depth, environmental conditions, and beach characteristics) and natural hatchling sex ratios should be taken into consideration. These results provide insight into the effectiveness of nest shading and watering as climate mitigation techniques and illustrate important points of consideration in the crafting of such strategies.     

Lack of Melanopsin Is Associated with Extreme Weight Loss in Mice upon Dietary Challenge

Metabolic disorders have been established as major risk factors for ocular complications and poor vision. However, little is known about the inverse possibility that ocular disease may cause metabolic dysfunction. To test this hypothesis, we assessed the metabolic consequences of a robust dietary challenge in several mouse models suffering from retinal mutations. To this end, mice null for melanopsin (Opn4^-/-), the photopigment of intrinsically photosensitive retinal ganglion cells (ipRGCs), were subjected to five weeks of a ketogenic diet. These mice lost significantly more weight than wild-type controls or mice lacking rod and cone photoreceptors (Pde6b^rd1/rd1). Although ipRGCs are critical for proper circadian entrainment, and circadian misalignment has been implicated in metabolic pathology, we observed no differences in entrainment between Opn4^-/- and control mice. Additionally, we observed no differences in any tested metabolic parameter between these mouse strains. Further studies are required to establish the mechanism giving rise to this dramatic phenotype observed in melanopsin-null mice. We conclude that the causality between ocular disease and metabolic disorders merits further investigation due to the popularity of diets that rely on the induction of a ketogenic state. Our study is a first step toward understanding retinal pathology as a potential cause of metabolic dysfunction.     

Zinc Prevents Sickness Behavior Induced by Lipopolysaccharides after a Stress Challenge in Rats

Sickness behavior is considered part of the specific beneficial adaptive behavioral and neuroimmune changes that occur in individuals in response to infectious/inflammatory processes. However, in dangerous and stressful situations, sickness behavior should be momentarily abrogated to prioritize survival behaviors, such as fight or flight. Taking this assumption into account, we experimentally induced sickness behavior in rats using lipopolysaccharides (LPS), an endotoxin that mimics infection by gram-negative bacteria, and then exposed these rats to a restraint stress challenge. Zinc has been shown to play a regulatory role in the immune and nervous systems. Therefore, the objective of this study was to examine the effects of zinc treatment on the sickness response of stress-challenged rats. We evaluated 22-kHz ultrasonic vocalizations, open-field behavior, tumor necrosis factor α (TNF-α), corticosterone, and brain-derived neurotrophic factor (BDNF) plasma levels. LPS administration induced sickness behavior in rats compared to controls, i.e., decreases in the distance traveled, average velocity, rearing frequency, self-grooming, and number of vocalizations, as well as an increase in the plasma levels of TNF-α, compared with controls after a stressor challenge. LPS also decreased BDNF expression but did not influence anxiety parameters. Zinc treatment was able to prevent sickness behavior in LPS-exposed rats after the stress challenge, restoring exploratory/motor behaviors, communication, and TNF-α levels similar to those of the control group. Thus, zinc treatment appears to be beneficial for sick animals when they are facing risky/stressful situations.     

Connecting Proline and γ-Aminobutyric Acid in Stressed Plants through Non-Enzymatic Reactions

The accumulation of proline (Pro) in plants exposed to biotic/abiotic stress is a well-documented and conserved response in most vegetal species. Stress conditions induce the overproduction of reactive oxygen species which can lead to cellular damage. In vitro assays have shown that enzyme inactivation by hydroxyl radicals (^·OH) can be avoided in presence of Pro, suggesting that this amino acid could act as an ^·OH scavenger. We applied Density Functional Theory coupled with a polarizable continuum model to elucidate how Pro reacts with ^·OH. In this work we suggest that Pro reacts favourably with ^·OH by H–abstraction on the amine group. This reaction produces the spontaneous decarboxylation of Pro leading to the formation of pyrrolidin-1-yl. In turn, pyrrolidin-1-yl can easily be converted to Δ¹-pyrroline, the substrate of the enzyme Δ¹-pyrroline dehydrogenase, which produces γ-aminobutyric acid (GABA). GABA and Pro are frequently accumulated in stressed plants and several protective roles have been assigned to these molecules. Thereby we present an alternative non-enzymatic way to synthetize GABA under oxidative stress. Finally this work sheds light on a new beneficial role of Pro accumulation in the maintenance of photosynthetic activity.     

Territoriality of Giant Otter Groups in an Area with Seasonal Flooding

Territoriality carries costs and benefits, which are commonly affected by the spatial and temporal abundance and predictability of food, and by intruder pressure. Giant otters (Pteronura brasiliensis) live in groups that defend territories along river channels during the dry season using chemical signals, loud vocalizations and agonistic encounters. However, little is known about the territoriality of giant otters during the rainy season, when groups leave their dry season territories and follow fish dispersing into flooded areas. The objective of this study was to analyze long-term territoriality of giant otter groups in a seasonal environment. The linear extensions of the territories of 10 giant otter groups were determined based on locations of active dens, latrines and scent marks in each season. Some groups overlapped the limits of neighboring territories. The total territory extent of giant otters was correlated with group size in both seasons. The extent of exclusive territories of giant otter groups was negatively related to the number of adults present in adjacent groups. Territory fidelity ranged from 0 to 100% between seasons. Some groups maintained their territory for long periods, which demanded constant effort in marking and re-establishing their territories during the wet season. These results indicate that the defense capacity of groups had an important role in the maintenance of giant otter territories across seasons, which may also affect the reproductive success of alpha pairs.     

Occupancy Modeling for Improved Accuracy and Understanding of Pathogen Prevalence and Dynamics

Most pathogen detection tests are imperfect, with a sensitivity < 100%, thereby resulting in the potential for a false negative, where a pathogen is present but not detected. False negatives in a sample inflate the number of non-detections, negatively biasing estimates of pathogen prevalence. Histological examination of tissues as a diagnostic test can be advantageous as multiple pathogens can be examined and providing important information on associated pathological changes to the host. However, it is usually less sensitive than molecular or microbiological tests for specific pathogens. Our study objectives were to 1) develop a hierarchical occupancy model to examine pathogen prevalence in spring Chinook salmon Oncorhynchus tshawytscha and their distribution among host tissues 2) use the model to estimate pathogen-specific test sensitivities and infection rates, and 3) illustrate the effect of using replicate within host sampling on sample sizes required to detect a pathogen. We examined histological sections of replicate tissue samples from spring Chinook salmon O. tshawytscha collected after spawning for common pathogens seen in this population: Apophallus/echinostome metacercariae, Parvicapsula minibicornis, Nanophyetus salmincola/ metacercariae, and Renibacterium salmoninarum. A hierarchical occupancy model was developed to estimate pathogen and tissue-specific test sensitivities and unbiased estimation of host- and organ-level infection rates. Model estimated sensitivities and host- and organ-level infections rates varied among pathogens and model estimated infection rate was higher than prevalence unadjusted for test sensitivity, confirming that prevalence unadjusted for test sensitivity was negatively biased. The modeling approach provided an analytical approach for using hierarchically structured pathogen detection data from lower sensitivity diagnostic tests, such as histology, to obtain unbiased pathogen prevalence estimates with associated uncertainties. Accounting for test sensitivity using within host replicate samples also required fewer individual fish to be sampled. This approach is useful for evaluating pathogen or microbe community dynamics when test sensitivity is <100%.     

Solution Structure of the HIV-1 Intron Splicing Silencer and Its Interactions with the UP1 Domain of Heterogeneous Nuclear Ribonucleoprotein (hnRNP) A1

Splicing patterns in human immunodeficiency virus type 1 (HIV-1) are maintained through cis regulatory elements that recruit antagonistic host RNA-binding proteins. The activity of the 3′ acceptor site A7 is tightly regulated through a complex network of an intronic splicing silencer (ISS), a bipartite exonic splicing silencer (ESS3a/b), and an exonic splicing enhancer (ESE3). Because HIV-1 splicing depends on protein-RNA interactions, it is important to know the tertiary structures surrounding the splice sites. Herein, we present the NMR solution structure of the phylogenetically conserved ISS stem loop. ISS adopts a stable structure consisting of conserved UG wobble pairs, a folded 2X2 (GU/UA) internal loop, a UU bulge, and a flexible AGUGA apical loop. Calorimetric and biochemical titrations indicate that the UP1 domain of heterogeneous nuclear ribonucleoprotein A1 binds the ISS apical loop site-specifically and with nanomolar affinity. Collectively, this work provides additional insights into how HIV-1 uses a conserved RNA structure to commandeer a host RNA-binding protein.