Extensive infiltration of brain tumors by microglia and macrophages is a hallmark of tumor progression, and yet the overall tumor microenvironment is characterized by an immunosuppressive phenotype. Here we identify esophageal cancer-related gene 4 (Ecrg4) as a novel thrombin-processed monocyte chemoattractant that recruits myeloid cells, promotes their activation, and leads to a blockade of tumor progression.
CT angiography (CTA) is often used for assessing patients with acute ischaemic stroke. Only limited observer reliability data exist. We tested inter- and intra-observer reliability for the assessment of CTA in acute ischaemic stroke.
Understanding life history and demographic variation among species within communities is a central ecological goal. Mortality schedules are especially important in ecosystems where disturbance plays a major role in structuring communities, such as coral reefs. Here, we test whether a trait-based, mechanistic model of mechanical vulnerability in corals can explain mortality schedules. Specifically, we ask whether species that become increasingly vulnerable to hydrodynamic dislodgment as they grow have bathtub-shaped mortality curves, whereas species that remain mechanically stable have decreasing mortality rates with size, as predicted by classical life history theory for reef corals. We find that size-dependent mortality is highly consistent between species with the same growth form and that the shape of size-dependent mortality for each growth form can be explained by mechanical vulnerability. Our findings highlight the feasibility of predicting assemblage-scale mortality patterns on coral reefs with trait-based approaches.
Current antiangiogenic therapies have led to the observation that such agents can lead to improved tumor vessel structure and function termed "vascular normalization" which reduces tumor burden. However, vessel normalization is a transient process, and patients often develop resistance/poor response to anti-vascular strategies that remains an important clinical challenge. Therefore, increasing effort has been made to better understand the cellular and molecular mechanisms of vascular normalization and its contribution to immunomodulation. Herein, we summarize the recent effort to better understand the cellular and molecular mechanisms of vascular normalization with a focus on preclinical genetic models. These studies remain important directions for a mechanistic understanding of the complexities of the maintenance of BBB integrity and the impact of its breakdown on tumor dissemination and pharmaco-distribution of therapeutics.
A 75-year-old man presented with a spontaneous right pneumothorax and pneumopericardium following right atrial pacemaker lead perforation, which is a rare complication, with only two case reports being documented in the literature to the authors' knowledge. It is important that radiologists be aware of cardiac lead perforations, as they may be the first to diagnose this complication.
Determining when corals reproduce has clear management and economic implications. Here we document the reproductive condition of corals in the genus Acropora on the island of Socotra in Yemen during February 2014. Twenty percent of colonies (n = 143) contained mature gametes and 28% had immature gametes indicating that spawning will occur in both February and March in 2014, confirming previous anecdotal reports of coral spawning at this time in Socotra. Acropora typically reproduce in synchrony with many other broadcast spawning scleractinian corals, and we therefore predict that many other species are reproductively active at this time of year.
Otitis media is one of the most common pediatric infections. While it is usually treated without difficulty, up to 20% of children may progress to long-term complications that include hearing loss, impaired speech and language development, academic underachievement, and irreversible disease. Hyperplasia of middle ear mucosa contributes to the sequelae of acute otitis media and is of important clinical significance. Understanding the role of growth factors in the mediation of mucosal hyperplasia could lead to the development of new therapeutic interventions for this disease and its sequelae.
Beliefs, cognitions, and behaviors relating to pain can be associated with a range of negative outcomes. In patients, certain beliefs are associated with increased levels of pain and related disability. There are few data, however, showing the extent to which beliefs of patients differ from those of the general population.
Many organisms have a complex life-cycle in which dispersal occurs at the propagule stage. For marine environments, there is growing evidence that high levels of recruitment back to the natal population (self-recruitment) are common in many marine organisms. For fish, swimming behavior is frequently invoked as a key mechanism allowing high self-recruitment. For organisms with weak-swimming larvae, such as many marine invertebrates, the mechanisms behind self-recruitment are less clear. Here, we assessed whether the combination of passive retention of larvae due to re-circulation processes near reefs, and the dynamics of settlement competence, can produce the high levels of self-recruitment previously estimated by population genetic studies for reef-building corals. Additionally, we investigated whether time to motility, which is more readily measurable than competence parameters, can explain the between-species variation in self-recruitment. We measured the larval competence dynamics of broadcast-spawning and brooding corals and incorporated these in a model of larval retention around reefs to estimate the potential for self-recruitment and assess its variation among species and reefs. Our results suggest that the larvae of many corals, even those with an obligate planktonic phase, develop with sufficient rapidity to allow high levels of self-recruitment, particularly for reefs with long water retention times. Time to motility explained 77-86% of the between-species variation in potential self-recruitment in scenarios with a realistic range of retention times. Among broadcast spawners, time to motility was strongly and positively correlated with egg size, i.e., broadcast spawner species with small eggs developed more rapidly and exhibited greater potential for self-recruitment. These findings suggest that, along with water retention estimates, easy-to-measure species traits, such as egg size and time to motility, may be good predictors of potential self-recruitment, and therefore may be used to characterize the spectrum of self-recruitment in corals.
Coral reef recovery from major disturbance is hypothesized to depend on the arrival of propagules from nearby undisturbed reefs. Therefore, reefs isolated by distance or current patterns are thought to be highly vulnerable to catastrophic disturbance. We found that on an isolated reef system in north Western Australia, coral cover increased from 9% to 44% within 12 years of a coral bleaching event, despite a 94% reduction in larval supply for 6 years after the bleaching. The initial increase in coral cover was the result of high rates of growth and survival of remnant colonies, followed by a rapid increase in juvenile recruitment as colonies matured. We show that isolated reefs can recover from major disturbance, and that the benefits of their isolation from chronic anthropogenic pressures can outweigh the costs of limited connectivity.
Demographic connectivity requires both the dispersal of individuals between sub-populations, and their subsequent contribution to population dynamics. For planktonic, non-feeding marine larvae, the capacity to delay settlement enables greater dispersal distances, but the energetic cost of delayed settlement has been shown to adversely impact post-settlement fitness in several taxa. Here, we assess whether delayed settlement influences mortality rates or growth rates for the first 6 weeks following settlement of the scleractinian coral, Acropora tenuis. Coral larvae that were settled at 2, 4, and 6 weeks after spawning, and then deployed in the field, showed negligible effects of delayed settlement on post-settlement survival and time to initial budding for colony formation. Between-cohort differences in budding rate appeared to be explained by temporal variation in the post-settlement acquisition of zooxanthellae. The potential for coral larvae to remain in the pelagic zone for increased periods of time with little to no effect on post-settlement survival and growth suggests that the capacity for delayed settlement is likely to have meaningful demographic consequences for broadcast-spawning reef-building corals, and that the predicted trade-off between delayed settlement and post-settlement fitness is less applicable to reef-building scleractinian corals than other taxa with non-feeding larvae.
We report an inverse relationship between expression of the orphan candidate tumor suppressor gene esophageal cancer related gene 4 (Ecrg4), and the mucosal epithelial cell response to infection in the middle ear (ME). First, we found constitutive Ecrg4 mRNA expression in normal, quiescent ME mucosa that was confirmed by immunostainning of mucosal epithelial cells and immunoblotting of tissue lysates for the 14 kDa Ecrg4 protein. Upon experimental ME infection, Ecrg4 gene expression rapidly decreased by over 80%, between 3 to 48 hrs, post infection. When explants of this infected mucosa were placed in culture and transduced with an adenovirus (AD) encoding Ecrg4 gene (ADEcrg4), the proliferative and migratory responses of mucosal cells were significantly inhibited. ADEcrg4 transduction of control explants from uninfected MEs had no effect on basal growth and migration. Over-expression of Ecrg4 in vivo, by pre-injecting MEs with ADEcrg4 48 hrs prior to infection, prevented the natural down-regulation of Ecrg4, reduced mucosal proliferation and prevented inflammatory cell infiltration normally observed after infection. Taken together, these data support a hypothesis that Ecrg4 plays a role in coordinating the inflammatory and proliferative response to infection of mucosal epithelium suggesting a possible mechanism for its putative anti-tumor activity.
The normal blood-brain barrier (BBB) consists of tight interendothelial cell junctions and adjacent astrocyte end feet separated by a basal lamina surrounding the endothelium. The interactions between the different cell types of BBB are disrupted in distinct patterns in the microenvironment of glioma. Malignant gliomas infiltrate the surrounding normal brain parenchyma; a process associated with vascular permeability (VP) and breakdown of the BBB. Herein, we describe methods to quantitatively measure glioma-induced vascular permeability, utilizing an orthotopic xenograft model of glioma.
The blood-brain barrier (BBB) is a multicellular vascular structure separating blood from the brain parenchyma that is composed of endothelial cells with tight intercellular junctions, surrounded by a basal lamina, astrocytes, and pericytes. Previous studies have generated detailed databases of the microvessel transcriptome; however, less information is available on the BBB at the protein level. In this study, we specifically focused on characterization of the membrane fraction of cells within the BBB to generate a more complete understanding of membrane transporters, tight junction proteins, and associated extracellular matrix proteins that are functional hallmarks of the BBB. We used Multidimensional Protein Identification Technology to identify a total of 1,143 proteins in mouse brain microvessels, of which 53% were determined to be membrane associated. Analyses of specific classes of BBB-associated proteins in the context of recent transcriptome reports provide a unique database to assess the relative contribution of genes at the level of both RNA and protein in the maintenance of normal BBB integrity.
The characterization of molecular responses following cerebral ischemia-induced changes in animal models capable of undergoing real-time analysis is an important goal for stroke research. In this study, we use transgenic mice to examine the activation of two different promoters in a firefly luciferase reporter mouse analyzable through a non-invasive bioluminescent imaging system. In the first model, we examine the middle cerebral artery occlusion (MCAO)-induced activation of Smad-binding elements (SBE), a downstream target of Smad 1/2/3 transcription factors, in which SBEs regulate the expression of the fluc reporter. We observed that MCAO induces a bilateral activation (i.e., both ipsilateral and contralateral brain hemispheres) of the SBE-luc reporter with a peak at 24 h. In the second model, we examined MCAO-induced activation of the osmolarity-sensitive promoter nuclear factor of activated T-cell 5 (NFAT5) and identified a peak reporter expression 72 h post-MCAO in the ipsilateral but not contralateral hemisphere. In each of these models, the assessment of post-MCAO fluc-expression provided both a quantitative measure (i.e., radiance in photons/sec/cm(2) /steradian) as well as qualitative localization of the molecular response following focal ischemic injury.
By virtue of its ability to regulate the composition of cerebrospinal fluid (CSF), the choroid plexus (CP) is ideally suited to instigate a rapid response to traumatic brain injury (TBI) by producing growth regulatory proteins. For example, Esophageal Cancer Related Gene-4 (Ecrg4) is a tumor suppressor gene that encodes a hormone-like peptide called augurin that is present in large concentrations in CP epithelia (CPe). Because augurin is thought to regulate senescence, neuroprogenitor cell growth and differentiation in the CNS, we evaluated the kinetics of Ecrg4 expression and augurin immunoreactivity in CPe after CNS injury. Adult rats were injured with a penetrating cortical lesion and alterations in augurin immunoreactivity were examined by immunohistochemistry. Ecrg4 gene expression was characterized by in situ hybridization. Cell surface augurin was identified histologically by confocal microscopy and biochemically by sub-cellular fractionation. Both Ecrg4 gene expression and augurin protein levels were decreased 24-72 hrs post-injury but restored to uninjured levels by day 7 post-injury. Protein staining in the supraoptic nucleus of the hypothalamus, used as a control brain region, did not show a decrease of auguin immunoreactivity. Ecrg4 gene expression localized to CPe cells, and augurin protein to the CPe ventricular face. Extracellular cell surface tethering of 14 kDa augurin was confirmed by cell surface fractionation of primary human CPe cells in vitro while a 6-8 kDa fragment of augurin was detected in conditioned media, indicating release from the cell surface by proteolytic processing. In rat CSF however, 14 kDa augurin was detected. We hypothesize the initial release and proteolytic processing of augurin participates in the activation phase of injury while sustained Ecrg4 down-regulation is dysinhibitory during the proliferative phase. Accordingly, augurin would play a constitutive inhibitory function in normal CNS while down regulation of Ecrg4 gene expression in injury, like in cancer, dysinhibits proliferation.
Drug delivery to the central nervous system requires the use of specific portals to enable drug entry into the brain and, as such, there is a growing need to identify processes that can enable drug transfer across both blood-brain and blood-cerebrospinal fluid barriers. Phage display is a powerful combinatorial technique that identifies specific peptides that can confer new activities to inactive particles. Identification of these peptides is directly dependent on the specific screening strategies used for their selection and retrieval. This chapter describes three selection strategies, which can be used to identify peptides that target the choroid plexus (CP) directly or for drug translocation across the CP and into cerebrospinal fluid.
The blood-brain barrier (BBB) is a monolayer of endothelial cells that is regulated by the proximity of a unique basement membrane and a tightly controlled molecular interaction between specialized subsets of cells including pericytes, astrocytes, and neurons. Working together, these cells form a neurovascular unit (NVU) that is dedicated to the local regulation of vascular function in the brain and BBB integrity. Accordingly, the intrinsic complexity of the cell-matrix-cell interactions of the NVU has made analyzing gene function in cell culture, tissue explants, and even animal models difficult and the inability to study gene function in the BBB in vivo has been a critical hurdle to advancing BBB research.Zebrafish has emerged as a premier vertebrate organism to model and analyze complex cellular interactions in vivo and genetic mechanisms of embryonic development. To this end, we provide a technical overview of the procedures that can be used in Zebrafish to analyze BBB integrity with a focus on the cerebrovasculature of adult fish where the BBB is now defined. The techniques that are used to measure the functional integrity, the cell biology, and the ultrastructure of the BBB include permeability assays, fluorescent imaging of reporter genes, and electron microscopy, respectively. Each can be applied to the functional analysis of mutant fish in ways that characterize the molecular sequelae to pathological insults that compromise BBB integrity. Due to the highly conserved nature of both the genetics and cell biology of zebrafish when compared with higher vertebrates, drug discovery techniques can be used in zebrafish models to complement drug development studies in other model systems.
Vagal nerve stimulation (VNS) can have a marked anti-inflammatory effect. We have previously shown that preinjury VNS prevented intestinal barrier breakdown and preserved epithelial tight junction protein expression. However, a pretreatment model has little clinical relevance for the care of the trauma patient. Therefore, we postulated that VNS conducted postinjury would also have a similar protective effect on maintaining gut epithelial barrier integrity.
The bladder exstrophy-epispadias-cloacal exstrophy complex is a spectrum of genitourinary malformations requiring multiple major reconstructive operations on each affected child. The need for surgical correction in this condition often continues through adolescence and into adulthood. Experience in caring for individuals with exstrophy-epispadias has taught us a great deal about the long-term functional, psychological, and social outcomes involved. Children undergoing repeated hospital admissions and extensive multiple operations have the potential for long-term adjustment problems with incontinence, ambulatory difficulties, psychological disturbance, sexual dysfunction, and issues surrounding self-esteem and social integration. By examining relevant published works from the world literature over the last 20 years, in this article we address with each of these areas and offers some insight into the ongoing issues.
The role of the Toll-like receptor 4 (TLR4), a component of the innate immune system, in the development of burn-induced acute lung injury (ALI) has not been completely defined. Recent data suggested that an intact TLR4 plays a major role in the development of organ injury in sterile inflammation. We hypothesized that burn-induced ALI is a TLR4-dependent process. Male C57BL/6J (TLR4 wild-type [WT]) and C57BL/10ScN (TLR4 knockout [KO]) mice were subjected to a 30% total body surface area steam burn. Animals were killed at 6 and 24 h after the insult. Lung specimens were harvested for histological examination after hematoxylin-eosin staining. In addition, lung myeloperoxidase (MPO) and intercellular adhesion molecule 1 immunostaining was performed. Lung MPO was measured by an enzymatic assay. Total lung keratinocyte-derived chemoattractant (IL-8) content was measured by enzyme-linked immunosorbent assay. Western blot was performed to quantify phosphorylated I?B?, phosphorylated nuclear factor ?B p65 (NF-?Bp65), and high mobility group box 1 expression. Acute lung injury, characterized by thickening of the alveolar-capillary membrane, hyaline membrane formation, intraalveolar hemorrhage, and neutrophil infiltration, was seen in WT but not KO animals at 24 h. Myeloperoxidase and intercellular adhesion molecule 1 immunostaining of KO animals was also similar to sham but elevated in WT animals. In addition, a reduction in MPO enzymatic activity was observed in KO mice as well as a reduction in IL-8 levels compared with their WT counterparts. Burn-induced ALI develops within 24 h after the initial thermal insult in our model. Toll-like receptor 4 KO animals were clearly protected and had a much less severe lung injury. Our data suggest that burn-induced ALI is a TLR4-dependent process.
The natural response to injury is dynamic and normally consists of complex temporal and spatial cellular changes in gene expression, which, when acting in synchrony, result in patent tissue repair and, in some instances, regeneration. However, current therapeutic regiments are static and most rely on matrices, gels and engineered skin tissue. Accordingly, there is a need to design next-generation grafting materials to enable biotherapeutic spatiotemporal targeting from clinically approved matrices. To this end, rather then focus on developing completely new grafting materials, we investigated whether phage display could be deployed onto clinically approved synthetic grafts to identify peptide motifs capable of linking pharmaceutical drugs with differential affinities and eventually, control drug delivery from matrices over both space and time.
Molecular phylogenetic studies on scleractinian corals have shown that most taxa are not reflective of their evolutionary histories. Based principally on gross morphology, traditional taxonomy suffers from the lack of well-defined and homologous characters that can sufficiently describe scleractinian diversity. One of the most challenging clades recovered by recent analyses is Bigmessidae, an informal grouping that comprises four conventional coral families, Faviidae, Merulinidae, Pectiniidae and Trachyphylliidae, interspersed among one another with no apparent systematic pattern. There is an urgent need for taxonomic revisions in this clade, but it is vital to first establish phylogenetic relationships within the group. In this study, we reconstruct the evolutionary history of Bigmessidae based on five DNA sequence markers gathered from 76 of the 132 currently recognized species collected from five reef regions in the central Indo-Pacific and the Atlantic.
CNS injury including stroke, infection, and tumor growth lead to astrogliosis, a process that involves upregulation of glial fibrillary acidic protein (GFAP) in astrocytes. However, the kinetics of astrogliosis that is related to these insults (i.e. tumor) is largely unknown.
The content and composition of cerebrospinal fluid (CSF) is determined in large part by the choroid plexus (CP) and specifically, a specialized epithelial cell (CPe) layer that responds to, synthesizes, and transports peptide hormones into and out of CSF. Together with ventricular ependymal cells, these CPe relay homeostatic signals throughout the central nervous system (CNS) and regulate CSF hydrodynamics. One new candidate signal is augurin, a newly recognized 14 kDa protein that is encoded by esophageal cancer related gene-4 (Ecrg4), a putative tumor suppressor gene whose presence and function in normal tissues remains unexplored and enigmatic. The aim of this study was to explore whether Ecrg4 and its product augurin, can be implicated in CNS development and the response to CNS injury.
Because the choroid plexus (CP) is uniquely suited to control the composition of cerebrospinal fluid (CSF), there may be therapeutic benefits to increasing the levels of biologically active proteins in CSF to modulate central nervous system (CNS) functions. To this end, we sought to identify peptides capable of ligand-mediated targeting to CP epithelial cells reasoning that they could be exploited to deliver drugs, biotherapeutics and genes to the CNS.
Dispersal influences ecological dynamics, evolution, biogeography, and biodiversity conservation, but models of larval dispersal in marine organisms make simplifying assumptions that are likely to approximate poorly the temporal dynamics of larval survival and capacity for settlement. In particular, larval mortality rates are typically assumed to be constant throughout larval life; and all larvae are frequently assumed to acquire and lose competence at the same time. To improve upon these assumptions, we here develop simple models of dispersal potential that incorporate rates of mortality, and acquisition and loss of settlement competence. We fit these models to empirical competence and survival data for five scleractinian coral species, to test the models ability to characterize empirical survival and competence patterns, and to estimate the dispersal potential implied by those patterns. The models fit the data well, incorporating qualitative features of competence and survival that traditional approaches to modeling dispersal do not, with important implications for dispersal potential. Most notably, there was high within-cohort variation in the duration of the competent period in all species, and this variation increases both self-recruitment and long-distance dispersal compared with models assuming a fixed competent period. These findings help to explain the seeming paradox of high genetic population structure, coupled with large geographic range size, observed in many coral species. More broadly, our approach offers a way to parsimoniously account for variation in competence dynamics in dispersal models, a phenomenon that our results suggest has important effects on patterns of connectivity in marine metapopulations.
Gliomas generally infiltrate the surrounding normal brain parenchyma, a process associated with increased vascular permeability (VP) and dysregulation of the blood-brain barrier (BBB). However, the molecular mechanisms underlying glioma-induced VP in the brain remain poorly understood. Using a conditional, endothelium-specific deletion of the focal adhesion kinase (FAK) in the mouse (FAK CKO), we show that FAK is critical for destabilization of the tumor endothelium in tumor-bearing mice, with mutant mice exhibiting a relatively normalized vasculature compared with wild-type mice (FAK WT). Tumor vessels in the FAK CKO mice displayed reduced VP compared with FAK WT mice, resulting in reduced tumor growth. Additionally, FAK CKO mice displayed partial restoration of cell-cell junction proteins in the tumor vessels and astrocyte-endothelium interactions in tumors, revealing an additional role of astrocytes in mediating tumor-induced VP. Together, these results provide genetic evidence that FAK is a mediator of tumor-induced VP in the brain. Our findings may help understand how therapeutics might be used to regulate specific cell-type interactions to restore BBB structure/function in cancer and perhaps other pathologic conditions.
Intestinal barrier breakdown following traumatic brain injury (TBI) is characterized by increased intestinal permeability, leading to bacterial translocation, and inflammation. The hormone ghrelin may prevent intestinal injury and have anti-inflammatory properties. We hypothesized that exogenous ghrelin prevents intestinal injury following TBI. A weight-drop model created severe TBI in three groups of anesthetized Balb/c mice. Group TBI: animals underwent TBI only; Group TBI/ghrelin: animals were given 10??g of ghrelin intraperitoneally prior and 1?h following TBI; Group sham: no TBI or ghrelin injection. Intestinal permeability was measured 6?h following TBI by detecting serum levels of FITC-Dextran after injection into the intact ileum. The terminal ileum was harvested for histology, expression of the tight junction protein MLCK and inflammatory cytokine TNF-?. Permeability increased in the TBI group compared to the sham group (109.7?±?21.8??g/mL vs. 32.2?±?10.1??g/mL; p?0.002). Ghrelin prevented TBI-induced permeability (28.3?±?4.2??g/mL vs. 109.7?±?21.8??g/mL; p?0.001). The intestines of the TBI group showed blunting and necrosis of villi compared to the sham group, while ghrelin injection preserved intestinal architecture. Intestinal MLCK increased 73% compared to the sham group (p?0.03). Ghrelin prevented TBI-induced MLCK expression to sham levels. Intestinal TNF-? increased following TBI compared to the sham group (46.2?±?7.1?pg/mL vs. 24.4?±?2.2?pg/mL p?0.001). Ghrelin reduced TNF-? to sham levels (29.2?±?5.0?pg/mL; p?=?NS). We therefore conclude that ghrelin prevents TBI-induced injury, as determined by intestinal permeability, histology, and intestinal levels of TNF-?. The mechanism for ghrelin mediating intestinal protection is likely multifactorial, and further studies are needed to delineate these possibilities.
In traumatic brain injury (TBI), severe disruptions occur in the choroid plexus (CP)-cerebrospinal fluid (CSF) nexus that destabilize the nearby hippocampal and subventricular neurogenic regions. Following invasive and non-invasive injuries to cortex, several adverse sequelae harm the brain interior: (i) structural damage to CP epithelium that opens the blood-CSF barrier (BCSFB) to protein, (ii) altered CSF dynamics and intracranial pressure (ICP), (iii) augmentation of leukocyte traffic across CP into the CSF-brain, (iv) reduction in CSF sink action and clearance of debris from ventricles, and (v) less efficient provision of micronutritional and hormonal support for the CNS. However, gradual post-TBI restitution of the injured CP epithelium and ependyma, and CSF homeostatic mechanisms, help to restore subventricular/subgranular neurogenesis and the cognitive abilities diminished by CNS damage. Recovery from TBI is facilitated by upregulated choroidal/ependymal growth factors and neurotrophins, and their secretion into ventricular CSF. There, by an endocrine-like mechanism, CSF bulk flow convects the neuropeptides to target cells in injured cortex for aiding repair processes; and to neurogenic niches for enhancing conversion of stem cells to new neurons. In the recovery from TBI and associated ischemia, the modulating neuropeptides include FGF2, EGF, VEGF, NGF, IGF, GDNF, BDNF, and PACAP. Homeostatic correction of TBI-induced neuropathology can be accelerated or amplified by exogenously boosting the CSF concentration of these growth factors and neurotrophins. Such intraventricular supplementation via the CSF route promotes neural restoration through enhanced neurogenesis, angiogenesis, and neuroprotective effects. CSF translational research presents opportunities that involve CP and ependymal manipulations to expedite recovery from TBI.
The enteric nervous system may have an important role in modulating gastrointestinal barrier response to disease through activation of enteric glia cells. In vitro studies have shown that enteric glia activation improves intestinal epithelial barrier function by altering the expression of tight junction proteins. We hypothesized that severe injury would increase expression of glial fibrillary acidic protein (GFAP), a marker of enteric glial activation. We also sought to define the effects of vagal nerve stimulation on enteric glia activation and intestinal barrier function using a model of systemic injury and local gut mucosal involvement. Mice with 30% total body surface area steam burn were used as model of severe injury. Vagal nerve stimulation was performed to assess the role of parasympathetic signaling on enteric glia activation. In vivo intestinal permeability was measured to assess barrier function. Intestine was collected to investigate changes in histology; GFAP expression was assessed by quantitative PCR, by confocal microscopy, and in GFAP-luciferase transgenic mice. Stimulation of the vagus nerve prevented injury-induced intestinal barrier injury. Intestinal GFAP expression increased at early time points following burn and returned to baseline by 24 h after injury. Vagal nerve stimulation prior to injury increased GFAP expression to a greater degree than burn alone. Gastrointestinal bioluminescence was imaged in GFAP-luciferase transgenic animals following either severe burn or vagal stimulation and confirmed the increased expression of intestinal GFAP. Injection of S-nitrosoglutathione, a signaling molecule released by activated enteric glia cells, following burn exerts protective effects similar to vagal nerve stimulation. Intestinal expression of GFAP increases following severe burn injury. Stimulation of the vagus nerve increases enteric glia activation, which is associated with improved intestinal barrier function. The vagus nerve may mediate the signaling that occurs from the central nervous system to the enteric nervous system following gastrointestinal injury.
Sexuality and reproductive mode are two fundamental life-history traits that exhibit largely unexplained macroevolutionary patterns among the major groups of multicellular organisms. For example, the cnidarian class Anthozoa (corals and anemones) is mainly comprised of gonochoric (separate sex) brooders or spawners, while one order, Scleractinia (skeleton-forming corals), appears to be mostly hermaphroditic spawners. Here, using the most complete phylogeny of scleractinians, we reconstruct how evolutionary transitions between sexual systems (gonochorism versus hermaphrodism) and reproductive modes (brooding versus spawning) have generated large-scale taxonomic patterns in these characters. Hermaphrodites have independently evolved in three large, distantly related lineages consisting of mostly reef-building species. Reproductive mode in corals has evolved at twice the rate of sexuality, while the evolution of sexuality has been heavily biased: gonochorism is over 100 times more likely to be lost than gained, and can only be acquired by brooders. This circuitous evolutionary pathway accounts for the prevalence of hermaphroditic spawners among reef-forming scleractinians, despite their ancient gonochoric heritage.
Severe injury can cause intestinal permeability through decreased expression of tight junction proteins, resulting in systemic inflammation. Activation of the parasympathetic nervous system after shock through vagal nerve stimulation is known to have potent anti-inflammatory effects; however, its effects on modulating intestinal barrier function are not fully understood. We postulated that vagal nerve stimulation improves intestinal barrier integrity after severe burn through an efferent signaling pathway, and is associated with improved expression and localization of the intestinal tight junction protein occludin.
Because the choroid plexus normally controls the production and composition of cerebrospinal fluid and, as such, its many functions of the central nervous system, we investigated whether ligand-mediated targeting could deliver genes to its secretory epithelium. We show here that when bacteriophages are targeted with epidermal growth factor, they acquire the ability to enter choroid epithelial cells grown in vitro as cell cultures, ex vivo as tissue explants or in vivo by intracerebroventricular injection. The binding and internalization of these particles activate EGF receptors on targeted cells, and the dose- and time-dependent internalization of particles is inhibited by the presence of excess ligand. When the phage genome is further reengineered to contain like green fluorescent protein or firefly luciferase under control of the cytomegalovirus promoter, gene expression is detectable in the choroid plexus and ependymal epithelium by immunohistochemistry or by noninvasive imaging, respectively. Taken together, these data support the hypothesis that reengineered ligand-mediated gene delivery should be considered a viable strategy to increase the specificity of gene delivery to the central nervous system and bypass the blood-brain barrier so as to exploit the biological effectiveness of the choroid plexus as a portal of entry into the brain.
Adult rat hypothalamo-pituitary axis and choroid plexus are rich in basic fibroblast growth factor (FGF2) which likely has a role in fluid homeostasis. Towards this end, we characterized the distribution and modulation of FGF2 in the human and rat central nervous system. To ascertain a functional link between arginine vasopressin (AVP) and FGF2, a rat model of chronic dehydration was used to test the hypothesis that FGF2 expression, like that of AVP, is altered by perturbed fluid balance.
Traumatic brain injury (TBI) causes gastrointestinal dysfunction and increased intestinal permeability. Regulation of the gut barrier may involve the central nervous system. We hypothesize that vagal nerve stimulation prevents an increase in intestinal permeability after TBI.
Toll-like receptor 4 (TLR-4) activation after sterile injury leads to organ dysfunction at distant sites. We have shown previously that intestinal barrier breakdown and alteration of tight junction proteins follows thermal injury; however, the role of TLR-4 in this process remains unclear. We hypothesized that increased intestinal permeability and barrier breakdown after burns is a TLR-4 dependent process; hence, knocking down the TLR-4 gene would have a protective effect on burn-induced intestinal dysfunction.
Intestinal injury owing to inflammation, severe trauma, and burn is a leading cause of morbidity and mortality. Currently, animal models employed to study the intestinal response to injury and inflammation depend on outdated methods of analysis. Given that these classic intestinal assays are lethal to the experimental animal, there is no ability to study the gut response to injury in the same animal over time. We postulated that by developing an in vivo assay to image intestinal injury using fluorescent dye, it could complement other expensive, time-consuming, and semiquantitative classic means of detecting intestinal injury. We describe a novel in vivo, noninvasive method to image intestinal injury using a charge-coupled device (CCD) camera that allows for serial visual and quantitative analysis of intestinal injury. Our results correlate with traditional, time-consuming, semiquantitative assays of intestinal injury, now allowing the noninvasive, nonlethal assessment of injury over time.
Intestinal barrier breakdown after severe burn can lead to intestinal inflammation, which may act as the source of the systemic inflammatory response. In vitro intestinal cell studies have shown that mitogen-activated protein kinase (MAPK) signaling is an important modulator of intestinal inflammation. We have previously observed that pentoxifylline (PTX) attenuates burn-induced intestinal permeability and tight junction breakdown. We hypothesized that PTX would limit intestinal barrier breakdown and attenuate inflammatory signaling via the MAPK pathway.
Because there are few reports using gene delivery in clinically-approved synthetic matrices, we examined the feasibility of using a noninvasive imaging system to study the kinetics of luciferase gene expression when delivered in an adenoviral vector. Using a mouse model of full thickness injury, we quantified the kinetics of gene expression, determined the optimal dose of particle delivery, and established the temporal importance of drug delivery in obtaining optimal gene expression. Specifically, we found that the ideal time to deliver adenovirus to a graft is during the early phase of graft wound closure (days 0-3 post-operatively) for a peak of gene expression to occur 7 days after delivery. Under these conditions, there is a saturating dose of 6 x 10(8) adenoviral particles per graft. In light of these findings, we examined whether the efficacy of delivery could be increased by modulating the composition of the grafts. When a collagen gene-activated matrix (GAM) containing basic fibroblast growth factor (FGF2) was compared to matrix alone, a significant increase in gene expression is observed when identical amounts of vector are delivered (p<0.05). Taken together, these results show how a noninvasive and quantitative assessment of gene expression can be used to optimize gene delivery and that the composition of matrices can dramatically influence gene expression in the wound bed.
In previous studies, we showed that gene activated matrices (GAMs) containing nonviral vectors successfully deliver genes to neurons after optic nerve and spinal cord injury. In the present study, we evaluated whether adenoviral vectors delivered within a GAM increase the efficiency of local gene delivery to injured CNS neurons. Lyophilized GAMs containing collagen and adenoviral vectors were assessed in vitro and in vivo.
Smoking is the largest single cause of preventable death and disease in Australia. This study describes smoking prevalence and the characteristics of rural smokers to guide general practitioners in targeting particular groups.
Loss of intestinal barrier function after burn injury allows movement of intraluminal contents across the mucosa, which can lead to the development of distant organ injury and multiple organ failure. Tight junction function is highly regulated by membrane-associated proteins including occludin and zonula occludens protein 1 (ZO-1), which can be modulated by systemic inflammation. We hypothesized that (1) burn injury leads to gut barrier injury, and (2) phosphodiesterase inhibition will attenuate these burn-induced changes. Male balb/c mice undergoing a 30% steam burn were randomized to resuscitation with normal saline or normal saline + pentoxifylline (PTX; 12.5 mg/kg). Intestinal injury was assessed by histological diagnosis and TNF-alpha levels using enzyme-linked immunosorbent assay. Intestinal permeability was assessed by measuring the plasma concentration of fluorescein isothiocyanate-dextran after intraluminal injection in the distal ileum. Occludin and ZO-1 levels were analyzed by immunoblotting and immunohistochemistry. Thirty percent total body surface area (TBSA) burn results in a significant increase in intestinal permeability. Treatment with PTX after burn attenuates intestinal permeability to sham levels. Burn injury resulted in a marked decrease in the levels of tight junction proteins occludin and ZO-1 at 6 and 24 h. The use of PTX after burn significantly decreases the breakdown of occludin and ZO-1. Pentoxifylline also attenuates the burn-induced increase in plasma and intestinal TNF-alpha. Confocal microscopy demonstrates that PTX attenuates the burn-induced reorganization of occludin and ZO-1 away from the tight junction. Pentoxifylline attenuates burn-induced intestinal permeability and decreases the breakdown and reorganization of intestinal occludin and ZO-1. Therefore, phosphodiesterase inhibition may be a useful adjunct strategy in the attenuation of burn-induced gut barrier injury.
Brachytherapy involves the therapeutic implantation of a radio-active seed source into, or close to, prostate cancer. We report the rare case of a 76-year-old man who presented with a prostate abscess after months of intractable pelvic pain following prostate cancer treatment with iodine-125 brachytherapy. Despite multiple investigations, the diagnosis was made only once the abscess discharged exudate per-urethra.
The normal BBB (blood-brain barrier) consists of a series of structures collectively known as neurovascular units, or NVU, that are composed of endothelial cells and astrocyte endfeet separated by a basal lamina at their interface. The integrity of the BBB and specifically endothelial tight junctions is maintained by interactions between these different components and the local microenvironment of the NVU. Central nervous system cancers such as gliomas disrupt the integrity of the BBB and this compromise is associated with increased tumor growth and invasion of the surrounding brain parenchyma. Because the relationship between glioma-induced BBB breakdown and glioma invasion remains poorly understood, and the host microenvironment can influence tumor cell migration, we used immunohistochemical techniques to characterize tumor associated BBB remodeling. Using an orthotopic xenograft model of glioma, we demonstrate that tumor cells induce specific changes in the composition of the basal lamina and in astrocytic components of the NVU. We suggest that these changes may be essential to understand the capacity of gliomas to regulate BBB integrity and as such, glioma invasion into brain parenchyma.
The use of engineered tissue for the treatment of a variety of acute to chronic wounds has become a clinical standard, and a better understanding of the cellular mechanisms of re-vascularization and barrier integrity could enhance clinical outcomes. Here, we focus on the characterization of the re-vascularization of acellular grafts such as Integra in an animal model to better understand the physiological properties of blood vessels growing in the collagen-glycosaminoglycan matrix vs. wound margins. While Integra has been extensively studied in pre-clinical models, the re-modeling mechanisms of the capillary bed under these matrices are not well understood. Therefore, our first objective was to quantify the kinetics of re-vascularization. The second objective was to assess changes in vascular permeability (VP) of the wound bed compared to normal adjacent skin. The third objective was to establish a non-invasive and quantitative assay for the measurement of VP to facilitate the rapid and reproducible characterization of vascular integrity. Using an excisional wound model in mice, we characterize the appearance, growth, and maturation of blood vessels in an Integra graft over 28 days after surgery. Initial appearance of blood vessels in the graft was observed at 7 days, with angiogenesis peaking between 7 and 14 days. The onset of VP coincided with the increase in re-vascularization of the wound bed and there was a sustained elevation of VP that declined to baseline by 28 days. We propose a non-invasive strategy to assess VP of the wound capillary bed will facilitate a better understanding of the cell and molecular basis of angiogenesis in wound healing.
Traumatic brain injury (TBI) can lead to several physiologic complications including gastrointestinal dysfunction. Specifically, TBI can induce an increase in intestinal permeability, which may lead to bacterial translocation, sepsis, and eventually multi-system organ failure. However, the exact mechanism of increased intestinal permeability following TBI is unknown. We hypothesized that expression of tight junction protein ZO-1 and occludin, responsible for intestinal architectural and functional integrity, will decrease following TBI and increase intestinal permeability. BALB/c mice underwent a weight drop TBI model following anesthesia. Brain injury was confirmed by a neurologic assessment and gross brain pathology. Six hours following injury, FITC-dextran (25 mg 4.4 kDa FITC-dextran) was injected into the intact lumen of the isolated ileum. Intestinal permeability was measured in plasma 30 min following injection, by using spectrophotometry to determine plasma FITC-dextran concentrations. Whole ileum extracts were used to measure expression of tight junction proteins ZO-1 and occludin by Western blot. TBI caused a significant increase in intestinal permeability (110.0 microg/mL +/-22.2) compared to sham animals (29.4 microg/mL +/- 9.7) 6 h after injury (p = 0.016). Expression of ZO-1 was decreased by 49% relative to sham animals (p < 0.02), whereas expression of occludin was decreased by 73% relative to sham animals (p < 0.001). An increase in intestinal permeability corresponds with decreased expression of tight junction proteins ZO-1 and occludin following TBI. Expression of intestinal tight junction proteins may be an important factor in gastrointestinal dysfunction following brain injury.
Coral bleaching caused by global warming is one of the major threats to coral reefs. Very recently, research has focused on the possibility of corals switching symbionts as a means of adjusting to accelerating increases in sea surface temperature. Although symbionts are clearly of fundamental importance, many aspects of coral bleaching cannot be readily explained by differences in symbionts among coral species. Here we outline several potential mechanisms by which the host might influence the bleaching response, and conclude that predicting the fate of corals in response to climate change requires both members of the symbiosis to be considered equally.
Under normal conditions, the intestinal mucosa acts as a local barrier to prevent the influx of luminal contents. The intestinal epithelial tight junction is comprised of several membrane associated proteins, including zonula occludens-1 (ZO-1) and occludin. Disruption of this barrier can lead to the production of pro-inflammatory mediators and ultimately multiple organ failure. We have previously shown that Pentoxifylline (PTX) decreases histologic gut injury and pro-inflammatory mediator synthesis. We hypothesize that PTX prevents the breakdown of ZO-1 and occludin in an in vitro model of immunostimulated intestinal cell monolayers.
Severe injury results in intestinal barrier dysfunction that may be responsible for significant morbidity and mortality. We postulated that mining a peptide library that was displayed on phage would identify peptide sequences that bind and internalize into the gut epithelium following injury.
Burn injury can result in loss of intestinal barrier function, leading to systemic inflammatory response syndrome and multiorgan failure. Myosin light chain kinase (MLCK), a tight junction protein involved in the regulation of barrier function, increases intestinal epithelial permeability when activated. Prior studies have shown that tumor necrosis factor (TNF)-alpha activates MLCK, in part through a nuclear factor (NF)-kappa B-dependent pathway. We have previously shown that pentoxifylline (PTX) decreases both TNF-alpha synthesis and NF-kappaB activation in models of shock. Therefore, we postulate that PTX will attenuate activation of the tight junction protein MLCK, which may decrease intestinal tight junction permeability after severe burn.
The aim of this article was to assess the level and prevalence of major chronic disease risk factors among rural adults. Two cross-sectional surveys were carried out in 2004 and 2005 in the southeast of South Australia and the southwest of Victoria. Altogether 891 randomly selected persons aged 25 to 74 years participated in the studies. Surveys included a self-administered questionnaire, physical measurements, and a venous blood specimen for lipid analyses. Two thirds of participants had cholesterol levels>or=5.0 mmol/L. The prevalence of high diastolic blood pressure (>or=90 mm Hg) was 22% for men and 10% for women in southeast of South Australia, and less than 10% for both sexes in southwest of Victoria. Two thirds of participants were overweight or obese (body mass index>or=25 kg/m2). About 15% of men and slightly less women were daily smokers. The abnormal risk factor levels underline the need for targeted prevention activities in the Greater Green Triangle region. Continuing surveillance of levels and patterns of risk factors is fundamentally important for planning and evaluating preventive activities.
Severe burn results in intestinal barrier breakdown, which may lead to the generation of a systemic inflammatory response and distant organ injury. Intestinal barrier integrity is regulated, in part, by the tight junction protein myosin light chain kinase (MLCK). Previous studies in cell culture have shown that activation of p38 MAPK plays an important role in modulating intestinal barrier function. We hypothesized that (1) severe burn up-regulates p38 MAPK activation and results in increased intestinal permeability via augmented expression of MLCK, and (2) inhibition of p38 MAPK will prevent the burn-induced increase in MLCK expression, resulting in improved intestinal barrier integrity.
The size structure of coral populations is the culmination of key demographic events, including recruitment, mortality and growth, thereby providing important insights to recent ecological dynamics. Importantly, the size structure of corals reflects both intrinsic (inherent life-history characteristics) and extrinsic (enhanced mortality due to chronic or acute disturbances) forcing on local populations, enabling post-hoc assessment of spatial and taxonomic differences in susceptibility to disturbance. This study examined the size structure of four locally abundant corals (Acropora downingi, Favia pallida, Platygyra daedalea, and massive Porites spp.) in two regions of the Persian Gulf: the southern Gulf (Dubai and Abu Dhabi) and eastern Gulf (western Musandam). Significant and consistent differences were apparent in mean colony sizes and size-distributions between regions. All corals in the southern Gulf were significantly smaller, and their size structure positively skewed and relatively more leptokurtic (i.e., peaky) compared to corals in the eastern Gulf. Sea surface temperatures, salinity, and the recent frequency of mass bleaching are all higher, in the southern Gulf, suggesting higher mortality rates and/or slower growth in these populations. Differences in size structure between locations were more pronounced than differences between species at each location, suggesting that extreme differences in environmental conditions and disturbance events have a greater influence on population dynamics in the Gulf than inherent differences in their life-history characteristics.
The coral genus Pocillopora is one of the few to include some species that broadcast spawn gametes and some species that brood larvae, although reports of reproductive mode and timing vary within and among species across their range. Notably, the ubiquitous Pocillopora damicornis has been described as both a brooder and spawner, although evidence of broadcast spawning is rare. Here, we report observations of broadcast-spawning in four species of Pocillopora on the Great Barrier Reef (GBR), including P. damicornis. All species spawned predictably during the early morning, two days following the full moon, and spawning was observed in multiple months over the summer period (November to February). Eggs and sperm were free-spawned concurrently. Eggs were negatively buoyant and contained Symbiodinium. This newfound knowledge on the mode, timing and regularity of broadcast spawning in Pocillopora spp. on the GBR brings us one step closer to elucidating the complex reproductive ecology of these species.
The effectiveness of marine protected areas depends largely on whether people comply with the rules. We quantified temporal changes in benthic composition, reef fish biomass, and fishing effort among marine park zones (including no-take areas) to assess levels of compliance following the 2005 rezoning of the government-controlled Karimunjawa National Park (KNP), Indonesia. Four years after the rezoning awareness of fishing regulations was high amongst local fishers, ranging from 79.5±7.9 (SE) % for spatial restrictions to 97.7±1.2% for bans on the use of poisons. Despite this high awareness and strong compliance with gear restrictions, compliance with spatial restrictions was weak. In the four years following the rezoning reef fish biomass declined across all zones within KNP, with >50% reduction within the no-take Core and Protection Zones. These declines were primarily driven by decreases in the biomass of groups targeted by local fishers; planktivores, herbivores, piscivores, and invertivores. These declines in fish biomass were not driven by changes in habitat quality; coral cover increased in all zones, possibly as a result of a shift in fishing gears from those which can damage reefs (i.e., nets) to those which cause little direct damage (i.e., handlines and spears). Direct observations of fishing activities in 2009 revealed there was limited variation in fishing effort between zones in which fishing was allowed or prohibited. The apparent willingness of the KNP communities to comply with gear restrictions, but not spatial restrictions is difficult to explain and highlights the complexities of the social and economic dynamics that influence the ecological success of marine protected areas. Clearly the increased and high awareness of fishery restrictions following the rezoning is a positive step. The challenge now is to understand and foster the conditions that may facilitate compliance with spatial restrictions within KNP and marine parks worldwide.
Scleractinian corals occur in symbiosis with a range of organisms including the dinoflagellate alga, Symbiodinium, an association that is mutualistic. However, not all symbionts benefit the host. In particular, many organisms within the microbial mucus layer that covers the coral epithelium can cause disease and death. Other organisms in symbiosis with corals include the recently described Chromera velia, a photosynthetic relative of the apicomplexan parasites that shares a common ancestor with Symbiodinium. To explore the nature of the association between C. velia and corals we first isolated C. velia from the coral Montipora digitata and then exposed aposymbiotic Acropora digitifera and A. tenuis larvae to these cultures. Three C. velia cultures were isolated, and symbiosis was established in coral larvae of both these species exposed to all three clones. Histology verified that C. velia was located in the larval endoderm and ectoderm. These results indicate that C. velia has the potential to be endosymbiotic with coral larvae.
Rapid access chest pain clinics have facilitated the early diagnosis and treatment of patients with coronary heart disease and angina. Despite this important service provision, coronary heart disease continues to be under-diagnosed and many patients are left untreated and at risk. Recent advances in imaging technology have now led to the widespread use of noninvasive computed tomography, which can be used to measure coronary artery calcium scores and perform coronary angiography in one examination. However, this technology has not been robustly evaluated in its application to the clinic.
The injured intestine is responsible for significant morbidity and mortality after severe trauma and burn; however, targeting the intestine with therapeutics aimed at decreasing injury has proven difficult. We hypothesized that we could use intravenous phage display technology to identify peptide sequences that target the injured intestinal mucosa in a murine model, and then confirm the cross-reactivity of this peptide sequence with ex vivo human gut. Four hours following 30% TBSA burn we performed an in vivo, intravenous systemic administration of phage library containing 10(12) phage in balb/c mice to biopan for gut-targeting peptides. In vivo assessment of the candidate peptide sequences identified after 4 rounds of internalization was performed by injecting 1×10(12) copies of each selected phage clone into sham or burned animals. Internalization into the gut was assessed using quantitative polymerase chain reaction. We then incubated this gut-targeting peptide sequence with human intestine and visualized fluorescence using confocal microscopy. We identified 3 gut-targeting peptide sequences which caused collapse of the phage library (4-1: SGHQLLLNKMP, 4-5: ILANDLTAPGPR, 4-11: SFKPSGLPAQSL). Sequence 4-5 was internalized into the intestinal mucosa of burned animals 9.3-fold higher than sham animals injected with the same sequence (2.9×10(5)vs. 3.1×10(4) particles per mg tissue). Sequences 4-1 and 4-11 were both internalized into the gut, but did not demonstrate specificity for the injured mucosa. Phage sequence 4-11 demonstrated cross-reactivity with human intestine. In the future, this gut-targeting peptide sequence could serve as a platform for the delivery of biotherapeutics.
Recurrent disturbances can have a critical effect on the structure and function of coral reef communities. In this study, long-term changes were examined in the hard coral community at Wanlitung, in southern Taiwan, between 1985 and 2010. In this 26 year interval, the reef has experienced repeated disturbances that include six typhoons and two coral-bleaching events. The frequency of disturbance has meant that species susceptible to disturbance, such as those in the genus Acropora and Montipora have almost disappeared from the reef. Indeed, almost all hard coral species have declined in abundance, with the result that total hard coral cover in 2010 (17.7%) was less than half what it was in 1985 (47.5%). In addition, macro-algal cover has increased from 11.3% in 2003 to 28.5% in 2010. The frequency of disturbance combined with possible chronic influence of a growing human population mean that a diverse reef assemblage is unlikely to persist on this reef into the future.
The Esophageal cancer-related gene-4 (Ecrg4) is a candidate tumor suppressor gene whose secreted protein product has been implicated in the development and progression of epithelial cancers, neuroprogenitor cell activation after central nervous system injury, cell senescence in neurodegeneration, and the survival of hematopoietic stem cells. Here, we investigated the temporal and spatial localization of Ecrg4 expression in healthy and injured mouse skin, and evaluated the biological activity of Ecrg4 using viral-mediated gene delivery in cutaneous wound healing models. Using in situ hybridization and immunohistochemistry, we found both Ecrg4 mRNA and its protein product localized to the epidermis, dermis, and hair follicles of healthy mouse skin. Upon cutaneous injury, Ecrg4 redistributed to the wound margins where gene microarray and quantitative RT-PCR showed an increased gene expression 5-10 days post-injury as a late phase injury response gene. Ecrg4 over-expression inhibited the directional migration of fibroblasts in modified Boyden chambers in vitro, but had no effect on rates of fibroblast proliferation. Ecrg4 over-expression in vivo at the wound margins delayed the rate of wound closure at 1 and 2 days after full-thickness punch injury. These findings point to the candidate tumor suppressor gene Ecrg4 as a novel, biologically active, constituent of skin and skin injury. The possibility that Ecrg4 serves as a wound termination factor during wound resolution is discussed.
A 48-year-old lady who presented with sepsis secondary to a pelvi-ureteric junction obstruction was treated with an extended course of piperacillin/tazobactam. Four days after completing the course she developed thrombocytopaenia. Intravenous immunoglobulin was required to bring her platelet count back to normal. In the absence of other causes the authors believe that a delayed reaction to piperacillin/tazobactam was the cause of her thrombocytopaenia.
Recent studies have shown that vagus nerve stimulation (VNS) can block the burn-induced systemic inflammatory response (SIRS). In this study we examined the potential for VNS to modulate vascular permeability (VP) in local sites (i.e. skin) and in secondary sites (i.e. lung) following burn. In a 30% total body surface area burn model, VP was measured using intravascular fluorescent dextran for quantification of the VP response in skin and lung. A peak in VP of the skin was observed 24h post-burn injury, that was blocked by VNS. Moreover, in the lung, VNS led to a reduction in burn-induced VP compared to sham-treated animals subjected to burn alone. The protective effects of VNS in this model were independent of the spleen, suggesting that the spleen was not a direct mediator of VNS. These studies identify a role for VNS in the regulation of VP in burns, with the translational potential of attenuating lung complications following burn.
Large surface area burn injuries lead to activation of the innate immune system, which can be blocked by parasympathetic inputs mediated by the vagus nerve. We hypothesized that vagal nerve stimulation (VNS) would alter the inflammatory response of peritoneal macrophages after severe burn injury. Male BALB/c mice underwent right cervical VNS before 30% total body surface area steam burn and were compared with animals subjected to burn alone. Peritoneal macrophages were harvested at several time points following injury and exposed to lipopolysaccharide (LPS) in culture conditions. The inflammatory response of peritoneal macrophages was measured by analyzing changes in nuclear factor ?B p65 phosphorylation using flow cytometry. We found that peritoneal macrophages isolated from mice subjected to burn injury were hyperresponsive to LPS challenge, suggesting burn-induced macrophage activation. We identified a protective role for VNS in blocking peritoneal macrophage activation. Analysis of the phosphorylation state of nuclear factor ?B pathway mediator, p65 Rel A, revealed a VNS-mediated reduction in p65 phosphorylation levels after exposure to LPS compared with burn alone. In combination, these studies suggest VNS mediates the inflammatory response in peritoneal macrophages by affecting the set point of LPS responsiveness.
The human open reading frame C2orf40 encodes esophageal cancer-related gene-4 (Ecrg4), a newly recognized neuropeptide-like precursor protein whose gene expression by cells in vitro, over-expression in mice in vivo, and knock-down in zebrafish affects cell proliferation, migration and senescence, progenitor cell survival and differentiation, and inflammatory function. Unlike traditionally secreted neuropeptide precursors, however, we find that Ecrg4 localizes to the epithelial cell surface and remains tethered after secretion. Here, we used cell surface biotinylation to establish that 14-kDa Ecrg4 localizes to the cell surface of prostate (PC3) or kidney (HEK) epithelial cells after transfection. Accordingly, this Ecrg4 is resistant to washing cells with neutral, high salt (2 M NaCl), acidic (50 mM glycine, pH 2.8), or basic (100 mM Na(2)CO(3), pH 11) buffers. Mutagenesis of Ecrg4 established that cell tethering was mediated by an NH(2)-terminus hydrophobic leader sequence that enabled both trafficking to the surface and tethering. Immunoblotting analyses, however, showed that different cells process Ecrg4 differently. Whereas PC3 cells release cell surface Ecrg4 to generate soluble Ecrg4 peptides of 6-14 kDa, HEK cells do neither, and the 14-kDa precursor resembles a sentinel attached to the cell surface. Because a phorbol ester treatment of PC3 cells stimulated Ecrg4 release from, and processing at, the cell surface, these data are consistent with a multifunctional role for Ecrg4 that is dependent on its cell of origin and the molecular form produced.
Coral reefs, one of the worlds most complex and vulnerable ecosystems, face an uncertain future in coming decades as they continue to respond to anthropogenic climate change, overfishing, pollution, and other human impacts [1, 2]. Traditionally, marine macroecology is based on presence/absence data from taxonomic checklists or geographic ranges, providing a qualitative overview of spatial shifts in species richness that treats rare and common species equally [3, 4]. As a consequence, regional and long-term shifts in relative abundances of individual taxa are poorly understood. Here we apply a more rigorous quantitative approach to examine large-scale spatial variation in the species composition and abundance of corals on midshelf reefs along the length of Australias Great Barrier Reef, a biogeographic region where species richness is high and relatively homogeneous . We demonstrate that important functional components of coral assemblages "sample" space differently at 132 sites separated by up to 1740 km, leading to complex latitudinal shifts in patterns of absolute and relative abundance. The flexibility in community composition that we document along latitudinal environmental gradients indicates that climate change is likely to result in a reassortment of coral reef taxa rather than wholesale loss of entire reef ecosystems.
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