Progress of inflammation depends on the balance between two biological mechanisms: pro-inflammatory and pro-resolving processes. Many extracellular and intracellular molecular components including cytokines, growth factors, steroids, neurotransmitters, and lipidergic mediators and their receptors contribute to the two processes, generated from cellular participants during inflammation. Fatty acid-derived mediators are crucial in directing the inflammatory phase and orchestrating heterogeneous reactions of participants such as inflamed cells, innate immune cells, vascular components, innervating neurons, etc. As well as activating specific types of receptor molecules, lipidergic mediators can actively control the functions of various ion channels via direct binding and/or signal transduction, thereby altering cellular functions. Lipid mediators can be divided into two classes based on which of the two processes they promote: pro-inflammatory, which includes prostaglandins and leukotrienes, and pro-resolving, which includes lipoxins, resolvins, and maresins. The research on the modulations of neuronal ion channels regarding the actions of the pro-inflammatory class has begun relatively earlier while the focus is currently expanding to cover the ion channel interaction with pro-resolvents. As a result, knowledge of inhibitory mechanisms by the pro-resolvents, historically seldom found for other known endogenous modulators or pro-inflammatory mediators, is accumulating particularly upon sensory neuronal cation channels. Diverse mechanistic explanations at molecular levels are being proposed and refined. Here we overviewed the interactions of lipidergic pro-resolvents with neuronal ion channels and outcomes from the interactions, focusing on transient receptor potential (TRP) ion channels. We also discuss unanswered hypotheses and perspectives regarding their interactions. Inflammation is a protective mechanism of the body against physical/chemical injury, or infection. The inflammatory reaction is associated with functional changes in diverse tissues and cell types. In the past, the progression of inflammation from its initiation through termination was believed to be dependent only on pro-inflammatory mechanisms on the molecular and cellular levels. That is, when pro-inflammatory mediators are upregulated, the inflammatory response may last or even deteriorate because of activations of the cellular players and their signal transduction. Conversely, when those are downregulated, the inflammatory response may passively subside. This indicates that pro-inflammatory mediators, secreted in response to insults, stimulate the participant cells and tissues to become or remain inflamed. In the absence of such stimulation, however, the cells and tissues passively recover from the inflamed state. In the early 2000's, this hypothesis began to be revised Serhan, . Resolut2+) concentrations can significantly affect intracellular signal transduction in many cell types. Moreover, ionic flux is essential for controlling neuronal excitability. Therefore, pro-resolvent interactions with cellular ion-permeating channels, if any, might be an important node for altering the inflammatory phase of cells. In fact, two axial mechanisms regarding such interactions have emerged. One involves ion channel modulation related to sensory neuronal excitability and the other involves that in mucus secreting pathways in the airway . These effects lead to beneficial outcomes with respect to fluid secretion in the airway  and the resultant changes in cellular function explain the molecular mechanism for many of the actions of pro-resolvents on innate immune cells and inflamed tissues. For example, RvE1  activates the GPR, chemR23, and antagonizes the type 1 leukotriene B4 receptor (BLT1). RvD1 (D designates DHA) activates GPR32 and lipoxin A4/Annexin-A1 receptor/formyl-peptide receptor 2 (ALX/FPR2). RvD5 has also been shown to activate GPR32. RvD2 activates GPR18 -inflamed animals. Since RvD1 is locally confined when injected and outcomes were measured at relatively acute time points, the pain relief observed in vivo appears to be mediated in the absence of intercellular actions such as recruitment or activation of innate immune cells.Of particular note, pro-resolvents seem to have a central role for endogenous inhibition of TRPA1. For instance, resolvin D1 (RvD1) strongly blocks TRPA1 activity , acidosis, abnormal hypertonicity, endogenous lipidergic pro-inflammatory messengers such as leukotrienes, and natural pungent compounds such as capsaicin and tarantula toxin evoke pain via direct structural challenges to TRPV1 protein, culminating in its electrical activation . Predictably, acute pain induced by capsaicin and inflammatory heat hyperalgesia, which are the Similarly, RvE1 and MaR1 have been found to inhibit TRPV1 activity and TRPV1-mediated pain behavior, but did not affect modalities involving TRPA1 , another D-series pro-resolving lipid, has also been shown to inhibit TRPV1 albeit one third of the potency of RvD2 , whereas the temperature threshold for TRPV1 activation is close to the range reported to actually cause pain perception in vivo administration of this compound has been shown to mitigate heat pain, a TRPV3-mediated sensory modality, but not mechanical pain under inflammatory conditions -RvD1) has been shown to specifically block TRPV3. The Albeit to a lesser extent than TRPV1 and TRPA1, TRPV4 is also a polymodal sensor ion channel. Elevated temperatures (>27–34°C), mechanical stretching caused by pressure or swelling, and endogenous pro-inflammatory mediators such as epoxyeicosatrienoic acids (EETs) and dimethylallyl pyrophosphate are able to activate TRPV4 , TRPV3 and TRPV4 have been identified as molecular targets of RvD1. However, it remains unclear whether TRPV3 or TRPV4 predominates over the RvD1 effect on the thermal phenotype.RvD1 is the only resolvin to date found to inhibit TRPV4. The Levine lab established an Using the same experimental approach described above for TRPA1 and TRPV3, an indirect ALX/FPR2-mediated GPR pathway has been excluded from the potential molecular interacting mechanism underlying these results. Although, it is unclear without binding assay, modulation via direct contact can be considered from what was recently hypothesized. It is suggested that chemical interactions between the lipid components of the bilayer membrane and the embedded regions of ion channel proteins are important for appropriate ion channel response and gating (Bavi et al., Nociceptor populations among primary sensory neurons propagate peripheral pathologic signals, including those originating from inflammation, to the central nervous system. Moreover, these neurons actively contribute to pro-inflammatory processes via neurogenic inflammation mechanisms, by interactively exchanging various intercellular mediators with innervating tissues. These are important steps in the development of chronic inflammatory pain. In terms of pain relief, the TRPs mentioned above are essentially focused on when resolution mechanisms are examined, since the activation of these ion channels explains the outset of nociceptor excitation and pathologic hyperactivity, which often exacerbate neurogenic inflammation. The actions of pro-resolving lipids through these sensory TRP modulations all seem to converge on pain alleviation.Postsynaptic dorsal horn neurons translate and relay signals received from the synaptic inputs of the central terminals of nociceptor neurons in the spinal cord (Choi et al., Upon RvE1 exposure, presynaptic ERK phosphorylation has also been shown to be suppressed. In addition, the frequency of spontaneous excitatory postsynaptic potentials, which reflects not only postsynaptic function but also the extent of presynaptic glutamate release, was also decreased. This finding may also explain the RvE1-mediated suppression of TRPV1-independent presynaptic TNF-α signals (Xu et al., in vivo treatment of these substances. Based on current efficacy and safety information, translational applications of these substances to some specific diseases are eagerly anticipated. Before this promise can be realized, it would be advantageous to establish proof of concept to clarify the relevant molecular mechanisms. The most ambiguous ones to date are the binding mode by which these pro-resolving lipids interact with channel proteins and the mechanisms how they initiate GPR-mediated signal transduction. Although, one putative lipid-interacting site has been predicted, it has not yet been established whether resolvins modify channel activity through this site. When GPR coupled, it has not been completely answered which GPR predominantly drives the effect and which intracellular signal deactivates the channel. Most studies have focused on the sensory circuit. TRPV channels are involved in plastic changes in other brain synapses (Shibasaki et al., Here we reviewed the effects of various pro-resolving lipids on the functions of four sensory TRP channels and NMDA receptors that convey nociceptive signals. These pro-resolvents commonly showed strong inhibitory actions on these ion channels, leading to a final outcome of pain resolution. It is perhaps surprising that a group of endogenous lipid derivatives from common precursors can contribute to the resolution of two different pathologic processes, pain and inflammation, via differential cellular targets and biological mechanisms. Immunology studies continue to confirm that the absence of adverse effects from 2+-activated anion channels is increased upon pro-resolvent treatment in the airway epithelium (Verriere et al., Lessons from studies of the airway or other tissues may help to frame a different paradigm. In the airway research field, rather than channel activation or inhibition, changes in the expression of an ion channel are a more important axis to define the effect of a pro-resolvent, which have received little attention in the neuroscience field. The activity of CaGC analyzed documented results and prepared the preliminary draft. SH supervised the study and finalized the manuscript. All authors read and approved the final manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Rapid fragmentation and degradation of large undisturbed habitats constitute major threats to biodiversity. Several studies have shown that populations in small and highly isolated habitat patches are prone to strong environmental and demographic stochasticity and increased risk of extinction. Based on community assembly theory, we predict recent rapid forest fragmentation to cause a decline in species and functional guild richness of forest birds combined with a high species turnover among habitat patches, and well defined dominance structures, if competition is the major driver of community assembly. To test these predictions, we analysed species co-occurrence, nestedness, and competitive strength to infer effects of interspecific competition, habitat structure, and species′ traits on the assembly of bird species communities from 12 cloud forest fragments in southern Kenya. Our results do not point to a single ecological driver of variation in species composition. Interspecific competition does not appear to be a major driver of species segregation in small forest patches, while its relative importance appears to be higher in larger ones, which may be indicative for a generic shift from competition-dominated to colonisation-driven community structure with decreasing fragment size. Functional trait diversity was independent of fragment size after controlling for species richness. As fragmentation effects vary among feeding guilds and habitat generalists, in particular, tend to decline in low quality forest patches, we plead for taking species ecology fully into account when predicting tropical community responses to habitat change. Habitat fragmentation has profound and mainly negative effects on the long-term viability of indigenous animal and plant species –2, in pa0Az with S0 being the average number of species per unit area and z being the scaling constant [A becomes Variation in species density, the average number of species per unit area, is closely related to the concept of species-area relationships (SAR). Given the common power function model of the SAR S = Sconstant  the specg effect  predictsg effect . Hence, Classic competition-based community assembly models  predict Mechanisms underlying community assembly are still discussed even after more than half a century of research in the field. Standard analyses of community assembly are based on species occurrence and absence, however, these data are only rarely linked to trait- and environmental variation. When aiming to uncover mechanisms and constraints behind the pattern of species co-existence, there is a clear need to link the geometry of species occurrences with environmental and species functional trait data which can be expected to replace classic co-occurrence analysis that have been dominated the field since the pioneering work of Diamond (1975) . Along tThe Taita bird community has been studied intensively over the past two decades, resulting in well supported knowledge about species richness, abundance, and ecological demands. Furthermore, the land use history of the Taita Hills is very well documented, and this combined information offers a strong framework to study how tropical avian communities are shaped in relation to species and landscape traits. Making use of this information, we here test the following three hypothesis: (i) Avian species density increases with decreasing fragment size, resulting in a high proportion of species surviving in forest fragments relative to intact forests; (ii) Species co-occurrence among small forest remnants is aggregated due to crowding effects; and (iii) Habitat specialists respond more strongly to habitat degradation compared to habitat generalists.2 and are geographically isolated from other mountain blocks to the south (90 km to the Usambara Mts.) and the north (80 km to the Chyulu Hills) (The Taita Hills cover an area of around 250 kmu Hills) . SemiariLand-cover information was derived from airborne true-colour images, converted to orthomosaics at a spatial resolution of 0.5 m . BrightnUnderstorey bird community metrics were derived from a long-term (1996 to 2010) bird ringing program using standard mist-netting procedures as described in Karr (1981) . Mist neFor each bird species we assembled data on body mass (g), average bill culmen length, depth and width (cm), tarsus, tail, and wing length (cm), and average hand-wing index. The dominant principal component of the three bill measures was used to assess bill characteristics, and the respective dominant eigenvector of the wing, tarsus, and tail measures served as a proxy to the type of locomotion . FollowiAnalyses were based on matrices containing species relative abundances with species in rows and forest fragments in columns. We composed matrices for each fragment separately and then grouped the three larger (> 90 ha) and nine smaller (<15 ha) fragments in two additional matrices. We calculated the following three metrics of species co-occurrence. First, we estimated species segregation among fragments (negative species associations) by the C-score , 47, 48 richness . Third, richness  using thrichness , 50, a mrichness  and Turnrichness . Source P, we used a null model approach and compared the observed co-occurrence metric scores with those obtained from 1000 null matrices each that were randomized using a null model that resamples the matrix with placement probabilities proportional to observed total abundances of rows and columns . SES scores should have values below –1.96 and above +1.96 at the two-sided 5% error level under the assumption that the respective null distribution is approximately normal. To account for multiple testing, all significance levels were Bonferroni corrected.For statistical inference of NODF, C-score, and betace model ). This ice model . Note thce model , raw FADC) and chose the best fitting competition matrix to assess the maximum impact of interspecific competition on community assembly. High values of rC point to the possibility that interspecific competition is a major driver for observed species distributions while low rC values point towards a minor impact of competition [Possible competitive interactions among species within the seven feeding guilds were assessed following Ulrich et al. (2014)  and Solipetition .S = 12.4A0.21;r2 = 0.44 , and to species richness as the continuous co-variate.Using the empirical species—area relationship 2 = 0.44  we assesWe recorded a total of 17,520 individuals from 69 bird species in the 12 Taita forest fragments , of whicColumba larvata, Phyllastrephus placidus, Phylloscopus ruficapillus, Turdus olivaceus, Zoothera gurneyi) in the smaller fragments.Average species density was independent of fragment isolation , forest A separate analysis based on forest specialist species only  also revP SES scores  and nine smaller fragments   revealedS scores  further S scores  were on Finally, we compared functional diversity between feeding guilds and between larger and smaller fragments , Table 3Within an island biogeographic framework , speciesThe counter-intuitive finding that species richness in small fragments was only 2% lower than in large ones, is in line with other studies that reported patch connectivity to be of higher importance than patch size , in partPrevious studies showed that bird assemblages strongly vary in composition with habitat area and structure (such as the degree of fragmentation), and predicted higher species richness in large, connected forest patches –69. Yet,Various studies have reported increasing mobility with decreasing habitat integrity –74. Yet,C = 0.36, We were surprised to see that functional trait identity was not linked to the pattern of species—co-occurrence. Species interactions are mediated by traits and classical community assembly theory predicts co-existing species to show low levels of trait similarity . Yet, weTotal trait space, as expressed by FAD, increases with species richness . HoweverIn conclusion, results of this study do not point to a single ecological driver of the observed variation in avian species composition among indigenous fragments of the Taita forest archipelago. Interspecific competition does not appear to be a major driver of species segregation, since in small forest fragments no single competitive strength hierarchy was able to predict at least a major part of the observed species abundances distributions. In larger fragments, however, the relative importance of competition might be higher. We did not find strong evidence for habitat filtering either, and community-wide species co-occurrences and joint occurrences between pairs of species were neither nested nor segregated, as would be expected if competition would be the main driver of species occurrences. The frequency of pairwise species segregation was even much below the level expected under random association. However, the large differences in species density and species richness between the two smallest fragments highlight that local peculiarities might heavily constrain species richness, irrespective of subsequent patterns of co-occurrences. Such variable, and partly opposing, responses of single guilds to fragmentation suggest that it is vital to take species ecology into consideration when predicting community-wide responses to habitat change in tropical forests.S1 Appendix(XLSX)Click here for additional data file.


HIV/AIDS, tuberculosis (TB), and malaria are 3 major global public health threats that undermine development in many resource-poor settings. Recently, the notion that positive selection during epidemics or longer periods of exposure to common infectious diseases may have had a major effect in modifying the constitution of the human genome is being interrogated at a large scale in many populations around the world. This positive selection from infectious diseases increases power to detect associations in genome-wide association studies (GWASs). High-throughput sequencing (HTS) has transformed both the management of infectious diseases and continues to enable large-scale functional characterization of host resistance/susceptibility alleles and loci; a paradigm shift from single candidate gene studies. Application of genome sequencing technologies and genomics has enabled us to interrogate the host-pathogen interface for improving human health. Human populations are constantly locked in evolutionary arms races with pathogens; therefore, identification of common infectious disease-associated genomic variants/markers is important in therapeutic, vaccine development, and screening susceptible individuals in a population. This review describes a range of host-pathogen genomic loci that have been associated with disease susceptibility and resistant patterns in the era of HTS. We further highlight potential opportunities for these genetic markers. HIV/AIDS, tuberculosis, and malaria are 3 major global public health threats causing substantial morbidity, mortality, negative socioeconomic impact, and human suffering . Whole-gIn the past, candidate gene studies have been used to identify disease susceptibility genetic loci for many major human infections. But with the advent of HTS approaches, many new loci have been and continue to be identified in diverse populations. A paradigm shift from candidate gene studies to GWAS and HTS has ushered in a “big data” genomic revolution era enabling us to redefine the genomic architecture of host-pathogen disease susceptibility. Both exome and whole-genome sequencing approaches are proving more successful and affordable. Host genetics influence clinical course of infectious diseases as well as genetic variation of the pathogens determining their survival in presence of selective pressure from the host and environment  and identify genetic markers of drug-resistant pathogens and parasites. Furthermore, HTS has given us unprecedented resolution of understanding the role of host genetics to infectious diseases susceptibility. This genomic revolution has generated data informative for understanding the frequency of many genetic traits, including those that cause disease susceptibility in African populations and populations of recent African descent .Pathogens have always been a major cause of human mortality, so they impose strong selective pressure on the human genome , 19. HTShttps://www.h3africa.org/consortium/projects/16-projects/89-cafgen). CAfGEN is specifically looking at both rapid and long-term HIV/AIDS progression status while utilizing a unique protocol that applies exome sequencing of AIDS extreme clinical phenotypes. The study designs and the protocols being utilized are valuable resources for future genomic research involving common infectious diseases. In this review, we describe the impact of HTS and genomics in understanding the human host-pathogen interaction.GWASs demand recruitment of large well-phenotyped clinical cohorts within appropriate study designs and settings. However, they are very expensive, and therefore few funding agencies are able to finance such studies yet they may offer unique opportunities to unveil genetically important signals. Currently, there are a growing number of communicable disease-specific research initiatives that are specifically interested in looking at the stages of the infections and GWAS of disease progression. A classic example is the Collaborative African Genomics Network (CAfGEN), a H3Africa-funded consortium probing host genetic factors that are important to the progression of HIV and HIV-TB infection in sub-Saharan African children  in genes and genomes can point to the evolutionary basis for differences among species and among races within a species . Many otβ-globin locus, G6PD deficiency, Duffy, ovalocytosis, ABO, and human leukocyte antigen confer resistance to malaria in the human host. Elevated frequencies of hemoglobinopathies such as thalassemia and sickle cell disease, which are caused by mutations at the β-globin locus, are maintained via balancing selection   AND Genome-wide association[Title])) where a disease-query was either a communicable or noncommunicable disease. Two search restrictions were set: publication date set (from 2005/01/01) and species (humans). A genomic database for all mutations identified to be conferring resistance to infectious diseases in different populations is a vital product of more than 10 years of GWAS. More studies should be appropriately designed to identify new potential infectious disease resistance-conferring mutations in human hosts. We searched the PubMed database for studies published since January 2005 using the terms “malaria,” “tuberculosis,” “HIV/AIDS,” “genome-wide association.” Search terms included combinations of ((Genomics and whole-genome sequencing have the capacity to greatly enhance knowledge and understanding of infectious diseases and clinical microbiology . Human gIt is less likely to find a population or individual who carries mutations conferring resistance to an infectious disease at every stage of the infection. Different individuals have different mutations that offer resistance to different stages of the infections. A genomic catalogue of these mutations identified through HTS, computational tools, and GWAS now combined with the rapidly growing genome-editing technology known as CRISPR/Cas9 will enable introduction of an array of disease-stage-specific resistance-conferring mutations. This will offer interventions at all levels of the disease process unlike traditional vaccines.


The surface design of titanium implants influences not only the local biological reactions but also affects at least the clinical result in orthopaedic application. During the last decades, strong efforts have been made to improve osteointegration and prevent bacterial adhesion to these surfaces. Following the rule of “smaller, faster, cheaper”, nanotechnology has encountered clinical application. It is evident that the hierarchical implant surface micro- and nanotopography orchestrate the biological cascades of early peri-implant endosseous healing or implant loosening. This review of the literature gives a brief overview of nanostructured titanium-base biomaterials designed to improve osteointegration and prevent from bacterial infection. In contrast to other metals such as cobalt-chrome-molybdenum (CoCrMo) or surgical steel (CrNiMo), it has almost no allergenic or immunogenic potential in vivo, showed excellent corrosion resistance and promotes osseointegration when applied to and fixed with osseous tissue. In addition, the low Young’s modulus of titanium and its alloys has biomechanical advantages since it results in smaller stress shielding compared to other biometals . In contTo date, the variety of surface modifications for titanium is extensive including mechanical (gritt blasting and attrition), chemical  and physical techniques (plasma-spraying) or a combination of these  Table 2Table 2.In contrast to its excellent osteoconductive properties, the low hardness of titanium exhibits a low wear resistance and limits its application for articulating parts of artificial joints. Another major disadvantage of most Ti surface designs is that they do only promote initial cell adherence followed by osseointegration but also promote bacterial adherence. The winner in the “race for the surface” (cell versus bacteria) decides if biomineralization leads to a solid anchoring of a total joint or bacterial growth and biofilm deposit leads to the clinical incidence of a periprosthetic infection (PI). The treatment of PI is demanding for both the orthopaedic surgeon and the patient. Even in uncomplicated cases, it is associated with two or more operations and requires mid-term immobilization of the patient. Moreover, PI results in relevant bone loss and soft tissue defects leading to reduced muscular function and may result in poor implant survivorship and disability of the patient. Especially multiresistant bacteria are problematic since their escape from eradication by antibiotics. Therefore, strong efforts have been made to develop innovative implant surface structures, which promote osseointegration as well as prevent PI. Since nanopatterning with dimensions smaller than 100 nm has entered orthopaedic research, one alternative is to design surface structures at the nanoscale, which are promising to reach these aims. This review of the literature summarizes recent data of nanostructured titanium implants with emphasis on orthopaedics.To predict the response of an osseous microenvironment to nanostructured titanium biomaterial in vivo, it is essential to get a brief insight into the complexity of the osseous cellular interactions in general and in the cell–surface interface in detail. Bones have multiple and various functions. The vital bone structure, by definition a rigid organ, is part of the hematopoietic system, stores minerals, supports and protects vulnerable organs, provides structure for the human skeleton and enables mobility. Depending on the constantly changing biomechanical requirements, bone adapts its structure. This is physiologically conditioned by a continuous balance of bone formation and degradation. This bone remodelling process is influenced by systemic  as well as local  stimuli . MoreoveIn contrast, osteoclasts maintain the homeostasis between bone formation and resorption as antagonists of the osteoblasts. They are characterized by multiple nuclei and are derived from the hematopoietic cell line. The homeostasis of bone formation and degradation is complex and predominantly regulated by the receptor activator of nuclear factor κβ (RANK) and its associated ligand. Receptor activator of nuclear factor κβ ligand (RANKL) does not only activate downstream pathways of osteoclastogenesis, but also modulates the homoeostasis of physiological as well as pathological osseous processes by linking to other signal paths . During In addition to the primarily bone forming cells, the osseous milieu contains its genuine and high specialized immune system. Nevertheless, the transition between immune and bone forming cells is fluent. The formation of osteolysis is commonly associated with the activity of osteoclasts as primarily bone-resorbing cells. In contrast, some studies showed that macrophages or multinuclear giant cells, which are ontogenetically closely linked to osteoclasts, are also capable of low-grade bone resorption . The macIt is evident that the osseous response depends not only on the biochemical composition of a biomaterial but also on the biomechanical characteristics of the implants surface. Topographical factors such as size, surface texture, macro- and microstructure, and shape of the implant play a crucial role in the cellular reaction . The bio2 while any small content of Ti2O3 and TiO depends on the local micro-environment -containing repeat of FN and acts as a primary receptor). FN also has growth factor binding regions and is able to fix growth factors for the corresponding receptors of the cellular membrane, e.g., synergistic effects on FN and bone morphogenic proteins (BMPs). Although rough titanium surfaces exhibit a better cellular attachment, some data indicate that smooth surfaces promote cellular spreading more than rough topography [Besides mesenchymal cells, neutrophils and macrophages colonize the implant, leading to the foreign body covering process and releasing cytokines and growth factors including BMP-2 to attract fibroblasts and osteoblasts. This phase is also accompanied with a quantitative and qualitative change of the protein layer between the cell and titanium surface. The interface biology of implants is complex. Especially the ubiquitous dimetric glycoprotein fibronectin (FN), with its two subunits, is involved in this process since it binds to cellular transmembrane integrins  bind to the negatively charged functional groups of the protein layer. One prominent example are the positive loaded histones from cellular fragments or from cytosol that bind to the interface of nanostructured titanium [The initial layer of predominantly negatively charged proteins onto the titanium surface neutralizes its charge during phase I and II. Afterwards also positive loaded extra- or intracellular proteins and ions  are dynamic structures and link the cytoskeletal network to the ECM, allowing cells to respond to the external environment . Therefore, mechanical forces at the implant-cellular interface can be translated by the cytoskeleton and second messenger activities into gene expression and posttranscriptional modification of expressed proteins [2+ influx, which is regulated by connexion 43 (Cx43), initiates similar cellular signals via gap junctions and is involved in osteoblast differentiation of MSCs [proteins . At thisproteins . Especia of MSCs .Phase V: Local changes of the pH and accumulation of Ca and PO4 lead to biomineralization and osteointegration of the titanium implant. Here, osteoblasts and osteoclasts are mainly involved. At last biomechanical forces induce bone remodelling.The detailed effects of micro- and nanoscale implant surface structures are not known and controversy discussed in the literature . Most inRegardless of the type of biomaterial, an initial inflammation response occurs in vivo but subsides within few days postoperatively. In contrast to this, the foreign body reaction, the extent of fibrosis and the potency for cellular or bacterial adherence is dependent on the biomaterial properties .The reaction of osteogenic bone cells to micro- and nanoscaled structured surfaces must first be elucidated in order to design the adequate coating for orthopaedic implants. In the following, the effects of microtopography, nanotopography, surface roughness, porosity and surface energy on osteogenic cell function and their consecutive influence on osseointegration are further described.The bone cells surrounded by physiological osseous milieu and its connected topographical architecture consist of nanoscaled structures. Osteocytes, hydroxyapatite crystals and collagen fibrils, as predominant elements of the bone matrix, are in the range of 50–300 nm in length and 0.5–5 nm in width. Therefore, the design of nanoscaled implant surfaces is derived from its organic standard and simulates equal growing conditions for osteogenic cells .However, even titanium implants are integrated close to the mineralized tissue. Some data suggest that titanium is separated by a very thin soft-tissue layer as a result of a weak foreign body reaction that prevents a direct contact its surface to the bone . In wellNevertheless, the extent of this fibrosis is crucial for the clinical result: since encapsulation of titanium implants is not desirable for orthopaedic implants as it cannot withstand the physical stresses of daily activities, micromovements may result in chronic implant loosening or fracture. Therefore, there is enough room for further improvements of titanium surface properties to decrease inflammatory response, avoid foreign body reaction and promote cellular attachment and biomineralization. Nanostructures are one promising technology to reach these aims.Cellular reactions  in vivo relay not only on the micrometre scale but also on the nanometre scale. Here, modifications of surface structures at the nanoscale level can improve the bioactivity and performance of titanium without changing its bioinert properties. Some of these effects are related to initial protein binding: shifting topography from the micro to the nanoscale increased surface energy leading to higher protein adsorption rates. In addition, nano-roughness alone influences the adhesion of osteoblasts and their spreading and proliferation (“rapid bone regeneration”) .In vivo, microstructured surfaces provided a better implant surface–bone contact and an increased mechanical stability. The higher is the roughness, the higher are the local surface eletrostatic charge density and adhesion energy. Thus, it is possible to generate nanostructured titanium surfaces with superhydrophilic properties which can accelerate bone healing in early stages ,77,78,79Meanwhile a large amount of diverse methods has been established to modify the surface structure of titanium implants. A typical example is the inorganic apatite coating, which can be controlled by plasmaspray techniques, electrodeposition or biomimetic precipitation of calcium phosphate (CaP immersion). During the last years, much more emphasis was put on the development of functionalized biomolecules onto titanium surfaces .One example is the immobilization of ECM proteins such as fibronectin, collagen, osteopontin or laminin by plasma polymerized and crosslinked allylamine (pp-AA)  or the c2 nanopores or nanotubes were formed . Modifications in anodization conditions  control the morphology such as diameter (15 nm–300 nm) or length of generated TiO2 tanotubes [Aiming for both optimization of osteointegration and prevention of bacterial adherence, a tremendous amount of research has been carried out into nanorough titanium implants. Especially defined nanostructures  are promising candidates to improve implant properties . To desianotubes .−1·V−1 vs. ~20 nmh−1·V−1 in LV regime) [The growth rate and mechanical strength can be improved by high-voltage (HV) hard anodization (growth rate in HV: 1.2 μmh regime) ,86.If the applied voltage exceeds the dielectric breakdown limit of the titanium oxide, the oxide will no longer be resistive to prevent further current flow and oxide growth. This leads to more gas evolution and sparking known as “Anodic Spark Deposition” (ASD), and “Micro-Arc Oxidation” (MAO) .2SO4, but fluoride solutions seem to be more useful by producing biologically-inspired nano-tubular structures. Another major advantage of anodization is that nanotubes from almost all Ti alloys can be generated . Interestingly nanotubes formed on Ti-6Al-4V by anodic oxidation have mainly been referred to as TiO2 nanotubes, while pure titanium oxide nanotubes are formed only on pure Ti [2 nanotubes are created, the nanotubes formed on Ti-6Al-4V by thermal oxidation were referred to as Ti-Al-V-O nanotubes.Among all electrolytes, the anodic oxide thickness is highest in H pure Ti ,89,90. S pure Ti . In cont2 nanotube structure accelerates growth in eukaryotic cells and reduces the ability of staphylococcal aggregate/biofilm formation. Nanotubes have a relevant size-effect on bone and other cells: Whereas diameters of 15–20 nm are optimal for increased cell adhesion, proliferation as well as for alkaline phosphatase (ALP) activity and bone matrix deposition [Besides morphological parameters, the degree of crystallinity changes the hydrophilic character of the nanotube. This has an impact on cell proliferation and osteointegration since nanostructured/hydrophilic surfaces show advantages ,93,94,95position ,99,100, position ,103,104.Typical structures are bone apatite which forms 10–20 nm long and 2–3 nm wide plates, whereas type I collagen fibres are ~200 nm long and 2–3 nm thick ,106.2 or ZrO2), for different crystallization states  and is also unchanged in different fluoride contents in the tubes. It was confirmed on several cell types such as mesenchymal stem cells, haematopoietic stem cells, endothelial cells, osteoblasts and osteoclasts [3) [This size-effect is valid for different substrate materials  ,108.The cause of the size effect is probably based on integrin clustering in the cell membrane leading to focal adhesion complexes. These have a diameter of about 10 nm which perfectly fit to nanotubular diameters of about 15 nm. In vivo, the cytotoxic effect of 100 nm is mitigated because larger nanotube dimensions show higher interaction energies, thus leading to higher protein adsorption which obviously influences cell behaviour . Even inIn addition, the cation-mediated attraction between extracellular matrix proteins  and the titanium surface is expected to be more efficient for a high surface charge density. This facilitated integrin-mediated osteoblast adhesion along the sharp convex edges or spikes of nanorough titanium surfaces where the magnitude of the negative surface charge density is the highest. In summary, Ti nanotubes with sharp edges and a diameter of approximately 15 nm should be the preferred surface for orthopaedic nanostructured implant. However, there are only few studies on whether in vitro effects of nanorough titanium can be transplanted to the in vivo situation.2 nanotubes  by pull-out experiments in rabbits [2 (TiO2-NT) [Some other investigators described improved bone bonding strength and osteointegration in nanotubular surfaces compared with grit blasted surfaces ,110. The rabbits ,112. MorTiO2-NT)  or Ti-24TiO2-NT) .In summary, most in vivo data suggest that especially the initial bone formation is enhanced in nanorough titanium surfaces . This stOne further option for changing surface properties of titanium is the implantation of bioactive ions such as Ca, Zn, H and Sr. Hegedus et al. found improved cell viability for Ti with implanted Ca or dual Ca + Si ion compared to pristine Ti substrate . This co2 nanotubes  [immobilization of epidermal growth factor (EGF) on 100 nm diameter TiOeration) ;○higher osteocalcin and osteopontin levels at 30 nm diameter ; and○enhanced osteogenic differentiation 15 nm diameter, chondrogenic differentiation on 100 nm ; andimmobilization of bone BMP-2 onto TiO nanotubes:vascular endothelial growth factor (VEGF) .More complex for getting official approval as a medical device is the covalent immobilization of biomolecules e.g., BMP-2 onto nanostructured Ti surfaces. Experimental data have shown that this can be a useful tool to control differentiation of mesenchymal stem cells (MSCs). Typical examples are:This functionalization could be performed directly, but usually is realized via self-assembled monolayers (SAMs), antibodies or peptides on the surface of nanostructures . The conEspecially the pharmacokinetics of Ti-bound BMPs is hard to control in vivo. This is relevant, since extensive concentration of local released BMP can induce osteolysis ,127,128.2+  pathways directly by modified surface structures as shown by Chakravorty et al. [However, it has been taken into account that the secondary structure of rhBMP-2 can change significantly dependent on the modified surface . These conformational changes will also influence the BMP-2 activity . Anothery et al. . Especiay et al. .The latest development of nanostructured titanium aims at osteopromotion and prevention of bacterial adhesion. One example is to design hierarchical architectures by a “layer-by-layer” (LbL) concept. Here, chitosan coated BSA nanoparticles (CBSA NPs) and oxidized alginate (OSA) showed features of nanostructures on titanium. The functional groups provide active sites for BMP-2-immobilization and promote bone formation. Furthermore, it was shown that the antibiotic vancomycin binds on the OSA. Therefore, these assembling nanoparticles might prevent infection during the bone healing process .However, the application of local antibiotics to nanostructured titanium for clinical application seems to be critical regarding pharmacological side effects including bacterial resistance, limitations in implant sterilization and official approval.2).More promising is the combination of a titanium surface structure with other agents such as HA or iodine as described by Tsuchiya et al. ,139. TheThe following section summarizes further anti-microbiological effects of nanotitanium structured surfaces and clinical strategies to treat infections.The demographic change in the Western world and prolonged life expectance have increased the rate of total joint revision operations dramatically . ImplantThe consequences of PI are an imprinting and traumatic experience for the patient. Radical debridement of soft tissue and bone during revision surgery leads to progressive bone loss and therefore loss of biomechanical stability. The revision operation increases the complication rate compared to primary implantation significantly . FurtherThe pathogenesis of PI or septic loosening is due to intraoperative contamination of the implant or the periprosthetic interface, which may become apparent in an early-onset acute local putrid infection, a Systemic Inflammatory Response Syndrome (SIRS) or in a “low-grade infection”. In addition to the direct contamination, a secondary infection of the prosthetic joint by bacteraemia e.g., after soft tissue infection, pneumonia or tooth extraction is also possible .The initial contamination of the biomaterials surface is followed by the development of a biofilm on the colonized surfaces. Biofilms are “aggregates of microorganisms in which cells are frequently embedded in a self-produced matrix of extracellular polymeric substances that are adherent to each other and/or a surface”. Typically, biofilms comprise many species and are complex systems with high cell densities . This biInnovative solutions for biofilm absorbing or antibacterial Ti surfaces were achieved by permanent binding of antibiotics , antimic3N4 surfaces are also capable of bacterial lysis. Thus, the topographical bactericidal effect is primarily dominated by its surface nanotopography and not only by its surface biochemistry [Escherichia coli on the surface by 40% in comparison with polished Ti in vitro [Mechanisms for these inherent antimicrobial properties are part of current research. It is hypothesised that defined nanotopographies are able to disrupt bacterial cell membranes and lead to a decreased bacterial viability. An effective microbial lysis is achieved by creating topographies which generate an adequate local stress across the microbial cell membrane. Dickson et al. defined the ideal space in between nanopillars in the range of 130–380 nm . In addihemistry . Sjöströin vitro .Staphylococcus aureus more effectively compared to less nanoscaled Ti surfaces [Mathew et al. showed that greater nanoscale surface roughness led to significantly decreased bacterial colonisation. In detail, implants coated with electrophoretic-deposited hydroxyapatite inhibited the growth of surfaces .Psaltoda claripennis) are covered by hexagonal arrays of nanopillars which are able to eliminate Gram-negative bacteria by its physical surface structure consistently [Escherichia coli, could be shown, whereas Gram-positive bacteria, e.g., Staphylococcus aureus, were not affected [Recently, a new potential antibacterial phenomenon based on solely nanotopographical properties has been found in locus-like insects. The wings of Clanger cicada  nanoparticles with fibrous scaffolds such as polycaprolactone (PCL) fabricated by electrospinning lead to protrusions mimicking the architecture of a rose stem. The branched nanoparticles  induced heterogeneous crystallization of the polymeric matrix. This three-dimensional composite enhances the mechanical strain and strength and offers excellent antibacterial activity, while supporting the growth of eukaryote cells . Bounded+ with bacterial membrane constituents. This causes structural changes, interrupts transmembrane electron transfer, oxidizes bacterial components and may at least induce bacterial death. In addition, Ag+ shows also cytotoxic effects such as mitochondrial dysfunction, disturbed membrane integrity, induction of reactive oxygen species, and interrupted adenosine triphosphate synthesis resulting in DNA lesions. The cytotoxic effects of Ag+ depend on the instability of Ag-based bactericides, such as the high mobility of Ag nanoparticles (NPs) or Ag-containing calcium titanate. This can be avoided by silver plasma immersion ion implantation (Ag-PIII) with atomic-scale heating (ASH). The three-dimensional, hierarchical structure of sand-blasted, large grit, and acid-etched (SLA) Ti surfaces showed optimal preconditions for silver plasma immersion ion implantation. These techniques showed not only a sufficient defence against multiple cycles of bacterial attacks, independently from silver release, but they are also nontoxic to eukaryotic cells such as bone marrow mesenchymal stem cells [The bactericidal effect of silver is based on interactions of Agem cells ,174,175.3+ acts also as a “Trojan horse” competiting with Fe3+ for binding to siderophores, thus interrupting crucial Fe-dependent metabolic pathways. One method embedding Ga into Ti surfaces is based on chemical and heating treatments resulting in a Ga-containing calcium titanate (Ga–CT) or gallium titanate (GT) surface. It was demonstrated that Ga–CT and GT interfaces exhibited very high antibacterial activity against multidrug resistant Acinetobacter baumannii (MRAB12) and inhibits bone resorption [Another strategy to combine antimicrobial and osteoprotective properties is the incorporation of gallium (Ga) ions in Ti surfaces. In contrast to Ag, Ga showed higher cytocompatibility since it can substitute Fe in many biological systems  and inhibits bone resorption. In bacteria Gasorption ,178,179.sorption  or poly sorption .Inspired by bacteriophages that use nanostructured proteins to invade bacteria, other investigators study the antibiotic role of nanostructures. Mimicking the two basic structural motifs of bacteriophages, spherical and rod-like polymer molecular brushes (PMBs) have been designed. However, the impact of these techniques for Ti implants is unclear in future ,183. HowHowever, for practical performance in orthopaedic surgery, the narrow valleys or slots of nanomaterials might be filled up with adsorbed proteins and other molecules within few minutes. Therefore, it also seems to be questionable if the antimicrobial effects of the nanostructures  will have a relevant and persisting effect on prokaryotes. Besides, appropriate mechanical strength of nanostructures is a requirement for clinical application. For some materials infinite element analysis is available to predict the mechanical behaviour . Especia2 nanoparticle [From our point of view, the future in nano-based orthopaedic Ti implants will stronger focus on NPs composites such as PLA electrospun fiber-TiOparticle  and alsoparticle . Besides+, Cd2+, Hg2+, Ni2+, etc. . These species and strain dependent strategies include:energy dependent efflux of toxic metals e.g., enzymatic transformations ;the expression of metal-binding proteins ;inhibition of ions to enter the cell (downregulated expression of membrane transport proteins); andpersisting phenotypes with slower growth or ceasing cell division .Many bacteria express metal-sensing transcriptional regulators allowing them to adapt to their environment rather quickly. Within minutes to hours, bacteria are able to escape toxic metals such as AgE. coli and Ag+ [In contrast to these initial physiological responses to toxins, which is described for  and Ag+ , there iIn contrast to today’s implants, future Ti surfaces will have more controlled surface design with less variation on the macro-, micro- and nanoscale. For many years, the pharmaceutical industry designed drugs on a molecular level. Future medical devices such as Ti implants might follow these developments to the benefit of orthopaedic patients. Rapid Prototyping (RP) Technologies , electron beam melting (EBM), selective laser melting (SLM) or 3D printing can fabricate porous Ti scaffolds with fully interconnected porous network and highly controllable porosity and pore size ,198,199.Understanding the molecular principles and interactions at the tissue-surface interface at different dimensional levels (porosity gradient) might help closing the gap between promising new designed Ti structures of the research stage  and the so-called “valley of death” of further translational and preclinical stages of implants. One example is to control cell performance to a certain extent by varying pore sizes ,200. Sma


The incidence of human papillomavirus- (HPV-) related oropharyngeal squamous cell carcinoma (OPSCC) has been rapidly increasing worldwide. HPV is reported in approximately 50% cases of OPSCC in Japan. However, there are few reports of synchronous bilateral HPV-positive tonsillar carcinoma, and, in almost all those cases, carcinoma was detected using positron emission tomography/computed tomography and/or bilateral tonsillectomy. We report the case of a 63-year-old male with bilateral tonsillar carcinoma detected using transoral endoscopic examination with narrow-band imaging (NBI). A biopsy of the bilateral tonsils revealed squamous cell carcinoma, which was demonstrated to be HPV-related using in situ hybridization and p16 immunohistochemistry. The patient was diagnosed as synchronous bilateral tonsillar carcinoma: T1 (2) N2b M0. He was treated with induction chemotherapy, bilateral radical tonsillectomy with neck dissection, and radiotherapy. To our knowledge, this is the first report of a synchronous bilateral tonsillar carcinoma detected using transoral NBI in the outpatient setting. Early diagnosis without the inspection under general anesthesia is beneficial for the patients with lymph node metastasis from unknown primary lesion. Cervical metastases from an unknown primary site account for approximately 5%–10% of the head and neck cancers . It is wThere is approximately a 4% incidence of synchronous secondary tumors in the head and neck area . A seconHead and neck squamous cell carcinoma (HNSCC) is strongly associated with lifestyle, including smoking and alcohol consumption, and the overall incidence has been declining in the United States in the past 20 years . This haRecently, we recommended using narrow-band imaging (NBI) for the detection of primary lesions . Transor2, day 1) and 5-Fluorouracil , the primary tumor showed a complete response (CR), and the neck lymph node showed a partial response (PR). The patient underwent bilateral radical tonsillectomy to confirm CR and underwent neck dissection. Histopathological examination revealed no primary tumor remnant in the bilateral tonsils and positive lymph node metastasis with extracapsular spread. Although we recommended the postoperative chemoradiotherapy covering the bilateral tonsillar beds and neck area, the patient refused the invasive treatment because of his job . He was so concerned with the adverse events like xerostomia and taste change that we indicated the postoperative radiation therapy alone to the left neck (54 Gy). Approximately 15 months after the radiation therapy, he developed a regional relapse in the right lymph node. Thus, he underwent right neck dissection and has been alive without disease for 3 years after the salvage surgery.A 63-year-old man presented with a 2-month history of a left neck mass. No obvious primary lesion was detected on routine otolaryngologic examination; however, fine-needle aspiration cytology revealed squamous cell carcinoma (SCC). He was referred to our hospital with an unknown primary neck metastasis. We sought the primary lesion throughout the pharynx; a small granular lesion on the right tonsil and a tiny mass on the left tonsillar pillar were recognized using transoral endoscopic examination with NBI . Both leJoseph et al. reported that bilateral HPV-related tonsillar carcinoma is caused by the same HPV-16 variant . In our Previous cervical cancer research has suggested three potential molecular mechanisms for the development of multifocal HPV-associated head and neck cancers .HPV-related tonsillar cancers arise from the basal layer of the tonsillar crypt, and the tumor is often covered by normal mucosa, not exposed to the luminal surface at the early stage. Hence, the tumor growing inside the palatine tonsils may not be easy to visualize if the mucosal defect is too small. Therefore, transoral examination with NBI is helpful for detecting the primary lesion in the tonsils, especially in HPV-related cases. This examination can be performed easily in the outpatient setting without any complicated preparations .In the case of lymph node metastasis from unknown primary lesion, bilateral tonsillectomy is recommended to detect the primary cancer in the tonsils. However, tonsillectomy needs general anesthesia, which may delay the diagnosis and treatment. In the present case, transoral NBI helped us diagnose the bilateral lesions at the outpatient clinic and initiate the treatment without delay.Synchronous bilateral tonsillar carcinomas are very rare, with only 20 cases reported in the English literature prior to this case; furthermore, only eight cases of HPV-related synchronous bilateral tonsillar carcinoma have been reported since 2010 . To the Transoral NBI examination is a useful method to detect the primary sites of unknown neck metastases. This is especially important in the era of increased incidence of HPV-related tonsillar cancers.


Posttraumatic stress disorder (PTSD), and combat-related PTSD in particular, has been associated with increased rates of cardiovascular disease, and cardiovascular-related death. However, less research has examined possible factors that may link PTSD to poorer cardiovascular health in combat veteran populations. The current pilot study investigated whether psychological symptomology and autonomic reactivity to emotional scripts would relate to poorer cardiovascular health in combat veterans without a current diagnosis of cardiovascular disease. Male veterans (N = 24), who served in combat since Operation Iraqi Freedom, completed a semi-structured interview and self-report measures to assess psychological symptomology. Autonomic reactivity, measured using heart rate variability , was obtained during script-driven imagery of emotional memories. Cardiovascular health was assessed using flow-mediated dilation (FMD) of the brachial artery. Correlational analyses and discriminant analysis were used to assess the relationship between psychological symptoms , autonomic reactivity to emotional scripts (HRV), and FMD. Overall, veterans in the current study showed poor cardiovascular health despite their relatively young age and lack of behavioral risk factors, with 15/24 exhibiting impaired FMD (FMD < 5%). Psychological symptomology was not associated with FMD; whereas autonomic reactivity to emotional  scripts was found to relate to FMD. Autonomic reactivity to negative scripts correctly classified 76.5% of veterans as having impaired versus normative FMD. Results from this pilot study highlight the importance of cardiovascular screening with combat veterans despite psychological diagnosis. Results also support the need for longitudinal research assessing the use of autonomic reactivity to emotionally valenced stimuli as a potential risk factor for poorer cardiovascular health. It is estimated that approximately 90 percent of adults within the United States will experience a traumatic event . Of thosPrevious research indicates that combat exposure , 11 and Most studies investigating the relationship between combat-related PTSD, and CV health have been epidemiological, investigating rates of diagnosed CV disease for those with and without combat exposure and PTSD. The studies that include assessment of CV health have most often relied on resting blood pressure, heart rate (HR), or lipid levels. Flow mediated dilation (FMD) provides unique and potentially predictive information concerning CV health. Specifically, FMD uses Doppler ultrasound to measure the vasodilatory response of an artery after an increase in luminal blood flow (flow within the artery), and is an assessment of endothelial function . FMD is Prior research suggests several possible links between PTSD and poorer CV health including dysreguation of the hypothalamic-pituitary-adrenal axis, which impacts corticosteroid release  and autonomic nervous system regulation . DysregThe current pilot study aimed to explore how psychological symptoms , autonomic reactivity (HRV) to emotional events, and combat-exposure may relate to FMD in a veteran sample. A focus on relatively young, and physically healthy combat veterans was to allow for examination of FMD prior to the development of CV disease or other chronic health problems. It was hypothesized that greater PTSD symptoms would relate to more impaired FMD. Further, we hypothesized that increased autonomic reactivity during emotional provocation  would significantly predict impaired FMD (FMD < 5% ), above Participants included male combat veterans  who served since the onset of Operation Iraqi Freedom (OIF), with varying levels of PTSD symptomatology . Exclusion criteria included active suicidal plan or intent, substance abuse within the past six months, schizophrenia or bipolar I disorder , historParticipants were recruited via advertisements in the general community  and on local college campuses , and by providing informational flyers to clinicians at local VA hospitals. This study was approved by the University of Kansas Medical Center and the University of Missouri—Kansas City Institutional Review Boards. All participants provided written, signed consent.The current study procedures described below were conducted across two study visits. During the first visit, participants completed a clinical assessment to assess symptoms of PTSD (described below). Participants were asked to return for a second study visit within six months (ranging from one week to six months) to complete the remainder of the clinical assessment, assessment of CV health, and the script-driven imagery task (described below). During the second study visit, participants completed the assessments in the following order: assessment of CV health, script-driven imagery task, and clinical assessment . Assessment of CV health was always conducted prior to script-driven imagery to minimize subject burden due to fasting, as well as to minimize potential carry-over effects of emotional responses to script-driven imagery that may have influenced the FMD response.Data collection began prior to the dissemination of Diagnostic and Statistical Manual– 5 . TherefoDepressive symptoms were assessed with the Beck Depression Inventory II (BDI-II ). The CoParticipants completed an anxiety-provoking task using script-driven imagery. This assessment is a slightly modified version of previously described script-driven imagery methods , 50. ParFlow-mediated dilation (FMD) was used to assess endothelial function, as an index of CV health. Participants were asked to refrain from food or caffeine for 12 hours and any medications prior to the procedure. Participants were asked to rest supine for 20 minutes prior to the start of the procedure in a temperature-controlled room (21–24 degrees C) that was quiet with dim lighting. We used a 9.0 Mhz linear array probe attached to a high-resolution ultrasound machine  to create an image of the brachial artery and blood flow. Our methods for the FMD procedure have been previously published –67.Briefly, once the brachial artery was identified, a stereotactic clamp was used to stabilize the ultrasound transducer and hold it in place during the procedure. A resting baseline for diameter and blood flow velocity was recorded for 60-seconds. An automated blood pressure cuff  was placed 2–3 cm below the anecubital fossa and was inflated above suprasystolic pressure to 220 mmHg for five minutes. Twenty seconds prior to cuff deflation, recording resumed for 3 minutes. All images were analyzed off-line using specialized software .Total severity scores calculated for the CAPS and the combat experiences sub-scale of the DRRI were the primary self-report variables of interest. We examined relationships with symptom severity rather than comparing dichotomous groups  due to the fact that all participants had experienced trauma (combat exposure) and PTSD symptoms were normally distributed across the sample. Total severity scores were also calculated for the BDI-II and AUDIT. Autonomic reactivity was calculated by subtracting the LF/HF ratio during the neutral script from the LF/HF ratio for positive (POS-NEU), negative (NEG-NEU) or combat (COMB-NEU) scripts. LF/HF was also measured during baseline (2-minutes prior to script onset), and recovery (5-minute post combat script) periods. All LF/HF variables were log-transformed prior to analyses due to non-normal distribution. Relationships between these variables, as well as FMD, were then explored using two-tailed Pearson’s correlations. Exploratory correlations between FMD and known CV risk factors including age, current smoking status, and BMI were also conducted to determine possible covariates for the primary analyses.p < 0.05 was considered significant. Effect sizes are reported to aid in the interpretation of findings and inform the design of future studies.The primary outcome measure for the dependent variable, FMD, was separated into a dichotomous measure of dilation less than or greater than five percent, with less than five percent indicating an impaired FMD response . Variablp > 0.10 for each variable). Excluding participants who were smokers did not alter study results.Descriptive analyses of the current sample are presented in p = 0.037), but not to LF/HF ratio to emotional scripts . The relationship between PTSD and LF/HF at baseline was trending with a moderate effect size . Combat experiences were not significantly related to LF/HF at baseline or LF/HF reactivity to emotional scripts . Self-reported PTSD symptoms , depressive symptoms , combat exposure  and LF/HF during recovery periods  were not related to FMD. Greater LF/HF reactivity to positive scripts related to higher FMD  and there were trends for LF/HF reactivity to negative  and combat  scripts (p = 0.058).PTSD symptoms related to combat experiences , correctly classifying 76.5% of all cases.LF/HF at baseline, as well as LF/HF reactivity to positive, negative and combat scripts were entered into a stepwise discriminate analysis. LF/HF ratio to negative events emerged as the only significant predictor of impaired FMD . This reduced FMD response was observed in our study population, despite their young age (M = 32.38), lack of significant alcohol and tobacco use, and moderate levels of PTSD symptoms. To our knowledge, two other studies have assessed the relationship between PTSD and FMD [A large percentage (62.5%) of the current sample exhibited impaired FMD, with vessel dilation less than five percent (mean FMD = 4.79). In contrast, the average FMD reported for healthy adults is closer to seven and a half percent , which i and FMD , 68. Vio and FMD . This re and FMD , similar and FMD . It is tFMD provides the unique opportunity to assess endothelial-dependent function, adding to information provided by other measures of CV health . The present results demonstrate the importance for physicians and healthcare providers to regularly assess for CV health with combat veterans (or other populations with high levels of trauma exposure), regardless of age or other demographic variables via measures of autonomic reactivity and/or FMD. Our findings highlight the importance of future research to identify factors that predict poor CV health in combat veterans and to identify targets for early intervention.Autonomic reactivity  to any emotionally valenced script  was found to positively relate to FMD (with a moderate to large effect size). This finding supports future investigations assessing the use of autonomic reactivity to emotional scripts as a predictor of CV health in populations with trauma history. Autonomic reactivity to negatively valenced stimuli emerged as the only significant predictor of impaired FMD. Interestingly, average LF/HF ratio to negative events was relatively lower than for neutral events suggesting decreased autonomic balance during neutral compared to negative events. This could reflect participant attempts to regulate autonomic arousal specifically during negatively valenced emotional provocation. It could be useful for future research to investigate whether one’s ability to purposefully down-regulate autonomic arousal may be an important contributor to chronic CV health, and a potential target for reducing impaired FMD.The current pilot results also suggest that measures of autonomic reactivity may be more powerful predictors of CV health in combat veterans than self-report measures of psychological symptoms or combat experiences. This may be due to sympathetic and parasympathetic reactivity being a more direct indicator of adrenal reactivity and stress hormone release  than PTSThe current findings suggest that autonomic reactivity to emotional provocation could potentially serve as a cost-efficient and objective screening measure for CV health. Moreover, this begs the question of whether interventions aimed at modifying autonomic reactivity could be beneficial in reducing impaired FMD. It is estimated that greater than 50% of CV disease reduction is attributed to changing CV risk factors . TherefoThe current pilot study was limited by a small sample size (N = 24), lack of a non-trauma-exposed control group, and a cross-sectional design. The current sample included male veterans who have served in combat since the onset of OIF, limiting generalizability of findings to other types of trauma. Additionally, the symptom severity reported by veterans in the current study was in the mild to moderate range, which may have limited our ability to identify relationships between symptom severity, FMD, and autonomic reactivity. Future research should assess the potential impact of PTSD on autonomic reactivity and FMD in treatment-seeking veterans with more severe PTSD symptomology. Lastly, FMD is specific to endothelial-dependent dilation. It would be important for future studies to incorporate several modes of vasodilation  to further understand relationships to trauma, PTSD, and autonomic reactivity.Our results suggest that young combat veterans exhibit impaired FMD, and that autonomic reactivity to emotionally valenced events is a potential predictor of CV health. Previous research has established relationships between hyperarousal symptoms associated with PTSD and parasympathetic function –52, 75,


Nevertheless, no small animal model developed so far is capable of maintaining the complete life cycle of Plasmodium parasites that infect humans. The ultimate goal is to develop humanized mouse systems in which a Plasmodium infection closely reproduces all stages of a parasite infection in humans, including pre-erythrocytic infection, blood stage infection and its associated pathology, transmission as well as the human immune response to infection. Here, we discuss current humanized mouse models and the future directions that should be taken to develop next-generation models for human malaria parasite research.Malaria parasite infection continues to inflict extensive morbidity and mortality in resource-poor countries. The insufficiently understood parasite biology, continuously evolving drug resistance and the lack of an effective vaccine necessitate intensive research on human malaria parasites that can inform the development of new intervention tools. Humanized mouse models have been greatly improved over the last decade and enable the direct study of human malaria parasites  Plasmodium parasites, causes more than 200 million clinical cases annually and is responsible for more than 400,000 deaths each year, mainly in children under the age of 5 and pregnant women living in the resource-poor countries of sub-Saharan Africa. In humans, the majority of malaria infections are caused by Plasmodium falciparum and Plasmo-dium vivax. In more temperate regions of the world, socioeconomic development, vector control programs, and the use of antimalarial chemotherapeutics have driven successful malaria elimination. However, declines in malaria infections have been slowest in tropical, resource-poor countries with a high malaria burden necessitating the development of new effective antimalaria therapeutics and vaccines that will prevent infection, disease, and onward transmission.Malaria, a disease caused by protozoan P. vivax infection, a subset of sporozoites form persistent liver stages (hypnozoites) that activate at later time points to cause relapsing blood stage (BS) infections  and RBCs, researchers have relied on in vitro infection models for human parasites to query malaria biology and identify targets of intervention. In this review, we will discuss how blood, tissue, and immune system-humanized mouse models can provide novel avenues to examine human malaria parasites. Humanized mice and their past use in malaria research have been reviewed recently  made it attractive to speculate that parasite behavior in the skin should be similar in Plasmodium species that infected humans. However, the recent identification of the development of exo-erythrocytic merozoites in the skin of P. berghei-infected mice  under the control of an albumin promoter in the livers of the highly immunodeficient Severe Combined Immune Deficiency mice (SCID Alb-uPA). Upon engraftment with huHeps, these mice become susceptible to infection with P. falciparum sporozoites and support complete liver stage development including the release of exo-erythrocytic merozoites that egress and invade human RBCs (huRBCs) ex vivo (fumarylacetoacetate hydrolase (FAH) in mice resulting in acute liver failure which can be rescued by administration of the drug, 2-(2-nitro-4-fluoromethlbenzoyl)-1,-3-cyclohexanedione (NTBC). Crossing these FAH−/− mice onto the severely immunocompromised C57BL/6 Rag2−/−IL2rγ−/− mouse generated FAH−/−Rag2−/−IL2rg−/− (FRG) mice. These mice have also been backcrossed onto the non-obese diabetic (NOD) background (FRGN), which additionally renders them hospitable to transplantation with CD34+ hematopoietic stem cells (HSCs)  mouse has been developed as yet another mo-del for P. falciparum PE infection . Traditional rodent malaria models have been critical to the iden-tification of host hepatocyte surface factors necessary for sporozoite invasion , 20. Rec ex vivo , 23. An s (HSCs) . Using Ns (HSCs) , are susrozoites , 27 and nfection . These melopment .P. yoelii and P. berghei have been extensively utilized because they allow a careful examination of PE immunity. Perhaps the greatest contribution of the rodent malaria models is in the examination of immunity and protection against an infectious sporozoite challenge after vaccination with whole, attenuated sporozoites. Attenuation was originally generated by gamma irradiation  where Plasmodium infection better mirrors observations made in humans. Numerous NHP species support infection by Plasmodium species that occur in simians and some NHP support direct infection with P. falciparum and P. vivax  and then subsequently challenged with a transgenic P. berghei parasite encoding the repeat regions of P. falciparum CSP. Immunized HIS-CD4/B mice exhibited high titers of circulating P. falciparum CSP antibodies and showed reduced parasite liver burden after challenge as compared with naïve controls  mice have been developed that enable the direct analysis of the HIS responses esponses . Huang aesponses , 58 for Plasmodium infection, Huang and colleagues also generated a humanized CD8 T cell mice referred to as HIS-CD8  susceptible to human Plasmodium liver infection and capable of driving functional human immune responses. However, such a model will not yet completely replicate human immunity because although liver-chimeric mice can be repopulated with high levels of huHeps, their liver sinusoidal endothelial cells, Kupffer cells, hepatic stellate cells, and cells of the myeloid lineage all remain of mouse origin. In rodent malaria systems, CD8+ dendritic cells have been shown to be critical to the generation of an effective immune response after whole sporozoite immunization  and enriched reticulocytes from UCB have been successfully used for in P. vivax in vitro BS invasion assays  family, variant antigens expressed on the surface of infected RBCs that interact with multiple host receptors, including ICAM1, CD36, E-selectin, and endothelial protein C receptor (EPCR)  . There aP. falciparum infection is cerebral malaria. It has been speculated that the targeting of different receptors via the expression of different PfEMP1 variants leads to tissue-specific sequestration of the parasites. In the brain microvasculature, the EPCR has been shown to play an important role in the sequestration of parasites  mice, where human fetal thymus and liver tissues as well as autologous HSCs are engrafted into the same mouse represent the most complete humanized mouse system to date (Plasmodium research will utilize the transplantation of CD34+ HSC cells to develop robust human immune and erythropoietic compartments and hepatocyte transplantation to ensure human liver chimerism. Ideally, this mouse will in addition harbor human receptors on endothelial cells so that the pathobiology of malaria BS infection can also be modeled. As it will take great effort to create such a complete and complex mouse model, a continued iterative cycle of basic parasitological discovery and immunology in rodent malaria models combined with studies in currently available, albeit imperfect huHep, HIS, and huRBC mice will further increase the predictive value of animal models for human clinical intervention.Conventional mouse model infections with rodent malaria parasites have critically contributed to our understanding of malaria parasite biology, pathogenesis, and immunology and have been important in malaria vaccine and drug discovery. However, differences between the genomes of the human-infective and rodent-infective  to date . CombiniNM and CS prepared the original drafts of this manuscript with subsequent revision by SK.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Rhabdomyosarcoma (RMS) is a malignant tumor of mesenchymal origin and comprises the largest category of soft-tissue sarcomas both in children and adolescents. From a pediatric oncology point of view, RMS has traditionally been classified into alveolar (ARMS) and embryonal (ERMS) subtypes. The anatomical localization of the tumor may vary, but commonly involve the head/neck regions, male and female urogenital tract or the trunk and extremities.Here, we report two challenging cases involving 17- and 9-years-olds males where diffuse and multiplex bone lesions suggested either a hematological disease or a primary bone tumor . Biopsies, proved a massive infiltration of the bone marrow cavity with rhabdomyosarcoma. In both cases, the ARMS subtype was confirmed using FOXO1 break-apart probes (FISH). Radiological examination could not identify primary soft tissue component in any localization at the time of diagnosis in either cases.Primary alveolar rhabdomyosarcoma of the bone as a subtype of ARMS, seems to be a distinct clinico-pathological entity with challenging diagnostic difficulties and different, yet better, biological behavior in comparison to soft tissue ARMS. However, it is difficult to be characterized or predict its prognosis and long-term survival as only sporadic cases (four) were reported so far. Rhabdomyosarcoma (RMS) is among the most common soft tissue sarcomas in childhood and adolescence with 4.5 new cases/1 million person/year in the USA and incidences in Europe share similar numbers . It is aAvailable data about primary bone ARMS is limited due to the fact that so far only four cases were found in literature reporting fusion-positive alveolar RMS confined to the bone marrow –10. ThusA 17-year-old male with Crohn’s disease in his medical history, presented with fever, weight loss and lower back pain; experienced over a period of 1–2 weeks. He was found to have elevated inflammatory markers and serious hypercalcemia with impaired renal function. Bone scintigraphy, lumbar spine and pelvic MRI revealed disseminated, diffuse infiltration of the bone marrow which primarily raised the suspicion of lymphoma Fig. . RepeateHistological examination of the second bone marrow biopsy (iliac crests) revealed solid sheets of tumor cells infiltrating the entire bone marrow replacing and expelling the normal hematopoietic cells. The highly cellular infiltrate showed no special arrangement, although fine fibro-vascular stroma could focally be identified. The monomorphic, poorly differentiated tumor cells had round, vesicular nuclei with fine chromatin content and were localized at the periphery of the cells, in an eccentric position Fig. . The cytA 9-year-old male was admitted to the hospital presented with recurrent fever, lower back and right lower limb pain, experienced over the period of a month. He was found to have mild anemia and elevated inflammatory markers. Imaging studies revealed disseminated multiplex bone lesions involving the entire vertebral column, pelvic bones, ribs, skull, the distal part of the right femur and the proximal part of the right tibia Fig. . These fThe biopsy sample that was taken from the tibia showed different morphological patterns and areas that made the overall histological picture misleading: among the bony trabeculae, a cellular tumor infiltrate could be identified and the cells were arranged in solid sheets replacing the normal hematopoiesis. Other foci of the tumor showed intramedullary cartilage islands around which spindle or ovoid shape tumor cells formed a cohesive structure; allowing mesenchymal chondrosarcoma diagnosis. Besides this, however, some areas of the tumor formed solid sheets of tumor islands that were divided by fine fibro-vascular stroma Fig. –d. The cAlveolar rhabdomyosarcoma is a high grade neoplasm that has the worst prognosis amongst other subtypes of RMSs , especially in fusion-positive cases when FOXO-1 gene is involved. It is generally known that the overall outcomes for patients with soft tissue ARMS is worse than in patients with ERMS - even with aggressive multimodal therapy . The proAlthough the previously reported four cases of primary ARMS (as well as our current two cases) show a better survival rate compared to its soft tissue counterpart, it still causes difficulties in precisely characterizing this tumor type. One reason is the low number of reported cases, while a major problem alongside this is that even data contained within medical literature is confusing with regards to ARMS classification. It distinguishes fusion-positive and fusion-negative cases; however, there is a tendency that fusion-negative cases should be considered in practical terms ERMS . Until tIn the case of (soft tissue) alveolar RMS it is known that it commonly infiltrates the bone marrow , causingThe natural history of primary alveolar RMS of bone may show individual variations, but our current cases, together with the other four reported ones –10, suggPrimary alveolar rhabdomyosarcoma of the bone as a subtype of ARMS, seems to be a distinct clinico-pathological entity with challenging diagnostic difficulties and different biological behavior when compared to soft tissue ARMS. More available data might be necessary to predict not only the course of the disease, but also to develop and set up further chemotherapeutical combinations that may increase the overall survival of the patients in the future.


The Fontan procedures, designed to treat paediatric patients with functional single ventricles, have markedly improved the patient’s survival into adulthood. The physiology of the Fontan circuit inevitably increases systemic venous pressure, which may lead to multi-system organ failure in the long-term follow-up. Fontan-associated liver disease (FALD) can progress to liver cirrhosis with signs of portal hypertension. Focal nodular hyperplasia-like nodules commonly develop in FALD. Imaging surveillance is often performed to monitor the progression of FALD and to detect hepatocellular carcinoma, which infrequently develops in FALD. Other abdominal abnormalities in post-Fontan patients include protein losing enteropathy and pheochromocytoma/paraganglioma. Given that these abdominal abnormalities are critical for patient management, it is important for radiologists to become familiar with the abdominal abnormalities that are common in post-Fontan patients on cross-sectional imaging.• Fontan procedure for functional single ventricle has improved patient survival into adulthood.• Radiologists should be familiar with unique imaging findings of Fontan-associated liver disease.• Focal nodular hyperplasia-like nodules commonly develop in Fontan-associated liver disease.• Hepatocellular carcinoma, protein-losing enteropathy, pheochromocytoma/paraganglioma may develop.The online version of this article (10.1007/s13244-018-0609-2) contains supplementary material, which is available to authorized users. The Fontan procedure was first introduced for patients with tricuspid atresia to allow systemic blood return to enter the pulmonary arteries without passage through a ventricle in 1968 by Fontan and Baudet . SubsequThe Fontan circuit allows the entire systemic venous return to enter the pulmonary vascular bed directly, bypassing the right ventricle. There are two ways to connect the systemic venous return to pulmonary arteries: either through the right atrium (namely atriopulmonary circulation) or by direct connection of superior vena cava (SVC) and inferior vena cava (IVC) to pulmonary arteries (namely cavopulmonary circulation). Cavopulmonary circulation is subdivided according to the type of IVC connection to the pulmonary artery either by intra-atrial lateral tunnel conduit or external cardiac conduit . Due to Fontan-associated liver disease (FALD) has been recognised as the disease process of liver structure and function resulting from the Fontan circulation, excluding other plausible causes such as viral hepatitis or alcohol toxicity. FALD is a broad term encompassing hepatic parenchymal change, hypervascular regenerative hepatic nodules and ultimately hepatic cirrhosis  . FALD iAlthough the imaging features of abdominal abnormalities after Fontan operation have been described in several publications mostly for the paediatric population –22, abdoAmong the gastrointestinal system, the liver is particularly vulnerable to the elevated systemic venous pressure in Fontan circulation compared to other organ systems for four principal reasons , 14. FirAdult patients with Fontan circulation are prone to developing FALD as they age. It frequently results in chronic hepatic congestion, liver cirrhosis, portal hypertension, focal nodular hyperplasia (FNH)-like nodules and even hepatocellular carcinoma (HCC) , 24–26. The severity of FALD is positively related to the duration of the Fontan circulation, elevated hepatic venous pressure, failing Fontan circulation and increased serological markers such as hyaluronic acid or gamma-glutamyltransferase , 29, 30.Given the clinical importance of FALD in adult patients with Fontan circulation, various non-invasive screening/surveillance protocols for FALD have been suggested regarding the timing, intervals, and modalities , 32, 33.The liver may appear normal or slightly hypoechoic on ultrasound (US) at the early stage of congestive hepatopathy. As the fibrosis develops, coarse heterogeneous hyperechoic parenchymal pattern and surface nodularity are visualised  Fig. 1aa1a. The In addition to the morphological changes of the liver, markedly heterogeneous hepatic enhancement with mosaic or reticular patterns, resulting from relatively slow and poor enhancement near the congested hepatic veins, is the most common imaging feature in FALD , 38. ConLarge regenerative hepatic nodules are commonly seen in adult patients with FALD. The pathophysiology of the development of the nodules is not fully elucidated, although it has been hypothesised that central venous hypertension transmitted directly to the hepatic peri-venular areas diminishes portal blood flow and promotes arterialisation –46. The Unfortunately, HCC can develop in adult patients with FALD , 46, 51 FNH-like nodules in FALD are often small and multiple, and show a predilection for the right lobe in the periphery (mostly within 2 cm of the liver surface) . On CEUSHCC typically shows , 51–57 aMRI using liver-specific contrast agent may be useful for the differentiation between FNH-like nodules and HCC, as HCC mostly show hypointensity in the hepatobiliary phase. DWI may be also useful in differentiating between HCC and FNH-like nodules. In our experience of a single case of HCC in FALD, HCC showed strong restricted diffusion, whereas FNH-like nodules did not show diffusion restriction, compatible with the study by Wells et al.  Figs. 33d and 4dOn Doppler US, normal hepatic vein shows a triphasic waveform, consisting of a hepatopetal (coming towards the liver) phase occurring during atrial systole and two hepatofugal phases related to atrial and ventricular diastole. Loss of the hepatopetal phase is typically considered to represent increased stiffness of the liver such as in hepatic fibrosis, steatohepatitis and cirrhosis –62.Fontan operation inevitably alters hepatic venous waveform on Doppler US. The pattern of hepatopetal phase in hepatic vein differs between patients with total cavopulmonary anastomosis  and with atriopulmonary connection –66 Fig.a–e. HepaPortal venous waveform on Doppler US normally shows slightly phasic hepatopetal flow with mildly decreased flow during inspiration due to transient compression of the compliant liver parenchyma and easily collapsible portal venules and hepatic sinusoids , 68. In On the arterial-phase CT with contrast injection through the upper extremity veins, dense opacification in the dependent portion of the IVC and hepatic veins is occasionally seen in post-Fontan patients. This is likely due to passive retrograde flow of the contrast material which is heavier than blood in the absence of effective right heart function  is characterised by enteric loss of proteins including albumin, immunoglobulins and clotting factors. Patients with PLE present with peripheral oedema, ascites, diarrhoea, weight loss and malabsorption. The reported incidence of PLE in Fontan patients ranges from 3 to 18%, with a reported mortality of 50% within 5 years after initial diagnosis –80. ElevEarly clinical diagnosis of PLE is often difficult because the patients may have a long asymptomatic period –82. As mPheochromocytoma and paraganglioma are neuroendocrine tumours arising from neural crest-derived cells or organs in the adrenal gland and along the central sympathetic and parasympathetic chains, respectively. There have been several anecdotal reports of the development of pheochromocytoma or paraganglioma in patients with congenital cyanotic heart disease after corrective or palliative cardiac surgery, including the Fontan procedure –90. SuggFontan procedures, designed to treat patients with functional single ventricles, have markedly improved the patient’s survival into adulthood. The physiology of the Fontan circuit inevitably increases systemic venous pressure, which may lead to multi-system organ failure in the long-term follow-up. FALD is most common among post-Fontan abdominal complications and can progress to liver cirrhosis with signs of portal hypertension. FNH-like nodules frequently develop in FALD as small multiple nodules with arterial-phase hyper-enhancement and show imaging findings similar to FNH. Imaging surveillance is often performed to monitor the progression of FALD to liver cirrhosis and to detect HCC, which infrequently develops in FALD. Extrahepatic abdominal abnormalities in post-Fontan patients include signs of portal hypertension such as ascites, splenomegaly and varices, protein losing enteropathy, and pheochromocytoma/paraganglioma. Given that these abdominal abnormalities are critical for patient management, it is important to be familiar with the typical imaging findings of the abnormalities on cross-sectional imaging.ESM 1(DOCX 7073 kb)