Ageing leads to dramatic changes in the physiology of many different tissues resulting in a spectrum of pathology. Nonetheless, many lines of evidence suggest that ageing is driven by highly conserved cell intrinsic processes, and a set of unifying hallmarks of ageing has been defined. Here, we survey reports of age-linked changes in basal gene expression across eukaryotes from yeast to human and identify six gene expression hallmarks of cellular ageing: downregulation of genes encoding mitochondrial proteins; downregulation of the protein synthesis machinery; dysregulation of immune system genes; reduced growth factor signalling; constitutive responses to stress and DNA damage; dysregulation of gene expression and mRNA processing. These encompass widely reported features of ageing such as increased senescence and inflammation, reduced electron transport chain activity and reduced ribosome synthesis, but also reveal a surprising lack of gene expression responses to known age-linked cellular stresses. We discuss how the existence of conserved transcriptomic hallmarks relates to genome-wide epigenetic differences underlying ageing clocks, and how the changing transcriptome results in proteomic alterations where data is available and to variations in cell physiology characteristic of ageing. Identification of gene expression events that occur during ageing across distant organisms should be informative as to conserved underlying mechanisms of ageing, and provide additional biomarkers to assess the effects of diet and other environmental factors on the rate of ageing. The ageing process encompasses progressive dysfunction in almost every organ in almost every organism, forming an immense challenge to modern medicine that seems to require many parallel interventions addressing an increasingly complex series of age-related conditions. A set of ‘Hallmarks of ageing’ have been defined, providing a framework for understanding ageing pathologies at the cellular and organismal level, but how these hallmarks arise and how cells respond to them often remains unclear , though these have required highly replicated sample sets at multiple time points or large cross-sectional meta-analyses to overcome confounding effects. Some very detailed meta-analyses have also directly addressed the question of whether conserved gene expression signatures can be detected across different cell types and organisms, identifying specific genes differentially expressed across datasets  all tissues regardless of their private age-linked gene expression programmes  and mitochondrial ribosomal proteins. This is observed in humans  ribosomal proteins and ribosome biogenesis factors. Ribosome biogenesis is a highly conserved pathway involving a host of proteins that act primarily in the nucleolus to process the four ribosomal RNAs and mediate temporally and spatially coordinated binding of ribosomal proteins  can represent small fold-changes (two-fold or less). This means that ribosome-related genes often miss differential gene expression cut-offs and fail to reach significance due to experimental and biological noise. Of course, these problems are addressed by improved statistical power, and it is reassuring that age-linked down-regulation of ribosomal protein and ribosome synthesis genes has been ubiquitously discovered by large cohort studies and meta-analyses in humans . This would imply that ribosome homeostasis is maintained in existing cells while fewer ribosomes are produced to supply new cells; however, the aforementioned reduction in ribosomal protein levels with age does not support this, and there have been many reports that translation efficiency decreases with age . Ageing is accompanied by an increase in the population of senescent cells, which have undergone an irreversible proliferative arrest in response to accumulated cellular damage, and this triggers the secretion of various cytokines, such as interleukin (IL)-6, IL-1α and IL-1β, chemokines, and proteases, which can promote inflammation  extend lifespan and delay ageing pathology substantially in worms, flies, mice and humans  pathway, which integrates nutrient availability and growth signalling, is ubiquitously associated with ageing in eukaryotes and mTOR inhibition by rapamycin has remarkable pro-longevity effects in disparate species (reviewed in Johnson et al. The existence of an age-linked stress response seems logical, particularly if ageing is accompanied by genetic and protein damage, and early transcriptomic studies in budding yeast reported evidence of this (Lesur and Campbell D. melanogaster, where remarkable similarities have been noted between gene expression changes that occur during ageing and those that result from oxidative stress or hypoxia (Landis et al. Drosophila cohorts have detected up-regulation of proteases and cytochrome p450 (Doroszuk et al. Much better evidence comes from D. mojavensis (Etges et al. Ercc1Δ/− repair deficient mutants, suggesting that transcription-blocking legions accumulate with age and bias the transcriptome (Vermeij et al. Even compared to other stress responses, evidence of increasing DNA repair is remarkably hard to find in transcriptomic studies. A handful of studies spanning a wide range of organisms report upregulation of repair genes (Etges et al. Overall, good evidence exists for age-associated occurrence of both protein and DNA damage (Lourenco dos Santos et al. There is evidence that the regulation of gene expression itself is affected by ageing, leading to intercellular heterogeneity in mRNA content that may represent noise. Bahar et al.  tested fIncreasing transcriptional heterogeneity and noise may be underpinned by changes in chromatin structure that occur during the ageing process. Promoter unmasking through progressive loss of histones with age is a major driver of gene expression noise in yeast (Hu et al. Defects in nuclear architecture and particularly in heterochromatin are associated with progerias including HGPS and Werner syndrome (Scaffidi and Misteli C. elegans. A recent study identified global defects in pre-mRNA splicing as a feature of ageing in C. elegans, and showed that the worm homologue of splicing factor 1, SFA-1, was required for lifespan extension by dietary restriction and modulation of the TORC1 pathway (Heintz et al. Changes in post-transcriptional processing of mRNA present another potential source of increased transcriptional variability. An early study reported a three-fold decrease in the proportion of cytosolic RNA that is polyadenylated in the livers of old rats compared with younger individuals (Yannarell et al. In summary, gene expression de-regulation represents a distinct transcriptional hallmark of cellular ageing that is likely to be directly associated with highly conserved age-induced changes in chromatin structure.Mitochondrial dysfunction hallmark of ageing, particularly given that upregulation of these genes is caused by many lifespan-extending treatments. Similarly, the impacts of Cellular senescence and Altered intracellular communication are well represented by dysregulation of immune system genes, consistent with this being perhaps the most prominent and reproducible phenotype of ageing in mammals.We compiled our list of hallmarks without direct comparison to the published hallmarks of ageing (Lopez-Otin et al. Loss of proteostasis hallmark is poorly reflected in gene expression datasets. As defined, this hallmark encompasses chaperones and responses to general protein mis-folding and damage for which there is limited evidence of change in the transcriptome, along with proteolytic pathways and autophagy that have only sporadically featured in reports of differential gene expression. It should be noted that changes in the inducibility of chaperones and damage response factors is a prominent feature of ageing in some and potentially all eukaryotes (see for example Labbadia and Morimoto In contrast, the Deregulated Nutrient Sensing clearly relates to the gene expression impacts of reduced growth factor signalling, but may also be the driver of reduced expression of protein synthesis factors given the tight regulation of ribosome biogenesis by nutrient signalling complexes such as TORC1. Nonetheless, the lifespan extension caused by deletion of ribosomal proteins in lower eukaryotes suggests that down-regulation of protein synthesis is more than a passive biomarker.Similarly, it is not clear which, if any, of the hallmarks of ageing can be directly attributed to the down-regulation of protein synthesis factors that is so clearly detected at the gene expression and proteomic levels. It may be that this is an effect rather than a cause; Genomic instability and Telomere attrition, however as discussed above, we see little evidence of a constitutive transcriptional response to DNA damage in ageing cells. The effects may well be indirect however; telomere attrition is the classic pathway to senescence and therefore telomere attrition may well contribute to the immune dysfunction gene expression hallmark. The de-differentiation and increased transcriptional noise observed in the transcriptome probably result from a combination of Genomic instability and Epigenetic alterations, which will have stochastic effects on gene expression at the individual cell level.Two hallmarks of ageing that might be expected to instigate a strong gene expression response are The identification of gene expression hallmarks associated with the cellular ageing process has multiple applications, despite the technical difficulties inherent in their detection. On one hand, characteristic patterns of age-linked gene expression change provide useful insights into the underlying drivers of ageing in diverse organisms, and will be very helpful in identifying outlier systems that do not reflect the normal progress of ageing. Furthermore, we envisage that gene expression differences will substantially precede ageing pathology, and therefore have significant potential as ageing biomarkers. Although substantial power is required to detect the hallmarks, this is not necessarily a problem given the rapidly decreasing cost of transcriptomic analysis, and although transcriptomic markers are unlikely to outperform well validated protein markers in studies focused on ageing, the generality of transcriptome analysis across studies in a broad range of fields should allow many additional datasets to be analysed for information on the ageing process. Discovering interventions that retard the ageing process as a whole is obviously a primary goal, and the ability to screen for beneficial effects in reduced timescales by transcriptomics should be extremely beneficial. Similar arguments can of course be made for the use of epigenetic ageing clocks, and we consider that the best approaches will integrate epigenetic and transcriptomic information, given that both are amenable to large studies.Drosophila showing that calorie content is not the primary driver of ageing and that extended lifespan can be combined with normal fecundity through appropriate choice of dietary amino acid composition (Mair et al. Benefits should not be restricted to purpose-made pharmaceutical studies and can also address lifestyle impacts on the ageing process. For example, it has long been known that dietary restriction is able to mitigate the physiological decline associated with ageing and extend lifespan to a significant extent. However, it is not clear that general dietary restriction is absolutely required to gain these benefits, with recent research in Transcriptomic analysis has clear limitations see “” section


The study of heat adaptation in military personnel offers generalizable insights into a variety of sporting, recreational and occupational populations. Conversely, certain characteristics of military employment have few parallels in civilian life, such as the imperative to achieve mission objectives during deployed operations, the opportunity to undergo training and selection for elite units or the requirement to fulfill essential duties under prolonged thermal stress. In such settings, achieving peak individual performance can be critical to organizational success. Short-notice deployment to a hot operational or training environment, exposure to high intensity exercise and undertaking ceremonial duties during extreme weather may challenge the ability to protect personnel from excessive thermal strain, especially where heat adaptation is incomplete. Graded and progressive acclimatization can reduce morbidity substantially and impact on mortality rates, yet individual variation in adaptation has the potential to undermine empirical approaches. Incapacity under heat stress can present the military with medical, occupational and logistic challenges requiring dynamic risk stratification during initial and subsequent heat stress. Using data from large studies of military personnel observing traditional and more contemporary acclimatization practices, this review article (1) characterizes the physical challenges that military training and deployed operations present (2) considers how heat adaptation has been used to augment military performance under thermal stress and (3) identifies potential solutions to optimize the risk-performance paradigm, including those with broader relevance to other populations exposed to heat stress. Heat stress, through occupational exposure to strenuous physical exercise and/or environmental extremes of heat and humidity, presents a perennial challenge  to militOver the last two decades, many Western militaries have conducted land-based campaigns in climatically-severe regions . These oRoutinely, the ‘special population’ fulfilling these roles is drawn from the general population of its parent nation and allied entities. Historically, the study of military personnel has provided insights into acclimatization mechanisms and strategies that have been generalizable to other occupational groups and the wider civilian population . The conIllustrated by research data from uniformed personnel observing traditional and more contemporary acclimatization practices – and with particular reference to recent United Kingdom (UK) military experience – this article (1) characterizes the physical challenges that modern training and operations present (2) considers how heat adaptation has been used or may be applied to protect and augment the health and performance of military personnel and (3) identifies potential solutions to optimize the risk-performance paradigm that operates both during and after heat acclimatization.No articles were dismissed in this review but, where appropriate, comment has been made on the limitations of the study presented, i.e., limited subject numbers and confounding methodological issues. Historically, based journals have been included where appropriate to indicate the limitations in current research. Use of the terms ‘military,’ ‘armed forces,’ ‘heat acclimation,’ ‘heat acclimatis(z)ation,’ heat adaptation and ‘heat illness’ were employed. Search engines included PubMed, Web of Knowledge and directly from the appropriate academic journals. Studies previously reported from our group’s work with the United Kingdom military are used to illustrate certain concepts, including novel unpublished data.In a volatile and increasingly complex world, governments of globally-facing nations foster contingency for a wide variety of overseas commitments . These mOperational capability may be impaired when unacclimatized personnel are required to travel at short-notice to a severe climate or to over-ride intrinsic or extrinsic thermal safeguards in the face of physical or tactical threats. For example, operational pressures may on occasion limit time or opportunity for full acclimatization, in favor of getting ‘boots on the ground’ with a view to achieving immediate military effects. From a physiological standpoint, however, inadequate or sub-optimal heat adaptation may have an array of consequences, the most feared of which is heat stroke and its associated morbidity and mortality . Heat stWhen an episode of heat illness occurs during, or soon after, strenuous physical exertion, it may be termed Exertional Heat Illness (EHI). This has been the predominant form of heat illness to affect British Army personnel in recent history, with EHI representing 96% of military cases reported by military doctors between 2009 and 2013  and contThe thermal challenge of military service raises questions over the optimal ways for parent organizations to exercise their duty of care and facilitate mission objectives. Heat adaptation is a key component of managing the overarching risk presented and, in the military as elsewhere, serves two key purposes: (1) the prevention of heat-related illnesses and (2) the improvement of physical performance and mental functioning. Crucially for military tasks, this may include improved decision-making under heat stress .Acquisition of the heat-adapted state occurs through repeated or continuous exposure to heat stress and accompanying elevation in Tc and may be achieved with artificial exposure to heat, by residence in a natural hot climate, or with physical training sufficient to raise Tc in less severe conditions . PhenotyBass and Henschel defined acclimatization as ‘the dramatic improvement in the ability to work in the heat which occurs within 4 to 7 days of first exposure’ . Over thHeat adaptation from exercise-heat stress is commonly induced through three mechanisms : (1) conOver the short term moderate intensity exercise appears to induce a more efficient physiological adaptation  in termsIsothermic or controlled hyperthermic acclimation commonly manipulates the endogenous thermal load under constant exogenous heat stress , but can2max and the number of days required to acclimatize  to promote an initial rest day, with no exercise permitted, in order to allow for transient factors specifically associated with preceding travel that may otherwise undermine heat tolerance . The impIn assessing the impact of progressive aerobic activity added into a program of ‘usual outdoor military activities,’ These data provide a basis to understand how a state of partially complete, or temporarily undermined, heat adaptation may risk more serious forms of heat illness; they would also help to explain the increased hospitalization risk in heat illness cases . One facAdaptations to heat exposure is never permanent. According to There are several factors, environmental and logistical, which may attenuate or even reverse the physiological effects of heat acclimatization. Whilst some of these factors are specific to the military others are relevant to other occupational groups including firefighters and humanitarian organizations. Core physiological changes in heat adaptation include increased sweat rate, sweating occurring at a lower temperature  and decrNumerous studies have attempted to capture the potential risk factors implicated in heat illness. Commonly cited precipitants include: alcohol , medicatHemorrhage remains the primary modality of battlefield death  commonlyDehydration often used synonymously with hypohydration  and is aL-arginine (L-citrulline (That acclimatized soldiers succumb to severe heat illness, or heat stroke , suggestarginine  and L-citrulline  which altrulline  which hatrulline . The efftrulline  expansiotrulline .Organizing deployments to allow time for military personnel to acclimate not only improves heat tolerance but can pay dividends in terms of improved physical and cognitive performance . HoweverCommonly an objective assessment of heat tolerance is used by assessing physiological responses to exercise in the heat . In athlSeveral novel, often technological, solutions, largely through the reduction of exogenous or endogenous heat stress, have been developed for the purposes of preventing heat illness and potentially improving athletic performance . Such adHeat stress remains a persistent problem for militaries and is likely, with climate change, to represent a challenge in years to come. Militaries are drawn from the populations they serve. The diverse constituents of militaries, coupled with the individual responses to heat acclimation and the varied environments in which militaries operate make generalizations difficult. Thermal stress in modern military training and operational deployments must be reviewed in this context. Heat adaptation remains a mainstay of augmenting physical and mental performance in the military. The ultimate singular goal of a military is to succeed in mission objectives whether in training, contingency operations, or warfighting. The consequences of not understanding what attenuates heat adaptation can pose an existential threat to militaries and their personnel regardless of the threat of heat illness. Heat adaptation in the military should not be reviewed in isolation but in the context of warfighting including wounding, the wearing of personal protective equipment, dehydration and febrile illnesses. Further work should ascertain the mechanisms by which physical fitness and acclimatization may protect against heat illness, over and above the heat syncope mitigated by plasma volume expansion. In the military this is challenging due to the obvious confounder that fitter individuals are more likely to be part of DCCC units and at increased risk of heat illness. Methods of being able to rapidly and safely heat adapt large numbers of varied military personnel, easily measure the adaptive changes and maintain, even manipulate, this adaptation remains a topic of ongoing research. The military remains a special population for thermal research.IP and MS researched, drafted, and edited the manuscript. DW provided editing and overarching review of content.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. All authors are serving members of the British Army.


Monitoring seasonal influenza epidemics is the corner stone to epidemiological surveillance of acute respiratory virus infections worldwide.This work aims to compare two sentinel surveillance systems within the Daily Acute Respiratory Infection Information System of Catalonia (PIDIRAC), the primary care ILI and Influenza confirmed samples from primary care (PIDIRAC-ILI and PIDIRAC-FLU) and the severe hospitalized laboratory confirmed influenza system (SHLCI), in regard to how they behave in the forecasting of epidemic onset and severity allowing for healthcare preparedness.Epidemiological study carried out during seven influenza seasons (2010–2017) in Catalonia, with data from influenza sentinel surveillance of primary care physicians reporting ILI along with laboratory confirmation of influenza from systematic sampling of ILI cases and 12 hospitals that provided data on severe hospitalized cases with laboratory-confirmed influenza (SHLCI-FLU). Epidemic thresholds for ILI and SHLCI-FLU  as well as influenza A (SHLCI-FLUA) and influenza B (SHLCI-FLUB) incidence rates were assessed by the Moving Epidemics Method.p-value Wilcoxon test = 0.039).Epidemic thresholds for primary care sentinel surveillance influenza-like illness (PIDIRAC-ILI) incidence rates ranged from 83.65 to 503.92 per 100.000 h. Paired incidence rate curves for SHLCI –FLU / PIDIRAC-ILI and SHLCI–FLUA/ PIDIRAC-FLUA showed best correlation index’ (0.805 and 0.724 respectively). Assessing delay in reaching epidemic level, PIDIRAC-ILI source forecasts an average of 1.6 weeks before the rest of sources paired. Differences are higher when SHLCI cases are paired to PIDIRAC-ILI and PIDIRAC-FLUB although statistical significance was observed only for SHLCI-FLU/PIDIRAC-ILI (The combined ILI and confirmed influenza from primary care along with the severe hospitalized laboratory confirmed influenza data from PIDIRAC sentinel surveillance system provides timely and accurate syndromic and virological surveillance of influenza from the community level to hospitalization of severe cases. Influenza seasonal epidemics occur yearly during each hemisphere’s winter season, whether it be in the Northern or Southern hemisphere. Acute respiratory infection caused by influenza virus ranges from mild to severe and can even cause death in at risk population such as the elderly and young infants. Annual epidemics are estimated to result in about 3 to 5 million cases of severe illness, and about 250,000 to 500,000 deaths worldwide. Hospitalization and death occur mainly among these high risk groups, which include also pregnant women and anyone with underlying medical conditions such as diabetes, obesity, cardiovascular or chronic obstructive pulmonary disease , 2.Influenza epidemics can cause an overload in the healthcare system because of high incidence rates of affected population, especially during peak illness periods. Influenza surveillance is made up information on morbidity and mortality as well as virological assessment of circulating influenza viruses. The resulting assessment and interpretation of these data can support public health authorities to implement special measure to strengthen capacity and preparedness which will in turn translate into action a reduction of morbidity and mortality. In most developed countries a network of sentinel physicians report cases attended for influenza-like illness (ILI), which is used as a proxy to estimate influenza virus circulation, and on the other hand collect samples for virological confirmation, identification of causative virus and describe predominant circulating type and subtype of influenza virus .In some countries community-based ILI reporting systems and indicators such as school and workplace absenteeism data are recorded weekly and have been used to estimate a proxy influenza intensity in combination with routine outpatient physician consultations and hospital influenza sentinel surveillance data , 5.Sentinel surveillance networks in primary care are a readily available and basic source of data that estimates disease burden of ILI cases as well as laboratory-confirmation of respiratory viruses, including influenza, by systematic sampling of ILI cases. From these data thresholds can be established to pinpoint the start and termination of influenza seasonal epidemics. Other relevant information gathered, such as patients’ vaccination status, will allow for vaccine effectiveness estimation . InfluenIn all, vaccine effectiveness is hampered and derives into low vaccine uptake. These facts underscore the importance of any surveillance data available whatsoever, making them essential for the detection and preparedness for any changes or unusual behavior of seasonal and non-seasonal influenza. However, a limitation to sentinel surveillance is the limited coverage of population within a geographical area and the bias towards young children who are more prone to seek medical assistance. During the 2009 influenza pandemic, the need to assess influenza severity became evident and from then on a number of European such as Spain, as well as other, countries introduced surveillance of severe disease and death due to influenza into their seasonal influenza surveillance systems –10.In Catalonia severe hospitalized confirmed influenza data from sentinel hospitals are informed on a weekly basis to the Daily Acute Respiratory Infection Sentinel Surveillance System of Catalonia (PIDIRAC) . IdentifHospital admissions for influenza and pneumonia through emergency departments provide some information on community cases, yet it is currently not easy to track a patient’s journey from community care (primary care consultations) into the hospital and the lack of denominator (source population) data is a major limitation of hospital-based surveillance .In October 2010, the Public Health Agency of Catalonia of the Department of Health in Catalonia, a region in the northeast of Spain with 7.5 million inhabitants, implemented the surveillance of severe hospitalized cases of confirmed influenza as a tool to complement information provided by the influenza sentinel system based on primary healthcare physicians implemented since 1998 [The PIDIRAC primary care network was composed by 60 sentinel physicians, general practitioners and pediatricians, covering 1% of the total population of Catalonia. Representativeness in terms of age, sex, and urban or rural setting was considered. Sentinel physicians reported ILI cases detected in their reference populations on a daily basis, following the European ILI case definition , 11. ForThe information collected in the PIDIRAC includes data on demographic, clinical and virological characteristics, seasonal vaccination status, chronic conditions, and pregnancy. Indicators provided by the PIDIRAC sentinel system for seven influenza seasons (2010–2011 to 2016–2017) was analyzed in the study. Indicators for this data source were the incidence rate of ILI syndromes (PIDIRAC-ILI) calculated according to weekly population under surveillance by each sentinel physician of the network that is reporting. Laboratory influenza confirmed isolates from reported ILI cases from primary care (PIDIRAC-FLU) were registered as weekly incidence rate positive to influenza. The latter indicator was separated as to influenza virus type A (PIDIRAC-FLUA) and type B (PIDIRAC-FLUB).The surveillance of Severe Hospitalized Laboratory Confirmed Influenza (SHLCI) was made up by twelve tertiary care hospitals representing a mean of 63.6% of the population under surveillance. Coverage increased from 53.3 to 94% of the total Catalan population in the 2015–2016 season. The catchment population for SHLCI surveillance was the population assigned to each hospital by the health system as a referral facility for any condition leading to hospitalization.The system is based on the notification of those laboratory confirmed influenza hospitalized cases admitted to the sentinel facility and who met the SHLCI case definition: Any case with clinical features compatible with influenza  , requiriOnly cases that were laboratory confirmed for influenza and met the SHLCI case definition were reported, regardless of whether they had been previously admitted to emergency room and transferred to ward or admitted to ward on initial consultation.Date of hospitalization and type and subtype of influenza virus were collected, other clinical and sociodemographic variables and outcomes, although collected, were not relevant for this study. We used data obtained from the surveillance of SHLCI for the same study period as in primary care surveillance for ILI, to calculate the cumulative hospitalization rates for each season and threshold for epidemic onset.Indicators for this data source were the incidence rate of severe hospitalized influenza laboratory confirmed cases (SHLCI -FLU) and separated as to influenza virus type A (SHLCI-FLUA) and type B (SHLCI-FLUB). For SHLCI-FLU and SHCLI-FLUA and FLUB the same method was applied to calculate epidemic thresholds using MEM system , 14.Epidemic thresholds for incidence rates by the Mobile Epidemics Method (MEM) were calculated and categorized as to intensity of epidemic  for the PIDIRAC-ILI and PIDIRAC-FLU indicators taking into account the seven epidemic seasons included in the study , 15.Epidemic curves obtained from both data sources were paired as PIDIRAC-ILI vs SHLCI-FLU, PIDIRAC-FLU vs SHLCI-FLU, PIDIRAC-FLUA vs SHLCI-FLUA and PIDIRAC-FLUB vs SHLCI-FLUB. For each pair, normal partial correlation were calculated at log scale. The week at which incidence rate exceeds estimated epidemic threshold level was used as an indicator to identify whether one source forecasts the onset of the epidemic activity with respect to the other. To calculate this factor by pairs the Wilcoxon signed rank test was carried out.To compare intensity of influenza epidemic observed by both data sources, correlation of intensity was calculated for each pair and the percentage of agreement within each epidemic threshold level. Correlation was defined as the proportion of weeks in which the observed intensity of activity for one source matches with the observed by the other source. To identify whether one source forecasts the onset of the epidemic with respect to the other for each season and for each indicator, the week at which incidence rate exceeds the estimated epidemic threshold level, this factor was studied by pairs according to Wilcoxon signed rank test.http://cran.r-project.org).The analysis was performed using the SPSS v.18 statistical package and the R v3.3.0 statistical software  although threshold level reached did not surpass the moderate level and did not reach high intensity level.Incidence rates of SHLCI-FLUA cases reached peak values of 15–20 cases/100,000 inhabitants, which are set within the moderate intensity level during epidemic weeks, while incidence rates of SHLCI-FLUB were less frequent and displayed lower incidence rates of 3–5 cases/100,000 inhabitants   Table .Table 2DThis study is based on surveillance data of ILI incidence rates from primary care sentinel surveillance, virology results from sampling by sentinel primary care physicians and severe laboratory-confirmed influenza (SHLCI) that required hospital admission during seven influenza epidemic seasons, from 2010 to 2017, in Catalonia. Data obtained from these seven epidemic seasons showed a good correlation between the two systems, primary care and SHLCI, although some differences in prompt detection of epidemic onset were observed according to predominant circulating influenza virus.Case definitions with a lower relative specificity for influenza, such as ILI can decrease the specificity of the model to detect an influenza season . This ocThe present study is an observational study with strengths and limitations. As a strength we can highlight that there are few studies investigating timeliness of sentinel primary care vs SHLCI to predict the onset of the epidemic allowing for on time preparedness to cope with the influenza epidemic and will allow for timely assessment of intensity and severity of an influenza epidemic.This study reveals the importance of both systems to understand yearly influenza epidemic behavior and to strengthen healthcare resource preparedness.Although MEM was primarily developed for modeling ILI data, it has been proven successful with SHLCI data showing that influenza severe cases’ waves correlate well with ILI, with higher epidemic thresholds and shorter range among intensity thresholds. Intensity, understood as the level of the population consultation rates or the percentage of ILI primary care consultations which is not synonymous of intensity or severity specific to influenza, but has proven to be one of the most reliable indicators to describe the impact of influenza on the population. On the other hand, SHLCI can complement primary care indicators giving a picture of the severity of the epidemic. Comparing intensities and severity across seasons is essential for the understanding of seasonal epidemic patterns and of future pandemics, and to assess control measures, such as the effectiveness of vaccination campaigns .As a limitation we consider the fact that a certain proportion of the population will not seek medical attention. Other complementary systems could possibly account for those who do not seek health care attention for ILI have been described such as Influweb or internet flu consultation trends such as Google flu trends, although these systems have, by themselves alone, low representability , 21.An epidemic threshold was determined for the ILI consultation rates reported by the sentinel physicians and for the SHLCI to assess the period of increased influenza epidemic earlier. Yet on time complete reporting of hospitalized severe cases is, at the best scenario, on a week’s delay, making this system unable to predict an overload in hospital resource demand with anticipation. Nevertheless the surveillance of SHLCI has the potential to reveal timely insights into epidemic severity as has been also pointed out by other authors –24.Besides, the voluntary participation of hospitals that could account for a selection bias. However, although a selection bias cannot be ruled out, the 12 participating tertiary hospitals cover 62% of population of Catalonia meaning that the effect can be assumed not to invalidate data.Another limitation is the representativeness of the samples tested in primary care by the systematic sampling of two first weekly ILI patients attended. Other circulating respiratory virus can mimic influenza and contribute to increase in ILI incidence rates, not so in SHLCI that are all influenza confirmed. Thus, increase in hospitalization of SHLCI reflects true influenza virus circulation, albeit a certain delay depending on circulating virus predominance. The implementation of a third hospital warning system based on data from emergency department records for laboratory confirmed influenza would be desirable to further improve timely information to health care for appropriate response.Another limitation to determining burden of hospitalization due to confirmed influenza is presented by the lack of the inclusion in the sentinel surveillance system of a data source for Influenza hospitalization. The use of other data systems such as hospital discharge data might spare the underestimation of the burden of laboratory confirmed influenza hospitalization . ImproveResults of two different surveillance systems during seven influenza seasons studied showed similar trends and were highly correlated with each other. The ILI consultation rates reported by the sentinel primary care physicians remain the basis of surveillance in Catalonia, because the system integrates epidemiological and virological information. Hospital data and virological data will remain very important to provide information on the severity, molecular and phenotypic characterization of the viruses, and whether the ILI incidence is truly related to influenza virus infections.The combined PIDIRAC Influenza sentinel surveillance system provides timely and accurate syndromic and virological surveillance of influenza from the community level to hospitalization of severe cases. The implementation of a hospital warning system based on data from emergency department records would be desirable as a complementary component of the combined outpatient-SHLCI scheme, allowing to inform control measures to lessen the burden of influenza in Catalonia and ensure access to quality timely information to health care for accurate preparedness.


Viruses will explore the impact of VSGs on the outcome of virus infection and the role of these genes within the infected cells and organism. We have focused on the most recent discoveries in VSG research, including the molecular biology of related virus–host interactions. Topics include studies on various steps of gene induction, innate immune responses to virus infection, and the mechanisms of virus immune evasion of related host defense pathways. The clinical presentation of VSG-driven pathology and strategies to use VSGs to cure chronic viral infections are also discussed. In this special Issue, we endeavored to assemble a collection of research papers and reviews that, together, will offer a comprehensive view on new areas of research pertaining to VSGs. Over the last few decades, accumulating evidence has demonstrated that numerous Interferon (IFN) stimulate genes (ISGs) can be directly upregulated following virus infection independent of IFN signaling. Indeed, whereas typical ISGs are driven by JAK-STAT signaling, other virally-stimulated genes are upregulated through the IRF3 and NF-κB pathways . These gMajzoub et al.  contribuDespite the recent progress in our understanding of these host defense responses, much more is left to be unraveled with respect to the regulation of antiviral responses triggered by distinct viral species. Clearly, IFN is a major component of host defense responses, but there are many ISGs whose antiviral activities remain to be clarified that can be regulated independently of IFN. These new genes would likely be ideal candidates to be targeted through therapeutic intervention as novel antiviral strategies.We hope this special issue provides a wealth of basic scientific data and introduces new concepts pertaining to ISGs that are regulated independently of IFN signaling.


Dynamic gain and loss of synapses is fundamental to healthy brain function. While Alzheimer’s Disease (AD) treatment strategies have largely focussed on beta-amyloid and tau protein pathologies, the synapse itself may also be a critical endpoint to consider regarding disease modification. Disruption of mechanisms of neuronal plasticity, eventually resulting in a net loss of synapses, is implicated as an early pathological event in AD. Synaptic dysfunction therefore may be a final common biological mechanism linking protein pathologies to disease symptoms. This review summarizes evidence supporting the idea of early neuroplastic deficits being prevalent in AD. Changes in synaptic density can occur before overt neurodegeneration and should not be considered to uniformly decrease over the course of the disease. Instead, synaptic levels are influenced by an interplay between processes of degeneration and atrophy, and those of maintenance and compensation at regional and network levels. How these neuroplastic changes are driven by amyloid and tau pathology are varied. A mixture of direct effects of amyloid and tau on synaptic integrity, as well as indirect effects on processes such as inflammation and neuronal energetics are likely to be at play here. Focussing on the synapse and mechanisms of neuroplasticity as therapeutic opportunities in AD raises some important conceptual and strategic issues regarding translational research, and how preclinical research can inform clinical studies. Nevertheless, substrates of neuroplasticity represent an emerging complementary class of drug target that would aim to normalize synapse dynamics and restore cognitive function in the AD brain and in other neurodegenerative diseases. A fundamental concept linking structural and synaptic plasticity is Hebbian learning , or consider what possibilities there are to study synaptic plasticity non-invasively in living humans. Preclinical studies are still valuable – but should be utilized more in the context of understanding pharmacokinetic-pharmacodynamic relationships of drug effects rather than being expected to predict drug efficacy.in vivo but have yet to be used routinely. Excitingly, recent work has shown that synaptic protein markers are secreted in exosomes and their levels are correlated to cognitive decline in FTD and AD  employs FDG-PET and cognitive endpoints to assess the effects of a methylene blue derivative on synaptic function. A second trial (NCT02167256) is also using similar FDG-PET and cognitive endpoints to assess the effects of the Src/Abl tyrosine kinase inhibitor saracatinib on synaptic function. Lastly, the SV2a PET ligand UCB-J is being used to measure the effects of the sigma2 receptor antagonist CT1812 on synaptic density in mild to moderate AD patients (NCT03493282). Hopefully positive findings in one or more of these studies will catalyze further research here.Recent work has greatly advanced our understanding of the direct and indirect means by which β-amyloid and tau pathologies may negatively impact synaptic function in AD. Considering AD as a disease of synaptic function and finding novel therapies that specifically target the synapse is an exciting avenue of research that complements traditional approaches in this field. Discovery and validation of synaptic therapies should focus on human disease rather than animal models wherever possible. An initiative taking such an approach is the Accelerating Medicines Partnership – Alzheimer’s Disease (AMP-AD) network, which via post-mortem gene expression and proteomic studies in AD patients, has flagged several synapse-related gene networks. These include a VGF-centered Alzheimer’s disease network, where the majority of genes regulate neuronal activity, synaptic plasticity, and cognitive function  or genesAll authors contributed intellectually to the manuscript. JJ and GG wrote the manuscript. All authors approved the manuscript. Figures created with BioRender.All authors are employees of Eli Lilly.


High-throughput shotgun metagenomics sequencing has enabled the profiling of myriad natural communities. These data are commonly used to identify gene families and pathways that were potentially gained or lost in an environment and which may be involved in microbial adaptation. Despite the widespread interest in these events, there are no established best practices for identifying gene gain and loss in metagenomics data. Horizontal gene transfer (HGT) represents several mechanisms of gene gain that are especially of interest in clinical microbiology due to the rapid spread of antibiotic resistance genes in natural communities. Several additional mechanisms of gene gain and loss, including gene duplication, gene loss-of-function events, and de novo gene birth are also important to consider in the context of metagenomes but have been less studied. This review is largely focused on detecting HGT in prokaryotic metagenomes, but methods for detecting these other mechanisms are first discussed. For this article to be self-contained, we provide a general background on HGT and the different possible signatures of this process. Lastly, we discuss how improved assembly of genomes from metagenomes would be the most straight-forward approach for improving the inference of gene gain and loss events. Several recent technological advances could help improve metagenome assemblies: long-read sequencing, determining the physical proximity of contigs, optical mapping of short sequences along chromosomes, and single-cell metagenomics. The benefits and limitations of these advances are discussed and open questions in this area are highlighted. Microbes are microscopic organisms that include prokaryotes (bacteria and archaea), viruses, fungi, and protists. These organisms, particularly prokaryotes and viruses, are known to rapidly adapt to novel abiotic and biotic environmental changes. The genetic bases for these adaptations have largely been identified by studying the genomes of isolated organisms of interest, which have greatly improved our understanding of the genetic bases of adaptations throughout microbial evolution .A substantial proportion of microbes in natural communities are currently uncultured  and acquNew genes can be acquired through three processes: 1) gene duplication and diversification, 2) the gain of a de novo gene , and 3) horizontal gene transfer (HGT). In addition to the gain of new genes, microbes are known to adapt to new environments through gene loss. All of these processes will be described in detail below.Although most MGS analyses are based on gene-centric methods, assembling MGS reads into genomes is one clear way to improve the identification of gene gain and loss events. The key challenge of this approach is that assembly errors can result in gene gain and loss events being falsely identified or missed. Several recent technological advancements could help overcome this barrier. These technologies include long-read sequencing, mapping the physical proximity and interactions of DNA fragments, optical mapping of short sequences along chromosomes, and single-cell metagenomics.Herein, the approaches applied so far to detect gene gain and loss events in MGS data are reviewed. This review largely focuses on identifying HGT events because these events are of primary interest within the field and there have been several recent methodological advances in HGT-detection in the context of metagenomes. However, methods to identify alternative processes of gene gain and loss will first be discussed . The bacAlthough HGT is the predominant mode of gene gain studied in MGS data, several other mechanisms of genic gain and loss are important to consider: gene duplication, gene loss, and de novo gene birth . The relGene duplication has long been known to be an important process underlying adaptation to novel environments . In prokRhodanobacter  is an element within plasmid DNA that specifies where the relaxase protein binds , which are phage-like particles that transfer DNA between cells. At least five families of GTAs have been identified in prokaryotes . Althoug species . It rema species . Two add species  and memb species , which a species .Numerous approaches have been developed for detecting HGT in genomics data , which aIn contrast, identifying potential HGT events in assembled contigs would be a source of direct evidence. The major challenge is that it is often only possible to assemble MGS reads into short contigs, which commonly represent a small fraction of complete genomes and can be enriched for assembly errors. Many existing methods for detecting HGT events in complete genomes cannot be applied to this fragmented data. This challenge is gradually being addressed as deeper sequencing read depths and improvements in related technologies have enabled higher-quality MAGs to be produced, as we discuss in the next sections. Nonetheless, methods specifically focused on poorly assembled MGS data have also been developed. These approaches for identifying putative HGT in both high and low-quality MAGs will be discussed below.The first major approach developed to detect HGT was based on comparing base and codon usage composition across genes within a genome . This geVerrucomicrobia to identify putative HGT events  is a software that implements a similar approach (manuscript in prep). The WAAFLE pipeline involves identifying the most similar matches in a pan-genome database for each gene in a contig. The tool then determines whether the genes in the contig can be explained entirely by a single species or if multiple species are needed to account for the contig gene content. This latter case is taken as putative evidence of HGT. The major strength of this approach is that multiple similarity matches are retained per gene in a contig. This information enables a conservative taxonomic-assignment approach to be employed where contigs can be classified as a single species, even if that species is not the best-hit for each gene, which is intended to reduce the number of false positives.For short MGS-derived contigs, one taxonomic-assignment approach has been proposed that involves running BLASTX  on genesAlthough the similarity-based HGT-detection methods described above use phylogenetic principles, they do not make direct use of phylogenetic trees to test for phylogenetic incongruencies. Testing for phylogenetic incongruencies refers to comparing a gene tree based on homologous sequences across taxa with the phylogenetic tree for those taxa. The growing number of prokaryotic genomes from pure cultures has enabled large-scale phylogenetic methods to be developed . SimilarThe methods for detecting HGT presented above approach the problem from a range of perspectives, which can result in strikingly different inferences of HGT. One example is a comparison of a compositional and a sequence similarity approach that resulted in fewer than 5% of HGT events in agreement . This drMetaCHIP is a recently published tool that is partially based on this idea, but intended specifically to identify HGT events between observed donor and recipient genomes in a natural community . This toAlthough the majority of HGT-detecting methods are focused on identifying transfers between different species, studying MAGs can also yield insight into population heterogeneity within a species. Because MAGs are based off the genomes of numerous bacterial cells in a community there is always some degree of genetic variation in the reads underlying MAGs. In addition, homologous recombination is known to occur between divergent strains, which can result in mosaic genomes with different gene blocks . Due to Leptospirillum and Ferroplasma species originating from acid mine drainage sites . The second obstacle preventing researchers from detecting HGT events is that these analyses require substantial bioinformatics expertise. Determining which approach to use is nontrivial and will largely depend on the biological question. For example, different methods are available if researchers are interested specifically in identifying hotspots of recombination within a single species  or identOne reassuring final point is that although different approaches to identify HGT in isolate genomes identify mainly nonoverlapping sets of genes , the genThe major challenge facing the identification of all the genic events described above is poor metagenome assemblies. This issue has recently been commented upon in the context of low-quality MAGs being added into public genome repositories . Either There are many metagenome assembly tools currently available, which are predominately based on De Bruijn graphs of overlapping k-mers . The outContig binning, where contigs from the same species or strain are grouped, is another key step when generating MAGs. Binning approaches typically group contigs based on sequence composition  and similar coverage of mapped reads . The mosEvaluating the quality of MAGs is a crucial step once the final contig bins have been generated and guidelines for how to categorize MAGs based upon quality metrics have recently been established . The twoImportantly, when reporting gene gain and loss events in a set of MAGs it would be important to also report the estimated completeness and contamination within these MAGs. In particular, it would be important to establish within a given MAG that more gene gain and loss events were inferred than are expected given the two quality scores. Ideally, manual validation of inferred gene gain and loss events would also be performed upon assemblies. At minimum this validation would include visually assessing the read coverage across an assembly at the site of the inferred gene gain or loss event. In practice, manually validating many events in this way would not be feasible, but it could be performed for a representative set.Several recent technologies have been developed which potentially could result in improved MAGs . These tThere are two promising long-read technologies currently available: Single-Molecule Real-Time  and OxfE. coli K-12 mG1655 genome at 99.5% base accuracy by first making multiple-sequence alignments of nanopore reads to correct read errors . Their method involves annealing fluorescent probes that bind specific short sequences in DNA. The DNA is then passed through narrow chambers that require a DNA molecule to be passed through in a straight line. The location of the fluorescent probes along genomic fragments is captured as DNA passes through the chamber. This approach has recently been applied to human samples to identify SVs and translocation events , results in sets of reads that are derived from the same genomic fragment. This information is especially useful for resolving SVs and repeats and for phasing variants , for which it will likely remain unfeasible to sequence at sufficient coverage to produce many high-quality MAGs in the near future.There are also many open questions about how to scan for HGT in assembled genomes. Currently, genomic context and potential transfer mechanisms are not directly integrated into HGT-detection pipelines. Automatically identifying corroborating evidence for a transfer, such as the presence of nearby prophage sequences, could help identify recent HGT events, which has been previously argued . The feaRegardless of which bioinformatics approach is used, inferences of gene gain and loss in MGS data sets will continue to improve as higher-quality MAGs are produced. The promising technologies outlined above are still in the early stages of application and there remain many open questions. For instance, whether optical mapping can be accurately applied to mixed communities remains unclear. Additional validation of this approach in conjunction with MGS data is necessary to determine whether it would actually improve MGS assemblies. In addition, there are several limitations specific to individual technologies, such as long-read sequencing and Hi-C requiring larger biomass samples. Benchmarking of all of these promising methods is required on natural communities of varying richness, complexity, and biomass to evaluate whether these methods should be differentially applied depending on sample-type or whether general best practices could be developed using only a subset of approaches.


The quality of cardiopulmonary resuscitation (CPR) has been shown to impact patient outcomes. However, post-CPR morbidity and mortality remain high, and CPR optimization is an area of active research. One approach to optimizing CPR involves establishing reliable CPR performance measures and then modifying CPR parameters, such as compressions and ventilator breaths, to enhance these measures. We aimed to define a reliable CPR performance measure, optimize the CPR performance based on the defined measure and design a dynamically optimized scheme that varies CPR parameters to optimize CPR performance.  We selected total blood gas delivery (systemic oxygen delivery and carbon dioxide delivery to the lungs) as an objective function for maximization. CPR parameters were divided into three categories: rescuer dependent, patient dependent, and constant parameters. Two optimization schemes were developed using simulated annealing method: a global optimization scheme and a sequential optimization scheme.  Variations of CPR parameters over CPR sequences (cycles) were analyzed. Across all patient groups, the sequential optimization scheme resulted in significant enhancement in the effectiveness of the CPR procedure when compared to the global optimization scheme. in vivo is needed before implementing these changes in actual practice. Our study illustrates the potential benefit of considering dynamic changes in rescuer-dependent parameters during CPR in order to improve performance. The advantage of the sequential optimization technique stemmed from its dynamically adapting effect. Our CPR optimization findings suggest that as CPR progresses, the compression to ventilation ratio should decrease, and the sequential optimization technique can potentially improve CPR performance. Validation  Cardiopulmonary resuscitation (CPR) involves delivering chest compressions and positive pressure ventilation to cardiac arrest victims to maintain circulatory blood flow and oxygen delivery . Studies2]) have been established as important factors for measuring CPR performance [Another approach to optimizing CPR involves defining and establishing reliable, appropriate CPR performance measures and then modifying CPR parameters to enhance these measures. Brain ischemia is a primary contributor to postarrest morbidity; thus, oxygen delivery and carbon dioxide elimination  is the compression speed, and x is the compression to ventilation ratio [Qmax is the maximum systemic blood flow Qs, and for carbon dioxide delivery to the lungs, Qmax is the maximum pulmonary blood flow Qp. On average, we assume that systemic blood flow is equal to pulmonary blood flow; hence, During CPR, the blood gas (oxygen and carbon dioxide) delivery to any organ in the body can be estimated using the following simplified equation :(1)DBG=Qon ratio . For sys2 and ∆CCO2, we need to start with pulmonary gas exchange equations presented in [fA) is equal to end-tidal gas,To calculate ∆COented in . The pres2O and s2CO are slopes of oxygen and carbon dioxide dissociation curves, respectively. In CPR-related parameters,R is the average ventilation rate, vt is the tidal volume, and vd is the dead-space volume. Solving for alveolar fractions, we getIn the steady-state condition, the oxygen and carbon dioxide balance in the lungs can be expressed using the following equations:Since changes in concentrations are small, they are in the linear range of dissociation curves, or in mathematical terms:Qs,max and Qp,max are maximum systemic and pulmonary blood flow, respectively. In steady-state CPR condition, Qs,max and Qp,max are equal to ensure balanced blood flow to the body and the lungs. The objective function we sought to maximize was the total blood gas delivery, defined as the summation of carbon dioxide delivery to the lung and systemic oxygen delivery. It should be noted that a differentially weighted sum of the two deliveries might also be a possible criterion.Finally, combining the above equations will result in mathematical expressions for systemic oxygen delivery and carbon dioxide delivery:fI2CO. Patient-dependent parameters are parameters that vary from patient to patient, such as Qmax. Constant parameters are fixed parameters, such as s2CO. In the optimization procedure, we varied the rescuer-dependent parameters for each patient  over the entire range of patients. The described parameters and their values are listed in In the next step, CPR parameters were divided into three categories: rescuer dependent, patient dependent, and constant parameters. Rescuer-dependent parameters are parameters that depend on rescuer performance, such as Next, two different optimization schemes were developed: a global optimization scheme and a sequential optimization scheme. Both of the methods used simulated annealing as the optimization technique. Simulated annealing is a computational intelligence technique that aims to solve combinatorial optimization problems to minimize the defined cost function which is characterized by a large number of solutions . The simf at a given temperature T is defined by the following equation:pϕ is the probability of occurrence of state ϕ, which could be calculated using the following equation:Unlike most of the optimization techniques which could be considered analogous to rapid cooling methods, simulated annealing is analogous to a process of slowly cooling a physical system by providing ample time in order to obtain states with globally minimum energy . As a reQmax, we sought to maximize the total blood gas delivery that is expressed by Equation  during CPR. During initiation of CPR, the rescuer should place more emphasis on chest compressions than on ventilation (high compression to ventilation ratio) and gradually decrease the compression to ventilation ratio and carry out the positive pressure ventilation more frequently.T for three different Qmax are presented in The results obtained for average ventilation time t for three different Qmax are presented in The results obtained for compression speed 1/In this study, we present an analysis of dynamic modeling of CPR parameters during CPR to improve CPR performance. While previous research efforts to optimize CPR have focused on finding the best compression to ventilation ratio and keeping the ratio constant during the CPR procedure, we propose a sequential optimization scheme to vary the rescuer-dependent parameters as needed during CPR sequences to optimize CPR performance.Our results suggest that as CPR progresses, the compression to ventilation ratio should decrease over time to optimize performance. One possible explanation for this observation in our model is that when CPR typically commences, blood oxygen content is sufficient for satisfactory systemic delivery and the carbon dioxide level has only started to rise above normal levels. As CPR progresses, oxygen decreases in the blood and carbon dioxide starts to accumulate, so ventilation becomes increasingly important. Hence, our model suggests that the optimal compression to ventilation ratio should begin higher than is currently practiced (greater than 30 compressions per ventilation) and then decrease over time.In this study, we followed to a large extent the methods as described in a conference paper . In thatin vivo, and they should not be used to adjust current CPR practices until such validation has been completed.There were several limitations to this study. First, we used one published systemic oxygen delivery model to develop analytical expressions for both systemic oxygen delivery and carbon dioxide delivery to the lungs , and we In summary, our study illustrates the potential benefit of considering dynamic changes in rescuer-dependent parameters during CPR in order to improve performance. Validation in an animal model and an assessment of generalizability to real data sets are important future steps.


Hepcidin is the major regulator of systemic iron metabolism, while the role of this peptide in the brain has just recently been elucidated. Studies suggest a dual role of hepcidin in neuronal iron load and inflammation. This is important since neuronal iron load and inflammation are pathophysiological processes frequently associated with neurodegeneration. Furthermore, manipulation of hepcidin activity has recently been used to recover neuronal damage due to brain inflammation in animal models and cultured cells. Therefore, understanding the mechanistic insights of hepcidin action in the brain is important to uncover its role in treating neuronal damage in neurodegenerative diseases. Iron is a crucial element for maintaining brain homeostasis. It is used by brain cells to synthesize myelin, produce neurotransmitters, and realize a wide range of important cellular biochemical reactions . But, irMost of systemic hepcidin is produced in hepatocytes via paracrine signaling from liver sinusoidal endothelial cells (LSECs) . LSECs rIron homeostasis in the brain must be finely regulated to serve the purposes of brain cells without damaging cell structures. There are different mechanisms that allow for this to occur; first, blood brain barrier (BBB) is a natural barrier that does not allow passing of iron with ease, which means that iron has to be transported transcellularly via TFR . The iroIt is believed that systemic hepcidin can cross blood barriers such as BBB . HepcidiIt has to be mentioned that models with liver hepcidin knockout do not result in significant brain iron dysmetabolism . This woIn vitro results from this study showed that ad-hepcidin and hepcidin peptide reduced brain iron uptake by decreasing TFR1, DMT1, and FPN expression in brain endothelial cells. So, how could hepcidin presence during ICH cause opposite results? During ICH brain iron is increased even many weeks after the insult  has been consistently observed by w levels . Data frw levels . Similarw levels .Hepcidin upregulation can be elicited by acute iron load in astrocytes and microglia, while in neurons this response seems to become more evident with higher doses of iron supplementation . On the Figure 1). The magnitude of cell-specific response to LPS is highest in glial cells and lowest in neurons , toll-like receptor 4 (TLR4), IL-6, STAT3 molecules  homologous protein (CHOP), which has also been involved in regulating liver hepcidin expression during endoplasmic reticulum (ER) stress . BMP/SMAIn vivo experiments during brain inflammation show that the use of iron chelation protects from brain iron overload, reduces microglial activation and improves cognitive functions in rodents by reducing levels of hepcidin and increasing levels of FPN in hippocampus . Treatment with hepcidin in these conditions protects neurons from oxidative stress  expression in astrocytes and microglia   and in isolated nuclei in Parkinson’s disease (PD) . In AD, in vivo models with mice have shown that Aβ oligomer accumulation in AD is related to TLR4 activity, which is in contrast with α-synuclein accumulation in PD, where the role of TLR4 is low or inexistent  aberrant response to different stimuli . This meexistent . Also, mexistent . Furtherexistent . These dAlzheimer’s disease and PD are diseases that increase in prevalence with age. It is believed that the ability of microglia and astrocytes to secure the homeostatic equilibrium for neurons is reduced with aging. The homeostatic disequilibrium might be a result of the genetic factors that are related with increased activity of inflammatory signals, decreased ability of microglia to clear oligomer accumulation and other factors . “PathogAlthough evidence for the role of inflammation and resultant iron load in neurodegenerative diseases is accumulating, it is of therapeutic importance to explain how do these pathogenic factors change the dynamic of cellular biochemistry. Data from animal models show that aging creates a favorable milieu for neuronal damage via iron load. In rats, levels of DMT1 and hepcidin are increased, while levels of FPN do not change . This woIn other diseases accompanied with neurodegeneration  iron deposition is also increased in different brain structures . ChangesStimulation and inhibition of hepcidin has been used in cultured cells and animal models to ameliorate brain damage. The rationale is evident; hepcidin affects FPN and iron import proteins, which means its manipulation can control cellular iron load. Furthermore, it has been shown that decreased activity or expression of FPN promotes neuronal damage . In animIn animal models with brain inflammation and increased oxidative stress, direct and indirect suppression of local and systemic hepcidin offers neuroprotection . But, inTable 1). The protective effect of acetylsalicylic acid is realized through increase in iron export in neuronal cells physiology. The source of hepcidin in the brain is local and probably systemic. It is believed that systemic hepcidin can pass through BBB, especially during BBB leakage (for example during inflammation). Neurons, glial cells, endothelial cells, and other brain cells may express hepcidin, although basal levels of brain hepcidin are low. They increase significantly during inflammation and most likely during brain iron load. The local molecular regulation of brain hepcidin expression is unfolding, while studies suggest that it bears resemblance with liver hepcidin regulation. Still, the details of this regulation await further clarification from future studies. Hepcidin produced by the brain cells in pathological conditions may limit iron transport through BMVEC, and thereby also limit neuronal iron load. High iron load is detrimental for neuronal function. In addition, high iron load can also affect glial cell activity by turning them into cells that promote neuronal damage. The observation of robust production of hepcidin by glial cells compared to neurons during changes in brain homeostasis is in-line with the already established role of glia as supportive, regulatory and protective cells in the brain milieu. During inflammatory conditions hepcidin promotes neuronal iron load, while the opposite occurs in non-inflammatory conditions. This observation is based on studies that have revealed the neuroprotective nature of hepcidin suppression during inflammation. This occurs due to the increase in iron import caused by inflammation, which is further exacerbated by FPN downregulation caused by hepcidin-dependent and hepcidin-independent mechanisms. On the other hand, use of ad-hepcidin during neuronal iron load (not caused by inflammation) protects these cells from oxidative stress. Intriguingly, hepcidin pretreatment also protects neurons from the deleterious effects of inflammation. It seems that pretreatment with hepcidin blocks the initiation of inflammation-induced biochemical cascade inside brain cells. This effect of hepcidin is not without precedent, because it has been observed in mice and human studies. Unfortunately, therapeutic implications of this physiological effect of hepcidin have not been studied in details, despite its potential crucial importance in brain and other pathologies. Still, pharmacologic manipulation of hepcidin is emerging as e new therapeutic territory in neurodegenerative diseases. Animal models have shown that the use of ad-hepcidin and suppression of hepcidin protects brain cells in models of neurodegenerative diseases. This has been further confirmed with the use of standard anti-inflammatory drugs such as acetylsalicylic acid or lycopene.Finally, considering the ever increasing importance of hepcidin for brain physiology, it is paramount for future studies to examine models with different cell specific hepcidin knockout in AD and PD that will reveal the temporal changes of hepcidin, but that will also define whether hepcidin is of primary importance for the pathophysiology of neurodegenerative diseases.The author confirms being the sole contributor of this work and has approved it for publication.The author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


In this perspective, we congratulate the international efforts to highlight critical challenges in brain tumor research through a recent Consensus Statement. We also illustrate the importance of developing more accurate and clinically relevant early translational in vitro brain tumor models—a perspective given limited emphasis in the Consensus Statement, despite in vitro models being widely used to prioritize candidate therapeutic strategies prior to in vivo studies and subsequent clinical trials. We argue that successful translation of effective novel treatments into the clinic will require investment into the development of more predictive early pre-clinical models. It is in the interest of researchers, clinicians, and ultimately, patients that the most promising therapeutic candidates are identified and translated toward use in the clinic. Highlighting the value of early pre-clinical brain tumor models and debating how such models can be improved is of the utmost importance to the neuro-oncology research community and cancer research more broadly. The recent Consensus Statement ‘Challenges to curing primary brain tumors ’, producUsing the example of targeting the DNA damage response (DDR) , effectiWhilst organoids represent a promising 3D culture system highlighted in the Consensus Statement, other clinically relevant 3D co-culture systems do exist ,5, and tAdditionally, worthy of consideration are the neurosurgical features of deriving human brain tumor cells used in the majority of pre-clinical models, whether in vitro  or in vivo (PDXs). Generally, these cells come from the resected tumor taken out at surgery, even though the infiltrative residual cells left inside patients after surgery are responsible for disease progression. For example, considering glioblastoma, which is characterized by extensive spatial and temporal heterogeneity ,9, it shHighlighting the value of early pre-clinical brain tumor models and debating how such models can be improved is essential to enable the identification and translation of novel treatment strategies that are most likely to be successful in clinical trials for patients with brain tumors. In cancer research, developing more clinically relevant in vitro models to better prioritize novel therapies worthy of further investigation is critical. The translational ‘pipeline’ may only ever be as successful as those treatments which enter it.


Transitions older adults experience post hospital discharge have primarily focused on the process of moving care from one setting to another . Older adults often experience a significant transition in terms of losing functional status after a hospital stay. Little is known about how older adults regain their functional ability, the type of work they engage in to recover, and conditions that influence their ability to work after a hospital stay. The objective of this Grounded Theory study was to understand strategies older adults use post discharge as they work to regain their functional status and what conditions facilitate or limit their ability to work toward returning to normal. A qualitative study was conducted. Adults aged 65 and older discharged from a large Midwestern teaching hospital (N = 14) were interviewed using in-depth one-on-one interviews. Data were analyzed using open, axial, and selective coding. Participants described key strategies they employed to regain their normal function following hospitalization and illness: doing exercises, expanding physical space, resuming prior activities and daily cares, and tracking improvement with benchmarks. Several conditions such as, presence of informal  and formal  support, perceived threats (relocation), and having poor physical or physiologic function, acted as barriers and facilitators to participants ability to work back to normal function. This study provides empirical data on work older adults engage in to transition back to normal function during the post discharge period. It presents opportunities for better supporting their work of regaining function.


Cellular senescence is now considered as a major mechanism in the development and progression of various diseases and this may include metabolic diseases such as obesity and type-2 diabetes. The presence of obesity and diabetes is a major risk factor in the development of additional health conditions, such as cardiovascular disease, kidney disease and cancer. Since senescent cells can drive disease development, obesity and diabetes can potentially create an environment that accelerates cell senescence within other tissues of the body. This can consequently manifest as age-related biological impairments and secondary diseases. Cell senescence in cell types linked with obesity and diabetes, namely adipocytes and pancreatic beta cells will be explored, followed by a discussion on the role of obesity and diabetes in accelerating ageing through induction of premature cell senescence mediated by high glucose levels and oxidised low-density lipoproteins. Particular emphasis will be placed on accelerated cell senescence in endothelial progenitor cells, endothelial cells and vascular smooth muscle cells with relation to cardiovascular disease and proximal tubular cells with relation to kidney disease. A summary of the potential strategies for therapeutically targeting senescent cells for improving health is also presented. The senescent state is a stress response which ensures that cell damage is removed through activation of the immune system. During this response, often induced by persistent DNA damage, cells lose their ability to proliferate; ensuring cells do not become cancerous. In addition, senescent cells develop an altered secretome consisting of pro-inflammatory factors, growth factors and proteases. Thus, senescent cells appear to mimic a wound healing response. Components of the senescent secretome function in attracting immune cells for the elimination of these damaged cells, thereby promoting the restoration of tissue homeostasis  is essential for normal intercellular communication required to maintain optimal tissue homeostasis and for providing structural and biochemical support to cells  and pro-inflammatory cytokines  are immobilised from the bone marrow into the peripheral blood in response to vascular damage via the release of growth factors (of which VEGF appears to be an important factor), cytokines and chemokines  observed in diabetics has also been shown to reduce the number and impair function of circulating EPC. Providing further support for the link between diabetes and EPC senescence, Imanishi et al.  demonstrVascular endothelial cells (EC) play an important role in regulating vascular tone (via nitric oxide activity), cellular adhesion and filtration, smooth muscle cell proliferation and thromboresistance (resistance to non-haemostatic blood clotting). As such, when endothelial cells become dysfunctional, the biological consequence can be multifaceted. Induction of cellular senescence is likely one mechanism promoting endothelial dysfunction.l-arginine (a substrate for NO synthesis) reduced the appearance of senescent-like cells and increased NO production, suggesting that administration of l-arginine could improve vascular function during CVD. Arginase 1 is an enzyme which catalyses the hydrolysis of arginine to urea and ornithine. A recent study showed that arginase 1 gene deletion or pharmacology inhibition protected against induction of EC senescence in diabetic mice  may interact with membrane receptors of cells that alter intracellular signalling and promoting the production of pro-inflammatory cytokines and ROS  expression, EC senescence and endothelial dysfunction associated with a reduction in glycogen synthase 3 beta (GSK-3β) activity and p-β-catenin  calcification through activating NF-kB and downregulating IGF1R  leading to the build-up of waste fluids in the blood  from becoming senescent. As mentioned above with regard to EC senescence, this may include the use of drugs which inhibit arginase 1 and SIRT1 activity. In addition to preventing cell senescence, it may one day be possible to reverse cell senescence. It has been suggested that the use of resveratrol-related compounds  can rescue cells from senescence, but further research is required (Latorre et al. Currently the use of drugs is justified only when a disease has manifested and is problematic. In addition, there are often specific drugs for specific diseases. If many different diseases have a common underlying cause, such as cell senescence, it may one day be possible to treat many different diseases with a single drug. However, rather than waiting for a disease to present itself, intermittent removal of senescent cells throughout our lifetime could be utilised as a preventative measure, ensuring long and healthy lives.


Endothelial biomarkers are gaining interest in the stratification of cardiovascular risk and early diagnosis of cardiotoxicity secondary to antineoplastic drugs. Interestingly, some drugs, such as anthracyclines, have been recently associated with vascular damage, which reveals the pivotal role of research in identifying biomarkers that could potentially be included into more specific cardiotoxicity risk scores. An extensive report of the incidences of cardiovascular adverse effects of oncologic drugs is presented, with the main purpose of highlighting not only the risk of developing heart failure but also the importance of associated vascular adverse effects  experienced by patients in the post-chemotherapy phase. Due to the ongoing research on a clinically relevant subject such as the cardiotoxic effects of oncologic drugs as exemplified by Finkelman et al. in the article “Arginine-Nitric Oxide Metabolites and Cardiac Dysfunction in Patients With Breast Cancer” , we wantThe most confusing aspect of the definition of cardiotoxicity is whether only clinical, clinical and subclinical, or only subclinical events should be considered . AnotherEndothelial biomarkers are gaining interest; it is known that bevacizumab decreases endothelial nitric oxide synthase activity, which may stimulate plasminogen activator inhibitor-1 expression, leading to an increased risk of hypertension . FinkelmHerrmann J et al. proposed a Cardiotoxicity Risk Score (CRS) that included a medication-related risk depending on incidence rates of cardiotoxicity . The cutAs depicted in Figure Through this manuscript, we would like to reinforce the idea that when addressing cardiotoxicity, not only should the myocardium be considered in our evaluation but also the endothelium. The latter could play an important role in how our patients face the early and late post-chemotherapy phase, thus helping to stratify patients' risks and optimize their management.


Over the past two decades it has become increasing clear that injury and loss of podocytes is an early and common clinical observation presented in many forms of glomerulopathy and chronic kidney disease. Identification of disease-causing monogenic mutations in numerous podocyte-expressed genes as well as studies conducted using preclinical animal models have shown that the podocyte plays a central role in establishing kidney dysfunction. In this review, we summarize current knowledge regarding the potential for podocyte-targeted therapies and give our view on how a deeper understanding of the molecular makeup of the podocyte will enable future therapeutic interventions. Specifically, we recount some of the currently described podocentric strategies for therapy and summarize the status and evolution of various model systems used to facilitate our understanding of the molecular and functional underpinnings of podocyte biology. Glomerular capillary tufts are responsible for renal ultrafiltration of blood and formation of primary urine. The filtration barrier is formed of three layers: highly fenestrated glomerular endothelial cells, the glomerular basement membrane (GBM) and podocyte cells with their foot processes. Mesangial cells, that are considered to act as pericytes in the glomerulus, are the third cellular component of glomerular tufts.Diabetic nephropathy (DN), one of the microvascular complications of diabetes, affects 20–40% of patients with diabetes mellitus . Renal gNumerous studies in transgenic mouse models developed to express/inactivate single genes exclusively in podocyte cells, with the use of podocyte-specific promoters, have validated the podocyte as an attractive therapeutic target of therapy in DN. For instance, kidneys, in which epidermal growth factor receptor (EGFR) has specifically been deleted in podocytes, are protected from diabetic injury and the progression of DN is interrupted . These sWhile the progression of acquired glomerular diseases, such as diabetic kidney disease, is a result of the convergence of multiple injurious pathways, a deep and thorough understanding of the pathomechanisms initiating and driving podocyte-specific hereditary diseases may hold promise to identify and discriminate among those potential targets that might be best suited to interrupt the development of more complex acquired glomerulopathies. Over 50 monogenic glomerulopathies have been described and understanding the molecular mechanisms of their causative roles in mediating podocyte dysfunction in glomerular disease will hopefully provide us with the knowledge to realize the potential for manipulating pleiotropic gene function in non-hereditary glomerular disease. Most of these monogenic targets themselves are not pharmacologically tractable candidates, yet understanding the responsible mechanisms accounting for their roles in disease etiology may very well identify key secondary mediators that can be manipulated. In the following section, we provide examples of various therapeutic strategies where the podocyte may be considered the principal target cell involved in the differing modes of action.Today, patients with DN are treated with either angiotensin-converting enzyme (ACE) inhibitors or angiotensin receptor blockers (ARB). Both drugs reduce the glomerular filtration pressure by primarily inhibiting efferent arteriole vasoconstriction. Based on animal and cell culture studies, there is considerable evidence suggesting that these drugs may also partially mediate their beneficial effects via direct actions on cells of the glomerulus, including podocytes , 6. HoweGlucocorticoids are the mainstay of treatment for minimal change disease, the most common cause of nephrotic syndrome in children. These patients typically respond robustly to this treatment as massive albuminuria disappears in a few days after onset of glucocorticoid treatment. This outcome has traditionally been considered to be due to immunosuppressive effects, although recently, two interesting studies suggest otherwise , 11. KupCalcineurin inhibitors have been shown to have direct effects on podocytes . This stper se in mediating the effects of rituximab in glomerular disease  is a transmembrane protein mostly known as a costimulatory receptor on antigen presenting cells. Reiser et al. reported in 2004 that B7-1 expression was induced in podocytes in a mouse model of proteinuria and that this had a pathogenic role as B7-1 deficient animals were protected from proteinuria . In 2013These initial encouraging results have been followed by many studies that have questioned the value of B7-1 expression in podocytes as a biomarker and as a therapeutic target. In patients with minimal change disease, FSGS and DN, no B7-1 expression was detected in podocytes , 20. In Gain-of-function mutations in the transient receptor potential cation channel TRPC6 are associated with the onset of familial forms of FSGS and increased expression of this channel has been described in acquired glomerular disorders including DN , 24. TRPMutations in enzymes involved in coenzyme Q10 biosynthesis, such as Coq2, Coq6 and aarF domain containing kinase 5 (ADCK4), have been identified by whole exome sequencing in few families with hereditary glomerular disorders –32. The Dysregulation of the actin cytoskeleton in podocyte foot processes is a central mediator in the pathogenesis of proteinuria. This is best highlighted by studies in hereditary proteinuric diseases as disease-causing mutations have been identified in several molecules involved either directly or indirectly in the regulation of the podocyte actin cytoskeleton , 34. TarDynamin is a GTPase regulating endocytosis and actin cytoskeleton assembly. Dynamin oligomerizes in an actin-dependent manner and promotes actin polymerization via direct interaction. It has been shown to be essential for podocyte structure and function. A small molecule, Bis-T-23, has previously been shown to augment dynamin oligomerization in cultured podocytes. Schiffer et al. treated various rodent models of proteinuria with Bis-T-23 and interestingly showed that the treatment ameliorated proteinuria and development of histopathological changes, supporting the idea that the podocyte actin cytoskeleton can be directly targeted to treat proteinuric diseases . They diWe recently identified an orphan G-protein coupled receptor (GPCR), Gprc5a, as a new highly podocyte-enriched protein that was significantly downregulated in patients with DN . ImportaAs a GPCR, Gprc5a, may represent a particularly interesting opportunity to directly target and pharmacologically manipulate podocyte signaling and health. However, Gprc5a is an orphan receptor that lacks the long N-terminal extracellular domain that is usually necessary for ligand binding and activation of downstream signaling via G-proteins of this class of receptor. In fact, we do not know today whether Gprc5a couples to G-proteins and to what intracellular signaling pathways it specifically couples and activates. Further studies are warranted to further understand the signaling role of this receptor in podocytes and whether manipulating such signaling may be podocyte protective.The examples given above provide a representative sample of the data supporting the potential therapeutic efficacy that might be afforded by targeting the molecular machinery of the podocyte. In the section below, we describe some of the model systems used to enable the molecular and functional characterization of podocyte biology.in vivo . Also very recently, the UK HARP-III trial designed to assess renal efficacy of Entresto (a combined ARB and neprilysin inhibitor), was shown to be without significant improvement on ACR and GFR vs. Irbesartan (ASN 2017 poster FR-PO1064). Perhaps the most interesting and promising example of drug repurposing comes from the SGLT2 class of inhibitors designed to facilitate blood glucose lowering in diabetes by preventing glucose reabsorption along the proximal tubule and promoting glucosuria. In addition to its glucose lowering activity, its kidney-specific MoA speaks in favor of a direct renal intrinsic mechanism where tubuloglomerular feedback is activated secondary to reduced distal tubule delivery of sodium. In this manner, a number of clinical trials are underway to determine whether SGLT2 inhibitors might become the first kidney-targeted therapeutic that is beneficial in treating CKD.As described earlier in this review, ample preclinical evidence supports the therapeutic potential of targeting the podocyte. Moving from proof of concept in rodent models of disease to clinical evaluation still has a long way to go. The molecular mechanisms of podocyte dysfunction in disease continue to be unraveled and there is reason to be optimistic that with continued research, novel therapeutic targets will be identified. FSGS orphan indications will hopefully pave the way to validating the clinical benefit of targeting unique molecular entities expressed by the podocyte. The hope and vision is that this will then pave the way to an expanded study of targeting the podocyte in DN.All authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.ML is employed by AstraZeneca. JP's research is supported by AstraZeneca.


Fibroblasts are the primary mesenchyme cell types that provide structural support during organ development and growth, and are the primary depositors of connective tissue matrix in response to injuries such as those occurring during skin scarring, tissue/organ fibrosis, systemic sclerosis, abdominal adhesions, just to name a few. Major efforts to study fibroblastic characteristics have centered on identifying surface markers that allow fibroblast purifications from tissues/organs.The glycoprotein CD90 is a widely expressed mesenchymal cell surface marker that is the subject of almost 1,000 publications (PubMed). It is present on mesenchymal stem cells  gene. The two fibroblast lineages either do or do not have a past history of En1 expression, and are referred to as “En1-Past” Fibroblasts (EPFs) and “En1-Naive” Fibroblasts (ENFs). Using genetic lineage tracing and live imaging tools, we demonstrated that EPFs are the primary contributors to the scar phenotype in the back in all injury models tested, including cutaneous wounding and irradiation-induced tissue fibrosis, whereas ENFs do not participate or contribute to scarring (Rinkevich et al., Second, CD90 does not discriminate between functionally unique fibroblast subtypes. Fibroblasts are functionally heterogeneous, both between and within anatomic skin locations. For example, the oral cavity skin quickly seals open wounds with minimal scarring, while back-skin responds to injuries with an opaque plug of dense connective tissue. Fibroblasts taken from various skin depths also diverge functionally (Sorrell and Caplan, Thirdly, by large scale screening of surface markers we found that CD90 does not mark a single defined skin fibroblast subtype, and this was recently confirmed by single-cell transcriptome analysis, in both human and mouse (Rinkevich et al., The functional role of CD90 on the surface of fibroblasts is not well understood, with just a handful of intimations. CD90 on cancer-associated fibroblasts has been shown to contribute to inflammation by promoting fibroblast release of IL-6, which promotes tumor progression (Shiga et al., + fibroblasts near the portal vein, suggesting these cells inhibit collagen degradation and promote accumulation of extracellular matrix (Katsumata et al., + stromal component in hepatocellular carcinoma (Sukowati et al., The relationship of CD90 to fibrosis is indeed far from clear. The loss of CD90 from lung fibroblasts is observed in idiopathic pulmonary fibrosis (Sanders et al., Defining fibroblast subsets based on their specific function could pave the way for clinical applications, and defining surface markers that mark specific classes of fibroblasts is a first step toward their cell enrichment. For examples, we have learned from previous studies that upper dermis fibroblasts promote the hair follicle regeneration, whereas lower dermis fibroblasts participate in wound healing (Driskell et al., We anticipate that fibroblasts exhibit a panoply of functionally distinct cell types, rivaling that of the hematopoietic system – different subsets carry different physiological functions and may display distinguishing surface markers. While the marker definition for hematopoietic subsets has been well established, the identification of markers for the various fibroblast subsets has just begun. The accumulating single cell transcriptome data on stromal cells from various tissues/organs including skin, may shed light on the unique markers for fibroblast subsets, and would pave the way toward cell enrichment strategies for clinical use.All authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.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.