Recent years have witnessed significant research interest surrounding the interaction among exercise, appetite and energy balance, which has important implications for health. The majority of exercise and appetite regulation studies have been conducted in males. Consequently, opportunities to examine sex-based differences have been limited, but represent an interesting avenue of inquiry considering postulations that men experience greater weight loss after exercise interventions than women. This article reviews the scientific literature relating to the acute and chronic effects of exercise on appetite control in men and women. The consensus of evidence demonstrates that appetite, appetite-regulatory hormone and energy intake responses to acute exercise do not differ between the sexes, and there is little evidence indicating compensatory changes occur after acute exercise in either sex. Limited evidence suggests women respond to the initiation of exercise training with more robust compensatory alterations in appetite-regulatory hormones than men, but whether this translates to long-term differences is unknown. Current exercise training investigations do not support sex-based differences in appetite or objectively assessed energy intake, and increasing exercise energy expenditure elicits at most a partial energy intake compensation in both sexes. Future well-controlled acute and chronic exercise studies directly comparing men and women are required to expand this evidence base. Obesity is a major risk factor for several chronic diseases, including type 2 diabetes mellitus and cardiovascular disease, and remains a significant global burden from a public health and economic standpoint ,2. WeighSimilar to many scientific fields, the majority of exercise and appetite regulation studies have traditionally focused research efforts on men. Consequently, much less is known about the interaction between exercise and appetite in women, and the opportunity to examine potential sex-based differences has been limited. A handful of exercise training studies have demonstrated that men experience greater reductions in body mass and body fat than women ,6,7, altExercise-induced changes in hormones implicated in appetite control and energy balance , glucagon-like peptide-1 (GLP-1), insulin, and leptin) may contribute to sex-based differences in body fat loss after exercise . AlthougThe purpose of this article is to review recent developments regarding appetite, appetite-regulatory hormone and energy intake responses to single bouts of exercise (acute responses) and exercise training (chronic responses) in men and women. Furthermore, this review will consider the potential implications of these findings for health and highlight important areas for future research.Appetite and energy intake are regulated at the physiological level by the neuroendocrine system, which involves complex interactions between central and peripheral mediated pathways ,17. AppeOf the short-acting appetite regulatory signals, ghrelin is unique as the only known gut peptide that is orexigenic, and is predominantly secreted into the circulation by the oxyntic glands of the stomach. Ghrelin exists in the circulation in two forms (acylated and unacylated) and, although only 10%–20% of circulating ghrelin is acylated ghrelin, this form is believed to be solely responsible for appetite stimulation .1–36 and PYY3–36), with PYY3–36 representing the most abundant and biologically active form. Concentrations of PYY are low in the fasted state and increase rapidly after meal intake, which highlights a potential role in meal termination and sensations of fullness between meals. Glucagon-like peptide-1 is also secreted from the intestinal L-cells in response to nutrient intake and similarly contributes to meal termination and satiety. It exists as an active (GLP7–36) and inactive (GLP9–37) form, with the active form rapidly degraded to its inactive form upon secretion into the circulation. The appetite-inhibiting effect of these hormones is further supported by studies demonstrating that peripheral administration of PYY3–36 . The. The8]. Scientific interest in potential sex-based differences in appetite regulation stems from initial evidence suggesting that men experience greater body mass and body fat reductions after exercise training than women ,6. FurthThe findings provided within the current literature from experiments focussing on weight loss can be useful in informing individuals about exercise and dietary approaches for health. In this regard, it has been established that exercise-induced energy deficits stimulate smaller changes in appetite, appetite-regulatory hormones and energy intake compared with dietary restriction in both men ,96 and wConsidering that exercise training appears to elicit at most a partial, but incomplete, compensation in energy intake in both sexes, it seems that men and women can endure prolonged periods in an exercise-induced energy deficit which may facilitate the development of a negative energy balance. Such incomplete compensation also supports evidence that larger exercise-induced energy deficits promote greater weight loss during an exercise intervention ,35. HoweAlthough a considerable body of literature has developed understanding of the relationship between exercise, appetite and weight control, there are only a few studies which have directly focused on sex-based differences. Additional research is required to expand the evidence base before definitive conclusions can be drawn. This should include different types of exercise and insights into the mechanisms governing appetite control, both of which appear sparse in the current literature. Studies investigating a wider array of appetite parameters, particularly appetite hormones beyond the initiation of exercise training, between men and women would also be welcomed. Furthermore, considering potential sex-based differences in non-homeostatic factors governing energy balance (e.g., neuronal responses , and cogThis review has demonstrated that appetite, appetite-regulatory hormone and energy intake responses to acute exercise-induced energy deficits are similar between men and women. Specifically, the consensus of evidence suggests that acute exercise transiently suppresses appetite, and does not stimulate compensatory changes in appetite, appetite-regulatory hormones or energy intake in the hours after acute exercise in either sex. Evidence derived from exercise training studies appear less conclusive, with limited evidence that women, but not men, respond to the initiation of exercise training with compensatory changes in appetite-regulatory hormones conducive to appetite stimulation. However, it is not known whether this change translates into long-term differences after a more sustained period of exercise training. Furthermore, evidence does not support a sex dimorphism in appetite or energy intake when assessed objectively, and increasing energy expenditure through exercise elicits at most a partial energy compensation in both sexes. Few studies have directly compared appetite, appetite-regulatory hormone and energy intake responses to acute and chronic exercise interventions between men and women. Therefore, these conclusions are supported by evidence drawn from the limited studies directly comparing the sexes and supplemented by those conducted in men and women separately. A better understanding of whether appetite, appetite-regulatory hormone and energy intake responses to exercise-induced energy deficits differ by sex may contribute to the development of more effective weight management strategies.


Early Childhood Education and Care (ECEC) settings have a pivotal role in shaping children’s dietary food habits by providing the contextual environment within which they develop these behaviours. This study examines systematic reviews for (1) the effectiveness of interventions to promote healthy eating in children aged 2–5 years attending centre-based childcare; (2) intervention characteristics which are associated with promoting healthy eating and; (3) recommendations for child-health policies and practices. An Umbrella review of systematic reviews was undertaken using a standardized search strategy in ten databases. Twelve systematic reviews were examined using validated critical appraisal and data extraction tools. Children’s dietary food intake and food choices were significantly influenced. Interventions to prevent obesity did not significantly change children’s anthropometric measures or had mixed results. Evidence was more convincing if interventions were multi-component, addressed physical activity and diet, targeted individual-level and environmental-level determinants and engaged parents. Positive outcomes were mostly facilitated by researchers/external experts and these results were not replicated when implemented in centres by ECEC providers without this support. The translation of expert-led interventions into practice warrants further exploration of implementation drivers and barriers. Based on the evidence reviewed, recommendations are made to inform child-health directed practices and policies. Good nutrition in early childhood is essential to ensure children reach their growth and developmental potential . FurtherConsiderable public health effort and research have been directed towards nutrition interventions in the home and the school setting ,14. HoweAlthough the home is still the primary influence ,22, centUmbrella reviews are becoming relatively common ,32 as a The primary aims of this Umbrella review are to examine previously published systematic reviews to determine (1) the effectiveness of interventions to promote healthy eating in children aged 2–5 years attending centre-based childcare; (2) intervention characteristics which are associated with successfully promoting healthy eating in pre-schoolers; and (3) recommendations for child-health directed policies and practices.To identify possible systematic reviews the online bibliographic databases Medline, Emcare , PsycINFO , Embase , CINAHL , Health Technology Assessment Database, ERIC, Scopus, Web of Science Core Collection, Joanna Briggs Institute (JBI) Evidence-Based Practice Database of Systematic Reviews and Cochrane Database of Systematic Reviews were searched for reviews published between January 2000 and September 2017. The search strategy is available as The following reviews were excluded: (1) studies with infants or studies where the children were attending compulsory schooling usually six years or older; (2) studies treating children for obesity or a clinical related condition; (3) studies using school, the home or settings which are not registered childcare; (4) studies in which dietary behaviour or dietary-related outcomes were secondary outcomes and not separately reported; (5) studies focused on low-income countries. Although the search strategy did not limit studies to particular countries, only systematic reviews relating to high-income countries as defined by the OECD (2017) were included because the childcare arrangements and practices are similar. An a priori protocol for the Umbrella review was registered with PROSPERO (CRD42017078749). To assess the methodological quality of the reviews and to determine the extent to which reviews had addressed the possibility of bias in the design, conduct and analysis, the Johanna Briggs Institute (JBI) Critical Appraisal Checklist for Systematic Reviews and Research Syntheses was used . This vaTo guide the extraction and synthesis of data from the selected studies and minimize the risk of author bias, a standardized tool, the JBI Data Extraction Form for Systematic Reviews and Research Synthesis  was emplp < 0.0005).The study selection process is summarised in Twelve systematic reviews were included in the final review and the quality assessment ratings are tabulated in Most of the primary studies in all of the reviews were randomised control trials (RCT) or cluster-RCT followed by case-control trials or quasi-experimental studies . ConcernThere was considerable heterogeneity between primary studies which precluded pooling of the data and meta-analysis or any systematic reviews undertaking Grading of Recommendations Assessment, Development and Evaluation (GRADE) . HeterogStudy findings favoured dietary effectiveness in most of the included reviews . AssesseSeven reviews focused on obesity-prevention and obesogenic behaviours (including diet-related behaviours). Despite reporting significant effects on BMI and other measures of adiposity for some primary studies, review authors concluded overall that diet-related interventions did not have a consistently positive impact . Two other reviews reported no significant changes in weight status . Ling, Robbins et al. (2016) and Zhou, Emerson et al. (2014) reported that the primary studies which significantly affected weight outcomes were multi-component interventions which addressed both dietary and physical activity behaviours. Mikkelsen, Husby et al. (2014) reported that single component interventions did not have a significant effect on children’s fruit and vegetable intake but five of six multi-component interventions did. Actively involved and engaged parents were also associated with consistently positive impacts on children’s weight status .More positive outcomes were seen in reviews assessing interventions directed at both environmental- and individual-level determinants of healthy eating behaviours. Most effective were multi-level interventions targeting environmental-level determinants including implementation support ,55,58,61Three reviews reported that the strongest effects came from interventions targeting environmental-level determinants. Bell and Golley (2015) examined 13 primary studies, with 12 reporting significant improvement in the food provided in centres (through food policy and changes in educators’ practices), the nutritional quality of menus and parental food provision of lunchboxes. Primary studies on interventions focusing on environmental-level factors reported positive outcomes including food and nutrition policies and the food environment, however few of these studies also reported on whether children’s dietary intake had changed as a result . Wolfendn = 15). Six primary studies attributed high parental engagement to the successful achievement of their primary outcome to effect changes in children’s weight .Half of the reviews reported an association between parental involvement and engagement, and achievement of objectives in ECEC interventions ,54,55,58Positive outcomes for healthy eating behaviours were mostly reported for interventions delivered by researchers or external experts ,52,55,56Eight reviews reported the number of included primary studies which used a theoretical framework ,57,58,61Most of the dietary-related interventions targeted educators’ behaviours and practices and included nutrition education and training sessions . EducatiEffective interventions involving children included interactive educational activities as part of the childcare curriculum ,54,57,58Active parental involvement included participation in any intervention component such as receiving written material, receiving regular newsletters, attending education sessions or workshops, completing homework tasks, participating in curriculum planning or participating in interactive hands-on activities such as cooking, growing vegetables or similar activities, with their children ,54,58,60Recommendations for practice and policy  includedSummarised recommendations for future research  includedThis Umbrella review investigated the effectiveness of interventions to promote healthy eating in children aged 2–5 years attending centre-based childcare. The aim was to also identify characteristics of successful interventions and list and summarise the most frequent recommendations for policy, practice and research. Overall, 12 systematic reviews of acceptable methodological quality were included examining 101 primary studies.Despite the considerable heterogeneity, the review findings supported the proposition that interventions to promote healthy eating in children aged 2–5 years attending centre-based childcare are effective. Successful interventions were multi-component, multi-level targeting both environmental and individual-level determinants of healthy eating behaviours. Multi-component interventions included educational strategies, changes to the centre-environment and policy. These findings are consistent with the conclusions of other Umbrella reviews for other settings ,66,67,68A common recommendation in the reviews was to underpin intervention design with theoretical frameworks and effective behavioural change theories, ideally components of Social Cognitive Theory  alongsidThe translation of changes in educators’ knowledge, practices and centre environment to children’s dietary behaviours was however not consistently observed ,54. MoreMore studies assessing the dietary-related outcomes from involving and engaging with parents are required. Even small levels of parental involvement were associated with better weight status outcomes ,53,54,55More research is needed to understand the interactions between educators and parents and the impact of collaborative parental engagement. With many children spending time in childcare and the premise that all food preferences are learnt, educators’ roles are crucial as very young children are dependent upon them not only to provide food but also to guide and shape their food preferences and dietary habits . QualitaThe impact of nutrition-related strategies to build the capacity of children is also an evidence gap. Findings emphasized the importance of targeting children with interactive education and hands-on experiences which are age-appropriate ,52. ThisThe impact of nutrition-related interventions and practices on children from low socio-economic areas is of particular interest. Many of the primary studies were directed at centres in low socio-economic areas or centres with a high proportion of children from disadvantaged families. The outcomes suggest that interventions supporting these populations could help reduce health inequalities . This obMissing from this Umbrella review was evidence of the sustainability of dietary-related interventions as few primary studies were implemented for more than a year and/or outcomes measured after the intervention. The recommended duration is at least one year, ideally 1–2 years ,58. NotaAlthough the more comprehensive the intervention, the more likely it is to be successful, comprehensiveness may affect feasibility and fidelity negatively and warrants further exploration. Ward, Welker et al. (2016) found an inverse relationship between comprehensiveness and positive outcomes. Furthermore, in an Umbrella review investigating community-based interventions promoting healthy eating and physical activity, multi-component interventions were not correlated with positive outcomes . MoreoveLastly, reviewers recommended that cost-effective studies be undertaken ,56,61. LBased on the data reported in the 12 reviews, reviewers cautioned that many primary studies were rated as weak or having insufficient information to permit evaluation. The actual effect of the intervention may therefore be smaller than the effects reported because of the low quality of reporting  and geneSome of the challenges identified by Pollock et al. (2017) and Ballard & Montgomery (2017) in their critique of the robustness of Umbrella reviews were encountered in this study. These challenges included primary studies overlapping between reviews and appearing in more than one review, and a mismatch between the scope of the systematic review being examined and the research question of the Umbrella review. Seven of the 12 reviews had a remit for obesity prevention rather than healthy eating as a primary outcome. Furthermore, the heterogeneity of the reviews and the assessment of insufficient information in three reviews precluded an evaluation of the quality of the research through the use of GRADE. This was compounded by the difficulty for the systematic reviews to apply GRADE or a meta-analysis for the same reason. To address these challenges, the primary studies which appeared in more than one review were identified and the extent of overlap considered. Moreover, in the data extraction stage, within these accepted reviews, only primary studies which met the scope of the Umbrella review were included, strengthening confidence in the findings. Mapping the overlapping primary studies reassured the authors that the search strategies were through and demonstrated consistency between reviewers. To further ensure methodological strength, the scope of the Umbrella review was limited to only those reviews where diet-related behaviours and measures were reported separately, PRISMA guided the search strategy and two validated tools were used to assess the quality and risk of bias of the reviews and to standardize data extraction . The conInterventions promoting healthy eating positively influence children’s dietary food patterns. Although environmental-level and individual-level determinants of healthy eating are impacted by centre-based interventions, these effects are not consistently translated to changes in children’s diet-related behaviours or anthropometrics as a measure of preventing obesity. Positive outcomes can be further strengthened with parental involvement and engagement, and multi-level, multi-component strategies are recommended. Comprehensiveness may, however, affected feasibility and fidelity negatively when enacted by end-users; therefore studies on existing interventions implemented by end-users are recommended. Meta-analysis and stronger study designs are called for but are often not feasible in the real world of childcare. Therefore the translation of research or expert-led interventions into practice warrants further qualitative exploration of implementation drivers and barriers with end-users. This understanding and end-user involvement may contribute to the sustainability of interventions which is rarely reported.The summarised findings and recommendations from this Umbrella review can inform child-health directed policies and practices. Based on the evidence, public health effort is warranted to support healthy eating interventions and practices in centre-based childcare. By incorporating multi-level and multi-component interventions into routine practices and extending this across the home and childcare setting, healthy food preferences and dietary-related behaviours can be influenced. More successful interventions require high levels of parental engagement, the use of behaviour change strategies and a focus on building the capacity of educators, children and parents.


Recent advances in the field of induced pluripotent stem cells (iPSCs) research have opened a new avenue for stem cell-based generation of vascular cells. Based on their growth and differentiation potential, human iPSCs constitute a well-characterized, generally unlimited cell source for the mass generation of lineage- and patient-specific vascular cells without any ethical concerns. Human iPSCs-derived vascular cells are perfectly suited for vascular disease modeling studies because patient-derived iPSCs possess the disease-causing mutation, which might be decisive for full expression of the disease phenotype. The application of vascular cells for autologous cell replacement therapy or vascular engineering derived from immune-compatible iPSCs possesses huge clinical potential, but the large-scale production of vascular-specific lineages for regenerative cell therapies depends on well-defined, highly reproducible culture and differentiation conditions. This review will focus on the different strategies to derive vascular cells from human iPSCs and their applications in regenerative therapy, disease modeling and drug discovery approaches. Induced pluripotent stem cells (iPSCs) are reprogrammed cells that have features similar to embryonic stem cells, such as self-regeneration without restriction and differentiation into different tissue or cell types, including vascular cells . ConnectArtificial reprogramming of somatic cells produces cells that maintain their great potential for medical use but research is associated with less ethical problems than embryonic stem cells resulting from the internal cell mass of blastocysts (embryoblasts) , 5. In pThe vascular system is essential for embryonic development and adult life and it is the first organ system to develop during vertebrate embryogenesis , 8. HereThe task of blood vessels involves transporting of all the nutrients as well as of their metabolites, e.g. of toxic waste. The goal is an optimal supply of all cells of the body and the blood circulation with vital substances –17. AnatVascular damage and dysfunction are implicated in the development of vascular diseases and cardiovascular pathological changes. Therefore, the therapeutically use of autologous cells for tissue regeneration is a ray of hope, which includes both local transplantation of the vascular cells to injured organs and the engineering of organs. Cell replacement therapy with vascular cells and vascular bypass grafts with autologous blood vessels or synthetic vascular grafts are the main treatments for certain cardiovascular diseases, such as coronary heart disease, aortic aneurysm, dissection, and peripheral vascular disease –23.The main limitations for the use of vascular cells for the treatment of cardiovascular diseases were the sources and amount of the cells. In general, their proliferative potential decreases with increasing donor age and intensive ex vivo expansion can limit their properties such as long term plasticity and result in a functional decline. In addition, adult cells may contain more DNA abnormalities  during the course of a lifetime –26. ThusThe unlimited proliferation potential of iPSCs and their capability to differentiation into virtually every cell type in the human body is of great significance to explore alternative cell sources capable of generating functional endothelial cells and SMC. Furthermore, the generation of structures to repair damaged endothelium for vascular regeneration as well as blood vessels en bloc were desired because endothelial cell regeneration is a slow and insufficient process , 32. TisBeside the abundant origins of iPSCs the potential to generate patient individualized vascular cells that bypass the immunogenicity and ethical issues are central advantages of using iPSCs as vascular cell source. However, a possible therapeutic use of pluripotent stem cells still holds medical risks, namely the potential to generate teratomas. Therefore, only donor cells that have reached a particular differentiation stage could be used, which means that the iPSCs must first be brought to an ordered differentiation path. Thus, a major obstacle for using human iPSCs for therapy or to model disease remains the lack of reliable, efficient and scalable protocols to differentiate functionally mature adult cell types.Based on progress in the research field, the present review aims to summarize the strategies and mechanisms of generating vascular cells through differentiating from human iPSCs, and to examine what this means for the potential application of cell therapy in the clinics.In mammalian development, vascular progenitors mainly emerge from the lateral and posterior mesoderm . Thus, vInduction of mesodermal differentiation can be achieved using conditions that promote self-aggregation of the iPSCs into three dimensional embryoid bodies (EB) or by the addition of mesoderm-inductive factors in chemically defined monolayer systems. The evolutionarily most ancient family members Nodal, Activin and BMP are members of the transforming growth factor β (TGFβ) superfamily of morphogens, which includes TGFβs, inhibins, bone morphogenetic proteins (BMPs), growth and differentiation factors (GDF), and others –36. CombNo matter which way of mesodermal induction is chosen, the different lineages of vascular cells emerge then from the proliferating and differentiating iPSC in the presence of growth factors, extracellular matrix molecules, growth factor inhibitors, small molecules, and neutralizing antibodies that can be used to promote enrichment of the endothelial cells or SMC or the lineage of choice. To identify the cell lineage of choice and to enrich the cells using some form of cell sorting and isolation (flow cytometry sorting or immunomagnetic separation), monoclonal antibodies to identify specific cell surface molecules or genetic tagging of the iPSC with lineage-specific fluorescent reporter systems are required. Subsequent additions of numerous growth factors (as described below) further promote vascular cell proliferation, differentiation and eventual maturation during the in vitro culture.Endothelial cells originate mainly from mesodermal progenitors . ShortlyVarious methods have already been disclosed to generate endothelial cells from human iPSCs , 49–52. Later on, more commonly endothelial lineage-committed cells could also be derived from EB formed by iPSCs. The first reported EB differentiation-based methods used high VEGF concentrations and resulted in less than 10% of CD31/PECAM1-positive cells after 2 weeks of culture –58. AlthNowadays, monolayer differentiation methods of feeder-free culture systems without the need of previous EB generation but with the combination of different culture substrates and chemical-defined conditions have been successfully applied to induce endothelial cells from iPSCs and were most commonly used. These improved methods resulted in an increase in differentiation fidelities, efficiencies and kinetics.l-glutamine, beta-mercaptoethanol, and human FGF2)  scaffolds in quiescent media (containing TGFβ1 and 0.5% FCS) and either incubated under static conditions or subjected to peristaltic flow in a bioreactor system for the remainder of the differentiation (day 30) [Obtaining vascular SMC with robust, mature, elastic fibers is a key obstacle in tissue-engineered blood vessels. Eoh et al., followed the Wanjare protocol to differentiate SMC from human iPSCs and placed these cells on scaffolds in a pulsatile flow bioreactor, resulting in vascular smooth muscle tissue with robust elastic fibers and enhanced functionality . Human i(day 30) . As SMC (day 30) .A straight forward approach concerning the clinical utility of iPSC-derived vascular cells was recently, reported by Lee et al. who enriched human iPSC-derived endothelial cells via a clinically compatible system . Herein,Clinical applications for vascular-related diseases (e.g. heart attack caused by atherosclerosis) commonly use the grafting of patients autologous arteries and veins, which are limited and often damaged due to the initial disease or advanced aging , 159. TiThe supportive action of an endothelial cell co-culture with the generated cells of interest for organogenesis was already reported , 172. TaAlthough non-immunogenic autologous endothelial cells as well as SMC isolated from patients themselves would be the first choice for vessel and/or vascularized tissue engineering, the numbers of cells obtained from small vessel biopsies remain limited and, moreover, these terminally differentiated cells bear a limited proliferation potential. Even the cells isolated from umbilical veins have limited proliferation potential , 174. ThThe integration of supporting vascular cells was further shown to improve maturation and functionality of engineered skeletal muscle tissue, finally resulting in a better engraftment upon in vivo implantation –170. CriThe therapeutic potential of human iPSCs is quite promising as they are patient-specific stem cells that could be derived from easily accessible sources of tissue—such as the donor’s skin or peripheral blood—and that do not face the immunological barrier or ethical concern which confront cells derived from human embryonic stem cells. With the appropriate differentiation protocols, iPSCs could then be used to generate vascular cells. Thus, the central advantages of iPSC-derived endothelial cells and SMC are their potential abundance by generating nearly unlimited numbers and their immune-privileged status as autologous tissue. Importantly, patient genomic information is maintained during the reprogramming and differentiation processes. Because iPSCs can be derived from subjects with genetic diseases, they represent an ideal vehicle to better understand a variety of diseases including heritable disorders, in particular the molecular mechanisms that dominate in vascular diseases, for screening of novel therapeutics and to foster vascular regeneration by cell replacement therapy or vascular grafts.Although iPSCs start a new era of regeneration medicine, the tumorigenesis risk might compromise their further clinical applications. In turn, for vascular regeneration already robust selection markers and refined experimental protocols have been established to guide human iPSCs reproducibly to a vascular lineage. Additional negative selection against remaining pluripotent cells could be an additional option, to limit the risk of teratoma formation. Concerning the patient-derived autologous iPSC-differentiated cells, the respective genetic background is a benefit for vascular disease modeling studies; but because the same genetic or acquired abnormalities that predisposed a patient to a particular disease will be persist in their iPSCs might result in dysfunctional vascular cells or iPSC and resulting vascular progenies with reduced differentiation capabilities.However, up to now, iPSC-derived vascular cells provide a reliable model system that enables basic vascular-biological research and molecular pharmacology studies with the potential to generate patient- specific disease models. In addition, the generation of clinically relevant numbers of vascular cells from human iPSC for use as a cell therapy in human subjects is already feasible.


During the past decade, studies of the mechanisms and functional implications of adult hippocampal neurogenesis (ahNG) have significantly progressed. At present, it is proposed that adult born neurons may contribute to a variety of hippocampal-related functions, including specific cognitive aspects and mood regulation. Several groups focussed on the factors that regulate proliferation and fate determination of adult neural stem/progenitor cells (NSC/NPC), including clinically relevant drugs. Opiates were the first drugs shown to negatively impact neurogenesis in the adult mammalian hippocampus. Since that initial report, a vast array of information has been collected on the effect of opiate drugs, by either modulating proliferation of stem/progenitor cells or interfering with differentiation, maturation and survival of adult born neurons. The goal of this review is to critically revise the present state of knowledge on the effect of opiate drugs on the different developmental stages of ahNG, as well as the possible underlying mechanisms. We will also highlight the potential impact of deregulated hippocampal neurogenesis on patients undergoing chronic opiate treatment. Finally, we will discuss the differences in the negative impact on ahNG among clinically relevant opiate drugs, an aspect that may be potentially taken into account to avoid long-term deregulation of neural plasticity and its associated functions in the clinical practice. Extensive experimental work demonstrated that new neurons can be generated in adult mammalian brain . One aduRecently a comprehensive review summarized data on how hippocampal function and related behavior may be modified by newborn neurons . At presA vast array of research investigated factors that regulate ahNG. External stimuli, including learning, environmental enrichment, and exercise have profound effects on proliferation/ differentiation of NPC as well as survival of their progeny . Aging, Extensive research efforts suggest that ahNG may be deregulated in several neuropsychiatric disorders, including major depressive disorder .An intringuing aspect of ahNG is its susceptibility to pharmacological modulation . AntidepEmerging evidence suggest that several psychoactive drugs result in molecular changes that may negatively affect different aspects of ahNG . These fOpiate drugs are powerful analgesics which are also among the most commonly abused addictive drugs. Clinical research suggested the occurrence of deficits in memory tasks, attention, verbal fluency and general cognitive performance in opiate addicts . ControvDepression often co-occurs with chronic pain and an emerging line of inquiry is also the association between opiate use and the risk of depression . In partAltogether several experimental data suggest that long-term opiate may produce maladaptive changes in brain structures involved in cognition and mood regulation such as the hippocampus. In morphine-dependent animals, mechanisms responsible for the disruption of long-term memory retention have been suggested, including effects on dendritic spine stability  and long+/GFAP-; neuroblasts which express doublecortin and the polysialylated form of neural cell adhesion molecule (PSA-NCAM) are referred to as Type-3 cells.Adult neurogenesis is a complex, multistage process which consists of a series of developmental events, namely proliferation, differentiation, migration, maturation and survival of NSC/NPC and their progeny. Based on their developmental stage, different cells participating in ahNG can be recognized and classified. Briefly, radial-glia-like stem cells, which are both glial fibrillary acidic protein (GFAP) and nestin positive are defined as Type-1 cells; Type-2 cells are neural progenitors which are highly proliferative and nestinOver the last few years, extensive progress has been made on how morphine can disrupt neurogenesis in the adult rodent hippocampus . The firin vivo, by largely reducing the number of 4-week-old BrdU-labeled cells in the dentate gyrus of drug-treated compared to control rats  and phosphorylated histone H3 (pHisH3), and these effects were antagonized by naloxone  reuptake . Since a progeny . In pres progeny . As a pr progeny . In linepocampus . AltogetCognitive dysfunction has been often reported in opiate drug abusers . Moreoveper se may induce profound changes in hippocampal plasticity, including deregulated hippocampal neurogenesis (Preclinical evidence summarized in this review support the idea that disruption of ahNG may, at least in part, contribute to the long-lasting effects produced by some opiates, like morphine, on cognition and mood regulation. Although preliminary, these preclinical data mark the importance of taking into account inhibition of ahNG as a potential long-term consequence of opiate use also in the clinical setting. This aspect appears even more relevant in consideration of the fact that chronic pain ogenesis .L-carnitine (While we wait for a definitive answer on any causal correlation between long term-opiate treatment and cognitive and emotional impairment in chronic pain patients, preclinical studies should be undertaken to increase our current understanding of the cellular and molecular effects of opiates on adult NPC and their progeny. Moreover, experimental work should be aimed at understanding differences between distinct opiates in their potential for disrupting ahNG. Ultimately these studies may help us to unravel different level of risk associated with distinct treatment options. In principle, drugs like methadone and tapentadol may result, after drug long-term treatment, in reduced or absent dysfunction in ahNG compared to morphine and other opiates. Alternatively, other analgesic drugs which do not reduce but rather increase ahNG in preclinical models like pregabalin/gabapentin  and acetarnitine  could bearnitine . We propVB and MG were involved in the concept, literature screening, and writing of the article, and approved it for publication.In the past, MG has received research grants from drug companies manufacturing analgesic drugs, including Angelini S.p.A, Grunenthal GmbH, Pfizer, Sigma Tau. The other 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.


During the last decade, novel immunotherapeutic strategies, in particular antibodies directed against immune checkpoint inhibitors, have revolutionized the treatment of different malignancies leading to an improved survival of patients. Identification of immune-related biomarkers for diagnosis, prognosis, monitoring of immune responses and selection of patients for specific cancer immunotherapies is urgently required and therefore areas of intensive research. Easily accessible samples in particular liquid biopsies (body fluids), such as blood, saliva or urine, are preferred for serial tumor biopsies.Although monitoring of immune and tumor responses prior, during and post immunotherapy has led to significant advances of patients’ outcome, valid and stable prognostic biomarkers are still missing. This might be due to the limited capacity of the technologies employed, reproducibility of results as well as assay stability and validation of results. Therefore solid approaches to assess immune regulation and modulation as well as to follow up the nature of the tumor in liquid biopsies are urgently required to discover valuable and relevant biomarkers including sample preparation, timing of the collection and the type of liquid samples. This article summarizes our knowledge of the well-known liquid material in a new context as liquid biopsy and focuses on collection and assay requirements for the analysis and the technical developments that allow the implementation of different high-throughput assays to detect alterations at the genetic and immunologic level, which could be used for monitoring treatment efficiency, acquired therapy resistance mechanisms and the prognostic value of the liquid biopsies. During the last years immunotherapies of cancer particularly immune checkpoint blockade inhibitors and adoptively transferred T cells have improved the outcome of tumor patients. Since the monitoring of tumor samples and immune responses prior and following these therapies are essential for the optimization of the mode of immunotherapy and for the selection of patients suitable for a specific immunotherapy the identification and systematic assessment of biomarkers has become an important issue for immunotherapy of cancer. In principle, liquid biopsies represent suitable samples not only for early detection, monitoring residual diseases, but also for monitoring immunotherapy responses and development of resistances, because multiple samples can be easily obtained prior, during and after immune and other therapies .Liquid biopsies as a source for tumor-derived information can be analyzed at the cellular, DNA, RNA, epigenetic, protein and metabolome levels. With the development and implementation of high-throughput technologies personalized targeted and/or tumor immunotherapies have become feasible. In the future, an individual antibody binding pattern of patients might be also used for the development of personalized immunotherapy and for the monitoring of immune responses. The ultimate goal to predict immunotherapeutic success is the identification of reliable prognostic and predictive biomarkers, which link immunity with patients’ outcome thereby allowing the selection of a group of patients, who will most likely benefit from a respective treatment regimen.The NIH/WHO international program on chemical safety has defined a biomarker as “any substance, structure, or process that can be measured in the body or its products and that can influence or predict the incidence of outcome or disease” .For monitoring immune cell responses with long term follow-up, body fluids, such as e.g. peripheral blood and serum/plasma, should be collected at different time points prior, during and after treatment until disease progression. In addition to conventional analysis of serum markers, the absolute lymphocyte count (ALC), subtypes of lymphocytes, composition of the immune cell repertoire, activity of different immune cell subpopulations, exosome and extracellular microvesicle (EV) profiles, expression of genes/microRNAs/proteins, cytokines, metabolites, chemokines, putative tumor-associated antigens (TAA), polymorphisms in Fc receptors, antibodies and the proliferative capacity of adoptively transferred immune cells could be also analyzed in primary tumors and/or body fluids Figure  3–9].–9.3–9].The integration of multiple high-throughput “omics” technologies including cancer genome sequencing studies that have catalogued the mutational landscape of human tumors , transcriptomics for analysis of peripheral blood mononuclear cells (PBMCs) and DNA/RNA analysis in circulating tumor cells (CTC) has helped to define biomarkers. Furthermore, different proteome-based technologies, such as the serologic evaluation of proteins and antibodies (PROTEOMEX), “bottom up” and “top down” proteomics, multi-parameter ELISAs, bead or chip based affinity proteomics and multispectral imaging have been implemented for diagnosis, immune monitoring, immune response assays and/or identification of novel (immuno) therapeutic targets. However, these different profiling methods have some advantages and disadvantages Table , which sEarly biomarker screening prior and during therapy could help to timely identify non-responders, and accordingly discontinue time- and cost-sensitive therapies in order to give patients the chance to switch to alternative treatment regimens. Such screening is mainly based on the identification of a biomarker signature rather than one particular biomarker, since a single biomarker cannot encompass all informations needed. A set of distinct biomarkers should be able to (i) discriminate between different tumor entities rather than merely between tumor patients and healthy individuals, (ii) discriminate different (early and advanced) tumor stages or subtypes and (iii) guide selection of efficient treatment options.Cell-free nucleic acids (cfNA) including DNA, RNA and Ago2-bound microRNAs (miRNAs), as well as CTC, exosomes and other EVs, protein panels, (auto)-antibodies, diverse metabolites, soluble mediators and changes in the frequency, composition and/or activity of (immune) cell populations are currently discussed as minimally-invasive biomarkers, which are easily accessible in blood samples from tumor patients to detect premalignant and early-stage cancers, to monitor response to treatment and development of therapy resistance –16.Recently, exosomes have gained interest to advance biomarker research, since these nanometer-scale vesicles are secreted by living cells and carry the full range of molecules, e.g. DNA, RNA, miRNAs, proteins, cytokines and chemokines from their originating cells . Due to therascreen EGF-R Plasma RGQ PCR Kit [Circulating cell-free and exosomal DNA and RNA in human blood is derived from different cellular sources, including blood and tumor cells. It is postulated that circulating tumor DNA (ctDNA) reflects the molecular changes occurring at the tumor site in real time . Indeed, PCR Kit  was succ PCR Kit  and can  PCR Kit . CtDNA a PCR Kit –26 and c PCR Kit . Due to  PCR Kit , 31. The PCR Kit . Analogo PCR Kit , 30.Circulating miRNAs in serum or plasma might be also used as biomarkers due to their high stability under storage, easy handling conditions and emerging expression signatures that are associated with cancer survival and therapy response . Plasma/Although more than 50 years ago CTCs in the periphery of solid tumor patients have been identified, CTCs are still under critical review concerning their usefulness as biomarkers in cancer patients . The bigSingle or panels of proteins  from serum that address soluble immune-regulatory molecules, such as cytokines, chemokines and co-stimulators , acute phighCD57+ lymphocytes regarding recurrence-free survival of patients with non-muscle-invasive bladder carcinoma after transurethral resection and intra-vesical IL-2 treatment [+) lymphocyte subsets in PBMCs are related to the increased survival of cancer patients already serving as biomarkers in myeloma [+ T cell cytokine responsiveness as measured by changes in the phosphorylation status of STAT molecules might serve as biomarker discriminating glioblastoma patients benefiting from a DC vaccine [Potentially of high importance are the absolute numbers and relative proportions of immune cell populations within the patients’ tumor and immune cell populations, in particular in blood, lymph nodes and the bone marrow. The significance of the immune cell composition, the frequency as well as the localization of immune cell subpopulations within the tumor quantified in the “Immunoscore”  has beenreatment . Other s myeloma . Functio vaccine .Several studies imply that cancers with higher mutation burdens, such as microsatellite instability associated with a greater likelihood of expressing neo-antigens, are most likely to respond to checkpoint blockade inhibitors . HoweverWhile the induction of immune-related adverse events (irAEs) is a serious concern of immune-checkpoint blockade inhibition observing an inflamed tumor environment as well as increased preexisting autoantibodies, parameters that might act as markers of immune responsiveness bear the potential as clinical biomarkers , 46. ThuSample preparation and generation is an important issue, since significant differences have been observed between distinct methods and consumables used for serum, plasma and immune cells obtained from PBMCs. In detail, the standby times until centrifugation (1 – 48 hrs), the blood collection method, nucleic acid preparation methods, exosome purification methods, methodical aspects for flow cytometry and the IgG purification from these samples could affect the analysis and result in reproducibility problems. Furthermore, if immune parameters are monitored, information about chemotherapy, anti-inflammatory drugs and hormone therapy is crucial, since these will most definitely affect the immune signatures. International consortia are currently establishing standard practices for blood collection and processing for circulating biomarkers  the standardization of assays and (ii) the harmonization of assays. The first requires the exact same assay with the same standard operating procedures (SOP) and well-trained personal is needed in all laboratories, which is hard in terms of costs and feasibility. The second, more accepted way of handling this issue is the harmonization that requires trained personal and standardized laboratory equipment, not the exact same assay kits, but positive quality controls that have to be implemented in every assay in all cooperating/participating laboratories and research centers . SuccessBiomarker development following a traditional stepwise process from cell culture via mouse models to patient's samples has helped, but also often failed in generating novel biomarkers with clinical relevance . To idenex vivo tumor micro tissue. A potential biomarker in this sense would have the capacity of a particular immune cell type to enter the tumor micro tissue or to change the effector function inside the tumor micro tissue upon a given immunotherapeutic intervention.To explore cell biology and drug efficacy, cell-based assays were employed, in which tumor cells were typically grown on two-dimensional plastic surfaces or as single cell suspensions. However, since cell biology is profoundly influenced by its microenvironment, cell-based assays should be rather performed using 3D cultures, since they reflect both extracellular matrix and cell-cell contacts, such as cell-matrix interactions. These techniques were successfully implemented to study tumor-stroma interactions in NSCLC or tumor immune cell infiltration , 52. Thiin vivo immunocompetent models over time. Thus, such model systems have the potential to lead to the identification of blood-borne biomarkers that predict patients’ responsiveness to immunotherapy.Very old, but still valid tools to study human diseases are experimental mouse models. Initially murine syngeneic mouse models or immune deficient human xenograft models were used to study new (immuno) therapeutic strategies to treat cancer. These models have major drawbacks since the tumors are either not of human origin or are human or even patient derived xenografts (PDX), but lack the immune system and the human stroma environment. In particular immunotherapies, such as checkpoint inhibitors like anti-PD1/anti-PD-L1 cannot be tested in immune deficient mice, since their intervention relies on the existing immune system. To circumvent these problems humanized mouse models have been established, in which the murine immune system is replaced by the human immune system using PBMCs or bone marrow stem cell transplantations . These hOther technologies are proteome-based, such as e.g. the Global Proteome Survey GPS that employs an oriented single chain antibody fragment immobilization approach with known antibodies and with context-independent motif specific (CIMS) antibodies that recognize peptides of hundreds of proteins allowing for a target discovery . One bigA very interesting additional family of analytes with the potency to serve as biomarkers in blood samples are metabolites  includinFurthermore, the use of auto-antibodies as biomarkers was exemplified in dissecting prostate cancer patients using a combination of different techniques such as protein arrays and Luminex that are spiked with auto-antigens (4012 or 3061). These results were validated on tissue microarrays (TMAs) and by qPCR of tissue samples. A different auto-antibody panel comprised of 165 antibodies (Ab) was found in serum of high when compared to low inflammation prostate cancer patients . Of noteIVD development in companies for biomarker identification is time consuming due to an inadequate understanding of the modes of action of therapeutic targets, inadequate preclinical models for discovery and validation in addition to the separation of the noise from real data and the cross-validation of a specific marker in an independent large cohort of patients. Another big challenge is the broad diversity in the patients’ cohort due to inter- and intra-heterogeneity of a particular disease –68. One In addition to blood as the most abundant liquid body fluid there are other body liquids to study biomarkers, such as ascites/pleural fluid, cerebrospinal fluid (CSF), bronchial alveolar lavage (BAL), urine and saliva/sputum Table . Among tCompared to blood saliva contains a similar variety of DNA, RNA, proteins, metabolites, and microbiota that can be compiled into a multiplex of cancer detection markers . Urine hThe most cost and effort effective way would certainly be to use standardized assays for biomarker analysis for a large number of patients. Since one faces a high patients’ cohort diversity, these standardized assays for a particular type of cancer disease might not be applicable. The profiling of blood samples might allow reducing the number of patients’ subgroups that require analyses of tissue biopsies , 78. Howin vitro data and in vivo preclinical mice data show re-expression of cancer testis antigens, improved anti-tumor immunity and better tumor control upon treatment with hypomethylating agents and/or histone deacetylation inhibitors. These data provide the rational for the use of hypomethylating agents and histone deacetylation inhibitors in melanoma patients. In the future, the methylation status should be investigated in liquid biopsies prior to individual therapy decision.Novel individual therapeutic options include the alteration of the epigenetic features of tumors, in particular histone modifications and DNA methylation. Interestingly, the methylation status of melanoma patients does correlate with the overall survival . FurtherProperly controlled tissue acquisition and multiplex immunohistochemical analysis might overcome issues related to pre-analytical and analytical variability leading to a more reproducible assessment of clinically meaningful biomarkers. However, this technology still has to be improved and optimized due to the limited tissue material available in the clinical setting. The analysis of multiple immune parameters in tissue biopsies by multispectral fluorescent immunohistochemistry is an important advancement and has recently been successfully implemented in some clinical studies , 69. Thiin vivo measurement of many critical molecular features of tumor lesions and microenvironment, such as metabolism, proliferation, hypoxia, cell death and immune cell infiltrate, which can assist the knowledge of how cancer immunotherapy works and also facilitates clinical decision making [The success rate of cancer immunotherapy is difficult to predict, as its efficacy often depends not only upon characteristics of the tumor lesions, but also of the tumor microenvironment involving immune cells and soluble mediators. Molecular and functional imaging with Positron Emission Tomography (PET) and Magnetic Resonance Imaging (MRI) allows repeated non-invasive n making .18F-FLT and 18F-FDG-PET were successfully used to study the kinetics and involvement of lymphocyte subsets in response to vaccination. These tracers incorporate in highly proliferative cells and are therefore not specific to cell types, but when locally applied e.g. in the tumor draining lymph node in conjunction with a DC vaccine, they reflect lymphocyte proliferation to vaccination. This technique allows an early discrimination between responding and non-responding patients in anti-cancer vaccination protocols and aids physicians in individualized decision-making processes [More conventional PET imaging tracers such as rocesses .+ and CD4+ T cells (use of cys-diabody and minibody) and CD11b+ myeloid cells (use of variable domain of a camelid heavy-chain antibody) are successfully used in preclinical mouse models and currently being developed for advanced PET imaging in clinical monitoring applications to cancer immunotherapy [New imaging tracers specifically targeting subsets of immune cells by the use of radiolabeled fragmented specific antibodies to CD8otherapy –87.One of the major challenges remains to establish the biological connection of peripheral biomarkers and the complex tumor immune contexture. There is still a debate of whether peripheral immune cells are representative for the anti-tumor immunity observed in the tumor microenvironment (TME). Particularly, the link between peripheral blood immune cells and the prognostic and predictive relevance of the spatial distribution of immune cells in the tumor tissue is still unclear . This isThe cellular sources of blood not only include the “classical” immune cells, but also CTCs of diverse tumor entities, hematopoietic stem cells, erythrocytes and platelets. In hemostasis, platelets have been demonstrated to be more present than effector cells and represent the major inflammatory cells mediating inflammasome signaling thereby affecting both innate and adaptive immune responses . This leRecently, the use of antibodies directed against different checkpoint targets, such as anti-PD-1 , anti-PD-L1  and anti-CTLA4 (Ipilimumab) has been successfully implemented for the treatment of a number of different tumor entities . For ant+ T cells with proliferative capacity [Alternatively to PD-L1 expression on tumor cells as a biomarker, one study found a significant correlation of high tumor PD-L1 expression on immune cells infiltrating the tumor as a predictive marker for therapy success . The succapacity , 87. Othin situ analysis of the primary site of interest as well as (ii) the spatial resolution of the TME known as the tumor landscape. Disadvantages of tissue biopsies are (i) the invasiveness of the method, (ii) lack of willingness of patients for the necessary surgery, (iii) serial assessment and of high importance, (iv) the tumor heterogeneity (intra and primary versus metastases). The advantage of blood samples are (i) minimal invasive, (ii) serial assessment, (iii) live monitoring and (iv) systemic view that may account of tumor heterogeneity (v) the speed and reduced cost of sample preparation and (vi) the easy access of blood material from healthy volunteers or patients’ with different diseases for comparison. The disadvantages on the other hand are (i) no spatial resolution of tumor microenvironment (ii) lack of knowledge whether the blood reflects the “true” host tumor interaction and (iii) uncertainty whether the blood sample overcomes the tumor heterogeneity issue  the e Figure . HoweverMore broadly, a comprehensive and integrated bio-sampling approach of tissues and PBMCs in the clinical setting and preclinical models may allow the identification of less invasive biomarkers that reflect clinically meaningful aspects of the immune tumor microenvironment, improving the triage and management of patients in cancer immunotherapy. Overall, new tools for tissue analysis, rigorous validation and standardization of methods will help to better understand the dynamic nature of immune-tumor interaction.The clinical implementation of blood-derived biomarkers to predict tumor progression, response to therapy, therapy resistance or even early diagnosis of tumor occurrence is certainly wanted and urgently required. Recent progress is promising and the identification of the “right” biomarker/biomarker panel to define different tumor entities, if possible at distinct disease stages, is emerging, but needs to overcome current challenges of sample preparation, standardization of techniques, and acceptance in clinical practice. Even though, prospective studies that validate the liquid biopsy data in independent patient cohorts are crucial and are still missing in most cases, the so far obtained biomarkers from retrospective studies using liquid biopsies are very encouraging Despite these advances there is still an urgent need of blood analysis as a potential surrogate for tissues samples as summarized in Figure


Aging has long been considered as the main risk factor for several neurodegenerative disorders including a large group of diseases known as tauopathies. Even though neurofibrillary tangles (NFTs) have been examined as the main histopathological hallmark, they do not seem to play a role as the toxic entities leading to disease. Recent studies suggest that an intermediate form of tau, prior to NFT formation, the tau oligomer, is the true toxic species. However, the mechanisms by which tau oligomers trigger neurodegeneration remain unknown. This review summarizes recent findings regarding the role of tau oligomers in disease, including release from cells, propagation from affected to unaffected brain regions, uptake into cells, and toxicity via mitochondrial dysfunction. A greater understanding of tauopathies may lead to future advancements in regards to prevention and treatment. Neurodegenerative diseases are a leading cause of death and disability affecting millions of elderly. As life expectancies rise, growing numbers of elderly become affected. This emerges as a major global issue as there are currently no adequate remedies for these diseases. A large group of these diseases, which are related to tau, a microtubule-associated protein, are known as tauopathies. Most notably, tauopathies include Alzheimer's disease (AD), progressive supranuclear palsy (PSP), Pick's disease, frontotemporal dementia (FTD), corticobasal degeneration, and variants of Parkinson's disease (PD) and Lewy body dementia , implying that tau dysfunction alone can cause neurodegeneration  cell internalization of soluble, uncoated tau via receptor-mediated endocytosis  does not desensitize the muscarinic receptors present on neurons of the hippocampus; hence, a repeat stimulus via tau increases intracellular calcium every time, thus altering intracellular calcium homeostasis and the following hyper-phosphorylation and misfolding of tau. Coupled with the fact that tau persists in the extracellular environment for a longer time than acetylcholine, a neurotoxic effect may occur. In other words, it is sensible to theorize that tauopathies progress via interaction of extracellular tau with M1 and M3 receptors on neurons leading to cytotoxic effects . APFs are similar to pore-forming protein toxins in that their properties lead to membrane disruption. A recent study showed the existence of tau APFs in human brain samples from patients with PSP and LBD as well as in mice brain samples which overexpressed mutated tau. The study discovered that APFs form after tau oligomer formation and bypass higher NFT aggregate formation. The findings showed that APF formation relies on mutations in tau, phosphorylation levels, and cell type (Lasagna-Reeves et al., Oligomeric tau intermediates decrease cell viability (Flach et al., Recently, data has shown that injected tau oligomers co-localize with the mitochondrial marker porin, suggesting a pathological relationship. In fact, tau oligomers might disrupt microtubule stability and trafficking, thus affecting organelle distribution. Mitochondria navigate long distances to provide for synaptic energy demand; therefore, inhibiting transport systems impairs energy production routes (Lasagna-Reeves et al., Also, data shows low levels of complex I in brain hemispheres injected with tau oligomers when compared to brains injected with monomers or fibrils. This implies that alterations of complex I subunit mRNA, minimization in protein levels of complex I subunits (Kim et al., Mitochondrial damage can lead to activation of the apoptotic pathway. Hemispheres injected with tau oligomers were found to have increased levels of caspase-9 activation (Lasagna-Reeves et al., Overall, evidence suggests that tau oligomers are the toxic entities in tau aggregation and that alteration of the mitochondrial membrane, minimization in complex I levels, and activation of the apoptotic-related caspase-9 can cause toxicity, impeding synaptic energy production (Lasagna-Reeves et al., Discovering the pathological role of tau oligomers within the brain along with related mechanisms of cellular tau oligomer secretion, propagation, and uptake will allow for a better understanding of tauopathies (Castillo-Carranza et al., SS, MG, and DC wrote the manuscript. DC and MG reviewed intellectual content. DC prepared the figures. SS, MG, DC, approved the final version of the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Progress in management has improved hospital mortality of patients admitted to the intensive care units, but also the prevalence of those patients needing weaning from prolonged mechanical ventilation, and of ventilator assisted individuals. The result is a number of difficult clinical and organizational problems for patients, caregivers and health services, as well as high human and financial resources consumption, despite poor long-term outcomes. An effort should be made to improve the management of these patients. This narrative review summarizes the main concepts in this field.There is great variability in terminology and definitions of prolonged mechanical ventilation.There have been several recent developments in the field of prolonged weaning: ventilatory strategies, use of protocols, early mobilisation and physiotherapy, specialised weaning units.There are few published data on discharge home rates, need of home mechanical ventilation, or long-term survival of these patients.Whether artificial nutritional support improves the outcome for these chronic critically ill patients, is unclear and controversial how these data are reported on the optimal time of initiation of parenteral vs enteral nutrition.There is no consensus on time of tracheostomy or decannulation. Despite several individualized, non-comparative and non-validated decannulation protocols exist, universally accepted protocols are lacking as well as randomised controlled trials on this critical issue. End of life decisions should result from appropriate communication among professionals, patients and surrogates and national legislations should give clear indications.Present medical training of clinicians and locations like traditional intensive care units do not appear enough to face the dramatic problems posed by these patients. The solutions cannot be reserved to professionals but must involve also families and all other stakeholders. Large multicentric, multinational studies on several aspects of management are needed. The increasing worldwide life expectancy results in high prevalence of patients suffering from chronic diseases and related “chronic critical illness” , 2. Up tThis article is a narrative review of randomized controlled trials (RCTs), observational studies, systematic reviews, and meta-analyses published between 1990 and 2017 in English, in PubMed, and Scopus databases using the keywords: Cronic Critical Illness, Noninvasive mechanical ventilation, Mechanical ventilation, Physiotherapy and ICU, Tracheostomy, Ventilator Assisted Individuals, Weaning mechanical ventilation, Weaning Unit. In the following pages the terms “Prolonged Weaning” and “PMV” will be used with the same meaning.A consensus-derived definition of patients suffering from chronic critical illness, included patients with one of six eligible clinical conditions: PMV, tracheostomy, stroke, traumatic brain injury, sepsis or severe wounds, and at least 8 days of ICU length of stay , 10. SevDiaphragm weakness is highly prevalent in critically ill patients. It may exist prior to ICU admission and may induce the need for MV but it also frequently develops during the ICU stay. Several risk factors for diaphragm weakness have been identified, among them sepsis and length and modalities of MV play central roles. Critical illness-associated diaphragm weakness is consistently associated with poor outcomes including increased ICU mortality, difficult weaning and PMV , 12.A long hospital stay and lack of response to or an inadequate level of appropriate therapy can lead to muscle wasting and weakness, deconditioning, recurrent symptoms and mood alterations . There aNational Association for Medical Direction of Respiratory Care (NAMDRC) [(NAMDRC) : “the neEuropean Respiratory Society (ERS) Task Force [sk Force : “the nesk Force , signifisk Force –22.Weaning according to New Definition (WIND) study [D) study : “succesD) study .There is great variability in terminology and definitions :NationalA population-based cohort study  in an ICVentilatory strategies;Weaning protocols;Early mobilisation and Physiotherapy,Specialised weaning units.There have been recent progresses in weaning from MV, among which:Progressive reduction in the level of assistance of Pressure Support Ventilation (PSV);Progressive longer periods of SBT through the tube;Syncronized Intermittent Mandatory Ventilation (SIMV: the patient can breath spontaneously between ventilator-delivered breaths).Neurally Adjusted Ventilatory Assist (NAVA).Noninvasive mechanical ventilation (NIV).High-flow oxygen (HFO).The most popular ventilatory strategies used to shorten and achieve a more succesful weaning from MV in the ICU are :ProgressIn the Nineties multicentric comparative studies in the ICUs gave conflicting results, reporting advantages with either PSV , or SBT Neurally Adjusted Ventilatory Assist is a mode recently introduced in the clinical use in which the ventilator applies an inspiratory positive pressure in proportion to the electrical activity of the diaphragm, the best available indicator of the neural drive to breathe . This mo:Should be used to facilitate weaning from MV in patients with hypercapnic ARF only in centres with adequate experience using NIV in this setting .Should not be used in the treatment of patients with established post-extubation ARF.Should be used to prevent post-extubation ARF in high-risk patients but not used to prevent post-extubation ARF in non-high-risk patients.Nava et al.  were theMore recently, the use of HFO compared with conventional oxygen therapy reduced the risk of re-intubation within 72 h in extubated patients at low risk for re-intubation . Among hA prospective multicenter RCT  in threeA study in PMV patients , confirmA prospective study included chronically critically ill patients admitted to Spanish respiratory care units . The weaTrials have demonstrated that application of protocols or guidelines for the weaning process may lead to a decrease in weaning time independent of the mode used, even better than automatic systems , 47. TheIn patients needing PMV a well defined protocol, independent of the modality used, was associated with a better outcome than uncontrolled clinical practice previously performed in the LWUs . A multiOverall, approximately 25% of PMV patients in the ICU develop generalized and persistent muscle weakness: approximately one million patients develop the ICU-acquired weakness syndrome  annually . Muscle Evidence of benefits from early mobilization and physiotherapy has progressed during the past 15 years with RCTs, systematic reviews , 54, andIneffective cough and secretion retention can play a significant role in weaning failure. Evaluation of cough strength by means of assessment of cough peak expiratory flow rate can predict extubation failure and may reduce ICU length of stay, expenditures, morbidity and mortality . Cough aRespiratory intermediate Intensive Care Units (RIICUs) within acute care hospitals  are lessAlternatively patients needing PMV may be transferred from acute care hospitals to specialised regional LWUs, often located within rehabilitation hospitals. These dedicated LWUs relieve pressure on ICU beds at lower costs, with specialized teams . Variable mortality and weaning success rates have been reported –66.More recently modalities of tele-monitoring have been proposed as a means to manage difficult-to-wean patients and VAIs .To take care of difficult-to-wean patients, recently the problem of appropriate ICUs utilisation has been faced by proposing different locations and modalities of assistance :RespiratThere are few published data on discharge home rates, use of NIV, or long-term survival in specialized LWUs. An early study  evaluateMalnutrition in critically ill patients is associated with poor outcomes, including impaired wound healing, higher rates of nosocomial infections, and all-cause mortality . NutritiWhether artificial nutritional support improves the outcome for critically ill patients is unclear . The admUse of tracheostomy seems to increase in patients needing PMV  and is nDecannulation is the final step of liberation from PMV. Despite several individualized, non-comparative and non-validated decannulation protocols exist, universally accepted protocols are lacking as well as RCTs on this critical issue. Presence of an intact sensorium, coordinated swallowing and protective coughing are often the minimum requirements for a successful decannulation . Also a In a prospective survey  in patieThese patients may require ethical end of life decisions such as withholding or withdrawing MV, appropriate symptoms management and adequate location outside the acute care hospital. Duration of weaning attempts and ICU length of stay should be defined on the basis of potential expected benefits: otherwise the lack of these elements may lead to a condition of treatment “oriented towards futility”. Appropriate palliative care for these patients has been associated with improved quality of life and reduction in intensive life-sustaining treatments  with impThere is a lack of wide scientific investigation in the field of prolonged mechanical ventilation. Future research should involve a shared definition of this condition for appropriate recruitment of patients. Large, multicentric, multinational RCT should be promoted and supported. We need studies on pathobiology and pathophysiology of these patients as distinct from acute critical illness patients. We also need well designed clinical RCT of different protocols of management on issues such as setting of MV, patient-ventilator interfaces, time of decannulation, nutritional support, sedation, drug therapy of symptoms and delirium, physical, psychological and cognitive long-term effects of prolonged mechanical ventilation. Costs/benefits of different facilities for care of these patients should be also evaluated in the frame of different health care systems Table .Table 3AThe clear side of progresses in management of ICU patients is improvement of hospital survival. The dark side is the increase in the number of difficult-to wean patients and related VAIs. Present medical training of clinicians and locations like ICU do not appear enough to face the dramatic problems  posed by these patients. Therefore the solutions cannot be reseved to professionals but must involve also families and all other stakeholders. We hope that this narrative review may contribute to offer patients, their families and associations, their caregivers and all interested stakeholders the occasion to sensitize governments and health services for the best management of these patients.


Leukocyte telomere length (TL) has been suggested as a marker of biological age in healthy individuals, but can also reflect inherited and acquired hematopoietic dysfunctions or indicate an increased turnover of the hematopoietic stem and progenitor cell compartment. In addition, TL is able to predict the response rate of tyrosine kinase inhibitor therapy in chronic myeloid leukemia (CML), indicates clinical outcomes in chronic lymphocytic leukemia (CLL), and can be used as screening tool for genetic sequencing of selected genes in patients with inherited bone marrow failure syndromes (BMFS). In tumor cells and clonal hematopoietic disorders, telomeres are continuously stabilized by reactivation of telomerase, which can selectively be targeted by telomerase-specific therapy. The use of the telomerase inhibitor Imetelstat in patients with essential thrombocythmia or myelofibrosis as well as the use of dendritic cell-based telomerase vaccination in AML patients with complete remissions are promising examples for anti-telomerase targeted strategies in hematologic malignancies. In contrast, the elevation in telomerase levels through treatment with androgens has become an exciting clinical intervention for patients with BMFS. Here, we review recent developments, which highlight the impact of telomeres and telomerase targeted therapies in hematologic dysfunctions. Telomeres consist of hundreds to thousands of repeated double strand hexanucleotides (in humans: (TTAGGG)n), a single-stranded overhang, as well as specific proteins that coat them (telosome or shelterin protein complex) ,2. TelomIn vertebrate cells, each cell division in vitro and in vivo is associated with the loss of 50–200 base pairs (bp) of telomeric TTAGGG repeats due to the “end-replication” problem . When ceTelomere shortening can be counteracted by telomerase, the telomere extending enzyme, which is embedded into the shelterin protein complex . TelomerHowever, hTERT expression in health and disease is a very delicate balance, and individuals with short telomeres due to failure in telomere maintenance mechanisms (telomeropathies) are prone to genomic instability and have an increased risk for organ failure. Telomeropathies, such as dyskeratosis congenita, can manifest in the BM , liver, and the lung, and their link to short telomeres can either be related to inherited genetic mutations or simply to an acquired pathophysiology of the disease. Androgenic anabolic steroids, which have been used to improve blood counts of patients with BMFS since the 1960s, concurrently slow telomere shortening or even sustain telomere elongation in leukocytes most likely by an elevation in telomerase expression levels in the underlying HSPC compartment . These aIn general, TL of peripheral blood cells decreases with age and is subject to considerable, genetically determined variation at any given age. When measured by quantitative fluorescence in situ and flow cytometry (flow-FISH)-based methodologies , which hBCR-ABL oncogene [Chronic myeloid leukemia (CML) is a clonal, myeloproliferative disorder that originates in the hematopoietic stem and progenitor compartment (HSPC) and that leads to a marked expansion of myeloid cells . CML orioncogene . In CML,oncogene . Leukocyoncogene . This leoncogene . These foncogene .During tyrosine kinase inhibitor (TKI) treatment, patients with Imatinib treatment duration > 144 days showed significantly longer telomeres in comparison to patients who recently started TKI treatment up to day 144 of treatment. Median telomere length in major remission was found to be significantly longer compared to patients without response to treatment measured either by cytogenetics, interphase FISH or quantitative RT-PCR . In concIn addition, it has been found, that in CML, TL analysis could indicate the severity of the disease and thus may provide important information for treatment decisions: when TL of newly diagnosed CML patients is evaluated before treatment, patients with the longest telomeres show a lower clinical risk profile in comparison to patients with shorter telomeres and may predict response to initial TKI therapy after 12 and 18 months according to ELN criteria . These dTelomerase activity is upregulated in CP-CML as compared to HSPCs from normal individuals and increases up to 50-fold in blastic phase CML. Telomerase activity therefore correlates with acute versus chronic forms of CML and disease progression ,59. In ap < 0.001) [HV) [HV unmutated cells have shorter telomeres than HV mutated cells, and longer telomeres are associated with increasing VH mutational load. TL therefore appears to be a very good surrogate marker for the IGHV mutational status and CLL patients with long telomeres and mutated IGVH had a significantly better prognosis than patients with short telomeres and unmutated IGVH [Chronic lymphocytic leukemia is characterized by the clonal expansion of abnormal lymphocytes (B cells) in the peripheral blood, which is in part dependent on several genetic and epigenetic alterations, B-cell receptor signaling, and the interaction between CLL cells and the tissue microenvironment ,61. TL m< 0.001) . In addi01) [VH) . VH unmuted IGVH . Furtherted IGVH , and patted IGVH . In manyted IGVH .hTERT transcript contains alternate splicing sites, and some mRNA variants of hTERT prevent the expression of a catalytically active (full length) telomerase with the possible result, that telomeric ends are not longer maintained [Telomerase expression and activity have both been investigated as biomarker for disease progression in CLL ,63. The intained . In a stintained  Quantifyintained . Whereasintained , the secintained .Myelodysplastic syndromes (MDS) are a heterogenous group of clonal diseases in HSPCs, which result in ineffective hematopoiesis and subsequent cytopenia and dysplasia . Patientn = 53), and for overall survival, there was no significant difference between high and low telomerase activity according to the log rank method. However, patients with high telomerase activity showed a significantly shorter interval for transformation to acute leukemia (p = 0.010), suggesting high telomerase activity as a poor prognostic marker in MDS.In a recent study , no signBone marrow failure syndromes (BMSF) are a heterogenous set of potentially fatal diseases that can develop at any stage of life. They are characterized by peripheral cytopenia in at least two of the three blood lineages and marrow hypoplasia. A subpopulation of patients with BMFS are typically characterized by ultra-short telomere length  due to altered telomere maintenance mechanism (telomeropathy) . TelomerIn contrast to the constitutional bone marrow failure observed in DKC patients, in acquired aplastic anemia (AA), HSPCs are destroyed by the patient’s own immune system . In ordeEventually, telomeres become critically short in cells with malignant potential. In the absence of mechanisms such as p53 signaling of DNA damage or activation of alternative modes of telomere maintenance, cells with critically short telomeres can continue to replicate and expand on a clonal level. Therefore, all patients with telomeropathies are prone to genomic instability and have an increased risk for cancer . For exaBecause of an inability to completely replicate their telomeres, chromosomes progressively shorten during each replication cycle. Telomere shortening is counteracted by the enzyme telomerase, which adds telomeric nucleotide sequences to the end of the chromosomes . TelomerThe catalytic subunit of human telomerase is a single copy gene that maps to chromosome 5p15.33, remarkably close to the telomere at the extreme terminus of chromosome 5p . TelomerJAK2) gene, the gene encoding the thrombopoietin receptor (MPL), and the calreticulin (CALR) gene [With the reactivation of hTERT expression, cancer cells have developed a basic difference compared to their precursors, which makes them one possible key to selectively target cancer cells by pharmacological approaches. Telomerase specific therapy is a very promising area of research and very effective when used in combination with other cancer therapies . By reduLR) gene ,50,51,52LR) gene .JAK2 mutation and showed a molecular response by reduction in JAK2 V617F mutant alleles. Furthermore, patients with calreticulin (CALR) and MPL mutations showed a reduced mutant allele burden, thereby providing evidence of an anticlonal activity of Imetelstat [Two phase II clinical trials reported a clinical response in MPN patients treated with the telomerase inhibitor GRN163L (Imetelstat) ,55. Imetetelstat . Similaretelstat . Howeveretelstat .Imetelstat, immunotherapy tries to exploit the high expression of telomerase in cancer cells. Spontaneous, antileukemic, cytotoxic T cells against telomerase-derived antigens have been shown to occur naturally in B-cell chronic lymphocytic leukemia [In addition to leukemia  giving aleukemia . AST-VACleukemia . The vacleukemia ,92. Thirleukemia . Telomerleukemia .n = 10), TERC (n = 7), DKC (n = 3) and RTEL (n = 1). The primary endpoint of the study was a 20% reduction in the rate of telomere attrition at 24 months. The study was stopped early after the first twelve patients all met the primary endpoint. All 27 patients were then assessed and unexpectedly almost all patients had a gain in telomere length with a mean increase of 386 base pairs [Shortened telomeres are a characteristic BMSF and can be due to heterozygous hTERT mutations, which compromise telomerase function by haploinsufficiency . Androgese pairs . Hematolse pairs . Althougse pairs , have nose pairs  provide With the confirmation of TL as an independent prognostic factor for overall survival in a recent meta-analysis, the concept of TL measurements in CLL has been proven to provide clinically relevant information. However, due the lack of standardization and the variability of methods to measure TL, the outcome of TL on other advanced myeloid cancers is not clear. Here, agreements are necessary that define the method of choice for measurements and its standardization.With the proof-of-concept data of Imetelstat in patients with ET and myelofibrosis, and the use of androgenic hormones to increase telomerase expression in patients with BMFS, telomerase targeted therapies provide important clinical interventions for hematological dysfunctions. In both cases, further randomized clinical studies are necessary to assess drug benefit-risks, determine the most effective dosing schedules and to clarify the mechanism of action.


Chronic eosinophilic inflammation is associated with tissue remodeling and fibrosis in a number of chronic T-helper 2 (Th2)-mediated diseases including eosinophilic esophagitis (EoE) and asthma. Chronic inflammation results in dysregulated tissue healing, leading to fibrosis and end organ dysfunction, manifesting clinically as irreversible airway obstruction in asthma and as esophageal rigidity, strictures, narrowing, dysmotility, dysphagia, and food impactions in EoE. Current therapies for EoE and asthma center on reducing inflammation-driven tissue remodeling and fibrosis with corticosteroids, coupled with symptomatic control and allergen avoidance. Additional control of Th2 inflammation can be achieved in select asthma patients with biologic therapies such as anti-IL-5 and anti-IL-13 antibodies, which have also been trialed in EoE. Recent molecular analysis suggests an emerging role for structural cell dysfunction, either inherited or acquired, in the pathogenesis and progression of EoE and asthma tissue remodeling. In addition, new data suggest that inflammation-independent end organ rigidity can alter structural cell function. Herein, we review emerging data and concepts for the pathogenesis of tissue remodeling and fibrosis primarily in EoE and relevant pathogenetic parallels in asthma, focusing additionally on emerging disease-specific therapies and the ability of these therapies to reduce tissue remodeling in subsets of patients. Allergic inflammation has the capacity to recruit eosinophils to the site of inciting stimulus. Prolonged eosinophil infiltration can contribute to significant tissue injury, leading to maladaptive tissue remodeling and fibrosis. We will focus primarily on eosinophilic disorders associated with robust tissue remodeling, specifically eosinophilic esophagitis (EoE) and its relevant pathogenetic parallels in asthma.The hypereosinophilic syndrome (HES)-associated tissue remodeling is arguably the most severe with cardiac damage leading to potential morbidity due to endomyocardial fibrosis. Asthma-associated airway remodeling occurs with epithelial denudation and goblet cell metaplasia, subepithelial fibrosis, angiogenesis, and smooth muscle hypertrophy . RemodelEosinophilic esophagitis is defined as a marked esophageal eosinophilic inflammation (≥15 eosinophils per high power field) that includes other inflammatory cells that likely contribute to remodeling such as mast cells, basophils, and adaptive as well as innate lymphoid cells –15. In tIn EoE, endoscopic features of remodeling vary between age groups. In children, features of esophageal pallor and furrows associate with histologic fibrosis and clinical dysphagia . In contAsthmatic airways demonstrate subepithelial fibrosis, with increased trichrome staining. The asthmatic epithelium demonstrates defective epithelial barrier function and loss of junctional proteins, with goblet cell metaplasia , 37. AirCurrent concepts of tissue remodeling have centralized on cellular and extracellular matrix responses to repetitive tissue injury and ineffective tissue regeneration in the context of chronic inflammation , 21, 45.via eotaxin-3/CCL26 and IL-5, further augmenting Th2 inflammation in the esophagus that can result in irreversible stricture formation  are elevated in EoE tissue biopsies , 91 and There is accumulating evidence that mechanical signals  alter the function of structural cells in the airway and esophagus in a manner that can be independent of, dependent on, or synergistic with, inflammation –95. Our Currently, there are no FDA-approved drugs indicated for the treatment of EoE. Some of the current treatment strategies and their effects on airway and esophageal remodeling are summarized below , 101.Topical esophageal corticosteroids constitute the most commonly utilized EoE therapy in children and adults. Similarly, inhaled corticosteroids are the most common agent used for persistent asthma. There has been relatively rapid accumulation of data for EoE since biopsies are procured regularly as part of disease monitoring. In contrast, airway biopsy is done in the context of clinical trials. Short-term studies in children have demonstrated that topical corticosteroids decrease fibrosis, VCAM-1, epithelial remodeling, subepithelial TGFβ1, and nuclear Smad2/3-positive cells in the subset of patients who have resolution of epithelial eosinophils following therapy . As suchSince EoE is a chronic disease, chronic therapy seems warranted. Studying a group of 32 children over a mean of 5 years (maximum of 10 years) treated with corticosteroids, Rajan and colleagues showed that children with EoE who persistently respond well to therapy have significantly less fibrosis and lower endoscopic scores than children who respond suboptimally to therapy , 111. InIn asthma, the effects of inhaled corticosteroids on remodeling and the best remodeling endpoint to follow are not entirely clear . This isEfficacy of topical corticosteroid therapy is dependent on mucosal drug delivery and esophageal mucosal contact time . SwallowIn children, dietary modification to remove allergen-derived antigenic stimulation has been shown to reverse subepithelial fibrosis in EoE , 128. InAnti-IL-5 blockade with mepolizumab is safe and achieves significant reduction in circulating peripheral eosinophils and inflamed tissue eosinophilia –131. EveAnti-IL-13 monoclonal antibody QAX576 significantly reduces esophageal eosinophilia and expression of EoE-related genes up to 6 months after treatment, but demonstrates only a trend for improved clinical symptoms . IL-13 bBoth EoE and asthma are diseases that involve robust tissue remodeling as part of the disease processes with resultant end organ dysfunction in a subset of subjects. In asthma, the clinical complication is irreversible airway obstruction. In EoE, it is stricture formation. One mechanism to this complication is prolonged, unbridled inflammation that can occur due to lack of therapeutic intervention or the failure of therapies to adequately control disease progression. The presumed inflammatory signals are from infiltrating cells that respond to alterations in structural cell physiology such as decreased barrier function and the onset of chemokine production. However, other signals such as mechanical changes in the airways due to airway rigidity and epithelial contraction during repeated rounds of bronchoconstriction drive structural cells such as epithelium to generate inflammatory signals that could propagate inflammation and be unresponsive to standard anti-inflammatory therapies such as corticosteroids.In addition to these issues, a number of additional considerations should be made when assessing the Th2-associated remodeling. The first issue is what parameters reflect remodeling most reliably? The second is the issue of pathogenic versus physiologic remodeling. The use of physiologic markers such as esophageal strictures or fixed airway obstruction likely represents an endgame of chronic disease and will likely be difficult to control. For this reason, one goal should be to find early markers of remodeling and control them. Of course, remodeling is also a normal process of wound healing that is necessary and required. What is not clear is how the shift from physiologic to pathogenic remodeling occurs. Possible explanations include disease duration, chronic inflammation, and/or mechanical signals such as tissue rigidity. Understanding the molecular mechanisms and the clinical phenotypes of these processes will be essential to better control allergic tissue remodeling and its consequences.QN and SA reviewed the literature and wrote the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.