Severe and recurrent infections of the respiratory tract in early childhood constitute major risk factors for the development of bronchial hyper-responsiveness and obstructive respiratory diseases in later life. In the first years of life, the vast majority of respiratory tract infections (RTI) leading to wheezing and asthma are of a viral origin and severity and recurrence are the consequence of a greater exposure to infectious agents in a period when the immune system is still relatively immature. Therefore, boosting the efficiency of the host immune response against viral infections seems to be a rational preventative approach. In the last decades it has been demonstrated that living in farm environments, i.e. early-life exposure to microbes, may reduce the risk of allergic and infectious disorders, increasing the immune response efficacy. These findings have suggested that treatment with bacterial lysates could promote a nonspecific immunomodulation useful in the prevention of recurrent RTIs and of wheezing inception and persistence. Experimental and clinical studies showing the reduction of RTI frequency and severity in childhood and elucidating the involved mechanisms can support this hypothesis. Respiratory tract infections (RTI) are amongst the leading causes of childhood morbidity and mortality globally , 2. ViruStructural and functional changes induced by respiratory viral infections in the airways during early childhood may promote self-perpetuating inflammatory mechanisms and, in predisposed individuals, a selection towards a Th2-type response that can lead to wheezing and asthma. Highly sensitive and specific tests, such as polymerase chain reaction, have been developed in the past few decades, allowing accurate detection of viruses associated with acute and recurrent wheezing illness , 18, 19.The increased susceptibility of infants and young children to respiratory infections is the result of the physiological immaturity of components of the systemic and local immune responses and, possibly, of sub-optimal complex inter-talk between microbiota and immune system effectors , 13, 14.Early-life gut microbiota development has proven to be a critical factor in protecting against infections –52. ReceSeveral bacterial lysates available for clinical use have shown to provide clinical benefit in patients with recurrent RTIs . We willThree OBL, administered by oral route, have shown to be effective in the prevention of viral infections in children: LW50020, RU 41740 and OM-85. The product that has been more extensively evaluated in recent, good quality studies is OM-85 , 60. StuDCs are major players in both innate and adaptive responses against viruses. Studies performed in vitro and ex vivo in experimental animals demonstrate that OBL can induce DC maturation, increasing the production of anti-viral cytokines/chemokines and the expression of surface molecules involved in antigen presentation , 60. In In human and murine blood monocyte cultures and in murine bone marrow-derived macrophage cultures, OBL increased cytokine  release, adhesion molecule  expression, oxygen radicals  production and killing of TNF-sensitive targets , 68–71. In addition to providing a physical and functional barrier, bronchial epithelial cells (BEC) are actively involved in the initiation of host reactions to viral infection, through the expression of various receptors and effector proteins on their surface and the release of anti-inflammatory agents , 80. ExpKlebsiella pneumoniae or Streptococcus pneumoniae [The possible “anti-viral” activities of OBL, shown by the in vitro studies, were confirmed in a mouse model of influenza virus infection Fig. a. In anieumoniae . Pre-treeumoniae . In addieumoniae . As compeumoniae . In conteumoniae . Their leumoniae . In a sieumoniae . In a dieumoniae . Furthereumoniae . In summeumoniae  and of Heumoniae , 81 as weumoniae , 83, 84.Several reviews and meta-analyses have shown that OBL can be effective in prevention of recurrent RTI of the upper and lower airways in pediatric populations , 85–89. OBLs appear to act on both the innate and the adaptive arms of the immune response, conferring efficacious broad immunoglobulin-related and cell-mediated immunity to the respiratory system. Additionally, experiments performed on BECs show that OBLs can also directly reduce viral infections, regardless of the engagement of the inflammatory and immuno-effector cells. Mechanistic data on the mode of action and results of clinical studies in pediatric patients can sustain the rationale for use of OBLs in the prophylaxis of viral RTIs and associated wheezing/asthma exacerbations. In addition to improving the quality of life of young patients and their families and to limiting the economic and social impact of these frequent respiratory disorders, the reduction of the frequency and severity of RTI is expected to decrease inappropriate use of antibiotics. Reduced prescription of antimicrobial drugs will have positive effects on the preservation of the physiologic gut microbiota composition and diversity, and importantly on reducing development of drug resistance. Ongoing (e.g. ORBEX) and future studies will show whether early OBL treatment can also be effective in prevention of long-term respiratory consequences of early and severe or recurrent RTI in childhood and will contribute to clarify the effect of OBLs both on immune system effectors and on microbiota and its metabolites.


Pharmacology can be defined as the study of the effects of chemical substances on the function of living organisms. Thus, pharmacology is essentially a translational discipline, bridging together biomedical and chemical sciences with the clinical practice. Therefore, the formation and growing of Pharmacological Societies around the world has been, and will continue to be, key in the development of integrated basic, experimental, and clinical sciences.This Research Topic aimed to celebrate the Chilean Society of Pharmacology (SOFARCHI) 40th anniversary by providing a diverse collection of in-depth reviews and original articles coming from members of our Society and colleagues around the world. SOFARCHI is a non-profit scientific society whose main objective is the promotion of research in pharmacology, from theoretical to experimental, and to clinical points of view. SOFARCHI was founded in 1978 with about 60 members who carried out teaching and research in pharmacology mainly in the central  and in southern zone  of Chile. Since 1978, SOFARCHI has held 41 annual congresses and currently has more than 140 members. In addition, SOFARCHI encourages the development and dissemination of pharmacology through promoting research among its members, sponsoring undergraduate and graduate courses, and fostering the creation of teaching books on different pharmacological topics. The Society also actively aids the Institute of Public Health, an agency of the Chilean state, in the technical evaluation of medicines that require sanitary registration. This highlights the commitment of SOFARCHI to promote the proper use of medicines in Chile, making known the main therapeutic and adverse effects of drugs to our fellow citizens. Lastly, the Society has developed a strong outreach program engaging our scientific work to high school students along the country.Over the last 10 years SOFARCHI has experienced important growth. The annual congress, our main yearly scientific activity, brings together more than 250 attendees, where about 70% of them are undergraduate and graduate students. Thus, SOFARCHI as a young society is always poised to contribute to Chile’s scientific and technological development, but also ready to collaborate with other partners in Latin America, and the world. Therefore, we believe that this research topic brings together many of the main lines of research carried out by Chilean pharmacologists. We are grateful to the Frontiers in Pharmacology Editorial team and many international experts from the field of pharmacology who have been able to support the research topic with their expertise, serving as editors and reviewers.We are glad that our topic generated an important global impact, reaching more than 50,000 article views and more than 12,000 downloads. The articles published in our research topic include reviews, mini-reviews, perspectives, brief research reports and original research articles. Our topic is grateful to the 240 authors that made their scientific contributions by submitting work of excellence to the Topic.Escobar et al.), on the chemical properties of amphetamine analogs with activity as monoamine oxidase inhibitors , on the contribution and relevance of P2X receptors in Alzheimer disease and in the toxicity of soluble Aβ peptide oligomers  and on novel strategies and perspectives for the treatment of dental pulp-derived pain . This group of contributions also included a detailed clinically-oriented review provided by Castillo et al., which summarizes the clinical experience of Chilean pharmacologists who have worked for more than 10 years with dexmedetomidine, a highly selective α2-adrenergic agonist with sedative and analgesic properties, with minimal respiratory effects. On the other hand, the Topic incorporated self-sustained minireviews summarizing and discussing the relevance of benzofurans as potential chemical scaffolds against Alzheimer disease , the effects of vitamin C on cancer cells and its therapeutic potential in cancer , novel drugs targeting pentameric ligand-gated ion channels as potential novel analgesics , the neurobiological basis of obsessive compulsive disorder (OCD) with focus on glutamate transporter 3 (EAAT3) , recent advances on therapies against non-infectious uveitis  and the beneficial effects of the mifepristone in metabolic syndromes .We would like to first highlight the broad thematic spectrum of the reviews and mini-reviews published in the issue, covering from molecular and cellular to pre-clinical and clinical pharmacology. This group of articles includes in-depth reviews on the contribution of the kappa opioid system to dopamine-related compulsive behaviors  significantly reduced the ethanol consumption of alcohol-preferring rats. Finally, and related to human hypertension, Gonzalez et al., elucidated a biochemical pathway which could underlie the antihypertensive effect of dietary potassium (K+) observed in human studies. Interestingly, many of the blood pressure lowering actions of K+ were seen in both hypertensive and normotensive rats, supporting a beneficial effect of dietary potassium in health and disease. Similarly, Vio et al., showed that a chronic high potassium diet down regulates key components of intratubular renin-angiotensin system. This down regulation could be a novel mechanism involved in the potassium-induced natriuresis that is beneficial for blood pressure and cardiovascular health. In the same field, Mondaca-Ruff et al., demonstrated a relationship between autophagy, the AT1R/RhoA/Rho Kinase-dependent pathway and Ang II-induced hypertrophy of vascular smooth muscle cells. Last, but not least, Figueroa et al., demonstrated that the pro-inflammatory phase of unilateral ureteral obstruction, a model of Chronic Kidney Disease, involves tubular damage with cortical upregulation of juxtaglomerular renin and medullary (pro)renin receptor downregulation. This scenario suggests a differential iRAS modulation as part of the mechanisms involved in the early stages of chronic kidney disease.Other original contributions to our topic covered relevant issues with impact on human health, such as chronic pain, cancer, addiction, hypertension, and chronic kidney disease. Regarding neural mechanisms underlying chronic pain, De La Fuente-Ortega et al., demonstrated that prenatal ethanol exposure affects iron homeostasis of specific brain areas , suggesting a critical role of iron homeostasis in maladaptive cognition observed in fetal alcohol spectrum disorder. In addition, Benavides-Rivas et al., described the role of enzymatic glutamate synthesis in the neurulation process using Xenopus oocytes. Finally, original research article of Sáez-Briones et al., studied the novel compound 2-Br-4,5-MDMA, a MDMA  analog brominated at C(2) of the aromatic ring. Interestingly, 2-Br-4,5-MDMA maintains the effects of MDMA as a substrate of the serotonin transporter (SERT). However, the bromination of MDMA modified the expected behavioral responses in rats, showing an absence of the classical MDMA-elicited behavioral responses including hyperlocomotion, enhanced active avoidance conditioning responses and increased social interaction.The thematic diversity of our topic also includes articles focused on developmental aspects and on medicinal chemistry. The original research contributed by In conclusion, we believe that the diversity of articles incorporated in our Research Topic reflects the integrative spirit of our scientific society and the translational character of the pharmacological sciences. In addition, we think that the new findings published show the growing quality of the Chilean science and highlight the rising contribution of our scientists to the global knowledge. We look forward to celebrate many years more of the SOFARCHI.All the authors have equally contributed to the work. The authors reviewed and approved the text for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Flavours and fragrances are volatile compounds of large interest for different applications. Due to their high tendency of evaporation and, in most cases, poor chemical stability, these compounds need to be encapsulated for handling and industrial processing. Encapsulation, indeed, resulted in being effective at overcoming the main concerns related to volatile compound manipulation, and several industrial products contain flavours and fragrances in an encapsulated form for the final usage of customers. Although several organic or inorganic materials have been investigated for the production of coated micro- or nanosystems intended for the encapsulation of fragrances and flavours, polymeric coating, leading to the formation of micro- or nanocapsules with a core-shell architecture, as well as a molecular inclusion complexation with cyclodextrins, are still the most used. The present review aims to summarise the recent literature about the encapsulation of fragrances and flavours into polymeric micro- or nanocapsules or inclusion complexes with cyclodextrins, with a focus on methods for micro/nanoencapsulation and applications in the different technological fields, including the textile, cosmetic, food and paper industries. Flavours and fragrances are a large class of compounds widely employed as additives in different technological fields, including food, cosmetics, textiles and others, mainly to ameliorate the olfactory and gustatory sensations of the product ,2. They To overcome these concerns, different encapsulation strategies have been applied, aiming to prevent the evaporation of volatile compounds and protect them from degradation . ThroughThe present review addresses the recent literature, mainly focusing on papers published between 2018 and 2020 related to the encapsulation of flavours and fragrances into polymeric capsules and inclusion complexes with CDs. Particular attention was devoted to the applications of these encapsulated systems in different technological fields, such as textiles, cosmetics, food and paper industries.Different methods have been reported in the literature for the encapsulation of flavours and fragrances in polymeric capsules . The choAlthough chemical methods for encapsulation are suitable for capsules formed by shells made of synthetic polymers, the so-defined physical-chemical and physical-mechanical methods can be employed both for natural and synthetic polymers. However, chemical methods are generally preferred for synthetic polymers, since, in most cases, they are more effective in controlling the size, the shape of the capsules and assuring a high loading capacity and encapsulation efficiency ,26. ThesCoacervation is a largely employed method since the 1950s for the micro- and nanoencapsulation of different compounds based on the physicochemical process of phase separation in which a polymeric dispersion can form a liquid polymer-rich phase, known as coacervate, at specific conditions ,32. TherPhase separation of a polymer from a colloidal dispersion can be also achieved using a vapour phase as the antisolvent, the so-called vapour-induced phase separation (VIPS). This technique has been widely employed for the preparation of films, membranes and hydrogels, but it has been recently proposed for the preparation of microcapsules. A complex mix of fragrances have been encapsulated in cellulose acetate microcapsules via the VIPS technique. The obtained capsules had a core-shell architecture, high encapsulation capacity and stability up to one year at room temperature, showing no fragrance diffusion without external stimuli at a dry state .Among physico-mechanical methods, the currently most employed for the encapsulation of flavours and fragrances is still spray-drying. It has been reported for flavour encapsulation that around 80–90% of the encapsulated products are obtained by spray-drying; then, by spray-chilling (5–10%), melt extrusion (2% to 3%) and melt injection (∼2%) .Specifically, spray-drying is one of the most common methods used for several reasons, such as equipment availability and simplicity, the possibility to use a wide variety of encapsulating agents, large-scale production, good efficiency and reduced processing costs ,42. On tSpray-drying has been extensively employed for the microencapsulation of EOs, using several wall materials, especially polysaccharides  or gums ,45. SpecSpray-chilling, also known as spray-cooling, spray-congealing or prilling, is another congener technique utilised for the microencapsulation of flavour compounds, especially when lipids are employed as wall materials ,50. Spraw/w of cinnamon oil and showed a continuous release of the major volatile components  for up to 28 days [Electrohydrodynamic processes such as electrospinning and electro-spraying can be also used for the encapsulation of flavours and fragrances , allowin 28 days . Recentl 28 days . Needlel 28 days  or the n 28 days . The obtMelt extrusion is another “traditional technique” employed in the past decades for the encapsulation of flavours and fragrances . It cons2 (sCO2) technologies have been employed to formulate particles or capsules with a wide variety of polymeric materials [2 can act either as a solvent, solute or antisolvent, giving rise to different techniques . CO2 methodologies are versatile and scalable, allowing a formulation process in a completely anhydrous medium, by obtaining noncontaminated products, high encapsulation efficiencies, customised particle properties and good scalability. The characteristics of the particles/capsules can be tuned by employing supercritical CO2 at different operating conditions  [2 as the solvent at a moderate pressure and temperature, was employed for the encapsulation of eucalyptol in polyethyleglycol/polycaprolactone microparticles and of Citrus aurantifolia EO in polyethylene glycol/lauric acid microparticles, demonstrating satisfactory entrapment efficiency and controlled release [Laurus nobilis EO) using the SFEE technique. A chemically modified food starch was used as the encapsulating agent. The efficiency of encapsulation by the SFEE process was found to be favoured by the increase in EO concentration and the final dried particles demonstrated to be effective in the protection of this highly volatile compound [Over the years, alternative novel methods have been investigated for the encapsulation of fragrances and flavours. Indeed, supercritical COaterials . In thesressure) . Recentl release ,77. sCO2compound .The formation of molecular inclusion complexes using CDs is a very common microencapsulation approach, widely investigated for different purposes ,80,81,82In the last years, the same research group has published several works reporting the encapsulation of molecular inclusion complexes of volatile compounds incorporated in polymeric fibres, mats or webs by electrospinning. A polymer-free electrospinning approach was applied on CD inclusion complexes to enhance the water solubility; improve the high temperature stability and control the release of carvacrol , tymol , camphorMicro- and nanocapsules/spheres, as well as molecular inclusion complexes with CDs, have been largely employed as protective carriers for aroma compounds  in different technological fields ,103. TheTextiles represent one of the most investigated applications for micro- and nanospheres/capsules encapsulating fragrances and aromas. These encapsulated volatile compounds have been employed for several years in textile-finishing processes, such as fabric conditioners to impart freshness and odour control ,106,107.The retention of fragrances and aromas, especially after washing or rubbing, depends on the penetration of microcapsules and nanocapsules into the spacing of textiles during the finishing process. To address this, in a work, a series of micro-/nanocapsules, with a size suitable for the pore spacing of cotton textiles and formed by citronella oil as the core material and chitosan as the wall material, was prepared through a microemulsion approach. These micro-/nanocapsules were applied on the textile through vacuum impregnation. The matching between the spacing of the pore sizes of cotton textiles and the sizes of micro-/nanocapsules was assessed via the retention of aromatic compounds in the finished cotton textiles after several washing cycles (washing durability). Indeed, the aromatic retention of cotton textiles finished by nanocapsules was much greater than the same textiles finished with microcapsules (28.84% vs. 1.55%) after 10 cycles of washing. The authors demonstrated that nanocapsules can penetrate better into the pores of the cotton textiles . To overOn the other side, the encapsulation of fragrances and aromas was pursued recently for the fabrication of “smart textiles” with additional functional properties , such asAmong the different classes of functional textiles, those with the most potential exploitation are the cosmetic textiles or cosmetotextiles. They are defined from the European Cosmetic Directive (76/768/EEC) Article l as “any textile product containing a substance or preparation that is released over time on different superficial parts of the human body, notably on human skin, and containing special functionalities such as cleansing, perfuming, changing appearance, protection, keeping in good condition or the correction of body odours” . In thesThe active ingredients, including fragrances and aromas, are generally encapsulated and released from the fabric to the skin upon the action of different triggering events, such as changes in the pH or temperature, sweating and rubbing . As for Botrytis cinerea and Alternaria alternata to improve the quality attributes and storage behaviours of peaches [Zingiber officinale EO were prepared using chitosan and carboxymethyl cellulose as shell materials to investigate the effects on the postharvest quality and prolonged the shelf-life of jujube fruits in terms of morphologic characteristics and some parameters as soluble solid contents, titratable acidity, the Red index and decay index and sensory quality [Another application in which the micro- and nanoencapsulation of fragrances and flavours research has been focused on is related to food . As for  peaches . Similar quality . The EO  quality .Active packaging represents a fashionable option to preserve the quality and prolong the shelf life of food products. It refers to packaging materials, which are not inert, and does not exert only a mechanical function of enclosing food, but they “actively” interact with the atmosphere inside the packaging or directly with food products ,155,156.The European Union (EU) Cosmetics Regulation defines a cosmetic product as “any substance or mixture used for external parts of the human body , teeth and mucous membranes of the oral cavity for cleaning them, perfuming them, changing their appearance and/or correcting body odors and/or protecting them or keeping them in good condition” . In the Fragrance ingredients are active ingredients commonly used in cosmetic products intended for the application to skin and hair with the purpose to release pleasant odours. In some cases, the products also labelled as “unscented” may contain fragrances to mask the unpleasant smell of other ingredients without giving a perceptible scent. The application of microencapsulation technology on the delivery of flavours and fragrances is a topic of relevant interest considering the need to improve the efficacy of a wide range of cosmetic (perfumes) and personal care  products . FragranDifferent examples can be found in the literature on the development of systems intended for the encapsulation of fragrances with cosmetic applications. Sansukcharearnpon et al. encapsulated six fragrances: camphor, citronellal, eucalyptol, limonene, menthol and 4-tert-butylcyclohexyl acetate using the solvent displacement method and different polymer blends of ethyl cellulose, hydroxypropyl methylcellulose and poly as polymeric carriers. The process gave a 40% fragrance loading capacity with an 80% of the encapsulation efficiency at the fragrance:polymer weight ratio of 1:1 . A more EOs and volatile compounds can be also encapsulated in CDs in order to improve their water solubility; avoid oxygen-, light- or heat-induced degradation and loss during processing and storage and to stabilise them against unwanted changes. Moreover, the use of CD–flavour inclusion complexes allows the use of very small amounts of flavours . 1-PhenyAnother application of the aroma and fragrances encapsulation is the design of aromatic paper or scented paper. Aromatic paper is intended to provide a pleasant surrounding atmosphere on the basis of the aromatherapy principles. In this regard, research has been focused on the development of wallpaper with the aim of providing comfortable sensations and to enhance the psychological and physical well-being ,213. SceLavender oil microcapsules were prepared with ABA-type triblock copolymer  and adsorbed onto the paper surface. The distribution of the microcapsules on the paper surface was homogeneous without degradation. The colour and gloss properties of the paper were also maintained in compliance with the standards . In anotRecently, encapsulated fragrances with an antibacterial effect were applied on paper. Specifically, vanillin was encapsulated in chitosan/poly(lactic-co-glycolic acid (PLGA) nanocapsules to prepare an aromatic wallpaper with an additional antibacterial action. Thanks to the presence of chitosan, the nanospheres showed an antibacterial effect against Gram positive and Gram negative, and the adhesion on the wallpaper was also enhanced . In anotFlavours and fragrances are compounds of great importance, widely employed in different products to improve the quality and ameliorate the satisfaction of the consumers. Encapsulation protects them from evaporation and chemical degradation, thereby controlling the release and allowing a simpler handling for processing. This strategy has enabled the use of flavours and fragrances for different technological applications, including the textiles, food, cosmetic and paper industries. Although research is still ongoing in this field, the encapsulation in core-shell polymeric nanoparticles, as well as the formation of molecular inclusion complexes between volatile compounds and CDs, are the most employed techniques in the experimental studies published in the last years. Both techniques resulted in being effective in encapsulating flavours, aroma compounds and EOs in a stable form suitable for different applications. Specifically, remarkable advances have been achieved for the encapsulation of these compounds or their molecular inclusion complexes in micro- or nanofibers/particles via electrodynamic processes. Among all technological fields in which core-shell-encapsulated flavours and fragrances find a relevant usage, the textiles and food packaging industries are the most investigated, despite other applications, such as paper production or coating, that can also benefit from the potential development of these micro- or nanosystems.


Ubiquitous exposure to endocrine-disrupting chemicals (EDCs) has caused serious concerns about the ability of these chemicals to affect neurodevelopment, among others. Since endocrine disruption (ED)-induced developmental neurotoxicity (DNT) is hardly covered by the chemical testing tools that are currently in regulatory use, the Horizon 2020 research and innovation action ENDpoiNTs has been launched to fill the scientific and methodological gaps related to the assessment of this type of chemical toxicity. The ENDpoiNTs project will generate new knowledge about ED-induced DNT and aims to develop and improve in vitro, in vivo, and in silico models pertaining to ED-linked DNT outcomes for chemical testing. This will be achieved by establishing correlative and causal links between known and novel neurodevelopmental endpoints and endocrine pathways through integration of molecular, cellular, and organismal data from in vitro and in vivo models. Based on this knowledge, the project aims to provide adverse outcome pathways (AOPs) for ED-induced DNT and to develop and integrate new testing tools with high relevance for human health into European and international regulatory frameworks. Worldwide, serious concern has arisen about human exposure to manufactured chemicals that can undoubtedly produce adverse health effects through disruption of the body’s endocrine system, known as endocrine-disrupting chemicals (EDCs) ,2. ThereA significant knowledge gap is how EDCs affect neurodevelopment, and endocrine disruption (ED)-induced developmental neurotoxicity (DNT) is hardly covered by the testing tools that are currently in regulatory use. The brain is among the most vulnerable organs with respect to toxic insults , in partYet, in the context of brain development, and hormonal involvement therein, there are considerable species differences ,18. For To address both the scientific and the methodological gaps, the Horizon 2020 research and innovation action ENDpoiNTs has been launched. ENDpoiNTs is part of the EURION cluster and includes 17 participants in Europe, USA, and Australia, integrating expertise in endocrine disruption and DNT and combines state-of-the-art in silico and in vitro tools, innovative experimental designs and technologies, and advanced biostatistics on human epidemiological and biomonitoring data. ENDpoiNTs will generate new knowledge about ED-induced DNT and aims to develop and improve in vitro, in vivo, and in silico models pertaining to ED-linked DNT outcomes for chemical testing . ENDpoiNThe project will also generate knowledge about species and sex differences and similarities for ED-induced DNT by integrating molecular, cellular, and organismal data from female and male in vitro (both human and rodent primary and/or pluripotent cell-based) and in vivo models. Furthermore, it will link these insights to human health by the use of appropriate test compounds in concentrations relevant for human exposure and by linking back from the experimental results to human epidemiology data.Based on this knowledge, the project aims to provide adverse outcome pathways (AOPs) for ED-induced DNT and to develop and integrate new testing tools with high relevance for human health into European and international regulatory frameworks.Widespread exposures to EDCs have been established by international biomonitoring programs e.g., HBM4EU) and show regional and temporal variations. Moreover, during a lifetime, humans are clearly exposed to multiple EDCs in complex mixtures from various routes and sources, and these mixtures include multiple chemical classes. Most importantly, exposure to EDCs, in particular during early life, has been clearly associated with a number of adverse health outcomes in EU and sh,9,10,11.The ENDpoiNTs project will include the establishment of human-relevant EDC exposure ranges from multiple international early life  cohorts and biomonitoring studies. The primary pregnancy cohort is the Swedish SELMA study, which includes over 2300 pregnant women and their children, with prenatal exposure to 54 environmental chemicals and neurodevelopmental effects monitored in children up to 7 years of age. Other pregnancy cohorts with neuro-related outcomes included in the project are the Dutch LINC and the Australian Barwon Infant Study (BIS) cohorts. Additionally, ENDpoiNTs will make use of publicly available EU-wide biomonitoring data through HBM4EU and the US CHEAR studies.The chemical compounds selected for testing within the project are based on previous results from the SELMA and the LINC cohorts in the H2020-funded EDC-MixRisk project and the FP7-funded DENAMIC project, respectively. It is a set of 27 known or suspected endocrine disruptors associated with human neurodevelopmental outcomes, such as cognition and behavior, in at least one of the projects, and comprises several pesticides, phthalates, bisphenols, perfluorinated compounds, and their key metabolites. The 27 parent compounds are listed in Both single EDCs and human-relevant mixtures of EDCs  will be tested.The endocrine signaling pathways frequently studied in the context of ED  play crucial roles in neurodevelopment and are therefore highly relevant for DNT-related endpoints . HoweverOne of the most thoroughly studied examples of hormonal programming of neurodevelopment is the role of sex steroids (estrogens and androgens) in the sexual differentiation of the brain. These hormones are known to be key drivers in the programming of sexual and socio-aggressive behaviors in males and females and in the development of sexually dimorphic, multifunctional brain structures such as the preoptic area, the ventromedial and arcuate nuclei of the hypothalamus, the bed nucleus of stria terminalis, and the amygdala ,23,24,25Thyroid hormone (TH) signaling is well-known for playing an essential role during brain development in all vertebrates ,28. In hThe glucocorticoid receptor (GR) is activated by its endogenous ligand cortisol and provides a vital negative feedback loop to modulate stress by downregulating the hypothalamic–pituitary–adrenal (HPA) axis . Yet, GRThe retinoic acid and retinoic X receptors (RAR and RXR) are activated by the vitamin A metabolites all-trans and 9-cis retinoic acid (RA), respectively. RA signaling is essential for embryonic development and plays crucial roles in anteroposterior and dorsoventral patterning of the neural plate and neural tube. Furthermore, RA signaling controls proliferation and differentiation of neuronal stem cells into neurons, astrocytes, and oligodendrocytes .LXRs have been shown to be involved in the differentiation of dopaminergic neurons [Also, the other identified receptors, such as the progesterone receptor (PR), the peroxisome proliferator-activated receptors (PPARs), the vitamin D3 receptor (VDR), the liver X/oxysterols receptors (LXRs), and the prostaglandin receptors play important roles in the developing brain. Progesterone promotes the proliferation of neural progenitor cells and dopaminergic neuronal differentiation in the developing CNS. Furthermore, it is involved in brain sexual differentiation and has been shown to stimulate myelination not only through increasing myelin synthesis in oligodendrocytes, but also by inducing the proliferation and promoting the differentiation of oligodendrocyte progenitor cells ,37. PPAR neurons , the mig neurons . Finally neurons . In huma neurons .In the EU and USA, identification of chemicals that have the potential to induce DNT is based on higher tiered animal testing. DNT testing is performed only rarely, upon evidence of neurotoxicity from acute or repeated dose toxicity studies in adult rodents, which is not always a reliable indicator for DNT ,42,43,44The ENDpoiNTs battery of testing methods is designed to capture the essential endpoints of ED-induced DNT relevant for humans, covering molecular, cellular, and organismal key events (KE) and outcomes crucial for neurodevelopmental processes both in vitro and in vivo. The models are of different origins and complexities, and focus mainly on processes in the central nervous system (CNS), with two exceptions that address early events in peripheral nervous system development. Wherever applicable and feasible, the in vitro and in vivo endpoints are designed to interrogate effects on brain regions that are relevant in the context of EDC impact on human health.The key events of brain development covered by the in vitro methods used in ENDpoiNTs have been chosen according to the available mechanistic insights pertaining to DNT see  42,43,44,45. TheThe endpoints will be probed by cell biological , biochemical , transcriptomic , and epigenomic readouts, both already established and newly identified within this project. Since various genes encoded on the X and Y chromosomes are involved in brain development, and the sexual differentiation of the brain results from an interaction between the genetic background and the actions of sex steroid hormones ,53,54, tIn a first step, the chosen in vitro models and endpoints will be tested for their responsiveness to the identified relevant endocrine pathways  with theTo ensure a smooth transition from development to validation of the methods, the following measures are implemented throughout the project: Each participant laboratory working with human induced pluripotent stem cells (hiPSCs) will generate their own hiPSC cell bank, comprising a ‘Master cell bank’ and a ‘Working cell bank’. Furthermore, for the individual assays that are established by the partners, standard operating procedures (SOPs) for all steps of each test method will be developed from each lab and then the SOPs will be exchanged amongst the in vitro partners of the project. The goal of this lab-to-lab test method transfer is to generate a proof-of-principle that the test methods are also working in a different laboratory, a prerequisite condition of the validation process for eventual regulatory acceptance of test methods. Finally, information on the individual test methods for ED-DNT will be collected in templates recently developed for in vitro DNT test methods from the OECD211 outline .Xenopus laevis frog. The non-mammalian vertebrate models are considered advantageous alternatives to traditional in vivo DNT testing because the fundamental mechanisms underlying the development and function of the nervous system are in concordance with those in mammalian species, including humans [The animal-based models which will be employed in the project are already part of the chemical testing guidelines recommended by the OECD and accepted internationally as standard methods for safety testing. These include rat, zebrafish, and the g humans . Furtherg humans . In addig humans ,42. WithTwo OECD level 4 tests, TG 421 (reproductive screening test) and TG 422 (combined 28-day/reproductive screening assay), have recently been updated with peripheral endpoints for EDCs in postnatal rat offspring. However, the brains of these offspring are not analyzed, but they could be studied with molecular markers predictive of ED-induced DNT. Molecular markers for EDC effects on neurodevelopment could also increase the predictive value and specificity of the DNT module in TG 443 (level 5), where, at present, behavioral tests and neuropathological assessment do not address endocrine-related effects, and where tests of cognitive functions, one of the main EDC targets disclosed by epidemiology in humans, are not mandatory. The developmental neurotoxicity TG 426 (level 4) is also not designed to specifically identify EDC-related effects on neurodevelopment. By integrating findings between the in vitro and in vivo models from molecular , cellular, morphological, and behavioral levels, the project will suggest a list of molecular endpoints for the amendment of existing in vivo test guidelines.(i)Transcriptomics. Transcriptomic analyses will assess the expression pattern changes of coding and non-coding RNAs, in the context of one or several DNT endpoints as a result of EDC exposure. Transcriptomics analyses will be performed in in vitro models and in specific brain areas of exposed rats, using RNA extracted both from bulk biological material and from single cells, to dissect population-specific pattern changes. Starting from the list of top regulated genes, functional annotation and data mining will be employed to gain knowledge on the affected cellular pathways and biological processes. Further analyses of regulatory motifs and associated master regulators will allow the reconstruction of gene networks and regulatory circuits affected by the exposures.(ii)Epigenomics. Epigenetic processes regulate temporal and spatial patterns of transcription and play a critical role in cell differentiation and tissue organization during development [elopment . Epigeneelopment  and thiselopment  and an ielopment ,57. Therelopment .(iii)Metabolomics. Metabolomics provides a functional readout of the physiologic state of an organism as determined by the sum of its genetic predisposition, regulation, protein abundance, and environmental influences. In ENDpoiNTs, targeted and non-targeted metabolomic approaches will be employed in exposed rats (hippocampus and MPO), zebrafish, and various in vitro models to link disruption of endocrine pathways with DNT endpoints. The aim is to understand the molecular mechanism of ED-induced DNT using exploratory and hypothesis-driven metabolomic pathway analyses, and to relate the affected molecular pathways to phenotypical, developmental, behavioral, and cognitive changes in the in vitro and in vivo assays.ENDpoiNTs will generate and integrate three different types of omics data:Additionally, tissue-imaging mass spectrometry will be used as a powerful technique to investigate the spatial distribution of metabolites in tissue and 3D structures and to relate it to the molecular changes and functional information in rat brain tissues, cortical brain organoids, and zebrafish. This will establish the spatial bridge between toxicant exposure in organisms (brain/organoids) and metabolite changes and provide a better understanding of the pathways and mechanisms affected.(a)signatures that can be used as “fingerprints” for exposures with a specific endocrine mode of action;(b)signatures that can predict certain KE and AO with relevance for DNT;(c)panels of epigenomic, transcriptomic, and metabolomic markers which can be used to predict EDC-induced developmental neurotoxicity in both in vitro and in vivo models.The transcriptomic, epigenomic, and metabolomic data generated will be analyzed and integrated across experimental models, exposures, and endpoints to identify:Within the ENDpoiNTs project, in silico screening models for identified endocrine targets will be developed to predict ED-linked DNT properties of new chemicals, and application of kinetic models will translate concentrations from the in vitro models to human exposures.To link the active concentration metric measured in vitro with an in vivo relevant effective dose, the integration of kinetics of both systems is required, and, in this context, the use of physiologically based toxicokinetic (PBTK) modeling is considered as a key element. PBTK models can simulate concentration–time profiles of substances at a specific target site in the body, whereas with reverse dosimetry, they can be used for the prediction of external effective doses in vivo starting from the in vitro active concentrations, i.e., the presumed target doses. The quantitative in vitro to in vivo extrapolation (QIVIVE) in this project will emphasize on prenatal exposures in humans, with the focus on the fetal brain as a critical target and an oral route of exposure. All QIVIVE extrapolations will correct for different protein binding patterns between serum and test medium and operate on equivalent nominal concentrations in serum and in vitro. Presumed target effect doses will be simulated for different intake exposure scenarios  to identify those intake patterns which lead to fetal serum concentration ranges similar to in vitro active concentrations.To develop initial in silico predictive models for first tier screening and compound identification, state of the art machine learning and deep learning methods  will be used. Various mathematical models will be developed for suspected molecular initiating events (MIEs) of importance in relation to the endocrine pathways. Receptor interactions will be based on data from public databases including, but not limited to, Tox21, ToxCast and PubChem Bioassay and the predictive models will be developed using a Mondrian conformal prediction approach to prevent information loss resulting from the highly unbalanced chemical datasets available. Similar models will be developed for prediction of blood–brain barrier penetration and brain tissue–blood partitioning, both to be used in conjunction with generic PBTK models to estimate neurotoxicity relevant exposure. The developed models will be compared to existing quantitative structure–activity relationship (QSAR) models, such as within the OECD QSAR toolbox, Danish QSAR, and VEGA, and will then be presented to OECD experts, ultimately to be reviewed by the QSAR management group.To ensure human relevance for the developed (in vitro) endpoints, epidemiological and biomonitoring data will be used as an information source for DNT effects of EDCs. The main objective is to link human-relevant EDC exposure ranges to DNT-related effects and markers using results from the ENDpoiNTs novel test tools through risk assessment metrics, for both single compounds and chemical mixtures.For single compounds, human-relevant levels of exposure will be linked to ED-related effects by constructing hazard quotients (HQ) per subject in multiple human studies. The HQ is the sum of each subject’s concentration of the EDC relative to the point of departure (POD) of the ED-related effect from the test system, where values above 1 are of concern. The percentage of subjects with HQ >1 will be reported per chemical and test system, thereby evaluating not only the extent of concern for human exposure, but also the sensitivity of test systems to the same chemical.In the case of mixtures, for those subjects determined to have sufficiently similar mixtures to the mixtures tested in vitro, a ‘similar mixture risk indicator’  will be constructed to determine the extent of the exposure relative to the POD from test systems. The calculated SMRI will be used in the analysis of epidemiology cohort studies to demonstrate the ED-related effects associated with higher index values. For comparison to the whole mixture strategy, when it is reasonable to assume additivity, a component-based strategy will be employed and a hazard index (HI) per subject from biomonitoring data will be constructed, e.g., .All three metrics  and the percentage of each that exceeds 1 (indicative of level of concern) will be used to compare exposures across EU regions and over time as EDC regulatory policies change.A number of AOPs related to DNT and neurotoxicity are published in the AOP-Wiki, yet only two are related to an endocrine mode of action involving TH signaling (AOP 8 and 152) . The maiENDpoiNTs integrates expertise in endocrine disruption and DNT and combines state-of-the-art in silico and in vitro tools, innovative experimental designs and technologies, and advanced biostatistics on human epidemiological and biomonitoring data to generate the necessary scientific insights into the correlative and causal links between endocrine disruption and DNT. Based on these insights, it will develop in silico and in vitro tools for chemical screening and novel molecular endpoints for existing animal-based test guidelines. While we do not expect that the developed methods and endpoints will have validated European Centre for the Validation of Alternative Methods (ECVAM) or OECD TG status at the end of the five years running time of the project, we will be able to provide robust, pre-validated tools based on scientific evidence and causal relationships established within the AOP framework. This will provide a base for future projects focusing on fully validating the developed methods and endpoints.The ultimate goal of ENDpoiNTs is to develop an integrated testing approach for ED-induced DNT and a strategy for its implementation into regulatory frameworks. The implementation strategy will be developed in the context of the EURION cluster that consists of eight sister projects advancing ED testing, and in close collaboration with the EURION international advisory panel with representatives from the OECD, the DG JRC (Joint Research Centre), and relevant European, national, and international  regulatory agencies. These interactions will provide regulatory guidance on the methods development in the project on the one hand, and an opportunity to continuously update regulators on the scientific findings on the other hand, which we expect will lead to faster regulatory acceptance and uptake of the developed integrated testing approach for ED-induced DNT.


Improving the quality of life for older adults diagnosed with dementia is a public health research priority. Reminiscence is one non-pharmacological technique used to address the behavioral and psychosocial problems associated with dementia. The uses of reminiscence in dementia care have not yet been integrated, synthesized, critically analyzed, or delineated in the literature. Whittemore and Knafl’s five-step method provided the framework for this integrative review. A comprehensive search of PubMed, CINAHL Plus, SCOPUS, and PsycINFO was undertaken. Articles published in English, focused on participants with a diagnosis of mild to moderate dementia, and with evidence of using reminiscence in dementia care were included. Twenty-six studies published between 2009 and 2019 that met the inclusion criteria were analyzed. Four themes emerged from the integrated findings: (a) recovery from the darkness of depressive symptoms; (b) enhancement of cognitive functions and filling the memory gap; (c) living a fulfilling life in late adulthood, and (d) fulfilling reminiscence functions. The critical appraisal process revealed mixed effectiveness of the use reminiscence on health outcomes in dementia care due to diverse types of reminiscence, different outcomes measures, different data collection toolkits, and a lack of a standardized reminiscence protocol among research studies. Results from this review provide a better understanding of the potential benefits of using reminiscence in dementia care. However, improving the methodological rigor of future studies is necessary to attain conclusive evidence of the effectiveness of using reminiscence in dementia care. Implications of these findings for gerontological education, practice, and research will be presented.


It is now clear that the most common solid cancer is squamous cell cancer (SCC) . This maCurrent conventional treatment regimens for SCC, including surgery, radiation, chemotherapy, targeted therapy and immunotherapy, are non-selective and are administered regardless of biomarkers . TherapyIn this Special Issue, Dr Darido brings together experts in the field of SCC to provide an overview of the current treatment advances. As we develop a better understanding of the limitations of current therapies, we expect to highlight new diagnostic, prognostic/predictive and therapy response/resistance biomarkers for interventional modalities in emerging immuno-oncology therapeutic areas, and areas of unmet clinical need. Future prospects enabling improved therapies will provide an exciting therapeutic roadmap for the control of SCC and will ultimately contribute to alleviating the huge burden of SCC in patients.


Mitophagy is the major degradation pathway, by which cells regulate mitochondrial number and integrity, in response to metabolic and physiological state. Research in mitophagy has grown exponentially over the last decade, revealing the intricate signaling pathways regulating mitophagy and the complexities of the molecular machinery involved in carrying out mitochondrial elimination. A diverse repertoire of mitophagy-related proteins has been discovered, highlighting an elaborate regulatory network of mitochondrial homeostasis that responds differentially to developmental, hormonal, and/or environmental signals. Moreover, the multi-layered crosstalk between mitophagy signaling pathways sustains energy metabolism, which is critical for tissues and organs homeostasis. Indeed, defective mitophagy causes accrual of dysfunctional mitochondria leading to bioenergetic stress, elevated ROS levels and pronounced inflammation that is accompanied by cellular and tissue degeneration. Thus, mitophagy is a pivotal contributor to cellular physiology, and tissue integrity, in addition to organismal development, healthspan, and survival.Frontiers in Cell and Developmental Biology includes a series of 11 articles that discuss recent advances in the field of mitophagy research and highlight challenges and outstanding questions, that need to be addressed before mitophagy modulation can be considered for the development of effective therapeutic interventions.The Research Topic on “Mitophagy in Health and Disease” in Ravanelli et al. discuss the critical role of the ubiquitin/proteasome system (UPS) in mitochondrial quality control. Alterations in the ubiquitination status of mitochondrial proteins contribute to the remodeling of the mitochondrial proteome in response to stress conditions. The authors discuss the tight crosstalk between the UPS and mitochondria that contributes to prevent proteostasis collapse and promote energy metabolism.Several molecular mechanisms have been identified that mediate mitochondrial removal in a cell type- and tissue-specific manner. In their review, Wang et al. surveys the regulation of mitochondrial removal by the phosphatase and tensin homolog (PTEN) isoforms. The authors introduce the molecular function of PTEN-short and PTEN-long proteins and their association with the mitophagic machinery. In addition, the authors highlight post-translational modifications as a central node of mitophagy, suggesting that their modulation can be used for the development of novel intervention approaches toward tackling mitochondrial-related disorders.The article by Ravanidis and Doxakis address the role of RNA-binding proteins (RBPs) in mitochondrial homeostasis. The authors describe the vital role of RBPs in the maintenance of mitochondrial metabolism through the regulation of mRNA splicing, stability, targeting to mitochondria and translation. Importantly, substantial evidence indicates that impaired expression, or mutations in RBPs, contribute to mitochondrial dysfunction that has been implicated in the development and progression of several age-associated neurodegenerative disorders.In their review, Zachari and Ktistakis survey the molecular mechanisms that govern mitochondrial degradation, with a particular focus on the early signaling events of mitophagy initiation and mitoautophagosome formation.Although several components of the mitophagic machinery have been uncovered, the origin of mitoautophagosomal membranes remains elusive. In their review, Ma et al. focus on how excessive mitochondrial damage can trigger innate immune responses and apoptotic cell death via BCL2 protein family members. The authors explore the molecular mechanisms that uphold mitochondrial homeostasis, including mitochondrial dynamics, mitochondrial biogenesis, and mitophagy among others, as well as, how these cellular events interfere with cell fate. Moreover, a relevant review by Joaquim and Escobar-Henriques discusses the pro-survival and pro-apoptotic role of mitophagy. The authors describe the involvement of mitofusins in mitophagy, through the modulation of mitochondrial morphology and endoplasmic reticulum (ER)-mitochondria contact sites. Finally, they summarize emerging findings, suggesting that impaired mitochondrial dynamics and mitophagy contribute to the pathogenesis of non-alcoholic liver disease.A growing body of evidence suggests an intricate communication between mitophagy and cell death pathways. In their review, Luo et al. discuss the contribution of mitophagy deregulation during aging to the homeostasis and viability of post-mitotic neurons and cardiomyocytes. Furthermore, the authors discuss recent studies that link impaired mitophagy to the development of neurodegenerative and cardiovascular pathologies.Several studies have revealed a progressive, age-related decline of mitophagic flux in multiple tissues, including as heart, kidney, liver, and brain. In their article, Xie C. et al. survey the role of defective mitophagy in the pathogenesis of Alzheimer disease (AD). Increasing evidence indicates that accumulation of damaged mitochondria, due to mitophagy impairment contributes to Aβ/Tau proteinopathies and stimulates persistent inflammation, causing neuronal loss, and cognitive decline. In addition, the authors discuss the potential use of mitophagy inducers, such as NAD+ precursor molecules and urolithin A, toward ameliorating aging, and AD pathological features.The article by Xie Y. et al. surveys the molecular pathways that govern mitochondrial elimination, focusing on the essential role of mitophagy receptors. Furthermore, the authors discuss the involvement of mitophagy receptors in tumorigenesis, and highlight the therapeutic potential of mitophagy modulation in cancer therapies.The review by Chen et al. delineate the molecular pathways and mechanisms of mitophagy in several model organisms, and discuss the significant contribution of mitophagy defects in age-related pathologies. In their article, Bakula and Scheibye-Knudsen introduce the term “mitophaging” pointing to the fundamental role of mitochondrial integrity in the maintenance of cellular fitness and organismal health. Both articles discuss interventions that target different steps in the process of mitophagy, by utilizing small molecular compounds as a means toward the development of novel, effective treatments against currently incurable pathologies.The last two articles discuss emerging findings that highlight the anti-aging properties of mitophagy. In their review, In closing this Editorial piece, we would like to thank all the authors and referees, for their valuable contributions, toward compiling this up-to-date and timely Research Topic on mitophagy in health and disease. Moreover, we hope that the collection of articles included in the topic will provide a useful point of reference and a stimulus for further research aiming to ultimately decipher the complex contributions of mitophagy to cellular and organismal homeostasis.KP and NT wrote the manuscript. KP, EF, and NT read and edited the manuscript. All authors contributed to the article and approved the submitted version.EF has CRADA arrangement with ChromaDex, and is consultant to Aladdin Healthcare Technologies, Vancouver Dementia Prevention Centre, and Intellectual Labs. The remaining 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.


The multifaceted key roles of cytokines in immunity and inflammatory processes have led to a high clinical interest for the determination of these biomolecules to be used as a tool in the diagnosis, prognosis, monitoring and treatment of several diseases of great current relevance . Therefore, the rapid and accurate determination of cytokine biomarkers in body fluids, cells and tissues has attracted considerable attention. However, many currently available techniques used for this purpose, although sensitive and selective, require expensive equipment and advanced human skills and do not meet the demands of today’s clinic in terms of test time, simplicity and point-of-care applicability. In the course of ongoing pursuit of new analytical methodologies, electrochemical biosensing is steadily gaining ground as a strategy suitable to develop simple, low-cost methods, with the ability for multiplexed and multiomics determinations in a short time and requiring a small amount of sample. This review article puts forward electrochemical biosensing methods reported in the last five years for the determination of cytokines, summarizes recent developments and trends through a comprehensive discussion of selected strategies, and highlights the challenges to solve in this field. Considering the key role demonstrated in the last years by different materials (with nano or micrometric size and with or without magnetic properties), in the design of analytical performance-enhanced electrochemical biosensing strategies, special attention is paid to the methods exploiting these approaches. Cytokines are soluble low-molecular-weight proteins secreted by cells (both immune and non-immune) responsible for regulation of host defense, tissue homeostasis, cell-to-cell communication, and inflammatory reactions. Cytokines are key biomolecules acting as mediators and modulators of the complex functional interactions and responses of the immune system .There is growing evidence that the quantification of cytokines allows immune and inflammatory responses to be predicted or monitored in a comprehensive and timely manner, thus providing clinically and immunologically useful information for the diagnosis and stratification of infectious, cancer and autoimmune diseases. Owing to the highly networked nature of their functions, multiplexed detection of different cytokines in a single sample is highly desirable to obtain a more complete and accurate picture of the process . This, tCytokines are small bioactive proteins (~6–70 kDa), secreted by immune and non-immune cells, which are considered core indicators of the functional status of the body. They are strongly associated with the immune system acting as signaling molecules at picomolar or nanomolar concentrations to regulate inflammation and modulate cellular activities . They afThese proteins act in concert with specific cytokine inhibitors and soluble cytokine receptors to regulate the human immune response. The binding of a cytokine ligand to its cognate receptor results in the activation of the receptor, which in turn triggers a cascade of signaling events that regulate various cellular functions, such as cell adhesion, phagocytosis, cytokine secretion, cell activation, cell proliferation, cell survival and cell death, apoptosis, angiogenesis and proliferation . CytokinChemokines, or “chemotactic cytokines,” make up a group of secreted proteins <40 kDa) which induce cell migration and include IL-8, MCP-1, MIP-1α, CC and CXC. They are involved in leukocyte chemoattraction and trafficking of immune cells to locations throughout the body. Chemokines are classified in: (i) homeostatic chemokines, which are involved in immune surveillance and navigation of cells through hematopoiesis, and are typically expressed constitutively; (ii) inflammatory chemokines, which are produced during infections or as a response to an inflammatory stimulus and facilitate an immune response by targeting cells of the innate and adaptive immune system ,42p-ABA-odifiers b [43] toodifiers .Moreover, immunoscaffolds reported for the determination of inflammatory cytokines and chemokines involved in click ,57 or el−1–ng mL−1 level, and were employed to perform the analysis of the target cytokines/receptors  in different human biofluids .The electrochemical immunoplatforms prepared using electrografting and click or electro-click chemistry exhibited analytical characteristics compatible with clinical applications providing LODs in the low pg mLRegarding nanomaterials, their multifunctional nature facilitates the improvement of several key features in electrochemical bioassays, including sample treatment, analyte capture, signal amplification and transduction. They have been widely used as electrode modifiers to improve the immobilization of bioreceptors and the charge transfer as well as advanced labels able to carry large amounts of electroactive reporters to amplify the electrochemical signals .2), magnetic and polymer (PPyNP) nanoparticles and carbon nanomaterials  either as single nanostructures or combined in hybrid nanostructures  as well as with other modifiers such as chitosan (CS), carboxymethylcellulose (CMC) and ionic liquids (ILs) displaying synergic properties.The nanostructures used in electrochemical biosensing strategies reported in the last five years for the determination of cytokines include metallic , metal oxide  in integrated immunoplatforms prepared at p-ABA grafted-SPCEs for the determination of IL-13Rα2 ,55. An e2 mesocrystal nanoarchitectures: a composite prepared from TiO2 (anatase) mesocages (AMCs) and a carboxy-terminated ionic liquid (CTIL) as a modifier of a GCE, and octahedral anatase TiO2 mesocrystals (OAMs) as a matrix for immobilizing acid phosphatase (ACP) and secondary antibody (Ab2) labeled with horseradish peroxidase (HRP)  sandwich-type immunosensor for the determination of IL-6 using two kinds of TiOse (HRP)  48]. Th. Th2 mes2O2/ferrocenemethanol system, in undiluted whole blood at the low pg mL−1 level with results in good agreement with those obtained using a commercial ELISA kit . Bi. Bi24]. −1 (TNF-α) and 0.95 pM (IL-6) and were applied to the determination in spiked human serum samples.Simple immunosensors with good analytical performance have been reported for the determination of TNF-α and IL-6 using sandwich and direct formats by covalent immobilization of capture antibodies on SAM-modified Au microarrays of thiolated cross-linkers able to bind primary amines through formation of amide bonds   and sulfBearing in mind that cytokines are involved in several of the diseases of concern to our society nowadays, the interest in their determination and in the implementation of competitive methodologies for this purpose will not wane. As already commented, it is essential to perform their multiplexed determination with tools able to satisfy the demands of the current clinic in terms of the simplicity of use, affordable cost and suitability for decentralized analysis by any user.Electrochemical biosensing technologies are evolving and maturing in amazing ways to meet many of these demands. Indeed, cytokines, as target biomarkers, are in vogue in electrochemical biosensing applications, as is evident by the upsurge in the number of research articles devoted to the determination of cytokines in the past few years.The incorporation of materials, other types of modifiers, chemicals and attractive bioassay formats into their design has played a decisive role in the opportunities and capabilities provided by electrochemical biosensing. In order to give a realistic picture of the state of the art, this review article discusses the significant advances and features achieved in the field through selected representative methods reported in the last five years. These methods show that both magnetic particles-based and integrated electrochemical bioplatforms exhibit compatible characteristics in terms of sensitivity and selectivity with the determination of these biomarkers in the clinic.The use of magnetic particles, particularly those of micrometric size, as solid supports for the implementation of bioassay configurations, combined with electrochemical transduction on SPEs has shown the suitability to perform multiomic determinations (simultaneous determination of IL-8 and its associated mRNA) and to analyze cytokines in small amounts of complex biological samples , in clinically actionable times and using simple test protocols.In the case of integrated formats, coupling with nanomaterials and a rational modification of electrode surfaces with suitable chemicals and modifiers play particular relevant roles. Electrografting of diazonium salts, click and electroclick chemistries provide a great versatility and ability for the functionalization of carbon nanomaterials and electrode surfaces, and in the incorporation of ethynyl-functionalized antibodies. The electrode functionalization, together with the use of SAMs of zwitterionic compounds, PC membrane-based off-matrix and (semi-)conducting polymers, makes it possible to impart anti(bio)fouling properties to biosensors, which is highly demanded to ensure their adequate performance in real world matrices. A variety of nanomaterials, either used singly, or combined in hybrid nanostructures or with other modifiers , have been employed as electrode modifiers to improve bioreceptor immobilization and charge transfer, as well as advanced labels to carry large numbers of electroactive reporters.−1, but in some cases also fg mL−1) and some of them have been successfully integrated into microfluidic devices. In addition, the methods have been applied to the determination of cytokines in un-pretreated whole blood and saliva, whole cells, cell lysates or secretions and paraffin tissue extracts.The reported bioelectroanalytical strategies exhibit high sensitivity . Furthermore, the application to real samples is still limited or has been addressed with a small number of samples and only at research level. Future efforts in this area should be carried out in close coordination with different actors in the clinic and society, and focused on exhaustive validations of both candidate biomarkers (existing and others that can be identified) and developed devices. This combined effort will be essential for the future implementation of these strategies in the market, clinic and daily life. At the same time as working on the validation of the biomarkers and the corresponding devices, one must not forget to continue advancing in the development of other strategies exploiting the use of other bioreceptors such as biomolecular switches and nanomaterials with mimicked enzyme activity (nanozymes). This could open the way for reagentless biodevices, with the capacity for almost real time determinations and of lower cost and more robust to variations in pH and temperature than those involving natural enzymes.Despite the many challenges lying ahead, we are pretty convinced that the continuous advancements in electrochemical biosensing technology will provide user-friendly but powerful tools suitable for adoption in real clinical settings, at which point screening programs and personalized medicine will become a reality, to assist in the diagnosis, prognosis and monitoring of diseases facing our society, by determining the cytokines involved in the autoimmune and inflammatory processes occurring during their starting and evolution.


Small island developing states in the Caribbean are among the most vulnerable countries on the planet to climate variability and climate change. In the last 3 decades, the Caribbean region has undergone frequent and intense heat waves, storms, floods, and droughts. This has had a detrimental impact on population health and well-being, including an increase in infectious disease outbreaks. Recent advances in climate science have enhanced our ability to anticipate hydrometeorological hazards and associated public health challenges. Here, we discuss progress towards bridging the gap between climate science and public health decision-making in the Caribbean to build health system resilience to extreme climatic events. We focus on the development of climate services to help manage mosquito-transmitted disease epidemics. There are numerous areas of ongoing biological research aimed at better understanding the direct and indirect impacts of climate change on the transmission of mosquito-borne diseases. Here, we emphasise additional factors that affect our ability to operationalise this biological understanding. We highlight a lack of financial resources, technical expertise, data sharing, and formalised partnerships between climate and health communities as major limiting factors to developing sustainable climate services for health. Recommendations include investing in integrated climate, health and mosquito surveillance systems, building regional and local human resource capacities, and designing national and regional cross-sectoral policies and national action plans. This will contribute towards achieving the Sustainable Development Goals (SDGs) and maximising regional development partnerships and co-benefits for improved health and well-being in the Caribbean. This Perspective article discusses progress towards bridging the gap between climate science and public health decision-making, highlighting the importance of multi-institutional and interdisciplinary collaborations to develop early warning systems for arboviral diseases in the Caribbean. Frequent and intense extreme climatic events in the Caribbean, such as heat waves, Saharan dust incursions, hurricanes, floods, and droughts, have a detrimental impact on human health and development in the Caribbean region . AdverseAedes spp. mosquitoes. The global incidence of dengue has increased 30-fold in the last 50 years, and the disease is now endemic in more than 120 countries in the tropics and subtropics [Aedes spp.) as dengue, was limited to Africa and Asia. Since then, these viral diseases have spread globally, following the path of dengue [Dengue is a viral infection transmitted to humans by female f dengue . Chikungf dengue . Zika ref dengue . To datef dengue . In 2019f dengue . In recef dengue . BetweenAedes aegypti is the primary vector of dengue, chikungunya, and Zika viruses, a mostly domestic, urban mosquito that lays its eggs in water-bearing containers in and around the home. Ae. aegypti thrive in warm and humid conditions. Warm temperatures with an optimum peak of around 28.5°C increase the risk of Ae. aegypti–transmitted diseases by speeding up mosquito development rates, reproduction, survival, biting rates, and viral replication in the mosquito [mosquito . The effIn this perspective, we discuss the state of the art in climate services for health in the Caribbean region, with a focus on early warning systems for mosquito-transmitted diseases, such as dengue, chikungunya, and Zika. We highlight the importance of multi-institutional and transdisciplinary collaborations at national, regional, and international levels for research-to-operations-to-research  innovation in the coproduction of climate services for health. These policy advances are of relevance to scientists and multisectoral decision-makers anticipating the impacts of climate extremes, variability, and climate change on health system capacity to respond to mosquito-borne disease outbreaks.There is a growing body of evidence on the impacts of climate extremes, variability, and climate change on human health in the Caribbean, including increases in acute asthma admission from Saharan dust clouds , increasAedes mosquitoes, and widespread intra and extraregional travel in the Caribbean [Better management of mosquito-borne diseases is a top priority for Caribbean small island developing states given the high socioeconomic burden, suitable climatic conditions for the proliferation of aribbean . Thanks aribbean , the CIMA recent study of climate and health stakeholders in Barbados and Dominica  identifiThe island of Barbados is among the 10 countries reporting the most cases of dengue in the Caribbean . It is aAe. aegypti mosquitoes if not properly covered and managed. This measure may have had the unintended consequence of increasing the overall risk of Aedes-transmitted diseases when rainfall is collected and stored. A link between temporary water storage and dengue risk has been recognised, but few studies have quantified the link or examined the impact of prolonged drought on dengue transmission.To mitigate the impacts of prolonged droughts, Barbados passed building regulations to mandate the construction of rainwater storage receptacles under large new buildings. However, these receptacles have the potential to become ideal larval habitat for To address this gap, Lowe and colleagues designed a coupled model framework to quantify the nonlinear and delayed impacts of climate factors, such as drought and extreme rainfall, on dengue transmission in Barbados from 1999 to 2016 . The modThis modelling approach, which infers the risk of dengue outbreaks given the cumulative effect of climate variations in the preceding months, is currently being extended to other countries and tested as an operational early warning tool. The CIMH routinely produces seasonal forecasts of the SPI, minimum temperature, and other climate indicators, with a lead time of up to 3 months. By incorporating operational seasonal climate forecasts in the dengue prediction model, probabilistic dengue outlooks could be issued at least 3 months in advance of any given month and updated on a monthly basis, by incorporating new meteorological observations and shorter lead climate forecasts. One avenue for communicating dengue outlooks is through the quarterly Caribbean Health Climatic Bulletin, coproduced by the CIMH, PAHO and CARPHA . The onlThe development and effective mainstreaming of climate services for health, such as climate-driven dengue early warning systems, require parallel efforts to increase the capacity of the climate and health sectors in the region. National health stakeholders in Dominica and Barbados identified significant gaps in the availability of financial resources to implement an operational early warning system and a self-reported lack of technical expertise in statistics, data science, and geographic information systems (GIS) . They alThere is a critical need to train and retain a cohort of climate and health practitioners. In particular, the health sector requires technical training in how to analyse and interpret basic climate information and integrate this information into health surveillance and planning, whereas the climate sector requires more knowledge about the decision priorities and needs of the health sector. Further to this, resources must be allocated to reconciling data holdings across sectors. Historical records are often inconsistent in format and primarily paper based, while decentralised collection and storage complicates archiving or digitising. Country personnel trained in data management, from collection to data entry to quality control to data sharing, may be a limiting factor. Following recommendations from the Global Vector Control Response , vector Capacity building needs are being addressed, in part, through the development of an online open access course on climate and health by a consortium of international experts led by WHO, PAHO, and WMO.Regional action on (1) the SDGs agenda, (2) the climate actions proposed by Caribbean small island developing states under the Paris Climate Agreement, (3) the strategic priority areas outlined by the 10-year Caribbean Community (CARICOM) Caribbean Cooperation in Health (CCH) (2016 to 2025) initiative , and (4)At the national level, countries in the Caribbean are developing and implementing their National Development Plans, National Adaptation Plans for climate change, and disaster risk reduction strategies. The PAHO has also led efforts to develop Health National Adaptation Plans focused on climate resilient health systems . Policy The Caribbean climate and health sectors are beginning to work together to attract the resources needed to increase local capacities to develop climate services for the health sector . However


The first epithelia to arise in an organism face the challenge of maintaining the integrity of the newly formed tissue, while exhibiting the behavioral flexibility required for morphogenetic processes to occur effectively. Epithelial cells integrate biochemical and biomechanical cues, both intrinsic and extrinsic, in order to bring about the molecular changes which determine their morphology, behavior and fate. In this review we highlight recent advances in our understanding of the various dynamic processes that the emergent epithelial cells undergo during the first seven days of mouse development and speculate what the future holds in understanding the mechanistic bases for these processes through integrative approaches. Epithelia are fundamental tissue structures comprising of sheets of interconnected cells that act as physical and chemical barriers between different tissues, or tissues and their surroundings. They are highly dynamic and can undergo characteristic changes in shape and size, driven by regionalised, coordinated cell behaviours. Epithelia are critical regulators of morphogenesis at multiple levels including tissues, organs, and ultimately the shape of the whole organism. During early mouse embryogenesis, cells differentiate into embryonic and extraembryonic tissues, giving rise to some of the first epithelia of the organism . The celEpithelia can be seen as the default type of eumetazoan cells as they represent the first organised tissue-type to arise during development. Apicobasal polarity is a defining feature of epithelia, whereby the component cells interact with the extracellular basal lamina along their basal side, while maintaining strong interactions with each other through intercellular junctions at their apical side. Blastomeres of the late 8-cell embryo possess proto-epithelial characteristics such as the accumulation of E-cadherin at cell-cell interfaces which, along with cortical actomyosin-mediated tension, helps to drive compaction –5. At thde novo epithelialisation is associated with fate commitment decisions whereby epiblast cells downregulate their naïve pluripotent genetic profile and commit to a primed state [in vitro, by a disengagement of the plasma membrane from the underlying actomyosin cortex [The ICM segregates into two epithelial tissues, the Epiblast and Primitive Endoderm (PrE) through a MAPK dependent pathway . The epied state . Both thed state . They arn cortex  and acqun cortex . The epin cortex ,32.As development progresses, heterogeneities emerge within epithelia of the early embryo. Recent single-cell molecular approaches have helped to unravel some of the transcriptomic, epigenetic and proteomic changes that precede the emergence of a diversity of overt behavioural differences among epithelial cells that are, in turn, responsible for subsequent morphological transformations –38. ThesCell-shape changes can initially result from exogenous mechanical forces generated through events such as expanding intra-embryonic cavities or changing topology of the surrounding tissue following implantation. These changes are stabilised by modifications to the cytoskeletal and adhesive architecture of cells through alterations to gene expression or the proteome . The resA notable example of cell-shape changes occurring within an epithelium is the conversion of a subset of VE cells, the distal visceral endoderm (DVE), from a squamous to a columnar epithelium. As the mouse embryo acquires its characteristic cup-shape with the expansion of the pro-amniotic cavity, the DVE is specified at the point of maximum curvature, at the distal extreme of the egg-cylinder . In addiGiven the crucial function of the AVE in axial patterning, the mechanism that induces the DVE would have to accommodate the inter-species variation in shapes seen between mammalian embryos. While the mouse embryo is ‘cylindrical’ in shape at this stage, the equivalent stage human embryo is ‘disk’ shaped . Cells sCell migration is a fundamental process that underlies many developmental events, enabling either the dispersal of cells to new locations or the reorganisation of cells within a tissue. For tissues that are specialised to act as barriers, epithelia are remarkably dynamic at the cellular level, facilitating widespread rearrangement and movement of cells.A striking example of migration of a sub-population of cells within the plane of an intact epithelial monolayer is the AVE, which collectively migrate from the distal-most tip of the egg-cylinder towards one side of the epiblast over the course of several hours . From thOver the last 15 years, a combination of live imaging and genetic knockouts have revealed that AVE migration is an active process, controlled by coordinated differences in behaviour in sub-regions of the VE ,55. KnocIn the blastocyst, the integrity of the epithelium itself can be a dynamic property. High blastocoel pressure leads to junctional leakages around cells undergoing mitosis, which, though momentarily disrupting epithelial integrity, helps to maintain the overall size of the blastocyst by restricting size oscillating within confined limits . In the It is unclear why such dispersal of cells during division is seen in some epithelia such as the epiblast, but not others, such as the VE. It is possible that it only occurs in epithelia subject to particular levels of stress, or a particular geometry . In the case of the epiblast, the dispersal of cells could have a selective advantage as it would prevent the localised clonal accumulation of cells with deleterious mutations, which could otherwise impact on epithelial integrity if they are collectively elimination during the wave of cell competition that occurs prior to gastrulation ,66. It cde novo pattering as the only means of generating new epithelia. In fact, the first migratory cell population in the embryo, the parietal endoderm (PE), is generated through delamination from the PrE, an epithelial tissue that arises from the ICM at ~E4.0. PE cells migrate along the inner surface of the TE, eventually contributing to extraembryonic structures of the foetus. Little is known about this first epithelial-to-mesenchymal transition (EMT) event, as the delamination of the PE coincides with the implantation of the embryo into the uterine wall, making it relatively intractable to experimentation.During homeostatic maintenance of tissues, interconversion of epithelial and mesenchymal cells is rare and seen mostly only under pathological circumstances. However, during development, transitions between the two states occur frequently, and are used as a means through which cells originating in epithelia can move over larger distances and reassembled at new anatomical locations. This reduces the reliance on However, this process is subsequently repeated multiple times during development, most notably at E6.25, during gastrulation. Here, cells in the pseudostratified epiblast undergo a fundamental cell fate choice, either delaminating to form the primitive streak through which cells move to give rise to the mesoderm and endoderm, or remain behind to commit to a default ectodermal differentiation pathway . Major spartial EMT [The mesenchymal endoderm progenitors, generated at gastrulation through EMT, later re-epithelialise through mesenchymal-to-epithelial transition (MET) to give rise to the definite endoderm (DE) which intercalates with the existing VE epithelium, collectively to form the gut tube . Recentltial EMT , which mtial EMT ,74.Epithelia are dynamic structures that must constantly reconcile two opposing imperatives; structural integrity and behavioural flexibility. Understanding how epithelial cells achieve this is important for understanding embryonic morphogenesis during development, and also in providing a route for identifying the molecular machinery relevant to pathological transformations. Great strides have been made in unravelling the molecular and cellular biology of embryonic epithelial behaviour. In the future, advances in imaging, biomechanical and biochemical perturbation and computational modelling approaches may help us connect the molecular to the biophysical modulation of epithelial dynamics.