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Significance: Nonhealing wounds are a significant burden for the health care system all over the world. Existing treatment options are not enough to promote healing, highlighting the urgent need for improved therapies. In addition, the current advancements in tissue-engineered skin constructs and stem cell-based therapies are facing significant hurdles due to the absence of a renewable source of functional cells. Recent Advances: Induced pluripotent stem cell technology (iPSC) is emerging as a novel tool to develop the next generation of personalized medicine for the treatment of chronic wounds. The iPSC provides unlimited access to various skin cells to generate complex personalized three-dimensional skin constructs for disease modeling and autologous grafts. Furthermore, the iPSC-based therapies can target distinct wound healing phases and have shown accelerating wound closure by enhancing angiogenesis, cell migration, tissue regeneration, and modulating inflammation. Critical Issues: Since the last decade, iPSC has been revolutionizing the field of wound healing and skin tissue engineering. Despite the current progress, safety and heterogeneity among iPSC lines are still major hurdles in addition to the lack of large animal studies. These challenges need to be addressed before translating an iPSC-based therapy to the clinic. Future Directions: Future considerations should be given to performing large animal studies to check the safety and efficiency of iPSC-based therapy in a wound healing setup. Furthermore, strategies should be developed to overcome variation between hiPSC lines, develop an efficient manufacturing process for iPSC-derived products, and generate complex skin constructs with vasculature and skin appendages.

Background Gene dosage imbalance caused by copy number variations (CNVs) is a prominent contributor to brain disorders. In particular, 15q11.2 CNV duplications and deletions have been associated with autism spectrum disorder and schizophrenia, respectively. The mechanism underlying these diametric contributions remains unclear. Methods We established both loss-of-function and gain-of-function mouse models of Cyfip1, one of four genes within 15q11.2 CNVs. To assess the functional consequences of altered CYFIP1 levels, we performed systematic investigations on behavioral, electrophysiological, and biochemical phenotypes in both mouse models. In addition, we utilized RNA immunoprecipitation sequencing (RIP-seq) analysis to reveal molecular targets of CYFIP1 in vivo. Results Cyfip1 loss-of-function and gain-of function mouse models exhibited distinct and shared behavioral abnormalities related to autism spectrum disorder and schizophrenia. RIP-seq analysis identified messenger RNA targets of CYFIP1 in vivo, including postsynaptic NMDA receptor (NMDAR) complex components. In addition, these mouse models showed diametric changes in levels of postsynaptic NMDAR complex components at synapses because of dysregulated protein translation, resulting in bidirectional alteration of NMDAR-mediated signaling. Importantly, pharmacological balancing of NMDAR signaling in these mouse models with diametric Cyfip1 dosages rescues behavioral abnormalities. Conclusions CYFIP1 regulates protein translation of NMDAR and associated complex components at synapses to maintain normal synaptic functions and behaviors. Our integrated analyses provide insight into how gene dosage imbalance caused by CNVs may contribute to divergent neuropsychiatric disorders.

This study aimed to biochemically and histopathologically investigate the effect of sunitinib on oxidative testicular damage induced by ischemia/reperfusion in rats.Experimental animals were divided into three groups of six rats each: testicular torsion-detorsion (TTD), sunitinib+testicular torsion-detorsion (STD), and sham control (SC). Sunitinib (25mg/kg) was administered orally to the STD group by gavage. Normal saline (0.9% NaCl) was administered orally to the TTD and control groups as the solvent. One hour after administration of sunitinib and 0.9% NaCl, all animal groups were done torsion-detorsion. Then, all the rats were killed by high-dose anesthesia, and their testicles were removed. Biochemical and histopathological examinations were performed on the removed testicular tissues.Malondialdehyde; it was observed that the results in the STD group were close to those of the SC group and statistically significant lower compared to the TTD group (p=0.001). The glutathione values were statistically significantly higher in the STD group compared to the TTD group (p0.001). Nuclear factor kappa B values, revealing a statistically significant difference between the TTD and STD groups (p0.001). The TNF-α levels were measured and indicating that the results of the STD group were statistically significantly lower than those of the TTD group (p0.001). Histopathologically, animal tissues given sunitinib were observed to resemble normal tissues.Sunitinib was shown to prevent histopathological changes in testicular tissue against ischemia/reperfusion damage.

Cell-cell communication through evolutionarily conserved signaling pathways governs embryonic development and adult tissue homeostasis. Deregulation of these signaling pathways has been implicated in a wide range of human diseases including cancer. One such pathway is the Hedgehog (Hh) pathway, which was originally discovered in Drosophila and later found to play a fundamental role in human development and diseases. Abnormal Hh pathway activation is a major driver of basal cell carcinomas (BCC) and medulloblastoma. Hh exerts it biological influence through a largely conserved signal transduction pathway from the activation of the GPCR family transmembrane protein Smoothened (Smo) to the conversion of latent Zn-finger transcription factors Gli/Ci proteins from their repressor (GliR/CiR) to activator (GliA/CiA) forms. Studies from model organisms and human patients have provided deep insight into the Hh signal transduction mechanisms, revealed roles of Hh signaling in a wide range of human cancers, and suggested multiple strategies for targeting this pathway in cancer treatment.

Middle age is increasingly accepted as a key period during which individuals are susceptible to the effect of environmental and lifestyle factors. Emerging research indicates that dietary factors play a crucial role in brain health and cognitive function, and studies in both animals and humans have demonstrated that dietary interventions can mitigate cognitive impairment. Specifically, magnesium has been shown to enhance learning and memory, and magnesium deficiency is associated with impaired hippocampal-dependent memory formation in animal studies. The aim of this study was to examine if supplementation with a magnesium-rich marine mineral blend (MMB) could alter middle-age-related cognitive impairment. Young and middle-aged rats were given access to a control diet or an experimental diet formulated with an MMB for 4 weeks before undergoing a series of behavioral assessments. Supplementation of MMB to middle-aged rats rescued a deficit in cognitive impairment, specifically a pattern separation paradigm that is sensitive to alterations in a type of brain plasticity called neurogenesis. It had no effect on general activity in the open field or performance on other hippocampal-associated tasks. Changes in cognitive function occur as a predictable consequence of aging. Research into whether modification of dietary factors, such as this MMB, may play a role in the prevention of age-related cognitive impairment warrants further investigation.

Introduction SARS-CoV-2 is the pathogen of COVID-19. The virus is composed of the spike, membrane and envelope. On physiological smell, odoriferous substances bind to proteins secreted by sustentacular cells in order to be processed by olfactory receptor neurons. Olfactory disorder is one of the main manifestations of COVID-19, however, research is still required to clarify the mechanism involved in SARS-CoV-2 induced anosmia. Objective This article aims to analyze current scientific evidence intended to elucidate the pathophysiological relationship between COVID-19 and the cause of olfactory disorders. Methods Pubmed, Embase, Scopus and ScienceDirect were used to compose this article. The research was conducted on November 24th, 2020. Original articles with experimental studies in human, animal and in vitro, short communications, viewpoint, published in the English language and between 2019 and 2020 were included, all related to the pathophysiological relationship between olfactory disorders and COVID-19 infection. Results Both human cell receptors ACE2 and TMPRSS2 are essential for the SARS-CoV-2 entrance. These receptors are mostly present in the olfactory epithelium cells, therefore, the main hypothesis is that anosmia is caused due to damage to non-neuronal cells which, thereafter, affects the normal olfactory metabolism. Furthermore, magnetic resonance imaging studies exhibit a relationship between a reduction on the neuronal epithelium and the olfactory bulb atrophy. Damage to non-neuronal cells explains the average recovery lasting a few weeks. This injury can be exacerbated by an aggressive immune response, which leads to damage to neuronal cells and stem cells inducing a persistent anosmia. Conductive anosmia is not sufficient to explain most cases of COVID-19 induced anosmia. Conclusion Olfactory disorders such as anosmia and hyposmia can be caused by COVID-19, the main mechanism is associated with olfactory epithelium damage, targeting predominantly non-neuronal cells. However, neuronal cells can also be affected, worsening the condition of olfactory loss.

Olfactory dysfunction is manifested in a wide range of neurological and psychiatric diseases, and often emerges prior to the onset of more classical symptoms and signs. From a behavioral perspective, olfactory deficits typically arise in conjunction with impairments of cognition, motivation, memory, and emotion. However, a conceptual framework for explaining the impact of olfactory processing on higher brain functions in health and disease remains lacking. Here we aim to provide circuit-level insights into this question by synthesizing recent advances in olfactory network connectivity with other cortical brain regions such as the prefrontal cortex. We will focus on social cognition as a representative model for exploring and critically evaluating the relationship between olfactory cortices and higher-order cortical regions in rodent models. Although rodents do not recapitulate all dimensions of human social cognition, they have experimentally accessible neural circuits and well-established behavioral tests for social motivation, memory/recognition, and hierarchy, which can be extrapolated to other species including humans. In particular, the medial prefrontal cortex (mPFC) has been recognized as a key brain region in mediating social cognition in both rodents and humans. This review will highlight the underappreciated connectivity, both anatomical and functional, between the olfactory system and mPFC circuitry, which together provide a neural substrate for olfactory modulation of social cognition and social behaviors. We will provide future perspectives on the functional investigation of the olfactory-mPFC circuit in rodent models and discuss how to translate such animal research to human studies.

Summary Background Polycystic ovary syndrome (PCOS) is a multifactorial hormonal disorder accompanied by impairment of endometrial function and structure. Pomegranate is recognized for its role in normalizing the female sex hormones in PCOS with little known about its effect on the accompanying endometrial histological alterations. Aim of the work To assess the possible ameliorative role of pomegranate juice extract (PJE) on endometrial injury in a rat model of PCOS. Material and methods Forty adult albino rats were equally divided into 4 groups; control, PJE-treated (400 mg/kg/day for 3 weeks), letrozole-treated (PCOS) (1 mg/kg/day for 3 weeks), and PJE & PCOS groups. Serum Follicle-stimulating hormone (FSH), Luteinizing hormone (LH), testosterone, estradiol, and tissue malondialdehyde (MDA) were assayed. Uterine samples were processed for histological staining with hematoxylin & eosin and Masson's trichrome stains, Ki67 and androgen receptor immunohistochemical staining, and scanning electron microscopy. Results PCOS group revealed a significant increase in serum FSH, LH, testosterone, estradiol, and tissue MDA. Uterine sections depicted various histological alterations in the endometrium with signs of inflammation. A significant increase in the endometrial collagen fiber content, as well as a significant upregulation in Ki67 and androgen receptor immunohistochemical expression were detected. Scanning electron microscopy showed a significant decrease in the mean number of pinopodes. Concomitant administration of PJE efficiently restored the studied biochemical, histological, and immunohistochemical parameters. Conclusion PJE ameliorated PCOS accompanying endometrial histological alterations through its antioxidant, anti-inflammatory, anti-fibrotic, anti-proliferative, and anti-androgenic effects most probably due to its polyphenols content.

[poster presented at iMarCo congress, abstract from the Molecular Ecology related article] All metazoans are in fact holobionts, resulting from the association of several organisms, and organismal adaptation is then due to the composite response of this association to the environment. Deciphering the mechanisms of symbiont acquisition in a holobiont is therefore essential to understanding the extent of its adaptive capacities. In cnidarians, some species acquire their photosynthetic symbionts directly from their parents (vertical transmission) but may also acquire symbionts from the environment (horizontal acquisition) at the adult stage. The Mediterranean snakelocks sea anemone, Anemonia viridis (Forskål, 1775), passes down symbionts from one generation to the next by vertical transmission, but the capacity for such horizontal acquisition is still unexplored. To unravel the flexibility of the association between the different host lineages identified in A.viridis and its Symbiodiniaceae, we genotyped both the animal hosts and their symbiont communities in members of host clones in five different locations in the North Western Mediterranean Sea. The composition of within-host–symbiont populations was more dependent on the geographical origin of the hosts than their membership to a given lineage or even to a given clone. Additionally, similarities in host–symbiont communities were greater among genets (i.e. among different clones) than among ramets (i.e. among members of the same given clonal genotype). Taken together, our results demonstrate that A.viridis may form associations with a range of symbiotic dinoflagellates and suggest a capacity for horizontal acquisition. A mixed-mode transmission strategy in A.viridis, as we posit here, may help explain the large phenotypic plasticity that characterizes this anemone.

Diets high in saturated fat (HFD) disrupt dopamine neurotransmission, whereas fasting alters tonic and phasic dopamine release to drive motivation and food consumption. However, functional compartments in the nucleus accumbens (NAc) influencing these effects are not well characterized, and sex comparisons have not been made. This study sought to determine whether consumption of a HFD, sex, or being fed versus fasted altered baseline dopamine release and reuptake throughout NAc subregions. Male and female C57BL/6 mice were fed a control diet or nutrient matched HFD for six weeks. Ex-vivo fast-scan cyclic voltammetry revealed females had significantly slower dopamine reuptake in the NAc core than males when fed ad lib control diet. Fasting enhanced dopamine release and reuptake in the NAc core but not the medioventral shell. Further, being fasted versus fed significantly increased dopamine release throughout the NAc core in control males but specifically promoted release and reuptake in only the ventrolateral core of HF-fed males, effects which were lacking in females. Finally, fasting promoted dopamine release and reuptake in the rostral NAc core of controls and more caudally in HFD groups. These data support that dopamine neurotransmission is heterogeneous in NAc subregions and suggest the ventrolateral core is responsive to energy state. Furthermore, a rostrocaudal gradient in the NAc core might control valence responses to fasting that could promote overeating after chronic HFD consumption.

A 57-year-old man with a history of hypertension was admitted to the emergency department with septic shock and was therefore intubated and transferred to our intensive care unit (ICU). The patient had a supracondylar knee fracture 4 months ago, and underwent open reduction, internal fixation with

While commendable strides have been made in reducing smoking initiation and improving smoking cessation rates, current available smoking cessation treatment options are still only mildly efficacious and show substantial interindividual variability in their therapeutic responses. Therefore, the primary goal of preclinical research has been to further the understanding of the neural substrates and genetic influences involved in nicotine's effects and reassess potential drug targets. Pronounced advances have been made by investing in new translational approaches and placing more emphasis on bridging the gap between human and rodent models of dependence. Functional neuroimaging studies have identified key brain structures involved with nicotine-dependence phenotypes such as craving, impulsivity, withdrawal symptoms, and smoking cessation outcomes. Following up with these findings, rodent-modeling techniques have made it possible to dissect the neural circuits involved in these motivated behaviors and ascertain mechanisms underlying nicotine's interactive effects on brain structure and function. Likewise, translational studies investigating single-nucleotide polymorphisms (SNPs) within the cholinergic, dopaminergic, and opioid systems have found high levels of involvement of these neurotransmitter systems in regulating the reinforcing aspects of nicotine in both humans and mouse models. These findings and coordinated efforts between human and rodent studies pave the way for future work determining gene by drug interactions and tailoring treatment options to each individual smoker.

Recent evidence suggests that about 30%of patients with mild to moderate Alzheimer’s disease (AD) without a known diagnosis of epilepsy may display epileptiform spikes during electroencephalographic (EEG) recordings. These abnormal discharges occur predominantly during sleep and may be associated with accelerated disease progression. Subclinical spikes may represent a relevant target for clinical drug interventions, and there is a clear unmet need for preclinical testing of novel disease modifying agents in suitable animal models. Transgenic rodent models of AD pathology exhibit various forms of epileptiform EEG activity related to the abnormal levels of amyloid species in the brain. Among them, large-amplitude cortical and hippocampal EEG spikes in mouse and rat AD models may be reminiscent of the subclinical epileptiform EEG spikes recorded in some AD patients. This article reports the recommendations of a multidisciplinary panel of experts on optimal EEG markers and experimental designs to measure and report epileptiform activities and their response to symptomatic and disease-modifying drugs in transgenic AD model rodents. These recommendations may harmonize future preclinical EEG studies in the drug discovery research and may increase the comparability of experimental outcomes and their translational clinical value.

Background: Circular RNAs (circRNAs) have recently emerged as crucial regulatory molecules in prostate cancer (PCa), but few researches focus on the effects of circRNAs on PCa radiosensitivity. The issue will be addressed in this study using circRNA Cyclin B2 (circ_CCNB2) as an object. Materials and Methods: All RNA and protein levels were severally examined using quantitative real-time polymerase chain reaction and Western blot. Colony formation assay and flow cytometry were implemented for detecting cell colony capacity and apoptotic cells, respectively. Cellular migration and invasion abilities were evaluated by transwell assay. The combination between potential target molecules was analyzed by dual-luciferase reporter and RNA immunoprecipitation (RIP) assays. The effect of circ_CCNB2 on PCa radiosensitivity in vivo was explored using xenograft models in mice. Results: Circ_CCNB2 was upregulated in irradiation-resistant PCa tissues and cells. Circ_CCNB2 knockdown had promoted effect on the radiosensitivity of irradiation-resistant PCa cells by inhibiting autophagy. Besides, circ_CCNB2 could directly sponge miR-30b-5p, and the promotion of circ_CCNB2 knockdown on PCa radiosensitivity was achieved by elevating miR-30b-5p. MiR-30b-5p enhanced the radiosensitivity of irradiation-resistant PCa cells through repressing the expression of its target kinesin family member 18A (KIF18A). Furthermore, circ_CCNB2 regulated the KIF18A level through targeting miR-30b-5p. Circ_CCNB2 downregulation facilitated PCa radiosensitivity in vivo through inhibiting autophagy by miR-30b-5p/KIF18A. Conclusions: In this study, knockdown of circ_CCNB2 was shown to promote PCa radiosensitivity through autophagy repression by miR-30b-5p/KIF18A axis, developing a molecular resistance mechanism of PCa radiotherapy and a feasible strategy to increase radiosensitivity.

Abstract. Folate and vitamin B12(V-B12) deficiencies are associated with metabolic diseases that may impair memory function. We hypothesized that folate and V-B12 may differently alter mild cognitive impairment, glucose metabolism, and inflammation by modulating the gut microbiome in rats with Alzheimer’s disease (AD)-like dementia. The hypothesis was examined in hippocampal amyloid-β infused rats, and its mechanism was explored. Rats that received an amyloid-β(25–35) infusion into the CA1 region of the hippocampus were fed either control(2.5 mg folate plus 25 μg V-B12/kg diet; AD-CON, n = 10), no folate(0 folate plus 25 μg V-B12/kg diet; AD-FA, n = 10), no V-B12(2.5 mg folate plus 0 μg V-B12/kg diet; AD-V-B12, n = 10), or no folate plus no V-B12(0 mg folate plus 0 μg V-B12/kg diet; AD-FAB12, n = 10) in high-fat diets for 8 weeks. AD-FA and AD-VB12 exacerbated bone mineral loss in the lumbar spine and femur whereas AD-FA lowered lean body mass in the hip compared to AD-CON(P

Aims TASK-1 (K2P3.1) two-pore domain potassium channels are atrial-specific and significantly upregulated in atrial fibrillation (AF) patients, contributing to AF-related electrical remodelling. Inhibition of TASK-1 in cardiomyocytes of AF patients was shown to counteract AF-related action potential duration shortening. Doxapram was identified as a potent inhibitor of the TASK-1 channel. In the present study, we investigated the antiarrhythmic efficacy of doxapram in a porcine model of AF. Methods and results Doxapram successfully cardioverted pigs with artificially induced episodes of AF. We established a porcine model of persistent AF in domestic pigs via intermittent atrial burst stimulation using implanted pacemakers. All pigs underwent catheter-based electrophysiological investigations prior to and after 14 d of doxapram treatment. Pigs in the treatment group received intravenous administration of doxapram once per day. In doxapram-treated AF pigs, the AF burden was significantly reduced. After 14 d of treatment with doxapram, TASK-1 currents were still similar to values of sinus rhythm animals. Doxapram significantly suppressed AF episodes and normalized cellular electrophysiology by inhibition of the TASK-1 channel. Patch-clamp experiments on human atrial cardiomyocytes, isolated from patients with and without AF could reproduce the TASK-1 inhibitory effect of doxapram. Conclusions Repurposing doxapram might yield a promising new antiarrhythmic drug to treat AF in patients. Translational perspective Pharmacological suppression of atrial TASK 1 potassium currents prolongs atrial refractoriness with no effects on ventricular repolarization, resulting in atrial-specific class III antiarrhythmic effects. In our preclinical pilot study the respiratory stimulant doxapram was successfully administered for cardioversion of acute AF as well as rhythm control of persistent AF in a clinically relevant porcine animal model.

Long non-coding RNAs (lncRNAs) are a new class of regulatory RNAs that play important roles in disease development and a variety of biological processes. Recent studies have underscored the importance of lncRNAs in the circadian clock system and demonstrated that lncRNAs regulate core clock genes and the core clock machinery in mammals. In this review, we provide an overview of our current understanding of how lncRNAs regulate the circadian clock without coding a protein. We also offer additional insights into the challenges in understanding the functions of lncRNAs and other unresolved questions in the field. We do not cover other regulatory ncRNAs even though they also play important roles; readers are highly encouraged to refer to other excellent reviews on this topic. Accepted version

Circadian rhythms are ~24 h cycles of behavior and physiology that are generated by a network of molecular clocks located in nearly every tissue in the body. In mammals, the circadian system is organized hierarchically such that the suprachiasmatic nucleus (SCN) is the main circadian clock that receives light information from the eye and entrains to the light-dark cycle. The SCN then coordinates the timing of tissue clocks so internal rhythms are aligned with environmental cycles. Estrogens interact with the circadian system to regulate biological processes. At the molecular level, estrogens and circadian genes interact to regulate gene expression and cell biology. Estrogens also regulate circadian behavior across the estrous cycle. The timing of ovulation during the estrous cycle requires coincident estrogen and SCN signals. Studies using circadian gene reporter mice have also elucidated estrogen regulation of peripheral tissue clocks and metabolic rhythms. This review synthesizes current understanding of the interplay between estrogens and the circadian system, with a focus on female rodents, in regulating molecular, physiological, and behavioral processes.

The circadian system regulates behavior and physiology in many ways important for health. Circadian rhythms are expressed by nearly every cell in the body, and this large system is coordinated by a central clock in the suprachiasmatic nucleus (SCN). Sex differences in daily rhythms are evident in humans and understanding how circadian function is modulated by biological sex is an important goal. This review highlights work examining effects of sex and gonadal hormones on daily rhythms, with a focus on behavior and SCN circuitry in animal models commonly used in pre-clinical studies. Many questions remain in this area of the field, which would benefit from further work investigating this topic.

Genetically encoded biological clocks are found broadly throughout life on Earth, where they generate circadian (about a day) rhythms that synchronize physiology and behavior with the daily light/dark cycle. Although the genetic networks that give rise to circadian timing are now fairly well established, our understanding of how the proteins that constitute the molecular ‘cogs’ of this biological clock regulate the intrinsic timing, or period, of circadian rhythms has lagged behind. New studies probing the biochemical and structural basis of clock protein function are beginning to reveal how assemblies of dedicated clock proteins form and evolve through post-translational regulation to generate circadian rhythms. This review will highlight some recent advances providing important insight into the molecular mechanisms of period control in mammalian clocks with an emphasis on structural analyses related to CK1-dependent control of PER stability.

The present review has analyzed the scientific literature, available in the PubMed and Scopus databases, in order to summarize the current state of diet anthocyanin research in breast cancer (BC) and colorectal cancer (CRC) animal models but also for up-to-date human studies. For CRC, 28 preclinical and 9 clinical studies were selected in line with our search query in science databases. In relation to BC, 14 preclinical and 5 clinical studies were selected. Remarkably, all the preclinical studies, to a greater or lesser degree, suggested a chemoprevention effect of anthocyanin in BC/CRC rodent models. These encouraging results from animal models are not extrapolated to the same degree to human studies where, from the similar theoretical daily doses of anthocyanins in these studies, the opposite results were reported. Nevertheless, it is worth mentioning that the anthocyanin doses in the human studies carried out recently are low if we consider the estimated exposure to anthocyanins issued by the European Food Safety Agency (EFSA) or extremely low if we consider with caution the human equivalent dose based on body surface area from the preclinical dosage regimes used. Therefore, although some clinical data has demonstrated an inverse relation between anthocyanin consumption and BC/CRC, this could, in fact, be more relevant if we increase the daily human anthocyanin dose (as observed in animal model dose-effect studies) while new toxicological data for this flavonoid subtype are brought to light.

Behavioral states naturally alternate between wakefulness and the sleep phases rapid eye movement and nonrapid eye movement sleep. Waking and sleep states are complex processes that are elegantly orchestrated by spatially fine-tuned neurochemical changes of neurotransmitters and neuromodulators including glutamate, acetylcholine, γ-aminobutyric acid, norepinephrine, dopamine, serotonin, histamine, hypocretin, melanin concentrating hormone, adenosine, and melatonin. However, as highlighted in this brief overview, no single neurotransmitter or neuromodulator, but rather their complex interactions within organized neuronal ensembles, regulate waking and sleep states. The neurochemical pathways presented here are aimed to provide a conceptual framework for the understanding of the effects of currently used sleep medications.

Basophils are granulocytes involved in parasite immunity and allergic diseases, known for their potent secretion of type 2 cytokines. Identifying their functions has proven to be controversial due to their relative rarity and their complex lineage phenotype. Here, we show that the expression of basophils lineage markers CD200R3 and FcεRIα is highly variable in inflammatory settings and hinders basophils identification by flow cytometry across multiple disease states or tissues. Fluorophore-conjugated antibody staining of these lineage markers strongly activates basophil type 2 cytokine expression, and represents a potential bias for coculture or in vivo transfer experiments. The Basoph8 is a mouse model where basophils specifically express a strong fluorescent reporter and the Cre recombinase. Basophils can be identified and FACS sorted unambiguously by their expression of the enhanced yellow fluorescent protein (eYFP) in these mice. We show that the expression of the eYFP is robust in vivo during inflammation, and in vitro on living basophils for at least 72 h, including during the induction of anaphylactoid degranulation. We bred and characterized the Basoph8xiDTR mice, in which basophils specifically express eYFP and the simian diphtheria toxin receptor (DTR). This model enables basophils conditional depletion relatively specifically ex vivo and in vivo during allergic inflammation and their detection as eYFP+ cells. In conclusion, we report underappreciated benefits of the commercially available Basoph8 mice to study basophils function.

Slow-cycling/dormant cancer cells (SCCs) have pivotal roles in driving cancer relapse and drug resistance. A mechanistic explanation for cancer cell dormancy and therapeutic strategies targeting SCCs are necessary to improve patient prognosis, but are limited because of technical challenges to obtaining SCCs. Here, by applying proliferation-sensitive dyes and chemotherapeutics to non-small cell lung cancer (NSCLC) cell lines and patient-derived xenografts, we identified a distinct SCC subpopulation that resembled SCCs in patient tumors. These SCCs displayed major dormancy-like phenotypes and high survival capacity under hostile microenvironments through transcriptional upregulation of regulator of G protein signaling 2 (RGS2). Database analysis revealed RGS2 as a biomarker of retarded proliferation and poor prognosis in NSCLC. We showed that RGS2 caused prolonged translational arrest in SCCs through persistent eukaryotic initiation factor 2 (eIF2α) phosphorylation via proteasome-mediated degradation of activating transcription factor 4 (ATF4). Translational activation through RGS2 antagonism or the use of phosphodiesterase 5 inhibitors, including sildenafil (Viagra), promoted ER stress-induced apoptosis in SCCs in vitro and in vivo under stressed conditions, such as those induced by chemotherapy. Our results suggest that a low-dose chemotherapy and translation-instigating pharmacological intervention in combination is an effective strategy to prevent tumor progression in NSCLC patients after rigorous chemotherapy.

Understanding T cell function in vivo is of key importance for basic and translational immunology alike. To study T cells in vivo, we developed a new knock-in mouse line, which expresses a fusion protein of granzyme B, a key component of cytotoxic granules involved in T cell-mediated target cell-killing, and monomeric teal fluorescent protein from the endogenous Gzmb locus. Homozygous knock-ins, which are viable and fertile, have cytotoxic T lymphocytes with endogeneously fluorescent cytotoxic granules but wild-type-like killing capacity. Expression of the fluorescent fusion protein allows quantitative analyses of cytotoxic granule maturation, transport and fusion in vitro with super-resolution imaging techniques, and two-photon microscopy in living knock-ins enables the visualization of tissue rejection through individual target cell-killing events in vivo. Thus, the new mouse line is an ideal tool to study cytotoxic T lymphocyte biology and to optimize personalized immunotherapy in cancer treatment., eLife digest Cytotoxic, or killer, T cells are a key part of the immune system. They carry a lethal mixture of toxic chemicals, stored in packages called cytotoxic granules. Killer T cells inject the contents of these granules into infected, cancerous or otherwise foreign cells, forcing them to safely self-destruct. In test tubes, T cells are highly efficient serial killers, moving from one infected cell to the next at high speed. But, inside the body, their killing rate slows down. Researchers think that this has something to do with how killer T cells interact with other immune cells, but the details remain unclear. To get to grips with how killer T cells work in their natural environment, researchers need a way to follow them inside the body. One approach could be to use genetic engineering to attach a fluorescent tag to a protein found inside killer T cells. That tag then acts as a beacon, lighting the cells up and allowing researchers to track their movements. Tagging a protein inside the cytotoxic granules would allow close monitoring of T cells as they encounter, recognize and kill their targets. But fluorescent tags are bulky, and they can stop certain proteins from working as they should. To find out whether it is possible to track killer T cells with fluorescent tags, Chitirala, Chang et al. developed a new type of genetically modified mouse. The modification added a teal-colored tag to a protein inside the granules of the killer T cells. Chitirala, Chang et al. then used a combination of microscopy techniques inside and outside of the body to find out if the T cells still worked. This analysis showed that, not only were the tagged T cells able to kill diseased cells as normal, the tags made it possible to watch it happening in real time. Super-resolution microscopy outside of the body allowed Chitirala, Chang et al. to watch the killer T cells release their toxic granule content. It was also possible to follow individual T cells as they moved into, and destroyed, foreign tissue that had been transplanted inside the mice. These new mice provide a tool to understand how killer T cells really work. They could allow study not only of the cells themselves, but also their interactions with other immune cells inside the body. This could help to answer open questions in T cell research, such as why T cells seem to be so much more efficient at killing in test tubes than they are inside the body. Understanding this better could support the development of new treatments for viruses and cancer.

Ca(2+)-activated Cl(−) currents have been observed in many physiological processes, including sensory transduction in mammalian olfaction. The olfactory vomeronasal (or Jacobson's) organ (VNO) detects molecular cues originating from animals of the same species or from predators. It then triggers innate behaviors such as aggression, mating, or flight. In the VNO, Ca(2+)-activated Cl(−) channels (CaCCs) are thought to amplify the initial pheromone-evoked receptor potential by mediating a depolarizing Cl(−) efflux. Here, we confirmed the co-localization of the Ca(2+)-activated Cl(−) channels anoctamin 1 (Ano1, also called TMEM16A) and Ano2 (TMEM16B) in microvilli of apically and basally located vomeronasal sensory neurons (VSNs) and their absence in supporting cells of the VNO. Both channels were expressed as functional isoforms capable of giving rise to Ca(2+)-activated Cl(−) currents. Although these currents persisted in the VNOs of mice lacking Ano2, they were undetectable in olfactory neuron-specific Ano1 knockout mice irrespective of the presence of Ano2. The loss of Ca(2+)-activated Cl(−) currents resulted in diminished spontaneous and drastically reduced pheromone-evoked spiking of VSNs. Although this indicated an important role of anoctamin channels in VNO signal amplification, the lack of this amplification did not alter VNO-dependent male–male territorial aggression in olfactory Ano1/Ano2 double knockout mice. We conclude that Ano1 mediates the bulk of Ca(2+)-activated Cl(−) currents in the VNO and that Ano2 plays only a minor role. Furthermore, vomeronasal signal amplification by CaCCs appears to be dispensable for the detection of male-specific pheromones and for near-normal aggressive behavior in mice.

Background Isolation of neurons from the adult mouse CNS is important in order to study their gene expression during development or the course of different diseases. New methods Here we present two different methods for the enrichment or isolation of neurons from adult mouse CNS. These methods: are either based on flow cytometry sorting of eYFP expressing neurons, or by depletion of non-neuronal cells by sorting with magnetic-beads. Results Enrichment by FACS sorting of eYFP positive neurons results in a population of 62.4% NeuN positive living neurons. qPCR data shows a 3–5fold upregulation of neuronal markers. The isolation of neurons based on depletion of non-neuronal cells using the Miltenyi Neuron Isolation Kit, reaches a purity of up to 86.5%. qPCR data of these isolated neurons shows an increase in neuronal markers and an absence of glial markers, proving pure neuronal RNA isolation. Comparison with existing methods Former data related to neuronal gene expression are mainly based on histology, which does not allow for high-throughput transcriptome analysis to examine differential gene expression. Conclusion These protocols can be used to study cell type specific gene expression of neurons to unravel their function in the process of damage to the CNS.

In the central nervous system, the type I transmembrane glycoprotein NG2 (nerve-glia antigen 2) is only expressed by pericytes and oligodendrocyte precursor cells (OPCs). Therefore, OPCs are also termed NG2 glia. Their fate during development has been investigated systematically in several genetically modified mouse models. Consensus exists that postnatal NG2 glia are restricted to the oligodendrocyte (OL) lineage, while, at least in the forebrain, embryonic NG2 glia could also generate astrocytes. In addition, experimental evidence for a neurogenic potential of NG2 glia in the early embryonic brain (before E16.5) has been provided. However, this observation is still controversial. Here, we took advantage of reliable transgene expression in NG2-EYFP and NG2-CreERT2 knock-in mice to study the fate of early embryonic NG2 glia. While pericytes were the main cells with robust NG2 gene activity at E12.5, only a few OPCs expressed NG2 at this early stage of embryogenesis. Subsequently, this proportion of OPCs increased from 3% (E12.5) to 11% and 25% at E14.5 and E17.5, respectively. When Cre DNA recombinase activity was induced at E12.5 and E14.5 and pups were analyzed at postnatal day 0 (P0) and P10, the vast majority of recombined cells, besides pericytes, belonged to the OL lineage cells, with few astrocytes in the ventral forebrain. In other brain regions such as brain stem, cerebellum, and olfactory bulb only OL lineage cells were detected. Therefore, we conclude that NG2 glia from early embryonic brain are restricted to a gliogenic fate and do not differentiate into neurons after birth.

Influenza is a global challenge, and future pandemics of influenza are inevitable. One of the lessons learned from past pandemics is that all pandemic influenza viruses characterized to date possess viral genes originating from avian influenza viruses (AIVs). During the past decades, a wide range of AIVs have overcome the species barrier and infected humans with different clinical manifestations ranging from mild illness to severe disease and even death. Understanding the mechanisms of infection in the context of clinical outcomes, the mechanism of interspecies transmission, and the molecular determinants that confer interspecies transmission is important for pandemic preparedness. Here, we summarize the epidemiology, virology, and pathogenicity of human infections with AIVs to further our understanding of interspecies transmission.

BACKGROUND: L-DOPA, the predominant therapy for Parkinson’s disease (PD) is associated with motor deficits after prolonged use. The nigrostriatal tract, a primary target of neurodegeneration in PD, contains abundant Vitamin-D receptors, suggesting a potential role for VD in the disease. Therefore, we tested the impact of Vitamin D(3) (V(3)) in a mouse model of PD. METHODS: PD was induced in adult male C57BL6 mice by a single intrastriatal injection of 6-hydroxydopamine. Two weeks post lesion, these mice received injections of a vehicle, VD(3), L-DOPA, or a combination of VD(3)/L-DOPA and compared with sham controls. Treatment lasted three weeks, during which motor-cognitive neurobehaviour was assessed. Five weeks post lesion, brains were collected and striatal levels of the following proteins assessed: tyrosine hydroxylase (TH), dopamine decarboxylase (DDC), monoamine oxidase (MAO-B), Catechol-O-methyl transferase (COMT), dopamine transporter (DAT), brain-derived neurotrophic factor (BDNF), microglia marker (CD11b), inflammation (IL-1β), apoptotic signaling (BAX) and oxidative stress (p47phox). RESULTS: Treatment with VD(3) attenuated behavioural deficits induced by 6-OHDA, protein associated with dopamine metabolism and biomarkers of oxidative stress. VD(3) significantly increased contralateral wall touches, exploratory motor and cognitive activities. VD(3) significantly enhanced the expression of TH, DAT, BDNF, while significantly reducing expression of MAO-B, CD11b, IL-I β and p47phox. CONCLUSION: VD(3) reversed some of the 6-OHDA induced changes in proteins involved in modulating the dopamine system, behavioural deficits and oxidative stress biomarkers. The data suggests that VD(3) might be beneficial in reducing L-DOPA dosage, thereby reducing problems associated with dosage and prolonged use of L-DOPA in PD management.

MicroRNAs (miRNAs) crucially modulate fundamental biologic processes such as angiogenesis. In the present study, we focused on the molecular function of miRNA-370-3p (miR-370) in regulating the angiogenic activity of endothelial cells (ECs). Transfection with miR-370 mimic (miR-370m) significantly inhibited the sprouting of human dermal microvascular EC (HDMEC) and HUVEC spheroids and mouse aortic rings, whereas miR-370 inhibitor (miR-370i) promoted sprout formation. Additional in vitro assays demonstrated the pleiotropic inhibitory effects of miR-370m on HDMEC proliferation, migration, and tube formation. Moreover, Matrigel plugs containing miR-370m-transfected HDMECs exhibited a reduced microvessel density after implantation into CD1 nude mice when compared with controls. In contrast, miR-370i exerted proangiogenic effects. Mechanistic analyses revealed that miR-370 directly targets smoothened (SMO) and down-regulates bone morphogenetic protein (BMP)-2 expression in HDMECs. Accordingly, inhibition of SMO by cyclopamine reversed miR-370i-induced HDMEC proliferation and migration. In addition, BMP-2 treatment counteracted miR-370m-suppressed tube formation of HDMECs, whereas blockade of BMP-2 with neutralizing antibody significantly inhibited miR-370i-induced tube formation. Taken together, these novel findings indicate that miR-370 is a potent inhibitor of angiogenesis, which directly targets SMO and BMP-2.-Gu, Y., Becker, V., Zhao, Y., Menger, M. D., Laschke, M. W. miR-370 inhibits the angiogenic activity of endothelial cells by targeting smoothened (SMO) and bone morphogenetic protein (BMP)-2.

Pancreatic islets are highly vascularized endocrine units. Accordingly, their adequate revascularization is of major importance for successful islet transplantation. The proteoglycan, nerve/glial antigen 2 (NG2) expressed in pericytes is a crucial regulator of angiogenesis. Therefore, we herein analyze whether this surface protein contributes to the revascularization of grafted islets. Islets were isolated from NG2+/+ (wild-type) and NG2-/- mice and their cellular composition was analyzed by immunohistochemical detection of insulin, glucagon, somatostatin and CD31. Moreover, insulin secretion was assessed by enzyme-linked immunosorbent assay (ELISA). In addition, isolated islets were transplanted into dorsal skinfold chambers of wild-type mice and their revascularization was determined by intravital fluorescence microscopy and immunohistochemistry. NG2+/+ and NG2-/- islets did not differ in their cellular composition and insulin secretion. However, transplanted NG2-/- islets exhibited a significantly lower functional capillary density and a reduced number of CD31-positive microvessels. These findings demonstrate that the loss of NG2 impairs the revascularization of transplanted islets, underlining the importance of this pericytic proteoglycan for islet engraftment.

Traumatic peripheral nerve injuries are common and socioeconomically highly relevant. Despite significant microsurgical advances, the results of surgical reconstruction are still far from optimal and the rate of life-long complications, such as impaired motor and sensory function or neuropathic pain, is high. Moreover, the regeneration of peripheral nerves is a complex and fragile process that is not yet completely understood. Hence, there is an urgent need to further elucidate the underlying biological mechanisms. Herein, we propose that the neural lymphatic vasculature and lymphangiogenesis play an essential role in both peripheral nerve injury and regeneration and discuss hypothetical mechanisms implementing the current literature. Finally, specific research approaches to test our hypothesis are introduced.

International audience; Lysyl oxidases are major actors of microenvironment and extracellular matrix (ECM) remodeling. These cross-linking enzymes are thus involved in many aspects of physiopathology, including tumor progression, fibrosis and cardiovascular diseases. We have already shown that Lysyl Oxidase-Like 2 (LOXL2) regulates collagen IV deposition by endothelial cells and angiogenesis. We here provide evidence that LOXL2 also affects deposition of other ECM components, including fibronectin, thus altering structural and mechanical properties of the matrix generated by endothelial cells. LOXL2 interacts intracellularly and directly with collagen IV and fibronectin before incorporation into ECM fibrillar structures upon exocytosis, as demonstrated by TIRF time-lapse microscopy. Furthermore, surface plasmon resonance experiments using recombinant scavenger receptor cysteine-rich (SRCR) domains truncated for the catalytic domain demonstrated their direct binding to collagen IV. We thus used directed mutagenesis to investigate the role of LOXL2 catalytic domain. Neither enzyme activity nor catalytic domain were necessary for collagen IV deposition and angiogenesis, whereas the SRCR domains were effective for these processes. Finally, surface coating with recombinant SRCR domains restored deposition of collagen IV by LOXL2-depleted cells. We thus propose that LOXL2 SRCR domains orchestrate scaffolding of the vascular basement membrane and angiogenesis through interactions with collagen IV and fibronectin, independently of the enzymatic crosslinking activity.

When the brain is in a pathological state, the content of lipid droplets (LDs), the lipid storage organelles, is increased, particularly in glial cells, but rarely in neurons. The biology and mechanisms leading to LD accumulation in astrocytes, glial cells with key homeostatic functions, are poorly understood. We imaged fluorescently labeled LDs by microscopy in isolated and brain tissue rat astrocytes and in glia‐like cells in Drosophila brain to determine the (sub)cellular localization, mobility, and content of LDs under various stress conditions characteristic for brain pathologies. LDs exhibited confined mobility proximal to mitochondria and endoplasmic reticulum that was attenuated by metabolic stress and by increased intracellular Ca2+, likely to enhance the LD–organelle interaction imaged by electron microscopy. When de novo biogenesis of LDs was attenuated by inhibition of DGAT1 and DGAT2 enzymes, the astrocyte cell number was reduced by ~40%, suggesting that in astrocytes LD turnover is important for cell survival and/or proliferative cycle. Exposure to noradrenaline, a brain stress response system neuromodulator, and metabolic and hypoxic stress strongly facilitated LD accumulation in astrocytes. The observed response of stressed astrocytes may be viewed as a support for energy provision, but also to be neuroprotective against the stress‐induced lipotoxicity., Main points Astroglial lipid droplets are dynamic organelles with confined mobility.Lipid droplet turnover is important for astrocyte survival/proliferation.Stressed astrocytes accumulate lipid droplets likely to provide energy and reduce lipotoxicity.

Influenza A viruses (IAVs) are the causative agents of one of the most important viral respiratory diseases in pigs and humans. Human and swine IAV are prone to interspecies transmission, leading to regular incursions from human to pig and vice versa. This bidirectional transmission of IAV has heavily influenced the evolutionary history of IAV in both species. Transmission of distinct human seasonal lineages to pigs, followed by sustained within-host transmission and rapid adaptation and evolution, represent a considerable challenge for pig health and production. Consequently, although only subtypes of H1N1, H1N2, and H3N2 are endemic in swine around the world, extensive diversity can be found in the hemagglutinin (HA) and neuraminidase (NA) genes, as well as the remaining six genes. We review the complicated global epidemiology of IAV in swine and the inextricably entangled implications for public health and influenza pandemic planning.

Decades of experimental and theoretical work support a now well-established theory that active dendritic processing contributes to the computational power of individual neurons. This theory is based on the high degree of electrical compartmentalization observed in the dendrites of single neurons in ex vivo preparations. Compartmentalization allows dendrites to conduct semi-independent operations on their inputs before final integration and output at the axon, producing a "network-in-a-neuron." However, recent in vivo functional imaging experiments in mouse cortex have reported surprisingly little evidence for strong dendritic compartmentalization. In this review, we contextualize these new findings and discuss their impact on the future of the field. Specifically, we consider how highly coordinated, and thus less compartmentalized, activity in soma and dendrites can contribute to cortical computations including nonlinear mixed selectivity, prediction/expectation, multiplexing, and credit assignment.

Background Nitrite stimulates insulin secretion from pancreatic β-cells; however, the underlying mechanisms have not been completely addressed. The aim of this study is to determine effect of nitrite on gene expression of SNARE proteins involved in insulin secretion from isolated pancreatic islets in Type 2 diabetic Wistar rats. Material and methods Three groups of rats were studied (n = 10/group): Control, diabetes, and diabetes + nitrite, which treated with sodium nitrite (50 mg/L) for 8 weeks. Type 2 diabetes was induced using a low-dose of streptozotocin (25 mg/kg) combined with high-fat diet. At the end of the study, pancreatic islets were isolated and mRNA expressions of interested genes were measured; in addition, protein expression of proinsulin and C-peptide in pancreatic tissue was assessed using immunofluorescence staining. Results Compared with controls, in the isolated pancreatic islets of Type 2 diabetic rats, mRNA expression of glucokinase (59%), syntaxin1A (49%), SNAP25 (70%), Munc18b (48%), insulin1 (56%), and insulin2 (52%) as well as protein expression of proinsulin and C-peptide were lower. In diabetic rats, nitrite administration significantly increased gene expression of glucokinase, synaptotagmin III, syntaxin1A, SNAP25, Munc18b, and insulin genes as well as increased protein expression of proinsulin and C-peptide. Conclusions Stimulatory effect of nitrite on insulin secretion in Type 2 diabetic rats is at least in part due to increased gene expression of molecules involved in glucose sensing (glucokinase), calcium sensing (synaptotagmin III), and exocytosis of insulin vesicles (syntaxin1A, SNAP25, and Munc18b) as well as increased expression of insulin genes.

Brain EGR1 (early growth response protein 1) overexpression aggravates focal ischemic brain injury, but its role in intracerebral hemorrhage (ICH) induced cerebral injury remains obscure. In this study, a rat ICH model was established by injecting type VII collagenase into the brain, and EGR1 knockdown reversed the increase of hematoma area, neurological function score, brain water content, blood–brain barrier (BBB) permeability, inflammation, p300 and retinoid a X receptor-α (RXRα) protein levels, as well as RXRα acetylation level induced by ICH. EGR1 expression was up-regulated in primary brain microvascular endothelial cells (BMECs), neurons, and astrocytes after ICH induction, and the up-regulation was most significant in BMECs. We also found that EGR1 promoted RXRα acetylation level by regulating p300 in BMECs. Subsequently, the BMECs were treated with OGD (oxygen glucose deprivation) plus hemin to simulate ICH condition. And silencing EGR1 rescued the upregulation of cell inflammation and the reduction of cell viability and TEER (transendothelial electric resistance) caused by OGD plus hemin via p300-mediated RXRα acetylation. Furthermore, the STAT3/NF-κB pathway was activated after treatment with OGD plus hemin, which was suppressed by silencing EGR1. And treatment with Stattic (an inhibitor of STAT3) restrained the effect of OGD plus hemin on NF-κB pathway activity, inflammation, cell viability and TEER. In conclusion, EGR1 increased RXRα acetylation level by regulating p300, thereby aggravating brain damage in ICH rat model and dysfunction in BMECs, which might through the STAT3/NF-κB pathway.

The hippocampal formation is critically involved in learning and memory and contains a large proportion of neurons encoding aspects of the organism’s spatial surroundings. In the medial entorhinal cortex (MEC), this includes grid cells with their distinctive hexagonal firing fields as well as a host of other functionally defined cell types including head direction cells, speed cells, border cells, and object-vector cells. Such spatial coding emerges from the processing of external inputs by local microcircuits. However, it remains unclear exactly how local microcircuits and their dynamics within the MEC contribute to spatial discharge patterns. In this review we focus on recent investigations of intrinsic MEC connectivity, which have started to describe and quantify both excitatory and inhibitory wiring in the superficial layers of the MEC. Although the picture is far from complete, it appears that these layers contain robust recurrent connectivity that could sustain the attractor dynamics posited to underlie grid pattern formation. These findings pave the way to a deeper understanding of the mechanisms underlying spatial navigation and memory.

AIMS: Atherosclerotic plaque hypoxia is detrimental for macrophage function. Prolyl hydroxylases (PHDs) initiate cellular hypoxic responses, possibly influencing macrophage function in plaque hypoxia. Thus, we aimed to elucidate the role of myeloid PHDs in atherosclerosis.METHODS & RESULTS: Myeloid specific PHD knockout (PHDko) mice were obtained via bone marrow transplantation (PHD1ko, PHD3ko) or conditional knockdown through lysozyme M-driven Cre recombinase (PHD2cko). Mice were fed high cholesterol diet for 6-12 weeks to induce atherosclerosis. Aortic root plaque size was significantly augmented 2.6-fold in PHD2cko, and 1.4-fold in PHD3ko compared to controls, but was unchanged in PHD1ko mice. Macrophage apoptosis was promoted in PHD2cko and PHD3ko mice in vitro and in vivo, via the HIF1α/BNIP3 axis. Bulk and single cell RNA data of PHD2cko bone-marrow-derived macrophages (BMDM) and plaque macrophages, respectively, showed enhanced HIF1α/BNIP3 signaling, which was validated in vitro by siRNA silencing. Human plaque BNIP3 mRNA was positively associated with plaque necrotic core size, suggesting similar pro-apoptotic effects in human. Further, PHD2cko plaques displayed enhanced fibrosis, while macrophage collagen breakdown by matrix metalloproteinases, collagen production and proliferation were unaltered. Instead, PHD2cko BMDMs enhanced fibroblast collagen secretion in a paracrine manner. In silico analysis of macrophage-fibroblast communication predicted SPP1 (osteopontin) signaling as regulator, which was corroborated by enhanced plaque SPP1 protein in vivo. Increased SPP1 mRNA expression upon PHD2cko was preferentially observed in foamy plaque macrophages expressing "triggering receptor expressed on myeloid cells-2" (TREM2hi) evidenced by single-cell RNA, but not in neutrophils. This confirmed enhanced fibrotic signaling by PHD2cko macrophages to fibroblasts, in vitro as well as in vivo.CONCLUSION: Myeloid PHD2cko and PHD3ko enhanced atherosclerotic plaque growth and macrophage apoptosis, while PHD2cko macrophages further activated collagen secretion by fibroblasts in vitro, likely via paracrine SPP1 signaling through TREM2hi macrophages.TRANSLATIONAL OUTLOOK: This study shows that myeloid PHD isoforms PHD2 and PHD3 worsen plaque characteristics and phenotype, such as plaque size, macrophage accumulation, apoptosis, and collagen accumulation in mice. We show both direct effects on macrophages and paracrine effects of macrophage PHD2 loss on vessel wall fibroblast populations. Broad spectrum-PHD inhibitors, e.g. Roxadustat, are currently being prescribed to chronic kidney disease patients, who are already at risk for cardiovascular disease. When considering this study and the pro-fibrotic and pro-apoptotic effects we report, broad PHD inhibition may therefore be sub-optimal and more targeted PHD inhibition of PHD1 should be considered.

Recombinant factor VIII (FVIII) products represent a life-saving intervention for patients with hemophilia A. However, patients can develop antibodies against FVIII that prevent its function and directly increase morbidity and mortality. The development of anti-FVIII antibodies varies depending on the type of recombinant product used, with previous studies suggesting that second-generation baby hamster kidney (BHK)-derived FVIII products display greater immunogenicity than do third-generation Chinese hamster ovary (CHO)-derived FVIII products. However, the underlying mechanisms responsible for these differences remain incompletely understood. Our results demonstrate that BHK cells express higher levels of the nonhuman carbohydrate α1-3 galactose (αGal) than do CHO cells, suggesting that αGal incorporation onto FVIII may result in anti-αGal antibody recognition that could positively influence the development of anti-FVIII antibodies. Consistent with this, BHK-derived FVIII exhibits increased levels of αGal, which corresponds to increased reactivity with anti-αGal antibodies. Infusion of BHK-derived, but not CHO-derived, FVIII into αGal–knockout mice, which spontaneously generate anti-αGal antibodies, results in significantly higher anti-FVIII antibody formation, suggesting that the increased levels of αGal on BHK-derived FVIII can influence immunogenicity. These results suggest that posttranslational modifications of recombinant FVIII products with nonhuman carbohydrates may influence the development of anti-FVIII antibodies.

Introduction: Aminoglycoside-induced acute kidney injury (AKI) is a pathology closely linked to oxidative and inflammatory reactions. Taking into account the previous reported antioxidant and anti-inflammatory effects of D-005, a lipid extract obtained from Cuban palm Acrocomia crispa (Arecaceae) fruits, this work aimed to evaluate the effects of D-005 on kanamycin-induced AKI. Methods: Male Wistar rats were divided into 7 groups: negative control (vehicle, Tween 65/H2O) and six groups treated with kanamycin to induce AKI: positive control (vehicle), D-005 (25, 100, 200, and 400 mg/kg) and grape seed extract (GSE, 200 mg/kg). D-005, vehicle, and GSE oral treatments were administered once daily for seven days, 1 h before kanamycin (500 mg/kg, i.p.). Serum uric acid and urea concentrations, renal histopathology, and oxidative markers (malondialdehyde (MDA), sulfhydryl (SH) groups, and catalase (CAT) activity) were assessed. Results: D-005 significantly reduced uric acid and urea levels, starting from D-005 100 mg/kg. Histopathologically, D-005, at all the tested doses, protected renal parenchyma structures (glomeruli, proximal tubules, and interstitium). These findings were accompanied by a significant reduction of MDA and SH group concentrations as well as restoration of CAT activity. The highest percentages of inhibition were obtained with the dose of 400 mg/kg. GSE, the reference substance, also prevented kanamycin-induced biochemical and histopathological changes, as well as reduced MDA and SH groups and restored CAT activity. Conclusion: The administration of repeated oral doses of D-005 significantly protected against kanamycin-induced AKI, which could be associated with the antioxidant and anti-inflammatory effects of this extract.

Aims: Nonsteroidal anti-inflammatory drugs, including ketoprofen, induce adverse effects within the gastrointestinal (GI)-tract. Hydrogen sulfide (H2S) is an antioxidative gaseous mediator contribu...

Background Obstructive sleep apnea, characterized by sleep fragmentation and chronic intermittent hypoxia (CIH), is a risk factor for Alzheimer’s disease (AD) progression. Recent epidemiological studies point to CIH as the best predictor of developing cognitive decline and AD in older adults with obstructive sleep apnea. However, the precise underlying mechanisms remain unknown. This study was undertaken to evaluate the effect of CIH on pathological human tau seeding, propagation, and accumulation; cognition; synaptic plasticity; neuronal network excitability; and gene expression profiles in a P301S human mutant tau mouse model of AD and related tauopathies. Methods We exposed 4- to 4.5-month-old male P301S and wild-type mice to an 8-week CIH protocol (6-min cycle: 21% O2 to 8% O2 to 21% O2, 80 cycles per 8 hours during daytime) and assessed its effect on tau pathology and various AD-related phenotypic and molecular signatures. Age- and sex-matched P301S and wild-type mice were reared in normoxia (21% O2) as experimental controls. Results CIH significantly enhanced pathological human tau seeding and spread across connected brain circuitry in P301S mice; it also increased phosphorylated tau load. CIH also exacerbated memory and synaptic plasticity deficits in P301S mice. However, CIH had no effect on seizure susceptibility and network hyperexcitability in these mice. Finally, CIH exacerbated AD-related pathogenic molecular signaling in P301S mice. Conclusions CIH-induced increase in pathologic human tau seeding and spread and exacerbation of other AD-related impairments provide new insights into the role of CIH and obstructive sleep apnea in AD pathogenesis.

Graphical abstract, Highlights • PLA2G16 is up-regulated in CRC, and high expression of PLA2G16 is associated with the advanced stages. • PLA2G16 promotes the malignant progression of CRC through the Hippo signaling pathway. • GCK exerts its anti-CRC effects by inhibiting the protein expression of PLA2G16. • Provide a new insights towards the development of effective therapeutic strategies for CRC treatment by targeting PLA2G16., Introduction Colorectal cancer (CRC) is a common malignant tumor with a high global incidence, metastasis rate and low cure rate. Changes in lipid metabolism-related genes can affect the occurrence and development of CRC, and may be a potential therapeutic target for CRC. Therefore, starting from lipid metabolism-related genes to find natural medicines for tumor treatment may become a new direction in CRC research. Objectives This study aimed to investigate the effect of PLA2G16, a key gene involved in lipid metabolism, on the biological function of CRC, and whether the anti-CRC effect of GCK is related to PLA2G16. Methods To explore the role of PLA2G16 in CRC in vitro and in vivo, we performed cell proliferation, migration, invasion and nude mice tumorigenesis assays. As for the mechanism, we designed RNA-seq analysis and verified by western blotting and immunofluorescence experiments. Subsequently, we found the anti-CRC effect of GCK is related to PLA2G16 through western blotting and rescue experiments. Results We showed that PLA2G16 was significantly higher in CRC tissues than the adjacent normal appearing tissues, and high PLA2G16 expression correlates with unfavorable prognosis of CRC patients. Further, PLA2G16 promoted the malignant progression of CRC by inhibiting the Hippo signaling pathway determined by RNA-seq analysis, and GCK exerted anti-CRC effects by inhibiting the protein expression of PLA2G16 in vitro and in vivo. Conclusion Our results suggested that PLA2G16 promote the malignant progression of CRC by inhibiting the Hippo signaling pathway and the anti-CRC effect of GCK is through inhibiting the protein expression of PLA2G16.

China banned the use of colistin as animal growth promoter in April 2017. Herein, we report the prevalence of mcr-1 in the intestine of healthy humans and risk factors associated with mcr-1 carriage after the implementation of the ban.We recruited 719 healthy volunteers from Shenzhen City from 1 March 2018 to 31 December 2019 to investigate the prevalence of mcr-1 in human intestine, and undertook a case-control study to ascertain the risk factors associated with the mcr-1-positive population. A further comparative study was conducted to identify differences between genetic characteristics of mcr-1-positive and mcr-1-negative Escherichia coli.Overall, 56 (7.8%, 95% CI 5.9%-10.0%, n = 719) individual faecal samples were positive for mcr-1, and prevalence of mcr-1 among individuals in 2019 (2.4%, 95% CI 8.7%-15.0%, 7/294) was significantly lower than that in 2018 (11.5%, 95% CI 1.0%-4.8%, 49/425) (p 0.0001). After the colistin ban, animal-derived food (pork and chicken meat) was no longer a risk factor for mcr-1 carriage in human intestine, whereas a higher intake of fish and seafood (75 g/day) and whole grains (150 g/day) was associated with higher and lower risk of mcr-1 carriage, respectively (OR 2.175, 95% CI 1.047-4.517; OR 0.045, 95% CI 0.004-0.567). Compared with mcr-1-negative E. coli, the mcr-1-positive E. coli had different patterns of resistance genes and genetic heterogeneity.Our study implicates aquatic food as beeing associated with mcr-1 carriage in the healthy population, even after the ban on colistin. Dietary modification (e.g. whole grains) may help to combat mcr-1-positive bacterial colonization of the gut.

Cardiovascular diseases are a common cause of death worldwide. Adult cardiomyocytes have limited regenerative capacity after injury, and there is growing interest in cardiac regeneration as a new therapeutic strategy. There are several limitations of induced pluripotent stem cell-based transplantation therapy with respect to efficiency and risks of tumorigenesis. Direct reprogramming enables the conversion of terminally differentiated cells into target cell types using defined factors. In most cardiac diseases, activated fibroblasts proliferate in the damaged heart and contribute to the progression of heart failure. In vivo cardiac reprogramming, in which resident cardiac fibroblasts are converted into cardiomyocytes in situ, is expected to become a new cardiac regenerative therapy. Indeed, we and other groups have demonstrated that in vivo reprogramming improves cardiac function and reduces fibrosis after myocardial infarction. In this review, we summarize recent discoveries and developments related to in vivo reprogramming. In addition, issues that need to be resolved for clinical application are described.

Background: FimH adhesin is proposed to enhance Escherichia coli kidney infection by acting with PapGII adhesin, but genetic epidemiology study and animal study have not been widely conducted to confirm this hypothesis. Methods: We compared the prevalence of adhesin gene and their coexistent pattern between upper and lower urinary tract infection (UTI) strains. fimH mutant (EC114FM), papGII mutant (EC114PM) and fimH/papGII double mutant (EC114DM) were constructed from a pylonephritogenic strain (EC114). We compared among these strains for the infection ability in bladders and kidneys of female BALB/c mice challenged transurethrally with these bacteria and assessed 1, 3, and 7 days after inoculation. Results: Strains carrying fimH-only genotype were significantly more prevalent in lower UTI (P

Tumor-associated macrophages (TAM) play a crucial role in promoting cancer progression. Upon cytokine stimulation, TAM preferentially polarize to the anti-inflammatory and pro-tumor M2 subtype. The mechanism underlying such preferential polarization remains elusive. Here, we report that macrophage-specific deletion of the SUMO-specific protease Sentrin/SUMO-specific protease 3 promotes macrophage polarization towards M2 in bone marrow-derived macrophage (BMDM) induced by interleukin 4 (IL-4)/IL-13 and in an ex vivo model (murine Py8119 cell line), as well as in a mouse orthotopic tumor model. Notably, Sentrin/SUMO-specific protease 3 (SENP3) loss in macrophages accelerated breast cancer malignancy in ex vivo and in vivo models. Mechanistically, we identified Akt Serine/threonine kinase 1 (Akt1) as the substrate of SENP3 and found that the enhanced Akt1 SUMOylation upon SENP3 loss resulted in Akt1 hyper-phosphorylation and activation, which facilitates M2 polarization. Analysis of clinical data showed that a lower level of SENP3 in TAM has a strong negative correlation with the level of the M2 marker CD206, as well as with a worse clinical outcome. Thus, increased Akt1 SUMOylation as a result of SENP3 deficiency modulates polarization of macrophages to the M2 subtype within a breast cancer microenvironment, which in turn promotes tumor progression.