Nafamostat reduces systemic inflammation in TLR7-mediated virus-like illness

Background The serine protease inhibitor nafamostat has been proposed as a treatment for COVID-19, by inhibiting TMPRSS2-mediated viral cell entry. Nafamostat has been shown to have other, immunomodulatory effects, which may be beneficial for treatment, however animal models of ssRNA virus infection are lacking. In this study, we examined the potential of the dual TLR7/8 agonist R848 to mimic the host response to an ssRNA virus infection and the associated behavioural response. In addition, we evaluated the anti-inflammatory effects of nafamostat in this model. Methods CD-1 mice received an intraperitoneal injection of R848 (200 μg, prepared in DMSO, diluted 1:10 in saline) or diluted DMSO alone, and an intravenous injection of either nafamostat (100 μL, 3 mg/kg in 5% dextrose) or 5% dextrose alone. Sickness behaviour was determined by temperature, food intake, sucrose preference test, open field and forced swim test. Blood and fresh liver, lung and brain were collected 6 h post-challenge to measure markers of peripheral and central inflammation by blood analysis, immunohistochemistry and qPCR. Results R848 induced a robust inflammatory response, as evidenced by increased expression of TNF, IFN-γ, CXCL1 and CXCL10 in the liver, lung and brain, as well as a sickness behaviour phenotype. Exogenous administration of nafamostat suppressed the hepatic inflammatory response, significantly reducing TNF and IFN-γ expression, but had no effect on lung or brain cytokine production. R848 administration depleted circulating leukocytes, which was restored by nafamostat treatment. Conclusions Our data indicate that R848 administration provides a useful model of ssRNA virus infection, which induces inflammation in the periphery and CNS, and virus infection-like illness. In turn, we show that nafamostat has a systemic anti-inflammatory effect in the presence of the TLR7/8 agonist. Therefore, the results indicate that nafamostat has anti-inflammatory actions, beyond its ability to inhibit TMPRSS2, that might potentiate its anti-viral actions in pathologies such as COVID-19.

Nafamostat reduces systemic inflammation in TLR7-mediated virus-like illness

Background: The serine protease inhibitor nafamostat has been proposed as a treatment for COVID-19, by inhibiting TMPRSS2-mediated viral cell entry. Nafamostat has been shown to have other, immunomodulatory effects, which may be beneficial for treatment, however animal models of ssRNA virus infection are lacking. In this study, we examined the potential of the dual TLR7/8 agonist R848 to mimic the host response to a ssRNA virus infection and the associated behavioural response. In addition, we evaluated the anti-inflammatory effects of nafamostat in this model. Methods: CD-1 mice received an intraperitoneal injection of R848 (200μg, prepared in DMSO, diluted 1:10 in saline) or diluted DMSO alone, and an intravenous injection of either nafamostat (100μL, 3mg/kg in saline) or saline. Sickness behaviour was determined by temperature, food intake, sucrose preference test, open field and forced swim test. Blood and fresh liver, lung and brain were collected 6 hours post-challenge to measure markers of peripheral and central inflammation by blood analysis and qPCR. Results: R848 induced a robust inflammatory response, as evidenced by increased expression of TNF, IFN-γ, CXCL1 and CXCL10 in the liver, lung and brain, as well as a sickness behaviour phenotype. Exogenous administration of nafamostat suppressed the hepatic inflammatory response, significantly reducing TNF and IFN-γ expression, but had no effect on lung or brain cytokine production. R848 administration depleted circulating leukocytes, which was restored by nafamostat treatment. Conclusions: Our data indicate that R848 administration provides a useful model of ssRNA virus infection, which induces inflammation in the periphery and CNS, and virus infection-like illness. In turn, we show that nafamostat has a systemic anti-inflammatory effect, in the presence of the TLR7/8 agonist. Therefore, the results indicate that nafamostat has anti-inflammatory actions, beyond its ability to inhibit TMPRSS2, that might potentiate its anti-viral actions in pathologies such as COVID-19.

Inflammatory mechanisms of mental illness: brain inflammatory response to interferon stimulation

AimsWe hypothesise that peripheral IFN stimulation results in a brain inflammatory response via pathways of neuroimmune communication which in turn results in sickness-behaviour and depressive phenotype. We aim to determine if peripheral IFN stimulation results in brain inflammatory response including upregulation of inflammatory cytokines and chemokines.BackgroundThere is increasing interest in the role of dysregulated immune function and inflammation in the pathogenesis of psychiatric disorders including mood disorders and dementias. Immune mechanisms offer a new approach to investigating mechanism in addition to offering hope for new avenues of treatment.Interferon (IFN) therapy in humans is known to be associated with a significant risk of developing depression, both during therapy and increasing risk of relapse in the years following exposure, yet the mechanism remains unclear. IFN stimulation in animal models may offer insights into this phenomenon, in addition to furthering our understanding the role of immune mechanisms in the development of psychiatric phenotypes.MethodMice (n. 42) were exposed to either IFN-alpha, IFN-gamma or vehicle control using either osmotic pump or intraperitoneal injection over the course of 7 days. Mice were scarificed, brains were dissected and RNA extracted. Inflammatory gene transcription within the brain was determined using real time quantitative polymerase chain reaction (RTqPCR). Absolute quantification was achieved using standard curves and reference gene. Statistical significance was determined using Mann-Whitney or ANOVA/Kruskal-Wallis depending on normality of data and number of groups.ResultIFNγ stimulation is associated with a significant brain upregulation of a number of inflammatory cytokines and chemokines including Il1β, Tnfα, Il10, Ifnγ, Ccl2, Ccl5, Ccl19, Cxcl10 and Ccr5. However, unexpectedly we did not find IFNα stimulation to associate with brain inflammatory transcriptional changes.ConclusionThis work demonstrates a brain inflammatory response to peripheral IFNγ stimulation. The inflammatory profile, including upregulated chemokines, suggests that recruitment of leukocytes across the blood brain barrier may be part of the immune response. Further experiments using existing tissues will explore if there are structural/cellular changes within the brain parenchyma. Further experiments within the group will seek to demonstrate if IFN treatment associates with sickness behaviour in order to determine if this is a clinically meaningful model. Suprisingly, we did not see similar changes in the IFNα treated group, which requires further investigation.Funding: University of Glasgow, The Sackler Trust

Sick and Tired: Polyparasitism, Food Stress and Sickness Behaviour in a Gregarious Mammal

Although sickness behaviour in response to non-lethal parasites has been documented in wild animals, it remains unclear how social and environmental stress might also shape an animal’s behavioural response to parasitism, nor do we know whether simultaneous infection with more than one parasite changes the way animals respond. Here, we combine physiological, environmental, behavioural and parasite measures to investigate behavioural responses to infection in wild vervet monkeys (Chlorocebus pygerythrus) living in a semi-arid region of South Africa. We quantified both activity budget and behavioural predictability to investigate the occurrence of sickness behaviour and infection with two non-lethal gastrointestinal parasite genera. Higher parasite load was linked to an increase in the time spent resting. However, the nature of the relationship with other behaviours was contingent on both the parasite genus in question, and how parasite species interacted, highlighting the importance of considering co-infection. Overall, food availability was the dominant predictor of behavioural change suggesting that, for monkeys living in a more extreme environment, coping with ecological stress may override the ability to modulate behaviour in response to other physiological stressors. Our findings provide insight into how animals living in harsh environments find ways to cope with parasite infection, avoidance, and transmission.

Wechselwirkungen zwischen neuroendokrinem System und Immunsystem bei chronisch-entzündlichen Systemerkrankungen

ZusammenfassungHormonelle und neuronale Signalwege können die Manifestation einer chronisch-entzündlichen Systemerkrankung entweder begünstigen oder verhindern. Bei bereits manifester Erkrankung modulieren Hormone und Neurotransmitter den Krankheitsverlauf, in dem sie die Krankheitsaktivität erhöhen oder abschwächen. Beispiele hierfür sind der entzündungshemmende Einfluss der körpereigenen und exogenen Glukokortikoide und die entzündungsfördernden Effekte von Stress bei chronisch-entzündlichen Systemerkrankungen. Bei chronisch-entzündlichen Systemerkrankungen ringt das aktivierte Immunsystem mit dem Gehirn und anderen Organsystemen um Energie, was zu vielfältigen Erkrankungsfolgen und Folgeerkrankungen führt: „sickness behaviour“ mit Fatigue-Symptomatik und depressiven Symptomen, Schlafstörungen, Anorexie, Fehl- und Mangelernährung, Knochenabbau, Muskelabbau und kachektische Fettsucht, Insulinresistenz mit Hyperinsulinämie (begleitet von einer Resistenz gegenüber dem Insulin-like growth factor 1), Dyslipidämie, Veränderungen der Steroidhormonachsen, Störungen der Hypothalamus-Hypophysen-Gonaden-Achse, erhöhter Sympathikotonus, herabgesetzte Aktivität des parasympathischen Nervensystems, arterielle Hypertonie und Volumenbelastung, Entzündungsanämie und zirkadiane Rhythmik der Symptomausprägung. Diese für die Patienten gravierenden Folgeerkrankungen, welche den chronisch-entzündlichen Systemerkrankungen inhärent sind, sollten konsequent therapiert werden.

Sexual maturity as risk for development of deviant behaviours in pig production systems with entire males

Male piglets are castrated primarily to avoid the unpleasant boar taint in meat, and additionally for the predisposition of castrates to accumulate fat and for their lower risk of developing unwanted behaviours. There are two main strategies available for withdrawing from surgical castration: one is immunocastration and the other is to raise entire male pigs or boars. Additionally, raising intact boars is more profitable because of the production of carcasses with lean meat and better feed conversion. Boars (compared to castrates) exhibit more aggressive, sexual, damaging social behaviour and reduced feeding behaviour with a lower prevalence of sickness behaviour as a result of good health and low susceptibility to chronic inflammation. In this review, the behaviours specific for boars as a result of sexual maturity are reviewed, with an overview of differences in the behaviour of surgically castrated barrows, immunocastrates and boars reared in group-housed systems. The raising of boars allows for good welfare of these animals in early life, but later, on reaching sexual maturity, the welfare of boars can be diminished because of their propensity to aggression and more mounting behaviour than castrates. Innovations in the breeding and management of boars are needed to improve their performance and to reduce welfare implications of these animals raised in social groups, and in particular to minimize deviant behaviours towards pen mates.