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Neonatal infection with neurotropic influenza A virus induces the kynurenine pathway in early life and disrupts sensorimotor gating in adult Tap1 −/− mice

Epidemiological studies suggest that early life infections may contribute to the development of neuropsychiatric disorders later in life. Experimental studies employing infections during neonatal life support this notion by reporting persistent changes in the behaviour of adult animals, including deficits in sensorimotor gating. We have previously described an induction of the kynurenine pathway in neonatal wild-type (WT) mice following a systemic infection with neurotropic influenza A/WSN/33 virus. Here, we use the same model of infection in both WT and Tap1-/- mice (expressing reduced levels of MHC class I) and study long-term effects of the infection on sensorimotor gating, as determined by measuring prepulse inhibition (PPI). Moreover, transcription of genes encoding enzymes in the kynurenine pathway and levels of kynurenic acid (KYNA), in the brain of Tap1-/- mice were investigated. In mice infected on postnatal day (P)3 or P4, the levels of several transcripts in the kynurenine pathway were altered at P7, P13 and P24. Transcripts encoding indoleamine-pyrrole 2,3-dioxygenase (IDO), degrading tryptophan in the first step of the kynurenine pathway were consistently up-regulated at all time-points investigated. The changes in transcript levels were accompanied by a transient elevation of KYNA in the brain of infected mice at P13. At age 5-6 months, neonatally infected Tap1-/-, but not WT, mice exhibited a reduction in PPI. The present data show that a neonatal infection targeting the brain can induce the kynurenine pathway and that such an infection can disrupt sensorimotor gating in adulthood in genetically vulnerable mice.

Induction of the kynurenine pathway by neurotropic influenza a virus infection

Glutamatergic NMDA (N‐methyl D‐aspartate) receptors play a critical role in brain development and neurotransmission. Kynurenic acid, an end product of tryptophan degradation along the kynurenine pathway, is an endogenous NMDA receptor antagonist. In the present study, the effects of neurotropic influenza A virus infection on the kynurenine pathway were investigated in mouse brain primary cell cultures and in mouse brain after infection on day 3 of postnatal life. Altered levels of transcripts encoding several key enzymes of the kynurenine pathway were observed in infected neuron and glial cell cultures. In vivo, changes in the levels of such transcripts in brain were observed on postnatal days 7 and 13 but not on day 24. On postnatal day 13, infiltrating T lymphocytes and increased levels of kynurenic acid were observed in the brains of the infected animals. Taken together, the present results indicate that central nervous system infections during early life can activate the entire kynurenine pathway. Such activation is likely to result in the generation of several bioactive metabolites, as supported by our finding of a transient increase of kynurenic acid. In light of its antagonistic actions on the NMDA receptor, kynurenic acid can potentially link infections with glutamatergic signaling in the developing brain.

Activation of kynurenine pathway in ex vivo fibroblasts from patients with bipolar disorder or schizophrenia: Cytokine challenge increases production of 3-hydroxykynurenine

Accumulating data suggest a causative link between immune stimulation, disturbed metabolism of tryptophan, and pathogenesis of bipolar disorder and schizophrenia. The goal of this study was to examine the production of kynurenic acid (KYNA), 3-hydroxykynurenine (3-HK) and the expression of kynurenine pathway enzymes involved in their synthesis and metabolism in cultured skin fibroblasts obtained from patients with bipolar disorder, schizophrenia or from healthy control individuals. The assessment was performed under basal conditions or following treatment with interferon (IFN)-γ, tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-6, or their combinations, in cells exposed to exogenous kynurenine. In both groups of patients, the baseline production of KYNA and 3-HK was increased, as compared to control subjects. Case-treatment analyses revealed significant interactions between bipolar case status and IL-1β, IL-6, IFN-γ + TNF-α, or IFN-γ + IL-1β, as well as between schizophrenia case status and IL-1β, IFN-γ + TNF-α, or IFN-γ + IL-1β, in terms of higher 3-HK. Noteworthy, no case-treatment interactions in terms of KYNA production were found. Observed changes did not appear to correlate with the expression of genes encoding kynurenine aminotransferases (KATs), kynureninase (KYNU) or kynurenine-3-monooxygenase (KMO). The single nucleotide polymorphisms (SNPs), rs1053230 and rs2275163, in KMO influenced KYNA levels yet did not explain the case-treatment discrepancies. In conclusion, our present findings indicate the utility of skin-derived fibroblasts for kynurenines research and support the concept of kynurenine pathway alterations in bipolar disorder and schizophrenia. The increase in ratio between neurotoxic 3-HK and neuroinhibitory/neuroprotective KYNA following exposure to cytokines may account for altered neurogenesis and structural abnormalities characteristic for both diseases.

Effects of pro-inflammatory cytokines on expression of kynurenine pathway enzymes in human dermal fibroblasts

BackgroundThe kynurenine pathway (KP) is the main route of tryptophan degradation in the human body and generates several neuroactive and immunomodulatory metabolites. Altered levels of KP-metabolites have been observed in neuropsychiatric and neurodegenerative disorders as well as in patients with affective disorders. The purpose of the present study was to investigate if skin derived human fibroblasts are useful for studies of expression of enzymes in the KP.MethodsFibroblast cultures were established from cutaneous biopsies taken from the arm of consenting volunteers. Such cultures were subsequently treated with interferon (IFN)-γ 200 U/ml and/or tumor necrosis factor (TNF)-α, 100 U/ml for 48 hours in serum-free medium. Levels of transcripts encoding different enzymes were determined by real-time PCR and levels of kynurenic acid (KYNA) were determined by HPLC.ResultsAt base-line all cultures harbored detectable levels of transcripts encoding KP enzymes, albeit with considerable variation across individuals. Following cytokine treatment, considerable changes in many of the transcripts investigated were observed. For example, increases in the abundance of transcripts encoding indoleamine 2,3-dioxygenase, kynureninase or 3-hydroxyanthranilic acid oxygenase and decreases in the levels of transcripts encoding tryptophan 2,3-dioxygenase, kynurenine aminotransferases or quinolinic acid phosphoribosyltransferase were observed following IFN-γ and TNF-α treatment. Finally, the fibroblast cultures released detectable levels of KYNA in the cell culture medium at base-line conditions, which were increased after IFN-γ, but not TNF-α, treatments.ConclusionsAll of the investigated genes encoding KP enzymes were expressed in human fibroblasts. Expression of many of these appeared to be regulated in response to cytokine treatment as previously reported for other cell types. Fibroblast cultures, thus, appear to be useful for studies of disease-related abnormalities in the kynurenine pathway of tryptophan degradation.

Neonatal infection with neurotropic influenza A virus affects working memory and expression of type III Nrg1 in adult mice

Epidemiological studies suggest that early life infections may contribute to the development of psychiatric disorders characterized by cognitive deficits. Here, we studied the effects of a neonatal influenza A/WSN/33 virus infection on locomotor activity, working memory and emotional behavior in adult mice. In addition to wild type mice, immunodeficient (Tap1(-/-)) mice lacking functional CD8(+) T cells, were included in the study to model the potential influence of a genetic deficit relating to virus clearance. Three to four months after the infection, infected Tap1(-/-) mice, but not wild type mice, exhibited deficits in working memory as well as increased rearing activity and anxiety. In the medial prefrontal cortices of these infected Tap1(-/-) mice reduced levels of type III Nrg1 transcripts were observed supporting a role for neuregulin 1 signaling in neuronal circuits involved in working memory. Virus replication, distribution or clearance did not differ between the two genotypes. The lack of CD8(+) T cells, however, appeared to contribute to a more pronounced glia response in Tap1(-/-) than in wild type mice. Thus, the present study suggest that the risk of developing deficits in cognitive and emotional behavior following a CNS infection during brain development is influenced by genetic variation in genes involved in the immune response.

Gene expression changes in brains of mice exposed to a maternal virus infection.

In this study, we tested the hypothesis that exposure to a maternal infection during fetal life can lead to the appearance of alterations in the brain later in life. C57BL/6 mice were infected intranasally with influenza A/WSN/33 virus on day 14 of gestation. The levels of transcripts encoding neuroleukin and fibroblast growth factor 5 were significantly elevated in the brains of the virus-exposed offspring at 90 and 280 days of age, but not at earlier time-points. For neuroleukin, this difference could also be observed at the protein level. Thus, a maternal influenza A virus infection can give rise to alterations in gene expression in the brain that become apparent only after a prepubertal latency period.

Behavioral disturbances in adult mice following neonatal virus infection or kynurenine treatment – Role of brain kynurenic acid

Exposure to infections in early life is considered a risk-factor for developing schizophrenia. Recently we reported that a neonatal CNS infection with influenza A virus in mice resulted in a transient induction of the brain kynurenine pathway, and subsequent behavioral disturbances in immune-deficient adult mice. The aim of the present study was to investigate a potential role in this regard of kynurenic acid (KYNA), an endogenous antagonist at the glycine site of the N-methyl-D-aspartic acid (NMDA) receptor and at the cholinergic α7 nicotinic receptor. C57BL/6 mice were injected i.p. with neurotropic influenza A/WSN/33 virus (2400 plaque-forming units) at postnatal day (P) 3 or with L-kynurenine (2×200 mg/kg/day) at P7-16. In mice neonatally treated with L-kynurenine prepulse inhibition of the acoustic startle, anxiety, and learning and memory were also assessed. Neonatally infected mice showed enhanced sensitivity to D-amphetamine-induced (5 mg/kg i.p.) increase in locomotor activity as adults. Neonatally L-kynurenine treated mice showed enhanced sensitivity to D-amphetamine-induced (5 mg/kg i.p.) increase in locomotor activity as well as mild impairments in prepulse inhibition and memory. Also, D-amphetamine tended to potentiate dopamine release in the striatum in kynurenine-treated mice. These long-lasting behavioral and neurochemical alterations suggest that the kynurenine pathway can link early-life infection with the development of neuropsychiatric disturbances in adulthood.

Influenza A virus transactivates the mouse envelope gene encoding syncytin B and its regulator, glial cells missing 1

Recently, two candidate analogs for human syncytin, denoted syncytins A and B, were identified in the murine genome. These were found to have expression patterns and functions similar to human syncytin. In addition, the identification of glial cells missing (GCM)-binding motifs in putative promoter regions of the mouse syncytins imply analogous regulation. Transcriptional modulation of syncytin by exogenous agents was recently suggested by studies reporting transactivation of syncytin in human cell lines following virus infections. The authors report that influenza A virus infection increased the levels of transcripts encoding Gcm1 and syncytin B, but not syncytin A, in NIH-3T3 cells as well as in mouse primary neurons or glia. Overexpression of human GCM1 in NIH-3T3 cells resulted in increased levels of transcripts encoding syncytin B but not syncytin A. Systemic administration of neurotropic influenza A virus resulted in a neuronal infection and increased levels of Gcm1-encoding transcripts in brains of young mice. The mouse may therefore be useful for studies on the expression and function of endogenous retroviral envelope genes and transcription factors regulating their expression in the placenta and brain during physiological or pathological conditions.

Endogenous Retroviruses and Human Neuropsychiatric Disorders

Schizophrenia and related disorders are devastating human neuropsychiatric disorders of complex etiology. Epidemiological and family studies indicate both genetic and environmental contributions to disease etiology.We propose that the altered expression of endogenous retroviruses such asHERV-Wcontribute to somecases of schizophrenia. We present both theoretical considerations and experimental evidence supporting this association. The further study of endogenous retroviral expression within the central nervous system might lead to new methods for the diagnosis and treatment of schizophrenia and related disorders.

109. Enhanced kynurenic acid levels and cytokine-induced production of 3-hydroxykynurenine in fibroblasts from bipolar or schizophrenic patients

Recent studies suggest that cytokine-related disturbances of kynurenine metabolism are involved in the pathogenesis of schizophrenia and bipolar disorder. Here, we examined the production of kynurenic acid (KYNA) and 3-hydroxykynurenine (3-HK) and the expression of kynurenine pathway enzymes under basal conditions and in the presence of pro-inflammatory cytokines in fibroblast cultures from patients with schizophrenia or bipolar disorder. Higher baseline production of KYNA and 3-HK was demonstrated in fibroblasts obtained from patients as compared with control subjects. Following exposure to interferon (IFN)-gamma, tumor necrosis factor (TNF)-alpha, interleukin (IL)1-beta, IL6, or combinations, 3-HK levels, but not KYNA, increased in the fibroblast cultures. After treatments with cytokines the mRNA expression of kynurenine aminotransferase (KAT) enzymes were decreased and kynurenase (KYNU) increased, kynurenine-3-monooxygenase (KMO) levels were not affected. Interaction analyses showed that cells from patients with schizophrenia or bipolar disorder respond differently to pro-inflammatory cytokines in terms of L-kynurenine metabolism as compared to control individuals. Potential associations with two polymorphisms in the KMO allele were also investigated. The single nucleotide polymorphisms (SNPs) rs1053230 and rs2275163 both had effects on KYNA levels but these SNPs did not explain the case-control differences in KYNA or 3-HK levels. In conclusion, our results provide evidence that the kynurenine pathway is abnormally affected by cytokine exposure in fibroblast cultures obtained from patients with schizophrenia or bipolar disorder.