Indoleamine 2, 3-dioxygenase is responsible for low stress tolerance after intracerebral hemorrhage.

In the chronic phase after intracerebral hemorrhage (ICH), the aftereffect-associated lowering of motivation burdens many patients; however, the pathogenic mechanism is unclear. Here, we revealed for the first time that indoleamine 2, 3-dioxygenase (IDO) expression and enzyme activity are increased in the collagenase-induced murine ICH model. IDO is a rate-limiting enzyme situated at the beginning of the kynurenine pathway and converts tryptophan, a source of serotonin (5-hydroxytryptamine; 5-HT), to kynurenine. In this study, we showed that IDO is localized in 5-HTergic neurons. After ICH, the synaptosomal 5-HT level decreased, but this effect was neutralized by subcutaneous injections of 1-methyl tryptophan (MT), a specific IDO inhibitor. These results suggest that ICH-induced IDO weakens the activity of 5-HTergic neurons. Accordingly, we next investigated whether the IDO increase contributes to the depression-like behaviors of ICH mice. The immobility times of tail suspension and forced swimming tests were significantly prolonged after ICH but shortened by the administration of 1-MT. In conclusion, the increased IDO after ICH was found to decrease 5-HT levels and subsequently reduce stress tolerance. These findings indicate that IDO is a novel therapeutic target for the ICH aftereffect-associated lowering of motivation.

Gadolinium causes M1 and M2 microglial apoptosis after intracerebral haemorrhage and exerts acute neuroprotective effects.

OBJECTIVES: Gadolinium (Gd) affects microglial polarization during remyelination. We previously reported that the suppression of proinflammatory microglia was neuroprotective in intracerebral haemorrhage (ICH). The objective of the present study was to investigate the effects of Gd on microglial polarization and neuronal injury after ICH. METHODS: Gadolinium was intraperitoneally administered to ICH mice prepared by an intrastriatal microinjection of collagenase type VII. The polarization of M1, 2a, b and c microglia was evaluated by real-time PCR using the respective markers. Changes in representative mRNAs were also confirmed by immunological methods. Neuroprotective effects were evaluated by counting NeuN-positive cells and a behavioural analysis. KEY FINDINGS: One day after ICH, the mRNA levels of proinflammatory M1 microglial markers, such as inducible nitric oxide synthase (iNOS), and anti-inflammatory M2 microglial markers, such as arginase1 (M2a, c), Ym1 (M2a), and transforming growth factor-ß (M2c), increased, while those of chemokine CCL1 (M2b) only increased after 3 days. Gd decreased the levels of all M1 and M2 markers. Arginase1 and iNOS protein levels also increased, and Gd reduced them due to apoptotic cell death. Gadolinium attenuated oedema, neuron loss, neurological deficits and the mortality rate without affecting haematoma sizes. CONCLUSIONS: Gadolinium induced M1 and M2 microglial apoptosis and exerted acute neuroprotective effects after ICH.

Bradykinin and interleukin-1ß synergistically increase the expression of cyclooxygenase-2 through the RNA-binding protein HuR in rat dorsal root ganglion cells.

Synergistic expression of cyclooxygenase-2 (COX-2) by interleukin-1ß (IL-1ß) and bradykinin (BK) in peri-sensory neurons results in the production of prostanoids, which affects sensory neuronal activity and responsiveness and causes hyperalgesia. To evaluate the effects of pro-inflammatory mediators on COX-2 expression, cultured rat dorsal root ganglion (DRG) cells were treated with IL-1ß and BK, which caused persistent increased COX-2 expression. Co-treatment increased COX-2 transcriptional activities in an additive manner by a COX-2 promoter luciferase assay. Immunoprecipitated HuR, an RNA-binding protein, in co-treated DRG cells contained more COX-2 mRNA than that of the control. The synergistic effects of IL-1ß and BK on COX-2 expression may be a result of RNA stabilization mediated by HuR in peri-sensory neurons. Multiple pro-inflammatory cytokines and mediators are produced during neurogenic inflammation and aberrant control of COX-2 mRNA turnover may be implicated in diseases including chronic inflammation, which results in inflammation-derived hyperalgesia around primary sensory neurons.

Shogaol but not gingerol has a neuroprotective effect on hemorrhagic brain injury: Contribution of the α, ß-unsaturated carbonyl to heme oxygenase-1 expression.

We investigated the effects of shogaol, which has an α, ß-unsaturated carbonyl group, and gingerol, which does not, on primary-cultured microglia to understand how the α, ß-unsaturated carbonyl interacts with Kelch-like ECH-associated protein (Keap)1. Shogaol (1 µM) but not the same concentration of gingerol significantly increased heme oxygenase (HO)-1 protein levels in cultured microglia without cytotoxicity. In addition, shogaol suppressed the release of the inflammation marker nitric oxide induced by 30 U/ml thrombin treatment. A docking simulation suggested that the α, ß-unsaturated carbonyl of shogaol but not gingerol interacts with Keap1. Nuclear import of nuclear factor E2-related factor 2 and increased binding of the HO-1 E2 enhancer support the docking-simulation prediction. The transcription inhibitor actinomycin D (0.1 µg/ml) markedly blocked the increase of HO-1 mRNA levels by shogaol. To evaluate whether the α, ß-unsaturated carbonyl can be used for intracerebral hemorrhage (ICH) therapy, we investigated the effect of shogaol on an in vivo mouse ICH model. Intracerebroventricular injection of 0.2 nmol shogaol increased striatal HO-1 protein levels and rescued ICH-induced neuron loss. Thus, the α, ß-unsaturated carbonyl is necessary for the interaction of compounds, such as shogaol, with Keap1, and these findings may be useful for screening novel ICH therapeutic agents that increase HO-1 expression.

Claudin domain containing 1 contributing to endothelial cell adhesion decreases in presence of cerebellar hemorrhage.

The claudin family comprises four-pass transmembrane proteins involved in the formation of tight junctions (TJs). Relatively recently, claudin domain containing (CLDND) 1, also known as claudin-25, was identified as a novel member of the claudin family. In the present study, we revealed that in the adult murine brain, CLDND1 is abundant in the cerebellum among common sites of intracerebral hemorrhage. Thus, the dynamics of CLDND1 after cerebellar hemorrhage were examined. Both CLDND1 mRNA and protein levels transiently decreased at 24 hr after hemorrhagic insult. For immunostaining, an anti-CLDND1 antibody that recognizes the specific epitope in the extracellular first loop was prepared. Dual immunohistochemical staining with CD31 using coronal cryosections of intact murine cerebellum tissue revealed that CLDND1 is expressed on endothelial cells. We therefore performed an in vitro permeability test using a human brain endothelial cell (HBEC) line to reveal whether CLDND1 contributes to cell adhesion like other claudins. CLDND1 was expressed on HBECs as well as in murine cerebellum tissue, and a strong signal was observed at TJs. RNA interference against CLDND1 decreased both the mRNA and protein levels without cytotoxicity. The permeability to small molecules, but not to large ones, across confluent HBECs increased on CLDND1 knockdown compared with mock-treated cells. These results suggest that the transient decrease of CLDND1 after cerebellar hemorrhage is responsible for low-molecular-weight selective vascular hyperpermeability. © 2017 Wiley Periodicals, Inc.