Ds-HMGB1 and fr-HMGB induce depressive behavior through neuroinflammation in contrast to nonoxid-HMGB1.

High mobility group box 1 (HMGB1) has been implicated as a key factor in several neuroinflammatory conditions. Our previous study suggested that the release of central HMGB1 acts as a late-phase mediator in lipopolysaccharide (LPS)-induced depression. Recent findings indicate that the redox state of HMGB1 is a critical determinant of its immunomodulatory properties. Here, we aimed to investigate the potential mechanisms that link the redox states of HMGB1 to depression in mice. Distinct redox forms of recombinant HMGB1 (rHMGB1) were used that included fully reduced HMGB (fr-HMGB1), which acted as a chemokine, and disulfide-HMGB1 (ds-HMGB1), which possessed cytokine activity. Fr-HMGB1 in vivo was partially oxidized into ds-HMGB1; thus, the mutant protein non-oxidizable chemokine-HMGB (nonoxid-HMGB1) was applied. Concurrent with depressive behavior induced by four-week stress exposure, the HMGB1 concentrations in the serum and cerebral cortex substantially increased. Therefore, a single dose of rHMGB1 (200ng/5µl/mice) or vehicle was administered to mice via intracerebroventricular (i.c.v.) injection. The receptor inhibitors of TLR4/RAGE/CXCR4 (TAK-242/FPS-ZM1/AMD3100) (3mg/kg) were intraperitoneally injected 30min prior to rHMGB1 treatment. Depressive-like behavior was measured 20h post i.c.v. injection. Administration of fr-HMGB1 prolonged the immobility duration in the tail suspension test (TST) and decreased sucrose preference. In addition to depressive behavior, the hippocampal TNF-α protein slightly increased. These depressive behaviors and upregulation of hippocampal TNF-α were alleviated or abrogated by pretreatment with the inhibitors AMD3100, FPS-ZM1, and TAK-242. Alternatively, nonoxid-HMGB1 failed to induce TNF-α protein or prolong the immobility duration. As expected, ds-HMGB1 administration substantially upregulated hippocampal TNF-α protein, increased the immobility time in the TST and decreased sucrose preference. Moreover, both glycyrrhizin and TAK-242 improved ds-HMGB1-induced depressive behavior. Furthermore, TAK-242 significantly blocked the upregulation of hippocampal TNF-α protein and protected hippocampal myelin basic protein from ds-HMGB1-induced reduction. These drugs had no effect on the total or central distance in the open field test. Collectively, this initial experiment demonstrates the role and receptor mechanisms of HMGB1 under different redox states on the induction of depressive-like behavior. Both ds-HMGB1 and fr-HMGB1 may induce depressive-like behavior in vivo mainly via neuroinflammatory response activation.

Rapid permissive action of dexamethasone on the regulation of blood pressure in a rat model of septic shock.

Glucocorticoids (GCs) play a vital role in the regulation of blood pressure by their permissive effects in potentiating vasoactive responses to catecholamines through glucocorticoid receptors. GCs achieve this function by controlling vascular smooth muscle tone. Clinically, low to moderate doses of GCs are generally used in the treatment of septic shock in recent years. GCs are now known to have both genomic and non-genomic effects. While genomic effects of GCs were well studied, few non-genomic effects were reported, much less the non-genomic mechanisms. One of the most important characters of their non-genomic effects is short latency. The aim of this study was to determine whether GCs can rapidly regulate blood pressure by their permissive action on norepinephrine (NE). Adrenalectomized rats were subjected to cecal ligation and puncture to induce septic shock. The septic rats displayed a significant decrease in the blood pressure response to NE. Dexamethasone (DEX) rapidly restores this hyporeactivity to NE in adrenalectomized septic rats. Further studies showed that DEX potentiates the NE-induced shrinkage and actin cytoskeleton rearrangement of single cell from mesenteric arteries in a short time. These findings suggest that GCs probably exert their permissive actions on the pressure response to NE through rapid non-genomic mechanisms. In this article, we found that as an adjunctive therapy for septic shock, the use of GCs may involve a rapid permissive action, and non-genomic effects of GCs may be involved in these processes.

Involvement of inflammasome activation in lipopolysaccharide-induced mice depressive-like behaviors.

AIMS: The NLRP3 inflammasome is a cytoplasmic multiprotein complex of the innate immune system that regulates the cleavage of interleukin-1ß and interleukin-18 precursors. It can detect a wide range of danger signals and trigger a series of immune-inflammatory reactions. There were plenty of studies indicating that activation of the immune system played pivotal roles in depression. However, the underlying mechanisms of immune-depression interactions remained elusive and there was no report about the involvement of inflammasome activation in depression. METHODS: We established an acute depression mouse model with lipopolysaccharide to explore the involvement of inflammasome activation in depression. RESULTS: The lipopolysaccharide-treated mice displayed depressive-like behaviors and pro-inflammatory cytokine interleukin-1ß protein and mRNA levels significantly increased. The NLRP3 inflammasome mRNA expression level also significantly elevated in depressed mice brain. Pretreatment with the NLRP3 inflammasome inhibitor Ac-YVAD-CMK significantly abrogated the depressive-like behaviors induced by lipopolysaccharide. CONCLUSION: These data suggest for the first time that the NLRP3 inflammasome is involved in lipopolysaccharide-induced mice depressive-like behaviors. The NLRP3 inflammasome may be a central mediator between immune activation and depression, which raises the possibility that it may be a more specific target for the depression treatments in the near future.

Neuropeptide Y induces secretion of high-mobility group box 1 protein in mouse macrophage via PKC/ERK dependent pathway.

Despite increasing evidence highlighting the role of NPY in the modulation of inflammatory reaction, surprisingly little is known about the direct effects of NPY on the release of proinflammatory mediators. In the present work, we have evaluated the effects of NPY on the release of TNF-α, IL-1ß, IL-6 and HMGB1 mediators in peritoneal macrophages. Our results demonstrate for the first time that NPY can directly induce active HMGB1 release and cytoplasmic translocation, while the production of TNF-α, IL-1ß and IL-6 is not affected. PKC and ERK pathway inhibitors can abolish the promotive effect of NPY on HMGB1 secretion. Thus, our results indicate that NPY might impact on the innate immune system by directly potentiating the HMGB1 release from the macrophage.

Dexamethasone induces rapid promotion of norepinephrine­mediated vascular smooth muscle cell contraction.

The aim of the present study was to identify the rapid effect of dexamethasone (Dex) on norepinephrine (NE)­mediated contraction of vascular smooth muscle cells (VSMCs) and to establish the underlying mechanism(s). Rat VSMCs were preincubated with lipopolysaccharide to simulate acute septic shock. Myosin light chain (MLC20) phosphorylation of VSMCs was detected by western blot analysis to observe the effects of Dex on NE­mediated contraction. Activation of the RhoA/ RhoA kinase (ROCK), extracellular signal­regulated kinase (ERK) and p38 signaling pathways was detected by western blot analysis to explore the mechanism. It was identified that Dex rapidly promoted NE­induced phosphorylation of MLC20 in VSMCs and this effect may be non­genomic. The RhoA/ROCK, ERK and p38 pathways were demonstrated to be important for the rapid effect of Dex­induced promotion of NE­mediated contraction in VSMCs. The present results indicate that Dex may rapidly reverse the hyporeactivity of vasoconstriction to NE in vitro and this effect may be mediated by specific non­genomic mechanisms through increased activation of the RhoA/ROCK, ERK and p38 signaling pathways.

Enhanced phosphorylation of MAPKs by NE promotes TNF-α production by macrophage through α adrenergic receptor.

The aim of this study was to investigate whether norepinephrine (NE) could regulate macrophage production of tumor necrosis factor alpha (TNF-α) by influencing the phosphorylation of mitogen-activated protein kinases (MAPKs). Primary macrophages from male BALB/c mice were applied to explore the mechanism by which NE influences the the secretion of TNF-α when macrophages were activated by lipopolysaccharides (LPS). We found that NE could increase crophage production of TNF-α when macrophages were activated by LPS, and this effect could be inhibited by α adrenergic antagonist phentolamine. Also, NE could increase the phosphorylation of c-Jun N-terminal kinase (JNK), extracellular signal-regulated kinases (ERK), and p38, through α receptor. Furthermore, JNK inhibitor SP600125, ERK inhibitor U0126, and p38 inhibitor SB203580 could all partially counteract NE's effect on the phosphorylation of MAPKs, as well as TNF-α production by macrophages. This study revealed that as macrophages were activated by LPS, NE promoted the secretion of inflammatory factors by increasing the phosphorylation of MAPKs through an α receptor-dependent pathway. Our results provide the evidence of a relationship between stress and diseases, as well as the mechanism by which stress induces or affects the inflammation-related diseases.

A novel strategy for development of glucocorticoids through non-genomic mechanism.

Glucocorticoids (GCs) are routinely believed to take effect through genomic mechanisms, which are also largely responsible for GCs' side effects. Beneficial non-genomic effects of GCs have been reported as being independent of the genomic pathway. Here, we synthesized a new type of GCs, which took effect mainly via non-genomic mechanisms. Hydrocortisone was conjugated with glycine, lysine and phenylalanine to get a bigger molecular structure, which could hardly go through the cell membrane. Evaluation of the anti-inflammatory efficacy showed that hydrocortisone-conjugated glycine (HG) and lysine could inhibit neutrophil degranulation within 15 min. HG could inhibit IgE-mediated histamine release from mast cells via a non-genomic pathway, and rapidly alleviate allergic reaction. Luciferase reporter assay showed that HG would not activate the glucocorticoid response element within 30 min, which verified the rapid effects independent of the genomic pathway. The work proposes a novel insight into the development of novel GCs, and provides new tools for experimental study on non-genomic mechanisms.

Neuropeptide Y promotes TGF-beta1 production in RAW264.7 cells by activating PI3K pathway via Y1 receptor.

OBJECTIVE: To examine the effect of neuropeptide Y (NPY) on TGF-beta1 production in RAW264.7 macrophages. METHODS: Enzyme linked immunosorbent assay (ELISA) was used to detect TGF-beta1 production. Cell counting kit 8 (CCK-8) was used to assay the viability of RAW264.7 cells. Western blot was used to detect the phosphorylation of PI3K p85. RESULTS: NPY treatment could promote TGF-beta1 production and rapid phosphorylation of PI3K p85 in RAW264.7 cells via Y1 receptor. The elevated TGF-beta1 production induced by NPY could be abolished by wortmannin pretreatment. CONCLUSION: NPY may elicit TGF-beta1 production in RAW264.7 cells via Y1 receptor, and the activated PI3K pathway may account for this effect.