Differential effect of restraint stress on MHC class II expression by murine peritoneal macrophages

The effect of restraint stress on the expression of MHC class II glycoproteins by peritoneal macrophages was evaluated. Restraint suppressed the expression of I-A by macrophages from mice that are susceptible to Mycobacterial infection. In contrast, restraint did not affect I-A expression by macrophages from resistant mice. The suppression of MHC class II expression required at least 8 h of restraint and recovered within 4 h after stress. The amount of restraint necessary to suppress I-A expression also resulted in higher levels of plasma corticosterone. Changes in I-A expression were under circadian rhythm control. The differences in the effect of restraint stress on expression of I-A by peritoneal macrophages from resistant and susceptible mice may, in part, be due to differences in the effect of corticosterone in MHC class II expression.     

Induction of major histocompatibility complex class II glycoproteins by interferon-gamma: attenuation of the effects of restraint stress.

The effect of restraint on the activation of macrophages was evaluated based on the induction of I-A expression following injection of viable Mycobacterium bovis (strain BCG) or treatment in vitro with recombinant interferon-gamma (rIFN-gamma). We found that restraint suppressed the induction of I-A expression when applied just prior to or at the same time as the injection of the microorganisms but had no effect if applied after the injection of the Mycobacteria. The effect of stress was attenuated by increasing the number of microorganisms or by incubating macrophages from stressed mice with higher doses of rIFN-gamma. The suppressive effect of restraint does not appear to be associated with uptake, processing or presentation of antigen but rather to an alteration in the response of the macrophages to rIFN-gamma.     

Induction of major histocompatibility complex class II glycoproteins by interferon-γ: attenuation of the effects of restraint stress

The effect of restraint on the activation of macrophages was evaluated based on the induction of I-A expression following injection of viable Mycobacterium bovis (strain BCG) or treatment in vitro with recombinant interferon-γ (rIFN-γ). We found that restraint suppressed the induction of I-A expression when applied just prior to or at the same time as the injection of the microorganisms but had no effect if applied after the injection of the Mycobacteria. The effect of stress was attenuated by increasing the number of microorganisms or by incubating macrophages from stressed mice with higher doses of rIFN-γ. The suppressive effect of restraint does not appear to be associated with uptake, processing or presentation of antigen but rather to an alteration in the response of the macrophages to rIFN-γ.     

Nerve growth factor production and expression of p75 by Schwann cells and neurofibroblasts in response to M. leprae infection and macrophage secretory products

This study describes the changes occurring in vitro in nerve growth factor (NGF) production and expression of p75 by murine Schwann cells and neurofibroblasts, following infection with Mycobacterium leprae and in the presence of macrophage secretory products, using a semiquantitative ELISA. These parameters are compared in two strains of mice, Swiss White (SW) and C57B1/6, as they differ in their response to M. leprae infection; C57B1/6 is the 'resistant' strain. On infection, NGF levels remained unaltered in Schwann cells from both strains, while fibroblasts from C57B1/6 strain showed an increase in NGF production. Expression of p75 by Schwann cells was decreased on infection in both strains of mice. In vivo , this opposing effect of infection on NGF production and p75 expression by Schwann cells and neurofibroblasts may result in suboptimal amounts of NGF reaching neurons of the affected leprous nerves. Macrophage secretory products suppressed the production of NGF by infected neurofibroblasts from SW strain mice and the expression of p75 in Schwann cells from both strains. These results indicate that macrophages do not assist in nerve repair in leprosy and the differences in response to macrophage secretory products in the two strains suggest that different mechanisms of nerve repair operate in SW and C57B1/6 mice and presumably in lepromatous and tuberculoid patients.     

Rantes secreted from macrophages disturbs skeletal muscle regeneration after cardiotoxin injection in Cbl-b-deficient mice

Deficiency of the Cbl-b ubiquitin ligase gene activates macrophages in mice. This study aimed to elucidate the pathophysiological roles of macrophages in muscle degeneration/regeneration in Cbl-b-deficient mice. We examined immune cell infiltration and cytokine expression in cardiotoxin-injected tibialis anterior muscle of Cbl-b-deficient mice. Ablation of the Cbl-b gene expression delayed regeneration of cardiotoxin-induced skeletal muscle damage compared with wild-type mice. CD8-positive T cells were still present in the damaged muscle on day 14 after cardiotoxin injection in Cbl-b-deficient mice, but there was dispersal of the same cells over that time-frame in wild-type mice. Infiltrating macrophages in Cbl-b-deficient mice showed strong expression of RANTES (regulated-on-activation, normal T cell expressed and secreted), a chemokine for CD8-positive T cells. In turn, a neutralizing antibody against RANTES significantly suppressed the infiltration of CD8-positive T cells into the muscle, resulting in restoration of the disturbed muscle regeneration. Cbl-b is an important regulatory factor for cytotoxic T-cell infiltration via RANTES production in macrophages.     

Effect of interleukin 1beta on the HPA axis in H1-receptor knockout mice

OBJECTIVES AND METHODS: Circulating cytokines such as interleukin-1 (IL-1), and tumor necrosis factor-alpha as well as lipopolysaccharide (LPS) are potent ACTH secretagogues, acting via stimulation of corticotropin-releasing hormone (CRH) and vasopressinergic neurons in the paraventricular nucleus of the hypothalamus (PVN). Histamine (HA) has been shown to stimulate ACTH secretion in rats, an effect in part mediated by CRH and arginine vasopressin (AVP). We have previously shown that inhibition of neuronal HA synthesis or central blockade of H(1) receptors (H(1)R) decreased the ACTH response to LPS in male rats. To further elucidate the role of neuronal HA in cytokine-induced activation of the HPA axis, we compared the effect of H(1)R knockout on IL-1beta-induced ACTH secretion in adult male mice. RESULTS: In H(1)R knockout mice, ACTH secretion increased from basal levels of 261 to 492 pmol/l in response to IL-1beta whereas the cytokine-induced ACTH secretion increased from 140 to 406 pmol/l in wild-type mice. Plasma corticosterone (CORT) rose from basal levels of 99 to 831 nmol/l in knockout mice upon IL-1beta stimulation, whereas in wild-type mice CORT levels rose from 112 to 841 nmol/l. There was no significant difference in IL-1beta-stimulated plasma ACTH or CORT levels between wild-type and knockout mice. Furthermore, there was no significant difference in basal or IL-1beta-stimulated hypothalamic levels of histamine and tele-methyl-histamine between wild-type and knockout mice. HDC gene expression was significantly lower in knockout mice than in wild-type mice both under basal and IL-1beta-stimulated conditions, while there were no significant differences in CRH gene expression in the PVN in knockout mice under basal and IL-1beta-stimulated conditions. Increased basal expression of AVP in the PVN of knockout mice was observed in this study compared to wild-type mice. CONCLUSION: We conclude that the lack of the gene for histamine H(1)R does not seem to be crucial for the ACTH and CORT response to IL-1beta, either due to possible functional compensation in the H(1)R knockout mouse or due to activation of pathways other than the neuronal histaminergic system.     

Activation of the hypothalamic-pituitary-adrenal axis differentially affects the anti-mycobacterial activity of macrophages from BCG-resistant and susceptible mice

The effect of hypothalamic-pituitary-adrenal (HPA) axis activation and exogenous glucocorticoids on the ability of splenic macrophages to control the growth of Mycobacterium avium was evaluated. We found that activation of the HPA axis by restraint stress or the addition of corticosterone increased the susceptibility of macrophages from mice that are innately susceptible to the in vivo growth of M. avium. In contrast, the ability of macrophages from innately resistant, congenic mice to control the growth of M. avium was not affected by HPA activation or the addition of corticosterone. The effect of restraint and of corticosterone on macrophage function was abrogated by either treating mice with the glucocorticoid receptor antagonist RU486 or the addition of the drug to cultures of macrophages. Activation of the HPA axis as well as the addition of corticosterone to cultures of macrophages resulted in a suppression of the production of tumor necrosis factor (TNF)-α and of reactive nitrogen intermediates by macrophages from both strains of mice. The lack of effect of HPA activation and of corticosterone on the mycobacterial resistance of macrophages from BCG-resistant mice, while at the same time suppressing the production of reactive nitrogen intermediates, appears to rule out a role for this antimicrobial pathway in innate resistance to mycobacterial growth.     

脑血管病患者外周血单个核细胞ACTH受体表达

采用放射配体分析及放射免疫测定法，检测了18例脑血管病患者外周血单个核细胞（PMNC）ACTH受体表达及血浆ACTH水平。结果显示：脑出血患者PMNCACTH受体表达水平及血浆ACTH含量均显著高于健康对照组（P<0.01），脑梗死患者血浆ACTH水平显著升高（P<0.01），但其PMNCACTH受体表达无明显变化（P>0.05）。上述患者免疫细胞活性检测结果表明，脑出血患者T细胞及NK细胞活性均较健康对照组明显降低（P<0.01），脑梗死患者只表现T细胞活性受抑制（P<005）。相关性分析显示，脑血管病患者PMNCACTH受体表达水平与T细胞和NK细胞活性呈负相关（P<0．01）。以上结果提示，脑血管病患者细胞免疫功能抑制效应可能是通过免疫细胞膜上ACTH受体介导的。     

Paradoxes of immunosuppression in mouse models of withdrawal

Previously, our laboratory showed that either abrupt (AW) or precipitated withdrawal (PW) from morphine led to profound suppression of murine splenic antibody responses to sheep red blood cells at 24 h post-withdrawal. In the present studies, we examined the immune mechanisms mediating suppression at that time point. A co-culture method was used to examine whether cells from withdrawn mice had (1) a deficit in function and/or (2) contained populations of suppressor cells. To examine the first hypothesis, cells from normal mice were co-cultured with cells from withdrawn mice in a 1:3 ratio (normal/withdrawn). To test the second hypothesis, the ratio was reversed. The results were paradoxical. Co-culture of cells in a 1:3 ratio showed that spleen cells from withdrawn mice had a deficit in macrophage function. Spleen cells from withdrawn mice also showed decreased mRNA levels of IL-1beta, IL-1-Ra, and TNF-alpha and a suppression of co-stimulatory molecule expression. To examine the second hypothesis, cells were co-cultured in a 3:1 ratio (normal/withdrawn). In this paradigm, spleen cells from abrupt withdrawn mice were shown to contain populations of both suppressor macrophages and B-cells. In vivo experiments carried out on mice 24 h post-withdrawal showed increased sensitivity to the lethal effects of LPS and increased production of TNF-alpha, implying a state of macrophage activation. Thus evidence for both suppressed and activated macrophages has been obtained in mice 24 h after abrupt withdrawal from morphine.     

Chromaffin cell function and structure is impaired in corticotropin-releasing hormone receptor type 1-null mice

Corticotropin-releasing hormone (CRH) is both a main regulator of the hypothalamic-pituitary-adrenocortical axis and the autonomic nervous system. CRH receptor type 1 (CRHR1)-deficient mice demonstrate alterations in behavior, impaired stress responses with adrenocortical insufficiency and aberrant neuroendocrine development, but the adrenal medulla has not been analyzed in these animals. Therefore we studied the production of adrenal catecholamines, expression of the enzyme responsible for catecholamine biosynthesis neuropeptides and the ultrastructure of chromaffin cells in CRHR1 null mice. In addition we examined whether treatment of CRHR1 null mice with adrenocorticotropic hormone (ACTH) could restore function of the adrenal medulla. CRHR1 null mice received saline or ACTH, and wild-type or heterozygous mice injected with saline served as controls. Adrenal epinephrine levels in saline-treated CRHR1 null mice were 44% those of controls (P<0.001), and the phenylethanolamine N-methyltransferase (PNMT) mRNA levels in CRHR1 null mice were only 25% of controls (P <0.001). ACTH treatment increased epinephrine and PNMT mRNA level in CRHR1 null mice but failed to restore them to normal levels. Proenkephalin mRNA in both saline- and ACTH-treated CRHR1 null mice were higher than in control animals (215.8% P <0.05, 268.9% P <0.01) whereas expression of neuropeptide Y and chromogranin B did not differ. On the ultrastructural level, chromaffin cells in saline-treated CRHR1 null mice exhibited a marked depletion in epinephrine-storing secretory granules that was not completely normalized by ACTH-treatment. In conclusion, CRHR1 is required for a normal chromaffin cell structure and function and deletion of this gene is associated with a significant impairment of epinephrine biosynthesis.     

B cells from glatiramer acetate-treated mice suppress experimental autoimmune encephalomyelitis

Multiple sclerosis (MS) is an autoimmune disease of the central nervous system (CNS) thought to be primarily mediated by T cells. However, emerging evidence supports an important role for B cells in the pathogenesis and inhibition of MS. Glatiramer acetate (GA), a Food and Drug Administration-approved drug for the treatment of MS, has a good safety profile. But GA's mechanism of action in MS is still elusive. In this study, we showed that B cells from GA-treated mice increased production of IL-10 and reduced expression of co-stimulatory molecules viz.: CD80 and CD86. B cells from GA-treated mice also diminished proliferation of myelin oligodendrocyte glycoprotein (MOG_35-55) specific T cells. Purified B cells transferred from GA-treated mice suppressed experimental autoimmune encephalomyelitis (EAE) in recipient mice compared with B cells transferred from mice treated with PBS or ovalbumin. The treatment effect of GA in EAE was abrogated in B cell-deficient mice. Transfer of B cells from GA-treated mice inhibited the proliferation of autoreactive T cells as well as the development of Th1 and Th17 cells but promoted IL-10 production in recipient mice. The number of peripheral CD11b⁺ macrophages in recipient mice also decreased after transfer of B cells from GA-treated mice; however, the number of dendritic cells and regulatory T cells remained unaltered. These results suggest that B cells are important to the protective effects of GA in EAE.     

Adrenocorticotropic hormone controls infantile spasms independently of cortisol stimulation

Infantile spasms constitute a severe seizure disorder unresponsive to standard anticonvulsants. Both prednisone and adrenocorticotropic hormone (ACTH) have produced remission of seizures in some patients. The mechanisms of action of these hormones are not known. Eight infants with infantile spasms were treated with prednisone for 2 weeks. This controlled the seizures in two patients. In the remaining six patients, prednisone was continued and ACTH was added. This treatment produced cessation of spasms in four patients. Serum prednisone and cortisol were measured at a number of points during treatment. In infants receiving prednisone and then prednisone plus ACTH, serum cortisol was suppressed to about one-quarter of baseline levels with the initiation of prednisone, and remained suppressed during ACTH administration. We conclude that ACTH can exert its effect on infantile spasms in the setting of adrenal suppression, and can act without stimulating endogenous cortisol production. A CNS site of action is suggested and should be sought.     

Pineal modulation of ACTH 1-17 effect upon murine corticosterone production

In tests of corticosterone production in vitro, aqueous pineal homogenate (APH) modulates the effect of a short-chain ACTH analogue, ACTH 1-17, added to adrenals from different circadian stages. Adrenal and pineal glands from female B6D2F1 mice, standardized on staggered LD 12:12 regimens, were obtained at the same clock-hour from each room, in order to cover 6 different circadian stages. Adrenals from each circadian stage were bisected and incubated with APH from the same circadian stage (isophasic incubation) or from one of the other 5 circadian stages (heterophasic incubation). ACTH 1-17 (0.05 IU) was added to each incubation medium. After 4 hours of incubation at 37 degrees C with 95% O2 and 5% CO2, the media were stored at -20 degrees C until corticosterone RIA were done. APH was found to have a statistically significant modulatory effect upon the stimulation by ACTH 1-17 of adrenal corticosterone production in vitro. This APH effect changed rhythmically as a function of circadian stage from amplification over no effect to attenuation, as a so-called feed-sideward.     

In vitro adrenergic modulation of cellular immune functions in experimental autoimmune encephalomyelitis

OBJECTIVE: To analyze the effects in vitro of alpha- and beta-adrenoceptor agonists on splenocyte proliferation and on proinflammatory cytokine production in splenocytes and peritoneal macrophages (MF) in different stages of EAE. METHODS: Splenocytes and peritoneal macrophages were harvested in the acute phase of EAE and in remission, and from controls. The beta-agonist terbutaline, the alpha(1)-agonist methoxamine, and the alpha(2)-agonist UK-14304 were added with ConA or lipopolysaccharide (LPS). TNF-alpha and IFN-gamma contents in supernatant and splenocyte proliferation were determined. RESULTS: Terbutaline and UK-14304 significantly suppressed TNF-alpha production by MF. However, EAE acute phase rats were resistant to the suppressive effect of UK-14304. Terbutaline significantly suppressed IFN-gamma and TNF-alpha production by splenocytes. EAE acute phase and remission animals showed reduced terbutaline-induced inhibition of IFN-gamma production. CONCLUSIONS: Disturbed sympathetic-immune communication in EAE is characterized by alterations in adrenergic sensitivity via both alpha- and beta-adrenergic pathways.     

ICAM-1 upregulation in the spinal cords of PLSJL mice with experimental allergic encephalomyelitis is dependent upon TNF-alpha production triggered by the loss of blood-brain barrier integrity

Urate (UA) selectively scavenges peroxynitrite-dependent radicals and suppresses CNS inflammation through effects that are manifested at the blood-brain barrier (BBB). ICAM-1 upregulation in the spinal cord tissues of myelin basic protein (MBP) immunized mice is selectively inhibited by UA treatment. In contrast, the expression of ICAM-1 and other adhesion molecules by circulating cells is unchanged. Moreover, TNF-alpha expression in the CNS tissues of MBP-immunized mice is suppressed by UA treatment but TNF-alpha-induced ICAM-1 expression on neurovascular endothelial cells is not. Therefore the effect of UA on ICAM-1 upregulation in the CNS tissues is likely due to its known contribution to the maintenance of BBB integrity in MBP-immunized mice which in turn inhibits cell invasion into the CNS and prevents TNF-alpha production in CNS tissues.     

TRPM2 contributes to inflammatory and neuropathic pain through the aggravation of pronociceptive inflammatory responses in mice

Accumulating evidence suggests that neuroimmune interactions contribute to pathological pain. Transient receptor potential melastatin 2 (TRPM2) is a nonselective Ca2?-permeable cation channel that acts as a sensor for reactive oxygen species. TRPM2 is expressed abundantly in immune cells and is important in inflammatory processes. The results of the present study show that TRPM2 plays a crucial role in inflammatory and neuropathic pain. While wild-type and TRPM2 knock-out mice showed no difference in their basal sensitivity to mechanical and thermal stimulation, nocifensive behaviors in the formalin test were reduced in TRPM2 knock-out mice. In carrageenan-induced inflammatory pain and sciatic nerve injury-induced neuropathic pain models, mechanical allodynia and thermal hyperalgesia were attenuated in TRPM2 knock-out mice. Carrageenan-induced inflammation and sciatic nerve injury increased the expression of TRPM2 mRNA in the inflamed paw and around the injured sciatic nerve, respectively. TRPM2 deficiency diminished the infiltration of neutrophils and the production of chemokine (C-X-C motif) ligand-2 (CXCL2), a major chemokine that recruits neutrophils, but did not alter the recruitment of F4/80-positive macrophages in the inflamed paw or around the injured sciatic nerve. Microglial activation after nerve injury was suppressed in the spinal cord of TRPM2 knock-out mice. Furthermore, CXCL2 production and inducible nitric oxide synthase induction were diminished in cultured macrophages and microglia derived from TRPM2 knock-out mice. Together, these results suggest that TRPM2 expressed in macrophages and microglia aggravates peripheral and spinal pronociceptive inflammatory responses and contributes to the pathogenesis of inflammatory and neuropathic pain.     

The role of the arginine vasopressin Avp1b receptor in the acute neuroendocrine action of antidepressants

In times of stress the hypothalamic-pituitary-adrenal (HPA) axis is activated and releases two neurohormones, corticotropin-releasing hormone (Crh) and arginine vasopressin (Avp), to synergistically stimulate the secretion of adrenocorticotropin hormone (ACTH) from the anterior pituitary, culminating in a rise in circulating glucocorticoids. Avp mediates its actions at the Avp V1b receptor (Avpr1b) present on pituitary corticotropes. Dysregulation of the stress response is associated with the pathophysiology of depression and a major treatment involves increasing the availability of monamines at the synaptic cleft. Acute administration of selective serotonin reuptake inhibitors (SSRI) and tricyclic antidepressants (TCA) has previously been shown to activate the HPA axis. The present study was undertaken to evaluate the involvement of the Avpr1b in the HPA axis response to acute SC administration of an SSRI (fluoxetine 10mg/kg) and a TCA (desipramine 10mg/kg). We measured plasma ACTH and corticosterone (CORT) levels and neuropeptide mRNA expression in the hypothalamic paraventricular nucleus (PVN) of Avpr1b knockout (KO) mice and wild-type controls. Fluoxetine and desipramine administration significantly attenuated plasma ACTH and CORT levels in male and female Avpr1b KO mice when compared to their wild-type counterparts. Avp, oxytocin (Oxt) and Crh mRNA expression in the PVN did not change in fluoxetine-treated male Avpr1b KO or wild-type mice. In contrast, fluoxetine treatment increased PVN Avp mRNA levels in female Avpr1b wild type but not KO animals. PVN Oxt mRNA levels increased in fluoxetine-treated female mice of both genotypes. The data suggests that the Avpr1b is required to drive the HPA axis response to acute antidepressant treatment and provides further evidence of a sexual dichotomy in the regulation of PVN Avp/Oxt gene expression following antidepressant administration.     

Retinoic acid enhances prostaglandin E2 production through increased expression of cyclooxygenase-2 and microsomal prostaglandin E synthase-1 in rat brain microglia

Retinoic acid (RA) is a well-known antiinflammatory agent. In this study, we show that RA has a dual effect on cyclooxygenase-2 (COX-2) expression in inflammatory activated microglia, the resident brain macrophages. After treatment of microglia with LPS or thrombin, COX-2 expression was induced in two phases, specifically, an initial increase at about 12 hr after stimulation followed by a decrease, and another increase at about 48-72 hr. However, PGE(2) and 15d-PGJ(2) were detected at about 12 hr, and the levels continuously increased thereafter. Interestingly, all-trans retinoic acid (ATRA) suppressed the expression of early-phase COX-2 but augmented late-phase COX-2 and inhibited iNOS in the whole time sequence. ATRA enhanced PGE(2) production but had little effect on 15d-PGJ(2). Moreover, ATRA selectively up-regulated the expression of a PGE(2) synthase, mPGES-1, but had little effect on the PGD(2) synthase, H-PGDS. The results collectively suggest that ATRA modulates microglial responses to inflammatory stimulators, particularly at the late phase, via enhancement of COX-2 expression and PGE(2) production.