Roles of GILZ in protein metabolism of L6 muscle cells exposed to serum from septic rats.

Sepsis is a complex inflammatory response to infection, associating with dramatic metabolic disorders. Although the mechanisms of immune response during sepsis have been largely clarified, current studies rarely pay attention to the disordered protein metabolism in sepsis. In this study, L6 rat skeletal muscle cells treated with serum from septic rats were used as an in vitro model for sepsis-like condition in skeletal muscle. We found that the expression of glucocorticoid-induced leucine zipper (GILZ) positively correlates with glucocorticoid receptor and negatively correlates with myosin heavy chain expression in L6 muscle cells upon septic serum induction. Moreover, we propose that GILZ may associate with cytokines such as TNF-α, IL-1ß as well as IL-10 to cooperatively modulate the glucocorticoid/glucocorticoid receptor-mediated regulation of protein metabolism during sepsis. So the present study provides a new approach and theoretical basis for further studies on the regulation of protein metabolism of skeletal muscle during sepsis.

Magnolol, a major bioactive constituent of the bark of Magnolia officinalis, exerts antiepileptic effects via the GABA/benzodiazepine receptor complex in mice.

BACKGROUND AND PURPOSE: The aim of this study was to evaluate the anti-convulsant effects of magnolol (6, 6', 7, 12-tetramethoxy-2, 2'-dimethyl-1-ß-berbaman, C18H18O2) and the mechanisms involved. EXPERIMENTAL APPROACH: Mice were treated with magnolol (20, 40 and 80 mg·kg(-1)) 30 min before injection with pentylenetetrazol (PTZ, 60 mg·kg(-1), i.p.). The anti-seizure effects of magnolol were analysed using seizure models of behaviour, EEG and in vitro electrophysiology and c-Fos expression in the hippocampus and cortex. KEY RESULTS: Magnolol at doses of 40 and 80 mg·kg(-1) significantly delayed the onset of myoclonic jerks and generalized clonic seizures, and decreased the seizure stage and mortality compared with those of the vehicle-treated animals. EEG recordings showed that magnolol (40 and 80 mg·kg(-1)) prolonged the latency of seizure onset and decreased the number of seizure spikes. The anti-epileptic effect of magnolol was reversed by the GABA(A)/benzodiazepine receptor antagonist flumazenil. Pretreatment with flumazenil decreased the effects of magnolol on prolongation of seizure latency and decline of seizure stage. In a Mg(2+)-free model of epileptiform activity, using multi-electrode array recordings in mouse hippocampal slices, magnolol decreased spontaneous epileptiform discharges. Magnolol also significantly decreased seizure-induced Fos immunoreactivity in the piriform cortex, dentate gyrus and hippocampal area CA1. These effects were attenuated by pretreatment with flumazenil. CONCLUSIONS AND IMPLICATIONS: These findings indicate that the inhibitory effects of magnolol on epileptiform activity were mediated by the GABA(A) /benzodiazepine receptor complex.

A genetic locus controlling aging-sensitive regression of B lymphopoiesis in an autoimmune-prone MRL/lpr strain of mice.

Aging readily affects immune system under the influence of environmental and/or intrinsic factors while accelerating the development of various immune disorders including autoimmune diseases. Little is known about molecular and cellular mechanisms connecting between immune senescence and development of autoimmune diseases. Here, we first show strain-specific and aging-sensitive onset of B-cell abnormality in a lupus-prone MRL/Mp.Fas(lpr) (MRL/lpr) strain of mice. This abnormality was characterized by the regression of B lymphopoiesis in the bone marrow of this strain. We next examined the association between the B-cell regression and onset of autoimmune diseases in aged (MRL/lpr x C3H/He.Fas(lpr)) F2 mice, in which pathologic phenotypes, such as glomerulonephritis, vasculitis, sialoadenitis and arthritis, variously developed. We also searched whole genome to identify genetic loci linked to the B-cell regression by using the same F2 mice. The B-cell regression manifested in the spleen of F2 mice was retrospectively evaluated by reverse transcriptase-based PCR quantification. The results demonstrated that the onset of autoimmune diseases in the F2 mice was not associated with the aging-sensitive B-cell regression. The genetic study identified a significant locus responsible for the B-cell regression in the vicinity of D5Mit233 (29 cM). This is first evidence for the presence of a genetic locus that affects B lymphopoiesis in an aging-sensitive manner.

Modafinil exerts a dose-dependent antiepileptic effect mediated by adrenergic alpha1 and histaminergic H1 receptors in mice.

Epilepsy is characterized by neuronal hyperexcitability and hypersynchronization. Disruption of electroencephalographically (EEG) synchronized epileptiform discharges may be a possible therapy for epilepsy. In the present study, to clarify the role of EEG desynchronization on epilepsy, we investigated the effect of modafinil, a potent wake-promoting substance with EEG desynchronization activity, on epilepsy in mice and clarified the receptors involved in the suppression of seizure caused by maximal electroshock (MES) and pentylenetetrazol (PTZ) kindling models. Modafinil given at 22.5, 45, and 90 mg/kg, i.p. significantly decreased the incidence of tonic hindleg extension in MES seizure models, and protected against PTZ-induced convulsive behaviors in a dose-dependent manner. In addition, modafinil at 180 mg/kg exerted an antiepileptic effect in the MES model; however, at the same dosage it increased the seizure stage in the PTZ-kindling model. The antiepileptic effect in both MES and PTZ models was antagonized by the adrenergic alpha(1) receptor antagonist terazosin, but not by the adrenergic alpha(2) receptor antagonist yohimbine or by dopaminergic receptor antagonists, SCH-23390 (for D(1) receptors) and haloperidol (for D(2) ones). Pyrilamine, a histaminergic H(1) receptor antagonist, counteracted the antiepileptic action of modafinil in the PTZ induced-kindling model, but not in the MES seizure model. Taken together, the present findings indicate that modafinil exerted its antiepileptic effect via adrenergic alpha(1) and histaminergic H(1) receptors, and might be of potential use in the treatment of epilepsy.

A novel autoimmune pancreatitis model in MRL mice treated with polyinosinic:polycytidylic acid.

In this study we established a new animal model for exploring the pathogenesis of autoimmune pancreatitis. We have found previously that MRL/Mp-+/+(MRL/+) mice develop pancreatitis spontaneously by an autoimmune mechanism but only when they are more than 34 weeks old. Because this disease might be a model of multi-factorial diseases controlled by genetic and environmental factors, beginning at 6 weeks old, we injected polyinosinic:polycytidylic acid (poly I:C) into MRL/+ mice and in addition, into MRL/Mp mice bearing the Fas deletion mutant gene, lpr (MRL/lpr). Poly I:C induced chronic severe pancreatitis in all the MRL/+ mice and to a lesser extent in the MRL/lpr mice by 18 weeks of age. There was no pancreatitis in control mice of both strains at the same age. Other than chronic pancreatitis, no severe autoimmune diseases were observed in MRL/+ mice. Immunohistochemical examinations revealed predominant infiltration of CD4+ T cells and Mac-2+ activated macrophages in the pancreatic lesions. Splenic expression of the mRNAs for TNF-alpha and IL-10, which is known to suppress the development of pancreatitis, were increased in both strains of mice. These findings suggest that an MRL strain of mice treated with poly I:C might be a good model for developing new approaches to the study of the pathogenesis of autoimmune pancreatitis.

Dominant localization of prostaglandin D receptors on arachnoid trabecular cells in mouse basal forebrain and their involvement in the regulation of non-rapid eye movement sleep.

Infusion of prostaglandin (PG) D(2) into the lateral ventricle of the brain induced an increase in the amount of non-rapid eye movement sleep in wild-type (WT) mice but not in mice deficient in the PGD receptor (DP). Immunofluorescence staining of WT mouse brain revealed that DP immunoreactivity was dominantly localized in the leptomeninges (LM) of the basal forebrain but that PGD synthase immunoreactivity was widely distributed in the LM of the entire brain. Electron microscopic observation indicated that DP-immunoreactive particles were predominantly located on the plasma membranes of arachnoid trabecular cells of the LM. The region with the highest DP immunoreactivity was clearly defined as bilateral wings in the LM of the basal forebrain located lateral to the optic chiasm in the proximity of the ventrolateral preoptic area, one of the putative sleep centers, and the tuberomammillary nucleus, one of the putative wake centers. The LM of this region contained DP mRNA 70-fold higher than that in the cortex as judged from the results of quantitative reverse transcription-PCR. PGD(2) infusion into the subarachnoid space of this region increased the extracellular adenosine level more than 2-fold in WT mice but not in the DP-deficient mice. These results indicate that DPs in the arachnoid trabecular cells of the basal forebrain mediate an increase in the extracellular adenosine level and sleep induction by PGD(2).

Arousal effect of orexin A depends on activation of the histaminergic system.

Orexin neurons are exclusively localized in the lateral hypothalamic area and project their fibers to the entire central nervous system, including the histaminergic tuberomammillary nucleus (TMN). Dysfunction of the orexin system results in the sleep disorder narcolepsy, but the role of orexin in physiological sleep-wake regulation and the mechanisms involved remain to be elucidated. Here we provide several lines of evidence that orexin A induces wakefulness by means of the TMN and histamine H(1) receptor (H1R). Perfusion of orexin A (5 and 25 pmol/min) for 1 hr into the TMN of rats through a microdialysis probe promptly increased wakefulness for 2 hr after starting the perfusion by 2.5- and 4-fold, respectively, concomitant with a reduction in rapid eye movement (REM) and non-REM sleep. Microdialysis studies showed that application of orexin A to the TMN increased histamine release from both the medial preoptic area and the frontal cortex by approximately 2-fold over the baseline for 80 to 160 min in a dose-dependent manner. Furthermore, infusion of orexin A (1.5 pmol/min) for 6 hr into the lateral ventricle of mice produced a significant increase in wakefulness during the 8 hr after starting infusion to the same level as the wakefulness observed during the active period in wild-type mice, but not at all in H1R gene knockout mice. These findings strongly indicate that the arousal effect of orexin A depends on the activation of histaminergic neurotransmission mediated by H1R.

Genetic dissection of vasculitis in MRL/lpr lupus mice: a novel susceptibility locus involving the CD72c allele.

An MRL/MpJ strain of mice bearing the Fas deletion mutant gene, lpr (MRL/lpr), composed of genomes derived from LG/J, AKR/J, C3H/Di and C57BL/6J mice, develops systemic vasculitis coincidentally with other collagen diseases, but a C3H/HeJ-lpr/lpr (C3H/lpr) strain does not. In a genome-wide screening of the MRL background genes mediating susceptibility to collagen diseases using N2 progeny mice MRL/lpr x (MRL/lpr x C3H/lpr)F1, we previously found that each collagen disease is controlled by a different set of genes. To clarify the candidate genes for vasculitis, we extended the linkage analysis of renal vasculitis to a larger number of N2 mice and to F2 intercross mice. Two distinct recessive susceptibility loci for vasculitis were mapped on chromosome (Chr) 4 at D4Mit89 and D4Mit147 in both progenies. The former was a novel locus for lupus phenotypes, which involved the MRL allele CD72(c) in contrast to the C3H allele CD72(b). The one on Chr 3 was a recessive locus which had an inhibitory effect on vasculitis. From their composition these loci seemed to be derived from AKR/J (for one) and LG/J (for another two) strains, and appeared to act in an additive manner on the development of vasculitis, indicating that vasculitis in MRL/lpr mice is inherited in a polygenic manner.

Genetic dissection of the complex pathological manifestations of collagen disease in MRL/lpr mice.

An MRL strain of mice bearing a Fas-deletion mutant gene, lpr, MRL/MpJ-lpr/lpr (MRL/lpr) develops collagen disease involving vasculitis, glomerulonephritis, arthritis and sialoadenitis, each of which has been studied as a model for polyarteritis, lupus nephritis, rheumatoid arthritis and Sjögren's syndrome, respectively. Development of such lesions seems dependent on host genetic background since the congenic C3H/HeJ-lpr/lpr (C3H/lpr) mice rarely develop them. To identify the gene loci affecting each lesion, a genetic dissection of these complex pathological manifestations was carried out. First, histopathological features in MRL/lpr, C3H/lpr, (MRL/lpr x C3H/lpr) F1 intercross, and MRL/lpr x (MRL/lpr x C3H/lpr) F1 backcross mice were analyzed. Genomic DNA of the backcross mice were subjected to association studies by Chi-squared analysis for determining which polymorphic microsatellite locus occurs at higher frequency among affected compared to unaffected individuals for each lesion. As a result, gene loci recessively associated with each lesion were mapped on different chromosomal positions. We concluded that each of these lesions in MRL/lpr mice is under the control of a different set of genes, suggesting that the complex pathological manifestations of collagen disease result from polygenic inheritance.

Effects of intraperitoneal Ringer's lactate instillation and infusion on postsurgical adhesion formation.

We compared the effects of continuous Ringer's lactate infusion and Ringer's lactate instillation on postsurgical adhesion formation in the rat model. The results show that intraperitoneal Ringer's lactate instillation reduces postsurgical adhesions. However, its efficacy is not improved by additional infusion of Ringer's lactate.