Altered sarcoplasmic reticulum function in rat diaphragm after high-intensity exercise

The present study examined the effects of acute high-intensity exercise on Ca(2+) uptake and release rates and Ca(2+)-adenosine triphosphatase (ATPase) activity of the sarcoplasmic reticulum (SR) from the costal diaphragm. The rats were run on a treadmill at an estimated requirement of 100% of maximal O2 consumption until fatigued (average time to exhaustion: 4.79 min). Muscle lactate and inorganic phosphate after exercise were increased by 65% (P < 0.05) and 35% (P < 0.05), respectively. With exercise, Ca(2+) uptake and release, which were detected in homogenates using the Ca(2+) fluorescent dye indo-1, were decreased by 24% (P < 0.05) and 22% (P < 0.05), respectively. The reduction in Ca(2+) uptake was paralleled by decreased activity of SR Ca(2+)-ATPase in both the absence and presence of Ca(2+) ionophore. These findings demonstrate that, in the diaphragm as well as in the locomotor muscles that have been explored in previous studies, the attenuations of the SR function is brought about by acute high-intensity exercise. These changes in the SR of the diaphragm may contribute, at least in part, to deteriorations in exercise tolerance and work productivity resulting from repetitive physical activities.     

Epigallocatechin Gallate, a Potential Immunomodulatory Agent of Tea Components, Diminishes Cigarette Smoke Condensate-Induced Suppression of Anti-Legionella pneumophila Activity and Cytokine Responses of Alveolar Macrophages

Even though cigarette smoking has been shown to suppress immune responses in the lungs, little is known about the effect of cigarette smoke components on respiratory infections. In the present study, the effects of cigarette smoke condensate (CSC) on bacterial replication in alveolar macrophages and the immune responses of macrophages to infection were examined. Furthermore, a possible immunotherapeutic effect of epigallocatechin gallate (EGCg), a major form of tea catechins, on the CSC-induced suppression of antimicrobial activity and immune responses of alveolar macrophages was also determined. The treatment of murine alveolar macrophage cell line (MH-S) cells with CSC significantly enhanced the replication of Legionella pneumophila in macrophages and selectively down-regulated the production of interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-α) induced by bacterial infection. The treatment of macrophages with EGCg not only overcame the CSC-induced suppression of antimicrobial activity but also strengthened the resistance of macrophages to infection. EGCg also markedly up-regulated the CSC-suppressed IL-6 and TNF-α production by macrophages in response to infection. The results of exogenous TNF-α treatment and neutralization treatment with anti-TNF-α and anti-gamma-interferon (IFN-γ) antibodies and the determination of IFN-γ mRNA levels indicate that CSC-suppressed macrophages can be activated by EGCg to inhibit L. pneumophila growth by up-regulation of TNF-α and IFN-γ production. Thus, this study revealed that CSC selectively alters the immune responses of macrophages to L. pneumophila infection and leads to an enhancement of bacterial replication in macrophages. In addition, the tea catechin EGCg can diminish such suppressive effects of CSC on alveolar macrophages.     

Abnormal distributions of callosal commissural and corticothalamic neurons in the cerebral neocortex of Shaking Rat Kawasaki

It is generally believed that haloperidol exerts its motor side effects and therapeutic effects mainly by antagonizing dopamine D(2) receptors in the striatum and the nucleus accumbens, respectively. Several neurotransmitters/modulators, including glutamate, acetylcholine, adenosine and histamine, affect dopaminergic activity in these centers. We have recently shown that N-methyl-D-aspartate receptor-mediated modulation of haloperidol-induced c-fos expression differs in functionally specific regions of the striatum and the nucleus accumbens. In the present study, the entire striatum and the nucleus accumbens were comprehensively examined for the pattern of modulation of haloperidol-induced c-fos expression by adenosine A(2), histamine H(3) and muscarinic receptor antagonists.Blockade of muscarinic and H(3) receptors resulted in a profound suppression of haloperidol-induced c-fos expression in the dorsolateral part of the striatum. In addition, the H(3) receptor antagonist suppressed the effects of haloperidol in the ventrolateral aspect of the striatum and the rostral parts of the medial striatum. Muscarinic receptor antagonists suppressed haloperidol-induced c-fos expression throughout the shell and in the mid-level of the core of the nucleus accumbens while A(2) and H(3) receptor antagonists did not.We found that the muscarinic and H(3) receptor antagonists suppress the induction of c-fos by haloperidol in the dorsolateral aspect of the striatum, an area implicated in the development of extrapyramidal motor symptoms following chronic haloperidol treatment. By contrast, haloperidol-induced c-fos expression in the nucleus accumbens, an area implicated in the therapeutic effects of haloperidol, was suppressed by the muscarinic receptor antagonist, but not by the H(3) receptor antagonist. Therefore we conclude that H(3) receptor modulation may provide a useful therapeutic target in future efforts to minimize neuroleptic-induced motor side effects.     

N-acetylcysteine fails to modulate the in vitro function of sarcoplasmic reticulum of diaphragm in the final phase of fatigue

AIM: In the present study, we tested the hypothesis whether N-acetylcysteine (NAC), a non-specific antioxidant, might influence fatigue by modulating Ca2+-handling capacity by the sarcoplasmic reticulum (SR). METHODS: In the presence (10 mm) or absence of NAC, bundles of rat diaphragm were stimulated with tetanic trains (350 ms, 30-40 Hz) at 1 train every 2 s for 300 s. SR functions, as assessed by SR Ca2+-uptake and release rates and SR Ca2+-ATPase activity, were measured in vitro on muscle homogenates. RESULTS: Following the 300-s stimulation, the force developed by NAC-treated muscles is approximately 1.8-fold higher (P < 0.05) than that of muscles without NAC treatment. Stimulation elicited an 18-30% depression in SR function (P < 0.05). Despite the differing degrees of fatigue between NAC-treated and non-treated muscles, SR functions in these muscles were reduced to similar extents. CONCLUSIONS: These results suggest that modulation of SR function measured in vitro may not be a major contributor to inhibition of diaphragmic fatigue with antioxidant, at least, in the final phase of fatigue where force output is remarkably reduced.     

Staphylococcal scalded skin syndrome in an extremely low‐birth‐weight neonate: Molecular characterization and rapid detection by multiplex and real‐time PCR of methicillin‐resistant Staphylococcus aureus

Background: Staphylococcal scalded skin syndrome (SSSS), caused by methicillin‐resistant Staphylococcus aureus (MRSA) producing exfoliative toxin (ET), is a life‐threatening infection for neonates in neonatal intensive care units (NICUs). SSSS in extremely low‐birth‐weight (ELBW) neonates is rare. A new class of MRSA (community‐acquired MRSA, CA‐MRSA) has been emerging in the community. The aim of this study was to characterize MRSA from an ELBW neonate with SSSS, and to develop rapid detection methods for SSSS‐associated and emerging pediatric MRSA. Methods: An ELBW infant in the NICU developed SSSS on day 16 after birth. Isolated MRSA was genetically characterized and compared with CA‐MRSA from bullous impetigo (biCA‐MRSA), which is positive for the ET and collagen‐adhesin (CNA) genes in many cases, and the Panton‐Valentine leucocidin (PVL) gene rarely. Specific primers and probes for five virulence genes (for ETA, ETB, ETD, PVL, CNA) were designed for multiplex polymerase chain reaction (PCR) and real‐time PCR. Results: MRSA strain H5 from SSSS exhibited the genotype (ST91, spa416[t375], agr3, SCCmecIVa, CoaI), and possessed the ETB and CNA genes, similar to ST91 biCA‐MRSA (albeit with a divergence). Multiplex PCR detected the ETB and CNA genes of strain H5, and real‐time PCR detected strain H5 at as low as 10² CFU/mL. The assays were 100% specific and 100% sensitive, for the five virulence genes. Conclusion: ETB‐positive ST91 MRSA, which was very similar to ST91 biCA‐MRSA, was isolated from an ELBW infant with SSSS. The multiplex and real‐time PCR assays specifically or quantitatively detected SSSS‐associated and emerging pediatric MRSA.     

The brain‐derived neutrophic factor val66met polymorphism and sudden unexpected infant death

Aim: Findings of hypoxia prior to death and involvement of a dysregulation of the serotonergic network in sudden infant death syndrome (SIDS) may indicate that brain‐derived neutrophic factor (BDNF) also is of importance with regard to sudden unexpected infant death. Based on this, the purpose of this study was to investigate the BDNF val66met polymorphism in SIDS cases, cases of infectious death and controls. Methods: The polymorphism was investigated in 163 SIDS cases, 34 cases of infectious death and 121 controls, using real‐time PCR and fluorescence melting curve analysis. Results: There were no differences in val66met genotype distribution between neither the SIDS cases nor the cases of infectious death and controls (p = 0.95 and p = 0.52, respectively). Conclusion: The study indicates that the val66met polymorphism is not important for sudden unexpected infant death. However, several other SNPs in the BDNF gene, as well as in other genes involved in this pathway, including G‐protein, have to be investigated to fully exclude any involvement of BDNF in SIDS.     

Dietary nucleic acid and intestinal microbiota synergistically promote a shift in the Th1/Th2 balance toward Th1-skewed immunity

BACKGROUND: Intestinal microbiota are known to play an important role in the establishment of oral tolerance, thereby protecting the organism from food allergies. Dietary intake of nucleic acid (NA) is also reported to have such an anti-allergic effect; however, one unsolved question is whether or not dietary NA would act through a process of toll-like receptor 9 signaling activated by DNA containing a CpG motif, a well-known sequence leading to immunostimulatory activity. In this study, we focused on the question of whether the addition of dietary NA lacking CpG motifs would allow continued modulation of the Th1/Th2 balance. METHODS: Germ free (GF) and Bifidobacterium-infantis-monoassociated BALB/c mice were maintained on either an NA-free casein diet or on an NA-supplemented casein diet for 4 weeks. Thereafter, both the in vivo anti-casein antibody levels and in vitro splenocyte cytokine secretion pattern were evaluated. RESULTS: Feeding with a casein diet elicited a substantial increase in the serum anti-casein-specific IgG1, IgG2a, and IgE levels of GF mice fed the NA free-diet. The in vitro cytokine production profile showed that enhanced IL-4 production in the GF mice fed the NA free-diet was markedly reduced by the supplementation with dietary NA in both the GF and B.-infantis-monoassociated mice. In addition, IFN-gamma secretion increased in the B.-infantis-reconstituted mice fed the diet containing NA. CONCLUSIONS: These results suggest that dietary intake of NA devoid of CpG motifs may prevent the development of allergies via acceleration of Th1-dominant immunity.     

IL-6 induced by double-stranded RNA augments allergic inflammation via suppression of Foxp3+ T-cell/IL-10 axis

Activation of innate immunity against viruses in the respiratory tracts affects the development of asthma. Most respiratory viruses generate double-stranded (ds)RNA during their replication. We recently showed that a low-dose administration of polyinosinic polycytidylic acid (poly IC), a mimetic of viral dsRNA, during allergen sensitization augments airway eosinophilia and hyperresponsiveness in mice via enhanced production of IL-13 from T cells. However, a phenotype of asthma under severer load of dsRNA remains unknown. d-galactosamine (d-GalN) is known as a strong sensitizer of poly IC. Mice were treated with poly IC plus d-GalN during allergen sensitization. A sublethal dose of poly IC/d-GalN augmented airway eosinophilia and CD4(+) T-cell accumulation in the lungs but not airway hyperresponsiveness. The augmented inflammation was associated with decreased IL-10 in the bronchoalveolar lavage fluid and decreased Foxp3(+) regulatory T cells in the lungs. Serum IL-6 was prominently higher in the mice treated with poly IC/d-GalN than in that with poly IC alone or d-GalN alone. Poly IC/d-GalN did not affect IL-17-producing T cells in the lungs. Poly IC/d-GalN failed to augment airway eosinophilia after anti-IL-10 receptor monoclonal antibody treatment during allergen challenge. Finally, anti-IL-6 receptor monoclonal antibody treatment before poly IC/d-GalN completely prevented the decrease of IL-10 and Foxp3(+) regulatory T cells and the augmentation of airway inflammation. These results indicate that enhanced production of IL-6 by poly IC/d-GalN induces the augmentation of allergic inflammation via suppression of Foxp3(+) regulatory T-cell/IL-10 axis. IL-6 may be a target for preventing asthma augmentation related to severe virus infection.     

Glycyrrhizin and isoliquiritigenin suppress the LPS sensor toll-like receptor 4/MD-2 complex signaling in a different manner

Recent evidences suggest that the extracts of plant products are able to modulate innate immune responses. A saponin GL and a chalcone ILG are representative components of Glycyrrhiza uralensis, which attenuate inflammatory responses mediated by TLRs. Here, we show that GL and ILG suppress different steps of the LPS sensor TLR4/MD-2 complex signaling at the receptor level. Extract of G. uralensis suppressed IL-6 and TNF-α production induced by lipid A moiety of LPS in RAW264.7 cells. Among various G. uralensis-related components of saponins and flavanones/chalcones, GL and ILG could suppress IL-6 production induced by lipid A in dose-dependent manners in RAW264.7 cells. Furthermore, elevation of plasma TNF-α in LPS-injected mice was attenuated by passive administration of GL or ILG. GL and ILG inhibited lipid A-induced NF-κB activation in Ba/F3 cells expressing TLR4/MD-2 and CD14 and BMMs. These components also inhibited activation of MAPKs, including JNK, p38, and ERK in BMMs. In addition, GL and ILG inhibited NF-κB activation and IL-6 production induced by paclitaxel, a nonbacterial TLR4 ligand. Interestingly, GL attenuated the formation of the LPS-TLR4/MD-2 complexes, resulting in inhibition of homodimerization of TLR4. Although ILG did not affect LPS binding to TLR4/MD-2, it could inhibit LPS-induced TLR4 homodimerization. These results imply that GL and ILG modulate the TLR4/MD-2 complex at the receptor level, leading to suppress LPS-induced activation of signaling cascades and cytokine production, but their effects are exerted at different steps of TLR4/MD-2 signaling.