Measuring neutrophil functions might be a good predictive marker of overtraining in athletes

In order to develop a predictive marker of overtraining in athletes, we examined the changes in neutrophil function [reactive oxygen species (ROS) production capability and phagocytic activity (PA)] for 10 male and 13 female judoists attending a training camp. Measurements were taken four times in total—immediately before and after a 2 h unified exercise loading (UEL) performed 1 day before (Pre‐Camp) and the day after the 7 day training camp (Post‐Camp). UEL‐mediated aspartate aminotransferase was higher at Post‐Camp than at Pre‐Camp in females but not in males. Post‐Camp leukocyte/neutrophil counts after the UEL significantly increased in females but not in males. The rate of change in C4 was significantly smaller in females than in males at Post‐Camp. Only ROS significantly decreased without any compensation (increase in PA) being made at Post‐Camp in females. In conclusion, this finding, namely that ROS significantly decreased only at Post‐Camp without any compensatory mechanism (increase in PA), would suggest that the training camp imposed greater loading on females than males. This consideration was supported by the atypical aspartate aminotransferase, leukocyte/neutrophil counts and C4 findings which were seen at Post‐Camp only in females. Therefore, regularly examining neutrophil functions such as ROS and PA might be a good preventative measure against overtraining in athletes participating in training camps. Copyright © 2008 John Wiley & Sons, Ltd.     

Role of interleukin-12 in determining differential kinetics of invariant natural killer T cells in response to differential burden of Listeria monocytogenes

Invariant (i) natural killer (NK) T cells are unique T lymphocytes expressing NKR-P1B/C (NK1.1), which recognize glycolipids, notably alpha-galactosylceramide (alpha-GalCer) presented by CD1d. The characteristic phenotype of these iNKT cells undergoes dramatic changes following Listeria monocytogenes infection, and interleukin (IL)-12 is involved in these alterations. Here we show that liver iNKT cells in mice are differentially influenced by the load of infection. Liver alpha-GalCer/CD1d tetramer-reactive (alpha-GalCer/CD1d(+)) T cells expressing NK1.1 became undetectable by day 2 following L. monocytogenes infection and concomitantly cells lacking NK1.1 increased regardless of the severity of infection. Whereas alpha-GalCer/CD1d(+)NK1.1(+) T cells remained virtually undetectable on day 4 following low-dose infection, considerable numbers of these cells were detected in high-dose-infected mice. Whereas numbers of IL-12 producers in the liver on day 4 post infection were comparable in low- and high-dose-infected mice without in vitro restimulation with heat-killed Listeria, those were more prominent in low-dose-infected mice than in high-dose-infected mice after restimulation despite the fact that higher numbers of macrophages and granulocytes infiltrated the liver in high-dose-infected mice than in low-dose-infected mice. Our results indicate that NK1.1 surface expression on iNKT cells is differentially modulated by the burden of infection, and suggest that a high bacterial load probably causes loss of IL-12 production.     

Activation-induced cytidine deaminase deficiency causes organ-specific autoimmune disease

Activation-induced cytidine deaminase (AID) expressed by germinal center B cells is a central regulator of somatic hypermutation (SHM) and class switch recombination (CSR). Humans with AID mutations develop not only the autosomal recessive form of hyper-IgM syndrome (HIGM2) associated with B cell hyperplasia, but also autoimmune disorders by unknown mechanisms. We report here that AID-/- mice spontaneously develop tertiary lymphoid organs (TLOs) in non-lymphoid tissues including the stomach at around 6 months of age. At a later stage, AID-/- mice develop a severe gastritis characterized by loss of gastric glands and epithelial hyperplasia. The disease development was not attenuated even under germ-free (GF) conditions. Gastric autoantigen -specific serum IgM was elevated in AID-/- mice, and the serum levels correlated with the gastritis pathological score. Adoptive transfer experiments suggest that autoimmune CD4+ T cells mediate gastritis development as terminal effector cells. These results suggest that abnormal B-cell expansion due to AID deficiency can drive B-cell autoimmunity, and in turn promote TLO formation, which ultimately leads to the propagation of organ-specific autoimmune effector CD4+ T cells. Thus, AID plays an important role in the containment of autoimmune diseases by negative regulation of autoreactive B cells.     

Ventilatory long-term facilitation in mice can be observed during both sleep and wake periods and depends on orexin.

Respiratory long-term facilitation (LTF) is a long-lasting (>1 h) augmentation of respiratory motor output that occurs even after cessation of hypoxic stimuli, is serotonin-dependent, and is thought to prevent sleep-disordered breathing such as sleep apnea. Raphe nuclei, which modulate several physiological functions through serotonin, receive dense projections from orexin-containing neurons in the hypothalamus. We examined possible contributions of orexin to ventilatory LTF by measuring respiration in freely moving prepro-orexin knockout mice (ORX-KO) and wild-type (WT) littermates before, during, and after exposure to intermittent hypoxia (IH; 5 x 5 min at 10% O2), sustained hypoxia (SH; 25 min at 10% O2), or sham stimulation. Respiratory data during quiet wakefulness (QW), slow wave sleep (SWS), and rapid-eye-movement sleep were separately calculated. Baseline ventilation before hypoxic stimulation and acute responses during stimulation did not differ between the ORX-KO and WT mice, although ventilation depended on vigilance state. Whereas the WT showed augmented minute ventilation (by 20.0 +/- 4.5% during QW and 26.5 +/- 5.3% during SWS; n = 8) for 2 h following IH, ORX-KO showed no significant increase (by -3.1 +/- 4.6% during QW and 0.3 +/- 5.2% during SWS; n = 8). Both genotypes showed no LTF after SH or sham stimulation. Sleep apnea indexes did not change following IH, even when LTF appeared in the WT mice. We conclude that LTF occurs during both sleep and wake periods, that orexin is necessary for eliciting LTF, and that LTF cannot prevent sleep apnea, at least in mice.     

Tissue-engineered smooth muscle cell and endothelial progenitor cell bi-level cell sheets prevent progression of cardiac dysfunction,microvascular dysfunction,and interstitial fibrosis in a rodent model of type 1 diabetes-induced cardiomyopathy

Background

Diabetes mellitus is a risk factor for coronary artery disease and diabetic cardiomyopathy, and adversely impacts outcomes following coronary artery bypass grafting. Current treatments focus on macro-revascularization and neglect the microvascular disease typical of diabetes mellitus-induced cardiomyopathy (DMCM). We hypothesized that engineered smooth muscle cell (SMC)-endothelial progenitor cell (EPC) bi-level cell sheets could improve ventricular dysfunction in DMCM.

Methods

Primary mesenchymal stem cells (MSCs) and EPCs were isolated from the bone marrow of Wistar rats, and MSCs were differentiated into SMCs by culturing on a fibronectin-coated dish. SMCs topped with EPCs were detached from a temperature-responsive culture dish to create an SMC-EPC bi-level cell sheet. A DMCM model was induced by intraperitoneal streptozotocin injection. Four weeks after induction, rats were randomized into 3 groups: control (no DMCM induction), untreated DMCM, and treated DMCM (cell sheet transplant covering the anterior surface of the left ventricle).

Results

SMC-EPC cell sheet therapy preserved cardiac function and halted adverse ventricular remodeling, as demonstrated by echocardiography and cardiac magnetic resonance imaging at 8 weeks after DMCM induction. Myocardial contrast echocardiography demonstrated that myocardial perfusion and microvascular function were preserved in the treatment group compared with untreated animals. Histological analysis demonstrated decreased interstitial fibrosis and increased microvascular density in the SMC-EPC cell sheet-treated group.

Conclusions

Treatment of DMCM with tissue-engineered SMC-EPC bi-level cell sheets prevented cardiac dysfunction and microvascular disease associated with DMCM. This multi-lineage cellular therapy is a novel, translatable approach to improve microvascular disease and prevent heart failure in diabetic patients.

     

On-site Direct Detection of Astaxanthin from Salmon Fillet Using Raman Spectroscopy

A new technology employing Raman spectroscopy is attracting attention as a powerful biochemical technique for the detection of beneficial and functional food nutrients, such as carotenoids and unsaturated fatty acids. This technique allows for the dynamic characterization of food nutrient substances for the rapid determination of food quality. In this study, we attempt to detect and measure astaxanthin from salmon fillets using this technology. The Raman spectra showed specific bands corresponding to the astaxanthin present in salmon and the value of astaxanthin (Raman band, 1518 cm^?1) relative to those of protein/lipid (Raman band, 1446 cm^?1) in the spectra increased in a dose-dependent manner. A standard curve was constructed by the standard addition method using astaxanthin as the reference standard for its quantification by Raman spectroscopy. The calculation formula was established using the Raman bands typically observed for astaxanthin (i.e., 1518 cm^?1). In addition, we examined salmon fillets of different species (Atlantic salmon, coho salmon, and sockeye salmon) and five fillets obtained from the locations (from the head to tail) of an entire Atlantic salmon. Moreover, the sockeye salmon fillet exhibited the highest astaxanthin concentration (14.2 mg/kg), while coho salmon exhibited an intermediate concentration of 7.0 mg/kg. The Raman-based astaxanthin concentration in the five locations of Atlantic salmon was more strongly detected from the fillet closer to the tail. From the results, a rapid, convenient Raman spectroscopic method was developed for the detection of astaxanthin in salmon fillets.     

Ammonia fermentation, a novel anoxic metabolism of nitrate by fungi.

The induction of fungal denitrification by Fusarium oxysporum requires a minimal amount of O(2), although excess O(2) completely represses this process (Zhou, Z., Takaya, N., Sakairi, M. A. C., and Shoun, H. (2001) Arch. Microbiol. 175, 19-25). Here we describe another metabolic mechanism of nitrate in fungal cells, termed ammonia fermentation, that supports growth under conditions more anoxic than those of denitrification. The novel nitrate metabolism of eukaryotes consists of the reduction of nitrate to ammonium coupled with the catabolic oxidation of electron donors to acetate and substrate-level phosphorylation. F. oxysporum thus has two pathways of dissimilatory nitrate reduction that are alternatively expressed in response to environmental O(2) tension. F. oxysporum prefers O(2) respiration when the O(2) supply is sufficient. We discovered that this fungus is the first eukaryotic, facultative anaerobe known to express one of three distinct metabolic energy mechanisms closely depending on environmental O(2) tension. We also showed that ammonia fermentation occurs in many other fungi that are common in soil, suggesting that facultative anaerobes are widely distributed among fungi that have been considered aerobic organisms.