Aerobic exercise training-induced alteration of gut microbiota composition affects endurance capacity.

This study aimed to clarify whether aerobic exercise training-induced alterations in the gut microbiota affect physiological adaptation with endurance exercise capacity. In study 1, ICR mice were randomly divided into three groups: vehicle intake + sedentary (V+S), vehicle intake + exercise training (V+Ex) and antibiotic intake + exercise training (AB+Ex). In the exercise training groups, treadmill running was performed for 8 weeks. During the exercise training intervention, the antibiotic-intake group freely drank water containing antibiotics. In study 2, ICR mice were randomly divided into three groups: Sham, transplantation of caecum microbiota from sedentary mice (Sed-CMT) and exercise training mice (Ex-CMT). In study 1, the treadmill running time to exhaustion, an index of maximal aerobic capacity, after aerobic exercise training in the V+Ex group was significantly longer than that in the V+S and AB+Ex groups. Gastrocnemius muscle citrate synthase (CS) activity and PGC-1α protein levels in the V+Ex group were significantly higher than in the V+S and AB+Ex groups. The bacterial Erysipelotrichaceae and Alcaligenaceae families were positively correlated with treadmill running time to exhaustion. In study 2, the treadmill running time to exhaustion after transplantation was significantly higher in the Ex-CMT group than in the Sham and Sed-CMT groups. Furthermore, CS activity and PGC-1α protein levels in the gastrocnemius muscle were significantly higher in the Ex-CMT group than in the Sham and Sed-CMT groups. Thus, gut microbiota altered by aerobic exercise training may be involved in the augmentation of endurance capacity and muscle mitochondrial energy metabolism. KEY POINTS: Aerobic exercise training changes gut microbiota composition, and the Erysipelotrichaceae and Alcaligenaceae families were among the altered gut bacteria. The gut microbiota was associated with endurance performance and metabolic regulator levels in skeletal muscle after aerobic exercise training. Continuous antibiotic treatment attenuated the increase in endurance performance, citrate synthase activity and PGC-1α levels in skeletal muscle induced by aerobic exercise training. Gut microbiota transplantation from exercise-trained mice improved endurance performance and metabolic regulator levels in recipient skeletal muscle, despite the absence of aerobic exercise training.

Icing after skeletal muscle injury with necrosis in a small fraction of myofibers limits inducible nitric oxide synthase-expressing macrophage invasion and facilitates muscle regeneration.

Growing evidence from animal experiments suggests that icing after skeletal muscle injury is harmful to muscle regeneration. However, these previous experimental models yielded massive necrotic myofibers, whereas muscle injury with necrosis in a small myofiber fraction (<10%) frequently occurs in human sports activities. Although macrophages play a proreparative role during muscle regeneration, they exert a cytotoxic effect on muscle cells through an inducible nitric oxide synthase (iNOS)-mediated mechanism. In this study, we established an animal injury model with necrosis limited to a small myofiber fraction and investigated the effect of icing on muscle regeneration with a focus on macrophage-related events. Icing after muscle injury of this model resulted in an enlarged size of regenerating myofibers compared with those in untreated animals. During the regenerative process, icing attenuated the accumulation of iNOS-expressing macrophages, suppressed iNOS expression in the whole damaged muscle, and limited the expansion of the injured myofiber area. In addition, icing increased the ratio of M2 macrophages within the injured site at an earlier time point than that in untreated animals. Following these phenomena in icing-treated muscle regeneration, an early accumulation of activated satellite cells within the damaged/regenerating area occurred. The expression level of myogenic regulatory factors, such as MyoD and myogenin, was not affected by icing. Taken together, our results suggest that icing after muscle injury with necrosis limited to a small fraction of myofibers facilitates muscle regeneration by attenuating iNOS-expressing macrophage invasion, limiting muscle damage expansion, and accelerating the accumulation of myogenic cells which form regenerating myofibers.

Antidepressant properties of voluntary exercise mediated by gut microbiota.

Although regular exercise has been reported to prevent depression, it has not been clarified whether the gut microbiota is involved in the factors that prevent depression through exercise. We investigated the effects of voluntary exercise on the gut microbiota and the prevention of depression-like behaviors using mice. C57BL/6 J male mice were subjected to 10 weeks of sedentary control or wheel running, then they were subjected to social defeat stress (SDS). Exercise attenuated that sucrose drinking was decreased by SDS treatment. Exercise increased the expression of Bdnf and decreased expression of Zo-1 and Claudin5 in the brain. Fecal Turicibacter, Allobaculum, and Clostridium sensu stricto, and propionate in the cecum were decreased by the exercise. Voluntary exercise-induced antidepressant properties might be partially caused by suppression of serotonin uptake into gut microbiota and increase the permeability of the blood-brain barrier via reduced propionate production.

Gain-of-function mutations in ALPK1 cause an NF-κB-mediated autoinflammatory disease: functional assessment, clinical phenotyping and disease course of patients with ROSAH syndrome.

OBJECTIVES: To test the hypothesis that ROSAH (retinal dystrophy, optic nerve oedema, splenomegaly, anhidrosis and headache) syndrome, caused by dominant mutation in ALPK1, is an autoinflammatory disease. METHODS: This cohort study systematically evaluated 27 patients with ROSAH syndrome for inflammatory features and investigated the effect of ALPK1 mutations on immune signalling. Clinical, immunologic and radiographical examinations were performed, and 10 patients were empirically initiated on anticytokine therapy and monitored. Exome sequencing was used to identify a new pathogenic variant. Cytokine profiling, transcriptomics, immunoblotting and knock-in mice were used to assess the impact of ALPK1 mutations on protein function and immune signalling. RESULTS: The majority of the cohort carried the p.Thr237Met mutation but we also identified a new ROSAH-associated mutation, p.Tyr254Cys.Nearly all patients exhibited at least one feature consistent with inflammation including recurrent fever, headaches with meningeal enhancement and premature basal ganglia/brainstem mineralisation on MRI, deforming arthritis and AA amyloidosis. However, there was significant phenotypic variation, even within families and some adults lacked functional visual deficits. While anti-TNF and anti-IL-1 therapies suppressed systemic inflammation and improved quality of life, anti-IL-6 (tocilizumab) was the only anticytokine therapy that improved intraocular inflammation (two of two patients).Patients' primary samples and in vitro assays with mutated ALPK1 constructs showed immune activation with increased NF-κB signalling, STAT1 phosphorylation and interferon gene expression signature. Knock-in mice with the Alpk1 T237M mutation exhibited subclinical inflammation.Clinical features not conventionally attributed to inflammation were also common in the cohort and included short dental roots, enamel defects and decreased salivary flow. CONCLUSION: ROSAH syndrome is an autoinflammatory disease caused by gain-of-function mutations in ALPK1 and some features of disease are amenable to immunomodulatory therapy.

Acetyl-L-carnitine attenuates Poly I:C-induced sickness behavior in mice.

Fatigue is accompanied by a decrease in physical activity or malaise, and might be reduced by acetyl-L-carnitine (ALC) administration. The purpose of this study was to investigate the preventive effects of ALC on Poly I:C-induced sickness behavior in mice. For the experiment, male C3H/HeN mice were used and treated with ALC for 5 days before Poly I:C administration. ALC administration attenuated the decrease in wheel behavior activity of mice at 24 h after Poly I:C administration and ALC-treated mice quickly recovered from the sickness behavior. The gene expression of brain-derived neurotrophic factor (BDNF) in the cerebrum and hippocampus, which is associated with physical activity, was higher in the ALC-treated group. Translocator protein 18kDa (TSPO), which has cytoprotective effects, was up-regulated in the cerebrum and hippocampus, suggesting that ALC suppressed the decrease in activity induced by Poly I:C treatment through enhancement of cytoprotective effects in the brain.

A simple questionnaire for the detection of testosterone deficiency in men with late-onset hypogonadism.

The Aging Males' Symptoms (AMS) score, developed to screen for late-onset hypogonadism (LOH), contains 17 questions regarding mental, physical, and sexual parameters. In the Japanese guidelines, a free testosterone (FT) <8.5 pg/mL is recommended for testosterone treatment. However, previous studies have shown no correlation between total AMS scores and testosterone concentration. We aimed to develop a better questionnaire for the detection of testosterone deficiency in men, for the diagnosis of LOH. In 234 Japanese men, aged 40-64 years, we analyzed the relationships of AMS with serum total testosterone (TT), FT, calculated FT (cFT), and calculated bioavailable testosterone (cBT), and identified useful questions for the detection of testosterone deficiency. Four scores, a decrease in muscular strength, a decrease in ability to perform sexually or the frequency, a decrease in the number of morning erections, and a decrease in sexual desire/libido, were negatively associated with two or more of the above four testosterone parameters, and the sum of these four scores (named the selective score) correlated with TT and cFT, independent of age. Statistical analysis revealed an association between insulin resistance and testosterone deficiency, and a higher selective score in smokers than non-smokers. Cubic function model analysis and logistic regression analysis revealed that selective scores ≥10 corresponded with the testosterone concentrations recommended for the diagnosis of LOH, including FT <8.5 pg/mL, independent of age, insulin resistance, and smoking. Thus, the selective score represents a simple and useful means for screening of testosterone deficiency in Japanese men, as an indicator of LOH.

Effect of dietary cellulose nanofiber and exercise on obesity and gut microbiota in mice fed a high-fat-diet.

Cellulose nanofiber (CN) consumption with exercise could be a potential strategy to control obesity. Here, we studied the effects of CN supplementation and voluntary exercise on obesity and gut microbiota in high-fat diet (HFD)-fed mice. Consumption of CN increased voluntary wheel running activity. CN intake and exercise together suppressed the increase in body weight and fat mass, and improved glucose tolerance. The fecal gut microbiota was analyzed by sequencing 16S ribosomal RNA genes. Principal component analysis revealed a shift in the microbiota composition resulting from exercise, but not from CN supplementation. Erysipelotrichaceae and Rikenellaceae decreased with exercise. Exercise also increased Ruminococcaceae, whereas exercise and CN intake together increased Eubacteriaceae. These two families are butyrate producers. Exercise increased the amount of acetate in the cecum. These results suggest that CN consumption improves exercise performance and exerts anti-obesity effects by modulating the balance of the gut microbiota.

Resistance training prevents muscle fibrosis and atrophy via down-regulation of C1q-induced Wnt signaling in senescent mice.

Increased complement component 1q (C1q) secretion with aging leads to muscle fibrosis and atrophy whereas resistance training attenuates circulating C1q levels. This study aimed to clarify whether resistance exercise-induced reduction of C1q secretion contributes to the inhibition of fibrosis and atrophy in aged muscles. Young (13-wk-old) and aged (38-wk-old) senescence-accelerated mouse prone 1 mice were randomly assigned to one of 4 groups: a young or aged sedentary control group, or a young or aged resistance training (climbing a ladder 3 d/wk for 12 wk) group. We found that resistance training ameliorated muscle fibrosis and atrophy in aged mice, concomitant with decreased circulating and muscle C1q levels and attenuated activation of muscle Wnt signaling (glycogen synthase kinase ß/ß-catenin), including ß-catenin in satellite (Pax7+/DAPI+) and fibroblast (vimentin+/DAPI+) cells. Furthermore, during muscle regeneration after mice were injured by cardiotoxin injection, we observed a reduction in circulating C1q levels, the inhibition of muscle fibrosis and repair, and decreased in the activation of muscle cytoplasmic and nuclear ß-catenin in aged mice from the resistance training group, but these effects were cancelled by a single preadministration of exogenous recombinant C1q. In addition, resistance training attenuated aging-related muscle loss concomitant with decreased expression of both muscle ring-finger protein 1 and muscle atrophy F-box in the muscle. Thus, resistance training-induced changes in circulating C1q levels may contribute to the prevention of muscle fibrosis and atrophy via muscle Wnt signaling in senescent mice.-Horii, N., Uchida, M., Hasegawa, N., Fujie, S., Oyanagi, E., Yano, H., Hashimoto, T., Iemitsu, M. Resistance training prevents muscle fibrosis and atrophy via down-regulation of C1q-induced Wnt signaling in senescent mice.

High sex hormone-binding globulin concentration is a risk factor for high fibrosis-4 index in middle-aged Japanese men.

Low endogenous testosterone and sex hormone-binding globulin (SHBG) concentrations have been reported to be associated with metabolic syndrome (MetS) and non-alcoholic fatty liver disease (NAFLD). However, little is known about the relationships between testosterone or SHBG and liver fibrosis in NAFLD. Thus, we aimed to clarify the relationships between serum testosterone or SHBG concentration and fibrosis-4 (FIB-4) index, a marker of liver fibrosis. Serum testosterone was assayed in various forms (total testosterone [TT], calculated free testosterone [cFT], calculated bioavailable testosterone [cbT], and SHBG) and metabolic markers were also measured in 363 Japanese men (mean age 51.1 ± 8.7 years) at routine health examinations. We then attempted to identify the factors contributing to liver fibrosis by investigating the associations between the metabolic markers, including testosterone, and FIB-4 index. People with a relatively high FIB-4 index (≥1.3) demonstrated lower cFT, cbT, homeostasis model assessment (HOMA)-ß, low-density lipoprotein-cholesterol, and blood urea nitrogen, but higher SHBG, than those with a lower FIB-4 index (<1.3). There were no significant differences in HbA1c, fasting glucose concentration, HOMA-R, or metabolic syndrome prevalence between the two groups. Binary regression analysis revealed that SHBG ≥52 nmol/L and cFT <8.0 ng/dL were statistically significant risk factors for FIB-4 index ≥1.3. Receiver operating characteristic analysis revealed that cFT <7.62 ng/dL (area under the curve [AUC] = 0.639) and SHBG ≥49.8 nmol/L (AUC = 0.649) were the strongest risk factors for FIB-4 index ≥1.3. In contrast to previous findings showing low SHBG concentrations in NAFLD, we provide evidence that high SHBG and low bioactive testosterone are associated with liver fibrosis.

Palmitoleic acid induces the cardiac mitochondrial membrane permeability transition despite the presence of L-carnitine.

Although palmitoleic acid (C16:1) is associated with arrhythmias, and increases in an age-dependent matter, the effects of L-carnitine, which is essential for the transport of long-chain fatty acids into the mitochondria, are unclear. It has been postulated that L-carnitine may attenuate palmitate (C16:0)-induced mitochondrial dysfunction and the apoptosis of cardiomyocytes. The aim of this study was to elucidate the activity of L-carnitine in the prevention of the palmitoleic acid-induced mitochondrial membrane permeability transition and cytochrome c release using isolated cardiac mitochondria from rats. Palmitoleoyl-CoA-induced mitochondrial respiration was not accelerated by L-carnitine treatment, and this respiration was slightly inhibited by oligomycin, which is an inhibitor of ATP synthase. Despite pretreatment with L-carnitine, the mitochondrial membrane potential decreased and mitochondrial swelling was induced by palmitoleoyl-CoA. In the presence of a combination of L-carnitine and tiron, a free radical scavenger, there was attenuated mitochondrial swelling and cytochrome c release following palmitoleoyl-CoA treatment. We concluded that palmitoleic acid, but not palmitate, induces the cardiac mitochondrial membrane permeability transition despite the presence of L-carnitine.

High antitumor activity of pladienolide B and its derivative in gastric cancer.

The antitumor activity of pladienolide B, a novel splicing inhibitor, against gastric cancer is totally unknown and no predictive biomarker of pladienolide B efficacy has been reported. We investigated the antitumor activity of pladienolide B and its derivative on gastric cancer cell lines and primary cultured cancer cells from carcinomatous ascites of gastric cancer patients. The effect of pladienolide B and its derivative on six gastric cancer cell lines was investigated using a MTT assay and the mean IC50 values determined to be 1.6 ± 1.2 (range, 0.6-4.0) and 1.2 ± 1.1 (range, 0.4-3.4) nM, respectively, suggesting strong antitumor activity against gastric cancer. The mean IC50 value of pladienolide B derivative against primary cultured cells from 12 gastric cancer patients was 4.9 ± 4.7 nM, indicative of high antitumor activity. When 18 SCID mice xenografted with primary cultured cells from three patients were administered the pladienolide B derivative intraperitoneally, all tumors completely disappeared within 2 weeks after treatment. Histological examination revealed a pathological complete response for all tumors. In the xenograft tumors after treatment with pladienolide B derivative, immature mRNA were detected and apoptotic cells were observed. When the expressions of cell-cycle proteins p16 and cyclin E in biopsied gastric cancer specimens were examined using immunohisctochemistry, positivities for p16 and cyclin E were significantly and marginally higher, respectively, in the low-IC50 group compared with the high-IC50 group, suggesting the possibility that they might be useful as predictive biomarkers for pladienolide B. In conclusion, pladienolide B was very active against gastric cancer via a mechanism involving splicing impairment and apoptosis induction.

Wet-type grinder-treated okara modulates gut microbiota composition and attenuates obesity in high-fat-fed mice.

Wet-type grinder (WG) is a nanofiber technology used to atomize dietary fiber-rich materials. WG-treated okara (WGO) exhibits high dispersion and viscosity similar to those of viscous soluble dietary fibers. Here, we studied the effect of WGO supplementation on obesity and gut microbiota composition in high-fat diet (HFD)-fed mice. WGO intake suppressed body weight gain and fat accumulation, improved glucose tolerance, lowered cholesterol levels, and prevented HFD-induced decrease in muscle mass. WGO supplementation also led to cecum enlargement, lower pH, and higher butyrate production. The bacterial 16S ribosomal RNA genes (16S rDNA) were sequenced to determine the gut microbiota composition of the fecal samples. Sequencing of bacterial 16S rDNA revealed that WGO treatment increased the abundance of butyrate producer Ruminococcus and reduced the abundances of Rikenellaceae, Streptococcaceae, and Prevotellaceae, which are related to metabolic diseases. Metabolomics analysis of the plasma of WGO- and cellulose-treated mice were conducted using ultra-high-performance liquid chromatography-mass spectrometry. Metabolic pathway analysis revealed that the primary bile acid biosynthesis pathway was significantly positively regulated by WGO intake instead of cellulose. These results demonstrate that WG is useful for improving functional properties of okara to prevent metabolic syndromes, including obesity, diabetes, and dyslipidemia.

Exercise training attenuates adipose tissue fibrosis in diet-induced obese mice.

Tissue fibrosis, such as that which occurs in obesity, is associated with chronic inflammatory diseases. Although regular exercise reduces adipose tissue inflammation, the mechanisms regulating the effects of exercise on adipose tissue fibrosis are unclear. This study aimed to clarify whether exercise training attenuates adipose tissue fibrosis with consequent reduction of extracellular matrix including collagens. Male C57BL/6J (4-week old) mice were randomly assigned to four groups that received a normal diet (ND) plus sedentary (n=8), an ND plus exercise training (n=8), a high-fat diet (HFD) plus sedentary (n=12), and an HFD plus exercise training (n=12). Mice were fed the ND or HFD from 4 to 20 weeks of age. The exercise groups were trained on a motorized treadmill for 60 min/day, 5 times/week over the same period. Histological hepatic fibrosis detected by Sirius red and α-smooth muscle actin staining were attenuated in HFD exercise mice compared with HFD sedentary mice. mRNA levels of transforming growth factor-ß and tissue inhibitors of metalloproteinase-1, major regulators of tissue fibrosis, were increased in HFD sedentary mice but were attenuated in HFD exercise mice. Similarly, adipose tissue from the HFD sedentary mice contained higher macrophages than adipose tissue from the ND mice, and this was also lowered by exercise training. These findings suggest that exercise training may be effective for attenuating adipose tissue inflammation in obesity.

Curcumin attenuates oxidative stress following downhill running-induced muscle damage.

Downhill running causes muscle damage, and induces oxidative stress and inflammatory reaction. Recently, it is shown that curcumin possesses anti-oxidant and anti-inflammatory potentials. Interestingly, curcumin reduces inflammatory cytokine concentrations in skeletal muscle after downhill running of mice. However, it is not known whether curcumin affects oxidative stress after downhill running-induced muscle damage. Therefore, the purpose of this study was to investigate the effects of curcumin on oxidative stress following downhill running induced-muscle damage. We also investigated whether curcumin affects macrophage infiltration via chemokines such as MCP-1 and CXCL14. Male C57BL/6 mice were divided into four groups; rest, rest plus curcumin, downhill running, or downhill running plus curcumin. Downhill running mice ran at 22 m/min, -15% grade on the treadmill for 150 min. Curcumin (3mg) was administered in oral administration immediately after downhill running. Hydrogen peroxide concentration and NADPH-oxidase mRNA expression in the downhill running mice were significantly higher than those in the rest mice, but these variables were significantly attenuated by curcumin administration in downhill running mice. In addition, mRNA expression levels of MCP-1, CXCL14 and F4/80 reflecting presence of macrophages in the downhill running mice were significantly higher than those in the rest mice. However, MCP-1 and F4/80 mRNA expression levels were significantly attenuated by curcumin administration in downhill running mice. Curcumin may attenuate oxidative stress following downhill running-induced muscle damage.

Exercise training attenuates hepatic inflammation, fibrosis and macrophage infiltration during diet induced-obesity in mice.

Nonalcoholic steatohepatitis, which is considered the hepatic event in metabolic syndrome, was recently associated with the innate immune system. Although regular exercise reduces hepatic injury markers like serum alanine aminotransferase (ALT) levels, the mechanisms regulating the effects of exercise on steatohepatitis are unclear. This study aimed to clarify whether exercise training suppresses hepatic injury, inflammation, and fibrosis by suppressing macrophage infiltration. Male C57BL/6J (4-week old) mice were randomly divided into four groups: normal diet (ND) control (n=7), ND exercise (n=5), high-fat diet and high-fructose water (HFF) control (n=11), and HFF exercise (n=11) groups. Mice were fed the ND or HFF from 4 to 20 weeks of age. The exercise groups were trained on a motorized treadmill for 60 min/day, five times/week. The nonalcoholic fatty liver disease (NAFLD) activity score and plasma ALT activity, indicators of liver injury, were increased in HFF control mice but were attenuated in HFF exercise mice. Hepatic inflammation, indicated by hepatic tumor necrosis factor (TNF)-α levels and hepatic resident macrophage infiltration, was significantly lower in HFF exercise mice than in HFF control mice. Hepatic fibrosis markers (histological hepatic fibrosis detected by Sirius red and α-smooth muscle actin staining and tissue inhibitor of matrix metalloproteinase-1 mRNA) were attenuated in HFF exercise mice compared with HFF control mice. These results suggest that exercise training reduces hepatic inflammation, injury, and fibrosis by suppressing macrophage infiltration.

LPS-induced TNF-α production is attenuated by intake with PHGG via gut microbial fermentation in mice.

OBJECTIVES: Intake of dietary fibers promotes the production of short-chain fatty acids (SCFAs), which can affect host inflammation via gut microbial fermentation. Although partially hydrolyzed guar-gum (PHGG) is a water-soluble dietary fiber with lower viscosity, its benefits in acute inflammation are yet to be determined. The aim of this study was to investigate the effect of PHGG intake on the lipopolysaccharide (LPS)-induced tumor necrosis factor (TNF)-α production. METHODS: Nine-wk-old male C3 H/HeN mice were used in this study, and they were randomly divided into control diet (CD) and CD + 5% PHGG (GGCD) groups. After a dietary intervention of 6 wk, LPS (1 mg/kg) was injected into the orbital vein. Plasma TNF-α concentration and SCFAs in cecum contents were then measured. Also, the effect of gut microbiota on LPS-induced TNF-α production was evaluated in PHGG-fed mice before and after antibiotic treatment. RESULTS: PHGG intake accelerated a dramatic suppression of LPS-induced TNF-α production (P < 0.01). PHGG-induced low pH in feces (P < 0.05) indicates that the gut microbiota induced high fermentation. Indeed, SCFAs in cecum contents of GGCD mice were significantly higher than in the CD group (P < 0.05). Furthermore, PHGG intake after antibiotic treatment did not induce the suppression of TNF-α. CONCLUSION: These results demonstrated that inflammation was inhibited by habitual PHGG ingestion, suggesting that this phenomenon might be associated with changes in gut microbiota-induced SCFAs production.

Protective action of L-carnitine on cardiac mitochondrial function and structure against fatty acid stress.

Cardiovascular risks are frequently accompanied by high serum fatty acid levels. Although recent studies have shown that fatty acids affect mitochondrial function and induce cell apoptosis, L-carnitine is essential for the uptake of fatty acids by mitochondria, and may attenuate the mitochondrial dysfunction and apoptosis of cardiocytes. This study aimed to elucidate the activity of L-carnitine in the prevention on fatty acid-induced mitochondrial membrane permeability transition and cytochrome c release using isolated cardiac mitochondria from rats. Palmitoyl-CoA-induced mitochondrial respiration that was observed with L-carnitine was inhibited with oligomycin. The palmitoyl-CoA-induced mitochondrial membrane depolarization and swelling were greatly inhibited by the presence of L-carnitine. In ultrastructural observations, terminally swollen and ruptured mitochondria with little or no distinguishable cristae structures were induced by treatment with palmitoyl-CoA. However, the severe morphological damage in cardiac mitochondria was dramatically inhibited by pretreatment with L-carnitine. Treatment with L-carnitine also attenuated 4-hydroxy-L-phenylglycine- and rotenone-induced mitochondrial swelling even when the L-carnitine could not protect against the decrease in oxygen consumption associated with these inhibitors. Furthermore, L-carnitine completely inhibited palmitoyl-CoA-induced cytochrome c release. We concluded that L-carnitine is essential for cardiac mitochondria to attenuate the membrane permeability transition, and to maintain the ultrastructure and membrane stabilization, in the presence of high fatty acid ß-oxidation. Consequently, the cells may be protected against apoptosis by L-carnitine through inhibition of the fatty acid-induced cytochrome c release.

Exercise-independent wheat-induced anaphylaxis caused by ω-5 gliadin in mice.

BACKGROUND: ω-5 Gliadin is known as a major allergen in wheat-dependent exercise-induced anaphylaxis. The characteristics that make ω-5 gliadin an allergen remain unclear. METHODS: Mice were sensitized by means of intraperitoneal injection of gliadin fractions together with the adjuvant alum. Anaphylactic responses were assessed by measuring body temperature and voluntary physical activity. Specific IgE levels in mouse serum were evaluated by ELISA. After oral administration of proteins to mice, concentrations of administered proteins in their portal blood were also analyzed by competitive inhibition ELISA. RESULTS: Oral administration of ω-5 gliadin evoked significant decreases in body temperature and physical activity of sensitized mice, whereas the gliadin fraction did not induce these effects at the same dose. These responses were exercise independent. ELISA analysis revealed that IgE antibodies from sensitized mice react to ω-5 gliadin with great efficacy. After oral administration of either the gliadin fraction or ω-5 gliadin, blood levels of ω-5 gliadin were much higher than those of the gliadin fraction. CONCLUSIONS: ω-5 Gliadin caused anaphylaxis in sensitized mice, whereas the gliadin fraction did not at the same dose. The anaphylactic response was exercise independent. It is likely that IgE of sensitized mice reacts strongly to ω-5 gliadin and that ω-5 gliadin is better absorbed from the gastrointestinal tract than other components of the gliadin fraction. These results indicated that ω-5 gliadin has prominent characteristics as an allergen and that exercise might be an indirect factor in anaphylaxis induction.

Impact of ω-5 gliadin on wheat-dependent exercise-induced anaphylaxis in mice.

The effects of ω-5 gliadin on wheat-dependent exercise-induced anaphylaxis (WDEIA) were investigated by using a mouse model. The gliadin fraction was prepared as a 70% ethanol-soluble solution, and ω-5 gliadin was purified by chromatography. Purified ω-5 gliadin was run on SDS-PAGE gel to reveal three bands with a molecular mass of 53-60 kDa and had the characteristic N-terminal sequence of ω-5 gliadin. The mice were sensitized to the gliadin fraction, and the anaphylactic response was assessed by measuring the body temperature and voluntary physical activity. An oral administration of ω-5 gliadin evoked a significant drop in both the body temperature and voluntary physical activity, similar to the effects of the whole gliadin fraction. ELISA and immunoblotting analyses revealed that the IgE expression from sensitized mice reacted most strongly to ω-5 gliadin. Taken together, these results indicate ω-5 gliadin to be a major allergen responsible for stimulating WDEIA in mice, with the characteristic potential for stimulating IgE production.

L-carnitine is essential to beta-oxidation of quarried fatty acid from mitochondrial membrane by PLA(2).

Mitochondrial beta-oxidation is an important system involved in the energy production of various cells. In this system, the function of L-carnitine is essential for the uptake of fatty acids to mitochondria. However, it is unclear whether or not endogenous respiration, ADP-induced O(2) consumption without substrates, is caused by L-carnitine treatment. In this study, we investigated whether L-carnitine is essential to the beta-oxidation of quarried fatty acids from the mitochondrial membrane by phospholipase A(2) (PLA(2)) using isolated mitochondria from the liver of rats. Intact mitochondria were incubated in a medium containing Pi, CoA and L-carnitine. The effect of L-carnitine treatment on ADP-induced mitochondrial respiration was observed without exogenous respiratory substrate. Increase in mitochondrial respiration was induced by treatment with L-carnitine in a concentration-dependent manner. Treatment with rotenone, a complex I blocker, completely inhibited ADP-induced oxygen consumption even in the presence of L-carnitine. Moreover, the L-carnitine dependent ADP-induced mitochondrial oxygen consumption did not increase when PLA(2) inhibitors were treated before ADP treatment. The L-carnitine-dependent ADP-induced oxygen consumption did contribute to ATP productions but not heat generation via an uncoupling system. These results suggest that L-carnitine might be essential to the beta-oxidation of quarried fatty acids from the mitochondrial membrane by PLA(2).