The enhancer HS2 critically regulates GATA-3-mediated Il4 transcription in T(H)2 cells.

GATA-3 is a master regulator of T helper type 2 (T(H)2) differentiation. However, the molecular basis of GATA-3-mediated T(H)2 lineage commitment is poorly understood. Here we identify the DNase I-hypersensitive site 2 (HS2) element located in the second intron of the interleukin 4 locus (Il4) as a critical enhancer strictly controlled by GATA-3 binding. Mice lacking HS2 showed substantial impairment in their asthmatic responses and their production of IL-4 but not of other T(H)2 cytokines. Overexpression of Gata3 in HS2-deficient T cells failed to restore Il4 expression. HS2 deletion impaired the trimethylation of histone H3 at Lys4 and acetylation of histone H3 at Lys9 and Lys14 in the Il4 locus. Our results indicate that HS2 is the target of GATA-3 in regulating chromosomal modification of the Il4 locus and is independent of the Il5 and Il13 loci.

Mina, an Il4 repressor, controls T helper type 2 bias.

T helper type 2 (T(H)2) bias, which is the propensity of naive CD4(+) T cells to differentiate into interleukin 4 (IL-4)-secreting T(H)2 cells, is a genetic trait that affects susceptibility to infectious, autoimmune and allergic diseases. T(H)2 bias correlates with the amount of IL-4 initially secreted by newly activated helper T cells that feeds back positively through the pathway of the IL-4 receptor and the transcription factors STAT6 and GATA-3 to drive T(H)2 development. Here we identify Mina, a member of the jumonji C (JmjC) protein family, as a genetic determinant of T(H)2 bias. Mina specifically bound to and repressed the Il4 promoter. Mina overexpression in transgenic mice impaired Il4 expression, whereas its knockdown in primary CD4(+) T cells led to Il4 derepression. Our findings collectively provide mechanistic insight into an Il4-regulatory pathway that controls helper T cell differentiation and genetic variation in T(H)2 bias.

Role of FK506-sensitive signals in asthmatic lung inflammation.

Asthma is airway inflammatory diseases caused by the activation of group 2 innate lymphoid cells (ILC2s) and type 2 helper T (TH2) cells. Cysteine proteases allergen cause tissue damage to airway epithelial cells and activate ILC2-mediated type 2 airway inflammation. FK506 is an immunosuppressive agent against calcium-dependent NFAT activation that is also effective against asthmatic inflammation. However, the effects of FK506 on cysteine protease allergen-mediated airway inflammation remain unclear. In this study, we investigated the suppressive effects of FK506 on airway inflammation. FK506 had a partial inhibitory effect on ILC2-dependent eosinophil inflammation and a robust inhibitory effect on T cell-dependent eosinophil inflammation in a cysteine protease-induced mouse asthma model. The infiltration of T1/ST2+ CD4 T cells in the lungs contributed to the persistence of eosinophil infiltration in the airway; FK506 completely inhibited the infiltration of T1/ST2+ CD4 T cells. In the initial phase, FK506 treatment targeted lung ILC2 activation induced by leukotriene B4 (LTB4)-mediated calcium signaling, but not IL-33 signaling. FK506 also inhibited the IL-13-dependent accumulation of T1/ST2+ CD4 T cells in the lungs of the later responses. These results indicated that FK506 potently suppressed airway inflammation by targeting ILC2 activation and T1/ST2+ CD4 T cell accumulation.

Strategies for Effective Home Modification in Older Adults.

There are various barriers to home modifications to prevent falls among the older population. Several strategies may be necessary to overcome these barriers and implement effective home modifications. The need for home modification should be assessed, which requires a home evaluation by a specialist. In Japan, welfare housing environment coordinators have been trained to provide advice on home modifications suitable for people with disabilities. In addition, in Japan, home assessment and advice on home modification before discharge from acute care hospitals for older people is allowed as a medical reimbursement, and a system for effective home modification is well established. Human resource training and medical policy arrangements on home modifications could improve the cost-effectiveness. In Japan, a system has been established to support the costs of home modification and environmental maintenance. Financial support has reduced the barrier to home modification. Fixed grab bars or shower chairs can be rented, which may be more cost-effective than purchasing them and may shorten the time required for installation. There may be psychological barriers to home modification for older population. Since many older people do not recognize the importance of home modification, promotion to convey the value of home modification may be necessary. Training of staff to engage in home modification, public financial support for modification, and ideas for reducing psychological hesitation may help to reduce the barriers for home modification and to enable effective home modification.

Essential Role of STAT3 Signaling in Hair Follicle Homeostasis.

Dominant-negative mutations associated with signal transducer and activator of transcription 3 (STAT3) signaling, which controls epithelial proliferation in various tissues, lead to atopic dermatitis in hyper IgE syndrome. This dermatitis is thought to be attributed to defects in STAT3 signaling in type 17 helper T cell　specification. However, the role of STAT3 signaling in skin epithelial cells remains unclear. We found that STAT3 signaling in keratinocytes is required to maintain skin homeostasis by negatively controlling the expression of hair follicle-specific keratin genes. These expression patterns correlated with the onset of dermatitis, which was observed in specific pathogen-free conditions but not in germ-free conditions, suggesting the involvement of Toll-like receptor-mediated inflammatory responses. Thus, our study suggests that STAT3-dependent gene expression in keratinocytes plays a critical role in maintaining the homeostasis of skin, which is constantly exposed to microorganisms.

[Case of bladder perforation due to the obturator nerve reflex during transurethral resection (TUR) of bladder tumor using the TUR in saline (Turis) system under spinal anesthesia].

Bladder perforation due to the obturator nerve reflex (ONR) is a serious complication during TUR of bladder tumor using the conventional TUR system; requiring monopolar electrocautery and non-conductive solution as perfusate. Recently, the TURis system, which employs bipolar electrocautery and physiological saline as perfusate, has been developed. Electrical resistance of physiological saline and human tissues are approximately 40 and 500 omega, respectively. Thus, theoretically, electrical current flows between the resection loop and the recovery electrode integrated in the outer sleeve of the endoscope, without forming electrical circuit in the patient's body; suggesting possible elimination of the ONR. Here we describe a case of bladder perforation during surgery using the TURis system; the ONR was exaggerated during the procedure to stop bleeding at the lateral wall using bipolar electrocautery. In addition to this case, there have been a few reports of the ONR during surgery using the TURis system, and it is reported that weak electrical current may pass through the patient's body in the TURis system. We consider that evaluation of the necessary precautions, such as the obturator nerve block, for the prevention of the ONR is important even in the surgery using the TURis system.

A randomized study assessing the impact of cilostazol on platelet function profiles in patients with diabetes mellitus and coronary artery disease on dual antiplatelet therapy: results of the OPTIMUS-2 study.

AIMS: Patients with type 2 diabetes mellitus (T2DM) have reduced platelet inhibition compared with non-diabetics following P2Y(12) receptor blockade. Whether inhibition of P2Y(12) signalling can be enhanced by adjunctive treatment with cilostazol in T2DM patients is unknown. The aim of this pilot study was to assess the functional impact of cilostazol in T2DM patients on standard aspirin and clopidogrel treatment. METHODS AND RESULTS: This was a prospective, double-blind, double-dummy, placebo-controlled, randomized, cross-over platelet function study. T2DM patients on dual antiplatelet therapy were assigned to receive cilostazol 100 mg or placebo twice daily for 14 days and afterwards crossed-over treatment assignments for another 14 days. Platelet function was performed at three time points: at baseline, 14 days after randomization, and 14 days after treatment cross-over. The P2Y(12) reactivity index, determined through flow cytometric assessment of the phosphorylation status of the vasodilator-stimulated phosphoprotein, was the primary endpoint measure. In addition to this flow cytometric evaluation, light transmittance aggregometry and VerifyNow testing were performed. A total of 25 T2DM patients were randomized; five patients discontinued treatment due to side effects. The P2Y(12) reactivity index was significantly lower following cilostazol treatment compared with placebo (36.3 +/- 20 vs. 59.9 +/- 16%; P = 0.0002). All other P2Y(12)-specific functional assessments showed enhanced inhibition of this signalling pathway following treatment with cilostazol. CONCLUSION: Adjunctive treatment with cilostazol in T2DM patients on standard dual antiplatelet therapy enhances inhibition of platelet P2Y(12) signalling.

Functional effects of high clopidogrel maintenance dosing in patients with inadequate platelet inhibition on standard dose treatment.

Updated guidelines on percutaneous coronary intervention recommend increasing the dose of clopidogrel to 150 mg in high-risk patients if <50% platelet inhibition is demonstrated. However, to date, the functional impact of this recommendation has been poorly explored. The aim of this study was to assess the functional implications associated with the use of clopidogrel 150 mg/day in patients with inadequate platelet inhibition while receiving standard 75 mg/day maintenance treatment. Patients with diabetes mellitus have a higher prevalence of inadequate clopidogrel-induced antiplatelet effects and stent thrombosis compared with those without diabetes and were selected for this analysis. Platelet inhibition was assessed using the VerifyNow P2Y12 assay in patients with type 2 diabetes receiving dual-antiplatelet therapy. Patients (n = 17) with <50% platelet inhibition were treated with clopidogrel 150 mg/day for 1 month. Adenosine diphosphate-induced aggregation and the P2Y12 reactivity ratio were also assessed. Platelet function profiles were compared with that of a control group (n = 17) with >or=50% inhibition. Platelet inhibition increased from 27.1 +/- 12% to 40.6 +/- 18% in patients treated with clopidogrel 150 mg/day (p = 0.009; primary end point). All other functional measures also showed enhanced clopidogrel-induced antiplatelet effects. The degree of platelet inhibition achieved after treatment with clopidogrel 150 mg/day varied broadly, and only 35% of patients yielded a degree of platelet inhibition >or=50%. Increasing the dose in patients with inadequate response to clopidogrel did not reach the same degree of antiplatelet effects as those achieved in patients with adequate response while receiving 75 mg/day. In conclusion, the use of a 150 mg maintenance dose of clopidogrel in patients with type 2 diabetes with <50% platelet inhibition is associated with enhanced antiplatelet effects. However, the antiplatelet effects achieved are nonuniform, and a considerable number of patients persist with inadequate platelet inhibition.

Medium-chain Triglycerides with Maltodextrin Increase Fat Oxidation during Moderate-intensity Exercise and Extend the Duration of Subsequent High-intensity Exercise.

Medium-chain triglycerides (MCT) are useful for increasing fat utilization during exercise. The highest rate of fat oxidation during submaximal exercise tends to precede the lactate threshold in untrained adults. In our previous study, blood lactate concentration was more than 4 mmol/L (onset of blood lactate) in recreational athletes during exercise at a workload corresponding to 60% peak O2 uptake (Vï½¥o2), which was below ventilation threshold. In the present study, we investigated the effect of 2 week of ingestion of food containing 6 g MCT on substrate oxidation during moderate-intensity (50% peak Vï½¥o2) exercise and high-intensity (70% peak Vï½¥o2) exercise in recreational athletes. For comparison, two experimental trials were conducted after participants had been administered isoenergic test foods (MCT-supplemented food with mainly maltodextrin-containing carbohydrate (MCT + CHO) or CHO) for 2 weeks, with a washout period between trials. Participants were instructed to perform cycle ergometer exercise at a workload corresponding to 50% peak Vï½¥o2 for 40 min followed by a workload corresponding to 70% peak Vï½¥o2 until exhaustion. Fat oxidation was significantly increased in the MCT + CHO trial (13.3 ± 2.7 g/40 min, mean ± SD, p < 0.05) during moderate-intensity exercise and the duration was extended significantly (23.5 ± 19.4 min, p < 0.05) during subsequent high-intensity exercise, compared with that observed in the CHO trial (fat oxidation; 11.7 ± 2.8 g/40 min, duration; 17.6 ± 16.1 min). In conclusion, continuous ingestion of 6 g MCT with maltodextrin could increase fat oxidation during moderate-intensity exercise and extend the duration of subsequent high-intensity exercise in recreational athletes, compared with the ingestion of isoenergic maltodextrin alone.

Effects of margarine containing medium-chain triacylglycerols on body fat reduction in humans.

We found previously that the ingestion of margarine containing medium-chain triacylglycerols (MCT) resulted in a significant increase in postprandial thermogenesis when compared with long-chain triacylglycerols (LCT). Diets that included margarine containing MCT and LCT were compared for 12 weeks in 73 subjects to investigate the effects on body weight, body fat, areas of subcutaneous and visceral fat, serum total cholesterols, triglycerides, lipoproteins, plasma glucose, serum insulin, total ketone bodies, and the activities of aspartate aminotransferase, alanine aminotransferase, and gamma-glutamyltranspeptidase. We conducted a double-blind, controlled study and used blended rapeseed oil and soybean oil (LCT) as a comparison. Two groups ingested 2100-2400 kcal/day of energy, 65-73 g/day of total fat, and 14 g/day of test margarine (5 g/day of MCT or LCT). The subjects on the MCT diet demonstrated significant decreases in body fat weight (- 3.8 +/- 2.4 kg vs - 2.4 +/- 1.7 kg; MCT vs LCT, mean +/- SD), subcutaneous fat (- 38.2 +/- 29.9 cm(2) vs - 22.6 +/- 19.3 cm(2)), and visceral fat (- 12.2 +/- 11.2 cm(2) vs - 1.6 +/- 12.8 cm(2)) after 12 weeks. There were no clinical differences in measured blood parameters. We suggest that the postprandial increase in thermogenesis and control of postprandial triglyceride levels may explain these results.

Comparison of diet-induced thermogenesis of foods containing medium- versus long-chain triacylglycerols.

The purpose of this study was to investigate the effect of 5-10 g of medium-chain triacylglycerols (MCT) on diet-induced thermogenesis in healthy humans. The study compared diet-induced thermogenesis after ingestion of test foods containing MCT and long-chain triacylglycerols (LCT), using a double-blind, crossover design. Eight male and eight female subjects participated in study 1 and study 2, respectively. In both studies, the LCT was a blend of rapeseed oil and soybean oil. In study 1, the liquid meals contained 10 g MCT (10M), a mixture of 5 g MCT and 5 g LCT (5M5L), and 10 g LCT (10L). In study 2, the subjects were given a meal (sandwich and clear soup) with the mayonnaise or margarine containing 5 g of MCT or LCT. Postprandial energy expenditure was measured by indirect calorimetry before and during the 6 h after ingestion of the test meals. Diet-induced thermogenesis was significantly greater after 5M5L and 10M Ingestion as compared to 10L ingestion. Ingestion of the mayonnaise or margarine containing 5 g MCT caused significantly larger diet-induced thermogenesis as compared to that of LCT. These results suggest that, in healthy humans, the intake of 5-10 g of MCT causes larger diet-induced thermogenesis than that of LCT, irrespective of the form of meal containing the MCT.

Dysregulation of suppressor of cytokine signaling 3 in keratinocytes causes skin inflammation mediated by interleukin-20 receptor-related cytokines.

Homeostatic regulation of epidermal keratinocytes is controlled by the local cytokine milieu. However, a role for suppressor of cytokine signaling (SOCS), a negative feedback regulator of cytokine networks, in skin homeostasis remains unclear. Keratinocyte specific deletion of Socs3 (Socs3 cKO) caused severe skin inflammation with hyper-production of IgE, epidermal hyperplasia, and S100A8/9 expression, although Socs1 deletion caused no inflammation. The inflamed skin showed constitutive STAT3 activation and up-regulation of IL-6 and IL-20 receptor (IL-20R) related cytokines, IL-19, IL-20 and IL-24. Disease development was rescued by deletion of the Il6 gene, but not by the deletion of Il23, Il4r, or Rag1 genes. The expression of IL-6 in Socs3 cKO keratinocytes increased expression of IL-20R-related cytokines that further facilitated STAT3 hyperactivation, epidermal hyperplasia and neutrophilia. These results demonstrate that skin homeostasis is strictly regulated by the IL-6-STAT3-SOCS3 axis. Moreover, the SOCS3-mediated negative feedback loop in keratinocytes has a critical mechanistic role in the prevention of skin inflammation caused by hyperactivation of STAT3.

Comparative quantitative analysis of BCR-ABL transcripts with the T315I mutant clone by polymerase chain reaction (PCR)-Invader method.

Drug resistance is a serious complication in the treatment of chronic myeloid leukemia (CML). The most common and best-characterized mechanism of secondary imatinib resistance in CML is the development of kinase domain mutations in the BCR-ABL gene. Second-generation tyrosine kinase inhibitors, such as dasatinib or nilotinib, overcome most of these mutations, but they are not effective against the T315I mutant. To determine whether these mutations contribute to clinical resistance, it is necessary to monitor the ratio of the mutant and wild-type forms. Here, we developed a polymerase chain reaction (PCR)-Invader assay for comparative quantitative analysis (qPI assay) of BCR-ABL transcripts with the T315I mutant clone. T315I ratios were calculated for the wild-type and mutant fold-over-zero (FOZ) values. In examination with 2 kinds of plasmids containing wild-type or T315I mutant PCR amplicons, mutant FOZ values were detected down to 1% of the total. The results of 12 serial samples from 2 patients (case A: Philadelphia-positive acute lymphoblastic leukemia and case B: CML) with the T315I mutant clone were compared with those of direct sequencing or 2 kinds of allele-specific oligonucleotide (ASO)-PCR. All samples showed the T315I mutation by qPI assay and ASO-PCR, and 10 samples showed it by direct sequencing. Significant correlation (correlation coefficient; r2 = 0.951) was noted between the qPI assay and quantitative ASO-PCR to analyze T315I mutant ratios. Thus, the qPI assay is a useful method for evaluating the T315I mutant clone in BCR-ABL transcripts.

Effect of ingestion of medium-chain triacylglycerols on moderate- and high-intensity exercise in recreational athletes.

Medium-chain triacylglycerols (MCT) are known to hydrolyze readily and completely to fatty acids and to be metabolized more easily by beta-oxidation than long-chain triacylglycerols (LCT). Therefore, we investigated the effect of 2 wk of ingestion of food containing a small amount (6 g) of MCT on energy metabolism during moderate-intensity exercise and high-intensity exercise in recreational athletes. For comparison, the subjects were administered food containing MCT or LCT for 14 d, and were instructed to perform cycle ergometer exercise at a workload corresponding to 60% peak O(2) uptake (VO(2)) for 40 min followed by a workload corresponding to 80% peak VO(2) until exhaustion. Blood lactate concentration, VO(2), VCO(2), and rating of perceived exertion (RPE) were measured at rest and during exercise. The exercise time to exhaustion at a workload corresponding to 80% peak VO(2) was significantly (p<0.05) longer in the MCT trial (10.2+/-7.6 min; mean+/-SD) than in the LCT trial (5.8+/-3.3 min). Blood lactate concentration and RPE during exercise were significantly (p<0.05) lower after ingestion of MCT-containing food. Fat oxidation rate was higher and carbohydrate oxidation rate was lower during exercise in the MCT trial than in the LCT trial, but the differences were not significant. These results indicate that the ingestion of MCT-containing food may suppress utilization of carbohydrate for energy production because of increased utilization of fatty acids for generating energy. In conclusion, our data suggest that short-term ingestion of food containing a small amount of MCT suppresses the increase in blood lactate concentration and RPE during moderate-intensity exercise and extends the duration of subsequent high-intensity exercise, at levels higher than those achieved by ingestion of LCT-containing food.

Effect of dietary medium- and long-chain triacylglycerols (MLCT) on accumulation of body fat in healthy humans.

We investigated whether a structured medium- and long-chain triacylglycerols (MLCT) diet could decrease accumulation of body fat in healthy humans. The study was conducted under a double-blind randomized design. Ninety-three subjects participated in this study. However, 10 subjects could not consume the specified meal, and one subject wished to opt out. Consequently, the study included 82 subjects. The experimental subjects consumed the test bread, which was made with 14 g of MLCT containing 1.7 g MCFA, daily at breakfast during the study period of 12 weeks, and the control subjects consumed bread made with long-chain triacylglycerols (LCT). All subjects consumed the same standard packaged meals. Body composition parameters were body weight, total body fat and abdominal fat, and blood analyses included serum cholesterol, triacylglycerols and phospholipids. Significant decreases of body weight, the amount of body fat, subcutaneous and visceral fat were noted in the MLCT group as compared with those of the LCT group for 12 weeks (P<0.05). Furthermore, a significant decrease in serum total cholesterol was noted in the MLCT group as compared with that of the LCT group at 8 weeks (P<0.05). However, other serum parameters were not different between the MLCT and LCT groups. The results suggest that the daily intake of MLCT diet could result in a reduction in body weight and in accumulation of body fat, and, moreover, it could reduce serum total cholesterol.