B-1a Cells Scavenge NETs to Attenuate Sepsis.

B-1a cells, a regulatory subset of B lymphocytes, produce natural IgM and IL-10. Neutrophil extracellular traps (NETs) play a crucial role in pathogen defense, but their excessive formation during sepsis can cause further inflammation and tissue damage. In sepsis, extracellular cold-inducible RNA-binding protein (eCIRP), a damage-associated molecular pattern, is released to induce NET formation. We hypothesize that B-1a cells clear NETs to prevent sepsis-induced injury. Sepsis in mice was induced by injecting 1 × 107 and 5 × 107 CFU E. coli intraperitoneally (i.p.). After 4 and 20 hours, we assessed the number of B-1a cells in the peritoneal cavity using flow cytometry. Our results showed that the number of peritoneal B-1a cells was significantly decreased in E. coli-sepsis mice. Importantly, replenishing B-1a cells via i.p. injection in sepsis mice significantly decreased NETs in peritoneal neutrophils. We also observed a decrease in serum inflammation and injury markers and a significant increase in overall survival rate in B-1a cell-treated septic mice. To understand the mechanism, we co-cultured bone marrow-derived neutrophils (BMDNs) with peritoneal B-1a cells in a contact or non-contact condition using an insert and stimulated them with eCIRP. After 4 hours, we found that eCIRP significantly increased NET formation in BMDNs. Interestingly, we observed that B-1a cells inhibited NETs by 67% in a contact-dependent manner. Surprisingly, when B-1a cells were cultured in inserts, there was no significant decrease in NET formation, suggesting that direct cell-to-cell contact is crucial for this inhibitory effect. We further determined that B-1a cells promoted NET phagocytosis and this was mediated through natural IgM, as blocking IgM receptor attenuated the engulfment of NETs by B-1a cells. Finally, we identified that following their engulfment, NETs were localized into the lysosomal compartment for lysis. Thus, our study suggests that B-1a cells decrease NET content in eCIRP-treated neutrophils and E. coli-sepsis mice.

Extracellular CIRP Induces Novel Nectin-2+ (CD112+) Neutrophils to Promote Th1 Differentiation in Sepsis.

Neutrophil heterogeneity represents different subtypes, states, phenotypes, and functionality of neutrophils implicated in sepsis pathobiology. Extracellular cold-inducible RNA-binding protein (eCIRP) is a damage-associated molecular pattern that promotes inflammation and alters neutrophil phenotype and function through TLR4. Nectin-2 or CD112 is an Ig-like superfamily member. CD112 serves as the ligand for DNAM-1 (CD226), which induces Th1 differentiation in naive CD4+ T cells. Th1 cells produce IFN-γ to fuel inflammation. CD112 is expressed mainly on APCs, but its expression in neutrophils is unknown. We hypothesize that eCIRP induces CD112 expression in neutrophils, promoting Th1 differentiation in sepsis. Incubation of neutrophils with recombinant murine (rm)CIRP significantly increased the gene and protein expression of CD112 in neutrophils. Anti-TLR4 Ab-treated neutrophils significantly decreased CD112+ neutrophils compared with controls upon rmCIRP stimulation. After 4 h of rmCIRP injection in mice, CD112+ neutrophils were significantly increased in the blood and spleen. At 20 h after cecal ligation and puncture-induced sepsis, CD112+ neutrophils were also significantly increased. Blood and splenic CD112+ neutrophils in septic CIRP-/- mice were much lower than in septic wild-type mice. Coculture of naive CD4 T cells with rmCIRP-treated (CD112+) neutrophils significantly increased IFN-γ-producing Th1 cells compared with coculture with PBS-treated neutrophils. CD112 Ab significantly attenuated Th1 differentiation induced by rmCIRP-treated neutrophils. Thus, eCIRP increases CD112 expression in neutrophils via TLR4 to promote Th1 differentiation in sepsis. Targeting eCIRP may attenuate sepsis by reducing Th1-promoting CD112+ neutrophils.

Force-velocity relationship profile of elbow flexors in male gymnasts.

BACKGROUND: The theoretical maximum force (F0), velocity (V0), and power (Pmax) of athletes calculated from the relationship between force and velocity (F-V relationship) and the slope of the F-V relationship, reflect their competitive and training activity profiles. Evaluating the F-V relationship of athletes facilitates categorizing the profiles of dynamic muscle functions in relation to long-term sport-specific training. For gymnastics, however, no studies have tried to examine the profiles of F-V relation and power output for upper limb muscles in relation to the muscularity, while the use of the upper extremities in this sport is very unique as described earlier. PURPOSE: It was hypothesized that the F-V relationship of the elbow flexion in gymnasts might be characterized by low capacity for generating explosive force, notably in terms of the force normalized to muscle size. METHODS: The F0, V0, and Pmax derived from the force-velocity relationship during explosive elbow flexion against six different loads (unloaded condition, 15, 30, 45, 60, and 75% of maximal voluntary isometric elbow flexion force (MVFEF)) for 16 gymnasts (GYM) and 22 judo athletes (JD). F0 and Pmax were expressed as values relative to the cross-sectional area index (CSAindex) of elbow flexors (F0/CSAindex and Pmax/CSAindex, respectively), which was calculated from muscle thickness in the anterior upper arm. The electromyogram (EMG) activities of the biceps brachii (BB) during the maximal isometric and dynamic tasks were also determined. RESULTS: There were no significant differences in CSAindex of elbow flexors between GYM and JD. MVFEF/CSAindex for GYM was significantly lower than that for JD. Force was linearly associated with velocity in the dynamic elbow flexion for all the participants (r =  - 0.997 to -0.905 for GYM, r =  - 0.998 to -0.840 for JD). F0, F0/ CSAindex, V0, Pmax, Pmax/CSAindex, and MVFEF were significantly lower in GYM than in JD. The activity levels of BB during the dynamic tasks tended to be lower in GYM than in JD at load of <45%MVC. CONCLUSION: Gymnasts cannot generate explosive elbow flexion force corresponding to their muscle size. This may be due to low neuromuscular activities during the maximal dynamic tasks against relatively low loads.

Profile of regional fat and fat-free soft tissue accumulation in male athletes.

BACKGROUND: It is unclear whether or not the breakpoint (BP), at which the proportion of each of fat mass (FM) and fat-free soft tissue mass (FFSTM) to body mass (BM) alter, exists in male athletes. We examined the hypothesis that in male athletes, the regional FM and FFSTM-BM relationships have a BP, but the body mass at BP (BMBP) differs among the arms, trunk, and legs. METHODS: By using a dual X-ray absorptiometry, whole-body and regional FMs and FFSTMs in the arms, trunk, and legs were estimated in 198 male athletes (20.8 ± 2.1 years; 1.73 ± 0.07 m; 72.7 ± 14.8 kg). To detect the BP in the relationship between each of FM and FFSTM and BM, a piecewise linear regression analysis was used. If a BP was detected in the corresponding relationship, the significant difference between the regression slopes above and below the BP was examined. RESULTS: The regression analysis indicated that the BMBP existed in the FM- and FFSTM-BM relationships regardless of region and whole body. For the whole body, BMBP was 81.8 kg for FM and 82.2 kg for FFSTM. In regional FM-BM relationships, BMBP was 80.5 kg for arms, 82.6 kg for trunk, and 63.3 kg for legs, and the regression slopes above the BMBP became higher than those below the BP, and vice versa in regional FFSTM-BM relationships (BMBP 104.6 kg for arms, 80.9 kg for trunk, and 79.0 kg for legs). The relative differences in the slopes between below and above BMBP in the regional FM-BM relationships were higher in the arms and trunk than in the legs, and those in the regional FFSTM-BM relationships in the legs than in the trunk. CONCLUSION: Whole-body and regional FM- and FFSTM-BM relationships for male athletes have breakpoints at which the proportion of the tissue masses to BM alters. The BMBP and differences in the distribution of regional FM and FFSTM around the breakpoint are region specific.

Genomic analysis of antibiotic resistance for Acinetobacter baumannii in a critical care center.

AIM: Acinetobacter baumannii is commonly associated with outbreaks and antibiotic-resistant nosocomial infection. This study aimed to determine the relationship between antibiotic resistance and genotypes of A. baumannii. METHODS: A study was undertaken in the critical care center (CCC) of Juntendo University Urayasu Hospital (Urayasu, Japan) between January 2012 and September 2015. Antimicrobial susceptibility tests were carried out according to the Clinical and Laboratory Standards Institute guidelines. All A. baumannii isolates were verified to carry carbapenemase genes and the ISA ba1 element using polymerase chain reaction. The genetic relationship of all A. baumannii isolates was determined by pulsed-field gel electrophoresis and multilocus sequence typing. RESULTS: During the study period, 1634 patients were admitted to the CCC. Acinetobacter baumannii was detected in 43 patients (average age, 58 ± 19 years; 67.4% men). Six patients were determined to be extensively drug-resistant A. baumannii and 21 patients determined to be multidrug-resistant A. baumannii. Antimicrobial susceptibility linked genotypes of A. baumannii. Molecular characterization by pulsed-field gel electrophoresis and multilocus sequence typing showed that closely related clones of A. baumannii had spread in the CCC. CONCLUSION: Resistance to antimicrobial drugs was significantly associated with certain A. baumannii genotypic types and molecular types. Thus, we might be able to predict whether the genotype has spread in the CCC or not when the susceptibility is examined, facilitating the appropriate isolation of patients.

Characteristics of indoor injuries in hotels compared to home among young children.

BACKGROUND: Although attention has recently been afforded to home injury prevention for young children, we often encounter young children who have experienced indoor injuries at places other than the home. We aimed to identify characteristics of unintentional indoor injuries that occurred when young children were not at home. METHODS: We retrospectively reviewed the medical records of young children (aged <6 years) with indoor injuries from January to June 2017. We classified patients injured in hotel rooms as the "Hotel group" and compared them to patients injured at home ("Home group"). RESULTS: Among 102 patients who met the study criteria, 33 patients (32.4%) were classified as the Hotel group. Falls were the most frequent cause of injury in both groups (Home, 56.5% versus Hotel, 87.9%). Falls from beds were more likely to happen in hotel rooms (1.4% versus 48.5%). In regard to fall-associated injuries, head and / or facial injury was most frequent in both groups (Home, 92.3% versus Hotel, 89.7%). A suture and follow-up were less likely in the Home group than in the Hotel group (18.8% versus 42.4%, 39.1% versus 69.7%, respectively). CONCLUSIONS: In hotel rooms, head and / or facial injuries due to falling from a bed were the most common types of injury, and they often needed more invasive procedures than home injuries. Injuries that tended to occur in hotel rooms are more predictable than home injuries. Modification of the surrounding environment has the potential to prevent unintentional injuries not only in the home environment but also in hotel rooms.

Body shape indices are predictors for estimating fat-free mass in male athletes.

It is unknown whether body size and body shape parameters can be predictors for estimating whole body fat-free mass (FFM) in male athletes. This study aimed to investigate whether body size and shape variables can be predictors for FFM in male athletes. Using a whole-body dual-energy X-ray absorptiometry scanner, whole body fat mass (FM) and FFM were determined in 132 male athletes and 14 sedentary males. The sample was divided into two groups: validation (N = 98) and cross-validation (N = 48) groups. Body height (BH), body mass (BM), and waist circumference at immediately above the iliac crest (W) were measured. BM-to-W and W-to-BH ratios were calculated as indices of body shapes. Stepwise multiple regression analysis revealed that BM/W and W/BH were selected as explainable variables for predicting FFM. The equation developed in the validation group was FFM (kg) = 0.883 × BM/W (kg/m) + 43.674 × W/BH (cm/cm)- 41.480 [R2 = 0.900, SEE (%SEE) = 2.3 kg (3.8%)], which was validated in the cross-validation group. Thus, the current results demonstrate that an equation using BM/W and W/BH as independent variables is applicable for predicting FFM in male athletes.

Synthesis of antimicrobial cyclodextrins bearing polyarylamino and polyalkylamino groups via click chemistry for bacterial membrane disruption.

Cyclodextrin derivatives are synthesized as membrane-disrupting agents via a microwave-assisted Huisgen reaction. Their ability to permeabilize bacterial membranes depends on the amino substituents and an appropriate balance of hydrophobicity and hydrophilicity, thus enabling the preparation of derivatives with selective toxicity against bacteria.

Structure and characterization of amidase from Rhodococcus sp. N-771: Insight into the molecular mechanism of substrate recognition.

In this study, we have structurally characterized the amidase of a nitrile-degrading bacterium, Rhodococcus sp. N-771 (RhAmidase). RhAmidase belongs to amidase signature (AS) family, a group of amidase families, and is responsible for the degradation of amides produced from nitriles by nitrile hydratase. Recombinant RhAmidase exists as a dimer of about 107 kDa. RhAmidase can hydrolyze acetamide, propionamide, acrylamide and benzamide with kcat/Km values of 1.14+/-0.23 mM(-1)s(-1), 4.54+/-0.09 mM(-1)s(-1), 0.087+/-0.02 mM(-1)s(-1) and 153.5+/-7.1 mM(-1)s(-1), respectively. The crystal structures of RhAmidase and its inactive mutant complex with benzamide (S195A/benzamide) were determined at resolutions of 2.17 A and 2.32 A, respectively. RhAmidase has three domains: an N-terminal alpha-helical domain, a small domain and a large domain. The N-terminal alpha-helical domain is not found in other AS family enzymes. This domain is involved in the formation of the dimer structure and, together with the small domain, forms a narrow substrate-binding tunnel. The large domain showed high structural similarities to those of other AS family enzymes. The Ser-cis Ser-Lys catalytic triad is located in the large domain. But the substrate-binding pocket of RhAmidase is relatively narrow, due to the presence of the helix alpha13 in the small domain. The hydrophobic residues from the small domain are involved in recognizing the substrate. The small domain likely participates in substrate recognition and is related to the difference of substrate specificities among the AS family amidases.

Novel catalytic activity of nitrile hydratase from Rhodococcus sp. N771.

Nitrile hydratase (NHase) from Rhodococcus sp. N771 is a non-heme iron enzyme catalyzing the hydration of various nitriles to the corresponding amides. We report a novel catalytic activity of NHase. When NHase was incubated with an large excess of commercially available isovaleronitrile, the charge transfer band from the sulfur ligand to the Fe atom shifted from 710 nm to 820 nm, but recovered within 4 min. Similar UV-Vis absorption changes were observed after the addition of isobutylisonitrile (iBuNC), a major impurity in commercially available isovaleronitrile, suggesting that NHase catalyzes the conversion of iBuNC to other compounds. The reaction product was identified as isobutylamine (iBuNH(2)) by liquid chromatography tandem mass spectrometry. NHase also converts t-butylisonitrile and 1,1,3,3,-tetramethylbutylisonitrile to the corresponding amines. Kinetic analysis of the conversion of iBuNC to iBuNH(2) showed a K(m) value comparable to that for nitriles, while the V(max) value was more than 10(5) times smaller than that for methacrylonitrile. This is the first report suggesting that NHase is a bifunctional enzyme catalyzing a reaction other than the hydration of nitriles.