Characterization of newly isolated thermotolerant bacterium Cupriavidus sp. CB15 from composting and its ability to produce polyhydroxyalkanoate from glycerol.

BACKGROUND: This study aimed to isolate a novel thermotolerant bacterium that is capable of synthesizing polyhydroxyalkanoate from glycerol under high temperature conditions. RESULTS: A newly thermotolerant polyhydroxyalkanoate (PHA) producing bacterium, Cupriavidus sp. strain CB15, was isolated from corncob compost. The potential ability to synthesize PHA was confirmed by detection of PHA synthase (phaC) gene in the genome. This strain could produce poly(3-hydroxybutyrate) [P(3HB)] with 0.95 g/L (PHA content 75.3 wt% of dry cell weight 1.24 g/L) using glycerol as a carbon source. The concentration of PHA was enhanced and optimized based on one-factor-at-a-time (OFAT) experiments and response surface methodology (RSM). The optimum conditions for growth and PHA biosynthesis were 10 g/L glycerol, 0.78 g/L NH4Cl, shaking speed at 175 rpm, temperature at 45 °C, and cultivation time at 72 h. Under the optimized conditions, PHA production was enhanced to 2.09 g/L (PHA content of 74.4 wt% and dry cell weight of 2.81 g/L), which is 2.12-fold compared with non-optimized conditions. Nuclear magnetic resonance (NMR) analysis confirmed that the extracted PHA was a homopolyester of 3-hydyoxybutyrate. CONCLUSION: Cupriavidus sp. strain CB15 exhibited potential for cost-effective production of PHA from glycerol.

Discovery of a novel series of medium-sized cyclic enteropeptidase inhibitors.

Enteropeptidase is located in the duodenum that involved in intestinal protein digestion. We have reported enteropeptidase inhibitors with low systemic exposure. The aim of this study was to discover novel enteropeptidase inhibitors showing more potent in vivo efficacy while retaining low systemic exposure. Inhibitory mechanism-based drug design led us to cyclize ester 2 to medium-sized lactones, showing potent enteropeptidase inhibitory activity and improving the ester stability, thus increasing fecal protein output in vivo. Optimization on the linker between two benzene rings resulted in discovery of ether lactone 6b, exhibiting further enhanced enteropeptidase inhibitory activity and long duration of inhibitory state. Oral administration of 6b in mice significantly elevated fecal protein output compared with the lead 2. In addition, 6b showed low systemic exposure along with low intestinal absorption. Furthermore, we identified the 10-membered lactonization method for scale-up synthesis of 6b, which does not require high-dilution conditions.

Feeding gastrostomy and duodenostomy using the round ligament of the liver versus conventional feeding jejunostomy after esophagectomy: a meta-analysis.

Esophageal cancer patients require enteral nutritional support after esophagectomy. Conventional feeding enterostomy to the jejunum (FJ) is occasionally associated with small bowel obstruction because the jejunum is fixed to the abdominal wall. Feeding through an enteral feeding tube inserted through the reconstructed gastric tube (FG) or the duodenum (FD) using the round ligament of the liver have been suggested as alternatives. This meta-analysis aimed to compare short-term outcomes between FG/FD and FJ. Studies published prior to May 2022 that compared FG or FD with FJ in cancer patients who underwent esophagectomy were identified via electronic literature search. Meta-analysis was performed using the Mantel-Haenszel random-effects model to calculate Odds Ratios (ORs) with 95% confidence intervals (CIs). Five studies met inclusion criteria to yield a total of 1687 patients. Compared with the FJ group, the odds of small bowel obstruction (OR 0.09; 95% CI, 0.02-0.33), catheter site infection (OR 0.18; 95% CI, 0.06-0.51) and anastomotic leakage (OR 0.53; 95% CI, 0.32-0.89) were lower for the FG/FD group. Odds of pneumonia, recurrent laryngeal nerve palsy, chylothorax and hospital mortality did not significantly differ between the groups. The length of hospital stay was shorter for the FG/FD group (median difference, -10.83; 95% CI, -18.55 to -3.11). FG and FD using the round ligament of the liver were associated with lower odds of small bowel obstruction, catheter site infection and anastomotic leakage than FJ in esophageal cancer patients who underwent esophagectomy.

[A Case of Pathological Complete Response Achieved with Neoadjuvant DCS Therapy in Gastric Cancer].

A 77-year-old man with appetite loss was referred to our hospital. Upper gastrointestinal endoscopy and computed tomography(CT)revealed advanced gastric cancer in the antrum with duodenal and pancreatic invasion. After 6 courses of neoadjuvant docetaxel, cisplatin, and S-1(DCS)therapy, CT revealed marked tumor shrinkage. Distal gastrectomy was performed. Histopathological examination showed no residual tumor cells or lymph node metastasis, and thus, finally, pathological complete response was considered to have been achieved. The patient was doing well and disease-free 3 years later. Thus, neoadjuvant DCS therapy can be a promising treatment option for borderline resectable advanced gastric cancer.

Improvement of the halotolerance of a Bacillus serine protease by protein surface engineering.

A moderately halotolerant serine protease was previously isolated from Bacillus subtilis from salted, fermented food. Eight mutation sites on the protein surface were selected for protein engineering based on sequence and structural comparisons with moderately halotolerant proteases and homologous non-halotolerant proteases. The newly constructed multiple mutants with substituted Asp and Arg residues were compared with the recombinant wild type (rApr) and the previously constructed mAla-8 substituted with Ala to analyze the contribution of protein surface charge to the salt adaptation of the protease. The three mutants showed >1.2-fold greater halotolerance than rApr. In addition, the mutants showed a broader range of pH stability than rApr, retaining >80% of their maximum activity in the pH range 5.0-11. The mutants also retained >75% of their activity after incubation for 1 h at pH 8.0 and 55°C or at pH 11.5 and 25°C. The Asp and Arg residues exchanged by multiple substitution probably played a role in increasing protein surface hydration and solubility in high salt conditions. This study illustrated that increasing a high proportion of the negative or positive charge on the surface of the Bacillus serine protease stably improved the protein's salt adaptation.

Chronic Exposure to SCO-267, an Allosteric GPR40 Full Agonist, Is Effective in Improving Glycemic Control in Rats.

Full agonist-mediated activation of free fatty acid receptor 1 (FFAR1/GPR40) alleviates diabetes in rodents. Considering that diabetes is a chronic disease, assessment of treatment durability of chronic exposure to a GPR40 full agonist is pivotal for treating patients with diabetes. However, the physiologic significance of chronic in vitro and in vivo exposure to GPR40 full agonists is largely unclear. Here, we evaluated the in vitro and in vivo effects of chronic treatment with SCO-267, a GPR40 full agonist, on signal transduction and glucose control. In vitro experiments showed that SCO-267 is an allosteric full agonist for GPR40, which activates the Gα q, Gα s, and Gα 12/13 pathways and ß-arrestin recruitment. The calcium signal response was largely sustained in GPR40-overexpressing CHO cells even after prolonged incubation with SCO-267. To evaluate the in vivo relevance of chronic exposure to GPR40 full agonists, SCO-267 (1 and 10 mg/kg) was administered once daily to neonatally streptozotocin-induced diabetic rats for 15-33 days, and glucose control was evaluated. After 15 days of dosing followed by the drug washout period, SCO-267 improved glucose tolerance, most likely by increasing insulin sensitivity in rats. After 33 days, repeated exposure to SCO-267 was highly effective in improving glucose tolerance in rats. Furthermore, chronic exposure to SCO-267 increased pancreatic insulin content. These results demonstrated that even after chronic exposure, SCO-267 effectively activates GPR40 in cells and rats, suggesting the clinical application of SCO-267 in treating chronic diseases including diabetes. SIGNIFICANCE STATEMENT: GPR40 is a validated therapeutic target for diabetes. This study showed that even after chronic exposure, SCO-267, an allosteric GPR40 full agonist, effectively activates GPR40 in cells and rats; these results suggest a durable efficacy of SCO-267 in patients.

Selective Acetyl-CoA Carboxylase 1 Inhibitor Improves Hepatic Steatosis and Hepatic Fibrosis in a Preclinical Nonalcoholic Steatohepatitis Model.

Acetyl-CoA carboxylase (ACC) 1 and ACC2 are essential rate-limiting enzymes that synthesize malonyl-CoA (M-CoA) from acetyl-CoA. ACC1 is predominantly expressed in lipogenic tissues and regulates the de novo lipogenesis flux. It is upregulated in the liver of patients with nonalcoholic fatty liver disease (NAFLD), which ultimately leads to the formation of fatty liver. Therefore, selective ACC1 inhibitors may prevent the pathophysiology of NAFLD and nonalcoholic steatohepatitis (NASH) by reducing hepatic fat, inflammation, and fibrosis. Many studies have suggested ACC1/2 dual inhibitors for treating NAFLD/NASH; however, reports on selective ACC1 inhibitors are lacking. In this study, we investigated the effects of compound-1, a selective ACC1 inhibitor for treating NAFLD/NASH, using preclinical in vitro and in vivo models. Compound-1 reduced M-CoA content and inhibited the incorporation of [14C] acetate into fatty acids in HepG2 cells. Additionally, it reduced hepatic M-CoA content and inhibited de novo lipogenesis in C57BL/6J mice after a single dose. Furthermore, compound-1 treatment of 8 weeks in Western diet-fed melanocortin 4 receptor knockout mice-NAFLD/NASH mouse model-improved liver hypertrophy and reduced hepatic triglyceride content. The reduction of hepatic M-CoA by the selective ACC1 inhibitor was highly correlated with the reduction in hepatic steatosis and fibrosis. These findings support further investigations of the use of this ACC1 inhibitor as a new treatment of NFLD/NASH. SIGNIFICANCE STATEMENT: This is the first study to demonstrate that a novel selective inhibitor of acetyl-CoA carboxylase (ACC) 1 has anti-nonalcoholic fatty liver disease (NAFLD) and anti-nonalcoholic steatohepatitis (NASH) effects in preclinical models. Treatment with this compound significantly improved hepatic steatosis and fibrosis in a mouse model. These findings support the use of this ACC1 inhibitor as a new treatment for NAFLD/NASH.

Enteropeptidase inhibitor SCO-792 effectively prevents kidney function decline and fibrosis in a rat model of chronic kidney disease.

BACKGROUND: Inhibiting enteropeptidase, a gut serine protease regulating protein digestion, suppresses food intake and ameliorates obesity and diabetes in mice. However, the effects of enteropeptidase inhibition on kidney parameters are largely unknown. Here, we evaluated the chronic effects of an enteropeptidase inhibitor, SCO-792, on kidney function, albuminuria and kidney pathology in spontaneously hypercholesterolaemic (SHC) rats, a rat chronic kidney disease (CKD) model. METHODS: SCO-792, an orally available enteropeptidase inhibitor, was administered [0.03% and 0.06% (w/w) in the diet] to 20-week-old SHC rats showing albuminuria and progressive decline in glomerular filtration rate (GFR) for five weeks. The effects of SCO-792 and the contribution of amino acids to these effects were evaluated. RESULTS: SCO-792 increased the faecal protein content, indicating that SCO-792 inhibited enteropeptidase in SHC rats. Chronic treatment with SCO-792 prevented GFR decline and suppressed albuminuria. Moreover, SCO-792 improved glomerulosclerosis and kidney fibrosis. Pair feeding with SCO-792 (0.06%) was less effective in preventing GFR decline, albuminuria and renal histological damage than SCO-792 treatment, indicating the enteropeptidase-inhibition-dependent therapeutic effects of SCO-792. SCO-792 did not affect the renal plasma flow, suggesting that its effect on GFR was mediated by an improvement in filtration fraction. Moreover, SCO-792 increased hydrogen sulphide production capacity, which has a role in tissue protection. Finally, methionine and cysteine supplementation to the diet abrogated SCO-792-induced therapeutic effects on albuminuria. CONCLUSIONS: SCO-792-mediated inhibition of enteropeptidase potently prevented GFR decline, albuminuria and kidney fibrosis; hence, it may have therapeutic potential against CKD.

Enteropeptidase inhibition improves obesity by modulating gut microbiota composition and enterobacterial metabolites in diet-induced obese mice.

Enteropeptidase is a transmembrane serine protease localized in the lumen of the duodenum that acts as a key enzyme for protein digestion. SCO-792 is an orally available enteropeptidase inhibitor that has been reported to have therapeutic effects on obesity and diabetes in mice. However, the mechanism underlying the therapeutic effect of SCO-792 has not yet been fully elucidated. In this study, we evaluated the role of gut microbiota on SCO-792-induced body weight (BW) reduction in high-fat diet-induced obese (DIO) mice. Chronic administration of SCO-792 substantially decreased BW and food intake in DIO mice. While the pair-fed study uncovered food intake-independent mechanisms of BW reduction by SCO-792. Interestingly, antibiotics-induced microbiota elimination in the gut canceled SCO-792-induced BW reduction by nearly half without affecting the anorectic effect, indicating the involvement of gut microbiota in the anti-obesity mechanism that is independent of food intake reduction. Microbiome analysis revealed that SCO-792 altered the gut microbiota composition in DIO mice. Notably, it was found that the abundance of Firmicutes decreased while that of Verrucomicrobia increased at the phylum level. Increased abundance of Akkermansia muciniphila, a bacterium known to be useful for host metabolism, was observed in SCO-792-treated mice. Fecal metabolome analysis revealed increased amino acid levels, indicating gut enteropeptidase inhibition. In addition, SCO-792 was found to increase the level of short-chain fatty acids, including propionate, and bile acids in the feces, which all help maintain gut health and improve metabolism. Furthermore, it was found that SCO-792 induced the elevation of colonic immunoglobulin A (IgA) concentration, which may maintain the microbiota condition, in DIO mice. In conclusion, this study demonstrates the contribution of microbiota to SCO-792-induced BW reduction. Enteropeptidase-mediated regulation of microbiota, enterobacterial metabolites, and IgA in the gut may coordinately drive the therapeutic effects of SCO-792 in obesity.

Enteropeptidase inhibition improves kidney function in a rat model of diabetic kidney disease.

AIM: To examine the effects of an enteropeptidase inhibitor, SCO-792, on kidney function in rats. MATERIALS AND METHODS: The pharmacological effects of SCO-792 were evaluated in Wistar fatty (WF) rats, a rat model of diabetic kidney disease (DKD). RESULTS: Oral administration of SCO-792 increased faecal protein content and improved glycaemic control in WF rats. SCO-792 elicited a rapid decrease in urine albumin-to-creatinine ratio (UACR). SCO-792 also normalized glomerular hyperfiltration and decreased fibrosis, inflammation and tubular injury markers in the kidneys. However, pioglitazone-induced glycaemic improvement had no effect on kidney variables. Dietary supplementation of amino acids (AAs), which bypass the action of enteropeptidase inhibition, mitigated the effect of SCO-792 on UACR reduction, suggesting a pivotal role for enteropeptidase. Furthermore, autophagy activity in the glomerulus, which is impaired in DKD, was elevated in SCO-792-treated rats. Finally, a therapeutically additive effect on UACR reduction was observed with a combination of SCO-792 with irbesartan, an angiotensin II receptor blocker. CONCLUSIONS: This study is the first to demonstrate that enteropeptidase inhibition is effective in improving disease conditions in DKD. SCO-792-induced therapeutic efficacy is likely to be independent of glycaemic control and mediated by the regulation of AAs and autophagy. Taken together with a combination effect of irbesartan, SCO-792 may be a novel therapeutic option for patients with DKD.

Impact of clinical characteristics of colonic diverticular bleeding in extremely elderly patients treated with direct oral anti-coagulant drugs: a retrospective multi-center study.

Since there were no available data about colonic diverticular bleeding in extremely elderly patients (>80 years old) treated with direct oral anticoagulants (DOACs), we tried to determine clinical characteristics in those with colonic diverticular bleeding taking DOACs and to compare clinical outcomes of those in DOAC-treated to those in warfarin-treated . We enrolled DOAC-treated (n = 20) and warfarin-treated (n = 23) extremely elderly patients with diverticular bleeding diagnosed by colonoscopy. We performed a retrospective review of patients' medical charts and endoscopic findings. We classified colonic diverticular bleeding based on endoscopic features due to modified previous study following three groups, type A (active bleeding), type B (non-active bleeding) and type C (bleeding suspected). Clinical outcomes such as number of recurrent bleeding, thrombotic events and mortality were estimated. There were no differences in endoscopical features and clinical characteristics between patients treated with DOAC and warfarin therapy. However, the number of recurrent bleeding, frequency of required blood transfusions and units of blood transfusion in warfarin-treated patients were significantly higher (p<0.05) compared to those in DOAC-treated groups. In addition, mortality and thrombotic events did not differ between DOAC- and warfarin-treated patients. Clinical outcomes suggest that DOACs can be recommended for extremely elderly patients with colonic diverticular disease.

The GPR40 Full Agonist SCO-267 Improves Liver Parameters in a Mouse Model of Nonalcoholic Fatty Liver Disease without Affecting Glucose or Body Weight.

Full agonism of G-protein-coupled receptor 40 (GPR40)/free fatty acid 1 receptor improves glycemic control in diabetic rodents. However, the effects of GPR40 full agonism on liver parameters are largely unknown. In the present study, we examined the effects of a GPR40 full agonist, SCO-267, on liver parameters in a nondiabetic mouse model with early-stage nonalcoholic fatty liver disease (NAFLD). SCO-267 was orally administered to mice, which were fed a choline-deficient, l-amino acid-defined, high-fat diet (CDAHFD), a mouse model for NAFLD. An oral dose of SCO-267 increased levels of circulating glucagon and glucagon-like peptide-1 in CDAHFD-fed mice. In a chronic-dose experiment, effects of SCO-267 were compared with those of a dipeptidyl peptidase-4 inhibitor (alogliptin) and a sodium glucose cotransporter 2 inhibitor (dapagliflozin). SCO-267 decreased liver triglyceride content, weight, collagen content, and plasma alanine aminotransferase (ALT) levels without affecting food intake or glucose levels in CDAHFD-fed mice. Furthermore, SCO-267 decreased levels of liver thiobarbituric acid reactive substances (TBARS), markers of oxidative stress. Alogliptin and dapagliflozin had no effect on liver weight or levels of triglyceride, collagen, plasma ALT, and liver TBARS. SCO-267 elevated mRNA levels of molecules with roles in mitochondrial function and ß-oxidation while inhibiting those with roles in lipogenesis, inflammation, reactive oxygen species generation, and fibrosis in the liver, all of which were less evident with alogliptin and dapagliflozin. This is the first study to show that the GPR40 full agonist SCO-267 improves liver parameters without affecting glucose or body weight in a mouse model of NAFLD. SIGNIFICANCE STATEMENT: Full agonism of GPR40/free fatty acid 1 receptor signaling stimulates islet and gut hormone secretions. The present study is the first to show the treatment effects of GPR40 full agonism on liver parameters in a mouse model for nonalcoholic fatty liver disease.

Bactericidal and Antiproliferative Effects of Peripheral Parenteral Nutrition Solutions with Sodium Bisulfite on Pathogenic Microorganisms in Catheter Lumens.

Catheter-related bloodstream infections (CRBSIs) due to pathogenic microorganisms pose a major threat to patients requiring parenteral nutrition (PN). Additives contained in medicines and foods have antiproliferative and bacteriostatic effects on pathogenic microorganisms. Therefore, PN solutions containing additives may also have an antibacterial effect. However, so far, there have been no reports on or observations of a PN solution with bactericidal activity. In this study, we assessed several nutrition solutions with antimicrobial activities and investigated their effects on pathogenic microorganisms colonizing catheter lumens. We selected the highly acidic Plas-Amino® (PA), which contains a large amount of sodium bisulfite as a preservative and potentially has an antimicrobial effect. In this study, we used the following pathogenic bacteria as the main causatives of CRBSIs: Staphylococcus aureus, Staphylococcus epidermidis, Bacillus cereus, Serratia marcescens, Pseudomonas aeruginosa, and Candida albicans. We then created a catheter lumen microorganism contamination model and evaluated the antibacterial effect of PA; we found that all bacteria in the control group grew significantly in the catheter lumen in a time-dependent manner at 48 and 72 h. On the other hand, we demonstrated that PA has bactericidal effects on S. aureus, S. epidermidis, B. cereus, S. marcescens, and P. aeruginosa in the catheter lumen and confirmed that it has a remarkable antiproliferative effect on C. albicans. Hence, we concluded that highly acidic PN solutions that contain a preservative like sodium bisulfite have bactericidal and growth inhibition effects on microorganisms in the catheter lumens of patients with CRBSIs and patients with totally implantable central venous access devices, in whom it is difficult to remove the catheter.

Locomotor activity and histological changes observed in a mouse model of knee osteoarthritis.

[Purpose] This study aimed to elucidate the changes in locomotor activity in a mouse model of knee osteoarthritis (OA). [Materials and Methods] Fourteen 20-week-old mice were divided into control and OA groups. Knee OA was surgically induced under anesthesia by destabilizing the meniscus. The OA group was reared normally for 8 weeks following surgery, during which OA was induced. Locomotor activity was measured every hour for 8 weeks using an infrared locomotor activity measurement device. Histological changes were evaluated according to the classification-system of Glasson. [Results] Locomotor activity in the OA group significantly decreased up to 2 weeks after surgery. Histological findings in the control group revealed an irregular cartilage surface in a portion of the tibia with no other abnormalities. Contrastingly, those in the OA group had eburnation of the medial femoral condyle, as well as fibrillation and fissures in the medial tibial plateau. Histological scores in the OA group were significantly higher than the control group. [Conclusion] Locomotor activity evaluations, in addition to histological scores and findings, are imperative for studies aiming to clarify the disease state and effect of interventions using mice models.

SCO-267, a GPR40 Full Agonist, Improves Glycemic and Body Weight Control in Rat Models of Diabetes and Obesity.

The GPR40/FFA1 receptor is a G-protein-coupled receptor expressed in the pancreatic islets and enteroendocrine cells. Here, we report the pharmacological profiles of (3S)-3-cyclopropyl-3-{2-[(1-{2-[(2,2-dimethylpropyl)(6-methylpyridin-2-yl)carbamoyl]-5-methoxyphenyl}piperidin-4-yl)methoxy]pyridin-4-yl}propanoic acid (SCO-267), a novel full agonist of GPR40. Ca2+ signaling and insulin and glucagon-like peptide-1 (GLP-1) secretion were evaluated in GPR40-expressing CHO, MIN6, and GLUTag cells. Hormone secretions and effects on fasting glucose were tested in rats. Single or repeated dosing effects were evaluated in neonatally streptozotocin-induced diabetic rats (N-STZ-1.5 rats), diet-induced obese (DIO) rats, and GPR40-knockout (Ffar1-/- ) mice. Treatment with SCO-267 activated Gq signaling in both high- and low-FFAR1-expressing CHO cells, stimulated insulin secretion in MIN6 cells, and induced GLP-1 release in GLUTag cells. When administered to normal rats, SCO-267 increased insulin, glucagon, GLP-1, glucose-dependent insulinotropic peptide, and peptide YY (PYY) secretions under nonfasting conditions. These results show the full agonistic property of SCO-267 against GPR40. Hypoglycemia was not induced in SCO-267-treated rats during the fasting condition. In diabetic N-STZ-1.5 rats, SCO-267 was highly effective in improving glucose tolerance in single and 2-week dosing studies. DIO rats treated with SCO-267 for 2 weeks showed elevated plasma GLP-1 and PYY levels, reduced food intake, and decreased body weight. In wild-type mice, SCO-267 induced GLP-1 secretion, food intake inhibition, and body weight reduction; however, these effects were abolished in Ffar1-/- mice, indicating a GPR40-dependent mechanism. In conclusion, SCO-267 stimulated islet and gut hormone secretion, improved glycemic control in diabetic rats, and decreased body weight in obese rats. These data suggest the therapeutic potential of SCO-267 for the treatment of diabetes and obesity.

SCO-792, an enteropeptidase inhibitor, improves disease status of diabetes and obesity in mice.

AIMS: Enteropeptidase is a serine protease localized on the duodenal brush border that catalyzes the conversion of inactive trypsinogen into active trypsin, thereby regulating protein breakdown in the gut. We evaluated the effects of SCO-792, a novel enteropeptidase inhibitor, in mice. MATERIALS AND METHODS: In vivo inhibition of enteropeptidase was evaluated via an oral protein challenge. Pharmacological effects were evaluated in normal mice, in diet-induced obese (DIO) mice and in obese and diabetic ob/ob mice. RESULTS: A single oral administration of SCO-792 inhibited plasma branched-chain amino acids (BCAAs) in an oral protein challenge test in mice, indicating in vivo inhibition of enteropeptidase. Repeated treatment with SCO-792 induced reduction in food intake and decrease in body weight in DIO and ob/ob mice. Plasma FGF21 levels were increased in SCO-792-treated DIO mice, an observation that was probably independent of reduction in food intake. Hyperglycaemia was markedly improved in SCO-792-treated ob/ob mice. A hyperinsulinaemic-euglycaemic clamp study revealed improved muscle insulin sensitivity in SCO-792-treated ob/ob mice. SCO-792 also improved plasma and liver lipid profiles and decreased plasma alanine transaminase, suggesting a potential treatment for liver diseases. Dietary supplementation with essential amino acids attenuated the effect of SCO-792 on reduction in food intake and decrease in body weight in normal mice, suggesting a pivotal role for enteropeptidase in these biological phenomena. CONCLUSIONS: SCO-792 inhibited enteropeptidase in vivo, reduced food intake, decreased body weight, increased insulin sensitivity, improved glucose and lipid control, and ameliorated liver parameters in mouse models with obesity and/or diabetes. SCO-792 may exhibit similar effects in patients.

Efficient bioconversion of enzymatic corncob hydrolysate into biomass and lipids by oleaginous yeast Rhodosporidium paludigenum KM281510.

This study aims to valorize of enzymatic corncob hydrolysate (ECH) for biomass and lipid productions via microbial bioconversion by the efficient oleaginous yeast Rhodosporidium paludigenum KM281510. Biomass (5.63 g/L), lipids (3.29 g/L), and lipid content (58% (g/g)) were observed by batch cultivation in shaking flask. The effect of total sugar concentration in ECH, agitation rate, temperature, and initial pH were investigated in both batch and fed-batch cultivations by shaking flask and 3.0 L airlift bioreactor. Biomass, lipids, and lipid content of 27.77 g/L, 20.27 g/L, and 70% (g/g) were obtained with 100 g/L total sugar (68 g/L glucose, 29 g/L xylose, and 3 g/L arabinose), pH 6.5, 25 °C, 6.0 vvm, for 7 days by batch cultivation in bioreactor. Surprisingly, production parameters were improved by fed-batch, wherein these promising high biomass (36.06 g/L), lipid production (25.12 g/L), and lipid productivity (2.52 g/L/d) values were achieved. Moreover, fed-batch cultivation promoted the utilization of xylose (2.5-times) and arabinose (3.4-times) higher than batch cultivation, achieving lipid content (70% (g/g)) with oleic acid (53%). These results would be helpful for understanding the comprehensive utilization of ECH, especially the pentose sugars, for growth and lipogenesis of oleaginous yeast as well as being a qualified biodiesel feedstock.

Daily and short-term application of joint movement for the prevention of infrapatellar fat pad atrophy due to immobilization.

[Purpose] To mobilize the knee joint during cast fixation and to determine whether infrapatellar fat pad changes can be prevented. [Materials and Methods] We randomly allocated Wistar rats into 3 groups as follows: normal group, raised in normal conditions (n=5); contracture group, immobilized with cast fixation (n=5); and prevention group, treated with joint movement during immobilization (n=5). We immobilized the right hindlimb using cast fixation. Joint movement in the prevention group was accomplished by repeatedly pulling the right hindlimb caudally and then returning the leg to the bent position for 10 minutes every day for 2 weeks. We used a metronome to maintain a constant speed, with one set lasting 2 seconds (1-second traction and 1-second return). [Results] The contracture group had adipose cells of various sizes and fibrosis in the infrapatellar fat pad. These changes were also found in milder forms in the prevention group. We found significant differences in the cross section of adipose cells and in knee extension restriction between the groups. [Conclusion] Promoting joint movement may not only have a therapeutic effect on adipose cells but also a preventative effect.

Sonication versus Tissue Sampling for Diagnosis of Prosthetic Joint and Other Orthopedic Device-Related Infections.

Current guidelines recommend collection of multiple tissue samples for diagnosis of prosthetic joint infections (PJI). Sonication of explanted devices has been proposed as a potentially simpler alternative; however, reported microbiological yield varies. We evaluated sonication for diagnosis of PJI and other orthopedic device-related infections (DRI) at the Oxford Bone Infection Unit between October 2012 and August 2016. We compared the performance of paired tissue and sonication cultures against a "gold standard" of published clinical and composite clinical and microbiological definitions of infection. We analyzed explanted devices and a median of five tissue specimens from 505 procedures. Among clinically infected cases the sensitivity of tissue and sonication culture was 69% (95% confidence interval, 63 to 75) and 57% (50 to 63), respectively (P < 0.0001). Tissue culture was more sensitive than sonication for both PJI and other DRI, irrespective of the infection definition used. Tissue culture yield was higher for all subgroups except less virulent infections, among which tissue and sonication culture yield were similar. The combined sensitivity of tissue and sonication culture was 76% (70 to 81) and increased with the number of tissue specimens obtained. Tissue culture specificity was 97% (94 to 99), compared with 94% (90 to 97) for sonication (P = 0.052) and 93% (89 to 96) for the two methods combined. Tissue culture is more sensitive and may be more specific than sonication for diagnosis of orthopedic DRI in our setting. Variable methodology and case mix may explain reported differences between centers in the relative yield of tissue and sonication culture. Culture yield was highest for both methods combined.

Characterization and mutation analysis of a halotolerant serine protease from a new isolate of Bacillus subtilis.

OBJECTIVES: A bacterial halotolerant enzyme was characterized to understand the molecular mechanism of salt adaptation and to explore its protein engineering potential. RESULTS: Halotolerant serine protease (Apr_No16) from a newly isolated Bacillus subtilis strain no. 16 was characterized. Multiple alignments with previously reported non-halotolerant proteases, including subtilisin Carlsberg, indicated that Apr_No16 has eight acidic or polar amino acid residues that are replaced by nonpolar amino acids in non-halotolerant proteases. Those residues were hypothesized to be one of the primary contributors to salt adaptation. An eightfold mutant substituted with Ala residues exhibited 1.2- and 1.8-fold greater halotolerance at 12.5% (w/v) NaCl than Apr_No16 and Carlsberg, respectively. Amino acid substitution notably shifted the theoretical pI of the eightfold mutant, from 6.33 to 9.23, compared with Apr_No16. The resulting protein better tolerated high salt conditions. CONCLUSIONS: Changing the pI of a bacterial serine protease may be an effective strategy to improve the enzyme's halotolerance.