Regioselective biooxidation of (+)-valencene by recombinant E. coli expressing CYP109B1 from Bacillus subtilis in a two-liquid-phase system

<p>Abstract</p> <p>Background</p> <p>(+)-Nootkatone (<b>4</b>) is a high added-value compound found in grapefruit juice. Allylic oxidation of the sesquiterpene (+)-valencene (<b>1</b>) provides an attractive route to this sought-after flavoring. So far, chemical methods to produce (+)-nootkatone (<b>4</b>) from (+)-valencene (<b>1</b>) involve unsafe toxic compounds, whereas several biotechnological approaches applied yield large amounts of undesirable byproducts. In the present work 125 cytochrome P450 enzymes from bacteria were tested for regioselective oxidation of (+)-valencene (<b>1</b>) at allylic C2-position to produce (+)-nootkatone (<b>4</b>) via <it>cis</it>- (<b>2</b>) or <it>trans</it>-nootkatol (<b>3</b>). The P450 activity was supported by the co-expression of putidaredoxin reductase (PdR) and putidaredoxin (Pdx) from <it>Pseudomonas putida </it>in <it>Escherichia coli</it>.</p> <p>Results</p> <p>Addressing the whole-cell system, the cytochrome CYP109B1 from <it>Bacillus subtilis </it>was found to catalyze the oxidation of (+)-valencene (<b>1</b>) yielding nootkatol (<b>2 </b>and <b>3</b>) and (+)-nootkatone (<b>4</b>). However, when the <it>in vivo </it>biooxidation of (+)-valencene (<b>1</b>) with CYP109B1 was carried out in an aqueous milieu, a number of undesired multi-oxygenated products has also been observed accounting for approximately 35% of the total product. The formation of these byproducts was significantly reduced when aqueous-organic two-liquid-phase systems with four water immiscible organic solvents – isooctane, <it>n</it>-octane, dodecane or hexadecane – were set up, resulting in accumulation of nootkatol (<b>2 </b>and <b>3</b>) and (+)-nootkatone (<b>4</b>) of up to 97% of the total product. The best productivity of 120 mg l^-1of desired products was achieved within 8 h in the system comprising 10% dodecane.</p> <p>Conclusion</p> <p>This study demonstrates that the identification of new P450s capable of producing valuable compounds can basically be achieved by screening of recombinant P450 libraries. The biphasic reaction system described in this work presents an attractive way for the production of (+)-nootkatone (<b>4</b>), as it is safe and can easily be controlled and scaled up.</p

NPC1L1 inhibitor ezetimibe is a reliable therapeutic agent for non-obese patients with nonalcoholic fatty liver disease

<p>Abstract</p> <p>Background</p> <p>We recently examined the distribution of abdominal fat, dietary intake and biochemical data in patients with nonalcoholic fatty liver disease (NAFLD) and found that non-obese NAFLD patients did not necessarily exhibit insulin resistance and/or dysregulated secretion of adipocytokines. However, dietary cholesterol intake was superabundant in non-obese patients compared with obese patients, although total energy and carbohydrate intake was not excessive. Therefore, excess cholesterol intake appears to be one of the main factors associated with NAFLD development and liver injury.</p> <p>Methods</p> <p>We reviewed a year of follow-up data of non-obese NAFLD patients treated with Niemann-Pick C1 like 1 inhibitor ezetimibe to evaluate its therapeutic effect on clinical parameters related to NAFLD. Without any dietary or exercise modification, 10 mg/day of ezetimibe was given to 8 patients. In 4 of 8 patients, ezetimibe was administered initially. In the remaining 4 patients, medication was switched from ursodeoxycholic acid to ezetimibe.</p> <p>Results</p> <p>In each patient, body mass index was maintained under 25 kg/m²during the observation period. Serum ALT levels significantly decreased within 6 months and in 4 patients levels reached the normal range (<30 U/L), which was accompanied with at least a 10% decrease in serum total cholesterol and LDL-cholesterol. However, ultrasonographic findings of fatty liver did not show obvious improvement for a year.</p> <p>Conclusion</p> <p>We conclude that the cholesterol absorption inhibitor ezetimibe can suppress hepatic injury in non-obese patients with NAFLD and that ezetimibe may offer a novel treatment for NAFLD.</p

Aggregation of scaffolding protein DISC1 dysregulates phosphodiesterase 4 in Huntington’s disease

Huntington’s disease (HD) is a polyglutamine (polyQ) disease caused by aberrant expansion of the polyQ tract in Huntingtin (HTT). While motor impairment mediated by polyQ-expanded HTT has been intensively studied, molecular mechanisms for nonmotor symptoms in HD, such as psychiatric manifestations, remain elusive. Here we have demonstrated that HTT forms a ternary protein complex with the scaffolding protein DISC1 and cAMP-degrading phosphodiesterase 4 (PDE4) to regulate PDE4 activity. We observed pathological cross-seeding between DISC1 and mutant HTT aggregates in the brains of HD patients as well as in a murine model that recapitulates the polyQ pathology of HD (R6/2 mice). In R6/2 mice, consequent reductions in soluble DISC1 led to dysregulation of DISC1-PDE4 complexes, aberrantly increasing the activity of PDE4. Importantly, exogenous expression of a modified DISC1, which binds to PDE4 but not mutant HTT, normalized PDE4 activity and ameliorated anhedonia in the R6/2 mice. We propose that cross-seeding of mutant HTT and DISC1 and the resultant changes in PDE4 activity may underlie the pathology of a specific subset of mental manifestations of HD, which may provide an insight into molecular signaling in mental illness in general

Genetic Polymorphisms of IL-17F and TRAF3IP2 Could Be Predictive Factors of the Long-Term Effect of Infliximab against Crohn’s Disease

Background. We aimed to identify certain genes related to response to infliximab (IFX) and biomarkers to predict the IFX effect for Japanese Crohn\u27s disease (CD) patients by performing an association study of single nucleotide polymorphisms (SNPs) in candidate genes in the interleukin-(IL-) 17 signaling pathway with response to IFX after 1 year of treatment. Methods. A total of 103 patients were divided into two groups, responders and nonresponders. Twenty-eight tag SNPs in 5 genes were genotyped. The frequencies of alleles and genotypes of each SNP were compared between responders and nonresponders in three different inheritance models. A genetic test was performed using a combination of the associated SNPs as biomarkers. Results. Multivariate logistic regression analysis indicated that the four variable factors, concomitant use of immunomodulators, penetrating disease, a G/G genotype of rs766748 in IL-17F, and a C/C or C/A genotype of rs1883136 in TRAF3IP2, independently contributed to response to IFX after 1 year of treatment. Genetic test using the polymorphisms of these genes perfectly predicted the responder and nonresponder CD patients with both concomitant use of immunomodulators and penetrating disease. Conclusion. IL17F and TRAF3IP2 are one of IFX-related genes, useful as biomarkers of IFX response, and may be target molecules for new therapeutic drugs

Efficacy of capillary pattern type IIIA/IIIB by magnifying narrow band imaging for estimating depth of invasion of early colorectal neoplasms

<p>Abstract</p> <p>Background</p> <p>Capillary patterns (CP) observed by magnifying Narrow Band Imaging (NBI) are useful for differentiating non-adenomatous from adenomatous colorectal polyps. However, there are few studies concerning the effectiveness of magnifying NBI for determining the depth of invasion in early colorectal neoplasms. We aimed to determine whether CP type IIIA/IIIB identified by magnifying NBI is effective for estimating the depth of invasion in early colorectal neoplasms.</p> <p>Methods</p> <p>A series of 127 consecutive patients with 130 colorectal lesions were evaluated from October 2005 to October 2007 at the National Cancer Center Hospital East, Chiba, Japan. Lesions were classified as CP type IIIA or type IIIB according to the NBI CP classification. Lesions were histopathologically evaluated. Inter and intraobserver variabilities were assessed by three colonoscopists experienced in NBI.</p> <p>Results</p> <p>There were 15 adenomas, 66 intramucosal cancers (pM) and 49 submucosal cancers (pSM): 16 pSM superficial (pSM1) and 33 pSM deep cancers (pSM2-3). Among lesions diagnosed as CP IIIA 86 out of 91 (94.5%) were adenomas, pM-ca, or pSM1; among lesions diagnosed as CP IIIB 28 out of 39 (72%) were pSM2-3. Sensitivity, specificity and diagnostic accuracy of the CP type III for differentiating pM-ca or pSM1 (<1000 μm) from pSM2-3 (≥1000 μm) were 84.8%, 88.7 % and 87.7%, respectively. Interobserver variability: κ = 0.68, 0.67, 0.72. Intraobserver agreement: κ = 0.79, 0.76, 0.75</p> <p>Conclusion</p> <p>Identification of CP type IIIA/IIIB by magnifying NBI is useful for estimating the depth of invasion of early colorectal neoplasms.</p

更年期女性の血清脂質に及ぼす閉経, トレーニング, 女性ホルモンの影響

Fifty-two women studies to determine the effect of menopause, training, estron, and estradiol on serum levels of total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), and triglyceride (TG). Subjects were assigned to one of two groups : trained endurance runners (average age=48.5±6.9 years ; training history=12.7±5.9 years ; training frequency per week=4.5±1.3 time ; training distance per day=8.9±4.7 km ; training duration ; n=24) ; control group (average age= 49.4±5.9 years ; n=28) . The post menopausal women undergone natural menopause. None of the subjects smoked, cardiovascular disease, or any from the hormonal replacement. Analysis of variance (ANOVA) indicated : 1) Significant difference between the traind group and the control group in measured of percentage of body fat (%BF : 21.8% ; 26.6%), (p<0.001) and the heart rate at ventilation threshold (HR at VT : 133.6bpm ; 116.5bpm) (p<0.001) and Vo_2 at VT (30.4ml/kg/min;18.9ml/kg/min) (p<0.001) ; and HDL-C (77.8mg/dl; 67.2mg/dl). 2) This %BF, HR at VT, Vo_2 at VT, and HDL-C tendency was identified when age differences on among the groups were statistically controlled. 3) In a simple correlation analysis, age showed a significant positively associated with TC (r=0.409), and %BF was positively associated with TC (r=0.579) and LDL-C (r=0.610), and BMI was positively associated with LDL-C (0.514), and significant positive correlation was found between HR at VT and HDL-C (r=0.514) in trained group. On the other hand, age was positively associated with TG (r=0.419) in control group, and %BF was negatively associated with HDL-C (r=-0.378). 4) Analysis of covariance, HDL-C was affected with training rather than menopause. 5) Estron was significantly and positively correlated with HDL-C (r=0.587) in post menopausal traind group. These data suggested that training prior to menopause had a positive effect on HDL-C levels. This effect continued with training following menopause and HDL-C levels were also maintained. This appears to controlled by hormones such as estrogen

An operative case of hepatic pseudolymphoma difficult to differentiate from primary hepatic marginal zone B-cell lymphoma of mucosa-associated lymphoid tissue

Hepatic pseudolymphoma (HPL) and primary hepatic marginal zone B cell lymphoma of mucosa-associated lymphoid tissue (MALT lymphoma) are rare diseases and the differential diagnosis between these two entities is sometimes difficult. We herein report a 56-year-old Japanese woman who was pointed out to have a space occupying lesion in the left lateral segment of the liver. Hepatitis viral-associated antigen/antibody was negative and liver function tests including lactic dehydrogenase, peripheral blood count, tumor markers and soluble interleukin-2 receptor were all within normal limit. Imaging study using computed tomography and magnetic resonance imaging were not typical for hepatocellular carcinoma, cholangiocarcinoma, or other metastatic cancer. Fluorodeoxyglucose-positron emission tomography examination integrated with computed tomography scanning showed high standardized uptake value in the solitary lesion in the liver. Under a diagnosis of primary liver neoplasm, laparoscopic-assisted lateral segmentectomy was performed. Liver tumor of maximal 1.0 cm in diameter was consisted of aggregation of lymphocytes of predominantly B-cell, containing multiple lymphocyte follicles positive for CD10 and bcl-2, consistent with a diagnosis of HPL rather than MALT lymphoma, although a definitive differentiation was pending. The background liver showed non-alcoholic fatty liver disease/early non-alcoholic steatohepatitis. The patient is currently doing well with no sign of relapse 13 months after the surgery. Since the accurate diagnosis is difficult, laparoscopic approach would provide a reasonable procedure of diagnostic and therapeutic advantage with minimal invasiveness for patients. Considering that the real nature of this entity remains unclear, vigilant follow-up of patient is essential

Bias corrections of GOSAT SWIR XCO₂ and XCH₄ with TCCON data and their evaluation using aircraft measurement data

We describe a method for removing systematic biases of column-averaged dry air mole fractions of CO2 (XCO2) and CH4 (XCH4) derived from short-wavelength infrared (SWIR) spectra of the Greenhouse gases Observing SATellite (GOSAT). We conduct correlation analyses between the GOSAT biases and simultaneously retrieved auxiliary parameters. We use these correlations to bias correct the GOSAT data, removing these spurious correlations. Data from the Total Carbon Column Observing Network (TCCON) were used as reference values for this regression analysis. To evaluate the effectiveness of this correction method, the tnzuncorrected/corrected GOSAT data were compared to independent XCO2 and XCH4 data derived from aircraft measurements taken for the Comprehensive Observation Network for TRace gases by AIrLiner (CONTRAIL) project, the National Oceanic and Atmospheric Administration (NOAA), the US Department of Energy (DOE), the National Institute for Environmental Studies (NIES), the Japan Meteorological Agency (JMA), the HIAPER Pole-to-Pole observations (HIPPO) program, and the GOSAT validation aircraft observation campaign over Japan. These comparisons demonstrate that the empirically derived bias correction improves the agreement between GOSAT XCO2/XCH4 and the aircraft data. Finally, we present spatial distributions and temporal variations of the derived GOSAT biases