Plasma prolactin axis shift from placental to pituitary origin in late prepartum mice.

The placenta secretes a prolactin (PRL)-like hormone PRL3B1 (placental lactogen II), a luteotropic hormone essential for maintaining pregnancy until labor in mice. A report from 1984 examined the secretion pattern of PRL3B1 in prepartum mice. In the current study, we found contradictory findings in the secretion pattern that invalidate the previous report. By measuring maternal plasma PRL3B1 and PRL every 4 hrs from gestational day 17 (G17), we newly discovered that maternal plasma PRL3B1 levels decrease rapidly in prepartum C57BL/6 mice. Interestingly, the onset of this decline coincided with the PRL surge at G18, demonstrating a plasma prolactin axis shift from placental to pituitary origin. We also found that maternal plasma progesterone regression precedes the onset of the PRL shift. The level of Prl3b1 mRNA was determined by RT-qPCR in the placenta and remained stable until parturition, implying that PRL3B1 peptide production or secretion was suppressed. We hypothesized that production of the PRL family, the 25 paralogous PRL proteins exclusively expressed in mice placenta, would decrease alongside PRL3B1 during this period. To investigate this hypothesis and to seek proteomic changes, we performed a shotgun proteome analysis of the placental tissue using data-independent acquisition mass spectrometry (DIA-MS). Up to 5,891 proteins were identified, including 17 PRL family members. Relative quantitative analysis between embryonic day 17 (E17) and E18 placentas showed no significant difference in the expression of PRL3B1 and most PRL family members except PRL7C1. These results suggest that PRL3B1 secretion from the placenta is suppressed at G18 (E18).

Prevention of Postoperative Cognitive Dysfunction by Minocycline in Elderly Patients after Total Knee Arthroplasty: A Randomized, Double-blind, Placebo-controlled Clinical Trial.

BACKGROUND: There are no effective pharmacologic interventions for preventing postoperative cognitive dysfunction in daily practice. Since the antibiotic minocycline is known to suppress postoperative neuroinflammation, this study hypothesized and investigated whether minocycline might have a preventive effect on postoperative cognitive dysfunction after noncardiac surgery. METHODS: This study included patients aged more than 60 yr undergoing total knee arthroplasty under general anesthesia. They were randomly assigned to minocycline and placebo groups, to orally receive 100 mg of minocycline or placebo twice daily from the day before surgery until the seventh day after surgery. Cognitive function was evaluated before surgery, and 1 week and 3 months after surgery, using a battery of four cognitive function tests, including Visual Verbal Learning Test, Trail Making Test, Stroop Color and Word Test, and Letter-Digit Coding Task. Additionally, 30 healthy volunteers were subjected to the same tests as the patients to examine the learning effect of repeated tests. The occurrence of postoperative cognitive dysfunction was judged from the results of the neurocognitive test battery, with consideration of the learning effect. The secondary endpoints were the effects of minocycline on postoperative delirium and postoperative pain. RESULTS: A total of 100 patients were randomized to the minocycline group, and 102 were randomized to the placebo group. The average age of patients was 75 yr. Evaluation showed no significant difference in the incidence of postoperative cognitive dysfunction between the minocycline and placebo groups at both 1 week (8 of 90 [8.9%] vs. 4 of 95 [4.2%]; odds ratio, 2.22 [95% CI, 0.64 to 7.65]; P = 0.240) and 3 months (15.3 of 90 [17.0%] vs. 15.3 of 95 [16.1%]; odds ratio, 1.07 [95% CI, 0.49 to 2.32]; P = 0.889) postoperatively. Missing data 3 months after surgery were corrected by the multiple imputation method. There were no differences between the two groups in postoperative delirium and postoperative pain. CONCLUSIONS: Minocycline is likely to have no preventive effect on postoperative cognitive dysfunction.

Surface Engineering of Top7 to Facilitate Structure Determination.

Top7 is a de novo designed protein whose amino acid sequence has no evolutional trace. Such a property makes Top7 a suitable scaffold for studying the pure nature of protein and protein engineering applications. To use Top7 as an engineering scaffold, we initially attempted structure determination and found that crystals of our construct, which lacked the terminal hexahistidine tag, showed weak diffraction in X-ray structure determination. Thus, we decided to introduce surface residue mutations to facilitate crystal structure determination. The resulting surface mutants, Top7sm1 and Top7sm2, crystallized easily and diffracted to the resolution around 1.7 Å. Despite the improved data, we could not finalize the structures due to high R values. Although we could not identify the origin of the high R values of the surface mutants, we found that all the structures shared common packing architecture with consecutive intermolecular ß-sheet formation aligned in one direction. Thus, we mutated the intermolecular interface to disrupt the intermolecular ß-sheet formation, expecting to form a new crystal packing. The resulting mutant, Top7sm2-I68R, formed new crystal packing interactions as intended and diffracted to the resolution of 1.4 Å. The surface mutations contributed to crystal packing and high resolution. We finalized the structure model with the R/Rfree values of 0.20/0.24. Top7sm2-I68R can be a useful model protein due to its convenient structure determination.

The influence of Javanese turmeric (Curcuma xanthorrhiza) on the pharmacokinetics of warfarin in rats with single and multiple-dose studies.

CONTEXT: Co-administration between warfarin (WF) and Curcuma xanthorrhiza Roxb. (Zingiberaceae) (CX) is found in Indonesian patients and need to be evaluated. OBJECTIVE: This study assesses the effect of concomitant administration of CX extract on the pharmacokinetics of WF in rats. MATERIALS AND METHODS: Wistar rats were divided into 4 groups (n = 6) and administered with 2% Pulvis Gummi Arabicum (PGA, control), fluconazole (FZ, 6 mg/kg), CX-1 (6 mg/kg) or CX-2 (18 mg/kg BW) for 7 days. For the single-dose study, at the 8th day, WF (1 mg/kg) was administered to all groups and blood samples were taken from 0.25 to 72 h. For the multiple-dose study, daily dose of WF was administered to all groups of rats and at the 7th to 9th day, the rats were treated with PGA, CX-1, CX-2 and FZ. Blood samples were withdrawn daily at 4 h after administration of WF from the 1st to 11th day. RESULTS: The area under the curve (AUC) of R- and S-WF in the CX-2 group was a significantly higher value compared to the control (77.54 vs. 35.27 mg.h/L for R-WF and 316.26 vs. 40.16 mg.h/L for S-WF; p < 0.05; Kruskal-Wallis method). The CX-2 administration also caused the increasing in the concentration level of R-WF (16%) and S-WF (27%) from the 7th to 9th day of administration. DISCUSSION AND CONCLUSIONS: The CX administration in a higher dose caused alteration on WF pharmacokinetics suggesting the need for clinical evaluation of the interaction between CX and WF.

Darunavir concentration in PBMCs may be a better indicator of drug exposure in HIV patients.

PURPOSE: The clinical efficacies of some antiretroviral drugs are known to not depend on its concentration in blood. To establish a method of dosage adjustment for darunavir (DRV) based on pharmacokinetic theory, we analyzed the correlation between DRV levels in peripheral blood mononuclear cells (PBMCs) and plasma. METHODS: The concentrations of DRV and ritonavir (RTV) in plasma and PBMCs of 31 samples obtained from 19 patients were analyzed. An in vitro kinetic study using MOLT-4 cells was performed to assess the contribution of RTV to the intracellular accumulation of DRV. RESULTS: DRV levels in PBMCs varied between 7.91 and 29.36 ng/106 cells (CV 37.5%), while those in plasma were greater. No significant correlation was found between the trough level of DRV in plasma and that in PBMCs (p = 0.575). The inter-day difference in DRV levels in PBMCs seemed smaller than that in plasma (- 41.6-23.0% vs - 83.3-109.1%). In the in vitro study, the elimination half-life of cellular efflux of DRV was 15.7 h in the absence of RTV and extended to 47.6 h in the presence of RTV. CONCLUSIONS: We found a poor correlation between intracellular DRV and plasma DRV levels in patients receiving highly active antiretroviral therapy. The efflux rate of DRV from cells was slow; therefore, the concentration of DRV in PBMCs may reflect average exposure to the drug and clinical efficacy.

Significant decreases in blood propofol concentrations during adrenalectomy for phaeochromocytoma.

AIM: The kinetics of propofol are influenced by cardiac output. The aim of this study was to examine changes in blood propofol concentrations during phaeochromocytoma surgery using target-controlled infusion (TCI) anaesthesia with propofol. METHODS: This is a prospective observational study. Ten patients with phaeochromocytoma who underwent unilateral adrenalectomy were included. Cardiac output was measured using an arterial pressure-based cardiac output analysis method. The target blood propofol concentrations were adjusted to maintain an approximate bispectral index (BIS) value of 40 before initiating surgery. The settings remained constant during surgery. Blood samples for propofol concentrations were collected from the radial artery at seven time points: two before tumour manipulation (T1, 2), two during tumour manipulation (T3, 4), and three after tumour vein ligation (T4-7). BIS values, the arterial pressure cardiac index (APCI) and haemodynamic parameters were measured at the same time points as the blood samples. The prop-ratio was calculated by dividing blood propofol concentrations by target concentrations of TCI. RESULTS: APCI increased during tumour manipulation and after tumour vein ligation. The prop-ratio was reduced significantly by approximately 40% and showed a significant negative correlation with APCI. BIS values increased significantly and showed a significant negative correlation with the prop-ratio. CONCLUSION: The increased APCI during tumour manipulation and after tumour vein ligation was associated with markedly reduced blood propofol concentrations. These results reveal that significant decreases in the anaesthetic effect may be observed in patients undergoing phaeochromocytoma surgery even if TCI anaesthesia is used with propofol.

Sex Differences in the Blood Concentration of Tacrolimus in Systemic Lupus Erythematosus and Rheumatoid Arthritis Patients with CYP3A5*3/*3.

The purpose of this study was to describe the impact of sex and cytochrome P450 3A5 (CYP3A5) variant on the blood concentration of tacrolimus in patients with systemic lupus erythematosus or rheumatoid arthritis. The blood concentration of tacrolimus (ng/mL) divided by the daily dose of tacrolimus (mg/day) and the patient's weight (kg) (C/D) was obtained from 55 patients. The C/D value was analysed according to genetic variation in CYP3A5 or ATP binding cassette subfamily B member 1 (ABCB1), sex, and age. The C/D value in the CYP3A5*3/*3 group was significantly higher than in the CYP3A5*1/*1 and *1/*3 groups (p < 0.05, effect size: d = 1.40). In the CYP3A5*3/*3 group, the concentration of tacrolimus was significantly higher in men than in women (p < 0.05, effect size: d = 1.78). Furthermore, in the CYP3A5*3/*3 group, the concentration of tacrolimus was significantly higher in women aged over 50 years than in women aged under 50 years (p < 0.05, effect size: d = 1.18). In contrast, ABCB1 genetic variations did not show any significant effect on the C/D value. Since the blood concentration of tacrolimus in patients with CYP3A5*3/*3 varies depending on sex and age, these factors should be considered when studying the difference of sex in CYP3A.

Dorsoventral asymmetry of photosynthesis and photoinhibition in flag leaves of two rice cultivars that differ in nitrogen response and leaf angle.

Rice is believed to show photosynthetic symmetry between adaxial and abaxial leaf sides. To verify this, we re-examined dorsoventral asymmetry in photosynthesis, chlorophyll fluorescence and anatomical traits in flag leaves of two Oryza sativa cultivars that differ in nitrogen (N) response and in leaf angle: 'Akenohoshi', a cultivar that can adapt to low-N (LN), with low leaf angle (more erect leaves), and 'Shirobeniya', a cultivar that is unable to adapt to LN, with higher leaf angle. Plants were grown under standard-N (SN) and LN conditions. LN leaves of both cultivars became more erect than SN, but LN Akenohoshi still had more erect ones than Shirobeniya. Contrary to results of previous studies, leaves of both cultivars showed an asymmetry in photosynthetic rate between adaxial and abaxial sides (higher on the adaxial side) under SN. SN leaves of both cultivars showed lower susceptibility to photoinhibition on the adaxial side than on the abaxial side. However, leaves of Akenohoshi showed less asymmetry in these traits under LN than under SN, whereas leaves of Shirobeniya had similar degrees of asymmetry in these traits under both SN and LN. Both cultivars also showed dorsoventral asymmetry in anatomical traits of mesophyll tissue regardless of N level, but the degree of asymmetry was lower in LN Akenohoshi. These data reveal that rice leaves exhibit dorsoventral asymmetry in photosynthetic and anatomical features, and that the degree of asymmetry varies with cultivar and N level. It is suggested that lower leaf angles (particularly in Akenohoshi) in the presence of LN represent a light acclimation to prevent photoinhibition.

Quantitative evaluation of biliary elimination of gadoxetate, a magnetic resonance imaging contrast agent, via geometrical isomer-specific transporting system in rats.

Gadoxetate, a magnetic resonance imaging contrast agent, is eliminated into bile. Gadoxetate geometrical isomers are chromatographically classified into two groups by differences between their ionic states (GIs-I and GIs-II; 65:35 w/w); however, the elimination mechanism of each isomer in vivo remains controversial. Thus, the contribution of carrier-mediated transport systems on the biliary elimination of gadoxetate was examined. Gadoxetate was injected intravenously into rats, and the time courses of the plasma concentrations and biliary elimination of GIs-I and GIs-II were examined by high-performance liquid chromatography techniques. The results showed that 34.7% of GIs-I (GIs-I(s); 22.6% of gadoxetate) was quickly eliminated into bile within 30 min after injection. The contents of the residual GIs-I (GIs-I(r)) and GIs-II in plasma similarly decreased according to a first-order elimination process (t1/2=23-27 min), and 64.0% of GIs-I(r) and GIs-II (49.6% of gadoxetate) was eliminated into the bile within 2 h after injection. There was no significant difference between the elimination half-lives of GIs-I(r) and GIs-II in rats. In conclusion, the geometrical isomer with specific conformation corresponding to 22.6% of gadoxetate was eliminated into bile in rats via a carrier-mediated transport system no later than 30 min after intravenous injection.

Blood Concentration of Cabazitaxel in a Patient Whose General Condition Worsened with Concomitant Use of Clarithromycin.

We encountered a case in which the general condition of a patient receiving cabazitaxel worsened with concomitant use of clarithromycin. Cabazitaxel is metabolized mainly by CYP3A4, and the frequency of adverse events is known to increase with increasing exposure. Although these drugs are not often quantified in daily practice, we quantified them because we considered it possible that the blood concentration of cabazitaxel had increased due to CYP3A4 inhibition of clarithromycin and that cabazitaxel-related adverse events had occurred. However, the concentration of cabazitaxel was not increased and we attributed the patient's deterioration to decreased tolerability of cabazitaxel. At least at a trough concentration of 70 ng/mL, which is the trough concentration when a normal dose of clarithromycin is administered, clarithromycin does not appear to have a significant effect on the blood concentration of cabazitaxel. This case suggests that the administration of the normal dose of clarithromycin might be relatively safe in patients receiving cabazitaxel.

Increased c­SRC expression is involved in acquired resistance to lenvatinib in hepatocellular carcinoma.

Lenvatinib, a multi-kinase inhibitor, serves a crucial role in the treatment of unresectable hepatocellular carcinoma (HCC). However, >50% of patients receiving lenvatinib therapy experience tumor growth or metastasis within 1 year, highlighting the need to address acquired resistance as a critical clinical challenge. To elucidate the factors associated with acquired resistance to lenvatinib, a lenvatinib-resistant HCC cell line (JHH-7_LR) was established by exposing a lenvatinib-sensitive HCC cell line, JHH-7, to lenvatinib. The changes in protein expression associated with the development of resistance were analyzed using a proteomic approach, detecting 1,321 proteins and significant changes in the expression of 267 proteins. Using Ingenuity Pathway Analysis bioinformatics software, it was revealed that the activity of multiple signaling pathways varied alongside the changes in expression of these proteins, and c-SRC was identified as a protein involved in a number of these signaling pathways, with its activity varying markedly upon the acquisition of resistance. When co-administering dasatinib, a c-SRC inhibitor, the partial restoration of lenvatinib sensitivity in the JHH-7_LR cell line was observed. The present study demonstrated that increased c-SRC expression was partially associated with HCC resistance to lenvatinib, suggesting that c-SRC inhibition could reduce the resistance of HCC to lenvatinib.

Target concentration achievement for efficacy and safety of patients with osteosarcoma treated with high-dose methotrexate based on individual pharmacokinetics: A retrospective study.

In the high-dose methotrexate (HD-MTX) treatment of patients with osteosarcoma, a dose-adjustment method using individual pharmacokinetic parameters (PK method) to optimize the concentration was developed in 2010. However, to the best of our knowledge, the clinical usefulness of the PK method has not been verified until now. In the present retrospective study, to assess the usefulness of the PK method, the achievement rate of an effective and safe concentration range was evaluated. A total of 43 patients with osteosarcoma who were administered HD-MTX therapy (43 first courses and 200 subsequent courses) were enrolled. The MTX dose in the first course was determined using a common method based on body surface area (BSA method); a total of 8-12 g/m2 was administered as an initial dose for 1 h and a maintenance dose for 5 h. In the subsequent courses, loading and maintenance doses were calculated by the PK method based on the serum MTX concentration profile of the previous course. The effective target concentration during 1-6 h after the start of MTX administration was 700-1,000 µmol/l, whereas the target safe MTX level was less than 10, 1 and 0.1 µmol/l at 24, 48 and 72 h, respectively. Notably, the rate of achieving the effective target concentration was significantly higher when using the PK method as compared to that when using the BSA method. The achievement rate of the safe target concentration at 24, 48 and 72 h when using the PK method was significantly higher. Additionally, the incidence of abnormal laboratory values of aspartate aminotransferase and alanine aminotransferase was significantly lower when using the PK method. Therefore, the PK method was suggested to be very useful in HD-MTX therapy for patients with osteosarcoma.

Risk of exposure of patients' family members to lenalidomide at home.

OBJECTIVES: Lenalidomide, a hazardous drug, has strict distribution controls. However, the risk of contamination with lenalidomide when patients take the drug has not been studied and the risk of drug exposure to people in the patient's living environment is unknown. Thus, we investigated the amount of lenalidomide that could be dispersed during the period between removal of the capsule and returning the used blister packages, and we considered the conditions under which lenalidomide could be dispersed and countermeasures. METHODS: The amount of lenalidomide contamination was measured on the outside of the unused blister packages returned by the patients, on the surface of the capsule, and on the inside of the package immediately after removal of the capsule. In addition, the amount of contamination was measured on the blister packages used by the patients and on the gloves worn by the pharmacists on receipt of the packages. Lenalidomide was analysed by liquid chromatography-tandem mass spectrometry. RESULTS: Lenalidomide amounts on the outside of the unused blister packages returned by the three patients were <10, <10, and 26.8 ng/pack, those on the capsule surface immediately after removal from the packages were 297, 388, and 297 ng/capsule, and those on the inside of packages immediately after removal of all capsules were 143, 184, and 554 ng/pack, respectively. A median of 15.6 ng/pack lenalidomide was detected on the surface of packages used by the patients (n=18). The lenalidomide remaining in the packages immediately after capsule removal (~200 ng/pack), except for the 15.6 ng/pack detected in the packages used by the patients, may have been dispersed in the patient's living environment (~90% or more). The maximum amount of lenalidomide on the surface of the packages used by the patients was over 2500 ng/pack. CONCLUSIONS: The amount of lenalidomide contamination per package was found to be at least 100 ng less after collection by the pharmacist than immediately after removal of the capsules. Therefore, it is recommended to clean the surrounding area and wash one's hands after taking the capsules.

Predicting cetuximab accumulation in KRAS wild-type and KRAS mutant colorectal cancer using 64Cu-labeled cetuximab positron emission tomography.

Overexpression of epidermal growth factor receptor (EGFR) is common in colorectal cancer. However, cetuximab as an EGFR-targeting drug is useful only for a subset of patients and currently no single predictor other than V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS) mutation status has been established. In the present study, we investigated cetuximab accumulation in colorectal tumors and major organs using (111)In-DOTA-cetuximab. We also evaluated the potential of positron emission tomography (PET) imaging of (64)Cu-DOTA-cetuximab. Colorectal tumor xenografts with a different EGFR expression level and KRAS mutation status were subjected to in vivo biodistribution study and PET imaging at 48 h post-injection of radiolabeled cetuximab. The EGFR expression levels on colorectal tumors were determined by ex vivo immunoblotting and ELISA. We found that KRAS wild-type tumors had significantly higher (111)In-DOTA-cetuximab accumulation than KRAS mutant tumors (P < 0.001). Based on KRAS mutation status, a strong correlation was found between (111)In-DOTA-cetuximab tumor uptake and EGFR expression level (KRAS wild type: r = 0.988; KRAS mutant: r = 0.829), and between (64)Cu-DOTA-cetuximab tumor uptake with EGFR expression level (KRAS wild type: r = 0.838; KRAS mutant: r = 0.927). Significant correlation was also found between tumor uptake of (111)In-DOTA-cetuximab and (64)Cu-DOTA-cetuximab (r = 0.920). PET imaging with (64)Cu-DOTA-cetuximab allowed clear visualization of tumors. Both radiolabeled cetuximab had effectively visualized cetuximab accumulation in colorectal tumors with a wide variety of EGFR expression levels and different KRAS mutation status as commonly encountered in the clinical setting. Our findings suggest that this radioimmunoimaging therefore can be clinically translated as an in vivo tool to predict cetuximab accumulation in colorectal cancer patients prior to cetuximab therapy.

Clinicopathological features of lung adenocarcinoma with KRAS mutations.

BACKGROUND: KRAS and epidermal growth factor receptor (EGFR) mutations are thought to play an important role in the carcinogenesis of lung adenocarcinoma. However, clinicopathological findings of KRAS mutated adenocarcinoma cases have not yet been fully clarified. The authors analyzed the relationship between the KRAS mutation and corresponding clinicopathological findings, focusing on nonmucinous and mucinous bronchioloalveolar elements. METHODS: EGFR and KRAS mutations were detected in DNA samples extracted from 182 surgically resected tissues of lung adenocarcinomas by the Smart Amplification Process. The relations between gene mutation status and clinicopathological features were analyzed. All adenocarcinoma cases were divided into bronchioloalveolar carcinoma (BAC), adenocarcinoma with bronchioloalveolar features, and adenocarcinoma without BAC components (non-BAC). BAC/adenocarcinoma with bronchioloalveolar features tumors were further assessed for the presence of mucinous features. RESULTS: EGFR and KRAS mutations were found in 76 and 30 cases, respectively. In the KRAS mutant group, BAC/adenocarcinoma with bronchioloalveolar features was found in 22 cases, which included 10 nonmucinous and 12 mucinous tumors. Of 19 cases with mucinous BAC/adenocarcinoma with bronchioloalveolar features, KRAS mutations were detected in 12, but no EGFR mutation was detected. In the KRAS mutant group, BAC/adenocarcinoma with bronchioloalveolar features had significantly earlier pathological stages and more favorable prognoses than did non-BAC. Mucinous BAC/adenocarcinoma with bronchioloalveolar features showed less smoking history than did nonmucinous BAC/adenocarcinoma with bronchioloalveolar features and non-BAC. Furthermore, transversion type KRAS mutations were more common in non-BAC. CONCLUSIONS: KRAS mutated adenocarcinomas can be divided into BAC/adenocarcinoma with bronchioloalveolar features and non-BAC types. Non-BAC adenocarcinoma is related to smoking history and has a poor prognosis. BAC/adenocarcinoma with bronchioloalveolar features adenocarcinoma, however, has a more favorable prognosis, and mucinous BAC/adenocarcinoma with bronchioloalveolar features has little relationship to smoking history.

Ammonia emission from rice leaves in relation to photorespiration and genotypic differences in glutamine synthetase activity.

BACKGROUND AND AIMS: Rice (Oryza sativa) plants lose significant amounts of volatile NH(3) from their leaves, but it has not been shown that this is a consequence of photorespiration. Involvement of photorespiration in NH(3) emission and the role of glutamine synthetase (GS) on NH(3) recycling were investigated using two rice cultivars with different GS activities. METHODS: NH(3) emission (AER), and gross photosynthesis (P(G)), transpiration (Tr) and stomatal conductance (g(S)) were measured on leaves of 'Akenohoshi', a cultivar with high GS activity, and 'Kasalath', a cultivar with low GS activity, under different light intensities (200, 500 and 1000 µmol m(-2) s(-1)), leaf temperatures (27·5, 32·5 and 37·5 °C) and atmospheric O(2) concentrations ([O(2)]: 2, 21 and 40 %, corresponding to 20, 210 and 400 mmol mol(-1)). KEY RESULTS: An increase in [O(2)] increased AER in the two cultivars, accompanied by a decrease in P(G) due to enhanced photorespiration, but did not greatly influence Tr and g(S). There were significant positive correlations between AER and photorespiration in both cultivars. Increasing light intensity increased AER, P(G), Tr and g(S) in both cultivars, whereas increasing leaf temperature increased AER and Tr but slightly decreased P(G) and g(S). 'Kasalath' (low GS activity) showed higher AER than 'Akenohoshi' (high GS activity) at high light intensity, leaf temperature and [O(2)]. CONCLUSIONS: Our results demonstrate that photorespiration is strongly involved in NH(3) emission by rice leaves and suggest that differences in AER between cultivars result from their different GS activities, which would result in different capacities for reassimilation of photorespiratory NH(3). The results also suggest that NH(3) emission in rice leaves is not directly controlled by transpiration and stomatal conductance.

Endosperm cell size reduction caused by osmotic adjustment during nighttime warming in rice.

High night temperature (HNT) often reduces yield in field crops. In rice, HNT during the ripening stage diminishes endosperm cell size, resulting in a considerable reduction in final kernel weight; however, little is known about the underlying mechanisms at cell level. In this study, we performed picolitre pressure-probe-electrospray-ionization mass spectrometry to directly determine metabolites in growing inner endosperm cells of intact seeds produced under HNT conditions, combining with 13C feeding and water status measurements including in situ turgor assay. Microscopic observation in the inner zone suggested that approximately 24.2% of decrease in cell expansion rate occurred under HNT at early ripening stage, leading to a reduction in cell volume. It has been shown that HNT-treated plants were subjected to mild shoot water deficit at night and endosperm cell turgor was sustained by a decline in osmotic potential. Cell metabolomics also suggests that active solute accumulation was caused by a partial inhibition of wall and starch biosynthesis under HNT conditions. Because metabolites were detected in the single cells, it is concluded that a partial arrest of cell expansion observed in the inner endosperms was caused by osmotic adjustment at mild water deficit during HNT conditions.

Fast and Inexpensive Phenotyping and Genotyping Methods for Evaluation of Barley Mutant Population.

To further develop barley breeding and genetics, more information on gene functions based on the analysis of the mutants of each gene is needed. However, the mutant resources are not as well developed as the model plants, such as Arabidopsis and rice. Although genome editing techniques have been able to generate mutants, it is not yet an effective method as it can only be used to transform a limited number of cultivars. Here, we developed a mutant population using 'Mannenboshi', which produces good quality grains with high yields but is susceptible to disease, to establish a Targeting Induced Local Lesions IN Genomes (TILLING) system that can isolate mutants in a high-throughput manner. To evaluate the availability of the prepared 8043 M3 lines, we investigated the frequency of mutant occurrence using a rapid, visually detectable waxy phenotype as an indicator. Four mutants were isolated and single nucleotide polymorphisms (SNPs) were identified in the Waxy gene as novel alleles. It was confirmed that the mutations could be easily detected using the mismatch endonuclease CELI, revealing that a sufficient number of mutants could be rapidly isolated from our TILLING population.

FLEND (nelarabine, fludarabine, and etoposide) for relapsed T-cell acute lymphoblastic leukemia in children: a report from Japan Children's Cancer Group.

Nelarabine is a key drug for T-cell acute lymphoblastic leukemia (T-ALL). Fludarabine and etoposide might have synergistic effect with nelarabine by inhibiting ribonucleotide reductase and by preparing cell cycle for G1/S phase, respectively. We had started phase 1/2 multicenter clinical trial of combination chemotherapy consisted of nelarabine, fludarabine, and etoposide (FLEND therapy) for children with relapsed/refractory T-ALL which has been conducted since October 2011. Although we could not complete this trial because of recruitment difficulties, we treated five children with first-relapsed T-ALL which were enrolled in the phase 1 dose escalation study of fludarabine and etoposide with nelarabine. No dose-limiting toxicity occurred, and frequent grade 3-4 toxicity was hematological toxicity and febrile neutropenia, as expected. There was no neurotoxicity. All 2 patients who received the target dose of FLEND, in which nelarabine (650 mg/m2), fludarabine (30 mg/m2), and etoposide (100 mg/m2) were administered for 5 consecutive days, were induced to complete remission. We concluded that FLEND might be safe and one of the promising combination chemotherapies to relapsed/refractory T-ALL.

L-Lactate dehydrogenase B may be a predictive marker for sensitivity to anti-EGFR monoclonal antibodies in colorectal cancer cell lines.

Recently, proteins derived from cancer cells have been widely investigated as biomarkers for predicting the efficacy of chemotherapy. In this study, to identify a sensitive biomarker for the efficacy of anti-epidermal growth factor receptor monoclonal antibodies (anti-EGFR mAbs), proteins derived from 6 colorectal cancer (CRC) cell lines with different sensitivities to cetuximab, an anti-EGFR mAb, were analyzed. Cytoplasmic and membrane proteins extracted from each CRC cell line were digested using trypsin and analyzed comprehensively using mass spectrometry. As a result, 148 and 146 peaks from cytoplasmic proteins and 363 and 267 peaks from membrane proteins were extracted as specific peaks for cetuximab-resistant and -sensitive CRC cell lines, respectively. By analyzing the proteins identified from the peptide peaks, cytoplasmic L-lactate dehydrogenase B (LDHB) was detected as a marker of cetuximab sensitivity, and it was confirmed that LDHB expression was increased in cetuximab-resistant CRC cell lines. Furthermore, LDHB expression levels were significantly upregulated with the acquisition of resistance to cetuximab in cetuximab-sensitive CRC cell lines. In conclusion, LDHB was identified as an important factor affecting cetuximab sensitivity using comprehensive proteome analysis for the first time.