ABSTRACT: BACKGROUND: Serum beta-cryptoxanthin levels are lower in overweight subjects than in normal subjects. Abnormalities of adipocytokine profiles in obesity subjects have been reported. There are several reports that serum beta-cryptoxanthin levels in them were relatively lower than normal subjects. OBJECTIVE: We hypothesize that supplementation of highly concentrated beta-cryptoxanthin improves serum adipocytokine profiles in obese subjects. This study tested the association between beta-cryptoxanthin intake and serum adipocytokine levels. METHODS: An intervention study consisted of a 3-week long before-and-after controlled trial, where beta-cryptoxanthin (4.7 mg/day) was given to 17 moderately obese postmenopausal women. RESULTS: The results indicated no significant changes in body weight or body mass index (BMI). Serum beta-cryptoxanthin levels increased significantly by 4-fold. Serum high molecular weight (HMW)-adiponectin levels increased significantly, while serum plasminogen activator inhibitor (PAI)-1 levels decreased. CONCLUSIONS: We concluded that increasing the intake of beta-cryptoxanthin to approximately 4 mg per day for 3 weeks may have beneficial effects on the serum adipocytokine status and consequently alleviate progression of metabolic syndrome.

Interleukin-6 (IL-6) is a principal proinflammatory cytokine inducing the acute phase response in various tissues, including liver. Here, we adopt the FD-LC-MS/MS method, consisting of fluorogenic derivatization (FD), separation by liquid chromatography (LC), and identification of proteins by LC-tandem mass spectrometry (MS/MS), to reveal how exposure to IL-6 alters temporally the intracellular secretory acute phase response (sAPR) proteins expressed in human hepatocytes as compared to non-exposure. Nine altered sAPR proteins were identified in cultures in response to IL-6. Seven of them (serum amyloid A protein, haptoglobin, fibrinogen α chain, fibrinogen β chain, fibrinogen γ chain, α(1)-acid glycoprotein and α(1)-antitrypsin) were significantly increased and two (β(2)-glycoprotein 1 and transferrin) were significantly decreased in response to IL-6. In addition, the transmission speed of transferrin might be much faster than the other sAPR proteins. These results suggest a different molecular mechanism for protein synthesis and the secretory pathway among the sAPR proteins. In this study, we observed the simultaneously and temporally altered expression of sAPR proteins which had been induced by exposure to IL-6 in human hepatocytes, in contrast to previous reports, in all of which the proteins were tested from the time they were secreted into the medium from the cells.

Innovative treatments of epileptic seizures are needed to improve the outcome of epilepsy. We studied the effect of pranlukast on seizure outcome in patients with intractable partial epilepsy. An open study was conducted to evaluate the clinical efficacy of 24-week pranlukast add-on therapy in 50 patients with intractable partial seizures. Serum concentrations of matrix metalloproteinase (MMP)-9 were determined using Biotrak Activity Assay System. Cytokines in cerebrospinal fluid (CSF) were measured by the BioPlex (BioRad) system and soluble TNF receptor1 (sTNFR1) in CSF was measured by the ELISA. Surface markers of lymphocytes in CSF were examined by cell-sorter. Seizure-free rate (SFR) was 13.6%, responder rate (RR) was 47.7%, and aggravation rate (AR) was 18.2% at the 13-24week period after starting pranlukast. In patients with increased serum MMP-9 before pranlukast therapy (baseline), comparison of paired serum levels showed a significant decrease after pranlukast therapy. Baseline CSF levels of IL-1β and IL-6 were elevated in patients compared with disease controls. Of four patients with paired data, three (including a responder to pranlukast) showed decreased pro-inflammatory cytokines (IL-1β, IL-6, and TNFα), and four showed decreased sTNFR1, after pranlukast treatment, and only a responder had markedly decreased frequency of CD8+ T cells in CSF. Pranlukast reduces seizure frequencies probably by pleiotropic effects including normalization of MMP-9 in sera, reduced leakage of pro-inflammatory cytokines into CNS, and inhibition of extravasation of leucocytes from brain capillaries. Further investigations by double-blind control study and animal models are warranted.

BACKGROUND: We aimed to identify a noninvasive predictor of portal venous pressure (PVP). METHODS: We directly measured the PVP in 40 consecutive patients who underwent direct percutaneous transhepatic portal vein puncture as part of the therapeutic management for liver diseases, and we evaluated the association of the PVP with noninvasive clinical parameters. The backgrounds of the liver were normal in 13 patients, chronic hepatitis in 17, and liver cirrhosis in ten. RESULTS: The mean PVP was 202 ± 114 mmH(2)O. In a multivariate linear regression analysis, the serum bile acid level and splenic volume showed independent positive correlations with the PVP (P < 0.001 and 0.002, respectively). The formula for estimating PVP was identified as follows: PVP (mmH(2)O) = serum bile acid (μmol/L) × 2.593 + splenic volume (cm(3)) × 0.416 + 65.929 (R (2) = 0.698). In a receiver operating characteristic (ROC) analysis, the AUC values of serum bile acid and splenic volume at a PVP of 200 mmH(2)O were 0.909 and 0.758, respectively. However, the AUC values of serum bile acid and splenic volume at a PVP of 250 mmH(2)O were 0.792 and 0.926, respectively, suggesting that the serum bile acid level and splenic volume are sensitive predictors of early and advanced portal hypertension, respectively. CONCLUSIONS: Combined measurements of the serum bile acid level and splenic volume may be useful to noninvasively assess the PVP prior to further invasive procedures.

NKG2D is a primary activating receptor that triggers cell-mediated cytotoxicity in NK cells against tumor and virus-infected cells. We previously identified the NKG2D haplotypes in the natural killer gene complex region on chromosome 12p: Two major haplotype alleles, LNK1 and HNK1, were closely related to low and high natural cytotoxic activity phenotypes, respectively. Furthermore, the haplotype of HNK1/HNK1 has revealed a decreased risk of cancer compared with LNK1/LNK1. In the present study, using flow cytometry, we evaluated the functional effects of NKG2D haplotypes and five htSNPs in terms of the cell-surface expression of NKG2D protein on NK and CD8 T cells of peripheral blood among 732 atomic-bomb survivors. NKG2D expression on NK cells showed significant increases, in the order of LNK1/LNK1, LNK1/HNK1 and HNK1/HNK1 haplotypes (p for trend =0.003), or with major homozygous, heterozygous, and minor homozygous genotypes for individual htSNPs (p for trend=0.02 ∼ 0.003). The same trend was observed for NKG2D expression on CD8 T cells. Our findings indicate that the NKG2D haplotypes are associated with the expression levels of NKG2D protein on NK and CD8 T cells, resulting in inter-individual variations in human cytotoxic response.

The exclusive reaction γp→K^{+}π^{-}Σ^{+} was measured for the first time using linearly polarized photons at beam energies from 1.85 to 2.96 GeV. Angular distributions in the rest frame of the K^{+}π^{-} system were fitted to extract spin-density matrix elements of the K^{*0} decay. The measured parity spin asymmetry shows that natural-parity exchange is dominant in this reaction. This result clearly indicates the need for t-channel exchange of the κ(800) scalar meson.

Nonsteroidal anti-inflammatory drugs (NSAIDs) are valuable agents; however, their use has been limited by their association with mucosal damage in the upper gastrointestinal tract. NSAIDs inhibit cyclooxygenase and consequently block the synthesis of prostaglandins, which have cytoprotective effects in gastric mucosa; these effects on prostaglandins have been thought to be major cause of NSAID-induced ulceration. However, studies indicate that additional NSAID-related mechanisms are involved in formation of gastric lesions. Here, we used a toxicoproteomic approach to understand cellular processes that are affected by NSAIDs in mouse stomach tissue during ulcer formation. We used fluorogenic derivatization-liquid chromatography-tandem mass spectrometry (FD-LC-MS/MS)-which consists of fluorogenic derivatization, separation and fluorescence detection by LC, and identification by LC-tandem mass spectrometry-in this proteomic analysis of pyrolic stomach from control and diclofenac (Dic)-treated mice. FD-LC-MS/MS results were highly sensitive; 10 differentially expressed proteins were identified, and all 10 were more highly expressed in Dic-treated mice than in control mice. Specifically, expression levels of 78kDa glucose-regulated protein (GRP78), heat shock protein beta-1 (HSP27), and gastrin were more than 3-fold higher in Dic-treated mice than in control mice. This study represents a first step to ascertain the precise actors of early NSAID-induced ulceration.

An individual in vitro culture system for bovine embryo needs to be developed for the study of embryo developmental competence. The objective of this study was to examine the effect of individual culture systems on the development of in vitro-matured-in vitro-fertilized bovine embryos. Two individual culture systems were compared. Cumulus-oocyte complexes (COC) were collected by aspiration of ovarian follicles (diameter, 2 to 5mm) obtained from a local abattoir. The COC were matured in TCM-199 supplemented with 5% calf serum and 0.02AUmL(-1) of FSH. Groups of 20 COC were incubated in 100-μL droplets of IVM media at 38.5°C under an atmosphere of 5% CO(2) for 20h. After 18h of gamete co-culture (3×10(6) spermmL(-1)), the presumptive zygotes were cultured in CR1aa medium supplemented with 5% calf serum at 38.5°C under an atmosphere of 5% CO(2), 5% O(2) and 90% N(2) for 9 days (fertilization=Day 0). The presumptive zygotes were randomly assigned to 1 of the following 3 treatments: single culture (1 zygote was cultured in a 5-μL droplet), well-of-the-well (WOW; Sugimura et al. 2010 Biol. Reprod. 83, 970-978) culture (25 zygotes were cultured individually in each 125-μL droplet) and control culture (25 zygotes were cultured in a 125-μL droplet). Embryo development was evaluated for cleavage and blastocyst rates, on Day 2 and Day 7 to 9 after IVF, respectively. The rates of development up to the blastocyst stage and total cell number in the blastocysts, determined by an air-drying method, were investigated. The cleavage and blastocyst rates were analysed by the chi-square test and the total cell numbers were analysed by ANOVA. The cleavage rates were significantly higher in the control and WOW groups than in the single-culture group (P<0.01) and the blastocyst rates were significantly lower in the single-culture group than in the control culture group (P<0.05; Table 1). The total cell numbers (mean±s.d.) of blastocysts did not significantly differ between the single culture (154.6±21.8), control culture (155.2±22.5) and WOW culture (159.8±27.0) groups. These results indicate that although the blastocyst rate was lower in the single-culture system than in the WOW or group culture system, in vitro-matured-in vitro-fertilized bovine embryos can be cultured using the single-culture system. Moreover, the quality of blastocysts developed by the single-culture system is the same as that of blastocysts developed using the other 2 culture systems.

More than 300000 embryos have been transferred all over the world (Stroud 2010 IETS Newsl. 27(4), 11-21). We have reported that embryos that showed the abnormal cleavage pattern at the first cell division can develop to the blastocyst stage (Somfai et al. 2010 J. Reprod. Dev. 56, 200-207). However, we have limited knowledge about the consequences of the pattern of first embryonic cleavage on their post-transfer developmental competence. The present study was conducted to determine the developmental competence of bovine blastocysts showing different cleavage patterns at their first cell division. Cumulus-oocyte complexes were collected by ovum pickup from Japanese Black cows and were subjected to in vitro maturation and IVF as reported previously (Imai et al. 2006 J. Reprod. Dev. 52, S19-S29 suppl). Inseminated oocytes were cultured in CR1aa medium supplemented with 5% calf serum covered by mineral oil at 38.5°C in 5% CO(2) in air with micro-droplets or 5% CO(2), 5% O(2) and 90% N(2). The kinetics of embryo development were analysed by time-lapse cinematography for 168h after IVF by using a Cultured Cell Monitoring System (CCM-M1.4ZS, Astec, Fukuoka, Japan). A total of 673 photographs of each embryo were taken (1 photograph in every 15min) during in vitro culture. Image stacks were analysed by the CCM-M1.4 software. Embryos were classified in 5 groups according to the pattern of first cleavage as normal cleavage (NC), direct cleavage from 1 cell to 3 to 4 blastomeres (3-4BL), unequal blastomeres (UB), multiple fragments (MF) and protrusion formation (PT). Blastocysts developing from each group were transferred into the ipsilateral uterine horn of each synchronized recipient on Day 7 or 8 after oestrus. Data on conception at Day 60, abortion and delivery were then recorded. Data were analysed by chi-square test and Student's t-test. In total, 43 embryos were transferred, 17 conceptions (39.5%) were established and 16 recipients (94.1%) were delivered. Only 1 abortion was detected at Day 223 in the NC group. The highest conception rate was observed in the NC group (55%, n=20) and the 3-4BL (n=12), UB (n=6) and PT (n=3) groups showed similar conception rates of 33.3%(1 implanted embryo belonged to 2 classes in UB and PT) and none of the embryos derived from the MF group (n=3) could cause conception. There was a significant difference (P<0.05) in conception rates between the NC group and totals of each of the other cleavage groups. No significant difference was found in gestation lengths and birth weights between the NC group (282.2±4.4 days, 30.6±3.8kg, respectively) and totals of each of the other cleavage groups (282.8±5.3 days, 30.3±1.9kg, respectively). These results indicate that embryos showing abnormal cleavage patterns at first cell division can develop to normal calves with normal gestation lengths and birth weights; however, their post-transfer viability is lower than for NC embryos.

In cattle, the prediction of embryonic viability after embryo transfer is an important research target. A previous study has indicated that the duration of the fourth cell cycle at the time of maternal-zygotic transition, which is involved in in vitro embryonic development, may be an indicator of blastocyst formation; this study showed that embryos with a short fourth cell cycle have a better potential of developing into blastocysts than those with a long fourth cell cycle (Lequarre et al. 2003 Biol. Reprod. 69, 1707-1713). However, the relationship between the fourth cell cycle duration and post-transfer viability of embryos is unclear. The aim of the present study was to examine the effect of the fourth cell cycle duration on embryo development after embryo transfer. Twenty-five IVF bovine embryos were cultured in well-of-the-well culture dishes contained 125 μ of CR1aa supplemented with 5% calf serum at 38.5°C in 5% O(2) and 5% CO(2) for 168h after insemination. In vitro development of the embryos was monitored using time-lapse cinematography (Sugimura et al. 2010 Biol. Reprod. 83, 970-978). We found that 61% of the blastocysts had a long fourth cell cycle (41.5±5.9h), which is commonly referred to as the lag phase, whereas the remaining embryos had a short fourth cell cycle (7.4±4.5h). All the embryos with a short fourth cell cycle exhibited a lag phase in the next cell cycle (32.9±6.6h). Moreover, embryos with a short fourth cell cycle were found to have a higher blastocyst rate (75.8%) than those with a long fourth cell cycle (48.1%; Student's t-test, P<0.01). However, embryonic cell number, apoptosis incidence, chromosomal abnormality and O(2) consumption were found to be identical between the 2 groups (Student's t-test, P>0.05). Real-time reverse-transcription PCR results of the individual blastocysts showed that the relative expression of 5 genes related to pregnancy reorganization, placentation and fetal growth-namely, CDX2, IFN-τ, PLAC8, AKR1B1 and IGF2R-did not differ between the 2 groups (Student's t-test, P>0.05). Furthermore, blastocysts derived from embryos with long (n=30) and short (n=19) fourth cell cycles were transferred into 49 recipient cows; we did not observe any difference between the long and short fourth cell cycles on the rates of pregnancy (long vs short fourth cell cycle, 30.0 vs 52.6%) and delivery (long vs short fourth cell cycle, 30.0 vs 47.4%; Yates' corrected chi-square test, P>0.10). These results show that blastocysts derived from embryos with either long or short fourth cell cycles have identical developmental competence after embryo transfer. Therefore, the fourth cell cycle duration during maternal-zygotic transition appears to be unavailable as the indicator of post-transfer viability of IVF bovine embryos.