Search

Raw egg white undergoes sol-gel transition by heat treatment, which changes it to an elastic gel. Here, protease treatment to render a new texture to heated egg white gel was applied. Protease-treated gels exhibited ductile flow without obvious rupture points. Transmission electron microscopy analysis showed that in protease-treated gels, protein aggregates were distributed more homogeneously compared with that observed in the untreated control, probably because ovalbumin was digested into small peptides as revealed by SDS-PAGE. The properties of the gel were evaluated by sensory tests and by measuring the movement of the masseter muscle, using surface electromyography. Results showed that maximum bite force and mastication duration were decreased for the protease-treated gels, which were evaluated as being softer, smoother, less elastic and better textured. Overall, our results indicate that protease-treated egg white gel has superior qualities and is easier to swallow than the untreated gel., Practical Applications: In the food industry, the use of egg white is limited compared with that of egg yolk and whole eggs. In this study, we performed protease treatment to generate a new food material with smoother and softer texture compared with heat treated egg white. Our findings may expand the consumption of egg white, which can be consumed by people with mastication and swallowing disorders, and reduce the waste of egg white as a surplus product.

The mucosal graft and flaps method (MG method) is a palatoplasty technique that was developed for the purpose of improving maxillary growth in patients with cleft palate. In the MG method, full-thickness buccal mucosa is grafted onto the raw surface created by pushback palatoplasty. The method is unlikely to result in severe scarring and has a favorable effect on maxillary growth. In addition, it is unlikely to result in oronasal fistula and provides good speech results. Overall, postive long-term treatment results have been obtained. Although the MG method is technically difficult and requires a lengthy surgery, the technique is considered to be effective for palate closure in terms of speech and maxillary growth.

Aim: Sorafenib is the standard systemic therapy for patients with advanced hepatocellular carcinoma (HCC). We aimed to assess the efficacy and safety of sorafenib therapy in very elderly patients aged 80 years and older with advanced HCC., Methods: In a retrospective multicenter study in Japan, we reviewed 185 patients (median age, 71 years; 82% male; 95% Child-Pugh class A) with advanced HCC who received sorafenib therapy. Data were compared between 24 (13%) patients aged 80 years and older and 161 (87%) patients aged less than 80 years. We used propensity score matching to adjust for differences between the two groups., Results: Median overall survival was 10.6 months in all patients: 11.7 months in patients aged 80 years and older and 10.5 months in those aged less than 80 years. There were no significant differences in overall survival, tumor response, and frequency and severity of drug-related adverse events between patients aged 80 years and older and those aged less than 80 years in both the entire study cohort and the propensity-matched cohort., Conclusion: Sorafenib may be effective and well tolerated, even in patients with advanced HCC who are aged 80 years and older, as well as those aged less than 80 years., (© 2014 The Japan Society of Hepatology.)

Bitterness-masking compounds were identified in a natural white mold cheese. The oily fraction of the cheese was extracted and further fractionated by using silica gel column chromatography. The four fractions obtained were characterized by thin-layer chromatography and nuclear magnetic resonance spectroscopy. The fatty acid-containing fraction was found to have the highest bitterness-masking activity against quinine hydrochloride. Bitterness-masking activity was quantitated using a method based on subjective equivalents. At 0.5 mM, the fatty acid mixture, which had a composition similar to that of cheese, suppressed the bitterness of 0.008% quinine hydrochloride to be equivalent to that of 0.0049-0.0060% and 0.5 mM oleic acid to that of 0.0032-0.0038% solution. The binding potential between oleic acid and the bitter compounds was estimated by isothermal titration calorimetry. These results suggest that oleic acid masked bitterness by forming a complex with the bitter compounds.

It remains to be clarified whether adoptive cellular therapy (ACT) in patients with advanced cancer, in whom strong immunosuppression and immune-escape mechanisms are established, has the potential to alter cytokine secretion from blood cells and affect the number of regulatory T cells (Tregs). In this study, the secretion of cytokines from peripheral blood cells and the number of peripheral blood Tregs were analyzed before and after ACT. Blood samples were collected from 109 consecutive cancer patients who received ACT, which consisted of anti-CD3 stimulated lymphokine-activated killer cells. For testing immune function, venous blood was obtained from patients before the start of therapy and after they had received 4 cycles of ACT. Of the 109 patients, 76 received ACT four times or more. All 109 blood samples at baseline and 76 follow-up samples were available. The secretion ability of various cytokines from peripheral blood cells was measured, as well as the number of peripheral blood Tregs. We found that the secretion ability of interferon (IFN)-γ and tumor necrosis factor (TNF)-α was enhanced significantly after treatment, while the number of Tregs and the ratio of Treg to CD4 was significantly decreased. Overall survival in patients with increased IFN-γ and TNF-α secretion after ACT was significantly longer. These findings suggest a potential therapeutic role for ACT in cancer immunotherapy.

We examined the effects of adoptive T-cell transfer (ACT) on the population of regulatory T cells (Tregs) in a mouse colorectal cancer transplant model. In an in vivo study, Treg populations in Balb/c mice colon26 transplant model after ACT were analyzed in peripheral blood, local lymph node, and tumor. In an in vitro study CD4+ cells were cultured in medium containing TGF-beta to induce Tregs. LAK cells were added or not in this Treg induction system. Treg induction after coculture with LAK was investigated. We also studied the role of IFN-gamma in the mechanism of Treg induction. Tregs in the draining lymph nodes and tumor were significantly suppressed by ACT. The induction of Tregs in vitro was inhibited by coculture with LAK cells. Furthermore, Tregs in the cultured cells were significantly inhibited by addition of exogenous IFN-gamma. Moreover, Tregs were increased by addition of IFN-gamma mAb. ACT may decrease Tregs in tumor-bearing hosts. One of the mechanisms is considered to be IFN-gamma inhibiting the induction of Tregs.

We cloned three novel papain-type cysteine proteases (CPs), triticain alpha, beta and gamma, from 1-d-germinating wheat seeds. Triticain alpha, beta and gamma were constituted with 461, 472 and 365 amino acid residues, respectively, and had Cys-His-Asn catalytic triads as well as signal and propeptide sequences. Triticain gamma contained a putative vacuole-sorting sequence. Phylogenetic analysis showed that these CPs were divided into mutually different clusters. Triticain alpha and gamma mRNAs were expressed in seeds at an early stage of maturation and at the stage of germination 2d after imbibition, while triticain beta mRNA appeared shortly after imbibition. The expression of mRNAs for triticain alpha and gamma was suppressed by uniconazol, a gibberellin synthesis inhibitor. All the three CP mRNAs were strongly expressed in both embryo and aleurone layers. These results suggest that triticain alpha, beta and gamma play differential roles in seed maturation as well as in digestion of storage proteins during germination.

An Aspergillus saitoi protease preparation, Molsin, was found to contain beta-glucosidase as well as protease activities. Application of Molsin to soybean curd improved its functionality by converting the contained isoflavone glycosides to their aglycones through beta-glucosidase, and also modified the rheological property into a creamy consistency through protease. The enzymatically modified soybean curd was characterized by a ductility flow having no particular rupture point.

We cloned a new cysteine proteinase of wheat seed origin, which hydrolyzed the storage protein gliadin almost specifically, and was named gliadain. Gliadain mRNA was expressed 1 day after the start of seed imbibition, and showed a gradual increase thereafter. Gliadain expression was suppressed when uniconazol, a gibberellin synthesis inhibitor, was added to germinating seeds. Histochemical detection with anti-gliadain serum indicated that gliadain was present in the aleurone layer and also that its expression intensity increased in sites nearer the embryo. The enzymological characteristics of gliadain were investigated using recombinant glutathione S-transferase (GST)-progliadain fusion protein produced in Escherichia coli. The GST-progliadain almost specifically digested gliadin into low molecular mass peptides. These results indicate that gliadain is produced via gibberellin-mediated gene activation in aleurone cells and secreted into the endosperm to digest its storage proteins. Enzymologically, the GST-progliadain hydrolyzed benzyloxycarbonyl-Phe-Arg-7-amino-4-methylcoumarin (Z-Phe-Arg-NH(2)-Mec) at K(m) = 9.5 microm, which is equivalent to the K(m) value for hydrolysis of this substrate by cathepsin L. Hydrolysis was inhibited by two wheat cystatins, WC1 and WC4, with IC(50) values of 1.7 x 10(-8) and 5.0 x 10(-8) m, respectively. These values are comparable with those found for GST-progliadain inhibition by E-64 and egg-white cystatin, and are consistent with the possibility that, in germinating wheat seeds, gliadain is under the control of intrinsic cystatins.

Two aspartic proteinase (AP) cDNA clones, WAP1 and WAP2, were obtained from wheat seeds. Proteins encoded by these clones shared 61% amino acid sequence identity. RNA blotting analysis showed that WAP1 and WAP2 were expressed in both germinating and maturing seeds. The level of WAP2 mRNA expression was clearly weaker than that of WAP1 in all tissues of seeds during germination and maturation. APs purified from germinating seeds were enzymatically active and digested the wheat storage protein, gluten. To elucidate the physiological functions of WAP1 and WAP2 in seeds, we investigated the localisation of WAP1 and WAP2 by in situ hybridisation. In germinating seeds investigated 24h after imbibition, both WAP1 and WAP2 were expressed in embryos, especially in radicles and shoots, scutellum, and the aleurone layer. In maturing seeds, WAP1 was expressed in the whole embryo, with slightly stronger expression in radicles and shoots. WAP1 was also expressed in the aleurone layer 3 weeks after flowering. Strong signals of WAP1 mRNA were detected in the whole embryo and aleurone layer 6 weeks after flowering. On the other hand, WAP2 was scarcely detected in seeds 3 weeks after flowering, and thereafter weak signals began to appear in the whole embryo. WAP1 and WAP2 were expressed widely in germinating and maturing seeds. Such diversity in site- and stage-specific expression of the two enzymes suggests their differential functions in wheat seeds.

Many plant aspartic proteinases (APs) are different from animal and microbial APs in that they contain a polypeptide insert, approximately 100 amino acids in length, in the C-terminal region. To interpret the significance of this insert, we constructed an expression system for rice AP oryzasin 1 by linking a pro-oryzasin 1 downstream of glutathione S-transferase (GST). GST-proOS1 expressed the highest degree of hemoglobin-hydrolytic activity when treated at pH 3.3 and incubated for 24 h at room temperature. We carried out a similar experiment using an insert-lacking proOS1 mutant, GST-DeltaproOS1, as the fusion protein, and found it to show similar activity. This result indicates that the insert is not involved in the production of AP activity. We then investigated the autolysis of the two proteins by Western blot analysis. GST-proOS1 was autolyzed into 67- and 64-kDa fragments, while GST-DeltaproOS1 autolyzed to 54- and 52-kDa products. GST-DeltaproOS1 clearly produced two molecular species early in the autolytic process, and not later than 3 h from the start, but no such clear result was observed in the case of GST-proOS1. This suggests that, although the presence of the plant AP-specific insert does not influence the enzyme activity by itself, it apparently has an effect on the autolysis of OS1.

Matrix metalloproteinase-II (MMP-II, 72-kd type IV collagenase, or gelatinase) is one of the gene families of zinc enzymes capable of degrading extracellular matrix molecules, and specifically of degrading type IV and V collagens, gelatin, fibronectin, and elastin. In this study, we used both the liver fibrosis model and the reversibility model of experimental cirrhosis to clarify how MMP-II participates in liver fibrosis of rats. To produce fibrosis model, rats received subcutaneous injections of CCl4 twice weekly for 7, 9, or 14 weeks. For the reversibility model, rats were treated with CCl4 three times a week for 8 weeks and killed at 3, 7, 14, 28, or 42 days after discontinuation of treatment. MMP-II gene expression was studied by Northern hybridization technique, and gelatinase activity of MMP-II was examined by zymography using gelatin substrate. At the same time, an immunohistochemical study using anti-type IV collagen antibody was carried out. In liver fibrosis model, nodule formation was established at 14 weeks. Immunodeposit of type IV collagen was increased in wide fibrous septa and was clearly observed along sinusoidal wall. Gene expression of MMP-II increased up to 7 to 12 times compared with that of controls, with the expression rate being maximum at an intermediate stage of fibrosis. Zymography showed the expressions of both 65-kd latent MMP-II, which is confirmed to be activated by adding p-aminophenylmercuric acetate, and 62-kd active MMP-II during fibrosis. The expression of both forms increased 13 to 28 times as the fibrosis progressed.(ABSTRACT TRUNCATED AT 250 WORDS)

Three cDNAs coding for monkey cytochrome P-450 (P450) 2C, 2E and 3A (MKmp13, MKj1 and MKnf2, respectively) were isolated from a lambda gt11 cDNA library of a liver from a 3-methylcholanthrene (3MC)-treated crab-eating monkey, using cDNA fragments for human P450 2C, 2E and 3A as respective probes. MKmp13 and MKnf2 were 1901 and 2032 bp long, containing entire coding regions for polypeptides of 490 and 503 residues, respectively. The deduced N-terminal amino acid sequences of MKmp13 and MKnf2 were identical with those of P450-MK1 and P450-MK2, which had been purified from liver microsomes of untreated and polychlorinated biphenyl (PCB)-treated crab-eating monkeys, respectively. MKj1 was 1508 bp long, encoding a polypeptide of 449 residues, which is presumed to lack N-terminal 45 residues as compared with the sequence for human P450 2E1. Northern blot analysis indicated that monkey P450 2C, 2E and 3A mRNAs were expressed constitutively in monkey livers. P450 2E and 3A mRNAs were induced by both 3MC and PCB, while P450 2C mRNA was induced only by PCB. The deduced amino acid sequences of four monkey cytochrome P-450 cDNAs, including P450 1A1 (MKah1) which we isolated previously, were more than 92% identical with those of corresponding human cytochrome P-450 cDNAs.

Clinical studies show that patients with liver cirrhosis associated with portal hypertension have a high incidence of duodenal ulcer and duodenitis. However, little information is available concerning pathophysiological process of such duodenal diseases in liver cirrhosis. Hemodynamics of the duodenal mucosa was studied in cirrhotics with esophageal varices (68 cases) and in noncirrhotics with non-ulcer dyspepsia (37 cases) as well. In each group, hemoglobin concentration in the peripheral venous blood was measured, and mucosal hemodynamics was examined in 4 regions of the duodenum by endoscopic reflectance spectrophotometer. No significant intergroup difference was noted in the mean age or sex ratio. Hemoglobin concentration in the peripheral venous blood was significantly lower (p less than 0.01) in the cirrhotics. There were no significant intergroup differences in duodenal mucosal blood volume. However, the cirrhotics showed significantly lower oxygen saturation of hemoglobin in all regions of the duodenum (p less than 0.01). These results show that the cirrhotics with esophageal varices had relative increase in blood volume and decrease in oxygen saturation of hemoglobin in the duodenal mucosa. Such microcirculatory disturbances seem to predispose liver cirrhosis patients to duodenal injury.

The purpose of this experiment was to examine the growth of condylar, epiphyseal, and spheno-occipital synchrondrosal cartilages cultured on medium BGJ. Materials from 12 neonatal rabbits were cultured for 7 days on this medium with the addition of 0, 10, or 20% fetal calf serum. Epiphyseal cartilages cultured with the addition of 20% fetal calf serum showed slightly better maintenance of in situ state, though little morphological changes were observed during 7 days. Spheno-occipital synchondrosal cartilages cultured with the addition of 10% or 20% fetal calf serum were histologically relatively similar to in situ state than those cultured without fetal calf serum, but no growth in length and width of these cartilages was observed. On the other hand, cultured condylar cartilages showed the least maintenance of histological and histochemical features. It was assumed that medium BGJ was not a recommendable medium for the organ culture of condylar cartilage.