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Vagal afferents regulate numerous physiological functions including arterial blood pressure, heart rate, breathing, and nociception. Cell bodies of vagal afferents reside in the inferior vagal (nodose) ganglia and their stimulation by various means is being considered as a way to regulate cardiorespiratory responses and control pain sensations. Stimulation of the nodose by exposure to infrared light is recently being considered as a precise way to elicit responses. These responses would likely involve the activity of temperature-sensitive membrane-bound channels. While papers have been published to track the expression of these transient receptor potential ion channels (TRPs), further studies are warranted to determine the in situ expression of the endogenous TRP proteins in the nodose ganglia to fully understand their pattern of expression, subcellular locations, and functions in this animal model. TRP ion channels are a superfamily of Na

Cholangiocarcinoma is an intractable carcinoma originating from the bile duct epithelium. To gain an understanding of the cell biology of cholangiocarcinoma, in vitro cell culture is valuable. However, well‑characterized cell lines are limited. In the present study, the morphology of the TK cholangiocarcinoma cell line was analyzed by three‑dimensional culture. Dispersed TK cells were injected into a gelatin mesh scaffold and cultivated for 3‑20 days. The morphology of the TK cells was investigated by phase‑contrast microscopy, optical microscopy, scanning electron microscopy (SEM) and transmission electron microscopy (TEM). TK cells were observed to proliferate three-dimensionally in the scaffold. The cells exhibited a globoid structure and attached to the scaffold. The SEM observation demonstrated typical microvilli and plicae on the surface of the structure. Light microscopy and TEM confirmed intercellular and cell‑to‑scaffold attachment in the three‑dimensional mesh. The culture also exhibited the formation of a duct-like structure covered by structured microvilli. In conclusion, three‑dimensional culture of TK cells demonstrated the morphological characteristics of cholangiocarcinoma in vitro. Production of high levels of carbohydrate antigen (CA)19‑9, CA50 and carcinoembryonic antigen was previously confirmed in the TK cell line. As a characteristic morphology was demonstrated in the present study, the TK cholangiocarcinoma cell line may be useful as an experimental model for further study of cholangiocarcinoma.

During cardiogenesis, a subset of epicardial cells undergoes epithelial–mesenchymal-transition (EMT) and the resulting epicardial-derived cells (EPDCs) contribute to the formation of coronary vessels. Our previous data showed hypoxia inducible factor-1α (HIF-1α) expression at specific sites within the epicardium and support a link between hypoxia inducible factors (HIFs) and the patterning of coronary vasculogenesis. To better understand the autocrine role of HIFs in the epicardium, we transduced adenovirus mediated expression of constitutively active HIF-1α (AdcaHIF1α) into the embryonic avian epicardium where the vascular precursors reside. We found that introducing caHIF1α into the epicardial mesothelium prevented EPDCs from proper migration into the myocardium. In vitro collagen gel assays and ex vivo organ culture data further confirmed that infection with AdcaHIF1α impaired the ability of EPDCs to invade. However, the proficiency of epicardial cells to undergo EMT was enhanced while the movement of EPDCs within the sub-epicardium and their differentiation into smooth muscle cells were not disrupted by caHIF1α. We also showed that the transcript level of Flt-1 (VEGFR1), which can act as a VEGF signaling inhibitor, increased several fold after introducing caHIF1α into epicardial cells. Blocking the activation of the VEGF pathway in epicardial cells recapitulated the inhibition of EPDC invasion. These results suggest that caHIF1α mediated up-regulation of Flt-1, which blocks the activation of the VEGF pathway, is responsible for the inhibition of EPDC myocardial migration. In conclusion, our studies demonstrate that HIF signaling potentially regulates the degree of epicardial EMT and the extent of EPDC migration into the myocardium, both of which are likely critical in patterning the coronary vasculature during early cardiac vasculogenesis. These signals could explain why the larger coronaries appear and remain on the epicardial surface.

Some foods from plant sources, particularly of Moringeceae origin, contained compounds that enhanced bacterial ice-nucleation activity. A hot water extract from mustard seeds was so potent in its enhancing activity that the compounds responsible could be isolated from the seeds as yellow crystals. By instrumental analyses, this enhancer was identified as 4-hydroxy-3-nitrophenylacetic acid. The addition of a purified preparation of this compound at 0.1-1 ppm to the cultivation medium for Xanthomonas campestris was effective for enhancing its ice-nucleation activity.

We have previously shown that estrogen receptor β (ERβ)-mediated up-regulation of quinone reductase (QR) is involved in the protection against estrogen-induced mammary tumorigenesis. Our present study provides evidence that the ERβ agonist, 2,3-bis-(4-hydroxy-phenyl)-propionitrile (DPN), and the selective estrogen receptor modulator tamoxifen (Tam), inhibit estrogen-induced DNA damage and mammary tumorigenesis in the aromatase transgenic (Arom) mouse model. We also show that either DPN or Tam treatment increases QR levels and results in a decrease in ductal hyperplasia, proliferation, oxidative DNA damage (ODD), and an increase in apoptosis. To corroborate the role of QR, we provide additional evidence in triple transgenic MMTV/QR/Arom mice, wherein the QR expression is induced in the mammary glands via doxycycline, causing a decrease in ductal hyperplasia and ODD. Overall, we provide evidence that up-regulation of QR through induction by Tam or DPN can inhibit estrogen-induced ODD and mammary cell tumorigenesis, representing a novel mechanism of prevention against breast cancer. Thus, our data have important clinical implications in the management of breast cancer; our findings bring forth potentially new therapeutic strategies involving ERβ agonists. Krishnamurthy, N., Hu, Y., Siedlak, S., Doughman, Y. Q., Watanabe, M., Montano, M. M. Induction of quinone reductase by tamoxifen or DPN protects against mammary tumorigenesis.

Recently, we found that mutation of the C-terminus of transcription factor hexamethylene bisacetamide-inducible protein 1 (HEXIM1) in mice leads to abnormalities in cardiovascular development because of aberrant vascular endothelial growth factor (VEGF) expression. HEXIM1 regulation of some genes has also been shown to be positive transcription elongation factor b (P-TEFb) dependent. However, it is not known whether HEXIM1 regulates VEGF in the mammary gland. We demonstrate that HEXIM1 regulates estrogen-induced VEGF transcription through inhibition of estrogen receptor-alpha recruitment to the VEGF promoter in a P-TEFb-independent manner in MCF-7 cells. Under hypoxic conditions, HEXIM1 inhibits estrogen-induced hypoxia-inducible factor-1 alpha (HIF-1alpha) protein expression and recruitment of HIF-1alpha to the hypoxia-response element in the VEGF promoter. In the mouse mammary gland, increased HEXIM1 expression decreased estrogen-driven VEGF and HIF-1alpha expression. Conversely, a mutation in the C-terminus of HEXIM1 (HEXIM1(1-312)) led to increased VEGF and HIF-1alpha expression and vascularization in mammary glands of heterozygous HEXIM1(1-312) mice when compared with their wild-type littermates. In addition, HEXIM1(1-312) mice have a higher incidence of carcinogen-induced mammary tumors with increased vascularization, suggesting an inhibitory role for HEXIM1 during angiogenesis. Taken together, our data provide evidence to suggest a novel role for HEXIM1 in cancer progression.

Lung maturation at the terminal sac stage of lung development is characterized by a coordinated increase in terminal sac formation and vascular development in conjunction with the differentiation of alveolar type I and type II epithelial cells. The Cited2-Tcfap2a/c complex has been shown to activate transcription of Erbb3 and Pitx2c during mouse development. In this study, we show that E17.5 to E18.5 Cited2-null lungs had significantly reduced terminal sac space due to an altered differentiation of type I and type II alveolar epithelial cells. In addition, E17 Cited2-null lungs exhibited a decrease in the number of apoptotic cells, contributing to the loss in airspace. Consistent with the phenotype, genes associated with alveolar cell differentiation and survival were differentially expressed in Cited2-null fetal lungs compared to those of wild-type littermates. Moreover, expression of Cebpa, a key regulator of airway epithelial maturation, was significantly decreased in Cited2-null fetal lungs. Cited2 and Tcfap2c were present on the Cebpa promoter in E18.5 lungs to activate Cebpa transcription. We propose that the Cited2-Tcfap2c complex controls lung maturation by regulating Cebpa expression. Understanding the function of this complex may provide novel therapeutic strategies for patients with respiratory distress syndromes.

Apoptosis repressor with caspase recruitment domain is expressed at high levels in brain and myogenic tissues, consistent with a role to inhibit apoptosis in the terminally differentiated cells. Expression of ARC in cancers is not known. In this study, we reported that ARC was highly expressed in various non-myogenic and non-neurogenic human and rat cancer cell lines. Unexpectedly, ARC was localized almost exclusively to the nuclei of cancer cells, which was unlike the cytoplasmic localization of ARC in non-cancer cells. Furthermore, nuclear ARC in cancer cells did not co-localize with nucleolus protein of 30kDa, an alternatively spliced ARC isoform. These findings indicate that ARC is distributed differently in cancer cells than non-cancer cells and thus might play a role in neoplastic transformation.

Hypoallergenic wheat flour produced by modification with cellulase and actinase showed inhibitory activity against ovalbumin permeation in an in vitro model by using the Caco-2 cell monolayer. The activity was found in the cellulase preparation used for producing the flour. An active compound was isolated by HPLC and identified as Trp-Ser-Asn-Ser-Gly-Asn-Phe-Val-Gly-Gly-Lys by 1H-NMR data and Edman degradation. The undecapeptide, some oligopeptides with the N-terminal sequences and Trp ethyl ester showed activity at 10(-7) M, acetyl Trp being active at 10(-2) M. These data suggest that the Trp residue without a free carboxyl group would be required for the inhibitory activity of ovalbumin absorption through the intestinal tract.

The role of apoptosis in cardiac morphogenesis has not been directly tested. Cardiomyocyte apoptosis is prevalent during the remodeling of the embryonic chicken cardiac outflow tract (OFT) in the transition from a single to a dual circulation. We tested the hypothesis that OFT cardiomyocyte apoptosis drives the shortening and rotation of the embryonic cardiac OFT and is required to achieve the mature ventriculo-arterial configuration. Chick embryos were treated with the peptide Caspase inhibitors zVAD-fmk or DEVD-cho at HH stages 15–20 (looped heart). Morphology of control and experimental embryos was assessed at HH stage 35, at which time the control hearts have developed a dual circulation. Infection of the hearts with a recombinant adenovirus expressing green fluorescent protein was used to follow the fate of the OFT cardiomyocytes. Affected embryos displayed abnormal persistence of a long infundibulum (OFT myocardial remnant) beneath the great vessels, indicating failure of OFT shortening. In some instances, the infundibulum connected both great vessels to the right ventricle in a side-by-side arrangement with transposition of the aorta, indicating a failure of rotation of the OFT, and modeling human congenital double outlet right ventricle. Defects were also observed at other sites in the heart where apoptosis is prevalent, such as in the formation of the cardiac valves and trabeculae. To more specifically target the apoptosis of the OFT cardiomyocytes, recombinant adenovirus was used to express the X-linked inhibitor of apoptosis protein in these cells. This resulted in an effect on outflow tract shortening and rotation similar to that of the peptide inhibitors, while the effects on the other cardiac structures were not observed. These results demonstrate that elimination of OFT cardiomyocytes by apoptosis is necessary for the proper formation of the ventriculo-arterial connections, and suggest apoptosis as a potential target of teratogens and genetic defects that are associated with congenital human conal heart defects.

Brown Norway rats were immunized with gluten, and then fed a diet containing hypoallergenic flour or an amino acid mixture. The rats were then made to inhale a solubilized gluten to induce gluten-specific bronchial asthma. The antibody levels in the serum of rats were measured by ELISA, and cell counts were done on cytospin preparations of bronchoalveolar lavage fluid. Body weight was decreased after allergen challenge in rats fed the amino acid mixture but not in rats fed the hypoallergenic flour. Antibody levels in the serum were significantly lower in rats fed hypoallergenic flour than in those fed the amino acid mixture. Differential cell counts in the bronchoalveolar lavage fluid showed that the numbers of eosinophils, lymphocytes, and neutrophils were significantly lower in rats fed the hypoallergenic flour than in those fed the amino acid mixture. These results suggest that hypoallergenic flour actively suppresses the allergic reactions, probably by inducing oral tolerance.

One class of the nuclear receptor AF-2 coactivator complexes contains the SRC-1/TIF2 family, CBP/p300 and an RNA coactivator, SRA. We identified a subfamily of RNA-binding DEAD-box proteins (p72/p68) as a human estrogen receptor alpha (hER alpha) coactivator in the complex containing these factors. p72/p68 interacted with both the AD2 of any SRC-1/TIF2 family protein and the hER alpha A/B domain, but not with any other nuclear receptor tested. p72/p68, TIF2 (SRC-1) and SRA were co-immunoprecipitated with estrogen-bound hER alpha in MCF7 cells and in partially purified complexes associated with hER alpha from HeLa nuclear extracts. Estrogen induced co-localization of p72 with hER alpha and TIF2 in the nucleus. The presence of p72/p68 potentiated the estrogen-induced expression of the endogenous pS2 gene in MCF7 cells. In a transient expression assay, a combination of p72/p68 with SRA and one TIF2 brought an ultimate synergism to the estrogen-induced transactivation of hER alpha. These findings indicate that p72/p68 acts as an ER subtype-selective coactivator through ER alpha AF-1 by associating with the coactivator complex to bind its AF-2 through direct binding with SRA and the SRC-1/TIF2 family proteins.

The development of food science in the near future probably depends on the advance in functional food science, the concept of which was proposed first in Japan nearly 15 years ago. The new science has been internationally distributed and accepted as conceptually being beyond nutrition. In Japan, however, it traced a unique path of progress in the form of a product-driven rather than concept-driven science. Actually, a number of substances and products with potential for disease risk reduction rather than simply for health maintenance have been investigated for their body-modulating functions. Some of them have been applied in practice to the industrialization of functional foods in terms of "foods for specified health uses" legally defined by new legislation. A variety of sophisticated methods have been introduced as well, including the so-called "XYZ" evaluation system, database construction for assessment of the function, and even the DNA microarray technique. The Ministry of Agriculture, Forestry, and Fisheries (MAFF) and the Ministry of Health and Welfare (MHW) also commenced their scientific as well as political activity, with its spread to industries which almost simultaneously began to vigorously investigate functional food products for enlargement of the food market. With all of this as a background, the Japan Liaison of the International Union of Food Science and Technology (IUFoST) hold a function food science symposium on behalf of related scientific bodies including the Japan Section of the International Life Science Institute (ILSI). This paper is an overview compiled from 12 presentations made in the symposium, with the aim of internationally publicizing the activity of functional food science in Japan.

Estrogen receptor (ER) alpha and beta mediate estrogen actions in target cells through transcriptional control of target gene expression. For 17beta-estradiol-induced transactivation, the N-terminal A/B domain (AF-1) and the C-terminal E/F domain (AF-2) of ERs are required. Ligand binding is considered to induce functional synergism between AF-1 and AF-2, but the molecular mechanism remains unknown. To clarify this synergism, we studied the role of reported AF-2 coactivators, p300/CREB binding protein, steroid receptor coactivator-1/transcriptional intermediary factor-2 (SRC-1/TIF2) family proteins and thyroid hormone receptor-associated protein-220/(vitamin D3 receptor-interacting protein- 205-(TRAP220/DRIP205) on the AF-1 activity in terms of synergism with the AF-2 function. We found that neither any of the SRC-1/TIF2 family coactivators nor TRAP220/DRIP205 is potent, whereas p300 potentiates the AF-1 function of both human ERalpha and human ERbeta. Direct interactions of p300 with the A/B domains of ERalpha and ERbeta were observed in an in vitro glutathione S-transferase pull-down assay in accordance with the interactions in yeast and mammalian two-hybrid assays. Furthermore, mutations in the p300 binding sites (56-72 amino acids in ERalpha and 62-72 amino acids in ERbeta) in the A/B domains caused a reduction in ligand-induced transactivation functions of both ERalpha and ERbeta. Thus, these findings indicate that ligand-induced functional synergism between AF-1 and AF-2 is mediated through p300 by its direct binding to the A/B regions of ERalpha and ERbeta.

A new polysaccharide with a molecular weight of 5.0 x 10(4) was isolated as a possible wheat allergen from a water-soluble fraction of flour by affinity chromatography and gel filtration. The isolated polysaccharide was found to be a possible wheat allergen, as it bound specifically to IgE antibodies in the sera of patients allergic to the water-soluble fraction of flour. Chemically, the sugar moiety of the polysaccharide consisted of D-glucose and D-mannose with beta-1,4-linkages in a molar ratio of 4.4:1. Since this mannoglucan is thought to be stable in our body, it would act as a remaining allergen to cause a long-lasting allergic reaction to wheat flour.

Two strains of measles virus IMA and SMD that were isdated using B95a cell line show hemadsorption negative. After adaptation to Vero cells, these strains gain the ability to agglutinate AGM-RBC and show hemadsorption positive. Sequence analysis of these two kinds of virus reveals that the two strains all have a Ser (HAD negative) to Gly (HAD positive) mutation at position 546 in the H protein. Site-directed mutagenesis and expression in COS cells were used to confirm that the mutation Ser→Gly is responsible for hemadsorption alteration. Then the monoclonal antibody against CD46 was used to identify that this mutation also governs the binding function of MV H protein to CD46 receptor. The data provide a new important amino acid of MV H protein that governs the hemadsorption and binding to CD46 receptor.

The development of the tubular heart into a complex four-chambered organ requires precise temporal and region-specific regulation of cell proliferation, migration, death and differentiation. While the regulatory mechanisms in heart morphogenesis are not well understood, increasing attention has focused on the homeodomain proteins, which are generally linked to morphogenetic processes. The homeodomain containing gene Gax has been shown to be expressed in heart and smooth muscle tissues. In this study, the Gax protein was detected in the nuclei of myocardial cells relatively late in chicken heart development, at a time when myocyte proliferation is declining. To test the hypothesis that the Gax protein functions as a negative regulator of cardiomyocyte proliferation, a replication-defective adenovirus was used to force its precocious nuclear expression during chicken heart morphogenesis. In experiments in which Gax- and β-galactosidase-expressing adenoviruses were co-injected, clonal expansion of myocytes was reduced, consistent with inhibition of myocyte proliferation. This effect on proliferation was corroborated by the finding that the percentage of exogenous Gax-expressing myocytes that were positive for the cell cycle marker PCNA decreased over time and was lower than in control myocytes. The precocious nuclear expression of Gax in tubular hearts resulted in abnormal heart morphology, including small ventricles with rounded apices, a thinned compact zone and coarse trabeculae. These results suggest a role for the Gax protein in heart morphogenesis causing proliferating cardiomyocytes to withdraw from the cell cycle, thus influencing the size and shape that the heart ultimately attains.

Differential tissue hypoxia drives normal cardiogenic events including coronary vessel development. This requirement renders cardiogenic processes potentially susceptible to teratogens that activate a transcriptional pathway that intersects with the hypoxia-inducible factor (HIF-1) pathway. The potent toxin 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is known to cause cardiovascular defects by way of reduced myocardial hypoxia, inhibition of angiogenic stimuli, and alterations in responsiveness of endothelial cells to those stimuli. Our working hypothesis is that HIF-1 levels and thus HIF-1 signaling in the developing myocardium will be reduced by TCDD treatment in vivo during a critical stage and in particularly sensitive sites during heart morphogenesis. This inadequate HIF-1 signaling will subsequently result in outflow tract (OFT) and coronary vasculature defects. Our current data using the chicken embryo model showed a marked decrease in the intensity of immunostaining for HIF-1α nuclear expression in the OFT myocardium of TCDD-treated embryos. This area at the base of the OFT is particularly hypoxic during normal development; where endothelial cells initially form a concentrated anastomosing network known as the peritruncal ring; and where the left and right coronary arteries eventually connect to the aortic lumen. Consistent with this finding, anomalies of the proximal coronaries were detected after TCDD treatment and HIF-1α protein levels decreased in a TCDD dose-dependent manner.

For application to the treatment of wheat-sensitive allergy, we developed a practical method for producing a haptenic peptide mixture, and evaluated its haptenic properties. Wheat flour was hydrolyzed with chymotrypsin to obtain a hydrolysate, and the diffusible fraction from the hydrolysate was subjected to gel-filtration. The resulting oligopeptide fraction could bind to wheat-specific IgE antibodies in the serum of patients allergic to wheat. Since this peptide mixture did not induce histamine release from basophils in these patients, it was concluded that the peptide mixture was composed of monovalent haptens. Histamine release from antigenstimulated basophils was almost completely inhibited when the basophils were preincubated with the haptenic peptide mixture. These results suggest that this haptenic peptide mixture can regulate the allergenic reaction in an epitope-specific manner.

The minimum primary structure of the IgE-binding epitope in wheat gluten was determined as Gln-Gln-Gln-Pro-Pro. The N-terminal glutamine and the two proline residues were essential for epitopic function. The occurrence of the second glutamine residue and acetylation of the N-terminal amino group were found to exert some auxiliary functions, whereas only a lesser contribution was expected for the third glutamine residue. It has also been confirmed that acetyl-Gln-Gln-Gln-Pro-Pro bound to wheat-specific IgE antibodies in the sera of patients allergic to wheat, although it did not induce histamine release from the basophils of these patients. Taken together, we concluded that the Gln-Gln-Gln-Pro-Pro motif constituted an IgE-binding rather than immunogenic epitope and also that acetyl-Gln-Gln-Gln-Pro-Pro might act as a hapten capable of binding to specific IgE molecules. The possibility exists that this peptide will have practical application to therapy for or prevention of wheat-sensitive allergy.

We investigated the murine antibody response to recombinant p17 (rp17) of human immunodeficiency virus type 1 (HIV-1) and the human antibody response directed to p17 in HIV-1 infection. Three large peptides covering residues 12-29, 53-87 and 87-115 of p17 were synthesized. The cysteine residues 57 and 87 of peptide 53-87 were reoxidized to form a disulfide bridge. Eighteen out of 19 murine monoclonal anti-rp17 antibodies had relatively high affinities (KA = 1.9 x 10(5)-1.4 x 10(8) M-1) with one of the 3 p17 peptides in the liquid phase. Each monoclonal antibody reacted only with one particular peptide and had no reactivity with the other 2 p17 peptides. All the monoclonal antibodies reacted with rp17 in the liquid phase with a reasonable degree of affinity (KA = 2.0 x 10(5)-1.8 x 10(7) M-1). Four HIV-1 carrier sera, which were positive in ELISA using rp17 as the antigen, reacted positively in an ELISA using 3 p17 peptides which were used to titrate murine monoclonal antibodies. Murine monoclonal antibodies having specificity for the 3 p17 peptides stained live HIV-1-infected cells by means of indirect membrane immunofluorescence, irrespective of their specificity. This suggests that the various portions of p17 (at least 3 regions of p17) were exposed on the surface of live infected cells, probably as short polypeptide chains.

Hypoallergenic wheat products in the form of pasta-like noodles and bread were fabricated. Wheat flour was added with a 0.6-fold weight of water dissolving collagenase to obtain hypoallergenic flour batter. Methods for producing hypoallergenic pasta-like noodles and bread from the batter were designed. In the noodle making, the batter was mixed with an oligosaccharide with a mild sweetness, a surfactant, and salt, and the mixture was extruded under heating. Both retorting and refrigerating processes were applied to the noodles. In the bread making a mixture of the batter, glucose, citric acid, sodium hydrogen carbonate and salt was baked at 180°C. The addition of glucose contributed to generation of faborable flavor and color.

Commercially available egg white was freeze-concentrated using bacterial ice nuclei, which improved its foaming properties. Baked meringue, angel cake, snow-jelly, and marshmallow were prepared using freeze-concentrated egg white foam and their physical properties were evaluated. We concluded that freeze-concentrated egg white can be used to make some foam foods.

The ice nucleation-active protein of Erwinia ananas IN-10 (inaA protein) was over-expressed as inclusion bodies in Escherichia coli in a yield of 15.3 mg of inaA protein from 60 mg of bacterial cells on a dry-matter basis. The inaA protein was purified from the inclusion bodies by solubilization with detergents to obtain a protein preparation free from sugar and lipid. This preparation had a distinct ice nucleation activity, indicating that the inaA protein per se is able to act as a nucleus.

This study aims to improve the stickiness and brightness of cooked rice from aged rice grains by removing its albumins and globulins with proteases. In newly–harvested grains, the proteins were easily removed by extraction with water which contained minerals from rice. In aged grains, however, the proteins were already denatured and extracted with difficulty. Proteases hydrolyzed even the denatured proteins to make them extractable. As a result, starch granules on the surface of grains were liberated. Some proteases also hydrolyzed a starch granule-associated protein, and consequently, made the starch highly gelatinizable by heating. The protease treatment of aged grains was effective for two main reasons: liberation of starch granules and removal of the granule-associated protein. During cooking of rice the liberated starch without the associated proteins gelatinized to give rise to stickiness. The gelatinized starch is considered to form a more isotropic film on the surface of every cooked rice grain to...

Xanthomonas campestris INXC-1 is able to freeze water at a subzero temperature higher than -5°C. High-pressure treatment at 300 MPa and 5°C for 5 min killed the cells without affecting their ice-nucleation activity. Egg white containing the pressurized cells began to freeze with a slight degree of supercooling. Ice crystals with a dendritic structure were formed when the egg white froze in the presence of the killed bacterial cells. The frozen egg white thawed more quickly than egg white frozen without the cells.

High-intensity fluorescent nanoparticles, quantum dots (QDs), have been applied to a wide range of biological studies and medical studies by taking advantage of their fluorescent properties. On the other hand, we have reported the specificity of JT-95 monoclonal IgM antibody, which recognizes the antigen of thyroid carcinomas. Here we show that the combination of QDs and JT-95 monoclonal antibody was applicable to Western blotting analysis, ELISA-like system, and fluorescent microscopic analysis of SW1736 thyroid carcinoma cell line. We have opened up the possibility that antibodies for higher specific recognition, even IgM, are applicable to the detection system with QDs.

A novel method for producing a low ovomucoid egg white preparation is proposed. Egg white powder (0.5 g) was dissolved in a 10-fold weight of distilled water and adjusted to pH 5, and ethanol was added to the solution at a final concentration of 20% (v/v). The mixture was vigorously stirred and centrifuged. The precipitate was washed three times with 20% ethanol (6.25 ml each), with about 65% of egg white proteins occurring in the precipitate. The use of ELISA demonstrated that 70% of ovomucoid was recovered from the supernatant fraction. However, functionally important proteins such as ovalbumin, ovotransferrin, and lysozyme still remained in the precipitate. These results may be due primarily to the much higher solubility of ovomucoid in this aqueous ethanol. Food quality evaluation showed that high whippability and foam stability were retained in the low ovomucoid preparation as in its material egg white. This product would thus be applicable as a new processed food for ovomucoid-sensitive allergic patients.

Tumor-infiltrating lymphocytes (TIL) were generated from 10 glioma specimens by using recombinant interleukin-2 and an anti-CD3 antibody (CD3 + TILs). We obtained more than 1 x 10(9) cells in 5 cases, more than 5 x 10(8) cells in 2 cases, and about 1 x 10(8) cells in 3 cases during three weeks of incubation from small specimens ranging in weight from 0.5 to 2.0 g. In 4 cases, TILs were expanded following stimulation with only rIL-2 (CD3-TILs). The growth rate of CD3-TILs was less than that of CD3 + TILs. Cytotoxicity of CD3 + TILs was lower than that of lymphokine-activated killer (LAK) cells in a standard 4h 51Cr release assay. Cold target inhibition was undertaken in three cases and specific cytotoxicity could be shown in only one case. CD3 + TILs mainly consisted of CD3-positive cells, ranging from 63.2 to 99.9%. The ratio of CD4-positive cells to CD8-positive cells was not constant. The expression of Leu 7 and CD16 was low. The present study did not confirm previous findings that TILs were more tumor-selective and potent than LAK cells. Furthermore, the results on in vitro antitumor activity of those cells were not necessarily consistent with the results on their clinical activity. Further careful work is necessary on the preparation of immunocytes and the subsequent adoptive immunotherapy.

Cited2 is a transcriptional modulator with pivotal roles in different biological processes. Cited2-deficient mouse embryos manifested two major defects in the developing eye. An abnormal corneal-lenticular stalk was characteristic of Cited2-/- developing eyes, a feature reminiscent of Peters' anomaly, which can be rescued by increased Pax6 gene dosage in Cited2-/- embryonic eyes. In addition, the hyaloid vascular system showed hyaloid hypercellularity consisting of aberrant vasculature, which might be correlated with increased VEGF expression in the lens. Deletion of Hif1a (which encodes HIF-1α) in Cited2-/- lens specifically eliminated the excessive accumulation of cellular mass and aberrant vasculature in the developing vitreous without affecting the corneal-lenticular stalk phenotype. These in vivo data demonstrate for the first time dual functions for Cited2:one upstream of, or together with, Pax6 in lens morphogenesis; and another in the normal formation of the hyaloid vasculature through its negative modulation of HIF-1 signaling. Taken together, our study provides novel mechanistic revelation for lens morphogenesis and hyaloid vasculature formation and hence might offer new insights into the etiology of Peters'anomaly and ocular hypervascularity.

To determine how estrogens are involved in the growth of endometrial cancer with varying degrees of differentiation, we investigated the status of p72, a novel specific coactivator for estrogen receptor α (ERα) activation function-1 (AF-1), AIB1, a steroid receptor coactivator amplified in breast cancer 1, erbB-2, a receptor tyrosine kinase, and ERα in endometrial cancer. Gene expression of ERα, p72, AIB1 and erbB-2 was measured in 26 samples of primary endometrial cancers by real-time RT-PCR, and their in vivo cellular effects on the transactivation function of ERα were examined by a transient expression assay. The mRNA levels of erbB-2 increased and those of ERα, p72 and AIB1 decreased with the loss of histological differentiation. Transient expression of p72, AIB1 and erbB-2 in human embryonic kidney 293T cells led to a synergistic promotion of the transactivation function of ERα in the presence of 17α-estradiol or 4-hydroxytamoxifen, an ERα AF-1 agonist/AF-2 antagonist, as a ligand. In conclusion, estrogen action through ERα AF-1 might be exerted by the increased expression of the coactivators p72 and AIB1, together with cross talk between erbB-2 and p72, to accelerate the transactivation of ERα AF-1 in endometrial cancer. These findings also suggest that the cooperative transactivation of ERα AF-1 by the overexpression of p72, AIB1 and erbB-2 might be involved in tamoxifen-stimulated growth of endometrial cancer.

The transcriptional modulator Cited2 is induced by various biological stimuli including hypoxia, cytokines, growth factors, lipopolysaccharide (LPS) and flow shear. In this study, we report that Cited2 is required for mouse fetal liver development. Cited2(-/-) fetal liver displays hypoplasia with higher incidence of cell apoptosis, and exhibits disrupted cell-cell contact, disorganized sinusoidal architecture, as well as impaired lipid metabolism and hepatic gluconeogenesis. Furthermore, we demonstrated the physical and functional interaction of Cited2 with liver-enriched transcription factor HNF4alpha. Chromatin immunoprecipitation (ChIP) assays further confirmed the recruitment of Cited2 onto the HNF4alpha-responsive promoters and the reduced HNF4alpha binding to its target gene promoters in the absence of Cited2. Taken together, this study suggests that fetal liver defects in mice lacking Cited2 result, at least in part, from its defective coactivation function for HNF4alpha.

Part of the salt in soy sauce formed no eutectic H2O•2NaCl crystals at sub-zero temperatures and remained in a freeze-concentrated product. NMR line width of (23)Na was broader in the concentrated soy sauce than in the material. A broad line width of (23)Na was also observed in an aqueous solution of NaCl and a non-diffusible soy sauce fraction. The data indicate that part of the salt in soy sauce interacted with its non-diffusible fraction and that such bound salt formed no eutectic crystals.

A methanol extract from yellow mustard seeds had antibacterial activity against Escherichia coli, Salmonella enteritidis, and Staphylococcus aureus. Two compounds with such activity were isolated from the extract. By instrumental analysis, the compounds were identified as 4-hydroxy-3-nitrophenylacetic and sinapic acids. Examination of the structure-activity relationship showed that the hydroxyl and nitro groups of the first compound were involved in the activity against all three species. The two methoxyl groups and the hydroxyl group in sinapic acid were effective against E. coli and all of the substituents of the benzene ring were effective against S. enteritidis. The presence of the propenoic group of the second compound was effective against S. aureus.

Hypoxia inducible factor-1 (HIF-1) initiates key cellular and tissue responses to physiological and pathological hypoxia. Evidence from in vitro and structural analyses supports a critical role for Cited2 in down-regulating HIF-1-mediated transcription by competing for binding with oxygen-sensitive HIF-1alpha to transcriptional co-activators CBP/p300. We previously detected elevated expression of HIF-1 target genes in Cited2(-/-) embryonic hearts, indicating that Cited2 inhibits HIF-1 transactivation in vivo. In this study, we show for the first time that highly hypoxic cardiac regions in mouse embryos corresponded to the sites of defects in Cited2(-/-) embryos and that defects of the outflow tract, interventricular septum, cardiac vasculature, and hyposplenia were largely rescued by HIF-1alpha haploinsufficiency. The hypoxia of the outflow tract and interventricular septum peaked at E13.5 and dissipated by E15.5 in wild-type hearts, but persisted in E15.5 Cited2(-/-) hearts. The persistent hypoxia and abnormal vasculature in the myocardium of interventricular septum in E15.5 Cited2(-/-) hearts were rescued with decreased HIF-1alpha gene dosage. Accordingly, mRNA levels of HIF-1-responsive genes were reduced in Cited2(-/-) embryonic hearts by HIF-1alpha heterozygosity. These findings suggest that a precise level of HIF-1 transcriptional activity critical for normal development is triggered by differential hypoxia and regulated through feedback inhibition by Cited2.