Evaluation of a receptor gene responsible for maternal blood IgY transfer into egg yolks using bursectomized IgY-depleted chickens.

In avian species, maternal immunoglobulin Y (IgY) is transferred from the blood to the yolks of maturing oocytes; however, the mechanism underlying this transfer is unknown. To gain insight into the mechanisms of maternal IgY transfer into egg yolks, IgY-depleted chickens were generated by removing the bursa of Fabricius (bursectomy) during egg incubation, and their egg production and IgY transport ability into egg yolks were determined. After hatching, blood IgY concentrations of the bursectomized chickens decreased gradually until sexual maturity, whereas those of IgA remained low from an early stage of growth (from at least 2 wk of age). Chickens identified as depleted in IgY through screening of blood IgY and IgA concentrations were raised to sexual maturity. At 20 wk of age, both blood and egg yolk IgY concentrations in the IgY-depleted group were 600-fold lower than those of the control group, whereas egg production did not differ between the groups. Intravenously injected, digoxigenin-labeled IgY uptake into the egg yolk was approximately 2-fold higher in the IgY-depleted chickens than in the controls, suggesting that IgY depletion may enhance IgY uptake in maturing oocytes. DNA microarray analysis of the germinal disc, including the oocyte nucleus, revealed that the expression levels of 73 genes were upregulated more than 1.5-fold in the IgY-depleted group, although we could not identify a convincing candidate gene for the IgY receptor. In conclusion, we successfully raised IgY-depleted chickens presenting a marked reduction in egg yolk IgY. The enhanced uptake of injected IgY into the egg yolks of the IgY-depleted chickens supports the existence of a selective IgY transport mechanism in maturing oocytes and ovarian follicles in avian species.

Higher incorporation of heterologous chicken immunoglobulin Y compared with homologous quail immunoglobulin Y into egg yolks of Japanese quail (Coturnix japonica).

In avian species, blood IgY is selectively incorporated into the yolks of maturing oocytes, although the precise mechanism is poorly understood. Our previous study showed that 22% of i.v.-injected heterologous chicken IgY (cIgY) was incorporated into egg yolks of Japanese quail (Coturnix japonica). However, it is not known whether homologous quail IgY (qIgY) can be more efficiently incorporated into quail egg yolks than cIgY. Therefore, we compared the uptakes of qIgY and cIgY i.v. administered into quail egg yolks and further characterized the uptakes of these 2 antibodies into quail ovarian follicles. Quail IgY and cIgY purified from the blood of the respective bird were labeled with digoxigenin, and their uptakes into quail egg yolks were determined by ELISA. Unexpectedly, total incorporation of the injected qIgY was only one-third of that of cIgY, although much more qIgY was left in blood compared with cIgY, suggesting that qIgY is the less preferable antibody as a transport ligand into quail egg yolks. On the other hand, deposition of the qIgY into heart, lung, liver, spleen, kidney, and ovarian follicular membrane was markedly higher than that of cIgY. Amino acid sequence analysis of 3 peptides derived from the trypsin-digested qIgY heavy chain revealed low homology between qIgY and cIgY. In conclusion, our results show that heterologous cIgY is more efficiently incorporated into quail egg yolks than homologous qIgY, possibly due to a distinctive antibody transport system existing in oocytes. The present results also may provide a new strategy for delivering useful proteinaceous substances into egg yolks in an attempt to produce designer eggs.

Ingestion of paddy rice increases intestinal mucin secretion and goblet cell number and prevents dextran sodium sulfate-induced intestinal barrier defect in chickens.

Paddy rice is a potential feed grain for chickens, whose strong gizzards can crush the hull. Here, we investigated whether paddy rice rich in hull-derived water-insoluble dietary fiber stimulates intestinal mucin secretion and production, as well as the possible involvement of paddy rice in intestinal barrier function. Layer male chicks at 7 d of age were divided into four groups according to the diet: corn, polished rice, brown rice, or paddy rice (650 g/kg diet), which they ate for 14 consecutive days. At 21 d of age, the birds were refed their experimental diets, and small intestinal mucin fractions were collected to determine intestinal mucin content. Small intestinal mucin secretion was induced most strongly in the paddy rice group (Experiment 1). The rank order of diet-induced mucin secretion was paddy rice > corn = brown rice > polished rice. Ileal MUC2 gene expression and ileal number of goblet cells were highest in the paddy rice group (Experiment 1). A study of bromodeoxy-U uptake into ileal epithelial cells indicated the increase in goblet cells in the paddy rice group was related to accelerate epithelial cell migration (Experiment 2). A single supplementation of isolated rice hulls without kernels increased MUC2 gene expression and goblet cell numbers (Experiment 3), suggesting the importance of the hull's bulk-forming capacity on mucin production. Finally, chicks fed corn or paddy rice were orally administered dextran sodium sulfate (DSS) to disrupt intestinal barrier function. In the DSS-treated birds, the intestinal permeability of fluorescein isothiocyanate dextran in the everted gut sacs was much lower in the paddy rice group than in the corn group (Experiment 4), showing that paddy rice protects against mucosal disruption. In conclusion, ingestion of paddy rice increases intestinal mucin secretion and production through enhanced MUC2 gene expression and epithelial turnover and prevents DSS-induced intestinal barrier defects in chickens.

Age-related variations in glycosylation of hydroxylysine in human and rat skin collagens.

The extent of glycosylation of hydroxylysine in human skin collagen rapidly decreased during maturation and then gradually increased in proportion to the age. This decrease of glycosylation observed during maturation was also confirmed in whole, soluble and insoluble collagens from rat skin. These findings may contribute to the investigations on the functional role of glycosylation and also on the mechanism of maturational as well as senile processes.

Lp(a) lipoprotein as a risk factor for coronary heart disease and cerebral infarction.

Attempts were made to prepare antisera monospecific for Lp(a) lipoprotein and to investigate the distribution of subjects according to plasma levels of Lp(a) in Japanese controls and patients with coronary heart disease or cerebral infarction. Positive plasma reactions to the double diffusion test for Lp(a) (Ouchterlony) were observed in 32.3% of the healthy Japanese subjects, which is similar to results previously reported in western countries. The plasma threshold level of 17 mg/dl was considered an appropriate point for dividing subjects into positive and negative groups depending on reactions to the double diffusion test. When subjects were divided into two groups at 17 mg/dl, a significant association was found between a high plasma level of Lp(a) and either coronary heart disease or cerebral infarction in the distribution of the cortical artery. These results suggest that Lp(a) may play an important part as a risk factor for coronary heart disease and cerebral infarction.

Lipoprotein abnormalities in survivors of cerebral infarction with a special reference to apolipoproteins and triglyceride-rich lipoproteins.

Serum lipid and apolipoprotein concentrations were measured in 37 male survivors of cerebral infarction (CI) and in 30 healthy controls. Both groups had similar total cholesterol levels, but the HDL-cholesterol level was significantly lower and the serum triglyceride level was significantly higher in the CI patients than in the controls. The ApoB level was significantly higher in the CI patients but there was no significant difference between the 2 groups in the levels of the other apolipoproteins (ApoA-I, A-II, C-II, C-III, and E). The HDL-cholesterol/ApoA-I ratio was significantly lower in the CI patients. Both the VLDL-triglyceride and VLDL-cholesterol levels were higher in the CI patients but the VLDL-cholesterol especially its cholesterol ester level was conspicuously high. A population of VLDL particles that bound to heparin on heparin-Sepharose columns was increased in the CI patients. We suggest that cholesterol ester is excessively transferred from HDL to VLDL during the disturbed catabolism of VLDL in CI patients.

Activation of rabbit platelets by Ca2+ influx and thromboxane A2 release in an external Ca(2+)-dependent manner by zooxanthellatoxin-A, a novel polyol.

1. Zooxanthellatoxin-A (ZT-A), a novel polyhydroxylated long chain compound, isolated from a symbiotic marine alga Simbiodinium sp., caused aggregation in rabbit washed platelets in a concentration-dependent manner (1-4 microM), accompanied by an increase in cytosolic Ca2+ concentration ([Ca2+]i). 2. ZT-A did not cause platelet aggregation or increase [Ca2+]i in a Ca(2+)-free solution, and Cd2+ (0.1-1 mM), Co2+ (1-10 mM) and Mn2+ (1-10 mM) inhibited ZT-A-induced aggregation. SK&F96365 (1-100 microM), a receptor operated Ca2+ channel antagonist, and mefenamic acid (0.1-10 microM), a non-specific divalent cation channel antagonist, inhibited platelet aggregation and the increase in [Ca2+]i induced by ZT-A. 3. Indomethacin (0.1-10 microM), a cyclo-oxygenase inhibitor, and SQ-29548 (0.1-10 microM), a thromboxane A2 (TXA2) receptor antagonist, inhibited platelet aggregation and the increase in [Ca2+]i induced by ZT-A. 4. Methysergide (0.01-1 microM), a 5-HT2 receptor antagonist, inhibited ZT-A-induced platelet aggregation but did not affect the increase in [Ca2+]i induced by ZT-A. 5. Tetrodotoxin (1 microM), a Na+ channel blocker and chlorpheniramine (1 microM), a H1-histamine receptor antagonist, neither affected ZT-A-induced platelet aggregation nor the increase in [Ca2+]i induced by ZT-A. 6. Genistein (1-100 microM), a protein tyrosine kinase inhibitor, and staurosporine (0.01-1 microM), a protein kinase C inhibitor, also inhibited ZT-A-induced platelet aggregation. 7. The present results suggest that ZT-A elicits Ca(2+)-influx from platelet plasma membranes. The resulting increase in [Ca2+]i subsequently stimulates the secondary release of TXA2 from platelets. Furthermore, the response to ZT-A may be associated with tyrosine phosphorylation.

Inverted T graft: novel technique using composite radial and internal thoracic arteries.

BACKGROUND: Multiple arterial revascularization is feasible because of the excellent long-term patency of the arterial grafts compared with venous grafts. We present a new operative technique for multiple arterial revascularization using composite radial and internal thoracic arterial grafts. METHODS: Between January and September 1997, 12 patients had coronary artery bypass grafting with inverted T graft. The indications for inverted T graft were aortic calcification in 4 patients, repeat coronary artery bypass grafting in 1 patient, and total arterial revascularization in 7 young patients. The inverted T graft was constructed by interconnecting the coronary arteries and radial artery with end-to-side and side-to-side anastomoses, and by anastomosing the internal thoracic artery to the side of radial artery. RESULTS: Overall, 38 distal anastomoses (average number per patient, 3.2) were made with an inverted T graft. There were no deaths or perioperative myocardial infarctions. Postoperative angiography disclosed that all of the anastomoses were patent. CONCLUSION: This technique allows multiple arterial revascularizations without technical difficulty. It is useful in patients with aortic calcification, repeat coronary artery bypass grafting patients, and young patients who are candidates for total arterial revascularization.

Control of c-fos expression in STC-1 cells by peptidomimetic stimuli.

Enteroendocrine cells respond to nutrient and non-nutrient stimuli in the gut lumen. The intestinal hormone cholecystokinin (CCK) is secreted in response to luminal fatty acids, amino acids, peptides and proteins. The peptidomimetic cephalosporins have been reported to provide model, stable, compounds with similar secretagogue activity to peptide. Putative luminal stimuli also influence transcriptional activity in enteroendocrine cells, but the mechanisms are uncertain. In the present study we have investigated the control of c-fos expression in STC-1 cells (an enteroendocrine cell line). Peptidomimetics stimulated calcium-dependent release of CCK, and increased intracellular calcium, phosphorylation of p42/44 mitogen-activated protein kinase (MAP kinase) and c-fos mRNA abundance. Hypotonic stress also increased p42/44 MAP kinase phosphorylation and c-fos mRNA, but not CCK release. The increase in c-fos mRNA was strikingly potentiated by peptidomimetics in hypotonic medium. Increased c-fos expression, but not CCK release, was suppressed by the MAP kinase (MEK) inhibitor PD98059, and by the tyrosine kinase inhibitor genistein. We conclude that in STC-1 cells, peptidomimetics act through the p42/44 MAP kinase pathway to increase c-fos expression but not exocytosis. Moreover, a putative non-nutritive stimulus, hypotonic stress, may interact with this pathway to enhance c-fos expression, independently of hormone release.

Tyrphostin 23 blocks phosphorylation of p42 but not p38 mitogen-activated protein kinase by zooxanthellatoxin-A.

Zooxanthellatoxin-A isolated from a symbiotic dinoflagellate, caused aggregation in rabbit platelets that was inhibited by genistein (50 microM) and tyrphostin 23 (500 microM). Zooxanthellatoxin-A increased tyrosine phosphorylation of 42-kDa proteins which were identified as p42 and p38 mitogen-activated protein kinase (MAPK) by immunoprecipitation. Tyrphostin 23 inhibited the tyrosine phosphorylation of p42 MAPK but not p38 MAPK. In contrast, genistein abolished zooxanthellatoxin-A-induced tyrosine phosphorylation of both p42 and p38 MAPK. The results suggest that tyrphostin 23 selectively inhibits tyrosine phosphorylation of p42 MAPK. The p38 MAPK tyrosine phosphorylation is not involved in zooxanthellatoxin-A-induced platelet aggregation.

Vasoconstriction induced by zooxanthellatoxin-B, a polyoxygenated long-chain product from a marine alga.

We found that zooxanthellatoxin-B from a symbiotic marine alga, Symbiodinium sp., caused a concentration-dependent contraction of the rabbit isolated aorta at concentrations of 10(-7)-10(-5) M. Verapamil (10(-6) M) and mefenamic acid (10(-5) M) significantly attenuated the contractile response to zooxanthellatoxin-B at lower concentrations (10(-7)-10(-6) M) but not at higher concentrations (3 X 10(-6)-10(-5) M). The response to zooxanthellatoxin-B was partly inhibited by phentolamine (10(-6) M), whereas it was potentiated by ouabain (10(-5) M). Tetrodotoxin (10(-6) M), methysergide (10(-6) M), chlorpheniramine (10(-6) M) or indomethacin (3 X 10(-6) M), however, did not affect it. The zooxanthellatoxin-B-induced contraction was abolished by incubation in Ca2+-free solution. The contractile response increased in a concentration-dependent fashion with Ca2+ (0.03 and 10 mM) or Sr2 + (0.10 and 10 mM). After treatment with verapamil (10(-6) or 5 X 10(-6) M), the concentration-contractile response curves for Ca2+ and Sr2+ in the presence of zooxanthellatoxin-B were shifted to the right in parallel. MgCl2 (10 mM) shifted the concentration-response curve for Ca2+ more markedly than did verapamil. Zooxanthellatoxin-B increased tissue Na+ and reduced tissue K+ contents in the aorta, suggesting that zooxanthellatoxin-B increases Na+ and K+ permeability across the plasma membrane. These results suggest that the zooxanthellatoxin-B-induced contraction of the aorta is caused mainly by a direct action on smooth muscle, i.e., an increase in Ca2+ permeability that occurs at least partly through voltage-sensitive Ca2+ channels as well as through nonselective cation channels in the cell membrane of smooth muscle.

Aspirin promotes hepatic triacylglycerol accumulation in essential fatty acid-deficient Japanese quail.

This study was conducted to investigate whether the effect of a high dose of aspirin on hepatic triacylglycerol content is altered by dietary essential fatty acids (EFA) in Japanese quail. The birds were given an EFA-free or EFA-adequate [containing 2% (w/w) linoleic acid] diet ad libitum from 7 to 24 days of age. On the final experimental day, the birds received vehicle or 800 mg aspirin/kg body weight intraperitoneally and were killed 4 h subsequently. In birds fed the EFA-free diet, hepatic triacylglycerol content was more than 2 times higher after aspirin compared with vehicle treatment; in contrast, aspirin had no affect in birds fed the EFA-adequate diet. Liver malic enzyme and phosphatidate phosphohydrolase activities, which are related to lipid synthesis, were not affected by dietary EFA or aspirin treatment. Liver carnitine palmitoyltransferase activity in the birds fed the EFA-free diet was significantly lower than that in the birds fed the EFA-adequate diet, but aspirin did not affect this activity. In groups given the EFA-free diet, peroxisomal beta-oxidation was increased by the aspirin treatment. We conclude that acute administration of aspirin to Japanese quail on an EFA-free diet induces hepatic triacylglycerol accumulation, and that changes in lipid synthesis and degradation do not contribute to this phenomenon.

Isolation of zooxanthellatoxins, novel vasoconstrictive substances from the zooxanthella Symbiodinium sp.

Isolation of zooxanthellatoxins, novel vasoconstrictive substances from the zooxanthella Symbiodinium sp. Toxicon 31, 371-376, 1993. New polyhydroxypolyenes with potent vasoconstrictive activity, zooxanthellatoxin-A and -B, were isolated from a symbiotic marine alga Symbiodinium sp. These compounds caused sustained contractions of isolated rabbit aorta at concentrations above 7 x 10(-7) M; this effect was abolished in Ca(2+)-free solution or in the presence of verapamil. Both compounds were relatively large molecules (mol. wt about 2900), containing a large number of oxygen atoms and olefinic carbons, thus differing from two other vasoconstrictive marine toxins, palytoxin and maitotoxin, in containing more olefins than palytoxin, and fewer ethereal rings than maitotoxin.

Factors regulating amylase secretion from chicken pancreatic acini in vitro.

In mammals, cholecystokinin regulates pancreatic exocrine secretion under physiological conditions. We have shown, however, that cholecystokinin at physiological concentrations does not induce pancreatic amylase secretion in birds. Therefore, we investigated the effects of various neurotransmitters and gut hormones on the pancreatic amylase secretory response in isolated chicken pancreatic acini. Acetylcholine (half-maximal stimulation at 800 nM) and vasoactive intestinal polypeptide (half-maximal stimulation at 40 pM) produced a concentration-dependent increase in amylase secretion at physiological concentrations. The combination of acetylcholine and vasoactive intestinal polypeptide produced an additive response in amylase secretion. Sodium nitroprusside, a spontaneous nitric oxide releaser, and bombesin, induced amylase secretion at concentrations greater than 10 nM and 100 nM, respectively. Gastrin and secretin increased amylase secretion at pharmacological concentrations (10 to 100 nM). Our findings suggest that neural regulation is important for pancreatic enzyme secretion in birds and the contribution of gut hormones seems to be physiologically unimportant.

Aspirin toxicity in chicks given diets deficient in linoleic acid.

The toxicity of dietary aspirin on growth rate and lipid metabolism was investigated under linoleic acid (LA; 18: 2n-6) deficient conditions. One-week-old chicks were given diets containing 0 or 2% LA with or without 0.4% aspirin, until 4 weeks of age. Growth was severely depressed by dietary aspirin when chicks were given the LA-free diet. The liver was enlarged by both the aspirin and LA deficiency. The aspirin treatment induced a significant increase of 18:0 and arachidonic acid (20: 4n-6) and a decrease of 18: 1n-9 in the liver. In chicks fed LA-free diets, the ratio of 20:3n-9/20: 4n-6, which was used as an indicator of LA deficiency, was suppressed by aspirin treatment. In conclusion, the present results suggest that aspirin toxicity is altered by dietary LA concentrations.

The structure of permetin A, a new polypeptin type antibiotic produced by Bacillus circulans.

The structure of permetin A(I), an antibiotic substance produced by Bacillus circulans AJ 3902, has been elucidated as a cyclic acyl peptide by means of the mass and nuclear magnetic resonance spectroscopic techniques. (Formula: see text.) The structure was found to be the same as polypeptin A(II) except that L-Thr in II is replaced by L-Ser in I. Details of the structural determination are given for the permetin A itself as well as for the hydrolyzed permetin A. (Formula: see text.)

New streptothricin-group antibiotics, AN-201 I, II and III. II. Chemical structures.

The new, belonging to the streptothricin-group antibiotics AN-201 I, II and III were found to be produced by a soil actinomycete identified as Streptomyces nojiriensis C-13. The chemical structures and the physical and spectroscopic properties of these compounds are reported here. On the basis of NMR and fast atom bombardment mass spectrometry (FAB-MS) spectra the antibiotics were identified as N beta-acetylated derivatives of streptothricins E, D and F.

New streptothricin-group antibiotics, AN-201 I and II. Screening, fermentation, isolation, structure and biological activity.

Two streptothricin-group antibiotics, AN-201 I and II, were newly discovered and isolated from the culture broth of Streptomyces nojiriensis C-13. These antibiotics were purified by IRC-50 (H+) and CM-Sephadex C-50 chromatography, and paper electrophoresis. Structural analysis of AN-201 I and II showed that they were N beta-acetylated derivatives of streptothricin E and D, respectively. They had antibacterial activities against several strains of Escherichia coli, Bacillus subtilis, Micrococcus luteus and Staphylococcus aureus, and showed a strong selective cytotoxic effect on 3T3 cells transformed with SV-40 as compared with their normal cells in a test system in vitro as well as in vivo.

Cholestyramine alters the lipid and energy metabolism of chicks fed dietary medium- or long-chain triacylglycerol.

The effects of cholestyramine, a bile acid binding polymer, on the lipid and energy metabolism of chicks given dietary medium-chain triacylglycerol (MCT) or long-chain triacylglycerol (LCT) were investigated. Chicks (from 8 to 17 days of age) were fed diets containing MCT or LCT at 200 g oil/kg diet with or without 2% cholestyramine under equalized feeding conditions. An adjusted LCT diet was formulated in order to supply another group with daily nutrients and dietary metabolizable energy (ME) equal to MCT groups, except for corn starch. ME intakes of chicks given MCT or LCT diets were reduced by cholestyramine; consequently, fat and energy retention was reduced, though the reduction was more drastic in chicks fed LCT. This was caused by a change in amounts of the fecal excretion of fat and bile acids. Cholestyramine enhanced the excretion of octanoic acid (8:0) in the feces, which suggests that bile acids are needed for 8:0 absorption. Cholestyramine affects the utilization of dietary MCT and LCT by lowering fat and energy retention in chicks. However, the effect of cholestyramine on MCT utilization was smaller than its effect on utilization of LCT.

Interactions between medium-chain and long-chain triacylglycerols in lipid and energy metabolism in growing chicks.

The combined effects of dietary medium-chain triacylglycerols (MCT) and long-chain triacylglycerols (LCT) on lipid and energy utilization in chicks were investigated. Corn oil was used as the LCT source, and trioctanoin (8:0) was used as the MCT source. The efficiency of dietary energy utilization (such as metabolizable energy values and fat and energy retention) decreased linearly as the level of MCT increased, but the efficiency of dietary protein utilization (protein retained per protein consumed) was not affected in a consistent manner. Fecal saturated and monounsaturated fatty acid composition was dependent on the dietary fatty acid composition, whereas fecal linoleic acid content was not. It is concluded that dietary MCT and LCT influence each other to some degree, with respect to protein and lipid metabolism in chicks. Moreover, in most cases, the nutritional characteristics of each triacylglycerol, including food efficiency and fat and energy retention, are independent of each other in growing chicks.