Absorption of Horseradish Peroxidase (HRP) In Vitro Across Bovine Jejunal and Ileal Epithelia Around the Time of Weaning

Using the everted sac methodology as well as an Ussing chamber, we investigated changes in the absorption of horseradish peroxidase (EC 1.11.1.7, HRP (40 kDa)) in jejunum and ileum segments isolated from male Holstein cattle around the time of weaning (6 to 15 wks old). By the everted sac method, HRP transport (HRP concentration on the serosal side sampled after a 60-min-incubation) at 15 wks of age was significantly greater than that at 6 wks of age, in both segments of the intestine. Absorption was not significantly different between the jejunum and ileum. Addition of Na^+/K^+ ATPase inhibitor (ouabain, 1 mM) did not cause any significant change in HRP absorption, whereas Na^+/H^+ anti-transporter inhibitor (amiloride, 1μM) significantly increased the absorption in both sacs at 8 wks of age. By the Ussing chamber method, there were no significant differences between the values for Jsm and Jms, while the Jnet value was nearly zero for both epithelia. In addition, the flux (Jms) of Lucifer yellow, a cell-membrane-impermeable fluorescence dye, was significantly greater at 6 than at 13 wks of age in the ileal epithelia, although the flux was significantly greater in the jejunal than the ileal epithelia at both ages. From these findings, we conclude that: 1) bovine jejunal and ileal epithelia are able to absorb a large molecule such as the HRP protein; 2) HRP transport occurs in a concentration-dependent manner and may in part be via a paracellular pathway; 3) the increased HRP transport shown at 15 wks of age may not be caused by an increased use of the paracellular pathway.horseradish peroxidaseabsorptiontransportcal

Map-guided surgery for atrial fibrillation

BackgroundAlthough current surgical procedures result in a high success rate for atrial fibrillation, they are not guided by electrophysiologic findings in individual patients and thus might include unnecessary incisions in some patients or be inappropriate for other patients. We sought to determine whether intraoperative mapping is beneficial for the surgical treatment of atrial fibrillation.MethodsA 256-channel 3-dimensional dynamic mapping system with custom-made epicardial patch electrodes was used to examine the atrial activation during atrial fibrillation and to determine the optimal procedure in 37 patients with continuous and 9 patients with intermittent atrial fibrillation intraoperatively.ResultsSurgical intervention for atrial fibrillation was not indicated in 3 patients in whom the atrial electrograms had a low voltage over a broad area. Concurrent, multiple, and repetitive activations arising from the pulmonary veins or left atrial appendage were observed in all patients. A simple left atrial procedure consisting of pulmonary vein isolation and left atrial incisions without any right atrial incisions was performed in 8 patients in whom the right atrial activation was passive, and all (100%) were cured of atrial fibrillation. The radial procedure was performed in the remaining 35 patients, and 31 (89%) of the patients were cured of atrial fibrillation. In this subset of patients, 10 exhibited reentrant or focal activation in the posterior left atrium between the right and left pulmonary veins and required an additional linear ablation on the posterior left atrium. The total amount of postoperative bleeding after the simple left atrial procedure was significantly less than after the radial procedure (378 ± 135 vs 711 ± 364 mL, P = .03). The right and left atrial transport functions were well preserved after both the radial and simple left atrial procedures.ConclusionIntraoperative mapping facilitates determining the optimal procedure for atrial fibrillation in each patient

RajiおよびK562細胞に対するナマズ卵レクチンと抗腫瘍薬との併用効果

Silurus asotus(catfish)lectin(SAL)is a 32 kDa protein belonging to the rhamnose-binding lectin(RBL)family. SAL also recognizes galactosylα-linked carbohydrate chains of not only glycoproteins but also glycosphingolipids, especially globotriaosylceramide(Gb3). in the previous study, we revealed that SAL increased uptake of doxorubicin(Dox)into Raji cells, and consequently enhanced cytotoxic effect of Dox. we studied whether SAL also enhances the cytotoxic effect of vinblastine and irinotecan in Gb3-expressing Raji or Gb3-negative K562 cells. Treatment of Raji cells with vinblastine(2.5μg/mL)and irinotecan(5μM)for 48 h caused decrease of cell viability to 69% and 68%, respectively. although cytotoxic effects of vinblastine and irinotecan were significantly increased in SAL-pretreated Raji cells, only a slight increase was observed in SAL-pretreated K562 cells with vinblastine. These results indicate that SAL enhances the cytotoxic effect of vinblastine or irinotecan in raji but not K562 cells

レスベラトロール処理細胞における抗腫瘍薬の効果

Chemo-resistance to anti-cancer drugs is a major obstacle in efforts to develop a successful treatment of cancer.Resveratrol(RSV)facilitates the cellular uptake of doxorubicin through an induced down-regulation of multidrug resistance -associated protein-1(MRP1)expression.We studied whether RSV enhances the cytotoxic effect of vinblastine,vincristine,irinotecan,and etoposide in MRP1-expressing Burkitt\u27s lymphoma Raji cells.Treatment with RSV caused decrease of cell viability in a time-and dose-dependent manner,and down-regulation of MRP1 expression in Raji cells.The cytotoxic effects of vinblastine,vincristine,and etoposide increased in RSV-pretreated Raji cells,but the effect of irinotecan did not change in the cells.These results indicate that RSV enhances the cellular uptake of vinblastine,and etoposide in Raji cell

ナマズ卵レクチンのクラスター形成と細胞内輸送

Rhamnose-binding lectins are widely found in fush eggs,and Silurus asotus lectin (SAL) isolated from catfish eggs having three carbohydrate recognition domains preferentially recognizes non-reducing end Galα-linked sugar chain.In the previous study,we revealed that SRL binds to globotriaosylceramide (Gb3) by the surface plasmon resonance analysis.However,its biological effect on cultured cells is still unclear.To investigate localization and trafficking of SAL in the renel adenocarcinoma ACHN cells,which express Gb3 on the cell surface,we prepared HiLyte Fluor 555-labeled SAL (HL-SAL).When ACHN cells were treated with HL-SAL at 4℃ for 5min, and at 37℃ for 24h,HL-SAL was distributed on the cell membrane and in the intracellular compartment,respectively.To trace the trafficking route of HL-SAL from cell surface to the intracellular compartment,the images of HL-SAL-treated live cells were obtained using confocal scanning microscopy.HL-SAL was clustered on ACHN cell surface,and furthermore,partially co-localised with transferrin in intracellular compartment.These results suggest that SAL induces alteration of Gb3 distribution on the membrane and migrates from the cell surface to the intracellular vesicles

キアンコウ(Lophius litulon)におけるレクチン活性の探索

Fish eggs or roe often contain rhamnose-binding lectins (RBLs) that have unique properties among animal lectins in the carbohydrate recognition and domain structure. To investigate new lectin activity in deep-sea fishes,we attempted to extract proteins from eggs (GFE) and liver (GFL) of goose fish,Lophius litulon. The crude saline-extracted fraction (SEF) was obtained from acetone-dried powders of GFE and GFL.Each SEF was separated by DEAE-ion exchange chromatography. Contrary to our expectation,any DEAE fractions had no hemaggulatination activity on rabbit erythrocytes. On the other hand,S-180 mouse ascites tumor cells were strongly aggutinated by GFL D100,D200 and SC fractions with the final concentrations required minimum agglutination,65.0, 98.75 and 112.5ug/mL,respectively. Whereas any typical ligand molecules including carbohydrates and glycoconjugates showed no inhibitory effect for such tumor cell agglutination,several protease inhibitors including phenylmethanesufonyl fluoride and EDTA could do that. agglutination was caused by their proteolytic activity on the tumor cell surface components.The results from additional separation by hydroxyapatite column chromatography,SDS-PAGE and zymography indicated that putative candidates possessing proteolytic activity might be ca. 100 or 45 kDa proteins

Glycosphingolipid-enriched microdomain中でGb3と会合する分子の探索

Silurus asotus (catfish) egg lectin (SAL) is a member of rhamnose-binding lectin family, and binds to globotriaosylceramide (Gb3) localized in a glycosphingolipid-enriched microdomain (GEM) on several tumor cell lines. It is known that GEM is involved in cell adhesion and has the capability of sending signals. We report here about proteins involved in GEM associating with Gb3 and mediating signal transduction system in Burkitt\u27s lymphoma Raji cells via SAL binding to Gb3. Although Gb3 molecule was sprinkled over the surface of Raji cells, SAL uniformly bound to the Raji cell surface. Sucrose density-gradient centrifugation method and fluorescence microscopic analysis revealed that not only CD20 and CD81 but also Lyn and Csk were distributed in GEM of Raji cells. The distribution of these molecules was not altered by the treatment with SAL at 4℃ for 30 ruin. Because CD20 was not co-precipitated with the anti-Gb3 antibody, CD20 was not associated with Gb3 on Raji cells. However, there was a 47kDa protein, which was specifically co-precipitated with anti-Gb3 antibody. These results suggest that the 47kDa protein is a candidate for Gb3-associating protein in Raji cells