Kyoichi Kagawa

Globin digest, acidic protease hydrolysate, inhibits dietary hypertriglyceridemia and Val-Val-Tyr-Pro, one of its constituents, possesses most superior effect

Globin digest (GD), prepared from globin by acidic protease treatment, suppressed the elevation of serum triglyceride level in not only total but also chylomicron fraction after oral administration of olive oil. By screening with this lowering activity, we concluded that Val-Val-Tyr-Pro (VVYP) would be most effective constituent having hypotriglyceridemic action in GD. The mode of their action was dose dependent and did not show species specificity. Neither the repression of peristaltic movement of intestine nor the delaying of gastric emptying was caused by intake of GD or VVYP, however, the excretion of administered lipid was much more than that of control. Furthermore, administration of GD caused more prominent activation of hepatic triglyceride lipase (HTGL) and the increase of hepatic free fatty acid (FFA) concentration in early phase after administration of fat. From these results, it could be elucidated that GD, and also VVYP, inhibited fat absorption from digestive tract and enhanced activity of HTGL, so that more rapid clearance of dietary hypertriglyceridemia was caused.

Anti-diabetic effects of globin digest and its active ingredient Leu-Ser-Glu-Leu in ICR mice, streptozotocin-induced diabetic mice and KK-Ay mice ☆

AIMS Leu-Ser-Glu-Leu (LSEL) is the main active ingredient of globin digest (GD) that has an anti-diabetic effect. Here, we investigated the anti-diabetic effect of LSEL for the first time. MAIN METHODS The anti-diabetic effects of GD and LSEL in ICR mice, streptozotocin (STZ)-induced diabetic mice and KK-Ay mice were examined. KEY FINDINGS GD and LSEL suppressed the elevation of blood glucose in an oral glucose tolerance test (OGTT) in ICR mice, STZ-induced diabetic mice and KK-Ay mice as well as in an oral sucrose tolerance test in ICR mice and in an insulin tolerance test (ITT) in KK-Ay mice. GD and LSEL decreased the blood glucose levels in the basal state in STZ-induced diabetic mice and KK-Ay mice. Furthermore, GD and LSEL elevated the serum insulin levels in an OGTT in ICR mice and KK-Ay mice and promoted the use of insulin in an ITT in KK-Ay mice. GD and LSEL increased the translocation or expression of the glucose transporter 4 in the muscle of ICR mice, STZ-induced diabetic mice and KK-Ay mice and increased the expression of the uncoupling protein 2 (UCP2) in the muscle of ICR mice. SIGNIFICANCE These results indicate that GD and LSEL control blood glucose through the promotion of glucose uptake in the muscle of the mice. The acceleration of glucose uptake by GD and LSEL may be controlled by the promotion of insulin secretion and the up-regulation of UCP2 expression. GD and LSEL seem to be useful for lowering the incidence of hyperglycemia.

Effect of glucocorticoids on calcium binding to plasma membranes of livers during regeneration

Abstract In the plasma membranes of normal and regenerating livers at 3rd, 7th and 10th day after partial hepatectomy, Hofstee plots of calcium(Ca) binding were biphasic. The number of the low affinity binding sites(nmoles/mg membrane protein) in the membranes of regenerating livers decreased from 370 for normal liver to 170 for 3rd day, 220 for 7th day and 460 for 10th day, but their association constants slightly increased. Glucocorticoids, hydrocortisone and fluocinolone, significantly enhanced Ca binding to the membranes of normal and regenerating livers. Especially, their actions to the regenerating livers were effective even on the low concentrations. These results indicate that glucocorticoids may restore the Ca binding in regenerating liver to the normal level.

Effect of globin digest on the liver injury and hepatic gene expression profile in galactosamine-induced liver injury in SD rats.

AIMS We investigated the effect of globin digest (GD) on the liver injury and hepatic gene expression profile in galactosamine (GalN)-induced liver injury. MAIN METHODS The effect of GD on the liver injury was examined by measuring the activities of serum transferases and hepatic antioxidant enzymes, histopathological analysis, gene expression profile, and proteins of the peroxisome proliferator-activated receptor alpha (PPARα) and met proto-oncogene (c-Met) in SD rats at 24 h after GalN administration. The effect of GD on the expression of PPARα and its target gene in AML-12 mouse hepatocytes was also examined. KEY FINDINGS GD suppressed the elevated activities of serum transferases in GalN-induced liver injury in SD rats. The thiobarbituric acid reactive substance content in GalN-injured liver was a decreasing tendency by GD. GD suppressed the increased oxidized glutathione content, and increased the decreased protein, reduced glutathione contents, and catalase activity in GalN-injured liver. GD may improve the antioxidant defense system and protein synthesis in GalN-injured liver. GD suppressed the elevated expression of the genes related to the inflammation, and decreased the histopathological grade value of inflammatory cell infiltration in GalN-injured liver. GD increased the expression of PPARα protein in GalN-injured liver, and also increased the expression of PPARα and its target gene in AML-12 hepatocytes. The total and phosphorylated c-Met proteins in GalN-injured liver were the increasing tendencies by GD. SIGNIFICANCE These findings indicate that GD has the hepatoprotective effect on GalN-induced liver injury in SD rats.

Activation of calcium-ATPase of liver plasma membrane by dextran sulfates.

Dextran sulfate (DS) with average molecular weight (AMW) of 20,000 and sulfur content of 18%, which has a high lipemia clearing activity, enhanced Ca2+ binding to the plasma membrane of rat liver, and the DS itself bound the membrane, whereas there was little binding of DS and Ca2+. Various DSs slightly activated Na+-K+-ATPase, but not Mg2+-ATPase activity of the membrane. These results suggest that DSs, especially with high AMW of 20,000, bind the plasma membrane, resulting in enhancements of the Ca2+ binding to there and Ca2+-ATPase activity.

Effects of globin digest and its active ingredient Trp-Thr-Gln-Arg on galactosamine/lipopolysaccharide-induced liver injury in ICR mice.

AIMS We investigated the effects of globin digest (GD) and its active ingredient Trp-Thr-Gln-Arg (WTQR) on galactosamine/lipopolysaccharide (GalN/LPS)-induced liver injury in imprinting control region (ICR) mice. MAIN METHODS The effects of WTQR and GD on the liver injury were examined by measuring the survival rate, serum aminotransferase activities, hepatic components, antioxidant enzyme activities, histopathological analysis, serum levels and hepatic gene expression of tumor necrosis factor-alpha (TNF-α), macrophage inflammatory protein-2 (MIP-2), and nitric oxide (NO) or inducible nitric oxide synthase (iNOS), and nuclear factor-kappa B (NF-κB) p65 content in GalN/LPS-treated ICR mice. RAW264 mouse macrophages were used to confirm the anti-inflammatory effects of WTQR and GD on the macrophages. KEY FINDINGS WTQR and GD increased the survival rate, suppressed the serum aminotransferase activities, serum levels and hepatic gene expression of TNF-α, MIP-2, and NO or iNOS, and nuclear NF-κB p65 content in GalN/LPS-treated mice; decreased the oxidized glutathione content, increased the superoxide dismutase activity, and decreased the histopathological grade values of the hepatocyte necrosis and lobular inflammation in GalN/LPS-injured liver; and suppressed the release levels and gene expression of TNF-α, MIP-2, and NO or iNOS, and nuclear NF-κB p65 content in LPS-stimulated RAW264 macrophages. WTQR and GD may improve the antioxidant defense system and inflammatory status in GalN/LPS-injured liver. SIGNIFICANCE These findings indicate that WTQR and GD have hepatoprotective effects on GalN/LPS-induced liver injury in ICR mice.

Transport and metabolism of dextran sulfates and the effect of calcium

Abstract The transport and metabolism of [35S]dextran sulfates (DSs) of various molecular weights and sulfur contents in rat intestinal mucosa, and the effects of calcium thereon, were investigated. The in vitro desulfation of DSs was observed largely in the microsomal fraction and was not influenced by the chain length of the glucosidic bond of DSs. The optimal pH for the desulfation of DS was about 5.0. The depolymerization of DSs prior to the desulfation was not significant even in the lysosomal fraction. The transport of DSs by everted sacs of intestine was increased significantly by the reduction of Ca2+ or the addition of ethylenediamine tetraacetic acid (EDTA) to the medium. After the intraduodenal administration of 10 mg/kg of [35S]DS of molecular weight 200,000 and sulfur content 18 per cent, the radioactivity in the lysosomal fraction was about 30 per cent of that in mucosal tissues, and decreased to 8 per cent with the simultaneous administration with EDTA (50 mg/kg). The radioactivity in the soluble fraction was also decreased. Lysosomal ATPase was activated by the DS or Ca2+, and inhibited by the simultaneous administration of EDTA. These findings suggest that DSs are transferred by pinocytosis and other mechanisms and are then desulfated mainly in microsomes followed by depolymerization. Further, Ca2+ may be necessary for the access of DSs to lysosomes which are involved in the metabolism of acid polysaccharides.