Fish oil suppresses obesity more potently in lean mice than in diet-induced obese mice but ameliorates steatosis in such obese mice.

This study sought to clarify the antiobesity effects of fish oil (FO) in terms of prevention and amelioration. An isocaloric diet composed of lard or FO was given to lean C57BL/6J mice for the study of prevention and high-fat diet-induced obese (DIO) mice for the study of amelioration for 4 weeks. Body weight gain and food efficiency were potently suppressed by FO in lean mice compared to lard diet-fed mice. Uncoupling protein-1 (UCP-1) expression in inguinal white adipose tissue (WAT) was also significantly induced by FO in lean mice. FO also suppressed body weight gain and food efficiency in DIO mice but did not reduce body weight. FO ameliorated liver steatosis in DIO mice by mildly inducing UCP-1 in inguinal WAT. FO suppressed obesity more potently in lean mice than in DIO mice but ameliorated steatosis in the DIO mice.

The role of increased FGF21 in VLDL-TAG secretion and thermogenic gene expression in mice under protein malnutrition.

Protein malnutrition promotes hepatic lipid accumulation in growing animals. In these animals, fibroblast growth factor 21 (FGF21) rapidly increases in the liver and circulation and plays a protective role in hepatic lipid accumulation. To investigate the mechanism by which FGF21 protects against liver lipid accumulation under protein malnutrition, we determined whether upregulated FGF21 promotes the thermogenesis or secretion of very-low-density lipoprotein (VLDL)-triacylglycerol (TAG). The results showed that protein malnutrition decreased VLDL-TAG secretion, but the upregulation of FGF21 did not oppose this effect. In addition, protein malnutrition increased expression of the thermogenic gene uncoupling protein 1 in inguinal white adipose and brown adipose tissue in an FGF21-dependent manner. However, surgically removing inguinal white adipose tissue did not affect liver triglyceride levels in protein-malnourished mice. These data suggest that FGF21 stimulates thermogenesis under protein malnutrition, but this is not the causative factor underlying the protective role of FGF21 against liver lipid accumulation.

Effects of pharmacological inhibition of plasminogen binding on liver regeneration in rats.

The fibrinolysis system is thought to play an important role in liver regeneration. We previously found that plasminogen (Plg) is localized to the cell surface of regenerating liver tissue as well as proliferating hepatocytes in vitro. Here, we investigated the significance of Plg binding to the cell surface during liver regeneration. Pre-administration of tranexamic acid (TXA), which is a competitive inhibitor of Plg binding, to hepatectomized rats mildly delayed restoration of liver weight in vivo. Although binding of Plg to the cell membrane decreased following TXA administration, TXA showed little effect on hepatocyte proliferation in rats. We also discovered that Plg treatment did not stimulate proliferation of primary rat hepatocytes in vitro. These results suggest that Plg/plasmin potentiates liver regeneration via a pathway distinct from those through which hepatocyte proliferation is stimulated.

The garlic-derived organosulfur compound diallyl trisulphide suppresses tissue factor function.

Tissue factor (TF) is a critical initiator of extrinsic coagulation that sometimes causes thromboembolism. Diallyl trisulphide (DATS) is a secondary metabolite of allicin generated in crushed garlic, with various pharmacological effects. This study aimed to clarify the effect of DATS on the extrinsic coagulation elicited by TF and arteriosclerosis. TF activity was measured using a clotting assay in TF-expressing HL60 cells. DATS inhibited TF activity in a dose-dependent manner. TF expression in TNF-α-stimulated human umbilical vein endothelial cells was examined using real-time PCR and western blotting. DATS inhibited TF mRNA and protein expression induced by TNF-α via inhibition of JNK signalling. The effect of DATS on arteriosclerosis was also examined in apolipoprotein E-deficient mice. DATS administration in these mice tended to decrease atherosclerotic lesion size. These results strongly suggest that DATS prevents thromboembolism triggered by atherosclerosis via the inhibition of plaque formation and TF function.

Very low-density lipoprotein receptor increases in a liver-specific manner due to protein deficiency but does not affect fatty liver in mice.

Very low-density lipoprotein receptor (VLDLR) is a member of the LDL receptor family that is involved in the uptake of VLDL into cells. Increased hepatic VLDLR under endoplasmic reticulum (ER) stress has been shown to cause fatty liver. In this study, the effect of dietary protein restriction on hepatic VLDLR and the role of VLDLR in fatty liver were investigated using Vldlr knockout (KO) mice. Growing wild-type (WT) and KO mice were fed a control diet containing 20% ​​protein or a low protein diet containing 3% protein for 11 days. In WT mice, the amount of hepatic Vldlr mRNA and VLDLR protein increased by approximately 8- and 7-fold, respectively, due to protein restriction. Vldlr mRNA and protein levels increased in both type 1 and type 2 VLDLR. However, neither Vldlr mRNA nor protein levels were significantly increased in heart, muscle, and adipose tissue, demonstrating that VLDLR increase due to protein restriction occurred in a liver-specific manner. Increased liver triglyceride levels during protein restriction occurred in KO mice to the same extent as in WT mice, indicating that increased VLDLR during protein restriction was not the main cause of fatty liver, which was different from the case of ER stress.

Diallyl Trisulfide Prevents Obesity and Decreases miRNA-335 Expression in Adipose Tissue in a Diet-Induced Obesity Rat Model.

SCOPE: Diallyl trisulfide (DATS), an organosulfur compound generates in crushed garlic, has various beneficial health effects. A growing body of evidence indicates that miRNAs are involved in the pathology of lifestyle diseases including obesity. The anti-obesogenic effect of garlic is previously reported; however, the effects of DATS on obesity, and the relationship between garlic compounds and the involvement of miRNA remains unclear. Here, the anti-obesogenic activity of DATS and the potential role of miRNA in a diet-induced obesity rat model are investigated. METHODS AND RESULTS: Oral administration of DATS suppressed body and white adipose tissue (WAT) weight gain in rats fed a high-fat diet compared with vehicle-administered rats. DATS lowered the plasma and liver triglyceride levels in obese rats, and decreased lipogenic mRNA levels including those of Srebp1c, Fasn, and Scd1 in the liver. DATS also suppressed de novo lipogenesis in the liver. Transcriptomic analyses of miRNA and mRNA in the epididymal WAT of obese rats using microarrays revealed that DATS decreased miRNA-335 expression and normalized the obesity-related mRNA transcriptomic signatures in epididymal WAT. CONCLUSION: The potent anti-obesogenic effects of DATS and its possible mechanism of action was clearly demonstrated in this study.

Garlic oil suppresses high-fat diet induced obesity in rats through the upregulation of UCP-1 and the enhancement of energy expenditure.

Garlic (Allium sativum L.) has long been used as a medicinal food. Indeed, garlic and its constituents have been shown to possess potent regulatory activities in bodily functions, including blood coagulation, lipid metabolism, immunity and xenobiotic metabolism. In this study, we aimed to examine the anti-obesity effects of garlic oil and to elucidate the possible underlying mechanisms. For this purpose, garlic oil (GO; 80 mg/kg body weight, p.o.) or corn oil alone as a vehicle-control were administered to male Sprague-Dawley rats every other day for 10 weeks. The results revealed that GO administration significantly reduced body weight gain and white adipose tissue (WAT) mass, which had been increased by feeding on the AIN-76-based high-fat diet (60% kcal fat). Expired gas analysis was performed at 9 weeks following the GO administration to calculate fuel oxidation. GO administration enhanced O2 consumption during the dark period (at night) and increased energy expenditure through fat oxidation during the light period (daytime); however, carbohydrate oxidation remained unaltered. Western blot analysis revealed that GO administration increased UCP1 protein expression in brown adipose tissue (BAT). On the whole, the findings of this study indicated that GO suppressed body weight gain and WAT mass in the rat model of high-fat diet-induced obesity by increasing UCP1 expression and by enhancing fat oxidation and energy expenditure.

Diallyl Trisulfide Inhibits Platelet Aggregation through the Modification of Sulfhydryl Groups.

Diallyl trisulfide (DATS) is a secondary metabolite of allicin, a volatile organosulfur flavoring compound generated by the crushing of garlic. These compounds have various medicinal effects such as antiplatelet activity. In this study, we demonstrated for the first time the cellular mechanism involved in the inhibition of platelet aggregation by DATS and dipropyl trisulfide (DPTS), which is a saturated analogue of DATS. Washed murine platelets were incubated with these sulfides, and platelet aggregation was evaluated by light transmission aggregometry. The amount of reaction products produced by DATS, DPTS, and glutathione (GSH) was measured using liquid chromatography-mass spectrometry. Compared with DPTS, DATS potently inhibited platelet aggregation induced by thrombin, U46619, and collagen. N-Ethylmaleimide (NEM), which is commonly used to modify sulfhydryl groups, also suppressed platelet aggregation. The reactivity of DATS with GSH was higher than that of DPTS. These data suggested that DATS inhibited platelet aggregation through the reaction of sulfhydryl groups.

Dietary protein restriction increases hepatic leptin receptor mRNA and plasma soluble leptin receptor in male rodents.

Leptin is an adipokine that regulates adipose tissue mass through membrane-anchored leptin receptor (Ob-R). Extracellular domain of Ob-R in plasma is called soluble leptin receptor (sOb-R), and is the main leptin-binding protein. Based on a previous DNA microarray analysis that showed induction of hepatic Ob-R mRNA in low-protein diet-fed mice, this study aimed to clarify the effect of dietary protein restriction on hepatic Ob-R mRNA and plasma sOb-R levels. First, the effect of protein restriction on hepatic Ob-R mRNA level was examined together with fasting and food restriction using male rats as common experimental model for nutritional research. Hepatic Ob-R mRNA level was increased by feeding low-protein diet for 7 d, although not significantly influenced by 12-h fasting and sixty percent restriction in food consumption. Then, effect of protein restriction on liver Ob-R and plasma sOb-R was investigated using male mice because specific sOb-R ELISA was more available for mice. Hepatic Ob-R mRNA level was also increased in protein restricted-mice although it did not increase in hypothalamus. Hepatic Ob-R protein was decreased, whereas plasma sOb-R was increased by protein restriction. Because the concentration of sOb-R increased without changing plasma leptin concentration, free leptin in plasma was significantly reduced. The direct effect of amino acid deprivation on Ob-R mRNA level was not observed in rat hepatoma cells H4IIE cultured in amino acid deprived medium. In conclusion, dietary protein restriction increased hepatic Ob-R mRNA, resulting in increased plasma sOb-R concentration, which in turn, reduces plasma free leptin level and may modulate leptin activity.

Branched-chain amino acid supplementation restores reduced insulinotropic activity of a low-protein diet through the vagus nerve in rats.

BACKGROUND: Previously, we reported that a low-protein diet significantly reduced insulin secretion in response to feeding within 1 h in rats, suggesting that the insulinotropic effect of dietary protein plays an important role in maintaining normal insulin release. The current study aimed to elucidate whether deficiency of certain amino acids could diminish the insulinotropic activity and to investigate whether reduced insulin secretion in response to a low-protein diet is restored by supplementation with certain amino acids. METHODS: First, we fed male Wistar rats (5-6 rats per group) with diets deficient in every single amino acid or three branched-chain amino acids (BCAAs); within 1-2 h after the onset of feeding, we measured the plasma insulin levels by using an enzyme-linked immunosorbent assay (ELISA). As insulin secretion was reduced in BCAA-deficient groups, we fed low-protein diets supplemented with BCAAs to assess whether the reduced insulin secretion was restored. In addition, we treated the pancreatic beta cell line MIN6 with BCAAs to investigate the direct insulinotropic activity on beta cells. Lastly, we investigated the effect of the three BCAAs on sham-operated or vagotomized rats to assess involvement of the vagus nerve in restoration of the insulinotropic activity. RESULTS: Feeding a low-protein diet reduced essential amino acid concentrations in the plasma during an absorptive state, suggesting that reduced plasma amino acid levels can be an initial signal of protein deficiency. In normal rats, insulin secretion was reduced when leucine, valine, or three BCAAs were deficient. Insulin secretion was restored to normal levels by supplementation of the low-protein diet with three BCAAs, but not by supplementation with any single BCAA. In MIN6 cells, each BCAA alone stimulated insulin secretion but the three BCAAs did not show a synergistic stimulatory effect. The three BCAAs showed a synergistic stimulatory effect in sham-operated rats but failed to stimulate insulin secretion in vagotomized rats. CONCLUSIONS: Leucine and valine play a role in maintaining normal insulin release by directly stimulating beta cells, and supplementation with the three BCAAs is sufficient to compensate for the reduced insulinotropic activity of the low-protein diet, through the vagus nerve.