Suppressive effects of processed aconite root on dexamethasone-induced muscle ring finger protein-1 expression and its active ingredients.

Processed aconite root (PA), the tuberous root of Aconitum carmichaelii prepared by autoclaving, is a crude drug used in Japanese traditional Kampo medicine and traditional Chinese medicine for the symptoms of kidney deficiency, that is related to the muscle atrophy in modern medicine. The objective of the present study is to evaluate the effectiveness of PA on muscle atrophy and to find its active ingredients using dexamethasone-induced muscle ring finger protein-1 (MuRF1) mRNA expression in murine myoblast C2C12 cells. Dexamethasone-induced MuRF1 expression was significantly suppressed by methanol-soluble part of boiling water extract of PA in a concentration-dependent manner with its IC50 value of 1.5 mg/ml. By the activity-guided fractionations of PA extract using the partition between organic solvents and its aqueous solution, the activity of PA did not transfer into the fraction containing aconitine-type diterpenoid alkaloids but into BuOH layer. Then, we found higenamine and salsolinol as the active ingredients in PA. Higenamine and salsolinol significantly suppressed dexamethasone-induced MuRF1 expression, and their IC50 values were 0.49 and 50 µM, respectively. The contents of higenamine and salsolinol in the decoctions of commercially available fourteen PA products are 0.12 and 14 µg/ml as the average values, and varied with the coefficient of variation (CV) values of 97 and 63%, respectively. Higenamine also significantly suppressed dexamethasone-induced mRNA expressions of muscle atrophy F-box protein (MAFbx)/atrogin1, casitas B-lineage lymphoma-b (Cbl-b), troponin, branched-chain amino acid aminotransferase 2 (BCAT2), and Bcl-2 binding and pro-apoptotic protein3 (Bnip3). Although the quality control of PA is regulated by the contents of diterpene alkaloids, salsolinol and higenamine can be used as the marker compounds to certificate the pharmacological activities of PA.

Maternal fish oil supplementation ameliorates maternal high-fructose diet-induced dyslipidemia in neonatal mice with suppression of lipogenic gene expression in livers of postpartum mice.

Maternal fructose consumption during pregnancy and lactation is associated with metabolic dysregulation in offspring. We tested the hypothesis that fish oil (FO) supplementation during pregnancy and lactation improves fructose-induced metabolic dysregulation in postpartum dams and offspring mice. We therefore aimed to determine the effects of FO supplementation on metabolic disruption in neonatal mice and dams induced by a maternal high-fructose diet (HFrD). The weight of the offspring of dams fed with HFrD on postnatal day 5 was significantly low, but this was reversed by adding FO to the maternal diet. Feeding dams with HFrD significantly increased plasma concentrations of triglycerides, uric acid, and total cholesterol, and decreased free fatty acid concentrations in offspring. Maternal supplementation with FO significantly suppressed HFrD-induced hypertriglyceridemia and hyperuricemia in the offspring. Maternal HFrD induced remarkable mRNA expression of the lipogenic genes Srebf1, Fasn, Acc1, Scd1, and Acly in the postpartum mouse liver without affecting hepatic and plasma lipid levels. Although expression levels of lipogenic genes were higher in the livers of postpartum dams than in those of nonmated mice, HFrD feeding increased the hepatic lipid accumulation in nonmated mice but not in postpartum dams. These findings suggest that although hepatic lipogenic activity is higher in postpartum dams than nonmated mice, the lipid consumption is enhanced in postpartum dams during pregnancy and lactation. Maternal FO supplementation obviously suppressed the expression of these lipogenic genes. These findings coincide with reduced plasma triglyceride concentrations in the offspring. Therefore, dietary FO apparently ameliorated maternal HFrD-induced dyslipidemia in offspring by suppressing maternal lipogenic gene expression and/or neonatal plasma levels of uric acid.

Flavones Inhibit LPS-Induced Atrogin-1/MAFbx Expression in Mouse C2C12 Skeletal Myotubes.

Muscle atrophy is a complex process that occurs as a consequence of various stress events. Muscle atrophy-associated genes (atrogenes) such as atrogin-1/MAFbx and MuRF-1 are induced early in the atrophy process, and the increase in their expression precedes the loss of muscle weight. Although antioxidative nutrients suppress atrogene expression in skeletal muscle cells, the inhibitory effects of flavonoids on inflammation-induced atrogin-1/MAFbx expression have not been clarified. Here, we investigated the inhibitory effects of flavonoids on lipopolysaccharide (LPS)-induced atrogin-1/MAFbx expression. We examined whether nine flavonoids belonging to six flavonoid categories inhibited atrogin-1/MAFbx expression in mouse C2C12 myotubes. Two major flavones, apigenin and luteolin, displayed potent inhibitory effects on atrogin-1/MAFbx expression. The pretreatment with apigenin and luteolin significantly prevented the decrease in C2C12 myotube diameter caused by LPS stimulation. Importantly, the pretreatment of LPS-stimulated myoblasts with these flavones significantly inhibited LPS-induced JNK phosphorylation in C2C12 myotubes, resulting in the significant suppression of atrogin-1/MAFbx promoter activity. These results suggest that apigenin and luteolin, prevent LPS-mediated atrogin-1/MAFbx expression through the inhibition of the JNK signaling pathway in C2C12 myotubes. Thus, these flavones, apigenin and luteolin, may be promising agents to prevent LPS-induced muscle atrophy.

Isoflavones derived from soy beans prevent MuRF1-mediated muscle atrophy in C2C12 myotubes through SIRT1 activation.

Proinflammatory cytokines are factors that induce ubiquitin-proteasome-dependent proteolysis in skeletal muscle, causing muscle atrophy. Although isoflavones, as potent antioxidative nutrients, have been known to reduce muscle damage during the catabolic state, the non-antioxidant effects of isoflavones against muscle atrophy are not well known. Here we report on the inhibitory effects of isoflavones such as genistein and daidzein on muscle atrophy caused by tumor necrosis factor (TNF)-α treatment. In C2C12 myotubes, TNF-α treatment markedly elevated the expression of the muscle-specific ubiquitin ligase MuRF1, but not of atrogin-1, leading to myotube atrophy. We found that MuRF1 promoter activity was mediated by acetylation of p65, a subunit of NFκB, a downstream target of the TNF-α signaling pathway; increased MuRF1 promoter activity was abolished by SIRT1, which is associated with deacetylation of p65. Of interest, isoflavones induced expression of SIRT1 mRNA and phosphorylation of AMP kinase, which is well known to stimulate SIRT1 expression, although there was no direct effect on SIRT1 activation. Moreover, isoflavones significantly suppressed MuRF1 promoter activity and myotube atrophy induced by TNF-α in C2C12 myotubes. These results suggest that isoflavones suppress myotube atrophy in skeletal muscle cells through activation of SIRT1 signaling. Thus, the efficacy of isoflavones could provide a novel therapeutic approach against inflammation-related muscle atrophy.