Design, synthesis, and biological evaluation of novel tetrahydroprotoberberine derivatives to reduce SREBPs expression for the treatment of hyperlipidemia.

Statins play an important role in the treatment of hyperlipidemia, but drug resistance and adverse effects greatly limits their application. To discover new lipid-lowering drugs, three different series of tetrahydroprotoberberine derivatives (THPBs) were designed and synthesized. These compounds were first tested for their effects on viability of HepG2 cells and 21 compounds with the percent of cell viability over 90% were further screened to evaluate their ability to reduce total cholesterol (TC) and triglyceride (TG) levels. Among these derivatives, two compounds displayed significant down-regulation both intracellular of TC and TG content, especially compound 49 exhibited the greatest efficacy. Mechanistically, compound 49 promoted proteasomal degradation of SREBPs. Importantly, compound 49 displayed superior bioavailability (F = 65.1%) and obvious efficacy in the treatment of high fat diet induced obesity in vivo. Therefore, compound 49 is a promising candidate to develop new treatment of hyperlipidemia.

Pseudoprotodioscin inhibits SREBPs and microRNA 33a/b levels and reduces the gene expression regarding the synthesis of cholesterol and triglycerides.

The extract of Dioscorea zingiberensis C.H. Wright rhizomes is found to be effective in the therapy of cardiovascular disease. Steroidal saponins make substantial contribution. Previous study has proposed that methylprotodioscin (MP) may promote cholesterol efflux by increasing ABCA1 expression. But the other main saponins ingredients are not referred to. The aim of the present work was to reveal the effect and mechanism of protodioscin (PD), MP and pseudoprotodioscin (PPD) on the synthesis-related gene expression of cholesterol and triglycerides. MTT assay apoptosis assay with annexin AV-APC and 7-AAD double staining were performed. MicroRNA assay and qRT-PCR were used to analyze the gene expression which regulates synthesis of cholesterol and triglycerides. Western blot was to demonstrate the levels of target proteins. Cholesterol efflux assay was executed to study the stimulative effect of saponins on cholesterol efflux. In Hep G2 cells, PPD increased ABCA1 protein and mRNA levels, and promoted the effluxion of ApoA-1-mediated cholesterol. The underlying mechanisms involved that PPD inhibited SREBP1c and SREBP2 transcription by decreasing microRNA 33a/b levels. This procedure reciprocally led to the increase of ABCA1 levels. In THP-1 macrophages, PPD showed the similar effect, which reduced HMGCR, FAS and ACC mRNA levels and promoted low density lipoprotein receptor by decreasing the PCSK9 levels. These studies demonstrated that PPD is a potential agent for cholesterol efflux, SREBPs and microRNA 33a/b inhibition, which related to the gene expression for the synthesis of cholesterol and triglycerides.

A lipid-free and insulin-supplemented medium supports De Novo fatty acid synthesis gene activation in melanoma cells.

While investigating the role played by de novo lipid (DNL) biosynthesis in cancer cells, we sought a medium condition that would support cell proliferation without providing any serum lipids. Here we report that a defined serum free cell culture medium condition containing insulin, transferrin and selenium (ITS) supports controlled study of transcriptional regulation of de novo fatty acid (DNFA) production and de novo cholesterol synthesis (DNCS) in melanoma cell lines. This lipid-free ITS medium is able to support continuous proliferation of several melanoma cell lines that utilize DNL to support their lipid requirements. We show that the ITS medium stimulates gene transcription in support of both DNFA and DNCS, specifically mediated by SREBP1/2 in melanoma cells. We further found that the ITS medium promoted SREBP1 nuclear localization and occupancy on DNFA gene promoters. Our data show clear utility of this serum and lipid-free medium for melanoma cancer cell culture and lipid-related areas of investigation.

Electroacupuncture ameliorates poloxamer 407-induced hyperlipidemia through suppressing hepatic SREBP-2 expression in rats.

AIMS: Acupuncture, particularly electroacupuncture (EA) has been shown to have the lipid-lowering effects, but not completely investigated. The present study was aimed to examine whether EA could attenuate poloxamer-407 (P-407)-induced hyperlipidemia in the rats and to investigate its potential mechanisms. MAIN METHODS: Rats received P-407 (0.4 g/kg, i.p.) to induce hyperlipidemia. EA was performed at ST36 and ST40 acupoints a total of three times with 12 h-interval starting 1 h before the P-407 injection at 0.6 mA intensity and 2 Hz frequency for 10 min. KEY FINDINGS: In P-407-induced hyperlipidemic rats, EA stimulation at ST36 and ST40 acupoints significantly lowered the serum levels of triglycerides, total cholesterol, LDL-cholesterol and atherogenic index, while markedly increasing the serum HDL-cholesterol levels. Meanwhile, hyperlipidemic rats had significantly higher expression of sterol regulatory element-binding protein (SREBP)-2, without any difference in SREBP-1 expression in the liver, as compared with normal ones. EA significantly attenuated the expression of SREBP-2 with a subsequent decrease in 3-hydroxy-3-methylglutaryl coenzyme A reductase and an increase in low-density lipoprotein receptor at both mRNA and protein levels in the liver of hyperlipidemic rats. These changes did not occur after electrical stimulation at a non-acupoint. SIGNIFICANCE: Taken together, our findings indicate that EA stimulation to P-407-induced hyperlipidemic rats improves the lipid abnormalities, which may be associated with regulation of the expression of key enzymes of cholesterol synthesis in the liver through modulation of SREBP-2.

An obesogenic diet enriched with blue mussels protects against weight gain and lowers cholesterol levels in C57BL/6 mice.

Obesity is linked to several health complications, such as cardiovascular disease, insulin resistance, and hypertension. Dyslipidemia in obesity is one of the prime causes for health complications. We have previously shown that blue mussels (BM) are a rich source of omega (n)-3 polyunsaturated fatty acids (PUFA) and increase the mRNA expression of peroxisome-proliferator activated receptor and adiponectin, thereby inducing anti-obesity and insulin sensitizing effects in vitro. However, the in vivo effects of BM on obesity and metabolic regulation are not known. We hypothesized that dietary intake of BM will prevent weight gain and improve lipid profile of C57BL/6 mice fed a high-fat diet (HFD). Mice were fed a HFD supplemented with 5% w/w BM (BM-HFD) for 4 weeks, and then switched to a HFD for 4 weeks. Mice fed a BM-HFD showed significantly lower body weight gain and abdominal fat, compared to the HFD. Furthermore, a BM-HFD significantly reduced plasma and hepatic total and low-density lipoprotein (LDL)-cholesterol, compared to HFD. The decrease in cholesterol levels coincided with inhibition of hepatic sterol regulatory element-binding protein-2 and HMG-CoA reductase mRNA expression, and an increase in LDL-receptor gene expression in the BM-HFD group, compared to the HFD group. In conclusion, our findings have established that BM reduces body weight gain in mice. BM may have potential to lower cholesterol levels by inhibiting cholesterol synthesis, thereby protecting against obesity and perhaps heart disease.

Synergic hypocholesterolaemic effect of n-3 PUFA and oestrogen by modulation of hepatic cholesterol metabolism in female rats.

n-3 PUFA such as EPA and DHA as well as oestrogen have been reported to decrease blood levels of cholesterol, but their underlying mechanism is unclear. The purpose of this study was to determine the effects of the combination of n-3 PUFA supplementation and oestrogen injection on hepatic cholesterol metabolism. Rats were fed a modified AIN-93G diet with 0, 1 or 2 % n-3 PUFA (EPA+DHA) relative to the total energy intake for 12 weeks. Rats were surgically ovariectomised at week 8, and, after 1-week recovery, rats were injected with 17ß-oestradiol-3-benzoate (E2) or maize oil for the last 3 weeks. Supplementation with n-3 PUFA and E2 injection significantly increased the ratio of the hepatic expression of phosphorylated AMP activated protein kinase (p-AMPK):AMP activated protein kinase (AMPK) and decreased sterol regulatory element-binding protein-2, 3-hydroxy-3-methylglutaryl coenzyme A reductase and proprotein convertase subtilisin/kexin type 9. Supplementation with n-3 PUFA increased hepatic expression of cholesterol 7α-hydroxylase (CYP7A1), sterol 12α-hydroxylase (CYP8B1) and sterol 27-hydroxylase (CYP27A1); however, E2 injection decreased CYP7A1 and CYP8B1 but not CYP27A1. Additionally, E2 injection increased hepatic expression of oestrogen receptor-α and ß. In conclusion, n-3 PUFA supplementation and E2 injection had synergic hypocholesterolaemic effects by down-regulating hepatic cholesterol synthesis (n-3 PUFA and oestrogen) and up-regulating bile acid synthesis (n-3 PUFA) in ovariectomised rats.

Polyphenol-rich black chokeberry (Aronia melanocarpa) extract regulates the expression of genes critical for intestinal cholesterol flux in Caco-2 cells.

Black chokeberry (Aronia melanocarpa) is a rich source of polyphenols. The hypolipidemic effects of polyphenol-rich black chokeberry extract (CBE) have been reported, but underlying mechanisms have not been well characterized. We investigated the effect of CBE on the expression of genes involved in intestinal lipid metabolism. Caco-2 cells were incubated with 50 or 100 µg/ml of CBE for 24 h for quantitative realtime polymerase chain reaction analysis. Expression of genes for cholesterol synthesis (3-hydroxy-3-methylglutaryl coenzyme A reductase and sterol regulatory element binding protein 2), apical cholesterol uptake (Niemann-Pick C1 Like 1 and scavenger receptor class B Type 1) and basolateral cholesterol efflux [ATP-binding cassette transporter A1 (ABCA1)] was significantly decreased by CBE compared with control. Western blot analysis confirmed that CBE inhibited expression of these proteins. In contrast, CBE markedly induced mRNA and/or protein levels of ABCG5 and ABCG8 that mediate apical cholesterol efflux to the intestinal lumen. Furthermore, CBE significantly increased mRNA and protein levels of low-density lipoprotein (LDL) receptor, and cellular LDL uptake. Expression of genes involved in lipid metabolism and lipoprotein assembly, including sterol regulatory element-binding protein 1c, fatty acid synthase and acyl-CoA oxidase 1, was significantly decreased by CBE in a dose-dependent manner. Concomitantly, CBE significantly increased sirtuin 1, 3 and 5 mRNA levels, while it decreased SIRT-2. Our data suggest that hypolipidemic effects of CBE may be attributed, at least in part, to increased apical efflux of LDL-derived cholesterol and to decreased chylomicron formation in the intestine; and specific isoforms of SIRT may play an important role in this process.