Nicotinate-curcumin improves NASH by inhibiting the AKR1B10/ACCα-mediated triglyceride synthesis.

BACKGROUND: Nonalcoholic steatohepatitis (NASH) is a prevalent chronic liver condition. However, the potential therapeutic benefits and underlying mechanism of nicotinate-curcumin (NC) in the treatment of NASH remain uncertain. METHODS: A rat model of NASH induced by a high-fat and high-fructose diet was treated with nicotinate-curcumin (NC, 20, 40 mg·kg- 1), curcumin (Cur, 40 mg·kg- 1) and metformin (Met, 50 mg·kg- 1) for a duration of 4 weeks. The interaction between NASH, Cur and Aldo-Keto reductase family 1 member B10 (AKR1B10) was filter and analyzed using network pharmacology. The interaction of Cur, NC and AKR1B10 was analyzed using molecular docking techniques, and the binding energy of Cur and NC with AKR1B10 was compared. HepG2 cells were induced by Ox-LDL (25 µg·ml- 1, 24 h) in high glucose medium. NC (20µM, 40µM), Cur (40µM) Met (150µM) and epalrestat (Epa, 75µM) were administered individually. The activities of ALT, AST, ALP and the levels of LDL, HDL, TG, TC and FFA in serum were quantified using a chemiluminescence assay. Based on the changes in the above indicators, score according to NAS standards. The activities of Acetyl-CoA and Malonyl-CoA were measured using an ELISA assay. And the expression and cellular localization of AKR1B10 and Acetyl-CoA carboxylase (ACCα) in HepG2 cells were detected by Western blotting and immunofluorescence. RESULTS: The results of the animal experiments demonstrated that NASH rat model induced by a high-fat and high-fructose diet exhibited pronounced dysfunction in liver function and lipid metabolism. Additionally, there was a significant increase in serum levels of FFA and TG, as well as elevated expression of AKR1B10 and ACCα, and heightened activity of Acetyl-CoA and Malonyl-CoA in liver tissue. The administration of NC showed to enhance liver function in rats with NASH, leading to reductions in ALT, AST and ALP levels, and decrease in blood lipid and significant inhibition of FFA and TG synthesis in the liver. Network pharmacological analysis identified AKR1B10 and ACCα as potential targets for NASH treatment. Molecular docking studies revealed that both Cur and NC are capable of binding to AKR1B10, with NC exhibiting a stronger binding energy to AKR1B10. Western blot analysis demonstrated an upregulation in the expression of AKR1B10 and ACCα in the liver tissue of NASH rats, accompanied by elevated Acetyl-CoA and Malonyl-CoA activity, and increased levels of FFA and TG. The results of the HepG2 cell experiments induced by Ox-LDL suggest that NC significantly inhibited the expression and co-localization of AKR1B10 and ACCα, while also reduced levels of TC and LDL-C and increased level of HDL-C. These effects are accompanied by a decrease in the activities of ACCα and Malonyl-CoA, and levels of FFA and TG. Furthermore, the impact of NC appears to be more pronounced compared to Cur. CONCLUSION: NC could effectively treat NASH and improve liver function and lipid metabolism disorder. The mechanism of NC is related to the inhibition of AKR1B10/ACCα pathway and FFA/TG synthesis of liver.

Lipids and lipid metabolism in cellular senescence: Emerging targets for age-related diseases.

Cellular senescence is a kind of cellular state triggered by endogenous or exogenous stimuli, which is mainly characterized by stable cell cycle arrest and complex senescence-associated secretory phenotype (SASP). Once senescent cells accumulate in tissues, they may eventually accelerate the progression of age-related diseases, such as atherosclerosis, osteoarthritis, chronic lung diseases, cancers, etc. Recent studies have shown that the disorders of lipid metabolism are not only related to age-related diseases, but also regulate the cellular senescence process. Based on existing research evidences, the changes in lipid metabolism in senescent cells are mainly concentrated in the metabolic processes of phospholipids, fatty acids and cholesterol. Obviously, the changes in lipid-metabolizing enzymes and proteins involved in these pathways play a critical role in senescence. However, the link between cellular senescence, changes in lipid metabolism and age-related disease remains to be elucidated. Herein, we summarize the lipid metabolism changes in senescent cells, especially the senescent cells that promote age-related diseases, as well as focusing on the role of lipid-related enzymes or proteins in senescence. Finally, we explore the prospect of lipids in cellular senescence and their potential as drug targets for preventing and delaying age-related diseases.

[Mechanism of astragaloside â £ in regulating autophagy of PC12 cells under oxygen-glucose deprivation by medicating Akt/mTOR/HIF-1α pathway].

This study explored the protective effect of astragaloside Ⅳ(AS-Ⅳ) on oxygen-glucose deprivation(OGD)-induced autophagic injury in PC12 cells and its underlying mechanism. An OGD-induced autophagic injury model in vitro was established in PC12 cells. The cells were divided into a normal group, an OGD group, low-, medium-, and high-dose AS-Ⅳ groups, and a positive drug dexmedetomidine(DEX) group. Cell viability was measured using the MTT assay. Transmission electron microscopy was used to observe autophagosomes and autolysosomes, and the MDC staining method was used to assess the fluorescence intensity of autophagosomes. Western blot was conducted to determine the relative expression levels of functional proteins LC3-Ⅱ/LC3-Ⅰ, Beclin1, p-Akt/Akt, p-mTOR/mTOR, and HIF-1α. Compared with the normal group, the OGD group exhibited a significant decrease in cell viability(P<0.01), an increase in autophagosomes(P<0.01), enhanced fluorescence intensity of autophagosomes(P<0.01), up-regulated Beclin1, LC3-Ⅱ/LC3-Ⅰ, and HIF-1α(P<0.05 or P<0.01), and down-regulated p-Akt/Akt and p-mTOR/mTOR(P<0.05 or P<0.01). Compared with the OGD group, the low-and medium-dose AS-Ⅳ groups and the DEX group showed a significant increase in cell viability(P<0.01), decreased autophagosomes(P<0.01), weakened fluorescence intensity of autophagosomes(P<0.01), down-regulated Beclin1, LC3-Ⅱ/LC3-Ⅰ, and HIF-1α(P<0.05 or P<0.01), and up-regulated p-Akt/Akt and p-mTOR/mTOR(P<0.01). AS-Ⅳ at low and medium doses exerted a protective effect against OGD-induced autophagic injury in PC12 cells by activating the Akt/mTOR pathway, subsequently influencing HIF-1α. The high-dose AS-Ⅳ group did not show a statistically significant difference compared with the OGD group. This study provides a certain target reference for the prevention and treatment of OGD-induced cellular autophagic injury by AS-Ⅳ and accumulates laboratory data for the secondary development of Astragali Radix and AS-Ⅳ.

Wnt5a/Ror2 promotes vascular smooth muscle cells proliferation via activating PKC.

INTRODUCTION: Abnormal proliferation of vascular smooth muscle cells (VSMCs) can cause various vascular diseases, such as atherosclerosis, restenosis, and pulmonary hypertension. However, the effect and underlying mechanism of Wnt5a on the proliferation of VSMCs remain unclear. Our study aimed to investigate whether Wnt5a/Ror2 promotes vascular smooth muscle cell proliferation via activating protein kinase C (PKC), thereby effectively alleviating vascular proliferative diseases. MATERIAL AND METHODS: The proliferation of HA-VSMC cell line was evaluated by CCK-8, EdU, and Plate clone formation assays. The Wnt5a gene knockdown and overexpression were carried out by standard methods. The interaction between Wnt5a and Ror2 was explored by co-immunoprecipitation. Western blotting and immunofluorescence were used to determine the expression levels of key proteins in VSMCs. RESULTS: The present study found that the expression of Wnt5a protein increased significantly in the proliferation of VSMCs stimulated by 10% serum in a time-dependent manner. Furthermore, the proliferative rate of VSMCs overexpressing Wnt5a was dramatically accelerated, whereas Wnt5a knockdown using siWnt5a reversed thisproliferative effect. Wnt5a up-regulated the expression of receptor tyrosine kinase-like orphan receptor 2 (Ror2) by binding to it. Further studies indicated that Wnt5a induces the PKC expression in VSMCs and knockdown of Wnt5a or Ror2 could inhibit PKC phosphorylation. CONCLUSIONS: Wnt5a could effectively promote the proliferation of VSMCs, which might be related to the binding of Wnt5a and Ror2 to activate PKC.

Ezetimibe and Cancer: Is There a Connection?

The high level of serum cholesterol caused by the excessive absorption of cholesterol can lead to hypercholesteremia, thus promoting the occurrence and development of cancer. Ezetimibe is a drug that reduces cholesterol absorption and has been widely used for the treatment of patients with high circulating cholesterol levels for many years. Mechanistically, ezetimibe works by binding to NPC1L1, which is a key mediator of cholesterol absorption. Accumulating data from preclinical models have shown that ezetimibe alone could inhibit the development and progression of cancer through a variety of mechanisms, including anti-angiogenesis, stem cell suppression, anti-inflammation, immune enhancement and anti-proliferation. In the past decade, there has been heated discussion on whether ezetimibe combined with statins will increase the risk of cancer. At present, more and more evidence shows that ezetimibe does not increase the risk of cancers, which supports the role of ezetimibe in anti-cancer. In this review, we discussed the latest progress in the anti-cancer properties of ezetimibe and elucidated its underlying molecular mechanisms. Finally, we highlighted the potential of ezetimibe as a therapeutic agent in future cancer treatment and prevention.

New dawn for cancer cell death: Emerging role of lipid metabolism.

BACKGROUND: Resistance to cell death, a protective mechanism for removing damaged cells, is a "Hallmark of Cancer" that is essential for cancer progression. Increasing attention to cancer lipid metabolism has revealed a number of pathways that induce cancer cell death. SCOPE OF REVIEW: We summarize emerging concepts regarding lipid metabolic reprogramming in cancer that is mainly involved in lipid uptake and trafficking, de novo synthesis and esterification, fatty acid synthesis and oxidation, lipogenesis, and lipolysis. During carcinogenesis and progression, continuous metabolic adaptations are co-opted by cancer cells, to maximize their fitness to the ever-changing environmental. Lipid metabolism and the epigenetic modifying enzymes interact in a bidirectional manner which involves regulating cancer cell death. Moreover, lipids in the tumor microenvironment play unique roles beyond metabolic requirements that promote cancer progression. Finally, we posit potential therapeutic strategies targeting lipid metabolism to improve treatment efficacy and survival of cancer patient. MAJOR CONCLUSIONS: The profound comprehension of past findings, current trends, and future research directions on resistance to cancer cell death will facilitate the development of novel therapeutic strategies targeting the lipid metabolism.

Celastrol ameliorates vascular neointimal hyperplasia through Wnt5a-involved autophagy.

Neointimal hyperplasia caused by the excessive proliferation of vascular smooth muscle cells (VSMCs) is the pathological basis of restenosis. However, there are few effective strategies to prevent restenosis. Celastrol, a pentacyclic triterpene, has been recently documented to be beneficial to certain cardiovascular diseases. Based on its significant effect on autophagy, we proposed that celastrol could attenuate restenosis through enhancing autophagy of VSMCs. In the present study, we found that celastrol effectively inhibited the intimal hyperplasia and hyperproliferation of VSMCs by inducing autophagy. It was revealed that autophagy promoted by celastrol could induce the lysosomal degradation of c-MYC, which might be a possible mechanism contributing to the reduction of VSMCs proliferation. The Wnt5a/PKC/mTOR signaling pathway was found to be an underlying mechanism for celastrol to induce autophagy and inhibit the VSMCs proliferation. These observations indicate that celastrol may be a novel drug with a great potential to prevent restenosis.

Celastrol suppresses lipid accumulation through LXRα/ABCA1 signaling pathway and autophagy in vascular smooth muscle cells.

The uptake of modified low-density lipoprotein (LDL) and the accumulation of lipid droplets induce the formation of vascular smooth muscle cells (VSMCs)-derived foam cells, thereby promoting the development and maturation of plaques and accelerating the progression of atherosclerosis. Celastrol is a quinine methide triterpenoid isolated from the root bark of traditional Chinese herb Tripterygium wilfordii. It possesses various biological properties, including anti-obesity, cardiovascular protection, anti-inflammation, etc. In the present study, we found that celastrol significantly reduced lipid accumulation induced by oxidized LDL (ox-LDL) in VSMCs. Mechanistically, celastrol up-regulated adenosine triphosphate-binding cassette transporter A1 (ABCA1) expression through activating liver X receptor α (LXRα) expression, which contributed to inhibit lipid accumulation in VSMCs. Meanwhile, celastrol decreased lipid accumulation by triggering autophagy in VSMCs. Therefore, these findings supported celastrol as a potentially effective agent for the prevention and therapy of atherosclerosis.

Effects of menthol and its interaction with nicotine-conditioned cue on nicotine-seeking behavior in rats.

RATIONALE: Increasing clinical evidence suggests that menthol, a significant flavoring additive in tobacco products, may contribute to smoking and nicotine dependence. Relapse to smoking behavior presents a formidable challenge for the treatment of tobacco addiction. An unresolved issue is whether the mentholation of tobacco products precipitates relapse to tobacco use in abstinent smokers. OBJECTIVES: The present study examined the effects of menthol on the perseverance and relapse of nicotine-seeking behavior in rats. METHODS: Male Sprague-Dawley rats were trained to press a lever for intravenous nicotine self-administration (0.03 mg/kg/infusion) under a fixed-ratio five schedule of reinforcement. Each nicotine infusion was signaled by the presentation of a sensory stimulus that was established as a discrete nicotine-conditioned cue. Five minutes prior to the sessions, the rats received an intraperitoneal injection of menthol (0.1 mg/kg) or vehicle. In the subsequent extinction test sessions, nicotine was unavailable with or without menthol and/or the nicotine-conditioned cue. The reinstatement tests were performed the following day after the extinction criterion was met. Menthol was also tested on food-seeking responses. In a subset of nicotine-trained rats, a transient receptor potential melastatin 8 (TRPM8) antagonist RQ-00203078 was given prior to menthol administration. RESULTS: Continued administration of menthol sustained responses on the previously active and nicotine-reinforced lever in the extinction tests. The readministration of menthol after extinction reinstated active lever responses. In both the extinction and the reinstatement tests, a combination of pre-session menthol administration and cue representation during the session produced a more robust behavioral effect than either menthol or the cue alone. No such effects of menthol was observed in food trained rats. RQ-00203078 did not change menthol effect on nicotine seeking. CONCLUSION: These data demonstrated that menthol specifically sustained and reinstated nicotine-seeking behavior, and this effect was independent of TRPM8 activity. These findings suggest that menthol in most tobacco products, even not menthol labeled, may contribute to the perseverance of and relapse to tobacco-seeking behavior.

Enhancing effect of menthol on nicotine self-administration in rats.

RATIONALE: Tobacco smoking is a leading preventable cause of premature death in the USA. Menthol is a significant flavoring additive in tobacco products. Clinical evidence suggests that menthol may promote tobacco smoking and nicotine dependence. However, it is unclear whether menthol enhances the reinforcing actions of nicotine and thus facilitates nicotine consumption. This study employed a rat model of nicotine self-administration to examine the effects of menthol on nicotine-taking behavior. METHODS: Male Sprague-Dawley rats were trained in daily 1-h sessions to press a lever for intravenous nicotine self-administration under a fixed-ratio 5 schedule of reinforcement. In separate groups, rats self-administered nicotine at four different doses (0.0075, 0.015, 0.03, and 0.06 mg/kg/infusion). Five minutes prior to the two test sessions, menthol (5 mg/kg) or its vehicle was administered intraperitoneally in all rats in a counterbalanced design within each group. In separate rats that self-administered 0.015 mg/kg/infusion nicotine, menthol dose-response function was determined. Menthol was also tested on food self-administration. RESULTS: An inverted U-shaped nicotine dose-response curve was observed. Menthol pretreatment shifted the nicotine dose-response curve to the left. The facilitating effect of menthol on the self-administration of 0.015 mg/kg/infusion nicotine was dose-dependent, whereas it produced similar effects at doses above the threshold of 2.5 mg/kg. Menthol tended to suppress the self-administration of food pellets. CONCLUSIONS: These data demonstrate that menthol enhances the reinforcing effects of nicotine, and the effect of menthol was specific to nicotine. The findings suggest that menthol directly facilitates nicotine consumption, thereby contributing to tobacco smoking.

Ezetimibe-mediated protection of vascular smooth muscle cells from cholesterol accumulation through the regulation of lipid metabolism-related gene expression.

BACKGROUND: Ezetimibe is a potent inhibitor of Niemann-Pick type C1-Like 1 and has been approved for the treatment of hypercholesterolemia. Our preliminary study showed that ezetimibe promotes cholesterol efflux from vascular smooth muscle cells (VSMCs). Our aim was to investigate the cellular mechanisms underlying the ezetimibe actions. METHODS AND RESULTS: Rat VSMCs were converted to foam cells by incubation with cholesterol:methyl-ß-cyclodextrin. The intracellular free cholesterol, total cholesterol, and the ratio of cholesteryl ester to total cholesterol were decreased after the incubation of VSMCs with different concentrations of ezetimibe (3, 10, 30, and 30 µmol/l) or treated with 30 µmol/l of ezetimibe for different time periods (6, 12, 24, and 48 h). Our results also showed that the expression of caveolin-1, liver X receptor α, and ATP-binding cassette transporter ABCA1 was enhanced, but the expression of nSREBP-1c was decreased in a concentration- and time-dependent manner. RNA interference was used to determine the roles of caveolin-1 and SREBP-1 in the lipid-lowering effect of ezetimibe. The results showed that caveolin-1 was involved in the regulation of intracellular cholesterol content, and the expression of caveolin-1 was repressed by SREBP-1. CONCLUSION: The present study indicates that ezetimibe protects VSMCs from cholesterol accumulation by regulating the expression of lipid metabolism-related genes.

Mesenchymal stem cell-derived microparticles: a promising therapeutic strategy.

Mesenchymal stem cells (MSCs) are multipotent stem cells that give rise to various cell types of the mesodermal germ layer. Because of their unique ability to home in on injured and cancerous tissues, MSCs are of great potential in regenerative medicine. MSCs also contribute to reparative processes in different pathological conditions, including cardiovascular diseases and cancer. However, many studies have shown that only a small proportion of transplanted MSCs can actually survive and be incorporated into host tissues. The effects of MSCs cannot be fully explained by their number. Recent discoveries suggest that microparticles (MPs) derived from MSCs may be important for the physiological functions of their parent. Though the physiological role of MSC-MPs is currently not well understood, inspiring results indicate that, in tissue repair and anti-cancer therapy, MSC-MPs have similar pro-regenerative and protective properties as their cellular counterparts. Thus, MSC-MPs represent a promising approach that may overcome the obstacles and risks associated with the use of native or engineered MSCs.

Ezetimibe suppresses cholesterol accumulation in lipid-loaded vascular smooth muscle cells in vitro via MAPK signaling.

AIM: To investigate the mechanisms of anti-atherosclerotic action of ezetimibe in rat vascular smooth muscle cells (VSMCs) in vitro. METHODS: VSMCs of SD rats were cultured in the presence of Chol:MßCD (10 µg/mL) for 72 h, and intracellular lipid droplets and cholesterol levels were evaluated using Oil Red O staining, HPLC and Enzymatic Fluorescence Assay, respectively. The expression of caveolin-1, sterol response element-binding protein-1 (SREBP-1) and ERK1/2 were analyzed using Western blot assays. Translocation of SREBP-1 and ERK1/2 was detected with immunofluorescence. RESULTS: Treatment with Chol:MßCD dramatically increased the cellular levels of total cholesterol (TC), cholesterol ester (CE) and free cholesterol (FC) in VSMCs, which led to the formation of foam cells. Furthermore, Chol:MßCD treatment significantly decreased the expression of caveolin-1, and stimulated the expression and nuclear translocation of SREBP-1 in VSMCs. Co-treatment with ezetimibe (3 µmol/L) significantly decreased the cellular levels of TC, CE and FC, which was accompanied by elevation of caveolin-1 expression, and by a reduction of SREBP-1 expression and nuclear translocation. Co-treatment with ezetimibe dose-dependently decreased the expression of phosphor-ERK1/2 (p-ERK1/2) in VSMCs. The ERK1/2 inhibitor PD98059 (50 µmol/L) altered the cholesterol level and the expression of p-ERK1/2, SREBP-1 and caveolin-1 in the same manner as ezetimibe did. CONCLUSION: Ezetimibe suppresses cholesterol accumulation in rat VSMCs in vitro by regulating SREBP-1 and caveolin-1 expression, possibly via the MAPK signaling pathway.

Inhibition of smooth muscle cell proliferation by ezetimibe via the cyclin D1-MAPK pathway.

Proliferation of vascular smooth muscle cells (VSMCs) contributes to the development of atherosclerosis. Ezetimibe is a new lipid lowering agent that inhibits cholesterol absorption. In the present study we attempted to investigate whether ezetimibe has any effect on VSMC proliferation and the potential mechanisms involved. Our data showed ezetimibe abrogated the proliferation and migration of primary rat VSMCs induced by Chol:MßCD. Mechanically, we found that ezetimibe was capable of abolishing cyclin D1, CDK2, phospho-Rb (p-Rb), and E2F protein expressions that were upregulated by Chol:MßCD treatment. In addition, Ezetimibe was able to reverse cell cycle progression induced by Chol:MßCD, which was further supported by its down-regulation of cyclin D1 promoter activity in the presence of Chol:MßCD. Furthermore, ezetimibe abrogated the increment of phospho-ERK1/2 (p-ERK1/2) and nuclear accumulation of ERK1/2 in VSMCs induced by Chol:MßCD. Inhibition of the MAPK pathway by using ERK1/2 inhibitor PD98059 attenuated the reduction effect of ezetimibe on the expressions of phosphor-MEK1 (p-MEK1), p-ERK1/2, and cyclin D1. Taken together our data suggest that ezetimibe inhibits Chol:MßCD-induced VSMCs proliferation and leads to cell cycle arrest at the G0/G1 phase by suppressing cyclin D1 expression via the MAPK signaling pathway. These novel findings support the potential pleiotropic effect of ezetimibe in cardiovascular disease.