Curcumin Upregulates LDL Receptor Expression via Sterol Regulatory Element Pathway in HepG2 Cells

Objective： Plasma low-density lipoprotein-cholesterol （LDL-C） is mainly taken up and cleared by hepatocellular LDL receptor （LDL-R）. LDL-R gene expression is regulated by the sterol regulatory element binding proteins （SREBPs）. Previous studies have shown that curcumin reduces plasma LDL-C and has hypolipidemic and anti-atherosclerotic effects. Herein, the authors investigated the effect of curcumin on LDL-R expression and its molecular mechanism in HepG2 cells.     

Evaluation of Antigenotoxic Effects of Plant Flavonoids Quercetin and Rutin on HepG2 Cells

The flavonoid quercetin and its derivative rutin were investigated for genotoxicity/antigenotoxicity activity in human hepatoma HepG2 cells using the comet assay. The extract cytotoxicity was evaluated using the trypan blue exclusion dye method with quercetin and rutin concentrations ranging from 0.1 to 200.0 μg/mL of culture medium. Three minor non‐cytotoxic concentrations were chosen to evaluate the genotoxicity and antigenotoxicity of the flavonoids (0.1, 1.0 and 5.0 μg/mL) through comet assay. The cultures were treated with three different concentrations of rutin or quercetin (genotoxicity) or their association with Aflatoxin B1 (AFB1), methyl methanesulfonate (MMS) or doxorubicin (DXR) (antigenotoxicity test) in three protocols: pre‐treatment, simultaneous treatment and post‐treatment. The cell cultures were also treated with 1% DMSO (control group), AFB1, MMS and DXR (positive‐control). Statistical analyses were performed using ANOVA and Dunnett's test (p ≤ 0.05). Quercetin at concentrations higher than 10.0 μg/mL or rutin higher than 50.0 μg/mL exhibited a cytotoxic effect on the cells, showing that quercetin is more cytotoxic than rutin. Furthermore, neither compound was able to induce genotoxicity in the concentrations evaluated. On the other hand, both flavonoids reduced DNA damage induced by AFB1, MMS and DXR in all treatment protocols. Copyright © 2011 John Wiley & Sons, Ltd.     

Palbinone from Paeonia suffruticosa Protects Hepatic Cells via Up‐regulation of Heme Oxygenase‐1

Paeonia suffruticosa has been traditionally employed for vitalizing blood circulation and alleviating liver and inflammatory diseases. The pathways by which palbinone (PB) isolated from P. suffruticosa mediates heme oxygenase‐1 (HO‐1) induction were investigated using the specific inhibitors for PI3K and mitogen activated protein kinases pathways. The effect of PB‐treatment on Nrf2 translocalization and HO‐1‐antioxidant response element (ARE) regulation was examined employing Western blot and luciferase assays. PB induced HO‐1 expression via the activation of Nrf2 in the hepatic cells, and ARE‐dependent genes were stimulated via the PB‐mediated Nrf2 activation. PB‐mediated HO‐1 expression could be involved with PI3K/Akt and ERK1/2 pathways. Our study suggests the mechanism by which PB induces HO‐1 expression in the hepatic cells. This might substantiate the traditional applications of P. suffruticosa for the treatment of oxidative stress‐related diseases including oxidant and inflammatory‐mediated vascular and liver diseases. Copyright © 2013 John Wiley & Sons, Ltd.     

桑叶生物碱、黄酮及多糖对HepG2细胞胰岛素抵抗的改善作用

目的:探讨桑叶生物碱、黄酮及多糖对Hep G2细胞胰岛素抵抗的改善及其作用机制。方法:采用高糖高胰岛素培养基诱导Hep G2细胞形成红外(IR)模型,葡萄糖氧化酶法检测正常组、模型组、罗格列酮组、生物碱组、黄酮组及多糖组的细胞葡萄糖消耗情况,并用RT-q PCR法检测C-Jun氨基端激酶(JNK)通路相关基因的表达,观察桑叶生物碱、黄酮及多糖改善Hep G2细胞胰岛素抵抗及其作用机制。结果:高糖及高胰岛素培养基培养Hep G2细胞,使细胞葡萄糖消耗率明显下降,造成胰岛素抵抗的细胞模型,给予桑叶生物碱、黄酮及多糖干预之后,细胞葡萄糖消耗率明显上升,同时下调JNK和上调IRS-1,AKT mRNA的表达。结论:桑叶生物碱、黄酮及多糖均可增加Hep G2细胞的葡萄糖消耗量,其改善胰岛素抵抗作用可能与调节JNK信号通路有关。     

槲皮素抑制人鼻咽癌CNE2细胞生长并诱导其凋亡的研究

目的:探讨槲皮素对人鼻咽癌CNE2细胞生长与凋亡的作用。方法:应用台盼蓝拒染法检测20,40,60,80μmol·L-1浓度槲皮素作用细胞24,48,72 h后的生长状况,流式细胞仪检测在以上浓度下作用细胞48 h后的细胞凋亡率,Western blot法检测以上浓度槲皮素作用细胞48 h后Bcl-2蛋白的表达情况。结果:各浓度槲皮素对人鼻咽癌CNE2细胞的生长有显著的抑制作用,呈明显的时间和剂量依赖性;经不同浓度的槲皮素作用48 h后,细胞凋亡率显著增加,当浓度在40μmol·L-1以上时,与对照组相比,差异具有统计学意义(P<0.01);Western blot法检测Bcl-2蛋白的表达随着槲皮素浓度的增加而降低,槲皮素浓度在40μmol·L-1以上时,与对照组相比,差异具有统计学意义(P<0.01)。结论:槲皮素可以有效抑制人鼻咽癌CNE2细胞生长,诱导细胞凋亡,其诱导该细胞凋亡的作用机制可能是通过下调Bcl-2蛋白的表达。     

姜黄素对人肝细胞HepG2中低密度脂蛋白受体基因表达影响的研究

 目的研究不同浓度姜黄素对人肝细胞HepG2中内源低密度脂蛋白受体(LDLR)表达的影响,在受体水平上探讨姜黄素的调脂作用机制。方法以荧光试剂标记配体法,利用流式细胞仪技术和荧光显微镜技术研究姜黄素对HepG2细胞LDLR表达活性的影响。利用细胞免疫技术、结合荧光显微镜技术流式细胞术研究姜黄素对HepG2细胞LDLR表达数量的影响。结果姜黄素可显著增加人肝细胞HepG2内源LDLR的表达(P<0.05)。结论姜黄素能够通过增加人肝细胞HepG2 LDLR的表达起到调节血脂和抗动脉粥样硬化的作用。     

β‐Eudesmol Induces JNK‐Dependent Apoptosis through the Mitochondrial Pathway in HL60 Cells

β‐eudesmol, a natural sesquiterpenol present in a variety of Chinese herbs, is known to inhibit the proliferation of human tumor cells. However, the molecular mechanisms of the effect of β‐eudesmol on human tumor cells are unknown. In the present study, we report the cytotoxic effect of β‐eudesmol on the human leukemia HL60 cells and its molecular mechanisms. The cytotoxic effect of β‐eudesmol on HL60 cells was associated with apoptosis, which was characterized by the presence of DNA fragmentation. β‐eudesmol‐induced apoptosis was accompanied by cleavage of caspase‐3, caspase‐9, and poly (ADP‐ribose) polymerase; downregulation of Bcl‐2 expression; release of cytochrome c from mitochondria; and decrease in mitochondrial membrane potential (MMP). Activation of c‐Jun N‐terminal kinases (JNK) mitogen‐activated protein kinases was observed in β‐eudesmol‐treated HL60 cells, and the inhibitor of JNK blocked the β‐eudesmol‐induced apoptosis, downregulation of Bcl‐2, and the loss of MMP. These data suggest that β‐eudesmol induces apoptosis in HL60 cells via the mitochondrial apoptotic pathway, which is controlled through JNK signaling. Copyright © 2012 John Wiley & Sons, Ltd.     

Quercetin reduced the formation of N‐acetyl‐p‐benzoquinoneimine,a toxic metabolite of paracetamol in rats and isolated rat hepatocytes

N‐acetyl‐p‐benzoquinoneimine (NAPQI) is toxic metabolite of paracetamol formed primarily by cytochrome P4502E1 (CYP2E1) metabolic pathway when administered at therapeutic doses or overdose. The influence of quercetin (flavonoid) on the bioactivation of paracetamol to NAPQI was investigated using rat liver microsomes and rats in vivo. Paracetamol (80 mg/kg) was administered orally without or with silymarin (100 mg/kg), a known inhibitor of CYP2E1, CYP3A4 and quercetin (10 and 20 mg/kg) to rats for 15 consecutive days. Area under the plasma concentration–time curve (AUC_0‐∞) and the peakplasma concentration (C_max) of paracetamol were dose‐dependently increased with quercetin (10 and 20 mg/kg) compared to paracetamol control group (p < 0.001). On the other hand, the AUC_0‐∞ and C_max of NAPQI were decreased significantly with quercetin. The same results were observed with silymarin also. The elevated liver and kidney functional enzymes/compounds were significantly reduced by quercetin and silymarin compared to paracetamol control group. The formation of NAPQI was reduced in the incubation samples in presence of quercetin in experiment using isolated rat hepatocytes. The presentstudy results revealed that quercetin might be inhibited the CYP2E1‐mediated metabolism of paracetamol; thereby decreased the formation of NAPQI and protected the liver and kidney.     

ERK1/2在黄芪多糖促进HepG2细胞凋亡中的作用

目的:阐明黄芪多糖与ERK1/2通路在HepG2细胞凋亡中的作用.方法:实验分为4组:空白对照组(control);B组,20 mg?L-1黄芪多糖3个剂量处理组(AP,20,40,80 mg?L-1黄芪多糖处理组( AP,40,80 mg?L-1),药物处理24 h后检测各指标.应用Western blot,MTT,TUNEL,线粒体膜电势检测等方法检测黄芪多糖对HepG2细胞生存凋亡及细胞中ERK1/2表达量的影响.结果:MTT与TUNEL表明黄芪多糖对HepG2细胞的凋亡具有促进作用,增加HepG2细胞的caspase-3的活性,降低(HepG2)细胞线粒体的膜电位,降低Bcl-2表达;黄芪多糖可以降低ERK1/2的表达,表明黄芪多糖的凋亡促进作用可能通过ERK1/2途径.结论:黄芪多糖可以通过抑制ERK1/2信号通路促进HepG2细胞的凋亡.     

槲皮素对自身免疫性前列腺炎模型COX-2及5-LOX表达的影响

目的:建立两种慢性自身免疫性前列腺炎模型,探讨槲皮素(Quercetin)对模型大鼠前列腺组织中炎症因子环氧化酶-2活性(COX-2)、5-脂氧酶(5-LOX)活性、排尿量及尿离子浓度的影响。方法:采用两种自身免疫性前列腺炎大鼠模型:雌二醇诱导的模型(M1)及纯化大鼠前列腺蛋白联合免疫佐剂诱导的模型(M2),分别给药30 d后测定大鼠0⁴h排尿量及尿离子浓度(M1);处死大鼠,取前列腺称重计算前列腺指数;取前列腺液镜下观察白细胞数及卵磷脂小体密度;HE染色制备前列腺组织病理切片并镜下观察;Western Blot法及免疫组化法测定前列腺组织内COX-2及5-LOX的表达。结果:两个模型中,槲皮素200 mg/kg给药组前列腺液内卵磷脂小体密度均明显增加,M1中槲皮素200 mg/kg给药组前列腺指数降低,M2中槲皮素200 mg/kg能明显降低前列腺液中的白细胞数;病理切片中均可见病变明显改善;两种模型中前列腺组织内COX-2及5-LOX表达明显被抑制;槲皮素200 mg/kg组对模型大鼠尿液中的Na+和Cl-的排出有明显的促进作用,但对K+的影响并不十分显著(M1);槲皮素(200 mg/kg、100 mg/kg)组能明显增加水负荷大鼠04h排尿量及尿离子浓度(M1);处死大鼠,取前列腺称重计算前列腺指数;取前列腺液镜下观察白细胞数及卵磷脂小体密度;HE染色制备前列腺组织病理切片并镜下观察;Western Blot法及免疫组化法测定前列腺组织内COX-2及5-LOX的表达。结果:两个模型中,槲皮素200 mg/kg给药组前列腺液内卵磷脂小体密度均明显增加,M1中槲皮素200 mg/kg给药组前列腺指数降低,M2中槲皮素200 mg/kg能明显降低前列腺液中的白细胞数;病理切片中均可见病变明显改善;两种模型中前列腺组织内COX-2及5-LOX表达明显被抑制;槲皮素200 mg/kg组对模型大鼠尿液中的Na+和Cl-的排出有明显的促进作用,但对K+的影响并不十分显著(M1);槲皮素(200 mg/kg、100 mg/kg)组能明显增加水负荷大鼠0⁴h的排尿量,与对照组比较差别具有统计学意义(M1)。结论:槲皮素可能是通过抑制前列腺组织内COX-2及5-LOX表达从而改善慢性前列腺炎并可能通过促进Na+和Cl-的排出改善模型大鼠排尿困难症状。     

槲皮素对U937细胞迁移和侵袭能力及MMP-2,MMP-9表达的影响

目的:观察槲皮素(quercetin)对人急性髓系白血病(human acute myeloid leukemia)U937细胞增殖、凋亡、黏附力、迁移和侵袭能力及基质金属蛋白酶-2(matrix metalloproteinase-2,MMP-2),基质金属蛋白酶-9(matrix metalloproteinase-9,MMP-9)表达的影响。方法:体外培养U937细胞,分别以0,10,20,40μmol·L-1槲皮素处理24,48,72 h,细胞计数试剂盒(cell counting kit-8,CCK-8)检测槲皮素对U937细胞增殖的抑制作用。实验随机分为空白组,槲皮素10,20,40μmol·L-1组。末端脱氧核苷酸转移酶介导的d UTP缺口末端标记测定法(TUNEL)检测U937细胞凋亡情况;细胞黏附实验检测U937细胞黏附力;划痕修复实验检测U937细胞迁移能力;transwell小室体外侵袭实验检测U937细胞侵袭能力;蛋白质印迹法(Western blot)检测U937细胞MMP-2,MMP-9蛋白表达。结果:槲皮素可抑制U937细胞的增殖。与空白组比较,槲皮素10,20,40μmol·L-1组明显升高U937细胞凋亡指数(P0.05,P0.01)。与空白组比较,槲皮素10,20,40μmol·L-1组明显降低U937细胞黏附率,迁移率,侵袭细胞数及MMP-2,MMP-9蛋白表达(P0.05,P0.01)。结论:槲皮素可显著抑制U937细胞迁移和侵袭能力,其机制可能与其抑制MMP-2和MMP-9表达有关。     

从自噬角度研究雷公藤甲素引起HepG2细胞肝毒性的机制

目的:探讨雷公藤甲素(TP)对HepG2细胞的过氧化损伤及对自噬的调控作用。方法:将质量浓度为0.003 2,0.016,0.08,0.4,2 g·L~(-1)的TP作用于体外培养的HepG2细胞48 h,另设空白组做对比,采用噻唑蓝(MTT)法检测细胞生长活性,采用酶联免疫吸附测定(ELISA)法检测超氧化物歧化酶(SOD),谷胱甘肽过氧化物酶(GSH-Px)的活性,采用蛋白质免疫印迹(Western blot)法检测自噬相关蛋白微管相关蛋白轻链3(LC3Ⅱ)与Beclin1蛋白的表达情况,用免疫荧光法检测自噬相关蛋白LC3的表达情况。结果:与空白组比较,随着TP质量浓度的增加,HepG2活性下降(P0.05,P0.01);检测细胞上清中SOD,GSH-Px的活性,均较空白组降低(P0.05,P0.01);Western blot表明TP可上调LC3Ⅱ与Beclin1蛋白水平的表达(P0.05,P0.01),免疫荧光法同样表明TP能够引起自噬相关蛋白LC3表达的增多(P0.05,P0.01),均具有一定的浓度依赖性。结论:一定质量浓度的TP可造成肝细胞损伤,其损伤机制可能与过氧化损伤以及过氧化损伤导致的自噬过度激活有关。     

白屈菜红碱诱导人肝癌细胞HepG2凋亡及机制探讨

目的:研究白屈菜红碱(CHE)体外诱导人肝癌细胞HepG2凋亡的作用与机制。方法:以肝癌HepG2细胞为研究对象,通过噻唑蓝(MTT)法测定CHE对细胞的增殖作用,荧光染料Hoechst 33285染色观察CHE对人肝癌细胞HepG2凋亡形态的变化,免疫印迹法(Western blot)及实时荧光定量PCR(Real-time PCR)检测B淋巴细胞瘤-2(Bcl-2),Bcl-2相关X蛋白(Bax),Bcl-2家族的抑凋亡蛋白Bcl-xl,促凋亡相关半胱氨酸蛋白酶-3(Caspase-3)蛋白和基因表达。结果:与溶剂组比较,CHE能显著抑制HepG2细胞的增殖,其6,12,24 h的半数抑制浓度(IC50)分别为12.98,10.53,11.21μmol·L~(-1),在Hoechst 33258染色结果中,CHE组出现细胞凋亡的典型特征;与溶剂组比较,高浓度CHE能明显上调Bax,Caspase-3的蛋白及mRNA表达,明显降低Bcl-xl蛋白及mRNA表达(P0.05)。结论:白屈菜红碱能抑制肝癌细胞HepG2的增殖,并能上调促凋亡蛋白和mRNA,下调抗凋亡蛋白和mRNA表达,进而诱导凋亡。     

Quercetin inhibited murine leukemia WEHI‐3 cells in vivo and promoted immune response

Enhanced flavonoid consumption is closely related with a reduced cancer incidence as shown in epidemiological studies. Quercetin (3,5,7,3′,4′‐pentahydroxylflavone) is one of the active components of flavonoids which exist in natural plants, particularly in onions and fruits. It was reported that quercetin induced apoptosis in human cancer cell lines, including human leukemia HL‐60 cells, but there is no available information as to its effects on leukemia cells in vivo. The purpose of the present studies was to focus on the in vivo effects of quercetin on leukemia WEHI‐3 cells. The effects of quercetin on WEHI‐3 cells injected into BALB/c mice were examined. Quercetin decreased the percentage of Mac‐3 and CD11b markers, suggesting that the differentiation of the precursors of macrophages and T cells was inhibited. There was no effect on CD3 levels but increased CD19 levels. Quercetin decreased the weight of the spleen and liver compared with the olive oil treated animals. Quercetin stimulated macrophage phagocytosis of cells isolated from peritoneum. Quercetin also promoted natural killer cell activity. Based on pathological examination, an effect of quercetin was observed in the spleen of mice previously injected with WEHI‐3 cells. Apparently, quercetin affects WEHI‐3 cells in vivo. Copyright © 2009 John Wiley & Sons, Ltd.     

Quercetin protects against cisplatin‐induced acute kidney injury by inhibiting Mincle/Syk/NF‐κB signaling maintained macrophage inflammation

Acute kidney injury (AKI) with high incidence and mortality is the main cause of chronic kidney disease. Previous studies have indicated that quercetin, an abundant flavonoid in plants, exhibited renoprotective role in AKI. However, the underlying mechanism is largely unknown. In this study, we try to explore whether quercetin protects against AKI by inhibiting macrophage inflammation via regulation of Mincle/Syk/NF‐κB signaling. The results demonstrated that quercetin can significantly inhibit expression and secretion of IL‐1β, IL‐6, and TNF‐α in LPS‐induced bone marrow‐derived macrophages (BMDMs) and reduce activity of Mincle/Syk/NF‐κB signaling in vitro. We also found that quercetin can strongly reduce the concentration of serum creatinine, BUN, IL‐1β, IL‐6, and TNF‐α in cisplatin‐induced AKI model. Furthermore, quercetin down‐regulated protein levels of Mincle, phosphorylated Syk and NF‐κB in kidney macrophages of AKI, as well as inhibited M1, up‐regulated M2 macrophage activity. Notably, the down‐regulation of LPS‐induced inflammation by quercetin was reversed after adding TDB (an agonist of Mincle) in BMDMs, suggesting that quercetin suppresses macrophage inflammation may mainly through inhibiting Mincle and its downstream signaling. In summary, these findings clarified a new mechanism of quercetin improving AKI‐induced kidney inflammation and injury, which provides a new drug option for the clinical treatment of AKI.