Hydrogen-rich water prevents lipid deposition in the descending aorta in a rat periodontitis model

Objective

Periodontitis has been causally linked to atherosclerosis, which is mediated by the oxidative stress. As hydrogen-rich water (HW) scavenges reactive oxygen species (ROS), we hypothesized that HW could prevent lipid deposition induced by periodontitis in the aorta. The aim of this study was to investigate the effects of HW on the initiation of atherosclerosis in a rat periodontitis model.

Design

Eighteen 8-wk-old male Wistar rats were divided into three groups of six rats; the periodontitis group, periodontitis + HW group and the no treatment (control) group. In the periodontitis and periodontitis + HW groups, periodontitis was induced using a ligature for 4 wk, while the periodontitis + HW group was given water containing 800–1000 μg/L hydrogen during the 4-wk experimental period.

Results

In the periodontitis group, lipid deposition in the descending aorta was observed. The periodontitis group also showed significant higher serum levels for ROS and oxidised low-density lipoprotein-cholesterol (ox-LDL) (1.7 and 1.4 times, respectively), and higher aortic expression levels of nitrotyrosine and hexanoyl-lysine (HEL) (7.9 and 16.0 times, respectively), as compared to the control group (p < 0.05). In the periodontitis + HW group, lipid deposition was lower. Lower serum levels of ROS and ox-LDL (0.46 and 0.82 times, respectively) and lower aortic levels of nitrotyrosine and HEL (0.27 and 0.19 times, respectively) were observed in the periodontitis + HW group than in the periodontitis group (p < 0.05).

Conclusions

HW intake may prevent lipid deposition in the rat aorta induced by periodontitis by decreasing serum ox-LDL levels and aortic oxidative stress.     

Oxygenized Low-Density Lipoprotein-Induced ASMC Dysregulation Depends on circ_0000345-Mediated Regulatory Mechanism

Aims: Vascular smooth muscle cells are key participants in atherosclerosis. Circular RNA hsa_circ_0000345 (circ_0000345) and miR-647 are related to oxygenized low-density lipoprotein (ox-LDL)-induced arterial smooth muscle cell (ASMC) dysregulation. However, the relationship between circ_0000345 and miR-647 in ox-LDL-induced ASMC dysregulation is unclear. Methods: Relative levels of circ_0000345, miR-647, and PAP-associated domain containing 5 (PAPD5) mRNA in AS patient’s serum and ox-LDL-induced ASMCs were detected via RT-qPCR. Gain-of-function experiments were utilized to analyze the effects of circ_0000345 upregulation on ox-LDL-induced cell proliferation, migration, invasion, and inflammatory response in ASMCs. The relationship between circ_0000345 or PAPD5 and miR-647 was validated by dual-luciferase reporter and RNA immunoprecipitation assays. Results: Circ_0000345 and PAPD5 were lowly expressed in AS patient’s serum and ox-LDL-induced ASMCs, while miR-647 expression had an opposing trend. Mechanistically, circ_0000345 was verified as a miR-647 sponge, and miR-647 overexpression impaired the inhibitory effects of circ_0000345 upregulation on ox-LDL-induced ASMC proliferation, migration, invasion, and inflammatory response. Further experiments demonstrated that PAPD5 was a miR-647 target, and circ_0000345 adsorbed miR-647 to mediate PAPD5 expression. Also, PAPD5 inhibition relieved miR-647 silencing-mediated suppression on ox-LDL-induced ASMC proliferation, migration, invasion, and inflammatory response. Conclusions: Circ_0000345 elevated PAPD5 expression via acting as a miR-647 sponge, resulting in alleviating ox-LDL-induced ASMC dysregulation. The study highlighted the critical role of circ_0000345 in AS.     

美乐托宁对ox-LDL诱导的人脐静脉血内皮祖细胞增殖、凋亡及Bcl-2表达的影响

目的:探讨美乐托宁(melatonin,Mel)对氧化低密度脂蛋白(oxidized low-density lipoprotein,ox-LDL)诱导的人脐静脉血内皮祖细胞(endothelial progenitor cell,EPC)增殖、凋亡及Bcl-2表达的影响.方法:密度梯度离心法获取人脐静脉血单个核细胞,培养7 d后,贴壁细胞分成7组:对照组以RPMI1640培养液培养;ox-LDL各浓度组(共3组)分别用含5,10,20 mg/L ox-LDL的RPMI 1640培养液孵育;Mel各浓度组(共3组)分别先用含0.5,1.0,2.0 mmol/L Mel的RPMI 1640培养液培养24h,然后移去含Mel的RPMI 1640培养液,加入含10 mg/L ox-LDL的RPMI 1640培养液孵育.采用MTT法检测EPC增殖能力;流式细胞仪检测细胞凋亡率;提取细胞RNA,采用逆转录一聚合酶链式反应(RT-PCR)技术检测Bcl-2 mRNA表达;采用免疫细胞化学法检测Bcl-2蛋白表达水平.结果:ox-LDL呈浓度依赖性抑制EPC增殖,诱导EPC凋亡;在加ox-LDL(10 mg/L)前加Mel干预,EPC增殖能力较ox-LDL组明显增强,细胞凋亡率明显降低;RT-PCR和免疫细胞化学法检测显示,ox-LDL组EPC的Bcl-2 mRNA和蛋白表达明显低于对照组,Mel组明显高于ox-LDL组(P＜0.01).结论:ox-LDL呈浓度依赖性诱导EPC凋亡、抑制EPC增殖,Mel能抑制ox-LDL的上述作用,其机制与上调Bcl-2表达有关.     

番石榴叶总黄酮抑制泡沫细胞形成及LOX-1表达的实验研究

目的 研究番石榴叶总黄酮抑制泡沫细胞形成及LOX-1表达的作用。方法 建立THP-1单核-巨噬细胞源性泡沫细胞模型,根据CCK-8法测定的番石榴叶总黄酮对该细胞的安全浓度范围,将配制的番石榴总黄铜溶液在其安全范围内选取低、中、高三个剂量组,以无血清培养液的空白对照组及ox-LDL模型组为对照。采用油红O染色观察细胞内脂质堆积情况,酶偶联比色法检测细胞内总胆固醇含量,Western blot法检测LOX-1的蛋白表达水平。结果 三个剂量组细胞内总胆固醇含量均低于模型组（P〈0.05）,并随番石榴叶总黄酮浓度升高而下降;三个剂量组的LOX-1蛋白表达均低于模型组（P〈0.05）,抑制作用随剂量升高而增强。结论 番石榴叶总黄酮通过抑制LOX-1的表达,减少巨噬细胞摄取ox-LDL,抑制泡沫细胞形成,从而起到抗动脉粥样硬化的作用。     

小檗碱对OX-LDL诱导的巨噬细胞泡沫化的作用

目的:观察小檗碱对巨噬细胞泡沫化的保护作用.方法:利用ox-LDL诱导小鼠巨噬细胞RAW264.7泡沫化,并在此过程中加入不同浓度小檗碱,通过油红O染色及胆固醇酯与总胆固醇的测定判断细胞泡沫化程度,评价小檗碱的作用效果.结果:与ox-LDL诱导的泡沫细胞组相比,小檗碱加入后细胞内脂质累积明显减少,而且泡沫化程度也降低了.结论:小檗碱能够降低ox-LDL诱导的巨噬细胞泡沫化程度.     

Kaempferol-induced GPER upregulation attenuates atherosclerosis via the PI3K/AKT/Nrf2 pathway

ContextThe effect of kaempferol, a regulator of oestrogen receptors, on atherosclerosis (AS) and the underlying mechanism is elusive.ObjectiveTo explore the effect and mechanism of kaempferol on AS.Methods and materialsIn vivo, C57BL/6 and apolipoprotein E (APOE)^–/– mice were randomly categorized into six groups (C57BL/6: control, ovariectomy (OVX), high-fat diet (HFD); APOE^–/–: OVX-HFD, OVX-HFD + kaempferol (50 mg/kg) and OVX-HFD + kaempferol (100 mg/kg) and administered with kaempferol for 16 weeks, intragastrically. Oil-Red and haematoxylin–eosin (HE) staining were employed to examine the effect of kaempferol. In vitro, human aortic endothelial cells (HAECs) were pre-treated with or without kaempferol (5, 10 or 20 μM), followed by administration with kaempferol and oxidized low-density lipoprotein (ox-LDL) (200 μg/mL). The effect of kaempferol was evaluated using flow cytometry, and TdT-mediated dUTP Nick-End Labelling (TUNEL).ResultsIn vivo, kaempferol (50 and 100 mg/kg) normalized the morphology of blood vessels and lipid levels and suppressed inflammation and apoptosis. It also activated the G protein-coupled oestrogen receptor (GPER) and PI3K/AKT/nuclear factor-erythroid 2-related factor 2 (Nrf2) pathways. In vitro, ox-LDL (200 μg/mL) reduced the cell viability to 50% (IC₅₀). Kaempferol (5, 10 or 20 μM) induced-GPER activation increased cell viability to nearly 10%, 19.8%, 30%, and the decreased cellular reactive oxygen species (ROS) generation (16.7%, 25.6%, 31.1%), respectively, consequently attenuating postmenopausal AS. However, the protective effects of kaempferol were blocked through co-treatment with si-GPER.ConclusionsThe beneficial effects of kaempferol against postmenopausal AS are associated with the PI3K/AKT/Nrf2 pathways, mediated by the activation of GPER.     

阿托伐他汀对ACS患者血清CTRP9的影响及其机制的研究

目的 探讨ACS患者血清脂肪细胞因子C1q/肿瘤坏死因子相关蛋白 9（CTRP9）与疾病的关系,并观察使用阿托伐他汀治疗后血清CTRP9水平的变化及可能的机制。 方法 选择102例ACS患者和70名正常对照者,检测2组血清CTRP9水平。再将102例ACS患者随机分为常规治疗组51例（M组）和阿托伐他汀组51例（A组）,后者在常规治疗基础上,加服阿托伐他汀20mg/d,治疗8周后观察2组患者血清CTRP9水平。并用ELISA法检测A、M组治疗前后的血清游离脂肪酸（FFA）、氧化低密度脂蛋白（ox-LDL）水平。 结果 与正常对照组相比,ACS患者血清 CTRP9水平显著降低（P＜0.05）,Logistic 回归分析表明,血清CTRP9是ACS患者的独立保护因子。与M组相比,治疗8周后,A组血清 CTRP9水平显著升高（P＜0.05）,血清FFA、ox-LDL水平明显下降（P＜0.05）,而M组血清 CTRP9、FFA、ox-LDL治疗前后表达水平无明显变化。 结论 ACS患者血清 CTRP9水平明显降低,CTRP9是ACS独立的保护因素,阿托伐他汀可升高ACS血清CTRP9 水平,其机制可能与下调血清FFA、ox-LDL有关。     

氧化低密度脂蛋白在尿酸致早期慢性肾脏疾病血管内皮损伤中的作用

目的：探讨氧化低密度脂蛋白（ox-LDL）在尿酸所致早期慢性肾脏疾病（CKD）血管内皮损伤中的作用。方法：30只雄性SD大鼠随机分为3组：A组（假手术组,n=10）,B组（右肾切除组,n=10）和C组（右侧肾切除＋氧嗪酸钾灌胃组,n=10）。检测血清尿酸（UA）、肌酐（Scr）、一氧化氮（NO）、内皮素-1（ET-1）、ox-LDL等指标。光镜观察血管壁病理损害。分析ox-LDL水平与UA、NO、ET-1、NO/ET-1比值的相关性。结果：（1）3组大鼠肾病理均未见明显病变,且血肌酐无明显差异,但C组血尿酸水平明显升高（P〈0.01）;（2）A组血管壁未见明显病变;C组可见内皮细胞脱落,内皮间隙增宽,细胞水肿,管壁炎症细胞聚集,血管中层平滑肌细胞增生明显,结构紊乱;B组病变与C组相似,但程度较轻。C组NO明显低下（P〈0.01）,ET-1明显增高（P〈0.05）,NO/ET-1比值明显降低（P〈0.01）;（3）单因素相关分析显示：尿酸与ET-1呈显著正相关（r=0.9374,P〈0.01）,与NO（r=-0.9462,P〈0.01）、NO/ET-1比值（r=-0.9230,P〈0.01）呈显著负相关;⑷C组ox-LDL水平显著升高（P〈0.05）,ox-LDL与尿酸（r=0.8479,P〈0.01）、ET-1（r=0.7900,P〈0.01）呈显著正相关,与NO（r=-0.7459,P〈0.05）、NO/ET-1比值（r=-0.7949,P〈0.01）呈显著负相关。结论：尿酸可致早期CKD血管内皮细胞损伤和功能紊乱,ox-LDL在其发生发展中可能起重要作用。     

孤儿核受体Nur77对氧化型低密度脂蛋白诱导的小鼠巨噬细胞系分泌炎症因子的影响

目的观察孤儿核受体Nur77对氧化型低密度脂蛋白（ox-LDL）诱导小鼠巨噬细胞分泌炎症因子的影响。方法建立稳定转染GFP-Nur77质粒的小鼠巨噬细胞系RAW 264.7,并以稳定转染绿色荧光蛋白（GFP）基因的细胞系RAW 264.7作为对照。将转染的细胞与ox-LDL（40 mg/L）共同孵育24 h后,采用ELISA法检测炎症因子TNF-α、IFNγ-、基质金属蛋白酶-9（MMP-9）、单核细胞趋化蛋白-1（MCP-1）和IL-6的表达。结果经ox-LDL刺激后,不同质粒DNA转染的两种细胞中,IFNγ-、TNF-α、MMP-9和MCP-1的浓度均升高（P<0.05）;但GFP-Nur77-RAW 264.7细胞的增长水平明显低于GFP-RAW 264.7细胞（P<0.05）。ox-LDL刺激前,GFP-RAW 264.7细胞分泌的4种炎症因子（TNF-α、IFN-γ、MMP-9和MCP-1）的浓度,分别为（19.00±1.69）ng/L、（99.10±3.09）ng/L、（0.14±0.02）ng/L和（23.96±3.57）ng/L;ox-LDL刺激后,分别上升为（27.22±0.38）ng/L、（549.90±1.81）ng/L、（0.45±0.06）ng/L和（49.31±3.66）ng/L。ox-LDL刺激前,GFP-Nur77-RAW 264.7和GFP-RAW 264.7细胞分泌的4种炎症因子的浓度无统计学的差异性。ox-LDL刺激后,4种炎症因子的浓度分别上升为（18.34±1.06）ng/L、（320.56±1.27）ng/L、（0.20±0.03）ng/L和（38.92±4.46）ng/L。而两种细胞（CTFP-Nur77-RAW 264.7细胞与GFP-RAW 264.7细胞）分泌IL-6的浓度未见显著差异。结论RAW 264.7细胞中Nur77表达的上调,能够显著减少ox-LDL诱导的炎症因子（IL-6除外）分泌,这可能是孤儿核受体Nur77心血管保护作用的重要机制之一。     

曲美他嗪抑制自噬减少中性粒细胞胞外核酸网形成

目的研究曲美他嗪(trimetazidine TMZ)对氧化低密度脂蛋白(oxidized low-density lipoprotein,ox-LDL)体外诱导中性粒细胞胞外核酸网(neutrophil extracellular traps,NETs)形成的影响及其与自噬的关系。方法密度梯度离心法提取小鼠骨髓中性粒细胞,ox-LDL体外干预建立NETs诱导模型;采用TMZ、LY294002、Rapamycin干预ox-LDL对NETs的诱导;免疫荧光法观察NETs标志物MPO-DNA的释放;PicoGreen试剂盒定量检测cfDNA/nets含量;Western blot检测髓过氧化物酶(MPO)、自噬相关蛋白Beclin-1、LC3b的表达。结果 ox-LDL以浓度-时间依赖的方式刺激中性粒细胞释放MPO-DNA复合物,使上清cfDNA/nets含量明显增高;细胞自噬相关蛋白LC3b、Beclin-1及MPO的蛋白水平亦随ox-LDL浓度增加上调; TMZ预处理能抑制NETs释放,降低LC3b、Beclin-1及MPO的蛋白表达,该结果可被PI3K通路阻断剂LY294002模拟,...