ox-LDL regulates proliferation and apoptosis in VSMCs by controlling the miR-183-5p/FOXO1

Fan Mingqiang,Huang Yinglong,Li Kunsheng,Yang Xiangxiang,Bai Jing,Si Qiaoke,Peng Zhengfei,Jia Chunwen,Zhang Qiangnu,Tao Ding 한국유전학회 2022 Genes & Genomics Vol.44 No.6

Background: microRNA-mRNA axes that are involved in oxidized low-density lipoprotein (ox-LDL)-induced vascular smooth muscle cells (VSMCs) proliferation/apoptosis imbalance need to be further investigated. Objective: To investigate the functional role of miR-183-5p/FOXO1 in VSMCs and its interaction with ox-LDL. Methods: RNA sequencing was used to detect transcriptome changes of VSMCs treated with ox-LDL. miR-183-5p and FOXO1 expression levels in VSMCs after ox-LDL treatment were assessed using qRT-PCR and western blotting. The regulatory effect of miR-183-5p on FOXO1 has been tried to prove using a dual-luciferase reporter assay. The functions of miR-183-5p, and FOXO1 were analyzed by CCK-8 assay and flow cytometry assay. The tissue samples or serum samples of high fat-feeding mice and carotid atherosclerosis patients were collected, and the levels of miR-183-5p/FOXO1 were analyzed. Results: RNA sequencing data showed 81 miRNAs including miR-183-5p was significantly changed after ox-LDL treatment in VSMCs. FOXO1, a miR-183-5p's potential target, was also down-regulated in ox-LDL treated cells. qRT-PCR and western blot found that expression of FOXO1 mRNA and protein significantly reduced in VSMCs treated with ox-LDL, accompanied by overexpression of miR-183-5p. miR-183-5p inhibited FOXO1 mRNA by binding to its 3' UTR. Interference miR-183-5p/FOXO1 could change proliferation/apoptosis imbalance in VSMCs under ox-LDL stimulation. Higher levels of miR-183-5p but reduced FOXO1 can be found in the thoracic aorta tissues of high fat-feeding mice. In serum samples from individuals with carotid atherosclerosis, Higher levels of miR-183-5p were observed. the miR-183-5p level was positively related to the level of serum ox-LDL in patients. Conclusions: Aberrant expression of miR-183-5p/FOXO1 pathway mediated ox-LDL-induced proliferation/apoptosis imbalance in VSMCs. The miR-183-5p/FOXO1 axis can potentially be utilized as the target in the treatment of patients with atherosclerosis.

MiR-590 Inhibits Endothelial Cell Apoptosis by Inactivating the TLR4/NF-κB Pathway in Atherosclerosis

Lei Yang,Chuanyu Gao 연세대학교의과대학 2019 Yonsei medical journal Vol.60 No.3

Purpose: Previous study has well documented the anti-apoptotic effects of miR-590 on oxidized low-density lipoprotein (ox-LDL)-treated endothelial cells (ECs). However, the mechanism underlying the anti-apoptotic effects of miR-590 in ox-LDL-treated ECsremains to be further addressed. Materials and Methods: ApoE-/- mice fed with a high-fat diet (HFD) and human aortic endothelial cells (HAECs) treated with ox-LDL were used as in vivo and in vitro models of atherosclerosis. The expressions of miR-590 and toll-like receptor 4 (TLR4) weredetected by quantitative real-time PCR and Western blot, respectively. Atherosclerotic lesion analysis was performed using Evansblue and hematoxylin-eosin staining. Cell proliferation was assessed by MTT assay. Apoptosis was examined using flow cytometryanalysis and Western blot analysis of Cleaved poly (ADP-ribose) polymerase (PARP) and Cleaved Caspase-3 levels. The effect ofmiR-590 on TLR4/nuclear factor kappa B (NF-κB) pathway was evaluated by Western blot. Binding between miR-590 and TLR4was confirmed by luciferase reporter assay and Western blot. Results: miR-590 was downregulated in the aorta tissues from HFD-fed apoE-/- mice and ox-LDL-treated HAECs. miR-590 overexpressioninhibited atherosclerotic lesion in HFD-induced apoE-/- mice and promoted proliferation and inhibited apoptosis of ox-LDL-treated HAECs. Additionally, TLR4 was identified as a direct target of miR-590 in ox-LDL-treated HAECs. Moreover, antimiR-590 reversed TLR4 knockdown-mediated promotion of cell proliferation and suppression of apoptosis in ox-LDL-treatedHAECs. miR-590 overexpression suppressed the TLR4/NF-κB pathway, and inhibition of the TLR4/NF-κB pathway promoted cellproliferation and impeded apoptosis in ox-LDL-treated HAECs. Conclusion: miR-590 promoted proliferation and blocked ox-LDL-induced apoptosis in HAECs through inhibition of the TLR4/NF-κB pathway.

Protection Effects of Allylmercaptan, Metabolite of Garlic on Endothelial Cell Injury Induced by Oxidized Low Density Lipoprotein

Seung Taek Yang(양승택) 한국생명과학회 2010 생명과학회지 Vol.20 No.11

Low density lipoprotein (LDL)의 산화는 동맥경화의 유발과 진행에 결정적 역할을 하는 것으로 알려져 있다. 본 연구에서는 마늘의 주요 대사산물인 allylmercaptan의 산화 low density lipoprotein에 의해 손상된 내피세포의 보호효과에 대하여 실험하였다. Allylmercaptan의 항산화 활성은 thiobarbituric acid substance (TBARS)로 측정하였다. Allylmercaptan은 0.1, 1 및 10 mM의 농도에서 Cu<SUP>2+</SUP>에 의해 유도된 LDL의 산화를 용량의존적으로 억제하였다. 폐동맥 내피세포를 37℃, 5% CO₂ 상태에서 24시간 동안 미리 배양시킨 후 세측한 다음 다시 24시간 동안 0.1 ㎎/ml oxidized LDL (ox-LDL)을 첨가하여 배양하였다. 이 때 ox-LDL이 Lactate dehydrogenase (LDH)의 방출과 glutathione (GSH)를 감소시키는 원인으로 세포막 손상의 지표로 LDH와 GSH 함량을 조사하였다. 본 실험 결과 allylmercaptan을 일정 농도 별로 endothelial cell에 첨가하여 배양했을 때 LDH의 방출과 GSH의 감소를 현저하게 억제하였다. Peroxide를 형광분석법으로 24 well plate에서 직접 측정한 결과 allylmercaptan이 폐동맥 내피세포 내에서 ox-LDL 유도 peroxide의 방출을 억제하였다. 그리고 allylmercaptan은 과산화수소의 소거능도 있었다. 본 실험결과 allylmercaptan은 ox-LDL 유도 폐동맥 내피세포를 보호할 수 있었으므로 allylmercaptan은 동맥경화의 예방에 유용할 것으로 생각된다. Oxidation of low density lipoprotein (LDL) has been recognized as an important role in the initiation and progression of atherosclerosis. In this study, effects of allylmercaptan, a major metabolite compound of garlic, was studied on endothelial cell injury induced by oxidized low density lipoprotein (ox-LDL). The antioxidative activity of allylmercaptan was investigated by monitoring a thiobarbituric acid substance (TBARS). Allylmercaptan inhibited LDL oxidation induced by Cu<SUP>2+</SUP> at concentrations of 0.1, 1 and 10 mM in a dose dependent manner. Lactate dehydrogenase (LDH) release, as an index of cell injury, and intracellular glutathione levels were determined. Pulmonary artery endothelial cells were preincubated with allylmercaptan at 37oC and 5% CO₂ for 24 hr, washed, and then exposed to 0.1 ㎎/ml oxidized LDL for 24 hr. Preincubation of endothelial cells with allylmercaptan significantly prevented the LDH release and depletion of GSH. Peroxides were measured directly in 24 well plates using a fluorometric assay. Allylmercaptan inhibited release of peroxides induced by ox-LDL in pulmonary artery endothelial cells. In a free system, allylmercaptan was shown to scavenge hydrogen peroxide. The data indicate that allylmercaptan can protect pulmonary artery endothelial cells from injury caused by oxidized LDL, and suggest that allylmercaptan may be useful for the prevention of atherosclerosis.

LncRNA MALAT1 protects human umbilical vein endothelial cells against ox-LDL triggered cell death through regulation of MGP

Xia Wang,Hongqin Zhao,Shaonan Yang,Xiaojun Shao,Shumin Nie,Xudong Pan 대한독성 유전단백체 학회 2019 Molecular & cellular toxicology Vol.15 No.3

Backgrounds: Metastasis associated lung adenocarcinoma transcript 1 (MALAT1) is an lncRNA that has been suggested as a key regulator in the onset of atherosclerosis (AS). This study described the role of MALAT1 in oxidized low density lipoprotein (ox-LDL)-induced endothelial cells death. Methods: Human umbilical vein endothelial cells (HUVECs) were subjected to ox-LDL, before which the expression of MALAT1 was overexpressed by transfection. CCK-8 assay, flow cytometer detection, and western blot were carried out to evaluate cell viability, apoptosis and autophagy. qRT-PCR and western blot analyses were performed to investigate the regulatory relationship between MALAT1, Matrix Gla protein (MGP) and mTOR signaling to decode the underlying mechanism. Results: Up-regulation of MALAT1 attenuated ox-LDLinduced HUVECs lose, as evidenced by the promoted cell viability, and the decreased apoptosis rate. This finding was coupled with the down-regulated p53, Bax, active-caspase-3, Beclin-1 and LC3-II, as well as the up-regulated Bcl-2 and p62. Meanwhile, MALAT1 upregulation promoted the phosphorylation of p70S6K and mTOR, and the expression of MGP. MGP up-regulation exhibited MALAT1-like propoties in preventing ox-LDL-induced cell death and mTOR deactivation. Of contrast, MGP silence affected HUVECs survival and mTOR signaling resulted in contrary impacts. Conclusion: The present work described that MALAT1 up-regulation prevented ox-LDL-mediated apoptosis and autophagy in HUVECs. The protective effects of MALAT1 might be partially via up-regulating MGP, which led to the activation of mTOR signaling.

복분자 미숙과 물추출물의 콜레스테롤 개선 효과

최혜란(Hye Ran Choi),이수정(Su Jung Lee),이정현(Jung-Hyun Lee),권지웅(Ji Wung Kwon),이희권(Hee Kwon Lee),정종태(Jong Tae Jeong),이태범(Tae-Bum Lee) 한국식품영양과학회 2013 한국식품영양과학회지 Vol.42 No.12

We investigated the effects of unripe black raspberry water extract (UBR-W) and oxidation-LDL treatment on cholesterol levels. Experiments using an established human hepatocellular carcinoma cell line (HepG2) showed a time-dependent increase in expression of LDL receptor after UBR-W treatment. Expression of LDL receptor-related genes, such as SREBP1 and 2, increased upon UBR-W treatment. However, expression of HDL-related genes was unaffected by UBR-W. HMG-CoA reductase activity was reduced by UBR-W treatment, whereas HMG-CoA mRNA expression significantly increased. In addition, the ApoB/ApoA1 mRNA level, which is a predictor of cardiovascular risk, was reduced in a time-dependent manner by UBR-W treatment. Macrophage-like cells (RAW 264.7) showed increased expression of ox-LDL-related genes, such as CD36, scavenger receptor-A, adipophilin, and PPAR-gamma, upon ox-LDL treatment compared to untreated control cells, and quantitative lipid analysis indicated a dramatic increase in lipid accumulation. However, UBR-W treatment significantly reduced expression of ox-LDL-related genes and largely prevented lipid accumulation. The results indicate that UBR-W mediates a cholesterol-lowering effect via inhibition of cholesterol synthesis and induction of LDL uptake through SREBP.

Characterization of oxidized phospholipids in oxidatively modified low density lipoproteins by nanoflow liquid chromatography-tandem mass spectrometry

Lee, J.Y.,Lim, S.,Park, S.,Moon, M.H. Elsevier 2013 Journal of chromatography A Vol.1288 No.-

Oxidized low density lipoproteins (Ox-LDLs) have an important role in the development of age-related vascular disease, such as atherosclerosis. Ox-LDLs are defined as oxidatively modified LDLs in the blood by enzymatic or non-enzymatic oxidation of phospholipids (PLs). For the characterization of Ox-LDLs at molecular level, oxidation patterns of oxidized PL (Ox-PL) products were systematically examined with standard PL molecules (16:0/22:6-PC, 18:0/22:6-PA, and 18:0/22:6-PG), by the formation of bilayer vesicles of each standard, followed by oxidation of PL vesicles using a Cu<SUP>2+</SUP> solution. This oxidative modification was applied to LDL standard materials. Nanoflow liquid chromatography-electrospray ionization-tandem mass spectrometry (nLC-ESI-MS/MS) analysis of the extracted PL mixtures resulted in the identification of 276 PLs both in the modified and non-modified LDL, including 139 Ox-PL species. Examination of the identified PL species from the standard LDL before and after oxidation supported the postulate that the specific location of an acyl chain of LPL can be exchanged between the sn-1 and -2 positions. This exchange occurs when the neighboring acyl chain is cleaved during oxidation and the polar head group of PL molecules can be dissociated to form PA molecules that result in the formation of various Ox-PA products. This study demonstrates that nLC-ESI-MS<SUP>n</SUP> can be utilized for the separation and structural characterization of complicated Ox-PL mixtures, including long chain products with hydroxylation or hydroperoxylation at an unsaturated acyl chain, and short chain products from the cleavage of unsaturated fatty acyl chains to form lysophospholipids (LPLs) or the truncation of an acyl chain into a shorter chain terminated with aldehyde or carboxylic acid.

<i>Eucommia ulmoides</i> leaf (EUL) extract enhances NO production in ox-LDL-treated human endothelial cells

Lee, Geum-Hwa,Lee, Hwa-Young,Choi, Min-Kyung,Choi, An-Hong,Shin, Tai-Sun,Chae, Han-Jung Elsevier 2018 Biomedicine & pharmacotherapy Vol.97 No.-

<P><B>Abstract</B></P> <P> <I>Eucommia ulmoides</I> leaves (EULs), referred to as Du-zhong, are utilized to lower blood pressure and improve liver and kidney tone, and also have been applied to cardiovascular disease in Korea, China, and Japan. Endothelial dysfunction, which is caused by endothelial nitric oxide synthase (eNOS) uncoupling, is an initial step in atherosclerosis. In this study, we investigated the protective effects of EUL aqueous extract against ox-LDL-induced eNOS uncoupling and its possible mechanisms in human umbilical vein endothelial cells (HUVECs). A EUL component, aucubin, was also applied to ox-LDL-exposed HUVECs. Whereas ox-LDL significantly decreased nitric oxide (NO) levels in HUVECs, EUL extract and aucubin led to significant recovery of NO levels. When treated with ox-LDL in the presence of EUL extracts or aucubin, O<SUB>2</SUB> <SUP>-</SUP> production was markedly reduced in HUVECs compared to treatment with ox-LDL alone. EUL extract and aucubin also led to recovery of phospho-eNOS Thr495 expression, a critical signaling component in eNOS uncoupling, suggesting that EUL has regulatory effects against eNOS uncoupling and might play preventive/regulatory roles against vascular endothelial dysfunction.</P> <P><B>Graphical abstract</B></P> <P>[DISPLAY OMISSION]</P>

개별 열량 소비 트레이닝이 비만 어린이의 산화-항산화체계에 미치는 영향

우진희 ( Jin Hee Woo ),강성훈 ( Sung Hwun Kang ) 한국스포츠정책과학원(구 한국스포츠개발원) 2008 체육과학연구 Vol.19 No.4

본 연구는 남자초등학생을 비만군(n=20, 나이=11.15±1.42세)과 정상군(n=10, 나이=10.70±0.95세)으로 나누어 개별 열량 소비 트레이닝을 12주간 실시하여 산화-항산화체계에 미치는 영향을 비교하는데 목적이 있다. 신체구성, 호흡순환기능, 혈청지질, 항산화효소, 조직손상 인자들을 분석하였으며, 연구결과 비만군은 체지방률, 수축기와 이완기 혈압, TG, LDL-c, ox-LDL, 8-OHdG 등이 정상군에 비해 유의하게 높다는 것과 이에 반하여 HDL-c, 항산화효소인 SOD 수치는 정상군과 비교하여 유의하게 낮게 나타났다. 또한 12주간의 개별 열량소비 트레이닝을 통해 정상군과 비만군 모두 ox-LDL 수치와 GPX활성이 유의하게 증가되었다. 따라서, 비만아동은 비만에 의한 조직손상과 항산화 방어 역량의 저하로 산화·항산화 체계의 불균형을 일으키며, 산화적 스트레스를 높여 혈관의 염증반응을 증가시켜 결국 혈관을 손상시키고 심혈관질환위험을 가속화시키는데 영항을 줄 수 있음을 시사한다. 또한 12주간의 개별열량 소비 트레이닝을 통하여 항산화효소 활성에 긍정적인 영향을 주어 체내의 산화·환원의 체계를 정상화시켜 심혈관계 스트레스 감소에 긍정적인 영향을 줄 수 있을 것으로 사료된다. The purpose of this study was to compare the differences between before and after 12 week individual calory consumption training on oxidation-antioxidation in obese children. All the subjects divided into obese training group(n=20, age=11.15±1.42) and normal training group(n=10, age=10.70±0.95). Before and after training, we measured that body composition, cardio-pulmonary function, serum lipid, antioxidant enzyme, tissue damage factors. There were differences in body fat, VO2 max, systolic and diastolic blood pressure, TG, HDL-c, LDL-c, ox-LDL, 8-OHdG, and SOD levels between groups. Also, there were differences in ox-LDL and GPX levels between pre and post. In conclusion, body fat, blood pressures, TG, LDL-c, ox-LDL, and 8-OHdG levels were higher and VO2 max, HDL-c, and SOD levels were lower in obese children than in normal children. But, GPX activity in obese children with regular exercise were found to be similar to those of normal children. These results indicated that increased cardiovascular stress and antioxidant imbalace begins in childhood in obesity. However, individual calory consumption training can enhance antioxidant capacity in obese children.

Methionine strengthens anti-inflammation of rice protein via depressing NF-κB activation and stimulating Msr expression in rats fed cholesterol-enriched diets

Zhengxuan Wang,Mingcai Liang,Bingxiao Liu,Lin Yang 한국식품과학회 2022 Food Science and Biotechnology Vol.31 No.6

Oxidized low-density lipoprotein (ox-LDL) is an inducer of inflammation. To elucidate the link of depression of ox-LDL accumulation and anti-inflammatory function of rice protein (RP) whether dependent on methionine availability, growing and adult rats were fed RP and methionine-supplemented RP (RM) under cholesterol-enriched dietary condition. After two weeks feeding, RP and RMs exerted the anti-inflammatory effects through up-regulating IL-10, while RP and RMs significantly reduced ox-LDL levels and effectively suppressed the expressions of inflammatory mediators (COX-2, IL-1β, IL-6, TNF-α, iNOS). The anti-inflammatory molecular mechanism was to inhibit NF-κB activation and to simulate methionine sulfoxide reductase expression. Results showed, under cholesterol-enriched dietary condition, the anti-inflammatory action can be induced by RP and enhanced by methionine in growing and adult rats. The present study reveals a link of the decreased ox-LDL accumulation with the anti-inflammatory function of RP, which is dependent on methionine availability and independent of dietary cholesterol.

The effect of oxidized low-density lipoprotein (ox-LDL) on radiation-induced endothelial-to-mesenchymal transition

Kim, Miseon,Choi, Seo-Hyun,Jin, Yeung Bae,Lee, Hae-June,Ji, Young Hoon,Kim, Joon,Lee, Yun-Sil,Lee, Yoon-Jin Informa Healthcare 2013 International journal of radiation biology Vol.89 No.5

<P><I>Purpose</I>: Radiation-induced cardiovascular disease is a potentially severe side-effect of thoracic radiotherapy treatment. Clinically, this delayed side-effect presents as a form of accelerated atherosclerosis several years after irradiation. As general endothelial dysfunction is known to be an initiating event in radiation-induced vascular damage, we examined the effects of radiation on endothelial cells in radiation-induced atherosclerosis.</P><P><I>Materials and methods</I>: The effects of radiation on human aortic endothelial cells (HAoEC) were assessed by immunoblotting and immunofluorescence assays. Radiation-induced phenotypic changes of endothelial cells (ECs) were examined using atherosclerotic tissues of irradiated apoprotein E null (ApoE<SUP>−/−</SUP>) mice.</P><P><I>Results</I>: Radiation induced the HAoEC to undergo phenotypic conversion to form fibroblast-like cells, called the endothelial-to-mesenchymal transition (EndMT), which leads to the upregulation of mesenchymal cell markers such as alpha-smooth muscle actin (α-SMA), fibroblast specific protein-1 (FSP-1), and vimentin, and downregulation of endothelial cell-specific markers such as CD31 and vascular endothelial (VE)-cadherin. Furthermore, compared with low-density lipoprotein (LDL), oxidized low-density lipoprotein (ox-LDL) significantly augmented radiation-induced EndMT in HAoEC. These fibrotic phenotypes of ECs were found in atherosclerotic tissues of irradiated ApoE<SUP>−/−</SUP> mice with increased levels of ox-LDL.</P><P><I>Conclusions</I>: Taken together, these observations suggest that ox-LDL accelerates radiation-induced EndMT and subsequently contributes to radiation-induced atherosclerosis, providing a novel target for the prevention of radiation-induced atherosclerosis.</P>