title: Phosphocreatine protects endothelial cells from oxidized low-density lipoprotein-induced apoptosis by modulating the PI3K/Akt/eNOS pathway.
abstract: Endothelial apoptosis triggered by oxidized low-density lipoprotein (oxLDL) can accelerate the progression of endothelial dysfunction atherosclerosis. Phosphocreatine (PCr) is a natural compound, which has been used in cardiac disease and cardiopulmonary resuscitation. However, its protective effects on atherosclerosis and its mechanism have not been clarified. In the present study, we investigated the anti-apoptotic effect of phosphocreatine in human umbilical vein endothelial cells (HUVECs) exposed to oxLDL and explored the possible mechanisms. HUVECs were pre-treated with 10-30 mM PCr and then stimulated with oxLDL. Cell morphology, cytotoxicity and apoptosis were evaluated by light microscopy, CCK assay, and flow cytometry respectively. Levels of Bax, Bcl-2, protein expression of protein kinase B (Akt), eNOS and caspase activities were assessed by Western blotting. Reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) were measured with fluorescent probes. Lactate dehydrogenase (LDH), malondialdehyde (MDA), nitric oxide (NO) and superoxide dismutase (SOD) contents were determined by spectrophotometer. Our results showed that PCr dose-dependently prevented oxLDL associated HUVEC cytotoxicity and apoptotic biochemical changes such as loss of MMP, LDH and MDA leakage and loss of SOD, decrease of Bcl-2/Bax protein ratio, activation of caspase-3 and 9, and ROS generation. In addition, the antiapoptotic effect of PCr was partially inhibited by a PI3K inhibitor (LY294002) and also enhanced p-Akt/Akt protein ratio, eNOS activation and NO production. In conclusion, our data show that the inhibition of oxLDL-induced endothelial apoptosis by PCr is due, at least in part to its anti-oxidant activity and its ability to modulate the PI3K/Akt/eNOS signaling pathway.
authors.affiliation: Department of Pharmacology, Dalian Medical University, West Section 9, South Road of Lushun, Dalian, 116044, China.
authors.forename: Anil
authors.initials: A
authors.lastname: Ahsan
authors.affiliation: Department of Pharmacology, Dalian Medical University, West Section 9, South Road of Lushun, Dalian, 116044, China.
authors.forename: Guozhu
authors.initials: G
authors.lastname: Han
authors.affiliation: Shanghai Huatuo Pharmaceutical Technology Development Limited Company, No. 590, Building 7, Layer 4, Road Ruiqing, Pudong New Area, Shanghai, China.
authors.forename: Junfang
authors.initials: J
authors.lastname: Pan
authors.affiliation: Department of Pharmacology, Dalian Medical University, West Section 9, South Road of Lushun, Dalian, 116044, China.
authors.forename: Shumin
authors.initials: S
authors.lastname: Liu
authors.affiliation: Biochemistry and Molecular Biology Department, Dalian Medical University, Dalian, China.
authors.forename: Arshad Ahmed
authors.initials: AA
authors.lastname: Padhiar
authors.affiliation: Department of Pharmacology, Dalian Medical University, West Section 9, South Road of Lushun, Dalian, 116044, China.
authors.forename: Peng
authors.initials: P
authors.lastname: Chu
authors.affiliation: Department of Pharmacology, Dalian Medical University, West Section 9, South Road of Lushun, Dalian, 116044, China.
authors.forename: Zhengwu
authors.initials: Z
authors.lastname: Sun
authors.affiliation: Department of Pharmacology, Dalian Medical University, West Section 9, South Road of Lushun, Dalian, 116044, China.
authors.forename: Zonghui
authors.initials: Z
authors.lastname: Zhang
authors.affiliation: Department of Pharmacology, Dalian Medical University, West Section 9, South Road of Lushun, Dalian, 116044, China.
authors.forename: Bin
authors.initials: B
authors.lastname: Sun
authors.affiliation: Department of Pharmacology, Dalian Medical University, West Section 9, South Road of Lushun, Dalian, 116044, China.
authors.forename: Jingjun
authors.initials: J
authors.lastname: Wu
authors.affiliation: Dalian University of Foreign Language, Dalian, China.
authors.forename: Aisha
authors.initials: A
authors.lastname: Irshad
authors.affiliation: Department of Pharmacology, Dalian Medical University, West Section 9, South Road of Lushun, Dalian, 116044, China.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Lin
authors.affiliation: Department of Pharmacology, Dalian Medical University, West Section 9, South Road of Lushun, Dalian, 116044, China.
authors.forename: Jinyong
authors.initials: J
authors.lastname: Peng
authors.affiliation: Department of Pharmacology, Dalian Medical University, West Section 9, South Road of Lushun, Dalian, 116044, China. tangzeyao@aliyun.com.
authors.forename: Zeyao
authors.initials: Z
authors.lastname: Tang
mesh_terms: D000975:Antioxidants / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3 / Q000378:metabolism; D053453:Caspase 9 / Q000378:metabolism; D002460:Cell Line; D002867:Chromones / Q000494:pharmacology; D061307:Human Umbilical Vein Endothelial Cells / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D008077:Lipoproteins, LDL / Q000378:metabolism*; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D009025:Morpholines / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010725:Phosphocreatine / Q000494:pharmacology*; D020011:Protective Agents / Q000494:pharmacology; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism* / Q000494:pharmacology; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013482:Superoxide Dismutase / Q000378:metabolism; D051028:bcl-2-Associated X Protein / Q000378:metabolism
chemical_list: D000975:Antioxidants; D002867:Chromones; D008077:Lipoproteins, LDL; D009025:Morpholines; D020011:Protective Agents; D019253:Proto-Oncogene Proteins c-bcl-2; D017382:Reactive Oxygen Species; D051028:bcl-2-Associated X Protein; C099252:oxidized low density lipoprotein; D010725:Phosphocreatine; D009569:Nitric Oxide; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C496314:NOS3 protein, human; D052250:Nitric Oxide Synthase Type III; D013482:Superoxide Dismutase; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt; D053148:Caspase 3; D053453:Caspase 9
keywords: Akt; Apoptosis; Endothelium; PCr; ROS; oxLDL