title: Caspase-1 Inflammasome Activation Mediates Homocysteine-Induced Pyrop-Apoptosis in Endothelial Cells.
abstract: Endothelial injury is an initial mechanism mediating cardiovascular disease.
  
  
  
  Here, we investigated the effect of hyperhomocysteinemia on programed cell death in endothelial cells (EC).
  
  
  
  We established a novel flow-cytometric gating method to define pyrotosis (Annexin V(-)/Propidium iodide(+)). In cultured human EC, we found that: (1) homocysteine and lipopolysaccharide individually and synergistically induced inflammatory pyroptotic and noninflammatory apoptotic cell death; (2) homocysteine/lipopolysaccharide induced caspase-1 activation before caspase-8, caspase-9, and caspase-3 activations; (3) caspase-1/caspase-3 inhibitors rescued homocysteine/lipopolysaccharide-induced pyroptosis/apoptosis, but caspase-8/caspase-9 inhibitors had differential rescue effect; (4) homocysteine/lipopolysaccharide-induced nucleotide-binding oligomerization domain, and leucine-rich repeat and pyrin domain containing protein 3 (NLRP3) protein caused NLRP3-containing inflammasome assembly, caspase-1 activation, and interleukin (IL)-1β cleavage/activation; (5) homocysteine/lipopolysaccharide elevated intracellular reactive oxygen species, (6) intracellular oxidative gradient determined cell death destiny as intermediate intracellular reactive oxygen species levels are associated with pyroptosis, whereas high reactive oxygen species corresponded to apoptosis; (7) homocysteine/lipopolysaccharide induced mitochondrial membrane potential collapse and cytochrome-c release, and increased B-cell lymphoma 2-associated X protein/B-cell lymphoma 2 ratio which were attenuated by antioxidants and caspase-1 inhibitor; and (8) antioxidants extracellular superoxide dismutase and catalase prevented homocysteine/lipopolysaccharide -induced caspase-1 activation, mitochondrial dysfunction, and pyroptosis/apoptosis. In cystathionine β-synthase-deficient (Cbs(-/-)) mice, severe hyperhomocysteinemia-induced caspase-1 activation in isolated lung EC and caspase-1 expression in aortic endothelium, and elevated aortic caspase-1, caspase-9 protein/activity and B-cell lymphoma 2-associated X protein/B-cell lymphoma 2 ratio in Cbs(-/-) aorta and human umbilical vein endothelial cells. Finally, homocysteine-induced DNA fragmentation was reversed in caspase-1(-/-) EC. Hyperhomocysteinemia-induced aortic endothelial dysfunction was rescued in caspase-1(-/-) and NLRP3(-/-) mice.
  
  
  
  Hyperhomocysteinemia preferentially induces EC pyroptosis via caspase-1-dependent inflammasome activation leading to endothelial dysfunction. We termed caspase-1 responsive pyroptosis and apoptosis as pyrop-apoptosis.
authors.affiliation: From the Centers for Metabolic Disease Research (H.X., Y.Z., Y.X., W.Y.Y., X.J., J.Y., X.Y., H.W.), Cardiovascular Research (X.S., X.Y., H.W.), Thrombosis Research (X.Y., H.W.), Departments of Pharmacology (X.Y., H.W.), Neuroscience (X.Q.), Temple University School of Medicine, Philadelphia, PA; Department of Cardiology, Sun Yixian Memorial Hospital, Zhongshan University School of Medicine, Guangzhou, China (Y.Z., J.W.); Department of Cardiology, Second Hospital of Nanchang University, Institute of Cardiovascular Disease in Nanchang University, Nan Chang, Jiang Xi, China (Y.X., X.C.); and Key Laboratory of Cardiovascular Disease and Molecular Intervention, Nanjing Medical University, Nanjing, China (Y.J.).
authors.forename: Hang
authors.initials: H
authors.lastname: Xi
authors.affiliation: From the Centers for Metabolic Disease Research (H.X., Y.Z., Y.X., W.Y.Y., X.J., J.Y., X.Y., H.W.), Cardiovascular Research (X.S., X.Y., H.W.), Thrombosis Research (X.Y., H.W.), Departments of Pharmacology (X.Y., H.W.), Neuroscience (X.Q.), Temple University School of Medicine, Philadelphia, PA; Department of Cardiology, Sun Yixian Memorial Hospital, Zhongshan University School of Medicine, Guangzhou, China (Y.Z., J.W.); Department of Cardiology, Second Hospital of Nanchang University, Institute of Cardiovascular Disease in Nanchang University, Nan Chang, Jiang Xi, China (Y.X., X.C.); and Key Laboratory of Cardiovascular Disease and Molecular Intervention, Nanjing Medical University, Nanjing, China (Y.J.).
authors.forename: Yuling
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: From the Centers for Metabolic Disease Research (H.X., Y.Z., Y.X., W.Y.Y., X.J., J.Y., X.Y., H.W.), Cardiovascular Research (X.S., X.Y., H.W.), Thrombosis Research (X.Y., H.W.), Departments of Pharmacology (X.Y., H.W.), Neuroscience (X.Q.), Temple University School of Medicine, Philadelphia, PA; Department of Cardiology, Sun Yixian Memorial Hospital, Zhongshan University School of Medicine, Guangzhou, China (Y.Z., J.W.); Department of Cardiology, Second Hospital of Nanchang University, Institute of Cardiovascular Disease in Nanchang University, Nan Chang, Jiang Xi, China (Y.X., X.C.); and Key Laboratory of Cardiovascular Disease and Molecular Intervention, Nanjing Medical University, Nanjing, China (Y.J.).
authors.forename: Yanjie
authors.initials: Y
authors.lastname: Xu
authors.affiliation: From the Centers for Metabolic Disease Research (H.X., Y.Z., Y.X., W.Y.Y., X.J., J.Y., X.Y., H.W.), Cardiovascular Research (X.S., X.Y., H.W.), Thrombosis Research (X.Y., H.W.), Departments of Pharmacology (X.Y., H.W.), Neuroscience (X.Q.), Temple University School of Medicine, Philadelphia, PA; Department of Cardiology, Sun Yixian Memorial Hospital, Zhongshan University School of Medicine, Guangzhou, China (Y.Z., J.W.); Department of Cardiology, Second Hospital of Nanchang University, Institute of Cardiovascular Disease in Nanchang University, Nan Chang, Jiang Xi, China (Y.X., X.C.); and Key Laboratory of Cardiovascular Disease and Molecular Intervention, Nanjing Medical University, Nanjing, China (Y.J.).
authors.forename: William Y
authors.initials: WY
authors.lastname: Yang
authors.affiliation: From the Centers for Metabolic Disease Research (H.X., Y.Z., Y.X., W.Y.Y., X.J., J.Y., X.Y., H.W.), Cardiovascular Research (X.S., X.Y., H.W.), Thrombosis Research (X.Y., H.W.), Departments of Pharmacology (X.Y., H.W.), Neuroscience (X.Q.), Temple University School of Medicine, Philadelphia, PA; Department of Cardiology, Sun Yixian Memorial Hospital, Zhongshan University School of Medicine, Guangzhou, China (Y.Z., J.W.); Department of Cardiology, Second Hospital of Nanchang University, Institute of Cardiovascular Disease in Nanchang University, Nan Chang, Jiang Xi, China (Y.X., X.C.); and Key Laboratory of Cardiovascular Disease and Molecular Intervention, Nanjing Medical University, Nanjing, China (Y.J.).
authors.forename: Xiaohua
authors.initials: X
authors.lastname: Jiang
authors.affiliation: From the Centers for Metabolic Disease Research (H.X., Y.Z., Y.X., W.Y.Y., X.J., J.Y., X.Y., H.W.), Cardiovascular Research (X.S., X.Y., H.W.), Thrombosis Research (X.Y., H.W.), Departments of Pharmacology (X.Y., H.W.), Neuroscience (X.Q.), Temple University School of Medicine, Philadelphia, PA; Department of Cardiology, Sun Yixian Memorial Hospital, Zhongshan University School of Medicine, Guangzhou, China (Y.Z., J.W.); Department of Cardiology, Second Hospital of Nanchang University, Institute of Cardiovascular Disease in Nanchang University, Nan Chang, Jiang Xi, China (Y.X., X.C.); and Key Laboratory of Cardiovascular Disease and Molecular Intervention, Nanjing Medical University, Nanjing, China (Y.J.).
authors.forename: Xiaojin
authors.initials: X
authors.lastname: Sha
authors.affiliation: From the Centers for Metabolic Disease Research (H.X., Y.Z., Y.X., W.Y.Y., X.J., J.Y., X.Y., H.W.), Cardiovascular Research (X.S., X.Y., H.W.), Thrombosis Research (X.Y., H.W.), Departments of Pharmacology (X.Y., H.W.), Neuroscience (X.Q.), Temple University School of Medicine, Philadelphia, PA; Department of Cardiology, Sun Yixian Memorial Hospital, Zhongshan University School of Medicine, Guangzhou, China (Y.Z., J.W.); Department of Cardiology, Second Hospital of Nanchang University, Institute of Cardiovascular Disease in Nanchang University, Nan Chang, Jiang Xi, China (Y.X., X.C.); and Key Laboratory of Cardiovascular Disease and Molecular Intervention, Nanjing Medical University, Nanjing, China (Y.J.).
authors.forename: Xiaoshu
authors.initials: X
authors.lastname: Cheng
authors.affiliation: From the Centers for Metabolic Disease Research (H.X., Y.Z., Y.X., W.Y.Y., X.J., J.Y., X.Y., H.W.), Cardiovascular Research (X.S., X.Y., H.W.), Thrombosis Research (X.Y., H.W.), Departments of Pharmacology (X.Y., H.W.), Neuroscience (X.Q.), Temple University School of Medicine, Philadelphia, PA; Department of Cardiology, Sun Yixian Memorial Hospital, Zhongshan University School of Medicine, Guangzhou, China (Y.Z., J.W.); Department of Cardiology, Second Hospital of Nanchang University, Institute of Cardiovascular Disease in Nanchang University, Nan Chang, Jiang Xi, China (Y.X., X.C.); and Key Laboratory of Cardiovascular Disease and Molecular Intervention, Nanjing Medical University, Nanjing, China (Y.J.).
authors.forename: Jingfeng
authors.initials: J
authors.lastname: Wang
authors.affiliation: From the Centers for Metabolic Disease Research (H.X., Y.Z., Y.X., W.Y.Y., X.J., J.Y., X.Y., H.W.), Cardiovascular Research (X.S., X.Y., H.W.), Thrombosis Research (X.Y., H.W.), Departments of Pharmacology (X.Y., H.W.), Neuroscience (X.Q.), Temple University School of Medicine, Philadelphia, PA; Department of Cardiology, Sun Yixian Memorial Hospital, Zhongshan University School of Medicine, Guangzhou, China (Y.Z., J.W.); Department of Cardiology, Second Hospital of Nanchang University, Institute of Cardiovascular Disease in Nanchang University, Nan Chang, Jiang Xi, China (Y.X., X.C.); and Key Laboratory of Cardiovascular Disease and Molecular Intervention, Nanjing Medical University, Nanjing, China (Y.J.).
authors.forename: Xuebin
authors.initials: X
authors.lastname: Qin
authors.affiliation: From the Centers for Metabolic Disease Research (H.X., Y.Z., Y.X., W.Y.Y., X.J., J.Y., X.Y., H.W.), Cardiovascular Research (X.S., X.Y., H.W.), Thrombosis Research (X.Y., H.W.), Departments of Pharmacology (X.Y., H.W.), Neuroscience (X.Q.), Temple University School of Medicine, Philadelphia, PA; Department of Cardiology, Sun Yixian Memorial Hospital, Zhongshan University School of Medicine, Guangzhou, China (Y.Z., J.W.); Department of Cardiology, Second Hospital of Nanchang University, Institute of Cardiovascular Disease in Nanchang University, Nan Chang, Jiang Xi, China (Y.X., X.C.); and Key Laboratory of Cardiovascular Disease and Molecular Intervention, Nanjing Medical University, Nanjing, China (Y.J.).
authors.forename: Jun
authors.initials: J
authors.lastname: Yu
authors.affiliation: From the Centers for Metabolic Disease Research (H.X., Y.Z., Y.X., W.Y.Y., X.J., J.Y., X.Y., H.W.), Cardiovascular Research (X.S., X.Y., H.W.), Thrombosis Research (X.Y., H.W.), Departments of Pharmacology (X.Y., H.W.), Neuroscience (X.Q.), Temple University School of Medicine, Philadelphia, PA; Department of Cardiology, Sun Yixian Memorial Hospital, Zhongshan University School of Medicine, Guangzhou, China (Y.Z., J.W.); Department of Cardiology, Second Hospital of Nanchang University, Institute of Cardiovascular Disease in Nanchang University, Nan Chang, Jiang Xi, China (Y.X., X.C.); and Key Laboratory of Cardiovascular Disease and Molecular Intervention, Nanjing Medical University, Nanjing, China (Y.J.). hongw@temple.edu.
authors.forename: Yong
authors.initials: Y
authors.lastname: Ji
authors.affiliation: From the Centers for Metabolic Disease Research (H.X., Y.Z., Y.X., W.Y.Y., X.J., J.Y., X.Y., H.W.), Cardiovascular Research (X.S., X.Y., H.W.), Thrombosis Research (X.Y., H.W.), Departments of Pharmacology (X.Y., H.W.), Neuroscience (X.Q.), Temple University School of Medicine, Philadelphia, PA; Department of Cardiology, Sun Yixian Memorial Hospital, Zhongshan University School of Medicine, Guangzhou, China (Y.Z., J.W.); Department of Cardiology, Second Hospital of Nanchang University, Institute of Cardiovascular Disease in Nanchang University, Nan Chang, Jiang Xi, China (Y.X., X.C.); and Key Laboratory of Cardiovascular Disease and Molecular Intervention, Nanjing Medical University, Nanjing, China (Y.J.).
authors.forename: Xiaofeng
authors.initials: X
authors.lastname: Yang
authors.affiliation: From the Centers for Metabolic Disease Research (H.X., Y.Z., Y.X., W.Y.Y., X.J., J.Y., X.Y., H.W.), Cardiovascular Research (X.S., X.Y., H.W.), Thrombosis Research (X.Y., H.W.), Departments of Pharmacology (X.Y., H.W.), Neuroscience (X.Q.), Temple University School of Medicine, Philadelphia, PA; Department of Cardiology, Sun Yixian Memorial Hospital, Zhongshan University School of Medicine, Guangzhou, China (Y.Z., J.W.); Department of Cardiology, Second Hospital of Nanchang University, Institute of Cardiovascular Disease in Nanchang University, Nan Chang, Jiang Xi, China (Y.X., X.C.); and Key Laboratory of Cardiovascular Disease and Molecular Intervention, Nanjing Medical University, Nanjing, China (Y.J.). hongw@temple.edu.
authors.forename: Hong
authors.initials: H
authors.lastname: Wang
mesh_terms: D000818:Animals; D020170:Caspase 1 / Q000378:metabolism*; D003541:Cystathionine beta-Synthase / Q000235:genetics / Q000378:metabolism; D006710:Homocysteine / Q000633:toxicity; D061307:Human Umbilical Vein Endothelial Cells / Q000378:metabolism*; D006801:Humans; D058847:Inflammasomes / Q000378:metabolism*; D053583:Interleukin-1beta / Q000378:metabolism; D008070:Lipopolysaccharides / Q000633:toxicity; D051379:Mice; D008810:Mice, Inbred C57BL; D000071199:NLR Family, Pyrin Domain-Containing 3 Protein / Q000378:metabolism; D000069292:Pyroptosis; D017382:Reactive Oxygen Species / Q000378:metabolism; D051028:bcl-2-Associated X Protein / Q000378:metabolism
chemical_list: D058847:Inflammasomes; D053583:Interleukin-1beta; D008070:Lipopolysaccharides; D000071199:NLR Family, Pyrin Domain-Containing 3 Protein; C492716:Nlrp3 protein, mouse; D017382:Reactive Oxygen Species; D051028:bcl-2-Associated X Protein; D006710:Homocysteine; D020170:Caspase 1; D003541:Cystathionine beta-Synthase
keywords: apoptosis; caspase-1; endothelial cells; homocysteine; pyroptosis