Resveratrol attenuates cigarette smoke induced endothelial apoptosis by activating Notch1 signaling mediated autophagy

Abstract Background Increasing evidence shows that endothelial apoptosis contributes to cigarette smoke (CS)-induced disease progression, such as chronic obstructive pulmonary disease (COPD). Our previous studies have validated Notch1 as an anti-apoptotic signaling in CS-induced endothelial apoptosis. Resveratrol (RESV) is a naturally occurring polyphenol that exhibits an anti-apoptotic activity in endothelial cells that exposed to many kinds of destructive stimulus. However, the effects of resveratrol on Notch1 signaling in CS-induced endothelial apoptosis have not yet been fully elucidated. Therefore, the aim of this study was to examine whether RESV can protect endothelial cells from CS-induced apoptosis via regulating Notch1 signaling. Methods Human umbilical vein endothelial cells (HUVECs) were pretreated with RESV for 2 h, followed by cotreatment with 2.5%CSE for 24 h to explore the role of RESV in CSE induced endothelial apoptosis. 3-methyladenine (3-MA) or rapamycin was used to alter autophagic levels. Lentivirus Notch1 intracellular domain (LV-N1ICD), γ-secretase inhibitor (DAPT) and Notch1 siRNA were used to change Notch1 expression. The expression of Notch1, autophagic and apoptotic markers were examined by Western blot and the apoptosis rate was detected by Flow cytometry analysis. Results Our results showed that activating autophagy reduced CSE-induced endothelial apoptosis, while blocking autophagy promoted cell apoptosis in HUVECs. RESV pretreatment attenuated the CSE-induced endothelial apoptosis and activated Notch1 signaling. RESV pretreatment also increased LC3b-II and Beclin1 production, decreased p62 and mTOR expression. 3-MA treatment inhibited autophagy and aggravated CSE induced apoptosis, while rapamycin promoted autophagy, led to a decrease in cell apoptosis. LV-N1ICD transfection upregulated autophagy and reduced apoptosis. However, this protective effect was abolished by 3-MA treatment. In cells treated with DAPT or Notch1 siRNA, autophagy was decreased, while apoptosis was increased. RESV partly rescued the DAPT or Notch1 siRNA induced apoptosis by activating Notch1 signaling. Conclusion In HUVECs, RESV attenuates CSE induced endothelial apoptosis by inducing autophagy in a Notch1-dependent manner.

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Resveratrol attenuates cigarette smoke induced endothelial apoptosis by activating Notch1 signaling mediated autophagy

10.21203/rs.3.rs-27268/v2 ◽

2020 ◽

Author(s):

Dandan Zong ◽

Xiang-ming Liu ◽

Jin-hua Li ◽

Ruo-yun Ouyang ◽

Ying-jiao Long ◽

...

Keyword(s):

Endothelial Cells ◽

Cigarette Smoke ◽

Cell Apoptosis ◽

Umbilical Vein ◽

Chronic Obstructive ◽

Dependent Manner ◽

Obstructive Pulmonary Disease ◽

Endothelial Apoptosis ◽

Notch1 Signaling ◽

Induced Apoptosis

Abstract Background Increasing evidences have showed that endothelial apoptosis contributes to cigarette smoke (CS)-induced disease progression, such as chronic obstructive pulmonary disease (COPD). Our previous studies have validated Notch1 as an anti-apoptotic signaling in cigarette smoke (CS)-induced endothelial apoptosis. Resveratrol (RESV) is a naturally occurring polyphenol that exhibits an anti-apoptotic activity in endothelial cells that exposed to many kinds of destructive stimulus. However, the effects of resveratrol on Notch1 signaling in CS-induced endothelial apoptosis have not yet been fully elucidated. Therefore, the aim of this study was to examine whether RESV can protect endothelial cells from cigarette smoke induced apoptosis via regulating Notch1 signaling. Methods Human umbilical vein endothelial cells (HUVECs) were pretreated with RESV for 2 h, followed by cotreatment with 2.5%CSE for 24h to explore the role of RESV in CSE induced endothelial apoptosis. 3-methyladenine (3-MA) or rapamycin was used to alter autophagic levels. Lentivirus Notch1 intracellular domain (LV-N1ICD) or γ-secretase inhibitor (DAPT) were used to change Notch1 expression. The expression of Notch1, autophagic and apoptotic markers were examined by Western blot and the apoptosis rate was detected by Flow cytometry analysis. Results Our results showed that activating autophagy reduced CSE-induced endothelial apoptosis, while blocking autophagy promoted cell apoptosis in HUVECs. RESV pretreatment attenuated the CSE-induced endothelial apoptosis and activated Notch1 signaling. RESV pretreatment also increased LC3b-II and Beclin1 production, decreased p62 and mTOR expression. 3-MA treatment inhibited autophagy and aggravated CSE induced apoptosis, while rapamycin promoted autophagy, led to a decrease in cell apoptosis. LV-N1ICD transfection upregulated autophagy and reduced apoptosis. However, this protective effect was abolished by 3-MA treatment. In cells treated with DAPT, autophagy was decreased, while apoptosis was increased. RESV partly rescued the DAPT induced apoptosis by activating Notch1 signaling. Conclusion In HUVECs, RESV attenuates CSE induced endothelial apoptosis by inducing autophagy in a Notch1-dependent manner.

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Resveratrol attenuates cigarette smoke induced endothelial apoptosis by activating Notch1 signaling mediated autophagy

10.21203/rs.3.rs-27268/v3 ◽

2020 ◽

Author(s):

Dandan Zong ◽

Xiang-ming Liu ◽

Jin-hua Li ◽

Ruo-yun Ouyang ◽

Ying-jiao Long ◽

...

Keyword(s):

Endothelial Cells ◽

Cigarette Smoke ◽

Cell Apoptosis ◽

Umbilical Vein ◽

Chronic Obstructive ◽

Dependent Manner ◽

Obstructive Pulmonary Disease ◽

Endothelial Apoptosis ◽

Notch1 Signaling ◽

Induced Apoptosis

Abstract Background: Increasing evidences have showed that endothelial apoptosis contributes to cigarette smoke (CS)-induced disease progression, such as chronic obstructive pulmonary disease (COPD). Our previous studies have validated Notch1 as an anti-apoptotic signaling in cigarette smoke (CS)-induced endothelial apoptosis. Resveratrol (RESV) is a naturally occurring polyphenol that exhibits an anti-apoptotic activity in endothelial cells that exposed to many kinds of destructive stimulus. However, the effects of resveratrol on Notch1 signaling in CS-induced endothelial apoptosis have not yet been fully elucidated. Therefore, the aim of this study was to examine whether RESV can protect endothelial cells from cigarette smoke induced apoptosis via regulating Notch1 signaling. Methods: Human umbilical vein endothelial cells (HUVECs) were pretreated with RESV for 2 h, followed by cotreatment with 2.5%CSE for 24h to explore the role of RESV in CSE induced endothelial apoptosis. 3-methyladenine (3-MA) or rapamycin was used to alter autophagic levels. Lentivirus Notch1 intracellular domain (LV-N1ICD) or γ-secretase inhibitor (DAPT) were used to change Notch1 expression. The expression of Notch1, autophagic and apoptotic markers were examined by Western blot and the apoptosis rate was detected by Flow cytometry analysis. Results: Our results showed that activating autophagy reduced CSE-induced endothelial apoptosis, while blocking autophagy promoted cell apoptosis in HUVECs. RESV pretreatment attenuated the CSE-induced endothelial apoptosis and activated Notch1 signaling. RESV pretreatment also increased LC3b-II and Beclin1 production, decreased p62 and mTOR expression. 3-MA treatment inhibited autophagy and aggravated CSE induced apoptosis, while rapamycin promoted autophagy, led to a decrease in cell apoptosis. LV-N1ICD transfection upregulated autophagy and reduced apoptosis. However, this protective effect was abolished by 3-MA treatment. In cells treated with DAPT, autophagy was decreased, while apoptosis was increased. RESV partly rescued the DAPT induced apoptosis by activating Notch1 signaling. Conclusion: In HUVECs, RESV attenuates CSE induced endothelial apoptosis by inducing autophagy in a Notch1-dependent manner.

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Resveratrol attenuates cigarette smoke induced endothelial apoptosis by activating Notch1 signaling mediated autophagy

10.21203/rs.3.rs-27268/v1 ◽

2020 ◽

Author(s):

Dandan Zong ◽

Xiang-ming Liu ◽

Jin-hua Li ◽

Ruo-yun Ouyang ◽

Ying-jiao Long ◽

...

Keyword(s):

Endothelial Cells ◽

Cigarette Smoke ◽

Cell Apoptosis ◽

Chronic Obstructive ◽

Dependent Manner ◽

Obstructive Pulmonary Disease ◽

Endothelial Apoptosis ◽

Notch1 Signaling ◽

Apoptotic Activity ◽

Induced Apoptosis

Abstract Background Endothelial apoptosis contributes to the pathogenesis of chronic obstructive pulmonary disease (COPD). Our previous studies have validated Notch1 as an anti-apoptotic signaling in cigarette smoke (CS)-induced endothelial apoptosis. Resveratrol (RESV) is a naturally occurring polyphenol that exhibits an anti-apoptotic activity in endothelial cells that exposed to many kinds of destructive stimulus. However, the effects of resveratrol on Notch1 signaling in CS-induced endothelial apoptosis have not yet been fully elucidated. Therefore, the aim of this study was to examine whether RESV can protect endothelial cells from cigarette smoke induced apoptosis via regulating Notch1 signaling. Methods HUVECs were pretreated with RESV for 2 h, followed by cotreatment with 2.5%CSE for 24h to explore the role of RESV in CSE induced endothelial apoptosis. 3-MA or rapamycin was used to alter autophagic levels. Lentivirus Notch1 intracellular domain (LV-N1ICD) or γ-secretase inhibitor (DAPT) were used to change Notch1 expression. The expression of Notch1, autophagic and apoptotic markers were examined by Western blot and the apoptosis rate was detected by Flow cytometry analysis. Results Our results showed that activating autophagy reduced CSE-induced endothelial apoptosis, while blocking autophagy promoted cell apoptosis in HUVECs. RESV pretreatment attenuated the CSE-induced endothelial apoptosis and activated Notch1 signaling. RESV pretreatment also increased LC3b-II and Beclin1 production, decreased p62 and mTOR expression. 3-MA treatment inhibited autophagy and aggravated CSE induced apoptosis, while rapamycin promoted autophagy, led to a decrease in cell apoptosis. LV-N1ICD transfection upregulated autophagy and reduced apoptosis. However, this protective effect was abolished by 3-MA treatment. In cells treated with DAPT, autophagy was decreased, while apoptosis was increased. RESV partly rescued the DAPT induced apoptosis by activating Notch1 signaling. Conclusion In HUVECs, RESV attenuates CSE induced endothelial apoptosis by inducing autophagy in a Notch1-dependent manner.

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Notch1 regulates endothelial apoptosis via the ERK pathway in chronic obstructive pulmonary disease

AJP Cell Physiology ◽

10.1152/ajpcell.00182.2017 ◽

2018 ◽

Vol 315 (3) ◽

pp. C330-C340 ◽

Cited By ~ 17

Author(s):

Dandan Zong ◽

Jinhua Li ◽

Shan Cai ◽

Shengdong He ◽

Qingqing Liu ◽

...

Keyword(s):

Chronic Obstructive Pulmonary Disease ◽

Endothelial Cells ◽

Notch Signaling ◽

Signaling Pathway ◽

Notch Signaling Pathway ◽

Chronic Obstructive ◽

Erk Pathway ◽

Obstructive Pulmonary Disease ◽

Endothelial Apoptosis ◽

Induced Apoptosis

The Notch signaling pathway plays critical role for determining cell fate by controlling proliferation, differentiation, and apoptosis. In the current study, we investigated the roles of the Notch signaling pathway in cigarette smoke (CS)-induced endothelial apoptosis in chronic obstructive pulmonary disease (COPD). We obtained surgical specimens from 10 patients with COPD and 10 control participants. Notch1, 2, and 4 express in endothelial cells, whereas Notch3 mainly localizes in smooth muscle cells. Compared with control groups, we found that the expression of Notch1, 3, and 4 decreased, as well as their target genes Hes1 and Hes2, while the expression of Notch2 and extracellular signal-regulated kinase (ERK)1/2 increased in COPD patients compared with controls, as well as in human pulmonary microvascular endothelial cells (HPMECs) when exposed to CS extract (CSE). Overexpression of Notch1 with N1ICD in HPMECs markedly alleviated the cell apoptosis induced by CSE. The ERK signaling pathway was significantly activated by CSE, which correlated with CSE-induced apoptosis. However, this activation can be abolished by N1ICD overexpression. Furthermore, treatment of PD98059 (ERK inhibitor) significantly alleviated CSE-induced apoptosis, as well as reduced the methylation of mitochondrial transcription factor A (mtTFA) promoter, which was correlated with CS-induced endothelial apoptosis. These results suggest that CS alters Notch signaling in pulmonary endothelial cells. Notch1 protects against CS-induced endothelial apoptosis in COPD through inhibiting the ERK pathway, while the ERK pathway further regulates the methylation of mtTFA promotor.

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Polydatin Prevents Methylglyoxal-Induced Apoptosis through Reducing Oxidative Stress and Improving Mitochondrial Function in Human Umbilical Vein Endothelial Cells

Oxidative Medicine and Cellular Longevity ◽

10.1155/2017/7180943 ◽

2017 ◽

Vol 2017 ◽

pp. 1-9 ◽

Cited By ~ 17

Author(s):

Ningbo Pang ◽

Tangting Chen ◽

Xin Deng ◽

Ni Chen ◽

Rong Li ◽

...

Keyword(s):

Oxidative Stress ◽

Endothelial Cells ◽

Mitochondrial Function ◽

Cell Apoptosis ◽

Umbilical Vein ◽

Ros Generation ◽

Mitochondrial Morphology ◽

Human Umbilical Vein ◽

Induced Apoptosis ◽

Umbilical Vein Endothelial Cells

Methylglyoxal (MGO), an active metabolite of glucose, has been reported to induce vascular cell apoptosis in diabetic complication. Polydatin (PD), a small natural compound from Polygonum cuspidatum, has a number of biological functions, such as antioxidative, anti-inflammatory, and nephroprotective properties. However, the protective effects of PD on MGO-induced apoptosis in endothelial cells remain to be elucidated. In this study, human umbilical vein endothelial cells (HUVECs) were used to explore the effects of PD on MGO-induced cell apoptosis and the possible mechanism involved. HUVECs were pretreated with PD for 2 h, followed by stimulation with MGO. Then cell apoptosis, reactive oxygen species (ROS) generation, mitochondrial membrane potential (MMP) impairment, mitochondrial morphology alterations, and Akt phosphorylation were assessed. The results demonstrated that PD significantly prevented MGO-induced HUVEC apoptosis. PD pretreatment also significantly inhibited MGO-induced ROS production, MMP impairment, mitochondrial morphology changes, and Akt dephosphorylation. These results and the experiments involving N-acetyl cysteine (antioxidant), Cyclosporin A (mitochondrial protector), and LY294002 (Akt inhibitor) suggest that PD prevents MGO-induced HUVEC apoptosis, at least in part, through inhibiting oxidative stress, maintaining mitochondrial function, and activating Akt pathway. All of these data indicate the potential application of PD for the treatment of diabetic vascular complication.

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Prohibitin protects against cigarette smoke extract-induced cell apoptosis in cultured human pulmonary microvascular endothelial cells

10.1101/2020.04.30.071589 ◽

2020 ◽

Author(s):

Yating Peng ◽

Zijing Zhou ◽

Aiyuan Zhou ◽

JiaXi Duan ◽

Hong Peng ◽

...

Keyword(s):

Endothelial Cells ◽

Cigarette Smoke ◽

Cell Apoptosis ◽

Cigarette Smoke Extract ◽

Microvascular Endothelial Cells ◽

Cigarette Smoke Exposure ◽

Dependent Manner ◽

Pulmonary Microvascular Endothelial Cells ◽

Mitochondrial Transcription Factor ◽

Microvascular Endothelial

Prohibitin is an evolutionarily conserved and ubiquitously expressed protein in eukaryocyte. It mediate many important roles in cell survival, apoptosis, autophagy and senescence. In the present study, we aimed to explore the role of prohibitin in cigarette smoke extract (CSE)-induced apoptosis of human pulmonary microvascular endothelial cells (HPMECs). For this purpose, HPMECs were trasfected with prohibitin and challenged with CSE. Our results showed that CSE exposure inhibited prohibitin expression in a dose-dependent manner in HPMECs. Overexpression of prohibitin could protect cell from CSE-induced injury by inhibiting CSE-induced cell apoptosis, inhibiting reactive oxygen species (ROS) production, increase mitochondrial membrane potential, increase the content of mitochondrial transcription factor A (mtTFA), IKKα/β phosphorylation and IκB-α degradation. CSE decreases prohibitin expression in endothelial cells and restoration of prohibitin expression in these cells can protect against the deleterious effects of CSE on mitochondrial and cells. We identified prohibitin is a novel regulator of endothelial cell apoptosis and survival in the context of cigarette smoke exposure.

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LncRNA-CCAT1/miR-152-5p is Involved in CSE-Induced Inflammation in HBE Cells Via Regulating ERK Signaling Pathway

10.21203/rs.3.rs-940217/v1 ◽

2021 ◽

Author(s):

Dan-dan Zong ◽

Xiang-ming Liu ◽

Jin-hua Li ◽

Ying-jiao Long ◽

Ruo-yun Ouyang ◽

...

Keyword(s):

Signaling Pathway ◽

Cigarette Smoke ◽

Specific Inhibitor ◽

Erk Signaling ◽

Luciferase Reporter ◽

Enzyme Linked Immunosorbent Assay ◽

Chronic Obstructive ◽

Dependent Manner ◽

Obstructive Pulmonary Disease ◽

Qrt Pcr

Abstract Background: Emerging studies have noted that dysregulated long non-coding RNAs (lncRNAs) are implicated in the pathological processes of chronic obstructive pulmonary disease (COPD). LncRNA colon cancer-associated transcript 1 (CCAT1) plays well-defined roles in the inflammatory progression. The study aims to figure out the effect and regulatory mechanism of CCAT1 in the cigarette smoke induced inflammation in COPD. Methods: The expression levels of CCAT1 and miR-152-3p were detected by quantitative real-time polymerase chain reaction (qRT-PCR). The inflammatory levels of IL-1β and IL-6 were evaluated by qRT-PCR and enzyme-linked immunosorbent assay (ELISA). Western blot was used for the measurement of ERK1/2, p-ERK1/2 protein levels. Luciferase reporter assay was performed for the target gene analysis.Results: CCAT1 was highly expressed in lung tissues of smokers with COPD compared with non-smokers without COPD samples. In human bronchial epithelial (HBE) cells, cigarette smoke extract (CSE) treatment led to an increase in CCAT1 expression in a dose- and time- dependent manner. Functional experiments showed that knockdown of CCAT1 ameliorated CSE-induced inflammation. Mechanistically, CCAT1 directly targeted miR-152-3p, and miR-197-3p overexpression reversed the pro-inflammatory effects of CCAT1 on HBE cells. Subsequently, miR-152-3p was found to regulate ERK signaling pathway. PD98059, ERK specific inhibitor, reversed miR-152-3p inhibition mediated inflammation in HBE cells. In addition, CCAT1 acted as a sponge for miR-152-3p to positively regulate ERK signaling pathway. Conclusion: Current findings suggest that CCAT1 promoted inflammation by activating ERK signal pathway via sponging miR‑152‑3p in CSE‑treated HBE cells. These results may provide a novel therapeutic target for alleviating cigarette smoke mediated airway inflammation.

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Endothelial Progenitor Cells as Pathogenetic and Diagnostic Factors, and Potential Targets for GLP-1 in Combination with Metabolic Syndrome and Chronic Obstructive Pulmonary Disease

International Journal of Molecular Sciences ◽

10.3390/ijms20051105 ◽

2019 ◽

Vol 20 (5) ◽

pp. 1105 ◽

Cited By ~ 3

Author(s):

Evgenii Skurikhin ◽

Olga Pershina ◽

Angelina Pakhomova ◽

Edgar Pan ◽

Vyacheslav Krupin ◽

...

Keyword(s):

Chronic Obstructive Pulmonary Disease ◽

Metabolic Syndrome ◽

Endothelial Cells ◽

Progenitor Cells ◽

Pulmonary Disease ◽

Endothelial Progenitor Cells ◽

Cigarette Smoke ◽

Chronic Obstructive ◽

Obstructive Pulmonary Disease

In clinical practice, there are patients with a combination of metabolic syndrome (MS) and chronic obstructive pulmonary disease (COPD). The pathological mechanisms linking MS and COPD are largely unknown. It remains unclear whether the effect of MS (possible obesity) has a major impact on the progression of COPD. This complicates the development of effective approaches for the treatment of patients with a diagnosis of MS and COPD. Experiments were performed on female C57BL/6 mice. Introduction of monosodium glutamate and extract of cigarette smoke was modeled to simulate the combined pathology of lipid disorders and emphysema. Biological effects of glucagon-like peptide 1 (GLP-1) and GLP-1 on endothelial progenitor cells (EPC) in vitro and in vivo were evaluated. Histological, immunohistochemical methods, biochemical methods, cytometric analysis of markers identifying EPC were used in the study. The CD31+ endothelial cells in vitro evaluation was produced by Flow Cytometry and Image Processing of each well with a Cytation™ 3. GLP-1 reduces the area of emphysema and increases the number of CD31+ endothelial cells in the lungs of mice in conditions of dyslipidemia and damage to alveolar tissue of cigarette smoke extract. The regenerative effects of GLP-1 are caused by a decrease in inflammation, a positive effect on lipid metabolism and glucose metabolism. EPC are proposed as pathogenetic and diagnostic markers of endothelial disorders in combination of MS with COPD. Based on GLP-1, it is proposed to create a drug to stimulate the regeneration of endothelium damaged in MS and COPD.

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The Protective Effect of Tetrahydroxystilbene Glucoside on High Glucose-Induced Injury in Human Umbilical Vein Endothelial Cells through the PI3K/Akt/eNOS Pathway and Regulation of Bcl-2/Bax

Journal of Vascular Research ◽

10.1159/000511035 ◽

2021 ◽

pp. 1-10

Author(s):

Jiankun Cui ◽

Bo Zhang ◽

Min Gao ◽

Baohai Liu ◽

Cong Dai ◽

...

Keyword(s):

Nitric Oxide ◽

Endothelial Cells ◽

Protective Effect ◽

High Glucose ◽

Umbilical Vein ◽

Vascular Complications ◽

Dependent Manner ◽

Human Umbilical Vein ◽

Induced Apoptosis ◽

Umbilical Vein Endothelial Cells

Endothelial dysfunction plays a central role in the pathogenesis of diabetic vascular complications. 2,3,5,4′-tetrahydroxystilbene-2-O-β-D-glucoside (TSG), an active component extracted from the roots of Polygonum multiflorum Thunb, has been shown to have strong antioxidant and antiapoptotic activities. In the present study, we investigated the protective effect of TSG on apoptosis induced by high glucose in human umbilical vein endothelial cells (HUVECs) and the possible mechanisms. Our data demonstrated that TSG significantly reversed the high glucose-induced decrease in cell viability, suppressed high glucose-induced generation of intracellular reactive oxygen species (ROS), the activity of caspase-3, and decreased the percentage of apoptotic cells in a dose-dependent manner. In addition, we found that TSG not only increased the expression of Bcl-2, while decreasing Bax expression, but also activated phosphorylation of Akt and endothelial nitric oxide synthase (eNOS) with subsequent nitric oxide production and ultimately reduced high glucose-induced apoptosis. However, the antiapoptotic effects of TSG were abrogated by pretreatment of the cells with PI3K inhibitor (LY294002) or eNOS inhibitor N<sup>G</sup>-L-nitro-arginine methyl ester, respectively. These results suggest that TSG inhibits high glucose-induced apoptosis in HUVECs through inhibition of ROS production, activation of the PI3K/Akt/eNOS pathway, and upregulation of the Bcl-2/Bax ratio, and thus may demonstrate significant potential for preventing diabetic cardiovascular complications.

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Purification, molecular cloning and mechanism of action of graminelysin I, a snake-venom-derived metalloproteinase that induces apoptosis of human endothelial cells

Biochemical Journal ◽

10.1042/bj3570719 ◽

2001 ◽

Vol 357 (3) ◽

pp. 719-728 ◽

Cited By ~ 20

Author(s):

Wen Bin WU ◽

Shin C. CHANG ◽

Ming-Yi LIAU ◽

Tur-Fu HUANG

Keyword(s):

Endothelial Cells ◽

Umbilical Vein ◽

Matrix Proteins ◽

Dependent Manner ◽

Single Chain ◽

Human Endothelial Cells ◽

Putative Precursor ◽

Induced Apoptosis ◽

Umbilical Vein Endothelial Cells ◽

Dose Dependent

Apoptosis, a programmed, physiological mode of cell death, is important in tissue homoeostasis. Here we report that a new metalloproteinase, graminelysin I, purified from Trimeresurus gramineus venom, induced apoptosis of human endothelial cells as examined by electrophoresis and flow cytometry. Graminelysin I contains only a metalloproteinase domain. It is a single-chain proteinase with a molecular mass of 27020Da. cDNA sequence analysis revealed that the disintegrin-like and cysteine-rich domains of the putative precursor protein of graminelysin I are likely to be processed post-translationally, producing the proteinase domain (graminelysin I). Graminelysin I cleaved the α chain of fibrinogen preferentially and cleaved the β chain either on longer incubation or at higher concentration. Graminelysin I inhibited the adhesion of human umbilical-vein endothelial cells (HUVECs) to immobilized fibrinogen and induced HUVECs detachment in a dose-dependent manner. These effects on HUVECs were abolished when graminelysin I was pretreated with EDTA. However, graminelysin I did not inhibit the adhesion of HUVECs to immobilized collagen. HUVECs were susceptible to death after treatment with graminelysin I when they were cultured on immobilized fibrinogen. In contrast, HUVECs were rather resistant to treatment with graminelysin I if they were cultured on immobilized collagen. Furthermore, graminelysin I induced apoptosis of HUVECs in a dose-dependent manner. Similarly, its apoptosis-inducing activity was blocked if it was treated with EDTA. These results suggest that the catalytic activity of graminelysin I on matrix proteins contributes to its apoptosis-inducing activity.

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