scholarly journals Activation of neutral sphingomyelinase 2 through hyperglycemia contributes to endothelial apoptosis via vesicle-bound intercellular transfer of ceramides

2021 ◽  
Author(s):  
Andreas Zietzer ◽  
Alina Lisann Jahnel ◽  
Marko Bulic ◽  
Katharina Gutbrod ◽  
Philip Düsing ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Cardiovascular Health ◽  
Spectrometric Analysis ◽  
Human Coronary Artery ◽  
Intercellular Transfer ◽  
Neutral Sphingomyelinase ◽  
Endothelial Apoptosis ◽  
Important Means ◽  
Induced Apoptosis ◽  
Ceramide Accumulation

Abstract Background Pro-apoptotic and pro-inflammatory ceramides are crucially involved in atherosclerotic plaque development. Local cellular ceramide accumulation mediates endothelial apoptosis, especially in type 2 diabetes mellitus, which is a major cardiovascular risk factor. In recent years, large extracellular vesicles (lEVs) have been identified as an important means of intercellular communication and as regulators of cardiovascular health and disease. A potential role for lEVs as vehicles for ceramide transfer and inductors of diabetes-associated endothelial apoptosis has never been investigated. Methods and Results A mass-spectrometric analysis of human coronary artery endothelial cells (HCAECs) and their lEVs revealed C16 ceramide (d18:1–16:0) to be the most abundant ceramide in lEVs and to be significantly increased in lEVs after hyperglycemic injury to HCAECs. The increased packaging of ceramide into lEVs after hyperglycemic injury was shown to be dependent on neutral sphingomyelinase 2 (nSMase2), which was upregulated in glucose-treated HCAECs. lEVs from hyperglycemic HCAECs induced apoptosis in the recipient HCAECs compared to native lEVs from untreated HCAECs. Similarly, lEVs from hyperglycemic mice after streptozotocin injection induced higher rates of apoptosis in murine endothelial cells compared to lEVs from normoglycemic mice. To generate lEVs with high levels of C16 ceramide, ceramide was applied exogenously and shown to be effectively packaged into the lEVs, which then induced apoptosis in lEV-recipient HCAECs via activation of caspase 3. Intercellular transfer of ceramide through lEVs was confirmed by use of a fluorescently labeled ceramide analogue. Treatment of HCAECs with a pharmacological inhibitor of nSMases (GW4869) or siRNA-mediated downregulation of nSMase2 abrogated the glucose-mediated effect on apoptosis in lEV-recipient cells. In contrast, for small EVs (sEVs), hyperglycemic injury or GW4869 treatment had no effect on apoptosis induction in sEV-recipient cells. Conclusion lEVs mediate the induction of apoptosis in endothelial cells in response to hyperglycemic injury through intercellular transfer of ceramides. Graphical abstract

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

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.

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

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.

SPECIFIC TOXICITY OF CALCIUM PHOSPHATE BIONS FOR HUMAN VENOUS AND ARTERIAL ENDOTHELIAL CELLS

2019 ◽  
pp. 53-61
Author(s):  
Д.К. Шишкова ◽  
Е.А. Великанова ◽  
В.Г. Матвеева ◽  
Ю.А. Кудрявцева ◽  
А.Г. Кутихин
Keyword(s):  
Endothelial Cells ◽  
Coronary Artery ◽  
Calcium Phosphate ◽  
Exposure Time ◽  
Internal Thoracic Artery ◽  
Human Coronary Artery ◽  
Hoechst 33342 ◽  
Ethidium Bromide Staining ◽  
Induced Apoptosis ◽  
Arterial Endothelial Cells

Цель исследования - оценка токсичности кальций-фосфатных бионов (КФБ) и магний-фосфатных бионов (МФБ) для культур эндотелиальных клеток. Методика. Эндотелиотоксичность бионов изучена при помощи добавления равных концентраций МФБ или КФБ к: 1) разреженным или конфлюэнтным культурам иммортализованных венозных эндотелиальных клеток человека линии EA.hy 926 с последующим культивированием в течение 24 ч или 4 ч соответственно; 2) конфлюэнтным культурам коммерческих первичных эндотелиальных клеток коронарной и внутренней грудной артерии человека с последующим культивированием в течение 24 ч. Эндотелиотоксические эффекты бионов оценивали при помощи сочетанного окрашивания клеток флюоресцентными красителями Hoechst 33342 и бромистым этидием, а также посредством колориметрического теста. Кроме того, методом проточной цитометрии оценивали пути и стадии гибели клеток вышеуказанных культур. Результаты. В отличие от МФБ, КФБ индуцировали гибель эндотелиальных клеток всех 3 линий путем апоптоза. Устойчивость культур к токсическому действию КФБ определялась степенью их конфлюэнтности (конфлюэнтные культуры более устойчивы чем разреженные) и типом клеточной линии (эндотелиальные клетки внутренней грудной артерии продемонстрировали большую устойчивость в сравнении с эндотелиальными клетками коронарной артерии). Заключение. Токсичность КФБ для культур эндотелиальных клеток специфична, то есть определяется их специфическим минеральным составом, а не общей для всех типов бионов корпускулярной природой. Добавление КФБ к конфлюэнтным культурам первичных артериальных эндотелиальных клеток и к иммортализованным венозным эндотелиальным клеткам вызывало их гибель, при этом экспозиция МФБ не оказывает значимого токсического действия. Эндотелиальные клетки внутренней грудной артерии менее чувствительны к воздействию КФБ в сравнении с эндотелиальными клетками коронарной артерии человека. Aim. To compare toxicity of calcium phosphate bions (CPB) and magnesium phosphate bions (MPB) for endothelial cells. Me-thods. To assess endothelial toxicity of the bions, we first added equal concentrations of either MPB or CPB to: 1) non-confluent or confluent cultures of immortalized human venous endothelial cells EA.hy 926 with the exposure time of 24 h or 4 h, respectively; 2) confluent cultures of commercially available primary human coronary artery and internal thoracic artery endothelial cells, with the exposure time of 24 h. Endothelial toxicity was then evaluated by combined Hoechst 33342 and ethidium bromide staining following fluorescence microscopy and by colorimetric cytotoxicity assay. In addition, we attempted to determine the pathway of bion-induced cell death utilizing flow cytometry. Results. In contrast to the MPB, CPB induced apoptosis of all studied endothelial cell lines. Resistance of endothelial cells to the CPB was defined by their confluence (confluent cultures demonstrated higher resistance), and cell type (internal thoracic artery endothelial cells were more resistant to the CPB as compared to the coronary artery endothelial cells). Conclusions. Endothelial toxicity of the CPB is defined by their specific mineral composition but not by their corpuscular nature, which is common for all nanoparticles. Addition of the CPB to the confluent cultures of primary human arterial cells and to immortalized human venous endothelial cells evoked their death. On the contrary, exposure to the MPB did not cause any toxic effects. Human internal thoracic artery endothelial cells are more resistant to the CPB in comparison with coronary artery endothelial cells.

scholarly journals Exendin-4 protects endothelial cells from lipoapoptosis by PKA, PI3K, eNOS, p38 MAPK, and JNK pathways

2013 ◽  
Vol 50 (2) ◽  
pp. 229-241 ◽  
Author(s):  
Özlem Erdogdu ◽  
Linnéa Eriksson ◽  
Hua Xu ◽  
Åke Sjöholm ◽  
Qimin Zhang ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
P38 Mapk ◽  
Experimental Studies ◽  
Receptor Activation ◽  
No Production ◽  
Ros Production ◽  
Human Coronary Artery ◽  
Glucagon Like Peptide 1 ◽  
Underlying Mechanisms ◽  
Induced Apoptosis

Experimental studies have indicated that endothelial cells play an important role in maintaining vascular homeostasis. We previously reported that human coronary artery endothelial cells (HCAECs) express the glucagon-like peptide 1 (GLP1) receptor and that the stable GLP1 mimetic exendin-4 is able to activate the receptor, leading to increased cell proliferation. Here, we have studied the effect of exendin-4 and native GLP1 (7–36) on lipoapoptosis and its underlying mechanisms in HCAECs. Apoptosis was assessed by DNA fragmentation and caspase-3 activation, after incubating cells with palmitate. Nitric oxide (NO) and reactive oxidative species (ROS) were analyzed. GLP1 receptor activation, PKA-, PI3K/Akt-, eNOS-, p38 MAPK-, and JNK-dependent pathways, and genetic silencing of transfection of eNOS were also studied. Palmitate-induced apoptosis stimulated cells to release NO and ROS, concomitant with upregulation of eNOS, which required activation of p38 MAPK and JNK. Exendin-4 restored the imbalance between NO and ROS production in which ROS production decreased and NO production was further augmented. Incubation with exendin-4 and GLP1 (7–36) protected HCAECs against lipoapoptosis, an effect that was blocked by PKA, PI3K/Akt, eNOS, p38 MAPK, and JNK inhibitors. Genetic silencing of eNOS also abolished the anti-apoptotic effect afforded by exendin-4. Our results support the notion that GLP1 receptor agonists restore eNOS-induced ROS production due to lipotoxicity and that such agonists protect against lipoapoptosis through PKA-PI3K/Akt-eNOS-p38 MAPK-JNK-dependent pathways via a GLP1 receptor-dependent mechanism.

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

2018 ◽  
Vol 315 (3) ◽  
pp. C330-C340 ◽  
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.

Protein kinase C iota mediates lipid-induced apoptosis of human coronary artery endothelial cells

2009 ◽  
Vol 78 (1) ◽  
pp. 40-44 ◽  
Author(s):  
K. Staiger ◽  
U. Schatz ◽  
H. Staiger ◽  
P. Weyrich ◽  
C. Haas ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Protein Kinase C ◽  
Coronary Artery ◽  
Protein Kinase ◽  
Human Coronary Artery ◽  
Kinase C ◽  
Protein Kinase C Iota ◽  
Induced Apoptosis

Rho protein inactivation induced apoptosis of cultured human endothelial cells

2002 ◽  
Vol 283 (4) ◽  
pp. L830-L838 ◽  
Author(s):  
Stefan Hippenstiel ◽  
Bernd Schmeck ◽  
Phillipe Dje N'Guessan ◽  
Joachim Seybold ◽  
Matthias Krüll ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Rho Gtpases ◽  
Human Umbilical Cord ◽  
Caspase 9 ◽  
Toxin B ◽  
Protein Levels ◽  
Endothelial Apoptosis ◽  
Clostridium Difficile Toxin B ◽  
Induced Apoptosis ◽  
Camp Elevation

Small GTP-binding Rho GTPases regulate important signaling pathways in endothelial cells, but little is known about their role in endothelial cell apoptosis. Clostridial cytotoxins specifically inactivate GTPases by glucosylation [ Clostridium difficile toxin B-10463 (TcdB-10463), C. difficile toxin B-1470 (TcdB-1470)] or ADP ribosylation ( C. botulinum C3 toxin). Exposure of human umbilical cord vein endothelial cells (HUVEC) to TcdB-10463, which inhibits RhoA/Rac1/Cdc42, or to C3 toxin, which inhibits RhoA, -B, -C, resulted in apoptosis, whereas inactivation of Rac1/Cdc42 with TcdB-1470 was without effect, suggesting that Rho inhibition was responsible for endothelial apoptosis. Disruption of endothelial microfilaments as well as inhibition of p160ROCK did not induce endothelial apoptosis. Exposure to TcdB-10463 resulted in activation of caspase-9 and -3 but not caspase-8 in HUVEC. Moreover, Rho inhibition reduced expression of antiapoptotic Bcl-2 and Mcl-1 and increased proapoptotic Bid but had no effect on Bax or FLIP protein levels. Caspase-3 activity and apoptosis induced by TcdB-10463 were abolished by cAMP elevation. In summary, inhibition of Rho in endothelial cells activates caspase-9- and -3-dependent apoptosis, which can be antagonized by cAMP elevation.

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