scholarly journals Functional Evidence for Biased Inhibition of G protein Signaling by YM-254890 in Human Coronary Artery Endothelial Cells

2020 ◽  
Author(s):  
Qianman Peng ◽  
Saud Alqahtani ◽  
Mohammed Zahid A Nasrullah ◽  
Jianzhong Shen
Keyword(s):  
Endothelial Cells ◽  
Coronary Artery ◽  
G Protein ◽  
G Proteins ◽  
Mapk Signaling ◽  
Selective Inhibitor ◽  
Functional Selectivity ◽  
Human Coronary Artery ◽  
Α Subunit ◽  
Camp Elevation

ABSTRACTSmall molecular chemicals targeting individual subtype of G proteins including Gs, Gi/o and Gq has been lacking, except for pertussis toxin being an established selective peptide inhibitor of the Gi/o protein. Recently, a cyclic depsipeptide compound YM-254890 isolated from culture broth of Chromobacterium sp. was reported as a selective inhibitor for the Gq protein by blocking GDP exchange of GTP on the α subunit of Gq complex. However, functional selectivity of YM-254890 towards various G proteins was not fully characterized, primarily due to its restricted availability before 2017. Here, using human coronary artery endothelial cells as a model, we performed a systemic pharmacological evaluation on the functional selectivity of YM-254890 on multiple G protein-mediated receptor signaling. First, we confirmed that YM-254890, at 30 nM, abolished UTP-activated P2Y2 receptor-mediated Ca2+ signaling and ERK1/2 phosphorylation, indicating its potent inhibition on the Gq protein. However, we unexpectedly found that YM-254890 also significantly suppressed cAMP elevation and ERK1/2 phosphorylation induced by multiple Gs-coupled receptors including β2-adrenegic, adenosine A2 and PGI2 receptors. Surprisingly, although YM-254890 had no impact on CXCR4/Gi/o protein-mediated suppression of cAMP production, it abolished ERK1/2 activation. Further, no cellular toxicity was observed for YM-254890, and it neither affected A23187- or thapsigargin-induced Ca2+ signaling, nor forskolin-induced cAMP elevation and growth factor-induced MAPK signaling. We conclude that YM-254890 is not a selective inhibitor for Gq protein; instead, it acts as a broad spectrum inhibitor for Gq and Gs proteins and exhibits a biased inhibition on Gi/o signaling, without affecting non-GPCR-mediated cellular signaling.

Inhibition of Pro-Inflammatory Cytokine Secretion by Select Antioxidants in Human Coronary Artery Endothelial Cells

2020 ◽  
Vol 90 (1-2) ◽  
pp. 103-112 ◽  
Author(s):  
Michael J. Haas ◽  
Marilu Jurado-Flores ◽  
Ramadan Hammoud ◽  
Victoria Feng ◽  
Krista Gonzales ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Ascorbic Acid ◽  
Coronary Artery ◽  
Inflammatory Cytokines ◽  
Inflammatory Cytokine ◽  
Culture Media ◽  
Cytokine Secretion ◽  
Human Coronary Artery ◽  
Tnf Α ◽  
Pro Inflammatory Cytokines

Abstract. Inflammatory and oxidative stress in endothelial cells are implicated in the pathogenesis of premature atherosclerosis in diabetes. To determine whether high-dextrose concentrations induce the expression of pro-inflammatory cytokines, human coronary artery endothelial cells (HCAEC) were exposed to either 5.5 or 27.5 mM dextrose for 24-hours and interleukin-1β (IL-1β), interleukin-2 (IL-2), interleukin-6 (IL-6), interleukin-8 (IL-8), and tumor necrosis factor α (TNF α) levels were measured by enzyme immunoassays. To determine the effect of antioxidants on inflammatory cytokine secretion, cells were also treated with α-tocopherol, ascorbic acid, and the glutathione peroxidase mimetic ebselen. Only the concentration of IL-1β in culture media from cells exposed to 27.5 mM dextrose increased relative to cells maintained in 5.5 mM dextrose. Treatment with α-tocopherol (10, 100, and 1,000 μM) and ascorbic acid (15, 150, and 1,500 μM) at the same time that the dextrose was added reduced IL-1β, IL-6, and IL-8 levels in culture media from cells maintained at 5.5 mM dextrose but had no effect on IL-1β, IL-6, and IL-8 levels in cells exposed to 27.5 mM dextrose. However, ebselen treatment reduced IL-1β, IL-6, and IL-8 levels in cells maintained in either 5.5 or 27.5 mM dextrose. IL-2 and TNF α concentrations in culture media were below the limit of detection under all experimental conditions studied suggesting that these cells may not synthesize detectable quantities of these cytokines. These results suggest that dextrose at certain concentrations may increase IL-1β levels and that antioxidants have differential effects on suppressing the secretion of pro-inflammatory cytokines in HCAEC.

scholarly journals Infection of Human Coronary Artery Endothelial Cells by Group B Streptococcus Contributes to Dysregulation of Apoptosis, Hemostasis, and Innate Immune Responses

2011 ◽  
Vol 2011 ◽  
pp. 1-8 ◽  
Author(s):  
Claudia Beyrich ◽  
Jürgen Löffler ◽  
Anna Kobsar ◽  
Christian P. Speer ◽  
Susanne Kneitz ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Coronary Artery ◽  
Group B Streptococcus ◽  
Gene Array ◽  
Neonatal Morbidity ◽  
Innate Immune ◽  
Human Coronary Artery ◽  
Chemotactic Protein ◽  
Early Onset Sepsis ◽  
Group B

Early onset sepsis due to group B streptococcus leads to neonatal morbidity, increased mortality, and long-term neurological deficencies. Interaction between septicemic GBS and confluent monolayers of human coronary artery endothelial cells (HCAECs) was analyzed by genome wide expression profiling. In total, 124 genes were differentially expressed (89 upregulated, 35 downregulated) based on a more than 3-fold difference to control HCAEC. Regulated genes are involved in apoptosis, hemostasis, oxidative stress response, infection, and inflammation. Regulation of selected genes and proteins identified in the gene array analysis was confirmed by Real-time RT-PCR assay (granulocyte chemotactic protein 2), ELISA (urokinase, cyclooxygenase 2, granulocyte chemotactic protein 1), and western blotting (Heme oxygenase1, BCL2 interacting protein) at various time points between 4 and 24 hours. These results indicate that GBS infection might influence signalling pathways leading to impaired function of the innate immune system and hemorrhagic and inflammatory complications during GBS sepsis.

scholarly journals Regulation of capillary tubules and lipid formation in vascular endothelial cells and macrophages via extracellular vesicle-mediated microRNA-4306 transfer

2018 ◽  
Vol 47 (1) ◽  
pp. 453-469 ◽  
Author(s):  
Ying Yang ◽  
Hui Luo ◽  
Can Zhou ◽  
Rongyi Zhang ◽  
Si Liu ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Coronary Artery ◽  
Vascular Endothelial Cells ◽  
Density Lipoprotein ◽  
Extracellular Vesicle ◽  
Luciferase Reporter ◽  
Vascular Endothelial ◽  
Human Coronary Artery ◽  
Lipid Formation

Objective This study aimed to examine regulation of capillary tubules and lipid formation in vascular endothelial cells and macrophages via extracellular vesicle-mediated microRNA (miRNA)-4306 transfer Methods Whole blood samples (12 mL) were collected from 53 patients, and miR-4306 levels in extracellular vesicles (EVs) were analyzed by reverse transcription-polymerase chain reaction. Human coronary artery vascular endothelial cells (HCAECs) and human monocyte-derived macrophages (HMDMs) were transfected with a scrambled oligonucleotide, an miR-4306 mimic, or an anti-miR-4306 inhibitor. The direct effect of miR-4306 on the target gene was analyzed by a dual-luciferase reporter assay. Results EV-contained miR-4306 released from HMDMs was significantly upregulated in coronary artery disease. Oxidized low-density lipoprotein (ox-LDL)-stimulated HMDM-derived EVs inhibited proliferation, migration, and angiogenesis abilities of HCAECs in vitro. However, ox-LDL-stimulated HCAEC-derived EVs enhanced lipid formation of HMDMs. The possible mechanism of these findings was partly due to EV-mediated miR-4306 upregulation of the Akt/nuclear factor kappa B signaling pathway. Conclusions Paracrine cellular crosstalk between HCAECs and HMDMs probably supports the pro-atherosclerotic effects of EVs under ox-LDL stress.

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