Journal: Cell

Article Title: Cannabinoid receptor 1 antagonist genistein attenuates marijuana-induced vascular inflammation

doi: 10.1016/j.cell.2022.04.005

Figure Lengend Snippet: (A) The effects of Δ9-THC on cell viability of human coronary artery endothelial cells (HCAECs), human umbilical vein endothelial cells (HUVECs), normal human cardiac fibroblasts-ventricular (NHCF-V), and human embryonic stem cell-derived cardiomyocytes (hESC-CMs). Cells were treated with increasing concentrations of Δ9-THC for 48 h, and cell viability was measured by the CellTiter-Glo luminescent cell viability assay.

Article Snippet: Cell culture We obtained the following: human umbilical vein endothelial cells (HUVECs), human coronary artery endothelial cells (HCAECs), normal human cardiac fibroblasts-ventricular (NHCF-V) cells, human erythroleukemia (HEL 92.1.7), and human neuroblastoma cells (SK-N-FI) from the American Type Culture Collection (ATCC).

Techniques: Derivative Assay, Cell Viability Assay

Journal: Journal of Translational Medicine

Article Title: Quinic acid regulated TMA/TMAO-related lipid metabolism and vascular endothelial function through gut microbiota to inhibit atherosclerotic

doi: 10.1186/s12967-024-05120-y

Figure Lengend Snippet: QA improved TMAO-induced inflammatory lesions and endothelial dysfunction in HCAECs. (A) CCK-8 was applied to detect the toxicity of QA on HCAECs. (B) CCK-8 was used to detect HCAECs proliferation. (C) The expression of COX-2, IL-6, E-selectin, ICAM-1, HMGB1 was detected by RT-qPCR. (D) The expression of p-P65, p-MAPK14 protein was detected by western blot. (E) HMGB1 levels were detected by ELISA. (F) The expression of ZO-2, VE-Cadherin and Occludin were detected by western blot. * P < 0.05 vs. Control, # P < 0.05 vs. TMAO

Article Snippet: To investigate the cytotoxicity of QA, human coronary artery endothelial cells (HCAECs, HUM-iCell-c006, iCell) were treated with 1, 2.5, 5, 10 and 20 μM QA.

Techniques: CCK-8 Assay, Expressing, Quantitative RT-PCR, Western Blot, Enzyme-linked Immunosorbent Assay, Control

Journal: Cellular signalling

Article Title: PAI-1 contributes to homocysteine-induced cellular senescence

doi: 10.1016/j.cellsig.2019.109394

Figure Lengend Snippet: Cultures of EA.hy926 endothelial cells (A) and HCAEC (B, C) were pretreated with TM5441 (10 μM) (A, B) or TM5A15 (10 μM) (C) in triplicate followed by Homocysteine (Hcy) treatment for 4–5 days. Whole cell extracts were prepared and equal amount of pooled proteins from three wells were subjected to Western blot analysis for senescence markers and regulators using specific antibodies as indicated (A–C). Bar represents mean ± sem. Quantitative data are shown on the right (A’-C’). The levels of at least 2–3 senescence markers were determined in repeat experiments. D, E. Whole cell extracts (HCAEC) were prepared from two separate experiments and equal amount of pooled proteins from three wells were subjected to Western blot analysis for senescence markers and regulators p53 and pERK1/2 (D), integrin β3 and PAI-1 (E) using specific antibodies. Quantitative data in the lower panel showing the levels of each regulator relative to loading control Actin (D’, E’).

Article Snippet: Endothelial cell culture: treatment with Hcy and small molecule inhibitors of PAI-1 Primary cultures of Human Coronary Artery Endothelial Cells (HCAEC) (Cell Applications; Cat # 300–05a) and EA.hy926 (ATCC cat #CRL-2922) were grown in MesoEndo Cell Growth Media and Dulbecco’s Modified Eagle Medium (DMEM) containing 10% fetal bovine serum and 1% penicillin and streptomycin respectively and maintained at 37 °C in a 5% CO 2 incubator.

Techniques: Western Blot, Control

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Three-Layered Silk Fibroin Tubular Scaffold for the Repair and Regeneration of Small Caliber Blood Vessels: From Design to in vivo Pilot Tests

doi: 10.3389/fbioe.2019.00356

Figure Lengend Snippet: Time dependence of the absolute cell numbers of HAAF (A) , HASMC (B) , and HCAEC (C) cells cultured on SilkGraft and on polystyrene. Absolute cell numbers were lower on SilkGraft than on polystyrene because the available surface area was reduced due to the use of the steel ring which kept the silk substrate under water. Total cell growth differences between cells cultured on SilkGraft or on polystyrene are expressed by the areas under the corresponding curves, the statistical levels of significance of which are: for HAAFs, P < 0.001; for HASMCs, P < 0.01; and for HCAECs, P < 0.001.

Article Snippet: Adult Human Coronary Artery Endothelial Cells (HCAECs), Human Aortic Smooth Muscle Cells (HASMCs), and Human Aortic Adventitial Fibroblasts (HAAFs) were provided by ScienCell Research Laboratories (Carlsbad, CA, USA).

Techniques: Cell Culture

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Three-Layered Silk Fibroin Tubular Scaffold for the Repair and Regeneration of Small Caliber Blood Vessels: From Design to in vivo Pilot Tests

doi: 10.3389/fbioe.2019.00356

Figure Lengend Snippet: Living cells density/mm 2 of apparent surface area after 21 days of in vitro culture.

Article Snippet: Adult Human Coronary Artery Endothelial Cells (HCAECs), Human Aortic Smooth Muscle Cells (HASMCs), and Human Aortic Adventitial Fibroblasts (HAAFs) were provided by ScienCell Research Laboratories (Carlsbad, CA, USA).

Techniques: In Vitro

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Three-Layered Silk Fibroin Tubular Scaffold for the Repair and Regeneration of Small Caliber Blood Vessels: From Design to in vivo Pilot Tests

doi: 10.3389/fbioe.2019.00356

Figure Lengend Snippet: Cumulative consumption of glucose and glutamine and release of lactate. Results were normalized per 10 3 cells. (A–C) The cumulative glucose consumption was higher for HAAFs ( P < 0.05) and HCAECs ( P < 0.001) seeded on SilkGraft, whereas it showed only marginal differences between the two substrates for HASMCs ( P > 0.05). (D–F) Glutamine consumption was lower for HAAFs ( P < 0.05) seeded on SilkGraft, similar for HASMCs ( P > 0.05) cultured on the two substrates, and significantly larger for HCAECs ( P < 0.001) grown on the silk substrate. (G,H) The cumulative amount of lactate released by HAAFs and HASMCs was the same whichever the substrate ( P > 0.05). Lactate release could not be assessed for HCAECs because the released lactate was re-uptaken and used for metabolic purposes. The statistical analysis of these data is shown in .

Article Snippet: Adult Human Coronary Artery Endothelial Cells (HCAECs), Human Aortic Smooth Muscle Cells (HASMCs), and Human Aortic Adventitial Fibroblasts (HAAFs) were provided by ScienCell Research Laboratories (Carlsbad, CA, USA).

Techniques: Cell Culture

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Three-Layered Silk Fibroin Tubular Scaffold for the Repair and Regeneration of Small Caliber Blood Vessels: From Design to in vivo Pilot Tests

doi: 10.3389/fbioe.2019.00356

Figure Lengend Snippet: Comparison of metabolic parameters ^* of the different cell types cultured on SilkGraft and polystyrene.

Article Snippet: Adult Human Coronary Artery Endothelial Cells (HCAECs), Human Aortic Smooth Muscle Cells (HASMCs), and Human Aortic Adventitial Fibroblasts (HAAFs) were provided by ScienCell Research Laboratories (Carlsbad, CA, USA).

Techniques: Comparison, Cell Culture

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Three-Layered Silk Fibroin Tubular Scaffold for the Repair and Regeneration of Small Caliber Blood Vessels: From Design to in vivo Pilot Tests

doi: 10.3389/fbioe.2019.00356

Figure Lengend Snippet: Relevant cytokines and chemokines secreted by each cell type cultured between 18 and 20 days on SilkGraft and polystyrene: HAFFs (A) , HCAECs (B) , and HASMCs (C) . Results of immunofluorescence intensities were normalized to 10 3 cells. IL-6: Interleukin-6; MCP-1: Monocyte chemoattractant protein-1; TIMP-2: Tissue inhibitor of metal proteinases-2; IP-10: Interferon gamma-induced protein-10; MCP-2: Monocyte chemoattractant protein-2; Eotaxin-1; RANTES: Regulated on activation normal T cell expressed and secreted; MIP-1β: Macrophage inflammatory protein-1β; TNF-β: Tumor necrosis factor-β; GM-CSF: Granulocyte-macrophage colony stimulating factor; IL-1α: Interleukin-1α; IL-1β: Interleukin-1β; ICAM-1: Intercellular adhesion molecule-1. The bars are the mean values of three independent experiments corrected for cell numbers. * P < 0.01; ** P < 0.001. SEMs, not shown, ranged between 5 and 10% of corresponding mean values.

Article Snippet: Adult Human Coronary Artery Endothelial Cells (HCAECs), Human Aortic Smooth Muscle Cells (HASMCs), and Human Aortic Adventitial Fibroblasts (HAAFs) were provided by ScienCell Research Laboratories (Carlsbad, CA, USA).

Techniques: Cell Culture, Immunofluorescence, Activation Assay

Journal: Frontiers in Cardiovascular Medicine

Article Title: A20/TNFAIP3 Increases ENOS Expression in an ERK5/KLF2-Dependent Manner to Support Endothelial Cell Health in the Face of Inflammation

doi: 10.3389/fcvm.2021.651230

Figure Lengend Snippet: Overexpression of A20 in human coronary artery endothelial cells (HCAEC) increases endothelial nitric oxide synthase (eNOS) expression and prevents its downregulation by TNF, while A20 knockdown decreases eNOS levels. (A) Relative eNOS mRNA (qPCR) normalized by mRNA levels of the housekeeping (HKG) gene 28S and expressed as fold change of non-treated (NT) Ctrl HCAEC, and (B) eNOS protein (WB) levels in non-transduced HCAEC (Ctrl) and in HCAEC transduced with rAd.A20 or control rAd.βgal (100 MOI), before and 24 h after treatment with TNF. β-Actin was used to correct for loading. (C) Relative eNOS and A20 mRNA levels (qPCR) in HCAEC non-transfected (Ctrl) and transfected with A20 siRNA or with AllStars negative control siRNA (Ctrl siRNA) for 24 h, normalized by the mRNA levels of the HKG gene cyclophilin A (CycA), and expressed as fold change of non-transfected Ctrl HCAEC. All data in (A–C) are presented as mean ± SEM of three to five independent experiments. Significance between groups was determined by one- or two-way ANOVA followed by the Tukey or Bonferroni multiple comparison post hoc test, respectively; * p < 0.05, ** p < 0.01, *** p < 0.001. Significance between NT and TNF-treated HCAEC was determined by an unpaired t test and depicted as hashtag in lieu of asterisk symbols in order to better differentiate between the two statistical methods used to analyze the data. # p < 0.05, ## p < 0.01, ### p < 0.001. (D) Relative eNOS and A20 mRNA levels in the aortae of 3–4-week-old A20 knockout (KO) and heterozygous (HT) mice, as well as wild-type (WT) littermates. Data are presented as mean ± SEM of 5–11 animals/group. Significance between groups was determined by one-way ANOVA followed by the Tukey multiple comparison post hoc test; * p < 0.05, ** p < 0.01, *** p < 0.001. (E) Representative eNOS immunohistochemistry (brown) in the brain of A20 wild-type (WT), heterozygous (HT), and knockout (KO) mice. Photomicrographs are representative of three animals per genotype, magnification = ×400.

Article Snippet: Human coronary artery endothelial cells (HCAEC) derived from six different donors from both genders were purchased from Lonza, Allendale, NJ, and cultured in EGM-2MV BulletKit medium.

Techniques: Over Expression, Expressing, Knockdown, Transduction, Control, Transfection, Negative Control, Comparison, Knock-Out, Immunohistochemistry

Journal: Frontiers in Cardiovascular Medicine

Article Title: A20/TNFAIP3 Increases ENOS Expression in an ERK5/KLF2-Dependent Manner to Support Endothelial Cell Health in the Face of Inflammation

doi: 10.3389/fcvm.2021.651230

Figure Lengend Snippet: A20 overexpression in HCAEC increases eNOS transcription in an ERK5-dependent manner. (A) mRNA levels of eNOS, KLF2, KLF4, and ERK5 were measured by qPCR in non-transduced HCAEC (Ctrl) and HCAEC transduced with rAd.A20 or control rAd.βgal at 100 MOI for 48 h. Data were normalized by the 28S HKG and expressed as mean ± SEM fold change of Ctrl ( n = 5–9). (B) eNOS, (D) KLF2, and (E) KLF4 mRNA and (C) eNOS protein levels were measured by qPCR, and WB in non-transduced HCAEC (Ctrl) and HCAEC transduced with rAd.A20 or control rAd.βgal at 100 MOI for 3 h prior to 24 h treatment with the ERK5 inhibitor, XMD8-92 (10 μM). Graphs in (B) , (D) , and (E) depict relative mRNA levels, normalized by the 28S HKG and expressed as mean ± SEM fold change of non-treated (NT) Ctrl ( n = 6–10). In (C) , β-actin was used to correct for loading. Densitometry results are presented as fold change of NT Ctrl cells ( n = 3). (F) KLF2 mRNA levels were measured by qPCR in non-transduced HCAEC (Ctrl) and in HCAEC transduced with rAd.A20 or control rAd.βgal at 100 MOI for 48 h prior to 24 h treatment with TNF (200 U/mL). Graphs depict relative KLF2 mRNA levels normalized by the 28S HKG and expressed as mean ± SEM fold change of non-treated (NT) Ctrl ( n = 3–5). * p < 0.05, ** p < 0.01, *** p < 0.001, as determined by two-way ANOVA followed by the Bonferroni post hoc test. Significance between NT and TNF-treated HCAEC was determined by an unpaired t test and depicted as hashtag in lieu of asterisk symbols in order to better differentiate between the two statistical methods used to analyze the data: # p < 0.05, ### p < 0.001.

Article Snippet: Human coronary artery endothelial cells (HCAEC) derived from six different donors from both genders were purchased from Lonza, Allendale, NJ, and cultured in EGM-2MV BulletKit medium.

Techniques: Over Expression, Transduction, Control

Journal: Frontiers in Cardiovascular Medicine

Article Title: A20/TNFAIP3 Increases ENOS Expression in an ERK5/KLF2-Dependent Manner to Support Endothelial Cell Health in the Face of Inflammation

doi: 10.3389/fcvm.2021.651230

Figure Lengend Snippet: A20 overexpression in HCAEC increases eNOS transcription in a KLF2-dependent manner. HCAEC were transduced with 100 MOI of shRNA-KLF2 or scramble shRNA-Ctrl for 24 h, then retransduced with rAd.A20 or control rAd.βgal at 200–250 MOI for 48 h or left non-transduced (Ctrl). Cell lysates were evaluated by qPCR for mRNA levels of (A) KLF2, (B) KLF4, and (C) eNOS. Graphs shown depict relative mRNA levels, normalized by the 28S HKG and expressed as mean ± SEM fold change of Ctrl shRNA-Ctrl cells ( n = 3–6) and by (D) WB for eNOS protein expression. GAPDH was used to correct for loading. Densitometry results are presented as fold change of Ctrl shRNA-Ctrl cells and expressed as mean ± SEM ( n = 4). * p < 0.05, ** p < 0.01, *** p < 0.001, as determined by two-way ANOVA followed by the Bonferroni post hoc test.

Article Snippet: Human coronary artery endothelial cells (HCAEC) derived from six different donors from both genders were purchased from Lonza, Allendale, NJ, and cultured in EGM-2MV BulletKit medium.

Techniques: Over Expression, Transduction, shRNA, Control, Expressing

Journal: Frontiers in Cardiovascular Medicine

Article Title: A20/TNFAIP3 Increases ENOS Expression in an ERK5/KLF2-Dependent Manner to Support Endothelial Cell Health in the Face of Inflammation

doi: 10.3389/fcvm.2021.651230

Figure Lengend Snippet: Overexpression of A20 in HCAEC modulates the expression levels of KLF2-dependent genes to increase EC homeostasis. mRNA levels of endothelin-1 (END1), chemokine C-C motif ligand 2 (CCL2)/monocyte chemoattractant protein-1 (MCP1), thrombomodulin (THBD), eNOS, KLF2, and A20 were measured by qPCR in non-transduced HCAEC (Ctrl) and HCAEC transduced with rAd.A20 or control rAd.βgal. Data were normalized by the 28S HKG and expressed as mean ± SEM fold change of Ctrl ( n = 8–10). * p < 0.05, ** p < 0.01, *** p <0.001, as determined by one-way ANOVA followed by Tukey post hoc test.

Article Snippet: Human coronary artery endothelial cells (HCAEC) derived from six different donors from both genders were purchased from Lonza, Allendale, NJ, and cultured in EGM-2MV BulletKit medium.

Techniques: Over Expression, Expressing, Transduction, Control

Journal: Frontiers in Cardiovascular Medicine

Article Title: A20/TNFAIP3 Increases ENOS Expression in an ERK5/KLF2-Dependent Manner to Support Endothelial Cell Health in the Face of Inflammation

doi: 10.3389/fcvm.2021.651230

Figure Lengend Snippet: A20 overexpression increases basal Ser-1177 eNOS phosphorylation in HCAEC. Representative WB of total eNOS and P-eNOS (Ser-1177) in non-transduced HCAEC (Ctrl) and HCAEC transduced with rAd.A20 or rAd.βgal at 100 MOI for 48 h; GAPDH was used to correct for loading. Densitometry results are presented as mean fold change of Ctrl ± SEM ( n = 4). * p < 0.05, as determined by one-way ANOVA followed by the Tukey post hoc test.

Article Snippet: Human coronary artery endothelial cells (HCAEC) derived from six different donors from both genders were purchased from Lonza, Allendale, NJ, and cultured in EGM-2MV BulletKit medium.

Techniques: Over Expression, Phospho-proteomics, Transduction

Journal: Cardiovascular Diagnosis and Therapy

Article Title: A novel animal model for vulnerable atherosclerotic plaque: dehydrated ethanol lavage in the carotid artery of rabbits fed a Western diet

doi: 10.21037/cdt-21-291

Figure Lengend Snippet: CCK8 assay of human coronary artery ECs. Left and right panels represent OD value and survival rate of EC, respectively, after treatment of HCAEC with dehydrated ethanol for 10 seconds. Cell proliferation was represented as cell quantity. Data are presented as mean±SD (n=6). EC, endothelial cell; OD, optical density; HCAEC, human coronary artery EC.

Article Snippet: Human coronary artery ECs (HCAEC; ScienCell Research Laboratories, Carlsbad, CA, USA) were treated with dehydrated ethanol for 10 seconds and then washed with phosphate-buffered saline (PBS) and EC medium (SciencCell).

Techniques: CCK-8 Assay

Journal: Cellular signalling

Article Title: Pharmacological inhibition of MALT1 protease activity suppresses endothelial activation via enhancing MCPIP1 expression

doi: 10.1016/j.cellsig.2018.05.009

Figure Lengend Snippet: (A) MALT1 protein levels in HUVEC, HAEC, HCAEC, HDMEC and HLMEC were measured by Western blot. (B) HUVECs were pretreated with DMSO or MI-2 (1 μM) for 30 minutes and then treated with or without inflammatory stimuli (10 ng/mL TNF α, 1 μ g/mL LPS or 10 ng/mL IL1 β ) for 8 hours. The protein levels of VCAM-1, ICAM-1 were detected by Western blot. The bands were quantified by Gel-Pro Analyzer software and presented as fold changes; n=3, *p<0.05 or **p<0.001 vs DMSO treatment. (C) HUVECs were pretreated with MI-2 (1 μM) for 30 minutes and then incubated with or without TNFα (10 ng/mL) for 4 hours. The relative mRNA levels of VCAM-1, ICAM-1, IL1β, TNFα MCP1 and CXCL1 were detected by QPCR using 2△△CT method. Data represent mean±SD, n=4. (D) HUVECs were transiently transfected with short interfering RNA targeting on MALT1 (si-MALT1) or nonspecific short interfering RNA (si-Control) by electroporation (Amaxa). Transfected cells were quiescent for 48 hours and then treated with or without TNFα (10 ng/mL) for 8 hours. Expression of MALT1 and VCAM-1 were detected using Western blot. Band intensity of VCAM-1 was quantified by Gel-Pro Analyzer software; n=3, **p<0.001 vs treated si-Control cells. (E&F) HUVECs were pretreated with DMSO or 1 μM of MLT-827 (E) or mepazine (F) for 30 minutes and then treated with or without inflammatory stimuli (10 ng/mL TNF) for 8 hours. The protein levels of VCAM-1, ICAM-1 were detected by Western blot. The bands were quantified by Gel-Pro Analyzer software and presented as fold changes; n=3, *p<0.05 or **p<0.001 vs DMSO treatment.

Article Snippet: The primary human vascular endothelial cells including human aortic ECs (HAEC), human coronary artery ECs (HCAEC), human dermal microvascular ECs (HDMEC), human lung microvascular ECs (HLMEC) and human umbilical vein ECs (HUVEC) were purchased from Lonza Walkersville Inc. (Walkersville, MD) and cultured in EGM or EGM2 medium according to the manufacturer’s instruction.

Techniques: Western Blot, Software, Incubation, Transfection, Small Interfering RNA, Control, Electroporation, Expressing