Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: Transcriptional Regulation of Endothelial Arginase 2 by Histone Deacetylase 2

doi: 10.1161/atvbaha.114.303685

Figure Lengend Snippet: Figure 2. Histone deacetylase (HDAC) inhibition upregulates arginase 2 (Arg2) activity, mRNA, and protein expression. Human aortic endothelial cells (HAECs) were incubated with 0.4 μmol/L of trichostatin A (TSA) for 18 hours and (A) arginase activity was measured in cell lysates using urea production assay11 and (B) Arg2 protein levels were determined using Western blot (WB). C, Dose-dependent (0, 0.1, 0.3, and 1 μmol/L) and (D) time-dependent (0, 2, 4, 8, and 18 hours) effects of TSA (1 μmol/L) on endothelial Arg2 protein expres- sion were determined using WB. E, HAECs were transduced with 15 MOI of adenoviruses encoding either nontargeted, Arg2 or Arg1 shRNA effect of TSA on total arginase activity was determined using the urea assay. Effect of TSA (1 μmol/L) on Arg2 mRNA levels was determined using (F) RT-polymerase chain reaction (PCR) and (G) real-time PCR in HAECs with increasing doses of TSA (0, 0.1, 0.2, and 1 μmol/L) or nicotinamide 0.1 mmol/L or 0.2 mmol/L (F only, lanes 4 and 5). H, mRNA levels were determined by real-time PCR in iso- lated mice aortas that were incubated with 200 nmol/L TSA for 18 hours. I, Arginase activity was determined from isolated segments of mice aorta exposed to TSA (200 nmol/L). J, Arginase activity was determined from HAECs exposed to TSA (1 μmol/L) or nicotinamide (5 mmol/L) for 30 minutes. *P<0.05 vs control (no TSA), $P<0.05 vs control (no TSA), #P<0.05 vs control (with TSA). WB and RT-PCR experi- ments were performed 3 to 4 times. Fold change in real-time PCR data indicates the differences in the ratio between Arg2 mRNA and 18s mRNA when the TSA-treated and control sample data are measured. IB indicates XXX; and RLU, XXX.

Article Snippet: To define and quantify the nature of TSA modulation of Arg2 expression, human aortic endothelial cells (HAECs) were treated with 400 nmol/L TSA for 18 hours as suggested by the manufacturer (Cell Signaling), and arginase activity and protein expression were then measured.

Techniques: Histone Deacetylase Assay, Inhibition, Activity Assay, Expressing, Incubation, Western Blot, Transduction, shRNA, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Isolation, Control, Reverse Transcription Polymerase Chain Reaction

Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: Transcriptional Regulation of Endothelial Arginase 2 by Histone Deacetylase 2

doi: 10.1161/atvbaha.114.303685

Figure Lengend Snippet: Figure 4. Effects of trichostatin A (TSA) on arginase transcription and promoter activity. A, Human aortic endothelial cells (HAECs) were pretreated with either 10 μg/mL of the transcriptional inhibitor actinomycin D (Act D) or 10 μg/mL of the protein translation inhibitor cyclo- heximide for 8 hours and cell lysates were then subjected to Western blot (WB) using antibodies against arginase 2 (Arg2) and GAPDH. B, Luciferase activity was determined in HAECs that were transfected with the pGL3 luciferase (LUC) reporter plasmid consisting of the −1 kb human ARG2 promoter and 3 different promoter fragments (as illustrated above the graph) and incubated in the presence of TSA. C, HAECs were transfected with pgL3.4 harboring the 1-kb Arg2 promoter and challenged with increasing doses of TSA. *P<0.05 vs con- trol, and WB results are representatives of 3 independent experiments. IB indicates XXX; and RLU, XXX.

Article Snippet: To define and quantify the nature of TSA modulation of Arg2 expression, human aortic endothelial cells (HAECs) were treated with 400 nmol/L TSA for 18 hours as suggested by the manufacturer (Cell Signaling), and arginase activity and protein expression were then measured.

Techniques: Activity Assay, Western Blot, Luciferase, Transfection, Plasmid Preparation, Incubation

Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: Transcriptional Regulation of Endothelial Arginase 2 by Histone Deacetylase 2

doi: 10.1161/atvbaha.114.303685

Figure Lengend Snippet: Figure 5. Attenuated nitric oxide production in trichostatin A (TSA)-treated human aortic endothelial cells (HAECs) was restored by arginase inhibition. HAECs were incubated with either TSA (1 μmol/L) alone or together with amino-2-borono-6-hexanoic acid (ABH; 1 mmol/L). A, Representative epifluorescence images and (B) quantitative analysis of DAF-DA fluorescence were used to assess nitric oxide (NO) production. C, A NO analyzer from Siever was used to measure byproducts of NO, mainly nitrite, in HAEC cell media that were incu- bated with either TSA alone or coincubated with TSA and ABH. D, Two hundred ninety-three cells that do not express native endothelial nitric oxide synthase (eNOS) were expressed with constitutively active eNOS constructs and incubated with either TSA alone or together with ABH. The DCFH-DiOxyQ-based fluorescent assay from Cell Biolabs was used to determine NO radicals in cell culture media. *P<0.05 vs control, #P<0.05 vs TSA alone, $P<0.05 vs TSA+ABH. DAF fluorescence images are representative of 3 independent experi- ments. CIeNOS indicates calcium-insensitive endothelial nitric oxide synthase; RFU, XXX; and RNS, XXX.

Article Snippet: To define and quantify the nature of TSA modulation of Arg2 expression, human aortic endothelial cells (HAECs) were treated with 400 nmol/L TSA for 18 hours as suggested by the manufacturer (Cell Signaling), and arginase activity and protein expression were then measured.

Techniques: Inhibition, Incubation, Fluorescence, Construct, Cell Culture, Control

Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: Transcriptional Regulation of Endothelial Arginase 2 by Histone Deacetylase 2

doi: 10.1161/atvbaha.114.303685

Figure Lengend Snippet: Figure 7. Histone deacetylase (HDAC) 2 is the specific HDAC isoform that regulates arginase 2 (Arg2) expression. Human aortic endothelial cells were transfected with increasing concentrations of (A) HDAC1, (B) HDAC3, (C) HDAC8, or (D) HDAC2 cDNAs, and Arg2 expression was determined by Western blot. E, HAECs that were transfected with HDAC2 siRNA (30 nmol/L) were subjected to immu- noblotting for Arg2 and HDAC2. Results are repre- sentatives of 3 to 4 independent experiments.

Article Snippet: To define and quantify the nature of TSA modulation of Arg2 expression, human aortic endothelial cells (HAECs) were treated with 400 nmol/L TSA for 18 hours as suggested by the manufacturer (Cell Signaling), and arginase activity and protein expression were then measured.

Techniques: Histone Deacetylase Assay, Expressing, Transfection, Western Blot

Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: Transcriptional Regulation of Endothelial Arginase 2 by Histone Deacetylase 2

doi: 10.1161/atvbaha.114.303685

Figure Lengend Snippet: Figure 8. Increased endothelial arginase 2 (Arg2) expression in response to oxidized low-density lipoprotein (OxLDL) involves promoter activation and histone deacetylase (HDAC) 2. A, Human aortic endothelial cells (HAECs) were exposed to 50 μg/mL of OxLDL for 18 hours and subjected to Western blot (WB) using Arg2 antibody. B, Luciferase activity was determined in HAECs that were transfected with pGL3 luciferase (LUC) reporter plasmids consisting of either −1 kb human ARG2 promoter or 1 of 3 different promoter fragments (as illustrated above the graph in Figure 4B) and incubated with 50 μg/mL of OxLDL for 18 hours. C, HAECs were transfected with an LUC reporter plasmid containing the −1-kb human ARG2 promoter, and cells were exposed to increasing doses of OxLDL (0, 25, 50, and 100 μg/mL) for 18 hours. Firefly luciferase activity was measured and normalized to Renilla luciferase. GFP and HDAC2 adenoviruses were added 8 hours after transfection with the luciferase constructs. D, Binding of HDAC2 to the Arg2 promoter was quantified using a ChIP assay. E, HAECs were exposed to increasing doses of OxLDL (0, 25, 50, and 100 μg/mL) for 18 hours and immunoblotted for HDAC2 and Arg2. F, HAECs with adenoviral-mediated HDAC2 overexpression or controls (Ad-GFP) were exposed to OxLDL, and expression levels of Arg2, HDAC2, and GAPDH (loading control) were determined with WB. *P<0.05 vs control. WB results are representatives of 3 to 4 inde- pendent experiments. IB indicates XXX; and RLU, XXX.

Article Snippet: To define and quantify the nature of TSA modulation of Arg2 expression, human aortic endothelial cells (HAECs) were treated with 400 nmol/L TSA for 18 hours as suggested by the manufacturer (Cell Signaling), and arginase activity and protein expression were then measured.

Techniques: Expressing, Activation Assay, Histone Deacetylase Assay, Western Blot, Luciferase, Activity Assay, Transfection, Incubation, Plasmid Preparation, Construct, Binding Assay, Over Expression, Control

Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: Transcriptional Regulation of Endothelial Arginase 2 by Histone Deacetylase 2

doi: 10.1161/atvbaha.114.303685

Figure Lengend Snippet: Figure 9. Oxidized low-density lipoprotein (OxLDL)-mediated impairment of vascular relaxation is ameliorated by histone deacetylase (HDAC) 2 overexpression. Isolated mice aortic rings were incubated in EBM2 media containing 100 MOI of either GFP or HDAC2 adeno- viruses. Twenty four hours post transduction, media was replaced with fresh media with or without OxLDL (50 μg/mL). After 48 hours, dose–response effects of acetylcholine (A) and sodium nitroprusside (SNP; B) on vascular relaxation were determined using wire myog- raphy. C, Mice aortic segments were transduced with 100 MOI of either GFP or HDAC2 adenoviruses and subjected to immunoblotting with anti-GFP and anti-HDAC2 antibodies. D, Human aortic endothelial cells (HAECs) overexpressing HDAC2 or GFP were subjected to an arginase activity assay. E, Nitric oxide production was determined using the DAF-FM DA fluorescence assay in HAECs that were transduced with either Ad-Arg2shRNA or Ad-HDAC2shRNA alone, or both reagents in combination. Nontargeted Ad-shRNA was used as a control. *P<0.05 vs control; n=6. RFU indicates XXX; and WT, wild type.

Article Snippet: To define and quantify the nature of TSA modulation of Arg2 expression, human aortic endothelial cells (HAECs) were treated with 400 nmol/L TSA for 18 hours as suggested by the manufacturer (Cell Signaling), and arginase activity and protein expression were then measured.

Techniques: Histone Deacetylase Assay, Over Expression, Isolation, Incubation, Transduction, Western Blot, Arginase Activity Assay, Fluorescence, shRNA, Control

Journal: Stem Cell Research & Therapy

Article Title: Extracellular vesicles from human adipose-derived stem cells relieve pain and inflammation in a rat model of knee osteoarthritis

doi: 10.1186/s13287-026-04932-7

Figure Lengend Snippet: Anti-inflammatory effect of hASC-EVs on IL-1β stimulated human chondrocytes (HC-OA). A Confocal images of HC-OAs treated with PKH67-labeled hASC-EVs, showing intracellular uptake. Scale bar, 20 μm. B – F The mRNA expression of TRPA1, COX-2, MMP-2, MMP-3, and MMP-9 was quantified using qPCR and normalized to GAPDH. Data are presented as means ± SD ( n = 3). ns, not significant ( p > 0.05), * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 vs. IL-1β only group

Article Snippet: Human OA chondrocytes (HC-OA; Cell Applications, USA) were derived from human articular cartilage obtained from donors diagnosed with osteoarthritis, as defined by the vendor.

Techniques: Labeling, Expressing

Journal: Advanced Healthcare Materials

Article Title: Plasma‐Polymerized Nanoparticles Presenting Fibrillin‐1 Drive Rapid Re‐Endothelialization of Vascular Grafts

doi: 10.1002/adhm.202503360

Figure Lengend Snippet: PPN‐PF8 functionalization of ePTFE improves endothelial attachment in vitro. A) Representative images of SEM and F‐actin staining. B) HCAEC attachment and C) spreading on ePTFE, ePTFE with passively bound PF8 (ePTFE‐PF8), PPN‐coated ePTFE (PPN), and PPN‐PF8 functionalized ePTFE, n = 3 per sample. Scale bar = 300 µm. * p < 0.05, *** p < 0.001 versus ePTFE.

Article Snippet: Human coronary artery endothelial cells (HCAEC; Cell Applications) were cultured in MesoEndo medium (Cell Applications) at 37 °C in 5% CO 2 .

Techniques: In Vitro, Staining

Journal: Advanced Healthcare Materials

Article Title: Plasma‐Polymerized Nanoparticles Presenting Fibrillin‐1 Drive Rapid Re‐Endothelialization of Vascular Grafts

doi: 10.1002/adhm.202503360

Figure Lengend Snippet: PPN‐PF8 functionalization of ePTFE improves endothelial proliferation in vitro. A) Representative images of SEM and F‐actin staining of HCAEC proliferation on days 1 and 3. B) HCAEC proliferation and C) spreading on ePTFE, ePTFE with passively bound PF8, PPN, and PPN‐PF8 functionalized ePTFE. * p < 0.05, *** p < 0.001 versus ePTFE on each day, Scale bar = 300 µm. n = 3 per sample.

Article Snippet: Human coronary artery endothelial cells (HCAEC; Cell Applications) were cultured in MesoEndo medium (Cell Applications) at 37 °C in 5% CO 2 .

Techniques: In Vitro, Staining