Journal: Microsystems & Nanoengineering

Article Title: Modular operation of microfluidic chips for highly parallelized cell culture and liquid dosing via a fluidic circuit board

doi: 10.1038/s41378-020-00216-z

Figure Lengend Snippet: a Live-cell fluorescence images (one image per four chambers) showing an overview of all the chambers after 3 days of culturing GFP-expressing HUVECs. b Live-cell fluorescence image of GFP-expressing HUVECs in chambers 6 and 7. c Fluorescence image of fixed HUVECs with the cell F-actin and nuclei stained with ActinRed and NucBlue, respectively. d HUVECs after seeding and subsequent monolayer formation. The cells were seeded at a high cell density (i) and confluent on day 1 (ii). The monolayer was still intact on day 3 (iii) but began to deteriorate on day 4 (iv). At this point, the medium in the supply vial was replaced. The cells showed signs of recovery on day 5 (v) as the monolayer started to reform. The red dots in (iv) and (v) mark the cells counted in the region of interest. e Cells in ten chambers were counted in regions where the monolayer had deteriorated on day 4 (98 h) and after 16 h of recovery (114 h). The black dots represent the mean cell count, and the error bars represent the standard deviation. The cell number increased by an average of 33% in these areas

Article Snippet: Live-cell images of the GFP-expressing HUVECs (Angio-Proteomie, USA) were taken with an EVOS FL cell imaging system using the GFP filter cube.

Techniques: Fluorescence, Expressing, Staining, Cell Counting, Standard Deviation

Journal: Frontiers in Cell and Developmental Biology

Article Title: MicroRNA-494 Regulates Endoplasmic Reticulum Stress in Endothelial Cells

doi: 10.3389/fcell.2021.671461

Figure Lengend Snippet: ER stress induces expression of the primary and mature forms of miR-494. Relative mRNA expression of ER stress responsive genes as measured using qRT-PCR. (A) spliced XBP1 and Total XBP1 and (B) DDIT3 (CHOP) mRNA levels in HUVECs treated with 5 μg/mL tunicamycin (TCN) over a time course. Relative expression of (C) mature miR-494 and (D) primary miR-494 (pri-miR-494) in HUVECs treated with TCN over a time course. Gene expression is normalized to GAPDH or U6 and mean fold change compared to vehicle control or time 0h is shown. Graphs are representative of one biological replicate from three independent replicates where values indicate mean ± standard deviation.

Article Snippet: For plasmid transfections, HUVECs were transfected using the Cytofect HUVECs transfection kit (Cat:TF200K, Cell Applications). pEGFP-ATF6 was a gift from Ron Prywes (Addgene plasmid # 32955) ( ).

Techniques: Expressing, Quantitative RT-PCR, Gene Expression, Control, Standard Deviation

Journal: Frontiers in Cell and Developmental Biology

Article Title: MicroRNA-494 Regulates Endoplasmic Reticulum Stress in Endothelial Cells

doi: 10.3389/fcell.2021.671461

Figure Lengend Snippet: miR-494 is a negative regulator of ER stress in vitro . Relative mRNA expression of ER stress responsive genes as measured by qRT-PCR. (A,B) DDIT3 (CHOP), (C,D) spliced XBP1 in HUVECs treated with 10 μg/mL TCN 48h after transfection with (A,C) miR-494 mimic or (B,D) miR-494 inhibitor. Gene expression is normalized to GAPDH and mean fold changes compared to control treatments are shown. (E,F) Simple Western blot analysis of HUVECs transfected with miR-494 mimic or control (24 h) (E) or miR-494 inhibitor or control (F) followed by TCN (10 μg/mL) for 24 h. (G) Cell viability in HUVECs as treated in (A) Vertical dotted red line indicates non-adjacent lanes. Graphs are mean + SEM fold changes of biological replicates from n = 3 independent experiments. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001 by two-tailed Student’s T -test.

Article Snippet: For plasmid transfections, HUVECs were transfected using the Cytofect HUVECs transfection kit (Cat:TF200K, Cell Applications). pEGFP-ATF6 was a gift from Ron Prywes (Addgene plasmid # 32955) ( ).

Techniques: In Vitro, Expressing, Quantitative RT-PCR, Transfection, Gene Expression, Control, Simple Western, Two Tailed Test

Journal: Frontiers in Cell and Developmental Biology

Article Title: MicroRNA-494 Regulates Endoplasmic Reticulum Stress in Endothelial Cells

doi: 10.3389/fcell.2021.671461

Figure Lengend Snippet: miR-494 regulates target genes in cell survival and DNA replication. (A) Venn diagram showing the number of downregulated target proteins in a Tandem Mass Tag labeled Mass Spectrometry profile from HUVECs treated with TCN or transfected with miR-494 compared to vehicle treatment or control miR respectively. (B) Fold-change (compared to respective controls) of protein or mRNA levels as assessed by Mass Spectrometry or qRT-PCR respectively for the six targets that were downregulated in both TCN and miR-494 groups. All six targets harbor miR-494 binding sites in their 3′UTRs. (C) Representative Simple Western blot showing survivin ( BIRC5 ) and GINS4 levels in HUVECs 24 h after miR-494 transfection followed by TCN treatment (24 h). Right panels show quantitation of biological replicates. ∗ P < 0.05, ∗∗ P < 0.01, by two-tailed Student’s T -test. (D) Immunofluorescence images showing survivin expression in HUVECs 24 h after miR transfection and/or TCN treatment. Right panel shows quantification via ImageJ of survivin fluorescence intensity in each cell. Each dot represents individual cells. Scale bar in white = 50 μm. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001 by one-way ANOVA with post hoc Tukey’s correction.

Article Snippet: For plasmid transfections, HUVECs were transfected using the Cytofect HUVECs transfection kit (Cat:TF200K, Cell Applications). pEGFP-ATF6 was a gift from Ron Prywes (Addgene plasmid # 32955) ( ).

Techniques: Labeling, Mass Spectrometry, Transfection, Control, Quantitative RT-PCR, Binding Assay, Simple Western, Quantitation Assay, Two Tailed Test, Immunofluorescence, Expressing, Fluorescence

Journal: American Journal of Translational Research

Article Title: Transplantation of HUVECs with genetically modified Nogo-B accelerates wound-healing in nude mice

doi:

Figure Lengend Snippet: Nogo-B affected HUVECs proliferation and tube formation by transfected with GV248-Nogo-B RNAi-GFP. (A) The cells grew well and shaped like spindles with no vacuole seen in the nucleus and were transfected under a fluorescence microscope and the whole cell emits fluorescence (200X). (B) Detection of the Nogo-B expression by Western Blot. Nogo-B was completely knocked down. (C) The effect of Nogo-B on proliferation and (D) tube-formation of HUVECs were measured by CCk-8 assay and tube formation assay. Data was shown as means ± SD, (C) n=3; (D) n=5; *P<0.05, **P<0.01, ***P<0.001. Scar bars: 100 μm.

Article Snippet: Human umbilical vein endothelial cells (HUVECs) were purchased from ScienCell (San Diego, USA) and cultured in endothelial cell medium (ECM, Gibco, Life Technologies) supplemented with 10% FBS.

Techniques: Transfection, Fluorescence, Microscopy, Expressing, Western Blot, CCK-8 Assay, Tube Formation Assay

Journal: American Journal of Translational Research

Article Title: Transplantation of HUVECs with genetically modified Nogo-B accelerates wound-healing in nude mice

doi:

Figure Lengend Snippet: Cytokines detected in HUVECs supernatant after knocked down Nogo-B. A. Cytokines in supernatant of HUVEC-NC and HUVEC-siNogo-B detected by antibody microarray. They were labeled by Cy3 and Cy5, which were two different fluorescent molecular. B. It showed the ratio of siNogo-B/NC from antibody microarray. EGF, VEGF-D, FGF-4 and Progranulin were increased obviously in the HUVEC-siNogo-B supernatant, whereas anti-angiogenesis cytokines MMP-19 and VEGI declined.

Article Snippet: Human umbilical vein endothelial cells (HUVECs) were purchased from ScienCell (San Diego, USA) and cultured in endothelial cell medium (ECM, Gibco, Life Technologies) supplemented with 10% FBS.

Techniques: Microarray, Labeling

Journal: American Journal of Translational Research

Article Title: Transplantation of HUVECs with genetically modified Nogo-B accelerates wound-healing in nude mice

doi:

Figure Lengend Snippet: HUVEC-siNogo-B-CM exhibited chemoattractive, mitogenic effects on fibroblasts. (A, B) The pictures showed the scratch test of fibroblasts in 24 h. The number of migrating fibroblasts in 50% HUVEC-siNogo-B group was significantly greater than that in 50% HUVEC-NC group in 24 h the migration (C) and proliferation (D) of fibroblasts were measured by Transwell Assay and CCk-8 Assay. The proliferative activity of fibroblasts increased markedly in 50% HUVEC-siNogo-B group as compared with 50% HUVEC-NC group after 4 days of co-culturing. Data was shown as means ± SD, n=3, *P<0.05, ***P<0.001. The scale bar is 500 µm.

Article Snippet: Human umbilical vein endothelial cells (HUVECs) were purchased from ScienCell (San Diego, USA) and cultured in endothelial cell medium (ECM, Gibco, Life Technologies) supplemented with 10% FBS.

Techniques: Migration, Transwell Assay, CCK-8 Assay, Activity Assay

Journal: American Journal of Translational Research

Article Title: Transplantation of HUVECs with genetically modified Nogo-B accelerates wound-healing in nude mice

doi:

Figure Lengend Snippet: The HUVECs-siNogo-B-CM promoted migration of epidermal cells. The migration of epidermal cells was measured by scratch test (A, B) and CCk-8 Assay (C). The epidermal cells in the 50% HUVEC-siNogo-B group was significantly greater than that in the 50% HUVEC-NC group in 12 h and 36 h. The scale bar is 500 µm. The epidermal cells in 50% HUVEC-siNogo-B group increased significantly faster after 4 days co-culturing. 5% FBS was treated as positive control and HUVEC-NC, DMEM as negtive control. Data was shown as means ± SD, n=3; *P<0.05, ***P<0.001. The scale bar is 500 µm.

Article Snippet: Human umbilical vein endothelial cells (HUVECs) were purchased from ScienCell (San Diego, USA) and cultured in endothelial cell medium (ECM, Gibco, Life Technologies) supplemented with 10% FBS.

Techniques: Migration, CCK-8 Assay, Positive Control, Control

Journal: American Journal of Translational Research

Article Title: Transplantation of HUVECs with genetically modified Nogo-B accelerates wound-healing in nude mice

doi:

Figure Lengend Snippet: HUVEC-siNogo-B could survive in 48 h after injected intradermally around the excisional wound on mice. The flow cytometry and the Immunofluorescence were performed to detected the survival of HUVEC-siNogo-B (GFP+) cells. The blue spots in the top left corner represents alive cells (A). The quantity of blue spots in two injection groups was much higher compared with vehicle control group. (B) Representative immunofluorescent staining of The GFP-labeled HUVECs. GFP+DAPI+ were survival cells by initially injected.

Article Snippet: Human umbilical vein endothelial cells (HUVECs) were purchased from ScienCell (San Diego, USA) and cultured in endothelial cell medium (ECM, Gibco, Life Technologies) supplemented with 10% FBS.

Techniques: Injection, Flow Cytometry, Immunofluorescence, Control, Staining, Labeling

Journal: American Journal of Translational Research

Article Title: Transplantation of HUVECs with genetically modified Nogo-B accelerates wound-healing in nude mice

doi:

Figure Lengend Snippet: Topical administration of HUVEC-siNogo-B accelerated wound healing on mice. A. The wound was treated with saline, HUVEC-NC and HUVEC-siNogo-B, examined at day 1, day 3, day 5, day 7, day 14 after wounding and digitally photographed. B. Quantitative analysis of wound closure demonstrating higher wound healing rate in the HUVEC-siNogo-B group. Data was shown as means ± SD, n=6; ***P<0.001.

Article Snippet: Human umbilical vein endothelial cells (HUVECs) were purchased from ScienCell (San Diego, USA) and cultured in endothelial cell medium (ECM, Gibco, Life Technologies) supplemented with 10% FBS.

Techniques: Saline

Journal: American Journal of Translational Research

Article Title: Transplantation of HUVECs with genetically modified Nogo-B accelerates wound-healing in nude mice

doi:

Figure Lengend Snippet: The HUVEC-siNogo-B increased callipary density in wound of nude mice. (A) Representative HE staining at day 7 after wounding. (B) Representative Immunohistochemistry staining of CD31 at day 7. Statistical analysis of CD31+ newly formed vessels showed that neovascularization was more obvious in HUVEC-siNogo-B treated wounds (C). Data was shown as means ± SD, n=5; ***P<0.001. The scale bar is 100 µm.

Article Snippet: Human umbilical vein endothelial cells (HUVECs) were purchased from ScienCell (San Diego, USA) and cultured in endothelial cell medium (ECM, Gibco, Life Technologies) supplemented with 10% FBS.

Techniques: Staining, Immunohistochemistry