dermal blood microvascular endothelial cells  (PromoCell)

Journal: bioRxiv

Article Title: VEGF counteracts shear stress-determined arterial fate specification during capillary remodeling

doi: 10.1101/2024.01.23.576920

Figure Lengend Snippet: a. HMVBECs were used to test the induction of arterial and Notch genes under the conditions of static (ST), ST with VEGF (20ng/ml), FSS (12dyn/cm 2 ) and FSS with VEGF. Gene expression was examined by quantitative RT-PCR followed by quantification relative to ST. b. FSS-induced expression of GJA4, GJA5 and Hey1 in HMVBECs with a gradient of VEGF. Gene expression was examined by quantitative RT-PCR followed by quantification relative to ST. FSS experiments were conducted using Vitroflo system. All conditions under FSS were in the same Vitroflo plate. c. HUVECs treated with siRNA were used to test FSS-induced expression of arterial genes and the effect of VEGF. d. A working model demonstrating VEGF signals through VEGFR2-PLCγ1 to counteract FSS-induced arterial specification. Student t-test was used for statistics. p<0.05 was determined as statistically significant. ns: not significant.

Article Snippet: Human microvascular blood endothelial cells (HMVBECs) (HDBECs, Promocell, #C-12211) were maintained in EGM2-MV full growth medium (Lonza).

Techniques: Expressing, Quantitative RT-PCR

Journal: bioRxiv

Article Title: VEGF counteracts shear stress-determined arterial fate specification during capillary remodeling

doi: 10.1101/2024.01.23.576920

Figure Lengend Snippet: a. HMVBECs were subjected to VEGF (20ng/ml) or FSS (12 dyn/cm 2 ) to test the activation of Notch signaling in a time course, with DMSO or γ-secretase inhibitor DAPT treatments. b. HMVBECs treated with siCTRL or siDll4 were subjected to VEGF and FSS to test the activation of Notch signaling. c-d. Working models demonstrating the mechanisms by which VEGF and FSS use to activate Notch signaling. VEGF activates the expression of Dll4, which consequently serves as a ligand to activate Notch signaling in a neighboring cell. By contrast, FSS does not rely on increasing Dll4 expression to activate Notch. Increase of Dll4 expression is a result rather than a cause of FSS-induced Notch activation. e. FSS-induced Notch signaling and expression of arterial genes. HMVBECs were subjected to FSS in a time course in the absence or presence of VEGF.

Article Snippet: Human microvascular blood endothelial cells (HMVBECs) (HDBECs, Promocell, #C-12211) were maintained in EGM2-MV full growth medium (Lonza).

Techniques: Activation Assay, Expressing

Journal: bioRxiv

Article Title: VEGF counteracts shear stress-determined arterial fate specification during capillary remodeling

doi: 10.1101/2024.01.23.576920

Figure Lengend Snippet: a. Pretreated with DMSO or 5uM DAPT for 3h, HMVBECs were subjected to FSS to induce arterial differentiation for 16h. Samples were analyzed by western blot. b. Representative retinal artery and branching arterioles at p5 from the Notch reporter mouse line CBF:H2B-Venus. Arterial ECs were labeled by immunostaining of Sox17 and colocalized with endogenous Venus signal in the nuclei. Arrow heads indicate Sox17 high /CBF-arterial ECs. Scale bars: 25 µm. c. In the background of DAPT-induced Notch inhibition (P1-3), arterial morphogenesis in P4 retina from control and PLCg1 iECKO mice. White arrow heads indicate arterial morphogenesis marked by Sox17 high ECs; yellow arrow heads indicate smooth muscle coverage. Abbreviations: A: artery; V: vein. Scale bars: 100 µm.

Article Snippet: Human microvascular blood endothelial cells (HMVBECs) (HDBECs, Promocell, #C-12211) were maintained in EGM2-MV full growth medium (Lonza).

Techniques: Western Blot, Labeling, Immunostaining, Inhibition, Control

Journal: Frontiers in Pharmacology

Article Title: Identification of kinases activated by multiple pro-angiogenic growth factors

doi: 10.3389/fphar.2022.1022722

Figure Lengend Snippet: Establishment of the Experimental System. (A) . Demonstration of Dermal Microvascular Endothelial Cell (DMEC) labeling. Fluorescent image of nuclear-localized mCherry overlaid on a phase contrast image of confluent DMECs. (B) . The proliferation of DMECs in full growth medium (dotted line) is exponential (solid line is a fitted exponential curve). (C) . Cell cycle analysis of relative DNA content determined by labeling live cells with Hoescht and imaged with a microscope. Full growth medium (left) produces a familiar distribution of cells while 24 h in Basal Proliferation Medium (BPM) (right) produces far fewer cells in S- and G2/M-phases. (D) . Demonstration of YOYO-1 dye to detect dead cells. (E) .) Example of nuclei (closed circles) and dead cells (open circles) over time in BPM. (F) . Cell death in BPM is countered by inhibition of caspase activity via 10 µM Q-VD-OPh.

Article Snippet: Human Dermal blood microvascular endothelial cells (DMECs) isolated from a single donor were purchased from Lonza (CC-2183).

Techniques: Labeling, Cell Cycle Assay, Microscopy, Inhibition, Activity Assay

Journal: Frontiers in Pharmacology

Article Title: Identification of kinases activated by multiple pro-angiogenic growth factors

doi: 10.3389/fphar.2022.1022722

Figure Lengend Snippet: Concentration-Dependent Population Dynamics of Dermal Microvascular Endothelial Cells (DMECs) in the Presence of Pro-Angiogenic Growth Factors. (A) . The number of cells ( N ) over time for four concentrations of FGF2. (B) . The number of dead cells ( M ) over time for the experiment shown in panel (A) . (C) . The proliferation rate, p , for the same experiment as in panels A,B. (D) . The birth rate, B , as a function of time. (E) . The death rate, D , as a function of time.

Article Snippet: Human Dermal blood microvascular endothelial cells (DMECs) isolated from a single donor were purchased from Lonza (CC-2183).

Techniques: Concentration Assay

Journal: Frontiers in Pharmacology

Article Title: Identification of kinases activated by multiple pro-angiogenic growth factors

doi: 10.3389/fphar.2022.1022722

Figure Lengend Snippet: The dose-response proliferative behavior of Dermal Microvascular Endothelial Cells (DMECs) to Three Pro-Angiogenic Growth Factors. The proliferation rate (A) , the birth rate (B) , and the death rate (C) for FGF2, VEGFA, and HGF over a range of concentrations. Note that the y -axes share the same scale to facilitate comparison of the relative magnitude of each.

Article Snippet: Human Dermal blood microvascular endothelial cells (DMECs) isolated from a single donor were purchased from Lonza (CC-2183).

Techniques: Comparison

Journal: Frontiers in Pharmacology

Article Title: Identification of kinases activated by multiple pro-angiogenic growth factors

doi: 10.3389/fphar.2022.1022722

Figure Lengend Snippet: Identification of kinases important for the proliferation rate of Dermal Microvascular Endothelial Cells (DMECs) in each pro-angiogenic growth factor studied. (A) . Magnitudes of the Kinome Regression (KIR) coefficients (i.e., influential kinases) for each identified growth factor. (B) . Venn diagram revealing the exclusivity and commonalities in the set of kinases identified for each growth factor. (C) . The list of the intersection of kinases implicated in all three growth factors. Kinases are ordered according to a rank obtained by summing each kinase’s ranks across all growth factors.

Article Snippet: Human Dermal blood microvascular endothelial cells (DMECs) isolated from a single donor were purchased from Lonza (CC-2183).

Techniques:

Journal: Frontiers in Pharmacology

Article Title: Identification of kinases activated by multiple pro-angiogenic growth factors

doi: 10.3389/fphar.2022.1022722

Figure Lengend Snippet: Orthogonal evidence supporting the role of kinases identified by Kinome Regression (KIR) Analysis. (A) . Targeting growth factor receptors with 5 nM of siRNA reduces the proliferation rate of Dermal Microvascular Endothelial Cells (DMECs) in the presence of cognate growth factors while minimizing reduction in proliferation in other growth factors. (B) . Targeting kinases with siRNA largely agrees with kinases implicated by KIR. Here, the effect is defined as the average over all three siRNAs used for each kinase and normalized to the no-siRNA control. Error bars are s. e.m. The asterisks indicate growth factors context in which the siRNA targeting a given kinase produced a statistically significant result ( p < 0.05 from multiple comparison testing).

Article Snippet: Human Dermal blood microvascular endothelial cells (DMECs) isolated from a single donor were purchased from Lonza (CC-2183).

Techniques: Control, Produced, Comparison

Journal: Microcirculation (New York, N.Y. : 1994)

Article Title: A bioengineered lymphatic vessel model for studying lymphatic endothelial cell-cell junction and barrier function

doi: 10.1111/micc.12730

Figure Lengend Snippet: (A) A schematic of an organotypic 3D lymphatic vessel model (LV-on-chip). Prox-1 (green) and CD31 (red) expression confirms lymphatic endothelial identity and cell morphology in the channel. (B) Morphologic changes in human dermal microvascular blood endothelial cells (BECs) with lymphatic endothelial cells (LECs) after one day of cell seeding. BECs become more contractile than LECs, forming a smaller vessel diameter compared to LECs. (C) BVs and LVs observed in mouse ear tissues. mLYVE-1, anti-mouse LYVE-1 antibody; mCD31, anti-mouse CD31 antibody. (D) Phalloidin (red) and anti-VE-cad (VE-cadherin) antibody (green) staining to visualize F-actin and adherens junctions. (E) Lymphatic and blood vessel barrier function. 70 kDa dextran was introduced into the vessel lumens and dextran diffusion was observed in real time under microscopy. Superimposed red dashed lines represent the edges of the vessel lumens. (F) Quantification of the permeability of BEC-generated engineered BVs and LEC-generated LVs. ** p = 0.0016, two tailed unpaired Student t-test, n = 5 per group. Data are expressed as mean ± S.E.M.

Article Snippet: Cell culture Primary human dermal microvascular lymphatic endothelial cells (LECs, neonatal) were purchased from Lonza.

Techniques: Expressing, Staining, Diffusion-based Assay, Microscopy, Permeability, Generated, Two Tailed Test