Journal: Nature Communications

Article Title: Expansion of functional personalized cells with specific transgene combinations

doi: 10.1038/s41467-018-03408-4

Figure Lengend Snippet: Overview of cell types immortalized in this study

Article Snippet: The following media were used for cultivation of fibroblasts: huFib medium (InSCREENeX GmbH), bone marrow stroma cells: huMSC medium (InSCREENeX GmbH), HUVECs: huVEC medium (InSCREENeX GmbH), HMVECs: huMEC medium (InSCREENeX GmbH), LECs: huLEC medium (InSCREENeX GmbH), chondrocytes: huChon medium (InSCREENeX GmbH), and osteoblasts: huOB maintenance medium (InSCREENeX GmbH).

Techniques:

Journal: Nature Communications

Article Title: Expansion of functional personalized cells with specific transgene combinations

doi: 10.1038/s41467-018-03408-4

Figure Lengend Snippet: Hepatocyte cell lines retain key hepatic properties. a Albumin and Glucose-6-phosphatase gene expression analysis of expandable hepatocytes cultured in standard 2D or 3D (9 days of cultivation as spheroids in spinner flasks) conditions and primary hepatocytes (pHep) either cultured in 3D conditions or freshly isolated (expression normalized to Gapdh, n = 3 independent experiments, except for pHep were n = 2 independent experiments, bars represent mean). b Immunofluorescence-based detection of Albumin and E-cadherin in e-mHepA expandable hepatocytes counterstained for nuclei with DAPI. Scale bar, 20 μm. c Glycogen storage in expandable hepatocytes and control fibroblasts as analyzed by Periodic Acid-Schiff staining. Representative bright field micrographs and quantification of multiple field of views are shown. Scale bar, 100 μm (mean, pooled data from n = 3 independent experiments, field of views analyzed in total 35 to 42). d Immunofluorescence micrograph of Albumin and Hnf4α in e-mHepB hepatocytes after 3D culture as spheroids for 9 days. Actin filaments and nuclei were counterstained with phalloidin and DAPI, respectively. Scale bar, 20 µm. e Induction of phase I metabolic enzyme expression in expandable hepatocytes cultured in 2D or 3D conditions and control fibroblasts in response to stimulation with 50 μM Dexamethasone or 2 μM 3-Methylcholanthrene for 72 h (expression normalized to Gapdh, n = 3 independent experiments, except for e-mHepB Cyp1a1 expression were n = 6 independent experiments, bars represent mean). f Luminescence-based detection of Phase I enzymatic activity in expandable hepatocytes and control fibroblasts after cells were treated as indicate above (activity normalized to 10 4 cells, pooled data from n = 2 independent experiments with n = 3 technical replicates each). g Staining of livers of Fah –/– / Rag2 –/– /Il2rg –/– (FRG) mice for FAH and GFP, 90 days after transplantation of GFP-tagged expandable hepatocytes. Scale bar, 100 µm

Article Snippet: The following media were used for cultivation of fibroblasts: huFib medium (InSCREENeX GmbH), bone marrow stroma cells: huMSC medium (InSCREENeX GmbH), HUVECs: huVEC medium (InSCREENeX GmbH), HMVECs: huMEC medium (InSCREENeX GmbH), LECs: huLEC medium (InSCREENeX GmbH), chondrocytes: huChon medium (InSCREENeX GmbH), and osteoblasts: huOB maintenance medium (InSCREENeX GmbH).

Techniques: Gene Expression, Cell Culture, Isolation, Expressing, Immunofluorescence, Control, Staining, Activity Assay, Transplantation Assay

Journal: Nature Communications

Article Title: Expansion of functional personalized cells with specific transgene combinations

doi: 10.1038/s41467-018-03408-4

Figure Lengend Snippet: Comparable phenotype of primary and expanded endothelial cells. a Phase contrast microscopy shows the expandable HUVEC cell line e-hUVEC-2 (MYC, ID1, and ID2). Scale bar 100 µm. b CD31 expression of human endothelial cell populations immortalized with three different gene sets as indicated. c Global gene expression analysis was performed on three different HUVEC lines (e-hUVEC-2, 8 and 9; in duplicate) (3, 4, 5, respectively) and two independent primary HUVEC populations (2) as well as four primary gingiva fibroblast populations (1). Expression data was processed with GeneSpring 11.5.1 software and a Standard Pearson Correlation was determined for each gene versus all other genes. The correlation heatmap depicts the pair-wise correlation coefficient between the given samples and displays the relationship between the different samples. The samples are clustered based on the pair-wise correlation coefficients between all entities. d Phenotypic stability of e-hUVEC-2 cells after 45 and 90 cumulative population doublings was evaluated by CD31 (also known as PECAM1) expression, acetylated LDL uptake, and eNOS activity. Gray fill: antibody isotype control; black outline: stained sample. MFI: median fluorescence intensity. e Immunofluorescence-based detection of CD31 and CD146 (also known as MCAM) in expandable HUVECs counterstained for nuclei with DAPI. Scale bars, 100 µm. f The phenotype and the functionality of cell line e-hUVEC-2 (cumulative population doubling 80) was compared to primary HUVECs based on flow cytometric analysis of CD31, TIE1, TIE2, and CD309 (also known as VEGFR2) expression. Gray fill: antibody isotype control; black outline: stained sample. MFI: median fluorescence intensity. g The angiogenic potential of primary HUVECs and e-hUVEC-2 was determined in vitro by a matrigel tube formation assay. Scale bars, 200 µm. h Spheroids in matrigel of primary HUVEC and e-hUVEC-2 were subcutaneously injected into Rag2 -/- Il2r g −/− mice. After two weeks the implants were dissected and stained for human CD31 (brown color). e-hUVEC-2 organized into human CD31 positive microvessels similar to primary HUVEC. Scale bars, 100 µm

Article Snippet: The following media were used for cultivation of fibroblasts: huFib medium (InSCREENeX GmbH), bone marrow stroma cells: huMSC medium (InSCREENeX GmbH), HUVECs: huVEC medium (InSCREENeX GmbH), HMVECs: huMEC medium (InSCREENeX GmbH), LECs: huLEC medium (InSCREENeX GmbH), chondrocytes: huChon medium (InSCREENeX GmbH), and osteoblasts: huOB maintenance medium (InSCREENeX GmbH).

Techniques: Microscopy, Expressing, Gene Expression, Software, Activity Assay, Control, Staining, Fluorescence, Immunofluorescence, In Vitro, Tube Formation Assay, Injection

Journal: Molecular Biology of the Cell

Article Title: A novel function of Tis11b/BRF1 as a regulator of Dll4 mRNA 3′-end processing

doi: 10.1091/mbc.E11-02-0149

Figure Lengend Snippet: Endogenous Tis11b protein targets and regulates Dll4 mRNA in endothelial cells. (A–C) Inhibition of endogenous Tis11b protein was achieved by transfection of HMVECd or HuAEC with one of the two different siRNAs targeting the human Tis11b sequence (T11b1, T11b2) or negative control siRNA (CTL). (A) Western blot analysis of Tis11b, Dll4, and tubulin in cell extracts from cells depleted of Tis11b protein (T11b1 or T11b2) and control cells (CTL). (B) Steady-state levels of Dll4 mRNA were quantified by RT-qrtPCR in cell extracts from siTis11b-treated cells (T11b1 or T11b2) or control cells (CTL). (C) Steady-state levels of Dll4 mRNA were quantified by RT-qPCR in Tis11b-depleted cell extracts (T11b1 or T11b2) or control extracts (CTL). Data are representative of at least three independent experiments. (D) RNP complexes were immunoprecipitated after in vivo cross-linking as described in Materials and Methods with anti-Tis11b antibodies (αTis11b) or nonimmune IgG (NI-IgGs). Presence of Dll4 or HPRT mRNA was detected by RT-PCR in the nonprecipitated extracts (Input) or in the immunoprecipitates.

Article Snippet: Primary endothelial cells (HMVECd and HuAEC) were purchased from LONZA (Basel, Switzerland) and cultivated according to the manufacturer's instructions.

Techniques: Inhibition, Transfection, Sequencing, Negative Control, Western Blot, Control, Quantitative RT-PCR, Immunoprecipitation, In Vivo, Reverse Transcription Polymerase Chain Reaction

Journal: Molecular Biology of the Cell

Article Title: A novel function of Tis11b/BRF1 as a regulator of Dll4 mRNA 3′-end processing

doi: 10.1091/mbc.E11-02-0149

Figure Lengend Snippet: Identification of potential binding sites for Tis11b protein in Dll4 mRNA. 3′-RACE experiments were performed on human endothelial microvascular RNA and 12 clones were sequenced. Sequence in gray background indicates nucleotides present in the NM_019074.2 refseq, boxed sequences are putative binding sites for Tis11b (ARE1 and ARE2), and underlined sequences are UGUA elements. Arrowheads indicate the positions of the added poly(A) tails determined from sequencing and the large arrowhead represents a hot spot found in 5 out of 12 clones.

Article Snippet: Primary endothelial cells (HMVECd and HuAEC) were purchased from LONZA (Basel, Switzerland) and cultivated according to the manufacturer's instructions.

Techniques: Binding Assay, Clone Assay, Sequencing

Journal: Toxicology Research

Article Title: Nanoparticle induced barrier function assessment at liquid–liquid and air–liquid interface in novel human lung epithelia cell lines † †Electronic supplementary information (ESI) available. See DOI: 10.1039/c9tx00179d

doi: 10.1039/c9tx00179d

Figure Lengend Snippet: Characterization of growth behavior: Growth curves of A549, hAELVi and huAEC cells show similar growth behavior for all cell types.

Article Snippet: CI-hAELVi (cat. no.: INS-CI-1015) and CI-huAEC (cat. no.: INS-CI-1011) cells were purchased from InSCREENeX GmbH (InSCREENeX GmbH, Germany).

Techniques:

Journal: The Journal of Physiological Sciences : JPS

Article Title: ABCG1 deficiency promotes endothelial apoptosis by endoplasmic reticulum stress-dependent pathway

doi: 10.1007/s12576-013-0281-8

Figure Lengend Snippet: Endothelial ABCG1 expression was correlated with cholesterol efflux and intracellular lipid content. a, b HUAEC were transfected with scrambled siRNA, ABCG1 siRNA and either pcDNA (empty vector) or a pReceiver-ABCG1 expression vector. ABCG1 mRNA (a) and protein expression (b) were measured by real time PCR and western blotting. c Cholesterol efflux to HDL was decreased in ABCG1-deficient EC (ABCG1−), and reversed in ABCG1 overexpression plasmid transfection (ABCG1+). d Intracellular cholesterol content was increased in ABCG1-deficient EC while decreased in ABCG1 overexpression plasmid transfection. TC total cholesterol, FC free cholesterol, CE cholesterol ester. Data represent mean ± SD (n = 3). *P < 0.05, **P < 0.001 vs. EC control

Article Snippet: Cell culture Human umbilical artery endothelial cells (HUAEC; ScienCell) were cultured in Endothelial Cell Medium (No.1001; ScienCell) in a CO 2 /O 2 incubator at 37 °C.

Techniques: Expressing, Transfection, Plasmid Preparation, Real-time Polymerase Chain Reaction, Western Blot, Over Expression, Control

Journal: The Journal of Physiological Sciences : JPS

Article Title: ABCG1 deficiency promotes endothelial apoptosis by endoplasmic reticulum stress-dependent pathway

doi: 10.1007/s12576-013-0281-8

Figure Lengend Snippet: Downregulation of ABCG1 promoted endothelial apoptosis. Annexin V-FITC/PI double-staining assay was used to quantify apoptosis in HUAEC by means of flow cytometry. Endothelial apoptosis includes early apoptotic (FITC +PI−) and late apoptotic (FITC +PI+). a Apoptosis in endothelial control, b increased apoptosis in ABCG1-deficient EC, c apoptosis in ABCG1-deficient EC transfected with ABCG1 plasmid, d apoptosis in endothelial control transfected with ABCG1 plasmid

Article Snippet: Cell culture Human umbilical artery endothelial cells (HUAEC; ScienCell) were cultured in Endothelial Cell Medium (No.1001; ScienCell) in a CO 2 /O 2 incubator at 37 °C.

Techniques: Double Staining, Flow Cytometry, Control, Transfection, Plasmid Preparation

Journal: The Journal of Physiological Sciences : JPS

Article Title: ABCG1 deficiency promotes endothelial apoptosis by endoplasmic reticulum stress-dependent pathway

doi: 10.1007/s12576-013-0281-8

Figure Lengend Snippet: Blocking ER stress reduced endothelial apoptosis in ABCG1-deficient EC. Compared with control cells (a), endothelial apoptosis was significantly induced in ABCG1-deficient EC (b). c Endothelial apoptosis was partly reversed in ABCG1-deficient EC treated with PBA. d Endothelial apoptosis was moderately but not statistically decreased in control cells treated with PBA. Endothelial apoptosis includes early apoptotic (FITC +PI−) and late apoptotic (FITC +PI+) as described in “Methods”

Article Snippet: Cell culture Human umbilical artery endothelial cells (HUAEC; ScienCell) were cultured in Endothelial Cell Medium (No.1001; ScienCell) in a CO 2 /O 2 incubator at 37 °C.

Techniques: Blocking Assay, Control

Journal: The Journal of Physiological Sciences : JPS

Article Title: ABCG1 deficiency promotes endothelial apoptosis by endoplasmic reticulum stress-dependent pathway

doi: 10.1007/s12576-013-0281-8

Figure Lengend Snippet: Cholesterol repletion and depletion were correlated with endothelial apoptosis and activation of ER stress. a Cyclodextrin (CD)-cholesterol loading of control cells increased FC and CE content. CD-cholesterol loading of ABCG1-deficient EC statistically increased CE content but not FC content. Cholesterol depletion with CD lowed FC and CE content in both endothelial control and ABCG1-deficient EC. b Cholesterol repletion induced endothelial apoptosis in endothelial controls. Cholesterol depletion significantly reversed endothelial apoptosis in ABCG1-deficient EC. Apoptosis was suppressed in ABCG1-deficient EC treated with U18666A, a cholesterol transport to ER inhibitor. c, d Cholesterol repletion increased the expression of GRP78 and CHOP, and cholesterol depletion reduced the expression of GRP78 and CHOP in ABCG1-deficient EC and control cells. e, f Addition of U18666A to ABCG1-deficient EC suppressed expression of GRP78 and CHOP. Data are mean ± SD (n = 3). *P = 0.05, **P < 0.001 vs. EC controls, # P < 0.001 vs. ABCG1-deficient EC. Representative images are shown for three independent experiments

Article Snippet: Cell culture Human umbilical artery endothelial cells (HUAEC; ScienCell) were cultured in Endothelial Cell Medium (No.1001; ScienCell) in a CO 2 /O 2 incubator at 37 °C.

Techniques: Activation Assay, Control, Expressing

Journal: International Journal of Molecular Sciences

Article Title: Venous and Arterial Endothelial Cells from Human Umbilical Cords: Potential Cell Sources for Cardiovascular Research

doi: 10.3390/ijms22020978

Figure Lengend Snippet: Morphology of endothelial cells. Human umbilical cord-derived endothelial cells from veins (HUVEC) or arteries (HUAEC) were seeded at a density of 15,000 cells/well at day 0, and cultivated for 2, 4, and 7 days. Phase contrast images were taken at 10-fold primary magnification. Cells developed the typical cobblestone-like morphology of mature endothelial cells with increasing proliferation time. Scale bar represents 100 µm.

Article Snippet: Pooled HUVEC (from 3 to 6 individual donors, Lonza, Cologne, Germany) and HUAEC (from 250 individual donors, PeloBiotech GmbH, Planegg, Germany) were cultivated in EGM-2 Bullet Kit containing endothelial cell culture medium supplemented with 2% ( v / v ) fetal bovine serum, vascular endothelial growth factor, basic fibroblast growth factor, human epithelial growth factor, insulin-like growth factor-1, hydrocortisone, heparin, ascorbic acid, gentamycin, and amphotericin B (Lonza) in a standard humidified incubator at 37 °C with 5% ( v / v ) CO 2 .

Techniques: Derivative Assay

Journal: International Journal of Molecular Sciences

Article Title: Venous and Arterial Endothelial Cells from Human Umbilical Cords: Potential Cell Sources for Cardiovascular Research

doi: 10.3390/ijms22020978

Figure Lengend Snippet: Endothelial cell viability and density. Human umbilical cord-derived endothelial cells from veins (HUVEC) or arteries (HUAEC) were seeded at a density of 15,000 cells/well at day 0, and cultivated for 2, 4, and 7 days. Cell viability staining detected living (green) and dead (red) cells ( A ). Quantification of vital cells showed that only a few cells died over time ( B ). The cell density, expressed as the number of vital adherent cells per mm 2 , increased over time ( C ). Fluorescent images were taken at 10-fold primary magnification. Scale bar represents 100 µm, and 50 µm in the magnified image sections in the upper right corner of each image. Squares with dashed lines indicate the image region that was magnified. Straight black lines indicate p ≤ 0.0099.

Article Snippet: Pooled HUVEC (from 3 to 6 individual donors, Lonza, Cologne, Germany) and HUAEC (from 250 individual donors, PeloBiotech GmbH, Planegg, Germany) were cultivated in EGM-2 Bullet Kit containing endothelial cell culture medium supplemented with 2% ( v / v ) fetal bovine serum, vascular endothelial growth factor, basic fibroblast growth factor, human epithelial growth factor, insulin-like growth factor-1, hydrocortisone, heparin, ascorbic acid, gentamycin, and amphotericin B (Lonza) in a standard humidified incubator at 37 °C with 5% ( v / v ) CO 2 .

Techniques: Derivative Assay, Staining

Journal: International Journal of Molecular Sciences

Article Title: Venous and Arterial Endothelial Cells from Human Umbilical Cords: Potential Cell Sources for Cardiovascular Research

doi: 10.3390/ijms22020978

Figure Lengend Snippet: Metabolic activity ( A ) and membrane integrity ( B ) of endothelial cells. Human umbilical cord-derived endothelial cells from veins (HUVEC) or arteries (HUAEC) were seeded at a density of 15,000 cells/well at day 0 and cultivated for 2, 4, and 7 days. The metabolic activity and membrane integrity were normalized to the number of vital adherent cells per mm 2 leading to decreasing values over time. Straight black lines indicate p ≤ 0.0058.

Article Snippet: Pooled HUVEC (from 3 to 6 individual donors, Lonza, Cologne, Germany) and HUAEC (from 250 individual donors, PeloBiotech GmbH, Planegg, Germany) were cultivated in EGM-2 Bullet Kit containing endothelial cell culture medium supplemented with 2% ( v / v ) fetal bovine serum, vascular endothelial growth factor, basic fibroblast growth factor, human epithelial growth factor, insulin-like growth factor-1, hydrocortisone, heparin, ascorbic acid, gentamycin, and amphotericin B (Lonza) in a standard humidified incubator at 37 °C with 5% ( v / v ) CO 2 .

Techniques: Activity Assay, Membrane, Derivative Assay

Journal: International Journal of Molecular Sciences

Article Title: Venous and Arterial Endothelial Cells from Human Umbilical Cords: Potential Cell Sources for Cardiovascular Research

doi: 10.3390/ijms22020978

Figure Lengend Snippet: Secretion of vasoactive substances by endothelial cells. Human umbilical cord-derived endothelial cells from veins (HUVEC) or arteries (HUAEC) were seeded at a density of 15,000 cells/well at day 0 and cultivated for 2, 4, and 7 days. The amounts of the sum of nitrite and nitrate ( A ), prostacyclin ( B ), and thromboxane B2 ( C ) were quantified by ELISA and normalized to the number of vital adherent cells per mm 2 . While less nitrite/nitrate and prostacyclin were released over time, the amount of thromboxane B2 remained constant. Straight black lines indicate p ≤ 0.0063.

Article Snippet: Pooled HUVEC (from 3 to 6 individual donors, Lonza, Cologne, Germany) and HUAEC (from 250 individual donors, PeloBiotech GmbH, Planegg, Germany) were cultivated in EGM-2 Bullet Kit containing endothelial cell culture medium supplemented with 2% ( v / v ) fetal bovine serum, vascular endothelial growth factor, basic fibroblast growth factor, human epithelial growth factor, insulin-like growth factor-1, hydrocortisone, heparin, ascorbic acid, gentamycin, and amphotericin B (Lonza) in a standard humidified incubator at 37 °C with 5% ( v / v ) CO 2 .

Techniques: Derivative Assay, Enzyme-linked Immunosorbent Assay

Journal: International Journal of Molecular Sciences

Article Title: Venous and Arterial Endothelial Cells from Human Umbilical Cords: Potential Cell Sources for Cardiovascular Research

doi: 10.3390/ijms22020978

Figure Lengend Snippet: Expression of actin fibers by endothelial cells. Human umbilical cord-derived endothelial cells from veins (HUVEC) or arteries (HUAEC) were seeded at a density of 15,000 cells/well at day 0 and cultivated for 2, 4, and 7 days. While most of the cells exhibited actin fibers throughout the cell body at day 2, an increasing number of cells with pronounced cortical actin fibers were present at day 4 and 7 (indicated by white arrows). Fluorescent images were taken at 20-fold primary magnification. Scale bar represents 50 µm, and 25 µm the in magnified image sections in the upper right corner of each image. Squares with dashed lines indicate the image region that was magnified.

Article Snippet: Pooled HUVEC (from 3 to 6 individual donors, Lonza, Cologne, Germany) and HUAEC (from 250 individual donors, PeloBiotech GmbH, Planegg, Germany) were cultivated in EGM-2 Bullet Kit containing endothelial cell culture medium supplemented with 2% ( v / v ) fetal bovine serum, vascular endothelial growth factor, basic fibroblast growth factor, human epithelial growth factor, insulin-like growth factor-1, hydrocortisone, heparin, ascorbic acid, gentamycin, and amphotericin B (Lonza) in a standard humidified incubator at 37 °C with 5% ( v / v ) CO 2 .

Techniques: Expressing, Derivative Assay

Journal: International Journal of Molecular Sciences

Article Title: Venous and Arterial Endothelial Cells from Human Umbilical Cords: Potential Cell Sources for Cardiovascular Research

doi: 10.3390/ijms22020978

Figure Lengend Snippet: Expression of von Willebrand factor (vWF) by endothelial cells. Human umbilical cord-derived endothelial cells from veins (HUVEC) or arteries (HUAEC) were seeded at a density of 15,000 cells/well at day 0 and cultivated for 2, 4, and 7 days. Cells from both sources expressed vWF at all time points. At day 4 and 7, some extracellular vWF fibers were observed (white arrows). Fluorescent images were taken at 20-fold primary magnification. Scale bar represents 50 µm, and 25 µm in the magnified image sections in the upper right corner of each image. Squares with dashed lines indicate the image region that was magnified.

Article Snippet: Pooled HUVEC (from 3 to 6 individual donors, Lonza, Cologne, Germany) and HUAEC (from 250 individual donors, PeloBiotech GmbH, Planegg, Germany) were cultivated in EGM-2 Bullet Kit containing endothelial cell culture medium supplemented with 2% ( v / v ) fetal bovine serum, vascular endothelial growth factor, basic fibroblast growth factor, human epithelial growth factor, insulin-like growth factor-1, hydrocortisone, heparin, ascorbic acid, gentamycin, and amphotericin B (Lonza) in a standard humidified incubator at 37 °C with 5% ( v / v ) CO 2 .

Techniques: Expressing, Derivative Assay

Journal: International Journal of Molecular Sciences

Article Title: Venous and Arterial Endothelial Cells from Human Umbilical Cords: Potential Cell Sources for Cardiovascular Research

doi: 10.3390/ijms22020978

Figure Lengend Snippet: Expression of collagen I-V by endothelial cells. Human umbilical cord-derived endothelial cells from veins (HUVEC) or arteries (HUAEC) were seeded at a density of 15,000 cells/well at day 0 and cultivated for 2, 4, and 7 days. While only traces of collagen were detected at day 2, the expression increased with increasing cultivation time, leading to large networks of collagen fibers at day 7 with cells from both sources. Fluorescent images were taken at 20-fold primary magnification. Scale bar represents 50 µm, and 25 µm in the magnified image sections in the upper right corner of each image. Squares with dashed lines indicate the image region that was magnified.

Article Snippet: Pooled HUVEC (from 3 to 6 individual donors, Lonza, Cologne, Germany) and HUAEC (from 250 individual donors, PeloBiotech GmbH, Planegg, Germany) were cultivated in EGM-2 Bullet Kit containing endothelial cell culture medium supplemented with 2% ( v / v ) fetal bovine serum, vascular endothelial growth factor, basic fibroblast growth factor, human epithelial growth factor, insulin-like growth factor-1, hydrocortisone, heparin, ascorbic acid, gentamycin, and amphotericin B (Lonza) in a standard humidified incubator at 37 °C with 5% ( v / v ) CO 2 .

Techniques: Expressing, Derivative Assay

Journal: International Journal of Molecular Sciences

Article Title: Venous and Arterial Endothelial Cells from Human Umbilical Cords: Potential Cell Sources for Cardiovascular Research

doi: 10.3390/ijms22020978

Figure Lengend Snippet: Expression of laminin by endothelial cells. Human umbilical cord-derived endothelial cells from veins (HUVEC) or arteries (HUAEC) were seeded at a density of 15,000 cells/well at day 0, and cultivated for 2, 4, and 7 days. While laminin was only intracellularly present at day 2, it was secreted into the extracellular space with increasing cultivation time, leading to a wide fleece-like network at day 7. Fluorescent images were taken at 20-fold primary magnification. Scale bar represents 50 µm, and 25 µm in the magnified image sections in the upper right corner of each image. Squares with dashed lines indicate the image region that was magnified.

Article Snippet: Pooled HUVEC (from 3 to 6 individual donors, Lonza, Cologne, Germany) and HUAEC (from 250 individual donors, PeloBiotech GmbH, Planegg, Germany) were cultivated in EGM-2 Bullet Kit containing endothelial cell culture medium supplemented with 2% ( v / v ) fetal bovine serum, vascular endothelial growth factor, basic fibroblast growth factor, human epithelial growth factor, insulin-like growth factor-1, hydrocortisone, heparin, ascorbic acid, gentamycin, and amphotericin B (Lonza) in a standard humidified incubator at 37 °C with 5% ( v / v ) CO 2 .

Techniques: Expressing, Derivative Assay