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Hlmec

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HLMECs are a type of primary human cell line derived from human lung microvascular endothelial cells. They are used for in vitro research and experimentation.

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Lab products found in correlation

Raw264, by American Type Culture Collection (3 mentions) Thp 1, by American Type Culture Collection (3 mentions) Hdmec, by Lonza (3 mentions) Hcaec, by Lonza (3 mentions) Human umbilical vein endothelial cells (huvec), by Lonza (3 mentions) Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (3 mentions) Egm 2mv, by Lonza (3 mentions) Rpmi 1640 medium, by Corning (2 mentions) Nucleofector kits for huvec, by Lonza (2 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions) Nucleofector device, by Lonza (1 mentions) Huvecs, by Lonza (1 mentions) Erk1 2, by Cell Signaling Technology (1 mentions) Jurkat, by American Type Culture Collection (1 mentions) Cos 7, by American Type Culture Collection (1 mentions) Actin sc 1616, by Santa Cruz Biotechnology (1 mentions) Nih3t3, by American Type Culture Collection (1 mentions) Phospho jnk, by Cell Signaling Technology (1 mentions) Phosphor p38, by Cell Signaling Technology (1 mentions) Phosphor erk1 2, by Cell Signaling Technology (1 mentions)

12 protocols using hlmec

1

Endothelial Cell Stimulation Assay

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All human primary vascular ECs (HAEC, HCAEC, HDMEC, HLMEC, and HUVEC) were purchased from Lonza Walkersville Inc., cultured in EGM or EGM2 medium according to the manufacturer instruction, and used for experiment in less than five passages. The human acute monocytic leukemia cell line THP-1 was obtained from American Type Culture Collection (ATCC) and maintained in RPMI 1640 medium (Corning) containing 10% FBS (Sigma-Aldrich). HeLa, A549, RAW264.7, and U937 were purchased from ATCC and cultured in Dulbecco's modified Eagle's medium supplemented with 10% FBS. Transient transfection of TRIM14 vector and siRNA into HUVECs was performed by electroporation using Nucleofector device (Lonza) and Nucleofector kits for HUVEC (Lonza) following the manufacturer’s instruction. After electroporation, cells were plated into 35 mm dishes and incubated for 24 h at 37°C, 5% CO2. Then, cells were treated with 10 ng/ml TNF-α or IL-1β for indicated times, and proteins from those cells were extracted and detected by western blot.
Huang X., Li Y., Li X., Fan D., Xin H.B, & Fu M. (2019). TRIM14 promotes endothelial activation via activating NF-κB signaling pathway. Journal of Molecular Cell Biology, 12(3), 176-189.
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2

Vascular Endothelial Cell Isolation and Culture

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All primary human vascular endothelial cells including HAEC, HCAEC, HDMEC, HLMEC and HUVEC were acquired from Lonza Walkersville Inc. (Walkersville, MD) and cultured in EGM or EGM2 medium according to the manufacturer’s instruction. In all the experiments, cells were used within five passages. The human acute monocytic leukemia cell line THP-1 was obtained from American Type Culture Collection (ATCC) and was grown in RPMI 1640 containing 10% fetal bovine serum. Other cell lines including CHO, Cos-7, NIH3T3, 293 T, HeLa, Jurkat, RAW264.7 and U937 were purchased from ATCC and cultured in DMEM supplemented with 10% fetal bovine serum. Human recombinant TNFα and LPS were purchased from Sigma (Saint Louis, MO). VCAM-1 (sc-13160), ICAM-1 (sc-1511-R), E-selectin (sc-14011), IKKα/β (sc-7607), and actin (sc-1616) antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, CA). Phospho-JNK, JNK, phospho-IKK, IKK, phosphor-ERK1/2, ERK1/2, phosphor-p38 and p38 antibodies were purchased from Cell Signaling Technology.
He H., Guo F., Li Y., Saaoud F., Kimmis B.D., Sandhu J., Fan M., Maulik D., Lessner S., Papasian C.J., Fan D., Jiang Z, & Fu M. (2016). Adiporedoxin suppresses endothelial activation via inhibiting MAPK and NF-κB signaling. Scientific Reports, 6, 38975.
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3

Culturing Human Lung Endothelial Cells

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Human lung microvascular endothelial cells (HLMEC; Lonza) were cultured as we described previously (6 (link)) and were grown to confluence in endothelial basic medium-2 (EBM-2; Lonza) supplemented with 10% fetal bovine serum (FBS), human recombinant epidermal growth factor, human recombinant insulin-like growth factor-1, human basic fibroblast growth factor, vascular endothelial growth factor, hydrocortisone, ascorbic acid, heparin, gentamicin, and amphotericin B. Endothelial cells (passages 6 −9) were serum-starved then treated with 10% lactated Ringers (LR) solution or 10% fibrinogen saline solution (final concentration in medium: 5 mg/ml). The fibrinogen used here was a highly purified preparation of fibrinogen derived from pooled human plasma and was produced by CSL Behring GmbH (RiaSTAP, Germany). For treatment of bleeding patients, the target blood concentrations of fibrinogen therapy are 1.5 to 2.0 mg/ml (10 (link)). In the present study, we used 5 mg/ml fibrinogen which is higher than the expected doses in vivo, but our previous results have shown that fibrinogen at 2.5, 5.0 and 10 mg/ml afforded similar protection to endothelial barrier (6 (link)).
Wu F., Chipman A., Dong J.F, & Kozar R.A. (2021). Fibrinogen activates PAK1/cofilin signaling pathway to protect endothelial barrier integrity. Shock (Augusta, Ga.), 55(5), 660-665.
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4

Culturing Human Lung Microvascular Endothelial Cells

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Human lung microvascular endothelial cells (HLMEC; Lonza, MD) were cultured as we described previously13 (link).
Wu F., Wang J.Y., Chao W., Sims C, & Kozar R.A. (2020). miR-19b targets pulmonary endothelial syndecan-1 following hemorrhagic shock. Scientific Reports, 10, 15811.
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5

Endothelial Cell Stimulation by TNFα

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All human primary vascular endothelial cells (HAEC, HCAEC, HDMEC, HLMEC, and HUVEC) were purchased from Lonza Walkersville Inc., cultured in EGM or EGM2 medium according to the manufacturer instruction, and used for experiment in less than five passages. The human acute monocytic leukemia cell line THP-1 was obtained from American Type Culture Collection (ATCC) and maintained in RPMI 1640 medium (Corning) containing 10% FBS (Sigma-Aldrich). HeLa, A549, RAW264.7, and U937 were purchased from ATCC and cultured in DMEM supplemented with 10% FBS. Transient transfection of TRIM65 vector and siRNA into HUVECs was performed by electroporation using Nucleofactor device (Lonza) and Nucleofector kits for HUVEC (Lonza) following the manufacturer’s instruction. After electroporation, cells were plated into 35 mm dishes and incubated for 24 h at 37°C, 5% CO2. Then, cells were treated with 10 ng/ml TNFα for indicated times and proteins from those cells were extracted and detected by western blotting.
Li Y., Huang X., Guo F., Lei T., Li S., Monaghan-Nichols P., Jiang Z., Xin H.B, & Fu M. (2019). TRIM65 E3 ligase targets VCAM-1 degradation to limit LPS-induced lung inflammation. Journal of Molecular Cell Biology, 12(3), 190-201.
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6

Vitamin K3 modulates SARS-CoV-2 protein effects

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HLMECs were obtained from Lonza Bioscience (Houston, TX, USA) and were cultured in EGMTM −2 MV Microvascular Endothelial Cell Growth Medium (Lonza Bioscience). HLMECs were pre-treated with various concentrations of vitamin K3 for 1 h, followed by the exposure of 1 μg/mL SARS-CoV-2 N protein for 8 h. THP-1 cells were cultured in RPMI1640 medium in a humidified incubator at 37 °C with 5% CO2. THP-1 cells were pre-treated with Vitamin K3 for 1 h and then exposed to 1 μg/mL SARS-CoV-2 E protein for 4 h.
Zhan Y., Zha D., Lin H., Mao X., Yang L., Huang H., He Z., Zhou S., Xu F., Qian Y, & Liu Y. (2023). Protective Role of Vitamin K3 on SARS-CoV-2 Structural Protein-Induced Inflammation and Cell Death. Pharmaceuticals, 16(8), 1101.
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7

Evaluation of 6,8-Diprenylgenistein in Tumor and Endothelial Cells

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SCCVII (mouse squamous cell carcinoma) cells were obtained from Dr. Han-Sin Jeong (Samsung Medical Center, Seoul, Korea) and maintained in RPMI-1640 medium (HyClone; GE Healthcare Life Sciences, Logan, UT, USA) added with 10% fetal bovine serum (FBS; HyClone) in a 5% CO2 humidified incubator at 37 °C. HLMECs (Lonza, Basel, Switzerland) were maintained in EGM-2 MV bullet kit medium (Lonza) containing 20% FBS in a 5% CO2 humidified incubator at 37 °C.
6,8-Diprenylgenistein (6,8-DG, purity more than 98%) was purchased from ChemFaces (cat no. CFN97935, Lot No. CFS201701, Wuhan, Hubei, China). A total of 10 mM stock solution (in DMSO) of 6,8-DG was prepared and protected from light at 4 °C and then diluted as required concentrations in cell culture medium or PBS.
Bae M.G., Hwang-Bo J., Lee D.Y., Lee Y.H, & Chung I.S. (2021). Effects of 6,8-Diprenylgenistein on VEGF-A-Induced Lymphangiogenesis and Lymph Node Metastasis in an Oral Cancer Sentinel Lymph Node Animal Model. International Journal of Molecular Sciences, 22(2), 770.
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8

Cell Culture Protocols for SARS-CoV-2 Research

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African green monkey kidney Vero E6 cell line (Istituto Zooprofilattico Sperimentale della Lombardia e dell’Emilia Romagna, Brescia, Italy) were cultured in Dulbecco’s Modified Eagle Medium (DMEM; Gibco, Thermo Fisher Scientific, Waltham, MA, USA) supplemented with 10% Fetal Bovine Serum (FBS; Gibco, Thermo Fisher Scientific). HL-mECs (Lonza Clonetics, Walkersville, MD, USA) were maintained in EGM-2 MV (Lonza, Basel, Switzerland) containing 10% FBS. A549 ACE2-positive (A549 ACE2+) cells, a kind gift from Dr. Stephen J Elledge (Harvard Medical School, Boston, MA, USA), were cultured in RPMI (Gibco, Thermo Fisher Scientific) supplemented with 10% FBS (Gibco, Thermo Fisher Scientific).
Bugatti A., Filippini F., Bardelli M., Zani A., Chiodelli P., Messali S., Caruso A, & Caccuri F. (2022). SARS-CoV-2 Infects Human ACE2-Negative Endothelial Cells through an αvβ3 Integrin-Mediated Endocytosis Even in the Presence of Vaccine-Elicited Neutralizing Antibodies. Viruses, 14(4), 705.
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9

Culturing Microvascular Endothelial Cells

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Human dermal microvascular endothelial cells (HMECs) were obtained from the Centers for Disease Control and Prevention (Atlanta, GA). HMECs were cultured in MCDB131 medium (Caisson’s Laboratories) supplemented with Fetal Bovine Serum (FBS) (10% v/v; Aleken Biologicals), epidermal growth factor (10 ng/mL, Thermo Fisher Scientific, Waltham, MA, USA), L-glutamine (10 mM, Thermo Fisher Scientific), and hydrocortisone (1 µg/mL, Sigma) [22 (link)]. Human cerebral microvascular endothelial cells (HCECs) were kindly provided by R. K. Yu and S. S. Dasgupta, Institute of Molecular Medicine and Genetics, Medical College of Georgia, Augusta, GA. These immortalized cell lines, which display typical morphological, phenotypic, and functional characteristics of microvascular endothelium, were grown in culture as recommended [23 (link),24 (link)]. Primary human lung microvascular ECs (HLMECs) were purchased from Lonza and maintained in culture according to the manufacturer’s instructions. All cell cultures were incubated and maintained at 37 °C in an incubator with 5% CO2.
Sahni A., Narra H.P., Patel J, & Sahni S.K. (2018). MicroRNA-Regulated Rickettsial Invasion into Host Endothelium via Fibroblast Growth Factor 2 and Its Receptor FGFR1. Cells, 7(12), 240.
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10

Culturing Vero E6 and HL-mECs

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African green monkey kidney Vero E6 cell line was obtained from Istituto Zooprofilattico Sperimentale della Lombardia e dell’Emilia Romagna (Brescia, Italy) and maintained in Dulbecco’s Modified Eagle Medium (DMEM; Gibco, Thermo Fisher Scientific, Waltham, MA, USA) supplemented with 10% fetal bovine serum (FBS; Gibco, Thermo Fisher Scientific). HL-mECs were purchased from Lonza Clonetics (Walkersville, MD, USA) and cultured in EGM-2 MV (Lonza, Basel, Switzerland) containing 10% FBS.
Bugatti A., Filippini F., Messali S., Giovanetti M., Ravelli C., Zani A., Ciccozzi M., Caruso A, & Caccuri F. (2023). The D405N Mutation in the Spike Protein of SARS-CoV-2 Omicron BA.5 Inhibits Spike/Integrins Interaction and Viral Infection of Human Lung Microvascular Endothelial Cells. Viruses, 15(2), 332.
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