Generation of Human Pulmonary Microvascular Endothelial Cell Lines

Krump‐Konvalinkova, Vera; Bittinger, Fernando; Unger, Ronald E.; Peters, Kirsten; Lehr, Hans‐Anton; Kirkpatrick, Charles James

doi:10.1038/labinvest.3780385

Cited by 143 publications

(111 citation statements)

References 43 publications

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“…4), indicating the dominant role of ␤ 2 -integrin and ICAM-1 interaction in mediating cell adhesion. In a typical test, for example, the baseline expression of E-selectin was quite low for quiescent HPMECs, and no significant up-regulation was found with 24-h TNF-␣-simulation (data not shown), which is consistent with previous results (30). Regardless of this, the impact of other adhesion molecules on cell adhesion should be taken into account in the future physiological studies.…”

Section: Figure 7 Binding Curve Of Pmns Adhesion To Quiescent Mda-mbsupporting

confidence: 79%

“…Human metastatic breast adenocarcinoma cell line MDA-MB-231 (purchased from the Cell Culture Center of Union Medical University, Beijing, China) was cultured in L-15 medium supplemented with 1 mM L-glutamine, 100 units/ml penicillin, 10 mg/ml streptomycin, and 10% FBS. Human pulmonary microvascular endothelial cell line HPMEC ST1.6R (a kind gift from Dr. C. J. Kirkpatrick, Institute of Pathology, Johannes-Gutenberg University, Mainz, Germany) was cultured as previously described (30). For adhesion frequency measurement, confluent cells were trypsinized and washed twice with fresh medium.…”

Section: Methodsmentioning

confidence: 99%

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Determining β2-Integrin and Intercellular Adhesion Molecule 1 Binding Kinetics in Tumor Cell Adhesion to Leukocytes and Endothelial Cells by a Gas-driven Micropipette Assay

Tong

Wang

et al. 2011

Journal of Biological Chemistry

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Interactions between polymorphonuclear neutrophils (PMNs) and tumor cells have been reported to facilitate the adhesion and subsequent extravasation of tumor cells through the endothelium under blood flow, both of which are mediated by binding ␤ 2 -integrin to intercellular adhesion molecule 1 (ICAM-1). Here the adhesions between human WM9 metastatic melanoma cells, PMNs, and human pulmonary microvascular endothelial cells (HPMECs) were quantified by a gas-driven micropipette aspiration technique (GDMAT). Our data indicated that the cellular binding affinity of PMN-WM9 pair was 3.9-fold higher than that of the PMN-HPMEC pair. However, the effective binding affinities per molecular pair were comparable between the two cell pairs no matter whether WM9 cells or HPMECs were quiescent or cytokine-activated, indicating that the stronger adhesion between PMN-WM9 pair is mainly attrib- Tumor metastasis requires the detachment of malignant cells from primary tumor, invasion through blood/lymph vessels, transmigration across the endothelium, and finally, adhesion to the host cells (1). The adhesion and subsequent extravasation of tumor cells through the vascular endothelium are critical steps in this complex cascade (2). Malignant melanoma is the most deadly skin cancer with high metastatic potential (3). Although the binding of such adhesion molecules as P-selectin (4, 5), Lu-ECAM-1 (6), or integrin ␣ 4 ␤ 1 (VLA-4) (7) to their ligands has been found to support the adhesion of melanoma tumor cells (TCs) 3 with endothelial cells (ECs), the expression of the molecules seems insufficient to support the direct adhesion between TCs and ECs in blood flow (8). Rather, the presence of polymorphonuclear neutrophils (PMNs) potentially enhances TCs adhesion to the ECs and induces the subsequent extravasation under flow conditions (9 -11). A "two-step adhesion" hypothesis was introduced that involves initial PMN tethering on ECs followed by subsequent TCs being captured by tethered PMNs (10). Another line of in vitro evidence has indicated independently that TCs are able to aggregate with PMNs in a shear flow (12), which proposes an alternative hypothesis that TCs interact first with PMNs to form aggregates in a near-wall region of vascular endothelium prior to binding to EC through PMNs. Thus, it is physiologically important to elucidate how TCs, PMNs, and/or ECs interact with each other and what their underlying molecular mechanisms are.Regulations of PMN-EC or PMN-TC adhesion have been extensively studied at the cellular and molecular levels. Tethering and rolling of leukocytes on the endothelium are initiated by selectins and stabilized by ␤ 2 -integrins, which is followed by PECAM-1-induced transmigration through the extracellular matrix (13,14). Tumor cells are found to interact with PMNs under shear flow in a cone-plate viscometer (12,(15)(16)(17). Evidently, the interactions between ␤ 2 -integrin (LFA-1 or Mac-1) expressed on PMNs and intercellular adhesion molecule 1 (ICAM-1) expressed on TCs or ECs dominate both PMN-EC a...

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Section: Figure 7 Binding Curve Of Pmns Adhesion To Quiescent Mda-mbsupporting

confidence: 79%

Section: Methodsmentioning

confidence: 99%

Determining β2-Integrin and Intercellular Adhesion Molecule 1 Binding Kinetics in Tumor Cell Adhesion to Leukocytes and Endothelial Cells by a Gas-driven Micropipette Assay

Tong

Wang

et al. 2011

Journal of Biological Chemistry

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“…Primary human lung microvascular endothelial cells (HMVEC-L) were obtained from Lonza. Immortalized lung microvascular endothelial cells (IMVEC-L) have been previously described and were a kind gift from Dr. Aron Fisher at the Institute for Environmental Medicine (17). HMVEC-L or IMVEC-L were seeded in six-well plates and grown to confluence.…”

Section: Preparation Of Rbcsmentioning

confidence: 99%

The receptor for advanced glycation end products mediates lung endothelial activation by RBCs

Mangalmurti

Friedman

Wang

et al. 2013

American Journal of Physiology-Lung Cellular and Molecular Phys

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The receptor for advanced glycation end products (RAGE) is a multiligand pattern recognition receptor implicated in multiple disease states. Although RAGE is expressed on systemic vascular endothelium, the expression and function of RAGE on lung endothelium has not been studied. Utilizing in vitro (human) and in vivo (mouse) models, we established the presence of RAGE on lung endothelium. Because RAGE ligands can induce the expression of RAGE and stored red blood cells express the RAGE ligand N ε -carboxymethyl lysine, we investigated whether red blood cell (RBC) transfusion would augment RAGE expression on endothelium utilizing a syngeneic model of RBC transfusion. RBC transfusion not only increased lung endothelial RAGE expression but enhanced lung inflammation and endothelial activation, since lung high mobility group box 1 and vascular cell adhesion molecule 1 expression was elevated following transfusion. These effects were mediated by RAGE, since endothelial activation was absent in RBCtransfused RAGE knockout mice. Thus, RAGE is inducibly expressed on lung endothelium, and one functional consequence of RBC transfusion is increased RAGE expression and endothelial activation. receptor for advanced glycation end products; red blood cell transfusion; endothelial cell; lung inflammation; red blood cells THE RECEPTOR FOR ADVANCED glycation end products (RAGE) is widely expressed on systemic endothelium and functions as a pattern recognition receptor for multiple ligands, including advanced glycation end products (AGEs), high mobility group box 1 (HMGB1), calgranulins (s100A12 and s100B), amyloid ␤-proteins, Mac-1, phosphotidylserine, and lipopolysaccharide (10,24,26,35). AGEs, a heterogenous group of adducts formed during pathological states, hyperglycemia, and periods of increased oxidative stress, lead to increased generation of reactive oxygen species (ROS) and inflammatory cytokines following ligation of RAGE expressed on vascular endothelium (25,31,32). We have previously shown that stored red blood cells (RBCs) that express the RAGE ligand N ε -carboxymethyl lysine (N ε -CML) trigger lung endothelial ROS generation that was attenuated by soluble receptor for advanced glycation end products (sRAGE), the extracellular ligandbinding domain of the receptor that acts as a decoy by binding RAGE ligands (19). This finding supports the hypothesis that RAGE ligands on the surface of stored RBCs promote activation of lung endothelium, thus contributing to the pathogenesis of lung injury in susceptible transfusion recipients.Because RAGE is abundantly expressed on alveolar epithelium, particularly type I epithelial cells, and is utilized as a specific marker of alveolar epithelial injury, the existence of RAGE on pulmonary endothelium has come into question (9,28,30). Indeed, prior studies failed to detect RAGE in lung endothelium (9, 28). However, the complexities of RAGE biology render it a difficult molecule to study, since RAGE is known to exist in several different isoforms, subject to cleavage by proteases, ra...

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“…Immortalized human pulmonary microvascular endothelial cells (HPMVEC clone ST1.6R) were generated as described previously (Krump-Konvalinkova et al, 2001) and cultured in Medium-199 supplemented with 15% FBS, glutamax, antibiotics, and endothelial cell growth supplement. Isolation, characterization, and propagation of mouse pulmonary microvascular endothelial cells (MPMVEC) from wild-type (C57BL/6) and gp91 phox gene-targeted mice have been previously described (Milovanova et al, 2006).…”

Section: Cell Culturementioning

confidence: 99%

Superoxide Flux in Endothelial Cells via the Chloride Channel-3 Mediates Intracellular Signaling

Hawkins

Madesh

Kirkpatrick

et al. 2007

MBoC

169

150

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.؊ resulted in rapid and concentration-dependent transient HE oxidation that was followed by a progressive and nonreversible increase in nuclear HE fluorescence. These fluorescence changes were inhibited by superoxide dismutase (SOD), the anion channel blocker DIDS, and selective silencing of the chloride channel-3 (ClC-3) by treatment with siRNA. Extracellular O 2 .؊ triggered Ca 2؉ release in turn triggered mitochondrial membrane potential alterations that were followed by mitochondrial O 2 .؊ production and cellular apoptosis. These "signaling" effects of O 2 .؊ were prevented by DIDS treatment, by depletion of intracellular Ca 2؉ stores with thapsigargin and by chelation of intracellular Ca 2؉ . This study demonstrates that O 2 .؊ flux across the endothelial cell plasma membrane occurs through ClC-3 channels and induces intracellular Ca 2؉ release, which activates mitochondrial O 2 .؊ generation.

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Generation of Human Pulmonary Microvascular Endothelial Cell Lines

Cited by 143 publications

References 43 publications

Determining β2-Integrin and Intercellular Adhesion Molecule 1 Binding Kinetics in Tumor Cell Adhesion to Leukocytes and Endothelial Cells by a Gas-driven Micropipette Assay

Determining β2-Integrin and Intercellular Adhesion Molecule 1 Binding Kinetics in Tumor Cell Adhesion to Leukocytes and Endothelial Cells by a Gas-driven Micropipette Assay

The receptor for advanced glycation end products mediates lung endothelial activation by RBCs

Superoxide Flux in Endothelial Cells via the Chloride Channel-3 Mediates Intracellular Signaling

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