Primary HAoECs (PromoCell) were treated for 7 days with recombinant human TNF (R&D Systems). Relative expression of CX3CL1 was measured by real-time PCR (Taqman Gene Expression Assays; ThermoFisher) and in supernatant by ELISA (R&D Systems). Magnetic-bead purified memory CD4 T cells (Memory CD4 T cell isolation kit, Miltenyi Biotec) were stimulated with recombinant human IL-15 (247-ILB; R&D Systems) for 2 days. Supernatants were tested for CX3CL1 and TNF protein by ELISA (R&D Systems). Supernatants were then diluted 1:2 and cultured with HAoECs. After 7 days, culture supernatants were harvested and tested by ELISA for CX3CL1.
Primary haoecs
Manufactured by PromoCell
Sourced in Germany
Primary HAoECs are human aortic endothelial cells derived from the aorta. They provide a model system for studying endothelial cell biology and function.
Automatically generated - may contain errors
Lab products found in correlation
Recombinant human tnf, by R&D Systems (1 mentions)
Taqman gene expression assay, by Thermo Fisher Scientific (1 mentions)
247 ilb, by R&D Systems (1 mentions)
Memory cd4 t cell isolation kit, by Miltenyi Biotec (1 mentions)
Gentamicin, by Thermo Fisher Scientific (1 mentions)
Pump system, by Ibidi (1 mentions)
Click it edu alexa fluor 488 imaging kit, by Thermo Fisher Scientific (1 mentions)
Mircury lnatm microrna target site blockers, by Qiagen (1 mentions)
2 protocols using primary haoecs
1
Regulation of Endothelial CX3CL1 Expression
2
Shear Stress Modulation of Endothelial Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Primary MAoECs (passage 3; PELOBiotech GmbH, Planneg, Germany) and primary HAoECs (passages 2, 3; Promocell, Heidelberg, Germany) were cultured using endothelial cell complete growth medium (Promocell) containing gentamicin (0.05 mg ml−1; ThermoFisher).
To grow the cells under different shear stresses, MAoECs were cultured in collagen-coated perfusion chambers (µ-Slides VI0.4, ibidi GmbH, Martinsried, Germany) and exposed to high shear stress (10 dyne cm−2) or low shear stress (5 dyne cm−2) for 48 h generated by the perfusion with EC complete medium (ibidi Pump System, ibidi GmbH) containing 5-ethynyl-2’-deoxyuridine (EdU, 10 µM final conc., Click-iT® EdU Alexa Fluor® 488 Imaging Kit, Life Technologies) and TSBs (TSBs in vivo ready) or control LNAs (both 50 nM final conc., Exiqon).
MAoECs were transfected with antisense oligonucleotides to block the interaction between miR-103 and lncWDR59 (TSB; 50 nM, miRCURY LNATM microRNA Target Site Blockers; Exiqon) or scrambled controls. To inhibit lncWDR59 function, murine and human ECs were transfected with lncWDR59 GapmeRs (50 nM, LNATM GapmeRs; Exiqon) that strongly induce the degradation of the lncWDR59 transcript in the nucleus of the ECs.
To grow the cells under different shear stresses, MAoECs were cultured in collagen-coated perfusion chambers (µ-Slides VI0.4, ibidi GmbH, Martinsried, Germany) and exposed to high shear stress (10 dyne cm−2) or low shear stress (5 dyne cm−2) for 48 h generated by the perfusion with EC complete medium (ibidi Pump System, ibidi GmbH) containing 5-ethynyl-2’-deoxyuridine (EdU, 10 µM final conc., Click-iT® EdU Alexa Fluor® 488 Imaging Kit, Life Technologies) and TSBs (TSBs in vivo ready) or control LNAs (both 50 nM final conc., Exiqon).
MAoECs were transfected with antisense oligonucleotides to block the interaction between miR-103 and lncWDR59 (TSB; 50 nM, miRCURY LNATM microRNA Target Site Blockers; Exiqon) or scrambled controls. To inhibit lncWDR59 function, murine and human ECs were transfected with lncWDR59 GapmeRs (50 nM, LNATM GapmeRs; Exiqon) that strongly induce the degradation of the lncWDR59 transcript in the nucleus of the ECs.
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