Human aortic endothelial cells (HAECs) were obtained commercially (GIBCO, C0065C or PromoCell, C-12271) and cultured according to the manufacturer’s standard protocol. HAECs were seeded at various density to meet the purposes of different studies, and maintained in EC medium containing growth supplements (EBM-2, Lonza, CC-3516, CC-4176 or PromoCell, C-22011) with 2% FBS. HAECs between passage 6 and 7 were used for experiments. For shear experiments, confluent HAECs in 100 mm dishes were exposed to steady laminar shear (LS, 15 dyn/cm2), or oscillatory shear (OS, ±5 dyn/cm2) conditions for 2 days using a cone-and-plate shear device as previously described [20 (link), 21 (link)]. For shear experiments with gain of function or loss of function components, the commercially available Ibidi pump system (Ibidi, Germany) was used to reduce reagent use, and set up according to manufacturer instructions. LS (15 dyn/cm2) or OS (±5 dyn/cm2) was applied for 2 days to 80% confluent HAECs cultured overnight on microchannel slides (µ-Slide I0.4 Luer, Ibidi, Germany) coated with 40 μg/ml collagen (Collagen Type I, BD Biosciences 35–4236).
Cc 4176
Manufactured by PromoCell
The CC-4176 is a laboratory equipment product. It is designed for conducting experimental and analytical tasks in a controlled laboratory environment. The core function of the CC-4176 is to provide a reliable and precise tool for researchers and scientists to carry out their work. No further details about its intended use or performance capabilities can be provided in an unbiased and factual manner.
Automatically generated - may contain errors
Lab products found in correlation
Pump system, by Ibidi (2 mentions)
Type 1 collagen, by BD (2 mentions)
C 12271, by PromoCell (2 mentions)
Cc 3516, by PromoCell (2 mentions)
C 22011, by PromoCell (2 mentions)
Slide 1 0.4 luer, by Ibidi (1 mentions)
μ slide i0.4 luer, by Ibidi (1 mentions)
Rnaimax transfection reagent, by Thermo Fisher Scientific (1 mentions)
Hcasmc, by PromoCell (1 mentions)
M csf, by Thermo Fisher Scientific (1 mentions)
Rnaimax, by Thermo Fisher Scientific (1 mentions)
Cc 3156, by Lonza (1 mentions)
3 protocols using cc 4176
1
Shear Stress Conditioning of Aortic Endothelial Cells
2
Human Aortic Endothelial Cell Shear Experiments
3
Silencing SASH1 in Human Aortic Endothelial Cells
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Human Aortic Endothelial Cells (HAECs) and Human Coronary Artery Smooth Muscle Cells (HCASMCs) were purchased from PromoCell. Cells were cultured in EGM2 medium (Lonza, CC-3156 & CC-4176) and smooth muscle cells growth medium (PromoCell, C-22162), respectively. All experiments were performed using cell passages 3 to 7. Human monocytes were isolated from human buffy coats (Etablissement Français du Sang, Rungis, France) and differentiated into macrophages as previously described 142 (link) . Macrophages were cultured for 6 days in RPMI 1640 medium with, 50 µg/ml gentamycin, 10% of fetal calf serum and 10ng/ml MCSF (Life Technologies, 11875-093 siRNA-mediated silencing of SASH1 Transfection of HAECs was performed using RNAiMAX (Life Technologies, 13778). Briefly, using RNAiMAX transfection reagent (Life technologies, 13778-100), two different siRNA targeting SASH1 (Ambion, S23572 and S23574) at a concentration of 20 nM were applied separately and compared to control siRNA (Ambion, 4390843). The medium was changed after 24h, total RNA and proteins were harvested 48h after transfection.
Efficiency of siRNA transfection and SASH1 knockdown was assessed by RT-qPCR and western blotting as described below.
Efficiency of siRNA transfection and SASH1 knockdown was assessed by RT-qPCR and western blotting as described below.
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