I am doing an in-vitro atherosclerotic study in my lab using HUVECs. I am using a commercial ox-LDL from Thermo Fischer (L34357) to induce the expression of inflammatory biomarkers such as ICAM-1, VCAM-1 and LOX-1. However, none of the concentrations I used, ranging from 50 to 200 ug/mL was able to induce a significant increase in these biomarkers as compared to the negative control. Does anyone have any experience of using the ox-LDL from Thermo in HUVECs and if it has worked for you?
Hao, Y., Xiong, R. and Gong, X., 2021. Memantine, NMDA receptor antagonist, attenuates ox-LDL-induced inflammation and oxidative stress via activation of BDNF/TrkB signaling pathway in HUVECs. Inflammation, 44(2), pp.659-670.
To validate whether the ox-LDL works I would propose to use an adequate positive control (e.g. TNF-alpha which should induce amongst other cytokines its own expression or LPS or PMA) to test responsiveness of HUVEC. For ref. see Circ J. 2009 Sep;73(9):1746-52. doi: 10.1253/circj.cj-99-0225; Sci Rep. 2017 Jan 27;7:41612. doi: 10.1038/srep41612; and Infect Immun. 1997 Aug;65(8):3337-44. doi: 10.1128/iai.65.8.3337-3344.1997.
When this works you should change the LOT or provider of our stimulus/compound.
If this do not work, you should use a new HUVEC vial/batch.
I have not tried ox-LDL for inflammation, only for EC labeling. Sorry.
4h of TNF 5ng/ml is enough to see an upregulation of those markers at RNA and protein levels. I have used it as a positive control for my experiments and works great. I hope it helps.
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