I've been trying to induce an inflammatory response using LPS (Sigma O127:B8 and O55:B5) using the immortalized human myometrial cells (TERT-HM) in our lab but have failed to get a significant IL6 or IL8 expression in all my experiments. Multiple studies have shown significant cytokine expression release with just 100ng/ml LPS concentration. However, I've gone as high as 5ug/ml LPS for 24hrs and still fail to obtain any significant expression. Can anyone who has previously worked with LPS suggest any tips how best to use LPS?
I've treated my cells with TNF-alpha and I obtain a reasonable inflammatory response. So I can confirm that my cell line seems to work fine. I am just not able to succeed to induce a similar response using LPS. I keep my LPS stock in -20 degree freezer and don't use freshly made media in every treatment.
I would appreciate any suggestion as I am out of options as to why I can not obtain any cytokine release in any dose-dependent or time-course treatments.
I'm using qPCR to analyze the mRNA expression.
Dear Dr Magdaleena Mbadhi
In human myometrial cells, the expression of TLR4 has been reported, although its levels may be relatively low compared to other cell types involved in the immune response like macrophages. How about trying the TLR4 expressions compared with the same exposure of TLR4?
Kind regards
AB Bayazid
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A.B Bayazid Thank you for your suggestion. Do you mean checking the expression of TLR4 instead of IL8 and IL6 with LPS treatments? Or treating the cells with a TLR4 compound instead? The aim of the project is to induce an infectious inflammatory response.
Dear Dr. Magdaleena Mbadhi
The less amount of TLR4 presence may cause problems. Hence, you can try different inflammatory stimuli (TNF-a, IFN-y, IL-1B, IL-6, etc.), that I have used.
Kind regards
AB Bayazid
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