I'm trying to interface between two phases of a project - the first set of transcriptomic measurements were collected from cells plated on dishes that were not pre-treated. The next phase involves gene knockdown protocols that suggest pre-treating petri dishes with collagen. I want to make sure that the protocols implemented in the second phase of the project are biologically consistent/relevant to the experiments from the first phase.
It's reasonable to assume that pre-treating with collagen will have an impact - I'm curious as to how significant that impact may be...Does anyone have any experience testing the effect of pre-treating petri-dishes with collagen/gelatin on HUVEC biology (primarily, gene expression)?
Thank you!
Does it really matter? Culture in vitro in a plastic dish will massively affect gene expression/phenotype compared to that in vivo, whether you culture or gelatin, collagen or anything else. Worrying about whether you should use gelatin or collagen is kind of meaningless really.
Hi,
I am not an expert regarding Huvec cells, but I guess that especially the Collagen treatment wil have an influence on the cells behavior. Cells can use other matrix proteins to interact with collagen and espcially most cell types I have used produce a more robust ECM on Collagen. So I think it will be not completely comparable, however I have no hands on experinece in this cell type
Hello,
Coating with gelatin or collagen will have a varied effects on gene expression of HUVECS. Further, even changing the concentration of collagen will result in changes in gene expression (our own unpublished observation)
At a vascular biology meeting several years ago, I have seen a poster of a student in the group of Hellmuth Augustin. They showed a comparative microarray analyses of endothelial cells (either HUVEC or microvascular, I am not 100% sure) plated on either collagen or gelatin and differences in gene expression were quite substantial, with at least 30% of all genes being differently regulated.
As some friendly suggestions of researchers, coating with gelatin or collagen will have a varied effects on gene expression of HUVECs. However, HUVECs can not adhere on the plastic (polystyrene) dish. It is important that you use primary culture or 2ndary culture in the media with serum without excessive growth factors (EGF, b-FGF, hydrocortisone, Heparin etc.) and endotoxin.
The simple answer is yes, yes it will impact the gene expression. Instead of trying to rationalize or predict the impact culturing on collagen could have, your best bet is to repeat the first phase under the culture conditions of the second phase. Or do a bridging study to gauge the significance, it'll save time in the long run.
Does it really matter? Culture in vitro in a plastic dish will massively affect gene expression/phenotype compared to that in vivo, whether you culture or gelatin, collagen or anything else. Worrying about whether you should use gelatin or collagen is kind of meaningless really.
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