I am interested to know why C6 is the famous cell line to be chosen to co-culture with bEnd.3 (endothelial cells).
As I am not an expert in BBB-models I can only speculate:
1. it could be that using a rat brain tumor cell line (glioma C6) was much easier to maintain, more reproducible than any (fresh?) preparation of inhomogeneous astrocytes.
2. it may depend on the original question, investigators wanted to address: for example, delivery of anti-cancer drugs through the blood brain barrier may make it more reasonable to use a brain tumor cell (although I doubt that many rat tumor cell lines really reflect human pathology).
3. I could think of many pharmacological questions where I expect your proposal of replacing the C6 glioma cells by any other cells could be good, but then I could ask why using bEnd.3 endothelioma cells in the first place (and not brain derived non-tumorigenic, "normal" endothelial cells - but that preparation should also not be too easy to prepare, and difficult to reproduce). Maybe using just two cell lines may work to address many questions, although using more natural cells might be better for other, more relevant questions.
Hello Robert. Thank you for your answer. However to answer your question on why bEnd.3 is chosen, it is largely due to personal interest. It is then supported by the few articles especially by Brown et al. 2007 which claimed that bEnd.3 has rapid growth, maintains BBB characteristics over repeated passages, forms the functional barriers, and amenable to molecular interventions.
Hello Nurul, I used bEnd.3 cells a lot, but not for the BBB model, just as confluent monolayers for rolling and adhesion experiments with leukocytes or tumor cells as a potential step in tumor metastasis, especially concerning the integrin VLA4, its counterreceptor VCAM-1 and the chemokine SDF-1; later I brought my model into a biophysics lab in Germany to investigate especially this mechanism at the so-called single-molecule level with atomic force microscopy (AFM). Unfortunately, after my independent project became successful, it was overtaken by the guest lab and others were promoted with it.
Concerning bEnd.3 cell culture, I do not know how much they can change from early passages with respect to maintaining the BBB, but they may not always grow as fast as expected. Indeed, in the biophysics lab, although it had a small and modern cell culture area, the lab was almost unable to keep these cells alive (as an unpaid guest scientist, I worked far away and had those cells maintained there). I lost probably several years, just because the lab lost the cells, again and again and again. Even the many frozen stocks I made were used without refreezing new aliquots. After three years I got some cells from another lab, but after loosing maybe 5 early stocks I only bought them, but again, the lab was unable to maintain those cells without me growing them.
I just want to stress, the cells can grow not as fast as expected, but they grow to confluence; I think other labs had trouble, too, to keep all the characteristics after several passages, but I usually had no problems within an experiment, except modulating the density of my receptor of interest, which in single-molecule experiments was just too high after regular activation of the cells, but I developed a reproducible way to use these cells even in my AFM experiments.
Good luck with your experiments!
Hi Robert. Thank you for sharing your experience on bEnd.3. It is good to know the worst case scenario that could happen to an experiment before actually initiating the experiment. I will keep your story in mind. Thanks!
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