Hi! I don't work with fibroblasts usually but with macrophages and could use some advice. I want to do some transwell co-culture experiments with macrophages and fibroblasts (primary, isolated from rheumatoid synovial tissue).
Are there reliable phenotypic markers for inflammatory activation of fibroblasts I can use?
If there are any good publications to give me a start, that would also be appreciated.
Thanks
Hefin
A huge amount of work was done on synovial fibroblast activation by macrophage products in the 1980s and early 1990s by people like Steven Krane and Jean-Michel Dayer. Synovial fibroblasts are probably embryologically different from other fibroblasts, belonging to a 'skeletal' type that can take on features of intimal fibroblasts (a distinct subpopulation on the synovial surface) bone marrow nurse cells and even follicular dendritic cells. Prostaglandin and metalloproteinase production were poopular markers of activation. Immunologically VCAM-1 and DAF (CD55) may be more interesting. It depends a lot on what hypothesis you are testing.
A problem with rheumatoid tissue is that it is hugely heterogeneous and freshly isolated cells may be in all sorts of states and include all sorts of subpopulations which will shift with hours and days in culture.
A simple indicator of activation can be morphology, although it is not easy to quantify - activated cells develop ramifying processes.
There is a pretty large literature from 25 years ago but the terminology is often confused. If I knew more precisely what idea you wanted to test I might be able to be more specific.
Thanks for your answer Jonathan, I know my question is vague sorry. We're interested in whether changes in macrophage phenotype can affect the function and inflammation-promoting phenotype of synovial fibroblasts. In the macrophage, I have a panel of surface antigens that I can use to infer how inflammatory they are. I was wondering if I can use antigens on the fibroblast as an indicator of their propensity to induce and propagate inflammation.
Inducing and propagating inflammation is a wide brief. In the 80s the focus was on tissue damage and I think that may have been a bit of a red herring. If you are interested in fibroblasts supporting further white cell infiltration and the formation of lymphoid follicles you might want to look up papers written by our group, including Leigh-R and Abrahams-V around 1995-8 and some later reviews of mine. My RG page should give a list of publications. Richard Leigh's paper on cultured synovial fibrolblasts responding to cytokines is perhaps the most relevant - we were using immunohistochemical analysis of surface antigens. If I had to put my money on the easiest and most relevant it would be VCAM-1 - since that is crucial as a survival signal for immigrant lymphoid cells.
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