Hi,
We recently set up an in vivo Treg suppression assay. We have 3 groups:
1. Transfer of 4 x 105 naive T cells (CD4+CD25-CD45RBhi) into Rag1-null hosts
2. Transfer of 4 x 105 naive T cells (CD4+CD25-CD45RBhi) in combination with 2 x 105 WT Tregs into Rag1-null hosts
3. Transfer of 4 x 105 naive T cells (CD4+CD25-CD45RBhi) in combination with 2 x 105 KO Tregs into Rag1-null hosts
Mice in the from the control and KO Treg groups lost a significant amount of weight (between 30-40% at day 23 following intravenous injection). This led us to think that the KO Tregs could not suppress the development of colitis (since this is the Fiona Powrie model).
However, while moderate colitis was observed in the control group, colitis was absent from the WT and KO groups. My question is why the mice from the KO group got so sick? We showed a significant amount of weight loss as well as colon shortening, which are both hallmark signs of colitis.
To note, we only used H&E staining to look for colitis. Is it possible that this might not pick up the severity (even though it is an accepted way of monitoring these mice)? Should I maybe do IHC with markers of inflammation (Ly6 molecules for granulocytes? CD31 for blood vessel dilation?). Also, could the route of T cell transfer affect the results? I injected the mice by tail vein. Should I try with IP? I can't seem to find a concensus on which injection is better?
Thanks for any help!
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