Infection and Immunity The origin and function of the FAP + cels in infected and uninfected wounds (360G-Wellcome-082869_Z_07_Z)

The aim of the project is to define the origin of the FAP _ cells in the tumour stroma. The first question to be asked is whether they are composed of haematopoietic cells, mesenchymal cells or a mixture of both. If they are haematopoietic in origin they could represent a case of a haematopoietic to mesenchymal transition as has been proposed for the stellate cells in the liver which are c045 but come from c045 progenitors (Miyata E et al. 2008). Transdifferentiation has also been proposed to occur from mesenchymal stem cells (MSCs) (Uccelli A et al. 2008) and it could be the case that these contribute c045 cells to the stroma. To answer this question OVAGFP BAC transgenic and non-transgenic mice would be sublethally irradiated and then would be reconstituted with sorted haematopoietic stem cells from non-transgenic or transgenic mice, respectively, differing at the thy1.1 locus. These mice would then be injected with LL2 or 816 tumour cells and the resulting tumours analysed for the presence of CD45+ and CD4s EGFP+ cells in the tumour stroma. This would allow the contribution of both non HSCs and HSCs to the FAP compartment to be ascertained. - Along with this lineage analysis further characterisation of the FAP cells would also be carried out. FACS-sorted GFP+ stromal cells could be further analysed to examine their surface markers. Recently it has been suggested that FAP _ marks human MSCs from the bone marrow (Bae S et al. 2008). This study was limited by the fact that it used commercially obtained MSCs and so it is unsure how well it applies to in vivo but it would be worth looking for GFP+ cells in the bone marrow. If these were found, they could be analysed for expression of CD45, indicating HSCs, or for several other markers as CD45.CD73.CD9 < rCD105+ cells would indicate an MSC origin. Cells from the tumours would also be analysed. To do this tumours would be obtained and single cell isolates prepared this could allow the phenotypes of the CD45+ and c045 populations to be better defined by other markers. An example would be looking for alpha smooth muscle actin which would define 'reactive' myofibroblasts in the stroma on CD45- cells. Finally functional studies would be performed to see what these cells were doing. They could function as APCs despite being CD45 , as has been reported for the stellate cells in the liver (Miyata E at al. 2008; Winau F et al. 2007). It would also be worth checking if they were anti-inflammatory for example through production of heme-oxygenase 1, or whether they produce retinoic acid which has been shown to be involved in inducing regulatory T-cells (Xiao S et al. 2008). This latter possibility again ties in with the hepatic stellate cells in the liver which also store a large amount of vitamin A (Reuben A, 2002).