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REPEATED TREATMENTS WITH BONE MARROW-DERIVED ALTERNATIVELY ACTIVATED MACROPHAGES INHIBIT COLITIS AND DO NOT ELICIT FIBROSIS
G Leung, A Wang, M Fernando, V Phan, D McKayUniversity of Calgary, Calgary, AB
Aims: Alternatively activated macrophages (AAMs) are associated with T helper-2 cell responses and the resolution of inflammation. We showed that adoptive transfer of AAM derived from peritoneal macrophages protected mice from dinitrobenzene sulphonic acid(DNBS)-induced colitis, as a proof-of-principle study for the treatment of inflammatory bowel disease (IBD). However, given that AAMs are also linked to the pathogenesis of pulmonary fibrosis, we sought to determine whether they were associated with adverse effects (specifically fibrosis) when mice were given multiple AAM treatments. We also tested whether bone marrow (BM)-derived macrophages were effective in protecting mice from DNBS colitis, as haematopoietic stem cells are a more efficient method of deriving macrophages from both experimental and clinical perspectives.
Methods: Macrophages from male Balb/c mice were obtained by culturing BM cells for seven days with macrophage colony stimulating factor (20 ng/ml), and alternative activation was elicited by a 48 h treatment with interleukin (IL)-4 and IL-13 (both 20 ng/ml). Macrophages (1 million) were given by intraperitoneal injection 6 h prior to the intra-rectal instillation of 3 mg DNBS in 100 µl 50% ethanol to induce colitis. Three cycles of DNBSħAAMs were performed at three weekly intervals and 72 h after the final treatments, mice were necropsied, colitis assessed by standard measures and fibrosis in the colon, small bowel, lung and liver assessed by collagen measurement, Masson's trichrome stain and Q-PCR for collagen and fibronectin.
Results: AAM differentiation was confirmed by increased mRNA and protein expression of arginase 1 and Ym1, established AAM markers. BM-derived AAMs significantly protected mice from DNBS-induced colitis, and while the severity of the disease decreased with each round of DNBS (as gauged by body weight changes), on each occasion AAM-treated mice fared better and on necropsy had lower disease activity scores, longer colons and less evidence of histological derangement than DNBS- only treated mice. Quantification of collagen, Masson's trichrome stain and Q-PCR revealed no evidence of increased fibrosis in the liver, lung, small intestine, or colon of AAM-treated mice.
Conclusions: Mice receiving BM-AAMs are protected against DNBS-induced colitis, and this benefit was sustained for three cycles of colitis and was not accompanied by evidence of fibrosis. We speculate that adoptive transfer of AAMs could be a viable anti-colitic therapy, that can remain effective over multiple rounds of treatments and, in this experimental paradigm, is not associated with the deleterious side-effects of collagen deposition and fibrosis.