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Thomas Ritter, Mikhail Nosov, Aideen Ryan, Oliver Treacy, Marese Cregg, Gerry Fahy, Mourice Morcos, Lisa O'Flynn; Bone Marrow Derived Dendritic Cells Prevent Corneal Allograft Rejection In The Rat. Invest. Ophthalmol. Vis. Sci. 2012;53(14):6188.
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© ARVO (1962-2015); The Authors (2016-present)
To study the role of in-vitro generated Bone Marrow derived Dendritic Cells (BMDC) after glucocorticoid treatment on corneal allograft survival in the rat.
BMDC were propagated from either Lewis (LEW) or Dark Agouti (DA) rat bone marrow precursors cells (1.5x106 cells/ml) in complete medium supplemented with rat GMCSF (5ng/µl) and IL-4 (5ng/µl). For glucocorticoid treatment of BMDCs, dexamethasone (Dexa) (10-6M) was added on d5 and d7 of a 10 day culture. BMDC and Dexa BMDC phenotype was characterised and analysed for expression of cell surface markers CD11b/c, MHC II, CD80, CD86 and His36 by flow cytometry. BMDC and Dexa BMDC antigen presenting cell function was examined in both antigen specific (Ovalbumin) and allo-antigen specific lymphocyte assays. Responder cells were analysed by FACS for proliferation and expression of lymphocyte activation markers CD25 or OX40. Moreover recall experiments were performed to study the mechanisms of immunomodulation in vivo. A fully allogeneic rat corneal transplantation model (DA to LEW) was used for in vivo studies. Day 10 syngeneic (LEW) alloantigen-pulsed or allogeneic (DA) Dexa BMDCs were harvested and 1x106cells/ml injected intravenously into LEW recipients 7 days prior to corneal transplant surgery. Graft survival, opacity and neovascularisation were monitored.
In vitro generated BMDCs have a semi-mature phenotype (moderate expression of MHC II, CD80 and CD86) and can be modified chemically to an immature phenotype with Dexa (low expression of MHC II, CD80 and CD86). BMDC and Dexa BMDC are capable of activating allogeneic lymphocytes but there is a reduced level of proliferation They can also efficiently present antigen and activate antigen specific T cells. In vivo untreated allogeneic control grafts were uniformly rejected (MST 13.8 ± 1.7d, n = 11). Injection of alloantigen-pulsed syngeneic BMDC had no effect on corneal allograft survival (MST 13 +/- 2.36d, n=6 and >30d n=1, spontaneous acceptance). In contrast, injection of allogeneic Dexa BMDC significantly prolonged corneal allograft survival (MST> 30d, n = 8). Opacity and neovascularisation scores were low in Dexa BMDC group compared to control groups.
Glucocorticoid treatment of allogeneic BMDCs is a potent way of inducing tolerogenic cells and represents a potential therapeutic approach for the prevention of corneal allograft rejection.
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