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D. Bauer, S. Wasmuth, J. Li, H. Li, K.–P. Steuhl, A. Heiligenhaus; Improvement of Herpetic Stromal Keratitis by Amniotic Membrane Transplantation: Study on the Influence on Lymphocytes . Invest. Ophthalmol. Vis. Sci. 2005;46(13):2788.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose:Transplantation of amniotic membrane (AMT) induces rapid reepithelization, reduction of stromal inflammation and ulceration in murine eyes with immunopathologic herpetic stromal keratitis (HSK). The study was performed to determine the role of T–lymphocytes in the process. Methods:BALB/c mice were chosen that had developed ulcerating HSK on day 14 after corneal HSV–1 infection. One group of mice was treated with tarsorrhapy, and human amniotic membrane were used as a patch in another group. Corneas were collected 48h post transplantation, and amounts of corneal Th1/Th2 cytokines, CRG–2 (IP–10), and IL–12 were determined. To study the direct effect of amniotic membranes on lymphocytes, splenocytes or draining lymph node (DLN) cells were co–cultured with amniotic membrane: the proliferative response to HSV–1–antigen or ConA was investigated by 3H–thymidine uptake assay; surface molecules of cells (CD25, CD69, MHC II) were measured by flow cytometry; cytokines (TH1/Th2) were measured by ELISA; MTT conversion of cells was strudied; signs for apoptosis were detected by Hoechst staining (33324), gel electrophoresis of DNA, and by Annexin V–PE staining (BD). Transwell inserts were used to determine the presence of cell modifying soluble factors in the AM. Results:AMT leads to rapid decrease of T–lymphocyte numbers in the mouse corneas with HSK. After AMT, the Th1 and Th2 cytokines, IL–12 and CRG–2 expression in the corneas were decreased. Splenocytes co–cultured with AM showed decreased antigen specific and unspecific 3H–thymidine uptake, decreased amounts of cytokines (IFN–g and IL–2) and reduction of surface molecules CD25, CD69 and MHC II. Cells were negative for MTT conversion after co–cultivation with AM and showed typical signs for apoptosis. Transwell co–cultivation of lymphocytes and AM revealed apoptosis of lymphotes. Conclusions:The data suggest that effector and helper functions of T–lymphocytes are influenced by amniotic membrane. The effect is at least in part due to soluble factors induced apoptosis of lymphocytes.
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