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Originally published In Press as doi:10.1167/iovs.08-3041 on February 28, 2009
 (Investigative Ophthalmology and Visual Science. 2009;50:3188-3198.)
© 2009 by The Association for Research in Vision and Ophthalmology, Inc.
doi:10.1167/iovs.08-3041
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Amniotic Membrane Transplantation Induces Apoptosis in T Lymphocytes in Murine Corneas with Experimental Herpetic Stromal Keratitis

Dirk Bauer,1 Susanne Wasmuth,1 Maren Hennig,1 Hanna Baehler,1 Klaus-Peter Steuhl,2 and Arnd Heiligenhaus1

1From the Department of Ophthalmology, Ophtha-Lab, St. Franziskus Hospital, Muenster, Germany; and the 2Department of Ophthalmology, University of Duisburg-Essen, Essen, Germany.

PURPOSE. To investigate the effect of human amniotic membrane transplantation (AMT) on T-cell immune response in murine corneas with herpetic stromal keratitis (HSK).

METHODS. Herpes simplex virus (HSV)-1–infected BALB/c mice with necrotizing HSK were treated with AMT. CD3+ cell apoptosis was determined in treated corneas and in vitro by flow cytometric analysis using the annexin V/7-AAD system. The effect of interleukin (IL)-2, cyclosporine, rapamycin, or Fas on T-cell survival was measured. Activation phenotype was measured by 3H-thymidine uptake and flow cytometry (CD25, CD69, major histocompatibility complex class II). Cytokine/chemokine secretion from amniotic membrane (AM)-treated corneas or draining lymph node cells was measured. The immune-modulating capacity of long-term AMT treatment and adoptive transfer of AM-treated splenocytes was tested.

RESULTS. After AMT, HSK and corneal inflammatory cell infiltration improved, and T-lymphocyte apoptosis occurred. T-cell apoptosis was also induced in vitro, independently of rIL-2, cyclosporine, rapamycin, or Fas. AMT-treated corneas and cultured lymphocytes had reduced IL-2, IL-10, IL-12, CRG-2, and CCL-2 content. Long-term AMT treatment decreased the proliferative response and type 1 helper T-cell cytokine level in draining lymph node cells. The improvement in HSK did not persist. Delayed-type hypersensitivity or HSV-1–specific cytotoxicity was not altered

CONCLUSIONS. The results suggest that murine HSK improves after AMT through reduced local T-helper cell immune responses by inducing apoptosis in T lymphocytes, independently of passive apoptosis or activation-induced cell death. AM also reduces local T-helper cytokine and chemokine levels but does not result in immune deviation. Immunologic memory against HSV-1 is not affected by AMT, and long-term protection or tolerance is not induced.






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