Influence of combined treatment of low dose rapamycin and cyclosporin A on corneal allograft survival.
OBJECTIVE
To analyze the immune modulatory effect of low-dose systemic treatment with rapamycin (Rapa) alone or in combination with cyclosporin A (CsA) in a high-responder corneal allograft model.


METHODS
A total of 80 C57BL/6 mice received corneal grafts from BALB/c donors. Recipients were treated with either CsA 3 mg/kg/day or Rapa 0.5 mg/kg/day monotherapy or received combined treatment. Immunomodulatory treatment was started on the day of surgery, and continued for 14 days. The frequency of CD4(+)CD25(+)Foxp3(+) T regulatory cells (Treg) in secondary lymphoid organs was measured by flow cytometry. Development of IFN-gamma producing alloreactive T cells was estimated by Elispot. In addition, corneal samples were subjected to real-time RT-PCR analysis for cytokine transcription.


RESULTS
Monotherapy with Rapa significantly delayed allograft rejection (13.4 +/- 1.34 days, p = 0.03). However, the combination of both, low-dose Rapa and CsA prolonged corneal allograft survival at a significantly higher level (MST = 17.1 +/- 1.37 days, p = 0.0001) than in the control group (MST = 11.2 +/- 1.91 days). Rapa monotherapy increased the frequency of CD4(+)CD25(+)Foxp3(+)Treg in draining lymph nodes, whereas addition of CsA reduced Tregs. Monotherapy with Rapa as well as combined treatment prevented development of IFN-gamma producing alloreactive T cells in spleen. Combined treatment resulted in down-regulation of intragraft CD3, IL-2, IFN-gamma and IL-10 transcription (p = 0.028, p = 0.027, p = 0.028 and p = 0.027 respectively).


CONCLUSIONS
Combined treatment with low-dose CsA and Rapa resulted in superior graft survival, and effectively modulated mRNA expression of inflammation and infiltration markers.
Stanojlovic
Svetlana
S
Department of Ophthalmology, Charité University Medicine, 10117, Berlin, Germany. svetlsta@scnet.rs
Schlickeiser
Stephan
S
Appelt
Christine
C
Vogt
Katrin
K
Schmitt-Knosalla
Isabela
I
Haase
Stefanie
S
Ritter
Thomas
T
Sawitzki
Birgit
B
Pleyer
Uwe
U
D000818:Animals; D003315:Cornea / Q000187:drug effects* / Q000276:immunology / Q000473:pathology; D016572:Cyclosporine / Q000008:administration & dosage*; D016207:Cytokines / Q000235:genetics; D004195:Disease Models, Animal; D004359:Drug Therapy, Combination; D005434:Flow Cytometry; D006085:Graft Survival / Q000187:drug effects* / Q000276:immunology; D007166:Immunosuppressive Agents / Q000008:administration & dosage*; D007274:Injections, Intraperitoneal; D015948:Keratoplasty, Penetrating; D008221:Lymphoid Tissue / Q000276:immunology; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D020123:Sirolimus / Q000008:administration & dosage*; D050378:T-Lymphocytes, Regulatory / Q000276:immunology; D014184:Transplantation, Homologous
D016207:Cytokines; D007166:Immunosuppressive Agents; D012333:RNA, Messenger; D016572:Cyclosporine; D020123:Sirolimus