The role of apoptosis in the pathogenesis of canine keratoconjunctivitis sicca: the effect of topical Cyclosporin A therapy.
OBJECTIVE
The exact etiology of dry eye is unknown but is believed to be multifactorial. Apoptosis has been implicated in the pathogenesis of autoimmune diseases such as Sjögren's syndrome (SS). This study attempted to gain a better understanding of the role of apoptosis and its regulation in the patho-physiology of dry eye. The therapeutic effect of immunomodulatory agents such as cyclosporin A (CsA) in the treatment of dry eye, particularly its impact on the level of apoptosis in the target tissues, is also investigated.


METHODS
A colony of dogs with spontaneous chronic idiopathic keratoconjunctivitis sicca (KCS) was maintained. Nictitans lacrimal gland (NLG), an accessory lacrimal gland, and conjunctival biopsies of the KCS and normal dogs were obtained before and after 12 weeks of treatment with 0.2% topical CsA ophthalmic emulsion b.i.d. (Allergan, Inc., Irvine, CA, U.S.A.). Tissues were prepared for the terminal deoxynucleotidyl transferase-mediated dUTP-digoxigenin nick end labeling (TUNEL) apoptosis assay and immunohistochemical analysis for various apoptosis mediators.


RESULTS
The TUNEL assay demonstrated that (i) the normal NLG and conjunctival epithelial cells exhibited a limited level of apoptosis; (ii) in KCS dogs, lacrimal acinar and conjunctival epithelial cells underwent an increased apoptosis, whereas the lymphocytes had a significantly lower level of apoptosis compared to those of the normal dogs; (iii) after topical CsA, apoptosis was induced in the lymphocytes and suppressed in the acinar and conjunctival epithelial cells in KCS dogs. Immunohistochemistry revealed that p53, fas, and fasL, but not bcl-2 were highly expressed in the target tissues of KCS dogs. The immunoreactivity of p53 was significantly decreased, whereas the bcl-2 level was increased after CsA administration.


CONCLUSIONS
The induction of epithelial cell apoptosis and the suppression of lymphocytic apoptosis in the NLG and ocular-surface tissues, such as conjunctiva of KCS dogs, indicates the important role of this phenomenon in the etiology of dry eye. Topical CsA appears to facilitate lymphocytic apoptosis and suppress epithelial cell apoptosis in the KCS dog. The differential expression of various apoptotic mediators after topical treatment implicates CsA in facilitating the reestablishment of the normal apoptotic balance, suggesting additional mechanisms by which CsA is therapeutic for dry-eye syndrome.
Gao
J
J
Department of Biological Science, Allergan, Inc., Irvine, California, USA. stern_michael@allergan.com
Schwalb
T A
TA
Addeo
J V
JV
Ghosn
C R
CR
Stern
M E
ME
D000287:Administration, Topical; D000818:Animals; D017209:Apoptosis; D003228:Conjunctiva / Q000187:drug effects / Q000378:metabolism / Q000473:pathology*; D016572:Cyclosporine / Q000008:administration & dosage / Q000627:therapeutic use*; D004247:DNA / Q000032:analysis; D004283:Dog Diseases / Q000188:drug therapy / Q000209:etiology* / Q000378:metabolism / Q000473:pathology; D004285:Dogs; D053222:Fas Ligand Protein; D005260:Female; D007124:Immunoenzyme Techniques / Q000662:veterinary; D007166:Immunosuppressive Agents / Q000008:administration & dosage / Q000627:therapeutic use*; D020287:In Situ Nick-End Labeling / Q000662:veterinary; D007638:Keratoconjunctivitis Sicca / Q000188:drug therapy / Q000209:etiology / Q000378:metabolism / Q000662:veterinary*; D007765:Lacrimal Apparatus / Q000187:drug effects / Q000378:metabolism / Q000473:pathology*; D008297:Male; D008562:Membrane Glycoproteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D011897:Random Allocation; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D019014:fas Receptor / Q000378:metabolism
D053222:Fas Ligand Protein; D007166:Immunosuppressive Agents; D008562:Membrane Glycoproteins; D019253:Proto-Oncogene Proteins c-bcl-2; D016159:Tumor Suppressor Protein p53; D019014:fas Receptor; D016572:Cyclosporine; D004247:DNA