June 2013
Volume 54, Issue 15
Free
ARVO Annual Meeting Abstract  |   June 2013
Mechanisms involved in conjunctival immunological tolerance breakdown by eye drop preservatives
Author Affiliations & Notes
  • Jeremías Galletti
    Immunology Laboratory, Institute of Experimental Medicine, National Academy of Medicine/CONICET, Buenos Aires, Argentina
    Ophthalmology, Hospital de Clínicas, University of Buenos Aires, Buenos Aires, Argentina
  • Laura Gabelloni
    Immunology Laboratory, Institute of Experimental Medicine, National Academy of Medicine/CONICET, Buenos Aires, Argentina
  • Florencia Sabbione
    Immunology Laboratory, Institute of Experimental Medicine, National Academy of Medicine/CONICET, Buenos Aires, Argentina
  • Pablo Chiaradia
    Ophthalmology, Hospital de Clínicas, University of Buenos Aires, Buenos Aires, Argentina
  • Mirta Giordano
    Immunology Laboratory, Institute of Experimental Medicine, National Academy of Medicine/CONICET, Buenos Aires, Argentina
  • Javier Casiraghi
    Ophthalmology, Hospital de Clínicas, University of Buenos Aires, Buenos Aires, Argentina
  • Footnotes
    Commercial Relationships Jeremías Galletti, None; Laura Gabelloni, None; Florencia Sabbione, None; Pablo Chiaradia, None; Mirta Giordano, None; Javier Casiraghi, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science June 2013, Vol.54, 2039. doi:
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    • Get Citation

      Jeremías Galletti, Laura Gabelloni, Florencia Sabbione, Pablo Chiaradia, Mirta Giordano, Javier Casiraghi; Mechanisms involved in conjunctival immunological tolerance breakdown by eye drop preservatives. Invest. Ophthalmol. Vis. Sci. 2013;54(15):2039.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract
 
Purpose
 

To evaluate the molecular and celular mechanisms involved in the disruptive immune effect of benzalkonium chloride (BAK) on the ocular surface.

 
Methods
 

A previously validated in vivo murine model was used (Mucosal Immunol. 2012 Jun 13. doi: 10.1038/mi.2012.44). In brief, Balb/c mice were given ovalbumin (OVA) alone or in combination with 0.01% BAK (B+O) daily for 5 days in both eyes, with and without nuclear factor κB (NFκB) pathway inhibitors: pyrrolidine dithiocarbamate (PDTC) and sulfasalazine (SSZ). On day 7, mice were immunized intraperitoneally with OVA in alum to evaluate the systemic response on day 14 as OVA-induced splenocyte proliferation by thymidine incorporation. To model in vitro the ocular surface epithelium, Pam212 cells were exposed for 15 min to different BAK concentrations (0.00001%-0.01%), then cultured for 96 h with syngeneic splenocytes and finally cell viability and expression of major histocompatibility complex class II (MHC-II) as activation marker were assayed.

 
Results
 

In the in vivo model, OVA instillation induced conjunctival tolerance (43±9% of control systemic response, p<0.05), in contrast to B+O (90±8%), which did not affect the subsequent cellular response. Addition of NFκB inhibitors restored tolerance in B+O mice (PDTC 39±3%, SSZ 54±8%, p<0.05) but did not significantly affect it in OVA mice (PDTC 31±5%, SSZ 69±5%, p<0.05). In the epithelial cultures, exposure to BAK decreased cell viability after 96 h but did not modify the MHC-II+ fraction (5% control, <8% BAK). Both γ interferon and splenocyte coculture increased MHC-II expression (15% and 19%, respectively, p<0.05). In BAK-exposed Pam212 cells, coculture with splenocytes increased even further the MHC-II+ fraction (24%-46% with increasing BAK concentrations, p<0.05). Epithelial pretreatment with PDTC or SSZ did not affect MHC-II expression in any case.

 
Conclusions
 

In the in vivo model, BAK-induced conjunctival tolerance breakdown could be reverted by the instillation of two known inhibitors of NFκB activation. Regarding the increased MHC II expression observed in conjunctival biopsies from preservative-exposed patients, BAK indirectly induced a similar change in cultured epithelial cells that involved their interaction with lymphocytes. These results suggest that NFκB modulation could have a therapeutic role in ocular surface immunology.

     
Keywords: 621 ocular irritants • 555 immunomodulation/immunoregulation • 553 immune tolerance/privilege  
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