Ex vivo adenovirus mediated gene transfection of human conjunctival epithelium

J. Shen, N. Taylor, L. Duncan, I. Kovesdi, J. T. Bruder, J. V. Forrester, A. D. Dick

Research output: Contribution to journalArticlepeer-review

Abstract

Aim - To investigate the efficacy of "ex vivo" adenoviral vector mediated gene transfection of human conjunctival epithelial cell as a possible route for gene therapy for the distribution of anti-inflammatory agents for the potential treatment of immune mediated ocular inflammatory disorders. Methods - Human conjunctival cells (HGs) were cultured with various concentrations of recombinant adenoviral vectors carrying a reporter gene LacZ, GFP, or an immunomodulating cytokine vIL-10. vIL-10 in culture supernatant was detected by sandwich ELISA and biological activity was assessed by suppression of ConA stimulated splenocyte proliferation. X-gal and GFP expression was assessed by histochemistry. Results - The extent of adenoviral vector mediated transfer of both reporter genes and vIL-10 was dose dependent. LacZ expression could be detected for at least 50 day after infection with multiple of infection (MOI) 200. Following AdCMVvIL-10 transduction, vIL-10 protein expression occurred between 4-6 days post-transduction, and was maintained at a detectable level for at least 1 month. Secreted vIL-10 showed biological activity, significantly inhibiting Con A induced splenocyte proliferation. Additionally, transfection of HCs with two Adv vectors, one carrying LacZ and the other carrying GFP, resulted in co-expression within a single cell. Conclusion - These results confirm previous successful adenoviral vector mediated gene transfer to HCs and further show that expression can be maintained. Furthermore the data show HCs can secrete biologically active vIL-10 that could be developed as a strategy to suppress immune mediated disorders. The successful co-transduction of HCs as described for other tissues, opens avenues to develop a multiple target gene therapy locally.

Original languageEnglish (US)
Pages (from-to)861-867
Number of pages7
JournalBritish Journal of Ophthalmology
Volume85
Issue number7
DOIs
StatePublished - 2001
Externally publishedYes

ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience

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