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Evaluation of delivery agents used for introduction of small interfering RNAs into feline corneal cells.
MedLine Citation:
PMID:  23363349     Owner:  NLM     Status:  In-Data-Review    
Objective-To evaluate agents used for delivery of small interfering RNAs (siRNAs) into feline corneal cells, toxicity of the delivery agents, and functionality of anti-feline herpesvirus 1 (FHV-1)-specific siRNA combinations. Sample-Feline primary corneal cells and 19 six-month-old colony-bred cats. Procedures-siRNA delivery into corneal cells via various delivery agents was evaluated via flow cytometric detection of labeled siRNAs. Cellular toxicity was evaluated with a proliferation assay. Functionality was tested via quantitative reverse transcriptase PCR assay, plaque assay, and flow cytometry. In vivo safety was evaluated with an ocular scoring method following topical application of delivery agents containing siRNAs into eyes. Corneal biopsy specimens were used to assess safety and uptake of siRNAs into corneal cells. Results-Use of 3 delivery agents resulted in > 95% transfection of primary corneal cells. Use of a peptide for ocular delivery yielded approximately 82% transfection of cells in vitro. In cultured corneal cells, use of the siRNA combinations resulted in approximately 76% to 89% reduction in FHV-1-specific mRNA, 63% to 67% reduction of FHV-1-specific proteins in treated cells, and 97% to 98% reduction in FHV-1 replication. The agents were nonirritating in eyes, caused no substantial clinical ocular signs, and were nontoxic. Histologically, corneal epithelium and stroma were normal in treated cats. However, none of the agents were effective in delivering siRNAs into the corneal cells in vivo. Conclusions and Clinical Relevance-The tested anti-FHV-1-specific siRNAs could potentially be used as a treatment for FHV-1 if a successful means of in vivo delivery can be achieved.
Rebecca P Wilkes; Dan A Ward; Kim M Newkirk; Joleen K Adams; Stephen A Kania
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Publication Detail:
Type:  Journal Article    
Journal Detail:
Title:  American journal of veterinary research     Volume:  74     ISSN:  1943-5681     ISO Abbreviation:  Am. J. Vet. Res.     Publication Date:  2013 Feb 
Date Detail:
Created Date:  2013-01-31     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  0375011     Medline TA:  Am J Vet Res     Country:  United States    
Other Details:
Languages:  eng     Pagination:  243-7     Citation Subset:  IM    
Department of Biomedical and Diagnostic Research, College of Veterinary Medicine, University of Tennessee, Knoxville, TN 37996.
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