Ophthalmology Commons^™

"Transduction Efficiency Of Aav 2/6, 2/8 And 2/9 Vectors For Delivering Genes In Human Corneal Fibroblasts ", Ajay Sharma, Arkasubhra Ghosh, Eric T. Hansen, Jason M. Newman, Rajiv R. Mohan

Pharmacy Faculty Articles and Research

In the present study, cellular tropism and relative transduction efficiency of AAV2/6, AAV2/8 and AAV2/9 vectors have been tested for the cornea using primary cultures of human corneal fibroblasts. The AAV6, AAV8 and AAV9 serotypes having AAV2 ITR plasmid encoding for alkaline phosphatase (AP) gene were generated by transfecting HEK293 cell line with pHelper, pARAP4 and pRep/Cap plasmids. Primary cultures of human corneal fibroblasts were exposed to AAV infectious particles at two different doses (1×105 and 2×105 MOI). Cytochemistry and enzyme assays were used to measure delivered transgene expression in samples collected at 4 and 30 hours after AAV infection …

Aav Serotype Influences Gene Transfer In Corneal Stroma In Vivo, Ajay Sharma, Jonathan C. K. Tovey, Arkasubhra Ghosh, Rajiv R. Mohan

Pharmacy Faculty Articles and Research

This study evaluated the cellular tropism and relative transduction efficiency of three AAV serotypes, AAV6, AAV8 and AAV9, for corneal gene delivery using mouse cornea in vivo and donor human cornea ex vivo. The AAV6, AAV8 and AAV9 serotypes having AAV2 plasmid encoding for alkaline phosphatase (AP) gene were generated by transfecting HEK293 cell line with pHelper, pARAP4 and pRep/Cap plasmids. Viral vectors (109 vg/μl) were topically applied onto mouse cornea in vivo and human cornea ex vivo after removing the epithelium. Human corneas were processed for transgene delivery at day 5 after viral vector application. Mouse corneas were harvested …

Isolation And Cultivation Of Equine Corneal Keratocytes, Fibroblasts And Myofibroblasts, Dylan G. Buss, Elizabeth A. Giuliano, Ajay Sharma, Rajiv R. Mohan

Pharmacy Faculty Articles and Research

Objective—To establish an in vitro model for the investigation of equine corneal wound healing. To accomplish this goal, a protocol to isolate and culture equine corneal keratocytes, fibroblasts and myofibroblasts was developed.

Animal material—Equine corneal buttons were aseptically harvested from healthy research horses undergoing humane euthanasia for reasons unrelated to this study. Slit-lamp biomicroscopy was performed prior to euthanasia by a board-certified veterinary ophthalmologist to ensure that all samples were harvested from horses free of anterior segment disease.

Procedure—Equine corneal stroma was isolated using mechanical techniques and stromal subsections were then cultured. Customized media at different culture conditions was used …

Defensins In Ocular Immunity, Minhao Wu

Wayne State University Dissertations

Corneal infection with P. aeruginosa results in corneal perforation in susceptible

B6, but not resistant BALB/c mice. This study explored their role mBD 1-4 in corneal

infection, and their potential synergy. Immunostaining and real-time RT-PCR data

demonstrated that their expression was either constitutive (mBD1 and mBD2) or

inducible (mBD3 and mBD4) in normal BALB/c and B6 corneas, and disparately

regulated in BALB/c vs B6 corneas after infection. Knock down studies using siRNA

treatment indicated that mBD2 and mBD3, but neither mBD1 nor mBD4, is required in

ocular defense. Moreover, in vivo studies demonstrated individual and combined effects

of mBD2 and …

Stromal Haze, Myofibroblasts, And Surface Irregularity After Prk, Marcelo V. Netto, Rajiv R. Mohan, Sunilima Sinha, Ajay Sharma, William Dupps, Steven E. Wilson

Pharmacy Faculty Articles and Research

The aim of this study was to investigate the relationship between the level of stromal surface irregularity after photorefractive keratectomy (PRK) and myofibroblast generation along with the development of corneal haze.

Variable levels of stromal surface irregularity were generated in rabbit corneas by positioning a fine mesh screen in the path of excimer laser during ablation for a variable percentage of the terminal pulses of the treatment for myopia that does not otherwise generate significant opacity. Ninety-six rabbits were divided into eight groups[.]

Slit lamp analysis and haze grading were performed in all groups. Rabbits were sacrificed at 4 hr …