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COCH transgene expression in cultured human trabecular meshwork cells and its effect on outflow facility in monkey organ cultured anterior segments

Authors
 Eun Suk Lee  ;  B’Ann T. Gabelt  ;  Jennifer A. Faralli  ;  Donna M. Peters  ;  Curtis R. Brandt  ;  Paul L. Kaufman  ;  Sanjoy K. Bhattacharya 
Citation
 INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE, Vol.51(4) : 2060-2066, 2010 
Journal Title
 INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE 
ISSN
 0146-0404 
Issue Date
2010
MeSH
Adenoviridae/genetics ; Adolescent ; Adult ; Animals ; Anterior Eye Segment ; Aqueous Humor/secretion* ; Blotting, Western ; Cells, Cultured ; Extracellular Matrix Proteins ; Female ; Fluorescent Antibody Technique, Indirect ; Gene Expression/physiology* ; Genetic Vectors ; Green Fluorescent Proteins/genetics ; Humans ; Macaca fascicularis ; Macaca mulatta ; Male ; Organ Culture Techniques ; Proteins/genetics* ; Trabecular Meshwork/metabolism* ; Transfection ; Transgenes/physiology*
Abstract
Purpose. To determine the effects of COCH transgene expression on cultured human trabecular meshwork (HTM) cell morphology and on outflow facility (OF) in monkey organ cultured anterior segments (MOCAS). Methods. An adenoviral (Ad) vector expressing both cochlin (COCH) and green fluorescent protein (GFP) (AdCOCHGFP) or GFP alone (AdGFP) was used to transduce cultured HTM cells (multiplicity of transduction, 2.8 and 28). COCH transgene expression in transduced HTM cells and the culture medium was verified by Western blot analysis and immunofluorescence detection 5 days after transduction. MOCAS were used to test the effect of Ad vectors (2.8 x 10(10) viral particles per segment) on OF. The morphology of transduced MOCAS was evaluated by light microscopy. Results. Western blot analysis showed a viral vector dose-dependent expression of cochlin in transduced cells and the culture medium. There was no notable morphologic change in transduced cells. In MOCAS, cochlin expression was detectable in the medium by 3 days after transduction. A 35% decrease in OF in AdCOCHGFP-transduced MOCAS was detected after 3 days, decreasing by 76% after 12 days when compared to control segments injected with AdGFP. Anterior segment pressure (ASP) more than doubled (P < 0.05) in segments injected with AdCOCHGFP at 12 days after transduction. Light microscopy revealed normal angle structures in transduced segments. Conclusions. Ad vector delivery of the COCH transgene resulted in cochlin expression in HTM cells and MOCAS. Cochlin expression was effective in decreasing OF and increasing ASP in MOCAS, suggesting possible involvement of cochlin in IOP elevation in vivo. COCH gene delivery has potential for use in developing a glaucoma model.
Files in This Item:
T201000634.pdf Download
DOI
10.1167/iovs.09-4521
Appears in Collections:
1. College of Medicine (의과대학) > Dept. of Ophthalmology (안과학교실) > 1. Journal Papers
Yonsei Authors
Lee, Eun Suk(이은석)
URI
https://ir.ymlib.yonsei.ac.kr/handle/22282913/100728
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