| First Author | Kuzmanovic M | Year | 2003 |
| Journal | Invest Ophthalmol Vis Sci | Volume | 44 |
| Issue | 8 | Pages | 3606-13 |
| PubMed ID | 12882814 | Mgi Jnum | J:151687 |
| Mgi Id | MGI:4355041 | Doi | 10.1167/iovs.02-1265 |
| Citation | Kuzmanovic M, et al. (2003) GFAP promoter drives Muller cell-specific expression in transgenic mice. Invest Ophthalmol Vis Sci 44(8):3606-13 |
| abstractText | PURPOSE: In an attempt to identify Muller cell-specific promoters and to better understand the gene regulatory mechanisms in retinal glial cells, the expression of the glial fibrillary acidic protein (GFAP) gene was studied in Muller cell cultures and in GFAP-enhanced green fluorescent protein (EGFP) transgenic mice. METHODS: A transfection assay of GFAP-luciferase constructs carrying a series of nested deletions was performed in an established Muller cell line. For in vivo analysis, transgenic mice were generated by injecting a construct carrying a 2.5-kb, 5' fragment of the mouse GFAP gene linked to the EGFP gene. Isolated retinas from transgenic mice were screened for GFP expression. Subsequently, the identity of the GFP-expressing cells was established by immunostaining cryostat sections of the retina with antibodies against Muller cell antigenic markers. Induction of the transgene and the endogenous GFAP gene was examined by injecting ciliary neurotrophic factor (CNTF) into the eye. RESULTS: The DNA transfection data suggested that proximal 5' sequences of the GFAP gene are sufficient to direct high-level reporter expression in Muller cell cultures. In transgenic mice, GFP fluorescence appeared in radially oriented processes that spanned almost the entire thickness of the retina. Immunostaining with antibodies to cellular retinaldehyde-binding protein (CRALBP) and glutamine synthetase showed that the GFP-expressing cells were Muller cells. GFP-expressing Muller cells were observed in the retinas of both albino and pigmented transgenic mice. In eyes injected with CNTF, both GFP and GFAP levels were highly elevated. These observations suggest that the 2.5-kb, 5' GFAP sequence can direct inducible reporter gene expression in Muller cells. In addition to Muller cells, a few GFP-labeled astrocytes were present in the adult retina. In the developing retina, GFP-expressing astrocytes were first present at the optic nerve head, and as development progressed, the cells gradually moved toward the periphery of the retina and acquired their adult, stellate morphology. CONCLUSIONS: The present study shows that the 2.5-kb, 5' flanking region of the mouse GFAP gene can be used to express GFP, and possibly other genes, specifically in Muller cells in the mouse retina. Furthermore, expression of the transgene can be upregulated by intravitreal injection of CNTF. |
