GFAP-driven GFP expression in activated mouse Müller glial cells aligning retinal blood vessels following intravitreal injection of AAV2/6 vectors.
BACKGROUND
Müller cell gliosis occurs in various retinal pathologies regardless of the underlying cellular defect. Because activated Müller glial cells span the entire retina and align areas of injury, they are ideal targets for therapeutic strategies, including gene therapy.


RESULTS
We used adeno-associated viral AAV2/6 vectors to transduce mouse retinas. The transduction pattern of AAV2/6 was investigated by studying expression of the green fluorescent protein (GFP) transgene using scanning-laser ophthalmoscopy and immuno-histochemistry. AAV2/6 vectors transduced mouse Müller glial cells aligning the retinal blood vessels. However, the transduction capacity was hindered by the inner limiting membrane (ILM) and besides Müller glial cells, several other inner retinal cell types were transduced. To obtain Müller glial cell-specific transgene expression, the cytomegalovirus (CMV) promoter was replaced by the glial fibrillary acidic protein (GFAP) promoter. Specificity and activation of the GFAP promoter was tested in a mouse model for retinal gliosis. Mice deficient for Crumbs homologue 1 (CRB1) develop gliosis after light exposure. Light exposure of Crb1(-/-) retinas transduced with AAV2/6-GFAP-GFP induced GFP expression restricted to activated Müller glial cells aligning retinal blood vessels.


CONCLUSIONS
Our experiments indicate that AAV2 vectors carrying the GFAP promoter are a promising tool for specific expression of transgenes in activated glial cells.
Aartsen
Wendy M
WM
Department of Neuromedical Genetics, Netherlands Institute for Neuroscience, an Institute of the Royal Netherlands Academy of Arts and Sciences, Amsterdam, The Netherlands.
van Cleef
Koen W R
KW
Pellissier
Lucie P
LP
Hoek
Robert M
RM
Vos
Rogier M
RM
Blits
Bas
B
Ehlert
Erich M E
EM
Balaggan
Kamaljit S
KS
Ali
Robin R
RR
Verhaagen
Joost
J
Wijnholds
Jan
J
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D015870:Gene Expression; D005822:Genetic Vectors / Q000008:administration & dosage* / Q000235:genetics*; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000235:genetics*; D006801:Humans; D007267:Injections; D051379:Mice; D009457:Neuroglia / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D012171:Retinal Vessels / Q000166:cytology*; D014161:Transduction, Genetic / Q000379:methods*; D019076:Transgenes / Q000235:genetics
D005904:Glial Fibrillary Acidic Protein; D049452:Green Fluorescent Proteins
Mol Ther. 2008 Jan;16(1):89-96
17955025
J Neurosci Methods. 2010 Jan 15;185(2):257-63
19850079
Mol Cell Neurosci. 2002 Sep;21(1):141-57
12359157
J Neurosci. 2002 Nov 1;22(21):9228-36
12417648
J Neurosci. 2007 Jan 17;27(3):564-73
17234588
Graefes Arch Clin Exp Ophthalmol. 2007 Jul;245(7):1019-25
17186258
Hum Mol Genet. 2006 Sep 15;15(18):2659-72
16885194
Prog Retin Eye Res. 2005 May;24(3):395-431
15708835
N Engl J Med. 2008 May 22;358(21):2240-8
18441370
Mol Ther. 2009 Dec;17(12):2096-102
19672248
Invest Ophthalmol Vis Sci. 2007 Apr;48(4):1844-52
17389520
N Engl J Med. 2008 May 22;358(21):2231-9
18441371
Eye (Lond). 2009 Oct;23(10):1980-4
19098702
Hum Mol Genet. 2006 Oct 15;15 Spec No 2:R235-43
16987889
Invest Ophthalmol Vis Sci. 2006 Aug;47(8):3586-94
16877433
Neuron. 2000 May;26(2):285-6
10839346
Nat Neurosci. 2000 Sep;3(9):873-80
10966617
Glia. 2007 Nov 1;55(14):1486-97
17705196
Nat Genet. 1999 Oct;23(2):217-21
10508521
Gene Ther. 2006 Aug;13(16):1191-7
16838031
Gene Ther. 2009 Apr;16(4):521-32
19092858
Prog Retin Eye Res. 2006 Jul;25(4):397-424
16839797
Invest Ophthalmol Vis Sci. 2009 Aug;50(8):3977-84
19387072
Proc Natl Acad Sci U S A. 2008 Sep 30;105(39):15112-7
18809924
PLoS One. 2009 Oct 14;4(10):e7467
19826483
Mol Ther. 2009 Dec;17(12):2088-95
19672246
N Engl J Med. 2009 Aug 13;361(7):725-7
19675341
Hum Mol Genet. 2003 Sep 1;12(17):2179-89
12915475
Prog Retin Eye Res. 2003 Jan;22(1):1-29
12597922
Br J Ophthalmol. 2003 Sep;87(9):1159-66
12928288
Vision Res. 1996 Sep;36(18):2979-94
8917798
Invest Ophthalmol Vis Sci. 2000 Mar;41(3):927-36
10711715
Curr Eye Res. 2002 Apr;24(4):305-12
12324870
Invest Ophthalmol Vis Sci. 2003 Aug;44(8):3606-13
12882814
J Cell Sci. 2004 Aug 15;117(Pt 18):4169-77
15316081
Hum Gene Ther. 1999 Jul 20;10(11):1885-91
10446928
Hum Mol Genet. 2006 Apr 15;15(8):1291-302
16520334
Vision Res. 2005 Dec;45(28):3512-9
16188288
Expert Opin Biol Ther. 2006 Dec;6(12):1279-94
17223737
Eye (Lond). 2008 Oct;22(10):1301-9
18327161
Invest Ophthalmol Vis Sci. 2008 Aug;49(8):3659-65
18469190
Hum Mutat. 2004 Nov;24(5):355-69
15459956
Gene Ther. 2008 Jun;15(11):849-57
18418417