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Retinal ischemia is a major cause of visual impairment and is associated with a high risk of subsequent ischemic stroke. The retina and its projections are easily accessible for experimental procedures and functional evaluation. We created and characterized a mouse model of global and transient retinal ischemia and provide a comprehensive chronologic profile of some genes that display altered expression during ischemia.

The alpha v beta (αvβ5) heterodimer has been implicated in many biological functions, including angiogenesis. We report the β5 gene expression pattern in embryonic and foetal mouse tissues as determined by Northern blotting and in situ hybridization. During the earliest stages, β5 mRNA is widespread in the mesoderm. During later developmental stages, it remains mostly confined to tissues of mesodermal origin, although probable inductive effects trigger shifts of β5 gene expression from some mesenchymatous to epithelial structures. This was observed in the teeth, skin, kidneys, and gut. Of physiological importance is the β5 labeling in the developing cardiovascular and respiratory systems and cartilages. Furthermore, early β5 gene expression was observed within the intra- and extraembryonic sites of hematopoiesis. This suggests a major role for β5 in the hematopoietic and angiogenic stem cells and thus in the development of the vascular system. Later, the β5 gene was expressed in endothelial cells of the vessels developing both by angiogenesis and vasculogenesis in the lung, heart, and kidneys. Moreover, the β5 hybridization signal was detected in developing cartilages but not in ossified or ossifying bones. β5-Integrin is a key integrin involved in angiogenesis, vasculogenesis, hematopoiesis, and bone formation.

The RCS rat presents an autosomal recessive retinal pigment epithelium dystrophy characterized by the outer segments of photoreceptors being phagocytosis-deficient. A systematic genetic study allowed us to restrict the interval containing the rdylocus to that between the markers D3Mit13 and D3Rat256. We report the chromosomal localization of the rat c-mergene in the cytogenetic bands 3q35-36, based on genetic analysis and radiation hybrid mapping. Using a systematic biocomputing analysis, we identified two strong related candidate genes encoding protein tyrosine kinase receptors of the AXLsubfamily. The comparison of their expression patterns in human and mice tissues suggested that the c-mergene was the best gene to screen for mutations. RCS rdy− and RCS rdy+ cDNAs were sequenced. The RCS rdy− cDNAs carried a significant deletion in the 5′ part of the coding sequence of the c-mergene resulting in a shortened aberrant transcript encoding a 20 amino acid peptide. The c-mergene contains characteristic motifs of neural cell adhesion. A ligand of the c-merreceptor, Gas6, exhibits antiapoptotic properties.

Based on the K8/JTS-1-mediated transfection technique, we developed an in vivo protocol for an efficient transfer of plasmid DNA to ocular cells. As determined with condensed plasmids containing reporter genes for either β-galactosidase (pcDNA-lacZ) or enhanced green fluorescent protein (pREP-EGFP), the immortalized human retinal epithelial cells RPE D407 and human embryonic kidney 293 cells can be transfected with typical efficiencies of 11 and 19%, respectively. Unlike 293 cells, RPE D407 cells had a reduced viability on transfection with both plasmids. In vivo, subretinal injections of DNA-K8/JTS-1 complexes revealed reporter gene expression in choroidal and RPE cells of normal pink-eyed Royal College of Surgeons (RCS) rats. The validity of this transfection technique in terms of retinal cell survival in RCS rats was then examined by using pREP-hFGF2 plasmid, which encodes the human basic fibroblast growth factor isoforms (hFGF2). Subretinal injection of pREP-hFGF2-K8/JTS-1 complexes into 3-week-old dystrophic RCS rat eyes reveals a delayed photoreceptor cell degeneration 60 days postinjection. In this case, the average analyzed field points with delayed cell dystrophy represent 14 to 17% of the retinal surface as compared with 2.6 and 4% in pREP5β and vehicle-injected eyes, respectively. Peptide-mediated in oculo transfection thus appears to be a promising technique for the treatment of retinal cell and photoreceptor degenerations.

Mutations in human BIGH3 (TGFB1), a gene identified after treatment of an adenocarcinoma cell line with TGF-β, have been observed in patients with granular Groenouw type I, Reis–Bu¨cklers, Thiel–Behnke, Avellino, and Lattice type I and IIIa, six autosomal dominant corneal dystrophies linked to chromosome 5q. In order to gain insight into the physiological role of this gene, we characterized the genomic structure of the mouse Bigh3 and its expression in murine embryos. The gene spans 30 kb on mouse chromosome 13 and has 17 exons. Embryonic expression of Bigh3 is observed in the mesenchyme of the first and second branchial archs as early as dpc 11.5 and is particularly strong in the mesenchyme of numerous tissues throughout all the development stages. In fetal eye, the expression is first seen at 11.5 dpc in the mesenchyme surrounding the optic stalk, extends toward the sclera and choroid by 14.3 dpc and reaches the cornea by 17.5 dpc. Because the physiological role of BIGH3/Bigh3 is still largely unknown, embryonic expression in organs like heart, vessels, and intestine may help to identify new functions which could be searched for in patients and in knock-out animal models. The characterization of the murine structure is a prerequisite for the making of such models.

• An argon fluoride excimer laser (193 nm) with a moving slit delivery system was used to perform anterior myopic keratomileusis in both eyes of 24 New Zealand white rabbits. Rabbits were killed immediately after ablation and at intervals up to 100 days. By slit-lamp microscopy, four rabbits at day 100 exhibited four clear corneas and four corneas had central, spotty, subepithelial haze. Light and electron microscopy documented corneal healing. In the early stages a transient acellular zone in the anterior stroma appeared over a period of three weeks, followed by an increased number of fibrocytes. In the corneas with opacification, focal areas of 20-μm-thick subepithelial scarring were present. An unexpected finding was transient damage to posterior stromal keratocytes and endothelial cells. The endothelium produced a layer of granular material that migrated anteriorly across Descemet's membrane. Immunochemistry at day 6 showed a marked staining for collagen IV, proteoglycans, fibronectin, and laminin.