Institute for Research in Ophthalmology

Sion, Switzerland

Institute for Research in Ophthalmology

Sion, Switzerland
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Abplanalp J.,University of Zürich | Abplanalp J.,ETH Zurich | Laczko E.,University of Zürich | Philp N.J.,Thomas Jefferson University | And 12 more authors.
Human Molecular Genetics | Year: 2013

Creatine transport has been assigned to creatine transporter 1 (CRT1), encoded by mental retardation associatedSLC6A8. Here,weidentifieda secondcreatine transporter (CRT2)knownasmonocarboxylate transporter 12 (MCT12), encoded by the cataract and glucosuria associated gene SLC16A12. A non-synonymous alteration inMCT12 (p.G407S) found in a patientwith age-related cataract (ARC) leads to a significant reduction of creatine transport. Furthermore, Slc16a12 knockout (KO) rats have elevated creatine levels in urine. Transport activity and expression characteristics of the two creatine transporters are distinct. CRT2 (MCT12)-mediated uptake of creatine was not sensitive to sodium and chloride ions or creatine biosynthesis precursors, breakdown product creatinine or creatine phosphate. Increasing pH correlated with increased creatine uptake. Michaelis-Menten kinetics yielded a Vmax of 838.8 pmol/h/oocyte and a Km of 567.4 μM. Relative expression in various human tissues supports the distinct mutation-associated phenotypes of the two transporters. SLC6A8 was predominantly found in brain, heart and muscle, while SLC16A12 was more abundant in kidney and retina. In the lens, the two transcripts were found at comparable levels.We discuss the distinct, but possibly synergistic functions of the two creatine transporters. Our findings infer potential preventivepower of creatine supplementation against the most prominent age-related vision impaired condition. © The Author 2013. Published by Oxford University Press. All rights reserved.


Produit-Zengaffinen N.,Institute for Research in Ophthalmology | Pournaras C.J.,University of Geneva | Schorderet D.F.,Institute for Research in Ophthalmology | Schorderet D.F.,University of Lausanne | Schorderet D.F.,Ecole Polytechnique Federale de Lausanne
Advances in Experimental Medicine and Biology | Year: 2014

The role played by autophagy after ischemia/reperfusion (I/R) in the retina remains unknown. Our study investigated whether ischemic injury in the retina, which causes an energy crisis, would induce autophagy. Retinal ischemia was induced by elevation of the intraocular pressure and modulation of autophagic markers was analyzed at the protein levels in an early and late phase of recovery. Following retinal ischemia an increase in LC3BII was first observed in the early phase of recovery but did not stay until the late phase of recovery. Post-ischemic induction of autophagy by intravitreal rapamycin administration did not provide protection against the lesion induced by the ischemic stress. On the contrary, an increase in the number of apoptotic cells was observed following I/R in the rapamycin treated retinas. © 2014, Springer Science+Business Media, LLC.


Boisset G.,Institute for Research in Ophthalmology | Schorderet D.F.,Institute for Research in Ophthalmology | Schorderet D.F.,University of Lausanne | Schorderet D.F.,Ecole Polytechnique Federale de Lausanne
Experimental Eye Research | Year: 2012

Ocular development is controlled by a complex network of transcription factors, cell cycle regulators, and diffusible signaling molecules. Together, these molecules regulate cell proliferation, apoptosis and specify retinal fate. In the zebrafish (. Danio rerio), hmx1 is a homeobox transcription factor implicated in eye and brain development. Hmx1 transcripts were detected in the nasal retina and lens as well as otic vesicles and pharyngeal arches by 24-32 hpf. Before this stage, transcripts were more uniformly expressed in the optic vesicle. Knockdown of hmx1 led to microphthalmia. Delayed withdrawal of retinal progenitors from the cell cycle resulting in retarded retinal differentiation was observed in morphant. The retina and brain also showed an increased cell death at 24 hpf. The polarized expression of hmx1 to the nasal part in the zebrafish retina strongly suggested an involvement in the nasal-temporal patterning. However, the key patterning genes tested so far were not regulated by hmx1. Altogether, these results suggest an important role for hmx1 in retinogenesis. © 2012 Elsevier Ltd.


Boulling A.,Institute for Research in Ophthalmology | Wicht L.,Institute for Research in Ophthalmology | Wicht L.,Ecole Polytechnique Federale de Lausanne | Schorderet D.F.,Institute for Research in Ophthalmology | And 2 more authors.
Molecular Vision | Year: 2013

Purpose: A homozygous mutation in the H6 family homeobox 1 (HMX1) gene is responsible for a new oculoauricular defect leading to eye and auricular developmental abnormalities as well as early retinal degeneration (MIM 612109). However, the HMX1 pathway remains poorly understood, and in the first approach to better understand the pathway's function, we sought to identify the target genes. Methods: We developed a predictive promoter model (PPM) approach using a comparative transcriptomic analysis in the retina at P15 of a mouse model lacking functional Hmx1 (dmbo mouse) and its respective wild-type. This PPM was based on the hypothesis that HMX1 binding site (HMX1-BS) clusters should be more represented in promoters of HMX1 target genes. The most differentially expressed genes in the microarray experiment that contained HMX1-BS clusters were used to generate the PPM, which was then statistically validated. Finally, we developed two genome-wide target prediction methods: one that focused on conserving PPM features in human and mouse and one that was based on the co-occurrence of HMX1-BS pairs fitting the PPM, in human or in mouse, independently. Results: The PPM construction revealed that sarcoglycan, gamma (35kDa dystrophin-associated glycoprotein) (Sgcg), teashirt zinc finger homeobox 2 (Tshz2), and solute carrier family 6 (neurotransmitter transporter, glycine) (Slc6a9) genes represented Hmx1 targets in the mouse retina at P15. Moreover, the genome-wide target prediction revealed that mouse genes belonging to the retinal axon guidance pathway were targeted by Hmx1. Expression of these three genes was experimentally validated using a quantitative reverse transcription PCR approach. The inhibitory activity of Hmx1 on Sgcg, as well as protein tyrosine phosphatase, receptor type, O (Ptpro) and Sema3f, two targets identified by the PPM, were validated with luciferase assay. Conclusions: Gene expression analysis between wild-type and dmbo mice allowed us to develop a PPM that identified the first target genes of Hmx1. © 2013 Molecular Vision.


Metrailler S.,Institute for Research in Ophthalmology | Schorderet D.F.,Institute for Research in Ophthalmology | Schorderet D.F.,University of Lausanne | Schorderet D.F.,Ecole Polytechnique Federale de Lausanne | And 2 more authors.
Experimental Eye Research | Year: 2012

RPE65-related Leber's congenital amaurosis (LCA) is a rod-cone dystrophy whose clinical outcome is mainly attributed to the loss of rod photoreceptors followed by cone degeneration. Pathogenesis in Rpe65 -/- mice is characterized by a slow and progressive degeneration of rods dependent on the constitutive activation of unliganded opsin. We previously reported that this opsin-mediated apoptosis of rods was dependent on Bcl-2-apoptotic pathway and Bax-induced pro-death activity. In this study, we report early initial apoptosis in the newly differentiated retina of Rpe65 -/- mice. Apoptotic photoreceptors were identified as rods and resulted from pathological phototransduction signaling. This wave of early apoptosis triggered Bcl-2-related pathway and Bax apoptotic activity, while activation of the caspases was not induced. Following cellular stress, multiple signaling pathways are initiated which either commit cells to death or trigger pro-survival responses including autophagy. We report that Bcl-2-related early rod apoptosis was associated with the upregulation of autophagy markers including chaperone-mediated autophagy (CMA) substrate receptor LAMP-2 and lysosomal hydrolases Cathepsin S and Lysozyme. This suggests that lysosomal-mediated autophagy may be triggered in response to early rod apoptosis in Rpe65-LCA disease. These results highlight that Rpe65-related primary stress induces early signaling events, which trigger Bax-induced-apoptotic pathway and autophagy-mediated cellular response. These events may determine retinal cell fate, progression and severity of the disease. © 2011 Elsevier Ltd.


Allaman-Pillet N.,Institute for Research in Ophthalmology | Oberson A.,Institute for Research in Ophthalmology | Schorderet D.F.,Institute for Research in Ophthalmology | Schorderet D.F.,University of Lausanne | Schorderet D.F.,Ecole Polytechnique Federale de Lausanne
Molecular Cancer Research | Year: 2015

Retinoblastoma is the most common pediatric intraocular neoplasm. While retinoblastoma development requires the inactivation of both alleles of the retinoblastoma tumor suppressor gene (RB1) in the developing retina, additional genomic changes are involved in tumor progression, which progressively lead to resistance of tumor cells to death. Therapeutics acting at very downstream levels of death signaling pathways should therefore be interesting in killing retinoblastoma cells. The BH3-only proteins promote apoptosis by modulating the interaction between the pro- and antiapoptotic members of the BCL2 protein family, and this effect can be recapitulated by the BH3 domains. This report analyzes the effect of various BH3 peptides, corresponding to different BH3-only proteins, on two retinoblastoma cell lines, Y79 and WERI-Rb, as well as on the photoreceptor cell line 661W. The BH3 peptide BIRO1, derived from the BCL2L11 death domain, was very effective in promoting Y79 and WERI-Rb cell death without affecting the 661W photoreceptor cells. This cell death was efficient even in absence of BAX and was shown to be caspase independent. While ROS production or AIF release was not detected from mitochondria of treated cells, BIRO1 initiated mitochondria fragmentation in a short period of time following treatment. Implications: The BIRO1 peptide is highly effective at killing retinoblastoma cells and has potential as a peptidomimetic. ©2014 AACR.


Hamann S.,Institute for Research in Ophthalmology | Hamann S.,Ecole Polytechnique Federale de Lausanne | Metrailler S.,Institute for Research in Ophthalmology | Schorderet D.F.,Institute for Research in Ophthalmology | And 4 more authors.
PLoS ONE | Year: 2013

α-Crystallins, initially described as the major structural proteins of the lens, belong to the small heat shock protein family. Apart from their function as chaperones, α-crystallins are involved in the regulation of intracellular apoptotic signals. αA- and αB-crystallins have been shown to interfere with the mitochondrial apoptotic pathway triggering Bax pro-apoptotic activity and downstream activation of effector caspases. Differential regulation of α-crystallins has been observed in several eye diseases such as age-related macular degeneration and stress-induced and inherited retinal degenerations. Although the function of α-crystallins in healthy and diseased retina remains poorly understood, their altered expression in pathological conditions argue in favor of a role in cellular defensive response. In the Rpe65-/- mouse model of Leber's congenital amaurosis, we previously observed decreased expression of αA- and αB-crystallins during disease progression, which was correlated with Bax pro-death activity and photoreceptor apoptosis. In the present study, we demonstrated that α-crystallins interacted with pro-apoptotic Bax and displayed cytoprotective action against Bax-triggered apoptosis, as assessed by TUNEL and caspase assays. We further observed in staurosporine-treated photoreceptor-like 661W cells stably overexpressing αA- or αB-crystallin that Bax-dependent apoptosis and caspase activation were inhibited. Finally, we reported that the C-terminal extension domain of αA-crystallin was sufficient to provide protection against Bax-triggered apoptosis. Altogether, these data suggest that α-crystallins interfere with Bax-induced apoptosis in several cell types, including the cone-derived 661W cells. They further suggest that αA-crystallin-derived peptides might be sufficient to promote cytoprotective action in response to apoptotic cell death. © 2013 Hamann et al.


le Carre J.,Institute for Research in Ophthalmology | Schorderet D.F.,Institute for Research in Ophthalmology | Schorderet D.F.,University of Lausanne | Schorderet D.F.,Ecole Polytechnique Federale de Lausanne | And 2 more authors.
Molecular Vision | Year: 2011

Purpose: In this study, we investigated the expression of the gene encoding β-galactosidase (Glb)-1-like protein 3 (Glb1l3), a member of the glycosyl hydrolase 35 family, during retinal degeneration in the retinal pigment epithelium (RPE)-specific 65-kDa protein knockout (Rpe65-/-) mouse model of Leber congenital amaurosis (LCA). Additionally, we assessed the expression of the other members of this protein family, including β-galactosidase-1 (Glb1), β- galactosidase-1-like (Glb1l), and β-galactosidase-1-like protein 2 (Glb1l2).Methods: The structural features of Glb1l3 were assessed using bioinformatic tools. mRNA expression of Glb-related genes was investigated by oligonucleotide microarray, real-time PCR, and reverse transcription (RT) -PCR. The localized expression of Glb1l3 was assessed by combined in situ hybridization and immunohistochemistry. Results: Glb1l3 was the only Glb-related member strongly downregulated in Rpe65-/- retinas before the onset and during progression of the disease. Glb1l3 mRNA was only expressed in the retinal layers and the RPE/choroid. The other Glb- related genes were ubiquitously expressed in different ocular tissues, including the cornea and lens. In the healthy retina, expression of Glb1l3 was strongly induced during postnatal retinal development; age-related increased expression persisted during adulthood and aging.Conclusions: These data highlight early-onset downregulation of Glb1l3 in Rpe65-related disease. They further indicate that impaired expression of Glb1l3 is mostly due to the absence of the chromophore 11-cis retinal, suggesting that Rpe65 deficiency may have many metabolic consequences in the underlying neuroretina. © 2011 Molecular Vision.


Escher P.,Institute for Research in Ophthalmology | Escher P.,University of Lausanne | Schorderet D.F.,Institute for Research in Ophthalmology | Schorderet D.F.,University of Lausanne | And 3 more authors.
Investigative Ophthalmology and Visual Science | Year: 2011

Purpose. To investigate the role of the myocyte enhancer factor 2 (Mef2) transcription factor family in retinal diseases, Mef2c expression was assessed during retinal degeneration in the Rpe65 -/- mouse model of Leber's congenital amaurosis (LCA). Mef2c-dependent expression of photoreceptor-specific genes was further addressed. Methods. Expression of Mef2 members was analyzed by oligonucleotide microarray, quantitative PCR (qPCR), and in situ hybridization. Mef2c-dependent transcriptional activity was assayed by luciferase assay in HEK293T cells. Results. Mef2c was the only Mef2 member markedly downregulated during retinal degeneration in Rpe65 -/- mice. Mef2c mRNA level was decreased by more than 2-fold at 2 and 4 months and by 3.5-fold at 6 months in retinas of Rpe65 -/- mice. Downregulation of Mef2c at the protein level was confirmed in Rpe65 -/- retinas. The decrease in Mef2c mRNA levels in the developing Rpe65 -/- retinas from postnatal day (P) 13 onward was concomitant with the decreased expression of the rodspecific transcription factors Nrl and Nr2e3. Nrl was further shown to drive Mef2c transcriptional activity, supporting a physiological role for Mef2c in the retina. In addition, Mef2c appeared to act as a transcriptional repressor of its own expression and the expression of the retina-specific retinal G-protein coupled receptor (Rgr), rhodopsin, and M-opsin genes. Conclusions. These findings highlight the early altered regulation of the rod-specific transcriptional network in Rpe65- related disease. They also indicate that Mef2c may act as a novel transcription factor involved in the development and the maintenance of photoreceptor cells. © 2011 The Association for Research in Vision and Ophthalmology, Inc.


Boulling A.,University of Lausanne | Boulling A.,Institute for Research in Ophthalmology | Escher P.,University of Lausanne
Experimental Eye Research | Year: 2016

Ex vivo electroporation of mouse retinas is an established tool to modulate gene expression and to study cell type-specific gene expression. Here we coupled ex vivo electroporation to luciferase reporter assays to facilitate the study of rod-photoreceptor-specific gene promoters. The activity of the rod-specific proximal bovine rhodopsin promoter was significantly increased in C57BL/6J wild-type retinas at postnatal days 1 and 7 by 3.4-fold and 8.7-fold respectively. In C57BL/6J Nr2e3rd7/rd7 retinas, where the rod photoreceptor-specific nuclear receptor Nr2e3 is not expressed, a significant increase by 2.5-fold was only observed at postnatal day 7. Cone-specific S-opsin promoter activity was not modulated in C57BL/6J wild-type and Nr2e3rd7/rd7 retinas. Taken together, we describe an easily implementable protocol to assess rod-specific promoter activity in a physiological context resembling that of the developing postnatal mouse retina. © 2016 Elsevier Ltd.

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