Retinitis pigmentosa: rod photoreceptor rescue by a calcium-channel blocker in the rd mouse.

Retinitis pigmentosa is an inherited degenerative disease of photoreceptors leading to blindness. A well-characterized model for this disease is provided by the retinal degeneration mouse, in which the gene for the rod cGMP phosphodiesterase is mutated, as in some affected human families. We report that D-cis-diltiazem, a calcium-channel blocker that also acts at light-sensitive cGMP-gated channels, rescued photoreceptors and preserved visual function in the retinal degeneration mouse. The long record of diltiazem prescription in cardiology should facilitate the design of clinical trials for some forms of retinitis pigmentosa.

Familial hematological malignancies: ASXL1 gene investigation.

PURPOSE: Familial aggregation among patients with several hematological malignancies has been revealed. This emphasizes the importance of genetic factors. Only few genes predisposing to familial hematological malignancies have been reported until now due to the low occurrence. We have described in previous study PRF1 and CEBPA variants that might contribute to the background of genetic factors, which encourage us to extend our investigations to other cooperating genes. The aim of this study is to determine whether germline additional sex combs-like 1 (ASXL1) gene mutations may be involved? METHODS/PATIENTS: In this study, we investigated the candidate gene ASXL1 by direct sequencing in 88 unrelated Tunisian and French families with aggregated hematological malignancies. RESULTS: We report a new p.Arg402Gln germline missense substitution in two related Tunisian patients which has not been previously described. We identified here this variant for the first time in non-Hodgkin lymphoma. The p.Arg402Gln variant was not found in 200 control chromosomes. In silico analysis has predicted potential deleterious effect on ASXL1 protein. CONCLUSIONS: From an extended candidate genes analyzed in the field of familial hematological malignancies, ASXL1 might be involved. This variant should be considered since a potential damaging effect was predicted by in silico analysis, with a view to develop functional assay in order to investigate the biological assessment.

Gangliosides and phospholipids of the membranes from bovine adrenal medullary chromaffin granules.

The lipid and ganglioside compositions of membranes of chromaffin granules isolated from bovine adrenal medulla have been investigated. The detailed lipid analysis revealed the presence of high levels of lysophosphatidylcholine, in agreement with previous studies, but also of sphingomyelin and plasmalogens. From these membranes, gangliosides have been extracted and separated by thin-layer chromatography and analysed. 95% of the total recovered gangliosides were hematosides (GM3), which migrated as three major species. Sugar analyses have been performed, as well as the fatty acid compositions. The three hematoside gangliosides appeared to differ on the basis of their fatty acid composition. Compared with the brain, chromaffin granule membranes showed a simple ganglioside composition, thus offering a good model for the study of the metabolism and the role of gangliosides. The simple ganglioside composition of chromaffin granule membranes has allowed us to state that there are 60 mol phospholipid and 30 mol cholesterol per mol ganglioside.

Effect of neuraminidase treatment on the topological distribution of phospholipids in chick brain microsomes.

The desialylation of chick brain microsomal membranes affects the transbilayer distribution of phospholipids. When intact microsomes were treated with neuraminidase, less phosphatidylcholine and sphingomyelin could be hydrolysed with phospholipase C under experimental conditions which allowed the hydrolysis of the phospholipids of the external leaflet only. In contrast, the accessibility of phosphatidylethanolamine and phosphatidylserine to the external probes (trinitrobenzene sulfonic acid or phospholipase C) was not affected. After neuraminidase treatment of a microsomal fraction, less phosphatidylcholine, newly synthesized through the cytidine pathway, could be hydrolysed by phospholipase C, whereas the reaction of newly synthesized phosphatidylethanolamine molecules with trinitrobenzene sulfonic acid was not affected. The results suggest that in biological membranes some choline phospholipid molecules may interact with the sialyl residue of sialocompounds. This interaction may contribute to the maintenance of phospholipid asymmetry in brain membranes.

Effect of exogenous gangliosides on the lipid composition of chick neurons in culture.

When exogenous gangliosides are added to the growth medium of neuronal cell cultures they are inserted into their plasma membranes and are afterwards metabolized in the cytoplasmic interior. The action of exogenous gangliosides brings important morphological and biochemical changes to neurons in culture. The present report shows that the treatment with exogenous gangliosides of a primary culture of chick neurons modified the distribution of fatty acids in phosphatidylinositol (PI), mainly that of arachidonic acid and the fatty acids of the (n - 3) series without affecting the other phospholipids. The composition of neutral lipids did not change but their content was increased up to 2-3-fold depending upon the concentration of gangliosides. The change of the growth medium from one containing fetal calf serum to a chemically defined one reduced dramatically the content of free fatty acids while the addition of gangliosides raised this content to normal levels. The increase in the amount of diacylglycerol (DG) confirmed the finding that gangliosides stimulate phosphoinositide degradation. Finally the fatty acid composition of DG suggests indirectly that this compound might be produced also by degradation of phosphatidylcholine and not only of PI.

A study on the topological distribution of phospholipids in microsomal membranes of chick brain using phospholipase C and trinitrobenzenesulfonic acid.

The transbilayer distribution of phospholipids in chicken brain microsomal membranes has been investigated using trinitrobenzenesulfonic acid and phospholipase C from Clostridium welchii. The exposure of intact microsomes to trinitrobenzenesulfonic acid showed that the labelling of aminophospholipids followed biphasic kinetics, indicating that these membranes contain a fast- and a slow-reacting pool of aminophospholipids. Use of microsomes radioiodinated on their surface led to the conclusion that the fast-reacting pool may be located on the outer leaflet of the microsomal vesicles. It contains about 35% of the phosphatidylethanolamine, 29% of the ethanolamine plasmalogens and 18% of the phosphatidylserine. The treatment of intact microsomes with the phospholipase C Cl. welchii produced the hydrolysis of 50% of the phospholipids without any loss of their permeability properties, indicating that they are not permeable to the hydrolase. Phospholipids extracted from the microsomes were hydrolyzed rapidly by the phospholipase C with the exception of phosphatidylserine and phosphatidylinositol. In intact microsomes about 90% of phosphatidylcholine, 32% of ethanolamine phospholipids and 60% of sphingomyelin were accessible to the phospholipase. These results suggest that the phospholipids have an asymmetric distribution in chicken brain microsomes, the external leaflet containing about 75% of the choline phospholipids and 25% of the aminophospholipids, whereas an opposite distribution is observed in the inner leaflet.

Ether lipid content of phosphoglycerides from the retina and brain of chicken and calf.

Phospholipid contents and compositions were determined for chicken and calf retinas, chicken brain and calf gray matter. Retinal phospholipid compositions differ from brain phospholipid compositions by including a higher percentage of choline phosphoglycerides and lower percentages of ethanolamine and serine phosphoglycerides. The proportion of sphingomyelin is lower in calf retina than in calf brain. Among the ethanolamine phosphoglycerides, the proportions of the alk-1-enylacyl (plasmalogen) type are lower and the proportions of alkylacyl and diacyl types are higher in retina than in brain. The alkyacyl glycerophosphoethanolamines accounted for 7.6% and 8.9% of the ethanolamine phosphoglycerides from chicken and calf retinas respectively. Lower proportions of plasmalogens in the choline phosphoglycerides were found in retina as compared with brain. The alkylacyl glycerylphosphocholines comprised 4.0% of the retinal choline phosphoglycerides. Overall, a smaller proportion of retinal phosphilipids than of brain phospholipids contained alkyl or alk-1-enyl ether groups and the ratio of alkyl groups to alk-1-enyl groups was greater in retina than in brain.

In vitro synthesis and transbilayer movement of phosphatidylethanolamine molecules labelled with different fatty acids in chick brain microsomes.

The transbilayer fatty acid distribution of diacylglycerophosphoethanolamine and the translocation of newly synthesized phosphatidylethanolamine molecules labelled with different fatty acids has been investigated in chick brain microsomes using trinitrobenzensulfonic acid. The determination of the fatty acid composition of diacylglycerophosphoethanolamine in both the outer and the inner leaflet of the microsomal vesicles revealed a similar distribution indicating that both leaflets share the same molecular species. The in vitro incorporation of radioactive fatty acids (16:0, 18:1 and 20:4(n-6] into ethanolamine phospholipids, known to be catalyzed by the lyosphosphatidylethanolamine acyl transferase, showed that the radioactive diacylglycerophosphoethanolamine molecules appeared first in the outer leaflet and were thereafter transferred to the inner leaflet. The apparent rate of translocation of the newly synthesized ethanolamine phospholipid molecules was the highest for those labelled with 16:0 and the lowest for those labelled with 20:4(n-6). The results indicate that the active site of the acyl-CoA:lysophosphatidylethanolamine acyltransferases is located on the outer leaflet of the microsomal vesicles and that the different newly synthesized molecular species of diacylglycerophosphoethanolamine may be translocated from the outer to the inner leaflet at different rates.

Familial invasive breast cancers: worse outcome related to BRCA1 mutations.

PURPOSE: Although all studies confirm that BRCA1 tumors are highly proliferative and poorly differentiated, their outcomes remain controversial. We propose to examine, through a cohort study, the pathologic characteristics, overall survival, local recurrence, and metastasis-free intervals of 40 patients with BRCA1 breast cancer. PATIENTS AND METHODS: A cohort of 183 patients with invasive breast cancer, treated at the Institut Curie and presenting with a familial history of breast and/or ovarian cancer, were tested for BRCA1 germ-line mutation. Tumor characteristics and clinical events were extracted from our prospectively registered database. RESULTS: Forty BRCA1 mutations were found among the 183 patients (22%). Median follow-up was 58 months. BRCA1 tumors were larger in size (P =.03), had a higher rate of grade 3 histoprognostic factors (P =.002), and had a higher frequency of negative estrogen (P =.003) and progesterone receptors (P =.002) compared with non-BRCA1 tumors. Overall survival was poorer for carriers than for noncarriers (5-year rate, 80% v 91%, P =.002). Because a long time interval between cancer diagnosis and genetic counseling artificially increases survival time due to unrecorded deaths, the analysis was limited to the 110 patients whose diagnosis-to-counseling interval was less than 36 months (19 BRCA1 patients and 91 non-BRCA1 patients). The differences between the BRCA1 and non-BRCA1 groups regarding overall survival and metastasis-free interval were dramatically increased (49% v 85% and 18% v 84%, respectively). Multivariate analysis showed that BRCA1 mutation was an independent prognostic factor. CONCLUSION: Our results strongly support that among patients with familial breast cancer, those who have a BRCA1 mutation have a worse outcome than those who do not.

In vitro and in vivo ethanolamine metabolism in rat brain: effect of time and aging.

The effect of the time in culture of foetal rat neurons and age on the incorporation of radioactive ethanolamine into methylated derivatives was investigated. Decreased incorporation of [3H]ethanolamine into its various methylated water-soluble and lipidic derivatives was observed in rat neurons cultures at 12 day in vitro (DIV) as compared to the 3rd and the 7th DIV. In vivo studies showed that there was a diminished labeling of methylated products in the older animals as compared to the younger ones. These in vitro and in vivo observations suggest a generalized decrease of N-methyltransferase activities during maturation and aging.

Breast cancer in elderly women: a retrospective analysis of combined treatment with tamoxifen and once-weekly irradiation.

PURPOSE: To evaluate retrospectively the efficacy of combined modality treatment (hormone therapy and hypofractionated radiotherapy) in a population of very elderly women with breast cancer. METHODS AND MATERIALS: Records on 70 patients of median age 81 years, treated between January 1988 and February 1994, whose median follow-up is now 36 months, have been evaluated. Information obtained included clinical stage at diagnosis, histology, tumor grading, hormone receptor levels, details of treatment, type of failure, survival data, and status at last follow-up examination. Treatment consisted of Tamoxifen 20 mg daily and a hypofractionated course of high dose-per-fraction once-weekly radiotherapy. In the majority of cases this consisted of seven exposures of 6.5 Gy (five to the involved breast, and two to the tumor bed) given over 6 weeks, on a 60Co unit. Nodes were treated when clinically involved, to a dose of 27.5-30 Gy in five to six fractions. RESULTS: At median follow-up of 36 months, the overall survival rate is 87% [confidence interval (CI) 78-95%], the disease specific survival rate is 88% (CI 80-96%), and 72% (CI 60-84%) of patients are free of disease. The local control rate at 36 months is 86% (CI 76-95%). When analyzed by T stage, 81% of T1 patients, 96% of T2 patients, 60% of T3 patients and, paradoxically 100% of T4 patients were in local control at 36 months, although at that point there were just four such patients available for consideration in the T4 group. Initial response to hormone therapy does not appear to be a predictive indicator for ultimate loco-regional control. There is a trend towards greater probability of loco-regional failure if total dose delivered to the breast is less than 35 Gy. CONCLUSIONS: Women of elderly age are often denied combined modality therapy, because of coexistant disease or fears held by the responsible physicians that elderly patients are unable to tolerate surgery or protracted courses of radiotherapy. Consequently, many are treated by tamoxifen alone with poor results. This study demonstrates that very high rates of loco-regional control are achievable using hormonal treatment combined with high dose-per-fraction once-weekly radiotherapy.

Survival and regeneration of adult human and other mammalian photoreceptors in culture.

PURPOSE: Fully mature neurons of central nervous system origin generally are considered unable to survive for extended periods of time in simple culture conditions. The authors report that adult and aged human, porcine, and rodent retinal neurons, including rod and cone photoreceptors, constitute an exception to this idea. METHODS: Cells were dissociated from human postmortem retinas, adult mammalian retinas, and selected brain regions and were seeded into tissue culture plates and left to develop as monolayer cultures for up to 2 months. A battery of antibody markers was used to identify the nature and morphology of the cells in vitro. RESULTS: Photoreceptor cell survival of rods and cones was observed routinely when the delay between the time of death until culture preparation was 50 hours or less, compatible with current eye bank practice. Two-week-old cultures were formed of rod photoreceptors, representing approximately 50% of neuronal cell types; cone photoreceptors, representing 5% to 30% of neuronal cell types; other retinal neurons (especially amacrine cells approximately 20%); and retinal glial cells, present in variable numbers. Glial cells were essential for long-term photoreceptor survival and neurite outgrowth. Adult mammalian brain neurons isolated under the same conditions did not survive. CONCLUSIONS: Fully adult human and other mammalian retinal neurons, including photoreceptors, exhibit remarkable plasticity in vitro, and such monolayer models may have applications in physiological, pharmacologic, and toxicologic studies of human and other mammalian retina.

Adult human retinal neurons in culture: Physiology of horizontal cells.

PURPOSE: Adult postmortem human retinal neurons in long-term monolayer cultures were recorded to characterize the voltage- and transmitter-gated currents in putative human horizontal cells (HCs). METHODS: Enzymatically and mechanically dissociated human retinal cells were seeded on polylysine and laminin- coated coverslips. Cells were identified by immunocytochemistry with cell type-specific antibodies and recorded with the patch-clamp technique. RESULTS: Immunostaining and responses to voltage steps confirmed the survival of various retinal cell types. Horizontal cells were identified by their specific glutamate-modulated anomalous rectifier K+ current conductance. This identification was further confirmed by subsequent immunolabeling of dye-labeled recorded cells with an anti-parvalbumin antibody that selectively stained HCs in frozen human retinal sections. Horizontal cells generated voltage-gated currents classically observed in HCs from fish to mammals: a transient outward K+ current, a sustained outward K+ current, and an L-type (Ca2+ current. Na+ currents were observed in only a few HCs. As in other species, glutamate, gamma-aminobutyric acid (GABA), and glycine generated responses mediated by the activation of kainate/(RS)-alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA), GABA(A), and glycine receptors, respectively. CONCLUSIONS: Various human retinal cell populations survive in vitro as indicated by immunolabeling with specific cell markers and by the diversity of responses to voltage steps. Human HCs exhibited extensive physiological similarities to HCs from other vertebrate species and a maintained expression of parvalbumin. These results constitute a comprehensive analysis of voltage- and transmitter-gated currents in a primate retinal neuron and validate the use of long-term monolayer culture of adult human neurons as a novel in vitro model for the study of human vision.

Simplified ganglioside composition of photoreceptors compared to other retinal neurons.

PURPOSE: The quantitative and qualitative ganglioside composition of retinal photoreceptor cells is unknown. The aim of this study was to analyze the lipid, especially ganglioside, make-up of photoreceptors compared to other retinal cells. METHODS: Retinas from adult normal rats were mechanically separated into outer (photoreceptors) and inner (other retinal neurons and glia) halves be planar vibratome sectioning. Total lipids were extracted, and each fraction (neural, phospholipids, and glycosphingolipids) was eluted sequentially by column chromatography and quantitated through high-performance thin layer chromatogram analysis. Similar analyses were performed on entire retinas from adult normal rats, adult dystrophic rats lacking photoreceptors (RCS-rdy-p+ strain), and isolated photoreceptor outer segments. RESULTS: Whereas phospholipids were distributed equally between the two halves, inner retina contained significantly more cholesterol (68% total) and gangliosides (74% total) than outer retina on a unit protein basis. The distribution on a percent molar basis of specific gangliosides also was significantly different between the two halves: Outer retina was dominated by GD3 (45% total ganglioside) and contained only trace amounts (<4%) of complex species (GT1b and GQ1b); inner retina was more typical of mature brain tissue exhibiting substantial amounts (approximately 25%) of more complex species. These data were supported by lipid compositional analyses of mutant photoreceptor-less retina. However, isolated outer segments resembled whole retina in containing higher levels of complex gangliosides. CONCLUSIONS: These data indicate that, compared to other central nervous system-derived neurons, photoreceptor cell body membranes exhibit a highly unusual simplified ganglioside composition. Such an unusual neuronal lipid composition may reflect structural adaptations to their specialized function.

Differential distribution of dystrophins in rat retina.

PURPOSE: Duchenne muscular dystrophy is frequently associated with a reduced amplitude of b-wave under scotopic conditions in the electroretinogram. This suggests that the dystrophin gene-encoded proteins play a role in retinal neurotransmission. The abnormal neurotransmission has been attributed to altered expressions of C-terminal products of the dystrophin gene in the outer plexiform layer, where photoreceptor cells form synapses with secondary neurons. The present study was undertaken to determine the cellular distribution of each member of the dystrophin superfamily in rat retina. METHODS: Examined in the study were the developmental pattern of dystrophins in rat retinae that exhibit inherited progressive photoreceptor degeneration; dystrophins messengers expression in the outer and the inner retina of normal rats, prepared by mechanical fractionation through the outer plexiform layer; and immunolocalization of dystrophin proteins and utrophin in normal and degenerated adult rat retinae, with several antibodies prepared against specific regions of the dystrophin superfamily. RESULTS: The results showed that Dp260 is exclusively localized in photoreceptor cells; Dp140 seems to be present in perivascular astrocytes; the exon 78 spliced isoform of Dp71 and the unspliced form are located in Müller glial cells and in perivascular astrocytes, respectively. Müller glial cells also contain utrophin. CONCLUSIONS: Although the role of these membrane cytoskeletal proteins remains to be elucidated in retina, the results support the hypothesis that b-wave reduction may be caused by molecular anomalies of C-terminal products of the dystrophin gene expressed in both neuron and glial cells.

Monosialoganglioside GM1 reduces ischemia--reperfusion-induced injury in the rat retina.

PURPOSE: Gangliosides are normal components of cell membranes, contribute to structural rigidity and membrane function, and have been shown to protect against various insults to the brain. This study evaluates the effect of exogenously administered monosialoganglioside GM1 on retinal damage induced by transient retinal ischemia and reperfusion. METHODS: Retinal ischemia was induced unilaterally in Long Evans rats by increasing intraocular pressure to 160 mm Hg for 60 minutes. GM1 (30 mg/kg, intraperitoneally) or buffer controls were administered at 48 hours, and 15 minutes before ischemia, and survival time after ischemia was either 8 or 15 days. The degree of retinal damage was assessed by histopathologic study according to Hughes' quantification of ischemic damage. RESULTS: Retinal ischemia led to significant reductions in thickness and cell number, principally in the inner retinal layers (30% to 80%), and to a lesser extent in the outer retinal layers (18% to 42%). Pretreatment with intraperitoneally injected monosialoganglioside GM1 conferred significant protection against retinal ischemic damage either 8 or 15 days after ischemic survival time. After 8 days reperfusion, the ischemic-induced loss in overall retinal thickness was reduced by 70%, and those of the inner nuclear and plexiform layers were reduced by 77% and 44%, respectively. Ischemic-induced ganglion cell, inner nuclear, and outer nuclear layer cell density losses were reduced by 45%, 40%, and 57%, respectively. After 15 days of reperfusion, approximately the same statistically significant differences could be observed in comparison with the 15-day ischemic--reperfusion group. CONCLUSIONS: Monosialoganglioside GM1 protects the rat retina from pressure-induced ischemic injury when administered intraperitoneally 2 days before insult. This protection afforded by GM1 can be observed even after 8 days or 15 days of reperfusion.

Selective excitotoxic degeneration of adult pig retinal ganglion cells in vitro.

PURPOSE: Excitotoxicity is proposed to play a prominent role in retinal ganglion cell (RGC) death ensuing from diseases such as glaucoma and ischemia, but cell culture studies have used tissue from newborn rodents, yielding conflicting data that implicate either N-methyl D-aspartate (NMDA) or non-NMDA glutamate (Glu) receptor-mediated pathways. Excitotoxic RGC death was examined in vitro in this study, using adult pigs, a large-animal model for human retina. METHODS: Adult pig retina (and for comparative purposes young and adult rat retina) were dissociated and maintained in monolayer culture. Medium was supplemented with Glu or pharmacologic agonists or antagonists, and surviving RGCs and other retinal neurons were quantified using specific immunolabeling methods. Electrophysiological responses to externally applied Glu of RGCs in culture were recorded using whole-cell patch-clamp techniques. RESULTS: Application of Glu led to selective, dose-dependent losses in large RGCs (maximal 37% decrease at 1 mM; median effective dose [ED50], approximately 80 microM) and neurite damage in surviving RGCs. Application of Glu agonists and Glu receptor subclass antagonists showed that large RGC death was mediated through both NMDA and non-NMDA receptor pathways. Small RGCs, amacrine cells, and all other retinal neurons were resistant to Glu-induced death. By comparison, rat retinal cultures displayed heightened RGC vulnerability to Glu, mediated exclusively by non-NMDA receptor-mediated pathways. Amacrine cells were unaffected by NMDA but were very sensitive to kainate application (>90% loss). Other retinal neurons were unaffected by any treatment. CONCLUSIONS: The molecular pathways underlying excitotoxic RGC death in vitro (non-NMDA or NMDA-preferring Glu receptors) vary among species and developmental stages. The selective elimination of adult pig large RGCs by NMDA receptor-mediated pathways more closely resembles human and animal glaucoma in vivo than other published culture models, providing a simplified experimental system for investigating the pharmacologic and toxicologic bases of glaucoma-like neuronal death.

Glial cell line-derived neurotrophic factor induces histologic and functional protection of rod photoreceptors in the rd/rd mouse.

PURPOSE: To evaluate the neuroprotective potential of glial cell line-derived neurotrophic factor (GDNF) in the retinal degeneration (rd/rd) mouse model of human retinitis pigmentosa. METHODS: Subretinal injections of GDNF were made into rd/rd mice at 13 and 17 days of age and electroretinograms (ERGs) recorded at 22 days. Control mice received saline vehicle injections or underwent no procedure. At 23 days of age, retinas from treated and control mice were fixed and processed for wholemount immunohistochemistry using an anti-rod opsin antibody, and rod numbers were estimated using an unbiased stereological systematic random approach. Subsequent to counting, immunolabeled retinas were re-embedded and sectioned in a transverse plane and the numbers of rods recalculated. RESULTS: Although ERGs could not be recorded from sham-operation or nonsurgical rd/rd mice at 22 days of age, detectable responses (both a- and b-waves) were observed in 4 of 10 GDNF-treated mice. Stereological assessment of immunolabeled rods at 23 days showed that control rd/rd retinas contained 41,880+/-3,890 (mean +/- SEM; n = 6), phosphate-buffered saline (PBS)-injected retinas contained 61,165+/-4,932 (n = 10; P < 0.001 versus control retinas) and GDNF-injected retinas contained 89,232+/-8,033 (n = 10; P < 0.001 versus control retinas, P < 0.002 versus PBS). This increase in rod numbers after GDNF treatment was confirmed by cell counts obtained from frozen sections. CONCLUSIONS: GDNF exerts both histologic and functional neuroprotective effects on rod photoreceptors in the rd/rd mouse. Thus rescue was demonstrated in an animal model of inherited retinal degeneration in which the gene defect was located within the rods themselves, similar to most forms of human retinitis pigmentosa. GDNF represents a candidate neurotrophic factor for palliating some forms of hereditary human blindness.

Selective transplantation of rods delays cone loss in a retinitis pigmentosa model.

BACKGROUND: Rod-cone retinal degenerations (retinitis pigmentosa) are typified by initial rod loss followed by secondary cone death. Rod death, predominantly caused by gene mutations expressed specifically in these cells, induces scotopic vision loss. Cone death, the overriding cause of blindness, has no current explanation. Disease progression and preliminary data suggest that cone survival depends on rods. OBJECTIVE: To establish whether rod transplantation into mutant rodless retinas could halt cone loss. METHODS: We transplanted pure sheets of rods isolated from normal-sighted mice into the subretinal space of recipient retinal degeneration mice lacking rods but possessing approximately 30% residual cones. Control animals were unoperated on or grafted with inner retinal cells from young normal donors, entire retinas from aged retinal degeneration mice, or gelatin. Two weeks after surgery, we quantified by an unbiased method the numbers of host retinal cones after immunolabeling with specific markers. RESULTS: Only mice receiving rod-rich transplants demonstrated statistically significant greater cone numbers, with rescue of 40% of host cones normally destined to die during this period. CONCLUSION: Cone survival depends specifically on rods. CLINICAL RELEVANCE: Such findings indicate that transplantation of rods could limit loss of cones, thus preserving useful vision in human retinitis pigmentosa. Arch Ophthalmol. 2000;118:807-811

Gangliosides and neurotrophic growth factors in the retina. Molecular interactions and applications as neuroprotective agents.

Polypeptide growth factors and gangliosides can both be considered as trophic agents involved in almost all stages of neural cell development, differentiation, survival, and pathology. In most cases their physiological roles are still not clear due to the considerable complexity in their regulation. Several growth factors [e.g., basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF)] and one species of ganglioside (GM1) have been shown to exert interactions with each other and also to exhibit neuroprotective effects against retinal ischemia in vivo and cerebral excitotoxicity in vitro. Different experimental models are used to investigate their relevance to ischemic and excitotoxic conditions in the retina, and it is shown that (1) both bFGF and EGF show very effective neuroprotection for rat retinal neurones exposed to toxic levels of glutamate or its nonphysiological agonist kainate in vitro; (2) GM1 (10(-5M) used under the same conditions does not afford protection; (3) retinal glial cells also suffer morphological perturbations following glutamate or kainate treatment, but this effect is dependent on neuron-glial interactions, indicating the existence of intermediate neuron-derived messenger molecules; (4) these glial changes can be corrected by posttreatment with either bFGF or EGF in vitro; (5) using an in vivo animal model involving anterior chamber pressure-induced ischemia in adult rats, it is shown that either pretreatment by intraperitoneal injection of GM1, or posttreatment by intraocular injection of the same ganglioside, reduces significantly histological damage to inner nuclear regions; and (6) in cultured retinal Müller glial cells the existence of molecular and metabolic interactions between both types of trophic factors is demonstrated. Hence both these groups of trophic molecules show interesting features for retinal ischemic treatment.