Levels of complement factor H-related 4 protein do not influence susceptibility to age-related macular degeneration or its course of progression.

Dysregulation of the alternative pathway (AP) of the complement system is a significant contributor to age-related macular degeneration (AMD), a primary cause of irreversible vision loss worldwide. Here, we assess the contribution of the liver-produced complement factor H-related 4 protein (FHR-4) to AMD initiation and course of progression. We show that FHR-4 variation in plasma and at the primary location of AMD-associated pathology, the retinal pigment epithelium/Bruch's membrane/choroid interface, is entirely explained by three independent quantitative trait loci (QTL). Using two distinct cohorts composed of a combined 14,965 controls and 20,741 cases, we ascertain that independent QTLs for FHR-4 are distinct from variants causally associated with AMD, and that FHR-4 variation is not independently associated with disease. Additionally, FHR-4 does not appear to influence AMD progression course among patients with disease driven predominantly by AP dysregulation. Modulation of FHR-4 is therefore unlikely to be an effective therapeutic strategy for AMD.

A single EFEMP1 mutation associated with both Malattia Leventinese and Doyne honeycomb retinal dystrophy.

Malattia Leventinese (ML) and Doyne honeycomb retinal dystrophy (DHRD) refer to two autosomal dominant diseases characterized by yellow-white deposits known as drusen that accumulate beneath the retinal pigment epithelium (RPE). Both loci were mapped to chromosome 2p16-21 (refs 5,6) and this genetic interval has been subsequently narrowed. The importance of these diseases is due in large part to their close phenotypic similarity to age-related macular degeneration (AMD), a disorder with a strong genetic component that accounts for approximately 50% of registered blindness in the Western world. Just as in ML and DHRD, the early hallmark of AMD is the presence of drusen. Here we use a combination of positional and candidate gene methods to identify a single non-conservative mutation (Arg345Trp) in the gene EFEMP1 (for EGF-containing fibrillin-like extracellular matrix protein 1) in all families studied. This change was not present in 477 control individuals or in 494 patients with age-related macular degeneration. Identification of this mutation may aid in the development of an animal model for drusen, as well as in the identification of other genes involved in human macular degeneration.

Variations in the complement regulatory genes factor H (CFH) and factor H related 5 (CFHR5) are associated with membranoproliferative glomerulonephritis type II (dense deposit disease).

INTRODUCTION: Membranoproliferative glomerulonephritis type II or dense deposit disease (MPGN II/DDD) causes chronic renal dysfunction that progresses to end stage renal disease in about half of patients within 10 years of diagnosis. Deficiency of and mutations in the complement factor H (CFH) gene are associated with the development of MPGN II/DDD, suggesting that dysregulation of the alternative pathway of the complement cascade is important in disease pathophysiology. SUBJECTS: Patients with MPGN II/DDD were studied to determine whether specific allele variants of CFH and CFHR5 segregate preferentially with the MPGN II/DDD disease phenotype. The control group was compromised of 131 people in whom age related macular degeneration had been excluded. RESULTS: Allele frequencies of four single nucleotide polymorphisms in CFH and three in CFHR5 were significantly different between MPGN II/DDD patients and controls. CONCLUSION: We have identified specific allele variants of CFH and CFHR5 associated with the MPGN II/DDD disease phenotype. While our data can be interpreted to further implicate complement in the pathogenesis of MPGN II/DDD, these associations could also be unrelated to disease pathophysiology. Functional studies are required to resolve this question.

An integrated hypothesis that considers drusen as biomarkers of immune-mediated processes at the RPE-Bruch's membrane interface in aging and age-related macular degeneration.

Age-related macular degeneration (AMD) is a blinding disease that afflicts millions of adults in the Western world. Although it has been proposed that a threshold event occurs during normal aging which leads to AMD, the sequelae of biochemical, cellular, and/or molecular events leading to the development of AMD are poorly understood. Although available data provide strong evidence that a significant proportion of AMD has a genetic basis, no gene(s) has yet been identified that causes a significant proportion of AMD. Moreover, no major molecular pathways involved in the etiology of this disease have been elucidated.Drusen, pathological deposits that form between the retinal pigmented epithelium (RPE) and Bruch's membrane, are significant risk factors for the development of AMD. In our view, the development of testable new hypotheses of drusen origins has been hindered significantly by the absence of a comprehensive profile of their molecular composition. In this review, we describe an integrated ultrastructural, histochemical, molecular biological, and biochemical approach to identify specific molecular pathways associated with drusen biogenesis. The implicit assumption underlying these recent investigations has been that a thorough understanding of the composition of drusen and source(s) of drusen-associated material is likely to provide fresh insight into the pathobiology underlying AMD. Significantly, these studies have revealed that proteins associated with inflammation and immune-mediated processes are prevalent among drusen-associated constituents. Transcripts that encode a number of these molecules have been detected in retinal, RPE, and choroidal cells. These data have also lead to the observations that dendritic cells, potent antigen-presenting cells, are intimately associated with drusen development and that complement activation is a key pathway that is active both within drusen and along the RPE-choroid interface. We propose herein a unifying hypothesis of drusen biogenesis that attempts to incorporate a large body of new and previously published structural, histochemical, and molecular data pertaining to drusen composition and development. This theory is put forth with the acknowledgment that numerous AMD genotypes may exist. Thus, only some aspects of the proposed hypothesis may be involved in any given AMD genotype. Importantly, this hypothesis invokes, for the first time, the potential for a direct role of cell- and immune-mediated processes in drusen biogenesis. We acknowledge that the proposed hypothesis clearly represents a paradigm shift in our conceptualization pertaining to pathways that participate in the development of drusen and age-related macular degeneration. It is our hope that other investigators will test, validate and/or refute various aspects of this hypothesis, and in so doing, increase our overall understanding of the biological pathways associated with early AMD.

Characterization of glucose transporter isoforms in the adult and developing human eye.

The expression of glucose transporter isoforms (Glut 1, Glut 3, Glut 4, and Glut 5) in the human eye was investigated at various ages ranging between 8 weeks gestation (first trimester) and adult using Western blot and immunohistochemical analyses. Glut 1 and Glut 3 expression and cellular localization patterns were similar to those of human brain. Glut 1 (50-kilodalton protein) was expressed by epithelial cells (retinal pigmented epithelium, choroidal, iridial, and pars planus), which form the blood-eye barrier, retinal Mueller cells, the lens fiber cells, iridial microvascular endothelial cells, and to a lesser extent by the outer segments of the photoreceptor cells in the adult eye. This pattern was conserved throughout development and was evident as early as 8 weeks gestation. In addition, the endothelial cells of vitreous hyaloid vessels expressed Glut 1 at 8 weeks gestation. Glut 3 (50 to 55-kilodalton protein) immunoreactivity was observed only in the adult inner synaptic layer of the retina. Neither Glut 4 nor Glut 5 was expressed in any occular tissue at any age examined. These results suggest that Glut 1 is the main glucose transporter of the human eye and that it is ontogenically conserved. In contrast, Glut 3 is associated with selective neuronal processes, and its expression is developmentally altered.

Expression and characterization of the IPM 150 gene (IMPG1) product, a novel human photoreceptor cell-associated chondroitin-sulfate proteoglycan.

The interphotoreceptor matrix (IPM) occupies the extracellular space between the apical surface of the retinal pigmented epithelium and the external limiting membrane of the neural retina. This space contains two chondroitin sulfate proteoglycans, designated IPM 150 and IPM 200, which are likely to effect retinal adhesion and photoreceptor survival. In an effort to characterize human IPM 150, several cDNA clones encoding its core protein have been isolated from a human retinal cDNA library. Translation of overlapping cDNA sequences yields a novel core protein with a predicted molecular mass of 89.3 kDa. Northern and dot-blot analyses as well as the isolation of expressed sequence tags demonstrate that IPM 150 mRNA is expressed not only in the neural retina but also in several other non-ocular tissues. In situ hybridization analyses indicate that, in the eye, IPM 150 mRNA is expressed specifically by cone and rod photoreceptor cells. Characterization of IPM 150 proteoglycan core protein and identification of its site of synthesis are important steps towards understanding the architecture and biology of the IPM.

Biochemical characterization of the major peanut-agglutinin-binding glycoproteins in vertebrate retinae.

Peanut agglutinin (PNA), a lectin that binds D-galactose-beta (1----3) N-acetyl-D-galactosamine disaccharide linkages, selectively labels cone photoreceptors in the retinae of a variety of species. PNA binds consistently to domains of the interphotoreceptor matrix associated with cone, but not rod, inner and outer segments, to cone cell body and axonal membranes, to cone synaptic pedicles, and to portions of the inner plexiform layer. In order to begin the characterization of the molecular species responsible for cone-specific PNA binding, chick, turkey, rat, dog, pig, monkey, and human retinal extracts were separated by SDS-polyacrylamide gel electrophoresis and probed with peroxidase-conjugated PNA. The results reveal the presence of six major groups of PNA-binding glycoproteins ranging from 30 to 88 kilodaltons. Most of these are shared by the seven species examined; however, some interspecies variation is present. Three groups, designated GP39/40, GP42/45, and GP60, are the most intensely labeled by PNA and are common to all species analyzed, while groups GP29/31 and GP88 are less intensely labeled and are present in most but not all of the species investigated. Labeling of the GP54 group is variable but is most consistently associated with extracts of rat and pig retinae. Trypsin treatment, which results in the loss of cone-associated PNA binding in the interphotoreceptor matrix, causes a visually detectable reduction in three of the six groups of PNA-binding glycoproteins in porcine retinal extracts. Of these, GP54 is the most sensitive, being undetectable on PNA-stained blots after only 5 minutes of enzyme exposure; GP88 and GP45 are less sensitive but both are markedly reduced after 15 minutes of trypsinization. Trypsin-sensitive molecules thus may be involved in the establishment of the cone-specific domains of interphotoreceptor matrix identified by PNA binding. These, as well as the other groups of PNA-binding molecules, are being utilized to develop more specific immunologic probes with which to further study of their distribution and function.

Ultrastructural visualization of primate cone photoreceptor matrix sheaths.

Glycoconjugates, including glycolipids, glycoproteins, and proteoglycans, are present in the plasma membrane of photoreceptor cells and in the interphotoreceptor matrix surrounding photoreceptor cell ellipsoids and outer segments. Although the precise function of these molecules is unknown, they may be important in mediating photoreceptor-pigment epithelial cell interactions, outer segment membrane assembly, and/or disc shedding. Lectins, affinity ligands for defined carbohydrate sequences, have proven particularly useful in studying the glycoconjugate composition of the interphotoreceptor matrix. The peanut lectin selectively binds to domains of the interphotoreceptor matrix surrounding cone ("cone matrix sheaths"), but not rod inner and outer segments. This is evidence for the existence of chemical and structural heterogeneity within the interphotoreceptor matrix. The studies described herein utilized ultrastructural pre-embedding histochemical labeling to assess whether, in addition to the surrounding interphotoreceptor matrix, peanut lectin binding is associated directly with that plasma membrane of cone inner and outer segments. This study confirms that ferritin-conjugated peanut agglutinin binds to cone matrix sheaths, and, in addition, provides ultrastructural evidence for the presence of binding to the plasma membrane surrounding cone inner and outer segments. The data suggest that cone membrane-associated peanut agglutinin-binding molecules may differ from those located within cone matrix sheaths.

Carbohydrate components recognized by the cone-specific monoclonal antibody CSA-1 and by peanut agglutinin are associated with red and green-sensitive cone photoreceptors.

Previous investigations have demonstrated that the monoclonal antibody CSA-1 and peanut agglutinin label specifically cone photoreceptor cells. In the present study, we compared the binding of CSA-1 and peanut agglutinin to that of the monoclonal antibodies COS-1 and OS-2, which have been shown to recognize the red/green- and blue-sensitive cone visual pigments, respectively. Using lectin and immunocytochemistry on serial semithin sections of the pig retina, we have demonstrated in the present study that both CSA-1 and peanut agglutinin label specifically the red-, and green-sensitive, but not the blue-sensitive cone cell outer segments. Peanut agglutinin does bind, however, to the cone matrix sheaths associated with all three types of cones. These observations support the idea that red-, and green-sensitive cone cells share some common molecular epitopes and may represent a differentiation line of cones, considerably different from that of blue-sensitive cones.

Vitronectin receptor expression and distribution at the photoreceptor-retinal pigment epithelial interface.

Laser scanning confocal microscopy was employed to map the distribution of integrin immunoreactivity at the photoreceptor-retinal pigment epithelial (RPE) interface of the primate retina, and to determine its relationship to the actin cytoskeleton. Immunolabeling using a polyclonal antibody to the human vitronectin receptor (VnR), a heterodimer containing the alpha v subunit in combination with either the beta 3 or beta 5 subunits, is detected primarily on the apical surface of the retinal pigment epithelium (RPE) in vivo and in vitro. It is also associated with the photoreceptor inner and outer segment cell surfaces. In contrast, immunolabeling using a polyclonal antibody to the human fibronectin receptor (FnR), a heterodimer containing the alpha 5 and beta 1 subunits, is detected principally on the basolateral surface of the RPE and is virtually absent in photoreceptors. A partial three-dimensional reconstruction of the anti-VnR labeling pattern in cone photoreceptors reveals cell surface labeling that originates at the level of the myoid just distal to the outer limiting membrane. It extends distally toward the ellipsoid and terminates at the level of the cone outer segment. Approximately 20-22 immunoreactive foci are distributed evenly around the perimeter of the cone ellipsoid. These foci correspond in number and location to the calycal processes that protrude from the distal portion of the ellipsoid. A double-labeling procedure, employing VnR antibody and a fluorescently labeled phallotoxin (phalloidin), was used to identify regions of VnR co-distribution with filamentous actin (F-actin). One such region includes the VnR-immunoreactive foci at the margins of the cone inner segments and the actin cables that course through the photoreceptor ellipsoid and terminate within the calycal processes. A second zone of co-distribution coincides with the actin-containing, circumferential bundle at the lateral borders of the RPE cells, and a third zone is associated with the apical microvilli of the RPE that ensheath cone outer segments. In order to help identify the specific subunits underlying VnR (alpha v beta 3/5) immunoreactivity, Northern blots of retinal-RPE RNA were probed with alpha 32P-cDNAs to the human alpha v, beta 3, and beta 5 subunits and additional immunolocalization studies were performed using integrin human alpha or beta subunit-specific antisera. The results from these studies strongly suggest that one or more integrins, containing the alpha v and/or beta 5 subunits, are expressed by the photoreceptors and RPE.(ABSTRACT TRUNCATED AT 400 WORDS)

Human ocular drusen possess novel core domains with a distinct carbohydrate composition.

Ocular drusen are extracellular deposits that form between the retinal pigmented epithelium (RPE) and Bruch's membrane. Although the presence of large and/or numerous drusen in the macula is a significant risk factor for development of age-related macular degeneration (AMD), a major cause of irreversible blindness, little is known about their origin or composition. We have expanded on our previous investigations related to drusen-associated glycoconjugates by examining lectin binding patterns after removal of terminal sialic acid residues. Strikingly, intense and distinct labeling of drusen subdomains is revealed by Arachea hypogea agglutinin (PNA) after neuraminidase treatment. PNA binding is confined to discrete domains within both hard and soft drusen. These "cores" are positioned centrally within drusen and are typically juxtaposed to Bruch's membrane. Only one core per druse is observed. PNA labeling of drusen cores does not co-localize with associated lipids and is abrogated by digestion with O-glycosidase but not N-glycosidase. The association of cores with small drusen suggests that they may participate in drusen biogenesis. (J Histochem Cytochem 47:1533-1539, 1999)

Immunohistochemical localization of carbonic anhydrase IV in capillaries of rat and human skeletal muscle.

We used polyclonal antisera raised in rabbits against membrane-bound rat lung and human lung carbonic anhydrase (CA) IV in immunofluorescence studies to stain cryosections of rat soleus and extensor digitorum longus (EDL) and several human skeletal muscles. There was strong specific staining of capillaries in all muscles investigated. Several techniques were applied to verify this result. (a) Serial sections were either incubated with anti-CA IV/FITC or processed for endothelial ATPase reaction. There was precise co-localization of antibody marked structures and ATPase stained capillaries. (b) Human muscle sections were double stained with anti-CA IV/TRITC and anti-von Willebrand factor (vWF)/FITC. vWF, a capillary marker, and CA IV were localized at identical sites. (c) The CAIV was released from capillaries by treatment with phosphatidylinositol specific phospholipase C, suggesting that the enzyme is anchored to the endothelial cell membrane via a phosphatidylinositolglycan anchor. (d) A rat hindlimb was perfused with diluted antiserum. Cryosections of perfused soleus and EDL processed for anti-rabbit IgG/FITC staining showed clear fluorescence associated with capillaries, indicating that the antigen was accessible from the capillary lumen. (e) Immune complexes formed during antiserum perfusion as described in d were precipitated from muscle homogenates. SDS-PAGE followed by immunoblotting showed that the predominant portion of total muscle CA IV was bound in these complexes and therefore must be located intravascularly.

Structural and compositional analyses of isolated cone matrix sheaths.

The cone matrix sheath is a biochemically and structurally distinct domain of the retinal interphotoreceptor matrix that is specifically associated with cone photoreceptor cell inner and outer segments. An aqueous extraction technique for the isolation of cone matrix sheath-enriched retinal fractions was developed and used to examine the structure and composition of this extracellular matrix domain. A complex substructural organization of the cone matrix sheath was observed. Many longitudinally oriented, filamentous structures extend the entire length of the sheath and terminate in filamentous rings at both the apical and basal ends. These longitudinal filaments are interconnected by a finer, anastomosing filamentous network. The basal rings of cone matrix sheaths are interconnected with similar rings of matrix material associated with rod photoreceptor inner segments. Gel electrophoresis and lectin blot analyses of cone matrix sheath-enriched fractions reveal the presence of 17- and 32-kilodalton bands, labeled by peanut lectin, that are likely to be components of the structural elements of cone matrix sheaths. Thus, structural elements potentially capable of mediating adhesion between the neural retina and the retinal pigmented epithelium are present in the cone matrix sheath and may be at least partially responsible for the observation that cone matrix sheaths are firmly adherent to the pigmented epithelium and the neural retina.

Characterization of molecules bound by the cone photoreceptor-specific monoclonal antibody CSA-1.

The cone-specific monoclonal antibody, CSA-1, selectively labels cone photoreceptors in the neural retina. Immunofluorescence assays reveal that CSA-1 binds cone outer and inner segments, cell bodies, axons and synaptic pedicles. The binding of the antibody is membrane-associated, not cytoplasmic. Several lines of evidence, including sensitivity to mild alkaline hydrolysis and galactosidic enzymes, suggest that the CSA-1 detected determinant is a galactose-containing carbohydrate moiety. Immunoblotting analyses show CSA-1 immunoreactivity with at least two molecules of 54,000 daltons that differ slightly in isoelectric point, and an additional, higher molecular weight species. CSA-1 should prove to be of value in studies of cone photoreceptor biology, in analyses of the development and differentiation of these cells, and in assessing their fate in various retinal pathologies.

Xyloside-induced disruption of interphotoreceptor matrix proteoglycans results in retinal detachment.

Unique domains of the retinal interphotoreceptor matrix (IPM), termed cone matrix sheaths, are composed largely of chondroitin 6-sulfate proteoglycan in most higher mammalian species. Recent investigations suggest that cone matrix sheaths participate in the maintenance of normal retinal attachment. To investigate the potential functional roles of IPM proteoglycans further, the synthesis of cone matrix sheath chondroitin 6-sulfate proteoglycan was perturbed in vivo. Intravitreal injections of p-nitrophenyl-beta-D-xylopyranoside (xyloside), a sugar that inhibits chondroitin sulfate proteoglycan synthesis, were administered to Yucatan micropigs. Their eyes were examined funduscopically and electroretinographically. At selected times, the eyes were enucleated and examined histochemically and immunohistochemically with various probes directed against cone photoreceptor cells and cone matrix sheaths. The IPM was affected selectively after xyloside administration; no inner retinal pathology or dysfunction was detected morphologically or electroretinographically. The degree of xyloside-induced perturbation was dependent on the duration of xyloside exposure and dose. It was classified into three stages, based on morphologic and histochemical criteria. Although all three stages could be observed in a given retina, a single stage typically predominated, depending on the particular dosage regimen. The early stage was characterized by IPM disruption, as evidenced by disorganization of chondroitin 6-sulfate and peanut agglutinin (PNA)-binding glycoconjugates. Cone photoreceptor cell outer segment degeneration and markedly decreased chondroitin 6-sulfate immunoreactivity distinguished the middle stage. During the late stage, there was a near complete absence of both immunoreactive chondroitin 6-sulfate and PNA-binding glycoconjugates in the IPM. Shallow retinal detachments that appeared funduscopically as patches of retinal whitening frequently were observed after moderate durations of xyloside exposure; these progressed peripherally with continued xyloside exposure. Histologically, the areas of retinal whitening corresponded to regions in which cone matrix sheaths were split transversely (ie, in a plane perpendicular to the longitudinal axes of the photoreceptor cell outer segments) or were separated completely from cone outer segments. Similar effects were not observed in control eyes. These results suggest that adhesion between the neural retina and retinal pigmented epithelium may be dependent, in part, on continuous synthesis of cone matrix sheath-associated proteoglycans and, potentially, other IPM proteoglycans. In addition, these proteoglycans appear to be necessary for the maintenance of cone photoreceptor cell outer segment integrity.

Retinal adhesiveness is weakened by enzymatic modification of the interphotoreceptor matrix in vivo.

The role of interphotoreceptor matrix (IPM) constituents in mediating adhesion between the retina and retinal pigment epithelium (RPE) was investigated by injecting specific enzymes into rabbit eyes either intravitreally or subretinally. Retinal adhesiveness was measured by peeling the retina from the pigment epithelium 1-3 days later and observing the amount of adherent pigment. Effects of enzymes on the IPM were monitored by observation of peanut agglutinin (PNA) binding to cone matrix sheaths; retinal and RPE toxicity was excluded by electroretinography and histology. Three enzymes that degrade glycosaminoglycans or saccharides known to be constituents of the IPM (chondroitinase ABC, neuraminidase, and testicular hyaluronidase) both weakened adhesion and altered PNA binding, although the effects on the cone matrix sheaths were different for each enzyme. An enzyme specific for hyaluronic acid (Streptomyces-derived hyaluronidase), which has not been identified as a major IPM constituent, had no effect on either adhesion or PNA binding. The authors conclude that IPM-associated glycoconjugates participate in retinal adhesion, although their precise composition, interaction with IPM components, and relationship to other mechanisms of adhesion remain to be determined.

Distribution of glycoconjugates in the human retinal internal limiting membrane.

The internal limiting membrane (ILM), or basal lamina, of the neural retina is located between the end feet of retinal Müller cells and the vitreous cortex. Recent studies of ILM substructure and histochemistry suggest that the ILM lamina rara externa, a region of the ILM likely to mediate vitreoretinal adhesion, is rich in noncollagen glycoconjugates. To characterize the spatial distribution and temporal expression of ILM glycoconjugates further, fetal and adult human retinas were examined using lectin histochemistry. Eyes from early fetal (13-15 weeks of gestation), midfetal (17-20 weeks of gestation), late fetal (23-26 weeks of gestation), full-term (38-41 weeks of gestation), early adult (17-20 yr of age), and late adult (57-84 yr of age) stages were examined. Although a number of significant observations pertaining to the distribution and temporal expression of ILM-associated glycoconjugates are described, two may have a significant relationship to vitreoretinal adhesion. In contrast to laminin and fibronectin, which are detectable throughout the retinal ILM at both young and older adulthood, erythrina cristagalii (ECA), a lectin with high binding affinity for galactose beta (1,3) N-acetyl-glucosamine, binds strongly to the ILM of young eyes but does not bind to the ILM of eyes from older donors. In addition, dolichos bifloris (DBA), a lectin with high binding affinity for N-acetylgalactosamine, does not bind to the ILM at early fetal stages, but binds at the midfetal stage, a time which corresponds with the onset of tertiary vitreous elaboration. More significantly, the DBA-binding glycoconjugate is neural retina ILM specific; it is not present in the basal lamina of the ciliary epithelium. Based on the known oligosaccharide composition of nonocular forms of human laminin, fibronectin, and collagen type I, and on the distribution of these molecules in the basal laminas of both the retina and ciliary epithelium, the authors conclude that the molecule bound by DBA has not been identified previously as a component of the ILM. Further characterization of ILM-associated molecules will be important to understand vitreoretinal adhesion and clinical disorders where adhesion is anomalous.

Interphotoreceptor matrix domains ensheath vertebrate cone photoreceptor cells.

The retinal interphotoreceptor matrix (IPM) occupies the space between the neural retina and the retinal pigmented epithelium (RPE), two neuroectoderm-derived epithelia. While the IPM appears to be a major route by which photoreceptor cells receive vital metabolic factors, relatively little is known concerning its structure and function. The studies reported here describe the presence of specialized domains of the IPM that ensheath cone, but not rod, inner and outer segments in pig, monkey, and human retinae. These cone extracellular matrix sheaths are chemically and structurally distinct from the remainder of the IPM as revealed by their specific binding of the lectin peanut agglutinin (PNA) and their structural stability during physical dissociation of the retina. Biochemical studies suggest that the PNA-binding components of the cone matrix sheaths are trypsin-sensitive glycoproteins. These structures may play a role in establishing a specialized microenvironment for cone photoreceptors, maintaining proper orientation of cone outer segments, and/or facilitating cone-RPE interactions.

Retinal adhesiveness in the monkey.

PURPOSE: To determine factors that influence retinal adhesion in the primate and compare these with previous data from the rabbit. METHODS: Retinal adhesiveness was studied in monkey eyes immediately after enucleation. The retina was peeled manually from the retinal pigment epithelium, and the amount of pigment that remained adherent to the retina was used as an index of adhesiveness. RESULTS: The rate of post mortem failure of retinal adhesiveness in the monkey was less than in the rabbit under similar conditions. However, as in the rabbit, adhesiveness was sensitive to temperature, pH, and the concentrations of calcium and magnesium, and subretinal injections of neuraminidase weakened adhesion beyond the injection sites. CONCLUSIONS: Mechanisms of retinal adhesion are similar in primates and rabbits.

Recovery of retinal adhesion after enzymatic perturbation of the interphotoreceptor matrix.

Previous investigations established that focal subretinal injections of neuraminidase, chondroitinase, and hyaluronidase in the rabbit lead to a diffuse loss of retinal adhesiveness beyond the site of injection. This loss of adhesiveness, measured by peeling of the retina immediately after enucleation, correlates with changes in the interphotoreceptor matrix (IPM), as monitored by lectin histochemistry. In this study, rabbits were evaluated during recovery of retinal adhesiveness after subretinal injections of neuraminidase and chondroitinase. Adhesion recovered steadily 5-20 days after chondroitinase injection. After administration of neuraminidase, adhesion remained low for approximately 14 days but recovered to normal by 20 days. The recovery of adhesiveness correlated closely with reestablishment of the normal distribution of peanut agglutinin-binding glycoconjugates in the IPM, one group of molecules thought to participate in retinal adhesion. Electroretinography and light microscopy showed no abnormalities in the retina or retinal pigment epithelium after recovery. These results suggest that IPM glycoconjugates participate in maintaining retinal adhesion.