Kazuhiko Unoki

Cytochrome P450 1B1 gene mutations in Japanese patients with primary congenital glaucoma1

PURPOSE To report a novel missense mutation and DNA polymorphism of the CYP1B1gene in Japanese patients with primary congenital glaucoma. METHODS A series of 11 unrelated patients with primary congenital glaucoma was examined. Patients were followed in the Kagoshima University Hospital between 1979 and 1998. DNA was extracted from leukocytes of the patients, their families, and unrelated healthy individuals. Amplicons spanning the coding regions of the CYP1B1 gene were examined by direct sequencing and enzyme-restriction detection. RESULTS In the 11 unrelated patients, besides the previously reported insertional mutation (1620 ins G), a novel missense mutation was identified at codons 444 to replace arginine with glutamine (R444Q) in one patient. The novel missense mutation cosegregated in the relevant family as an autosomal recessive pattern and was not found in other patients or control individuals. In addition, five polymorphic sites were found at codons 48, 119, 330, 432, and 449. These polymorphic alleles did not cosegregate with the disease, and they were found in healthy individuals as well. CONCLUSIONS Approximately 20% of Japanese patients with primary congenital glaucoma may be affected by mutations in the CYP1B1 gene. Further studies are justified to explore whether a relationship exists between the phenotypic expressivity of the disease and the type of mutation.

Preferential recognition of synthetic peptides from HTLV-I gp21 envelope protein by HLA-DRB1 alleles associated with HAM/TSP (HTLV-I-associated myelopathy/tropical spastic paraparesis)

To determine CD4+ T-cell epitopes of HTLV-I-envelope protein recognized by the HLA alleles associated with HAM/TSP, we established 20 CD4+ T-cell lines from peripheral blood mononuclear cells (PBMCs) of naive healthy donors using a panel of synthetic peptides spanning the entire length of HTLV-I-envelope proteins, gp46 and gp21. We quantitated the precursor frequencies of HTLV-1-envelope specific CD4+ T-cells and analyzed epitope specificity in the context of HLA alleles. The precursor frequencies ranged from 3.0 to 10.6 per 10(7) PBMCs in the naive healthy donors. The CD4+ T-cell epitopes of HTLV-I-envelope protein were clustered in amino acids 76 to 90, 136 to 160, 171 to 185 and 196 to 210 of gp46, and in amino acids 366 to 400 and 436 to 485 of gp21. The CD4+ T-cell epitopes of gp21 were preferentially recognized by HLA-DRB1 0101 and 1502 which were known to be associated with HAM/TSP. Thus, it was suggested that HTLV-I gp21 might contain the major CD4+ T-cell epitopes recognized by HLA-DRB1 alleles of HAM/TSP.

Ultracytochemical demonstration of glycogen in cone, but not in rod, photoreceptor cells in the rat retina

The presence of native glycogen in photoreceptor cells of the rat retina has not been identified in the literature. We have studied this ultracytochemically. After perfusion with glutaraldehyde fixative, the eyes were enucleated, and the retinal tissues, postfixed with OsO4, were embedded in epoxy resin. Some tissues were treated with saliva before postfixation. Ultrathin sections, stained by the periodic acid-thiocarbohydrazide-silver proteinate (PA-TCH-SP) method or with uranyl acetate and lead citrate, were examined by electron microscopy. On routinely stained sections, glycogen particles seemed to be absent in the cytoplasmic matrix of the photoreceptor cells because they were indistinguishable from the numerous ribosomes. This was due to a similarity in size and electron density. After PA-TCH-SP staining, fine electron-dense reaction products appeared on small cytoplasmic particles (but not on ribosomes) in the inner segments, perikarya and synaptic terminals of a subpopulation of photoreceptor cells. These particles, 15-25 nm in diameter, were identified as beta-particles of glycogen because of their susceptibility to enzyme digestion. The glycogen-rich photoreceptor cells were thought to be cone cells by reasons of their morphological features, such as synaptic terminals, nuclei and outer segments. These results suggest that the cone, but not the rod, photoreceptor cells in the rat contain abundant glycogen.

Early structural changes during spontaneous closure of idiopathic full-thickness macular hole determined by optical coherence tomography: a case report

BackgroundSpontaneous closure of an idiopathic full-thickness macular hole has been reported to occasionally occur. However, the cells involved in plugging the macular hole have not been determined conclusively. We aimed to report the early structural changes that occur during a spontaneous closure of an idiopathic full-thickness macular hole determined by spectral-domain optical coherence tomography.Case presentationA 71-year-old Japanese man with an idiopathic full-thickness macular hole and subclinical posterior vitreous detachment in the left eye was followed. Three weeks after the identification of the macular hole, optical coherence tomography showed tissue that protruded from the interior wall of the macular hole at the level of the external limiting membrane toward the center of the macular hole. Five months after the first examination, he returned with improvements of his visual symptoms, and the macular hole was closed by a thin retinal tissue which included the restored external limiting membrane that bridged across the macular hole. However, the inner segment/outer segment junction line was not intact and the fovea was detached. Two months later, optical coherence tomography showed an almost normal foveal configuration with an essentially restored inner segment/outer segment junction line and foveal reattachment.ConclusionOur results suggest that Müller cells proliferate and/or extend at the level of the end of the external limiting membrane to form a tissue bridge across the macular hole associated with the external limiting membrane restoration first of all. This leads to the adhesion of other retinal layers and resolution of the foveal detachment.

Hyperreflective dots surrounding the central retinal artery and vein in optic disc melanocytoma revealed by spectral domain optical coherence tomography

PurposeTo report findings of optic disc melanocytoma (ODM) obtained using spectral domain optical coherence tomography (SD OCT), with special reference to the central retinal artery and vein surrounded by hyperreflective dots.MethodsRetrospective review of five eyes of five patients with ODM. Demographic information, ophthalmic examination including best-corrected visual acuity, dilated funduscopic examination, and SD OCT images were evaluated.ResultsDome-shaped, darkly pigmented tumors were seen ophthalmoscopically in the optic discs of all eyes. On OCT, the first branches of the central retinal artery and/or vein were well defined as oblique sections of tubular structures with a perivascular distribution of hyperreflective dots in the elevated retina (nerve fiber layer) over the tumor. The portions where these vessels turn toward the retina were displaced more anteriorly than those of eyes without ODM. Hyperreflective dots of various sizes were also observed in elevated retinas over the tumors, which shadowed and obscured the subjacent tissue in all eyes.ConclusionsSD OCT provides higher definition images of ODM relating to the branches of the central retinal artery/vein, revealing anterior displacement of vessels and perivascular distribution of hyperreflective dots that suggest melanophages and/or tumor cells or proteins and/or lipid deposits.

Midkine from Various Sources in Constant Light-Induced Photoreceptor Degeneration of the Rat

Midkine (MK) is a heparin binding growth factor that is expressed temporally during the early stages of retinoic acid-induced differentiation of embryonal carcinoma cells1,2. MK protein produced by L-cell which had been transformed stably with a mouse MK expression vector, enhances the survival and neurite outgrowth of cultured embryonic neurons and promote mitosis in some fibroblast cell lines3–5. In constant light induced retinal degeneration, we demonstrated that the intravitreal injection of MK from L-cell show significant rescue effects on photoreceptor damage by both histological and electrophysiological methods6,7. These results indicate that MK has neurotrophic activity or survival promoting activity in retina of the rat.

Specific binding of peanut agglutinin to foveal and peripheral cone photoreceptors of monkey retina.

peanut agglutinin, a lectin with specific affinity for galactose beta-(1----3)-N-acetyl-galactosamine disaccharides, showed binding to the slender cones in the rod-free fovea as well as to the conical cones in the periphery of the monkey (Macaca fuscata) retina, but not to the rods, as revealed by light-microscopic cytochemistry with horseradish-peroxidase-conjugated lectin.

Focal choroidal excavation with changes in shape and alterations of inner retina during long follow-up in an eye with polypoidal choroidal vasculopathy.

A focal choroidal excavation in the macula was first described by Jampol and collegaues in 2006. This clinical entity is characterised by an excavation of the choroid as detected by optical coherence tomography (OCT). Focal choroidal excavation is seen ophthalmoscopically as a yellowish spot or a pigmentary alteration and its appearance in the OCT images is relatively stable. It is occasionally associated with central serous chorioretinopathy, polypoidal choroidal vasculopathy and choroidal neovascularisation. The pathogenesis and natural history of focal choroidal excavation are not known mainly because of the short follow-up periods of the earlier studies and absence of histopathology. We followed a case of focal choroidal excavation associated with polypoidal choroidal vasculopathy for eight years and found features that have not been reported. We report the findings and discuss how they are related to the pathogenesis of focal choroidal excavation.

Glycohistochemical Study of Light-Induced Retinal Degeneration

We are interested in the physiological roles of the sialoglycoconjugates associated with rod photoreceptor cells, which specifically metabolize sialic acids by balancing sialyltransferase with sialidase (1, 2). Defects of O- and N-linked sialoglycoconjugates may cause retinal dysplasia and retinal degeneration, respectively, in humans (3). Retinal degeneration can be experimentally induced in rats by modification of sialic acids of retinal sialoglycoconjugates (4, 5), but the pathomechanism underlying this degenerative process remains to be clarified.

Distribution of Glycosyltransferase in Bovine Eyes

The activities of sialyl-, fucosyl- and galactosyltransferase were measured in membrane preparations from bovine eyes. Radioactive sugars from CMP-N-neuraminic acid, GDP-fucose and UDP-galactose were incorporated into the endogenous and exogenous acceptors. The enzymatic activities of all three glycosyltransferases were found to be high in the neural retina and retinal pigment epithelium, moderate in the cornea and uvea, and low in the lens and sclera, except for a high galactosyltransferase activity in the cornea.