Kenji Kitahara

Induction of nestin, Ki-67, and cyclin D1 expression in Müller cells after laser injury in adult rat retina.

PurposeThe purpose was to examine the expression of nestin, Ki-67, and cyclin D1 in Müller cells after laser injury in adult rat retina.MethodsThe right eyes of adult Brown Norway rats were treated with laser photocoagulation. The eyes were removed 3, 7, and 14 days after laser treatment. The retinas were investigated immunocytochemically by confocal microscopy. Agarose-embedded sections were immunostained with antibodies to nestin, vimentin, glial fibrillary acidic protein (GFAP), glutamate-aspartate transporter (GLAST), rhodopsin, Ki-67, and cyclin D1. Cell death was examined using terminal deoxynucleotidyl transferase-mediated uridine 5′-triphosphate-biotin nick end labeling (TUNEL) assay on agarose sections.ResultsNestin expression was induced in Müller cells following laser injury. In addition, Ki-67 and cyclin D1 expression was found in the nuclei of Müller cells after the treatment. TUNEL assay demonstrated that Müller cells were not labeled; hence these cells were not apoptotic.ConclusionsThese results suggest that dedifferentiation and proliferation of Müller cells can be induced by laser injury in adult rat retina.

Dichromatism in macaque monkeys.

Old World primates have trichromatic vision because they have three types of cone photoreceptor, each of which is maximally sensitive to short, middle or long wavelengths of light1. Although a proportion of human males (about 8% of caucasians, for example) have X-chromosome-linked colour-vision abnormalities2, no non-human Old World primates have been found to be colour-vision defective3,4. We have tested 3,153 macaque monkeys but found only three dichromats, a frequency that is much lower than in humans.Habitat loss and fragmentation remain the greatest threats to the worlds biodiversity. The local extinction of plant species from habitat fragments is common, although the reasons for this are not fully understood. Fragmentation is known to influence both birth- and death-related processes, but the disruption of plant reproduction, especially pollination and seed production, is thought to be particularly important. The effects of fragmentation on post-pollination processes such as seed dispersal and germination have rarely been explored experimentally. Here I show that seeds planted in forest fragments are less likely to germinate than those in continuous forest. This finding can have negative demographic consequences because it reduces the emergence of seedlings.

Vision: Dichromatism in macaque monkeys

Old World primates have trichromatic vision because they have three types of cone photoreceptor, each of which is maximally sensitive to short, middle or long wavelengths of light. Although a proportion of human males (about 8% of caucasians, for example) have X-chromosome-linked colour-vision abnormalities, no non-human Old World primates have been found to be colour-vision defective. We have tested 3,153 macaque monkeys but found only three dichromats, a frequency that is much lower than in humans.

CYP4V2 Mutations in Two Japanese Patients with Bietti’s Crystalline Dystrophy

Bietti’s crystalline dystrophy (BCD) is an autosomal-recessive retinal dystrophy characterized by numerous glistening intraretinal dots scattered over the fundus, particularly in the posterior pole. The purpose of this study was to report mutations in the CYP4V2 gene (encoding a ubiquitously-expressed 525-amino acid sequence belonging to the CYP450 family) and to investigate the impact of the mutation on pre-mRNA splicing. DNA and RNA analyses were conducted using blood samples from two unrelated Japanese patients with BCD (a 46-year-old female and a 52-year-old male). In the female patient, a homozygous deletion/insertion mutation (g.IVS6–8_–1delc.802_810del/insGC) including the 3´-acceptor splice site was identified. Reverse transcription-PCR analysis revealed that the complete length of exon 7 (186 bp), is skipped, resulting in the in-frame deletion mutation (p.V268_E329del). Conversely, the male patient was a compound heterozygote for the deletion/insertion and novel nonsense (p.W340X) mutations. Clinically, the female patient had decreased visual acuity, constriction of visual fields, severely reduced amplitudes in both rod and cone electroretinograms (ERGs). Despite being 6 years older, the male patient presented with milder clinical manifestations having good visual acuity and substantial amplitudes in both rod and cone ERGs. Because the CYP4V2 truncated protein with the p.W340X mutation lacks 186 amino acids at the C-terminus, if expressed, it retains 62 amino acids encoded in exon 7, which are important for enzymatic activity. In the male patient, expression of both mutant alleles may compensate for the malfunction of each mutated protein and could explain why a milder form of BCD results from compound heterozygosity.

MR tractography with diffusion tensor imaging in clinical routine

Using MRI, we demonstrated that the depiction of the cerebral white matter fiber tracts has become a routine procedure. Diffusion tensor (DT) sequences may be analyzed with combined volume analysis and tractography extraction software, giving indirect visualization of white matter connections. We obtained DT data from 20 subjects with normal MR imaging and five patients presenting cerebral diseases such as brain tumors, multiple sclerosis and stroke, with five patients explored on two different MR scanners. Data were transferred to dedicated workstations for anatomical realignment, determination of voxel eigenvectors and calculation of fiber tract orientations in a region of interest. In all subjects, axonal directions underlying the main neuronal pathways could be delineated. Comparisons between diseased regions and contralateral areas demonstrated changes in voxel anisotropy in injured regions, revealing possible preferential fiber orientations within diffuse T2 hyperintensities. Rapid data processing allows imaging of the normal and diseased fiber pathways as part of the routine MRI examination. Therefore, it appears that whenever white matter disease is suspected a tractography can be performed with this fast and simple method that we proved to be reliable and reproducible

Autosomal dominant familial exudative vitreoretinopathy in two Japanese families with FZD4mutations (H69Y and C181R)

Background:Familial exudative vitreoretinopathy (FEVR) is a hereditary disorder characterized by impaired vascularization of parts of the peripheral retina. Autosomal dominant FEVR (adFEVR), a major form of FEVR and assigned to chromosome 11q13–23 (EVR1) locus, is caused by deletion mutations in the C-terminal region of the frizzled-4 (FZD4) gene. This paper describes the clinical phenotype of adFEVR in two Japanese families with two different mutations in the FZD4gene. Methods:We encountered three Japanese patients with adFEVR and studied them using mutation analysis of the FZD4gene with PCR, sequencing, and a restriction enzyme digestion. Results:Two previously unreported missense mutations, p.H69Y and p.C181R, were identified in the N-terminal extracellular region of two of the patients. This region was highly conserved among other vertebrate species and FZD family members, unlike the C-terminal region. Co-segregation analysis revealed that all affected individuals carried one of these mutations, while unaffected individuals did not. The mutations were not detected in normal individuals (n = 120). The affected individuals had mild to severe retinal abnormalities. Conclusions: FZD4mutations in either the N- or C-terminal region underlie adFEVR, which indicates that FZD4plays an important role in retinal angiogenesis. Analysis of FZD4mutations in families with adFEVR is useful for genetic counseling and for early diagnosis.

Compound heterozygous CNGA3 mutations (R436W, L633P) in a Japanese patient with congenital achromatopsia

Congenital achromatopsia is a stationary retinal disorder with autosomal recessive inheritance that is characterized by loss of color discrimination, low visual acuity, photophobia, and nystagmus. This disorder has been shown to be associated with CNGA3, CNGB3, and GNAT2 mutations, and the frequency of mutations in the CNGA3 gene (encoding alpha subunit of the cone-specific cGMP-gated cation channel) was 23-33% in European populations. The aim of this study was to test the hypothesis that CNGA3 mutations are also responsible for congenital achromatopsia in Japanese patients. DNA from venous blood samples from a total of 14 patients from 13 Japanese pedigrees was prepared. Mutation screening of the CNGA3 gene was performed using direct sequencing and PCR-single-strand conformation polymorphism analysis. Compound heterozygous missense mutations (p.R436W and p.L633P, the latter of which was novel) were identified in one patient only, a 22-year-old female. Neither of these two mutations was found in 150 Japanese control individuals. The patients parents and sister carried one of these mutations each but were not affected. No mutations in the CNGB3 or GNAT2 genes were identified in the patient. Clinically, best-corrected visual acuity was 0.1 in both eyes. No specific findings were obtained in funduscopy. Optical coherence topography revealed a normal foveal thickness but a 20% decrease in parafoveal thickness. Ganzfeld full-field electroretinograms (ERGs) showed normal responses in rod and mixed rod-plus-cone ERGs but no response in cone or 30-Hz flicker ERGs. Spectral sensitivity on a white background revealed a curve with only one peak at around 500 nm, which fits the absorption spectrum of human rhodopsin. L633, conserved among vertebrate orthologs of human CNGA3, is a hydrophobic residue forming part of the carboxy-terminal leucine zipper (CLZ) domain, which is functionally important in the mediation of intracellular interactions. To our knowledge, this is the first report of a Japanese complete achromat with CNGA3 mutations, and of any patient with a missense mutation within the CLZ domain. The outcome suggests low frequency (7%, 1/14) of CNGA3 mutations in Japanese patients.

An Analysis of the Farnsworth-Munsell 100-Hue Test

For the analysis of the bipolarity patterns by the Farnsworth-Munsell 100-Hue test, the entire colour circle was divided into four, i.e. from quadrant I to IV. The sub-total of the error score of each quadrant was calculated by revolving the divided line for one cycle.

Variations in long- and middle-wavelength-sensitive opsin gene loci in crab-eating monkeys

We analyzed variations in long (L)- and middle (M)-wavelength-sensitive opsin gene loci in crab-eating monkeys. Unlike humans, most monkeys have a single L and a single M gene. Two variant genotypes, one with only one opsin gene (dichromatic) and one with tandemly arrayed multiple genes, were also found in the monkeys. However, the frequency of the former was 0.47%, and that of the latter was 5% in the monkeys, while 2% and 66%, respectively, in Caucasian males. The two variants were found only in Java Island, Indonesia, and South Thailand, respectively. The data suggest that the frequency of each genotype is different among Old World primates.

Tenon's Capsule Anaesthesia for Cataract Surgery with IOL Implantation*

Objective: To evaluate a small dosage of Tenons capsule anaesthesia as a less invasive substitute for retrobulbar anaesthesia in cataract surgery and intraocular lens (IOL) implantation. Study Design: Replacing the retrobulbar anaesthetic technique with the Tenons capsule anaesthetic technique from April 1990. Setting: The Jikei University School of Medicine Department of Ophthalmology. Patients: Over 1500 patients requiring cataract surgery were given the Tenons capsule method of anaesthesia. Main Outcome Measures: To show that complications attributable to retrobulbar anaesthesia would be less frequent with the Tenons capsule method. Results: There is no suppression of ocular motility and no pain caused by the anaesthesia. Neither is there an elevation of blood pressure or arrhythmia. There are no complications either in the method of anaesthesia, or in the surgery. There is a definite sedative effect obtained with a small dosage (0.5–1.0 cc), and this effect is prolonged for 60 minutes or longer. Conclusion: Tenons capsule anaesthesia is a very useful substitute for conventional retrobulbar anaesthesia, and is easy for inexperienced surgeons to use.