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Curr Eye Res. Author manuscript; available in PMC 2009 July 29.
Published in final edited form as:
Curr Eye Res. 2009 May; 34(5): 395–400.
doi: 10.1080/02713680902859639.
PMCID: PMC2717950
NIHMSID: NIHMS125938
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Autofluorescence Imaging and Phenotypic variance in a Sibling pair with Early Onset Retinal Dystrophy due to defective CRB1 function
Joaquin Tosi,1 Ilene Tsui,2 Luiz Lima,3 Nan Kai Wang,1 and Stephen H. Tsang1
1 Bernard & Shirlee Brown Glaucoma Laboratory, Departments of Ophthalmology, Columbia University, New York, NY
2 Wellesley College
3 Vitreous, Retina, Macula Consultants of New York, and The LuEsther T. Mertz Retinal Research Center, Manhattan Eye, Ear, and Throat Hospital, New York, NY
Corresponding Author: S.H.Tsang
1Brown Glaucoma Laboratory, Department of Pathology & Cell Biology, College of Physicians and Surgeons, Columbia University, New York, NY 10032, Email: sht2/at/columbia.edu. Telephone: 001-212-342-1189. FAX: 001-212-305-4987
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Abstract
Purpose
To phenotype two siblings with autosomal recessive early onset retinal dystrophy due to CRB1 mutations.
Methods
Autofluorescence (AF) imaging, high resolution optical coherence tomography (OCT), and full-field electroretinography (ERG) were performed. The results of DNA sequencing from polymerase chain reaction (PCR) products of the CRB1 gene were obtained from hospital records.
Results
Two siblings, 14-years-old and 17-years-old, were compound heterozygote for 749 del Ser and C948Y mutations in the gene encoding CRB1. AF imaging documented the preservation of retinal pigment epithelium (RPE) along the arterioles. High resolution OCT showed abnormally thick retinae with increased lamination.
Conclusion
Leber congenital amaurosis caused by CRB1 is a unique form of early onset retinal dystrophy because it spares the para-arteriolar RPE and causes abnormal retinal lamination with thickening. These findings, detectable with AF imaging and high resolution OCT, can be combined with electrophysiology and genetic testing to molecularly classify retinal degenerations efficiently.
The Crumbs gene (CRB1) encodes a large transmembrane protein required for maintenance of apical-basal epithelial polarity and is involved in stalk membrane formation during photoreceptor morphogenesis (Pellikka et al., 2002). Mutations in CRB1, the human homolog of Drosophila Crumbs, cause autosomal recessive retinitis pigmentosa (Richard et al., 2006); (den Hollander et al., 2001). Clinically, it is known to manifest with sparing of para-arteriolar retinal pigment epithelium (Vallespin et al., 2007), Coats-like exudative vasculopathy, and as an early onset retinal dystrophy (also known as Leber congenital amaurosis) (Massey-Harroche et al., 2007).
Herein, we describe genotype-phenotype correlations of two patients with CRB1 RP using recently available imaging techniques.
Genetic Analyses
Patients’ and their parents’ DNA were screened on the arRP array (http://www.asperophthalmics.com/ARRPgenetest.htm) containing mutant alleles of 8 known arRP genes: AIPL1, CRB1, CRX, GUCY2D, RPE65, RPGRIP1, MERTK and LRAT (Asper Ophthalmics). The slides were imaged with the Genorama QuattroImager and the sequence variants were identified by Genorama genotyping software.
Segregation analysis showed that all family members are heterozygous carriers. Father is a heterozygote carrier for 749 del Ser, Mother is a heterozygote carrier for C948Y. Both Cases 1 and 2 are compound heterozygous for 749 del Ser and C948Y.
Case 1
A 14-year-old boy was referred for the evaluation of Crumbs associated early onset retinal dystrophy. He was genetically tested elsewhere and known to be a compound heterozygote for 749 del Ser and C948Y mutations. On exam, the patient had horizontal sensory nystagmus. His best corrected visual acuity was count fingers at 2 feet in the right eye and 20/200 in the left eye. Axial length was 21.37 mm in the right and 21.53mm in the left. The anterior segment examination was normal.
Dilated fundus examination showed slightly pale optic nerves, macular atrophy, and nummular pigment clumping in both eyes. There were also diffuse drusen like deposits in the posterior pole. There was no cystoid macular edema or Coats-like exudation. There was para-arteriolar RPE preservation observed funduscopically and demonstrated with AF (Figure 1Fig. 1). Cirrus-OCT (Carl Zeiss, Dublin, CA, USA) showed abnormal inner retinal lamination and loss of photoreceptors (Figure 1Fig. 1). There is an increased retinal thickness in the nasal retina compared to controls (Figure 3Fig. 3).
Fig. 1
Fig. 1
Fig. 1
Fundus photograph of a 14-year-old boy with early onset retinal dystrophy due to mutations in the CRB1 gene. Posterior pole shows prominent xanthophyll as well as macular pigment mottling. The disc looks fairly normal and blood vessels are slightly attenuated. (more ...)
Fig. 3
Fig. 3
Fig. 3
Increased nasal retinal thickness in patients with mutations in the CRB1 gene. The thickness of the total retina and the distance between RPE and NFL in the nasal and temporal boundary of the ring were measured using Photoshop caliper. The same parameters (more ...)
Full field ERG was performed using Dawson-Trick-Litzkow (DTL) recording electrodes and Ganzfeld stimulation according to International Society for Clinical Electrophysiology of Vision (ISCEV) standards (Figure 4Fig. 4). Scotopic responses were extinguished and the photopic cone responses were difficult to evaluate secondary to photomyoclonic reflex.
Fig. 4
Fig. 4
Fig. 4
Full-field electroretinogram (ERG) findings were consistent with severe generalized rodcone dysfunction affecting the rod system more than the cone system due to compound heterozygote 749 del Ser and C948Y mutations in the gene encoding CRB1. ISCEV standardized (more ...)
Case 2
The 17-year-old sister of the above patient was also a compound heterozygote for 749 del Ser and C948Y mutations, as confirmed with old records from another institution. She had horizontal sensory nystagmus and best-corrected acuity vision was 20/200 in the right eye and 20/400 in the left eye. Axial length was 19.98 mm in the right and 19.76 in the left. Anterior segment exam was unremarkable.
Dilated fundus examination showed optic nerve pallor, macular atrophy, and peripheral intraretinal pigment migration less severe than her brother (Figure 2Fig. 2). There were a few drusenoid deposits in the posterior pole. Fundus AF imaging showed RPE preservation around the arterioles. There is an increased retinal thickness in the nasal retina compared to controls (Figure 3Fig. 3).
Fig. 2
Fig. 2
Fig. 2
Right eye fundus photograph of a 17-year-old girl with early onset retinal dystrophy due to mutations in the CRB1 gene. Posterior pole exhibits macular atrophy and pigment mottling. The disc shows some pallor. Peripheral intraretinal pigmentation, RPE (more ...)
Full field ERGs showed generalized retinal dysfunction affecting rods more than cones (Figure 4Fig. 4). Scotopic rod specific ERG appeared to be extinguished. Maximal ERG a- and b- wave amplitudes were 70 microvolts and 86 microvolts in the right; 51 microvolts and 78 microvolts in the left. Transient photopic ERG b-wave amplitudes were 28 microvolts in the right and 37 microvolts in the left. Burian-Allen contact lens electrodes were used for 30 Hz flicker ERG and obtained the values of 25 microvolts 30 Hz flicker response. Transient photopic ERG obtained with Burian-allen contact electrodes showed 40 microvolts in the right eye.
Gene specific therapeutics for early onset retinal dystrophy patients are currently undergoing clinical trials (Jacobson et al., 2003); (Jacobson et al., 2006); (Downs et al., 2007); (Koenekoop et al., 2007); (Bainbridge et al., 2008); (Maguire et al., 2008). The key to gene-based treatments is efficient and accurate genotyping (Stone, 2007a); (Stone, 2007b). Therefore, it is essential to phenotype and genotype individuals with retinal dystrophies for genetic counseling and enrollment into treatment trials. The CRB1 gene is known to function in eye development and mutations result in the loss of photoreceptor signaling. CRB1 mutant retinae are thicker and have a coarse lamination pattern similar to patterns observed during stages of normal human retinal development (Stone, 2007a), as opposed to other inherited retinal degenerations. CRB1 gene product interacts with members of the peripheral membrane-associated guanylate kinase (MAGUK) family membrane protein known as palmitoylated 5 protein (MPP5), MPP3 and MPP4 to form a scaffold at the retinal outer limiting membrane. (Kantardzheiva et al., 2006; Kantardzheiva et al., 2005; Stöhr et al., 2003; van Rossum et al., 2006).
Additionally, in this sibling pair, the younger brother manifested more severe retinal dysfunction than the older sister. This demonstrates variance in expressivity of compound heterozygotes for 749 del Ser and C948Y mutations in the gene encoding CRB1 gene.
AF is most commonly used to study peripapillary and macular RPE. To our knowledge, this is the first application of AF imaging that shows para-arteriolar RPE preservation in Crumbs associated RP. Another distinct clinical feature of CRB1 mutant retinae is the abnormally thicker retina with a coarse lamination pattern that appears on OCT.
In summary, this case small series uses fundus AF and high resolution OCT to demonstrate para-arteriolar sparing of RPE and abnormal retinal lamination. These clinical features, distinguishing Crumbs RP from other retinal dystrophies, should prompt screening for the CRB1 gene. Molecular diagnosis of RP is an important tool in the era of emerging therapeutic options.
Acknowledgments
We are grateful to Neeco Palmer and the Photography Department of Columbia University for their support. SHT is a Burroughs-Wellcome Program in Biomedical Sciences Fellow, and is supported by the Charles Culpeper Scholarship, Foundation Fighting Blindness, Hirschl Trust, Schneeweiss Stem Cell Fund, Joel Hoffmann Foundation, Jonas Family Fund, Jahnigen-Hartford-American Geriatrics Society, Eye Surgery Fund, Crowley Research Fund, and Bernard Becker-Association of University Professors in Ophthalmology-Research to Prevent Blindness (RPB).
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