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D A Saperstein

Publications and source records attributed to D A Saperstein.

8 recordsLinked to original sources
PubMed2001

Prevalence of mutations causing retinitis pigmentosa and other inherited retinopathies.

Inherited retinopathies are a genetically and phenotypically heterogeneous group of diseases affecting approximately one in 2000 individuals worldwide. For the past 10 years, the Laboratory for Molecular Diagnosis of Inherited Eye Diseases (LMDIED) at the University of Texas-Houston Health Science Center has screened subjects ascertained in the United States and Canada for mutations in genes causing dominant and recessive autosomal retinopathies. A combination of single strand conformational analysis (SSCA) and direct sequencing of five genes (rhodopsin, peripherin/RDS, RP1, CRX, and AIPL1) identified the disease-causing mutation in approximately one-third of subjects with autosomal dominant retinitis pigmentosa (adRP) or with autosomal dominant cone-rod dystrophy (adCORD). In addition, the causative mutation was identified in 15% of subjects with Leber congenital amaurosis (LCA). Overall, we report identification of the causative mutation in 105 of 506 (21%) of unrelated subjects (probands) tested; we report five previously unreported mutations in rhodopsin, two in peripherin/RDS, and one previously unreported mutation in the cone-rod homeobox gene, CRX. Based on this large survey, the prevalence of disease-causing mutations in each of these genes within specific disease categories is estimated. These data are useful in estimating the frequency of specific mutations and in selecting individuals and families for mutation-specific studies.

Amino Acid Substitution↗
PubMed2001

Gene transfer mediated by recombinant baculovirus into mouse eye.

PURPOSE: To determine the efficiency of baculoviruses (BVs) to transfer recombinant genes in vivo into murine ocular tissues. METHODS: Recombinant (r)BVs carrying fluorescent protein (FP) cDNA under the control of cytomegalovirus (CMV) immediate early promoter were constructed. Initially, cultured HEK293 and ARPE19 cells were infected with these rBVs and analyzed for efficiency and stability of transgene expression. The rBV-CMV green (G)FP was also injected into the intravitreal and subretinal space of mouse eye. Mice were periodically analyzed to determine the efficiency and stability of expression by histologic examination under fluorescence microscopy. The effect of rBV-CMV-GFP on the physiology of the retina was analyzed by electroretinography. RESULTS: cDNAs encoding fluorescent proteins were efficiently transduced in HEK293 and ARPE19 cells in vitro. GFP expression in vivo was observed exclusively in retinal pigment epithelial (RPE) cells after subretinal injections. Intravitreal injections of rBV resulted in GFP expression in the corneal endothelium, lens, RPE, and retina. GFP expression was observed for up to 14 days after injection. The infiltration of macrophages, observed 2 days after injection in the area of GFP transduction, had dissipated by day 8 after injection. No alteration in ERG responses was observed 6 weeks after injection of rBV-CMV-GFP. CONCLUSIONS: BV efficiently transduces cultured RPE cells and many cell types in vivo in the eye, including endothelial, epithelial, and neuronal cells. BV may be a useful vector for transferring genes in cultured cells and in vivo into ocular tissue.

Animals↗
PubMed2000

Lim2(To3) transgenic mice establish a causative relationship between the mutation identified in the lim2 gene and cataractogenesis in the To3 mouse mutant.

PURPOSE: Lim2 is the gene encoding the ocular lens-specific intrinsic membrane protein MP19. We previously reported finding a single nonconservative G->T transversion in exon two of the Lim2 gene. This mutation was linked to the cataract in the To3 (Total opacity number 3) mouse mutant, confirming Lim2 as an ideal candidate gene for the To3 cataract. The aim of the present study was to substantiate a causative relationship between the mutation in the Lim2 gene and cataractogenesis in the To3 mouse mutant. To this end a Lim2To3 transgene cassette was engineered and introduced into fertilized normal mouse embryos to test its ability to induce cataractogenic lens development. METHODS: A Lim2 genomic clone was isolated and purified from a murine 129/SvJ genomic library. A restriction endonuclease map of the gene was generated using classical Southern techniques. The murine Lim2 promoter was characterized by transfecting primary chicken lens epithelial cells with Lim2 promoter-CAT reporter constructs and assaying promoter activity and specificity. This genomic clone was then used in conjunction with PCR to generate a Lim2To3 transgene cassette. After sequencing of the PCR engineered portion, the Lim2To3 transgene was then used to generate Lim2To3 transgenic mice via pronuclear injection. Founder mice and their offspring from outcrosses and intercrosses were characterized by ophthalmic examination, PCR and Southern DNA analysis, RT-PCR mRNA analysis, and histology of lens sections. RESULTS: Two mice, from independent microinjections, were identified as positive for presence of the Lim2To3 transgene cassette as well as presence of bilateral congenital cataracts and reduced eye size and mass. One of these founders was incapable of germline transmission of the transgene to offspring and was not characterized further. The other was capable of germline transmission and was characterized as described above. PCR DNA analysis revealed a perfect concordance between presence of the Lim2To3 transgene cassette and congenital cataract in offspring of this founder. Transgenic hemizygotes exhibited cataract and a reduction in eye and lens size and mass, while transgenic "homozygotes" presented with a more severe cataract and microphthalmic reduction in eye and lens size and mass. Southern analysis revealed approximately 2 copies of the transgene cassette integrated into a single chromosomal site in the founder and all hemizygous offspring. RT-PCR analysis revealed a very low ratio of Lim2To3 transgenic mRNA compared to endogenous normal Lim2. Finally, histology revealed that lens development was abnormal in mutant transgenic animals by embryonic day E15. By E19, just prior to birth, gross disorganization of secondary fibers was observed in mutants. CONCLUSIONS: These transgenic experiments firmly establish a causative relationship between the previously identified mutation in the Lim2 gene and cataractogenesis in the To3 mouse mutant. The low levels of mutant mRNA produced by the transgene cassette as compared to endogenous levels of normal Lim2 mRNA provides evidence that this dominant mutation results in a mutant MP19 protein with altered function rather than simply loss of function.

Animals↗
PubMed1997

Exogenous Neisseria meningitidis endophthalmitis.

PURPOSE: To report a case of Neisseria meningitidis endophthalmitis in association with a leaking filtering bleb and to consider antibiotic prophylaxis of those people with whom the patient had contact. METHOD: We treated an 81-year-old man who had a chronic, leaking filtering bleb and who developed exogenous N meningitidis endophthalmitis. RESULT: N meningitidis endophthalmitis was controlled with antibiotic therapy. Antibiotic prophylaxis for those with whom the patient had contact was not recommended. CONCLUSION: In this case, the N meningitidis strain was not considered invasive because the bacteria apparently entered the eye through a leaky filtering bleb and not through the bloodstream. Recovery of noninvasive N meningitidis does not require prophylaxis for patient contacts. In cases of endogenous or idiopathic N meningitidis endophthalmitis, antibiotic prophylaxis of close patient contacts may be warranted.

Aged↗
PubMed1991

Independent segregation of fundus albipunctatus and the transthyretin (prealbumin) gene.

A defect in the visual cycle has been suspected in patients with fundus albipunctatus (FALB), rendering genes encoding visual cycle components etiologic candidates. One such component is the retinoid and thyroxine transport protein transthyretin (TTR, prealbumin) which in the eye is synthesized only in the retinal pigment epithelium and is believed to play a role in retinal retinoid transport. The authors established polymerase chain reaction conditions that allow rapid assay of TTR alleles as defined by MspI and Fnu4HI restriction fragment length polymorphisms. In a candidate gene analysis of an affected family, they demonstrate independent segregation of the TTR and FALB disease loci. These results exclude the possibility that a TTR gene defect causes FALB in this family.

Alleles↗