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M L Katz

Publications and source records attributed to M L Katz.

At least 19 recordsLinked to original sources
PubMed2004

Comparison of conventional and microwave-assisted processing of mouse retinas for transmission electron microscopy.

Conventional fixation and processing of mammalian retinal tissues for transmission electron microscopic (TEM) examination is slow and produces ultrastructural artefacts in the photoreceptor cell layer. Among these artefacts are gaps between photoreceptor outer segment disc membranes and between photoreceptor cells in the region of the retina where the cell nuclei are located. A study was undertaken to determine whether a much more rapid microwave-assisted fixation and processing protocol would have an effect on the quality of ultrastructural preservation of the retina, particularly on the photoreceptor cell artefacts. The overall ultrastructural preservation of the retina was similar for the conventional and microwave-assisted techniques. However, the magnitudes of the photoreceptor artefacts were significantly reduced when microwave irradiation was used during primary fixation and processing. It is clear that, at least for the retina, employing microwave irradiation during specimen preparation for TEM results in superior ultrastructural preservation with a substantial reduction in the time required for sample preparation.

Animals↗
PubMed2001

Mouse gene knockout models for the CLN2 and CLN3 forms of ceroid lipofuscinosis.

The childhood neuronal ceroid-lipofuscinoses (NCLs) are autosomal-recessively inherited neurodegenerative disorders that result in severe cognitive decline and premature death. The genetic bases for a number of different forms of NCL, including those designated CLN2 and CLN3, have now been determined. However, the mechanisms by which the gene defects cause the disease pathology are not known and no effective treatments for these disorders have been developed. To provide tools for studying the mechanisms underlying the disease pathologies and for screening potential therapeutic interventions, work is under way to develop mouse models for the CLN2 and CLN3 disorders. Targeted gene replacement was used to generate mice in which the murine orthologue of the CLN3 gene has been knocked out. Mice that are homozygous for the Cln3 knockout allele develop a number of pathological features similar to those that occur in the human disorder. Among these are accumulation of autofluorescent lysosomal storage bodies, behavioural abnormalities, retinal degeneration, and premature death. On a mixed strain genetic background, the appearance of these symptoms was quite variable, suggesting that other genes can modify the effects of CLN3 mutations. Work to develop a similar mouse gene knockout model for the CLN2 disorder is well under way. Chimaeric mice have been developed with cells that carry an induced mutation in the mouse orthologue of the CLN2 gene that would prevent synthesis of a functional CLN2 protein in mice that are homozygous for the mutation. Mice will be developed that are homozygous for this mutation, and these animals will be evaluated for the development of pathologies similar to those that occur in the human disorder.

Aminopeptidases↗
PubMed2001

Assessment of dietary therapies in a canine model of Batten disease.

The neuronal ceroid lipofuscinoses (NCLs) are inherited neurodegenerative diseases that occur in a number of animal species, including dogs. A study was conducted to determine whether the resupply of nutrients lost in NCL English Setter dogs would modify the course of the disease. Carnitine and polyunsaturated fatty acids have been reported to be reduced in NCL English Setters. Therefore, the normal laboratory diets of NCL dogs were supplemented with carnitine, fish oil and corn oil and the disease progression was compared with that of an untreated litter mate. The following specific prognostic indicators of NCL were monitored: cognitive function, brain atrophy, brain glucose metabolism and lifespan. Carnitine, with or without lipid supplements, dramatically delayed the progression of cognitive decline in NCL dogs. When fish oil and corn oil only were supplied, brain atrophy was reduced. A combination of all three supplements preserved cognitive function and increased lifespan by 10%. However, brain glucose hypometabolism and cerebral atrophy were not reduced. The results in this study indicated that the effectiveness of therapeutic interventions can be assessed by non-invasive methods at a relatively early stage of the disease process. Our study suggests that dietary supplementation with carnitine is a promising new approach for delaying or preventing the cognitive decline in dogs, and perhaps, with human NCL patients.

Animals↗
PubMed2001

Effect of Rpe65 knockout on accumulation of lipofuscin fluorophores in the retinal pigment epithelium.

PURPOSE: In all mammalian species examined to date the retinal pigment epithelium (RPE) has been found to accumulate autofluorescent lysosomal storage bodies (lipofuscin) during senescence. Substantial evidence indicates that retinoids in the RPE-retina complex play a major role in RPE lipofuscin formation. Indeed, at least one RPE lipofuscin fluorophore is derived in part from vitamin A aldehyde. However, the precise mechanisms by which retinoids modulate RPE lipofuscin accumulation have not been elucidated. In mice without a functional Rpe65 gene, isomerization of all-trans- to 11-cis-retinol is blocked. Experiments were performed to determine whether this impairment of retinoid metabolism alters RPE lipofuscin accumulation. METHODS: RPE lipofuscin fluorophore content was compared in 12- to 13-month-old Rpe65(+/+), Rpe65(+/-), and Rpe65(-/-) mice. Lipofuscin fluorophore content was determined using quantitative fluorometric measurements. RPE lipofuscin content was also estimated with quantitative ultrastructural techniques. RESULTS: In the Rpe65(-/-) mice, RPE lipofuscin fluorophore accumulation was almost abolished. In addition, a significantly reduced accumulation of lipofuscin fluorophores was also observed in the Rpe65(+/-) animals. The inability of the RPE of)Rpe65(-/-) mice to supply 11-cis-retinal from the RPE to the retinal photoreceptors was accompanied by a massive accumulation of lipid droplets in the RPE that appeared to contain substantial amounts of retinoids. CONCLUSIONS: These findings indicate that formation of RPE lipofuscin fluorophores is almost completely dependent on a normal visual cycle. The absence of retinal (both all-trans and 11-cis) in Rpe65 knockout mice drastically reduced formation of lipofuscin fluorophores in these animals. Even an excessive accumulation of retinyl fatty acid esters in the RPE of Rpe65 knockout mice did not contribute to lipofuscin accumulation.

Animals↗
PubMed2000

Histopathologic and immunocytochemical analysis of the retina and ocular tissues in Batten disease.

PURPOSE: To describe the pathophysiologic features of retinal degeneration in Batten disease (juvenile neuronal ceroid lipofuscinosis [JNCL]) caused by mutations in the CLN3 gene. STUDY DESIGN: Comparative human tissue study. MATERIALS: The retina and other ocular tissues of a 22-year-old man with JNCL were compared with the same tissues of a healthy 30-year-old man. DNA from whole blood and RNA from retina were used for genotype analysis. METHODS: The retinas, corneas, conjunctiva, and ciliary body were processed for histopathologic and immunofluorescence analysis. Genomic DNA was subjected to polymerase chain reaction (PCR) and nucleotide sequence analyses. Reverse transcriptase/PCR and sequence analysis were performed on retinal RNA. RESULTS: The JNCL donor was heterozygous for a approximately 1 kb deletion in CLN3, as found in most JNCL patients. The other allele had a single base pair deletion in exon 6 that resulted in a frame shift. Gross pathology of the JNCL retina resembled that in retinitis pigmentosa, including deposits of bone spicule pigment. Histopathologic studies revealed loss of neurons from all retinal layers. Immunofluorescence labeling with antibodies to rhodopsin, recoverin, and cone opsin demonstrated degenerate rods and cones with short outer segments in the far periphery. Autofluorescent lipopigment granules were prominent in ganglion cells and some cells of the inner nuclear layer, but not in the photoreceptors. The retinal pigment epithelium (RPE) had fewer lipofuscin granules than the control specimen. Increased numbers of lipofuscin granules were found in the epithelia of the ciliary body and conjunctiva, but not in the cornea of the JNCL eye. CONCLUSIONS: Immunofluorescence studies revealed degenerate rods and cones in the far periphery. Lipofuscin granules were decreased in the RPE, consistent with loss of photoreceptor outer segments. The novel finding that degenerate photoreceptors did not contain autofluorescent inclusions suggests that granule accumulation may not precede photoreceptor degeneration in JNCL. The presence of normal photoreceptor proteins in the degenerate rods and cones suggests that these cells may be capable of functional regeneration if a therapy for Batten disease is developed.

Adult↗
PubMed2000

Retinal vascular changes induced by the oxidative stress of alpha-tocopherol deficiency contrasted with diabetic microangiopathy.

It has been proposed that oxidative tissue damage is involved in the development of diabetic angiopathies. To evaluate this hypothesis, experiments were conducted to identify the retinal vessel changes induced by the oxidative stress related to alpha-tocopherol deficiency and examine possible similarities with the lesions characteristic of diabetic retinopathy. Twenty-one-day-old male Fisher 344 albino rats were divided randomly to receive a basal, chemically defined diet either with (adequate group) or without (deficient group) alpha-tocopherol. After 6 and 8 months, some rats (n = 3 per group) were killed and the eyes removed. In order to evaluate cell integrity and localization of lipofuscin-specific autofluorescence by light and fluorescence microscopy, some of the retinas were prepared for cryostat-sections while others were digested by elastase to isolate intact retinal vasculatures. After 8 and 14 months, the central retina of one eye per rat (n = 6 to 8 per group) was examined by electron microscopy for retinal capillary basement membrane (RCBM) thickening and other ultrastructural changes. At 6 and 8 months, the deficient rats exhibited extensive shortening and disarray of rod outer segments (ROS), marked loss of photoreceptor cells, and pronounced increases in the numbers of granules with lipofuscin-specific autofluorescence in the retinal pigment epithelium (RPE) and retinal vessels. At 14 months, the ultrastructure revealed that the damage to ROS involved disruption of membranes and that the capillary lipofuscin was contained mainly within the endothelial cells. Membrane remnants were found in the lipofuscin granules of both the RPE and retinal vessels. In addition, there was an increase in RCBM thickness (98.7 +/- 2.6 nm vs. 86.9 +/- 2.9 nm). RCBM thickening was the only finding common with diabetic retinopathy, and the thickening was 13.6%, significantly less than that reported in diabetic rat models with 8 and 14 months durations (34% and 53.1%, respectively). Capillary lipofuscin accumulation, which was prominent in the deficient rats, is not notable in diabetes. Both the moderate RCBM thickening and marked lipofuscin accumulations seen in alpha-tocopherol-deficient rats were similar to changes occurring in the aging process, though more pronounced. The spectrum of microangiopathies characteristic of diabetic retinopathy did not develop in alpha-tocopherol-deficient rats. These findings suggest that oxidative damage, though probably involved, is unlikely to play a predominant role in the development of diabetic retinal microangiopathies.

Animals↗
PubMed1999

Senescence of the retinal pigment epithelium.

Senescence of human cells has largely been studied as an in vitro phenomenon resulting from replicative exhaustion. The literature contains many studies of retinal pigment epithelium (RPE) cells which document replicative senescence. Several studies by Burke and others illustrate the relationship between donor age and replicative lifespan, the relationship between geographical location of RPE in the posterior pole and replicative lifespan, and the phenomena of altered cellular morphology and decreased culture saturation density for senescent RPE cells. Other studies have focused on the alterations of the expression of specific genes or the alteration of enzymatic activities during the senescence of RPE cells in vitro. Recently, a technique utilizing a histochemical staining procedure for beta galactosidase has been developed which identifies senescent cells. Normal beta galactosidase histochemistry which identifies the lysosomal form of the enzyme is performed at pH 4.0, while senescence-associated beta galactosidase activity is observed at pH 6.0 and is observed in the cytoplasm. We have studied the replicative senescence of human RPE cells in vitro using this procedure and have also measured the length of chromosomal telomeres to identify the aging of cultures in vitro. Our results show that RPE cultures accumulate beta galactosidase positive cells as a function of the number of population doublings and that these data correlate with the shortening of chromosomal telomeres to a functional limit observed for many human cell types at senescence. We have also recently extended this work to the development of a senescence-associated beta galactosidase procedure for observing senescent RPE cells in vivo. Basically, the same histochemical procedure is used with a post-staining bleaching step to clearly visualize staining in the RPE. Our first studies were performed on globes from Rhesus monkeys at a variety of ages from 1 year to 29 years of age. The results show the accumulation of beta galactosidase positive cells in the older monkey eyes. We have also examined several human eyes in an attempt to observe whether any relationship exists between beta galactosidase staining and age, pathology (diabetes, basal laminar deposits), and geographical location (macula vrs. periphery). These studies represent a first effort to determine if senescent RPE are present in vivo. It will be important to extend these studies so that these data might be expressed on a quantitative bases.

Aging↗
PubMed1999

A mouse gene knockout model for juvenile ceroid-lipofuscinosis (Batten disease).

The human hereditary ceroid-lipofuscinoses are a group of autosomal recessively inherited diseases characterized by massive accumulations of autofluorescent lysosomal storage bodies in the cells of many tissues and by neuronal degeneration throughout the central nervous system. There are a number of clinically and genetically distinct forms of ceroid-lipofuscinosis, the most common of which is the juvenile type, also known as Batten disease and CLN3. To study the mechanisms that lead to pathology in CLN3 and to evaluate potential therapies, a mouse model has been generated by targeted disruption of the mouse ortholog of the CLN3 gene (Cln3). As in affected humans, mice homozygous for the disrupted Cln3 allele show accumulation of autofluorescent storage material in neurons and other cell types. The storage material consists of membrane-bounded intracellular inclusions with ultrastructural features typical of the ceroid-lipofuscinoses. The accumulation of this storage material validates the Cln3 knockout mice as a model for the human disorder.

Animals↗
PubMed1999

Long-term variations in cyclic light intensity and dietary vitamin A intake modulate lipofuscin content of the retinal pigment epithelium.

Experiments were conducted to determine whether the intensity of cyclic light exposure to the retina over a long period of time affects retinoid-dependent accumulation of lipofuscin in the retinal pigment epithelium (RPE). Albino rats were maintained from weaning on diets either containing (+A) or lacking (-A) retinyl palmitate, which can be metabolized to the retinoids involved in the visual cycle. Animals in each dietary group were divided between bright (L) and dim (D) cyclic light treatments. Thus, the experiments employed the following four treatment groups: +A/D, +A/L, -A/D, and -A/L. After 6, 12, and 15 months from the start of the treatments, animals in each group were killed for quantitative determination of: 1) retinal photoreceptor densities; 2) RPE lipofuscin content; and 3) RPE lipofuscin fluorescence intensity. Animals in the L groups had a lower volume of RPE lipofuscin than those in the D groups fed the same diet. Among the -A rats, this reduced lipofuscin volume could be attributed to a light-enhanced depletion of vitamin A from the retina and an accompanying loss of photoreceptor cells. In the +A animals, however, there were no differences in photoreceptor densities between the D and L groups. In the -A rats, the volume of RPE lipofuscin decreased between 6 and 15 months of age, whereas it increased in the +A animals. In contrast, lipofuscin fluorescence intensity increased between 6 and 15 months of age in all four treatment groups. However, in the +A rats, the fluorescence intensity was lower in the L than in the D group at all three ages. In the -A groups, light level had no effect on lipofuscin fluorescence intensity. At all three ages, fluorescence intensity was lower in the -A animals than in +A rats. Thus, at light intensities below those that induce acute retinal degeneration, long-term exposure to higher intensity light inhibits the age-related increase in RPE lipofuscin volume. A decrease in the volume of RPE lipofuscin after the retina is depleted of vitamin A suggests that lipofuscin is turned over, and that RPE lipofuscin content is determined by a balance between the rates at which lipofuscin is formed and at which it is eliminated from the RPE. An age-related increase in lipofuscin-specific fluorescence intensity after vitamin A depletion from the retina suggests that lipofuscin fluorophores may continue to form slowly from retinoids that have been modified such that they can no longer enter the visual cycle.

Animals↗
PubMed1999

Characterization and chromosomal mapping of a mouse ortholog of the late-infantile ceroid-lipofuscinosis gene CLN2.

Late-infantile ceroid-lipofuscinosis (CLN2) is an autosomal recessively inherited, neurodegenerative disease in humans. The CLN2 locus has been mapped to Chromosome (Chr) 11p15, and its sequence and genomic organization have recently been reported. In the present study, the cDNA sequence, exon/intron organization, and chromosomal localization of a mouse ortholog of the CLN2 gene are described. The mouse cDNA contains an open reading frame that predicts a protein product of 562 amino acids. The mouse and human coding regions are 86% and 88% identical at the nucleic acid and amino acid levels, respectively. One less codon appears in the mouse cDNA when compared with the human ortholog. The mouse gene (Cln2) spans more than 6 kb and consists of 13 exons separated by introns ranging in size from 111 to 1259 bp. Length polymorphism in an (AC)(n) microsatellite in intron 3 of the mouse Cln2 gene was used to perform segregation analysis with The Jackson Laboratory DNA Panel Mapping Resource. On the basis of this analysis, the Cln2 gene was localized to a region of mouse Chr 7 that corresponds to human Chr 11p15. Characterization of the mouse Cln2 gene will facilitate generation of a mouse model for late-infantile ceroid-lipofuscinosis by gene targeting and identification of functionally important regions of the Cln2 protein.

Amino Acid Sequence↗
PubMed1999

Ocular manifestations of equine motor neuron disease.

The characteristics of the ocular manifestations of equine motor neuron disease (EMND) are described. Forty-two horses with histories, clinical signs and necropsies compatible with EMND were the subjects of this study. Ophthalmoscopic lesions that varied in severity were found in 40 of 42 horses and appeared as a distinct pigmented reticulated pattern at the tapetal-nontapetal junction or throughout the fundus, depending upon severity. The pattern colours ranged from yellow brown to black. Areas of hyperreflectivity formed mosaic patterns in the tapetal fundus. ERG B-wave amplitudes were usually at least 50% reduced and many animals showed extinguished amplitudes. None of the horses had apparent visual impairment. Histopathologically, all 42 horses had retinal pigment epithelial (RPE) congestion with ceroid-lipofuscin. Retinal degeneration was variable even within the eyes. Thin layer chromatography (TLC) analysis of the RPE and neural retina identified both green and orange emitting fluorescent compounds not found in normal horses. All unsupplemented horses had plasma vitamin E levels <1.0 microg/ml. The potential significance of this report is the pathognomonic role the ocular manifestations exhibit in helping to diagnose equine motor neuron disease.

Animals↗
PubMed1999

Reversible accumulation of lipofuscin-like inclusions in the retinal pigment epithelium.

PURPOSE: The amounts of autofluorescent lysosomal storage bodies, known as lipofuscin, increase during senescence in the retinal pigment epithelia (RPEs) of mammalian eyes. This increase in lipofuscin content may result from a failure of the RPE to dispose of any lipofuscin constituents once they have formed. Alternatively, the RPE may eliminate lipofuscin but at a rate insufficient to prevent its accumulation. Experiments were conducted to distinguish between these two possibilities. METHODS: Albino rats were given intravitreal injections of the protease inhibitor leupeptin, which induces a rapid accumulation of lipofuscin-like inclusions in the RPE. The amount of these inclusions in the RPE was monitored as a function of time after the leupeptin treatment with quantitative ultrastructural analysis. In addition, the intensity of lipofuscin-specific fluorescence in the RPE was monitored over the same time period with the use of quantitative microfluorometry. These parameters were also followed in untreated control eyes of age-matched animals. RESULTS: A single leupeptin injection resulted in a rapid massive accumulation of electron-dense inclusion bodies in the RPE. These inclusions appeared to be derived primarily from phagocytosed photoreceptor outer segments. Accompanying the accumulation of these inclusions was a significant increase in lipofuscin-specific fluorescence in the RPE. Over a 12-week period after the leupeptin treatment, the amounts of inclusion material and the fluorescence intensities returned to normal levels. CONCLUSIONS: These findings suggest that the age-related increase in RPE lipofuscin content results from an imbalance in the rates of lipofuscin formation and disposal rather than from a complete absence of a disposal mechanism. The results imply that turnover of lipofuscin constituents may be rapid relative to the animals' life span. Thus, it may be possible to slow or reverse the age-related increase in RPE lipofuscin content by either inhibiting the processes involved in lipofuscin formation or enhancing the disposal processes.

Aging↗
PubMed1998

Coding sequence and exon/intron organization of the canine CLN3 (Batten disease) gene and its exclusion as the locus for ceroid-lipofuscinosis in English setter dogs.

Hereditary ceroid-lipofuscinosis in English setters has been proposed to be the canine equivalent of human juvenile ceroid-lipofuscinosis, which results from defects in the CLN3 gene. Analyses were performed to determine whether the disease in English setters is also the consequence of a CLN3 gene mutation. Canine CLN3 cDNA was found to contain a 1,314-bp open reading frame predicting a derived amino acid sequence which is 89%, 85%, and 84% identical to the predicted amino acid sequences for the human, mouse, and rabbit CLN3 proteins, respectively. The canine gene has sixteen exons. No differences were detected when cDNA nucleotide sequences from an English setter with ceroid-lipofuscinosis and from a normal dog were compared. Moreover, alleles of the canine CLN3 gene distinguished by an intragenic marker segregated independently from the disease in an English setter family, eliminating CLN3 as the locus for the canine disease. A ceroid-lipofuscinosis-affected Tibetan terrier was homozygous for a Gly70Glu CLN3 variant; however, this allele is common in dog breeds considered free of ceroid-lipofuscinosis.

Amino Acid Sequence↗
PubMed1998

Altered mitochondrial function in canine ceroid-lipofuscinosis.

The neuronal ceroid-lipofuscinoses (NCL) are a group of autosomal recessively inherited neurodegenerative disorders characterized by progressive dementia, neuronal atrophy, and premature death. The late infantile and juvenile types of NCL show massive accumulation of mitochondrial ATP synthase subunit c protein in both mitochondria and lysosomes. The specific accumulation of this mitochondrial protein suggests that mitochondrial function may be impaired in the NCL diseases. Therefore, a study was conducted to determine whether oxidative phosphorylation is altered in liver mitochondria from English setters with NCL, an animal model in which there is also massive accumulation of the subunit c protein. The ADP/O ratios were significantly depressed in affected and carrier dogs, suggesting that the disease mutation led to a partial uncoupling of oxidative phosphorylation. On the other hand, ADP-stimulated respiration rates were higher than normal in both carriers and affected dogs. The increased respiration rates were highest in the carriers, and may reflect a compensatory response to the reduced efficiency of oxidative phosphorylation. Accompanying the increased respiration rates were elevations in mitochondrial ADP content with the elevation being greater in the carriers than in the affected dogs. This suggests that the increased respiration rates may be due, at least in part, to enhanced ADP uptake by the mitochondria. In the carriers, the enhanced respiration rate may be sufficient to offset the reduced efficiency of oxidative phosphorylation. In the affected animals, which had lower respiration rates than the carriers, the enhanced respiration rates may not be sufficient to offset the reduced efficiency of oxidative phosphorylation. Impaired mitochondrial function may therefore contribute to the disease pathology.

Adenosine Diphosphate↗
PubMed1998

Visual sensitivity and interphotoreceptor retinoid binding protein in the mouse: regulation by vitamin A.

Interphotoreceptor retinoid binding protein (IRBP) is a retinoid and fatty acid binding glycoprotein secreted by rod and cone photoreceptors in all vertebrates. IRBP is believed to serve as a carrier for retinoids in the bleaching and regeneration cycle of rhodopsin. IRBP protein has been found to be decreased in vitamin A-deprived rats; it is rapidly recovered after retinol repletion. To understand the mechanism for this recovery, we determined whether vitamin A affects transcription and translation of the IRBP gene. Wild-type and transgenic mice harboring the IRBP promoter-CAT reporter fusion gene were maintained on a retinol-deficient diet supplemented with retinoic acid (-A) or on a control diet (+A) for up to 60 wk postweaning. Some of the -A mice were given retinol repletion for 7 days (-A+A). Electroretinography analysis revealed alterations in waveform and a 2 log unit decrease in b-wave sensitivity in the -A mice over a broad range of stimulus wavelengths. Retinol repletion effected a full recovery. Immunochemistry showed a significant decrease in the immunogold-labeled IRBP between the retinal pigment epithelium and the outer segments of the -A mice compared with +A and -A+A mice. Northern blots showed no differences in the amounts of IRBP or CAT mRNA between these three treatment groups. These results suggest that the regulation of IRBP by retinol is not transcriptional.

Animals↗