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A Spector

Publications and source records attributed to A Spector.

At least 19 recordsLinked to original sources
PubMed1992

Na,K-ATPase of cultured bovine lens epithelial cells: H2O2 effects.

Na,K-ATPase function was studied in cultured bovine lens epithelial cells under confluent and non-confluent conditions. The affinity of the Na,K-ATPase for the cardiac glycoside, ouabain, differs between the confluent and non-confluent cultures. The confluent cells have a higher affinity for ouabain than do the non-confluent cells. The ouabain affinity of the confluent cells is similar to that for the Na,K-ATPase isolated from the bovine axolemma and the bovine lens cortex. The ouabain affinity of the non-confluent cells is similar to that for the Na,K-ATPase of the renal medulla and bovine lens epithelium. Similar results are not found with confluent and non-confluent MDCK cells. H2O2 treatment of confluent and non-confluent lens epithelial cell cultures has differing effects on the Na,K-ATPase function. In the confluent cell preparations, H2O2 affects K(+)-dependent dephosphorylation of the intermediate phosphoenzyme. In the non-confluent preparations. H2O2 appears to inhibit K(+)-occlusion.

Animals↗
PubMed1991

Phosphorylation of alpha-crystallin B in Alexander's disease brain.

The phosphorylation of alpha-crystallin B was studied in homogenates of autopsy samples of brain tissue from patients with Alexander's disease, a condition characterized by over-expression of this protein. After incubation in the presence of [gamma-32P]ATP and cAMP the homogenates were analyzed by two-dimensional electrophoresis, (isoelectric focusing followed by SDS-PAGE). Three major polypeptides having the same molecular weight as bovine lens alpha-crystallin B and pIs 7.1, 6.9 and 6.7 were detected in the Coomassie blue stained gels. These three polypeptides were recognized by an alpha-crystallin B-specific antiserum in Western blots. The polypeptides with pIs 7.1 and 6.7 co-migrated in isoelectric focusing gels with bovine lens alpha B and its phosphorylated form alpha Bp, respectively. Radioautography of the two-dimensional gels demonstrated the presence of 32P in the most acidic polypeptide. The results demonstrate the occurrence of alpha B phosphorylation in Alexander's disease brain tissue.

Blotting, Western↗
PubMed1991

The calf gamma crystallins--a Raman spectroscopic study.

The solution structures of the four major components of bovine lens gamma-crystallin, gamma s, gamma II, gamma III and gamma IV are compared using Raman spectroscopy. The spectral region sensitive to the vibrational frequencies of aromatic and sulfur containing residues and to the backbone skeletal stretching modes (500-1000 cm-1), and that reflecting secondary structure (1,000-1,700 cm-1) are strikingly similar in all four gamma-crystallin fractions. These similarities are indicative of the dominant anti-parallel beta sheet structure common to all the gamma-crystallins. A comparison of the ratios of the Raman intensities at 850 cm-1 and 830 cm-1 (I850/I830), an empirical measure of the degree of hydrogen bonding of phenolic hydroxyl groups, suggests that the tyrosine residues in all the gamma-crystallin fractions are moderately hydrogen bonded. Distinct differences in the solution structures of the gamma-crystallins were observed in the higher energy end of the vibrational Raman spectra. The sulfhydryl stretching frequencies for the gamma-crystallins exhibit complex splitting patterns in the 2,500-2,600 cm-1 region. These patterns are due to the competing effects of hydrogen bonding and S-pi interactions with neighboring aromatic residues. All five proteins exhibit multiple, but distinct, thiol frequencies, suggesting that the microenvironments of the cysteine residues in these proteins are significantly different.

Animals↗
PubMed1991

Traumatic osteoarthrosis of the ankle joint secondary to ankle fractures.

Arthritis, in one form or another, is one of the most common disorders that affect man. Whether the etiology is systemic, neurologic, or traumatic, the one common finding is the destruction of joint cartilage. Although there have been massive amounts of literature concerning osteoarthritis, little has been written about arthritis of the ankle joint. Current concepts in the pathology, terminology, symptomatology, and treatment of the arthritic ankle are discussed.

Ankle Injuries↗
PubMed1990

The apparent molecular size of native alpha-crystallin B in non-lenticular tissues.

The apparent molecular size of the native alpha-crystallin B in cytosol preparations from rat heart, brain and retina was determined by gel permeation chromatography, detecting the protein by immunochemical assay (ELISA), using an alpha-crystallin specific antiserum. Native alpha-crystallin from cytosol preparations of rat lens cortex was used as a reference. alpha-Crystallin B present in all three cytosol preparations from non-lenticular tissues eluted in a single symmetrical peak, with the same elution volume as alpha-crystallin from lens cortex cytosol preparations, corresponding to an apparent average molecular size of 0.8 x 10(6) Da. No other species could be detected. The results indicate that the alpha-crystallin aggregates characterized by an apparent average molecular mass of 0.8 x 10(6) Da, and considered to be the native, physiological form of the protein in the lens, are indeed not specific to lens tissue. Furthermore, the size of these alpha-crystallin aggregates is independent of their polypeptide composition. Aggregates found in the lens, composed of alpha A and alpha B polypeptides and their respective phosphorylated forms alpha Ap and alpha Bp, are similar in size to those found in heart, brain and retina, containing the alpha B but not the alpha A polypeptide.

Animals↗
PubMed1990

Some aspects of Dr Kinoshita's contributions to lens protein chemistry.

A review of some of Dr Kinoshita's contributions to our understanding of lens protein and glutathione biochemistry is presented. Particular emphasis is placed on Dr Kinoshita's work involved with the relationship of carbohydrate metabolism and the maintenance of reduced glutathione, the question of the biological function of glutathione in the lens, the effect of oxidative stress provided by diamide and azoester on glutathione, membrane pump function and protein and also ascorbate and H2O2 effects on Na+, K(+)-ATPase. The importance of oxidative stress was recognized early by Dr Kinoshita and he has continued to make significant contributions in this area as illustrated by his work with Dr Zigler on posterior subcapsular cataracts and with Drs Garland and Zigler on mixed function oxidation. It is concluded that Dr Kinoshita's overall contributions in the areas mentioned above have been broad and of considerable importance.

Animals↗
PubMed1990

Hydrogen peroxide-induced DNA damage in bovine lens epithelial cells.

The present investigation was undertaken to determine the types and extent of DNA damage resulting from incubation of primary cultures of bovine lens epithelial cells with hydrogen peroxide. Significant numbers of DNA single-strand breaks were detected by alkaline elution after exposure to as little as 25 microM H2O2 for 5 min at 37 degrees C. The extent of single-strand breakage was concentration dependent and linear from 25 to 200 microM H2O2. The observed single-strand breaks appear primarily due to the action of the hydroxyl radical via a Fenton reaction as both an iron chelator, 1,10-phenanthroline and OH. scavengers, including DMSO, KI and glycerol, significantly inhibited the DNA-damaging effect of H2O2. Diethyldithiocarbamate, an inhibitor of superoxide dismutase, further potentiated the DNA-damaging effects of H2O2, presumably by increasing the steady-state concentration of Fe2+. DNA-protein cross-linking was not observed. In addition, significant levels of 5,6-saturated thymine residues or pyrimidine dimers were not detected after modification of the alkaline elution methodology to allow the use of either E. coli endonuclease III or bacteriophage T4 endonuclease V, respectively. No double-strand breaks were detected after incubation of epithelial cell cultures with H2O2 concentrations of up to 400 microM for 10 min and subsequent neutral filter elution. Since, in vivo, the lens epithelium contains populations of both quiescent and dividing cells, the degree of susceptibility to oxidative damage was also studied in actively growing and plateau-phase cultures. Reduced levels of single-strand breakage were observed when plateau-phase cultures were compared to actively growing cells. In contrast, essentially no differences in repair rates were noted at equitoxic doses of H2O2. The above results suggest that lens epithelial cells may be particularly sensitive to oxidative damage and thus are a good model system in which to study the effects of oxidative stress.

Animals↗
PubMed1990

Ultraviolet light induced DNA damage and repair in bovine lens epithelial cells.

DNA damage caused by UV-B and UV-A irradiation and the rate of repair of such damage was quantitated in bovine lens epithelial cell cultures using a modified alkaline elution methodology. Two enzymes, bacteriophage T4 endonuclease V, which cleaves at the site of pyrimidine dimers, and E. coli endonuclease III, which cleaves at the site of thymine glycols, were utilized. Pyrimidine dimers were not detected after UV-A irradiation of lens cultures with up to 400 J/m2. In contrast, after exposure to as little as 2 J/m2 of UV-B irradiation, large numbers of pyrimidine dimers were observed. At higher fluences, thymine glycols were also found. Significant levels of DNA-DNA crosslinking were suggested by reduced rates of elution of DNA from cells treated with both UV-B irradiation and H2O2 in comparison to treatment with H2O2 alone. Protein-DNA crosslinks, in contrast, were not observed. The rate of repair of UV-B induced DNA damage was quantitated by harvesting cells at various times after the UV-B exposure. Single-strand breaks were never observed immediately after UV-B exposure but appeared later during the repair phase. In contrast to the repair of H2O2 induced DNA damage, which is largely completed within 30 min of exposure, more than 50% of the UV-B light induced DNA damage remained unrepaired five hours after exposure. This difference between the rate of repair of H2O2 and UV-B induced DNA damage could provide valuable insights into the nature of DNA damaging agents in the lens environment and may reflect underlying differences in the potential for epithelial cell DNA mutation in response to various DNA damaging insults.

Animals↗
PubMed1990

Prevalence of age-related cataract in Ganzi and Shanghai. The Epidemiological Study Group.

It has been reported that the prevalence of cataract is much higher in plateau than that in plain regions. But the data are incomplete and can not be compared because of different examination methods and different diagnostic criteria. A small group of ophthalmologists was sent to Ganzi, Sichuan Province at the altitude of 3.393 m, and to Qinpu in the suburb of Shanghai which is at the sea level. In Ganzi 3,905 (91.97%) people including 822 Hans and 3,083 Zang and in Qinpu 3,101 (93.20%) were studied. Age-related cataract was diagnosed when there are cortical or nuclear, or mixed opacities; and aphakia due to previous extraction of age-related cataract. Direct method was used to standardize the data. The prevalence of age-related cataract in Ganzi is about 82% higher than that in Qinpu in all age groups; from age 16-60, the prevalence of age-related cataract in Ganzi is about 180% (in Hans) and 160% (in Zangs) higher than that in Qinpu. There is no significant difference in the prevalence of age-related cataract between the ethnic groups in Ganzi. In Ganzi, the sunshine hours per year are 1.28 times that in Qinpu, and the global radiation in Ganzi is 1.47 times that in Qinpu. The higher prevalence of cataract in Ganzi than in Qinpu may be due to the greater exposure to the sunshine on the Tibetan plateau. The differences in prevalence between males and females and between indoor and outdoor work are discussed.

Adolescent↗
PubMed1989

The dephosphorylation of lens alpha-crystallin A chain.

The present communication reports the presence of a phosphoprotein phosphatase activity in bovine lens preparations which dephosphorylates alpha Ap, the phosphorylated form of alpha A, one of the alpha-crystallin polypeptides, in a Ca2+/calmodulin dependent manner. The activity was found in soluble preparations from epithelial cells but it could not be detected in similar preparations from fiber cells. A 60,000 Mr calmodulin binding polypeptide and a 15,000 Mr polypeptide found in the epithelial cell preparations comigrated in SDS-PAGE with the A and B subunits of bovine brain calcineurin (phosphoprotein phosphatase 2B) respectively. The 15,000 Mr was specifically recognized by an anti-bovine brain calcineurin antiserum. Bovine brain calcineurin was as effective in dephosphorylating alpha Ap as the lens preparations. Thus, it is likely that the activity present in the lens is related to this enzyme. The results indicate that the lens specific polypeptide alpha A may be subject to metabolic control through phosphorylation and dephosphorylation pathways regulated by cAMP and calcium respectively. Changes in the activities of these pathways appear to occur during differentiation of the lens epithelial cell and may be related to gene regulation during the differentiation process.

Animals↗
PubMed1989

UV laser photodamage to whole lenses.

Lenses from rat or calf were exposed in vitro to UV radiation from a nitrogen laser operated at 337.1 nm or from an excimer laser operated at 3.8 nm. Visible light transmission was monitored during calf lens irradiations at 308 nm and found to decrease. Proteins were extracted from the irradiated rat or calf lenses, separated into water soluble and insoluble fractions, and analysed using SDS-PAGE. Comparison of these gels with dark controls showed that, following photolysis, there was loss of polypeptide material in the 20-30 kDa region and concomitant formation of polymers at 40 and 60 kDa, and at greater than 100 kDa in calf lens (308 nm irradiation) and rat lenses (337.1 nm irradiation) in vitro. In addition, there was evidence for formation of lower molecular weight polypeptides at 10 kDa in the protein from irradiated rat lenses. The rat SDS-PAGE gels were challenged against anti-calf gamma crystallin serum. There was clear evidence that the polymeric material, in the water insoluble protein fraction from the 337.1 nm photolyzed rat lenses was derived in part from gamma crystallin. The macromolecular changes detected in these photolyzed rat and calf lens proteins were similar to those previously reported to accompany aging in the human lens. Biochemical changes of the type observed in UV irradiated rat and calf lenses may be responsible for the loss of visible light transmission seen in calf lenses.

Animals↗
PubMed1989

Repair of H2O2-induced DNA damage in bovine lens epithelial cell cultures.

H2O2 concentrations only slightly higher than normal physiological levels found in the lens and aqueous fluid produce a significant number of DNA single-strand breaks in lens epithelial cell cultures. In this investigation, the repair of DNA damaged by short-term, H2O2-induced oxidation was examined in bovine lens epithelial cell cultures. Repair was rapidly initiated and was almost completed in 30 min. A drop in NAD concentration was associated with the DNA damage. 3-Aminobenzamide inhibition of poly(ADP-ribose) polymerase, an enzyme believed to be stimulated by DNA oxidation and involved in DNA repair, prevented the loss of NAD. In contrast, a similar drop in ATP concentration was only slightly lessened by the presence of this inhibitor. Inhibition of the polymerase by 3-aminobenzamide primarily affected only the early recovery period. Overall, recovery occurred almost as effectively in the presence of the inhibitor as in its absence. Preincubation of lens cultures with o-phenanthroline, an iron chelator, prevented the drop in NAD levels associated with DNA damage. Since a hydroxyl radical is produced from H2O2 by a Fenton type reaction, this result supports the concept that the H2O2-induced oxidation of DNA is caused by hydroxyl radical. In contrast, peroxide-induced loss of activity of a cytosolic enzyme, glyceraldehyde-3-phosphate dehydrogenase, was unaffected by the presence of o-phenanthroline, suggesting direct H2O2 oxidation of this enzyme. The results of these experiments suggest that lens epithelium contains enzymes that rapidly repair single-strand DNA breaks induced by H2O2 insult.

Animals↗
PubMed1989

The synthesis of glutathione peroxidase analogs.

While it is recognized that oxidation has a major role in the development of cataract, few compounds have been developed which effectively reduce the potential for oxidative insult. A major oxidant confronting the lens is H2O2. Few compounds capable of metabolizing H2O2 have been synthesized. This communication reports that a number of compounds with glutathione peroxidase activity have been developed. Some of these compounds are 10 fold more active than Ebselen, previously recognized as the synthetic compound with the greatest GSH peroxidase activity.

Azoles↗
PubMed1979

An extrinsic membrane polypeptide associated with high-molecular-weight protein aggregates in human cataract.

A 43,000-dalton polypeptide has been isolated from the high-molecular-weight disulfide-rich fraction of the water-insoluble protein of human cataractous lenses. On the basis of immunochemical reactivity and fluorescent antibody binding, this polypeptide is localized in the membrane region of the lens cell. This observation suggests an interaction between the soluble lens proteins and membrane-associated polypeptides in the formation of large protein aggregates which may cause cataract.

Antibody Specificity↗