Biomedical subjects

Dayu Li

Publications and source records attributed to Dayu Li.

11 recordsLinked to original sources

PubMed2004

Protein profile of aging and its retardation by caloric restriction in neural retina.

Aging is a slow, gradual deterioration process of an organism. The only experimental intervention, which can reliably retard aging and age-related degenerative diseases, is dietary caloric restriction (CR). To gain insight into the mechanism of CR intervention, we have investigated the protein profile of aging and its retardation by CR in the neural retina of Brown Norway (BN) rats using the comprehensive proteomic approach. We found that the intensities of 18 proteins decreased significantly with age. CR intervention can completely prevent seven of them, and partially protect eight of them, from such age-related declines. The major protein targets protected by CR intervention appear to be glycolytic enzymes and molecular chaperones. These data are the first to suggest that CR may retard the age-related degeneration of retina by maintaining sufficient glucose metabolism, by ensuring proper protein folding, and/or by preventing protein denaturation in the neural retina.

Aging↗

PubMed2004

Dietary caloric restriction may delay the development of cataract by attenuating the oxidative stress in the lenses of Brown Norway rats.

Dietary caloric restriction (CR) is the only experimental intervention that can reliably retard the development of cataract in a normal animal model. Here we have studied the possible mechanisms by which CR retards the age-related degeneration of the lens of Brown Norway rats. We have found that CR slowed protein insolubilization and blunted declines of the total soluble thiols, protein thiols, reduced glutathione and ascorbic acid levels in the lenses of old BN rats. From the lens protein point of view, the development of cataract in rat lenses has 3 stages: (1) the precipitation of gamma-crystallin, (2) the insolubilization of beta-crystallin, and (3) the final precipitation of alpha-crystallin which was saturated with other denatured lens proteins. A similar sequence is also observed when the lens proteins are subjected to oxidative stress in vitro. These data are the first to suggest that CR may retard the age-related degeneration of the lens by attenuating the oxidative stress in the lens. Since oxidative stress is likely a main cause of human cataract, CR intervention may be relevant to humans as well.

Aging↗

PubMed2004

Cluster analysis of genes with significant change in expression in cells conditioned to survive TBOOH.

Immortal murine lens epithelial cells, alphaTN4-1 have been conditioned to survive H2O2, H cells, or TBOOH, T cells, at concentrations that will cause cataract in vitro. Since H cells are killed by TBOOH but T cells survive H2O2, it is of interest to examine the gene expression of these cell lines. We now report the results of cluster analysis of genes whose expression is significantly changed by TBOOH. The analysis has revealed a small group of antioxidative defense genes that contribute to the survival of T and H cells when exposed to oxidative stress.

Animals↗

PubMed2004

Comparison of characteristics of peroxide-conditioned immortal human lens-epithelial cell lines with their murine counterparts.

Previously, this laboratory has reported the characteristics of murine immortal lens-epithelial cells (alphaTN4-1) conditioned to survive either H2O2 or tertiary butyl hydroperoxide (TBOOH) stress. This communication now describes similar observations upon human HLE-B3 cells. It was found that the human cells are more sensitive to peroxides than their murine counterpart. Similar to the murine cells, conditioning to TBOOH endows the HLE-B3 cells with resistance to H2O2 but unlike the murine cells, conditioning to H2O2 gives the human cells resistance to TBOOH. Furthermore, while withdrawal of TBOOH stress from TBOOH-conditioned alphaTN4-1 cells causes a loss of resistance to this peroxide but not H2O2, with human cells resistance to both peroxides is retained. Examination of the antioxidative defense (AOD) enzyme activities show an extraordinary increase in catalase activity and significant augmentation of most other enzymes assayed in all conditioned human cell lines. In contrast, it was previously found that only catalase and glutathione-S-transferase have considerable increases in activity in the murine lines. However, in most cases, the AOD enzyme activity in murine-control cells is about 2-fold higher than in human control cells. The gene expression of human TBOOH-conditioned (Thum) and control (Chum) lines were also examined utilizing microarray analysis. Surprisingly, no significant change in gene expression was found for any of the prominent AOD enzymes. Such results differ from the response of murine cells where many AOD enzymes have increased expression. These observations suggest while the same AOD enzymes may be utilized in both murine and human lens-epithelial cells, the levels at which they are maintained and the manner in which they are recruited in response to stress may differ.

Animals↗

PubMed2004

DeltaFosB expression and cataract.

DeltaFosB is a truncated form of a FosB transcription factor, which is created by alternative splicing. Previous work has shown that transgenic mice expressing DeltaFosB both in the retina and in the lens developed a posterior subcapsular cataract resulting from the misalignment of the fibres in the suture region. In the previous study, it was not clear whether DeltaFosB expression was required in both tissues to produce the cataract. Therefore, DeltaFosB expression targeted to either the lens or the retina was undertaken in order to clarify the contribution of each tissue to cataract development. For lens expression, the R2betaB1DeltaFosB construct was synthesized (R2, an enhancer; betaB1, a chicken betaB1 crystallin gene promoter fragment). For the retina, RhoDeltaFosB was prepared. As a promoter, the bovine rhodopsin upstream region was used. DeltaFosB expression in heterozygote animals was monitored by Western blotting. Cataract development in heterozygotes of R2betaB1DeltaFosB transgenics and in both heterozygotes and homozygotes of RhoDeltaFosB transgenics was followed by slitlamp examination. The transgenic mice prepared with RhoDeltaFosB expressed DeltaFosB only in the retina and showed no sign of lens opacity. One line of the R2betaB1DeltaFosB transgenic was found to have expression only in the lens and developed posterior subcapsular cataract. We concluded that retinal expression of DeltaFosB is not sufficient to cause cataract while expression exclusively in the lens produces posterior subcapsular cataract.

Animals↗

PubMed2004

Matrix metalloproteinases are down-regulated in rat lenses exposed to oxidative stress.

Matrix metalloproteinases are important biological effectors of tissue remodelling. Increased MMP expression occurs during injury, inflammation, cellular transformation, and oxidative stress. Oxidative stress in the lens, a causal factor in cataractogenesis, has been shown to induce MMP secretion. The objective of this study was to assess the expression of MMPs and their regulators in an oxidative stress model of cataract, where epithelial cell death and cortical fibre cell swelling occurs in rat lenses after exposure to riboflavin, oxygen, and light. Two time points (4 and 7 hr of exposure) were chosen in order to compare transparent lenses with partially opaque lenses. MMP activity, protein, and mRNA levels were measured. The results show that MMP-2, MMP-9, MT1-MMP, and MT3-MMP are down-regulated by oxidative stress and that the down-regulation is most likely due to reduced gene transcription. In contrast, genes for catalase, glutathione peroxidase, and GAPDH are essentially unaffected, while beta-actin mRNA and protein levels are markedly increased at both time points. The down-regulation of MMPs occurs in lenses still seemingly transparent after 4 hr of exposure, indicating that reduced MMP activity is a relatively early response to the oxidative stress. Moreover, in our model system, MMP inhibition, not induction, is associated with cataractogenesis.

Actins↗

PubMed2004

The effect of stress withdrawal on gene expression and certain biochemical and cell biological properties of peroxide-conditioned cell lines.

Maturity onset cataract is a disease that afflicts >25% of the U.S. population over 65. Oxidative stress is believed to be a major factor in the development of this disease and peroxides are suspected to be prominent stressing agents. To elucidate mechanisms involved in the protection of cells against oxidative stress, immortal murine lens epithelial cells (alphaTN4-1) have been conditioned to survive lethal concentrations of either tertiary butyl hydroperoxide, TBOOH (a lipid peroxide prototype) (T cells), or H2O2 (H cells). It was found that T cells survived exposure to H2O2 but H cells were killed by TBOOH. In this communication, biological characteristics of the T cells are reported. It is shown that the T cell's ability to survive TBOOH is lost if the cells are grown in the absence of this peroxide (denoted as T- cells). By comparing the differential gene expression of 12,422 genes and ESTs from T and T- and the unconditioned control cells, 16 genes were found that may account for the loss of resistance to TBOOH. They include 5 glutathione-S-transferases, superoxide dismutase 1, zeta crystallin, a NADPH quinone reductase, as well as genes involved in detoxifying aldehydes, controlling iron metabolism, and degrading toxic lipoproteins.

Animals↗

PubMed2003

Caloric restriction retards age-related changes in rat retina.

The neural retina of mammals consists of light sensitive photoreceptors and connecting neural cells that receive and send visual signal to the brain. Dietary caloric restriction (CR) is the only experimental intervention that can reliably retard the age-related degeneration of the retina in a normal mammalian model. Here, we studied the effect of CR on various biochemical parameters in the retina of male Brown Norway rats at different ages. We found that CR slowed the age-dependent protein insolubilization, blunted the declines in the total soluble thiols, and reduced glutathione and ascorbic acid levels in neural retina. We also observed that CR retarded the age-related decline in the levels of taurine, a vital amino acid in neural retina. These data are the first to implicate that CR may retard the age-related degeneration of retina by attenuating the oxidative stress and/or by sustaining the pool of protective factors in the neural retina.

Aging↗

PubMed2003

Peroxide toxicity in conditioned lens epithelial cells--evaluation of multi-defense systems.

Immortal murine lens epithelial cells which were conditioned to survive peroxide stress were found to have a remarkable increase in catalase activity as well as lesser changes in a number of other antioxidative defense systems [Invest. Ophthalmol. Vis. Sci. 43 (2002) 3251]. Furthermore, the gene expression of hundreds of other genes was altered. In order to determine the relative importance of catalase, other enzyme systems which maintain the reducing environment of the cell and the involvement of Fenton chemistry, an analysis of the effect of inhibiting catalase, disruption of the cells' reducing environment by inhibition of GSSG reductase (GR) and chelation of metal ion was investigated. It was found that inhibition of catalase caused peroxide resistant cells to die within 48-72 hr when exposed to normally tolerated concentrations of peroxide. If 1,10-phenanthroline (OP), an effective metal ion chelator was present, the cells were not affected by catalase inhibition and survived peroxide stress. Peroxide vulnerable unconditioned control cells were similarly protected by the chelator. The results demonstrate that H2O2 itself has minimal toxicity and that it is the products resulting from interaction with metal ion that produces lethal toxicity. In stark contrast, however, metal chelation did not protect the cells when GR was inhibited by BCNU. Examination of non-protein thiol (NP-SH), which is primarily GSH, indicated that rapid and extensive oxidation occurred almost immediately after exposure to peroxide under all conditions. However, NP-SH returns to the normal range in the conditioned cells even though later cell death is observed in some cases, suggesting fatal damage during the period when the cell is exposed to an oxidizing environment. Examination of DNA damage by alkaline elution indicated that H2O2 caused little observed strand breakage in peroxide resistant cells even if catalase is inhibited, suggesting that such cells have developed other systems to protect DNA and that H2O2 induced death is probably not related to DNA single strand breaks. In contrast, unconditioned cells (C cells) show extensive H2O2 induced DNA damage which is prevented by OP. Thus, depending on the conditions, DNA damage may contribute to cell death. The overall results indicate that the conditioned cell lines are not simply dependent on catalase activity but have developed a complex defense which includes GSH dependent systems and possibly more effective regulation of metal ion concentrations to resist oxidative stress.

Animals↗

PubMed2002

The effect of H2O2 and tertiary butyl hydroperoxide upon a murine immortal lens epithelial cell line, alphaTN4-1.

The effect of comparable concentrations of H(2)O(2) and tertiary butyl hydroperoxide (TBHP) upon an immortal murine lens epithelial cell line was examined as part of an ongoing effort to delineate differences in the mechanism by which these peroxides cause cell death. Both compounds result in cell death of normal, unconditioned cells within 24hr. It was found that with similar conditions, TBHP conditioned alphaTN4-1 cells survive H(2)O(2) stress while H(2)O(2) conditioned cells are killed by TBHP. To better understand how these peroxides act, their effect upon unconditioned cells has been investigated. Both peroxides cause a rapid loss of GSH and disruption of pump activity as illustrated by (14)C-choline transport and (86)Rb uptake. While H(2)O(2) exposure resulted in extensive DNA damage, TBHP had a minimal effect. DNA damage caused by H(2)O(2) was shown to activate polyADP-ribosyl polymerase (PARP), leading to depletion of NAD and ATP. H(2)O(2) induced cell death could be delayed by addition of 3-aminobenzamide (3AB), an inhibitor of PARP. ATP levels in cells subjected to H(2)O(2) were also maintained by the presence of 3AB. H(2)O(2) stress also disrupted glycolysis and mitochondrial activity but these parameters were not affected by TBHP. TBHP induced cell death, under the relatively mild conditions used in this work, appears to be caused by membrane disruption and loss of a reducing environment.

Adenosine Triphosphate↗

PubMed2002

Differential amplification of gene expression in lens cell lines conditioned to survive peroxide stress.

PURPOSE: The response of lens systems to oxidative stress is confusing. Antioxidative defense systems are not mobilized as expected, and unanticipated defenses appear important. Therefore, mouse lens cell lines conditioned to survive different peroxide stresses have been analyzed to determine their global changes in gene expression. METHODS: The immortal mouse lens epithelial cell line alphaTN4-1 was conditioned to survive 125 microM H2O2 (H cells) or a combination of both 100 microM tertiary butyl hydroperoxide (TBHP) and 125 microM H2O2 (HT cells), by a methodology previously described. The total RNA was isolated from the different cell lines and analyzed with oligonucleotide mouse expression microarrays. Four microarrays were used for each cell line. Microarray results were confirmed by real-time RT-PCR. RESULTS: A new cell line resistant to both 125 microM H2O2 and 100 micro M TBHP was developed, because cells resistant to H2O2 were killed by TBHP. Analysis of classic antioxidative enzyme activities showed little change between cells that survive H2O2 (H) and those that survive H2O2 and TBHP (HT). Therefore, the global change in gene expression in these cell lines was determined with gene expression microarrays. The fluorescent signal changes of the genes within the three cell lines, H, HT, and control (C), were analyzed by statistical methods including Tukey analysis. It was found that from the 12,422 gene fragments and expressed sequence tags (ESTs) analyzed--based on a one-way ANOVA with a stringent cutoff of one false positive per 1000 genes and correcting for microarray background and noise--approximately 950 (7.6%) genes had a significant change in expression in comparing the C, H, and HT groups. A small group of antioxidative defense genes were found in this population, including catalase, members of the glutathione (GSH)-S-transferase family, NAD(P)H menadione oxidoreductase 1, and the ferritin light chain. The remaining genes are involved in a broad spectrum of other biological systems. In the HT versus H comparison, only a few genes were found that had increased expression in the HT line compared with expression in the H line, including GSH-S-transferase alpha 3 and hephaestin. Many genes that are frequently considered antioxidative defense genes, including most of the GSH peroxidases, unexpectedly showed little change. CONCLUSIONS: An unusual and generally unexpected small group of antioxidative defense genes appear to have increased expression in response to H2O2 stress. Cell lines resistant to H2O2 do not appear to survive challenge with another type of peroxide, TBHP, a lipid peroxide prototype. However, acquisition of TBHP resistance by H cells was found to be accompanied by significantly amplified expression of only a few additional antioxidative defense genes. Many of the amplified genes do not appear to be involved with antioxidative systems, reflecting the complexity of the cells' response to oxidative stress.

Animals↗