Modulation of base excision repair by low density lipoprotein, oxidized low density lipoprotein and antioxidants in mouse monocytes

Kuang-Hua Chen, Deepak K. Srivastava2, Rakesh K. Singhal3, Sam Jacob1, Ahmed E. Ahmed1 and Samuel H. Wilson2,4

Sealy Center for Molecular Science and
1 Department of Pathology, University of Texas Medical Branch, Galveston, TX 77555,
2 Laboratory of Structural Biology, National Institute of Environmental Health Sciences, Research Triangle Park, NC 27709 and
3 Department of Pediatrics, The New York Hospital–Cornell Medical Center, New York, NY 10021, USA


    Abstract
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 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
In the present study, we found that oxidized low density lipoprotein, but not low density lipoprotein, down-regulated base excision repair activity in extracts of mouse monocyte cell line PU5-1.8. An enzyme required in this pathway, DNA polymerase ß, was also down-regulated. In contrast, treatment of monocytes with a combination of ascorbate and {alpha}-tocopherol up-regulated base excision repair activity and expression of DNA polymerase ß. Co-treatment of monocytes with antioxidants plus oxidized low density lipoprotein prevented down-regulation by oxidized low density lipoprotein. Oxidative DNA damage, as measured by 8-hydroxyguanine accumulation in genomic DNA, was found in cells treated with oxidized low density lipoprotein; 8-hydroxyguanine was not found in the cells treated with low density lipoprotein, antioxidants or oxidized low density lipoprotein plus antioxidants. These results establish a linkage between the DNA base excision repair pathway, oxidative DNA damage and oxidized low density lipoprotein treatment in mouse monocytes. Since oxidized low density lipoprotein is implicated in chronic disease conditions such as atherogenesis, these findings facilitate understanding of genetic toxicology mechanisms related to human health and disease.

Abbreviations: BER, base excision repair; ß-pol, DNA polymerase ß; LDL, low density lipoprotein; LPS, lipopolysaccharide; Ox-LDL, oxidized low density lipoprotein; 8-oxo-G, 8-hydroxyguanine; ROS, reactive oxygen species.


    Introduction
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Coronary artery disease is the major cause of morbidity and mortality in the United States and most Western countries. Even though the precise molecular mechanism and sequence of events involved in the initiation and progression of this disease remain to be elucidated, there is a wealth of evidence demonstrating that atherosclerotic lesions result from an excessive inflammatory response mediated by reactive oxygen species (ROS) (15), which are known to have deleterious effects on cells, including causing lipid peroxidation and oxidative DNA damage (68).

Increased deposition of oxidatively modified low density lipoprotein (LDL) is a key step in the atherogenic process. Macrophages recognize and internalize oxidized low density lipoprotein (Ox-LDL) via `scavenger' receptors, rather than the normal LDL receptor, which results in its unregulated uptake and can eventually cause transformation into foam cells (911). Although multiple pathways probably govern LDL modification, a cell-mediated oxidative pathway functioning via ROS is likely to be the most significant (9,1215). Ox-LDL is cytotoxic, inducing cell injury and death in endothelial cells, smooth muscle cells and foam cells (16,17). Ox-LDL's toxicity to vascular cells theoretically could play an important role in atherosclerotic lesion development. Recently, DNA fragmentation and free-radical-induced lipid peroxidation have been implicated as mechanisms of Ox-LDL cytotoxicity (18,19). Lipid peroxidation is known to induce oxidative DNA damage (20,21). Taken together, these results suggest that oxidative DNA damage may be one of the underlying mechanisms of vascular cell death caused by Ox-LDL in atherosclerotic lesions. These oxidative DNA lesions are efficiently and quantitatively repaired primarily by the base excision repair (BER) pathway, which is generally considered to be a constitutively expressed repair system in mammalian cells (7,2225). Therefore, enzymes involved in the BER pathway play a key role in allowing the cell to respond to oxidative DNA damage. If oxidative DNA damage is left unrepaired, deleterious consequences ensue, including mutation, DNA fragmentation, altered gene expression and cell death.

Antioxidants have been widely used for the prevention of atherosclerosis due to their function as free radical scavengers that decrease the initiation and propagation of fatty acid oxidation. Strong epidemiological evidence links dietary antioxidants to a decrease in LDL oxidation and to the reduced clinical expression of atherosclerosis; use of the antioxidants, such as ascorbate, {alpha}-tocopherol and probucol, in animal model studies have shown that the anti-oxidation mechanism accounts for the inhibitory effect of these agents on atherogenesis (2629). The antioxidant protective mechanism against oxidative damage is accomplished by quenching free radicals or reacting with their products (30,31). Therefore, antioxidants can decrease the amount of oxidative damage to DNA, proteins and other macromolecules. Although observational epidemiological studies have produced some intriguing results, they have not unequivocally established that a high intake of antioxidants leads to decreased risk of cardiovascular disease (3234). Thus, intensive study of the molecular mechanisms of prevention of atherosclerotic lesion formation by antioxidants is warranted. In addition, the response to oxidative stress is considered to be linked to carcinogenesis, and therefore this area of research has relevance to cancer biology as well.

Previous studies by Penn et al. (3537) indicated that plaque DNA from both patients and experimental animals displayed alterations similar to those described for genomic DNA from many tumors. They proposed that both atherosclerotic plaque formation and tumorigenesis share common mechanisms and suggest that genetic mutations play a role in the etiology of atherosclerotic plaque formation. Recently, we found that several oxidative stress-inducing agents up-regulated the protein level of DNA polymerase ß (ß-pol), a key enzyme in the BER pathway, and also induced BER activity in mouse monocytes (38). Hence, the BER system is subject to oxidative stress-related up-regulation in monocytes. Here, we determined the effect of LDL, Ox-LDL and antioxidants on the ß-pol level, BER activity and amount of oxidative DNA damage in mouse monocytes. Our results show that treatment with Ox-LDL caused a decrease in both ß-pol level and in vitro BER activity, while LDL treatment had no effect. This Ox-LDL-induced down-regulation of DNA BER activity correlates with an accumulation of 8-hydroxyguanine (8-oxo-G) in cellular DNA. Co-treatment of monocytes with antioxidants (ascorbate and {alpha}-tocopherol) plus Ox-LDL prevent down-regulation of BER activity and accumulation of 8-oxo-G. Implications of these findings are discussed.


    Materials and methods
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Materials
Low density lipoprotein, cupric sulfate, Chelex-100, creatine phosphokinase, diTris-phosphocreatine, nicotinamide adenine dinucleotide, proteinase K, ribonuclease, DNA polymerase I, DNase I and alkaline phosphatase were obtained from Sigma (St Louis, MO). Dulbecco's modified Eagle medium (DMEM), F-10 nutrient mixture (Ham), heat-inactivated fetal bovine serum and gentamycin were purchased from Gibco BRL–Life Technologies (Gaithersburg, MD). Fetal bovine serum was obtained from Hyclone Laboratories (Logan, UT). dNTPs were obtained from Pharmacia LKB Biotechnology (Piscataway, NJ). [{alpha}-32P]dCTP was from DuPont NEN (Boston, MA). 8-oxo-G and dG were obtained from Wako Pure Chemical Industry (Houston, TX). Oligonucleotides were purchased from Operon Technologies (Alameda, CA). Mouse monocyte cell line (PU5-1.8) was obtained from American Type Culture Collection (Rockville, MD).

Cell culture
To determine the effect of LDL, Ox-LDL, ascorbate and {alpha}-tocopherol on ß-pol expression, BER activity and DNA lesion formation, a mouse monocyte cell line (PU5-1.8) was grown in a humidified atmosphere with 10% CO2 in DMEM containing gentamycin (50 µg/ml), L-glutamine (4 mM) and heat-inactivated fetal bovine serum (10%) at 37°C. For experiments, cells were split by manual disruption, centrifuged, resuspended in F-10 nutrient mixture (Ham), plated at a concentration of 4x106 cells/100 mm dish and grown overnight. At the initiation of the experiment, cells were replenished with fresh F-10 nutrient mixture and treated with LDL, Ox-LDL, ascorbate and {alpha}-tocopherol alone or a combination of these agents as described in the text. Control cells were treated with medium only. The data are from three independent experiments with a single result of a typical experiment shown.

For testing the effect of LDL, Ox-LDL, ascorbate and {alpha}-tocopherol on the level of 8-oxo-G in mouse monocyte genomic DNA, cells were treated with the reagents at the following concentrations: LDL, 0.5 µg; Ox-LDL, 0.5 µg; ascorbate, 40 µM; {alpha}-tocopherol, 2 µM. Cells were incubated for 4 or 8 h prior to isolation of genomic DNA samples and 8-oxo-G determination was carried out as described below.

Preparation of oxidized low density lipoprotein
Oxidation of LDL was carried out with CuSO4·5H2O. LDL (1 mg/ml) was first dialyzed twice in 500 ml phosphate buffered saline (PBS) at 4°C for 2 h and then treated with 5 µM CuSO4·5H2O at 37°C for 24 h. Chelex-100 was used to remove CuSO4·5H2O. The modification of LDL was confirmed by the relative electrophoretic mobility (39).

Western blotting analysis
The ß-pol protein level was determined by western blotting as described previously (38,40). Briefly, after treatment, control and treated cell-free extracts were prepared as follows. Cells were scraped and centrifuged at 500 g for 5 min at 4°C, washed once with ice-cold PBS, lysed in RIPA buffer (10 mM Tris–HCl, pH 7.5, 150 mM NaCl, 0.5% sodium deoxycholate, 1% Triton X-100, 0.1% SDS, 1 mM EDTA, 1 mM dithiothreitol, 1 mM PMSF and 2.7 µg/ml aprotinin), and the lysates were microcentrifuged at 11 000 r.p.m. for 10 min to remove cellular debris. Cell-free supernatant fractions were stored at –70°C for later analysis of ß-pol. Total cellular protein (20 µg) was applied to each lane, separated by 12.5% SDS–PAGE and electrotransferred to a nitrocellulose membrane. ß-Pol was measured by incubating the membrane with mouse monoclonal 18 S antibody to ß-pol followed by antibody to mouse immunoglobulin G (IgG) conjugated to horseradish peroxidase (HRP) (38,40). Immobilized HRP activity was detected by enhanced chemiluminescence.

Base excision repair assay
To measure BER activity, cell-free extracts were prepared by resuspending cells in PBS containing 1 mM PMSF, 1 mM dithiothreitol, 2.7 µg/ml aprotinin and lysing by four cycles of freeze/thawing. Cell supernatant fractions were prepared by centrifugation as described above. Total cellular protein (5 µg) was used for BER determination according to the method of Singhal et al. (41). The standard reaction mixture (20 µl) contained 100 mM Tris–HCl, pH 7.5, 5 mM MgCl2, 1 mM dithiothreitol, 0.1 mM EDTA, 2 mM ATP, 0.5 mM NAD, 5 mM diTris-phosphocreatine, 10 U creatine phosphokinase, 40 nM duplex oligonucleotide and 0.3 µM [{alpha}-32P]dCTP (specific activity 6.6x106 d.p.m./pmol). Reactions were incubated at 25°C for 10 min and stopped by adding a `stopping solution' (0.01% xylene cyanol, 0.01% bromophenol blue and 80% formamide) to the reaction mixture. After incubation at 75°C for 5 min, DNA was resolved by electrophoresis on 15% polyacrylamide gel containing 7 M urea, 89 mM Tris, 89 mM boric acid and 2 mM EDTA, pH 8.0. Gels were dried and autoradiographed at –70°C. BER activity is indicated by the level of [32P]dCMP incorporation into DNA, either 51 or 22 nucleotides long.

Analysis of 8-hydroxyguanine
HPLC analysis of 8-oxo-G was performed according to the methods described previously (42). Monocytes were incubated in 1.2 ml digestion buffer (10 mM Tris–HCl, pH 8.0, 100 mM NaCl, 25 mM EDTA, 5% SDS, 0.1 mg/ml proteinase k and 10 µg/ml RNase) at 37°C for 18 h. After digestion, NaCl was added to a final concentration of 1 M followed by centrifugation at 4000 r.p.m. for 15 min. The supernatant fraction was extracted twice with chloroform:isoamyl alcohol (24:1) and DNA was precipitated by the addition of one-half volume of 7.5 M ammonium acetate and two volumes of ethanol. The mixture was kept at –20°C for 2 h and centrifuged. DNA was resuspended in 10 mM Tris–HCl pH 7.4 and the quantity and purity was determined. Samples were digested with DNase I (40 U/100 µg DNA) in the presence of 10 mM MgSO4 at 37°C for 30 min. The P1 nuclease digestion was carried out in the same buffer at 37°C for 2 h with the addition of 0.5 M sodium acetate (pH 7.0) to lower the pH to 7.0 and 1 M ZnSO4 to a final concentration of 1 mM. Alkaline phosphatase (2.5 U/100 µg DNA) was added to the mixture and incubated for an additional 30 min. Acetone (5 ml, HPLC grade) was added to the mixture to precipitate enzymes. Solvent was evaporated under nitrogen gas at temperatures not exceeding 30°C. The residues were dissolved in a small volume of mobile phase solution (100 mM sodium acetate, pH 5.2, 5% methanol) and filtered through a 0.2 µm membrane prior to HPLC analysis using a model 580 dual piston pump and a model 5200A coulochem II electrochemical detector (ESA, Bedford, MA). The potentials were set at +850 mV for the guard cell, +100 mV for the conditioning cell, and +500 and +800 mV for electrodes 1 and 2, respectively for the analytical cell. 8-oxo-G and dG were separated on a YMCB-02-3 C8 base-deactivated 3µ, 4.6 mmx15 cm column (YMC, Wilmington, NC) under isocratic conditions by a mobile phase containing 100 mM sodium acetate, pH 5.2 (adjusted with HPLC grade phosphoric acid) at a flow rate of 1 ml/min. The chromatograms were collected and analyzed by chromjet dual channel integrator (Spectra physics, Fremont, CA). Concentrations of 8-oxo-G, obtained from channel 1 that linked to electrode 1 at a potential of +500 mV, were expressed relative to the concentrations of dG, obtained from channel 2 that linked to electrode 2 at a potential of +800 mV.


    Results
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 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Effects of antioxidants on ß-pol level
Since antioxidants such as ascorbate and {alpha}-tocopherol can reverse oxidative stress and have been used to reduce atherogenesis caused by Ox-LDL (2629), the effect of these antioxidants on ß-pol expression was determined in mouse monocytes. Cells were treated with either ascorbate (40 µM) or {alpha}-tocopherol (2 µM). ß-Pol was induced about 2-fold throughout the 8 h period of treatment with either ascorbate or {alpha}-tocopherol (Figure 1Go). In contrast, when cells were exposed to a combination of ascorbate (40 µM) and {alpha}-tocopherol (2 µM), ß-pol protein level increased 3.5-fold after 1 h and the highest level of induction was 5.5-fold at 4 h.



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Fig. 1. Effect of ascorbate, {alpha}-tocopherol or ascorbate plus {alpha}-tocopherol on the ß-pol level in mouse monocytes. Results are expressed as fold-increase over control in ß-pol protein level as measured by quantitative immunoblotting. Cells were treated with ascorbate (40 µM) ({circ}) or {alpha}-tocopherol (2 µM) ({triangleup}) or ascorbate (40 µM) plus {alpha}-tocopherol (2 µM) ({blacksquare}) and harvested at the indicated time. ß-Pol level was determined using the crude cell extract as described under Materials and methods.

 
Effect of Ox-LDL, LDL, ascorbate and {alpha}-tocopherol on ß-pol protein level
To test the effect of Ox-LDL on the ß-pol protein level in mouse monocytes, cells were exposed to 0.5 µg/ml Ox-LDL. Ox-LDL at this concentration did not have an effect on cell viability as measured by trypan blue exclusion (data not shown). The results indicated that the ß-pol level was reduced after 1 h exposure to Ox-LDL (Figure 2Go); ß-pol level decreased to ~50 and ~20% of the control level (without treatment) after exposure to Ox-LDL for 4 and 8 h, respectively.



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Fig. 2. Ascorbate plus {alpha}-tocopherol prevents down-regulation of ß-pol by Ox-LDL. ß-Pol level is expressed as percentage of the control. Cells were treated with Ox-LDL (0.5 µg/ml) alone ({circ}) or Ox-LDL (0.5 µg/ml) plus ascorbate (40 µM) and {alpha}-tocopherol (2 µM) ({blacksquare}). Cells were harvested at the indicated time. ß-Pol protein level was determined as described in Figure 1Go.

 
To determine whether the down-regulation of ß-pol is specific for Ox-LDL, the effect of non-oxidized LDL on the ß-pol level was determined. Cells were exposed to 0.5 µg/ml LDL and the ß-pol level was analyzed. LDL had no effect on the ß-pol protein level during 8 h of treatment (data not shown).

Since antioxidants both up-regulate ß-pol level in monocytes and potentially inactivate ROS, we studied whether treatment of antioxidants would influence the ß-pol down-regulation by Ox-LDL. Monocytes were co-treated with Ox-LDL and both antioxidants, and then the ß-pol level was analyzed. The results indicated that co-treatment with ascorbate and {alpha}-tocopherol prevented the down-regulation of ß-pol by Ox-LDL and resulted in a modest increase in ß-pol level as compared with the control (Figure 2Go).

Effect of ascorbate, {alpha}-tocopherol, LDL and Ox-LDL on BER
Previous studies demonstrated that ß-pol carries out short patch gap-filling DNA synthesis in an in vitro BER assay with an oligonucleotide substrate (41). Since ascorbate plus {alpha}-tocopherol and Ox-LDL differentially modulate expression of ß-pol in monocytes, we treated monocytes with the antioxidants or with Ox-LDL alone and also with antioxidants plus Ox-LDL and then examined BER activity in the crude cell-free extract. A 51 base pair oligonucleotide containing a G:U base pair at position 22 was used as substrate to study BER activity in vitro (41). BER activity was determined from the amounts of both the fully repaired 51 base pair molecule containing dCMP instead of dUMP and the unligated single nucleotide replacement intermediate (i.e. 22 nucleotide molecule with dCMP at position 22). With the untreated cell extract, most of the BER product corresponded with the 22 nucleotide-long unligated molecule, indicating that the mouse monocyte cell extract was relatively deficient in DNA ligase activity compared with bovine testis nuclear extract (41) or mouse fibroblast extract (43). The data show that ascorbate plus {alpha}-tocopherol up-regulated BER (Figure 3Go). This was especially evident from the amount of 51 nucleotide product formed. In contrast, Ox-LDL down-regulated BER (Figure 4Go). Further, co-treatment with ascorbate plus {alpha}-tocopherol prevented the down-regulation of BER by Ox-LDL (Figure 4Go). In contrast, non-oxidized LDL did not affect BER activity (data not shown).



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Fig. 3. Effect of ascorbate plus {alpha}-tocopherol on the in vitro BER activity of mouse monocyte extracts. Cells were treated with ascorbate (40 µM) plus {alpha}-tocopherol (2 µM) and harvested at the indicated time (h). Experiments were conducted as described under Materials and methods. An autoradiogram of a denaturing gel is shown. The two BER products are indicated: 51mer, the 51 nucleotide-long product of the overall BER reaction, and 22mer, the 22 nucleotide-long intermediate representing the product of single nucleotide gap filling replacement synthesis prior to ligation (i.e. [{alpha}-32P]dCMP incorporation).

 


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Fig. 4. Effect of time of treatment with Ox-LDL or Ox-LDL plus ascorbate/{alpha}-tocopherol on in vitro BER activity of mouse monocyte extracts. (A) Control, without treatment; (B) cells treated with Ox-LDL (0.5 µg/ml) alone; (C) cells treated with Ox-LDL (0.5 µg/ml) plus both antioxidants [ascorbate (40 µM) and {alpha}-tocopherol (2 µM)]. The two BER products, 51mer and 22mer, are as described in Figure 3Go. The period of treatment is shown at the bottom in hours.

 
Effect of LDL and Ox-LDL on DNA damage
To determine whether there is a correlation between the level of oxidative genomic DNA damage and the down-regulation of BER activity, monocytes were treated with LDL and Ox-LDL and analyzed for oxidative DNA damage by measuring the 8-oxo-G level. 8-oxo-G level (2.3x10–5) was significantly higher in monocytes exposed to Ox-LDL for 8 h than in untreated control cells or in cells treated with Ox-LDL for 4 h (1–3x10–6). Since antioxidants prevented down-regulation of ß-pol and BER by Ox-LDL (Figures 2 and 4GoGo), we examined the effect of antioxidants on the level of oxidative DNA damage in the cells. We found that ascorbate plus {alpha}-tocopherol prevented the Ox-LDL-induced accumulation of 8-oxo-G (8 h exposure) and that the antioxidants alone (4 or 8 h exposure) had no effect on the level of oxidative DNA damage as compared with the control.


    Discussion
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 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
It is increasingly evident that Ox-LDL contributes to the development of atherosclerosis. Ox-LDL is toxic in vitro to all cell types found in atherosclerotic lesions, including endothelial cells, smooth muscle cells and macrophages (16,17). The mechanism of toxicity is not known; however, it has been suggested that Ox-LDL generates ROS (4446). It is known that ROS cause oxidative DNA damage in cells. A previous study by Reid et al. (47) showed that a murine macrophage cell line exposed to a high concentration (100 µg/ml) of Ox-LDL for 24 h demonstrated DNA fragmentation and apoptosis. These results suggest that Ox-LDL may cause DNA damage and that apoptosis might be a mechanism of macrophage cell death in atherosclerotic lesions in the arterial wall (47).

The results presented in this paper demonstrate that Ox-LDL, but not LDL, down-regulates DNA BER activity in extracts of mouse monocytes and results in accumulation of oxidative DNA damage in genomic DNA. Co-treatment with antioxidants plus Ox-LDL prevents these effects, suggesting that an oxidative mechanism is involved in the Ox-LDL effect. Since unrepaired oxidative DNA damage may be harmful to cells, our findings provide useful insight into the potential mechanism of atherosclerotic plaque formation. Penn et al. (3537) have proposed that somatic cell mutations are involved in the etiology of atherosclerosis and in macrophage death (apoptosis) in advancing atherosclerotic plaques.

In these experiments, cells were treated with a low concentration (0.5 µg/ml) of Ox-LDL, that is more likely to be physiologically relevant than the high concentration (100 µg/ml) used by Reid et al. (47). Oxidative DNA damage was detected after 8 h of exposure to Ox-LDL; however, no 8-oxo-G was found after 4 h exposure. The DNA damage and the increased level of 8-oxo-G/dG are in agreement with earlier reports (4850). These results suggest that either the cellular DNA repair capacity is sufficient to repair the DNA damage after a 4 h exposure, or that the time required for Ox-LDL to have an effect on DNA damage is >4 h. Accumulation of oxidative DNA damage in mouse monocytes treated with Ox-LDL could be the result of the combination of oxidative stress and down-regulation of the ß-pol level and BER activity (Figures 2 and 4GoGo) since BER is the major pathway for the repair of oxidative DNA damage (7,2225). It would be interesting to determine whether Ox-LDL treatment would result in accumulation of a higher level of oxidative DNA damage and occur earlier in monocytes with a ß-pol gene deletion created by a tissue specific knock-out procedure (51).

The oxidative hypothesis of atherosclerosis presumes that compounds which limit vascular oxidative stress should also limit atherogenesis and atherosclerosis. It has been shown that ascorbate and {alpha}-tocopherol act synergistically to reduce free radicals through a mechanism by which ascorbate regenerates {alpha}-tocopherol, which in turn acts as the primary antioxidant (30). Therefore, both antioxidants have been widely used in animal models and human studies in an attempt to prevent atherosclerosis (2629). We examined the question of whether both antioxidants could prevent the down-regulation of ß-pol level and BER activity in monocytes treated with Ox-LDL. In contrast to the down-regulation of ß-pol and BER observed in the presence of Ox-LDL, ascorbate plus {alpha}-tocopherol protects against this down-regulation of ß-pol (Figure 2Go) and BER (Figure 4Go). The mechanism by which antioxidants prevent the down-regulation of ß-pol and BER by Ox-LDL could be the reduction of Ox-LDL to LDL by the antioxidants before cellular uptake. It is also possible that ascorbate and {alpha}-tocopherol could function in intracellular compartments to reduce Ox-LDL to LDL. Our results suggest that the latter mechanism is most practical since a combination of both antioxidants not only prevented the down-regulation of ß-pol and BER by Ox-LDL, but also up-regulated ß-pol and BER (Figures 1–4GoGoGoGo).

In summary, these results demonstrate a linkage between Ox-LDL exposure, reduced DNA repair capacity and increased oxidative DNA damage in monocytes. Understanding the function of DNA repair in monocytes exposed to Ox-LDL may provide new insights into oxidant-induced atherosclerosis.


    Acknowledgments
 
This work was supported in part by the NIH grant (ES-06492). We thank T.Y.Rawson for expert technical assistance, as well as L.Pipper and Dr F.He for preparation of figures.


    Notes
 
4 To whom correspondence should be addressed Email: wilson5{at}niehs-nih.gov Back


    References
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 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 

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Received August 31, 1999; revised January 12, 2000; accepted January 21, 2000.