Nicholas School of the Environment and Earth Sciences, Duke University, Durham, North Carolina 277080328
1 To whom correspondence should be addressed at Box 90328, Duke University, Research and Science Drives, Durham, NC 277080328. Fax: (919) 668-1799. E-mail: jonf{at}duke.edu.
Received September 28, 2004; accepted January 10, 2005
![]() |
ABSTRACT |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Key Words: PCB126; MAPK; ARE; HepG2; oxidative stress; signal transduction.
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
The specific molecular mechanism underlying PCBs' toxicity has not been resolved. It has been suggested that the molecular responses associated with exposure to coplanar PCBs are due to the interaction of these compounds with the aryl hydrocarbon receptor (AhR), and subsequent activation of the cytochrome P450 1A1 subfamily (Endo et al., 2003; Hennig et al., 1999
; Rushmore and Kong, 2002
; Safe and Krishnan, 1995
; Toborek et al., 1995
). Induction of CYP1A1 can lead to oxidative stress via the generation of reactive oxygen species (ROS) (Ramadass et al., 2003
; Stohs, 1990
). Recent studies suggest that oxidative stress may be involved in PCBs and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) toxicity (Hassoun et al., 2000
, 2002
; Howard et al., 2003
; Ramadass et al., 2003
). In contrast, noncoplanar PCBs exhibit AhR-independent effects, or antagonism of AhR-mediated CYP 1A1 induction (Kietz and Fischer, 2003
; Suh et al., 2003
).
There has been accumulating evidence supporting a role for ROS as second messengers that modulate gene transcription. Elevated levels of ROS may affect stress-responsive signal transduction cascades, including the mitogen-activated protein kinases (MAPKs) pathways. MAPKs are components of conserved signal transduction pathways that have been shown to participate in cellular differentiation, division, movement, and death (Schaeffer and Weber, 1999). MAPKs are protein serine/threonine kinases that include extracellular signal regulated kinases (ERKs), Jun N-terminal kinases or stress-activated protein kinases (JNKs), and p38. These kinases are activated in response to extracellular stimuli through phosphorylation by kinase kinases (Davis, 1995
).
PCB congeners have been shown to modulate the activities of stress-responsive signal transduction pathways. Exposure to PCB77 (3,3',4,4'-tetrachlorobiphenyl) increases JNK activity in cultured endothelial cells. This activity is modulated by intracellular glutathione content (Slim et al., 2000). Canesi et al. (2003)
reported that two ortho-substituted noncoplanar PCBs, PCB47 (2,2',4,4'-tetrachlorobiphenyl) and PCB153 (2,2',4,4',5,5'-hexachlorobiphenyl), can alter immune functions in mussel (Mytilus galloprovincialis Lam.) hemocytes; possibly by increasing p38 and JNK phosphorylation.
PCB126 (3,3', 4,4',5-pentacholorobiphenyl) is a non-ortho-chlorinated, coplanar congener, that has toxicities similar to those of TCDD. PCB126 has been shown to affect transcription via the AhR (Fukuzawa et al., 2003; Hassoun et al., 2001
). It has also been shown that exposure to PCB126 induces ROS in echinoderm cells and in mammalian hepatic and brain tissues (Coteur et al., 2001
; Hassoun et al., 2001
, 2002
). The ability of PCB126 to modulate the activities of stress-responsive signal transduction pathways and activating protein-1 (AP-1) has not been previously investigated. In the present study, we investigated the mechanism of PCB126 toxicity and the relation among PCB126 exposure, intracellular oxidative stress, and changes in MAPKs signaling in the human hepatoma cell line HepG2.
![]() |
MATERIALS AND METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Cytotoxicity assays were performed as previously described (Shokri et al., 2000). Briefly, cells were seeded onto 48-well tissue culture dishes in complete MEM media, and incubated for 24 h. After the 24-h incubation, various concentrations of PCB126 (in DMSO) were added, and the incubation continued for an additional 24 h. To determine cell viability, supernatants were removed; cells were rinsed with PBS, and then incubated for 3 h in neutral red solution in complete MEM. Cells were subsequently washed and then fixed with calcium chloride (1%, w/v) in formaldehyde (0.5%, v/v). To extract the dye from fixed cells, cells were lysed with acetic acid (1%, v/v) in 50% (v/v) ethanol. Optical density was measured at 540 nm. Positive controls, representing 100% cytotoxicity, consisted of cells exposed to distilled water. Cells incubated in complete MEM in the absence of PCB126 represented 100% viability, and were included as a negative control. Each treatment was performed in triplicate, and the experiments were repeated three times.
Transient transfection and reporter gene assay.
HepG2 cells were seeded onto 24-well dishes at a density of 2 x 104 cells/well, 24 h prior to transfection. The level of oxidative-stress-responsive transcription was measured using a chloramphenicol acetyltransferase (CAT) reporter gene that contains the antioxidant response element (ARE) from the rat NQO1 gene (pQR-ARE). pQR-ARE consists of a single copy of the ARE from NQO1 fused to a rat glutathione-S-transferase minimal promoter. As a negative control, a two-nucleotide mutant form of the ARE, which is unresponsive to oxidative stress was used (pM1) (Favreau and Pickett, 1991, 1995
). The pM1 plasmid is identical to pQR-ARE with the exception of a mutation of the GC box in the consensus ARE sequence (GTGACNNNGC).
Cells were transfected using LipofectAMINE according to the manufacturer's instructions (Life Technologies). pSV-ßGal plasmid (Promega) was cotransfected as an internal control to normalize for transfection efficiency. After 45 h incubation, the transfection mixture was replaced with the complete medium, and cells were incubated for an additional 24 h. In experiments where glutathione was depleted, cells were exposed to 0.5 mM buthionine sulfoximine (BSO) for 16 h prior to the addition of PCB126. After 24-h treatments with PCB126, CAT concentrations and ß-galactosidase activities were determined by sandwich ELISA using a CAT-ELISA kit (Roche Biotechnologies) and the ß-Galactosidase Enzyme Assay System (Promega), respectively. All assays were performed in triplicate, the experiments were repeated at least three times, and CAT protein levels were normalized by ß-galactosidase activity. The fold induction of the treated sample is calculated over that of the vehicle control.
Western immunoblot analysis.
HepG2 cells were exposed to 10 or 25 µM PCB126 for 24 h. Hydrogen peroxide (500 µM) was used as a positive control (reviewed in Torres and Forman, 2003). Cell lysates were prepared by combining PBS-washed cells with 1x SDS sample buffer; consisting of 62.5 mM TrisHCl (pH 6.8), 2% (w/v) SDS, 10% (v/v) glycerol, 50 mM DTT, and 0.1% (w/v) bromophenol blue. This mixture was sonicated for 1015 sec, to shear DNA and to reduce sample viscosity, and then heated at 95100°C for 5 min. Proteins were resolved on SDSpolyacrylamide gels and electrotransferred to Immobilon-P transfer membrane (Millipore). Immunoblotting was performed using polyclonal antibodies against phosphorylated and nonphosphorylated MAPKs (Cell Signaling Technology), and HPR-conjugated secondary antibodies (Amersham). For c-Jun, antibodies specific to phosphorylated c-Jun at serine 63 and 73 were used. As a loading control, blots were probed with an anti-mouse polyclonal antibody to ß-actin (Abcam). Antigen-antibody complexes were visualized using chemiluminescence (Cell Signaling Technology, Amersham), and then exposing the membrane to X-ray film. Target protein levels were measured by densitometry using Scion Image software (Scion Corporation). Data are expressed as fold induction of treated samples over that of the vehicle controls, normalized to the level of actin. Each experiment was repeated at least three times.
Kinase activity assays.
Kinase activity assays were performed using commercial assay kits by following the manufacturer's instructions (Cell Signaling Technology). Briefly, HepG2 cells were exposed to 25 µM PCB126 for 4 and 24 h, after which they were lysed in buffer consisting of 20 mM Tris, pH 7.4, 150 mM NaCl, 1 mM EDTA, 1 mM EGTA, 1% Triton, 2.5 mM sodium pyrophosphate, 1 mM ß glycerolphosphate, 1 mM Na3VO4, 1 µg/ml leupeptin, and 1 mM PMSF. Lysates were then sonicated four-times for 5 sec, centrifuged for 10 min at 14,000 xg and the supernatants collected. The kinase of interest was then isolated by precipitation. For JNK, N-terminal c-Jun (189) fusion protein bound to glutathione Sepharose beads was used. For ERK, immobilized phospho-p44/42 MAPK (Thr202/Tyr204) monoclonal antibody cross-linked to agarose beads was used. For p38, a monoclonal antibody to phospho-p38 (Thr180/Tyr182) was used. After precipitation, beads were washed twice with lysis buffer, and then twice with kinase buffer (25 mM Tris, pH 7.5, 5 mM ß-glycerolphosphate, 2 mM DTT, 0.1 mM Na3VO4, 10 mM MgCl2). Kinase reactions containing 250 µg of protein, 10 µCi of [-32P]-ATP and 25 µM of non-radioactive ATP were incubated at 30 °C for 45 min. The reactions were then terminated by the addition of 3 x SDS sample buffer. Samples were heated to 95100°C for 5 min, centrifuged at 14,000 xg for 2 min. Proteins were resolved by SDSpolyacrylamide gel electrophoresis. The amounts of 32P incorporated into the substrates were determined by PhosphoImager analysis using ImageQuant software (Molecular Dynamics). The data are expressed as fold-induction of treated samples over that of vehicle controls. Each experiment was repeated at least three times.
Statistical analysis.
All statistical analyses were performed using StatView software (SAS Institute Inc., Cary, NC). The results are presented as the mean ± standard mean error of three separate experiments. The significance of mean differences was detected by analysis of variance (ANOVA) followed by Fisher's Protected Least Squares Differences post hoc test for individual comparisons. The criterion for statistical significance was set at p < 0.05.
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Effect of PCB126 on ARE-Mediated Transcription
Transient transfection analyses, using the rat NQO1-based CAT reporter plasmids, pQR-ARE and pM1, were performed to investigate the transcriptional response of HepG2 cells after 24-h exposure to PCB126. Exposure to PCB126 increased ARE-mediated transcription 1.51-fold at 25 µM (p < 0.0001) and 1.48-fold at 10 µM (p < 0.0001) (Fig. 1). These results suggest that PCB126 can affect transcription through an oxidative stress response mechanism.
|
Effect of PCB126 on c-Jun Phosphorylation
The results from the reporter gene experiments indicate that PCB126 affects ARE-mediated transcription. The level of activity of AP-1 is controlled by the level of c-Jun phosphorylation. Thus, the effect of a 24-h exposure to PCB126 on the level of c-Jun phosphorylation was determined. PCB126 significantly increased c-Jun phosphorylation at both ser 63 and ser73 (Fig 2). At a PCB126 concentration of 10 µM, there was a 1.79-fold increase in phosphorylation and 2.09-fold increase at 25 µM (p = 0.0338) at ser73. Similarly, there was a 2.27-fold increase at 10 µM and 2.59-fold increase at 25 µM (p = 0.0338) on ser63.
|
|
|
|
Effect of PCB126 on MAPK Activity
The effect of PCB126 exposure on MAPK activities in HepG2 cells was examined following 4- and 24-h exposures to 25 µM PCB126. MAPKs can be activated by various chemicals including phenolic antioxidants and some PCBs, after short exposures (30 min to 5 h) (Canesi et al., 2003; Frigo et al., 2003
; Slim et al., 2000
; Yu et al., 1997
). PCB126 exposure did not cause a significant change in the activity of JNK; 1.2-fold after 24-h and 1.14-fold after 4-h (Fig. 6). ERK activity significantly increased by 1.49-fold (p = 0.0126) following 4-h PCB126 exposure (Fig. 7). After 24 h, ERK activity was similar to that of control. PCB126 exposure also caused a significant increase in p38 activity by 2.25-fold after 24-h exposure (p = 0.0071) and 3.14-fold after 4-h exposure (p = 0.0003) (Fig. 8). These results demonstrate that PCB126 can significantly activate ERK and p38. These observations are consistent with the effect of PCB126 on MAPK phosphorylation presented above.
|
|
|
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
The diverse array of diseases associated with PCB exposure suggests that multiple mechanisms may be involved in the toxicity of this class of environmental contaminants. A significant amount of research has focused on the effects of PCBs on the AhR pathway, or their role as endocrine disruptors (Binelli and Provini, 2003; Endo et al., 2003
; Hennig et al., 1999
; Olea et al., 1998
; Rushmore and Kong, 2002
; Safe and Krishnan, 1995
; Toborek et al., 1995
; Whaley et al., 2001
). Although the ability of PCB126 to induce a TCCD-like transcriptional response via the AhR pathway has been described, there is a paucity of data examining the capacity of this PCB congener to affect alternative signal transduction pathways.
Exposure to PCBs is correlated with a decrease in antioxidant defenses and an increase in the levels of ROS (Coteur et al., 2001; Hassoun et al., 2001
, 2002
; Twaroski et al., 2001
). ROS may be produced as a by-product of PCB-induced cytochrome P450 activity (Ramadass et al., 2003
; Stohs, 1990
). Alternatively, PCBs may be enzymatically oxidized to semiquinones or quinones, which can undergo redox cycling (McLean et al., 2000
; Srinivasan et al., 2002
). Thus, PCB toxicity may be mediated through oxidative stress, or the generation of ROS (Hassoun et al., 2000
, 2002
; Howard et al., 2003
; Ramadass et al., 2003
; Twaroski et al., 2001
).
To monitor the effects of chemical and biological toxicants on oxidative-stress-responsive transcription, reporter genes that contain antioxidant response elements (ARE) are routinely used. The ARE is a cis-acting, DNA sequence originally characterized in the promoters of the genes coding rat and mouse glutathione S-transferase Ya subunits (GST-Ya) (Rushmore et al., 1991). AREs are present in the promoters of the genes that encode for phase II detoxifying enzymes. AREs mediate the expression and coordinated induction of the NQO1 (Favreau and Pickett, 1991
) and GST Ya subunit genes in response to antioxidants, xenobiotics, oxidants, and hydrogen peroxide (Dalton et al., 1999
; Jeyapaul and Jaiswal, 2000
; Rushmore et al., 1991
). Exposure to PCB126 caused a significant increase in ARE-mediated transcription in HepG2 cells. Depletion of glutathione, which increases a cell's susceptibility to oxidative stress, prior to PCB126 exposure, enhanced the increase in oxidative stress inducible transcription.
Oxidative stress can cause an increase in AP-1 activity, i.e., the binding of the c-fos/c-jun heterodimer to AP-1 promoter elements (Dalton et al., 1999; Tharappel et al., 2002
; Turpaev, 2002
). It has been suggested that AP-1 may be an activator of ARE-mediated transcription (Bergelson et al., 1994
; Dalton et al., 1999
; Jaiswal, 1994
). The AP-1 family of transcription factors consists of homodimers and heterodimers of Jun, Fos, ATF2, or bZIP. AP-1 activity is controlled, in part by the level of c-Jun phosphorylation (Karin, 1995
). c-Jun is involved in mediating transcriptional activation elicited by extracellular stimuli and regulates the expression of genes involved in cell growth and differentiation (Pulverer et al., 1991
; Smeal et al., 1991
). Disruption or inappropriate activation of the AP-1 transcription complex could result in adverse health effects including cancer progression, neurodegenerative diseases, and inflammatory responses (Guo et al., 2002
; Hunot et al., 2004
; Luo et al., 1998
; Young et al., 2003
). Exposure of HepG2 cells to PCB126 resulted in a significant increase in c-Jun phosphorylation at both ser73 and ser63. The observed increase in c-Jun phosphorylation is consistent with the increase in ARE-mediated transcription. The PCB126-induced increase in c-Jun phosphorylation was not associated with an increase in the steady state level of c-Jun protein. The activation of c-Jun by PCB126 suggests a mechanistic link between PCB exposure and the etiology of several diseases.
Changes in the level of c-Jun phosphorylation are controlled by the activities of MAPKs. Activated MAPKs phosphorylate c-Jun on ser73 and ser63, which increases the stability of AP-1 heterodimers and potentates its transcriptional activity (Karin et al., 1997; Pulverer et al., 1991
; Smeal et al., 1991
, 1992
). The role of several different MAPKs in regulating AP-1 activity in response to a diverse array of extracellular stimuli has been investigated (Karin, 1995
; Karin et al., 1997
; Shaulian and Karin, 2002
). Since c-Jun is reported to be a common downstream effector of JNK, ERK, and p38 (Frigo et al., 2003
; Waskiewicz and Cooper, 1995
), it was reasonable to hypothesize that these kinases mediate PCB126 inducible increases in c-Jun phosphorylation in HepG2 cells. To determine which MAPKs are affected, changes in the levels of phosphorylation and the cognate activity of MAPKs following PCB126 exposure were investigated. PCB126 exposure caused a significant increase in ERK1/2 and p38 phosphorylation and kinase activity. The effects of PBC126 on c-Jun, ERK, and p38, and its limited effect on JNK are comparable to those observed following exposure to hydrogen peroxide, an archetypical cellular ROS (Clerk et al., 1998
; Guyton et al., 1996
; Kurata, 2000
). These results indicate that PCB126 is capable of activating MAPKs in HepG2 cells. In addition, they suggest that the MAPKs pathway is involved in PCB126 toxicity.
The toxicity of PCB126 has been shown to be similar to that of TCDD (Fukuzawa et al., 2003; Hassoun et al., 2001
). PCB126 has approximately 10% of TCDD's liver tumor-promoting potency in rats (Hemming et al., 1995
). Furthermore, the affect of PCB126 on MAPK signal transduction pathways is similar to that observed for TCDD. In Jurket and Hepa-1 cells, TCDD exposure caused an increase in the levels of JNK and ERK1/2 phosphorylation, without affecting p38 (Kwon et al., 2003
; Tan et al., 2002
). In additional to affecting the activity of c-Jun, ERK and p38 can also affect the activities of multiple transcription factors, either directly or indirectly, including PPAR
, ER STAT, Myc, Elk-1, CREB, SRF, Max, CHOP MEF2, and ATF2. Thus, PCB126-mediated activation of MAPK signal transduction pathways could have multiple and sometime contradictory effects on cell growth, differentiation, development, or apoptosis.
In summary, our results indicate that PCB126 can activate transcription by an AhR-independent mechanism. They suggest a model where PCB126 exposure increases ERK and p38 phosphorylation and activity. This increase in activity may be due to elevated levels of oxidative stress or other mechanism. The activated MAPKs subsequently increase c-Jun phosphorylation that increases AP-1 activity, which results in transcription of genes that are regulated via AP-1 consensus sequences. PCB126-activated MAPKs and AP-1 could affect the transcription of multiple stress-response and growth regulatory genes. It has been recently shown that exposure to PCB126 in rodents affects the steady-state level of expression of multiple genes. These genes are regulated by AhR-dependent and AhR-independent processes (Vezina et al., 2004). The inappropriate activation of these processes and genes could lead to the disruption of normal cell growth and the etiology of pathologies.
![]() |
ACKNOWLEDGMENTS |
---|
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Bergelson, S., Pinkus, R., and Daniel, V. (1994). Induction of AP-1 (Fos/Jun) by chemical agents mediates activation of glutathione S-transferase and quinone reductase gene expression. Oncogene 9, 565571.[ISI][Medline]
Binelli, A., and Provini, A. (2003). POPs in edible clams from different Italian and European markets and possible human health risk. Mar. Pollut. Bull. 46, 879886.[CrossRef][ISI][Medline]
Bosetti, C., Negri, E., Fattore, E., and La Vecchia, C. (2003). Occupational exposure to polychlorinated biphenyls and cancer risk. Eur. J. Cancer Prev. 12, 251255.[CrossRef][ISI][Medline]
Canesi, L., Ciacci, C., Betti, M., Scarpato, A., Citterio, B., Pruzzo, C., and Gallo, G. (2003). Effects of PCB congeners on the immune function of Mytilus hemocytes: Alterations of tyrosine kinase-mediated cell signaling. Aquat. Toxicol. 63, 293306.[CrossRef][ISI][Medline]
Clerk, A., Michael, A., and Sugden, P. H. (1998). Stimulation of multiple mitogen-activated protein kinase sub-families by oxidative stress and phosphorylation of the small heat shock protein, HSP25/27, in neonatal ventricular myocytes. Biochem. J. 333 (3), 581589.[ISI][Medline]
Coteur, G., Danis, B., Fowler, S. W., Teyssie, J. L., Dubois, P., and Warnau, M. (2001). Effects of PCBs on reactive oxygen species (ROS) production by the immune cells of Paracentrotus lividus (Echinodermata). Mar. Pollut. Bull. 42, 667672.[CrossRef][ISI][Medline]
Dalton, T. P., Shertzer, H. G., and Puga, A. (1999). Regulation of gene expression by reactive oxygen. Annu. Rev. Pharmacol. Toxicol. 39, 67101.[CrossRef][ISI][Medline]
Davis, R. J. (1995). Transcriptional regulation by MAP kinases. Mol. Reprod. Dev. 42, 459467.[CrossRef][ISI][Medline]
Endo, F., Monsees, T. K., Akaza, H., Schill, W. B., and Pflieger-Bruss, S. (2003). Effects of single non-ortho, mono-ortho, and di-ortho chlorinated biphenyls on cell functions and proliferation of the human prostatic carcinoma cell line, LNCaP. Reprod. Toxicol. 17, 229236.[CrossRef][ISI][Medline]
Favreau, L. V., and Pickett, C. B. (1991). Transcriptional regulation of the rat NAD(P)H:quinone reductase gene. Identification of regulatory elements controlling basal and inducible expression by planar aromatic compounds and phenolic antioxidants. J. Biol. Chem. 266, 45564561.
Favreau, L. V., and Pickett, C. B. (1995). The rat quinone reductase antioxidant response element. Identification of the nucleotide sequence required for basal and inducible activity and detection of antioxidant response element-binding proteins in hepatoma and non-hepatoma cell lines. J. Biol. Chem. 270, 2446824474.
Frigo, D. E., Tang, Y., Beckman, B. S., Scandurro, A. B., Alam, J., Burow, M. E., and McLachlan, J. A. (2004). Mechanism of AP-1-mediated gene expression by select organochlorines through the p38 MAPK pathway. Carcinogenesis 25, 249261.
Fukuzawa, N. H., Ohsako, S., Nagano, R., Sakaue, M., Baba, T., Aoki, Y., and Tohyama, C. (2003). Effects of 3,3',4,4',5-pentachlorobiphenyl, a coplanar polychlorinated biphenyl congener, on cultured neonatal mouse testis. Toxicol. In Vitro 17, 259269.[ISI][Medline]
Golden, R., Doull, J., Waddell, W., and Mandel, J. (2003). Potential human cancer risks from exposure to PCBs: A tale of two evaluations. Crit. Rev. Toxicol. 33, 543580.[ISI][Medline]
Guo, R. F., Lentsch, A. B., Sarma, J. V., Sun, L., Riedemann, N. C., McClintock, S. D., McGuire, S. R., Van Rooijen, N., and Ward, P. A. (2002). Activator protein-1 activation in acute lung injury. Am. J. Pathol. 161, 275282.
Guyton, K. Z., Liu, Y., Gorospe, M., Xu, Q., and Holbrook, N. J. (1996). Activation of mitogen-activated protein kinase by H2O2. Role in cell survival following oxidant injury. J. Biol. Chem. 271, 41384142.
Hardell, L., Van Bavel, B., Lindstrom, G., Carlberg, M., Eriksson, M., Dreifaldt, A. C., Wijkstrom, H., Starkhammar, H., Hallquist, A., et al. (2004). Concentrations of polychlorinated biphenyls in blood and the risk for testicular cancer. Int. J. Androl. 27, 282290.[CrossRef][ISI][Medline]
Hassoun, E. A., Li, F., Abushaban, A., and Stohs, S. J. (2000). The relative abilities of TCDD and its congeners to induce oxidative stress in the hepatic and brain tissues of rats after subchronic exposure. Toxicology 145, 103113.[CrossRef][ISI][Medline]
Hassoun, E. A., Li, F., Abushaban, A., and Stohs, S. J. (2001). Production of superoxide anion, lipid peroxidation and DNA damage in the hepatic and brain tissues of rats after subchronic exposure to mixtures of TCDD and its congeners. J. Appl. Toxicol. 21, 211219.[CrossRef][ISI][Medline]
Hassoun, E. A., Wang, H., Abushaban, A., and Stohs, S. J. (2002). Induction of oxidative stress in the tissues of rats after chronic exposure to TCDD, 2,3,4,7,8-pentachlorodibenzofuran, and 3,3',4,4',5- pentachlorobiphenyl. J. Toxicol. Environ. Health A 65, 825842.[CrossRef][ISI][Medline]
Hemming, H., Bager, Y., Flodstrom, S., Nordgren, I., Kronevi, T., Ahlborg, U. G., and Warngard, L. (1995). Liver tumour promoting activity of 3,4,5,3',4'-pentachlorobiphenyl and its interaction with 2,3,7,8-tetrachlorodibenzo-p-dioxin. Eur. J. Pharmacol. 292, 241249.[Medline]
Hennig, B., Slim, R., Toborek, M., and Robertson, L. W. (1999). Linoleic acid amplifies polychlorinated biphenyl-mediated dysfunction of endothelial cells. J. Biochem. Mol. Toxicol. 13, 8391.[CrossRef][ISI][Medline]
Howard, A. S., Fitzpatrick, R., Pessah, I., Kostyniak, P., and Lein, P. J. (2003). Polychlorinated biphenyls induce caspase-dependent cell death in cultured embryonic rat hippocampal but not cortical neurons via activation of the ryanodine receptor. Toxicol. Appl. Pharmacol. 190, 7286.[CrossRef][ISI][Medline]
Howsam, M., Grimalt, J. O., Guino, E., Navarro, M., Marti-Rague, J., Peinado, M. A., Capella, G., and Moreno, V. (2004). Organochlorine exposure and colorectal cancer risk. Environ. Health Perspect. 112, 14601466.[ISI][Medline]
Hunot, S., Vila, M., Teismann, P., Davis, R. J., Hirsch, E. C., Przedborski, S., Rakic, P., and Flavell, R. A. (2004). JNK-mediated induction of cyclooxygenase 2 is required for neurodegeneration in a mouse model of Parkinson's disease. Proc. Natl. Acad. Sci. U.S.A. 101, 665670.
Jaiswal, A. K. (1994). Jun and Fos regulation of NAD(P)H: quinone oxidoreductase gene expression. Pharmacogenetics 4, 110.[ISI][Medline]
Jeyapaul, J., and Jaiswal, A. K. (2000). Nrf2 and c-Jun regulation of antioxidant response element (ARE)-mediated expression and induction of gamma-glutamylcysteine synthetase heavy subunit gene. Biochem. Pharmacol. 59, 14331439.[CrossRef][ISI][Medline]
Karin, M. (1995). The regulation of AP-1 activity by mitogen-activated protein kinases. J. Biol. Chem. 270, 1648316486.
Karin, M., Liu, Z., and Zandi, E. (1997). AP-1 function and regulation. Curr. Opin. Cell Biol. 9, 240246.[CrossRef][ISI][Medline]
Kietz, S., and Fischer, B. (2003). Polychlorinated biphenyls affect gene expression in the rabbit preimplantation embryo. Mol. Reprod. Dev. 64, 251260.[CrossRef][ISI][Medline]
Kurata, S. (2000). Selective activation of p38 MAPK cascade and mitotic arrest caused by low level oxidative stress. J. Biol. Chem. 275, 2341323416.
Kwon, M. J., Jeong, K. S., Choi, E. J., and Lee, B. H. (2003). 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD)-induced activation of mitogen-activated protein kinase signaling pathway in Jurkat T cells. Pharmacol. Toxicol. 93, 186190.[CrossRef][ISI][Medline]
Luo, Y., Umegaki, H., Wang, X., Abe, R., and Roth, G. S. (1998). Dopamine induces apoptosis through an oxidation-involved SAPK/JNK activation pathway. J. Biol. Chem. 273, 37563764.
McLean, M. R., Twaroski, T. P., and Robertson, L. W. (2000). Redox cycling of 2-(x'-mono, -di, -trichlorophenyl)- 1, 4- benzoquinones, oxidation products of polychlorinated biphenyls. Arch. Biochem. Biophys. 376, 449455.[CrossRef][ISI][Medline]
Muto, T., Wakui, S., Imano, N., Nakaaki, K., Takahashi, H., Hano, H., Furusato, M., and Masaoka, T. (2002). Mammary gland differentiation in female rats after prenatal exposure to 3,3',4,4',5-pentachlorobiphenyl. Toxicology 177, 197205.[CrossRef][ISI][Medline]
Olea, N., Pazos, P., and Exposito, J. (1998). Inadvertent exposure to xenoestrogens. Eur. J. Cancer Prev. 7 (Suppl. 1), S17S23.[ISI][Medline]
Pavuk, M., Cerhan, J. R., Lynch, C. F., Schecter, A., Petrik, J., Chovancova, J., and Kocan, A. (2004). Environmental exposure to PCBs and cancer incidence in eastern Slovakia. Chemosphere 54, 15091520.[CrossRef][ISI][Medline]
Pulverer, B. J., Kyriakis, J. M., Avruch, J., Nikolakaki, E., and Woodgett, J. R. (1991). Phosphorylation of c-jun mediated by MAP kinases. Nature 353, 670674.[CrossRef][ISI][Medline]
Ramadass, P., Meerarani, P., Toborek, M., Robertson, L. W., and Hennig, B. (2003). Dietary flavonoids modulate PCB-induced oxidative stress, CYP1A1 induction, and AhR-DNA binding activity in vascular endothelial cells. Toxicol. Sci. 76, 212219.
Rier, S. E. (2002). The potential role of exposure to environmental toxicants in the pathophysiology of endometriosis. Ann. N.Y. Acad. Sci. 955, 201212; discussion 230 232, 396406.
Rushmore, T. H., and Kong, A. N. (2002). Pharmacogenomics, regulation and signaling pathways of phase I and II drug metabolizing enzymes. Curr. Drug Metab. 3, 481490.[ISI][Medline]
Rushmore, T. H., Morton, M. R., and Pickett, C. B. (1991). The antioxidant responsive element. Activation by oxidative stress and identification of the DNA consensus sequence required for functional activity. J. Biol. Chem. 266, 1163211639.
Safe, S., and Krishnan, V. (1995). Cellular and molecular biology of aryl hydrocarbon (Ah) receptor-mediated gene expression. Arch. Toxicol. Suppl. 17, 99115.[Medline]
Safe, S. H. (1994). Polychlorinated biphenyls (PCBs): Environmental impact, biochemical and toxic responses, and implications for risk assessment. Crit. Rev. Toxicol. 24, 87149.[ISI][Medline]
Schaeffer, H. J., and Weber, M. J. (1999). Mitogen-activated protein kinases: Specific messages from ubiquitous messengers. Mol. Cell. Biol. 19, 24352444.
Shaulian, E., and Karin, M. (2002). AP-1 as a regulator of cell life and death. Nat. Cell Biol. 4, E131E136.[CrossRef][ISI][Medline]
Shokri, F., Heidari, M., Gharagozloo, S., and Ghazi-Khansari, M. (2000). In vitro inhibitory effects of antioxidants on cytotoxicity of T-2 toxin. Toxicology 146, 171176.[CrossRef][ISI][Medline]
Slim, R., Toborek, M., Robertson, L. W., Lehmler, H. J., and Hennig, B. (2000). Cellular glutathione status modulates polychlorinated biphenyl-induced stress response and apoptosis in vascular endothelial cells. Toxicol. Appl. Pharmacol. 166, 3642.[CrossRef][ISI][Medline]
Smeal, T., Binetruy, B., Mercola, D., Grover-Bardwick, A., Heidecker, G., Rapp, U. R., and Karin, M. (1992). Oncoprotein-mediated signalling cascade stimulates c-Jun activity by phosphorylation of serines 63 and 73. Mol. Cell. Biol. 12, 35073513.[Abstract]
Smeal, T., Binetruy, B., Mercola, D. A., Birrer, M., and Karin, M. (1991). Oncogenic and transcriptional cooperation with Ha-Ras requires phosphorylation of c-Jun on serines 63 and 73. Nature 354, 494496.[CrossRef][ISI][Medline]
Srinivasan, A., Robertson, L. W., and Ludewig, G. (2002). Sulfhydryl binding and topoisomerase inhibition by PCB metabolites. Chem. Res. Toxicol. 15, 497505.[CrossRef][ISI][Medline]
Stohs, S. J. (1990). Oxidative stress induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Free Radic. Biol. Med. 9, 7990.[ISI][Medline]
Suh, J., Kang, J. S., Yang, K. H., and Kaminski, N. E. (2003). Antagonism of aryl hydrocarbon receptor-dependent induction of CYP1A1 and inhibition of IgM expression by di-ortho-substituted polychlorinated biphenyls. Toxicol. Appl. Pharmacol. 187, 1121.[CrossRef][ISI][Medline]
Tan, Z., Chang, X., Puga, A., and Xia, Y. (2002). Activation of mitogen-activated protein kinases (MAPKs) by aromatic hydrocarbons: Role in the regulation of aryl hydrocarbon receptor (AHR) function. Biochem. Pharmacol. 64, 771780.[CrossRef][ISI][Medline]
Tharappel, J. C., Lee, E. Y., Robertson, L. W., Spear, B. T., and Glauert, H. P. (2002). Regulation of cell proliferation, apoptosis, and transcription factor activities during the promotion of liver carcinogenesis by polychlorinated biphenyls. Toxicol. Appl. Pharmacol. 179, 172184.[CrossRef][ISI][Medline]
Toborek, M., Barger, S. W., Mattson, M. P., Espandiari, P., Robertson, L. W., and Hennig, B. (1995). Exposure to polychlorinated biphenyls causes endothelial cell dysfunction. J. Biochem. Toxicol. 10, 219226.[Medline]
Torres, M., and Forman, H. J. (2003). Redox signaling and the MAP kinase pathways. Biofactors 17, 287296.[ISI][Medline]
Turpaev, K. T. (2002). Reactive oxygen species and regulation of gene expression. Biochemistry (Mosc.) 67, 281292.[CrossRef][ISI][Medline]
Twaroski, T. P., O'Brien, M. L., and Robertson, L. W. (2001). Effects of selected polychlorinated biphenyl (PCB) congeners on hepatic glutathione, glutathione-related enzymes, and selenium status: Implications for oxidative stress. Biochem. Pharmacol. 62, 273281.[CrossRef][ISI][Medline]
Vezina, C. M., Walker, N. J., and Olson, J. R. (2004). Subchronic exposure to TCDD, PeCDF, PCB126, and PCB153: Effect on hepatic gene expression. Environ. Health Perspect. 112, 16361644.[ISI][Medline]
Waskiewicz, A. J., and Cooper, J. A. (1995). Mitogen and stress response pathways: MAP kinase cascades and phosphatase regulation in mammals and yeast. Curr. Opin. Cell Biol. 7, 798805.[CrossRef][ISI][Medline]
Whaley, D. A., Keyes, D., and Khorrami, B. (2001). Incorporation of endocrine disruption into chemical hazard scoring for pollution prevention and current list of endocrine disrupting chemicals. Drug Chem. Toxicol. 24, 359420.[CrossRef][ISI][Medline]
Yoshizawa, K., Walker, N. J., Jokinen, M. P., Brix, A. E., Sells, D. M., Marsh, T., Wyde, M. E., Orzech, D., Haseman, J. K., and Nyska, A. (2005). Gingival carcinogenicity in female Harlan Sprague-Dawley rats following two-year oral treatment with 2,3,7,8-tetrachlorodibenzo-p-dioxin and dioxin-like compounds. Toxicol. Sci. 83, 6477.
Young, M. R., Yang, H. S., and Colburn, N. H. (2003). Promising molecular targets for cancer prevention: AP-1, NF-kappa B and Pdcd4. Trends Mol. Med. 9, 3641.[CrossRef][ISI][Medline]
Yu, R., Tan, T. H., and Kong, A. T. (1997). Butylated hydroxyanisole and its metabolite tert-butylhydroquinone differentially regulate mitogen-activated protein kinases. The role of oxidative stress in the activation of mitogen-activated protein kinases by phenolic antioxidants. J. Biol. Chem. 272, 2896228970.
Zeiger, M., Haag, R., Hockel, J., Schrenk, D., and Schmitz, H. J. (2001). Inducing effects of dioxin-like polychlorinated biphenyls on CYP1A in the human hepatoblastoma cell line HepG2, the rat hepatoma cell line H4IIE, and rat primary hepatocytes: Comparison of relative potencies. Toxicol. Sci. 63, 6573.