Prevention of the epoxy resin-based root canal sealers-induced cyclooxygenase-2 expression and cytotoxicity of human osteoblastic cells by various antioxidants

Cyclooxygenase-2 (COX-2) is an inducible enzyme believed to be responsible for prostaglandin synthesis at the site of inflammation. Recently, the activation of COX-2 expression may be one of the important pathogenesis of root canal sealers-induced periapical inflammation. However, little is known about whether chemical interaction can modulate the epoxy resin-based root canal sealers-induced cytotoxicity as well as COX-2 expression. The aim of the present study was to investigate the effects of antioxidants catalase, superoxide dismutase (SOD), and N-acetyl-L-cysteine (NAC) on AH26- and Topseal-induced COX-2 mRNA gene and cytotoxicity in human osteoblastic cell line U2OS cells. The results showed that both epoxy resin-based root canal sealers were cytotoxic to U2OS cells in a concentration-dependent manner (p<0.05). AH26 and Topseal were found to induce COX-2 mRNA gene expression in U2OS cells. The addition of glutathione (GSH) precursor NAC led to decrease the induction of COX-2 mRNA gene expression and cytotoxicity by both AH26 and Topseal (p<0.05). However, catalase and SOD lacked the ability to prevent AH26-and Topseal-induced cytotoxicity and COX-2 mRNA gene expression (p>0.05). Taken together, the activation of COX-2 mRNA gene expression may be one of the pathogenesis of epoxy resin-based root canal sealers-induced periapical inflammation. In addition, GSH depletion, but not the attack of oxygen free radicals, could be the mechanism for epoxy resin-based root canal sealers-induced cytotoxicity and COX-2 mRNA gene expression. Factors that induce GSH synthesis may appear useful in preventing cell damage mediated by epoxy resin-based root canal sealers.     

Root canal sealers cause cytotoxicity and oxidative damage in hepatocytes

Elutable substances and degradation products from root canal sealers may gain access to periodontal tissue in many ways. The aim of this study was to investigate damage and oxidative stress caused by treatment with root canal sealers in primary cultured rat hepatocytes. Four root canal sealers were used: Canals, Sealapex, AH 26, and AH Plus. Toxicity was detected by assessing lactate dehydrogenase (LDH) leakage, and oxidative stress was determined by analysis of glutathione (GSH) levels. One-way analysis of variance was performed on the data. As the concentrations of sealer were increased, LDH leakage increased (p < 0.01). Increased GSH depletion accompanied LDH leakage increase. GSH levels were reduced to zero at a concentration of 0.05% for Canals and 0.10% for AH 26. These results confirmed the hypothesis that root canal sealers can cause cytotoxicity in hepatocytes. The Canals and AH 26 sealers can also cause oxidative stress in hepatocytes.     

Protective effect of NAC on formaldehyde-containing-ZOE-based root-canal-sealers-induced cyclooxygenase-2 expression and cytotoxicity in human osteoblastic cells

Cyclooxygenase-2 (COX-2) is an inducible enzyme believed to be responsible for prostaglandin synthesis at site of inflammation. Recently, the activation of COX-2 expression may be one of the important pathogenesis of root-canal-sealers-induced periapical inflammation. However, little is known about whether chemical interaction can modulate the COX-2 expression and cytotoxicity induced by formaldehyde-containing-ZOE-based root canal sealers. The aim of this study was to investigate the effects of antioxidants such as catalase, superoxide dismutase (SOD), and N-acetyl-L-cysteine (NAC) on N2- and endomethasone-induced COX-2 mRNA gene and cytotoxicity in human osteoblastic cell line U2OS cells. Our data demonstrated that both formaldehyde-containing-ZOE-based root canal sealers were found to induce COX-2 mRNA gene expression in U2OS cells. The addition of glutathione (GSH) precursor NAC led to decrease the induction of COX-2 mRNA gene expression and cytotoxicity by both N2 and Endomethasone (p < 0.05). However, catalase and SOD lacked the ability to prevent cytotoxicity and COX-2 mRNA gene expression induced by N2 and Endomethasone (p > 0.05). The data presented here demonstrated that the activation of COX-2 mRNA gene expression may be one of the pathogenesis of formaldehyde-containing-ZOE-based root-canal-sealers-induced periapical inflammation. In addition, GSH depletion, but not the attack of oxygen free radicals, could be the mechanism for cytotoxicity and COX-2 mRNA gene expression induced by formaldehyde-containing-ZOE-based root canal sealers. NAC appears as a useful agent in protecting cell damage mediated by formaldehyde-containing-ZOE-based root canal sealers.     

Cytotoxicity of formaldehyde on human osteoblastic cells is related to intracellular glutathione levels

Formaldehyde that leaches out of formaldehyde-releasing root canal sealers, specifically from setting material extruded into the periapical region may participate in the development of periapical inflammation or the continuation of a pre-existing periapical lesion. However, the effects of formaldehyde on human osteoblasts have not been investigated. The aim of this study was to evaluate the mechanisms of cytotoxicity of formaldehyde on human osteoblastic cell line U2OS in vitro. Cytotoxicity and cell proliferation assays were performed to elucidate the adverse effects of formaldehyde on U2OS cells. Formaldehyde demonstrated a cytotoxic effect to U2OS cells in a dose-dependent manner (p<0.05). The 50% inhibition concentration of formaldehyde was about 3 mM. Formaldehyde also inhibited cell proliferation during a 3-day culture period (p<0.05). To determine whether glutathione (GSH) levels were important in the cytotoxicity of formaldehyde, we pretreated cells with the GSH precursor, 2-oxothiazolidine-4-carboxylic acid (OTZ) to boost thiol levels, or buthionine sulfoximine (BSO) to deplete GSH. The addition of OTZ acted as a protective effect on the formaldehyde-induced cytotoxicity (p<0.05). In contrast, the addition of BSO enhanced the formaldehyde-induced cytotoxicity (p<0.05). Taken together, the levels of formaldehyde tested inhibited cell growth and proliferation on U2OS cells. Formaldehyde has significant potential for periapical toxicity. These inhibitory effects were associated with intracellular GSH levels.     

Sealing properties of different resin-based root canal sealers

In this study, a comparison of the sealing properties of three different resin-based sealers, AH26, Diaket, and EndoREZ with those of the root canal sealer zinc oxide-eugenol-based U/P Root Canal Sealer was performed. Eighty extracted single-rooted human mandibular first premolar teeth were used. The coronal part of each tooth was removed at the apical 17-mm of roots. The root canals were prepared to a size 7 with ProFile 0.06 taper Series 29 rotary instruments in a crown-down manner before lateral condensation of gutta-percha with the tested sealers (n = 20). A fluid filtration method was used for quantitative evaluation of apical leakage. Statistical analysis revealed that there were no significant differences between the resin-based sealers groups (AH26, Diaket, and EndoREZ) (p > 0.05). U/P Root Canal Sealer, zinc oxide-eugenol-based sealer, leaked significantly more than other tested groups (p < 0.05). Under the limitations of this study, resin-based root canal sealers were found more effective in sealing root canals than the zinc oxide-eugenol-based sealer.     

Inhibitory effects of root canal sealers on the expression of inducible nitric oxide synthase in lipopolysaccharide-stimulated murine macrophage cells

Excessive production of nitric oxide (NO) is associated with inflammation. In the present study, we examined the effects of root canal sealers (N2 Universal, Sealapex, and AH26) on NO production and inducible NO synthase (iNOS) expression in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages. Root canal sealers decreased NO synthesis in LPS-induced RAW 264.7 macrophages in a dose-dependent manner. RT-PCR and Western blot analysis demonstrated markedly lower levels of iNOS mRNA and protein in LPS-activated macrophage cells treated with root canal sealers compared with untreated cells. From these results, we conclude that root canal sealers do not inhibit NO synthesis by direct inhibition of the enzyme, but rather through inhibition of iNOS mRNA expression (leading to a decrease in iNOS protein expression). Our data, therefore, suggest that root canal sealers may be an effective inhibitor of LPS-induced inflammatory effects in macrophage cells. Further in vitro and in vivo studies are necessary to confirm the effects of root canal sealers on the inflammatory processes.     

Assessment of the genotoxicity of resin and zinc-oxide eugenol-based root canal sealers using an in vitro mammalian test system.

Endodontic sealers frequently are placed in direct contact with living tissues. Thus root canal sealers should have good biocompatibility. However, the pathobiologic effects, especially genotoxicity, of various root canal sealers widely used in dentistry have not been studied systematically on eukaryotic cells. The aim of this study was to examine the genotoxic potential of resin and zinc oxide-eugenol-based root canal sealers using a mammalian test system. In this study, tetrazolium bromide reduction assay, DNA precipitation assay, and DNA fragmentation analysis were performed to investigate the cytotoxic and genotoxic effects of four different root canal sealers on cultured V79 cells. The results show that all the root canal sealers tested are cytotoxic to V79 cells. The toxicity decreased in the order of N2 > AH26 > AHplus > Canals. In addition, N2, AH26, and AHplus exhibited genotoxicity by causing DNA single-strand breaks and digestion of genomic DNA. However, N2 is the most toxic root canal sealer among those tested. Taken together, these findings suggest that the sealers evaluated in this study show different toxic effects depending on the types and components. Root canal sealers containing formaldehyde and bisphenol A diglyether proved to be not only cytotoxic but also genotoxic. This mammalian test system might be used routinely for evaluation of the genotoxicity of dental materials in future investigations.     

Induction of c-fos and c-jun protooncogenes expression by formaldehyde-releasing and epoxy resin-based root-canal sealers in human osteoblastic cells.

An important requirement for a root-canal sealer is biologic compatibility; most evaluations have focused on general toxicological and local tissue irritating properties. There is only scant information about mutagenicity or carcinogenicity testing for root-canal sealer. It has been shown that c-fos and c-jun are induced rapidly by a variety of chemical and physical stimuli. Numerous works have extensively investigated the induction mechanisms of c-fos and c-jun protooncogenes by these agents; however, little is known about the induction of cellular signaling events and specific gene expression after cell exposure to root-canal sealers. Therefore, we used osteoblastic cell line U2-OS to examine the effect of zinc-oxide eugenol-based (N2 and Endomethasome), epoxy resin-based (AH Plus), and calcium hydroxide-based (Sealapex) root-canal sealers on the expression of c-fos and c-jun protooncogenes to understand in more detail the molecular mechanisms of root-canal sealer-induced genotoxicity. The cytotoxicity decreased in an order of N2 > Endomethasome > AH Plus > Sealapex. In addition, N2, Endomethasome, and AH Plus rapidly induced c-jun and c-fos mRNA levels in cells. However, Sealapex did not induce c-jun and c-fos mRNA expression at detectable levels all time points. Taken together, persistent induction of c-jun and c-fos protooncogenes by formaldehyde-releasing and epoxy resin-based root-canal sealers may be distributed systemically via apex to cause some unexpected adverse effects on human beings. These data should be taken into consideration when choosing a root-canal sealer.     

莱菔硫烷诱导胰岛系INS-1细胞血红素氧合酶1的表达****◆○

背景：莱菔硫烷可用于氧化应激相关疾病的治疗,血红素氧合酶是一种催化血红素降解的应激蛋白,已经成为预防氧化攻击的首选研究靶标之一。目的：观察核因子E2相关因子2激动剂莱菔硫烷对大鼠胰岛细胞系INS-1细胞血红素氧合酶1蛋白表达作用及细胞保护机制。方法：体外培养INS-1细胞,先用3 μmol/L莱菔硫烷进行干预培养3 h,再分别加入不同的胰岛素抵抗诱导剂葡萄糖氧化酶、地塞米松和葡萄糖胺刺激建立胰岛素抵抗细胞模型。结果与结论：3 μmol/L莱菔硫烷处理INS-1细胞中血红素氧合酶1的表达增加(P < 0.05),其效应在4 h后达到高峰(P < 0.05)。3 μmol/L莱菔硫烷的预处理可逆转由胰岛素抵抗诱导剂导致的血红素氧合酶1表达下调(P < 0.05)。而且在葡萄糖胺处理的INS-1细胞中,莱菔硫烷对血红素氧合酶1的表达改善与磷酸化PKB的表达上调具有正相关性(P < 0.05, r = 0.23)。结果证实,莱菔硫烷可能通过诱导核因子E2相关因子2介导的血红素氧合酶1表达,增强胰岛细胞抗氧化防御功能和抗损伤信号系统,从而达到拮抗胰岛素抵抗诱导剂对胰岛细胞的损伤作用。     

The influence of glutathione on redox regulation by antioxidant proteins and apoptosis in macrophages exposed to 2-hydroxyethyl methacrylate (HEMA)

Resin monomers like 2-hydroxyethyl methacrylate (HEMA) disturb cell functions including responses of the innate immune system, mineralization and differentiation, or induce cell death via apoptosis. These phenomena are associated with oxidative stress and a reduction in the concentration of the antioxidant glutathione (GSH), resulting in imbalanced redox homeostasis. Thus far, the precise mechanism of how resin monomers interfere with cellular redox regulation is unknown. The present study provides insight into the induction of apoptosis and the differential expression of antioxidant enzymes depending on the availability of GSH. Buthionine sulfoximine (BSO) was used to inhibit GSH synthesis, while 2-oxothiazolidine-4-carboxylate (OTC), and N-acetylcysteine (NAC) as prodrugs supported GSH synthesis in RAW264.7 mouse macrophages exposed to HEMA (0-8 mm) for 24 h. The level of GSH was significantly decreased after cells were preincubated with BSO, and the formation of reactive oxygen species (ROS) increased in cultures subsequently exposed to HEMA. Apoptosis was drastically increased by BSO in HEMA-exposed cell cultures as well, but OTC and NAC retracted HEMA-induced cell death. These results show that dental monomer-induced apoptosis is causally related to the availability of GSH. The hydrogen peroxide decomposing enzymes glutathione peroxidase (GPx1/2) and catalase were differentially regulated in HEMA-exposed cultures. Expression of GPx1/2 was inhibited by HEMA and further reduced in the presence of BSO. SOD1 (superoxide dismutase) expression was inhibited in the presence of HEMA, and was decreased to an even greater extent by BSO, possibly due to H(2)O(2)-feedback inhibition. The expression of catalase was considerably up-regulated in HEMA-exposed cultures, implying that H(2)O(2) is the type of ROS that is significantly increased in monomer-exposed cells. OTC and NAC counteracted the effect of HEMA on GPx1/2, SOD1, and catalase expression. HO-1 (heme oxygenase) expression was strongly enhanced by HEMA, suggesting the need for further antioxidants like bilirubin to support enzyme activities that directly regulate H(2)O(2) equilibrium. Expression of the oxidoreductase thioredoxin (TRX1), the second major thiol-dependent antioxidant system in eukaryotic cells, was slightly reduced, while the oxygen-sensing protein HIF-1α was downregulated in HEMA-exposed cell cultures. These results indicate that cells and tissues actively respond to monomer-induced oxidative stress by the differential expression of enzymatic antioxidants.     

The biocompatibility evaluation of epoxy resin-based root canal sealers in vitro.

The cytotoxic and mutagenic effects of epoxy resin-based root canal sealer AH26 and AH-Plus were determined in vitro. Root canal sealers were eluted for 24 h in dimethyl sulfoxide (DMSO) and diluted in culture medium. Cytotoxic effects were assessed using the MTT [tetrazolium dye, 3-(4.5-dimethylthiazol-2-yl)-2.5-diphenyltetrazolium bromide, C18H16N5SBr] assay for mitochondrial enzyme activity and also the cell viability. Genotoxicity assays were assessed using the alkaline single cell gel electrophoresis assay (comet assay) for DNA damage measurement. Result indicated that both the AH26 and AH-Plus sealers exhibited a dose-dependent increase in astrocyte toxic effects. Additionally, dose-dependent astrocyte DNA damage was also noted for both sealers. Therefore, these epoxy resin-based sealers, AH26 and AH-Plus demonstrated both cytotoxicity and genotoxicity in vitro.     

Glutathione deficiency intensifies ischaemia-reperfusion induced cardiac dysfunction and oxidative stress.

The efficacy of glutathione (GSH) in protecting ischaemia-reperfusion (I-R) induced cardiac dysfunction and myocardial oxidative stress was studied in open-chest, stunned rat heart model. Female Sprague-Dawley rats were randomly divided into three experimental groups: (1) GSH-depletion, by injection of buthionine sulphoxamine (BSO, 4 mmol kg(-1), i.p.) 24 h prior to I-R, (2) BSO injection (4 mmol kg(-1), i.p.) in conjunction with acivicin (AT125, 0.05 mmol kg(-1), i.v.) infusion 1 h prior to I-R, and (3) control (C), receiving saline treatment. Each group was further divided into I-R, with surgical occlusion of the main left coronary artery (LCA) for 30 min followed by 20 min reperfusion, and sham. Myocardial GSH content and GSH : glutathione disulphide (GSSG) ratio were decreased by approximately 50% (P < 0.01) in both BSO and BSO + AT125 vs. C. Ischaemia-reperfusion suppressed GSH in both left and right ventricles of C (P < 0.01) and left ventricles of BSO and BSO + AT125 (P < 0.05). Contractility (+dP/dt and -dP/dt) in C heart decreased 55% (P < 0.01) after I and recovered 90% after I-R, whereas +/-dP/dt in BSO decreased 57% (P < 0.01) with ischaemia and recovered 76 and 84% (P < 0.05), respectively, after I-R. For BSO + AT125, +/-dP/dt were 64 and 76% (P < 0.01) lower after ischaemia, and recovered only 67 and 61% (P < 0.01) after I-R. Left ventricular systolic pressure in C, BSO and BSO + AT125 reached 95 (P > 0.05) 87 and 82% (P < 0.05) of their respective sham values after I-R. Rate-pressure double product was 11% (P > 0.05) and 25% (P < 0.05) lower in BSO and BSO + AT125, compared with Saline, respectively. BSO and BSO + AT125 rats demonstrated significantly lower liver GSH and heart Mn superoxide dismutase activity than C rats after I-R. These data indicate that GSH depletion by inhibition of its synthesis and transport can exacerbate cardiac dysfunction inflicted by in vivo I-R. Part of the aetiology may involve impaired myocardial antioxidant defenses and whole-body GSH homeostasis.     

Redox regulation of p38 MAPK activation and expression of ICAM-1 and heme oxygenase-1 in human alveolar epithelial (A549) cells

We have explored the potential role of redox events in p38 mitogen-activated protein kinase (MAPK) activation and their relevance to the inducible expression of intercellular adhesion molecule-1 (ICAM-1) and heme oxygenase-1 (HO-1) in A549 cells. Tumor necrosis factor-alpha (TNFalpha) and hydrogen peroxide (H2O2) both activated p38, but only TNFalpha activated nuclear factor-kappaB (NF-kappaB). N-Acetyl-L-cysteine (20 mM) inhibited both H2O2- and TNFalpha-induced p38 phosphorylation (14 +/- 7 and 37 +/- 4% of control, respectively). The mitochondrial complex I and III inhibitors, rotenone and antimycin A, and allopurinol partially inhibited H2O2- but not TNFalpha-induced p38 activation. However, rotenone and antimycin A augmented intracellular oxidative stress measured by dichlorofluorescein fluorescence. TNFalpha, but not H2O2, induced ICAM-1 in A549 cells, which was attenuated by a proteasome inhibitor, but not by the p38 MAPK inhibitor SB203580. In contrast, hemin and hemoglobin, but neither TNFalpha nor H2O2, caused efficient HO-1 expression. However, hemin had no effect on p38 activation and SB203580 did not influence hemin-induced HO-1 protein expression. Collectively, these data suggest that p38 is a cytokine- and oxidative stress-responsive pathway in A549 cells. Whereas NF-kappaB appears crucial in ICAM-1 induction, p38 activation itself is not sufficient to confer HO-1 expression and may not be involved in HO-1 and ICAM-1 induction in A549 cells.     

Mycoplasma fermentans MALP-2 Induces Heme Oxygenase-1 Expression via Mitogen-Activated Protein Kinases and Nrf2 Pathways To Modulate Cyclooxygenase 2 Expression in Human Monocytes

Heme oxygenase-1 (HO-1) is a stress-inducible rate-limiting enzyme in heme degradation that confers cytoprotection against oxidative injury and performs a vital function in the maintenance of cell hemostasis. Increasing numbers of reports have indicated that mycoplasma-derived membrane lipoproteins/lipopeptides, such as macrophage-activating lipopeptide-2 (MALP-2), function as agents that stimulate the immune system by producing various inflammatory mediators, such as cytokines and cyclooxygenase 2 (COX-2), which play roles in the pathogenesis of inflammatory responses during mycoplasma infection. Here, we report that MALP-2 induced HO-1 mRNA and protein expression and upregulated HO-1 enzyme activity in THP-1 cells. Specific inhibitors of mitogen-activated protein kinases (MAPKs), SB203580, PD98059, and SP600125, significantly abolished HO-1 expression. In addition, MALP-2 also induced NF-E2-related factor 2 (Nrf2) translocation, and the silencing of Nrf2 expression in THP-1 cells decreased the levels of MALP-2-mediated HO-1 expression. Furthermore, COX-2 protein expression levels were upregulated in THP-1 cells in response to MALP-2, and transfection with small interfering RNAs of HO-1 significantly increased COX-2 accumulation. These results demonstrate that MALP-2 induces HO-1 expression via MAPKs and Nrf2 pathways and, furthermore, that MALP-2-induced COX-2 expression was modulated by HO-1 in THP-1 cells.     

Glutathione deficiency intensifies ischaemia‐reperfusion induced cardiac dysfunction and oxidative stress

The efficacy of glutathione (GSH) in protecting ischaemia‐reperfusion (I‐R) induced cardiac dysfunction and myocardial oxidative stress was studied in open‐chest, stunned rat heart model. Female Sprague–Dawley rats were randomly divided into three experimental groups: (1) GSH‐depletion, by injection of buthionine sulphoxamine (BSO, 4 mmol kg^–1, i.p.) 24 h prior to I‐R, (2) BSO injection (4 mmol kg^–1, i.p.) in conjunction with acivicin (AT125, 0.05 mmol kg^–1, i.v.) infusion 1 h prior to I‐R, and (3) control (C), receiving saline treatment. Each group was further divided into I‐R, with surgical occlusion of the main left coronary artery (LCA) for 30 min followed by 20 min reperfusion, and sham. Myocardial GSH content and GSH : glutathione disulphide (GSSG) ratio were decreased by ?50% (P < 0.01) in both BSO and BSO + AT125 vs. C. Ischaemia‐reperfusion suppressed GSH in both left and right ventricles of C (P < 0.01) and left ventricles of BSO and BSO + AT125 (P < 0.05). Contractility (+dP/dt and –dP/dt) in C heart decreased 55% (P < 0.01) after I and recovered 90% after I‐R, whereas ±dP/dt in BSO decreased 57% (P < 0.01) with ischaemia and recovered 76 and 84% (P < 0.05), respectively, after I‐R. For BSO + AT125, ±dP/dt were 64 and 76% (P < 0.01) lower after ischaemia, and recovered only 67 and 61% (P < 0.01) after I‐R. Left ventricular systolic pressure in C, BSO and BSO + AT125 reached 95 (P > 0.05) 87 and 82% (P < 0.05) of their respective sham values after I‐R. Rate‐pressure double product was 11% (P > 0.05) and 25% (P < 0.05) lower in BSO and BSO + AT125, compared with Saline, respectively. BSO and BSO + AT125 rats demonstrated significantly lower liver GSH and heart Mn superoxide dismutase activity than C rats after I‐R. These data indicate that GSH depletion by inhibition of its synthesis and transport can exacerbate cardiac dysfunction inflicted by in vivo I‐R. Part of the aetiology may involve impaired myocardial antioxidant defenses and whole‐body GSH homeostasis.     

Intracellular glutathione regulates tumour necrosis factor-α-induced p38 MAP kinase activation and RANTES production by human bronchial epithelial cells

BACKGROUND: RANTES plays an important role in the production of allergic inflammation of the airway through its chemotactic activity for eosinophils. The cellular reduction and oxidation (redox) changes are involved in the activation of p38 mitogen-activated protein (MAP) kinase and the induction of cytokine expression. It has previously been shown that tumour necrosis factor (TNF)-MA activates p38 mitogen-activated protein (MAP) kinase to produce cytokine, including RANTES, that N-acetylcysteine (NAC) attenuates cytokine production by human bronchial epithelial cells (BECs), and that sensitivity to TNFalpha is inversely correlated with cellular redox state. However, a role of cellular redox regulated by intracellular glutathione (GSH) in TNFalpha-induced p38 MAP kinase activation and p38 MAP kinase-mediated RANTES production by human BECs has not been determined. OBJECTIVE: Human BECs were exposed to NAC or buthionine sulfoximine (BSO). TNFalpha-induced p38 MAP kinase activation and p38 MAP kinase-mediated RANTES production by human BECs were then examined in order to clarify these issues. RESULTS: The results showed that: NAC attenuated TNFalpha-induced p38 MAP kinase activation and RANTES production; SB 203580 as the specific inhibitor of p38 MAP kinase activity attenuated TNF-alpha-induced RANTES production; BSO facilitated TNF-alpha-induced p38 MAP kinase activation and RANTES production; SB 203580 attenuated BSO-mediated facilitation of TNF-alpha-induced RANTES production; and the intracellular GSH increased in NAC-treated cells, whereas the intracellular GSH was reduced in BSO-treated cells. CONCLUSIONS: These results indicate that cellular redox regulated by GSH is critical for TNF-alpha-induced p38 MAP kinase activation and p38 MAP kinase-mediated RANTES production by human BECs.