Expression of gamma-aminobutyric acid type A receptor beta3 subunit in murine cleft palate induced by 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin / 中华口腔医学杂志

Objective@#To investigate the expression of gamma-aminobutyric acid type A receptor beta3 subunit (GABRB3) on cleft palate in C57BL/6J mice induced by 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD).@*Methods@#Sixty C57BL/6J pregnant mice on gestation day (GD) 10.5 were divided into two groups: one group was administered through gastric tubes one dose of 28 μg/kg TCDD (experimental group) and the other group was administered through gastric tubes one dose of 5.6 ml/kg corn oil (control group). Embryos were removed by cesarean section from pregnant mice during the palatal formation stage (GD 13.5-17.5) and the palatal tissue studied in morphological and histological observation. The relative mRNA and protein expression of GABRB3 was measured by real-time quantitative PCR and Western blotting. Localization of GABRB3 protein was measured by immunohistochemistry or immunofluorescence.@*Results@#The incidence of cleft palate at GD17.5 was 100% in experimental group and there was no cleft palate occurred in the control group (0); elevation of palatine processes in experimental group was completed on GD15.5 which was clearly delayed by a day compared with that in control group. On GD14.5-GD17.5, the mRNA expression (0.561±0.073, 0.728±0.104, 0.782±0.137, 0.686±0.145) and protein expression (0.288±0.013, 0.404±0.017, 0.399±0.012, 0.307±0.010) in the experimental group were significantly lower than the control group mRNA expression (0.818±0.088, 0.865±0.086, 1.021±0.054, 1.163±0.179) and protein expression (0.481±0.017, 0.456±0.009, 0.474±0.016, 0.529±0.015)(P<0.05). Immunohistochemistry and immunofluorescence showed that GABRB3 was mainly expressed in the mesenchymal cells and medial edge epithelium.@*Conclusions@#TCDD delayed palatal shelf elevation and eventually led to cleft palate may be associated with a decrease in GABRB3. GABRB3 may play an important role in the elevation and fusion phases of the palate development.

The beneficial effects of Montelukast against 2 3 7 8 tetrachlorodibenzo dioxin toxicity in female reproductive system in rats

ABSTRACT PURPOSE: To determine the toxic effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on reproductive system and the beneficial effects of Montelukast (ML) with histological and biochemical analysis. METHODS: Rats were randomly divided into four equal groups (control, TCDD, ML and TCDD+ML). Tissue samples were collected on day 60 and oxidative status and histological alterations were analyzed. RESULTS: The results showed a significant increase in oxidative and histological damage on uterine and ovarian tissues. Otherwise, the oxidative and histological damages caused by TCDD were prevented with ML treatment. CONCLUSION: The toxic effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin on female reproductive system were reversed with Montelukast treatment. Therefore, we claimed that ML treatment might be useful for TCDD toxicity.

Study on the balance rebuilding of regulatory T cells/Th17 cells by 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin for the treatment of rheumatoid arthritis / 中华风湿病学杂志

Objective To investigate the regulatory T cells (Treg) induced by 2, 3, 7, 8-tetrachloro-dibenzo-p-dioxin (TCDD) on the aryl hydrocarbon receptor activator from na?ve T cells, in collagen-induced arthritis (CIA)-rheumatoid arthritis (RA) mouse in the correction of rheumatoid arthritis pathological process on Th17/Treg cell imbalance. Methods ① Rat CIA model was established by bovine Ⅱ collagen injection. The rats were divided into 3 groups: the normal group (N), the CIA model group (C), and the TCDD group (T).② The percentage of Th17 and Treg from peripheral blood mononuclear cells (PBMCs) were analyzed with flow cytometry (FCM). Th17 and Treg cells related cytokine including interleukin (IL)-1β, IL-6, IL-17A, IL-23, tumor necrosis factor (TNF)-α and transforming growth factor (TGF)-β were measured with enzyme linked immunosorbent assay (ELISA). ③ The changes of the arthritis parameters, and radiological of T/NT before and after treatment were observed. ④ After two weeks of treatment, the rats were killed. The swollen joints were used for pathological assessment and arthritic pathology injury score was evaluated. ⑤ All data were analyzed by one-way analysis of variance (ANOVA) test and t test. Results In group C:When the second immunization was compared with the first, the percentage of Th17 cells was significantly increased [(2.99 ±0.16)%, (4.02 ± 0.33)%, t=-6.82, P<0.01], while Treg cells were significantly reduced [(2.67 ±0.57)%, (1.79 ±0.39)%, t=3.11, P=0.000], Th17/Treg was significantly increased (1.14 ±0.14, 2.27 ±0.39, t=-4.53, P=0.039). In group T, the percentage of Th17 cells were significantly reduced [(2.69±0.24)%, (1.21±0.25)%, t=5.12, P<0.01] before and after treatment, while Treg cells were significantly increased [(2.76 ±0.36)%, (3.78 ±0.22)%, t=-6.24, P<0.01], Th17/Treg was significantly reduced (1.08±0.07, 0.21±0.05, t=-11.92, P=0.027). In group C:When the second immunization was compared with the first, the Th17 cells related cytokines were significantly increased inclu-ding IL-1 [(96±12) pg/ml, (153±18) pg/ml, t=-13.24, P<0.01], IL-6 [(246±14) pg/ml, (295±15) pg/ml, t=-9.41, P=0.000], IL-17 [(76 ±14) pg/ml, (99 ±16) pg/ml, t=-5.78, P<0.01], IL-23 [(85 ±9) pg/ml, (102 ±11 ( pg/ml, t=-11.71, P<0.01], TNF-α [(303 ±12) pg/ml, (345 ±17) pg/ml, t=-15.56, P=0.007], while Treg cells related cytokines TGF-β was significantly reduced [(42 ±7) ng/ml, (35 ±8) ng/ml, t=7.52, P=0.012]. Th17 cells related cytokines were significantly reduced including IL-1 [(93 ±10) pg/ml, (79 ±8) pg/ml, t=10.52, P<0.01], IL-6 [(245 ±11) pg/ml, (151 ±8) pg/ml, t=19.23, P<0.01], IL-17 [(76 ±10) pg/ml, (62 ±9) pg/ml, t=7.37, P=0.005], IL-23 [(84 ±11) pg/ml, (38 ±6) pg/ml, t=14.48, P=0.000], TNF-α [(294 ±8) ng/ml, (195 ±9) ng/ml, t=23.35, P<0.01], while Treg cells related cytokine TGF-β was significantly increased (41 ±8, 61 ±10, t=-10.61, P=0.000. Conclusion TCDD can increase Treg cells, reduce Th17 cells, so the Th17/Treg cell in RA patho-genesis could be re-balanced.

Serum 2,3,7,8-Tetrachlorodibenzo-p-dioxin Levels and Their Association With Age, Body Mass Index, Smoking, Military Record-based Variables, and Estimated Exposure to Agent Orange in Korean Vietnam Veterans / 예방의학회지

OBJECTIVES: The aim of this study was to examine the levels of serum 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and evaluate their association with age, body mass index, smoking, military record-based variables, and estimated exposure to Agent Orange in Korean Vietnam veterans. METHODS: Serum levels of TCDD were analyzed in 102 Vietnam veterans. Information on age, body mass index, and smoking status were obtained from a self-reported questionnaire. The perceived exposure was assessed by a 6-item questionnaire. Two proximity-based exposures were constructed by division/brigade level and battalion/company level unit information using the Stellman exposure opportunity index model. RESULTS: The mean and median of serum TCDD levels was 1.2 parts per trillion (ppt) and 0.9 ppt, respectively. Only 2 Vietnam veterans had elevated levels of TCDD (>10 ppt). The levels of TCDD did not tend to increase with the likelihood of exposure to Agent Orange, as estimated from either proximity-based exposure or perceived self-reported exposure. The serum TCDD levels were not significantly different according to military unit, year of first deployment, duration of deployment, military rank, age, body mass index, and smoking status. CONCLUSIONS: The average serum TCDD levels in the Korean Vietnam veterans were lower than those reported for other occupationally or environmentally exposed groups and US Vietnam veterans, and their use as an objective marker of Agent Orange exposure may have some limitations. The unit of deployment, duration of deployment, year of first deployment, military rank, perceived self-reported exposure, and proximity-based exposure to Agent Orange were not associated with TCDD levels in Korean Vietnam veterans. Age, body mass index and smoking also were not associated with TCDD levels.

Effect of Selective Cyclooxygenase 2 Inhibitor in TCDD Pre-exposed Thyroid Papillary Carcinoma Cell Line

BACKGROUND: Cyclooxygenase 2 (COX-2) is related to carcinogenesis and progression of cancer. COX-2 has been detected in thyroid cancer. This suggests that COX-2 inhibitor may be useful to control the growth of thyroid cancer cells as well as the progression of thyroid cancer. Tetrachlorodibenzodioxin (TCDD), acting as an inflammatory cytokine, directly induces the expression of COX-2. We examine whether TCDD controls the effect of COX-2 inhibitor on thyroid cancer cells. METHODS: The effects of TCDD and celecoxib on thyroid papillary carcinoma cell line (SNU790) were examined using cell proliferation and fluorescence-activated cell sorting analysis. Western blot analysis was performed to determine the expressed COX-2 levels and the cell cycle-related proteins. The matrix metalloproteinase-2 (MMP-2) expression and gelatinolytic activity were examined using real time-polymerase chain reaction and zymography. RESULTS: TCDD directly induced the growth of SNU790 and the expression of cyclin D1, cyclin A, cyclin E, p21 and COX-2. Celecoxib suppressed the growth of SNU790 and the expression of cyclin D1 and cyclin E. Celecoxib reduced the MMP-2 expression and the gelatinolytic activity, but those effects were decreased in the SNU790 by either pre-treatment with TCDD or co-treatment with TCDD and celecoxib. CONCLUSIONS: Celocoxib effect is directly reduced depending on the exposure to TCDD. TCDD exposure should be considered in the treatment with Celecoxib.

Signaling pathway for 2,3,7,8-tetrachlorodibenzo- p-dioxin-induced TNF-alpha production in differentiated THP-1 human macrophages

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD), a prototypic halogenated aromatic hydrocarbon (HAH), is known as one of the most potent toxicants. At least a part of its toxic effects appears to be derived from its ability to induce TNF-alpha production. However, the signaling pathway of TCDD that leads to TNF-alpha expression has not been elucidated. In this study, we investigated the signaling mechanism of TCDD-induced TNF-alpha expression in PMA-differentiated THP-1 macrophages. TCDD induced both mRNA and protein expression of TNF-alpha in a dose- and time-dependent manner. Alpha-Naphthoflavone (NF), an aryl hydrocarbon receptor (AhR) inhibitor, prevented the TCDD-induced expression of TNF-alpha at both mRNA and protein levels. Genistein, a protein tyrosine kinase (PTK) inhibitor, and PD153035, an EGFR inhibitor, also blocked the increase of TNF-alpha expression by TCDD, indicating the role of EGFR in TCDD-induced TNF-alpha expression. On the other hand, PP2, a c-Src specific inhibitor, did not affect TCDD-induced TNF-alpha expression. EGFR phosphorylation was detected as early as 5 min after TCDD treatment. TCDD-induced EGFR activation was AhR-dependent since co-treatment with alpha-NF prevented it. ERK was found to be a downstream effector of EGFR activation in the signaling pathway leading to TNF-alpha production after TCDD stimulation. Activation of ERK was observed from 30 min after TCDD treatment. PD98059, an inhibitor of the MEK-ERK pathway, completely prevented the TNF-alpha mRNA and protein expression induced by TCDD, whereas inhibitors of JNK and p38 MAPK had no effect. PD153035, an EGFR inhibitor, as well as alpha-NF significantly reduced ERK phosphorylation, suggesting that ERK activation by TCDD was mediated by both EGFR and AhR. These results indicate that TNF-alpha production by TCDD in differentiated THP-1 macrophages is AhR-dependent and involves activation of EGFR and ERK, but not c-Src, JNK, nor p38 MAPK. A signaling pathway is proposed where TCDD induces sequential activation of AhR, EGFR and ERK, leading to the increased expression of TNF-alpha.