Effects of superoxide dismutase and polyclonal tumor necrosis factor-alpha antibodies on chloroacetate-induced cellular death and superoxide anion production by J774.A1 macrophages

Dichloroacetate (DCA) and trichloroacetate (TCA) are by-products that are formed during the process of water chlorination and have been previously shown to induce superoxide anion (SA) production and cellular death when added to J774.A1 macrophage cultures. In this study, the effects of superoxide dismutase (SOD) and polyclonal tumor necrosis factor-alpha (TNF-alpha) antibodies on DCA- and TCA-induced SA production and cellular death have been tested on the J774.A1 macrophage cultures. TCA and DCA were added to different cultures either alone, each at a concentration of 16 mM, or in combination with SOD (2-12 units/ml), or with TNF-alpha antibodies (10 and 25 units/ml). Cells were incubated for 48 h, after which cellular death/viability, lactate dehydrognase (LDH) leakage by the cells, and SA production by the cells were determined. While TCA and DCA caused significant cellular toxicity, indicated by reduction in cellular viability and increases in LDH leakage and SA production, SOD addition resulted in significant reduction of the effects induced by the compounds. On the other hand, addition of TNF-alpha antibodies to the DCA- and TCA-treated cultures resulted in significant reduction of DCA- but not TCA-induced cellular death and SA production by the cells. Although these results suggest a significant role for SA in DCA- and TCA-induced cellular death, they may also suggest two different mechanisms for the chloroacetate-induced SA production by the cells.     

The induction of oxidative stress and cellular death by the drinking water disinfection by-products,dichloroacetate and trichloroacetate in J774.A1 cells

The in vitro toxicity of the drinking water disinfection by products dichloroacetate (DCA) and trichloroacetate (TCA) were studied using the J774A.1 macrophage cell line. DCA and TCA were added to cell cultures at concentrations ranging between 8-32 mM and incubated for 24, 36 and 60 h. DCA and TCA effects on cellular viability, lactate dehydrogenase (LDH) release and superoxide anion (SA) production by the cells, as well as superoxide dismutase (SOD) activities of the cells were determined. DCA and TCA caused time- and concentration-dependent increases in cellular death, in LDH release and production of SA by the cells. The compounds also caused modulations in SOD activities of the cells, with increases observed at the lower concentrations and/or shorter periods of incubations and suppression with the higher concentrations and/or longer periods of incubation. The results of the study indicate that DCA and TCA induce macrophage activation and that the activation is associated with cellular toxicity. Also, DCA and TCA are found to be equitoxic to J774.A1 cells.     

The effect of a single dose of tumor necrosis factor alpha inhibitor on gut permeability in rats during exposure to a heat stress

Heat stroke is a life threatening illness characterized by a core body temperature of >40 °C, delirium and convulsions, and often results in multi-organ dysfunction, due to the release of endotoxin through the intestinal wall into the circulation. While playing a major role in the gastrointestinal tract permeability changes seen in Crohn's disease, it is not clear whether tumor necrosis factor alpha (TNF-α) mediates the increase in intestinal permeability and the release of endotoxin into the circulation in heat stroke. The aim of the present study was to determine the acute effects of a single dose of TNF-α antibody on gut permeability in rats during heat stress. Fifty-five Sprague-Dawley rats (28 male and 27 female) were treated with either saline or infliximab (a monoclonal antibody to TNF-α), anesthetized with pentobarbitone (50 mg kg⁻¹) and then exposed to either normothermic conditions or an ambient temperature of between 41 and 42 °C for 70 min. Fluorescent isothiocyanate labeled dextrans (FITC-dextrans) were administered intragastrically as a marker of intestinal permeability. Liver enzymes, endotoxin and TNF-α were analyzed in the blood. Exposure to a heat stress significantly increased intestinal permeability to FITC-dextrans compared to the controls (P<0.05). Infliximab did not have an effect on the intestinal permeability to the FITC-dextrans. Heat stress had no significant effect on liver enzymes or endotoxin concentration versus controls (P>0.05). TNF-α was not detectable in any of the samples. TNF-α did not mediate the release of endotoxin into the circulation after an acute bout of heat stroke.     

Differential effects of tumor necrosis factor and asbestos fibers on manganese superoxide dismutase induction and oxidant-induced cytotoxicity in human mesothelial cells

We compared induction of manganese superoxide dismutase (MnSOD) by asbestos fibers and tumor necrosis factor (TNF) using cultures human mesothelial cells. Transformed pleural mesothelial cells (MET 5A) were exposed for 48 h to amosite asbestos fibers (2 g/cm²), to TNF (10 Ng/ml), and to the combination of these two. TNF and amosite+TNF caused significant MnSOD mRNA upregulation. Similarly MnSOD specific activity was increased by TNF (290% increase) and the amosite+TNF combination (313% increase) but not by amosite alone. In cell injury experiments, amosite and amosite+TNF exposures caused significant cell membrane injury when assessed by lactate dehydrogenase release, which was 31% and 57% higher than in the unexposed cells. However, only the amosite+TNF combination caused significant depletion of cellular high-energy nucleotide when expressed as percentage of [¹⁴C]denine labeling in cellular high-energy nucleotides. The nucleotide levels were 91.5 ± 2.0% in the unexposed cells, 89.9 ± 3.9% in amosite-exposed cells, 90.1 ± 2.2% in TNF-exposed cells, and 79.8 ± 9.4% in amosite+TNF-exposed Amosite+TNF-exposed cells were also most sensitive to menadione (20 mol/L, 2 h), a compound which generates superoxide radicals intracellularly. In conclusion, our data suggests that in human mesothelial cells inflammatory cytokines but not asbestos fibers alone can cause MnSOD induction. In this study, however amosite asbestos+TNF treatment rendered these cells more vulnerable to oxidant-induced cell damage despite elevated MnSOD activity.Abbreviations MnSOD manganese superoxide dismutase - TNF tumor necrosis factor - LDH lactate dehydrogenase     

A novel molecule ‘shati’ increases dopamine uptake via the induction of tumor necrosis factor‐α in pheochromocytoma‐12 cells

The psychostimulant properties of methamphetamine (METH) are associated with an increase in extracellular dopamine (DA) levels in the brain, via facilitation of DA’s release from pre‐synaptic nerve terminals and inhibition of its reuptake through DA transporter. Recently, we have demonstrated that tumor necrosis factor‐α (TNF‐α) increases DA uptake and inhibits METH dependence. Moreover, we have clarified ‘shati’ identified in the nucleus accumbens of mice treated with METH is involved in METH dependence. In the present study, we investigated the effects of TNF‐α on DA uptake in PC12 cells and established a PC12 cell line transfected with a vector containing shati cDNA to examine the precise mechanism behind the role of shati in DA uptake. Moreover, we examined the relationship between shati and TNF‐α. TNF‐α increased DA uptake via the mitogen‐activated protein kinase kinase pathway and inhibited the METH‐induced decrease in DA uptake in PC12 cells. Transfection of the vector containing shati cDNA into PC12 cells, induced the expression of shati and TNF‐α mRNA, accelerated DA uptake, and inhibited the METH‐induced decrease in DA uptake. These results suggest that the functional roles of shati in METH‐regulated behavioral changes are mediated through inhibition of the METH‐induced decrease in DA uptake via TNF‐α.     

Tumour necrosis factor‐α‐induced protein 8‐like 2 is a novel regulator of proliferation,migration, and invasion in human rectal adenocarcinoma cells

Tumour necrosis factor‐α‐induced protein 8‐like 2 (TIPE2) is a tumour suppressor in many types of cancer. However, the mechanism of action of TIPE2 on the growth of rectal adenocarcinoma is unknown. Our results showed that the expression levels of TIPE2 in human rectal adenocarcinoma tissues were higher than those in adjacent non‐tumour tissues. Overexpression of TIPE2 reduced the proliferation, migration, and invasion of human rectal adenocarcinoma cells and down‐regulation of TIPE2 showed reverse effects. TIPE2 overexpression increased apoptosis through down‐regulating the expression levels of Wnt3a, phospho (p)‐β‐Catenin, and p‐glycogen synthase kinase‐3β in rectal adenocarcinoma cells, however, TIPE2 knockdown exhibited reverse trends. TIPE2 overexpression decreased autophagy by reducing the expression levels of p‐Smad2, p‐Smad3, and transforming growth factor‐beta (TGF‐β) in rectal adenocarcinoma cells, however, TIPE2 knockdown showed opposite effects. Furthermore, TIPE2 overexpression reduced the growth of xenografted human rectal adenocarcinoma, whereas TIPE2 knockdown promoted the growth of rectal adenocarcinoma tumours by modulating angiogenesis. In conclusion, TIPE2 could regulate the proliferation, migration, and invasion of human rectal adenocarcinoma cells through Wnt/β‐Catenin and TGF‐β/Smad2/3 signalling pathways. TIPE2 is a potential therapeutic target for the treatment of rectal adenocarcinoma.