Apoptosis is induced in Chlamydia trachomatis infected HEp-2 cells by the addition of a combination innate immune activation compounds and the inhibitor wedelolactone

Problem: Innate immune activation of human cells, for some intracellular pathogens, is advantageous for vacuole morphology and pathogenic viability. It is unknown whether innate immune activation is advantageous to Chlamydia trachomatis viability. ----- ----- Method of study: Innate immune activation of HEp-2 cells during Chlamydia infection was conducted using lipopolysaccharide (LPS), polyI:C, and wedelolactone (innate immune inhibitor) to investigate the impact of these conditions on viability of Chlamydia. ----- ----- Results: The addition of LPS and polyI:C to stimulate activation of the two distinct innate immune pathways (nuclear factor kappa beta and interferon regulatory factor) had no impact on the viability of Chlamydia. However, when compounds targeting either pathway were added in combination with the specific innate immune inhibitor (wedelolactone) a major impact on Chlamydia viability was observed. This impact was found to be due to the induction of apoptosis of the HEp-2 cells under these conditions. ----- ----- Conclusion: This is the first time that induction of apoptosis has been reported in C. trachomatis-infected cells when treated with a combination of innate immune activators and wedelolactone.

Cytotoxicity evaluation of three pairs of hexabromocyclododecane (HBCD) enantiomers on Hep G2 cell

Hexabromocyclododecanes (HBCDs) are additive brominated flame retardants mainly used in plastics and textiles. At the present time, these compounds are found in almost all environmental and human samples. In order to evaluate the environmental safety and health risk of HBCDs, the enantiomerically pure alpha-, beta-, and gamma-HBCD were prepared using high performance liquid chromatography (HPLC) on a PM-P-CD column and the cytotoxicities of their enantiomers were evaluated in Hep G2 cells. Results from the 3-(4,5-dimethylthioazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), resazurin reduction and lactate dehydrogenase (LDH) release assays showed a good agreement that the order of cytotoxicity was gamma-HBCD >= beta-HBCD > alpha-HBCD, and that significantly lower cell viability and higher LDH release were observed in all (+)-enantiomers ((+) alpha-, (+) beta- and (+) gamma-HBCD) than the corresponding (-)-forms ((-) alpha-, (-) beta- and (-) gamma-HBCD). Additionally, the formation of reactive oxygen species (ROS) induced by these HBCD enantiomers were detected. The positive correlation between the LDH release and ROS formation demonstrated that the toxic mechanism might be mediated by oxidative damage. These results suggest that environmental and human health risks of HBCDs must be evaluated at the level of individual enantiomers. (C) 2008 Published by Elsevier Ltd.

Apoptosis induction on human hepatoma cells Hep G2 of decabrominated diphenyl ether (PBDE-209)

Polybrominated diphenyl ethers (PBDEs) are an important class of halogenated organic brominated flame retardants. Because of their presence in abiotic and biotic environments widely and their structural similarity to polychlorinated biphenyls (PCBs), concern has been raised on their possible adverse health effects to humans. This study was designed to determine the anti-proliferative, apoptotic properties of decabrominated diphenyl ether (PBDE-209), using a human hepatoma Hep G2 line as a model system. Hep G2 cells were cultured in the presence of PBDE-209 at various concentrations (1.0-100.0 mu mol/L) for 72 h and the percentage of cell viability was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. The results showed that PBDE-209 inhibited the cells viability in time and concentration-dependent characteristics at concentrations (10.0-100.0 mu mol/L). We found that anti-proliferative effect of PBDE-209 was associated with apoptosis on Hep G2 cells by determinations of morphological changes, cell cycle and apoptosis. Mechanism study showed that PBDE-209 could increase the generation of intracellular reactive oxygen species (ROS) concentration-dependently. Antioxidant N-acetylcyteine partially inhibited the increase of ROS. The mechanism for its hepatoma-inhibitory effects was the induction of cellular apoptosis through ROS generation. In addition, activity of lactate dehydrogenase (LDH) release increased when the cells incubated with PBDE-209 at various concentrations and times. These results suggested that PBDE-209 had the toxicity activity of anti-proliferation and induction of apoptosis in tumor cells in vitro. (c) 2007 Elsevier Ireland Ltd. All rights reserved.

Predicted glycosyl transferase genes located outside the HEP island are required for formation of heterocyst envelope polysaccharide in Anabaena sp strain PCC 7120

During maturation, heterocysts form an envelope layer of polysaccharide, called heterocyst envelope polysaccharide (HEP), whose synthesis depends on a cluster of genes, the HEP island, and on an additional, distant gene, hepB, or a gene immediately downstream from hepB. We show that HEP formation depends upon the predicted glycosyl transferase genes all4160 at a third locus and alr3699, which is adjacent to hepB and is cotranscribed with it. Mutations in the histidine kinase genes hepN and hepK appear to silence the promoter of hepB and incompletely down-regulate all4160.

低剂量辐射诱导hep G2细胞克隆存活和细胞周期适应性反应

以低剂量γ射线(0.05Gy)预照射人肝癌细胞hepG2,8h后再用高剂量(3Gy)照射,测定了细胞的克隆存活率和细胞周期。结果表明,低剂量辐射预处理可诱导hepG2细胞产生克隆存活适应性反应,并且有助于细胞通过G2/M期阻滞;低剂量辐射诱导的克隆存活适应性反应与增强的通过细胞周期阻滞的能力之间有一定的相关性。

Determination of rare-earth impurities in ultra-highly pure Er2O3 and Ho2O3 by [HEH(HEP)] extraction chromatography-atomic emission spectrometry

This article applied [HEH(HEP)] (2-ethyhexyldrogen-2-ethylhexyl phosphonate)extraction chromatography to separate 14 rare earth impurities from ultra-highly pure Er2O3 and Ho2O3, and then the impurities were determined by atomic emission spectrometry. The average percentage recovery for each element is in the range of 70%similar to 140%. The relative standard deviations of the determination are +/-3.3%similar to 2.2%. This method can be applied to the determination of the trace amounts of rare eath impurities in Er2O3 and Ho2O3 with a purity of 99.999 9%-99.999 99%.

Hole-type gaseous detectors for medical imaging and HEP experiments

This work presents the results on the development of a high pressure Xe gaseous detector envisaging medical imaging. The detector uses two VUV photosensors operating face-to-face, based on the CsI-MHSP with position discrimination capability. The known effect of the charge gain decrease with the gas pressure is compensated with a light gain using the electroluminescence process. Studies of signal amplitude, energy and position resolution are presented. On the second part, gaseous detectors based on THGEM where studied in order to fulfill the requirements of present and future high energy physics experiments. The work is focused on the application of THGEM in RICH detectors elements: a triple THGEM detector using CsI photocathodes in Ne mixtures was suggested. The ion backflow to the photocathode remains a concern and a limitation; to minimize it, the application of a new hole-structure, THCOBRA, was suggested. Preliminary results foresee good perspectives for the successful application of THCOBRA in ion back-flow suppression. This work contains several methods and measurements of the CsI photocathodes behaviour in radiation gaseous detectors. A long discussion on its issues and possible solutions are presented.

The role of intimin in the adherence of enterohaemorrhagic Escherichia coli (EHEC) O157:H7 to HEp-2 tissue culture cells and to bovine gut explant tissues

Intimin, an outer membrane protein encoded by eaeA, is a key determinant for the formation of attaching and effacing (AE) lesions by enterohaemorrhagic Escherichia coli (EHEC). To investigate the role of intimin in adherence, the eaeA gene was insertionally inactivated in three EHEC O157:H7 strains of diverse origin. The absence or presence of intimin did not correlate with the extent of adhesion of mutant or wild-type O157:H7 in tissue culture and neonatal calf gut tissue explant adherence assays. Adherence of the eaeA mutants to HEp-2 cells was diffuse with no evidence of intimate attachment whereas wild-type bacteria formed microcolonies and AE lesions. Intimin-independent adherence to neonatal calf gut explants was demonstrated by eaeA mutants and wild-type strains which adhered in the greatest numbers to colon but least well to rumen tissue. These results confirm that intimin is necessary for intimate attachment and that additional adherence factors are involved in intimin-independent adherence.

Activation of the selenopritein SEPS1 gene expression by pro-inflammatory cytokines in Hep G2 cells

SEPS1 (also called selenoprotein S, SelS) plays an important role in the production of inflammatory cytokines and its expression is activated by endoplasmic reticulum (ER) stress. In this report, we have identified two binding sites for the nuclear factor kappa B in the human SEPS1 promoter. SEPS1 gene expression, protein levels and promoter activity were all increased 2–3-fold by TNF-α and IL-1β in HepG2 cells. We have also confirmed that the previously proposed ER stress response element GGATTTCTCCCCCGCCACG in the SEPS1 proximate promoter is fully functional and responsive to ER stress. However, concurrent treatment of HepG2 cells with IL-1β and ER stress produced no additive effect on SEPS1 gene expression. We conclude that SEPS1 is a new target gene of NF-κB. Together with our previous findings that SEPS1 may regulate cytokine production in macrophage cells, we propose a regulatory loop between cytokines and SEPS1 that plays a key role in control of the inflammatory response.

In vitro cytotoxic activity of Baccharis dracunculifolia and propolis against HEp-2 cells

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

In vitro Cytotoxic Effect of Brazilian Green Propolis on Human Laryngeal Epidermoid Carcinoma (HEp-2) Cells

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Evaluation of the antigenotoxicity of polysaccharides and beta-glucans from Agaricus blazei, a model study with the single cell gel electrophoresis/Hep G2 assay

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Shiitake (Lentinula edodes (Berkeley) Pegler) extracts as a modulator of micronuclei induced in HEp-2 cells

Shiitake (Lentinula edodes (Berkeley) Pegler) is one of the most consumed mushrooms, for both therapeutic purposes and as food, therefore, the study of its biological properties is of great interest for producers and consumers. Aqueous extracts of the shiitake mushroom (L. edodes (Berkeley) Pegler) were evaluated by the micronucleus test (MN) in HEp-2 cells in vitro, to analyze their possible mutagenic and antimutagenic activities. None of the three extract concentrations tested (0.5, 1.0 and 1.5 mg/mL) presented mutagenicity at any of the preparation temperatures (4 degrees C, 22 +/- 2 degrees C and 60 degrees C). In the antimutagenicity evaluation, all extract concentrations at all preparation temperatures presented a strong protective activity for the HEp-2 cells in response to the alkylating agent methyl methanesulfonate (MMS) in the different treatment protocols: pre-treatment, simultaneous treatment and post-treatment. The extracts prepared at 22 +/- 2 degrees C presented the lowest frequencies of MN in the evaluations of mutagenicity and antimutagenicity, indicating these as the best option for potential therapeutic use. (c) 2006 Elsevier Ltd. All rights reserved.