Determination of the Efficacy of Ozone Treatment Systems Using a Gap Junction Intercellular Communication Bioassay

Upham, Brad L.; Yao, Jehng Jyun.; Trosko, James E.; Masten, Susan J.

doi:10.1021/es00012a006

Cited by 34 publications

(31 citation statements)

References 36 publications

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“…Acetonitrile is noncytotoxic and noninhibitory on GJIC up to a concentration of 2% (v/v). 41 Acetonitrile was used as the vehicle control and the volumes added to the cell cultures were typically between 5 and 25 ll (0.25-1.25%). Lucifer yellow was loaded into the cells by making 2 or 3 scrape lines on the monolayer with a sharp scalpel.…”

Section: Gap Junctional Intercellular Communicationmentioning

confidence: 99%

Cigarette smoke components inhibited intercellular communication and differentiation in human pancreatic ductal epithelial cells

Tai

Upham

Olson

et al. 2007

Intl Journal of Cancer

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Smoking is a well-documented risk factor for the development of pancreatic adenocarcinoma. Although the most abundant polycyclic aromatic hydrocarbons (PAHs) in cigarette smoke are methylated anthracenes and phenanthrenes, the epigenetic toxicity of these compounds has not been extensively studied. We previously showed that methylanthracenes, which possess a bay-like structure, affect epigenetic events such as an induced release of arachidonic acid, inhibition of gap junctional intercellular communication (GJIC) and induction of mitogen-activated protein kinases in a pluripotent rat liver epithelial stem cell line. Anthracenes with no bay-like structures were inactive. These biological effects are all molecular events associated with the promotional phase of cancer. A human immortalized, nontumorigenic pancreatic ductal epithelial cell line, H6c7, was examined to study the epigenetic toxicity of PAHs related to pancreatic cancer by using scrape-loading dye transfer, immunostaining, RT-PCR and telomerase assay methods. H6c7 cells were GJIC-incompetent and exhibited high telomerase activity when grown in growth factor and hormonesupplemented medium. In the presence of the cAMP elevating drugs (forskolin and IBMX) the cells became GJIC competent and expressed connexins. Telomerase activity was also decreased by cAMP elevating drug treatment. After induction of cAMP, 1-methylanthracene with bay-like structures inhibited GJIC, whereas the 2-methylanthracene lacking a bay-like structure had no effect on GJIC. Telomerase activity remained high in 1-methylanthracene treatment but not with 2-methylanthracene. These results indicate that a prominent component of cigarette smoke, namely methylanthracenes with distinct structural configurations, could be a potential etiological agent contributing to the epigenetic events of pancreatic cancer. ' 2007 Wiley-Liss, Inc.Key words: connexin 36; telomerase; PAH; gap junction Pancreatic cancer is the fourth most common cause of cancerrelated death in the United States. 1,2 Incidence in the developed world parallels the United States, and in undeveloped nations the incidence is lower, which could be due to underreporting. 3 There are no early screening tests and no universally-effective treatment options, making this disease one of the most fatal cancers known, with a 5-year survival rate of less than 5%. 1,3-5 A recent article even suggested no patients are actually ''cured'' of this disease. 6 Although several risk factors, such as diet, have been associated with pancreatic cancer, cigarette smoking is the only unequivocal risk factor that has been identified. 1,7-9 The incidence of pancreatic cancer is 50-90% higher among blacks in the United States than whites. This disparity has been linked primarily to cigarette smoking-and to a lesser extent diabetes-in men, 10 again indicating the significance of tobacco smoke in the incidence of pancreatic cancer.Although cigarette smoke has been identified as one etiological cause of pancreatic cancer, the underlying molecular mechanism ca...

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Section: Gap Junctional Intercellular Communicationmentioning

confidence: 99%

Cigarette smoke components inhibited intercellular communication and differentiation in human pancreatic ductal epithelial cells

Tai

Upham

Olson

et al. 2007

Intl Journal of Cancer

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“…The scrape loading/dye transfer technique (described in [27]) was used to estimate the clones' coupling capacity. The dye was transferred to adjacent cells through gap junction for 5 min in the osteoblastic cells and 3 min in the WB-F344 cells when using the 457 Da-Lucifer yellow (LY), or 10 min for both cell lines when using LY conjugated to cadavarine biotin-X (649 Da).…”

Section: Measurement Of Gjicmentioning

confidence: 99%

Reduced gap junctional intercellular communication and altered biological effects in mouse osteoblast and rat liver oval cell lines transfected with dominant‐negative connexin 43

Upham

Suzuki

Chen

et al. 2003

Molecular Carcinogenesis

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Gap junctional intercellular communication (GJIC) maintains normal growth and differentiation of cells in a tissue. The intercellular molecules traversing gap junctions are largely unknown, but the molecular weight (MW) cutoff is normally 1200 Da. No differences in dye transfer were observed in normal or vector controls of WB-F344 rat liver epithelial or mouse osteoblastic MC3T3-E1 cells with either Lucifer Yellow (LY) with a MW of 457 Da (LY-457) or LY with a MW of 649 Da (LY-649). Transfection of a dominant negative-connexin 43 (Cx43) gene decreased GJIC (>50%) when LY-649 was used, however, normal GJIC was observed in both cell lines when LY-457 was used. Therefore, the MW cut off in these clones was considerably less than the wild type. The dominant negative clones of the MC3T3-E1 cells exhibited over 90% less alkaline phosphatase (ALPase) activity and calcium deposition after the induction of differentiation. Similarly, dominant negative Cx43 inhibited gene expression of ALPase and bone sialoprotein but not osteocalcin in MC3T3-E1. WB-F344 cells normally exhibit a biphasic response to 12-O-tetradecanoylphorbol-13-acetate (TPA) where inhibition of GJIC recovers after 2 h, but the dominant negative clones showed no recovery from inhibition of GJIC by TPA. Dominant negative Cx43 also inhibited the formation of network-like structures by WB-F344 cells on Matrigel. These results demonstrate that the dominant negative gene transfected into cell types containing the wild-type connexins result in diminished channel sizes, thus allowing the determination of whether distinct biological endpoints, i.e., differentiation, are dependent upon either small or high MW intercellular signals.

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“…To test every subtle change in existing environmental remediation strategies using in vivo assays would certainly be prohibitively costly and inhumane, and to limit in vitro assays to the measurement of genotoxicity would exclude the accurate assessment of risks to human health considering that many chemicals, including many carcinogens, are epigenetically toxic (16,17). Furthermore, monitoring the levels of known toxicants using analytical chemical techniques is not an acceptable alternative to eliminating in vivo experiments because simply removing the parent compound does not always ensure a safe environment: Toxic by-products could result from the chemical or biologic transformation of the parent compound (64). An example of such a situation was demonstrated by Upham et al (38,64), in which selected PAHs were oxidatively removed using ozone but the resulting mixture became more epigenetically toxic than the parent compound.…”

Section: Carcinogenesis As a Multistage Multimechanism Process Involvmentioning

confidence: 99%

“…Furthermore, monitoring the levels of known toxicants using analytical chemical techniques is not an acceptable alternative to eliminating in vivo experiments because simply removing the parent compound does not always ensure a safe environment: Toxic by-products could result from the chemical or biologic transformation of the parent compound (64). An example of such a situation was demonstrated by Upham et al (38,64), in which selected PAHs were oxidatively removed using ozone but the resulting mixture became more epigenetically toxic than the parent compound.…”

Section: Carcinogenesis As a Multistage Multimechanism Process Involvmentioning

confidence: 99%

Modulated Gap Junctional Intercellular Communication as a Biomarker of PAH Epigenetic Toxicity: Structure-Function Relationship

Upham¹,

Weis²,

Trosko³

1998

Environmental Health Perspectives

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Cancer is a multistage multimechanism process involving gene and/or chromosomal mutations (genotoxic events), altered gene expression at the transcriptional, translational, and posttranslational levels (epigenetic events), and altered cell survival (proliferation and apoptosis or necrosis), resulting in an imbalance of the organism's homeostasis. Maintenance of the organism's homeostasis depends on the intricate coordination of genetic and metabolic events between cells via extracellular and intercellular communication mechanisms. The release of a quiescent cell, whether normal or premalignant, from the suppressing effects of communicating neighbors requires the downregulation of intercellular communication via gap junctions, thereby allowing factors that control intracellular events to exceed a critical mass necessary for the cell to either proliferate or undergo apoptosis. Therefore, determining the role an environmental pollutant must play in the multistage carcinogenic process includes mechanisms of epigenetic toxicity such as the effects of a compound on gap junctional intercellular communication (GJIC). A classic example of a class of compounds in which determination of carcinogenicity focused on genotoxic events and ignored epigenetic events is polycyclic aromatic hydrocarbons (PAHs). The study of structure-activity relationships of PAHs has focused exclusively on the genotoxic and tumorinitiating properties of the compound. We report on the structure-activity relationships of two-to four-ringed PAHs on GJIC in a rat liver epithelial cell line. PAHs containing a bay or baylike region were more potent inhibitors of GJIC than the linear PAHs that do not contain these regions. These are some of the first studies to determine the epigenetic toxicity of PAHs at the epigenetic level.Environ Health Perspect 1 06(Suppl 4): 975-981 (1998). http.//ehpnetl.niehs.nih.gov/docs/ 1998/Suppl4/975-981upham/abstract.html

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Determination of the Efficacy of Ozone Treatment Systems Using a Gap Junction Intercellular Communication Bioassay

Cited by 34 publications

References 36 publications

Cigarette smoke components inhibited intercellular communication and differentiation in human pancreatic ductal epithelial cells

Cigarette smoke components inhibited intercellular communication and differentiation in human pancreatic ductal epithelial cells

Reduced gap junctional intercellular communication and altered biological effects in mouse osteoblast and rat liver oval cell lines transfected with dominant‐negative connexin 43

Modulated Gap Junctional Intercellular Communication as a Biomarker of PAH Epigenetic Toxicity: Structure-Function Relationship

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