I explored the effect of genetic (cloning) and non-genetic (arrangement of the method and quality of culture serum) factors upon variability of the results obtained by using the "kiss of death" method for evaluation of intercellular communication in Chinese hamster cells V79-4. Cloning had neither quantitative (intraspecies communication) nor qualitative (interspecies communication) effects. Of the non-genetic factors, the time interval of application of transferred agent (thioguanine) after plating the cells and the agent concentration were investigated. These factors did not influence significantly the results. Intercellular communication was more intensive in a less rich medium containing bovine serum than in a richer medium containing foetal calf serum. On the basis of the results obtained I conclude that the higher variability of intercellular communication values determined by the "kiss of death" method is not due to an error in the experimental method used but probably reflects the real changes in these values that occur with time in cell populations.

• MeSH
• buněčné klony MeSH
• buněčné linie MeSH
• F faktor MeSH
• krev MeSH
• mezibuněčná komunikace * MeSH
• mezibuněčné spoje fyziologie   MeSH
• thioguanin farmakologie   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• thioguanin MeSH

Humans are exposed to phthalates released from plastics, cosmetics, or food on a daily basis. Phthalates have low acute liver toxicity, but their chronic exposures could induce molecular and cellular effects linked to adverse health outcomes, such as liver tumor promotion or chronic liver diseases. The alternation of gap junctional intercellular communication (GJIC) and MAPK-Erk1/2 pathways in liver progenitor or oval cells can disrupt liver tissue homeostatic mechanisms and affect the development and severity of these adverse outcomes. Our study with 20 different phthalates revealed their structurally dependent effects on liver GJIC and MAPK-Erk1/2 signaling in rat liver WB-F344 cell line with characteristics of liver oval cells. The phthalates with a medium-length side chain (3-6 C) were the most potent dysregulators of GJIC and activators of MAPK-Erk1/2. The effects occurred rapidly, suggesting the activation of non-genomic (non-transcriptional) mechanisms directly by the parental compounds. Short-chain phthalates (1-2 C) did not dysregulate GJIC even after longer exposures and did not activate MAPK-Erk1/2. Longer chain (≥7 C) phthalates, such as DEHP or DINP, moderately activated MAPK-Erk1/2, but inhibited GJIC only after prolonged exposures (>12 h), suggesting that GJIC dysregulation occurs via genomic mechanisms, or (bio)transformation. Overall, medium-chain phthalates rapidly affected the key tissue homeostatic mechanisms in the liver oval cell population via non-genomic pathways, which might contribute to the development of chronic liver toxicity and diseases.

• Klíčová slova
• MAP-kinases Erk1/2 activation, gap junctional intercellular communication, gap junctions, hepatotoxicity, non-genomic mechanism, oval cells, phthalates, progenitor cells,
• MeSH
• buněčné linie MeSH
• játra cytologie   účinky léků   metabolismus   MeSH
• krysa rodu Rattus MeSH
• kyseliny ftalové aplikace a dávkování   chemie   toxicita   MeSH
• MAP kinasový signální systém účinky léků   MeSH
• mezerový spoj účinky léků   MeSH
• mezibuněčná komunikace účinky léků   MeSH
• viabilita buněk účinky léků   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• vztahy mezi strukturou a aktivitou MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• kyseliny ftalové MeSH

The frequencies of adverse outcomes associated with male reproductive health, including infertility and testicular cancer, are increasing. These adverse trends are partially attributed to increased exposure to environmental agents such as endocrine-disrupting chemicals (EDCs). This study addresses effects on EDCs on adjacent prepubertal Sertoli TM4 cells, specifically on 1) testicular gap junctional intercellular communication (GJIC), one of the hallmarks of non-genotoxic carcinogenicity, 2) GJIC building blocks connexins (Cx), and 3) mitogen-activated protein kinases MAPKs. We selected eight representatives of EDCs: organochlorine chemicals such as pesticides dichlorodiphenyltrichloroethane, lindane, methoxychlor, and vinclozolin, industrial chemicals bisphenol A and 2,2',4,4',5,5'-hexachlorobiphenyl, and components of personal care products, triclocarban and triclosan. EDCs rapidly dysregulated GJIC in Sertoli TM4 cells mainly via MAPK p38 and/or Erk1/2 pathways by the intermediate hyper- or de-phosphorylation of Cx43 (Ser368, Ser282) and translocation of Cx43 from the plasma membrane, suggesting disturbed intracellular trafficking of Cx43 protein. Surprisingly, EDCs did not rapidly activate MAPK Erk1/2 or p38; on the contrary, TCC and TCS decreased their activity (phosphorylation). Our results indicate that EDCs might disrupt testicular homeostasis and development via testicular GJIC, junctional and non-junctional functions of Cx43 and MAPK-signaling pathways in Sertoli cells.

• Klíčová slova
• Connexins, Endocrine-disrupting chemicals, Gap junctional intercellular communication, Reproductive toxicity, Sertoli cells, Testicular tumors,
• MeSH
• endokrinní disruptory * metabolismus   MeSH
• fosforylace MeSH
• konexin 43 genetika   metabolismus   MeSH
• lidé MeSH
• mezerový spoj metabolismus   MeSH
• mezibuněčná komunikace MeSH
• testikulární nádory * metabolismus   MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• endokrinní disruptory * MeSH
• konexin 43 MeSH

Inhalation exposures to polycyclic aromatic hydrocarbons (PAHs) have been associated with various adverse health effects, including chronic lung diseases and cancer. Using human bronchial epithelial cell line HBE1, we investigated the effects of structurally different PAHs on tissue homeostatic processes, namely gap junctional intercellular communication (GJIC) and MAPKs activity. Rapid (<1 h) and sustained (up to 24 h) inhibition of GJIC was induced by low/middle molecular weight (MW) PAHs, particularly by those with a bay- or bay-like region (1- and 9-methylanthracene, fluoranthene), but also by fluorene and pyrene. In contrast, linear low MW (anthracene, 2-methylanthracene) or higher MW (chrysene) PAHs did not affect GJIC. Fluoranthene, 1- and 9-methylanthracene induced strong and sustained activation of MAPK ERK1/2, whereas MAPK p38 was activated rather nonspecifically by all tested PAHs. Low/middle MW PAHs can disrupt tissue homeostasis in human airway epithelium via structure-dependent nongenotoxic mechanisms, which can contribute to their human health hazards.

• Klíčová slova
• Gap junctional intercellular communication, Human bronchial epithelial cell line, Methylated anthracenes, Mitogen-activated protein kinases, Nongenotoxic mechanisms, Polycyclic aromatic hydrocarbons,
• MeSH
• bronchy cytologie   MeSH
• buněčné linie MeSH
• epitelové buňky účinky léků   fyziologie   MeSH
• lidé MeSH
• mezerový spoj účinky léků   MeSH
• mezibuněčná komunikace účinky léků   MeSH
• mitogenem aktivované proteinkinasy metabolismus   MeSH
• polycyklické aromatické uhlovodíky toxicita   MeSH
• viabilita buněk účinky léků   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• mitogenem aktivované proteinkinasy MeSH
• polycyklické aromatické uhlovodíky MeSH

Ambient air pollution and smoking are well-documented risk factors for male infertility. Prevalent air pollutants and cigarette smoke components, polycyclic aromatic hydrocarbons (PAHs), are environmental and occupational toxicants that act as chemicals disrupting endocrine regulation and reproductive potential in males. Testicular gap junctional intercellular communication (GJIC) is critical for normal development and function of testicular tissue, thus we assessed GJIC as a process potentially targeted by PAHs in testes. Lower MW PAHs with a bay or bay-like region rapidly dysregulated GJIC in Leydig TM3 cells by relocalization of major testicular gap junctional protein connexin 43 (Cx43) from plasma membrane to cytoplasm. This was associated with colocalization between Cx43 and ubiquitin in intracellular compartments, but without any effect on Cx43 degradation rate or steady-state Cx43 mRNA levels. A longer exposure to active PAHs decreased steady-state levels of full-length Cx43 protein and its 2 N-truncated isoforms. Inhibition of GJIC by PAHs, similarly to a prototypic GJIC-inhibitor TPA, was mediated via the MAP kinase-Erk1/2 and PKC pathways. Polycyclic aromatic hydrocarbon-induced GJIC dysregulation in testes was cell-type-specific because neither PAH dysregulated GJIC in Sertoli TM4 cells, despite PAHs were rapidly taken up by both Leydig TM3 as well as Sertoli TM4 cells. Because TPA effectively dysregulated GJIC in both testicular cell types, a unique regulator of GJIC targeted by PAHs might exist in Leydig TM3 cells. Our results indicate that PAHs could be a potential etiological agent contributing to reproductive dysfunctions in males through an impairment of testicular GJIC and junctional and/or nonjunctional functions of Cx43.

• Klíčová slova
• Cx43 truncated isoforms, connexins, endocrine disruptors, gap junctional intercellular communication, polycyclic aromatic hydrocarbons, testicular cells,
• MeSH
• buněčné linie MeSH
• endokrinní disruptory chemie   toxicita   MeSH
• fosforylace MeSH
• konexin 43 genetika   metabolismus   MeSH
• Leydigovy buňky účinky léků   metabolismus   patologie   MeSH
• mezerový spoj účinky léků   metabolismus   patologie   MeSH
• mezibuněčná komunikace účinky léků   MeSH
• mitogenem aktivované proteinkinasy metabolismus   MeSH
• myši MeSH
• polycyklické aromatické uhlovodíky chemie   toxicita   MeSH
• Sertoliho buňky účinky léků   metabolismus   patologie   MeSH
• signální transdukce MeSH
• viabilita buněk účinky léků   MeSH
• zátoková oblast polycyklických aromatických uhlovodíků MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• endokrinní disruptory MeSH
• GJA1 protein, mouse MeSH Prohlížeč
• konexin 43 MeSH
• mitogenem aktivované proteinkinasy MeSH
• polycyklické aromatické uhlovodíky MeSH

The inhibition of intercellular gap junctional communication (IGJC) by alkyl ethers (ethylene glycol, monomethyl ether, polyethylene glycol 1,000 and polyethylene glycol 6000) was examined using V79 Chinese hamster cells in vitro. Ethylene glycol and monomethyl ether inhibited IGJC very strongly, whilst the other agents inhibited IGJC only insignificantly. When the cells were treated with the combination of two agents, ethylene glycol and monomethyl ether, a significant increase in the inhibition of IGJC occurred. This was probably the result of potentiation rather than an addition effect. The effect of ethylene glycol was antagonized by dibutyryl cyclic adenosine monophosphate (DbcAMP). This effect was most intensive when the cells were treated with both agents at the same time and, in other experimental combinations, the effect was lower but also significant. Caffeine did not influence IGJC either in combination with DbcAMP or by itself.

• MeSH
• buněčné linie MeSH
• časové faktory MeSH
• dibutyryl cyklický AMP antagonisté a inhibitory   farmakologie   toxicita   MeSH
• ethylenglykol MeSH
• ethylenglykoly antagonisté a inhibitory   toxicita   MeSH
• kofein farmakologie   MeSH
• křečci praví MeSH
• lékové interakce MeSH
• methylethery antagonisté a inhibitory   toxicita   MeSH
• mezibuněčná komunikace účinky léků   MeSH
• mezibuněčné spoje účinky léků   fyziologie   MeSH
• polyethylenglykoly toxicita   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• zvířata MeSH
• Check Tag
• křečci praví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• srovnávací studie MeSH
• Názvy látek
• dibutyryl cyklický AMP MeSH
• ethylenglykol MeSH
• ethylenglykoly MeSH
• kofein MeSH
• methylethery MeSH
• polyethylenglykoly MeSH

Polycyclic aromatic hydrocarbons (PAHs) are a broad class of ubiquitous environmental pollutants with known or suspected carcinogenic properties. Tumor promotion is a cell-proliferative step of cancer that requires the removal of cells from growth suppression via the inhibition of gap-junctional intercellular communication (GJIC). Inhibition of GJIC measured with an in vitro WB-F344 rat liver epithelial cell system was used to assess the relative potencies of 13 PAHs suggested by the U.S. Environmental Protection Agency (EPA) as the principal contaminants and 22 other PAHs, most of them identified in environmental samples. Maximal inhibition of GJIC was detected after 30 min of exposure, followed by a recovery in intercellular communication after an additional 30 min of exposure, suggesting a transient character of inhibition. Although microM concentrations of PAHs were required to reach the inhibition level equal to the model tumor promoter phorbol 12-myristate 13-acetate (IC50 = 8 nM), 12 of the PAHs under study were found to be strong inhibitors of GJIC (strongest effects were observed with fluoranthene, picene, 5-methylchrysene and nine additional PAHs). The other nine PAHs, including benzo[a]pyrene, inhibited GJIC only up to 50-75% of the control level. Interestingly, several high molecular weight PAHs with known strong carcinogenic properties possessed only weak (dibenzopyrenes) or no inhibition potency (dibenzofluoranthenes, naphtho[2,3-a]pyrene and benzo[a]perylene). Based on the IC50 values related to the reference PAH benzo[a]pyrene, we suggested arbitrary values of inhibition equivalency factors (GJIC-IEFs) ranging from 0 (noninhibiting PAHs) to 10.0 (strongest inhibitors), suitable for the purposes of environmental risk assessment.

• MeSH
• Agentura Spojených států pro ochranu životního prostředí normy   MeSH
• buněčné linie MeSH
• epitel účinky léků   MeSH
• játra cytologie   účinky léků   MeSH
• karcinogeny toxicita   MeSH
• krysa rodu Rattus MeSH
• látky znečišťující životní prostředí toxicita   MeSH
• mezerový spoj účinky léků   MeSH
• mezibuněčná komunikace účinky léků   MeSH
• molekulární struktura MeSH
• polycyklické aromatické uhlovodíky chemie   toxicita   MeSH
• tetradekanoylforbolacetát toxicita   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Spojené státy americké MeSH
• Názvy látek
• karcinogeny MeSH
• látky znečišťující životní prostředí MeSH
• polycyklické aromatické uhlovodíky MeSH
• tetradekanoylforbolacetát MeSH

Despite intensive research into toxic bloom-forming cyanobacteria, the majority of their metabolites remain unknown. The present study explored in detail a novel bioactivity identified in cyanobacteria, i.e. inhibition of gap junctional intercellular communication (GJIC), a marker of tumor promotion. The extracellular mixture (exudate) of the cyanobacterial strain Cylindrospermopsis raciborskii (SAG 1.97) was fractionated by semi-preparative reversed phase HPLC, and the fractions assessed for their potencies to inhibit GJIC. Two non-polar fractions that significantly inhibited GJIC were further fractionated, tested and analyzed using multiple mass spectrometric methods. Investigations led to the identification of a putative chemical compound (molecular formula C18H34O3, m/z 299.2581 for the [M+H](+) ion) responsible for observed bioactivities. Specific inhibitors of signaling pathways were used to screen for biochemical mechanisms beyond GJIC inhibition, and the results indicate the involvement of ERK1/2 kinases via a mechanism related to the action of epidermal growth factor EGF but clearly distinct from other anthropogenic tumor promoters like polychlorinated biphenyls or polycyclic aromatic hydrocarbons. The chemical and in vitro toxicological characterizations of the newly described metabolite provide important insights into the still poorly understood health impacts of complex toxic cyanobacterial blooms and indicate that currently applied monitoring practices may underestimate actual risks.

• Klíčová slova
• Cyanobacteria, Exudate, Fractionation, Tumor promotion,
• MeSH
• bakteriální toxiny farmakologie   MeSH
• buněčné linie MeSH
• Cylindrospermopsis metabolismus   MeSH
• krysa rodu Rattus MeSH
• mezerový spoj účinky léků   MeSH
• mezibuněčná komunikace účinky léků   MeSH
• viabilita buněk účinky léků   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• bakteriální toxiny MeSH

Ethylene glycol (EG) has been previously shown to inhibit gap junctional intercellular communication. In this paper we examine conditions under which the effect of EG on gap junctional communication is assessed by metabolic cooperation assay. The later, after the start of metabolic cooperation assay, EG was added, the lower its inhibitory effect was. If treatment with EG began 12 h or even later after plating of cells, no significant effect on gap junctional communication was observed. Short EG treatments (2-8 h) induced a reversible inhibition of cell-to-cell communication, provided that the cells could communicate freely after the drug was removed. However, if further cell-to-cell communication was excluded, the effect of short exposures was irreversible. Using Scrape loading method we observed that after a 60 min exposure to EG the standard gap junctional intercellular communication was completely restored in a few hours.

• MeSH
• biotest metody   MeSH
• časové faktory MeSH
• CHO buňky účinky léků   metabolismus   MeSH
• ethylenglykol farmakologie   MeSH
• kokultivační techniky MeSH
• křečci praví MeSH
• mezerový spoj účinky léků   fyziologie   MeSH
• mezibuněčná komunikace * MeSH
• thioguanin metabolismus   farmakologie   MeSH
• zvířata MeSH
• Check Tag
• křečci praví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• ethylenglykol MeSH
• thioguanin MeSH

The behavior of cells during the assessment of gap junctional intercellular communication (GJIC) by metabolic cooperation assay was studied, either under standard conditions, or with the simultaneous addition of an inhibitor of GJIC-ethylene glycol, and eventually plus dibutyryl cyclic adenosine monophosphate. Generally, the behavior of cells in the course of the assay did not depend significantly on the combination of drugs used. Cell locomotion during the assay was minimal, and consequently the results depended, almost solely on the probability of mutual contact between donor and recipient cells during the seeding. The first significant decrease in recovery of recipient cells, that is a detectable GJIC, was observed 30 min after the assay had started. Final results of the assay depended on the first 12 h of the experiment, in which the maximum decrease in the recovery of recipient cells, that is the maximum GJIC, was reached. The longer duration of the assay did not affect the results significantly.

• MeSH
• buněčné linie MeSH
• Cricetulus MeSH
• dibutyryl cyklický AMP farmakologie   MeSH
• ethylenglykol MeSH
• ethylenglykoly farmakologie   MeSH
• fibroblasty cytologie   účinky léků   metabolismus   MeSH
• křečci praví MeSH
• mezerový spoj účinky léků   fyziologie   MeSH
• mezibuněčná komunikace účinky léků   fyziologie   MeSH
• mitóza MeSH
• protinádorové antimetabolity farmakologie   MeSH
• thioguanin farmakologie   MeSH
• zvířata MeSH
• Check Tag
• křečci praví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• dibutyryl cyklický AMP MeSH
• ethylenglykol MeSH
• ethylenglykoly MeSH
• protinádorové antimetabolity MeSH
• thioguanin MeSH