AIMS: Hofbauer cells (HBCs) are placental macrophages playing various roles during normal and complicated pregnancies, and of the latter, chorioamnionitis is the most frequent. METHODS: In placenta with chorioamnionitis, we examined immunohistochemical expression profiles of IL-1β, IL-10, and their potential regulators, CYP2C8 and soluble epoxide hydrolase (sEH), in Hofbauer cells and compared the results with our previously published data for normal placenta. RESULTS: We found that the expression profiles of the studied proteins in Hofbauer cells in chorioamnionitis differs from normal placenta. In chorioamnionitis, HBCs showed a moderate expression of IL-1β together with a weak expression of IL-10 and CYP2C8. Contrary to normal placenta, HBCs in chorioamnionitis express sEH. We demonstrated a moderate positive correlation between the expression of CYP2C8 and sEH in chorioamnionitis (Spearman r = 0.5654), suggesting enhanced degradation of anti-inflammatory epoxyeicosatrienoic acids. Moreover, the relations of IL-1β and IL-10 to CYP2C8, previously described in normal placenta, disappeared. Furthermore, a weak expression of anti-inflammatory IL-10 in chorioamnionitis was accompanied by change in circularity of HBCs (Spearman r = 0.8193). CONCLUSION: Taken together, these findings suggest a possible alteration of the anti-inflammatory role of HBCs and its regulation in chorioamnionitis.

• MeSH
• antiflogistika MeSH
• chorioamnionitida * MeSH
• cytochrom P450 CYP2C8 * metabolismus   MeSH
• epoxid hydrolasy * metabolismus   MeSH
• imunomodulace MeSH
• interleukin-10 MeSH
• lidé MeSH
• placenta metabolismus   MeSH
• těhotenství MeSH
• Check Tag
• lidé MeSH
• těhotenství MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

Metabolic deactivation by cytochrome P450 (CYP) is considered a potential mechanism of anticancer drug resistance. However, this hypothesis is predominantly based on indirect pieces of evidence and/or is influenced by interfering factors such as the use of multienzymatic models. Thus, an experimental approach for its verification is needed. In the present work, we employed HepG2 cells transduced with CYP enzymes involved in docetaxel, paclitaxel and vincristine metabolism to provide mechanistic evidence on their possible roles in resistance to these chemotherapeutic agents. Using MTT proliferation tests, we showed that overexpression of CYP3A4 resulted in decreased antiproliferative activity of 1 μM docetaxel (by 11.2, 23.2 and 22.9% at 24, 48 and 72 h intervals, respectively), while the sensitivity of CYP3A4-transduced cells was restored by co-administration of ketoconazole. Paclitaxel exhibited differential efficacy in CYP2C8- and empty vector-transduced cells (significant differences between 10.9 and 24.4% for 0.01, 0.1 and 1 μM concentrations), but neither montelukast nor clotrimazole was capable of affecting this asymmetry. Finally, the pharmacological activity of vincristine was not influenced by CYP3A4 or CYP3A5 overexpression. In the follow-up caspase activation assays, docetaxel was confirmed to be a victim of CYP3A4-mediated resistance, which is, at least partly, brought by impaired activation of caspases 3/7, 8 and 9. In summary, our data demonstrate that CYP3A4-mediated metabolic deactivation of docetaxel might represent a significant mechanism of pharmacokinetic resistance to this drug. In contrast, the possible role of CYPs in resistance to paclitaxel and vincristine has been disconfirmed. Importantly, the expression of CYP3A4 in HepG2_CYP3A4 cells is comparable to that in primary hepatocytes and HepaRG cells, which suggests that our results might be relevant for in vivo conditions, e.g., for hepatocellular carcinoma. Thus, our data may serve as a valuable in vitro background for future in vivo studies exploring the area of intratumoural metabolism-based drug resistance.

• MeSH
• antitumorózní látky farmakologie   MeSH
• buňky Hep G2 MeSH
• chemorezistence fyziologie   MeSH
• cytochrom P-450 CYP3A metabolismus   MeSH
• cytochrom P450 CYP2C8 metabolismus   MeSH
• cytostatické látky farmakologie   MeSH
• hepatocyty účinky léků   metabolismus   MeSH
• lidé MeSH
• metabolická clearance účinky léků   MeSH
• metabolická inaktivace účinky léků   MeSH
• nádorové buněčné linie MeSH
• proliferace buněk účinky léků   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

OBJECTIVES: Cytochromes P450 (CYPs) are heme enzymes oxygenating a broad range of substrates. Their activity is dependent on the presence of a suitable electron donor (eukaryotic NADPH:CYP oxidoreductase or cytochrome b5). The Escherichia naturally contain no CYPs and no NADPH:CYP oxidoreductase, however it was reported that some CYPs heterologously expressed in E. coli may exist in the ferrous form. A small bacterial flavoprotein, flavodoxin is considered to be responsible for reduction some of these CYPs. METHODS: The reduction state of several human CYPs expressed in the intact living E. coli cells was examined. In addition, molecular dynamics and steered molecular dynamics simulations were performed to predict and compare affinity of flavodoxin toward selected CYPs. RESULTS: We determined the reduction state of five human CYPs heterologously expressed in E. coli. The computationally predicted stabilities of CYP-flavodoxin complexes correlate with the percentage of reduced CYPs in bacterial cells. The mean electron transfer distance within optimized complexes was also related to the percentage of reduced CYPs. CONCLUSION: Depending on the resting state, the CYPs heterologously expressed in E. coli could be divided into two groups; CYP2C8, 2C9, 3A4 are in E. coli present mainly in the oxidized form; while CYP1A1, 1A2, 2A6, 2A13, 2B6, 2D6 are found predominantly in the reduced form. We found a significant correlation between the stability of CYP-flavodoxin complexes and the percentage of reduced CYPs in bacteria. Hence, the naturally expressed flavodoxin is probably responsible for reduction of a larger group of human CYPs in bacterial cells.

• MeSH
• aromatické hydroxylasy metabolismus   MeSH
• cytochrom P-450 CYP1A1 metabolismus   MeSH
• cytochrom P-450 CYP1A2 metabolismus   MeSH
• cytochrom P-450 CYP2D6 metabolismus   MeSH
• cytochrom P-450 CYP3A metabolismus   MeSH
• cytochrom P450 CYP2A6 metabolismus   MeSH
• cytochrom P450 CYP2B6 metabolismus   MeSH
• cytochrom P450 CYP2C8 metabolismus   MeSH
• cytochrom P450 CYP2C9 metabolismus   MeSH
• Escherichia coli MeSH
• flavodoxin metabolismus   MeSH
• geneticky modifikované organismy MeSH
• lidé MeSH
• NADPH-cytochrom c-reduktasa metabolismus   MeSH
• oxidace-redukce MeSH
• systém (enzymů) cytochromů P-450 metabolismus   MeSH
• železité sloučeniny metabolismus   MeSH
• železnaté sloučeniny metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH