CYP epoxygenases Dotaz Zobrazit nápovědu
INTRODUCTION: The success of pregnancy depends on the regulation of immunological processes in the placenta. Important mediators of an immune response include pro- and anti-inflammatory interleukins which may be regulated by CYP epoxygenases and their metabolites. The relation between interleukins and CYP epoxygenases expression in human placenta has not yet been studied vastly. MATERIAL AND METHODS: We investigated the expression patterns of IL-1β and IL-10 in embryonic (n=8), early foetal (n=16) and term (n=7) human placenta tissue by an immunohistochemical method and evaluated the results by Kruskal-Wallis test. The obtained data was correlated using Spearman's correlation coefficient to our previously published data of CYP epoxygenases expression in the same samples. To confirm that Hofbauer cells express IL-10 and IL-1β as well as CYP2C8 and IL-10 together, and thus there is a relation between proteins of interests, we used multiplex immunofluorescent staining. RESULTS: The expression of IL-1β decreased with gestational age in cytotrophoblast, syncytiotrophoblast, as well as in Hofbauer cells whilst IL-10 decreased in cytotrophoblast, remained at the same levels in syncytiotrophoblast and increased in Hofbauer cells. In trophoblast cells, we found a statistically significant positive correlation between the expression of CYP2J2 and CYP2C9 with IL-1β, whereas there was no relation between IL-10 and any of the tested CYP epoxygenases. In Hofbauer cells, we found a significant positive correlation between CYP2C8 and IL-10 and a significant negative correlation between CYP2C8 and IL-1β. CONCLUSION: Our results showed that the exact role and relation of interleukins and CYP epoxygenases and their metabolites is dependent on their respective cellular context. Because of IL-10, IL-1β, as well as HBCs play a role in various pathological conditions, further investigation of the exact role of CYP epoxygenase, interleukins and their relations is needed.
- MeSH
- cytochrom P450 CYP2C8 MeSH
- interleukin-1 MeSH
- interleukin-10 * MeSH
- lidé MeSH
- placenta * MeSH
- těhotenství MeSH
- trofoblasty MeSH
- Check Tag
- lidé MeSH
- těhotenství MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- Publikační typ
- abstrakt z konference MeSH
There is growing evidence that some members of cytochrome P450 enzymes contribute to regulation of normal prenatal development. CYP epoxygenases (CYP2C and CYP2J subfamilies) convert arachidonic acid into four regioisomeric epoxyeicosatrienoic acids (EETs), biologically active molecules involved in mitogenesis and cell signaling. Almost nothing is known about localization of their expression in tissues during human prenatal development. The spatio-temporal expression pattern of CYP2C8, CYP2C9, CYP2C19 and CYP2J2 in human embryonic/fetal intestines, liver, and kidney was investigated by immunohistochemical method. CYP epoxygenases are expressed already in early stages of development in these embryonic/fetal tissues (as early as 7th week of IUD in the intestines, 5th week of IUD in the liver, and 6th week of IUD in the kidney). In kidney, CYP epoxygenases are expressed in the metanephrogenic blastema (but not in the uninduced mesenchyme) and in the tubular system. In the intestines, diverse CYP epoxygenases distribution along crypt-villus axis could suggest role in cell differentiation. Moreover, we detected higher CYP2J2 level in these organs than in adult tissue samples.
- MeSH
- časové faktory MeSH
- cytochrom P450 CYP2C8 genetika MeSH
- embryo savčí enzymologie MeSH
- imunohistochemie MeSH
- játra embryologie enzymologie MeSH
- ledviny embryologie enzymologie MeSH
- lidé embryologie MeSH
- střeva embryologie enzymologie MeSH
- systém (enzymů) cytochromů P-450 genetika metabolismus MeSH
- vývojová regulace genové exprese * MeSH
- Check Tag
- lidé embryologie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
CYP2C and CYP2 J enzymes, commonly named as cytochrome P450 (CYP) epoxygenases, convert arachidonic acid to four regioisomeric epoxyeicosatrienoic acids (EETs), biologically active eicosanoids with many functions in organism. EETs are rapidly hydrolysed to less active dihydroxyeicosatrienoic acids (DHETs) by soluble epoxide hydrolase (sEH). We investigated spatio-temporal expression pattern of CYP2C8, CYP2C9, CYP2 J2 and sEH in normal human placenta by immunohistochemical method. In the villous trophoblast, CYP2C8 was the most abundant protein. Its expression is higher than the CYP2C9 and CYP2 J2 in the cytotrophoblast in the embryonic stage of development and remains higher in syncytiotrophoblast of term placenta. Unlike to CYP2C8, CYP2C9 and CYP2 J2 expression decrease in term placenta. sEH expression increases with gestation age and is strictly limited to cytotrophoblast in embryonic and foetal stages of the development. Moreover, CYP2C8 shows more intensive staining than the other protein monitored in Hofbauer cells in villous stroma. Specific information regarding the exact role of EETs and DHETs functions in a normal placenta is still unknown. Based on CYP epoxygenases and sEH localization and well known information about the functions of placental structures during development, we suggest that these enzymes could play different roles in various cell populations in the placenta. As the placenta is absolutely crucial for prenatal development, arachidonic acid is essential part of human nutrient and CYP epoxygenases expression can be affected by xenobiotics, further investigation of the exact role of CYP epoxygenases, sEH, and their metabolites in normal pregnancy and under pathological conditions is needed.
- MeSH
- cytochrom P450 CYP2C8 biosyntéza MeSH
- cytochrom P450 CYP2C9 biosyntéza MeSH
- lidé MeSH
- placenta cytologie enzymologie MeSH
- regulace genové exprese enzymů fyziologie MeSH
- systém (enzymů) cytochromů P-450 biosyntéza MeSH
- těhotenské proteiny biosyntéza MeSH
- těhotenství metabolismus MeSH
- Check Tag
- lidé MeSH
- těhotenství metabolismus MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
CYP epoxygenases metabolize arachidonic acid into four regioisomers of epoxyeicosatrienoic acids (EETs) which are hydrolysed into their corresponding diols by soluble epoxide hydrolase (sEH). EETs are very biologically active molecules. They promote proliferation and inhibit apoptosis as well as numerous other functions within organisms. Peroxisome proliferator-activated receptor α (PPARα) play role in regulation of CYP epoxygenases and sEH. PPARα is the ligand-dependent transcriptional factor which is activated by various compounds, including fibrates. The latter are widely used in clinical practice. This study investigates the changes in expression of CYP2C8, CYP2J2, and sEH in HEK293, HepG2, and HT-29 cell lines after fibrate treatment using two different incubation times. The results demonstrate that the effect of fibrates on arachidonic acid-metabolizing enzymes expression is concentration-dependent. Although CYP2C8 expression is downregulated by the fibrates treatment, the results reveal that changes in CYP2J2/sEH ratio are closely associated with cell proliferation and could explain the differing proliferation response of cells to different concentrations of fibrates.
- MeSH
- buňky Hep G2 MeSH
- buňky HT-29 MeSH
- cytochrom P450 CYP2C8 biosyntéza genetika MeSH
- deriváty kyseliny fibrové farmakologie MeSH
- epoxid hydrolasy biosyntéza MeSH
- HEK293 buňky MeSH
- hypoglykemika farmakologie MeSH
- kyseliny arachidonové metabolismus MeSH
- lidé MeSH
- PPAR alfa účinky léků metabolismus MeSH
- regulace genové exprese enzymů účinky léků MeSH
- systém (enzymů) cytochromů P-450 biosyntéza genetika MeSH
- vztah mezi dávkou a účinkem léčiva MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Fibrates belong to a group of ligands of peroxisome proliferator-activated receptor alpha (PPARα), which play a role in the regulation of CYP epoxygenases and soluble epoxide hydrolase (sEH), key enzymes in the metabolism of biologically highly active epoxyeicosatrienoic acids (EETs). We demonstrated that low doses of fibrates stimulate proliferation of the MCF7 cell line, while high doses suppress it. The increase in cell proliferation was accompanied by an increase in CYP epoxygenases and decrease in sEH levels. The overall level of PPARα remained same after low-dose fibrate stimulation; however, there was a significant shift of the receptor to the cell nucleus. PPARα expression was further demonstrated by immunohistochemistry in both carcinoma and healthy breast tissue samples both in the cytoplasm and in the nuclei. We have also observed higher nuclear PPARα positivity in tumor tissues. Although our results obtained for MCF7 cells suggest the potential role of PPARα in cell proliferation, we did not find an association between nuclear localization of PPARα and the expression of proliferation marker Ki-67 in tumor tissues. The exact role of PPARα in carcinogenesis still remains unclear.
- MeSH
- antigen Ki-67 metabolismus MeSH
- deriváty kyseliny fibrové farmakologie MeSH
- epoxid hydrolasy metabolismus MeSH
- geny erbB-2 * MeSH
- inhibiční koncentrace 50 MeSH
- lidé MeSH
- MFC-7 buňky MeSH
- nádory prsu metabolismus patologie MeSH
- PPAR alfa metabolismus MeSH
- proliferace buněk účinky léků MeSH
- pyrimidiny farmakologie MeSH
- receptory pro estrogeny metabolismus MeSH
- receptory progesteronu metabolismus MeSH
- rozpustnost MeSH
- subcelulární frakce metabolismus MeSH
- systém (enzymů) cytochromů P-450 metabolismus MeSH
- transport proteinů účinky léků MeSH
- viabilita buněk účinky léků MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
There is growing evidence that soluble epoxide hydrolase (sEH) may play a role in cell differentiation. sEH metabolizes biologically highly active and generally cytoprotective epoxyeicosatrienoic acids (EETs), generated from arachidonic acid metabolism by CYP epoxygenases (CYP2C and CYP2J subfamilies), to less active corresponding diols. We investigated the effect of sEH inhibitor (TPPU) on the expression of villin, CYP2C8, CYP2C9, CYP2J2, and sEH in undifferentiated and in vitro differentiated HT-29 and Caco2 cell lines. The administration of 10 μM TPPU on differentiated HT-29 and Caco2 cells resulted in a significant decrease in expression of villin, a marker for intestinal cell differentiation. It was accompanied by a disruption of the brush border when microvilli appeared sparse and short in atomic force microscope scans of HT-29 cells. Although inhibition of sEH in differentiated HT-29 and Caco2 cells led to an increase in sEH expression in both cell lines, this treatment had an opposite effect on CYP2J2 expression in HT-29 and Caco2 cells. In addition, tissue samples of colorectal carcinoma and adjacent normal tissues from 45 patients were immunostained for sEH and villin. We detected a significant decrease in the expression of both proteins in colorectal carcinoma in comparison to adjacent normal tissue, and the decrease in both sEH and villin expression revealed a moderate positive association. Taken together, our results showed that sEH is an important player in intestinal cell differentiation.
Embryonic and tumour cells are able to protect themselves against various harmful compounds. In human pathology, this phenomenon exists in the form of multidrug resistance (MDR) that significantly deteriorates success of anticancer treatment. Cytochromes P450 (CYPs) play one of the key roles in the xenobiotic metabolism. CYP expression could contribute to resistance of cancer cells to chemotherapy. CYP epoxygenases (CYP2C and CYP2J) metabolize about 20% of clinically important drugs. Besides of drug metabolism, CYP epoxygenases and their metabolites play important role in embryos, normal body function, and tumors. They participate in angiogenesis, mitogenesis, and cell signaling. It was found that CYP epoxygenases are affected by peroxisome proliferator-activated receptor α (PPARα). Based on the results of current studies, we assume that PPARs ligands may regulate CYP2C and CYP2J and in some extent they may contribute to overcoming of MDR in patients with different types of tumours.
- MeSH
- biologické modely MeSH
- chemorezistence * MeSH
- lidé MeSH
- mnohočetná léková rezistence * MeSH
- receptory aktivované proliferátory peroxizomů metabolismus MeSH
- systém (enzymů) cytochromů P-450 metabolismus MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
INTRODUCTION: Cytochrome (CYP) epoxygenases (CYP2C and CYP2J) and soluble epoxide hydrolase (sEH) participate in the metabolism of arachidonic acid and may also have a potential role in enterocyte differentiation. The first critical step in the study of intestinal cell differentiation is the determination of a suitable in vitro model, which must be as similar as possible to the conditions of a living organism. It is known that HT-29 and Caco2 cell lines derived from human colorectal carcinomas can differentiate into enterocyte-like cells in appropriate culture conditions. MATERIAL AND METHODS: We tested 4 different approaches of enterocyte-like differentiation and determined the most appropriate culture conditions for each model. Subsequently, the changes in the expression of CYP epoxygenases and sEH in undifferentiated and differentiated cells were measured by In-Cell ELISA. These results were compared with immunohistochemical profiles of expression of CYP epoxygenases and sEH in samples of human embryonic and fetal intestines as well as adult duodenum and colon. RESULTS: Our results show that sodium butyrate (NaBt)-differentiated HT-29 cells and spontaneously differentiated Caco2 cells resemble CYP epoxygenases and sEH profiles, corresponding with different types of intestines. CONCLUSION: Our study revealed that the most suitable models for the study of the role of CYP epoxygenases and sEH expression in differentiation of intestinal epithelium are NaBt-differentiated HT-29 cells and spontaneously differentiated Caco2 cells.
- MeSH
- buněčná diferenciace * MeSH
- buňky HT-29 MeSH
- Caco-2 buňky MeSH
- enterocyty enzymologie MeSH
- epoxid hydrolasy metabolismus MeSH
- kyselina arachidonová metabolismus MeSH
- lidé MeSH
- střeva cytologie embryologie MeSH
- střevní sliznice * embryologie metabolismus MeSH
- systém (enzymů) cytochromů P-450 metabolismus MeSH
- techniky in vitro MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
BACKGROUND: Fibrates are widely used hypolipidemic drugs, which serve as ligand of peroxisome proliferator-activated receptor α (PPARα). Recently, they have also been considered as potential anticancer agents. We studied effect of fibrates treatment on cell proliferation, expression of CYP2J2 and concomitant changes in expression of cell cycle regulatory proteins in three different human cell lines: HEK293, HepG2, and HT-29. METHODS: We used WST-1 viability test, western blot and immunocytochemistry for detection of proteins of interests and analysis of cell cycle. RESULTS: Our results showed that at lower concentrations of all tested fibrates, viability of all tested cell lines is increased, whereas at higher concentrations, repression is apparent. Unfortunately, the viability of tested cells is predominantly increased in a range of concentration which is reached in patient plasma. This phenomenon is accompanyed by elevation of CYP2J2, increased number of cyclin E-positive cells and decreased number of Cdc25A-positive cells in all tested cell lines, and elevated cyclin A expression in HepG2 and HT-29. These changes are concentration-dependent. We suppose that increased level of CYP2J2 could explain enhanced cell proliferation in lower concentration of fibrates. CONCLUSION: Based on our results, we suggested there is no anti-cancer effect of fibrates in tested carcinoma cell lines.
