Inhibition of soluble epoxide hydrolase (sEH) appears to be promising for the treatment of many diseases. Studies have focused on the beneficial effects of epoxyeicosatrienoic acids (EETs), which are sEH substrates. However, our recent studies have shown that the sEH activity is crucial for the proper intestinal cell differentiation. In this recent study, we investigated the impact of TPPU, an inhibitor of sEH, on the colon cancer cell lines Caco2 and HT-29. We analysed the changes in the expression of the cytoskeletal protein ezrin and the phosphorylated protein kinase p38 (p-p38). Our results showed a decrease in ezrin expression in differentiated cells and an increase in p-p38 expression after TPPU treatment. Immunocytochemical staining revealed a higher staining intensity of p-p38 in the nuclei of HT-29 cells following TPPU treatment. Immunohistochemical staining was performed on human samples of normal colon tissue, grade 2 tumours, and embryonal/foetal tissues. The staining intensity of ezrin in tumours was reduced in the surface area compared to the crypts. Additionally, we observed the translocation of p-p38 expression from the cytoplasm to the nucleus during differentiation. The tumour samples exhibited higher levels of p-p38 in the cytoplasm, similar to normal undifferentiated tissue. To observe the disruption of the cytoskeleton after TPPU treatment, confocal microscopy was used. It was found that β-actin associated with ezrin forms clusters under the plasma membranes. All of these results are significant because sEH inhibitors are being tested in clinical trials, but they could cause an unexpected adverse effects.
- MeSH
- buněčná diferenciace * účinky léků MeSH
- buňky HT-29 MeSH
- Caco-2 buňky MeSH
- cytoskeletální proteiny * metabolismus MeSH
- epoxid hydrolasy * antagonisté a inhibitory metabolismus MeSH
- fenylmočovinové sloučeniny farmakologie MeSH
- inhibitory enzymů farmakologie MeSH
- lidé MeSH
- mitogenem aktivované proteinkinasy p38 metabolismus MeSH
- nádory tračníku * farmakoterapie patologie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
INTRODUCTION: Inflammation of the placenta is harmful to both the fetus and the mother. Inflammation is strongly associated with diabetes, a common complication of pregnancy. Hofbauer cells (HBCs), unique immune system cells of fetal origin in the placenta, play complex roles, including growth of placental villi and their branching, stromal remodelling, and angiogenesis. METHODS: Our study investigated the expression of IL-1β, IL-10, CYP2C8, CYP2C9, CYP2J2 and sEH in HBCs from patients with type 1 diabetes mellitus (T1DM) and gestational diabetes mellitus (GDM) compared to healthy controls using immunohistochemistry. We also assessed the structure of the villus stroma using Masson ́s trichrome. RESULTS: In T1DM, HBCs showed inflammatory activation characterised by increased IL-1β and decreased CYP epoxygenase expression compared to normal placentas. Conversely, significant inflammation in HBCs appeared less likely in GDM, as levels of IL-1β and CYP epoxygenases remained stable compared to normal placentas. However, GDM showed a significant increase in sEH expression. Both types of diabetes showed delayed placental villous maturation and hypovascularisation, with GDM showing a more pronounced effect. CONCLUSION: The expression profiles of IL-1β, CYP epoxygenases and sEH significantlly differ between controls and diabetic placentas and between T1DM and GDM. These facts suggest an association of the CYP epoxygenase-EETs-sEH axis with IL-1β expression as well as villous stromal hypovascularisation. Given the stable high expression of IL-10 in both controls and both types of diabetes, it appears that immune tolerance is maintained in HBCs.
- MeSH
- diabetes mellitus 1. typu * metabolismus MeSH
- gestační diabetes * MeSH
- interleukin-10 metabolismus MeSH
- lidé MeSH
- placenta metabolismus MeSH
- těhotenství MeSH
- zánět metabolismus MeSH
- Check Tag
- lidé MeSH
- těhotenství MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Intestinal epithelial differentiation is a highly organised process. It is influenced by a variety of signalling pathways and enzymes, such as the PI3K pathway and soluble epoxide hydrolase (sEH) from arachidonic acid metabolism. We investigated the changes in the expression of enzymes and lipid messenger from the PI3K pathway, including PTEN, during intestinal cell differentiation in vitro using HT-29 and Caco2 cells and compared them with immunohistochemical patterns of these proteins in human colon. To investigate the possible crosstalk between the PI3K pathway and sEH, we treated HT-29 and Caco2 cells with the sEH inhibitor TPPU. Administration of TPPU to differentiated cells decreased the expression of PTEN, thus reversing the change in its expression observed during cell differentiation. In addition, multiplex immunofluorescence staining confirmed the relationship between the expression of PTEN and villin, a marker of intestinal cell differentiation, ranging from a moderate correlation in undifferentiated cells to a very strong correlation in differentiated cells treated with TPPU. Furthermore, we confirm that PTEN and sEH mirrored their expression patterns in samples of prenatal and adult human intestine compared to tumours using immunohistochemical staining. Taken together, it appears that PTEN and sEH cooperate in the process of intestinal cell differentiation. A better understanding of the crosstalk between the PI3K pathway and sEH and its consequences for cell differentiation is highly desirable, as several sEH inhibitors are under clinical investigation for the treatment of various diseases.
Fibrates are widely used hypolipidaemic agents that act as ligands of the peroxisome proliferator-activated receptor α (PPARα). p38 is a protein kinase that is mainly activated by environmental and genotoxic stress. We investigated the effect of the PPARα activators fenofibrate and WY-14643 and the PPARα inhibitor GW6471 on the levels of activated p38 (p-p38) in the colorectal cancer cell lines HT-29 and Caco2 in relation to their differentiation status. Fibrates increased p-p38 in undifferentiated HT-29 cells, whereas in other cases p-p38 expression was decreased. HT-29 cells showed p-p38 predominantly in the cytoplasm, whereas Caco2 cells showed higher nuclear positivity. The effect of fibrates may depend on the differentiation status of the cell, as differentiated HT-29 and undifferentiated Caco2 cells share similar characteristics in terms of villin, CYP2J2, and soluble epoxide hydrolase (sEH) expression. In human colorectal carcinoma, higher levels of p-p38 were detected in the cytoplasm, whereas in normal colonic surface epithelium, p-p38 showed nuclear positivity. The decrease in p-p38 positivity was associated with a decrease in sEH, consistent with in vitro results. In conclusion, fibrates affect the level of p-p38, but its exact role in the process of carcinogenesis remains unclear and further research is needed in this area.
- MeSH
- buněčná diferenciace MeSH
- Caco-2 buňky MeSH
- deriváty kyseliny fibrové farmakologie MeSH
- hypolipidemika * farmakologie MeSH
- lidé MeSH
- PPAR alfa * metabolismus MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
BACKGROUND: In Turner syndrome (TS), fluorescent in situ hybridization (FISH) karyotyping offers an alternative to classical karyotyping. OBJECTIVE: We tested the added value of FISH karyotyping from lymphocytes (mesodermal origin), buccal cells (ectodermal origin), and a rear-tongue smear (endodermal origin) to determine the 45,X cell line fraction and its impact on patient phenotype. DESIGN AND PATIENTS: Classical karyotyping and three FISH assays were done in 153 girls and women previously diagnosed with TS in four university hospitals. The 45,X cell line fraction was determined for each method and correlated with the major phenotypic signs. RESULTS: Classical karyotyping identified 45,X/46,XX mosaicism in 77/153 subjects (50%), 45,X monosomy in 52/153 (34%), and other karyotypes in 24/153 (16%). FISH from lymphocytes verified 45,X in 47/52 original cases, whereas 4/52 had 45,X/46,XX and 1/52 45,X/47,XYY mosaicism. The 45,X cell line fraction was higher in FISH from lymphocytes compared to classical karyotyping (median 86.4% vs. 70.0%; p < 0.001), while there was no difference for FISH from buccal or rear-tongue smear cells. The mean 45,X cell line fraction was more abundant in patients with several of the characteristic phenotypic signs compared to patients without them (p < 0.01), but the predictive power was insufficient. CONCLUSION: FISH analysis confirmed the findings of classical karyotyping; only a few 45,X monosomy cases were reclassified as mosaics. The 45,X cell line fraction did not show clinically meaningful prediction of the phenotype. FISH analysis of buccal or rear-tongue epithelial cells may be a non-inferior, less invasive alternative to classical karyotyping.
We investigated the effects of PPARα activators fenofibrate and WY-14643 as well as the PPARα inhibitor GW6471 on the PI3K/Akt/PTEN pathway of intestinal cell differentiation. Our previous study showed that all these compounds increased the expression of villin, a specific marker of intestinal cell differentiation in HT-29 and Caco2 cells. Our current results confirmed the central role of lipid messenger phosphatidylinositol-4,5-bisphosphate (PIP2), a known player in brush border formation, in mediating the effects of tested PPARα ligands. Although all tested compounds increased its levels, surprisingly, each of them affected different PIP2-metabolizing enzymes, especially the levels of PIP5K1C and PTEN. Moreover, we found a positive relationship between the expression of PPARα itself and PIP2 as well as PIP5K1C. By contrast, PPARα was negatively correlated with PTEN. However, the expression of antigens of interest was independent of PPARα subcellular localization, suggesting that it is not directly involved in their regulation. In colorectal carcinoma tissues we found a decrease in PTEN expression, which was accompanied by a change in its subcellular localization. This change was also observed for the regulatory subunit of PI3K. Taken together, our data revealed that fenofibrate, WY-14643, and GW6471 affected different members of the PI3K/Akt/PTEN pathway. However, these effects were PPARα-independent.
- Publikační typ
- časopisecké články MeSH
Musculus masseter je párový čtyřhranný vícezpeřený masivní žvýkací sval, rozepjatý od jařmového oblouku po úhel a dolní části ramene dolní čelisti. Má zásadní význam v estetice obličeje, je důležitým znakem pohlavního dimorfismu. Je to jeden z nejsilnějších žvýkacích svalů, tvořený převážně červenými svalovými vlákny. Společně s musculus pterygoideus medialis provádí elevaci mandibuly, v rámci pterygomaseterového poutka. Kromě toho se podílí na protruzi, retruzi a laterotruzi mandibuly. Hraje zásadní roli v etiopatogenezi kraniomandibulárních parafunkcí, dislokaci zlomenin mandibuly, relapsu po ortognátních operacích. Je velmi dobře vyšetřitelný palpačně i sonograficky, terapeuticky jej lze dobře ovlivnit přímými i nepřímými fyzioterapeutickými technikami.
Being composed mainly of red muscle fibers, the masseter is one of the strongest muscles of mastication. The masseter muscle is a paired, quadrangular multipennate in structure. It originates from zygomatic arch and runs to angle and lower part of the mandibular ramus. It has a crucial role in facial aesthetics representing an important sign of sexual dimorphism. The masseter helps with mandibular protrusion, retrusion and laterotrusion; together with medial pterygoid muscle it provides mandibular elevation, as a part of pterygomasseteric sling. When in dysfunction, the masseter has a crucial role in etiopathogenesis of craniomandibular parafunctions, dislocations of mandibular fractures and relapses after orthognathic surgical approaches. Examination of the masseter can be carried out by palpation and sonography. Therapeutically, the masseter can be well influenced by both direct or indirect physiotherapeutical techniques.
- MeSH
- lidé MeSH
- musculus masseter * anatomie a histologie fyziologie inervace krevní zásobení MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- přehledy MeSH
- MeSH
- barvení a značení MeSH
- buněčné jádro * MeSH
- fluorescenční barviva MeSH
- fluorescenční protilátková technika MeSH
- indoly * MeSH
- lidé MeSH
- propidium MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- dopisy MeSH
- práce podpořená grantem MeSH
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
AIMS: Turner syndrome is the only chromosome monosomy that is postnatally compatible with life. The reported incidence of TS is 1 in 2500 liveborn girls. The phenotype of these girls is highly variable, with cardiac abnormalities being life-threatening defects. The aim of the study was to reveal the possible influence of the parental origin of the X chromosome in these patients on a selected phenotype that is associated with Turner syndrome. Selected symptoms and parameters were: a bicuspid aortic valve, aortic coarctation, lymphoedema, pterygium colli, coeliac disease, thyroiditis, otitis media, diabetes mellitus 2, renal abnormalities, spontaneous puberty, and IVF. METHODS: The X chromosome haplotype was determined for a group of 45,X patients verified by native FISH. A molecular diagnostic method based on the detection of different lengths of X chromosome-linked STR markers using the Argus X-12 QS kit was used to determine the X haplotype. RESULTS: Our results, analysed by Fisher's exact (factorial) test, suggest independence between the maternal/paternal origin of the inherited X chromosome and the presence of the anomalies that were studied (P=1 to P=0.34). CONCLUSION: In the group of 45,X patients, who were precisely selected by means of the native FISH method, no correlation was demonstrated with the parental origin of the X chromosome and the observed symptom.
- MeSH
- chromozom X MeSH
- fenotyp MeSH
- haplotypy MeSH
- lidé MeSH
- Turnerův syndrom * genetika MeSH
- vrozené srdeční vady * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
