RESUMEN
Although some clinical studies have reported increased mitochondrial respiration in patients with fatty liver and early nonalcoholic steatohepatitis (NASH), there is a lack of in vitro models of nonalcoholic fatty liver disease (NAFLD) with similar findings. Despite being the most commonly used immortalized cell line for in vitro models of NAFLD, HepG2 cells exposed to free fatty acids (FFAs) exhibit a decreased mitochondrial respiration. On the other hand, the use of HepaRG cells to study mitochondrial respiratory changes following exposure to FFAs has not yet been fully explored. Therefore, the present study aimed to assess cellular energy metabolism, particularly mitochondrial respiration, and lipotoxicity in FFAtreated HepaRG and HepG2 cells. HepaRG and HepG2 cells were exposed to FFAs, followed by comparative analyses that examained cellular metabolism, mitochondrial respiratory enzyme activities, mitochondrial morphology, lipotoxicity, the mRNA expression of selected genes and triacylglycerol (TAG) accumulation. FFAs stimulated mitochondrial respiration and glycolysis in HepaRG cells, but not in HepG2 cells. Stimulated complex I, IIdriven respiration and ßoxidation were linked to increased complex I and II activities in FFAtreated HepaRG cells, but not in FFAtreated HepG2 cells. Exposure to FFAs disrupted mitochondrial morphology in both HepaRG and HepG2 cells. Lipotoxicity was induced to a greater extent in FFAtreated HepaRG cells than in FFAtreated HepG2 cells. TAG accumulation was less prominent in HepaRG cells than in HepG2 cells. On the whole, the present study demonstrates that stimulated mitochondrial respiration is associated with lipotoxicity in FFAtreated HepaRG cells, but not in FFAtreated HepG2 cells. These findings suggest that HepaRG cells are more suitable for assessing mitochondrial respiratory adaptations in the developed in vitro model of earlystage NASH.
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Enfermedad del Hígado Graso no Alcohólico , Humanos , Células Hep G2 , Mitocondrias , Respiración , Línea Celular , Ácidos Grasos no Esterificados , TriglicéridosRESUMEN
Aim: This investigation was designed to synthesize half-sandwich Rh(III) and Ru(II) complexes and study their antiproliferative activity in human cancer cell lines. Materials & methods: Nine compounds were prepared and tested by various assays for their anticancer activity and mechanism of action. Results: Hit Rh(III) complex 6 showed low-micromolar potency in cisplatin-sensitive (A2780) and -resistant (A2780cis) ovarian carcinoma cell lines, promising selectivity toward these cancer cells over normal lung fibroblasts and an unprecedented mechanism of action in the treated cells. DNA synthesis was decreased and CDKN1A expression was upregulated, but p21 expression was not induced. Conclusion: Rh complex 6 showed high antiproliferative activity, which is induced through a p21-independent mechanism of action.
Nine rhodium(III)and ruthenium(II) complexes were developed and screened for their anticancer activity on a panel of human cancer cell lines. The best-performing rhodium(III) complex (6) showed high activity in ovarian cancer cells, including the variant resistant to the conventional anticancer drug cisplatin, while it was less effective towards non-cancerous lung fibroblasts. In cancer cells, compound 6 induced a modification of the cell cycle connected with a significant decrease in DNA synthesis, which was not observed for cisplatin. The effect of 6 on the expression of proteins related to the cell cycle modification was analysed by quantitative PCR and western blot in cancer cells and the results indicated a p21-independent mode of anticancer action, which is different from cisplatin.
RESUMEN
In the presence of carboxypeptidase, the hydrolytically stable complex [Os(η6-pcym)(L2)Cl]PF6 (2) partially released the bioactive substituent indomethacin, bound through the amide bond to the chelating 2-(1,3,4-thiadiazol-2-yl)pyridine-based moiety of L2. Stability in the presence of other relevant biomolecules (GSH, NADH, GMP) and cancer cell viability were also studied.
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Antineoplásicos , Complejos de Coordinación , Antineoplásicos/química , Carboxipeptidasas A , Línea Celular Tumoral , Complejos de Coordinación/química , Complejos de Coordinación/farmacología , Indometacina/farmacología , LigandosRESUMEN
INTRODUCTION: The inflammatory process in Crohn's disease (CD) is closely associated with the formation of reactive oxygen species. Antioxidant enzymes can play an important role in the outcome of CD and may influence postoperative recurrence in these patients. The aim of our study was to evaluate gene expression of intracellular antioxidant enzymes in surgically resected intestinal specimens of patients with CD, both in macroscopically normal and in inflamed tissue. METHODS: A total of 28 patients referred for elective bowel resection were enrolled in the study. Full-thickness small intestinal specimens were investigated. Gene expression of antioxidant enzymes - superoxide dismutase (SOD), glutathione peroxidase (GPX), glutathione reductase (GSR) - was evaluated both in macroscopically normal and inflamed samples. RESULTS: There were significantly lower levels of SOD1 mRNA (p = 0.007) and GSR mRNA (p = 0.027) in inflamed tissue compared to macroscopically normal areas. No significant differences were found between affected and non-affected intestinal segments in mRNA for SOD2, SOD3 and GPX. CONCLUSIONS: Our pilot data clearly showed that the gene expression of major antioxidant enzymes is not a uniform mechanism in the pathogenesis of Crohn's disease. Topically decreased gene expression of SOD1 and GSR might facilitate the segmental tissue injury caused by reactive oxygen species.
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Antioxidantes , Enfermedad de Crohn , Expresión Génica , Superóxido Dismutasa-1 , Enfermedad de Crohn/genética , Enfermedad de Crohn/metabolismo , Glutatión Peroxidasa/genética , Humanos , Intestinos , ARN Mensajero/genética , Especies Reactivas de Oxígeno , Superóxido Dismutasa/genética , Superóxido Dismutasa-1/genética , Superóxido Dismutasa-1/metabolismoRESUMEN
Mitochondria play an essential role in the pathogenesis of nonalcoholic fatty liver disease (NAFLD). Previously, we found that succinate-activated respiration was the most affected mitochondrial parameter in mice with mild NAFLD. In this study, we focused on the role of succinate dehydrogenase (SDH) in NAFLD pathogenesis. To induce the progression of NAFLD to nonalcoholic steatohepatitis (NASH), C57BL/6J mice were fed a Western-style diet (WD) or control diet for 30 weeks. NAFLD severity was evaluated histologically and the expression of selected proteins and genes was assessed. Mitochondrial respiration was measured by high-resolution respirometry. Liver redox status was assessed using glutathione, malondialdehyde, and mitochondrial production of reactive oxygen species (ROS). Metabolomic analysis was performed by GC/MS. WD consumption for 30 weeks led to reduced succinate-activated respiration. We also observed decreased SDH activity, decreased expression of the SDH activator sirtuin 3, decreased gene expression of SDH subunits, and increased levels of hepatic succinate, an important signaling molecule. Succinate receptor 1 (SUCNR1) gene and protein expression were reduced in the livers of WD-fed mice. We did not observe signs of oxidative damage compared to the control group. The changes observed in WD-fed mice appear to be adaptive to prevent mitochondrial respiratory chain overload and massive ROS production.
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Dieta Occidental , Mitocondrias Hepáticas/metabolismo , Enfermedad del Hígado Graso no Alcohólico/etiología , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Oxidación-Reducción , Estrés Oxidativo , Ácido Succínico/metabolismo , Animales , Apoptosis , Biomarcadores , Respiración de la Célula , Modelos Animales de Enfermedad , Susceptibilidad a Enfermedades , Fibrosis , Metaboloma , Metabolómica/métodos , Ratones , Enfermedad del Hígado Graso no Alcohólico/patología , Succinato Deshidrogenasa/metabolismoRESUMEN
Pancracine, a montanine-type Amaryllidaceae alkaloid (AA), is one of the most potent compounds among natural isoquinolines. In previous studies, pancracine exhibited cytotoxic activity against diverse human cancer cell lines in vitro. However, further insight into the molecular mechanisms that underlie the cytotoxic effect of pancracine have not been reported and remain unknown. To fill this void, the cell proliferation and viability of cancer cells was explored using the Trypan Blue assay or by using the xCELLigence system. The impact on the cell cycle was determined by flow cytometry. Apoptosis was evaluated by Annexin V/PI and by quantifying the activity of caspases (-3/7, -8, and -9). Proteins triggering growth arrest or apoptosis were detected by Western blotting. Pancracine has strong antiproliferative activity on A549 cells, lasting up to 96 h, and antiproliferative and cytotoxic effects on MOLT-4 cells. The apoptosis-inducing activity of pancracine in MOLT-4 cells was evidenced by the significantly higher activity of caspases. This was transmitted through the upregulation of p53 phosphorylated on Ser392, p38 MAPK phosphorylated on Thr180/Tyr182, and upregulation of p27. The pancracine treatment negatively altered the proliferation of A549 cells as a consequence of an increase in G1-phase accumulation, associated with the downregulation of Rb phosphorylated on Ser807/811 and with the concomitant upregulation of p27 and downregulation of Akt phosphorylated on Thr308. This was the first study to glean a deeper mechanistic understanding of pancracine activity in vitro. Perturbation of the cell cycle and induction of apoptotic cell death were considered key mechanisms of pancracine action.
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Adenocarcinoma del Pulmón/patología , Proliferación Celular/efectos de los fármacos , Compuestos Heterocíclicos de 4 o más Anillos/farmacología , Leucemia/patología , Neoplasias Pulmonares/patología , Células A549 , Alcaloides/aislamiento & purificación , Alcaloides/farmacología , Amaryllidaceae/química , Antineoplásicos Fitogénicos/aislamiento & purificación , Antineoplásicos Fitogénicos/farmacología , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Células Hep G2 , Compuestos Heterocíclicos de 4 o más Anillos/aislamiento & purificación , Humanos , Células MCF-7RESUMEN
Gastrointestinal side effects of donepezil, including dyspepsia, nausea, vomiting or diarrhea, occur in 20-30% of patients. The pathogenesis of these dysmotility associated disorders has not been fully clarified yet. Pharmacokinetic parameters of donepezil and its active metabolite 6-O-desmethyldonepezil were investigated in experimental pigs with and without small intestinal injury induced by dextran sodium sulfate (DSS). Morphological features of this injury were evaluated by a video capsule endoscopy. The effect of a single and repeated doses of donepezil on gastric myoelectric activity was assessed. Both DSS-induced small intestinal injury and prolonged small intestinal transit time caused higher plasma concentrations of donepezil in experimental pigs. This has an important implication for clinical practice in humans, with a need to reduce doses of the drug if an underlying gastrointestinal disease is present. Donepezil had an undesirable impact on porcine myoelectric activity. This effect was further aggravated by DSS-induced small intestinal injury. These findings can explain donepezil-associated dyspepsia in humans.
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Donepezilo/farmacocinética , Tracto Gastrointestinal/patología , Tracto Gastrointestinal/fisiopatología , Indanos/metabolismo , Metaboloma , Complejo Mioeléctrico Migratorio , Piperidinas/metabolismo , Estómago/fisiopatología , Animales , Endoscopía Capsular , Sulfato de Dextran , Donepezilo/química , Donepezilo/farmacología , Femenino , Tracto Gastrointestinal/efectos de los fármacos , Metaboloma/efectos de los fármacos , Complejo Mioeléctrico Migratorio/efectos de los fármacos , Estómago/efectos de los fármacos , PorcinosRESUMEN
S100 proteins are involved in the pathogenesis of sporadic colorectal carcinoma through different mechanisms. The aim of our study was to assess tissue mRNA encoding S100 proteins in patients with non-advanced and advanced colorectal adenoma. Mucosal biopsies were taken from the caecum, transverse colon and rectum during diagnostic and/or therapeutic colonoscopy. Another biopsy was obtained from adenomatous tissue in the advanced adenoma group. The tissue mRNA for each S100 protein (S100A4, S100A6, S100A8, S100A9, S100A11 and S100P) was investigated. Eighteen biopsies were obtained from the healthy mucosa in controls and the non-advanced adenoma group (six individuals in each group) and thirty biopsies in the advanced adenoma group (ten patients). Nine biopsies were obtained from advanced adenoma tissue (9/10 patients). Significant differences in mRNA investigated in the healthy mucosa were identified between (1) controls and the advanced adenoma group for S100A6 (p = 0.012), (2) controls and the non-advanced adenoma group for S100A8 (p = 0.033) and (3) controls and the advanced adenoma group for S100A11 (p = 0.005). In the advanced adenoma group, differences between the healthy mucosa and adenomatous tissue were found in S100A6 (p = 0.002), S100A8 (p = 0.002), S100A9 (p = 0.021) and S100A11 (p = 0.029). Abnormal mRNA expression for different S100 proteins was identified in the pathological adenomatous tissue as well as in the morphologically normal large intestinal mucosa.
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Adenoma/patología , Calgranulina A/metabolismo , Calgranulina B/metabolismo , Proteínas de Ciclo Celular/metabolismo , Neoplasias Colorrectales/patología , ARN Mensajero/metabolismo , Proteína A6 de Unión a Calcio de la Familia S100/metabolismo , Proteínas S100/metabolismo , Adenoma/genética , Adenoma/metabolismo , Anciano , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Proteínas de Unión al Calcio/genética , Proteínas de Unión al Calcio/metabolismo , Calgranulina A/genética , Calgranulina B/genética , Estudios de Casos y Controles , Proteínas de Ciclo Celular/genética , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/metabolismo , Femenino , Estudios de Seguimiento , Humanos , Masculino , Persona de Mediana Edad , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Proyectos Piloto , Pronóstico , ARN Mensajero/genética , Proteína A6 de Unión a Calcio de la Familia S100/genética , Proteína de Unión al Calcio S100A4/genética , Proteína de Unión al Calcio S100A4/metabolismo , Proteínas S100/genéticaRESUMEN
Maladaptation of mitochondrial oxidative flux seems to be a considerable feature of nonalcoholic fatty liver disease (NAFLD). The aim of this work was to induce NAFLD in mice fed a Western-style diet (WD) and to evaluate liver mitochondrial functions. Experiments were performed on male C57BL/6J mice fed with a control diet or a WD for 24 weeks. Histological changes in liver and adipose tissue as well as hepatic expression of fibrotic and inflammatory genes and proteins were evaluated. The mitochondrial respiration was assessed by high-resolution respirometry. Oxidative stress was evaluated by measuring lipoperoxidation, glutathione, and reactive oxygen species level. Feeding mice a WD induced adipose tissue inflammation and massive liver steatosis accompanied by mild inflammation and fibrosis. We found decreased succinate-activated mitochondrial respiration and decreased succinate dehydrogenase (SDH) activity in the mice fed a WD. The oxidative flux with other substrates was not affected. We observed increased ketogenic capacity, but no impact on the capacity for fatty acid oxidation. We did not confirm the presence of oxidative stress. Mitochondria in this stage of the disease are adapted to increased substrate flux. However, inhibition of SDH can lead to the accumulation of succinate, an important signaling molecule associated with inflammation, fibrosis, and carcinogenesis.
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Peroxidación de Lípido , Mitocondrias Hepáticas/metabolismo , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Tejido Adiposo/metabolismo , Tejido Adiposo/patología , Animales , Dieta Alta en Grasa/efectos adversos , Glutatión/metabolismo , Hígado/metabolismo , Hígado/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Enfermedad del Hígado Graso no Alcohólico/etiología , Succinato Deshidrogenasa/metabolismoRESUMEN
BACKGROUND: Memantine, currently available for the treatment of Alzheimer's disease, is an uncompetitive antagonist of the N-methyl-D-aspartate type of glutamate receptors. Under normal physiologic conditions, these unstimulated receptor ion channels are blocked by magnesium ions, which are displaced after agonist-induced depolarization. In humans, memantine administration is associated with different gastrointestinal dysmotility side effects (vomiting, diarrhoea, constipation, motor-mediated abdominal pain), thus limiting its clinical use. Mechanism of these motility disorders has not been clarified yet. Pigs can be used in various preclinical experiments due to their relatively very similar gastrointestinal functions compared to humans. The aim of this study was to evaluate the impact of a single and repeated doses of memantine on porcine gastric myoelectric activity evaluated by means of electrogastrography (EGG). METHODS: Six adult female experimental pigs (Sus scrofa f. domestica, mean weight 41.7±5.0 kg) entered the study for two times. The first EGG was recorded after a single intragastric dose of memantine (20 mg). In the second part, EGG was accomplished after 7-day intragastric administration (20 mg per day). All EGG recordings were performed under general anaesthesia. Basal (15 minutes) and study recordings (120 minutes) were accomplished using an EGG stand (MMS, Enschede, the Netherlands). Running spectral analysis based on Fourier transform was used. Results were expressed as dominant frequency of gastric slow waves (DF) and power analysis (areas of amplitudes). RESULTS: Single dose of memantine significantly increased DF, from basic values (1.65±1.05 cycles per min.) to 2.86 cpm after 30 min. (p = 0.008), lasting till 75 min. (p = 0.014). Basal power (median 452; inter-quartile range 280-1312 µV^2) raised after 15 min. (median 827; IQR 224-2769; p = 0.386; NS), lasting next 30 min. Repetitively administrated memantine caused important gastric arrhythmia. Basal DF after single and repeated administration was not different, however, a DF increase in the second part was more prominent (up to 3.18±2.16 after 15 and 30 min., p<0.001). In comparison with a single dose, basal power was significantly higher after repetitively administrated memantine (median 3940; IQR 695-15023 µV^2; p<0.001). Next dose of 20 mg memantine in the second part induced a prominent drop of power after 15 min. (median 541; IQR 328-2280 µV^2; p<0.001), lasting till 120 min. (p<0.001). CONCLUSIONS: Both single and repeated doses of memantine increased DF. Severe gastric arrhythmia and long-lasting low power after repeated administration might explain possible gastric dysmotility side effects in the chronic use of memantine.
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Antagonistas de Aminoácidos Excitadores/efectos adversos , Enfermedades Gastrointestinales/inducido químicamente , Motilidad Gastrointestinal/efectos de los fármacos , Memantina/efectos adversos , Estómago/efectos de los fármacos , Administración Oral , Enfermedad de Alzheimer/tratamiento farmacológico , Animales , Modelos Animales de Enfermedad , Electromiografía , Antagonistas de Aminoácidos Excitadores/administración & dosificación , Femenino , Enfermedades Gastrointestinales/diagnóstico , Enfermedades Gastrointestinales/fisiopatología , Motilidad Gastrointestinal/fisiología , Humanos , Memantina/administración & dosificación , Estómago/fisiopatología , Sus scrofaRESUMEN
A series of six indenyl molybdenum compounds bearing a thiophenyl function in the side chain were prepared and characterized by analytical and spectroscopic methods. The structures of [(η5-C9H6CH2C4H3S)(η3-C3H5)Mo(CO)2] and [(η5-C9H6CH2C4H3S)Mo(CO)2(bpy)][BF4] were determined by single-crystal X-ray diffraction. The compounds bearing N,N-chelating ligands exhibit increased cytotoxic activity against human leukemia cell lines MOLT-4; up to two orders of magnitude lower IC50 values were observed compared to analogues with unsubstituted indenyl, which clearly demonstrates the strong effect of the indenyl ligand modification on the biological activity of the molybdenum(ii) compounds. The highest cytostatic potential was observed for the complex bearing 4,7-diphenyl-1,10-phenanthtoline [(η5-C9H6CH2C4H3S)Mo(CO)2(Ph2phen)][BF4] with IC50 (MOLT-4) = 0.19 ± 0.02 µM. Detailed regulation of the molecular and cellular mechanism by this derivative was investigated on the lung carcinoma cell line A549 and compared with the lung fibroblast cell line MRC-5. Rather unusual differences in the effects on tumor and non-tumor cell lines provide a unique insight into the cytostatic action of molybdenum(ii) complexes.
RESUMEN
Ten new vanadocene complexes bearing N,N'-chelating ligands were prepared, characterized, and their cytotoxicity toward a panel of cancer cells was measured. Structures of four vanadocene compounds were determined by single crystal X-ray diffraction analysis. Complexes containing 1,2-bis(phenylimino)acenaphthene (bian) and 1,2-bis(4-methoxyphenylimino)acenaphthene (4-MeO-bian) exhibit higher cytotoxicity than those with dipyrido[3,2-a:2',3'-c]phenazine (dppz) and (E)-N-((pyridin-2-yl)methylene)benzenamine (pyma). In light of the finding, cytotoxic mechanisms of two highly effective complexes [(η5-C5H4Me)2V(bian)][OTf]2 (3b) and [(η5-C5H4Me)2V(4-MeO-bian)][OTf]2 (4b) against human A549 lung adenocarcinoma cells were investigated by following membrane leakage of intracellular lactate dehydrogenase, Trypan Blue staining and activation of tumor protein p53 (p53). Evaluated complexes have a potent dose-dependent antiproliferative activity, causing cell cycle redistribution by the increased accumulation of cells in the G2 and S phase. In accord with the observed cell cycle deceleration, cyclin-dependent kinase inhibitor-interacting protein 1 (p21WAF1/Cip1), extracellular signal-regulated kinases 1 and 2 (ERK1/2), Checkpoint kinase 1 (Chk1), Checkpoint kinase 2 (Chk2) and their phosphorylated forms Chk1 at serine 345 and Chk2 at threonine 68 increased. In the cells exposed to complexes, dose- and time-dependent apoptotic process is initiated by the activation of the initiator caspase 8, followed by activation of effector caspase 3/7 and phosphatidylserine externalization. Moreover, because of treatment, A549 cells activate prosurvival mitogen-activated protein kinases (MAPK) signaling and up-regulate antiapoptotic protein B-cell lymphoma (Bcl-2), thereby promoting evasion of cell death. Both complexes exhibited considerably higher cytotoxic effect than the reference anticancer drug cis-platin and the cytotoxicity was more pronounced at higher treatment time.
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Antineoplásicos/farmacología , Quelantes/farmacología , Complejos de Coordinación/farmacología , Antineoplásicos/síntesis química , Antineoplásicos/química , Apoptosis/efectos de los fármacos , Caspasas/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Quelantes/síntesis química , Quelantes/química , Complejos de Coordinación/síntesis química , Complejos de Coordinación/química , Ensayos de Selección de Medicamentos Antitumorales , Puntos de Control de la Fase G2 del Ciclo Celular/efectos de los fármacos , Humanos , Ligandos , Estructura Molecular , Vanadio/químicaRESUMEN
Histone modifications play a key role in the epigenetic regulation of gene transcription in cancer cells. Histone acetylations are regulated by two classes of enzymes, histone acetyltransferases (HATs) and histone deacetylases (HDACs). HDACs are increased in ovarian carcinomas and they are involved in carcinogenesis and resistance to chemotherapeutic agents. In our study we investigated anticancer effect of HDAC inhibitor sodium butyrate (NaBu) on cisplatin-sensitive and cisplatin-resistant ovarian cell lines A2780 and A2780cis. A2780 and A2780cis were treated with NaBu alone or in combination with cisplatin (CP). NaBu inhibited the growth of both cell lines and enhanced cytotoxic effect of CP. Exposure to NaBu for 24 h induced cell cycle arrest. The expressions of EMT-related genes and proteins were further investigated by qPCR and western blot analysis. Loss of E-cadherin has been shown to be crucial in ovarian cancer development. We found that NaBu dramatically induce expression of E-cadherin gene (CDH1) and protein levels in A2780 and A2780cis. We investigated correlation between transcription and methylation of CDH1gene. Methylation level analysis in 32 CpG sites in CDH1 gene (promoter/exon1 regions) was performed using bisulfite NGS (Next Generation Sequencing). We found that cisplatin-resistant cell line A2780cis cells differ from their cisplatin-sensitive counterparts in the CDH1 methylation. Methylation in A2780cis cells is elevated compared to A2780. However, NaBu-induced expression of CDH1 was not accompanied by CDH1 demethylation. NaBu treatment induced changes in expression of EMT-related genes and proteins. Interestingly E-cadherin zinc finger transcriptional repressor SNAIL1 was upregulated in both cell lines. Mesenchymal marker vimentin was downregulated. Matrix metalloproteases (MMPs) are necessary for pericellular proteolysis and facilitate migration and invasion of tumour cells. NaBu induced mRNA expression of MMPs, mild changes in activities of gelatinases MMP2 and MMP9 were detected. Our data demonstrate that NaBu sensitizes cisplatin-resistant ovarian cancer cells, re-established E-cadherin expression, but it was not able to reverse the EMT phenotype completely.
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Ácido Butírico/farmacología , Cisplatino/farmacología , Transición Epitelial-Mesenquimal/efectos de los fármacos , Transición Epitelial-Mesenquimal/genética , Antígenos CD/genética , Antineoplásicos/farmacología , Cadherinas/genética , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Resistencia a Antineoplásicos/genética , Epigénesis Genética/efectos de los fármacos , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Marcadores Genéticos , Código de Histonas/efectos de los fármacos , Inhibidores de Histona Desacetilasas/farmacología , Humanos , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/genética , Neoplasias Ováricas/patologíaRESUMEN
Differential diagnosis between benign and malignant biliary stenosis can be difficult in clinical practice. Histology of biopsy specimens is often indeterminate. Laboratory markers (serum bilirubin > 75 µmol/L, carbohydrate antigen 19-9 > 400 U/mL) and the length of stenosis (>15 mm) can be helpful but are not specific enough. The aim of this study was to investigate bile acids in liver bile of patients with benign and malignant biliary stenosis and controls without stenosis. A total of 73 patients entered the study: 7 subjects with benign biliary stenosis (6 men, 1 woman; 68 ± 13 years old), 21 with malignant biliary stenosis (15 men, 6 women; 72 ± 14 years old), and 45 patients without biliary stenosis (22 men, 23 women; 70 ± 13 years old); out of those, 25 subjects have and 20 do not have choledocholithiasis. Twenty-three different bile acids were investigated by high-performance liquid chromatography/mass spectrometry. Serum total bilirubin was significantly higher in patients with malignant biliary stenosis compared with nonstenotic controls (p = 0.005). Significant relationship (r > 0.7) was found between several pairs of bile acids. Significantly lower bile acid concentrations in malignant biliary stenosis compared to controls without stenosis were found for GLCA (p = 0.032), GUDCA (p = 0.032), GCDCA (p = 0.006), GDCA (p = 0.031), GHCA (p = 0.005), TUDCA (p = 0.044), and TDCA (p = 0.036). Significant difference in cholic acid was found between benign and malignant stenosis (p = 0.022). Analysis of bile acids might be helpful in the differential diagnosis of malignant and benign biliary stenosis. More patients need to be enrolled in further studies so that the real diagnostic yield of bile acids can be determined.
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Analysis of Exhaled breath condensate (EBC) is a re-discovered approach to monitoring the course of the disease and reduce invasive methods of patient investigation. However, the major disadvantage and shortcoming of the EBC is lack of reliable and reproducible standardization of the method. Despite many articles published on EBC, until now there is no clear consensus on whether the analysis of EBC can provide a clue to diagnosis of the diseases. The purpose of this paper is to investigate our own method, to search for possible standardization and to obtain our own initial experience. Thirty healthy volunteers provided the EBC, in which we monitored the density, pH, protein, chloride and urea concentration. Our results show that EBC pH is influenced by smoking, and urea concentrations are affected by the gender of subjects. Age of subjects does not play a role. The smallest coefficient of variation between individual volunteers is for density determination. Current limitations of EBC measurements are the low concentration of many biomarkers. Standardization needs to be specific for each individual biomarker, with focusing on optimal condensate collection. EBC analysis has a potential become diagnostic test, not only for lung diseases.
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Pruebas Respiratorias/métodos , Cloruros/metabolismo , Proteínas/metabolismo , Urea/metabolismo , Adulto , Factores de Edad , Anciano , Biomarcadores/análisis , Biomarcadores/metabolismo , Cloruros/análisis , Femenino , Voluntarios Sanos , Humanos , Concentración de Iones de Hidrógeno , Masculino , Persona de Mediana Edad , Proyectos Piloto , Proteínas/análisis , Estándares de Referencia , Valores de Referencia , Factores Sexuales , Fumar/metabolismo , Manejo de Especímenes , Urea/análisisRESUMEN
Myofibroblast expansion is a critical event in the pathogenesis of liver fibrosis. The activation of hepatic stellate cells (HSC) to myofibroblast (MFB) results in the enhanced production of extracellular matrix (ECM). In this study, we explored the effect of acidic fibroblast growth factor (FGF-1) treatment on a transforming growth factor (TGF-ß1) induced MFB conversion. We used HSC-T6 cell line, which represents well-established model of activated HSC. These cells strongly expressed α-smooth muscle actin (α-SMA) and fibronectin (FN-EDA) after stimulation with TGF-ß1, which is a stimulus for MFB differentiation and ECM production. FGF-1 reduced proteins expression to levels comparable with untreated cells. Mild repression of secreted gelatinases was seen in culture media after FGF-1 treatment. The exposure of cells to collagen gel leads to changes in cell morphology and in expression of MFB markers. Lack of α-SMA in cells embedded to collagen gel was detected. When stimulated with TGF-ß1, the cells increased expression of FN-EDA, but not α-SMA. Although the cells on plastic and in collagen gel show different properties, FGF-1 reduced expression of FN-EDA in both conditions. Disrupting TGF-ß1 signalling pathway represents a potential strategy for the treatment of fibrosis. We showed that FGF-1 could antagonize signals initiated by TGF-ß1.
Asunto(s)
Diferenciación Celular/efectos de los fármacos , Factor 1 de Crecimiento de Fibroblastos/farmacología , Expresión Génica/efectos de los fármacos , Células Estrelladas Hepáticas/fisiología , ARN Mensajero/análisis , Factor de Crecimiento Transformador beta/metabolismo , Actinas/genética , Actinas/metabolismo , Animales , Línea Celular , Colágeno/farmacología , Fibronectinas/genética , Fibronectinas/metabolismo , Miofibroblastos , Ratas , Factor de Crecimiento Transformador beta/genética , Factor de Crecimiento Transformador beta/farmacologíaRESUMEN
Epigallocatechin gallate (EGCG) is an antioxidant found in green tea. In this study, male Wistar rats were subjected either to partial hepatectomy (PHx), or a sham operation (LAP). Twenty-four hours after surgery, hepatocytes were isolated and treated with various concentrations of EGCG for up to 72 h. We then measured markers of cell viability, oxidative stress, DNA synthesis, and caspase activity. Morphological criteria, cell viability tests, and albumin synthesis revealed toxicity starting at 10 µmol/L. DNA synthesis was higher in hepatocytes isolated from rats after PHx and inhibited by EGCG. Furthermore, EGCG increased the activity of caspases 3 and 7, seen more in hepatocytes from PHx rats. In conclusion, EGCG at a concentration of 10 µmol/L was toxic for hepatocytes isolated from both PHx and LAP rats.
Asunto(s)
Antioxidantes/toxicidad , Catequina/análogos & derivados , Hepatectomía , Hepatocitos/efectos de los fármacos , Animales , Antioxidantes/administración & dosificación , Caspasa 3/metabolismo , Caspasa 7/metabolismo , Catequina/administración & dosificación , Catequina/toxicidad , Supervivencia Celular/efectos de los fármacos , ADN/biosíntesis , Relación Dosis-Respuesta a Droga , Hepatocitos/enzimología , Hepatocitos/metabolismo , Masculino , Estrés Oxidativo/efectos de los fármacos , Cultivo Primario de Células , Ratas , Ratas WistarRESUMEN
BACKGROUND: Two-thirds partial hepatectomy (PHx) is an established model for the study of liver regeneration after resection. This process is accompanied by oxidative stress. AIMS: In our study, we tested the effect of epigallocatechin gallate (EGCG), a green tea antioxidant, on the early phase of liver regeneration after PHx. METHODS: Male Wistar rats were divided into five groups: (I) laparotomy + water for intraperitoneal injections, (II) laparotomy + EGCG 50 mg/kg body weight, (III) PHx + water for injections, (IV) PHx + EGCG 20 mg/kg and (V) PHx + EGCG 50 mg/kg, for 3 consecutive days. The rats were killed 24 h after surgery. Biochemical analysis of rat sera was performed. Histological samples were stained with hematoxylin & eosin and bromodeoxyuridine (BrdU). In hepatectomized rats, we also measured plasma malondialdehyde, tissue malondialdehyde, glutathione and cytokines levels, the activity of caspases 3/7, expression of Nqo-1 and HO-1 genes at the mRNA level, and expression of p21, p-p27 and p-p53 genes at the protein level. RESULTS: We observed lower accumulation of BrdU in group V when compared to groups III and IV. The activity of caspases 3/7 and expression of p-p53 were lower in group V than in groups III and IV. Tissue levels of IL-6 were lower in group V when compared to group III. Significant differences were not noted in other parameters. CONCLUSIONS: Administration of EGCG did not stimulate early phase liver regeneration in rats after PHx. There was even lower DNA synthesis in the group treated with a high dose of EGCG.