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1.
Folia Biol (Praha) ; 66(5-6): 155-160, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-34087971

RESUMEN

Early mouse neural stem cells (NSCs) first appear in embryonic day E5.5 and express pluripotency markers Oct4, Sox2, Nanog and early neural marker Sox1. Early NSCs are a good model for understanding the role of various pathways that control initial neural commitment. However, a protocol for differentiation of mouse embryonic stem cells (ESCs) into early NSCs by adherent monolayer culture has not yet been established. Hence, in this study, we identified the combination of growth factors and small molecules that differentiated mouse ESCs into early NSCs and supported their proliferation. Leukaemia inhibitory factor (LIF) was the first factor to be tested and it was found that ESCs can differentiate into early neurogenic lineage in the presence of LIF. However, we found that the induction is weaker in the presence of LIF as compared to cells differentiated in its absence. GSK-3 inhibitor, along with BMP and TGF-ß pathway inhibitor (dual SMAD inhibition), are commonly used to sequentially direct ESCs towards NSCs. However, when we used this combination, mouse ESCs failed to differentiate into early NSCs. We observed that by adding Wnt inhibitor to the combination of GSK-3 inhibitor, BMP inhibitor, TGF-ß inhibitor and LIF, it was possible to differentiate ESCs into early NSCs. qRT-PCR analysis of early NSCs illustrated that they expressed key pluripotency genes Oct4 and Nanog, albeit at levels lower than non-differentiated ESCs, along with early neural markers Sox1 and Pax6.


Asunto(s)
Glucógeno Sintasa Quinasa 3 , Células-Madre Neurales , Animales , Proteínas Morfogenéticas Óseas , Diferenciación Celular , Células Madre Embrionarias , Factor Inhibidor de Leucemia/farmacología , Ratones
2.
Folia Biol (Praha) ; 64(5-6): 155-166, 2018.
Artículo en Inglés | MEDLINE | ID: mdl-30938672

RESUMEN

The cellular components of the satellite cell niche participate in the regulation of skeletal muscle regeneration. Beside myogenic cells at different developmental stages, this niche is formed by cells of the immune system, the interstitial connective tissue and the vascular system. Unambiguous determination of the origin of these cell types could contribute to optimization of the cell-based therapy of skeletal muscle disorders. In our work, we intravenously transplanted mouse GFP+ unseparated bone marrow cells into whole-body lethally irradiated immunocompetent mice four weeks before cardiotoxin-induced injury of the recipients' skeletal muscles. Seven and 28 days after the toxin injection, the injured regenerating and contralateral intact muscles were examined for identification of GFP+ bone marrow-derived cells by direct fluorescence, protein immunohistochemistry and immunogold transmission electron microscopy. In both the intact and injured muscles, GFP positivity was determined in immune cells, mainly in macrophages, and in interstitial spindleshaped cells. Moreover, in the injured muscles, rare GFP+ endothelial cells of the blood vessels and newly formed myotubes and muscle fibres were present. Our results confirmed the ability of bone marrowderived cells to contribute to the cellular component of the satellite cell niche in the intact and regenerating skeletal muscle. These cells originated not only from haematopoietic stem cells, but obviously also from other stem or progenitor cells residing in the bone marrow, such as multipotent mesenchymal stromal cells and endothelial progenitors.


Asunto(s)
Células de la Médula Ósea/citología , Regeneración/fisiología , Células Satélite del Músculo Esquelético/citología , Nicho de Células Madre , Animales , Fluorescencia , Proteínas Fluorescentes Verdes/metabolismo , Ratones Endogámicos C57BL , Músculo Esquelético/lesiones , Músculo Esquelético/patología , Músculo Esquelético/ultraestructura
3.
Adv Exp Med Biol ; 944: 19-25, 2017.
Artículo en Inglés | MEDLINE | ID: mdl-27826888

RESUMEN

Non-tuberculous mycobacteria (NTM) are species other than those belonging to the Mycobacterium tuberculosis complex and do not cause leprosy. NTM are generally free-living organisms that are ubiquitous in the environment. There have been more than 140 NTM species identified to-date. They can cause a wide range of infections, with pulmonary infections being the most frequent (65-90 %). There is growing evidence that the incidence of NTM lung diseases and associated hospitalizations are on the rise, mainly in regions with a low prevalence of tuberculosis. A crucial clinical problem remains the evaluation of NTM significance in relation to the disease, especially in regard to the colonization of the respiratory tract in patients with residual lesions after tuberculosis or bronchiectasis. Clinical and radiographic pictures of mycobacteriosis, as well as therapy, have often similarities to those of tuberculosis. The treatment regimen should be individualized. In addition to antituberculotics, antibiotics are used more frequently. The most common mycobacteria causing lung disease in Slovakia are Mycobacterium avium and Mycobacterium abscessus.


Asunto(s)
Infecciones por Mycobacterium no Tuberculosas/diagnóstico , Infecciones por Mycobacterium no Tuberculosas/tratamiento farmacológico , Micobacterias no Tuberculosas/efectos de los fármacos , Antibacterianos/uso terapéutico , Antituberculosos/uso terapéutico , Humanos , Incidencia , Mycobacterium/efectos de los fármacos , Infecciones por Mycobacterium no Tuberculosas/epidemiología , Infecciones por Mycobacterium no Tuberculosas/microbiología , Prevalencia , Infecciones del Sistema Respiratorio/diagnóstico , Infecciones del Sistema Respiratorio/tratamiento farmacológico , Infecciones del Sistema Respiratorio/epidemiología , Infecciones del Sistema Respiratorio/microbiología , Eslovaquia/epidemiología
4.
Adv Exp Med Biol ; 905: 87-95, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-26987338

RESUMEN

The guinea pig sensitized by ovalbumin is the most widely used model to study cough experimentally, as the neurophysiology of the vagus nerve in the guinea pig is closest to humans. Nonetheless, the choice of the antigen remains questionable, which influences the translation of results into clinical medicine. The present study seeks to develop an alternative model of cough study using house dust mite sensitization (HDM). Thirty guinea pigs were divided into the HDM group, ovalbumin (OVA) group, and control group based on their cough response to 0.4 M citric acid. In the HDM group animals were sensitized by 0.25 %HDM aerosol, which they inhaled for 5 min over 5 days, followed by inhalation of 0.5 %HDM in the same protocol. Sensitization was confirmed by a skin test. Symptoms of allergic rhinitis were induced by intranasal application of 15 µl 0.5 %HDM and cough challenges with citric acid were performed. Airway resistance was measured in vivo by Pennock's method. We found that both HDM and OVA-sensitized groups showed a significantly enhanced nasal reactivity and cough response compared with controls. The airway resistance data did not show significant differences. We conclude that the HDM cough model replicates functional aspects of the OVA model, which may make it an alternative to the latter. However, the superiority of the HDM model for experimental cough studies remains to be further explored.


Asunto(s)
Tos/inmunología , Modelos Animales de Enfermedad , Cobayas , Inmunización/métodos , Ovalbúmina/inmunología , Pyroglyphidae/inmunología , Hipersensibilidad Respiratoria/inmunología , Resistencia de las Vías Respiratorias/inmunología , Animales , Ácido Cítrico , Masculino , Pruebas Cutáneas
5.
Adv Exp Med Biol ; 921: 61-70, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-27130219

RESUMEN

Phosphodiesterases (PDEs) are enzymes involved in the degradation of cAMP and cGMP. Selective PDE4 inhibitors (e.g., roflumilast) are effective in therapy of chronic obstructive pulmonary disease associated with neutrophil inflammation. The aim of this study was to evaluate the effects of a selective PDE4 inhibitor, YM976, on citric acid-induced cough, in vivo and in vitro airway smooth muscle reactivity to histamine, and on inflammatory mediators in ovalbumin-sensitized guinea pigs, with experimentally induced eosinophil inflammation. The YM976 was administered intraperitoneally at a dose of 1.0 mg/kg once daily for 7 days. Sensitization with ovalbumin led to a significant increase in the number of coughs, and in vivo and in vitro airway reactivity. Also, increased plasma levels of IL-4, IL-5, and PAF were observed, with a significant increase in the differential count of eosinophils in both blood and bronchoalveolar lavage fluid. The YM976 suppressed the number of coughs, the airway reactivity in tracheal tissue strips, and the IL-4 level. The findings indicate that PDE4 inhibition by YM976 exerts antitussive and anti-inflammatory effects in guinea pigs with ovalbumin-induced eosinophilic inflammation.


Asunto(s)
Hiperreactividad Bronquial/tratamiento farmacológico , Tos/tratamiento farmacológico , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/química , Ovalbúmina/toxicidad , Inhibidores de Fosfodiesterasa 4/farmacología , Piridinas/farmacología , Pirimidinonas/farmacología , Sistema Respiratorio/efectos de los fármacos , Animales , Hiperreactividad Bronquial/inducido químicamente , Hiperreactividad Bronquial/patología , Tos/inducido químicamente , Tos/patología , Cobayas , Masculino
6.
Adv Exp Med Biol ; 935: 53-62, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-27334733

RESUMEN

Phosphodiesterases (PDEs) represent a super-family of 11 enzymes hydrolyzing cyclic nucleotides into inactive 5' monophosphates. Inhibition of PDEs leads to a variety of cellular effects, including airway smooth muscle relaxation, inhibition of cellular inflammation, and immune responses. In this study we focused on theophylline, a known non-selective inhibitor of PDEs. Theophylline has been used for decades in the treatment of chronic inflammatory airway diseases. It has a narrow therapeutic window and belongs to the drugs whose plasma concentration should be monitored. Therefore, the main goal of this study was to evaluate the plasma theophylline concentration and to determine its relevance to pharmacological effects after single and longer term (7 days) administration of theophylline at different doses (5, 10, 20, and 50 mg/kg) in guinea pigs. Airway hyperresponsiveness was assessed by repeated exposure to ovalbumin. Theophylline reduced specific airway resistance in response to histamine nebulization, measured in a double chamber body plethysmograph. A decrease in tracheal smooth muscle contractility after cumulative doses of histamine and acetylcholine was confirmed in vitro. A greater efficacy of theophylline after seven days long treatment indicates the predominance of its anti-inflammatory activity, which may be involved in the bronchodilating action.


Asunto(s)
Resistencia de las Vías Respiratorias/efectos de los fármacos , Antiinflamatorios/farmacología , Hiperreactividad Bronquial/tratamiento farmacológico , Broncodilatadores/farmacología , Inflamación/tratamiento farmacológico , Teofilina/farmacología , Animales , Hiperreactividad Bronquial/inducido químicamente , Hiperreactividad Bronquial/patología , Broncoconstricción/efectos de los fármacos , Cobayas , Inflamación/inducido químicamente , Inflamación/patología , Masculino , Contracción Muscular/efectos de los fármacos , Ovalbúmina/toxicidad , Pletismografía Total
7.
Folia Biol (Praha) ; 62(5): 203-206, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-27978415

RESUMEN

Ensuring mycoplasma-free cell culture is of prime importance as they severely affect cellular characteristics leading to experimental artefacts and spurious results. Various methods persist for mycoplasma detection; out of the whole array of methods polymerase chain reaction (PCR) is the most favoured one because it is highly sensitive, specific and quick. The PCR-based detection procedure involves three steps: cell culture supernatant collection, DNA isolation, and PCR. We have modified this procedure so that cell culture supernatant can directly be used for PCR without the need for DNA extraction. This modification makes the procedure quicker and more sensitive because loss of mycoplasma DNA is prevented and this loss becomes more significant when the level of mycoplasma contamination is very low.


Asunto(s)
Contaminación de ADN , Mycoplasma/genética , Mycoplasma/aislamiento & purificación , Reacción en Cadena de la Polimerasa/métodos , Animales , Línea Celular , Humanos , Ratones
8.
Adv Exp Med Biol ; 835: 15-22, 2015.
Artículo en Inglés | MEDLINE | ID: mdl-25252893

RESUMEN

Tuberculosis currently belongs to rare respiratory diseases in Slovakia. However, the emergence and spread of multi-drug resistant tuberculosis (MDR-TB) and extensively drug-resistant tuberculosis (XDR-TB) are major challenges for global tuberculosis control, since the treatment of resistant forms creates both medical and financial problems. Cultivation methods of diagnosis are time-consuming, many times exceeding the time of the initial phase of tuberculosis treatment. Therefore, in the presented study we compared the standard procedures, based on the cultivation of mycobacteria and subsequent drug susceptibility testing to antituberculotics, with molecular-genetic methods using PCR diagnostic kits. The molecular-genetic testing enables to obtain direct and fast evidence of Mycobacterium tuberculosis, with genomic verification of resistance to the most important anti-tuberculosis drugs - isoniazid and rifampicin in MDR-TB, and ethambutol, aminoglycosides, and fluoroquinolones in XDR-TB. In 2012-2013, we confirmed 19 cases of drug-resistant tuberculosis in Slovakia. The resistance to rifampicin was confirmed in all strains with both methods. In two cases, the molecular-genetic testing did not show resistance to isoniazid, as confirmed by conventional cultivation. Furthermore, two strains demonstrating susceptibility in conventional microbiological testing to ethambutol and five strains to fluoroquinolones were verified as actually being resistant using a PCR method. Rapid diagnosis and identification of MDR-TB or XDR-TB strains using molecular-genetic testing is an essential tool for the timely and appropriate drug treatment and prevention of spread of drug resistant strains.


Asunto(s)
Antituberculosos/farmacología , ADN Bacteriano/genética , Farmacorresistencia Bacteriana Múltiple/genética , Tuberculosis Extensivamente Resistente a Drogas/diagnóstico , Mycobacterium tuberculosis/efectos de los fármacos , Tuberculosis Pulmonar/diagnóstico , Aminoglicósidos/farmacología , Medios de Cultivo , Etambutol/farmacología , Tuberculosis Extensivamente Resistente a Drogas/tratamiento farmacológico , Tuberculosis Extensivamente Resistente a Drogas/epidemiología , Tuberculosis Extensivamente Resistente a Drogas/microbiología , Fluoroquinolonas/farmacología , Genotipo , Humanos , Isoniazida/farmacología , Pruebas de Sensibilidad Microbiana , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/crecimiento & desarrollo , Reacción en Cadena de la Polimerasa , Rifampin/farmacología , Eslovaquia/epidemiología , Tuberculosis Pulmonar/tratamiento farmacológico , Tuberculosis Pulmonar/epidemiología , Tuberculosis Pulmonar/microbiología
9.
Adv Exp Med Biol ; 838: 11-8, 2015.
Artículo en Inglés | MEDLINE | ID: mdl-25310957

RESUMEN

Chronic inflammatory diseases, associated with airway obstruction and cough, are usually treated with bronchodilating and anti-inflammatory drugs. Inhibition of phosphodiesterases (PDE) leads to both of these effects and influences apoptosis of immune cells. In chronic obstructive pulmonary disease, roflumilast, a selective PDE4 inhibitor, has been recently approved for pharmacotherapy. The aim of this study was to evaluate the effect of long-term administration of roflumilast in experimental allergic inflammation in guinea pigs. Male adult guinea pigs were used in the study. There were four experimental groups sensitized with ovalbumin for 14 days and thereafter treated per os, by inhalation, and intraperitoneally for 7 days with roflumilast or vehicle. A control group was left without sensitization. Roflumilast reduced specific airway resistance after nebulization of histamine, as measured in a double-chamber whole-body plethysmograph. This effect was confirmed in in vitro organ bath, with significant decreases in tracheal and lung smooth muscle contractility after cumulative doses of histamine. Suppression of hematological and immunological markers of inflammation and enhanced apoptosis in animals treated with roflumilast points to the possibility of a beneficial effect of roflumilast in allergic inflammation.


Asunto(s)
Resistencia de las Vías Respiratorias/efectos de los fármacos , Aminopiridinas/farmacología , Benzamidas/farmacología , Hiperreactividad Bronquial/tratamiento farmacológico , Pulmón/efectos de los fármacos , Inhibidores de Fosfodiesterasa 4/farmacología , Administración por Inhalación , Administración Oral , Animales , Apoptosis/efectos de los fármacos , Hiperreactividad Bronquial/inducido químicamente , Hiperreactividad Bronquial/inmunología , Hiperreactividad Bronquial/fisiopatología , Líquido del Lavado Bronquioalveolar/citología , Líquido del Lavado Bronquioalveolar/inmunología , Ciclopropanos/farmacología , Cobayas , Histamina/farmacología , Leucocitos Mononucleares/efectos de los fármacos , Leucocitos Mononucleares/inmunología , Leucocitos Mononucleares/patología , Pulmón/inmunología , Pulmón/fisiopatología , Masculino , Contracción Muscular/efectos de los fármacos , Músculo Liso/efectos de los fármacos , Ovalbúmina , Pletismografía Total , Cultivo Primario de Células , Técnicas de Cultivo de Tejidos
10.
Adv Exp Med Biol ; 838: 1-10, 2015.
Artículo en Inglés | MEDLINE | ID: mdl-25252904

RESUMEN

Nitric oxide (NO) effects in airways are influenced by the activity of NO-synthase isoforms and NO metabolism. Inducible NO-synthase (iNOS), which produces large amounts of NO, is active during the inflammatory process. NO quickly reacts, producing reactive oxygen species (ROS). In this study we attempted to detect the expression of iNOS and markers of ROS in the airway hyperreactivity (AHR) condition. The study was performed in guinea pigs, divided into four groups. Two groups were treated with the non-selective inhibitor of NO-synthase L-NAME. The other two groups were used as controls. Exhaled NO was monitored in vivo, AHR was assessed both in vivo and in vitro, and the expression of iNOS in lung homogenate, and oxidative stress markers were measured in the venous blood. L-NAME significantly affected the AHR only in in vitro condition, blocked the expression of iNOS in control but not in sensitized animals, and decreased the level of exhaled NO. The results concerning the oxidative stress markers are equivocal. The study confirmed that NO is involved in the regulation of AHR; the effects being mediated via iNOS and ROS activity.


Asunto(s)
Hiperreactividad Bronquial/metabolismo , Inhibidores Enzimáticos/farmacología , NG-Nitroarginina Metil Éster/farmacología , Óxido Nítrico Sintasa de Tipo II/genética , Óxido Nítrico/biosíntesis , Animales , Hiperreactividad Bronquial/inducido químicamente , Hiperreactividad Bronquial/tratamiento farmacológico , Hiperreactividad Bronquial/fisiopatología , Espiración , Expresión Génica/efectos de los fármacos , Cobayas , Pulmón/efectos de los fármacos , Pulmón/metabolismo , Pulmón/fisiopatología , Masculino , Óxido Nítrico/metabolismo , Óxido Nítrico Sintasa de Tipo II/antagonistas & inhibidores , Óxido Nítrico Sintasa de Tipo II/metabolismo , Ovalbúmina , Estrés Oxidativo , Especies Reactivas de Oxígeno/metabolismo
11.
Adv Exp Med Biol ; 832: 59-67, 2015.
Artículo en Inglés | MEDLINE | ID: mdl-25300685

RESUMEN

Meconium aspiration in newborns causes lung inflammation and injury, which may lead to meconium aspiration syndrome (MAS). In this study, the effect of the antioxidant N-acetylcysteine on respiratory and inflammatory parameters were studied in a model of MAS. Oxygen-ventilated rabbits were intratracheally given 4 mL/kg of meconium (25 mg/mL) or saline. Thirty minutes later, meconium-instilled animals were administered N-acetylcysteine (10 mg/kg; i.v.), or were left without treatment. The animals were oxygen-ventilated for additional 5 h. Ventilatory pressures, oxygenation, right-to-left pulmonary shunts, and leukocyte count were measured. At the end of experiment, trachea and lung were excised. The left lung was saline-lavaged and a total and differential count of cells in bronchoalveolar lavage fluid (BAL) was determined. Right lung tissue strips were used for detection of lung edema (expressed as wet/dry weight ratio) and peroxidation (expressed by thiobarbituric acid-reactive substances, TBARS). In lung and tracheal strips, airway reactivity to acetylcholine was measured. In addition, TBARS and total antioxidant status were determined in the plasma. Meconium instillation induced polymorphonuclear-derived inflammation and oxidative stress. N-acetylcysteine improved oxygenation, reduced lung edema, decreased polymorphonuclears in BAL fluid, and diminished peroxidation and meconium-induced airway hyperreactivity compared with untreated animals. In conclusion, N-acetylcysteine effectively improved lung functions in an animal model of MAS.


Asunto(s)
Acetilcisteína/farmacología , Lesión Pulmonar Aguda/tratamiento farmacológico , Antiinflamatorios/farmacología , Antioxidantes/farmacología , Síndrome de Aspiración de Meconio/tratamiento farmacológico , Edema Pulmonar/tratamiento farmacológico , Lesión Pulmonar Aguda/inducido químicamente , Lesión Pulmonar Aguda/metabolismo , Lesión Pulmonar Aguda/fisiopatología , Animales , Líquido del Lavado Bronquioalveolar/química , Líquido del Lavado Bronquioalveolar/citología , Modelos Animales de Enfermedad , Humanos , Recién Nacido , Inyecciones Intravenosas , Intubación Intratraqueal , Recuento de Leucocitos , Peroxidación de Lípido/efectos de los fármacos , Pulmón/efectos de los fármacos , Pulmón/fisiopatología , Meconio , Síndrome de Aspiración de Meconio/metabolismo , Síndrome de Aspiración de Meconio/fisiopatología , Estrés Oxidativo , Edema Pulmonar/inducido químicamente , Edema Pulmonar/metabolismo , Edema Pulmonar/fisiopatología , Conejos , Respiración Artificial , Sustancias Reactivas al Ácido Tiobarbitúrico/metabolismo , Tráquea/efectos de los fármacos , Tráquea/fisiopatología
12.
Adv Exp Med Biol ; 832: 35-43, 2015.
Artículo en Inglés | MEDLINE | ID: mdl-25300682

RESUMEN

Anti-inflammatory drugs are increasingly used for treatment of neonatal meconium aspiration syndrome (MAS), but their adverse effects are poorly known. Therefore, the aim of this study was to evaluate the effects of the antioxidant N-acetylcysteine on cardiovascular parameters in an animal model of MAS. Oxygen-ventilated rabbits were intratracheally instilled 4 mL/kg of meconium suspension (25 mg/mL) or saline. Thirty minutes later, meconium-instilled animals were given N-acetylcysteine (10 mg/kg, i.v.) or the same volume of saline. Changes in cardiovascular parameters (blood pressure, heart rate, and heart rate variability) were recorded over a 5-min course of solution administration, over 5 min after its end, and then hourly for 5 h. Oxidation markers (thiobarbituric acid-reactive substances (TBARS) and total antioxidant status) and aldosterone, as a non-specific marker of cardiovascular injury, were determined in plasma. Meconium instillation did not evoke any significant cardiovascular changes, but induced oxidative stress and elevated plasma aldosterone. N-acetylcysteine significantly reduced the mentioned markers of injury. However, its administration was associated with short-term increases in blood pressure and in several parameters of heart rate variability. Considering these effects of N-acetylcysteine, its intravenous administration in newborns with MAS should be carefully monitored.


Asunto(s)
Acetilcisteína/farmacología , Lesión Pulmonar Aguda/tratamiento farmacológico , Antiinflamatorios/farmacología , Antioxidantes/farmacología , Síndrome de Aspiración de Meconio/tratamiento farmacológico , Lesión Pulmonar Aguda/sangre , Lesión Pulmonar Aguda/inducido químicamente , Lesión Pulmonar Aguda/fisiopatología , Aldosterona/sangre , Animales , Presión Sanguínea/efectos de los fármacos , Modelos Animales de Enfermedad , Frecuencia Cardíaca/efectos de los fármacos , Humanos , Recién Nacido , Inyecciones Intravenosas , Intubación Intratraqueal , Pulmón/efectos de los fármacos , Pulmón/fisiopatología , Meconio , Síndrome de Aspiración de Meconio/sangre , Síndrome de Aspiración de Meconio/fisiopatología , Estrés Oxidativo , Conejos , Respiración Artificial , Sustancias Reactivas al Ácido Tiobarbitúrico/metabolismo
13.
Neoplasma ; 61(3): 305-8, 2014.
Artículo en Inglés | MEDLINE | ID: mdl-24824932

RESUMEN

The primary cilium is a solitary, sensory, non-motile microtubule-based structure that arises from the centrosome and is projected from the surface of most human cells. The objective of the current pilot study was to conduct an investigation of presence and frequency of cilia in gastrointestinal stromal tumors (GIST).The presence of primary cilia in GIST was evaluated in 9 patients, including 8 primary tumors and 1 liver metastasis. In 2 patients the presence of primary cilia was evaluated not only in the primary tumor, but also in recurrence: in 1 patient in recurrence without previous treatment with imatinib and in 1 patient in recurrence after treatment with imatinib. The primary cilia of GIST cells were immunofluorescently stained with primary monoclonal anti-acetylated tubulin alpha antibody and cell nuclei with DAPI.We observed 9985 nuclei of cells of GISTs and 425 primary cilia in total. The median of frequency of primary cilia in cells of GISTs in all examined samples was 4.26%, in primary tumors was 4.32% and in metastases was 3.64%, respectively. This pilot study provides the evidence of the presence of primary cilia in GISTs in different organs. Primary cilia were identified in all examined cases of GIST, including primary tumors, metastases and recurrent lesions without and with previous treatment with imatinib.


Asunto(s)
Cilios/patología , Neoplasias Gastrointestinales/patología , Tumores del Estroma Gastrointestinal/patología , Adulto , Anciano , Femenino , Humanos , Masculino , Persona de Mediana Edad
14.
Adv Exp Med Biol ; 756: 341-7, 2013.
Artículo en Inglés | MEDLINE | ID: mdl-22836652

RESUMEN

As inflammation plays an important role in the pathogenesis of neonatal meconium aspiration syndrome (MAS), anti-inflammatory agents including inhibitors of phosphodiesterases (PDE) are increasingly used in the treatment. To evaluate side effects of PDE inhibitors, this study analyzed changes in blood pressure, heart rate (HR) and heart rate variability (HRV) during and after intravenous aminophylline in the animal model of MAS. Oxygen-ventilated rabbits were given meconium intratracheally (25 mg/ml, 4 ml/kg) or saline. Thirty minutes later, the animals were treated by intravenous aminophylline (Syntophyllin, 2 mg/kg) or saline (sham-treated controls). A second dose of the treatment was given 2 h later. During (5 min) and immediately after (5 min) the treatment, and during 5 h after the treatment, mean blood pressure in the femoral artery (MAP), HR and HRV were evaluated. In meconium-instilled animals, increases in MABP, HR, and HRV were observed already 5 min after aminophylline administration, while in saline-instilled animals aminophylline increased HR and caused inconsistant changes in HRV parameters compared to sham-treated animals. Within 5 h after the treatment administration, MAP, HR, and HRV parameters gradually returned to the initial values. Concluding, intravenous aminophylline may lead to acute cardiovascular changes. Thus, if aminophylline is used for treatment of MAS, its possible cardiovascular effects should be considered, particularly in patients with cardiovascular instability.


Asunto(s)
Aminofilina/toxicidad , Broncodilatadores/farmacología , Broncodilatadores/toxicidad , Sistema Cardiovascular/efectos de los fármacos , Síndrome de Aspiración de Meconio/tratamiento farmacológico , Aminofilina/uso terapéutico , Animales , Presión Sanguínea/efectos de los fármacos , Broncodilatadores/uso terapéutico , Modelos Animales de Enfermedad , Frecuencia Cardíaca/efectos de los fármacos , Humanos , Recién Nacido , Pulmón/efectos de los fármacos , Meconio , Inhibidores de Fosfodiesterasa/uso terapéutico , Inhibidores de Fosfodiesterasa/toxicidad , Conejos
15.
Folia Biol (Praha) ; 59(5): 188-97, 2013.
Artículo en Inglés | MEDLINE | ID: mdl-24280141

RESUMEN

Head and neck cancer is one of the most common cancers in Europe. Many current anti-cancer treatments, including ionizing radiation, induce apoptosis via DNA damage. Unfortunately, such treatments are non-selective to cancer cells and produce similar toxicity in normal cells, including adult stem cells. One of the fundamental properties of an adult stem cell is that it does not have any tissue-specific structures that allow it to perform specialized functions. However, under certain stimuli, unspecialized adult stem cells can give rise to specialized cells to generate replacements for cells that are lost during one's life or due to injury or disease. Nevertheless, specialization of stem cells must be controlled by specific milieu and also initiated at the proper time, making the entire process beneficial for tissue recovery and maintaining it for a long time. In this paper we assess whether irradiated dental pulp stem cells have maintained open their options to mature into specialized cells, or whether they have lost their unspecialized (immature) state following irradiation. Our findings showed radiation-induced premature differentiation of dental pulp stem cells towards odonto-/osteoblast lineages in vitro. Matrix calcification was visualized from Day 6 or Day 9 following irradiation of cells expressing low or high levels of CD146, respectively.


Asunto(s)
Diferenciación Celular/efectos de la radiación , Senescencia Celular/efectos de la radiación , Pulpa Dental/citología , Radiación Ionizante , Células Madre/citología , Células Madre/efectos de la radiación , Antígeno CD146/metabolismo , Recuento de Células , Proliferación Celular/efectos de la radiación , Supervivencia Celular/efectos de la radiación , Células Cultivadas , Humanos , Cinética , Osteogénesis/efectos de la radiación , Factores de Tiempo
16.
Int Endod J ; 45(5): 401-12, 2012 May.
Artículo en Inglés | MEDLINE | ID: mdl-22142405

RESUMEN

AIM: To determine the response of dental pulp stem cells (DPSCs) to DNA-damaging cytostatic cisplatin and compare it with the response of normal human dermal fibroblasts (HDFs). METHODOLOGY: Dental pulp stem cells were exposed to 5, 10, 20 or 40 µmol L(-1) of cisplatin. The proliferation of affected cells was assessed by a Z2 Counter and viability was assessed by means of a Vi-Cell XR using Trypan blue exclusion staining. Cell cycle analysis and induction of apoptosis were performed by flow cytometry. Induction of apoptosis was determined by monitoring the activities of caspases. The expression of proteins was detected by electrophoresis and Western blotting. The descriptive statistics of the results was analyzed by Student's t-test. RESULTS: Dental pulp stem cells had a greater genotoxic stress response to cisplatin compared to HDFs. All three main Mitogen-activated protein kinases (MAPK) families - extracellular signal-regulated kinases (ERK), c-Jun-N-terminal kinase (JNK) and p38 were activated after treatment of DPSCs with cisplatin. The activation of MAPK pathways was not observed in HDFs exposed to cisplatin. The exposure of DPSCs and HDFs to cisplatin provoked an increase in p53 and p21 expression and p53 phosphorylation of serine 15. Higher concentrations of cisplatin reduced the viability of DPSCs and HDFs and induced the activation of caspases 3/7 and 9. CONCLUSION: Dental pulp stem cells had a greater genotoxic stress response to cisplatin compared to HDFs. Cisplatin in higher concentrations triggered activation of MAPK and apoptosis in DPSCs but not in HDFs.


Asunto(s)
Cisplatino/toxicidad , Citostáticos/toxicidad , Pulpa Dental/citología , Ectodermo/citología , Células Madre Mesenquimatosas/efectos de los fármacos , Apoptosis/efectos de los fármacos , Caspasa 3/efectos de los fármacos , Caspasa 7/efectos de los fármacos , Caspasa 8/efectos de los fármacos , Caspasa 9/efectos de los fármacos , Técnicas de Cultivo de Célula , Ciclo Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/efectos de los fármacos , Pulpa Dental/efectos de los fármacos , Ectodermo/efectos de los fármacos , Activación Enzimática/efectos de los fármacos , Quinasas MAP Reguladas por Señal Extracelular/efectos de los fármacos , Fibroblastos/efectos de los fármacos , Humanos , Proteínas Quinasas JNK Activadas por Mitógenos/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Mutágenos/toxicidad , Fosforilación , Inhibidores de Proteínas Quinasas/farmacología , Serina/efectos de los fármacos , Piel/citología , Piel/efectos de los fármacos , Proteína p53 Supresora de Tumor/efectos de los fármacos , Proteínas Quinasas p38 Activadas por Mitógenos/efectos de los fármacos
17.
Vnitr Lek ; 58(12): 938-42, 2012 Dec.
Artículo en Checo | MEDLINE | ID: mdl-23427952

RESUMEN

The primary cilium is a mechanosensory, solitary, non-motile microtubule-based structure that in the quiescent phase of the cell cycle projects from the surface of the majority of human cells, including embryonal, stem and mesenchymal cells, fibroblasts, myoblasts, cardiomyocytes, vascular smooth muscle and endothelial cells. Primary cilia are in increased frequency also present on the surface of endothelial cells in atherosclerotic predilection sites, lipoid streaks and dots and atheromatous plaques. The primary cilium is formed from the mother centriole. Primary cilia are currently studied in mechanobiology of cardiovascular apparatus and their role in cell migration, cell cycle control and atherogenesis. The aim of this paper is to provide a review of the current knowledge on the primary cilia of cells of cardiovascular apparatus.


Asunto(s)
Cilios/fisiología , Células Endoteliales/ultraestructura , Músculo Liso Vascular/ultraestructura , Miocitos Cardíacos/ultraestructura , Cilios/ultraestructura , Células Endoteliales/fisiología , Humanos , Músculo Liso Vascular/fisiología , Miocitos Cardíacos/fisiología
18.
Folia Biol (Praha) ; 57(6): 232-41, 2011.
Artículo en Inglés | MEDLINE | ID: mdl-22264717

RESUMEN

The recently discovered capacity of bone marrow cells (BMCs) to contribute to injury-induced skeletal muscle regeneration has brought new possibilities in the treatment of skeletal muscle diseases. However, a suitable method of BMC transplantation usable for such therapy has to be established. In this work, recipient mice were intramuscularly injected with cardiotoxin, then whole-body lethally irradiated to eradicate satellite cells in their injured tibialis anterior (TA) muscles and to suppress haematopoiesis, and subsequently intravenously transplanted with lacZ+ BMCs with the aim to investigate the role of exogenous BMCs in response to skeletal muscle injury. Seven to 33 days after grafting, recipient TA muscles were examined to detect donor-derived X-gal+ cells and analysed by quantitative PCR. In injured recipients' muscles, X-gal positivity was identified 14 and 33 days after grafting in some infiltrating neutrophils and macrophages, infrequently in fibroblasts of endomysium, and in many large multinucleated cells (devoid of myogenic markers desmin and nestin) resembling foreign body giant cells situated in the vicinity of necrotic muscle fibres. qPCR confirmed the presence of transplanted lacZ+ BMCs in injured recipients' muscles. Our results proved the ability of intravenously transplanted adult BMCs to settle in injured muscles and generate blood cells that infiltrated endomysium and took part in the cleaning reaction. After inhibition of endogenous myogenesis, BMCs were not able to participate in formation of new muscle fibres due to persisting necrosis of degenerated muscle fibres. Instead, BMCs attempted to resorb necrotic structures, which confirmed the indispensable role of bone marrow-derived macrophages in skeletal muscle regeneration.


Asunto(s)
Células de la Médula Ósea/citología , Trasplante de Médula Ósea , Músculo Esquelético/lesiones , Músculo Esquelético/patología , Animales , Células de la Médula Ósea/metabolismo , ADN/metabolismo , Glucuronidasa/metabolismo , Inmunohistoquímica , Ratones , Ratones Endogámicos C57BL , Irradiación Corporal Total
19.
Bratisl Lek Listy ; 112(3): 131-5, 2011.
Artículo en Inglés | MEDLINE | ID: mdl-21452764

RESUMEN

In this study the effects of non-selective PDE inhibitors (theophylline and theobromine) and selective inhibitors of PDE 1, 3, 4 and 5 on cough, induced by citric acid, were evaluated. Inhalation of citric acid aerosol was used for cough provocation in healthy and ovalbumin-sensitized guinea pigs and the number of cough efforts was registered after visual and acoustic control by a skilled observer, with subsequent evaluation of airflow changes in a double chamber whole body plethysmograph. The pre-treatment with theophylline and theobromine (10 mg/kg b.w. intraperitoneally) decreased the number of cough efforts evoked by inhalation of citric acid aerosol (0.6 mol/l) in both healthy and ovalbumin-sensitized animals. The selective inhibitors (all 1 mg/kg b.w. intraperitoneally) of PDE1 (vinpocetin), PDE3 (cilostazol), and PDE4 (citalopram) showed antitussive effects in healthy guinea pigs. Conversely, the antitussive potential of PDE1 (vinpocetin), PDE4 (citalopram), and PDE5 (zaprinast) was observed in ovalbumin-sensitized animals. We conclude that the administration of non-selective PDE inhibitors influenced the citric acid-induced cough both in healthy and ovalbumin-sensitized guinea pigs, indicating the participation of a bronchodilating action and suppression of airway hyperreactivity in the cough suppression. With selective inhibitors, PDE4 inhibition seems to be the most effective in cough suppression, confirming its positive effects tested in chronic airway inflammatory diseases associated with bronchoconstriction and cough (Fig. 6, Ref. 27).


Asunto(s)
Antitusígenos/farmacología , Tos/fisiopatología , Inhibidores de Fosfodiesterasa/farmacología , Reflejo/efectos de los fármacos , Teobromina/farmacología , Teofilina/farmacología , Animales , Hiperreactividad Bronquial/inmunología , Hiperreactividad Bronquial/fisiopatología , Ácido Cítrico , Tos/inducido químicamente , Cobayas , Masculino , Ovalbúmina/inmunología
20.
Physiol Res ; 69(Suppl 3): S523-S532, 2020 12 31.
Artículo en Inglés | MEDLINE | ID: mdl-33476174

RESUMEN

Mental disorders affect 10-20 % of the young population in the world. Major depressive disorder (MDD) is a common mental disease with a multifactorial and not clearly explained pathophysiology. Many cases remain undetected and untreated, which influences patients' physical and mental health and their quality of life also in adulthood. The aim of our pilot study was to assess the prediction value of selected potential biomarkers, including blood cell counts, blood cell ratios, and parameters like peroxiredoxin 1 (PRDX1), tenascin C (TNC) and type IV collagen (COL4) between depressive pediatric patients and healthy peers and to evaluate a short effect of antidepressant treatment. In this study, 27 young depressive patients and 26 non-depressed age-matched controls were included. Blood analyses and immunological assays using commercial kits were performed. Platelet count was the only blood parameter for which the case/control status was statistically significant (p=0.01) in a regression model controlling for the age and gender differences. The results from ELISA analyses showed that the case/control status is a significant predictor of the parameters PRDX1 (p=0.05) and COL4 (p=0.009) in respective regression model considering the age and gender differences between MDD patients and controls. A major finding of this study is that values of platelet count, monocyte to lymphocyte ratio, white blood cell, and monocyte counts were assessed by the Random Forest machine learning algorithm as relevant predictors for discrimination between MDD patients and healthy controls with a power of prediction AUC=0.749.


Asunto(s)
Trastorno Depresivo Mayor/epidemiología , Adolescente , Biomarcadores/análisis , Estudios de Casos y Controles , Trastorno Depresivo Mayor/diagnóstico , Trastorno Depresivo Mayor/psicología , Femenino , Humanos , Masculino , Proyectos Piloto , Calidad de Vida , Eslovaquia/epidemiología
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