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 TiempoRESUMEN
The aim of our work was to evaluate mechanisms leading to radiosensitization of MOLT-4 leukemia cells following valproic acid (VA) treatment. Cells were pretreated with 2 mM VA for 24 h followed by irradiation with a dose of 0.5 or 1 Gy. The effect of both noxae, alone and combined, was detected 1 and 24 hours after the irradiation. Induction of apoptosis was evaluated by a flow cytometry. The extent of DNA damage was further determined by phosphorylation of histone H2AX using confocal microscopy. Changes in protein expression were identified by SDS-PAGE/immunoblotting. Two-millimolar VA increased apoptosis induction after irradiation as well as phosphorylation of H2AX and provokes an increase in the level of p53 and its phosphorylation at Ser392 in 4 h post-irradiation. Likewise, p21 protein reached its maximal expression in 4 h after the irradiation of VA-treated cells. Twenty four hours later, only the p53 phosphorylated at Ser15 was detected. At the same time, the protein mdm2 (negative regulator of p53) was maximally activated. The 24-hour treatment of MOLT-4 leukemia cells with 2 mM VA results in radiosensitizing, increases apoptosis induction, H2AX phosphorylation, and also p53 and p21 activation.
Asunto(s)
Inhibidores Enzimáticos/farmacología , Histonas/metabolismo , Fármacos Sensibilizantes a Radiaciones/farmacología , Ácido Valproico/farmacología , Apoptosis/efectos de los fármacos , Apoptosis/efectos de la radiación , Línea Celular Tumoral , Daño del ADN/efectos de los fármacos , Daño del ADN/efectos de la radiación , Reparación del ADN/efectos de los fármacos , Rayos gamma , Regulación de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica/efectos de la radiación , Histonas/genética , Humanos , Leucemia/metabolismo , Fosforilación/efectos de los fármacos , Fosforilación/efectos de la radiación , Proteínas Proto-Oncogénicas c-mdm2/genética , Proteínas Proto-Oncogénicas c-mdm2/metabolismo , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismo , Proteínas de Unión al GTP rho/genética , Proteínas de Unión al GTP rho/metabolismoRESUMEN
The relationship between signal pathways MEK1/2-ERK1/2 and ATM-p53 in the response to DNA damage is not well understood. The aim of our study was to investigate the effect of mitoxantrone and two protein kinase inhibitors - caffeine (inhibitor of ATM kinase) and U0126 (inhibitor of MEK1/2 kinase) - on MOLT-4 and Jurkat leukaemic cell lines. In this work we show that the inhibition of MEK1/2 is associated with an increased mortality of cells after mitoxantrone treatment. Inhibition of ATM by caffeine delayed mitoxantrone-induced cell death in MOLT-4 cells. Mitoxantrone itself induced cell-cycle arrest and accumulation of the cells in late S and G2/M phase. Inhibition of ATM, but not of MEK1/2, abrogated mitoxantrone-induced cell-cycle arrest. Inhibition of MEK1/2 did not change mitoxantroneinduced up-regulation of p53 and p21, but inhibition of ATM markedly decreased up-regulation of p53 and p21, and p53 phosphorylation on serine 15 and serine 392. It can be concluded that: 1) mitoxantrone- induced phosphorylation of p53 on serine 15 and serine 392 is ATM dependent and MEK1/2-ERK1/2 independent. 2) ATM inhibition by caffeine prevents G2 cell arrest and in p53-positive cells MOLT-4 delays the onset of mitoxantrone-induced cell death. 3) Inhibition of MEK1/2-ERK1/2 cascade potentiates the cytostatic effect of mitoxantrone regardless of the p53 status.
Asunto(s)
Antineoplásicos/farmacología , Apoptosis , Proteínas de Ciclo Celular/antagonistas & inhibidores , Proteínas de Unión al ADN/antagonistas & inhibidores , Proteína Quinasa 1 Activada por Mitógenos/antagonistas & inhibidores , Proteína Quinasa 3 Activada por Mitógenos/antagonistas & inhibidores , Mitoxantrona/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Supresoras de Tumor/antagonistas & inhibidores , Proteínas de la Ataxia Telangiectasia Mutada , Butadienos/farmacología , Ciclo Celular , Proteínas de Ciclo Celular/genética , Línea Celular Tumoral , Proteínas de Unión al ADN/genética , Fase G2/efectos de los fármacos , Humanos , Células Jurkat , Proteína Quinasa 3 Activada por Mitógenos/genética , Nitrilos/farmacología , Proteínas Serina-Treonina Quinasas/genética , Transducción de Señal , Proteínas Supresoras de Tumor/genéticaRESUMEN
Cellular response to ionizing radiation-induced damage depends on the cell type and the ability to repair DNA damage. Some types of cells undergo apoptosis, whereas others induce a permanent cell cycle arrest and do not proliferate. Our study demonstrates two types of response of embryonic diploid fibroblasts WI-38 to ionizing radiation. In the WI-38 cells p53 is activated, protein p21 increases, but the cells are arrested in G2 phase of cell cycle. Some of the cells die by apoptosis, but in remaining viable cells p16 increases, senescence associated DNA-damage foci occur, and senescence-associated beta-galactosidase activity increases, which indicate stress-induced premature senescence.