RESUMEN
Glioblastoma multiforme (GBM), a malignant brain tumor with a dismal prognosis, shows a high level of chemo- and radioresistance and, therefore, attempts to sensitize glioma cells are highly desired. Here, we addressed the question of whether artesunate (ART), a drug currently used in the treatment of malaria, enhances the killing response of glioblastoma cells to temozolomide (TMZ), which is the first-line therapeutic for GBM. We measured apoptosis, necrosis, autophagy and senescence, and the extent of DNA damage in glioblastoma cells. Further, we determined the tumor growth in nude mice. We show that ART enhances the killing effect of TMZ in glioblastoma cell lines and in glioblastoma stem-like cells. The DNA double-strand break level induced by TMZ was not clearly enhanced in the combined treatment regime. Also, we did not observe an attenuation of TMZ-induced autophagy, which is considered a survival mechanism. However, we observed a significant effect of ART on homologous recombination (HR) with downregulation of RAD51 protein expression and HR activity. Further, we found that ART is able to inhibit senescence induced by TMZ. Since HR and senescence are pro-survival mechanisms, its inhibition by ART appears to be a key node in enhancing the TMZ-induced killing response. Enhancement of the antitumor effect of TMZ by co-administration of ART was also observed in a mouse tumor model. In conclusion, the amelioration of TMZ-induced cell death upon ART co-treatment provides a rational basis for a combination regime of TMZ and ART in glioblastoma therapy.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Neoplasias Encefálicas/patología , Senescencia Celular/efectos de los fármacos , Glioma/patología , Recombinación Homóloga/efectos de los fármacos , Animales , Artemisininas/farmacología , Artesunato , Muerte Celular/efectos de los fármacos , Dacarbazina/análogos & derivados , Dacarbazina/farmacología , Humanos , Ratones , Ratones Desnudos , Temozolomida , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The columnar phenotype of apple trees (Malus x domestica) is characterized by a compact growth habit with fruit spurs instead of lateral branches. These properties provide significant economic advantages by enabling high density plantings. The columnar growth results from the presence of a dominant allele of the gene Columnar (Co) located on chromosome 10 which can appear in a heterozygous (Co/co) or homozygous (Co/Co) state. Although two deep sequencing approaches could shed some light on the transcriptome of columnar shoot apical meristems (SAMs), the molecular mechanisms of columnar growth are not yet elaborated. Since the influence of phytohormones is believed to have a pivotal role in the establishment of the phenotype, we performed RNA-Seq experiments to study genes associated with hormone homeostasis and clearly affected by the presence of Co. Our results provide a molecular explanation for earlier findings on the hormonal state of columnar apple trees. Additionally, they allow hypotheses on how the columnar phenotype might develop. Furthermore, we show a statistically approved enrichment of differentially regulated genes on chromosome 10 in the course of validating RNA-Seq results using additional gene expression studies.
Asunto(s)
Regulación de la Expresión Génica de las Plantas/fisiología , Genes de Plantas/genética , Malus/fisiología , Reguladores del Crecimiento de las Plantas/fisiología , Alelos , Cromosomas de las Plantas/genética , Biología Computacional , Expresión Génica , Secuenciación de Nucleótidos de Alto Rendimiento , Homeostasis , Malus/genética , Malus/crecimiento & desarrollo , Meristema/genética , Meristema/crecimiento & desarrollo , Meristema/fisiología , Modelos Moleculares , Análisis de Secuencia por Matrices de Oligonucleótidos , Fenotipo , Reguladores del Crecimiento de las Plantas/genética , Brotes de la Planta/genética , Brotes de la Planta/crecimiento & desarrollo , Brotes de la Planta/fisiología , ARN de Planta/química , ARN de Planta/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Análisis de Secuencia de ARN , Transcriptoma , ÁrbolesRESUMEN
The differentiation of CD4(+) or CD8(+) T cells following priming of naive cells is central in the establishment of the immune response against pathogens or tumors. However, our understanding of this complex process and the significance of the multiple subsets of differentiation remains controversial. Gene expression profiling has opened new directions of investigation in immunobiology. Nonetheless, the need for substantial amount of biological material often limits its application range. In this study, we have developed procedures to perform microarray analysis on amplified cDNA from low numbers of cells, including primary T lymphocytes, and applied this technology to the study of CD4 and CD8 lineage differentiation. Gene expression profiling was performed on samples of 1000 cells from 10 different subpopulations, defining the major stages of post-thymic CD4(+) or CD8(+) T cell differentiation. Surprisingly, our data revealed that while CD4(+) and CD8(+) T cell gene expression programs diverge at early stages of differentiation, they become increasingly similar as cells reach a late differentiation stage. This suggests that functional heterogeneity between Ag experienced CD4(+) and CD8(+) T cells is more likely to be located early during post-thymic differentiation, and that late stages of differentiation may represent a common end in the development of T-lymphocytes.