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SUMMARY: This study is to investigate the effect of survivin down-regulation by Egr1-survivin shRNA combined with radiotherapy on the apoptosis and radiosensitivity of esophageal squamous cell carcinoma ECA109 and KYSE150 cells. ECA109 and KYSE150 cells were transfected with Egr1-survivin shRNA, and then treated with radiotherapy. After 24 h, the mRNA and protein levels of Egr1-survivin were detected by qPCR and Western-Blot. Cell cycle and apoptosis were detected by flow cytometry. Western blot also detected levels of cleavaged Caspase 3 and Caspase 9. YM155 was used as a positive control to inhibit survivin expression. The levels of survivin mRNA and protein in ECA109 and KYSE150 cells treated with Egr1-survivin shRNA combined with radiotherapy were significantly lower than those of the blank control group, the empty vector control group, and, the YM155 + radiotherapy group (P<0.05). Meanwhile, after survivin down-regulation, the ratio of G2 to S phase of ECA109 and KYSE150 cells increased significantly, leading to significant G2 and S phase arrest. Additionally, apoptosis of ECA109 and KYSE150 cells increased significantly (P <0.01). Further, protein levels of cleavaged Caspase 3 and Caspase 9 significantly increased in Egr1-survivin shRNA combined with radiotherapy group. Egr1-survivin shRNA combined with radiotherapy can down-regulate survivin expression, which further increases the apoptosis, and enhances the radiosensitivity of ECA109 and KYSE150 cells.
Este estudio tuvo como objetivo investigar el efecto de la regulación negativa de survivina por el shRNA de Egr1-survivina combinado con radioterapia sobre la apoptosis y la radiosensibilidad del carcinoma de células escamosas de esófago Células ECA109 y KYSE150. Las células ECA109 y KYSE150 se transfectaron con shRNA de survivina Egr1 y luego se trataron con radioterapia. Después de 24 h, los niveles de ARNm y proteína de Egr1-survivina se detectaron mediante qPCR y Western-Blot. El ciclo celular y la apoptosis se detectaron mediante citometría de flujo. La transferencia Western también detectó niveles de Caspasa 3 y Caspasa 9 escindidas. Se usó YM155 como control positivo para inhibir la expresión de survivina. Los niveles de ARNm y proteína de survivina en células ECA109 y KYSE150 tratadas con shRNA de survivina Egr1 combinado con radioterapia fueron significativamente más bajos que los del grupo control en blanco, el grupo control de vector vacío y el grupo de radioterapia YM155 + (P <0,05). Mientras tanto, después de la regulación negativa de survivina, la proporción entre las fases G2 y S de las células ECA109 y KYSE150 aumentó significativamente, lo que llevó a una detención significativa de las fases G2 y S. Además, la apoptosis de las células ECA109 y KYSE150 aumentó significativamente (P <0,01). Además, los niveles de proteína de Caspasa 3 y Caspasa 9 escindidas aumentaron significativamente en el shRNA de Egr1- survivina combinado con el grupo de radioterapia. El shRNA de survivina de Egr1 combinado con radioterapia puede regular negativamente la expresión de survivina, lo que aumenta aún más la apoptosis y mejora la radiosensibilidad de las células ECA109 y KYSE150.
Assuntos
Humanos , Neoplasias Esofágicas/terapia , Survivina , Carcinoma de Células Escamosas do Esôfago/terapia , Radiossensibilizantes , Tolerância a Radiação , RNA Mensageiro , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/radioterapia , Transfecção , Regulação para Baixo , Western Blotting , Apoptose , Terapia Combinada , RNA Interferente Pequeno , Linhagem Celular Tumoral/efeitos da radiação , Proteína 1 de Resposta de Crescimento Precoce , Caspase 3 , Caspase 9 , Reação em Cadeia da Polimerase em Tempo Real , Citometria de Fluxo , Carcinoma de Células Escamosas do Esôfago/genética , Carcinoma de Células Escamosas do Esôfago/radioterapiaRESUMO
BACKGROUND: Anaplastic thyroid cancer (ATC) represents a rare lethal human malignancy with poor prognosis. Multimodality treatment, including radiotherapy, is recommended to improve local control and survival. Valproic acid (VA) is a clinically available histone deacetylase inhibitor with a well-documented side effect profile. In this study, we aim to investigate the combined effect of VA with photon irradiation in vitro. METHODS: Anaplastic thyroid cancer cells (8505c) were used to investigate the radiosensitizing effect of VA. RESULTS: VA sensitized cells to photon irradiation. VA increased radiation-induced apoptosis and radiation-induced DNA damage measured by γH2AX foci induction. Furthermore, VA prolonged γH2AX foci disappearance over time in irradiated cells and decreased the radiation-induced levels of mRNA of key DNA damage repair proteins of the homologous recombination (HR) and the nonhomologous end joining (NHEJ) pathways. CONCLUSIONS: VA at a clinically safe dose enhance the radiosensitivity of 8505c cells through an increase in radiation-induced apoptosis and a disruption in the molecular mechanism of HR and NHEJ DNA damage repair pathways.
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Carcinoma Anaplásico da Tireoide , Neoplasias da Glândula Tireoide , Humanos , Ácido Valproico/farmacologia , Histonas/metabolismo , Carcinoma Anaplásico da Tireoide/tratamento farmacológico , Carcinoma Anaplásico da Tireoide/genética , Linhagem Celular Tumoral , Neoplasias da Glândula Tireoide/tratamento farmacológico , Neoplasias da Glândula Tireoide/genética , Neoplasias da Glândula Tireoide/radioterapia , Dano ao DNARESUMO
PURPOSE: Esophageal squamous cell carcinoma is associated with high morbidity and mortality rate for which radiotherapy is the main treatment modality. Niraparib, a Poly (ADP-ribose) polymerase 1 inhibitors (PARPi) was previously reported to confer radiosensitivity in different malignancies including non-small cell lung cancer. In this study, we assessed the in vivo ability of niraparib in conferring radiosensitivity to esophageal squamous cell carcinoma cells. MATERIALS AND METHODS: In this study, KYSE-30 and KYSE-150 cell lines were selected as in vivo esophageal squamous cell carcinoma models. The experimental groups were: niraparib tosylate alone, radiotherapy alone, control (no intervention), and combination therapy (radiotherapy + niraparib tosylate). Cell cytotoxicity assay, colony formation assay, flow cytometry, immunofluorescence, Western blotting, immunohistochemistry, lentivirus transfection analysis, and xenograft models were used for confirming radiosensitizing ability of niraparib and to investigate the possible cellular mechanism involved in radiosensitization. RESULTS: The colony formation efficiency of the combination group was significantly much lower than that of the single radiation group (P < 0.01). Cell cytotoxicity assay demonstrated a significant reduction in proliferation of irradiated cells after treatment with niraparib tosylate compared to niraparib tosylate alone (P < 0.01). Cell apoptosis significantly increased in the combination group compared to either niraparib tosylate or radiotherapy alone (P < 0.01). Rate of tumor suppression rate was significantly high in the combined treatment group (P < 0.01) but, significantly decreased in nude mice. Western blot and lentivirus infection model suggested overexpression of FANCG genes to confer radiosensitivity. CONCLUSION: These results suggest that the synergistic effect of niraparib tosylate and radiation may be related to the down-regulation of FANCG.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Esofágicas , Carcinoma de Células Escamosas do Esôfago , Neoplasias Pulmonares , Radiossensibilizantes , Animais , Apoptose , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Linhagem Celular Tumoral , Neoplasias Esofágicas/tratamento farmacológico , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/radioterapia , Carcinoma de Células Escamosas do Esôfago/tratamento farmacológico , Carcinoma de Células Escamosas do Esôfago/patologia , Carcinoma de Células Escamosas do Esôfago/radioterapia , Humanos , Indazóis , Neoplasias Pulmonares/tratamento farmacológico , Camundongos , Camundongos Nus , Piperidinas , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Inibidores de Poli(ADP-Ribose) Polimerases/uso terapêutico , Tolerância a Radiação , Radiossensibilizantes/farmacologia , Radiossensibilizantes/uso terapêuticoRESUMO
INTRODUCTION AND OBJECTIVES: Circular RNA La Ribonucleoprotein 1B (circ-LARP1B) was reported to serve as an oncogene in many types of cancers. Radiotherapy (RT) is an important element of the multimodal treatment concept in malignancies. Here, this work aimed to investigate the role of circ-LARP1B in the tumorigenesis and radiosensitivity of hepatocellular carcinoma (HCC). PATIENTS OR MATERIALS AND METHODS: Quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot were used to detect the expression of genes and proteins. In vitro experiments were conducted using cell counting Kit-8 (CCK-8), colony formation, EDU, transwell, and tube formation assays, respectively. Dual-luciferase reporter assay was employed to identify the target relationship between miR-578 and circ-LARP1B or IGF1R (insulin-like growth factor 1 receptor). In vivo assay was performed using murine xenograft model. RESULTS: Circ-LARP1B was highly expressed in HCC tissues and cells, and high expression of circ-LARP1B was closely associated with poor prognosis. Functional experiments demonstrated that circ-LARP1B silencing impaired cell proliferation, invasion, angiogenesis and reduced radioresistance in vitro. Mechanistically, circ-LARP1B could competitively bind with miR-578 to relieve the repression of miR-578 on the expression of its target gene IGF1R. Further rescue assay confirmed that miR-578 inhibition reversed the inhibitory effects of circ-LARP1B knockdown on HCC cell malignant phenotypes and radioresistance. Moreover, miR-578 overexpression restrained tumorigenicity and enhanced radiosensitivity in HCC cells, which were attenuated by IGF1R up-regulation. Besides that, circ-LARP1B knockdown impeded tumor growth and enhanced irradiation sensitivity in HCC in vivo. CONCLUSIONS: Circ-LARP1B knockdown restrained HCC tumorigenicity and enhanced radiosensitivity by regulating miR-578/IGF1R axis, providing a new target for the treatment of HCC.
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Carcinoma Hepatocelular , Neoplasias Hepáticas , MicroRNAs , Animais , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/radioterapia , Linhagem Celular Tumoral , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/radioterapia , Camundongos , MicroRNAs/genética , MicroRNAs/metabolismo , RNA Circular/genética , Tolerância a Radiação/genética , Receptor IGF Tipo 1/genética , Receptor IGF Tipo 1/metabolismoRESUMO
PURPOSE: Patients submitted to radiotherapy (RT) may present in their healthy tissues surrounding the treated tumor, some typical acute inflammatory reactions induced by ionizing radiation (IR). The manifestation of inflammatory processes is a result of exacerbation of the immune system, as a response to radiation exposure, and this can be a limiting factor for RT protocols. To counteract this, some thiazolidinediones, such as LPSF/GQ-16, may be useful for modulating the patient's radioinduced inflammatory response in normal tissues. In this context, the present work aims to evaluate the activity of LPSF/GQ-16 on the levels of cytokines and the expression of the gene PPARγ in mononuclear cells irradiated in vitro, to analyze the immunomodulatory activity of the molecule and its action on radiomitigation. MATERIALS AND METHODS: For this, blood samples from eight donors were collected and irradiated with 2 Gy, then the PBMC (peripheral blood mononuclear cells) were cultured and treated with LPSF/GQ-16. The levels of cytokines TNF-α, IFN-γ, IL-2 and IL-4 were quantified by CBA, while the genes of TNF-α, IFN-γ and PPARγ were analyzed by RT-PCR. RESULTS: LPSF/GQ-16 significantly reduced the expression of proinflammatory cytokines (IFN-γ and TNF-α) in irradiated and nonirradiated groups. There was no significant reduction of anti-inflammatory cytokines (IL-2 and IL-4) by LPSF/GQ-16. The mRNA expression of PPAR-γ, IFN-γ and TNF-α in the presence of LPSF/GQ-16 was higher in the nonirradiated sample. CONCLUSION: LPSF/GQ-16 showed effective activity after irradiation, with an important immunomodulatory activity in irradiated PBMCs.
Assuntos
PPAR gama , Tiazolidinedionas , Citocinas/genética , Expressão Gênica , Humanos , Interleucina-2 , Interleucina-4 , Leucócitos Mononucleares , PPAR gama/genética , Fator de Necrose Tumoral alfaRESUMO
PURPOSE: This study set out to probe into the effect and mechanism of miR-144-3p on radiosensitivity of gastric cancer (GC) cells. METHODS: Cancer tissue and paracancerous tissue of GC patients admitted to our hospital were collected, their miR-144-3p expression was tested, GC cells were transfected, and survival and biological behavior of those cells under radiation were detected. RESULTS: After detection, miR-144-3p expression was down-regulated in GC tissue, while ZEB1 was up-regulated. There was no remarkable difference in the survival fraction of cells in each group before receiving radiation, but that of tumor cells decreased obviously (p < 0.05) after radiation exposure. Survival fraction of cells overexpressing miR-144-3p or silencing ZEB1 decreased more obviously, while the inhibition of miR-144-3p or overexpressing ZEB1 was weaker. Biological behavior of cells under 6 Gy radiation was detected. It was found that miR-144-3p overexpression or silencing ZEB1 dramatically inhibited the proliferation activity of GC cells under 6 Gy radiation, increased the levels of pro-apoptotic Bax and caspase-3 proteins (p < 0.05) and decreased the anti-apoptotic protein Bcl-2 level (p < 0.05), resulting in an increase in the apoptosis rate of cells. miR-144-3p was confirmed to be ZEB1 targeting site by dual luciferase report. Moreover, rescue experiments prove that it can increase the radiosensitivity of GC cells by regulating ZEB1 expression. CONCLUSION: miR-144-3p expression was down-regulated in GC, and it can increase the radiosensitivity of those cells by inhibiting ZEB1 expression.
Assuntos
MicroRNAs/metabolismo , Tolerância a Radiação , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/radioterapia , Homeobox 1 de Ligação a E-box em Dedo de Zinco/metabolismo , Apoptose , Caspase 3/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos da radiação , Sobrevivência Celular , Regulação para Baixo , Feminino , Mucosa Gástrica/metabolismo , Inativação Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Doses de Radiação , Transfecção/métodos , Regulação para Cima , Homeobox 1 de Ligação a E-box em Dedo de Zinco/genética , Proteína X Associada a bcl-2/metabolismoRESUMO
Poly (ADP-ribose) polymerase (PARP) inhibitors (iPARPs) have shown efficacy in homologous recombination (HR) deficiency patients with advanced castration resistant prostate cancer and have shown a radiosensitizing effect in preclinical and early clinical trials. Preclinical data in prostate cancer cells suggest a similar cytotoxic effect with half the radiation dose under the effect of Olaparib or Rucaparib irrespective of HR status. Due to the biologic synergy of radiotherapy (RT) and iPARPs, the risk of recurrence of high-risk prostate cancer and the morbidity associated with prostate cancer local treatment, this interesting strategy seems promising, and a better understanding of the clinical implications remains to be elucidated.
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Resumen: La protección gonadal ha sido durante largo tiempo un importante factor para abordar el requisito "tan bajo como sea razonablemente posible" ALARA. El presente artículo realiza una revisión sistemática sobre la utilidad de la protección gonadal en la radiografía de pelvis en la cual se han destacado los factores que tienen que ver con su realización diaria, es decir, la dosis de radiación involucrada, la radiosensibilidad de las gónadas, la posición del protector gonadal y el factor psicológico de la población en relación con su uso. La incorporación de equipamientos modernos, con dosis y protocolo optimizados, transforma el beneficio de la protección en un tema al menos debatible. Mientras algunos siguen respaldando la medida, otros organismos y autores ya no la respaldan. Es necesario reconsiderar prácticas actuales fundadas en consensos científicos que pueden estar obsoletos y considerar el factor de cambio cultural basados en estos nuevos consensos para su implementación, sin generar mayor preocupación en la población.
Abstract: Gonadal protection has long been an important factor in addressing the ALARA "as low as reasonably possible" requirement. This article performs a systematic review on the usefulness of gonadal protection in pelvic radiography, in which the factors that have to do with its daily performance have been highlighted, that is, radiation dose involved, the radiosensitivity of the gonads, the position of the gonadal shielding and the psychological factor of the population in relation to its use. The incorporation of modern equipment, with optimized dose and protocols, transforms the benefit of gonad shielding into an issue that is at least debatable. While some continue to support the measure, other agencies and authors no longer support it. It is necessary to reconsider current practices based on scientific consensus that may be obsolete and consider the factor of cultural change based on these new consensus for its implementation, without generating major concern in the population.
Assuntos
Humanos , Criança , Pelve/diagnóstico por imagem , Proteção Radiológica/métodos , Gônadas/efeitos da radiação , Doses de Radiação , Raios X , RadiografiaRESUMO
BACKGROUND: Previous studies have identified that patients with EGFR mutations tend to have better responses to targeted therapy, as well as chemotherapy; however, the effect of genetic alterations in terms of radiotherapy (RT)-related outcomes has not been fully assessed. We studied the impact of common non-small cell lung cancer (NSCLC) genetic alterations (EGFR, ALK and KRAS) in relation to objective response rate (ORR) to RT in patients with brain metastases. METHODS: From 2009-2015, 153 patients with an available genotyping status were treated with whole-brain irradiation (WBI) before receiving systemic therapy. Primary outcome was ORR; secondary outcomes included intracranial progression-free survival (IPFS) and overall survival (OS). RESULTS: Overall, ORR was 47.1%. ORR to RT varied significantly according to molecular status: EGFR (64.5%) ALK (54.5%) KRAS (20%) and WT (35.4%) (P = 0.001). EGFR mutation was the only independently associated factor for response to WBI (RR 3.52 [95% CI 1.6-7.7]; P = 0.002). Median IPFS was 10.8 months [95% CI 8.2-13.5] overall; however, IPFS also varied significantly according to molecular status: EGFR (18.2 months), ALK (18.4 months), KRAS (6.0 months) and WT (8.7 months) (P < 0.0001). OS for EGFR, ALK, KRAS and WT patients was 36.6, 32.2, 15.5 and 22.4 months, respectively (P = 0.014). Intracranial-ORR (HR 0.4 [95% CI 0.2-0.6], P < 0.001) and mutation status (HR 0.7 [95% CI 0.6-0.9], P < 0.042) were independently associated with a higher OS. CONCLUSIONS: RT response varies as per tumor molecular status. The presence of EGFR mutations favors the organ-specific response to RT, and is associated with longer OS in patients with NSCLC and BM. KEY POINTS: This study addressed for the first time the difference in radiotherapy-related outcomes in patients with different genotypes of non-small cell lung cancer (NSCLC) before they received systemic therapy. Results show that response to radiotherapy varies as per tumor molecular status, particularly EGFR-mutated tumors, have a favorable response to radiotherapy, contrary to KRAS-mutated tumors.
Assuntos
Quinase do Linfoma Anaplásico/genética , Neoplasias Encefálicas/mortalidade , Carcinoma Pulmonar de Células não Pequenas/mortalidade , Regulação Neoplásica da Expressão Gênica/efeitos da radiação , Mutação , Proteínas Proto-Oncogênicas p21(ras)/genética , Adenocarcinoma de Pulmão/genética , Adenocarcinoma de Pulmão/mortalidade , Adenocarcinoma de Pulmão/patologia , Adenocarcinoma de Pulmão/radioterapia , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/genética , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/radioterapia , Neoplasias Encefálicas/secundário , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Carcinoma Pulmonar de Células não Pequenas/radioterapia , Receptores ErbB/genética , Feminino , Seguimentos , Rearranjo Gênico , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/mortalidade , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/radioterapia , Masculino , Pessoa de Meia-Idade , Prognóstico , Estudos Prospectivos , Radioterapia/mortalidade , Taxa de SobrevidaRESUMO
RESUMEN El mejoramiento genético convencional en frijol común resulta difícil debido a que presenta una base genética estrecha y muy estable. En este sentido, la combinación de la mutagénesis y el cultivo de tejidos, es una alternativa para inducir variabilidad genética en la búsqueda de tolerancia a factores bióticos y abióticos. Es por ello, que el presente trabajo tuvo como objetivo determinar el efecto de diferentes explantes irradiados en la regeneración in vitro de frijol común (Phaseolus vulgaris L.) cultivar "ICA Pijao". Se aplicaron radiaciones gamma en callos, en el nudo cotiledonal con un cotiledón (NC-1) con dosis de 0, 10, 20, 30, 40, 50 y 60 Gy y semillas con 0, 100, 200, 300 y 400 Gy. Se evaluó el porcentaje de germinación, longitud de las raíces, porcentaje de explantes que formaron callos, masa fresca (g) de los callos, número de brotes por callo y el número de brotes con raíces. La radiación gamma provocó una disminución en la masa fresca del callo y NC-1. Los callos y el NC-1 solamente formaron brotes con las dosis de 10 y 20 Gy, pero estos fueron hiperhíricos. Los resultados demostraron que la semilla irradiada fue el explante con el que se logró la regeneración in vitro de plantas con hojas definidas, por lo que se recomienda como explante inicial para el uso combinado de mutagénesis y regeneración in vitro de plantas para el cultivar P. vulgaris "ICA Pijao" a través de la organogénesis indirecta.
ABSTRACT Conventional breeding in common bean is difficult because they have a close and very stable genetic base. In this connection the combination of mutagenesis and tissue culture is an alternative to induce genetic variability in the search for tolerance to biotic and abiotic factors. For this reason, the present study aimed to determine the effect of different irradiated explants in the in vitro regeneration of common bean (Phaseolus vulgaris L.) cultivar "ICA Pijao". To do this, were applied doses gamma radiation in callus, cotiledonary node with one cotyledon (NC-1) with doses of radiation 0, 10, 20, 30, 40, 50 and 60 Gy and seeds with 0, 100, 200, 300 and 400 Gy. The length of roots in the germinated seeds, fresh mass (g) of the callus and the number of shoots per callus were determined. The gamma radiation caused a decrease in the fresh weight of callus and NC-1. The callus and NC-1, irradiated with doses of 10 and 20 Gy they formed buds but these were hyperhydric. Results demonstrated that the irradiated seed was the explant with which it was achieved regeneration of shoots with leaves defined, so it is recommended as initial explant for combined use of mutagenesis and in vitro regeneration of plants for P. vulgaris cultivar "ICA Pijao" via organogenesis indirect.
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BACKGROUND: Glioma is the most prevalent malignant tumor in human central nervous systems. Recently, the development of resistance to radiotherapy in glioma patients markedly vitiates the therapy outcome. MiR-153-3p has been reported to be closely correlated with tumor progression, but its effect and molecular mechanism underlying radioresistance remains unclear in glioma. METHODS: The expression of miR-153-3p was determined in radioresistant glioma clinical specimens as well as glioma cell lines exposed to irradiation (IR) using quantitative real-time PCR. Cell viability, proliferation and apoptosis were then evaluated by MTT assay, colony formation assay, Flow cytometry analysis and caspase-3 activity assay in glioma cells (U87 and U251). Tumor forming was evaluated by nude mice model in vivo. TUNEL staining was used to detect cell apoptosis in nude mice model. The target genes of miR-153-3p were predicted and validated using integrated bioinformatics analysis and a luciferase reporter assay. RESULTS: Here, we found that miR-153-3p was down-regulated in radioresistant glioma clinical specimens as well as glioma cell lines (U87 and U251) exposed to IR. Enhanced expression of miR-153-3p promoted the radiosensitivity, promoted apoptosis and elevated caspase-3 activity in glioma cells in vitro, as well as the radiosensitivity in U251 cell mouse xenografs in vivo. Mechanically, B cell lymphoma-2 gene (BCL2) was identified as the direct and functional target of miR-153-3p. Moreover, restoration of BCL2 expression reversed miR-153-3p-induced increase of radiosensitivity, apoptosis and caspase-3 activity in U251 cells in vitro. In addition, clinical data indicated that the expression of miR-153-3p was significantly negatively associated with BCL2 in radioresistance of glioma samples. CONCLUSIONS: Our findings suggest that miR-153-3p is a potential target to enhance the effect of radiosensitivity on glioma cells, thus representing a new potential therapeutic target for glioma.
Assuntos
Genes bcl-2/fisiologia , Glioma/genética , MicroRNAs/fisiologia , MicroRNAs/efeitos da radiação , Tolerância a Radiação/genética , Adulto , Análise de Variância , Western Blotting , Caspase 3/análise , Linhagem Celular Tumoral , Proliferação de Células/efeitos da radiação , Sobrevivência Celular/efeitos da radiação , Regulação para Baixo , Feminino , Citometria de Fluxo , Regulação Neoplásica da Expressão Gênica , Marcação de Genes/métodos , Genes bcl-2/efeitos da radiação , Glioma/radioterapia , Humanos , Marcação In Situ das Extremidades Cortadas , Masculino , MicroRNAs/análise , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase em Tempo Real , Fatores de TempoRESUMO
Radiotherapy is one of the leading treatments for clinical cancer therapy. External beam radiotherapy has been proposed as an adjuvant treatment for patients bearing differentiated thyroid cancer refractory to conventional therapy. Our purpose was to study the combined effect of HDAC inhibitors (HDACi) and ionizing irradiation in thyroid cancer cell lines (Nthy-ori 3-1, WRO, TPC-1 and 8505c). HDACi radiosensitized thyroid cancer cells as evidenced by the reduction of survival fraction, whereas they had no effect in the normal cells. HDACi enhanced radiation-induced cell death in WRO cells. Gamma-H2AX foci number increased and persisted long after ionizing exposure in the HDACi-treated cells (WRO and TPC-1). Moreover, the expression of the repair-related gene Ku80 was differentially modulated only in the cancer cells, by the compounds at the protein and/or mRNA levels. We present in vitro evidence that HDACi can enhance the radiosensitivity of human thyroid cancer cells.
Assuntos
Ácido Butírico/farmacologia , Inibidores de Histona Desacetilases/farmacologia , Tolerância a Radiação/efeitos dos fármacos , Neoplasias da Glândula Tireoide/patologia , Ácido Valproico/farmacologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos da radiação , Morte Celular/efeitos dos fármacos , Morte Celular/efeitos da radiação , Linhagem Celular Tumoral , Dano ao DNA , Enzimas Reparadoras do DNA/genética , Enzimas Reparadoras do DNA/metabolismo , Raios gama , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos da radiação , Histonas/metabolismo , Humanos , Tolerância a Radiação/efeitos da radiação , Neoplasias da Glândula Tireoide/genéticaRESUMO
Enhanced radiosensitivity at low doses of ionizing radiation (IR) (0.2 to 0.6 Gy) has been reported in several cell lines. This phenomenon, known as low doses hyper-radiosensitivity (LDHRS), appears as an opportunity to decrease toxicity of radiotherapy and to enhance the effects of chemotherapy. However, the effect of low single doses IR on cell death is subtle and the mechanism underlying LDHRS has not been clearly explained, limiting the utility of LDHRS for clinical applications. To understand the mechanisms responsible for cell death induced by low-dose IR, LDHRS was evaluated in DLD-1 human colorectal cancer cells and the expression of 80 microRNAs (miRNAs) was assessed by qPCR array. Our results show that DLD-1 cells display an early DNA damage response and apoptotic cell death when exposed to 0.6 Gy. miRNA expression profiling identified 3 over-expressed (miR-205-3p, miR-1 and miR-133b) and 2 down-regulated miRNAs (miR-122-5p, and miR-134-5p) upon exposure to 0.6 Gy. This miRNA profile differed from the one in cells exposed to high-dose IR (12 Gy), supporting a distinct low-dose radiation-induced cell death mechanism. Expression of a mimetic miR-205-3p, the most overexpressed miRNA in cells exposed to 0.6 Gy, induced apoptotic cell death and, more importantly, increased LDHRS in DLD-1 cells. Thus, we propose miR-205-3p as a potential radiosensitizer to low-dose IR.
RESUMO
BACKGROUND: Glioma is the most prevalent malignant tumor in human central nervous systems. Recently, the development of resistance to radiotherapy in glioma patients markedly vitiates the therapy outcome. MiR-153-3p has been reported to be closely correlated with tumor progression, but its effect and molecular mechanism underlying radioresistance remains unclear in glioma. METHODS: The expression of miR-153-3p was determined in radioresistant glioma clinical specimens as well as glioma cell lines exposed to irradiation (IR) using quantitative real-time PCR. Cell viability, proliferation and apoptosis were then evaluated by MTT assay, colony formation assay, Flow cytometry analysis and caspase-3 activity assay in glioma cells (U87 and U251). Tumor forming was evaluated by nude mice model in vivo. TUNEL staining was used to detect cell apoptosis in nude mice model. The target genes of miR-153-3p were predicted and validated using integrated bioinformatics analysis and a luciferase reporter assay. RESULTS: Here, we found that miR-153-3p was down-regulated in radioresistant glioma clinical specimens as well as glioma cell lines (U87 and U251) exposed to IR. Enhanced expression of miR-153-3p promoted the radiosensitivity, promoted apoptosis and elevated caspase-3 activity in glioma cells in vitro, as well as the radiosensitivity in U251 cell mouse xenografs in vivo. Mechanically, B cell lymphoma-2 gene (BCL2) was identified as the direct and functional target of miR-153-3p. Moreover, restoration of BCL2 expression reversed miR-153-3p-induced increase of radiosensitivity, apoptosis and caspase-3 activity in U251 cells in vitro. In addition, clinical data indicated that the expression of miR-153-3p was significantly negatively associated with BCL2 in radioresistance of glioma samples. CONCLUSIONS: Our findings suggest that miR-153-3p is a potential target to enhance the effect of radiosensitivity on glioma cells, thus representing a new potential therapeutic target for glioma.
Assuntos
Humanos , Masculino , Feminino , Adulto , Pessoa de Meia-Idade , Tolerância a Radiação/genética , Genes bcl-2/fisiologia , MicroRNAs/efeitos da radiação , MicroRNAs/fisiologia , Glioma/genética , Fatores de Tempo , Regulação para Baixo , Regulação Neoplásica da Expressão Gênica , Sobrevivência Celular/efeitos da radiação , Western Blotting , Análise de Variância , Marcação de Genes/métodos , Genes bcl-2/efeitos da radiação , Marcação In Situ das Extremidades Cortadas , MicroRNAs/análise , Linhagem Celular Tumoral , Proliferação de Células/efeitos da radiação , Caspase 3/análise , Reação em Cadeia da Polimerase em Tempo Real , Citometria de Fluxo , Glioma/radioterapiaRESUMO
AIMS: Polymorphisms in cell cycle genes are considered prognostic as radiosensitivity markers in patients with head and neck squamous cell carcinoma. Therefore, we aimed to investigate the relationship of ATM 5557G>A, ATM IVS62 + 60G>A, TP53 215G>C, BCL2-938C>A, TGFß-509C>T, and TGFß 29C>T with radiotherapy response. MATERIALS AND METHODS: Genotyping was performed by polymerase chain reaction followed by restriction fragment length polymorphism in 210 patients with oral cavity/oropharyngeal carcinoma and 101 patients with laryngeal tumors. RESULTS: In irradiated oral cavity/oropharyngeal tumors, the ATM IVS62 + 60G>A AA genotype significantly increased local recurrence risk (odds ratio [OR] = 4.43; confidence interval [CI] = 1.22-16.13) and the BCL2-938C>A C allele and the TGFß-509C>T T allele were associated with worse disease-specific survival (hazard ratio [HR] = 0.46; CI = 0.24-0.90 and HR = 2.20; CI = 1.12-4.29, respectively). In irradiated laryngeal carcinoma, the TGFß 29C>T C allele was associated with increased local recurrence risk (OR = 0.09; CI = 0.02-0.53), death rate (OR = 0.18; CI = 0.04-0.86), and worse local disease-free and disease-specific survival rates (HR = 0.13; CI = 0.03-0.59 and HR = 0.21; CI = 0.07-0.60, respectively), while the BCL2-938C>A C allele was related to a worse disease-specific survival (HR = 0.32; CI = 0.12-0.83). DISCUSSION: These results can help individualize treatment according to a patient's genetic markers. We demonstrated that ATM IVS62 + 60G>A, TGFß 29C>T, TGFß-509C>T, and BCL2-938C>A can function as biomarkers of tumor radiosensitivity, being candidates for a predictive genetic profile of radiotherapy response.
Assuntos
Biomarcadores Tumorais/genética , Neoplasias de Cabeça e Pescoço/genética , Adulto , Idoso , Proteínas Mutadas de Ataxia Telangiectasia/genética , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Carcinoma de Células Escamosas/genética , Feminino , Genótipo , Neoplasias de Cabeça e Pescoço/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Polimorfismo de Nucleotídeo Único/genética , Prognóstico , Regiões Promotoras Genéticas/genética , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Radioterapia , Taxa de Sobrevida , Fator de Crescimento Transformador beta1/genética , Fator de Crescimento Transformador beta1/metabolismo , Resultado do TratamentoRESUMO
BACKGROUND: miR-22 has been shown to be frequently downregulated and act as a tumor suppressor in multiple cancers including breast cancers. However, the role of miR-22 in regulating the radioresistance of breast cancer cells, as well as its underlying mechanism is still not well understood. METHODS: The expressions of miR-22 and sirt1 at mRNA and protein levels were examined by qRT-PCR and Western Blot. The effects of miR-22 overexpression and sirt1 knockdown on cell viability, apoptosis, radiosensitivity, γ-H2AX foci formation were evaluated by CCK-8 assay, flow cytometry, colony formation assay, and γ-H2AX foci formation assay, respectively. Luciferase reporter assay and qRT-PCR analysis were performed to confirm the interaction between miR-22 and sirt1. RESULTS: miR-22 was downregulated and sirt1 was upregulated at both mRNA and protein levels in breast cancer cells. miR-22 overexpression or sirt1 knockdown significantly suppressed viability, induced apoptosis, reduced survival fraction, and increased the number of γ-H2AX foci in breast cancer cells. Sirt1 was identified as a target of miR-22 and miR-22 negatively regulated sirt1 expression. Ectopic expression of sirt1 dramatically reversed the inhibitory effect of miR-22 on cell viability and promotive effect on apoptotic rates and radiosensitivity in breast cancer cells. CONCLUSIONS: miR-22 suppresses tumorigenesis and improves radiosensitivity of breast cancer cells by targeting sirt1, providing a promising therapeutic target for breast cancer.
Assuntos
Neoplasias da Mama/radioterapia , MicroRNAs/metabolismo , Tolerância a Radiação , Sirtuína 1/metabolismo , Apoptose/genética , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular , Regulação para Baixo , Feminino , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Histonas/metabolismo , Humanos , Dosagem Radioterapêutica , Sirtuína 1/genéticaRESUMO
ABSTRACT This study aimed to evaluate the radiosensitivity of castor bean seeds after applications of different doses of Cobalt60 gamma radiation. Seeds were pre-soaked for 24 hours in distilled water and then irradiated with 50, 100, 150, and 200 Gy, except the control. Sowing was performed in trays, which contained soil as substrate and were maintained in a greenhouse. The electrical conductivity, emergence, emergence speed index, growth parameters and activities of antioxidant enzymes (superoxide dismutase, ascorbate peroxidase, and catalase) were evaluated in the leaves and roots of castor bean seedlings. Gamma radiation did not affect the electrical conductivity of the seeds; however, at a dose of 200 Gy, the emergence and emergence speed index of the seedlings was negatively affected. An analysis of the morphophysiological parameters revealed a reduction in seedling size as the radiation dose increased. There was a significant increase in superoxide dismutase and ascorbate peroxidase activities at higher radiation doses in the leaves, but not in roots. Thus, the analysis of all the variables suggests a response pattern as to the morphophysiological and biochemical changes of castor bean seedlings due to the increase of gamma radiation, which may serve as a tool for generating greater genetic variability.
Assuntos
Ricinus/efeitos da radiação , Sementes/efeitos da radiação , Radioisótopos de Cobalto , Germinação/efeitos da radiação , Raios gama , Ricinus/crescimento & desenvolvimento , Sementes/crescimento & desenvolvimento , Relação Dose-Resposta à RadiaçãoRESUMO
BACKGROUND: miR-22 has been shown to be frequently downregulated and act as a tumor suppressor in multiple cancers including breast cancers. However, the role of miR-22 in regulating the radioresistance of breast cancer cells, as well as its underlying mechanism is still not well understood. METHODS: The expressions of miR-22 and sirt1 at mRNA and protein levels were examined by qRT-PCR and Western Blot. The effects of miR-22 overexpression and sirt1 knockdown on cell viability, apoptosis, radiosensitivity, γ-H2AX foci formation were evaluated by CCK-8 assay, flow cytometry, colony formation assay, and γ-H2AX foci formation assay, respectively. Luciferase reporter assay and qRT-PCR analysis were performed to confirm the interaction between miR-22 and sirt1. RESULTS: miR-22 was downregulated and sirt1 was upregulated at both mRNA and protein levels in breast cancer cells. miR-22 overexpression or sirt1 knockdown significantly suppressed viability, induced apoptosis, reduced survival fraction, and increased the number of γ-H2AX foci in breast cancer cells. Sirt1 was identified as a target of miR-22 and miR-22 negatively regulated sirt1 expression. Ectopic expression of sirt1 dramatically reversed the inhibitory effect of miR-22 on cell viability and promotive effect on apoptotic rates and radiosensitivity in breast cancer cells. CONCLUSIONS: miR-22 suppresses tumorigenesis and improves radiosensitivity of breast cancer cells by targeting sirt1, providing a promising therapeutic target for breast cancer.
Assuntos
Humanos , Feminino , Tolerância a Radiação , Neoplasias da Mama/radioterapia , MicroRNAs/metabolismo , Sirtuína 1/metabolismo , Dosagem Radioterapêutica , Neoplasias da Mama/metabolismo , Histonas/metabolismo , Regulação para Baixo , Regulação Neoplásica da Expressão Gênica , Sobrevivência Celular , Apoptose/genética , Linhagem Celular Tumoral , Técnicas de Silenciamento de Genes , Sirtuína 1/genéticaRESUMO
BACKGROUND: To determine the effects of endostatin on vascular growth factor receptor 2 (VEGFR2) expression in non-small cell lung cancer (NSCLC) cells and the mechanisms underlying its radiosensitizing effect. METHODS: VEGFR2 mRNA levels were determined in different NSCLC cell lines using qRT-PCR. RT-PCR and Western blot assays were used to assess the expression of mRNA and proteins. The radiosensitivity of the cells was determined by colony-formation assays; and cell apoptosis and cell cycle distribution were determined by flow cytometry. RESULTS: VEGFR2 mRNA levels differed among the five NSCLC cell lines (P < 0.01), with the highest expression in Calu-1 cells and lowest in A549 cells. Endostatin significantly inhibited the growth of Calu-1 cells (P < 0.01) (IC20 = 296.5 µg/ml), and the expression of VEGFR2 and HIF-1α (P < 0.05). Phosphorylation of protein kinase B (Akt), extracellular signal-regulated kinases 1/2 (ERK1/2), and p38 were significantly lower in endostatin-treated cells than control (P < 0.05). Endostatin enhanced the radiosensitivity of Calu-1 cells to SER = 1.38 and induced apoptosis (P < 0.01) and G2/M blockage (P < 0.01). However, endostatin had limited effects on A549 cells. Compared with Calu-1 cells, there was not significantly effects on cell radiosensitivity (SER = 1.09). CONCLUSIONS: Endostatin induces apoptosis and enhances radiosensitivity of the VEGFR2 high-expressing cell line Calu-1, but it has a limited effect on the VEGFR2 low-expressing cell line A549.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Endostatinas/farmacologia , Neoplasias Pulmonares/genética , Radiossensibilizantes/farmacologia , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Morte Celular/efeitos dos fármacos , Morte Celular/genética , Linhagem Celular Tumoral , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/genética , Raios gama , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Neoplasias Pulmonares/patologia , Tolerância a Radiação/efeitos dos fármacos , Tolerância a Radiação/genéticaRESUMO
Cervical cancer is the second common cause of cancer deaths in women worldwide. Radioresistancy of cancer is a principal cause of treatment impairing. Inhibitor of apoptosis proteins (IAPs) widely block apoptosis against apoptotic stimuli, including current chemo- and radiation therapies. Apollon, a membrane of IAP, can support cells against apoptosis and is over expressed in some treatment-resistant cancer cells. The aim of this study was to evaluate the effects of apollon knockdown on induction of apoptosis and also its potential for enhancement of radiosensitvity on hela cells. plasmid encoding shRNA which has been confirmed its effect against apollon, transfected into hela cells. Consequent effects on the level of P53 , Bax and BAK analyzed by real time PCR. Apoptotic phenotype of transfected cells was monitored by Tunnel assay. Viability of hela cells after radiotherapy was analyzed by MTT assay. shRNA1 effectively increased transcription of p53, Bax and BAK and induced apoptosis phenotype of treated hela cells. Radiosensitivity of transfected cell was increased after knock-down of apollon obviously. Apollon knockdown induces apoptosis in hela cell . Also it can be as new molecular target for radio-sensitizing strategies in these cells. So, apollon can be a potentially considerable therapeutic object for cervical cancer.