Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 9 de 9
Filtrar
1.
Journal of Experimental Hematology ; (6): 45-48, 2013.
Artículo en Chino | WPRIM | ID: wpr-325215

RESUMEN

This study was purposed to explore the changes of possible angiogenetic factors other than VEGF after inhibition of NHE1 and their related mechanisms. The K562 cells were treated by NHE1 specific inhibitor cariporide, the angiogenesis factors after inhibition of NHE1 were screened by using protein chip, the IL-8 expression level after cariporide treatment was detected by real-time quantitative PCR; the K562 cells with stable interference of NHE1 were constructed, the IL-8 expression level after interference of NHE1 was detected by real-time quantitative PCR; the p38 phosphorylation level in K562 cells treated with cariporide was detected by Western blot. After treatment of K562 cells with p38 inhibitor SB203580, the IL-8 expression level was decreased by real-time quantitative PCR. The results of protein chip showed that IL-8 expression decreased after cariporide treatment. Real-time quantitative PCR confirmed this inhibitory effect. The p38 phosphorylation level increased after cariporide treatment. The down-regulation of IL-8 expression induced by cariporide treatment was partially restored after K562 cells were treated with p38 inhibitor SB203580. It is concluded that the inhibition of NHE1 can inhibit IL-8 expression through up-regulation of p38 phosphorylation.


Asunto(s)
Humanos , Proteínas de Transporte de Catión , Regulación hacia Abajo , Guanidinas , Farmacología , Imidazoles , Farmacología , Interleucina-8 , Metabolismo , Células K562 , Fosforilación , Piridinas , Farmacología , Intercambiador 1 de Sodio-Hidrógeno , Intercambiadores de Sodio-Hidrógeno , Sulfonas , Farmacología , Proteínas Quinasas p38 Activadas por Mitógenos , Metabolismo
2.
Journal of Experimental Hematology ; (6): 1341-1345, 2012.
Artículo en Chino | WPRIM | ID: wpr-325263

RESUMEN

This study was aimed to investigate whether the inhibition of NHE1 activity and intracellular acidification can reverse resistance of leukemia cells to the imatinib and to explore downstream signal molecule networks of BCR/ABL in the cells of chronic myelocytic leukemia (CML) patients. The mRNA and protein expression of P-glycoprotein (Pgp) and the drug accumulation were assayed after acidifying the primary leukemia cells of patients or K562/DOX and K562/G01 cells. The effects of intracellular acidification of primary leukemia cells on the phosphorylation level changes of ERK1/2 and p38 MAPK were analyzed by Western blot. The results showed that the intracellular concentration of drugs in the advanced patients increased and the sensitivity of K562/DOX and K562/G01 cells to imatinib was enhanced after intracellular acidification or treatment with NHE1 inhibitor cariporide. With downregulation of intracellular pH, the phosphorylation of p38 MAPK decreased in advanced patients and the phosphorylation of ERK1/2 increased within 3 min and then decreased after 30 min. SB203580, the specific inhibitor of p38 MAPK, displayed a synergistic effect with the inhibitor of NHE1 to downregulate the mRNA and protein expression of Pgp. It is concluded that the inhibiton of NHE1 can significantly decrease the protein expression of Pgp in K562/DOX and K562/G01 cells, increase the accumulation of Rhodamine123 and doxorubicin in the cells of advanced patients and enhance the sensitivity of cells to imatinib in which the p38 MAPK signal transduction pathways involves.


Asunto(s)
Humanos , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP , Metabolismo , Benzamidas , Farmacología , Proteínas de Transporte de Catión , Metabolismo , Resistencia a Antineoplásicos , Inhibidores Enzimáticos , Farmacología , Regulación Leucémica de la Expresión Génica , Mesilato de Imatinib , Imidazoles , Farmacología , Células K562 , Sistema de Señalización de MAP Quinasas , Piperazinas , Farmacología , Piridinas , Farmacología , Pirimidinas , Farmacología , Intercambiador 1 de Sodio-Hidrógeno , Intercambiadores de Sodio-Hidrógeno , Metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos , Metabolismo
3.
Journal of Experimental Hematology ; (6): 570-573, 2011.
Artículo en Chino | WPRIM | ID: wpr-313942

RESUMEN

This study was aimed to investigate the expression level of CIAPIN1 mRNA in leukemia patients and explore its significance in leukemias. The fresh bone marrow was collected from 112 newly diagnosed leukemia patients, the total RNA was extracted by means of TRIzoL, the cDNA was synthesized, the expression of CIAPIN1 mRNA was detected by real-time quantitative PCR using β-actin as internal reference; 10 normal healthy persons were selected as controls. The results showed that the expression of CIAPIN1 mRNA was statistically higher in acute myeloid leukemia (AML), acute lymphocytic leukemia (ALL) and chronic phase chronic myeloid leukemia (CML) patients than that in normal persons (p < 0.05); but there was no statistical difference between chronic lymphocytic leukemia (CLL) and normal persons (p > 0.05). It is concluded that the CIAPIN1 gene higher expresses in MNC of newly diagnosed leukemia patients, up-regulation of CIAPIN1 expression may play an important role in pathogenesis of leukemia.


Asunto(s)
Adulto , Femenino , Humanos , Masculino , Persona de Mediana Edad , Adulto Joven , Células de la Médula Ósea , Metabolismo , Estudios de Casos y Controles , Regulación Leucémica de la Expresión Génica , Péptidos y Proteínas de Señalización Intracelular , Genética , Leucemia , Genética , Metabolismo , Patología , Leucemia Linfocítica Crónica de Células B , Genética , Leucemia Mielógena Crónica BCR-ABL Positiva , Genética , Leucemia Mieloide Aguda , Genética
4.
Journal of Experimental Hematology ; (6): 656-660, 2011.
Artículo en Chino | WPRIM | ID: wpr-313922

RESUMEN

This study was aimed to investigate the effects of anti-CD44 mAb A3D8 on proliferation and apoptosis of AML cells, to explore the mechanism of ERK1/2 and Bim in this process. Effect of the anti-CD44 mAb A3D8 on the HL-60 cell proliferation was assayed with MTT method, the change of mitochondrial transmembrane potential of HL-60 cells was analyzed by flow cytometry. The mRNA expression of Bim was determined by real-time quantitative RT-PCR. Western blot was used to detect the protein expression of p-ERK1/2. The results showed that mAb A3D8 could remarkably inhibit the proliferation capacity of the HL-60 cells in a dosage- and time-dependent ways. The mitochondrial transmembrane potential in HL-60 cells treated with A3D8 (3.0 µg/ml) was significantly decreased as compared with the control cells. Furthermore, the mRNA expression of Bim was much higher than that in controls. Expression of the p-ERK was much lower than that of the controls. It is concluded that anti-CD44 mAb A3D8 can inhibit the proliferation and induce the apoptosis of HL-60 cells, mechanism of which is enhancing the expression of Bim via inhibiting p-ERK1/2.


Asunto(s)
Humanos , Anticuerpos Monoclonales , Alergia e Inmunología , Farmacología , Apoptosis , Proteínas Reguladoras de la Apoptosis , Metabolismo , Proteína 11 Similar a Bcl2 , Proliferación Celular , Regulación Leucémica de la Expresión Génica , Células HL-60 , Receptores de Hialuranos , Alergia e Inmunología , Proteínas de la Membrana , Metabolismo , Proteína Quinasa 1 Activada por Mitógenos , Metabolismo , Proteína Quinasa 3 Activada por Mitógenos , Metabolismo , Proteínas Proto-Oncogénicas , Metabolismo , Regulación hacia Arriba
5.
Journal of Experimental Hematology ; (6): 661-665, 2011.
Artículo en Chino | WPRIM | ID: wpr-313921

RESUMEN

This study was purposed to investigate the effect of hypoxia microenvironment on K562 leukemic cell differentiation, and characteristics of NHE1 involvement in this process. The K562 cells were treated with hypoxia-mimical agent CoCl₂ or under actual hypoxia culture, and the specific NHE1 inhibitor Cariporide was used to inhibit NHE1 activity. The fluorescent probe BCECF was used for pH(i) measurements. Gene expression was analyzed by RT-PCR. The morphological characteristics was determined by Wright's staining. Signaling pathways were detected by Western blot using phosphospecific antibodies. The results indicated that the hypoxia or mimetic hypoxia favored K562 cells differentiation with up-regulation of C/EBPα. Moreover, treatment with Cariporide under hypoxia synergistically enhanced leukemia cell differentiation. Treatment with Cariporide increased levels of phosphorylated ERK5 and P38 mitogen-activated protein kinase (MAPK). It is concluded that the hypoxia or mimetic hypoxia can induce the differentiation of K562 cells, the inhibition of NHE1 activity can promote the hypoxia-induced K562 cell differentiation. The enhancement of hypoxia-induced K562 differentiation by Cariporide via MAPK signal pathway suggests a possible therapeutic target of NHE1 under hypoxia microenvironment in the treatment of leukemias.


Asunto(s)
Humanos , Proteínas de Transporte de Catión , Metabolismo , Diferenciación Celular , Hipoxia de la Célula , Células K562 , Sistema de Señalización de MAP Quinasas , Intercambiador 1 de Sodio-Hidrógeno , Intercambiadores de Sodio-Hidrógeno , Metabolismo
6.
Journal of Experimental Hematology ; (6): 666-670, 2011.
Artículo en Chino | WPRIM | ID: wpr-313920

RESUMEN

The aim of this study was to investigate the effect of intracellular acidification on accumulation of rhodamine 123 (rh123) in non-mature cells with none or low expression of multidrug resistance MDR1. The expression of MDR1 mRNA was detected by real-time quantitative RT-PCR. Confocal laser microscopy was used to determine the calibration curve of intracellular acidification (pHi). MTT assay was used to detect the cytotoxicity of intracellular acidification on HL-60, MSC and CD34(+) cells from umbilical cord blood. Flow cytometry was applied to measure the influence of intracellular acidification. The results indicated that the intracellular acidification had no obvious cytotoxicity on HL-60, MSC and CD34(+) cells. The acidification resulted in the increased rhodamine 123 accumulation in HL-60, MSC and CD34(+) cells without P-gp activity. Moreover, the more primitive cells, the less accumulation of intracellular Rh123 were observed. It is concluded that the intracellular acidification can reverse the MDR of HL-60, MSC and CD34(+) cells.


Asunto(s)
Humanos , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP , Metabolismo , Antígenos CD34 , Metabolismo , Células de la Médula Ósea , Biología Celular , Metabolismo , Fenómenos Fisiológicos Celulares , Resistencia a Múltiples Medicamentos , Resistencia a Antineoplásicos , Sangre Fetal , Biología Celular , Metabolismo , Células HL-60 , Células Madre Mesenquimatosas , Biología Celular , Metabolismo
7.
Journal of Experimental Hematology ; (6): 857-860, 2011.
Artículo en Chino | WPRIM | ID: wpr-313878

RESUMEN

This study was aimed to investigate the expression of Na(+)/H(+) exchanger 1 (NHE1) in K562 and HL-60 cells undergoing DNA damage induced by etoposide and to elucidate the regulating mechanism. Real-time quantitative PCR (RQ-PCR) and Western blot methods were used to determine the expression of NHE1 in K562 cells after the treating with etoposide. Meanwhile, the flow cytometry was used to detect the apoptosis of leukemic cells. The luciferase reporter vector containing NHE1 promoter was constructed to measure relative luciferase activity after treating with different etoposide concentrations. The results showed that the mRNA and protein of NHE1 increased in accordance with apoptosis ratio in HL-60 cells after treated with etoposide (p < 0.05), but no such obvious increase in K562 cells. Treatment with NHE1 specific inhibitor could block etoposide induced alkalization and reduce the apoptosis ratio of HL-60 cells. The expression pattern and apoptosis alteration was not similar in K562 cells. Relative luciferase activity of reporter vector containing NHE1 promoter however increased in K562 cells after treated with etoposide. It is concluded that the expression of NHE1 is up-regulated in the process of apoptosis of HL-60 cells induced by etoposide and depends on the pHi increasing caused by NHE1 up-regulation which is not found in K562 cells although the transcriptional activity increased.


Asunto(s)
Humanos , Apoptosis , Proteínas de Transporte de Catión , Metabolismo , Daño del ADN , Etopósido , Células HL-60 , Células K562 , Regiones Promotoras Genéticas , Intercambiador 1 de Sodio-Hidrógeno , Intercambiadores de Sodio-Hidrógeno , Metabolismo
8.
Journal of Experimental Hematology ; (6): 122-126, 2010.
Artículo en Chino | WPRIM | ID: wpr-328560

RESUMEN

The objective of this study was to compare the effects between knocking-out Sam68 gene by homologous recombination method and silencing the gene by siRNA silencing technique in DT40 cell line. Gene targeting technique was used to isolate Sam68 gene-deleted chicken DT40 cells. Meanwhile, Sam68 gene silencing cells was obtained by using stable expression of siRNA plasmid pSilencer-Sam68. Then, the function of these two cell lines were analyzed by comparing with wild-type DT40 cell line. The results showed that the growth retardation in Sam68 gene knocked-out cell line was observed due to elongation of the G2/M phase, but which could not be found in Sam68 gene silencing cell line. It is concluded that in accordance with study of protein function in living cells, use of gene knockout technique for cell line can provide the experimental results more real than those resulting from gene silence technique.


Asunto(s)
Animales , Línea Celular Tumoral , Pollos , Regulación Neoplásica de la Expresión Génica , Técnicas de Inactivación de Genes , Silenciador del Gen , Marcación de Gen , Plásmidos , ARN Interferente Pequeño , Genética , Transfección
9.
Chinese Journal of Hematology ; (12): 605-609, 2009.
Artículo en Chino | WPRIM | ID: wpr-314452

RESUMEN

<p><b>OBJECTIVE</b>To investigate the impact of intracellular acidification (IA) on drug resistance of leukemia cells with high P-glycoprotein (P-gp) expression, and to provide a new method for the reversing of multidrug resistance (MDR).</p><p><b>METHODS</b>Real-time PCR was used to determine the expression level of mdr1 gene, and the leukemia cells with high P-gp expression were selected. The specific inhibitor of Na+/H+ exchanger 1 and the "high K+" buffer were used to acidify the cells, and the confocal laser microscopy was used to determine the intracellular pH (pHi) and effect of IA on the accumulation of doxorubicin. The MTT method was used to determine the effect of IA on the cell viability. The flow cytometry was used to detect the effect of IA on the P-gp function, and Western blotting was used to determine the effect of IA on the expression of P-gp.</p><p><b>RESULTS</b>The pHi was decreased to 7.0, and compared with that of control the mdr1 mRNA expression was decreased to (53.2+/-11.0)% after 1 h, and to (16.6+/-7.0)% after 3 h treatment. The P-gp expression was decreased to (56.0+/-9.0)% of the control after 3 h treatment. The accumulation of Rh123 was 71.03+/-0.47 at pHi 7.0, which was increased obviously as compared to the control group 20.07+/-0.39. The increased accumulation of doxorubicin was also observed by confocal laser microscopy.</p><p><b>CONCLUSION</b>The expression and function of P-gp on the patients cells are inhibited by IA.</p>


Asunto(s)
Humanos , Subfamilia B de Transportador de Casetes de Unión a ATP , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP , Genética , Metabolismo , Supervivencia Celular , Doxorrubicina , Farmacocinética , Resistencia a Antineoplásicos , Guanidinas , Farmacología , Concentración de Iones de Hidrógeno , Leucemia , Quimioterapia , Metabolismo , ARN Mensajero , Genética , Sulfonas , Farmacología , Células Tumorales Cultivadas
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA