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1.
Braz. j. biol ; 84: e251336, 2024. graf
Article in English | LILACS, VETINDEX | ID: biblio-1355879

ABSTRACT

Abstract Bulbine natalensis and Chorophytum comosum are potential medicinal source for the treatment of cancers. Chronic myeloid leukaemia is a hematopoietic stem cells disorder treated by tyrosine kinase inhibitors but often cause recurrence of the leukaemia after cessation of therapy, hence require alternative treatment. This study determines the anti-cancer effect of leaf, root and bulb methanolic and aqueous extracts of B. natalensis and C. comosum in chronic human myelogenous leukaemia (K562) cell line by MTT, Hoechst bis-benzimide nuclear and annexin V stain assays. The root methanolic extract of B. natalensis and C. comosum showed a high cytotoxicity of 8.6% and 16.7% respectively on the K562 cell line at 1,000 μg/ml concentration. Morphological loss of cell membrane integrity causing degradation of the cell and fragmentation were observed in the root methanolic extract of both plants. A high apoptosis (p < 0.0001) was induced in the K562 cells by both leaf and root extracts of the C. comosum compared to the B. natalensis. This study shows both plants possess apoptotic effect against in vitro myelogenous leukaemia which contributes to the overall anti-cancer properties of B. natalensis and C. comosum to justify future therapeutic applications against chronic myelogenous leukaemia blood cancer.


Resumo Bulbine natalensis Baker e Chorophytum comosum (Thunb.) Jacques são potenciais fontes medicinais para o tratamento de cânceres. A Leucemia Mieloide Crônica (LMC) é um distúrbio das células-tronco hematopoiéticas que é tratado com inibidores da tirosina quinase, mas frequentemente, causa recorrência da leucemia após a interrupção da terapia, portanto, requer um tratamento alternativo. Este estudo determinou o efeito anticancerígeno de extratos metanólicos e aquosos de folha, raiz e bulbo de B. natalensis e C. comosum na linhagem celular de leucemia mieloide humana crônica (K562) por ensaios de MTT, Hoechst bis-benzimida nuclear e anexina V. O extrato metanólico da raiz de B. natalensis e C. comosum apresentou alta citotoxidade de 8,6% e 16,7% respectivamente, na linhagem celular K562 com a concentração de 1,000 μg / ml. Perda morfológica da integridade da membrana celular causando degradação dos núcleos, citoplasma e encolhimento celular foi observada no extrato metanólico da raiz de ambas as plantas. Uma alta apoptose (p <0,0001) foi induzida nas células K562 por extratos de folhas e raízes de C. comosum em comparação com B. natalensis. Este estudo mostrou que ambas as plantas possuem efeito apoptótico contra leucemia mieloide in vitro que contribui para as propriedades anticâncer gerais de B. natalensis e C. comosum para justificar futuras aplicações terapêuticas contra câncer de sangue de LMC.


Subject(s)
Humans , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/drug therapy , Xanthorrhoeaceae , Apoptosis , K562 Cells
2.
Article in Chinese | WPRIM | ID: wpr-928677

ABSTRACT

OBJECTIVE@#To investigate the drug resistant related FOXO3/Bcl-6 signaling pathway in K562/G cell line and its related microRNA(miRNA) mechanisms.@*METHODS@#The drug resistance potency of imatinib on K562/G was detected by MTT assay. The expression of FOXO3 and Bcl-6 proteins in K562 and K562/G cells was detected by Western blot. Real-time PCR (RT-PCR) was used to detect the expression of FOXO3 and Bcl-6 mRNA. The miRNA expression profiling in K562 and K562/G cells was analyzed by microarray technique, and the miRNA targeted to FOXO/Bcl-6 signaling pathway was identified.@*RESULTS@#The expression of FOXO3 and Bcl-6 protein was significantly increased in K562/G cells as compared with that in K562 cells (P<0.01), the expression level of Bcl-6 mRNA showed no increase in K562/G cells. However, FOXO3 mRNA was up-regulated in K562/G cells (P<0.05). MiRNA microarray results showed that 109 miRNAs were expressed differentially in K562 and K562/G cells. The expression of 81 miRNAs were up-regulated while 28 miRNAs were down-regulated. Through reverse prediction by bioinformatics, miR-6718-5p, miR-5195-5p, miR-4711-3p, miR-4763-5p, miR-4664-5p and miR-3176 were related to FOXO/Bcl-6 signaling pathway.@*CONCLUSION@#The FOXO3/Bcl-6 signaling pathway contributes to imatinib resistance in K562/G cell line, and the miRNA expression profiles showed significant differences between K562/G and K562 cells.


Subject(s)
Forkhead Box Protein O3/genetics , Humans , Imatinib Mesylate/pharmacology , K562 Cells , MicroRNAs/genetics , RNA, Messenger , Signal Transduction
3.
Int. j. morphol ; 40(5): 1276-1283, 2022. ilus, graf
Article in Spanish | LILACS-Express | LILACS | ID: biblio-1405294

ABSTRACT

RESUMEN: Las bacteriocinas son péptidos antimicrobianos de síntesis ribosomal secretadas por bacterias. Dentro de estas destaca nisina que posee potenciales usos en terapias antibióticas, como biopreservante de alimentos y probióticos. También se ha descrito que nisina posee citotoxicidad sobre líneas celulares neoplásicas, pero existe poca información de su efecto sobre células tumorales sanguíneas. Debido al potencial uso que presenta nisina, es relevante determinar la toxicidad que presenta sobre líneas celulares tumorales del tipo sanguíneo. Para esto, se realizaron ensayos de actividad hemolítica sobre eritrocitos humanos y de toxicidad sobre células mononucleares de sangre periférica humanas, determinándose que nisina no posee efecto citotóxico sobre este tipo de células normales humanas sanguíneas. Se realizaron también, ensayos de citotoxicidad con líneas celulares tumorales (K562 y U937), con el fin de determinar dosis, tiempo de exposición y selectividad en el efecto tóxico de nisina sobre las células tumorales humanas. Estos ensayos muestran que nisina presenta actividad citotóxica sobre líneas celulares K562 y U937 a las 72 h de exposición, a una concentración de 40 µg/mL, que corresponde a 100 veces la concentración mínima inhibitoria (MIC) usada para su acción sobre bacterias. Al comparar el efecto de nisina sobre células mononucleares de sangre periférica humanas con las líneas tumorales linfoides y mieloides (K562 y U937 respectivamente), se observa un efecto selectivo de nisina sobre las células tumorales sanguíneas.


SUMMARY: Bacteriocins are antimicrobial peptides of ribosomal synthesis secreted by bacteria. Among these, nisin stands out, which has potential uses in antibiotic therapies, as a food bio preservative and probiotics. Nisin has also been reported to have cytotoxicity on neoplastic cell lines, but there is little information on its effect on blood tumor cells. Due to the potential use that nisin presents, it is relevant to determine the toxicity it presents on tumor cell lines of the blood type. For this, hemolytic activity tests were carried out on human erythrocytes and toxicity on human peripheral blood mononuclear cells, determining that nisin does not have a toxic effect on this type of normal human blood cells. Cytotoxicity tests were also carried out with tumor cell lines (K562 and U937), to determine dose, exposure time and selectivity in the toxic effect of nisin on human tumor cells. These tests show that nisin shows cytotoxic activity on K562 and U937 cell lines at 72 h of exposure, at a concentration of 40 µg / mL, which corresponds to 100 times the minimum inhibitory concentration (MIC) used for its action on bacteria. When comparing the effect of nisin on human peripheral blood mononuclear cells with lymphoid and myeloid tumor lines (K562 and U937 respectively), a selective effect of nisin on blood tumor cells is observed.


Subject(s)
Humans , Cell Line, Tumor/drug effects , Anti-Bacterial Agents/pharmacology , Nisin/pharmacology , Staphylococcus aureus/drug effects , Bacteriocins/pharmacology , In Vitro Techniques , Microbial Sensitivity Tests , Cell Survival/drug effects , K562 Cells/drug effects , U937 Cells/drug effects
4.
Int. j. high dilution res ; 20(2/3): 34-43, June 4, 2021.
Article in English | LILACS, HomeoIndex | ID: biblio-1396358

ABSTRACT

Natural killer (NK) cells are among the first in defense of the innate immune system by eliminating a variety of abnormal or stressed cells such as cancer cells or virus-infected cells. Individuals who exhibit low cytolytic NK cell activity are believed to be at higher risk of viral infection, tumorigenesis, and various other diseases of the immune system. Therefore, restoration of impaired NK cell function might be an essential step in immunostimulatory therapy of immunocompromised patients. Bacillus firmus is a non-pathogenic gram-positive bacterium of the environment, which possesses various immunomodulatory properties in vitro and in vivo. This retrospective study reports on the effect of B. firmus on the activity of NK cells in vitro. Basal cytolytic NK cell activity against tumor cells among peripheral blood mononuclear cells (PBMCs) of routine patients was determined in a standardized NK cell cytotoxicity assay. The impact of cultivation of PBMCs with B. firmus preparation Bacillus firmus e volumine ex muris cellulae (Bacillus firmus (evc)) 6x on tumor cell killing by NK cells was monitored in relation to basal NK cell activity. This study showed that stimulation of PBMCs with Bacillus firmus (evc) 6x in vitro led to a significant increase in NK cell function. Substantial improvement in cytolytic NK cell activity (more than 1.3-fold of basal activity) was much more pronounced for patients with compromised NK cell function. Due to its immunostimulatory mode of action, Bacillus firmus (evc) may be of particular importance in therapy of patients with NK cell deficiency.


Subject(s)
Killer Cells, Natural , K562 Cells , Bacillus firmus/immunology
5.
Article in Chinese | WPRIM | ID: wpr-880136

ABSTRACT

OBJECTIVE@#To investigate the regulatory effects of RBM47 on HMGA2 and the function of RBM47 in human chronic myeloid leukemia cell K562.@*METHODS@#K562 cells were transduction by the overexpressed and knockdown RBM47 lentiviral vector. CCK-8 assay was used to detect the effect of RBM47 on the proliferation of K562 cells. Flow cytometry assay was used to detect the effect of RBM47 on the cell cycle progression of K562 cells. RNA immunoprecipitation assay was used to detect the association between RBM47 and HMGA2 mRNA. RT-qPCR was used to detect the effects of RBM47 on the stability of HMGA2 mRNA. Western blot was used to evaluate the effect of RBM47 on HMGA2 protein expression.@*RESULTS@#The overexpressed RBM47 could inhibit the proliferation and cell cycle progression of K562 cells. However, the inhibitation of RBM47 could improve the proliferation and cell cycle progression of K562 cells. RBM47 combined with HMGA2 mRNA could promote the degradation of HMGA2 mRNA. Thus, the overexpressed RBM47 could decrease the expression of HMGA2 protein in K562 cells.@*CONCLUSION@#RNA binding protein RBM47 can inhibit the proliferation of K562 cells by regulating HMGA2 expression.


Subject(s)
Apoptosis , Cell Proliferation , HMGA2 Protein/genetics , Humans , K562 Cells , RNA, Messenger/genetics , RNA-Binding Proteins/genetics
6.
Article in Chinese | WPRIM | ID: wpr-880091

ABSTRACT

OBJECTIVE@#To investigate the effect of tumor necrosis factor death receptor (DR) 4 demethylation to the proliferation and apoptosis of myeloid leukemia K562 cells.@*METHODS@#The logarithmic phase of K562 cells were treated by desitabine (DCA) at 0, 0.8, 1.6 and 3.2 μmol/L, and the cells were divided into control group, DCA low dose group, DCA medium dose group and DCA high dose group respectively. The cells in control group were treated by tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) 0.5 μg/ml for 24 h, and the cells were divided into TRAIL group. The cells in DCA high dose group were treated by TRAIL 0.5 μg/ml for 24 h, and were divided into DCA high dose + TRAIL group. Methylation-specific polymerase chain reaction (MS-PCR) was used to measure the methylation status of the DR4 gene promoter in the control group and DCA low, medium and high dose groups. Real-time fluorescent quantitative polymerase chain reaction (qRT-PCR) and Western blot were used to determine the relative expression of DR4 mRNA and protein in the control group and DCA low, medium and high dose groups. Dime- thylthiazole (MTT) method was used to determine the inhibition rate of cell proliferation of the cells in control group, DCA high dose group, TRAIL group, DCA high dose + TRAIL group. Flow cytometry was used to determine the apoptotic rate of the cells in control group, DCA high dose group, TRAIL group, DCA high dose + TRAIL group.@*RESULTS@#The cells in the control group were methylation-positive, the brightness of the methylation bands of the cells in the DCA low, medium, and high dose groups was gradually decreased to disappear, and the DCA high dose group showed negative for methylation. The relative expression of DR4 mRNA and protein in the control group, DCA low, medium and high dose groups was increased sequentially (r=0.624, 0.704). The inhibition rate of cell proliferation of the cells in the control group, DCA high dose group, TRAIL group, DCA high dose + TRAIL group was increased sequentially (r=0.653, 0.754, 0.709, 0.725) at 24, 48 and 72 h.@*CONCLUSION@#DCA can reverse the methylation level of DR4 gene promoter in ML K562 cells and up-regulate the expression of DR4, which may enhance the proliferation inhibition and apoptosis promotion effects of TRAIL on K562 cells.


Subject(s)
Apoptosis , Cell Line, Tumor , Cell Proliferation , Demethylation , Humans , K562 Cells , Leukemia, Myeloid , Receptors, TNF-Related Apoptosis-Inducing Ligand/metabolism , TNF-Related Apoptosis-Inducing Ligand/metabolism
7.
Article in Chinese | WPRIM | ID: wpr-880085

ABSTRACT

OBJECTIVE@#The present study was to evaluate the anti-tumor effects of acidic RNA protein complex (FA-2-b-β) extracted from the wild edible Qinba mushroom in inducing of apoptosis and immunoregulation of tumor cell.@*METHODS@#Cell proliferation inducing rate of FA-2-b-β to K562 cell was measured using CCK-8. Apoptosis rate was detected by using flow cytometry. Chronic myeloid leukemia model was developed by tail vein injection/subcutaneous inoculation of K562 cells in NCG mice. The tumor burden of mice was observed. The general condition of the mice was monitored twice daily. The peripherivcal full blood counts of mice was tested daily. RT-qPCR and Western blot was FA-2-b-β performed to determine involvement of apoptotic-related gene and protenin, Immunofluorescence and immunohistochemistry was used to detected the expression of CD3, CD4 and CD8.@*RESULTS@#The proliferation and apoptosis of K562 cell could be inhibitied and induced by FA-2-b-β, there was 100% successful in the tumor formation in vivo, after treated by drug for 21 days there were significantly increased peripheral leucocytes, but decreased hemoglobin of mice treated by FA-2-b-β as compared with those in control group. The CD3, CD4 and CD8 showed positive in mice, and the propotation was imbalance, but it showed reserved after treated by FA-2-b-β.@*CONCLUSION@#FA-2-b-β is strong anti-leukemia effect in vitro and in vivo, suggesting the traditional Chinese medicine maybe contribute to the anti-cancer and immunoregulation research.


Subject(s)
Agaricales , Animals , Apoptosis , Cell Proliferation , Humans , K562 Cells , Leukemia, Myelogenous, Chronic, BCR-ABL Positive , Mice
8.
Article in Chinese | WPRIM | ID: wpr-880076

ABSTRACT

OBJECTIVE@#To construct an acute myeloid leukemia cell line stably expressing CD123-CLL1 so as to provide an "in vitro" model for studying the role of CD123 and CLL-1 in leukemia and the treatment targeting CD123 and CLL-1.@*METHODS@#The recombinant plasmid of lentivirus was constructed by synthesizing CD123 and CLL-1 sequences and PCR homologous recombination. The lentivirus vector was packaged by three-plasmid packaging system. After collecting the supernatant of lentivirus, the virus titer was determined by quantitative PCR. K562 leukemia cells were collected and transtected with virus supernatant. Leukemia cell line stably expressing the target gene were screened by purinomycin. The expression levels of CD123 and CLL-1 were detected by RT-PCR and flow cytometry.@*RESULTS@#The lentiviral vector was successfully constructed, and identified by agarose gel electrophoresis and gene sequencing, then the virus titer of the supernatant was up to 5.81×10@*CONCLUSION@#Lentiviral vector expressing CD123-CLL1 has been successfully constructed, and K562 leukemia cell line stably expressing CD123 and CLL-1 has been successfully obtained.


Subject(s)
Cell Line, Tumor , Genetic Vectors , Humans , Interleukin-3 Receptor alpha Subunit , K562 Cells , Lentivirus/genetics , Leukemia, Lymphocytic, Chronic, B-Cell/genetics , Plasmids , Transfection
9.
Article in Chinese | WPRIM | ID: wpr-880033

ABSTRACT

OBJECTIVE@#To explore the effects of costunolide on the proliferation and apoptosis of human chronic myeloid leukemia drug resisitant cell line K562/ADR and its mechanism.@*METHODS@#The proliferation of the cells was assessed by CCK-8 assay, while flow cytometry was used to detect the apoptosis of the cells. The related-proteins were detected by using Western blot.@*RESULTS@#The proliferation of K526/ADR cells was significantly inhibited by costunolide in a dose-dependent manner (r=0.9886) after treated by 0.01, 0.1, 0.25, 0.5, 1, 2.5, 5, 10, 25, 50 and 100 μmol/L costunolide for 72 h, and IC@*CONCLUSION@#Costunolide could inhibit the proliferation and apoptosis of K562/ADR cells through regulation of PI3K/AKT pathway.


Subject(s)
Apoptosis , Cell Proliferation , Humans , K562 Cells , Phosphatidylinositol 3-Kinases , Proto-Oncogene Proteins c-akt , Sesquiterpenes
10.
Article in English | WPRIM | ID: wpr-888781

ABSTRACT

In this study, three new germacranolide sesquiterpenes (1-3), together with six related known analogues (4-9) were isolated from the whole plant of Carpesium cernuum. Their structures were established by a combination of extensive NMR spectroscopic analysis, HR-ESIMS data, and ECD calculations. The anti-leukemia activities of all compounds towards three cell lines (HEL, KG-1a, and K562) were evaluated in vitro. Compounds 1-3 exhibited moderate cytotoxicity with IC


Subject(s)
Antineoplastic Agents, Phytogenic/pharmacology , Asteraceae/chemistry , Drug Screening Assays, Antitumor , Humans , K562 Cells , Phytochemicals/pharmacology , Sesquiterpenes, Germacrane/pharmacology
11.
Journal of Experimental Hematology ; (6): 1387-1393, 2021.
Article in Chinese | WPRIM | ID: wpr-922270

ABSTRACT

OBJECTIVE@#To investigate the inhibitory effects of novel phosphodiesterase 4 inhibitor ZL-n-91 to the proliferation of leukemia cells L1210 and K562.@*METHODS@#CCK-8 method was used to detect the effect of ZL-n-91 to the proliferation of L1210 and K562 cells, and the proliferation rate, IC@*RESULTS@#ZL-n-91 showed a significant inhibitory effect to the proliferation of leukemia cells L1210 and K562 in a dose-dependent manner (P<0.001). After treated by ZL-n-91, the leukemia cells L1210 and K562 in the S-phase in cell cycle decreased significantly compared with those in control group (P<0.01). The apoptosis of leukemia cells L1210 and K562 could be induced by ZL-n-91 (P<0.001), and the expression level of apoptosis related protein BAX significantly increased. In the animal experiment, the result showed that ZL-n-91 could significantly inhibit the growth of subcutaneously transplantation tumor (P<0.05).@*CONCLUSION@#The novel phosphodiesterase 4 inhibitor ZL-n-91 can effectively inhibit the proliferation of leukemia cells L1210 and K562, which has the potential of anti-leukemia drug development.


Subject(s)
Animals , Cell Proliferation , Humans , K562 Cells , Leukemia , Mice , Mice, Nude , Phosphodiesterase 4 Inhibitors/pharmacology
12.
Journal of Experimental Hematology ; (6): 1028-1033, 2021.
Article in Chinese | WPRIM | ID: wpr-888514

ABSTRACT

OBJECTIVE@#To investigate the role of petroleum ether extract of Rhizoma Amorphophalli (SLG) in inhibiting proliferation and promoting apoptosis and differentiation of leukemia K562 cells.@*METHODS@#K562 cells were processed by SLG and PD98059 which was the ERK signaling pathway blocker. Then cell vitality was tested by MTT. Cell apoptosis rate and positive percentage of antigen expression related with differentiation were detected by flow cytometry. The protein expression levels of ERK1/2 and pERK1/2 were detected by Western blot.@*RESULTS@#The proliferation activity of K562 was reduced by 50, 100, 200 mg/L SLG in a concentration dependent manner (r=0.9997). The apoptosis rate and positive expression rate of CD11b, CD14 and CD42b which were related with differentiation were raised by SLG, as well as the expression of pERK1/2, while PD98059 could reverse the promoting effect of SLG on apoptosis and differentiation partially.@*CONCLUSION@#SLG can inhibit the proliferation and promote apoptosis and differentiation of K562 cells through ERK signaling pathway.


Subject(s)
Alkanes , Apoptosis , Cell Proliferation , Humans , K562 Cells , Petroleum , Plant Extracts/pharmacology
13.
Journal of Experimental Hematology ; (6): 1768-1774, 2021.
Article in Chinese | WPRIM | ID: wpr-922332

ABSTRACT

OBJECTIVE@#To investigate the effect of silencing DNA methyltransferase 1(DNMT1) to the methylation of the promoter of the tumor suppressor gene wnt-1 (WIF-1) in human chronic myeloid leukemia (CML) cells.@*METHODS@#DNMT1 siRNAi plasmid was constructed and DNMT1 siRNAi was transfected into CML K562 cells. RT-PCR and Western blot were used to detect the expression of DNMT1 gene and related protein, and methylation PCR was used to detect WIF-1 gene promoter methylation level. The trypan blue exclusion and MTT assay were used to detect the cell proliferation, flow cytometry were used to detect the cell apoptosis rate, colony formation assay was used to detect cell colony formation ability. Expression of Wnt/β- catenin and its downstream signaling pathway proteins were detected by Western blot after DNMT1 gene was silenced.@*RESULTS@#The expression level of DNMT1 mRNA and its related protein in the experimental group were significantly lower than those in the control group and negative control group (P<0.05). After 72 hours of successful transfection, the WIF-1 gene in the control group and negative control group were completely methylated, while in the experimental group, the methylation level significantly decreased. The results of MSP showed that the PCR product amplified by the unmethylated WIF-1 primer in the experimental group increased significantly,while by the methylated WIF-1 primer decreased significantly. After 48 h of transfection, the OD value, viable cell number and colony formation of the cells in experimental group were significantly lower than those in the negative control group and the control group (P<0.05). The apoptosis rate of the cells in experimental group was significantly higher than those in the negative control group and control group (P<0.05). The expression levels of β- actin, myc, cyclin D1 and TCF-1 in K562 cells in the experimental group were significantly lower than those in the negative control group and control group (P<0.05).@*CONCLUSION@#Silencing DNMT1 gene can inhibit the proliferation and promote the apoptosis of K562 cells. The mechanism may be related to reverse the hypermethylation level of the WIF-1 gene promoter, thereby inhibit the activity of the Wnt/β- catenin signaling pathway.


Subject(s)
Adaptor Proteins, Signal Transducing/metabolism , DNA Methylation , Humans , K562 Cells , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/genetics , Repressor Proteins/metabolism
14.
Journal of Experimental Hematology ; (6): 1714-1718, 2021.
Article in Chinese | WPRIM | ID: wpr-922323

ABSTRACT

OBJECTIVE@#To investigate the regulation of chronic myelogenous leukemia (CML) imatinib resistant genes, in order to improve the therapeutic effect of CML imatinib resistant patients.@*METHODS@#The human CML cell line K562 and imatinib-resistant K562 cells (K562/G01) were collected, and transcriptome of the cells were achieved by RNA-seq. The sequencing data were analyzed by using standard procedures.@*RESULTS@#Compared with K562 cells, 464 genes were significantly changed in K562/G01 cells, including 163 up-regulated and 301 down-regulated genes. The GO function annotation analysis and KEGG pathway analysis results showed that the differentially expressed genes were mainly involved in biological processes such as oxidative phosphorylation, localization to protein organelle, ribonucleoprotein complex biogenesis and so on. Gene Set Enrichment Analysis (GSEA) plots showed that 5 gene-sets were up-regulated in K562/G01 significantly, including the pathway of TGF-beta, mTOR and CML.@*CONCLUSION@#CML imatinib resistance is associated with oxidative phosphorylation, during which the pathway of TGF-beta and mTOR are significantly up-regulated.


Subject(s)
Drug Resistance, Neoplasm , Gene Expression Profiling , Humans , Imatinib Mesylate/pharmacology , K562 Cells , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/genetics
15.
Journal of Experimental Hematology ; (6): 1704-1709, 2021.
Article in Chinese | WPRIM | ID: wpr-922321

ABSTRACT

OBJECTIVE@#To investigate the effect of ursane triterpenoids 3β,19α-dihydroxyursu-12-ene-23,28-dicarboxylic acid (Rotundioic acid, RA) on the sensitivity of adriamycin-resistant K562 cells (K562/ADM Cell) anti-tumor drug, and to explore the effect and mechanism of RA on the multidrug resistance of K562/ADM cells.@*METHODS@#CCK-8 method was used to detect the effect of RA on the sensitivity of K562 cells and K562/ADM cells to anti-tumor drug. Reverse transcription-polymerase chain reaction (RT-PCR) and Western blot were used to detect the expression level of mRNA and the protein in K562 and K562/ADM cells, and the effect of RA on the expression of MDR1 mRNA and P-gp in K562/ADM cells was also detected; Western blot was used to detect the expression of p-JNK, p-p38 and p-ERK1/2 in K562/ADM cells.@*RESULTS@#RA could increased the sensitivity of K562/ADM cells to adriamycin(the reversal factor was 1.61 times), the difference showed statistically significantly (P<0.05); the resistance factor of K562/ADM to ADM was 41.76 times. The expression of MDR1 mRNA in K562 cells was extremely low, and the protein product P-glycoprotein (P-gp) was almost not expressed; MDR1 mRNA and P-gp in K562/ADM cells were highly expressed; RA could down-regulate the expression levels of MDR1 and P-gp in K562/ADM cells. In addition, RA could upregulate the phosphorylation levels of p38 and ERK1/2 in K562/ADM cells, but it has no effect on the expression of p-JNK.@*CONCLUSION@#RA may participate in the regulation of MAPK signaling pathway by upregulating the expression levels of p-p38 and p-ERK1/2 in K562/ADM cells, and thus inhibit the transcription and translation levels of MDR1, and finally reverse the multidrug resistance of leukemia cells.


Subject(s)
ATP Binding Cassette Transporter, Subfamily B , ATP Binding Cassette Transporter, Subfamily B, Member 1 , Drug Resistance, Multiple , Drug Resistance, Neoplasm , Humans , K562 Cells
16.
Journal of Experimental Hematology ; (6): 1081-1085, 2020.
Article in Chinese | WPRIM | ID: wpr-827157

ABSTRACT

OBJECTIVE@#To investigate the effect of sphingosine-1-phosphate receptor 2 (S1PR2) specific antagonist JTE-013 on the proliferation of human chronic myeloid leukemia (CML) cell line K562.@*METHODS@#K562 cells were treated with JTE-013 (0, 0.5, 1, 5, 10, 20 μmol/L) for 24 and 48 hours respectively, CCK8 assay was used to detect the cell viability. K562 cells were treated with JTE-013 (0, 5, 10, 20 μmol/L) for 24 hours, propidium iodide (PI) DNA staining was used to analyze the cell cycle, Western blot was used to determine the levels of P21 and Cyclin D1 protein expression.@*RESULTS@#JTE-013 inhibited the proliferation of CML cell line K562 in a dose dependent manner (r=-0.971). The proliferation rate of CML cells showed that the activity of CML cells decreased gradually with the increase of JTE-013 concentration (r=-0.971). The detection demonstrated that JTE-013 suppressed tumor cell proliferation through cell cycle arrest in G/G phase. Further detection of the protein expressions of G phase regulators showed that level of P21 increased, and expression of Cyclin D1 decreased.@*CONCLUSION@#JTE-013, a S1PR2 antagonist, can inhibit the proliferation of human CML K562 cells, which may be achieved by arresting the cells in G/G phase.


Subject(s)
Apoptosis , Cell Proliferation , Humans , K562 Cells , Leukemia, Myelogenous, Chronic, BCR-ABL Positive , Pyrazoles , Pyridines , Receptors, Lysosphingolipid , Sphingosine-1-Phosphate Receptors
17.
Journal of Experimental Hematology ; (6): 1096-1104, 2020.
Article in Chinese | WPRIM | ID: wpr-827155

ABSTRACT

OBJECTIVE@#To compare the expression of miR-199a-5p between ADM-resistant AML cell (K562/ADM)and ADM-sensitive AML cell (K562), and to investigate the effect of miR-199a-5p on regulating AML drug resistance as well as its molecular mechanism.@*METHODS@#MTT method was used to detect the proliferation inhibition effect of ADM on K562 and K562/ADM cells, the IC was calculated. miR-199a-5p expression in cell lines (K562 and K562/ADM) and bone marrow sample (refractory/relapsed AML patients and complete remission AML patients) was detected by RT-qPCR. K562/ADM and K562 cells were transfected by miR-199a-5p mimic and miR-199a-5p inhibitor respectively to ensure that miR-199a-5p expression in K562/ADM cells was increased and that in K562 cells was decreased. Then proliferation inhibition effect of ADM on both cells was detected by CCK-8 and mRNA and protein DRAM1 expression in both cells was measured by real time RT-PCR and Western blot respectively. Dual luciferase reporter assay was used to detect wether there were direct binding sites between miR-199a-5p and DRAM1 3' UTR. CCK-8 was used to measure the proliferation inhibition effect of ADM on K562/ADM cells when DRAM1 was downregulated by siRNA.@*RESULTS@#The IC of ADM for K562/ADM and K562 cells was 146.14±0.079 and 3.08±0.056 μg/ml respectively. As compared with patients in complete remission group, MiR-199a-5p expression in refractory/ relapsed AML patients significantly decreased, and the MiR-199a-5p expression in K562/ADM cells was also dramatically downregulated, compared with K562 cells (P<0.05). When the expression of miR-199a-5p was upregulated in K562/ADM cells, the proliferation inhibition effect of ADM on cells elevated and both DRAM1 mRNA and protein expressions decreased. Conversely, when miR-199a-5p expression was downregulated in K562 cells, the proliferation inhibition effect of ADM on cells obviously reduced and both DRAM1 mRNA and protein expression increased (P<0.05). Dual luciferase reporter Assay showed a direct interaction between miR-199a-5p and its binding site within DRAM1 mRNA. Both DRAM1 mRNA and protein expression in K562/ADM were markedly higher than those in K562 cells (P<0.05). The ADM chemosensitivity of K562/ADM cells was improved significantly when DRAM1 expression was downregulated (P<0.05).@*CONCLUSION@#miR-199a-5p is downregulated in chemoresistant AML cells. miR-199a-5p expression plays an important role in regulating the sensitivity of AML cells to ADM treatment. DRAM1 is a functional target gene for miR-199a-5p modulating AML chemoresistance.


Subject(s)
Animals , Doxorubicin , Humans , K562 Cells , Leukemia, Myeloid, Acute , Male , MicroRNAs , RNA, Messenger
18.
Journal of Experimental Hematology ; (6): 1137-1143, 2020.
Article in Chinese | WPRIM | ID: wpr-827150

ABSTRACT

OBJECTIVE@#To investigate the effects of CPEB4 on the migration and cycle of K562 cells and the changes of protein molecules that may be involved in the regulatory mechanism.@*METHODS@#Western blot was used to detect the expression of CPEB4 in normal leukocytes and K562 cells. The overexpression plasmid pcDNA3.1(+)-His-CPEB4, silencing plasmid pPLK+Puro-CPEB4 shRNA were transfected into K562 cells by electroporation so as to change CPEB4. The transfection efficiency was detected by Western blot. Finally, the migration and cycle of different cells were detected by Transwell chamber and flow cytometry.Western blot was used to detect the expression changes of MMP2, MMP9, CDK4, CyclinD1 and P21 proteins.@*RESULTS@#Compared with normal white blood cells, the expression of CPEB4 protein in K562 cells was significantly enhanced (P<0.01); Compared with the control group, CPEB4-silenced K562 cells showed that the cell migration ability was significantly enhanced (P<0.01); G/G phase cell ratio reduced, G/M phase cell ratio increased, and cell cycle progression accelerated(P<0.01), The expression levels of MMP2 (P<0.05), MMP9 (P<0.05), CDK4 (P<0.01), CyclinD1 (P<0.01) proteins increased significantly. The expression level of P21 protein significantly decreased (P<0.01). The migration ability of K562 cells after CPEB4 overexpression was decreased (P<0.01), the cell ratio of G/G phase in the cell cycle increased, the cell proportion of S phase decreased and the cell cycle progression was arrested at G/G phase (P<0.01). The expression of P21 protein increased, MMP2 , MMP9, CDK4, CyclinD1 protein expression decreased significantly(P<0.05-0.01).@*CONCLUSION@#CPEB4 can inhibit the migration of K562 cells and arrest cell cycle progression at G/G phase. Its mechanism may be related with regulating the exprossion of MMP2, MMP9, CDK4, CyclinD1 and P21 proteins.


Subject(s)
Apoptosis , Cell Cycle , Cell Proliferation , Humans , K562 Cells , Leukemia, Myelogenous, Chronic, BCR-ABL Positive , RNA-Binding Proteins
19.
Journal of Experimental Hematology ; (6): 1167-1170, 2020.
Article in Chinese | WPRIM | ID: wpr-827145

ABSTRACT

OBJECTIVE@#To investigate the effect of chidamide on the killing activity of NK (Natural killer cell, NK) cells targeting K562 cells and its related mechanism.@*METHODS@#K562 cells were pretreated with chidamide at different concentrations and cocultured with NK cells at different effect-target ratios. The killing effect of chidamide on K562 cells by NK cells, the expression of natural killer group 2 member D (NKG2D) ligands and apoptosis rate of K562 cells were detected by flow cytometry.@*RESULTS@#The killing sensitivity of NK cells to K562 cells could be enhanced by chidamide. The expression of ULBP2 on K562 cell surface could be up-regulate, however, the expression of ULBP1 and MICA/MICB showed no statistically difference as compared with control group. Chidamide showed no obvious cytotoxicity to K562 cells.@*CONCLUSION@#Chidamide can significantly improve killing efficiency of NK cells on K562 cells, which may be related to the up-regulation of ULBP2 expression.


Subject(s)
Aminopyridines , Benzamides , GPI-Linked Proteins , Histocompatibility Antigens Class I , Humans , Intercellular Signaling Peptides and Proteins , K562 Cells , Killer Cells, Natural , Allergy and Immunology , NK Cell Lectin-Like Receptor Subfamily K
20.
Article in English | WPRIM | ID: wpr-880597

ABSTRACT

OBJECTIVES@#To investigate the effect of adriamycin (ADM), idelalisib or ADM and their combination on cell proliferation and intracellular concentration of ADM, and to explore the reversal effect of idelalisib on drug resistance to ADM.@*METHODS@#The K562 and K562/ADM cells were respectively treated with ADM and idelalisib at different concentrations. The 50% inhibitory concentration (IC@*RESULTS@#The cell survival rates were significantly decreased in a dose-dependent manner when the cells were treated with different doses of ADM (0.001-10.000 mg/L ). The IC@*CONCLUSIONS@#Idelalisib exerts effect on inhibition of the proliferation in myeloid leukemia K562 and K562/ADM cells, which may partially reverse the drug resistance of K562/ADM cells to ADM. The mechanisms for the effect of idelalisib may be related to increasing the accumulation of ADM and inducing the cell apoptosis in the K562 and K562/ADM cells.


Subject(s)
ATP Binding Cassette Transporter, Subfamily B, Member 1 , Cell Proliferation , Doxorubicin/pharmacology , Drug Resistance, Multiple , Drug Resistance, Neoplasm , Humans , K562 Cells , Leukemia, Myeloid , Purines , Quinazolinones
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