Effects of Methionine Restriction on Proliferation, Cell Cycle, and Apoptosis of Human Acute Leukemia Cells / 中国实验血液学杂志

OBJECTIVE@#To investigate the effects of methionine restriction on proliferation, cell cycle and apoptosis of human acute leukemia cells.@*METHODS@#Cell Counting Kit-8 (CCK-8) assay was used to detect the effect of methionine restriction on HL-60 and Jurkat cells proliferation. The effect of methionine restriction on cell cycle of HL-60 and Jurkat cells was examined by PI staining. Annexin V-FITC / PI double staining was applied to detect apoptosis of HL-60 and Jurkat cells following methionine restriction. The expression of cell cycle-related proteins cyclin B1, CDC2 and apoptosis-related protein Bcl-2 was evaluated by Western blot assay.@*RESULTS@#Methionine restriction significantly inhibited the proliferation of HL-60 and Jurkat cells in a time-dependent manner (HL-60: r =0.7773, Jurkat: r =0.8725), arrested the cells at G2/M phase (P < 0.001), and significantly induced apoptosis of HL-60 and Jurkat cells (HL-60: P < 0.001; Jurkat: P < 0.05). Furthermore, Western blot analysis demonstrated that methionine restriction significantly reduced the proteins expression of Cyclin B1 (P < 0.05), CDC2 (P < 0.01) and Bcl-2 (P < 0.001) in HL-60 and Jurkat cells.@*CONCLUSION@#Acute leukemia cells HL-60 and Jurkat exhibit methionine dependence. Methionine restriction can significantly inhibit the proliferation, promote cell cycle arrest and induce apoptosis of HL-60 and Jurkat cells, which suggests that methionine restriction may be a potential therapeutic strategy for acute leukemia.

Study on the Regulation of Chidamide on CD8+T Cells in T-cell Acute Lymphoblastic Leukemia / 中国实验血液学杂志

OBJECTIVE@#To explore the regulatory effect of chidamide on CD8+ T cells in T-cell acute lymphoblastic leukemia.@*METHODS@#The expression levels of CXCL9 and CXCL3 mRNA in Jurkat cells, lymphocytes treated with chidamide and lymphocytes co-cultured with chidamide-treated Jurkat cells were detected by fluorescence quantitative PCR. The proportion of CD8+ T cells in lymphocytes treated with chidamide and lymphocytes co-cultured with chidamide-treated Jurkat cells was determined by flow cytometry.@*RESULTS@#Chidamide upregulated CXCL9 mRNA expression in Jurkat cell line in a dose-dependent manner (r=0.950). The mRNA expression of CXCL9 in chidamide 5 μmol/L group was 164 times higher than that in control group. Chidamide upregulated CXCL9 mRNA expression in lymphocytes, but the up-regulated level was significantly lower than that in Jurkat cell line treated with the same concentration of chidamide. Co-culture with chidamide treated Jurkat cells upregulated the proportion of CD8+ T cells in lymphocytes.@*CONCLUSION@#In T-cell acute lymphoblastic leukemia, chidamide may increase the concentration of CXCL9 in the tumor microenvironment by up-regulating the expression of CXCL9 in tumor cells, leading to an increase in the number of CD8+ T cells.

Deficiency of two-pore segment channel 2 contributes to systemic lupus erythematosus via regulation of apoptosis and cell cycle / 中华医学杂志(英文版)

BACKGROUND@#Systemic lupus erythematosus (SLE) is a complex autoimmune disease, and the mechanism of SLE is yet to be fully elucidated. The aim of this study was to explore the role of two-pore segment channel 2 (TPCN2) in SLE pathogenesis.@*METHODS@#Quantitative reverse transcription polymerase chain reaction (qRT-PCR) was used to detect the expression of TPCN2 in SLE. We performed a loss-of-function assay by lentiviral construct in Jurkat and THP-1 cell. Knockdown of TPCN2 were confirmed at the RNA level by qRT-PCR and protein level by Western blotting. Cell Count Kit-8 and flow cytometry were used to analyze the cell proliferation, apoptosis, and cell cycle of TPCN2-deficient cells. In addition, gene expression profile of TPCN2-deficient cells was analyzed by RNA sequencing (RNA-seq).@*RESULTS@#TPCN2 knockdown with short hairpin RNA (shRNA)-mediated lentiviruses inhibited cell proliferation, and induced apoptosis and cell-cycle arrest of G2/M phase in both Jurkat and THP-1 cells. We analyzed the transcriptome of knockdown-TPCN2-Jurkat cells, and screened the differential genes, which were enriched for the G2/M checkpoint, complement, and interleukin-6-Janus kinase-signal transducer and activator of transcription pathways, as well as changes in levels of forkhead box O, phosphatidylinositol 3-kinase/protein kinase B/mechanistic target of rapamycin, and T cell receptor pathways; moreover, TPCN2 significantly influenced cellular processes and biological regulation.@*CONCLUSION@#TPCN2 might be a potential protective factor against SLE.

Seed oil of Brucea javanica induces apoptosis through the PI3K/Akt signaling pathway in acute lymphocytic leukemia Jurkat cells / 中国天然药物

Brucea javanica oil emulsion (BJOE) has been used to treat tumor in China for more than 40 years. However, its components and effectiveness in the treatment of acute lymphocytic leukemia (ALL) and its mechanism of anti-cancer activity remain unknown. In the current study, high-performance liquid chromatography-evaporative light scattering detector (HPLC-ELSD) was used to analyze the components of BJOE. Then, the anti-leukemia effects of BJOE were examined both in vitro and in vivo using ALL Jurkat cells and the p388 mouse leukemia transplant model, respectively. The primary ALL leukemia cells were also used to confirm the anti-leukemia effects of BJOE. The apoptotic-related results indicated that BJOE induced apoptosis in Jurkat cells and were suggestive of intrinsic apoptotic induction. Moreover, BJOE inhibited Akt (protein kinase B) activation and upregulated its downstream targets p53 and FoxO1 (forkhead box gene, group O-1) to initiate apoptosis. The activation of GSK3β was also involved. Our findings demonstrate that BJOE has anti-leukemia effects on ALL cells and can induce apoptosis in Jurkat cells through the phosphoinositide3-kinase (PI3K) /Akt signaling pathway.

Effect of Down-Regulation of LncRNA-HOTAIRM1 to Proliferation, Apoptosis and KIT/AKT Pathway of Jurkat Cells / 中国实验血液学杂志

OBJECTIVE@#To observe the effects of down-regulation of long non-coding RNA HOX antisense intergenic RNA myeloid 1 (LncRNA-HOTAIRM1) to the proliferation and apoptosis of Jurkat in human leukemia T lymphocytes, and explore its mechanism.@*METHODS@#Jurkat cells were cultured in vitro and randomly divided into control group, HOTAIRM1 siRNA-NC group and HOTAIRM1 siRNA group; the expressions of LncRNA-HOTAIRM1 mRNA, KIT receptor tyrosine kinase (KIT) mRNA and serine threonine kinase (AKT) mRNA in Jurkat cells were detected by real-time fluorescence quantification (RT-qPCR); the proliferation of Jurkat cells in each groups was detected by CCK-8 method; the apoptosis of Jurkat cells in each groups was detected by Annexin V-FITC/PI double staining; the expressions of KIT, AKT, p-KIT, p-AKT, B-lymphoma-2 gene (BCL-2) and Caspase-3 were detected by Western blot.@*RESULTS@#Compared with the cells in the control group and HOTAIRM1 siRNA-NC group, the expression level of LncRNA-HOTAIRM1 mRNA, cell survival rate, expression levels of KIT mRNA, AKT mRNA, p-KIT, p-AKT and BCL-2 proteins in Jurkat cells in HOTAIRM1 siRNA group were significantly lower (P<0.05), while the expression level of Cleared Caspase-3 protein and Jurkat cell apoptosis rate were significantly higher (P<0.05).@*CONCLUSION@#LncRNA-HOTAIRM1 may inhibit Jurkat cell proliferation and induce apoptosis through KIT/AKT signaling pathway.

Effect of Regulating A20 Expression on NF-κB Expression and Biological Characteristics of Jurkat Cells / 中国实验血液学杂志

OBJECTIVE@#To explore the effect of regulating A20 expression on NF-κB and biological characteristics of Jurkat cells with glucocorticoid (GC) resistance.@*METHODS@#CCRF CEM and Jurkat cells were treated with dexamethasone (DEX) at concentrations of 100、10、1、0.1、0.01 and 0.001 μmol/L, and cultured for 24、48 and 72 h. The proliferation inhibition rate of Jurkat cell was detected by CCK-8. A20 plasmid was constructed, A20-siRNA was designed and synthesized, and transfected into Jurkat cells by liposome. CCK-8 was used to detect the proliferation rates of Jurkat cells in different concentrations of DEX group, DEX combined with A20 plasmid group and A20-siRNA group. The mRNA expression level of NF-κB was detected by RT-qPCR, the protein expression level of NF-κB was detected by Western blot, and the apoptosis of Jurkat cells was examined by flow cytometry.@*RESULTS@#The inhibitory effects of DEX at different concentrations on the growth of CCRF CEM cells were time-dependent (r=0.984, P＜0.05) and concentration-dependent (r=0.966, P＜0.05). At the point of 24 hour, the IC approached 1 μmol/L in CCRF CEM cells. Great large differences began to appear between 1 and 10 μmol/L, the proliferation rate of Jurkat cells treated with 1 μmol/L DEX did not show a significant change. Therefore, 1 μmol/L was selected as control group. The cell proliferation rate of A20 plasmid transfection combined with different concentrations of DEX group was lower than that of DEX group and A20-siRNA combined with DEX group. After transfection of A20 plasmid, the expression level of NF-κB was significantly lower than that of control group (P＜0.05), and the apoptotic rate was significantly higher than that of control group (P＜0.05). After transfection of Jurkat cells with A20-siRNA, the expression level of NF-κB was significantly higher than that of control group (P＜0.05). The apoptotic rate of cells in A20-siRNA group was not significantly changed (P＞0.05).@*CONCLUSION@#Jurkat cells are resistant to DEX. A20 overexpression combined with DEX can increase sensitivity of Jurkat cells with GC resistance and decrease the proliferation rate of Jurkat cells, down-regulate the expression level of NF-κB and promote the apoptosis of Jurkat cells.

Dihydroartemisinin Induces Apoptosis of Human Acute T Lymphocytic Leukemia Cells by Activating Oxidative Stress / 中国实验血液学杂志

OBJECTIVE@#To investigate the effects of dihydroartemisinin (DHA) on the proliferation and apoptosis of human T-cell acute lymphoblastic leukemia (T-ALL) Jurkat cell.@*METHODS@#The effects of DHA on the proliferation of Jurkat cells and the recovery of DHA-inhibited cell viability by N-acetyl-L-cysteine (NAC) were examined by CCK-8 assay. Flow cytometry was performed to analyze the cell apoptosis and generation of reactive oxygen species (ROS). Western-blot was used to detected protein expression of DNA damage-related genes, as well as apoptosis-associated genes, respectively.@*RESULTS@#DHA inhibited the proliferation of Jurkat cells, and shows a concentration-dependent manner(r =0.936), and NAC could partially restore the activity of DHA on cell proliferation inhibition. With the increase of drug concentration, the apoptosis rate (r =0.946) and ROS accumulation was increased (r =0.965). Western blot showed that the protein expressions of DNA damage-related gene γ-H2AX and apoptosis-related genes p53, c-Caspase3, BAX and cPARP were significantly increased, and BCL-2 protein expression was decreased.@*CONCLUSION@#DHA can induce ROS production in Jurkat cells, which can cause DNA damage, activate the P53 apoptotic pathway, and promote apoptosis of cells.

Effect of Down-Regulating the CD59 by RNAi Lentivirus on the Expression of Acute T-lineage Leukemia Jurkat Cell Line / 中国实验血液学杂志

OBJECTIVE@#To analyze the effect of down-regulating the CD59 gene expression by RNAi lentivirus as vector on Jurkat cell line of acute T-lineage leukemia.@*METHODS@#The expression of CD59 in Jurkat cell line of acute T-line leukemia was induced to decrease by RNAi lentivirus as vector. The transfection of RNA lentivirus and the localization of CD59 molecule were analyzed by laser confocal technique. The relative expression of CD59 gene in blank control, negative control and RNAi lentivirus transfected group was detected by real-time fluorescence quantitative PCR, and the enzyme-linked immunosorbent assay was used to detect the expression of TNF-β and IL-3 in supernatants of cultured cells in 3 groups. The expression levels of apoptosis-related molecules including Caspase-3, Survivin, BCL-2 and BCL-2-associated X protein (BAX) were measured by Western blot.@*RESULTS@#The transfection efficiency for Jurkat cells was higher than 90%. CD59 was mainly located on the cell membrane. Compared with the blank control group and the negative control group, the expression level of CD59 mRNA and protein in the RNAi lentivirus transfected group significantly decreased (P<0.05). Compared with the blank control group and the negative control group, the expression of TNF-β and IL-3 in the RNAi lentivirus transfected group were significantly higher and lower (P<0.05) respectively. The expression levels of Survivin and BCL-2 in the RNAi lentivirus transfected group were significantly lower than those in the blank control group and the negative control group, while the expression levels of Caspase-3 and BAX in the RNAi lentivirus transfected group were significantly higher than those in the blank control group and the negative control group (P< 0.05).@*CONCLUSION@#The down-regulation of CD59 gene expression induced by RNAi lenti-virus can decrease the expression of proliferation and differentiation-promoting molecule such as IL-3 and increase the expression of TNF-related factor in Jurkat cell line of acute T-lineage leukemia, which also can increase the expression of apoptosis-related proteins such as Caspase-3 and BAX, and decrease the expression of anti-apoptosis-related proteins such as Survivin and BCL-2.

Apoptosis-Inducing Effect of Ginsenoside Rh2 on Human Acute T Lymphoblastic Leukemia Jurkat Cells and Its Mechanism / 中国实验血液学杂志

OBJECTIVE@#To investigate the apoptosis-inducing effect of Ginsenoside (Rh2) on human acute T lymphoblastic leukemia Jurkat cells and it mechamism.@*METHODS@#The effects of different concentration of Rh2 (0, 10 , 20, 40 and 80 µg/ml) on the proliferation activity of Jurkat cells were detected by methyl thiazolyl tetrazolium (MTT) method, and the semi-inhibitory concentration (IC) of Rh2 on Jurkat cells at 48 h was calculated. Microscopy and Hoechst 33258 fluorescence staining were used to observe the apoptosis of Jurkat cells treated with IC Rh2 for 48 h. And then, the cell experiment was divided into 4 groups: control, Rh2 (IC), PI3K inhibitor LY294002 (50 µmol/l) and Rh2 (IC) + LY294002 (50 µmol/l). After synchronous culture for 48 h, the apoptosis and cycle changes of Jurkat cells were detected by using PI single staining and Annexin V-FITC/PI double staining, respectively. Western blot was used to detect the expression level of apoptosis-related protein BAX, BCL-2, Cleaved-Caspasase 3, cell cycle-related protein Cyclin D1 and PI3K/AKT signaling pathway-related protein AKT and p-AKT.@*RESULTS@#Rh2 (10-80 µg/ml) inhibited the Jurkat cell proliferation in a dose-time dependent manner (r = 0.999, P＜0.01; r = 0.991; P＞0.05), accompanied by obvious morphological changes of apoptosis cells. Flow cytometry showed that compared with the control group, the cell apoptosis rate in Rh2 or LY294002 group significantly increased, and the cell cycle was mostly blocked in G0/G1 phase. However, the cell apoptosis and cell cycle block in Rh2+LY294002 group were more significant than that in Rh2 and LY294002 group. Western blot showed that compared with the control group, Rh2 significantly promoted the expression of BAX and Cleaved-Caspasase 3, inhibited the expression of BCL-2, Cyclin D1 and p-AKT, furthermore LY294002 significantly promoted this effect.@*CONCLUSION@#Rh2 can induce the apoptosis of Jurkat cells in time-dose dependent manner, moreover, Rh2 also can result in an obvious block of Jurkat cells at G0/G1, that may be closely related to a series of apoptotic signaling cascades mediated by Rh2 inhibiting PI3K/AKT pathway.

β-arrestin 1 Promotes Senescence of Acute Lymphoblastic Leukemia Jurkat Cells / 中国实验血液学杂志

OBJECTIVE@#To investigate the effect of β-arrestin1 gene on senescence of T-ALL cells and its possible mechanism.@*METHODS@#The bone marrow specimens of T-ALL patients and controls were collected, the expression of β-arrestin1 and β-arrestin1 in the T-ALL patients was detected by RT-PCR and Western blot, respectively, and the relation of β-arrestin1 expression with the clinical pathologic characteristics and the prognosis of T-ALL patients was analyzed statistically. The stable Jurkat cell line with knocked down or overexpressed β-arrestin1 was constructed, the CCK method was used to detect the Jurkat cell number, the β-gal staining was used to analyze the effect of β-arrestin1 on senescence of Jurkat cells, the cross analysis of RNA-Seg data and KEGG data was performed for screening the possible signaling pathway, and Western blot was performed for varifying the key sites of signaling pathway.@*RESULTS@#The β-arrestin1 expression in specimens of T-ALL patients decreased (P＜0.01), moreover the β-arrestin1 expression negatively related with peripheral blood cell number (r=-0.601), the blasts in peripheral blood (r=-0.516) and extramedullary infiltration (r=-0.359), while positively related with the response to chemotherapy (r=0.393). The detection of stable Jurkat cell line with knocked-down and overexpressed β-arrestin1 found that the β-arrestin 1 could decrease the Jurkat cell number and accelarate the senescence of Jurkat cells (P＜0.05). The cross analysis of RNA-Seg data and KEGG data showed that the senescence of T-ALL cells may be regulated via RAS-P16-PRb-E2F1 by β-arrestin 1. Western bolt confirmed that β-arrestin1 promoted the expression of Ras and p16, and decreased the expression of pRB and E2F1 (P＜0.05).@*CONCLUSIONS@#β-arrestin1 accelerates the senescence of Jurkat cells via Ras-p16-pRb-E2F1, and delays the progression in T-ALL, which may provide a new hypothesis for the pathogenesis of T-ALL.

Advances in application of Jurkat cell model in research on infectious diseases / 中国当代儿科杂志

Infectious diseases can be caused by multiple pathogens, which can produce specific immune response in human body. The immune response produced by T cells is cellular immunity, which plays an important role in the anti-infection process of human body, and can participate in immunological protection and cause immunopathology. The outcome of various infectious diseases is closely related to cellular immune function, especially the function of T cells. Jurkat cells belong to the human acute T lymphocyte leukemia cell line. Jurkat cell model can simulate the function T lymphocytes, so it is widely used in the in vitro studies of T cell signal transduction, cytokines, and receptor expression, and can provide reference and guidance for the treatment of various infectious diseases and the research on their pathogenesis. The Jurkat cell model has been widely used in the in vitro studies of viral diseases and atypical pathogens, but parasitic infection studies using the Jurkat cell model are still rare. This article reviews advances in the application of Jurkat cell model in the research on infectious diseases.

Immunomodulatory effects of ethanol extract of germinated ice plant (Mesembryanthemum crystallinum) / 한국실험동물학회지

The purpose of this study was to investigate the immunomodulatory activity of ice plant (Mesembryanthemum crystallinum) extract (IPE) in vitro and in vivo. Raji (a human B cell line) and Jurkat (a human T cell line) cells were treated with various doses of IPE and cell proliferation was measured by WST assay. Results showed that IPE promoted the proliferation of both Raji and Jurkat cells in a dose-dependent manner. IPE also enhanced IL-6 and TNF-α production in macrophages in the presence of lipopolysaccharide (LPS), although IPE alone did not induce cytokine production. Moreover, IPE treatment upregulated iNOS gene expression in macrophages in a time- and dose-dependent manner and led to the production of nitric oxide in macrophages in the presence of IFNγ. In vivo studies revealed that oral administration of IPE for 2 weeks increased the differentiation of CD4+, CD8+, and CD19+ cells in splenocytes. These findings suggested that IPE has immunomodulatory effects and could be developed as an immunomodulatory supplement.

IL-12 and IL-23 Production in Toxoplasma gondii- or LPS-Treated Jurkat T Cells via PI3K and MAPK Signaling Pathways

IL-12 and IL-23 are closely related in structure, and have been shown to play crucial roles in regulation of immune responses. However, little is known about the regulation of these cytokines in T cells. Here, we investigated the roles of PI3K and MAPK pathways in IL-12 and IL-23 production in human Jurkat T cells in response to Toxoplasma gondii and LPS. IL-12 and IL-23 production was significantly increased in T cells after stimulation with T. gondii or LPS. T. gondii and LPS increased the phosphorylation of AKT, ERK1/2, p38 MAPK, and JNK1/2 in T cells from 10 min post-stimulation, and peaked at 30–60 min. Inhibition of the PI3K pathway reduced IL-12 and IL-23 production in T. gondii-infected cells, but increased in LPS-stimulated cells. IL-12 and IL-23 production was significantly reduced by ERK1/2 and p38 MAPK inhibitors in T. gondii- and LPS-stimulated cells, but not in cells treated with a JNK1/2 inhibitor. Collectively, IL-12 and IL-23 production was positively regulated by PI3K and JNK1/2 in T. gondii-infected Jurkat cells, but negatively regulated in LPS-stimulated cells. And ERK1/2 and p38 MAPK positively regulated IL-12 and IL-23 production in Jurkat T cells. These data indicate that T. gondii and LPS induced IL-12 and IL-23 production in Jurkat T cells through the regulation of the PI3K and MAPK pathways; however, the mechanism underlying the stimulation of IL-12 and IL-23 production by T. gondii in Jurkat T cells is different from that of LPS.

Oridonin inhibits proliferation of Jurkat cells via the down-regulation of Brg1 / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To investigate the effect of oridonin on the human acute lymphocytic leukemia cell line Jurkat and its mechanism.</p><p><b>METHODS</b>Jurkat cells were cultured in vitro and treated with various concentrations (0, 1.25, 2.5, 5, and 10 μmol/L) of oridonin for different lengths of time (24, 48, and 72 hours). The proliferation of Jurkat cells was analyzed by MTT assay. The changes in nuclear morphology were evaluated by fluorescence microscopy at 12 hours after treatment with various concentrations of oridonin. The expression levels of Brg1, P53, and C-myc were determined by semi-quantitative Western blot in Jurkat cells treated with various concentrations of oridonin for 24 hours or 5 μmol/L oridonin for various lengths of time (0, 2, 6, 12, and 24 hours). The expression levels of P53 and C-myc and proliferation of Jurkat cells were evaluated after Brg1 expression was knocked down by Brg1-specific siRNA.</p><p><b>RESULTS</b>Compared with the control group, the proliferation of oridonin-treated Jurkat cells was significantly inhibited in a concentration- and time-dependent manner (P<0.05). According to the florescence microscopic analysis, oridonin treatment led to nuclear pyknosis in Jurkat cells. Compared with the control group, Jurkat cells treated with 5 μmol/L oridonin had reduced expression of Brg1 and C-myc but elevated expression of P53. Brg1 knock-down led to a significant reduction in proliferation of Jurkat cells (P<0.05), up-regulated expression of P53, and down-regulated expression of C-myc.</p><p><b>CONCLUSIONS</b>Oridonin can inhibit the proliferation of Jurkat cells, probably via the Brg1 signaling pathway.</p>

Expression of β-integrin family members in children with T-cell acute lymphoblastic leukemia / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To study the expression of β-integrin family members in children with T-cell acute lymphoblastic leukemia (T-ALL) and their significance.</p><p><b>METHODS</b>Quantitative real-time PCR analyses were performed to assess the expression levels of β-integrin family members in bone marrow samples from 22 children with newly-diagnosed T-ALL and 21 controls (16 children with non-malignant hematologic disease and 5 healthy donors with bone marrow transplantation). Jurkat cells were treated with integrin inhibitor arginine-glycine-aspartate (Arg-Gly-Asp, RGD) peptide. The cell viability and apoptosis rate were determined by CCK8 assay and flow cytometry respectively.</p><p><b>RESULTS</b>The mRNA levels of integrins β, β, and βwere significantly lower in children with T-ALL than in controls (P<0.05). In T-ALL patients, high integrin βexpression was associated with lower white blood cell counts (<100×10/L), minimal residual disease (MRD) positivity, and day 33 bone marrow negative remission (P<0.05). In T-ALL patients, higher integrin βexpression was associated with relapse of T-ALL (P<0.05). Based on survival curve analysis, higher integrin βexpression was related to lower event-free survival and overall survival rates. RGD peptide treatment inhibited the proliferation of Jurkat cells and increased their apoptosis rate (P<0.05).</p><p><b>CONCLUSIONS</b>β-Integrin may play a role in the occurrence and development of T-ALL by affecting cell proliferation and apoptosis. The expression of integrin β5 is closely related to the risk of relapse of T-ALL. The expression of integrin β3 is closely related the treatment response and prognosis of T-ALL.</p>

Effect of BRD4 Inhibitor JQ1 on Proliferation Inhibition and Apotosis Induction in Jurkat Cells / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To investigate the effect and possible mechanism of bromo-domain inhibitors (JQ1) on proliferation inhibition and inducing apoptosis of acute T lymphocyte leukemia cell line (Jurkat) .</p><p><b>METHODS</b>Jurkat cell line was treated by JQ1 at different concentrations. MTT was used to detect the cell proliferation inhibition rate. The flow cytometry with AnnexinV-FITC/PI fluorescence staining was used to detect the changes of apoptosis rate, and real-time fluorescent quantitative PCR was used to detect c-Myc/Notch1 gene expression levels.</p><p><b>RESULTS</b>With the increasing of drug concentration and prolonging of time, the inhibitory rate of Jurkat cell growth was enhanced in time-dose dependent manner; Jurkat cells was treated by 0.8, 1.6, and 4 µ mol/LJQ1 for 48 h and 72 h, the cell apoptosis rate was enhanced with the increase of drug concentration and prolonging of time, and the difference was statistically different in comparison with the control group(P<0.05); PCR detection indicated that Notch1 and c-Myc mRNA expression was reduced in 48 h after JQ1 treatment, which was statistically different from the control group,(P<0.05) .</p><p><b>CONCLUSION</b>JQ1 can effectively inhibit the growth of Jurkat cell line, and potentially induce apoptosis through Notch1 and c-Myc gene. Hence JQ1 may be one of new methods used to treat T-ALL.</p>

Effect of Tyrosine Phosphorylation Sites of Oncogenic Protein NPM-ALK on Cell Cycle and Its Related Mechanisms / 中国实验血液学杂志

<p><b>UNLABELLED</b>Objective：To explore the effect of tyrosine phosphorylation sites Tyr644 and Tyr664 in oncogenic protein NPM-ALK on cell cycle and its related mechanisms.</p><p><b>METHODS</b>Transiently transfected 293T cells and stably transfected Jurkat cells were used for analysis of cell cycle and protein after the transfection with the constructed recombinant plasmid pEGFP-N1, pEGFP-N1-NPM-ALK and pEGFP-N1-NPM-ALK(644, 664); soft agar assay for colony formation was performed to examine the different carcinogenicity of stable cell lines; cell viability of stable cell lines was examined by CCK-8 after the treatment with PPP.</p><p><b>RESULTS</b>The S arrest occurred in both NPM-ALK(644,664) transfected 293T and Jurkat cells; the susceptibility of NPM-ALK transfected Jurkat cells to PPP was highest among the 3 stable cell lines; the phosphorylated levels of AKT, ERK and STAT3 were decreased in NPM-ALK(644,664) cells compared with the NPM-ALK ones. Additionally, the double mutation induced the increase of CDK2 and the decrease of P27 (P<0.05).</p><p><b>CONCLUSION</b>The mutation of Tyr644 and 664 sites in NPM-ALK can induce cell cycle arrest in S phase and lower susceptibility to PPP that may be related with the phosphorylation change of cell growth related molecules in the downstream of NPM-ALK.</p>

Promoter methylation status of SFRP genes and induced apoptosis by demethylation in Jurkat cells / 中华血液学杂志

<p><b>OBJECTIVE</b>To study the promoter methylation status of SFRP genes and the effect of 5- aza- 2'- deoxycytidine (5- Aza- CdR)induced apoptosis via Wnt/β- catenin pathway by demethylation in Jurkat cells.</p><p><b>METHODS</b>Jurkat cells were treated with different concentrations of 5- Aza- CdR. The cell proliferation level of Jurkat cells was detected by MTT assay. Apoptosis was evaluated by flow cytometry. Methylation- spcific PCR (MSP) was used to determine the methylation status of SFRP genes. The expressions of SFRP genes were detected by real time fluorescence quantitative PCR. The mRNA expression levels of survivin, c- myc and cyclin- D1 were analyzed by RT- PCR. Western blot was used to detect the levels of β-catenin protein.</p><p><b>RESULTS</b>Compared with control group, the different concentrations of 5-Aza-CdR could significantly inhibit the proliferation of Jurkat cells in a time-dose dependent manner (P<0.05). After being treated by 5- Aza- CdR for 48 hours, the cell early apoptosis rate in experiment group was significantly higher than that in control group (P<0.05). The promoters of SFRP1, SFRP2, SFRP4, SFRP5 genes were hypermethylation state in the control group, after being treated by 5-Aza-CdR for 72 hours, the brightness of SFRP1, SFRP2, SFRP4, SFRP5 genes' methylation strips weakened in a dose- dependent manner. SFRP mRNA expression increased (P<0.05) when 5- Aza- CdR concentration increased, and the level of β- catenin protein was dampened in a dose- dependent manner (P<0.05). As compared to the control group, the mRNA expressions of associated apoptosis genes survivin, c-myc and cyclin- D1, respectively were obviously down- regulated in a dose- dependent manner (P<0.05).</p><p><b>CONCLUSION</b>The effect of demethylation could up- regulate SFRP genes expressions by reversing its hypermethylation and induced apoptosis by down-regulation of β-catenin and associated apoptosis genes.</p>

Antiproliferative effect of silencing LSD1 gene on Jurkat cell line and its mechanism / 中华血液学杂志

<p><b>OBJECTIVE</b>To investigate the effect of silencing LSD1 gene by RNA interference on the proliferation, apoptosis on human lymphocytic leukemia Jurkat cell line and its mechanism.</p><p><b>METHODS</b>The hairpin- like oligonucleotide sequences targeting LSD1 gene was transfected into Jurkat cells by lipofectamine(TM) 2000. The LSD1 mRNA and protein were detected by RQ- PCR and Western blot. Cell growth was determined by MTT. Cell apoptosis was analyzed by flow cytometry. The expression of Bcl-2, Bax, procaspase- 3, and histone H3K4me, H3K4me2, H3K4me3, Act- H3, H3K9me were detected by Western blot.</p><p><b>RESULTS</b>LSD1 mRNA was markedly suppressed by the shRNA targeting LSD1. LSD1 shRNA suppressed the proliferation and induced cells apoptosis of Jurkat cells. The cell apoptotic rate was (41.34±3.58)%, (3.45±1.54)%, (1.76±0.52)% in LSD1 shRNA, Neg-shRNA and Blank respectively, the difference among them was statistically significant (P<0.05). LSD1 shRNA down- regulated the expressions of Bcl- 2 and procaspase- 3, and up- regulated the expression of Bax. The methylation of H3K4me1, me2 and acetylation of Act- H3 improved without change of the methylation of H3K4me3.</p><p><b>CONCLUSIONS</b>Deplete of LSD1 gene maybe through modifying the methylation of histone H3K4 to promote the cell apoptosis and inhibit cell growth in Jurkat cell line.</p>

Tal1 promotes proliferation of acute lymphoblastic leukemia Jurkat cells in vitro / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the role of Tal1 gene, which is aberrantly expressed in 40%-60% of patients with T lymphocytic leukemia (T-ALL), in the proliferation of T-ALL cells.</p><p><b>METHODS</b>We established stable Jurkat-siTal1 and Jurkat-T1 cell lines by trasnfecting T-ALL Jurkat cells with lentiviral vectors to knock-down or overexpress Tal1. Jurkat cells transfected with negative control siRNAs for Tal1 knock-down (Jurkat-mock1) and over-expression(Jurkat-mock2) served as the control cells. The proliferation of the cells lines was assessed using CCK-8 assay, and the cell cycle distribution was determined by flow cytometry. The mRNA and protein expressions of cyclin-dependent kinase inhibitor 2 (CDKN2A) and cyclin-dependent kinase inhibitor 1 (CDKN2B) were measured by real-time RT-PCR and Western blotting, respectively.</p><p><b>RESULTS</b>Jurkat-T1 cells showed more active proliferation in vitro than Jurkat-mock2 cells, while Jurkat-siTal1 cells showed slower growth than Jurkat-mock1 cells. In Jurkat-T1 cells, G0/G1 phase cells were decreased and S phase cells increased compared with Jurkat-mock2 cells, and Jurkat-siTal1 cells showed increased G0/G1 phase cells and decreased S phase cells compared with Jurkat-mock1 cells. Real-time RT-PCR and Western blotting showed that Tal1 inhibited the cellular expression of CDKN2A and CDKN2B at both mRNA and protein levels.</p><p><b>CONCLUSION</b>Tal1 promotes the growth and the transition from G0/G1 phase to S phase in T-ALL cells Jurkat by inhibiting the expressions of G0/G1 and S phase negative regulatory proteins CDKN2A and CDKN2B.</p>