Chemokine CXCL12-Induced Calcium Response of Jurkat T Cells under Fluid Shear Stress / 医用生物力学

Objective To investigate the mechanism of mechano-chemical coregulation in chemokine-induced calcium response of Jurkat T cells under fluid shear stress (FSS). Methods By using parallel-plate flow chamber combined with fluorescence microscope, the calcium response of Jurkat T cells on CXCL12 was observed to extract the corresponding characteristic parameters under static or flow state, with or without extracellular Ca2+, respectively. Results Immobilized CXCL12 could induce firm adhesion of the circulating Jurkat T cells, and the arrested cells increased with the increase of CXCL12 concentration. Force could trigger the calcium response of Jurkat T cells and sharply raised the activation ratio from 4% up to 75% when the FSS increased from 0 to 20 mPa. Under 20 mPa FSS, extracellular Ca2+ could stimulate quickly the calcium response by shortening the delay time (about 23 s), and enhance calcium intensity by prolonging the climbing time (about 7 s) and half time (about 20 s). Conclusions The cooperation between FSS and extracellular Ca2+ would accelerate and enhance CXCL12-mediated-calcium response of Jurkat T cells, which indicated a fast mechanosensitive pathway through ‘extracellular calcium influx-intracellular calcium store release’. The research results would contribute to understanding the process of T cells activation and providing the clue for relevant pathological and drug research.

Catharanthus roseus aqueous extract induces apoptosis, cell cycle and alters the associated gene expression in Jurkat cells

Abstract@#Introduction: The alkaloids present in Catharanthus roseus (C. roseus), vinblastine and vincristine are important anticancer agents that cause cell cycle arrest and apoptosis in various types of cell lines. However, there is no previous reports that emphasized the clear mechanisms of anticancer exerted by a crude aquoeus extract of C. roseus although it has been historically used to treat various diseases. Methods: The cytotoxicity effects of C. roseus aqueous extract on Jurkat cells were evaluated by annexin/PI staining, caspase 3/7 assay, JC-1 assay and cell cycle assay. Gene expression profiling was performed by using SmartChip Real-Time PCR system to evaluate the expression profiles of oncology-related genes of Jurkat cells treated with C. roseus aqueous extract. Results: Flow cytometry analysis revealed that the extract has caused S-phase arrest and associated with apoptosis through the externalization of phosphatidylserine and depletion of mitochondrial membrane potential in time-dependent manner. The apoptosis mechanism was mediated through the activation of caspase 3/7. From the gene expression analysis, 8 differentially regulated genes were associated with apoptosis which were CDKN1C, CHI3L2, BIRC8, GFER, ID3-1, BBC3-2, TRAF4 and VCAN. Meanwhile, 7 differentially regulated genes were associated with cell cycle progression which were PIMI-1, CDKN1C, SKP1A, CDC25C, LTBP1, CCNG2 and RBL1. Conclusion: The recent data may facilitate the identification of specific targeting pathways induced by the extract. The information obtained may be used as diagnostic tools, prognostic markers, and predictors of response to C. roseus treatment especially for this particular type of cancer.

Effect and mechanism of Solanine on proliferation and apoptosis of Jurkat cells / 中华实用儿科临床杂志

Objective To explore the effect of proliferation and apoptosis of Solanine on acute T lymphocyte leukemia (T-ALL) Jurkat cells and its mechanism.Methods After treated with different concentrations of Solanine,the proliferation of Jurkat cells was detected by CCK-8 assay,and the effect of Solanine on apoptosis of Jurkat cells were detected by flow cytometry.The expression of Bcl-2 and Bax in Jurkat cells were detected by Western blot,and the expression levels of Bcl-2 mRNA and Bax mRNA were detected by real-time fluorescence quantitative polymerase chain reaction.Results CCK-8 assay showed that Solanine significantly inhibited the proliferation of Jurkat cells in a dose-and time-dependent manner.The results of flow cytometry showed that the apoptosis rates of Jurkat cells treated with Solanine for 24 h were (2.40-± 0.98) ％,(28.43-± 4.86) ％,(41.56-± 1.87) ％,respectively,in a dose-dependent manner.Western blot showed that Solanine could increase the expression of Bax and decrease the expression of Bcl-2 in Jurkat cells,and they all were dose-dependent.Conclusion Solanine can significantly inhibit the proliferation and induce apoptosis of Jurkat cells.The mechanism is related to the up-regulation of Bax expression and down-regulation of Bcl-2 expression.

IL-12 reverses inhibitory effect of cisplatin on immune function of human NK cells and its mechanism / 中国肿瘤生物治疗杂志

@# Objective: To investigate the reverse effect and mechanism of IL-12 on chemotherapeutic medicine suppressing the immune function of NK cells. Methods: Purified NK cells were stimulated with PMAplus Ionomycin in the presence or absent of Cisplatin (DDP) and IL-12. The levels of IFN-γ and TNF-α in culture supernatants were detected by enzyme-linked immunosorbent assay (ELISA); The content of IFN-γ and TNF-α, TRAIL (TNF-related apoptosis inducing ligand) and transcription factors including T-bet and p-STAT-4 in NK cells were analyzed by Flow cytometry. The cytotoxicity of purified NK cells (pretreated with/without chemotherapeutics and IL-12 for 48 h) to Jurkat cells was measured by Flow cytometry. Results: Chemotherapeutics significantly inhibited the production of IFN-γ, TNF-α and the expression of TRAILin NK cells, which were significantly reversed by IL-12 (P<0.05 or P<0.01). Further study revealed that chemotherapeutics down-regulated while IL-12 reversed the expression of p-STAT4 to restore cytokine production. In addition, DDP also inhibited but IL-12 recovered the cytotoxicity of NK cells against tumor cells by inducing the expression of TRAIL (P<0.05 or P<0.01). Conclusion: Chemotherapeutics inhibited the cytotoxicity of NK cells and its secretion of cytokines (IFNγ and TNF-α), which were reversed by IL-12 via up-regulating TRAIL and p-STAT-4; this might provide experimental evidence for the clinical application of IL-12 for rebuild the immune function of tumor patients receiving chemotherapy.

Effect of down-regulation of CD59 gene on apoptosis of acute T lymphocyte Jurkat cell lines / 中国免疫学杂志

Objective:To study the change of related molecular about apoptosis we reduce the expression of CD59 on acute T lymphocyte Jurkat cell lines .Methods: RNA interference (RNAi) was used to reduce the expression of CD59 gene by lentivirus,confocal was applyed to observe the transfection efficiency and the location of CD59 molecular then Real-time-PCR and Western blot were used to select the most effective group to do the rest experiment;Western blot was used to detect the change of expression about Bcl-2,Bax,Caspase-3 and Survivin;ELISA was used to investigate the expression of IL-3 and TNF-β.Results: Confocal observed each group′s transfection efficiency over 90%,CD59 molecules were mainly located in cell membrane;Real-time-PCR and Western blot showed group A had the best down-regulation efficiency;we defined RNAi-CD59-A as experimental group for subsequent experiments named RNAi-CD59;the experimental group can enhance the expression of Bax,caspase-3 (P<0.05),inhibit the expression of Bcl-2 and Survivin(P<0.05);ELISA showed that the expression of IL-3 in the down-regulation group increase(P<0.05),the expression of TNF-β decrease (P<0.05).Conclusion: Down-regulation CD59 can promote the expression of apoptosis molecular in acute leukemia Jurkat cell lines restrain the expression of proliferation molecular.

Promotion of immune response by soluble Tim-3 in vitro and therapeutic potential / 军事医学

Objective To evaluate the role of recombinant human soluble Tim-3 (hTim-3-Fc) in regulating immune response.Methods Soluble hTim-3 was incubated with human macrophage cell line U 937, human T cell line Jurkat and normal human PBMC before cytokines secreted by or expressed in different immune cells were analyzed using ELISA , RT-PCR and Western-blotting, respectively.Results Soluble hTim-3 significantly promoted the activation of different immune cells.Our data showed that IL-8 secretion by U937 cells, IL-2 secretion by Jurkat cells , IL-2 and IFN-γsecretion by human PBMCs were all significantly increased .In addition , soluble hTim-3 significantly increased the IFN-α2 and IFN-β1 mRNA expression in U937, Jurkat and PBMCs and increased the phosphorylation of stat-1 in Jurkat and U937 cells.Conclusion Recombinant soluble hTim-3 can significantly promote the activation of immune cells in vitro, which shows its therapeutic potential .

Evaluation of Cytotoxic Activity of Sargassum vulgare From the Lebanese Coast Against Jurkat Cancer Cell Line.

The discovery of cancer drugs that effectively destroy cancer cells or stop their growth without toxicity to normal cells is a challenge to enhance the therapeutic effects and reduce side effects. Many papers have highlighted the implication of marine algae that show anticancer activity. In this report, we assessed the cytotoxic activity of two different crude extracts from Sargassum vulgare (Sargassaceae), a marine brown algae collected from the Lebanese coast, against Jurkat human cancer cell line using trypan blue exclusion test. Both extracts, water: ethanol extract and chloroform: ethanol extract, showed cytotoxic activity against Jurkat cancer cell line with IC50 values of 136.907 μg/ mL and 49.056 μg/ mL, respectively after 72 hours of treatment. Further research designed to isolate and identify novel and efficient anticancer drug candidates from these seaweed extracts need to be explored.

Antioxidant Potential of Polyphenol Rich Extract from Hibiscus sabdariffa.

Aims: Hibiscus sabdariffa is a medicinal plant species that is consumed for its health benefits in Africa, therefore this study investigated the antioxidant properties of Hibiscus polyphenolic rich extract (HPE), prepared from Hibiscus sabdariffa. Place and Duration of Study: School of Biomolecular Sciences, Liverpool John Moores University, Byrom Street, Liverpool, United Kingdom, between June 2009 and December 2010. Methodology: The antioxidant assays evaluated the scavenging abilities of HPE: Firstly against superoxide ions generated during the xanthine oxidase mediated breakdown of xanthine to uric acid. Secondly against ABTS (2,2-azino-bis-(3-ethylbenzthiazoline 6-sulfonic acid)) radical cation generated by filtering a solution of ABTS through manganese dioxide powder and potassium persulphate. Finally metal chelation ability of HPE against Iron ions (Fe2+) induced oxidative damage in cultured Jurkat-T cells was also assessed. Results: The results showed that 1.0% and 2.5% (v/v) diethyl ether extract of HPE significantly inhibited superoxide ions by 42.35 and 100.00% respectively. The extract also inhibited uric acid production, which suggest that components of HPE inhibit xanthine oxidase activity. In addition, it was found that HPE scavenge ABTS radical cations in dose dependent manner. HPE inhibited Fe2+-mediated lipid peroxidation in cultured Jurkat-T cells supplemented with 0.5 mg/ml and 1.0 mg/ml of HPE by 19.67% and 31.69% respectively, metal chelation ability was identified as a potential mechanism behind this observed reduction. Conclusions: HPE is rich in different phenolic compounds; therefore strong antioxidant potential of HPE observed in this study may be related to their polyphenolic constituents. This study demonstrated that Hibiscus sabdariffa is an efficient antioxidant plant species in vitro and may be beneficial in reducing oxidative damage to lipid and thus prevent or reduce the development and progression of free radical mediated diseases.

Effect of over expression of C-terminal Src kinase binding protein (Cbp) and Cbp palmitoylation to proliferation of T-leukemia Jurkat cell in nude mice / 中国免疫学杂志

Objective:To establish T lineage leukemia Jurkat cell mice model of over expression of C-terminal Src kinase binding protein( Cbp ) and Cbp palmitoylation and to research the effect of Cbp and Cbp palmitoylation to proliferation of Jurkat cell.Methods:Virus transfected cell of neg-EGFP,Cbp-EGFP and Cbp-m-EGFP were used in mice model.24 female BALB/c-nu mice were randomly divided into blank control group,empty virus control group,over expression of CBP group and Cbp palmitoylation group, 6 mice in each group.The nude mice were weighed in 0,1,2,3,4,5 weeks.The amount of white blood cell in peripheral blood were counted in 0, 1, 2, 3, 4, 5 weeks.The proliferation of Jurkat cell in peripheral blood of mice were observed by laser confocal microscope.The pathological changes of liver were observed using HE staining.The proliferation of Jurkat cell in the bone marrow and peripheral blood of mice were detected with flow cytometry.Results:The weight of mice in over expression of Cbp group was less than that in blank control group,but higher than that in empty virus control group and Cbp palmitoylation group.The weight of mice in Cbp palmitoylation group was less than that in blank control group,empty virus control group and over expression of Cbp group.The amount of white blood cell in peripheral blood and proliferation of Jurkat cell in liver, bone marrow and peripheral blood of mice in over expression of Cbp group was higher than that in blank control group, but less than that in empty virus control group and Cbp palmitoylation group.The amount of white blood cell in peripheral blood and proliferation of Jurkat cell in liver, bone marrow and peripheral blood of mice in Cbp palmitoylation group was higher than that in blank control group,empty virus control group and over ex-pression of Cbp group.Conclusion:Over expression of Cbp and Cbp palmitoylation in T lineage leukemia Jurkat cell mice model was established.Over expression of Cbp has inhibitory effect on the proliferation of Jurkat cell.Cbp palmitoylation has promotable effect on the proliferation of Jurkat cell.

Molecular cloning, in vitro expression and bioactivity of TRAIL (TNFSF10) gene from finless porpoises / 生物工程学报

To construct soluble TNF related apoptosis inducing ligand (TRAIL) expression system and investigate the effect of the expression product on tumor cell. It may provide valuable information for research into the immune system of the finless porpoise. The full-length cDNA of TRAIL (designated fTRAIL) was cloned from the total RNA of the finless porpoises blood using RT-PCR techniques and then the extracellular soluble fragments of fTRAIL (designated fsTRAIL) was ligated into pET43.1a. Recombinant soluble fTRAIL (pET43.1a-fsTRAIL) fused with Nus-his tag was efficiently expressed in Escherichia coli BL21 (DE3) and the Nus-His-fsTRAIL protein was purified. The expression of Nus-His-fsTRAIL was verified by Western blotting. In vitro, the effects of the purified Nus-His-fsTRAIL protein on Jurkat and HeLa cells were etected by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrasodium bromide (MTT) assay, TrypanBlue and Flow Cytometry analysis. The expression system pET43.1a-fsTRAIL was constructed and Nus-His-fsTRAIL protein was expressed successfully. In vitro, the Nus-His-fsTRAIL protein was able to inhibit the proliferation and induce apoptosis of Jurkat and HeLa cells in a dose-dependent manner. The Nus-His-fsTRAIL protein has anti-tumor activity against Jurkat and HeLa cells in vitro.

Immune cytotoxic effect of trichloroethylene in Jurkat T cells / 中国职业医学

OBJECTIVE: To explore the immune cytotoxic effect and the maximum non-effect dose of trichloroethylene( TCE) on Jurkat T cells in vitro. METHODS: i) Naive and activated Jurkat T cells were treated with different concentrations of TCE( 0. 10, 0. 50, 1. 00, 2. 00, 5. 00, 10. 00 mmol / L). Phorbol-12-myristate-13-acetate and ionomycin were used as agonist. No TCE was used in the control group and dimethyl sulfoxide( DMSO) was used as the solvent group. The morphology of Jurkat T cells was observed using a light microscope and the survival rate of Jurkat T cells was investigated using CCK-8 essay after cells were cultured for 24,48 and 72 hours. ii) Nave and activated Jurkat T cells were treated with different concentrations of TCE( 0. 00,0. 02,0. 20,2. 00 mmol / L). The apoptosis of cells was detected using flow cytometry and the level of interleukin-2( IL-2) in supernatant was detected using enzyme linked immunosorbent assay after cells were cultured for 24,48 and 72 hours. RESULTS: i) Cytotoxic effect was observed after cells were exposed to 10. 00 mmol / L TCE for 24 hours. Cells dispersed,cell volume diminished,cell membrane ruptured,cytoplasm condensed and increased outflow of intercellular organelles. The effect of interaction between exposure dose and exposure time was statistically significant on cell survival rate( P < 0. 01). Compared with the control and DMSO groups at the same time points,there were no significant differences in the 0. 10,0. 50,1. 00 and 2. 00 mmol / L TCE treatment groups in cell survival rates in three different time points( P > 0. 05),while the cell survival rates of 5. 00 and 10. 00 mmol / L TCE treatment groups were significantly decreased( P < 0. 01). ii) When TCE concentration was 0. 00-2. 00 mmol / L,there were no significant differences in the main effect of exposure dose and interactions of between exposure dose and cell type or exposure time on cell apoptosis rate( P > 0. 05). Compared with the same time points and groups of naive Jurkat T cells,the levels of IL-2 of activated Jurkat T cells were significantly increased( P < 0. 01). In the three different time points,the level of IL-2 of activated Jurkat T cells increased in accordance with the TCE exposure dose,showing a dose-effect relationship( P < 0. 01). The level of IL-2 of activated Jurkat T cells increased in accordance with TCE exposure time,showing a time-effect relationship( P < 0. 01). CONCLUSION:s TCE at the level of 2. 00 mmol / L had no observed effect in Jurkat T cells. High doses of TCE( ≥5. 00 mmol / L) showed cytotoxic damages to naive and activated Jurkat T cells and low doses of TCE( ≤2. 00 mmol / L) could stimulate activated Jurkat T cells secrete IL-2 in a dosedependent and time-dependent manner.

Effects of celastrol on the proliferation and apoptosis of human leukemia cells HL-60 and Jurkat / 中国药科大学学报

@#To determine the effects of celastrol on the proliferation and apoptosis of cell lines HL-60 and Jurkat in human leukemia. Human leukemia cell lines were treated with celastrol at different concentrations. The inhibitory rate of cell proliferation was detected by MTT assay; the apoptosis rate was detected by AnnexinV-FITC/PI double staining; cell cycle was observed by PI staining; cell morphology was observed by transmission electron microscope(TEM). Cell proliferation was inhibited by celastrol, with IC50 of(0. 46±1. 05)μmol/L and(0. 88±1. 13)μmol/L at 24 h. Celastrol induced cell apoptosis in a dose-dependent manner. The cell cycle distribution of G1 phase rate increased, S phase rate decreased(P< 0. 05)with typical cell apoptosis-induced morphological changes. Results showed that celastrol could significantly inhibit cell proliferation and induce apoptosis in human leukemia cell lines of HL-60 and Jurkat.

Effects of high expression of Csk-binding protein on morphology and biological function of Jurkat cells / 中国组织工程研究

BACKGROUND:The linkage and synergistic effect of adaptor proteins can effectively regulate signal transduction of T cel s, which can form a limit or amplification cascade to realize the complex immune function of T cel s. C-terminal Src kinase (Csk)-binding protein (Cbp) is an adaptor protein, which mainly exert the negative feedback regulation of Src kinase activity. This negative feedback effect depends on Y317 of Cbp, which may be involved in the SH2 domain of Csk. OBJECTIVE:To explore the effects of high expression of Cbp on ultrastructure and related biological function of Jurkat cel s. METHODS:The virus particles were constructed with expressing enhanced green fluorescent protein (EGFP) only and Cbp-EGFP fusion protein to transfect Jurkat cel s. There were untransfected group (Jurkat group), negative control group (transfected with expression of EGFP virus only), and Cbp group (transfected with Cbp-EGFP virus). RESULTS AND CONCLUSION:Confocal microscope showed that cel transfection efficiency was more than 95%and Cbp was located on the cel membrane. Optical microscope showed after transfection with Cbp-EGFP virus, more Jurkat cel s shrunk, with poor size uniformity. Apoptosis detection showed that after transfection with Cbp-EGFP virus, the number of apoptotic and necrotic cel s was greatly increased. Cbp mRNA expression was increased, Csk expression was decreased obviously and lymphocyte-specific protein tyrosine kinase expression was increased. So, in Jurkat cel s, the high expression of Cbp can decrease the uniformity of cel s and increase the necrosis cel s, thus inhibiting the signal transduction.

Mechanism of ouabain inducing the apoptosis of T lymphoblastic leukemia Jurkat cells / 医学研究生学报

Objective Ouabain is a cardiotonic steroid that can induce the apoptosis of many tumorous cells .This study was to investigate the anti-tumor mechanisms of ouabain by observing its effects on the apoptosis of T lymphoblastic leukemia Jurkat cells and the expressions of hTERT and c-myc mRNA and protein . Methods Jurkat cells were treated with ouabain at the concentrations of 50 and 100 nmol/L for 24 and 48 hours, and those treated with 1 ×PBS served as the control .Then the apoptosis rate of the cells was detected by flow cytometry after Annexin V/PI staining, the expressions of hTERT and c-myc mRNA determined by RT-PCR, and those of hTERT and c-myc protein by Western blot . Results The apoptosis rates of the Jurkat cells in the 50 and 100 nmol/L oua-bain groups were (5.67 ±3.71)%and (9.63 ±4.83)%respectively at 24 hours, and (19.67 ±4.55)%and (37.60 ±11.89)%at 48 hours, significantly higher than (4.23 ±1.01)%in the PBS control group at 48 hours (P<0.05).Compared with the control, the expressions of hTERT and c-myc mRNA were decreased by 200%and those of hTERT and c-myc protein by 224%and 400%, re-spectively, at 48 hours (P<0.05).There was a positive correlation between the reduction of the mRNA levels and that of the protein levels of hTERT and c-myc (P<0.05). Conclusion Ouabain can down-regulate the mRNA and protein expressions of hTERT and c-myc, which may be one of the mechanisms of its induction of the apoptosis of Jurkat T lymphocyte leukemia cells .

CD59 has a positive effect on LAT induced T lymphocytes activation / 中国免疫学杂志

Objective:To investigate the function of CD 59 in LAT induced T lymphocytes'proliferation and activation.Methods:Transfected LAT-GFP recombinant lentiviral vectors into Jurkat cells and established a fusion-protein stable express cell line ( Jurkat-GFP ).Junket-GFP cells were transfected with pSUPER-siCD59 plasmids by electroporetion or stimulated by anti-CD59 antibody.The cellular locations of CD 59 and LAT were observed under fluorescence microscope with the immunofluorescence cytochem -istry.The cells proliferation were measured by MTT assay.Furthermore,Western blot was used to detect the total and phosphorylation levels of several down-stream proteins after T cell activated .Results: Jurkat-GFP cells successfully transfected with pSUPER-siCD59 plasmids showed lower fluorescence staining.CD59 and LAT distributed uniformly on the cell surface before stimulated with anti-CD59 antibody and formed clusters once upon stimulation.Jurkat-GFP cells stimulated with anti-CD59 antibody showed a higher level of pro-liferation and protein phosphorylation ,compared with the others.Conclusion:CD59 contributed to LAT induced signaling transduction of T lymphocytes ,and stimulated CD59 molecule partly promoted T cell activation.

Specific cytotoxicity of MUC1 chimeric antigen receptor-engineered Jurkat T cells against hepatocellular carcinoma / 第二军医大学学报

Objective: To investigate the specific cytotoxicity of MUC1-specific chimeric antigen receptor (CAR)-engineered Jurkat T cells against hepatocellular carcinoma (HCC) cells. Methods: The expression cassettes of both the first and the third generation MUC1-specific CAR gene (i.e. MUC1-CAR and G3MUC1-CAR) were constructed and cloned into lentivirus transfer plasmids, respectively. Then the obtained recombinant lentiviruses carrying the MUC1-specific CAR gene and hrGFP reporter genes were used to infect Jurkat T cells in vitro to establish MUC1-sepcific CAR-engineered Jurkat cells (i.e. CAR-T cells). Subsequently, the assays of CCK-8, ELISA, and LDH were used to detect the cell proliferation, IL-2 secretion and killing effect of the CAR-T cells, respectively. Results: We successfully constructed the expression cassettes of MUC1-sepcific CAR gene and the corresponding recombinant lentivirus, established the MUC1-specific CAR-engineered Jurkat T cells. Both types of MUC1-specific CARs-engineered Jurkat T cells could recognize MUC1 molecule and specifically kill MUC1 over-expressed HCC cells while left normal hepatic cells almost undamaged. In addition, the cell proliferation, the secretion of IL-2 and killing effect of the G3MUC1-CAR-engineered Jurkat T cells was significantly superior to the MUC1-CAR-engineered counterpart (P<0.05 or 0.01). Conclusion: The MUC1-sepcific CAR-engineered Jurkat T cells can specifically kill MUC1 over-expressed HCC cells.

Citotoxicidad in vitro de extractos laticiferos de Calotropis procera (Aiton) W.T. Aiton y Pedilanthus tithymaloides (L.) Poit / In vitro cytotoxicity of laticifers extracts from Calotropis procera (Aiton) WT and Pedilanthus tithymaloides (L) Poit

In order to assess the anticancer action of extracts obtained by latex from Calotropis procera and Pedilanthus tithymaloides, samples were collected from adult plants. Soluble proteins were extracted with 16 uL of 50 mM sodium acetate pH 5/ug integral latex and centrifugation at 16,000 x g for 15 min, the supernatant was named "latex crude extract" (LCE). The "latex methanolic extract" (LME) was obtained on dried latex. Both extracts were tested in vitro by cytotoxic and cytostatic activity in Jurkat cell cultures. Cellular viability, proliferation, necrosis and apoptosis were evaluated. LCE and LME of C. procera were found with cytotoxic and cytostatic activity after 24 incubation hours (p < 0,05) with doses from 1ug/mL. The LCE and LME of P. tithymaloides presented cytotoxic effect (p < 0,05) from 50 ug/mL and from 1ug/mL, respectively.

Con el objetivo de evaluar el potencial anticanceroso de extractos de látex de Calotropis procera y Pedilanthus tithymaloides se colectaron muestras de plantas adultas. Las proteínas solubles fueron extraídas con 16 uL de acetato de sodio 50 mM pH 5/ug de látex integral y centrifugación a 16.000 x g durante 15 min, denominándose al sobrenadante “extracto crudo de látex” (ECL). El “extracto metanólico de látex” (EML) se obtuvo sobre látex deshidratado. Ambos extractos fueron probados en su actividad citotóxica y citostática in vitro sobre cultivos de células Jurkat. Se realizaron estudios de viabilidad, proliferación, necrosis y apoptosis celular. El ECL y el EML de C. procera presentaron actividad citotóxica y citostática después de 24 y 48 horas de incubación (p < 0,05) con dosis desde 1 ug/mL. Los ECL y EML de P. tithymaloides presentaron efectos citotóxicos (p < 0,05) a partir de 50 ug/mL y desde 1 ug/mL respectivamente.

Identification and function analysis of a novel transcript 630 in acute T-cell lymphoblastic leukemia Jurkat cell lines / 中华实用儿科临床杂志

Objective To identify and analyze a novel transcript 630 in acute T-cell lymphoblastic leukemia cell (T-ALL) Jurkat cell lines.Methods RNA-sequencing was applied to transcriptome profiling.The novel transcript 630 was identified by reverse transcription polymerase chain reaction (RT-PCR) combined with sequencing.The expression level of the novel transcript was detected in some tumor cell lines and normal cells.The sequences for small interfering RNA (siRNA) against 630 were designed and synthesized.Then,the effects of siRNA were determined by real-time PCR.Apoptotic cells and cell cycle analysis were investigated with the flow cytometry after siRNA-mediated knockdown of 630.Results The novel transcript 630 was identified.High expression level of 630 was detected in Jurkat E6-1,HL60 and human umbilical vein endothelial(HUVE) cells.The knockdown of transcript 630 induced a decrease of cellular proliferation.Conclusions The novel transcript 630 is one of the novel transcripts in Jurkat cells.The decreased expression level of transcript 630 can suppress the proliferation of leukemia cells.The novel transcript 630 might contribute to the treatment of T-ALL.

MicroRNA profiling of tacrolimus-stimulated Jurkat human T lympocytes / 대한외과학회지

PURPOSE: This study investigated the Jurkat T cell line expresses cytotoxicity when treated with different concentrations of FK506, and analyzed the expression pattern of microRNA when stimulated by FK506 using the microRNAs microarray, as well as the expression pattern of a gene that is related to the differentiation, activation and proliferation of T cells after being affected by the change of microRNAs. METHODS: To investigate the effects of FK506 on microRNA expression, we purified total RNA of Jurkat cells treated with 20 microM FK506 for 72 hours and used to analyze microRNA profiling by using Agilent's chip. RESULTS: These results demonstrated that treatment with FK506 markedly induced the down-regulation of 20 microRNAs as well as the up-regulation of 20 microRNAs in a time-dependent manner. The genes that down-regulated by FK506 include let-7a*, miR-20a*, and miR-487a. Otherwise miR-202, miR-485-5p, and miR-518c* are gradually up-regulated in expression. Sanger Institute and DAVIDs bioinformatics indicated that microRNAs regulated the several transcriptomes including nuclear factor of activated T cell-related, T cell receptor/interleukin-2 signaling, and Ca(2+)-calmodulin-dependent phosphatase calcineurin pathways. CONCLUSION: As a result of treating FK506 to a Jurkat cell line and running the microRNA microarray, it was found that FK506 not only took part in the suppression of T cell proliferation/activation by inhibiting calcineurin in Jurkat apoptosis, but also affected the microRNAs that are involved in the regulation of various signal transduction pathways.

Mechanism of escin sodium in inhibition of human leukemia Jurkat cells proliferation / 中草药

Objective To investigate the effects of escin sodium on proliferation of human leukemia Jurkat cells and its possible mechanism. Methods The reduction of cellular viability was determined by MTT assay. Hoechst 33258 staining, DNA fragmentation assay, FITC-Annexin V-FITC/PI staining assay, and cytometric analysis were used to confirm the features of apoptosis and cell cycle. Western blotting assays were performed to explore the apoptotic pathway. Results Escin sodium inhibited the proliferation of Jurkat cells in both dose- and time-dependent manners. The morphological apoptosis, DNA fragmentation pattern, and the percentage of Annexin V+/PI- (early apoptosis) cells were markedly increased in escin sodium-treated Jurkat cells. Escin sodium activated Caspase-8, Caspase-9, and Caspase-3, degraded poly (ADP-ribose) polymerase (PARP), and attenuated Bcl-2 expression. Conclusion Escin sodium could inhibit the proliferation of Jurkat cells via the induction of apoptosis effectively.