Journal: iScience

Article Title: CRISPR-Cas12a-integrated transgenes in genomic safe harbors retain high expression in human hematopoietic iPSC-derived lineages and primary cells

doi: 10.1016/j.isci.2023.108287

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Article Snippet: Jurkat, Childhood T-cell acute lymphoblastic leukemia line , DSMZ-German Collection of Microorganisms and Cell Cultures GmbH , Jurkat; DSMZ ACC 282; RRID:CVCL_0065.

Techniques: Virus, Selection, Recombinant, Knock-Out, Clone Assay, Plasmid Preparation, Modification, Staining, Generated, Software, Flow Cytometry

Journal: Scientific Reports

Article Title: CRISPRi for specific inhibition of miRNA clusters and miRNAs with high sequence homology

doi: 10.1038/s41598-022-10336-3

Figure Lengend Snippet: Expression of miRNAs encoded within mir-17–92 cluster upon dCas9-KRAB-mediated inhibition of this locus. ( A ) Normalized expression of miR-17-5p, miR-18a-3p, miR-19a-3p, miR-20a-5p and miR-19b-3p upon the use of three most effective sgRNAs targeting putative TSS of mir-17–92 cluster in JURKAT cell line. ( B ) Normalized expression of miR-17-5p, miR-18a-3p, miR-19a-3p, miR-20a-5p and miR-19b-3p upon the use of three most effective sgRNAs targeting putative TSS of mir-17–92 cluster in ALL-SIL cell line. Scr—scrambled control. * p < 0.05; ** p < 0.01; *** p < 0.001.

Article Snippet: DND-41 and Jurkat T-cell acute lymphoblastic leukemia cell lines were purchased from the Leibniz Institute DSMZ—German Collection of Microorganisms and Cell Cultures.

Techniques: Expressing, Inhibition, Control

Journal: Scientific Reports

Article Title: CRISPRi for specific inhibition of miRNA clusters and miRNAs with high sequence homology

doi: 10.1038/s41598-022-10336-3

Figure Lengend Snippet: Expression of miRNAs encoded within mir-106a-363 cluster upon dCas9-KRAB-mediated inhibition of this locus. ( A ) Normalized expression of miR-106a-5p, miR-20b-5p, miR-19b-3p and miR-363-3p upon the use of three most effective sgRNAs targeting putative TSS of mir-106a-363 cluster in JURKAT cell line. ( B ) Normalized expression of miR-106a-5p, miR-20b-5p, miR-19b-3p and miR-363-3p upon the use of three most effective sgRNAs targeting putative TSS of mir-106a-363 cluster in ALL-SIL cell line. Scr—scrambled control. * p < 0.05; ** p < 0.01; *** p < 0.001.

Article Snippet: DND-41 and Jurkat T-cell acute lymphoblastic leukemia cell lines were purchased from the Leibniz Institute DSMZ—German Collection of Microorganisms and Cell Cultures.

Techniques: Expressing, Inhibition, Control

Journal: Scientific Reports

Article Title: CRISPRi for specific inhibition of miRNA clusters and miRNAs with high sequence homology

doi: 10.1038/s41598-022-10336-3

Figure Lengend Snippet: Comparison of specificity of miRZip and dCas9-KRAB approaches towards selective silencing of miR-130a-3p and miR-130b-3p. ( A ) Normalized expression of miR-130a-3p and miR-130b-3p upon the use of miRZip vector encoding shRNAs targeting miR-130a-3p or miR-130b-3p as compared do scrambled control miRZip vector (Scr) in JURKAT cell line. ( B ) Normalized expression of miR-130a-3p and miR-130b-3p upon the use of dCas9-KRAB system and sgRNAs targeting mir-130a TSS as compared do scrambled control (Scr) in JURKAT cell line. ( C ) Normalized expression of miR-130a-3p and miR-130b-3p upon the use of dCas9-KRAB system and sgRNAs targeting mir-130a TSS as compared do scrambled control (Scr) in JURKAT cell line. * p < 0.05; ** p < 0.01; *** p < 0.001.

Article Snippet: DND-41 and Jurkat T-cell acute lymphoblastic leukemia cell lines were purchased from the Leibniz Institute DSMZ—German Collection of Microorganisms and Cell Cultures.

Techniques: Comparison, Expressing, Plasmid Preparation, Control

Journal: BMC Microbiology

Article Title: IRAK-M alters the polarity of macrophages to facilitate the survival of Mycobacterium tuberculosis

doi: 10.1186/s12866-017-1095-2

Figure Lengend Snippet: IRAK-M over-expression resulted in higher bacterial load, while IRAK-M interference resulted in lower bacterial load in M. tb infected cells. Lentiviruses that over-expresses (IRAK-M-GFP-Lentivirus, OE) or interferes with (IRAK-M-RNAi-GFP-Lentivirus, KD) IRAK-M expression, were constructed. a Lentiviruses of negative control (NC) and OE were introduced into Jurkat cells. Cells were examined by light microscope and fluorescent microscope at 96 h post-transfection. More than 80% of Jurkat cells expressed GFP (200×, scale bar 50 μm). b Lentiviruses of NC and KD were introduced in U937 cells. Cells were examined by light microscope and fluorescent microscope at 96 h post-transfection. More than 90% of U937 cells expressed GFP (200×, scale bar 100 μm). c Expression of IRAK1-4 in Jurkat and U937 cells was evaluated by Western blot. d - g Bacterial load in Jurkat ( d , e ) and U937 cells ( f , g ) challenged with virulent M. tuberculosis H37Rv strain (MOI 10) were analyzed by acid-fast staining and colony forming units (CFU). Cells were infected with lentivirus for 96 h, challenged with H37Rv for 5 h and washed three times with PBS. d , f At 24 h post-infection, 1 × 10 6 cells were resuspended in 20 μl PBS and fixed in 4% para-formaldehyde for 15 min on the slides. Acid-fast staining (AF) was performed and arrows indicated AF-positive bacteria in the cells (1000×, scale bar 10 μm). e , g For determination of CFU, at 5 and 24 h post-infection, 1 × 10 5 cells were washed aseptically, homogenized and plated at 10-fold serial dilutions on Middlebrook 7H11 agar. CFU numbers per plate were counted to evaluate the bacterial load 3 to 4 weeks later. The bacterial load in cells of different groups was expressed as Log10 CFU ± SEM (**, p < 0.01; ***, p < 0.001)

Article Snippet: Human monocytic leukemia line U937 (ATCC® CRL-1593.2) and human T-cell acute lymphoblastic leukemia line Jurkat (ATCC® TIB-152TM) were cultured at 37 °C in CO 2 incubator (CCL-170B-8, ESCO, Singapore) in RPMI-1640 medium, supplemented with 10% fetal bovine serum, 100 U/ml penicillin and 100 μg/ml streptomycin.

Techniques: Over Expression, Infection, Expressing, Construct, Negative Control, Light Microscopy, Microscopy, Transfection, Western Blot, Staining, Bacteria

Journal: PLoS Neglected Tropical Diseases

Article Title: Mitochondria and lipid raft-located F O F 1 -ATP synthase as major therapeutic targets in the antileishmanial and anticancer activities of ether lipid edelfosine

doi: 10.1371/journal.pntd.0005805

Figure Lengend Snippet: ( A ) L . panamensis promastigotes were untreated (Control) or treated with 10 μM edelfosine at the indicated times, and cells with disrupted ΔΨ m (DiOC 6 (3) low ) and ROS production (HE→Eth) were measured by flow cytometry. The numbers in each quadrant refer to the percentages of cells in each population. ( B ) L . panamensis promastigotes untreated (Control) and treated with edelfosine (EDLF) for 3 h were incubated with 2 μM HE and 10 μg/ml Hoechst 33342, and then analyzed by fluorescence microscopy. ( C ) L . panamensis promastigotes and ( D ) Jurkat cells were preincubated with 10 μg/ml CsA for 1 h, or with 10 mM NAC or 10 mM GSH for 2 h, and then incubated in the absence or presence of 10 μM edelfosine for 9 h. After treatment, the percentage of hypodiploid cells was analyzed by flow cytometry. Untreated control cells were run in parallel. ( E ) L . panamensis promastigotes and ( F ) Jurkat cells were preincubated with 10 μM rotenone, 5 mM malonate, 10 μM antimycin A, 1.5 mM azide, 50 μM CCCP or with 1 and 10 μM oligomycin ( L . panamensis and Jurkat cells, respectively) for 1 h, and then incubated in the absence or presence of 10 μM edelfosine for 9 h. After treatment, cells producing ROS were quantified by flow cytometry. Untreated control cells were run in parallel. Data shown are means ± SD or representative of three independent experiments performed. Asterisks denote that the differences between the indicated groups (C and D) and with control cells (E and F) are statistically significant. (*) P <0.05. (**) P <0.01.

Article Snippet: We also found that edelfosine was very efficient in promoting cell death in additional human leukemic cell lines, including human T-cell acute lymphoblastic leukemia (T-ALL) cell lines Jurkat (53.4 ± 6.2% apoptosis) and CEM-C7H2 (58.2 ± 5.9% apoptosis).

Techniques: Control, Flow Cytometry, Incubation, Fluorescence, Microscopy

Journal: PLoS Neglected Tropical Diseases

Article Title: Mitochondria and lipid raft-located F O F 1 -ATP synthase as major therapeutic targets in the antileishmanial and anticancer activities of ether lipid edelfosine

doi: 10.1371/journal.pntd.0005805

Figure Lengend Snippet: ( A ) L . panamensis promastigotes and T-cell leukemia Jurkat cells were untreated (Control) or pretreated with MCD, and then incubated in the absence or presence of 10 μM edelfosine for 24 h. Percentage of hypodiploid cells were measured by flow cytometry. ( B ) L . panamensis promastigotes and T-cell leukemia Jurkat cells were untreated (Control) or pretreated with MCD and then incubated with 10 μM [ 3 H]edelfosine for 1 h. Drug uptake was determined as shown in the Materials and Methods section. Data shown are means ± SD of three independent experiments performed. Asterisks denote that the differences between the indicated groups are statistically significant. (**) P <0.01. (***) P <0.001.

Article Snippet: We also found that edelfosine was very efficient in promoting cell death in additional human leukemic cell lines, including human T-cell acute lymphoblastic leukemia (T-ALL) cell lines Jurkat (53.4 ± 6.2% apoptosis) and CEM-C7H2 (58.2 ± 5.9% apoptosis).

Techniques: Control, Incubation, Flow Cytometry

Journal: PLoS Neglected Tropical Diseases

Article Title: Mitochondria and lipid raft-located F O F 1 -ATP synthase as major therapeutic targets in the antileishmanial and anticancer activities of ether lipid edelfosine

doi: 10.1371/journal.pntd.0005805

Figure Lengend Snippet: ( A ) Jurkat cells untreated (Control) and treated with 10 μM edelfosine for 9 h were lysed in 1% Triton X-100 and subjected to discontinuous sucrose density gradient centrifugation. Individual fractions were subjected to SDS-PAGE, and location of GM1 was determined using CTx B subunit conjugated with horseradish peroxidase. ( B ) Proteins from lipid rafts of untreated control and edelfosine-treated Jurkat cells were subjected to two-dimensional gel electrophoresis followed by MALDI-TOF analysis. Mitochondrial F O F 1 -ATP synthase β subunit is indicated by an arrow. ( C ) Mass spectrum of the tryptic peptides of the F O F 1 -ATP synthase β subunit spot. Mass value (m/z) and putative amino acid position assignments are indicated above peaks. ( Inset ) Peptide coverage map of human F O F 1 -ATP synthase β subunit; the peptides used for identification are highlighted in bold characters and underlined. ( D ) Jurkat cells were untreated (Control) or preincubated with 10 μM oligomycin for 1 h and then incubated in the absence or presence of 10 μM edelfosine for 9 h, and ΔΨ m disruption (Low ΔΨ m ) and DNA breakdown (hypodiploids) were evaluated. Data shown are means ± SD or representative of three independent experiments. Asterisks denote that the differences between the indicated groups are statistically significant. (**) P <0.01.

Article Snippet: We also found that edelfosine was very efficient in promoting cell death in additional human leukemic cell lines, including human T-cell acute lymphoblastic leukemia (T-ALL) cell lines Jurkat (53.4 ± 6.2% apoptosis) and CEM-C7H2 (58.2 ± 5.9% apoptosis).

Techniques: Control, Gradient Centrifugation, SDS Page, Two-Dimensional Gel Electrophoresis, Electrophoresis, Incubation, Disruption

Journal: Molecular bioSystems

Article Title: Metabolomics identifies the intersection of phosphoethanolamine with menaquinone-triggered apoptosis in an in vitro model of leukemia

doi: 10.1039/c5mb00237k

Figure Lengend Snippet: Table 1

Article Snippet: Expansion cell culture protocols The human acute T cell lymphoblastic leukemia cell line Jurkat (ATCC CRL-2570) was purchased from the American Type Culture Collection (ATCC) and cultured in RPMI-1640 with 2mM L-glutamine supplemented with 10% fetal bovine serum (FBS) and 1% antibiotic and antimycotic (AA) solution in T25 flasks.

Techniques: Control