Journal: Translational Oncology

Article Title: Exogenous dihomo-γ-linolenic acid triggers ferroptosis via ACSL4-mediated lipid metabolic reprogramming in acute myeloid leukemia cells

doi: 10.1016/j.tranon.2024.102227

Figure Lengend Snippet: ACSL4 mediates DGLA-induced ferroptosis. ( a ) Western blot showing ACSL4 protein levels in Kasumi-1 cells expressing negative control sgRNA (sgNC) or ACSL4-targeting sgRNA (sgACSL4). ( b ) Cell viability of Kasumi-1 cells expressing sgNC or sgACSL4 treated with erastin for 24 h. ( c ) Cell viability of Kasumi-1 cells expressing sgNC or sgACSL4 treated with DGLA for 24 h. ( d - f )The levels of lipid ROS ( d ), MDA ( e ), and Fe 2+ ( f ) in Kasumi-1 cells expressing sgNC or sgACSL4 treated with DGLA (20 µM) for 24 h. ( g ) TEM images of Kasumi-1 cells expressing sgNC or sgACSL4 treated with DGLA (20 μM) for 24 h. Scale bars: upper panel, 1 μm; lower panel, 500 nm. White arrows indicate normal mitochondria, red arrows indicate abnormal mitochondrial structure, yellow arrows indicate lipid droplets. Data are shown as mean ± SD, n = 3 biologically independent experiments, ** p < 0.01, *** p <0.001.

Article Snippet: The corresponding sgRNAs were designed and cloned into the vector, and negative control sgRNAs (sgNC) were generated as controls (Genechem, China).

Techniques: Western Blot, Expressing, Negative Control

Journal: Translational Oncology

Article Title: Exogenous dihomo-γ-linolenic acid triggers ferroptosis via ACSL4-mediated lipid metabolic reprogramming in acute myeloid leukemia cells

doi: 10.1016/j.tranon.2024.102227

Figure Lengend Snippet: ACSL4 reprograms DGLA-associated lipids. ( a ) Lipids were analyzed by LC-MS/MS. Heatmap shows lipids fold-changes in Kasumi-1 cells expressing sgNC or sgACSL4 treated with DGLA (20 µM). ( b ) Distribution of fatty acid chains of species from lipids in ( a ) that were down-regulated upon DGLA treatment. ( c ) Pie chart, by PUFA chain subclass, showing proportions of each lipid species in the down-regulated lipids upon DGLA treatment. ( d ) Pie charts, by phospholipid subclass, showing proportions of lipid species containing DGLA that were identified in ( a ). Data are shown as mean ± SD, n = 3 biologically independent experiments.

Article Snippet: The corresponding sgRNAs were designed and cloned into the vector, and negative control sgRNAs (sgNC) were generated as controls (Genechem, China).

Techniques: Liquid Chromatography with Mass Spectroscopy, Expressing

Journal: Translational Oncology

Article Title: Exogenous dihomo-γ-linolenic acid triggers ferroptosis via ACSL4-mediated lipid metabolic reprogramming in acute myeloid leukemia cells

doi: 10.1016/j.tranon.2024.102227

Figure Lengend Snippet: Deletion of ACSL4 inhibits the anticancer activity of DGLA in vivo . ( a ) Hypodermic injection of Kasumi-1 cells stably transfected with sgNC or sgACSL4 into BALB/c nude mice and treated with DGLA or vehicle, established subcutaneous xenograft tumors (n = 6 mice/group). ( b ) Tumor images show the relative sizes of the xenograft tumors formed by Kasumi-1 cells expressing sgNC or sgACSL4 and treated with DGLA or vehicle on day 18. ( c ) Changes in tumor volumes over time for mice implanted with Kasumi-1 cells expressing sgNC or sgACSL4 and treated with DGLA or vehicle. ( d - e ) Measures of tumor volume ( d )and tumor weight ( e ) values of mice at the study endpoint. ( f - h ) The levels of lipid ROS ( f ), MDA ( g ), and Fe 2+ ( h ) in xenograft tumors treated with vehicle or DGLA for 18 days (n = 6 tumor samples from different animals per group). ( i ) TEM images of mitochondria ultrastructure in xenograft tumors treated with DGLA or vehicle for 18 days. Scale bars: upper panel, 1 μm; lower panel, 500 nm. White arrows indicate normal mitochondria, red arrows indicate abnormal mitochondrial structure, yellow arrows indicate lipid droplets. Data are shown as mean ± SD, ** p < 0.01, *** p <0.001.

Article Snippet: The corresponding sgRNAs were designed and cloned into the vector, and negative control sgRNAs (sgNC) were generated as controls (Genechem, China).

Techniques: Activity Assay, In Vivo, Injection, Stable Transfection, Transfection, Expressing

Journal: bioRxiv

Article Title: Deletion of the transcriptional regulator TFAP4 accelerates c-MYC-driven lymphomagenesis

doi: 10.1101/2022.12.19.520971

Figure Lengend Snippet: A, Schematic representation of hematopoietic reconstitution of lethally irradiated recipient mice with fetal liver derived donor HSPCs. Fetal liver cells from double transgenic Eμ-MYC/Cas9 E13.5 donor embryos (an abundant source of HSPCs) were lentivirally transduced with vectors encoding BFP or CFP as a marker and sgRNAs targeting Tfap4 (sgTfap4 ), a positive sgRNA targeting Trp53 (sgTrp53 ) or a negative non-targeting control sgRNA ( sgControl ). These transduced donor fetal liver cells were then transplanted into lethally irradiated C57BL/6-Ly5.1 recipient mice. Tumour-free survival of recipient mice was measured as days post-transplantation. Hematopoietic tissues and peripheral blood were harvested from tumour burdened recipient mice for further analysis. B, Kaplan-Meier survival curve showing tumour-free survival of mice transplanted with either of two vectors containing a sgTfap4 , a positive control sgTrp53 or a negative sgControl . N indicates number of sick mice/number of mice transplanted with sgRNA transduced HSPCs for each sgRNA. Median survival post transplantation in days is indicated in brackets. **** P < 0.0001.

Article Snippet: A vector constitutively expressing a negative control sgRNA targeting human NLRC5 (sgRNA-1 5’-GCTGCAGAAGTGTCAGCTCCAGG) and the BFP tag was also obtained from the Merck CRISPR glycerol stock arrayed whole mouse genome sgRNA library, and this vector was used for the pre-leukemic analyzes.

Techniques: Irradiation, Derivative Assay, Transgenic Assay, Transduction, Marker, Transplantation Assay, Positive Control