Journal: Nucleic Acids Research

Article Title: Antisense oligonucleotide-mediated correction of CFTR splicing improves chloride secretion in cystic fibrosis patient-derived bronchial epithelial cells

doi: 10.1093/nar/gkaa490

Figure Lengend Snippet: CFTR c.3718-2477C>T aberrant splicing and correction with a splice-switching antisense oligonucleotide. ( A ) Schematic of the aberrant splicing caused by the CFTR c.3718-2477C>T mutation in intron 22. The C>T mutation creates a de novo 5′ splice site resulting in the insertion into the mRNA of an 84 nucleotide pseudoexon (ψ-Ex) that has an in-frame stop codon. An ASO designed to mask the CFTR c.3718-2477C>T de novo splice site redirects splicing to the canonical splice site, restoring wild-type (WT) mRNA. ( B ) Sequence alignment of ASO-ψ to the ψ-Ex 5′ splice site found in CFTR c.3718-2477C>T. ( C ) RT-PCR analysis of CFTR splicing in a lymphoblast cell line from a CF patient homozygous for the CFTR c.3718-2477C>T mutation following transfection with vehicle only (–), a control, non-targeted ASO (ASO-C) or ASO-ψ targeting the ψ-Ex 5′ splice site. Cells were untreated (−) or treated (+) with puromycin before RNA isolation. GAPDH was analyzed as a control for total cellular RNA abundance. Splicing was quantified as the percent of RNA transcripts with ψ-Ex [(ψ-Ex/(ψ-Ex+WT))x100] and is shown below each lane.

Article Snippet: The human patient lymphoblast cell line (GM11860; Coriell Institute) was grown in RPMI media supplemented with 15% fetal bovine serum (FBS).

Techniques: Mutagenesis, Sequencing, Reverse Transcription Polymerase Chain Reaction, Transfection, Control, Isolation

Journal: Pathology and Oncology Research

Article Title: Knockdown of GPSM1 Inhibits the Proliferation and Promotes the Apoptosis of B-Cell Acute Lymphoblastic Leukemia Cells by Suppressing the ADCY6-RAPGEF3-JNK Signaling Pathway

doi: 10.3389/pore.2021.643376

Figure Lengend Snippet: Downregulation of GPSM1 suppressed cell proliferation, induced cell cycle arrest and promoted apoptosis in BALL-1 and Reh cells. (A) Expression of GPSM1 mRNA in the human B lymphoblast cell line HMy2.CIR and several human leukemia cell lines (BALL-1, Jurkat and Reh). GAPDH was used as an internal reference for normalization. * p < 0.05 vs. the HMy2.CIR group. (B) GPSM1 protein expression in the human B lymphoblast cell lineHMy2.CIR and several human leukemia cell lines (BALL-1, Jurkat and Reh) was investigated by western blotting. (C) The grayscale value of the GPSM1 protein band was quantified by Tanon Image software, and GAPDH was used as an internal reference. * p < 0.05 vs. the HMy2.CIR group. (D) Relative mRNA expression in sh-Con- and sh-GPSM1-transfected BALL-1 and Reh cells. (E) GPSM1 protein expression in sh-Con- and sh-GPSM1-transfected BALL-1 and Reh cells. (F) The grayscale value of the GPSM1 protein band was quantified by Tanon Image software, and GAPDH was used as an internal reference. * p < 0.05, compared with the sh-Con group. (G) The CCK-8 assay was used to evaluate the cell proliferation ability of BALL-1 and Reh cells transfected with sh-GPSM1 and sh-Con on four consecutive days. * p < 0.05, ** p < 0.01, compared with the sh-Con group. (H,I) The cell cycle was examined by PI staining using flow cytometry. (J) Cell apoptosis rates were assessed by flow cytometry after staining cells with annexin V-PE/7-AAD. Nonapoptotic cells: annexin V-PE − /7-AAD − , early apoptotic cells: annexin V-PE + /7-AAD − , late apoptotic cells: annexin V-PE + /7-AAD + . PE, phycoerythrin; 7-AAD, 7-amino-actinomycin D. (K) The total apoptosis rate is the sum of the early apoptosis rate and the late apoptosis rate. * p < 0.05, compared with the sh-Con group. All data are expressed as the mean ± SD.

Article Snippet: A human T-cell lymphoblastic leukemia cell line (Jurkat) was obtained from Procell Life Science and Technology Co., Ltd. A human lymphoblastic leukemia cell line (Reh) was obtained from Shanghai Genechem Co., Ltd (Shanghai, China).

Techniques: Expressing, Western Blot, Software, Transfection, CCK-8 Assay, Staining, Flow Cytometry

Journal: Pathology and Oncology Research

Article Title: Knockdown of GPSM1 Inhibits the Proliferation and Promotes the Apoptosis of B-Cell Acute Lymphoblastic Leukemia Cells by Suppressing the ADCY6-RAPGEF3-JNK Signaling Pathway

doi: 10.3389/pore.2021.643376

Figure Lengend Snippet: Downregulation of GPSM1 suppressed cell proliferation, induced cell cycle arrest and promoted apoptosis in BALL-1 and Reh cells. (A) Expression of GPSM1 mRNA in the human B lymphoblast cell line HMy2.CIR and several human leukemia cell lines (BALL-1, Jurkat and Reh). GAPDH was used as an internal reference for normalization. * p < 0.05 vs. the HMy2.CIR group. (B) GPSM1 protein expression in the human B lymphoblast cell lineHMy2.CIR and several human leukemia cell lines (BALL-1, Jurkat and Reh) was investigated by western blotting. (C) The grayscale value of the GPSM1 protein band was quantified by Tanon Image software, and GAPDH was used as an internal reference. * p < 0.05 vs. the HMy2.CIR group. (D) Relative mRNA expression in sh-Con- and sh-GPSM1-transfected BALL-1 and Reh cells. (E) GPSM1 protein expression in sh-Con- and sh-GPSM1-transfected BALL-1 and Reh cells. (F) The grayscale value of the GPSM1 protein band was quantified by Tanon Image software, and GAPDH was used as an internal reference. * p < 0.05, compared with the sh-Con group. (G) The CCK-8 assay was used to evaluate the cell proliferation ability of BALL-1 and Reh cells transfected with sh-GPSM1 and sh-Con on four consecutive days. * p < 0.05, ** p < 0.01, compared with the sh-Con group. (H,I) The cell cycle was examined by PI staining using flow cytometry. (J) Cell apoptosis rates were assessed by flow cytometry after staining cells with annexin V-PE/7-AAD. Nonapoptotic cells: annexin V-PE − /7-AAD − , early apoptotic cells: annexin V-PE + /7-AAD − , late apoptotic cells: annexin V-PE + /7-AAD + . PE, phycoerythrin; 7-AAD, 7-amino-actinomycin D. (K) The total apoptosis rate is the sum of the early apoptosis rate and the late apoptosis rate. * p < 0.05, compared with the sh-Con group. All data are expressed as the mean ± SD.

Article Snippet: A human T-cell lymphoblastic leukemia cell line (Jurkat) was obtained from Procell Life Science and Technology Co., Ltd. A human lymphoblastic leukemia cell line (Reh) was obtained from Shanghai Genechem Co., Ltd (Shanghai, China).

Techniques: Expressing, Western Blot, Software, Transfection, CCK-8 Assay, Staining, Flow Cytometry

Journal: Genes and Environment

Article Title: Weight of evidence approach using a TK gene mutation assay with human TK6 cells for follow-up of positive results in Ames tests: a collaborative study by MMS/JEMS

doi: 10.1186/s41021-021-00179-1

Figure Lengend Snippet: Differentially expressed proteins (DEP) identified in TK6 cells exposed to 4-nitroanthranilic acid. a A total of 1078 DEPs that differentially expressed compared to control groups were identified based on a log 2 fold change of ≥2. Effects of treatment time are greater than that of dose. b A total of 168 specific DEPs that differentially expressed compared to other groups were further extracted from the 420 DEPs in 24 h 800 μg/mL group based on a log 2 fold change of ≥1

Article Snippet: The human lymphoblast cell line TK6 was purchased from the Japanese Collection of Research Bioresources cell bank and the American Type Culture Collection.

Techniques: Control

Journal: Genes and Environment

Article Title: Weight of evidence approach using a TK gene mutation assay with human TK6 cells for follow-up of positive results in Ames tests: a collaborative study by MMS/JEMS

doi: 10.1186/s41021-021-00179-1

Figure Lengend Snippet: Target gene ontology analysis for TK6 cells exposed to 4-nitroanthranilic acid for 24 h. Whereas no significance was found in GO:0006974 (cellular response to DNA damage stimulus), GO:0006281 (DNA repair), and GO:0006979 (response to oxidative stress), statistically significance was found in GO:0008631 ~ intrinsic apoptotic signaling pathway in response to oxidative stress among DEPs in 24 h 800 μg/mL group and specific DEPs in 24 h 800 μg/mL group, indicating involvement with oxidative stress

Article Snippet: The human lymphoblast cell line TK6 was purchased from the Japanese Collection of Research Bioresources cell bank and the American Type Culture Collection.

Techniques:

Journal: Genes and Environment

Article Title: Weight of evidence approach using a TK gene mutation assay with human TK6 cells for follow-up of positive results in Ames tests: a collaborative study by MMS/JEMS

doi: 10.1186/s41021-021-00179-1

Figure Lengend Snippet: Possible mechanisms of oxidative stress produced by the redox reaction of amino- and nitro compounds in human TK6 cells

Article Snippet: The human lymphoblast cell line TK6 was purchased from the Japanese Collection of Research Bioresources cell bank and the American Type Culture Collection.

Techniques: Produced