ABSTRACT
@# Objective: To investigate the effects of Wilms’tumor 1-associating protein (WTAP) on proliferation, migration and invasion of human lung adenocarcinoma A549 cells. Methods: Human lung adenocarcinoma cell line A549 and HEK293T cells were chosen for this study. Two sets of shWTAP interference sequences were designed to construct lentiviral vector plasmid. Human lung adenocarcinomaA549 cells were infected after packaging lentivirus in HEK293T cells, and the control group was transfected with 277 empty vector plasmid. The mRNAand protein expression levels of WTAPinA549 cells were detected by qPCR and WB. Changes in proliferation, migration and invasion of A549 cells were detected by BrdU assay, cell scratch healing assay and Transwell assay, respectively. Results: Two plasmids, shWTAP-1 and shWTAP-2, were successfully constructed. Compared with the control group, the mRNA and protein expression levels of WTAP were significantly down-regulated inA549 cells with WTAP knockdown (both P<0.05), and the proliferation, migration and invasion ability of cells were significantly decreased (all P<0.05). Conclusion: Knockdown of WTAP significantly inhibited the proliferation, migration and invasion of human lung adenocarcinoma A549 cells. The expression of WTAP gene is associated with the occurrence and development of lung adenocarcinoma. WTAP may be a potential target for the diagnosis and treatment of lung adenocarcinoma.
ABSTRACT
@#[Abstract] Objective: To establish a chimeric antigen receptor（CAR）modified T cells specifically targeting CD19 molecule (CD19CAR-T cells) and to testify their in vitro killing effect on target cells. Methods: CD19-CAR fragments yielded by PCR were constructed into pCDH-GFP lentiviral vectors by molecular cloning technology. The packaged lentiviral particles were transducted into CD3+ T cells of donors. Transduction efficiency was measured by flow cytometry and PCR. The in vitro cytotoxicity of obtained CD19CAR-T cells against CD19+ Ramos cells was tested by 7-AAD staining. Results: The amplification folds of CD3+ T cells increased to (78.8± 23.2) folds after in vitro culture for 10 days, and about (58.3±5.4)% cells expressing GFP.About (57.4±9.3)% CD19+Ramos cells were specifically killed by the CD19-CAR-T cells in vitro at the E∶T ratio of 5∶1. Conclusion: This study successfully established an effective method for constructing and amplifying CD19-CAR-T cells in vitro, which showed profound efficiency and specific cytotoxity against CD19+ Ramos cells.And this report might provide an experimental evidence for clinical treatment of CD19+ B cell neoplasmas.