Construction of human recombinant lentiviral vector of GP73and its expression in DC2.4 cells

Objective:This study aimed to prepare Golgi protein-73(GP73)-DC vaccine by constructing human lentiviral vector of GP73and transfecting this vector in vitro into a murine dendritic cell line DC 2.4.This will lay the foundation for further study on targeted therapy of hepatocellular carcinoma using GP73-DC vaccine.Methods:Human GP73 gene was cloned into lentivirus GV358vector and validated by polymerase chain reaction(PCR)and sequencing.The validated GV358-GP73vector was then cotransfected with pHelper 1.0plasmid into 293Tcells,in order to prepare lentivirus GP73vector.Finally,the protein expression of GP73 was achieved by transfecting the lentivirus GP73vector into DC 2.4cells in vitro and determined by Western blotting.Results:PCR and sequencing confirmed that the GV358-GP73lentivirus vector was constructed correctly,and the titer level reached 2×108TU/mL.After transfection in DC 2.4cells,the GP73protein level was significantly enhanced.Conclusion:The lentivirus GV538-GP73vector was constructed successfully,and could be effectively expressed in DC 2.4cells.

A Study on the Mechanism of Bruceine D in the Treatment of Non-small Cell Lung Cancer H1299 Cells

Background:Non-small cell lung cancer(NSCLC)is considered one of the leading causes of cancer-related death.Despite the availability of drugs for the treatment of NSCLC,the need for the development of novel agents with high efficiency and fewer adverse effects remains unmet.The natural compound bruceine D(BD)is widely recognized for its notable anti-inflammatory,antiparasitic,and hypoglycemic activities.However,it is unclear whether BD can be used as a novel agent for NSCLC treatment.Materials and Methods:MTT and colony formation assays were used to assess the antiproliferative effect of BD on NSCLC cells.Wound healing and transwell assays were performed to determine the effect of BD on the migration and invasion of H1299 cells,respectively.Western blotting assay was used to detect the expression levels of proteins.Results:We demonstrated that BD significantly inhibited the proliferation of H1299,A549,and H226 cells with respective IC50 values of 6.06±0.52,7.15±0.90,and 7.21±0.75μM.In addition,BD suppressed colony formation of H1299 cells in a dose-dependent manner.Following treatment with BD,the migration and invasive capabilities of H1299 cells were significantly inhibited in a dose-and time-dependent manner.Moreover,the results of Western blotting demonstrated that BD treatment resulted in the upregulation of the protein expression of E-cadherin and downregulation of the expression of N-cadherin,twist,snail,integrinαv,integrinβ4,matrix metalloproteinase-7,andβ-catenin proteins.Conclusion:BD inhibits proliferation,migration,and invasion of NSCLC cells;therefore,BD may be considered for its potential in adjuvant therapy for NSCLC.