Abstract:

BACKGROUND: In the study of intervertebral disc tissue engineering, the intervertebral disc cells modified by gene engineering have the advantages of fast growth and secreting more extracellular matrix as the seed cells. Investigating the infection efficiency of intervertebral disc nucleus pulposus (NP) cells infected by lentivirus will have practical value.
OBJECTIVE: To detect the infection condition and infection parameter of human NP cells infected by different titers of green fluorescence protein mediated by lentivirus. To provide the experimental evidence for the lentivirus infection as gene vector to the human NP cells.
METHODS: Human NP cells were isolated and cultured from two idiopathic scoliosis patients via enzyme digestion. High titer of lentivirus-mediated green fluorescence protein (GFP) gene was constructed by gene reconstitution technique. The passage 2 NP cells were infected with different multiplicities of infection (MOIs), and green fluorescence was observed using an inverted fluorescence microscope. The infection efficiency was detected by flow cytometry.
RESULTS AND CONCLUSION: After 4 days, the infection efficiency of the passage 2 NP cells infected with different MOIs (1, 10, 50 and 100) was 32.1%, 41.1%, 54.2% and 86.8%, respectively. The expression of GFP in the NP cells was maintained above 60% until the passage 5. The GFP gene mediated by lentivirus was effectively expressed in human NP cells for a long time period. It may provide evidence that lentivirus-mediated GFP gene is highly effectively expressed in human NP cells.