Release of replication-deficient retroviruses from a packaging cell line: Interaction with glioma tumor spheroids in vitro
✍ Scribed by Frits Thorsen; Therese Visted; Paulina Lehtolainen; Seppo Ylä-Herttuala; Rolf Bjerkvig
- Publisher
- John Wiley and Sons
- Year
- 1997
- Tongue
- French
- Weight
- 267 KB
- Volume
- 71
- Category
- Article
- ISSN
- 0020-7136
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✦ Synopsis
The present study describes how various growth conditions affect gene expression and virus production from a retroviral packaging cell line (Liz 9), grown as monolayers and as multicellular spheroids. In addition, to study the direct interaction between packaging cells and tumor tissue of glioma origin, Liz 9 spheroids were confronted with tumor spheroids derived from a human glioma cell line, GaMg. The results show a progressive gene transfer into the tumor tissue, with 9% transfection efficacy after 5 days of co-culture. In comparison, no gene transfer was observed when the Liz 9 spheroids were confronted with normal brain-cell aggregates. The Liz 9 spheroids established from early-passage cultures (passages 7-14) showed limited growth during 28 days, whereas those initiated from late-passage monolayer cultures (passages 39-49) showed extensive growth. Flow-cytometric DNA profiles of monolayers and of spheroids indicated no difference in cell-cycle distribution or ploidy between early and late passages. A cell-viability assay using scanning confocal microscopy revealed mostly viable cells in the Liz 9 spheroids, with only a few dead cells scattered within the structures. The lacZ-gene expression was maintained in early-and in latepassage cultures. In comparison, in Liz 9 early-passage monolayers, the virus titer was 3.1 3 10 4 6 0.4 3 10 4 CFU/ml, whereas no virus titer was found in late-passage cultures. The virus titer from the Liz 9 spheroids was found to be between 10 3 and 10 4 CFU/ml. It is concluded that the virus production from packaging cells may vary, depending on passage number and tissue-culture conditions. In the present study, this is demonstrated by a complete loss in virus titer during prolonged culture of packaging cells. In addition, the 3-dimensional confrontation system described allows direct visualization of how packaging cells interact with tumor tissue. Thus, the co-culture system represents a model for studying the efficiency of packaging cells in transfecting heterogeneous tumor tissue in vitro.