Observation of measles virus cell-to-cell spread in astrocytoma cells by using a green fluorescent protein-expressing recombinant virus
Observation of measles virus cell-to-cell spread in astrocytoma cells by using a green fluorescent protein-expressing recombinant virus
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DOI：
10.1128/jvi.73.11.9568-9575.1999
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发表时间：
1999-11-01
影响因子：
5.4
通讯作者：
Rima, BK
中科院分区：
文献类型：
--
作者：
Duprex, WP;McQuaid, S;Rima, BK
一种表达增强型绿色荧光蛋白（EGFP）的重组麻疹病毒（MV）已被拯救。该病毒MVeGFP从位于编码麻疹病毒核衣壳蛋白的基因之前的额外转录单位表达报告基因。重组病毒感染人星形细胞瘤细胞（GCCM）。免疫细胞化学（ICC）和EGFP自发荧光显示，EGFP是细胞感染的早期和非常敏感的指标。经常观察到EGFP阳性和ICC阴性的细胞。共聚焦显微镜用于间接观察MV感染的GCCM细胞，并随后在真实的时间内跟踪细胞间的扩散。这些星形细胞瘤细胞具有延伸的突起，在许多情况下这些突起密切相关。这些过程似乎在细胞间传播中起重要作用，并且观察到MVeGFP在感染周围细胞中利用它们。在预期为均匀单层的细胞间扩散中观察到异质性。在组织培养中，物理限制控制了在广泛的细胞融合后形成的合胞体的完整性。当大约50个细胞融合时，合胞体迅速解体，许多受感染的细胞脱落。观察到残留的粘附EGFP阳性细胞继续参与周围细胞的感染或保持EGFP阳性但不再参与MV感染向邻近细胞的传播。
A recombinant measles virus (MV) which expresses enhanced green fluorescent protein (EGFP) has been rescued. This virus, MVeGFP, expresses the reporter gene from an additional transcription unit which is located prior to the gene encoding the measles virus nucleocapsid protein. The recombinant virus was used to infect human astrocytoma cells (GCCM). Immunocytochemistry (ICC) together with EGFP autofluorescence showed that EGFP is both an early and very sensitive indicator of cell infection. Cells that were EGFP-positive and ICC-negative were frequently observed. Confocal microscopy was used to indirectly visualize MV infection of GCCM cells and to subsequently follow cell-to-cell spread in real time. These astrocytoma cells have extended processes, which in many cases are intimately associated. The processes appear to have an important role in cell-to-cell spread, and MVeGFP was observed to utilize them in the infection of surrounding cells. Heterogeneity was seen in cell-to-cell spread in what was expected to be a homogeneous monolayer. In tissue culture, physical constraints govern the integrity of the syncytia which are formed upon extensive cell fusion. When around 50 cells were fused, the syncytia rapidly disintegrated and many of the infected cells detached. Residual adherent EGFP-positive cells were seen to either continue to be involved in the infection of surrounding cells or to remain EGFP positive but no longer participate in the transmission of MV infection to neighboring cells.