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Temporal Analysis of the Nuclear-to-cytoplasmic Translocation of a Herpes Simplex Virus 1 Protein by Immunofluorescent Confocal Microscopy
Published on: November 4, 2018
[Temporal expression of HCMV IE1 and pp65 in human glioma U(251) cells]
Jian-Hua Li1, Ying Fu, Li-Yu Chen
1Department of Microbiology, Xiangya School of Medicine, Central South University, Changsha 410078, China.
Objective:
To determine whether U(251) cells are permissive for human cytomegalovirus (HCMV), and to investigate the characteristics of temporal expression of proteins IE1 and pp65.
Methods:
U(251) cells were infected with HCMV, and then the cells were observed under the transmission electronic microscope, and the viral nucleic acid was detected by PCR, and the expression levels of IE1 and pp65 were analyzed by immunohistochemical assay with anti-IE1 monoclonal antibody and anti-pp65 monoclonal antibody at various time spost infection.
Results:
Morphological changes of the infected cells appeared under the transmission electron microscope. The viral nucleic acid was detected successfully by PCR. The expression of IE1 was detected firstly at 4h post infection, and reached a peak within 14h, and then decreased. The incoming pp65 was detected at 1h, the low expression levels of pp65 were detected firstly at 4h, and they could remain relatively constant through 96 h, but the maximum expression occurred at 120 h.
Conclusion:
Human glioma U(251) cells are permissive for HCMV, the temporal cascade of HCMV gene expression can be observed in the infected U(251) cells, but it is delayed obviously in the human fibroblast.
Insights
Human glioma U(251) cells support human cytomegalovirus (HCMV) replication. Temporal expression of HCMV proteins IE1 and pp65 was observed, though delayed compared to human fibroblasts.
Area of Science:
- Virology
- Cell Biology
- Neuroscience
Context:
- Human cytomegalovirus (HCMV) is a ubiquitous herpesvirus with significant clinical implications, particularly in immunocompromised individuals.
- Understanding HCMV tropism and replication dynamics in different cell types is crucial for developing antiviral strategies.
- Glioma cells, such as U(251), represent a potential niche for viral persistence.
Purpose:
- To ascertain the permissivity of U(251) glioma cells to HCMV infection.
- To characterize the temporal expression patterns of key HCMV proteins, IE1 and pp65, in U(251) cells.
Summary:
- U(251) cells were infected with HCMV and analyzed using transmission electron microscopy, PCR, and immunohistochemistry.
- HCMV infection induced observable morphological changes and viral nucleic acid presence.
- IE1 expression peaked within 14 hours post-infection, while pp65 showed a delayed but sustained expression pattern, peaking at 120 hours.
Impact:
- Demonstrates that U(251) glioma cells are permissive to HCMV infection.
- Reveals a distinct temporal cascade of HCMV IE1 and pp65 protein expression in U(251) cells.
- Highlights a potential delay in HCMV gene expression kinetics in U(251) cells compared to human fibroblasts, suggesting cell-type-specific regulatory mechanisms.
