Related Experiment Videos
Morphological changes in HAV-infected Frp/3 cells and immunolocalization of HAAg
A Tinari1, F M Ruggeri, M Divizia
1Department of Ultrastructures, Istituto Superiore di Sanità, Rome, Italy.
Abstract:
Electron and immunoelectron microscopic studies were carried out on HAV-infected Frp/3 cells. The infection led to a distinctive cytopathic effect (CPE) arising on day 3 up to the complete detachment of monolayers on day 7. Infected cells exhibited progressive modifications, beginning from the formation of long helical polyribosomes. Subsequently, hypertrophy, cisternal dilatation and degranulation of the RER could be observed. Furthermore, the formation of concentric membranous bodies (CMB), large myelin-like structures and annulate lamellae could be revealed at later times of infection. 24-27 nm virus-like particles were observed within cytoplasmic vesicles or outside extensively degenerated cells. Indirect immunoperoxidase staining were used to localize HAV antigen (HAAg) in thin sectioned infected Frp/3 cells. Vesicular inclusion bodies, often seen to contain electron-lucent particles, resulted darkly stained as well as tracts of the RER and myelin-like structures. Negatively stained preparations from cell lysates revealed small clusters of HAV particles which sometimes appeared to be still associated with residual membrane fragments. Our findings seem to suggest that HAV replication occurs in close association with cytoplasmic membranes and a direct involvement of the RER seems to be demonstrated.
Insights
Hepatitis A virus (HAV) infection causes significant cell damage in Frp/3 cells, marked by changes in the endoplasmic reticulum and the formation of virus particles. Replication appears closely linked to cellular membranes, particularly the RER.
Area of Science:
- Hepatology
- Virology
- Cell Biology
Background:
- Hepatitis A virus (HAV) is a significant human pathogen.
- Understanding HAV replication mechanisms is crucial for developing antiviral strategies.
- HAV infection induces characteristic cytopathic effects in host cells.
Purpose of the Study:
- To investigate the ultrastructural changes in HAV-infected Frp/3 cells.
- To elucidate the role of cellular membranes in HAV replication.
- To localize HAV antigen (HAAg) within infected cells.
Main Methods:
- Electron microscopy (EM) and immunoelectron microscopy (IEM) were employed.
- Frp/3 cells were infected with HAV and analyzed at various time points.
- Indirect immunoperoxidase staining was used for antigen localization.
Main Results:
- Distinct cytopathic effects (CPE) were observed, including cell detachment.
- Progressive modifications in the rough endoplasmic reticulum (RER) were noted, such as polyribosome formation, hypertrophy, dilatation, and degranulation.
- Virus-like particles (24-27 nm) were found in cytoplasmic vesicles and extracellularly.
- HAV antigen (HAAg) was localized to vesicular inclusion bodies, RER, and myelin-like structures.
- HAV replication appears associated with cytoplasmic membranes, with a demonstrated involvement of the RER.
Conclusions:
- HAV infection induces profound ultrastructural alterations in Frp/3 cells.
- The RER and cytoplasmic membranes are likely critical sites for HAV replication.
- These findings provide insights into the intracellular lifecycle of HAV.