Related Experiment Videos
Differential down-regulation of CD95 or CD95L in chronically HIV-infected cells of monocytic or lymphocytic origin:
M Pinti1, J Pedrazzi, F Benatti
1Department of Internal Medicine, Infectious Disease Clinics, University of Modena, Reggio Emilia School of Medicine, Italy.
Insights
Human immunodeficiency virus type 1 (HIV-1) infection affects CD95/CD95L expression differently in lymphoid and monocytic cells. HIV-1 alters CD95L transcription in lymphoid cells and CD95 expression in monocytic cells, suggesting varied anti-apoptotic strategies.
Area of Science:
- Immunology
- Virology
- Cell Biology
Background:
- Chronic human immunodeficiency virus type 1 (HIV-1) infection impacts cellular functions, including apoptosis regulation.
- CD95 (Fas receptor) and its ligand CD95L (FasL) are key mediators of apoptosis.
- Understanding HIV-1's modulation of CD95/CD95L is crucial for deciphering viral pathogenesis and host immune responses.
Purpose of the Study:
- To investigate the expression of CD95 and CD95L in HIV-1 infected lymphoid (ACH-2) and monocytic (U1) cell lines.
- To determine if HIV-1 infection differentially affects CD95/CD95L expression based on cell lineage.
- To elucidate the transcriptional and post-transcriptional mechanisms underlying these changes.
Main Methods:
- Flow cytometry was used to analyze plasma membrane expression of CD95 and CD95L.
- Quantitative competitive RT-PCR was developed and employed to measure CD95L mRNA levels.
- Cell lines used included ACH-2 (HIV-1 infected lymphoid), U1 (HIV-1 infected monocytic), A301 (parental lymphoid), and U937 (parental monocytic).
Main Results:
- CD95 expression was similar on A301 and ACH-2 cells, but decreased on U1 cells.
- Plasma membrane CD95L was undetectable; intracellular CD95L was present in A301 and ACH-2 cells, but not U937 or U1 cells.
- ACH-2 cells showed significantly reduced CD95L mRNA compared to A301 cells, indicating transcriptional inhibition by HIV-1.
- U1 cells exhibited decreased CD95 expression, while ACH-2 cells showed reduced CD95L expression.
Conclusions:
- Chronic HIV-1 infection differentially regulates CD95/CD95L expression in lymphoid and monocytic cells.
- In lymphoid cells, HIV-1 inhibits CD95L expression at the transcriptional level, potentially impairing cytotoxic functions.
- In monocytic cells, HIV-1 reduces CD95 expression, possibly as an anti-apoptotic survival mechanism within viral reservoirs.
Abstract:
We analysed the expression of CD95/CD95L in two widely used models for studying the cellular effects of chronic infection with human immunodeficiency virus type 1 (HIV-1), i.e. ACH-2 cells, derived from the lymphocytic cell line A301, and U1, derived from monocytic U937 cells. A301 and ACH-2 mounted the same amount of plasma membrane CD95, while U1 had a consistent decrease in CD95 expression. Using different antibodies, we failed to detect the plasma membrane form of its ligand, CD95L, but we could see the intracellular presence of that molecule in A301 cells and, to a lesser extent, in ACH-2 cells, but not in U937 or U1 cells. To confirm the cytofluorimetric data and quantify the expression of CD95L at the RNA level, we developed a quantitative competitive RT-PCR assay. The HUT78 cell line had about 50,000 copies mRNA/1000 cells, three times more after induction with a phorbol ester and ionomycin. ACH-2 expressed about 400- (basal) or 10- (induced) fold less CD95L mRNA than the parental cell line A301; U937 and U1 were below the limit of detection. In cells of lymphoid origin (ACH-2) chronic HIV infection inhibits the expression of CD95L, the phenomenon occurring at the transcriptional level. In cells of monocytic origin (U1) the infection decreases the plasma membrane expression of CD95. This suggests that HIV could trigger different anti-apoptotic strategies which likely depend upon the cell line which is infected. In monocytic cells which act as a viral reservoir, the expression of the molecule whose binding triggers apoptosis decreases, while in lymphoid cells, capable of exerting cytotoxicity, the expression of a molecule which induces apoptosis is reduced.