Normal and tumor-bearing host macrophage responses: variability in accessory function, surface markers, and cell-cycle kinetics
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D Askew1, A D Yurochko, C J Burger
1Department of Biology, Virginia Polytechnic Institute and State University, Blacksburg 24061.
Abstract:
Normal and tumor-bearing host (TBH) peritoneal macrophage (M phi) responses to in vitro lipopolysaccharide (LPS) treatment were measured by assessing functional and phenotypic changes. Both normal and TBH untreated M phi suppressed mixed lymphocyte reaction (MLR) reactivity at all concentrations. Normal host M phi treated with LPS for 3 h were suppressive at all concentrations. TBH M phi treated with LPS for 3 h were not suppressive in the MLR until more than 5% were added. Surprisingly, 24 h treatment of normal and TBH M phi with LPS induced cells that significantly enhanced MLR reactivity when added at 2% or 5%. These cells were not suppressive until a 20% M phi concentration was reached. LPS treatment of normal and TBH M phi changed the percentage of cells expressing the surface markers Mac-1, -2, -3, and Ia as determined by flow cytometry. Normal host peritoneal M phi treated with LPS for 3 h had decreased Mac-1 and -3 expression, but there was no change in Mac-2 or Ia. Plating for 24 h did not change the percentage of M phi expressing Mac-1, -3, or Ia but did cause an increase in Mac-2+ M phi. Treatment of normal host M phi with LPS for 24 h led to a decrease in Mac-1+ and Ia+ M phi, no change in Mac-3+ M phi, but an increase in Mac-2+ M phi. LPS treatment of TBH M phi for 3 h decreased the number of Mac-1+ M phi, but Mac-2+, -3+, or Ia+ M phi numbers did not change. Plating TBH M phi for 24 h caused a decrease in the number of Mac-1+ M phi, no change in Mac-3+ or Ia+ M phi, but an increase in Mac-2+ M phi. Treatment with LPS for 24 h led to no change in the number of Mac-1+, -3+, or Ia+ TBH M phi, but Mac-2+ M phi increased. The phenotypic and functional changes after LPS treatment led us to ask if these changes were detectable at the level of DNA and RNA. Flow cytometric analysis of acridine orange-stained M phi was used to measure DNA and RNA levels. This analysis determines M phi cell-cycle kinetics and estimates their RNA synthesis. In normal host M phi, a 3-h LPS treatment caused a decrease of cells in G0/G1 but an insignificant change in RNA levels.(ABSTRACT TRUNCATED AT 250 WORDS)
Insights
Lipopolysaccharide (LPS) alters macrophage function, with short-term treatment suppressing immune responses and long-term treatment enhancing them. These changes in peritoneal macrophages from normal and tumor-bearing hosts correlate with altered surface marker expression.
Area of Science:
- Immunology
- Cell Biology
Background:
- Peritoneal macrophages (M phi) play crucial roles in immune responses.
- Tumor-bearing hosts (TBH) exhibit altered immune cell functions.
- Lipopolysaccharide (LPS) is a potent immune stimulant.
Purpose of the Study:
- To investigate the functional and phenotypic changes in peritoneal macrophages from normal and TBH after LPS treatment.
- To assess the impact of LPS treatment duration on macrophage activity.
Main Methods:
- In vitro LPS treatment of normal and TBH peritoneal macrophages.
- Mixed lymphocyte reaction (MLR) assays to measure functional changes.
- Flow cytometry to analyze surface marker expression (Mac-1, -2, -3, Ia).
- Acridine orange staining and flow cytometry for DNA and RNA levels.
Main Results:
- Short-term (3h) LPS treatment of normal macrophages suppressed MLR, while TBH macrophages showed reduced suppression.
- Long-term (24h) LPS treatment of both normal and TBH macrophages enhanced MLR reactivity.
- LPS treatment altered the expression of macrophage surface markers, with varying effects based on treatment duration and host status.
- Short-term LPS treatment decreased cells in G0/G1 phase in normal macrophages with minimal RNA level changes.
Conclusions:
- Macrophage responses to LPS are complex and depend on treatment duration and host tumor status.
- Phenotypic changes, including surface marker expression, accompany functional alterations.
- Further investigation into DNA and RNA level changes may elucidate underlying mechanisms.