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The effects of acetylated low-density lipoproteins on fluid-phase pinocytosis by macrophages
1Division of Biomedical Sciences, King's College London, England.
Abstract:
A simple method has been set up to measure the rate of fluid-phase pinocytosis in resident mouse peritoneal macrophages in culture. The method uses 125I-labelled polyvinylpyrrolidone as a nondegradable marker of fluid-phase pinocytosis. The accumulation of 125I-labelled polyvinylpyrrolidone by the cells was directly proportional to its concentration in the culture medium up to at least 200 micrograms/ml. The estimates of the rate of fluid-phase pinocytosis were reproducible within each experiment (coefficient of variation 8.5%) but varied between individual experiments. Fluid-phase pinocytosis was undetectable at 4 degrees C and reduced greatly at 37 degrees C by metabolic inhibitors and 1 mM ZnSO4. High concentrations of human acetylated low-density lipoproteins, which are taken up rapidly by macrophages, decreased the rate of fluid-phase pinocytosis by up to about 70%. The inhibition was seen after only 2 h of incubation. Unmodified low-density lipoproteins, which are taken up only slowly by macrophages, did not usually inhibit fluid-phase pinocytosis (in fact, they sometimes increased it). Modified low-density lipoprotein uptake, leading to massive lipid accumulation in macrophages in the arterial wall, has been postulated to be involved in the pathogenesis of atherosclerosis. This study raises the possibility that the rate of fluid-phase pinocytosis in these lipid-laden arterial macrophages may be reduced.
Insights
Researchers developed a method to measure fluid-phase pinocytosis in macrophages using a radiolabeled marker. Acetylated low-density lipoproteins significantly reduced this process, suggesting a potential role in atherosclerosis.
Area of Science:
- Cell Biology
- Biochemistry
- Immunology
Background:
- Pinocytosis is a key cellular process for internalizing extracellular fluid and solutes.
- Macrophages play critical roles in immunity and tissue homeostasis, including lipid metabolism.
- Altered pinocytosis in macrophages is implicated in various pathological conditions, including atherosclerosis.
Purpose of the Study:
- To establish a reproducible method for quantifying fluid-phase pinocytosis in cultured mouse macrophages.
- To investigate the effects of modified and unmodified low-density lipoproteins (LDL) on pinocytosis rates.
- To explore the potential implications of altered pinocytosis in the context of atherosclerosis pathogenesis.
Main Methods:
- Utilized 125I-labelled polyvinylpyrrolidone (PVP) as a non-degradable marker for fluid-phase pinocytosis.
- Measured the accumulation of 125I-PVP by resident mouse peritoneal macrophages in vitro.
- Assessed the impact of temperature, metabolic inhibitors, ZnSO4, and different LDL modifications on pinocytosis rates.
Main Results:
- A dose-dependent accumulation of 125I-PVP was observed, establishing a reliable assay.
- Fluid-phase pinocytosis was significantly inhibited by metabolic inhibitors, ZnSO4, and notably, by acetylated LDL (up to 70%).
- Unmodified LDL did not inhibit, and sometimes enhanced, fluid-phase pinocytosis.
Conclusions:
- A robust method for measuring macrophage fluid-phase pinocytosis was developed.
- Rapid uptake of acetylated LDL markedly impairs fluid-phase pinocytosis in macrophages.
- Reduced pinocytosis in lipid-laden macrophages could contribute to the pathogenesis of atherosclerosis.