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Updated: Jan 13, 2026
Isolation of Murine Peritoneal Macrophages to Carry Out Gene Expression Analysis Upon Toll-like Receptors Stimulation
Published on: April 29, 2015
THP-1-derived polarized macrophages differ by drug transporter transcriptomics, P-glycoprotein activity and
Katharina Hamburg1, Cindy Bay1, Jürgen Burhenne1
1Internal Medicine IX - Department of Clinical Pharmacology and Pharmacoepidemiology Medical Faculty Heidelberg, Heidelberg University Hospital, Im Neuenheimer Feld 410, 69120, Heidelberg, Germany.
Abstract:
Inferior efficacy of antibiotics (e.g. rifampicin) in M2 macrophage-dominant tuberculosis granuloma might be related to poor drug uptake into these cells, potentially mediated by altered expression and activity of drug transporters such as P-glycoprotein (P-gp, encoded by ABCB1). Consequently, THP-1 cells were differentiated (200 nM phorbol 12-myristate 13-acetate; 72 h) and polarized to M1 (50 ng/mL lipopolysaccharide, 20 ng/mL interferon-gamma; 48 h) or M2 (20 ng/mL interleukin-4, interleukin-13; 48 h) macrophages. Then, quantitative polymerase-chain reaction array-based transcriptional analysis, flow cytometry, and ultra-performance liquid chromatography coupled to tandem mass spectrometry were used to evaluate the impact of differentiation and polarization on the expression levels of 84 drug transporter genes, the efflux activity of P-gp, and the cellular rifampicin uptake kinetics. ABCB1 was enhanced 166-fold during M1 polarization and 55-fold during M2 polarization. P-gp efflux activity in M2 cells was 1.55-fold higher than in M1 cells (P < 0.05). Rifampicin uptake into M2 cells was lower than into M1 cells after 2 h (P < 0.01) or 4 h exposure (P = 0.06) to 0.05 µM rifampicin. Together, when compared to M1 macrophages, M2 cells exhibit a considerably altered fingerprint of drug transporter expression levels, including an enhanced expression and activity of P-gp, being accompanied by lowered rifampicin uptake.
Insights
Tuberculosis drug efficacy is reduced in M2 macrophages due to higher P-glycoprotein (P-gp) activity, which limits rifampicin uptake. This study investigated drug transporter changes in M1 versus M2 macrophages.
Area of Science:
- Pharmacology
- Immunology
- Cell Biology
Background:
- Antibiotic efficacy in tuberculosis (TB) granulomas, particularly those rich in M2 macrophages, is often suboptimal.
- This may stem from reduced drug penetration into M2 macrophages, potentially influenced by drug transporter activity, such as P-glycoprotein (P-gp).
Purpose of the Study:
- To investigate the impact of macrophage polarization (M1 vs. M2) on drug transporter expression and activity.
- To determine how these changes affect the cellular uptake of rifampicin, a key anti-TB drug.
Main Methods:
- THP-1 cells were differentiated and polarized into M1 or M2 macrophages.
- Quantitative polymerase-chain reaction arrays, flow cytometry, and ultra-performance liquid chromatography-tandem mass spectrometry were employed.
- Evaluated expression of 84 drug transporter genes, P-gp efflux activity, and rifampicin cellular uptake kinetics.
Main Results:
- ABCB1 (encoding P-gp) expression significantly increased during both M1 (166-fold) and M2 (55-fold) polarization.
- M2 macrophages exhibited 1.55-fold higher P-gp efflux activity compared to M1 macrophages.
- Rifampicin uptake was significantly lower in M2 macrophages than in M1 macrophages after 2h or 4h exposure.
Conclusions:
- M2 macrophages display a distinct drug transporter profile compared to M1 macrophages.
- Enhanced P-gp expression and activity in M2 macrophages contribute to reduced rifampicin uptake.
- These findings highlight potential mechanisms underlying the inferior efficacy of antibiotics in M2 macrophage-dominant TB granulomas.
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