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Updated: Aug 8, 2026
Isolation Protocol of Mouse Monocyte-derived Dendritic Cells and Their Subsequent In Vitro Activation with Tumor Immune Complexes
Published on: May 31, 2018
Interleukin-1 and tumour necrosis factor production by human monocytoid cells: study on a single cell level
A Simbirtsev1, E Prokopieva, E Ivanova
1Research Institute of Highly Pure Biochemicals, St. Petersburg, Russia.
Insights
Normal monocytes and U-937 cells produce Interleukin-1 beta (IL-1 beta) upon LPS stimulation, with monocytes responding faster. Tumor Necrosis Factor alpha (TNF alpha) production requires cell differentiation, indicating distinct regulatory pathways for these key inflammatory cytokines.
Area of Science:
- Immunology
- Cell Biology
- Cytokine Research
Background:
- Monocytoid cells, including monocytes and cell lines like U-937, THP-1, and HL-60, are crucial in immune responses.
- Interleukin-1 beta (IL-1 beta) and Tumor Necrosis Factor alpha (TNF alpha) are key pro-inflammatory cytokines.
- Understanding the regulation of IL-1 beta and TNF alpha production in different monocytoid cell types is vital for immunology research.
Purpose of the Study:
- To investigate the production of IL-1 beta and TNF alpha in normal human monocytes and transformed monocytoid cell lines (U-937, THP-1, HL-60).
- To compare the kinetics of cytokine synthesis and secretion between normal monocytes and cell lines following stimulation.
- To explore the conditions required for TNF alpha induction and its relationship with IL-1 beta.
Main Methods:
- Biological assays and cytokine-specific ELISA were employed to quantify IL-1 beta and TNF alpha.
- Immunocytochemical methods, including immunoperoxidase staining, were used for single-cell level analysis.
- Cell lines were stimulated with lipopolysaccharide (LPS) and phorbol 12-myristate 13-acetate (PMA) to induce cytokine production.
Main Results:
- Quiescent monocytes and cell lines lacked intracellular IL-1 beta and TNF alpha.
- LPS induced IL-1 beta synthesis in ~90% of monocytes, 15-20% of U-937, 3-5% of THP-1, and none of HL-60 cells.
- Monocytes exhibited faster IL-1 beta synthesis kinetics (1-2 hours) compared to cell lines (4-6 hours).
- Proliferating U-937 cells did not synthesize IL-1 beta; TNF alpha required PMA-induced differentiation.
- Recombinant IL-1 beta and TNF alpha showed limited induction of the other cytokine in differentiated U-937 cells.
Conclusions:
- Human monocytes and specific cell lines exhibit differential regulation of IL-1 beta and TNF alpha production.
- LPS is a potent inducer of IL-1 beta in monocytes and some cell lines, with distinct kinetic profiles.
- TNF alpha production in U-937 cells is dependent on differentiation, suggesting complex regulatory mechanisms involving cell state and external stimuli.
Abstract:
Human IL-1 beta and TNF alpha production by normal and transformed monocytoid cells was studied using biological assays, cytokine specific ELISA and by immunocytochemical methods on a single cell level. Quiescent human blood monocytes and cultured in vitro transformed human monocytoid cell lines U-937, THP-1 and HL-60 did not contain IL-1 beta and TNF alpha in their cytoplasm. IL-1 beta synthesis and secretion was induced by LPS stimulation in nearly 90% monocytes, 15-20% U-937, 3-5% THP-1 and in no HL-60 cells. Normal human blood monocytes had a more rapid kinetics of IL-1 beta synthesis. IL-1 beta positive cells stained with antibodies to human IL-1 beta appeared at 1-2 hours after LPS application, while in monocytic cell lines only after 4-6 hours. Using immunoperoxidase staining of U-937 cells pulse labelled with 3H-thymidine, it was shown that proliferating cells did not synthetize IL-1 beta. Instead of IL-1 beta, TNF alpha could be induced by LPS in U-937 cells only after preliminary differentiation with PMA. Recombinant IL-1 beta induced a very low level of TNF alpha production in PMA-treated cells. Similarly recombinant TNF alpha alone induced IL-1 beta synthesis only in a few U-937 cells.
