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Updated: Jun 16, 2026
Differentiation of Functional Osteoclasts from Human Peripheral Blood CD14+ Monocytes
Published on: January 27, 2023
Effect of interleukin-32gamma on differentiation of osteoclasts from CD14+ monocytes
Yong-Gil Kim1, Chang-Keun Lee, Ji Seon Oh
1University of Ulsan, Asan Medical Center, Seoul, Korea.
Objective:
Interleukin-32 (IL-32) induces various inflammatory molecules in human monocytes and differentiation of monocytes into macrophage-like cells. This study was undertaken to evaluate the effects of IL-32gamma, the most biologically active isoform, on the differentiation and activation of osteoclasts.
Methods:
CD14+ monocytes were obtained from healthy volunteers, and samples of synovial tissue and synovial fluid were obtained from patients with rheumatoid arthritis (RA) and patients with osteoarthritis (OA). The concentration and expression levels of IL-32gamma in RA and OA samples were evaluated by enzyme-linked immunosorbent assay and immunoblotting, respectively. To examine the osteoclastogenic effects and functional activities, isolated monocytes were treated with either IL-32gamma or IL-17 in the presence or absence of soluble RANKL (sRANKL) on a culture system and on Osteologic disks. The expression of RANKL and osteoprotegerin (OPG) messenger RNA (mRNA) in RA fibroblast-like synoviocytes (FLS) was measured using reverse transcription-polymerase chain reaction (PCR) and real-time PCR.
Results:
The concentration and expression levels of IL-32gamma were higher in the RA samples than in the OA samples. Upon costimulation with sRANKL, the osteoclast count and resorbed area increased more significantly in the IL-32gamma-stimulated cultures than in those stimulated with IL-17. In the IL-32gamma-treated group without sRANKL stimulation, osteoclasts were differentiated, but the cells displayed low resorption activity. In RA FLS, RANKL mRNA expression increased in the presence of both IL-32gamma and IL-17. However, transcription of OPG decreased following IL-32gamma stimulation, resulting in a significant increase in the RANKL:OPG ratio.
Conclusion:
Our results suggest that IL-32gamma is a potent mediator of active osteoclast generation in the presence of sRANKL. Moreover, this novel cytokine creates more favorable conditions for osteoclastogenesis in the RA joint by increasing the RANKL:OPG ratio in FLS.
Insights
Interleukin-32gamma (IL-32gamma) promotes osteoclast generation and activity, particularly in rheumatoid arthritis (RA) joints. This cytokine increases the RANKL:OPG ratio, favoring bone erosion in inflammatory conditions.
Area of Science:
- Immunology
- Rheumatology
- Cell Biology
Background:
- Interleukin-32 (IL-32) is known to induce inflammatory molecules and monocyte differentiation.
- IL-32gamma is the most biologically active isoform of IL-32.
- Osteoclasts play a critical role in bone resorption and are implicated in inflammatory joint diseases.
Purpose of the Study:
- To investigate the effects of IL-32gamma on osteoclast differentiation and activation.
- To compare the effects of IL-32gamma with IL-17 on osteoclastogenesis.
- To assess the impact of IL-32gamma on the RANKL/OPG system in rheumatoid arthritis.
Main Methods:
- Collected CD14+ monocytes from healthy volunteers and synovial samples from RA and osteoarthritis (OA) patients.
- Quantified IL-32gamma concentration and expression using ELISA and immunoblotting.
- Stimulated monocytes with IL-32gamma or IL-17, with or without soluble RANKL (sRANKL), to assess osteoclastogenesis and resorption.
- Measured RANKL and osteoprotegerin (OPG) mRNA expression in RA fibroblast-like synoviocytes (FLS) via RT-PCR and real-time PCR.
Main Results:
- IL-32gamma levels were elevated in RA samples compared to OA samples.
- IL-32gamma significantly enhanced osteoclast differentiation and resorption in the presence of sRANKL, surpassing IL-17 effects.
- IL-32gamma alone induced osteoclast differentiation with limited resorption.
- In RA FLS, IL-32gamma increased RANKL mRNA and decreased OPG mRNA, elevating the RANKL:OPG ratio.
Conclusions:
- IL-32gamma is a potent mediator of osteoclast generation, especially when co-stimulated with sRANKL.
- IL-32gamma promotes osteoclastogenesis in the RA joint by modulating the RANKL:OPG ratio in FLS.
- This cytokine may contribute to bone erosion in rheumatoid arthritis.
