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NanoUPLC-MS(E) proteomic analysis of osteoclastogenesis downregulation by IL-4
Mirna S Freire1, Ana Paula C Cantuária2, Stella M F Lima3
1Centro de Análises Proteômicas e Bioquímicas, Pós-graduação em Ciências Genômicas e Biotecnologia, Universidade Católica de Brasília, SGAN 916N, Av. W5, Campus II, Modulo C, Brasília, DF, Brazil; Programa de Doutorado da Rede Centro-Oeste, Universidade de Brasília, Campus Universitário Darcy Ribeiro, Brasília, DF, Brazil.
Abstract:
Bone resorption is an important factor in bone homeostasis and imbalance can cause several diseases. In osteoimmunology, IL-4 has been described as an important factor in promoting M2 macrophage profile. In order to shed some light on the effect of IL-4 on osteoclast precursors in the presence of RANKL, cytokines and nitric oxide (NO) production and the proteomic profile were analyzed. The presence of IL-4 in in vitro osteoclastogenesis provides production of TNF-α, IL-1α, IL-1β, IL-10 and IL-12 at basal cell levels. Regarding NO production, IL-4 was sufficient to increase the basal NO levels. Proteomic analyses identified 877 global proteins. IL-4 in in vitro RANKL-mediated osteoclastogenesis leads to the expression of 118 proteins. The presence of rIL-4 in in vitro rRANKL-mediated-osteoclastogenesis downregulated this process. However, the proteomics findings in the osteoclastogenesis demonstrated a much greater effect on osteoclast precursor cells than on RANKL-differentiated osteoclasts. These results suggest that the main effect of IL-4 in pre-osteoclast cells leads to a M2 macrophage activation, and this probably contributed to a reduction in osteoclastogenesis when both stimuli were used. This study noticed that IL-4 plays an important regulatory role in bone homeostasis due to its suppressive potential of precursor osteoclast cells.
Insights
Interleukin-4 (IL-4) suppresses osteoclast precursor cells, impacting bone homeostasis. This study reveals IL-4’s regulatory role in bone by reducing osteoclast formation through M2 macrophage activation.
Area of Science:
- Osteoimmunology
- Bone Biology
- Cellular Proteomics
Background:
- Bone resorption is crucial for skeletal homeostasis; its dysregulation contributes to various diseases.
- Interleukin-4 (IL-4) is known to promote the M2 macrophage phenotype.
- Understanding IL-4's role in osteoclastogenesis is vital for bone health research.
Purpose of the Study:
- To investigate the effects of IL-4 on osteoclast precursors.
- To analyze cytokine and nitric oxide (NO) production under IL-4 influence.
- To determine the proteomic profile changes during IL-4-mediated osteoclastogenesis.
Main Methods:
- In vitro osteoclastogenesis model using osteoclast precursors.
- Stimulation with IL-4 and Receptor Activator of Nuclear Factor kappa-B Ligand (RANKL).
- Measurement of cytokine and NO levels; comprehensive proteomic analysis.
Main Results:
- IL-4 induced production of TNF-α, IL-1α, IL-1β, IL-10, and IL-12 in basal cells.
- IL-4 significantly increased basal nitric oxide (NO) levels.
- Proteomic analysis revealed 118 differentially expressed proteins in IL-4 treated cells, with a pronounced effect on precursor cells, downregulating RANKL-mediated osteoclastogenesis.
Conclusions:
- IL-4 primarily affects osteoclast precursor cells, promoting M2 macrophage activation.
- This M2 activation contributes to reduced osteoclastogenesis when IL-4 and RANKL are co-administered.
- IL-4 exhibits a significant regulatory role in bone homeostasis via suppression of precursor osteoclast cells.
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