Hippuric acid and 3-(3-hydroxyphenyl) propionic acid inhibit murine osteoclastogenesis through RANKL-RANK independent pathway

Authors: ZHAO, HAIJUN - Arkansas Children'S Nutrition Research Center (ACNC); LAZARENKO, OXANA - Arkansas Children'S Nutrition Research Center (ACNC); CHEN, JIN-RAN - Arkansas Children'S Nutrition Research Center (ACNC)

Submitted to: Journal of Cellular Physiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/4/2019
Publication Date: 7/4/2019
Citation: Zhao, H., Lazarenko, O.P., Chen, J. 2019. Hippuric acid and 3-(3-hydroxyphenyl) propionic acid inhibit murine osteoclastogenesis through RANKL-RANK independent pathway. Journal of Cellular Physiology. https://doi.org/10.1002/jcp.28998.
DOI: https://doi.org/10.1002/jcp.28998

Interpretive Summary: Previously published results indicated that diets supplemented with 10% whole blueberry (BB) powder had robust effects on promoting bone formation. We hypothesized that the positive effect of BB diet on bone resulted from BB-derived phytonutrients (plant-derived compounds from diet), e.g., phenolic acids (such as hippuric acid (HA) and 3-(3-hydroxyphenyl) propionic acid (3-3-PPA)). Appropriate bone development is dependent upon a balance between bone formation (due to osteoblasts) and bone resorption (due to osteoclasts). We investigated if HA and 3-3-PPA inhibit osteoclast bone resorption in multiple bone cell culture systems. It was found that HA and 3-3-PPA dose-dependently inhibited osteoclast differentiation. The effects of HA and 3-3-PPA on osteoclast formation did not utilize classic, known osteoclast formation pathways. These results indicate that dietary BB may exert positive effects on bone health through the actions of the phytonutrients HA and 3-3-PPA, which serve to dampen the activities of the cells that are involved in bone breakdown. This information highlights the potential importance of certain polyphenolic-rich fruits to bone health, and points to some of the mechanisms by which these foods impact bone.

Technical Abstract: Nutritional factors influence bone development. Previous studies demonstrated that bone mass significantly increased with suppressed bone resorption in early life of rats fed with AIN-93G semi-purified diets supplemented with 10% whole blueberry (BB) powder for 2 weeks. However, the effects of increased phenolic acids in animal serum due to this diet on bone and bone resorption were unclear. This in vitro and in ex vivo study examined the effects of phenolic hippuric acid (HA) and 3-(3-hydroxyphenyl) propionic acid (3-3-PPA) on osteoclastic cell differentiation and bone resorption. We cultured murine osteoclast (macrophage) cell line, RAW 264.7 cells, and hemato-poietic osteoclast progenitor cells (isolated from 4-week-old C57BL6/J mice) with 50 ng/ml of receptor activator of nuclear factor '-' ligand (RANKL). Morphologic studies showed decreased osteoclast number with treatment of 2.5% mouse serum from BB diet–fed animals compared with those treated with serum from standard casein diet–fed mice in both RAW 264.7 cell and primary cell cultures. HA and 3-3-PPA, but not 3–4-PPA, had dose-dependent suppressive effects on osteoclastogenesis and osteoclast resorptive activity in Corning osteo-assay plates. Signaling pathway analysis showed that after pretreatment with HA or 3-3-PPA, RANKL-stimulated increase of osteoclastogenic markers, such as nuclear factor of activated T-cells, cytoplasmic 1 and matrix metallopeptidase 9 gene/protein expression were blunted. Inhibitory effects of HA and 3-3-PPA on osteoclastogenesis utilized RANKL/RANK independent mediators. The study revealed that HA and 3-3-PPA significantly inhibited osteoclastogenesis and bone osteoclastic resorptive activity.