miR-324 mediates bone homeostasis through the regulation of osteoblast and osteoclast differentiation and activity

Hayman, D. J.; Lin, H.; Prior, A.; Charlesworth, G.; Johnson de Sousa Brito, F. M.; Hao, Y.; Patel, K.; Soul, J.; Clark, I. M.; Pirog, K. A.; Barter, M. J.; van 't Hof, R. J.; Young, D. A.

doi:10.1101/2023.07.10.548366

bioRxiv · 10.1101/2023.07.10.548366

miR-324 mediates bone homeostasis through the regulation of osteoblast and osteoclast differentiation and activity

Hayman, D. J.·Lin, H.·Prior, A.·Charlesworth, G.·Johnson de Sousa Brito, F. M.·Hao, Y.·Patel, K.·Soul, J.·Clark, I. M.·Pirog, K. A.·Barter, M. J.·van 't Hof, R. J.·Young, D. A.

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Abstract

microRNAs (miRNAs) are non-coding RNAs which modulate the expression of other RNA molecules. One miRNA can target many transcripts, allowing each miRNA to play key roles in many biological pathways. miR-324 is a miRNA previously implicated in bone and cartilage maintenance, defects of which result in common age-related diseases, such as osteoporosis or osteoarthritis (OA). In global miR-324-null mice cartilage damage was increased in both surgically and ageing-induced OA, despite minimal changes to the cartilage transcriptome, with few predicted miR-324 targets dysregulated. However, micro-computed tomography and histology demonstrated that global miR- 324-null the mice had an increase in bone mineral density, trabecular thickness and cortical thickness, with many parameters increasing with age. The bone marrow of miR-324-null mice also had reduced lipid content while and in vivo TRAP staining revealed a decrease in osteoclasts, with histomorphometry demonstrating an increased rate of bone formation in miR-324-null mice. Ex vivo assays revealed that the high bone mass phenotype of the miR-324-null mice resulted from increased osteoblast activity and decreased osteoclastogenesis. RNA-seq and qRT-PCR followed by miR-324 target prediction and validation in osteoblasts, osteoclasts and bone marrow macrophages identified the osteoclast fusion regulator Pin1 as a miR-324 target in the osteoclast lineage and the master osteogenic regulator Runx2 as a target of miR-324-5p in osteoblasts, the in vitro overexpression of which recapitulated the increased osteogenesis and decreased adipogenesis phenotype observed in vivo. These data point to important roles of miR-324 in skeletal biology with altered bone homeostasis in miR-324-null mice potentially causal for the increased cartilage damage observed during OA and ageing. Elucidation of pathways regulated by miR-324 offer promise for the treatment of bone diseases such as osteoporosis.

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Hayman, D. J., Lin, H., Prior, A., Charlesworth, G., Johnson de Sousa Brito, F. M., Hao, Y., Patel, K., Soul, J., Clark, I. M., Pirog, K. A., Barter, M. J., van 't Hof, R. J., Young, D. A.. 2023-07-10. miR-324 mediates bone homeostasis through the regulation of osteoblast and osteoclast differentiation and activity. https://doi.org/10.1101/2023.07.10.548366

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