Department of Spine Surgery, Qilu Hospital of Shandong University, Jinan, Shandong, China.
Department of Microorthopaedics, Affiliated Hospital of Shandong University of Traditional Chinese Medicine, Jinan, Shandong, China.
Clin Transl Med. 2023 Sep;13(9):e1369. doi: 10.1002/ctm2.1369.
The imbalance between osteoblasts and osteoclasts may lead to osteoporosis. Osteoblasts and osteoclasts have different energy requirements, with aerobic glycolysis being the prominent metabolic feature of osteoblasts, while osteoclast differentiation and fusion are driven by oxidative phosphorylation.
By polymerase chain reaction as well as Western blotting, we assayed coactivator-associated arginine methyltransferase 1 (CARM1) expression in bone tissue, the mouse precranial osteoblast cell line MC3T3-E1 and the mouse monocyte macrophage leukaemia cell line RAW264.7, and expression of related genes during osteogenic differentiation and osteoclast differentiation. Using gene overexpression (lentivirus) and loss-of-function approach (CRISPR/Cas9-mediated knockout) in vitro, we examined whether CARM1 regulates osteogenic differentiation and osteoblast differentiation by metabolic regulation. Transcriptomic assays and metabolomic assays were used to find the mechanism of action of CARM1. Furthermore, in vitro methylation assays were applied to clarify the arginine methylation site of PPP1CA by CARM1.
We discovered that CARM1 reprogrammed glucose metabolism in osteoblasts and osteoclasts from oxidative phosphorylation to aerobic glycolysis, thereby promoting osteogenic differentiation and inhibiting osteoclastic differentiation. In vivo experiments revealed that CARM1 significantly decreased bone loss in osteoporosis model mice. Mechanistically, CARM1 methylated R23 of PPP1CA, affected the dephosphorylation of AKT-T450 and AMPK-T172, and increased the activities of phosphofructokinase-1 and pructose-2,6-biphosphatase3, causing an up-regulation of glycolytic flux. At the same time, as a transcriptional coactivator, CARM1 regulated the expression of pyruvate dehydrogenase kinase 3, which resulted in the inhibition of pyruvate dehydrogenase activity and inhibition of the tricarboxylic acid cycle, leading to a subsequent decrease in the flux of oxidative phosphorylation.
These findings reveal for the first time the mechanism by which CARM1 affects both osteogenesis and osteoclast differentiation through metabolic regulation, which may represent a new feasible treatment strategy for osteoporosis.
成骨细胞与破骨细胞之间的失衡可能导致骨质疏松症。成骨细胞和破骨细胞有不同的能量需求，成骨细胞的有氧糖酵解是其显著的代谢特征，而破骨细胞的分化和融合则由氧化磷酸化驱动。
通过聚合酶链反应和 Western blot 法，我们检测了骨组织、小鼠颅前成骨细胞系 MC3T3-E1 和小鼠单核巨噬细胞白血病细胞系 RAW264.7 中的共激活因子相关精氨酸甲基转移酶 1（CARM1）表达，以及成骨分化和破骨分化过程中的相关基因表达。通过体外基因过表达（慢病毒）和功能丧失（CRISPR/Cas9 介导的敲除）方法，我们研究了 CARM1 是否通过代谢调节来调控成骨分化和破骨分化。转录组学分析和代谢组学分析用于寻找 CARM1 的作用机制。此外，体外甲基化实验用于阐明 CARM1 对 PPP1CA 的精氨酸甲基化位点。
我们发现 CARM1 将成骨细胞和破骨细胞的葡萄糖代谢从氧化磷酸化重编程为有氧糖酵解，从而促进成骨分化并抑制破骨分化。体内实验表明，CARM1 显著减少了骨质疏松症模型小鼠的骨丢失。在机制上，CARM1 甲基化 PPP1CA 的 R23，影响 AKT-T450 和 AMPK-T172 的去磷酸化，并增加磷酸果糖激酶-1 和果糖-2,6-二磷酸酶 3 的活性，导致糖酵解通量上调。同时，作为转录共激活因子，CARM1 调节丙酮酸脱氢酶激酶 3 的表达，导致丙酮酸脱氢酶活性抑制和三羧酸循环抑制，随后氧化磷酸化通量减少。
这些发现首次揭示了 CARM1 通过代谢调节影响成骨和破骨分化的机制，这可能代表了骨质疏松症的一种新的可行治疗策略。
