Isosakuranetin inhibits subchondral osteoclastogenesis for attenuating osteoarthritis via suppressing NF-κB/CXCL2 axis

Abstract

• CMC/TOFMS method, a biological affinity chromatographic system, was utilized to screen potential constituents from a natural plant. • We for the first time identified Isosakuranetin (ISN) from Rhizoma anemarrhenae (Zhimu) via the CMC/TOFMS method. • ISN inhibits subchondral osteoclastogenesis in osteoarthritis (OA) mice model, thereby ameliorating articular cartilage deterioration. • ISN ameliorated abnormal angiogenesis in the subchondral bone marrow and alleviated osteoclast-associated pain in OA mice model. • NF-κB/CXCL2 axis was required for ISN targeting of osteoclastogenesis. As the most predominant form of arthritis, osteoarthritis (OA) is featured with irreversible progress and involvement of the whole joint. Since OA onset, abnormal mechanical load initiates excessive osteoclastogenesis, evolving a rapid turnover of subchondral bone, cyst creation, synovitis, cartilage degradation, and ultimately resulting in joint failure. Additionally, aberrant vascularization and nociceptive pain are invoked by osteoclast-induced angiogenesis and sensory innervation in the subchondral bone. Rhizoma anemarrhenae (Zhimu) has been extensively demonstrated to show multiple pharmacological effects including anti-inflammation, anti-aging, and immunomodulation. Herein, Broussonin a (BRA), Markogein (MAN), and Isosakuranetin (ISN) derived from Rhizoma anemarrhenae, were initially discovered for their affinity with Bone marrow mononuclear cell (BMMC) membranes using the Cell membrane chromatography/Time of flight mass spectrometry (CMC/TOFMS) method, while only ISN exerted a significant inhibitory effect on RANKL-induced osteoclastogenesis in BMMC in vitro . Intriguingly, we disclosed that ISN blunted the overactivation of Tartrate-resistant acid phosphatase positive (TRAP+) osteoclasts in subchondral bone in OA mice, as indicated by enhanced bone volume/total volume (BV/TV), trabecular number (Tb.N), and trabeculae thickness ( Tb.Th ), as well as diminished trabecular pattern factor ( Tb.pf ). Treatment with ISN also impaired aberrant angiogenesis and nociceptive reaction in the subchondral bone marrow. Moreover, ISN hindered the loss of articular cartilage proteoglycan and lowered the Osteoarthritis Research Society International (OARSI) grade, boosting the expression amount of Aggrecan (ACAN) and Collagen II (COL II) positive cells while reducing Matrix metalloproteinase 13 (MMP-13) positive cells. For mechanisms, We verified that ISN hampered subchondral osteoclastogenesis by blocking nuclear factor kappa light chain enhancer of activated B cells (NF-κB) signaling and C-X-C Motif Chemokine Ligand 2 (CXCL2) stimulation. Taken together, we reveal that ISN impedes the progression of OA by preventing hyperactivated subchondral osteoclastogenesis via suppressing the NF-κB/CXCL2 axis.

Topics & Concepts