Icariin promotes osteogenic differentiation by upregulating alpha-enolase expression.

Osteogenic differentiation is a crucial biological process for maintaining bone remodelling. Aerobic glycolysis is the main source of energy for osteogenic differentiation. Alpha-enolase (Eno1), a glycolytic enzyme, is a therapeutic target for numerous diseases. Icariin, a principal active component of the traditional Chinese medicine Epimedium grandiflorum, can stimulate osteogenic differentiation. Here, we aimed to determine if icariin promotes osteogenic differentiation via Eno1. Icariin (1 µM) significantly promoted osteogenic differentiation of MC3T3-E1 cells. Icariin upregulated Eno1 protein and gene expressions during osteogenic differentiation. Moreover, ENOblock, a specific inhibitor of Eno1, markedly inhibited icariin-induced osteogenic differentiation. Futhermore, western blot assay showed that Eno1 might mediate osteogenic differentiation through the BMP/Smad4 signalling pathway. Collectively, Eno1 could be a promising drug target for icariin to regulate osteogenic differentiation.

Role of the major histocompatibility complex class II protein presentation pathway in bone immunity imbalance in postmenopausal osteoporosis.

Bone immunity regulates osteoclast differentiation and bone resorption and is a potential target for the treatment of postmenopausal osteoporosis (PMOP). The molecular network between bone metabolism and the immune system is complex. However, the molecular mechanism underlying the involvement of the major histocompatibility complex class II (MHC-II) molecule protein presentation pathway in PMOP remains to be elucidated. The MHC-II molecule is a core molecule of the protein presentation pathway. It is combined with the processed short peptide and presented to T lymphocytes, thereby activating them to become effector T cells. T-cell-derived inflammatory factors promote bone remodeling in PMOP. Moreover, the MHC-II molecule is highly expressed in osteoclast precursors. MHC-II transactivator (CIITA) is the main regulator of MHC-II gene expression and the switch for protein presentation. CIITA is also a major regulator of osteoclast differentiation and bone homeostasis. Therefore, we hypothesized that the MHC-II promotes osteoclast differentiation, providing a novel pathogenic mechanism and a potential target for the treatment of PMOP.

Roles of Altered Macrophages and Cytokines: Implications for Pathological Mechanisms of Postmenopausal Osteoporosis, Rheumatoid Arthritis, and Alzheimer's Disease.

Postmenopausal osteoporosis (PMOP) is characterized by the uncoupling of bone resorption and bone formation induced by estrogen deficiency, which is a complex outcome related to estrogen and the immune system. The interaction between bone and immune cells is regarded as the context of PMOP. Macrophages act differently on bone cells, depending on their polarization profile and secreted paracrine factors, which may have implications for the development of PMOP. PMOP, rheumatoid arthritis (RA), and Alzheimer's disease (AD) might have pathophysiological links, and the similarity of their pathological mechanisms is partially visible in altered macrophages and cytokines in the immune system. This review focuses on exploring the pathological mechanisms of PMOP, RA, and AD through the roles of altered macrophages and cytokines secretion. First, the multiple effects on cytokines secretion by bone-bone marrow (BM) macrophages in the pathological mechanism of PMOP are reviewed. Then, based on the thought of "different tissue-same cell type-common pathological molecules-disease pathological links-drug targets" and the methodologies of "molecular network" in bioinformatics, highlight that multiple cytokines overlap in the pathological molecules associated with PMOP vs. RA and PMOP vs. AD, and propose that these overlaps may lead to a pathological synergy in PMOP, RA, and AD. It provides a novel strategy for understanding the pathogenesis of PMOP and potential drug targets for the treatment of PMOP.

Effects of Icariin on Modulating Gut Microbiota and Regulating Metabolite Alterations to Prevent Bone Loss in Ovariectomized Rat Model.

Postmenopausal osteoporosis (PMOP) is an estrogen deficiency-induced bone loss, which has been shown an association with an altered gut microbiota (GM). Gut microbiota-bone axis has been recognized as a crucial mediator for bone homeostasis. Icariin (ICA) is an effective agent to delay bone loss by regulating the bone homeostasis. Thus, we hypothesize that ICA can prevent bone loss by modulating GM and regulating metabolite alterations. The effects of ICA on bone metabolism improvement in ovariectomized (OVX) rats and their relationships with the GM and fecal metabolites were investigated. Micro-computed tomography (micro-CT) and hematoxylin-eosin (HE) staining showed a typical bone boss in OVX group, while ICA or estradiol (E2) administration exhibited positive effects on bone micro-architecture improvement. The GM such as Actinobacteria, Gammaproteobacteria, Erysipelotrichi, Erysipelotrichales, Enterobacteriales, Actinomycetales, Ruminococcus and Oscillospira significantly correlated to serum bone Gla-protein (BGP), receptor activator of nuclear factor-κB (RANK), receptor activator of nuclear factor-κB ligand (RANKL), osteoprotegerin (OPG) and tartrate resistant acid phosphatase (TRACP). Further t-test revealed a substantial variation of the GM and fecal metabolites in different treatments. Among them, Lachnoclostridium, Butyricimonas, Rikenella, Paraprevolla, Adlercreutzia, Enterorhabdus, Anaerovorax, Allobaculum, Elusimicrobium, Lactococcus, Globicatella and Lactobacillus were probably the key microbial communities driving the change of bile acid, amino acid and fatty acid, thereby leading to an improvement of PMOP. The significant up-regulation of L-Saccharopine, 1-Aminocyclohexadieneacid and linoleic acid after ICA administration suggested important contributions of amino acid and fatty acid metabolisms in the prevention and treatment of PMOP. Taken together, our study has provided new perspectives to better understand the effects of ICA on PMOP improvement by regulating GM and the associated fecal metabolites. Our findings contribute to develop ICA as a potential therapy for PMOP.

Verification of pain-related neuromodulation mechanisms of icariin in knee osteoarthritis.

Knee osteoarthritis (KOA) is a common disease with no specific treatment. Icariin (ICA) is considered an agent for KOA. This study aimed to confirm the pain-related neuromodulation mechanisms of ICA on KOA. Three experiments were designed: (1) verifying the therapeutic effects of ICA in vivo and in vitro, (2) exploring the potential pain-related neuromodulation pathways involved in ICA treatment by functional magnetic resonance imaging (fMRI) and virus retrograde tracing (VRT) and (3) confirming the pain-related targets by tandem mass tag (TMT)-based quantitative proteomics and bioinformatic analyses. Experiment 1 verified the efficacy of ICA in OA animal and cell models. Experiment 2 found a series of brain regions associated with KOA reversed by ICA treatment, indicating that a pain-related hypothalamic-mediated neuromodulation pathway and an endocannabinoid (EC)-related pathway contribute to ICA mechanisms. Experiment 3 explored and confirmed four pain-related genes involved in KOA and ICA treatment. We confirmed the key role of pain-related neuromodulation mechanisms in ICA treatment associated with its analgesic effect. Our findings contribute to considering ICA as a novel therapy for KOA.

Identification of the Key Role of NF-κB Signaling Pathway in the Treatment of Osteoarthritis With Bushen Zhuangjin Decoction, a Verification Based on Network Pharmacology Approach.

This study aimed to identify whether the NF-κB signaling pathway plays a key role in the treatment of osteoarthritis (OA) with Bushen Zhuangjin Decoction (BZD) based on a typical network pharmacology approach (NPA). Four sequential experiments were performed: 1) conventional high-performance liquid chromatography (HPLC), 2) preliminary observation of the therapeutic effects of BZD, 3) NPA using three OA-related gene expression profiles, and 4) verification of the key pathway identified by NPA. Only one HPLC-verified compound (paeoniflorin) was identified from the candidate compounds discovered by NPA. The genes verified in the preliminary observation were also identified by NPA. NPA identified a key role for the NF-κB signaling pathway in the treatment of OA with BZD, which was confirmed by conventional western blot analysis. This study identified and verified NF-κB signaling pathway as the most important inflammatory signaling pathway involved in the mechanisms of BZD for treating OA by comparing the NPA results with conventional methods. Our findings also indicate that NPA is a powerful tool for exploring the molecular targets of complex herbal formulations, such as BZD.