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IntroductionFerroptosis plays a crucial role in the progression of postmenopausal osteoporosis. Anemarrhena asphodeloides Bunge/Phellodendron chinense C.K. Schneid (AA/PC) is the core herb pair in traditional Chinese medicines formulae for postmenopausal osteoporosis treatment. However, the synergistic effects, and mechanisms, of AA/PC on alleviating ferroptosis and postmenopausal osteoporosis remain unclear.MethodsThe goal herein was to analyze the effective ingredients and molecular mechanisms of AA/PC in the treatment of osteoporosis through serum pharmacochemistry, network pharmacology, metabolomics analysis, and pharmacodynamics evaluation. A bilateral ovariectomized (OVX) mouse model was established.Results and DiscussionMicron-scale computed tomography analysis showed that AA/PC increased bone mineral density in OVX mice. The effects of AA/PC were better than AA or PC alone on inhibiting the bone resorption marker nuclear factor of activated T-cells 1. Furthermore, five absorbable compounds were detected in serum: mangiferin, magnoflorine, berberine, timosaponin BIII, and timosaponin AIII. Network pharmacology showed these compounds had close relationship with seven ferroptosis targets. Importantly, compared with AA or PC alone, the AA/PC herb pair exerted better effects on regulating crucial ferroptosis pathways, including the system xc-/glutathione/glutathione peroxidase 4, transferrin receptor/ferritin, and acyl-CoA synthetase long chain family member 4/polyunsaturated fatty acids signaling pathways. These results indicate that AA/PC exerts synergistic effects on regulating glutathione synthesis, iron homeostasis, and lipid metabolism in ferroptosis. This work lays the foundation for further development and use of AA/PC herb pair for preventing and treating postmenopausal osteoporosis.

Bones serve mechanical and defensive functions, as well as regulating the balance of calcium ions and housing bone marrow.. The qualities of bones do not remain constant. Instead, they fluctuate throughout life, with functions increasing in some situations while deteriorating in others. The synchronization of osteoblast-mediated bone formation and osteoclast-mediated bone resorption is critical for maintaining bone mass and microstructure integrity in a steady state. This equilibrium, however, can be disrupted by a variety of bone pathologies. Excessive osteoclast differentiation can result in osteoporosis, Paget’s disease, osteolytic bone metastases, and rheumatoid arthritis, all of which can adversely affect people’s health. Osteoclast differentiation is regulated by transcription factors NFATc1, MITF, C/EBPα, PU.1, NF-κB, and c-Fos. The transcriptional activity of osteoclasts is largely influenced by developmental and environmental signals with the involvement of co-factors, RNAs, epigenetics, systemic factors, and the microenvironment. In this paper, we review these themes in regard to transcriptional regulation in osteoclastogenesis.

Age-related hepatic lipid accumulation has become a major health problem in the elderly population. Specnuezhenide (SPN) is a major active iridoid glycoside from an edible herb Fructus Ligustri Lucidi, which is commonly used for preventing age-related diseases. However, the beneficial effects of SPN on age-related liver injury remain unknown. This study aimed to reveal the effect of SPN on age-related hepatic lipid accumulation and the underlying mechanism. D-galactose (D-gal)-induced aging mice were treated with vehicle or SPN for 12 weeks. Treatment of SPN decreased lipid accumulation and inflammation in the liver of D-gal–induced mice. Untargeted and targeted metabolomics showed that the SPN could regulate the bile acid (BA) synthesis pathway and restore the BA compositions in serum, livers, and feces of the D-gal–induced mice. Furthermore, SPN enhanced the protein and mRNA levels of hepatic BAs synthesis enzymes cytochrome P45027A1, cytochrome P4507A1, cytochrome P4507B1, and cytochrome P4508B1. Meanwhile, SPN alleviated D-gal-induced gut dysbiosis and reversed the proportions of microbes associated with bile salt hydrolase activity, including Lactobacillus, Ruminiclostridium, and Butyrivibrio. Our study revealed that SPN attenuated age-related hepatic lipid accumulation by improving BA profiles via modulating hepatic BA synthesis enzymes and gut microbiota.

Natural alkaloids are polycyclic, nitrogen-containing, and basic compounds obtained from plants. In this review, the advances in bioactive alkaloids with respect to their chemical structures, herbal sources, and effects for the prevention and treatment of osteoporosis are discussed. Anti-osteoporosis alkaloids are classified into six categories based on the chemical structure, namely, isoquinoline alkaloids, quinolizidine alkaloids, piperidine alkaloids, indole alkaloids, pyrrolizidine alkaloids and steroidal alkaloids. They promote mesenchymal stem cells differentiation, improve osteoblast proliferation, stimulate osteoblast autophagy and suppress osteoclast formation. These natural alkaloids can regulate multiple signaling pathways, including interrupting the tumor necrosis factor receptor associated factor 6- receptor activator of nuclear factor kappa B interaction, inhibiting the nuclear factor kappa B pathway in osteoclasts, activating the p38 mitogen-activated protein kinases pathway in osteoblasts, and triggering the wingless and int-1 pathway in mesenchymal stem cells. This review provides evidence and support for novel drug and clinical treatment of osteoporosis using natural alkaloids.

Natural products have proven their value as drugs that can be therapeutically beneficial in the treatment of various diseases. However, most natural products have low solubility and poor bioavailability, which pose significant challenges. To solve these issues, several drug nanocarriers have been developed. Among these methods, dendrimers have emerged as vectors for natural products due to their superior advantages, such as a controlled molecular structure, narrow polydispersity index, and the availability of multiple functional groups. This review summarizes current knowledge on the structures of dendrimer-based nanocarriers for natural compounds, with a particular focus on applications in alkaloids and polyphenols. Additionally, it highlights the challenges and perspectives for future development in clinical therapy.

Bitter acids (BA) from Humulus lupulus L. (hops) are known for beer brewing and have been proved to have activities on reliving oxidative stress and osteoporosis. This study firstly determined the effects of BA on senile osteoporosis (SOP) in D-galactose induced aging mice. BA significantly increased femoral bone mineral density, serum levels of alkaline phosphatase, osteocalcin as well as decreased serum tartrate-resistant acid phosphatase contents in D-galactose induced mice. BA also promoted proliferation, differentiation, mineralization and inhibited cell senescence, apoptosis in D-galactose treated osteoblasts. Furthermore, BA attenuated D-galactose induced oxidative stress through the activation of nuclear translocation of Nrf2 and the increase of intracellular HO-1, NQO1 expressions. In conclusion, BA might attenuate osteoporosis in aged mice through activating Nrf2/HO-1/NQO1 signaling pathway. As important components in beer, BA have potential in the development of drugs for SOP treatment.

Alzheimer’s disease (AD) and osteoporosis (OP) are progressive degenerative diseases caused by multiple factors, placing a huge burden on the world. Much evidence indicates that OP is a common complication in AD patients. In addition, there is also evidence to show that patients with OP have a higher risk of AD than those without OP. This suggests that the association between the two diseases may be due to a pathophysiological link rather than one disease causing the other. Several in vitro and in vivo studies have also proved their common pathogenesis. Based on the theory of traditional Chinese medicine, some classic and specific natural Chinese medicines are widely used to effectively treat AD and OP. Current evidence also shows that these treatments can ameliorate both brain damage and bone metabolism disorder and further alleviate AD complicated with OP. These valuable therapies might provide effective and safe alternatives to major pharmacological strategies.

As aging progresses, β-amyloid (Aβ) deposition and the resulting oxidative damage are key causes of aging diseases such as senior osteoporosis (SOP). Humulus lupulus L. (hops) is an important medicinal plant widely used in the food, beverage and pharmaceutical industries due to its strong antioxidant ability. In this study, APP/PS1 mutated transgenic mice and Aβ-injured osteoblasts were used to evaluate the protective effects of hops extracts (HLE) on SOP. Mice learning and memory levels were assessed by the Morris water maze. Mice femurs were prepared for bone micro-structures and immunohistochemistry experiments. The deposition of Aβ in the hippocampus, cortex and femurs were determined by Congo red staining. Moreover, protein expressions related to antioxidant pathways were evaluated by Western blotting. It was found that HLE markedly improved learning abilities and ameliorated memory impairment of APP/PS1 mice, as well as regulated antioxidant enzymes and bone metabolism proteins in mice serum. Micro-CT tests indicated that HLE enhanced BMD and improved micro-architectural parameters of mice femur. More importantly, it was discovered that HLE significantly reduced Aβ deposition both in the brain and femur. Further in vitro results showed HLE increased the bone mineralization nodule and reduced the ROS level of Aβ-injured osteoblasts. Additionally, HLE increased the expression of antioxidant related proteins Nrf2, HO-1, NQO1, FoxO1 and SOD-2. These results indicated that Humulus lupulus L. extract could protect against senior osteoporosis through inhibiting Aβ deposition and oxidative stress, which provides a reference for the clinical application of hops in the prevention and treatment of SOP.

Scopoletin, a typical example of a coumarin compound, exists in several Artemisia species and other plant genera. However, the systemic metabolic effects induced by scopoletin remain unclear. In the present study, we evaluated the metabolic profiles in scopoletin-exposed zebrafish embryos using UHPLC-Q-Obitrap-HRMS combined with multivariate analysis. Compared with the control group, 33 metabolites in scopoletin group were significantly upregulated, while 27 metabolites were significantly downregulated. Importantly, scopoletin exposure affected metabolites mainly involved in phosphonate and phosphinate metabolism, vitamin B6 metabolism, histidine metabolism, sphingolipid metabolism, and folate biosynthesis. These results suggested that scopoletin exposure to zebrafish embryos exhibited marked metabolic disturbance. This study provides a perspective of metabolic impacts and the underlying mechanism associated with scopoletin exposure.

Bone nonunion remains a challenge during the treatment of bone defect accompanied with infection. Epimedii herba is a widely used medicine in clinic to enhance bone healing. Epimedin C and icariin are the major flavonoid glycosides from Epimedii herba. Although the effects of these compounds have been reported, their different absorption and utilisation by osteoblasts remain unclear. In the present study, lipopolysaccharide (LPS)-induced osteoblast was adopted as the model cell to evaluate the effects of epimedin C and icariin. The intracellular and extracellular drug concentrations within 24 h were assayed by pipette tip solid-phase extraction and high-performance liquid chromatography, respectively. MTT, alkaline phosphatase (ALP) and calcified nodule staining were performed to identify and evaluate the effects of epimedin C and icariin on LPS-induced osteoblasts. The regulatory roles of epimedin C and icariin in the bone morphogenetic protein-2 (BMP-2)/Runt-related transcription factor 2 (Runx2) signalling pathway were investigated. The results revealed that epimedin C and icariin were not efficiently absorbed by LPS-induced osteoblasts. Nevertheless, they still had high utilisation efficiency after entering the cells. ALP activity, mineralisation and osteoblasts proliferation were enhanced by a high concentration of epimedin C and icariin. The suppressed expression of BMP-2 and Runx2 in LPS-induced osteoblasts was up-regulated significantly after treatment with epimedin C and icariin. These findings firstly illustrated the behavior of epimedin C and icariin from Epimedii herba on LPS-induced osteoblasts and the regulatory property on the expression of key genes and proteins of the BMP-2/Runx2 signalling pathway, which might be helpful for better understanding flavonoids’ mechanism to enhance bone repair and improving their future application.

Erxian decoction (EXD), a traditional Chinese medicine formula, has been used for treatment of osteoporosis for many years. The purpose of this study was to investigate the pharmacological effect of EXD in preventing osteoblast apoptosis and the underlying mechanism of prevention. Putative targets of EXD were predicted by network pharmacology, and functional and pathway enrichment analyses were also performed. Evaluations of bone mineral density, serum estradiol level, trabecular area fraction, serum calcium levels, and tumor necrosis factor (TNF)-α levels in ovariectomized rats, as well as cell proliferation assays, apoptosis assays, and western blotting in MC3T3-E1 osteoblasts were performed for further experimental validation. Ninety-three active ingredients in the EXD formula and 259 potential targets were identified. Functional and pathway enrichment analyses indicated that EXD significantly influenced the PI3K-Akt signaling pathway. In vivo experiments indicated that EXD treatment attenuated bone loss and decreased TNF-α levels in rats with osteoporosis. In vitro experiments showed that EXD treatment increased cell viability markedly and decreased levels of caspase-3 and the rate of apoptosis. It also promoted phosphorylation of Akt, nuclear translocation of transcription factor NF-erythroid 2-related factor (Nrf2), and hemeoxygenase-1 (HO-1) expression in TNF-α-induced MC3T3-E1 cells. Our results suggest that EXD exerted profound anti-osteoporosis effects, at least partially by reducing production of TNF-α and attenuating osteoblast apoptosis via Akt/Nrf2/HO-1 signaling pathway.

Vancomycin is an effective antibiotic for treatment of bone infection caused by Staphylococcus aureus, however, a high local concentration of vancomycin might induce a delay in bone union. Icariin has been reported to suppress osteoclastogenes and promote osteogenesis. Our study aimed to investigate the effect of icariin on bone repair after anti-infection treatment in vivo and to explore the resisting effect of icariin on rat calvarial osteoblasts (ROBs) inhibited with high doses of vancomycin. Rabbits with bone infection of S. aureus were treated with implanted vancomycin-calcium sulfate (VCS) and icariin at 10.86 mg/kg/day for consecutive 8 weeks. Micro-CT, morphology, blood biochemistry were evaluated. In addition, ROBs were treated with vancomycin and icariin at different doses. Cell proliferation and differentiation capabilities, BMP2, Runx2, OPG, RANKL mRNA levels and protein expression were assessed. The results indicated that high dose of vancomycin significantly decreased bone mass and inhibited osteocalcin secretion; icariin increased these indicators compared with the single vancomycin treatment. Over 0.1 mg/mL of vancomycin inhibited the proliferation and differentiation of ROBs, while icariin resisted the inhibition of vancomycin by regulating cell cycle and promoting the Alkaline phosphatase (ALP) activity. Moreover, icariin promote bone formation by up-regulating BMP2/Runx2 and OPG/RANKL pathways. Icariin exhibited osteoplastic properties on osteoblasts that had been inhibited with high doses of vancomycin. Therefore, icariin is helpful for post-infection treatment of bone infection.

Natural antioxidants are widely used in the life sciences. Phellinus igniarius is a historically used natural antioxidant containing a variety of active compounds. Phenols, particularly Inoscavin A and Hypholomine B, are found in the high concentrations. Better quantitative methods are needed to perform quality control in order to support further research of this mushroom. An ultra-performance liquid chromatography method coupled to photodiode-array detection and an electrospray ionization tandem mass spectrometry method (UPLC-PAD-MS) was developed to simultaneously quantify Inoscavin A and Hypholomine B levels in the medicinal fungus Phellinus igniarius. The two compounds were quantified using UPLC-PAD and UPLC-MS. The methods were accurate (mean accuracy for spiked matrix ranged from 101.5% to 105.8%), sensitive (limit of detection ranged from 0.28 to 1.14 mg L-1) and precise (the relative standard deviations ranged from 0.13 to 2.8%). Inoscavin A and Hypholomine B were purified using high-speed counter-current chromatography (HSCCC), structural evaluated to meet the request of standard substances. UPLC separation was performed on a reversed-phase C18 column using gradient elution with acetonitrile and 0.1% formic acid over 10 min. The developed method was successfully applied to determine Inoscavin A and Hypholomine B in twelve Phellinus igniarius samples of different origins and the results showed that it was suitable for the analysis of these active components in Phellinus igniarius samples.