Structural elucidation and anti-diabetic osteoporotic activity of an arabinogalactan from Phellodendron chinense Schneid.

Phellodendron chinense Schneid. was widely used as a medicinal herb for the treatment of diabetic osteoporosis in China. In this study, an arabinogalactan, named as PPCP-1, was isolated from the bark of Phellodendron chinense Schneid., and purified by DEAE-cellulose DE52 and Sephacryl S-200 HR column chromatography. The structure of PPCP-1 was characterized as a repeating unit consisting of →3)-ß-d-Galp-(1→, →3,6)-ß-d-Galp-(1→, →5)-α-l-Araf-(1→, →4)-α-d-Glcp-(1→, →3)-α-d-Glcp-(1→, →4)-α-d-Manp-(1→ with branches of →5)-α-l-Araf-(1→, →3,5)-α-l-Araf-(1→ and terminal α-l-Araf. Pharmacologically, the oral administration of PPCP-1 preserved osteoporosis associated with hyperglycemia by inhibiting α-glucosidase activity, improving glucose tolerance, decreasing the accumulation of advanced glycation end products (AGEs), as well as down-regulating the expression of receptor for AGEs in tibias of streptozotocin-induced diabetic rats. Collectively, the present study suggested that the arabinogalactan PPCP-1 from Phellodendron chinense Schneid. might potentially be used as functional foods for bone health and/or developed for drug discovery for alleviating diabetic osteoporosis.

Timosaponin AIII attenuates inflammatory injury in AGEs-induced osteoblast and alloxan-induced diabetic osteoporosis zebrafish by modulating the RAGE/MAPK signaling pathways.

BACKGROUND: Advanced glycation end products (AGEs) deposition causes inflammatory injury in osteoblasts and contributes to diabetic osteoporosis. The receptor for advanced glycation end product/mitogen-activated protein kinase pathway (RAGE/MAPK) signaling pathway is closely linked to the pathogenesis of diabetic osteoporosis. Timosaponin AIII, a steroidal saponin isolated from Anemarrhena asphodeloides Bunge (Asparagaceae), shows anti-inflammatory and anti-osteoporosis effects. PURPOSE: The present study was aimed to investigate the therapeutic effects of timosaponin AIII on diabetic osteoporosis and whether its effect is dependent on protecting osteoblasts against AGEs-induced injury via RAGE/MAPK signaling suppression. METHODS: An alloxan-induced diabetic osteoporosis zebrafish model was applied to investigate the effects of timosaponin AIII in vivo, and alendronate was used as a positive control. Moreover, related mechanisms were explored in primary rat osteoblasts. Molecular docking was applied to investigate the interactions between timosaponin AIII and RAGE. RESULTS: Timosaponin AIII treatment reversed alloxan-induced reduction in the mineralized area of the larvae head skeleton, accompanied by a decreased level of triglyceride and total cholesterol in the zebrafish. Additionally, AGEs significantly influenced RAGE expression, alkaline phosphatase activity, interleukin 1ß expression, interleukin 6 expression, and tumor necrosis factor-α expression, and increased cell apoptosis. Timosaponin AIII significantly downregulated AGEs-induced interleukin 1ß, interleukin 6, and tumor necrosis factor-α levels, and upregulated alkaline phosphatase and osteocalcin levels. Timosaponin AIII also significantly reduced the expression of RAGE and had additive effects on downstream P38, extracellular signal-regulated kinase and c-Jun N-terminal kinase in AGEs-induced osteoblast. Molecular docking predicted that hydrogen and hydrophobic interactions occurred between timosaponin AIII and RAGE. CONCLUSION: These data clarified that timosaponin AIII attenuates diabetic osteoporosis via a novel mechanism involved suppressing the RAGE/MAPK signaling pathway. Our finding highlights the potential value of timosaponin AIII as an anti-diabetic osteoporosis agent.

Integrated pathological cell fishing and network pharmacology approach to investigate main active components of Er-Xian decotion for treating osteoporosis.

ETHNOPHARMACOLOGICAL RELEVANCE: Oxidative damage to osteoblasts was a key factor in the development of osteoporosis. Er-Xian Decotion (EXD) is widely used in China for the treatment of osteoporosis, which has a variety of antioxidant active ingredients. EXD may be an important source of protection against oxidative damage in osteoblasts, but the anti-osteoporotic active components of EXD is currently unclear. AIM OF THE STUDY: This work established an effective and reliable drug screening method to find main active ingredients in EXD (M-EXD) that can protect osteoblasts against oxidative stress and achieve anti-osteoporosis effects. MATERIALS AND METHODS: H2O2-induced osteoblast cell fishing with UHPLC-QTOF/MS was firstly used to discover the potential active components from EXD. Afterword, the EXD compound-osteoporosis target network was constructed using network pharmacology, thus potentially anti-osteoporosis ingredients were founded, and their combination were defined as the M-EXD. Finally, pharmacology effects of M-EXD was evaluated by ovariectomized rats, prednisolone induced-zebrafish and H2O2-induced osteoblasts. RESULTS: 40 candidate active ingredients in EXD were initially screened out via pathological cell fishing. According to network pharmacology result, M-EXD consisted of 13 ingredients since they had a close relationship with 65 osteoporosis-related targets. Pharmacological evaluation showed that M-EXD significantly ameliorated oxidative stress in H2O2-induced osteoblast model, evidently reversed the activity of ALP, ROS, GSH-px, NO and MDA compared with the model group. M-EXD showed better anti-oxidative activities than individual ingredients, presenting obvious synergetic effects. In osteoporosis rat and zebrafish models, M-EXD also demonstrated good anti-osteoporotic properties by mitigating the osteoporosis bone loss and increasing serum bone morphogenetic protein 2, and reversing osteocalcin expression in bone tissue. It significantly ameliorated oxidative stress in the in-vivo models. Moreover, M-EXD and EXD showed similar anti-osteoporotic and anti-oxidative properties, while the rest components of EXD had no satisfactory anti-osteoporotic efficacy. CONCLUSIONS: Our work successfully identified the main active components in EXD, which could represent the efficacy of EXD on treating osteoporosis, and meanwhile, it also provided an effective strategy to investigate active ingredients from natural medicines, which might be helpful for drug development and application.

[Rapid screening of anti-osteoporosis active ingredients from Liuwei Dihuang Decoction by osteoblast membrane chromatography/ultra-high performance liquid chromatography-time of flight mass spectrometry].

A method was established for determination of potential anti-osteoporosis ingredients from Liuwei Dihuang Decoction by osteoblast cell membrane chromatography/ultra-performance liquid chromatography/time-of-flight mass spectrometry (CMC/UPLC-TOF/MS). The osteoblasts were used as the source of cell membrane for the preparation of CMC stationary phase. An osteoblast CMC column (10 mm×2 mm) was prepared by coating silica gel (0.05 g) with cell membrane. The active ingredients in the aqueous extract of Liuwei Dihuang Decoction (90 g/L) were first screened by CMC. Water was used as the mobile phase, and the flow rate was 0.20 mL/min. Then, the eluates of the CMC were qualitatively analyzed by UPLC-TOF/MS using a WATERS ACQUITY UPLC BEH C18 column (10 mm×2 mm). Acetonitrile-water was used as the mobile phase at a flow rate of 0.40 mL/min. This method could quickly and selectively identify 16 potential anti-osteoporotic active ingredients from Liuwei Dihuang Decoction. Due to high catalpol content in Liuwei Dihuang Decoction and its good affinity with CMC column, catapol was selected for in vivo and in vitro pharmacological examinations. It was found that catalpol (1-10 µmol/L) could significantly promote the proliferation of osteoblasts in a dose-dependent manner. It also significantly increased the area of bone staining in osteoporosis zebrafish model. The osteoblast CMC/UPLC-TOF/MS method could quickly screen the anti-osteoporosis active ingredients in complex traditional Chinese medicine prescriptions, and had the advantages of simple operation, high efficiency and high sensitivity.