Tetramethylpyrazine Protects Against Glucocorticoid-Induced Apoptosis by Promoting Autophagy in Mesenchymal Stem Cells and Improves Bone Mass in Glucocorticoid-Induced Osteoporosis Rats.

Glucocorticoid-induced osteoporosis (GIOP) is a widespread clinical complication due to the common use of glucocorticoids. Excess glucocorticoids induce apoptosis of bone marrow-derived mesenchymal stem cells (BMSCs), which have been shown to play an increasingly important role in the pathogenesis and therapy of osteoporosis. Tetramethylpyrazine (TMP), an extract from one of the most recognized herbs in traditional Chinese medicine (Chuanxiong), has been reported to have antiapoptotic properties. In this study, we tested whether TMP protects rat BMSCs following exposure to glucocorticoids in vitro and in vivo. We treated BMSCs with different concentrations of TMP (50, 100, or 200 µM) and exposed them to 10-6 M dexamethasone (Dex) for 48 h in vitro. Our data showed that TMP inhibited Dex-induced cytotoxicity and protected BMSCs from apoptosis. Interestingly, further results demonstrated that TMP prevented apoptosis in BMSCs by promoting autophagy in an AMP-activated protein kinase (AMPK) and mammalian target of rapamycin (mTOR) pathway-dependent manner. In addition, calcein fluorescence double labeling and microcomputed tomography scanning indicated that 12 weeks of TMP administration augmented bone formation and protected trabecular bone mass in GIOP rats. We also discovered that first-passage BMSCs isolated from the TMP treatment group had a lower rate of apoptosis and a higher light chain 3 (LC3)-II/LC3-I ratio than the GIOP group. Our findings demonstrate for the first time that TMP can protect BMSCs from exposure to excess glucocorticoids by promoting autophagy through AMPK/mTOR pathway and might be an effective agent for the prevention and treatment of GIOP.

Effects of Ginkgo Biloba Extract EGb-761 on Neuropathic Pain in Mice: Involvement of Opioid System.

Neuropathic pain is considered as one of the most difficult types of pain to manage with conventional analgesics. EGb-761 is extracted from leaves of Ginkgo biloba and has analgesia and anti-inflammatory properties. This study aimed to examine the effect of EGb-761 on chronic constriction injury (CCI)-induced neuropathic pain behaviors, including thermal hyperalgesia and mechanical allodynia, and to explore the possible mechanisms underlying this action. To this end, CCI mice were intraperitoneally injected with EGb-761 (10, 20, 40, and 80 mg/kg), and thermal hyperalgesia, mechanical allodynia, cytokines, and mu-opioid receptor expression were measured. Results showed that EGb-761 attenuated thermal hyperalgesia and mechanical allodynia dose-dependently and the best delivery time window was from day 7 to day 14 after CCI. Additionally, EGb-761 treatment significantly decreased pro-inflammatory cytokines and enhanced mu opioid receptor (MOR) expression in the sciatic nerve. Moreover, the opioid antagonist naloxone prevented the effect of EGb-761 on thermal hyperalgesia and mechanical allodynia but did not influence the effect of EGb-761 on inflammatory cytokines. In conclusion, this study suggests that the potential of EGb-761 as a new analgesic for neuropathic pain treatment, and opioid system may be involved in the EGb-761-induced attenuation of thermal hyperalgesia and mechanical allodynia. Copyright © 2016 John Wiley & Sons, Ltd.

Gastrodin: an ancient Chinese herbal medicine as a source for anti-osteoporosis agents via reducing reactive oxygen species.

Increased levels of reactive oxygen species (ROS) are a crucial pathogenic factor of osteoporosis. Gastrodin, isolated from the traditional Chinese herbal agent Gastrodia elata, is a potent antioxidant. We hypothesized that gastrodin demonstrates protective effects against osteoporosis by partially reducing reactive oxygen species in human bone marrow mesenchymal stem cells (hBMMSCs) and a macrophage cell line (RAW264.7 cells). We investigated gastrodin on osteogenic and adipogenic differentiation under oxidative stress in hBMMSCs. We also tested gastrodin on osteoclastic differentiation in RAW264.7 cells. Hydrogen peroxide (H2O2) was used to establish an oxidative cell injury model. Our results showed that gastrodin significantly promoted the proliferation of hBMMSCs, improved some osteogenic markers, reduced lipid generation and inhibited the mRNA expression of several adipogenic genes in hBMMSCs. Moreover, gastrodin reduced the number of osteoclasts, TRAP activity and the expression of osteoclast-specific genes in RAW264.7 cells. Gastrodin suppressed the production of reactive oxygen species in both hBMMSCs and RAW264.7 cells. In vivo, we established a murine ovariectomized (OVX) osteoporosis model. Our data revealed that gastrodin treatment reduced the activity of serum bone degradation markers, such as CTX-1 and TRAP. Importantly, it ameliorated the micro-architecture of trabecular bones. Gastrodin decreased osteoclast numbers in vivo by TRAP staining. To conclude, these results indicated that gastrodin shows protective effects against osteoporosis linking to a reduction in reactive oxygen species, suggesting that gastrodin may be useful in the prevention and treatment of osteoporosis.

Ophiopogonin D: A new herbal agent against osteoporosis.

Excessive reactive oxygen species (ROS) play an important role in the development of osteoporosis. Ophiopogonin D (OP-D), isolated from the traditional Chinese herbal agent Radix Ophiopogon japonicus, is a potent anti-oxidative agent. We hypothesized that OP-D demonstrates anti-osteoporosis effects via decreasing ROS generation in mouse pre-osteoblast cell line MC3T3-E1 subclone 4 cells and a macrophage cell line RAW264.7 cells. We investigated OP-D on osteogenic and osteoclastic differentiation under oxidative status. Hydrogen peroxide (H2O2) was used to establish an oxidative damage model. In vivo, we established a murine ovariectomized (OVX) osteoporosis model. Then, we searched the molecular mechanism of OP-D against osteoporosis. Our results revealed that OP-D significantly promoted the proliferation of MC3T3-E1 cells and improved some osteogenic markers. Moreover, OP-D reduced TRAP activity and the mRNA expressions of osteoclastic genes in RAW264.7 cells. OP-D suppressed ROS generation in both MC3T3-E1 and RAW264.7 cells. OP-D treatment reduced the activity of serum bone degradation markers, including CTX-1 and TRAP. Further research showed that OP-D displayed anti-osteoporosis effects via reducing ROS through the FoxO3a-ß-catenin signaling pathway. In summary, our results indicated that the protective effects of OP-D against osteoporosis are linked to a reduction in oxidative stress via the FoxO3a-ß-catenin signaling pathway, suggesting that OP-D may be a beneficial herbal agent in bone-related disorders, such as osteoporosis.

Protective effects of myricitrin against osteoporosis via reducing reactive oxygen species and bone-resorbing cytokines.

Oxidative stress is a crucial pathogenic factor in the development of osteoporosis. Myricitrin, isolated from Myrica cerifera, is a potent antioxidant. We hypothesized that myricitrin possessed protective effects against osteoporosis by partially reducing reactive oxygen species (ROS) and bone-resorbing cytokines in osteoblastic MC3T3-E1 cells and human bone marrow stromal cells (hBMSCs). We investigated myricitrin on osteogenic differentiation under oxidative stress. Hydrogen peroxide (H2O2) was used to establish an oxidative cell injury model. Our results revealed that myricitrin significantly improved some osteogenic markers in these cells. Myricitrin decreased lipid production and reduced peroxisome proliferator-activated receptor gamma-2 (PPARγ2) expression in hBMSCs. Moreover, myricitrin reduced the expression of receptor activator of nuclear factor kappa-B ligand (RANKL) and IL-6 and partially suppressed ROS production. In vivo, we established a murine ovariectomized (OVX) osteoporosis model. Our results demonstrated that myricitrin supplementation reduced serum malondialdehyde (MDA) activity and increased reduced glutathione (GSH) activity. Importantly, it ameliorated the micro-architecture of trabecular bones in the 4th lumbar vertebrae (L4) and distal femur. Taken together, these results indicated that the protective effects of myricitrin against osteoporosis are linked to a reduction in ROS and bone-resorbing cytokines, suggesting that myricitrin may be useful in bone metabolism diseases, particularly osteoporosis.

Ginsenoside-Rb2 displays anti-osteoporosis effects through reducing oxidative damage and bone-resorbing cytokines during osteogenesis.

Reactive oxygen species (ROS) are a significant pathogenic factor of osteoporosis. Ginsenoside-Rb2 (Rb2), a 20(S)-protopanaxadiol glycoside extracted from ginseng, is a potent antioxidant that generates interest regarding the bone metabolism area. We tested the potential anti-osteoporosis effects of Rb2 and its underlying mechanism in this study. We produced an oxidative damage model induced by hydrogen peroxide (H2O2) in osteoblastic MC3T3-E1 cells to test the essential anti-osteoporosis effects of Rb2in vitro. The results indicated that treatment of 0.1 to 10µM Rb2 promoted the proliferation of MC3T3-E1 cells, improved alkaline phosphatase (ALP) expression, elevated calcium mineralization and mRNA expressions of Alp, Col1a1, osteocalcin (Ocn) and osteopontin (Opn) against oxidative damage induced by H2O2. Importantly, Rb2 reduced the expression levels of receptor activator of nuclear factor kappa-B ligand (RANKL) and IL-6 and inhibited the H2O2-induced production of ROS. The in vivo study indicated that the Rb2 administered for 12weeks partially decreased blood malondialdehyde (MDA) activity and elevated the activity of reduced glutathione (GSH) in ovariectomized (OVX) mice. Moreover, Rb2 improved the micro-architecture of trabecular bones and increased bone mineral density (BMD) of the 4th lumbar vertebrae (L4) and the distal femur. Altogether, these results demonstrated that the potential anti-osteoporosis effects of Rb2 were linked to a reduction of oxidative damage and bone-resorbing cytokines, which suggests that Rb2 might be effective in preventing and alleviating osteoporosis.

Protection by salidroside against bone loss via inhibition of oxidative stress and bone-resorbing mediators.

Oxidative stress is a pivotal pathogenic factor for bone loss in mouse model. Salidroside, a phenylpropanoid glycoside extracted from Rhodiola rosea L, exhibits potent antioxidative effects. In the present study, we used an in vitro oxidative stress model induced by hydrogen peroxide (H(2)O(2)) in MC3T3-E1 cells and a murine ovariectomized (OVX) osteoporosis model to investigate the protective effects of salidroside on bone loss and the related mechanisms. We demonstrated that salidroside caused a significant (P<0.05) elevation of cell survival, alkaline phosphatase (ALP) staining and activity, calcium deposition, and the transcriptional expression of Alp, Col1a1 and Osteocalcin (Ocn) in the presence of H(2)O(2). Moreover, salidroside decreased the production of intracellular reactive oxygen species (ROS), and osteoclast differentiation inducing factors such as receptor activator of nuclear factor-kB ligand (RANKL) and IL-6 induced by H(2)O(2). In vivo studies further demonstrated that salidroside supplementation for 3 months caused a decrease in malondialdehyde (MDA) and an increase in reduced glutathione (GSH) concentration in blood of ovariectomized mouse (P<0.05), it also improved trabecular bone microarchitecture and bone mineral density in the fourth lumbar vertebra and distal femur. Our study indicated that the protection provided by salidroside in alleviating bone loss was mediated, at least in part, via inhibition of the release of bone-resorbing mediators and oxidative damage to bone-forming cells, suggesting that salidroside can be used as an effective remedy in the treatment or prevention of osteoporosis.

Tanshinone IIA attenuates the inflammatory response and apoptosis after traumatic injury of the spinal cord in adult rats.

BACKGROUND: Spinal cord injury (SCI), including immediate mechanical injury and secondary injury, is associated with the inflammatory response, apoptosis and oxidative stress in response to traumatic injury. Tanshinone IIA (TIIA) is one of the major extracts obtained from Salvia miltiorrhiza BUNGE, which has anti-inflammatory and anti-apoptotic effects on many diseases. However, little is known about the effects of TIIA treatment on SCI. Therefore, the aim of the present study is to evaluate the pharmacological action of TIIA on secondary damage and the underlying mechanisms of experimental SCI in rats. METHODOLOGY/PRINCIPAL FINDINGS: SCI was generated using a weight drop device on the dorsal spinal cord via a two-level T9-T11 laminectomy. SCI in rats resulted in severe trauma, characterized by locomotor disturbance, edema, neutrophil infiltration, the production of astrocytes and inflammatory mediators, apoptosis and oxidative stress. TIIA treatment (20 mg/kg, i.p.) after SCI induced significant effects: (1) improved motor function (Basso, Beattie and Bresnahan scores), (2) reduced the degree of tissue injury (histological score), neutrophil infiltration (myeloperoxidase activity) and the expression of astrocytes, (3) inhibited the activation of SCI-related pathways, such as NF-κB and MAPK signaling pathways, (4) decreased the production of pro-inflammatory cytokines (TNF-α, IL-1ß, and IL-6) and iNOS, (5) reduced apoptosis (TUNEL staining, and Bcl-2 and caspase-3 expression) and (6) reversed the redox state imbalance. CONCLUSIONS/SIGNIFICANCE: The results clearly show that TIIA has a prominent protective effect against SCI through inhibiting the inflammatory response and apoptosis in the spinal cord tissue after SCI.

Hyperbaric oxygen preconditioning ameliorates anxiety-like behavior and cognitive impairments via upregulation of thioredoxin reductases in stressed rats.

The present study examined the protective effect of hyperbaric oxygen preconditioning (HBO-PC) and the role of thioredoxin reductase (TrxR) in a post-traumatic stress disorder (PTSD)-induced rat model by using single prolonged stress (SPS). Rats were randomly divided into Sham, HBO, SPS and HBO+SPS groups. HBO-PC was conducted by exposing rats to 100% oxygen at 2.5atm absolute for 1h each day for 5 consecutive days. SPS was performed 24h after the last HBO-PC conditioning event. At 1h, 6h, 12h, 24h and 72h after SPS, TrxR mRNA expression was analyzed in the hippocampus; Nissl and TUNEL staining were performed at 72h after SPS. The results indicated that HBO-PC was able to significantly preserve viable neurons in the CA1 subfield of hippocampus following SPS exposure, as evidenced by reduced amounts of CA1 neuronal apoptosis. Furthermore, HBO-PC upregulate the expression of TrxR-1 and TrxR-2 mRNA in the hippocampus at 6h and 12h after SPS exposure and ameliorated anxiety-like behavior and cognitive impairments normally induced by SPS. Taken together, these findings suggest that HBO-PC is beneficial for the improvement of anxiety-like behavior and cognitive impairments induced by SPS exposure, and this effect might be associated with inhibition of neuronal apoptosis via upregulation of TrxR in stressed rats.

[Effects of Yunnan Baiyao on peri-operative bleeding of patients undergoing cervical open-door laminoplasty: a multi-center randomized double-blind placebo-control trial].

OBJECTIVE: To investigate the effects of Yunnan Baiyao on peri-operative bleeding of the patients undergoing cervical open-door laminoplasty. METHODS: 197 patients undergoing cervical open-door laminoplasty were randomly divided into 2 groups Yunnan Baiyao group (receiving Yunnan Baiyao 500 mg three times daily for 5 days) and placebo group. The amounts of intra-operative and post-operative bleeding were compared, and the side effect of Yunnan Baiyao was also analyzed. RESULTS: Statistical analysis showed that the amount of intra-operative bleeding of the Yunnan Baiyao was 350 ml +/- 190 ml, significantly lower than that of the placebo group (443 ml +/- 266 ml, P < 0.05). There were no statistical differences in the amount of postoperative bleeding and side effect rate between the two groups. CONCLUSION: Yunnan Baiyao is effective and safe in reducing the amount of intra-operative bleeding of cervical open-door laminoplasty.