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Chondrocytes that are embedded within the growth plate or the intervertebral disc are sensitive to environmental stresses, such as inflammation and hypoxia. However, little is known about the molecular signalling pathways underlining the hypoxia-induced mitochondrial dysfunction and apoptosis in chondrocytes. In the present study, we firstly examined the hypoxia-induced apoptosis, mitochondrial dysfunction and the activation of cyclic adenosine monophosphate (cAMP) response element-binding protein (CREB) signalling in human chondrocyte cell line, C28/I2 and then investigated the regulatory role of RhoA, a well-recognized apoptosis suppressor, in such process, with gain-of-function strategy. Our results indicated that hypoxia induced apoptosis and inhibited CREB phosphprylation in chondrocytes, meanwhile, the dysfunctional mitochondria with up-regulated mitochondrial superoxide and reactive oxygen species (ROS) levels, whereas with a reduced mitochondrial membrane potential (MMP) and Complex IV activity were observed in the hypoxia-treated C28/I2 cells. However, the overexpressed RhoA blocked the hypoxia-mediated reduction in CREB phosphprylation and inhibited the apoptosis induction, along with an ameliorated mitochondrial function in the hypoxia-treated C28/I2 cells. In conclusion, the present study confirmed the reduced CREB phosphorylation, along with the apoptosis induction and mitochondrial dysfunction in the hypoxia-treated chondrocyte cells. And the overexpression of RhoA ameliorated the hypoxia-induced mitochondrial dysfunction and apoptosis via blocking the hypoxia-mediated reduction in CREB phosphorylation. [ABSTRACT FROM AUTHOR]

Inflammation and cytokines have been recognized to correlate with intervertebral disc (IVD) degeneration (IDD), via mediating the development of clinical signs and symptoms. However, the regulation mechanism remains unclear. We aimed at investigating the regulatory role of interleukin (IL)β and high mobility group box 1 (HMGB1) in the inflammatory response in human IVD cells, and then explored the signalling pathways mediating such regulatory effect. Firstly, the promotion to inflammatory cytokines in IVD cells was examined with ELISA method. And then western blot and real time quantitative PCR were performed to analyse the expression of toll-like receptors (TLRs), receptors for advanced glycation endproducts (RAGE) and NF-κB signalling markers in the IL-1β- or (and) HMGB1-treated IVD cells. Results demonstrated that either IL-1β or HMGB1 promoted the release of the inflammatory cytokines such as prostaglandin E2 (PGE2), TNF-α, IL-6 and IL-8 in human IVD cells. And the expression of matrix metalloproteinases (MMPs) such as MMP-1, -3 and -9 was also additively up-regulated by IL-1β and HMGB1. We also found such additive promotion to the expression of TLR-2, TLR-4 and RAGE, and the NF-κB signalling in intervertebral disc cells. In summary, our study demonstrated that IL-1β and HMGB1 additively promotes the release of inflammatory cytokines and the expression of MMPs in human IVD cells. The TLRs and RAGE and the NF-κB signalling were also additively promoted by IL-1β and HMGB1. Our study implied that the additive promotion by IL-1β and HMGB1 to inflammatory cytokines and MMPs might aggravate the progression of IDD. [ABSTRACT FROM AUTHOR]

Background: Mesenchymal stem cells (MSCs) can differentiate into chondroblasts, adipocytes, or osteoblasts under appropriate stimulation. Ghrelin, an endogenous ligand for the growth hormone secretagogue receptor (GHSR), stimulates growth hormone (GH) secretion, and has both orexigenic and adipogenic effects. This study sought to understand the potential involvement of members of MAPK serine/threonine kinases in the ghrelin-induced growth of rabbit MSCs ( rBMSC). Methods: We applied various concentrations of ghrelin to cultured rBMSC and observed the growth rate of the cells by MTT, changes in the phosphorylation state of ERK1/2, JNK and p38, and the expression levels of ALP, Runx2, and Osterix by wetern blot. Results: We found that the growth and osteogenic differentiation of ghrelin-treated rBMSC are promoted primarily by phosphorylated ERK1/2, and that this phosphorylation, as well p38 phosphorylation, is mediated by GHSR. Conclusions: Our study suggests that ghrelin promotes the growth and osteogenic differentiation of rBMSC primarily through the ERK1/2 pathway. [ABSTRACT FROM AUTHOR]

Mesenchymal stromal progenitor cells (MSCs) are multipotent progenitors that can be isolated from numerous tissues. MSCs can undergo osteogenic differentiation under proper stimuli. We have recently demonstrated that bone morphogenetic protein 9 (BMP9) is one of the most osteogenic BMPs. As one of the least studied BMPs, BMP9 has been shown to regulate angiogenesis in endothelial cells. However, it is unclear whether BMP9-regulated angiogenic signaling plays any important role in the BMP9-initiated osteogenic pathway in MSCs. Here, we investigate the functional role of hypoxia-inducible factor 1α (HIF1α)-mediated angiogenic signaling in BMP9-regulated osteogenic differentiation of MSCs. We find that BMP9 induces HIF1α expression in MSCs through Smad1/5/8 signaling. Exogenous expression of HIF1α potentiates BMP9-induced osteogenic differentiation of MSCs both in vitro and in vivo. siRNA-mediated silencing of HIF1α or HIF1α inhibitor CAY10585 profoundly blunts BMP9-induced osteogenic signaling inMSCs. HIF1α expression regulated by cobalt-induced hypoxia also recapitulates the synergistic effect between HIF1α and BMP9 in osteogenic differentiation. Mechanistically, HIF1α is shown to exert its synergistic effect with BMP9 by inducing both angiogenic signaling and osteogenic signaling in MSCs. Thus, our findings should not only expand our understanding of the molecular basis behind BMP9-regulated osteoblastic lineage-specific differentiation, but also provide an opportunity to harness the BMP9-induced synergy between osteogenic and angiogenic signaling pathways in regenerative medicine. [ABSTRACT FROM AUTHOR]

Background: Nucleus pulposus (NP) cell senescence is an important cellular feature within the degenerative disc. It is known that a very acidic niche exists in the degenerative disc, which participates in regulating disc cell viability and matrix metabolism. Objective: The present study was aimed to investigate the role and potential signaling transduction pathway of an acidic pH in regulating NP cell senescence. Methods: Rat NP cells were cultured in an acidic pH of 7.2 close to that in a healthy disc (Control group) or in an acidic pH of 6.2 close to that in a severe degenerative disc (Experiment group) for 10 days. Additionally, the experimental NP cells were incubated along with the inhibitor SB203580 to analyze the role of p38 MAPK pathway in this process. Results: Compared with the control NP cells, experimental NP cells showed a suppressed cell proliferation potency, an increased G0/G1 phase fraction whereas a decreased S-phase fraction and a declined telomerase activity, an up-regulated expression of senescence-related molecules (p16 and p53), and a down-regulated expression of matrix-related moleucles (aggrecan and collagen II). Further analysis showed that inhibition of the p38 MAPK pathway partly reversed effects of acidic pH of 6.2 on the experimental NP cells. Conclusion: The very acidic niche identified in a severe degenerative disc promotes NP cell senescence through regulating the p38 MAPK pathway. The present study provides a new mechanism that drives NP cell senescence during disc degeneration. [ABSTRACT FROM AUTHOR]

Bone-like hydroxyapatite/poly amino acid (BHA/PAA) is a potential bone repair material. Rifapentine-loaded poly(lactic-co-glycolic acid) microspheres (RPMs) are bioactive and efficient controlled-release delivery systems used in vitro. The aim of the present study was to investigate the in vivo drug release characteristics of RPM-loaded BHA/PAA on a rabbit model of bone defect. RPM was combined with BHA/PAA to obtain the drug-loaded, slow-releasing bioactive material. Bone defects were generated in New Zealand white rabbits and the rabbits were then implanted with RPM-loaded BHA/PAA. High-performance liquid chromatography (HPLC) was used to determine the concentrations of rifapentine in the plasma and the local muscle tissues of the treated rabbits. Hematoxylin and eosin (H&E) staining and biochemical analyses were performed to elucidate potential side effects of RPM-loaded BHA/PAA on the heart, liver and kidney histopathology and functions of the treated rabbits. The biocompatibility and osteogenic ability of RPM-loaded BHA/PAA was evaluated by H&E staining. The results demonstrated that the material was completely degraded and absorbed at 12 weeks following implantation and new trabecular bone and cartilage tissues had formed. The in vivo release tests revealed that RPM-loaded BHA/PAA exhibited sustained release profiles of rifapentine and the drug concentration in the muscle tissues remained higher than the minimum inhibitory concentration of rifapentine against Mycobacterium tuberculosis for as long as 12 weeks. In addition, RPM-loaded BHA/PAA had no long-term side effects to the heart, liver and kidney of the treated rabbits. In conclusion, the present study demonstrated that RPM-loaded BHA/PAA slowly and continuously released rifapentine in vivo and exhibited no side effects on heart, liver and kidney tissues and function. Furthermore, RPM-loaded BHA/PAA promoted new bone formation, while it was gradually degraded and absorbed. The present study provided a theoretical basis for the potential advancement in developing novel treatments for osteoarticular tuberculosis. [ABSTRACT FROM AUTHOR]

Osteoporosis is a disease of the skeleton that is characterized by the loss of bone mass and degeneration of bone microstructure, resulting in an increased risk of fracture. Oxidative stress, which is known to promote oxidative damage to mitochondrial function and also cell apoptosis, has been recently indicated to be implicated in osteoporosis. However, there are few agents that counteract oxidative stress in osteoporosis. In the present study, the protective effects of allicin against the oxidative stress-induced mitochondrial dysfunction and apoptosis were investigated in murine osteoblast-like MC3T3-E1 cells. The results demonstrated that allicin counteracted the reduction of cell viability and induction of apoptosis caused by hydrogen peroxide (H2O2) exposure. The inhibition of apoptosis by allicin was confirmed by the inhibition of H2O2-induced cytochrome c release and caspase-3 activation. Moreover, the inhibition of apoptosis by allicin was identified to be associated with the counteraction of H2O2-induced mitochondrial dysfunction. In addition, allicin was demonstrated to be able to significantly ameliorate the repressed phosphoinositide 3-kinase (PI3K)/AKT and cyclic adenosine monophosphate response element-binding protein (CREB)/extracellular-signal-regulated kinase (ERK) signaling pathways by H2O2, which may also be associated with the anti-oxidative stress effects of allicin. In conclusion, allicin protects osteoblasts from H2O2-induced oxidative stress and apoptosis in MC3T3-E1 cells by improving mitochondrial function and the activation of PI3K/AKT and CREB/ERK signaling. The present study implies a promising role of allicin in oxidative stress-associated osteoporosis. [ABSTRACT FROM AUTHOR]

In the last 30 years, the 5-year-survival rate of patients with osteosarcoma has not improved as a result of the low prevalence and large tumor heterogeneity. Therefore, the development of novel drugs for the treatment of osteosarcoma is urgently required. The present study aimed to identify potential novel drugs for the treatment of osteosarcoma, thus used β-catenin as a target and performed high content screening. In a total of 14 botanical extracts assessed, resveratrol markedly downregulated the expression of β-catenin and significantly inhibited MG-63 cell proliferation. CCK-8 assay was used to confirm the anti-osteosarcoma effect of resveratrol and flow cytometry and western blotting were performed to analyze the underlying mechanisms of the proapoptotic effects of resveratrol. β-catenin is a vital member of the canonical Wnt signaling pathway and, therefore, the target genes of this pathway were further analyzed. The results of this analysis demonstrated that resveratrol suppressed the MG-63 cells by inhibiting the canonical Wnt signaling pathway. [ABSTRACT FROM AUTHOR]

Background: Perioperative pain after total knee arthroplasty (TKA) may seriously affect the rapid recovery of patients. The purpose of this study was to assess whether the combined use of adductor canal block (ACB) and local infiltration anesthesia (LIA) can further reduce postoperative pain and improve early functional recovery. Materials and methods: PubMed, Web of Science, EMBASE, and Cochrane Central Register of Controlled Trials were systematically searched for randomized controlled trials (RCTs) comparing ACB + LIA and LIA alone in primary TKA. The primary outcomes were visual analog scale (VAS) scores at rest and walking, morphine consumption, range of motion (ROM) at 24 and 48 h postoperatively and distance walked. The secondary outcomes were the length of stay, the incidence of nausea and vomiting, and the total complications. Subgroup analyses were performed on the VAS at rest and walking, morphine consumption, and distance walked at 24, 48, and 72 h postoperatively. Results: A total of 10 RCTs involving 797 patients were enrolled in this meta-analysis. The results demonstrated that the combined application of ACB + LIA had a lower resting VAS at 24 h postoperatively (p = 0.02) and the walking score at 24 (p = 0.0002) and 48 h (p = 0.02) postoperatively compared with LIA alone. Similarly, the combined ACB + LIA group also had less morphine consumption at 48 h postoperatively (p = 0.0005) and had a higher ROM score at 24 h (p = 0.01) postoperatively compared to the LIA group. There were no statistical differences in length of stay, distance walked, and incidence of nausea and vomiting. Conclusion: The current meta-analysis showed that ACB + LIA significantly reduced postoperative walking pain and morphine consumption and promoted rapid recovery in the early postoperative period. There is no statistical difference in the length of stay and ROM after 72 h in the two groups. [ABSTRACT FROM AUTHOR]

Background: Osteophytes that form adjacent to degenerated disc have osteogeic potential. Studies suggest that their formation is associated with mesenchymal precursors arising from the chondrosynovial junction. This study is aimed to determine the cellular aging and osteogenic differentiation potential of osteophyte-derived mesenchymal cells (oMSCs) when compared to patient-matched bone marrow stromal cells (bMSCs). Materials and Methods: oMSCs and bMSCs were isolated from tissue samples during anterior cervical discectomy and fusion surgery. Extensive expansion of cell cultures was performed and early and late passage cells (P4 and P9, respectively) were used to study cell senescence and telomerase activity. Furthermore, osteogenic differentiation was applied to detect their osteogenic capacity. Results: The proliferation capacity of oMSCs in culture was superior to that of bMSCs and these cells readily underwent osteogenic differentiation. Our results showed that oMSCs had higher telomerase activity in late passages compared with bMSCs, although there was no significant difference in the telomerase activity in the early passages in either cell types. The telomerase activity was detectable only in early passage oMSCs and not in bMSCs. Conclusions: Our results indicate that oMSCs retain a level of telomerase activity in vitro, which may account for the relatively greater longevity of these cells, compared to bMSCs. Furthermore, when compared to bMSCs, oMSCs maintained a higher proliferative capacity and the same osteogenic capacity, which may offer new insights of tissue formation. [ABSTRACT FROM AUTHOR]

SUMMARY: STUDY DESIGN Normal rats and paraplegic rats with and injured spinal cord were used to study bacterial translocation from gut.OBJECTIVE To investigate whether bacterial translocation from gut occurs after spinal cord injury.SUMMARY OF BACKGROUND DATA It has been demonstrated that trauma and operation can lead to gastrointestinal paralysis; disturbance of gastrointestinal motility following trauma may cause bacterial overgrowth in gastrointestinal tract and increase the incidence of bacterial translocation. However, bacterial translocation from gut after spinal cord injury has not been studied.METHODS Under aseptic manipulation, samples of blood were collected for bacterial cultures and endotoxin determination. In the meantime, samples of mesenteric lymph node, spleen, and liver were collected for bacterial culture. The jejunum and ileum were observed by light and electron microscope.RESULTS Endotoxemia and bacterial translocation appeared 24 hours and 48 hours correspondingly after spinal cord injury complicated with paraplegia.CONCLUSION Bacterial translocation from gut would occur after spinal cord injury in rats, which indicated that antibiotics should be administered to paraplegic patients with spinal cord injury as soon as possible to prevent potential bacterial translocation. [ABSTRACT FROM AUTHOR]

Mini: The anatomical relationship of the extrapedicular approach with the spinal nerve and its branches was researched in cadavers. Three types of extrapedicular paths were simulated: puncture inside the medial branch (MB), puncture outside the MB, and puncture outside the lateral branch (LB). At L1-L3, puncture outside the LB could avoid damage to these nerves. [ABSTRACT FROM AUTHOR]

Wingless/integrated (Wnt) signaling has emerged as a major mechanism for promoting bone formation and a target pathway for developing bone anabolic agents against osteoporosis. However, the downstream events mediating the potential therapeutic effect of Wnt proteins are not fully understood. Previous studies have indicated that increased glycolysis is associated with osteoblast differentiation in response to Wnt signaling, but direct genetic evidence for the importance of glucose metabolism in Wnt-induced bone formation is lacking. Here, we have generated compound transgenic mice to overexpress Wnt family member 7B (Wnt7b) transiently in the osteoblast lineage of postnatal mice, with or without concurrent deletion of the glucose transporter 1 (Glut1), also known as solute carrier family 2, facilitated glucose transporter member 1. Overexpression of Wnt7b in 1-mo-old mice for 1 wk markedly stimulated bone formation, but the effect was essentially abolished without Glut1, even though transient deletion of Glut1 itself did not affect normal bone accrual. Consistent with the in vivo results, Wnt7b increased Glut1 expression and glucose consumption in the primary culture of osteoblast lineage cells, and deletion of Glut1 diminished osteoblast differentiation in vitro. Thus, Wnt7b promotes bone formation in part through stimulating glucose metabolism in osteoblast lineage cells. [ABSTRACT FROM AUTHOR]

Background: Intervertebral disc degeneration is a pathological process that involves an inflammation response. As a classical cellular feature, several studies have demonstrated that inflammation can promote nucleus pulposus (NP) cell apoptosis. Therefore, attenuation of NP cell apoptosis may be a potential way to retard disc degeneration. Objective: The present study was aimed to investigate the protective effects of osteogenic protein-1 (OP-1) against NP cell apoptosis in an inflammation environment, and the potential signaling transduction pathway. Methods: Rat NP cells were cultured in medium with or without inflammatory cytokine tumor necrosis factor (TNF)-α for 6 days. The exogenous TNF-α was added into the medium to investigate its protective effects. NP cell apoptosis was evaluated by cell apoptosis ratio, caspase-3 activity, gene/protein expression of apoptosis-related molecules (Bcl-2, Bax, and caspase-3). Additionally, the intracellular reactive oxygen species (ROS) content and activity of the NF-κB pathway were also analyzed. Results: Compared with the control NP cells, TNF-α significantly increased cell apoptosis ratio, caspase-3 activity, gene/protein expression of Bcl-2, Bax and caspase-3, ROS content, and activity of the NF-κB pathway. However, OP-1 partly attenuated these effects in NP cells treated with TNF-α. Conclusion: OP-1 is effective in attenuating TNF-α-caused NP cell apoptosis, and the ROS/NF-κB pathway may be the potential signaling transduction pathway. The present study indicates that OP-1 may be helpful to inhibit inflammation-mediated disc degeneration. [ABSTRACT FROM AUTHOR]

Background: Osteosarcoma (OS) is well-known for poor prognosis due to its high incidence of proliferation and metastasis. Researches have provided valuable insights into the tumorigenesis of S100A9 in some cancers. We aimed to understand the expression level, functions and mechanisms of S100A9 in human osteosarcoma for the first time.Methods: The expression of S100A9 protein was detected in 120 human osteosarcoma tissues and 40 normal human bone tissues using tissue microarrays analysis. The knockdown of S100A9 induced by RNA interference (RNAi) method in three osteosarcoma cell lines (U2OS, 143B, MG63) was applied to analyze the effects of S100A9 on cell proliferation, cell cycle distribution, migration, invasion and xenotransplanted tumors. Moreover, MAPK-ERK1/2, MAPK-p38, NF-κB-p65, NF-κB-p50, p21, p27, CDK2 and CDK4 were tested.Results: The expression of S100A9 was increased in human osteosarcoma issues and was positively correlated with clinical classification and survival rate. Down-regulation of S100A9 inhibited OS cellular proliferation, migration, invasion and cell cycle S phase in vitro and suppressed tumor formation in vivo with the reduction on PCNA and Ki67 proliferation index. Our data also demonstrated that knockdown of S100A9 repressed the protein levels of phospho-ERK1/2, phospho-p50, phospho-p65 except phospho-p38, and prompted up-regulation of p21 and p27 leading to inactivation of cyclin dependent kinase 2(CDK2) and cyclin dependent kinase 4(CDK4).Conclusions: S100A9 might be a significant role for predicting osteosarcoma prognosis and down-regulation of S100A9 could be used as a potential target for gene therapy. [ABSTRACT FROM AUTHOR]

The aim of the present study was to explore the effect on nitric oxide (NO) content and osteocyte apoptosis of steroid-induced avascular necrosis of the femoral head (SANFH) in an animal model of SANFH. A total of 40 Japanese white rabbits, 5 months of age and weighing 2.5±0.5 kg, were randomly divided into groups A (hormone + endotoxin group), B (endotoxin + normal saline group), C (normal saline + hormone group) and D (control group). Following the establishment of the model, a blood sample was taken from the heart of each animal and centrifuged; the levels of NO in the serum were detected. The bilateral femoral heads were conventionally dissected, fixed, decalcified and stained with hematoxylin and eosin. Subsequently, the empty bone lacunae were counted under an optical microscope. Changes in osteocyte morphology were observed using electron microscopy and osteocyte apoptosis was detected with a terminal deoxynucleotidyl transferase dUTP nick end labeling assay. The percentage of empty bone lacunae in group A was significantly higher compared with that in groups B, C and D (P<0.01); however, there was no significant difference in percentage among groups B, C and D. The NO content in group A was significantly higher compared with that in groups B, C and D (P<0.01); however, there was no significant difference in NO content among groups B, C and D. The osteocyte apoptosis index in group A was significantly higher compared with that in the other groups (P<0.01); there was no significant difference among groups B, C and D. NO content was positively correlated with osteocyte apoptosis index (r=0.707). Thus, the present study found that NO content and the osteocyte apoptosis index were increased in SANFH, and that they play an important role in SANFH. The content of NO was positively correlated with the osteocyte apoptosis index, indicating that NO induces apoptosis. [ABSTRACT FROM AUTHOR]

Although classified as benign, giant cell tumors of the bone (GCTB) may be aggressive, recur and even metastasize to the lungs. In addition, the pathogenesis and histogenesis remain unclear; thus, the driving factors behind the strong tumor growth capacity of GCTB require investigation. In the present study, the expression levels of hypoxia-inducible factor (HIF)-1α and vascular endothelial growth factor (VEGF), which are promoted by hypoxic conditions, were determined in 22 sacral GCTB samples using immunohistochemistry and western blot analysis. Furthermore, CD34 expression was analyzed using these methods. The correlation between HIF-1α or VEGF expression and the tumor microvessel density (MVD) was then determined. The results demonstrated that HIF-1α, VEGF and CD34 were overexpressed in the 22 sacral GCTB specimens, and overexpression of HIF-1α and VEGF correlated with the tumor MVD. Thus, the present study has provided novel indicators for the tumor growth capacity of GCTBs. [ABSTRACT FROM AUTHOR]

The article presents a study which investigated the impact of sacral nerve stimulation to treat and prevent bacterial translocation from the gut and endotoxemia in rabbits following an acute and complete spinal cord injury (CSI). It was found that SCI-induced bacterial translocation and endotoxemia cause dysfunction in the intestinal tract that could increase the incidence of systemic inflammatory response syndrome (SIRS) and multiple organ dysfunction syndrome (MODS).