microRNA-19a protects osteoblasts from dexamethasone via targeting TSC1.

Activation of mTOR complex 1 (mTORC1) could protect human osteoblasts from dexamethasone. Tuberous sclerosis complex 1 (TSC1) is mTORC1 upstream inhibitory protein. We demonstrate here that microRNA-19a ("miR-19a", -3p) targets the 3' untranslated regions of TSC1 mRNA. Expression of miR-19a downregulated TSC1 in OB-6 osteoblastic cells and primary human osteoblasts. miR-19a activated mTORC1 and protected human osteoblasts from dexamethasone. mTORC1 inhibition, by RAD001 or Raptor shRNA, almost completely abolished miR-19a-induced osteoblast cytoprotection against dexamethasone. Knockdown of TSC1 by targeted shRNA similarly induced mTORC1 activation and protected osteoblasts. Moreover, miR-19a activated mTORC1-dependent NF-E2-related factor 2 (Nrf2) signaling and inhibited dexamethasone-induced reactive oxygen species production in osteoblasts. Together, miR-19a protects human osteoblasts from dexamethasone possibly via targeting TSC1-mTORC1 signaling.

Activation of AMP-activated protein kinase by compound 991 protects osteoblasts from dexamethasone.

Dexamethasone (Dex) induces direct cytotoxicity to cultured osteoblasts. The benzimidazole derivative compound 991 ("C991") is a novel and highly-efficient AMP-activated protein kinase (AMPK) activator. Here, in both MC3T3-E1 osteoblastic cells and primary murine osteoblasts, treatment with C991 activated AMPK signaling, and significantly attenuated Dex-induced apoptotic and non-apoptotic cell death. AMPKα1 knockdown (by shRNA), complete knockout (by CRISPR/Cas9 method) or dominant negative mutation (T172A) not only blocked C991-mediated AMPK activation, but also abolished its pro-survival effect against Dex in osteoblasts. Further studies showed that C991 boosted nicotinamide adenine dinucleotide phosphate (NADPH) activity and induced mRNA expression of NF-E2-related factor 2 (Nrf2)-regulated genes (heme oxygenase-1 and NADPH quinone oxidoreductase 1). Additionally, C991 alleviated Dex-induced reactive oxygen species (ROS) production in osteoblasts. Notably, genetic AMPK inhibition reversed the anti-oxidant actions by C991 in Dex-treated osteoblasts. Together, we conclude that C991 activates AMPK signaling to protect osteoblasts from Dex.

MicroRNA-200a activates Nrf2 signaling to protect osteoblasts from dexamethasone.

Treatment with dexamethasone in human osteoblasts leads to oxidative stress and cell injures. NF-E2-related factor 2 (Nrf2) is a key anti-oxidant signaling. We want to induce Nrf2 activation via microRNA-mediated silencing its suppressor Keap1. Our results show that microRNA-200a ("miR-200a") expression depleted Keap1, causing Nrf2 protein stabilization in OB-6 osteoblastic cells. Reversely, the miR-200a anti-sense led to Keap1 upregulation and Nrf2 degradation. miR-200a expression activated Nrf2 signaling, which inhibited dexamethasone-induced reactive oxygen species production and OB-6 cell death/apoptosis. Keap1 shRNA also activated Nrf2 and protected OB-6 cells from dexamethasone. Importantly, miR-200a was in-effective in Keap1-silenced (by shRNA) OB-6 cells. In the primary human osteoblasts, Keap1 silence by targeted-shRNA or miR-200a protected cells from dexamethasone. Significantly, miR-200a level was decreased in necrotic femoral head tissues, which was correlated with Keap1 mRNA upregulation. Together, miR-200a expression activates Nrf2 signaling and protects human osteoblasts from dexamethasone.

Activation of Nrf2 by MIND4-17 protects osteoblasts from hydrogen peroxide-induced oxidative stress.

MIND4-17 is a recently developed NF-E2-related factor 2 (Nrf2) activator, which uniquely causes Nrf2 disassociation from Keap1. Here, we showed that pretreatment with MIND4-17 significantly inhibited hydrogen peroxide (H2O2)-induced viability reduction of primary osteoblasts and OB-6 osteoblastic cells. Meanwhile, MIND4-17 inhibited both apoptotic and non-apoptotic osteoblast cell death by H2O2. MIND4-17 treatment induced Keap1-Nrf2 disassociation, causing Nrf2 stabilization, accumulation and nuclear translocation in osteoblasts, leading to transcription of several Nrf2-dependent genes, including heme oxygenase 1 (HO-1), NAD(P)H quinone oxidoreductase 1 (NQO1), γ-glutamylcysteine synthetase modifier subunit (GCLM) and catalytic subunit (GCLC). Additionally, MIND4-17 largely attenuated H2O2-reactive oxygen species (ROS) production, lipid peroxidation and DNA damages. Nrf2 knockdown by targeted short hairpin RNA (shRNA) exacerbated H2O2-induced cytotoxicity in OB-6 osteoblastic cells, and nullified MIND4-17-mediated cytoprotection against H2O2. Meanwhile, Keap1 shRNA took over MIND4-17's actions and protected OB-6 cells from H2O2. Together, MIND4-17 activates Nrf2 signaling and protects osteoblasts from H2O2.

Assessment of GSK1904529A as a promising anti-osteosarcoma agent.

The insulin growth factor-I receptor (IGF1R) signaling is a key mechanism for osteosarcoma (OS) cell proliferation. GSK1904529A is a novel small molecule IGF1R kinase inhibitor. Its activity against OS cells was tested. In both established OS cell lines (Saos-2 and MG-63) and primary human OS cells, treatment with GSK1904529A (at nM concentrations) significantly inhibited cell proliferation. At the molecular level, GSK1904529A almost completely blocked IGF1R activation in OS cells, and inhibited downstream AKT-ERK activation. IGF1R silence by targeted shRNA also inhibited AKT-ERK activation and Saos-2 cell proliferation. Significantly, GSK1904529A was unable to further inhibit proliferation of IGF1R-silenced Saos-2 cells. In vivo, GSK1904529A administration orally inhibited Saos-2 tumor growth in nude mice. Together, these results suggest that targeting IGF1R by GSK1904529A inhibits OS cell growth in vitro and in vivo.

[Selection of suitable procedure in the treatment of degenerative lumbar scoliosis based upon imaging studies].

OBJECTIVE: To compare the clinical efficacies of two different procedures in the treatment of degenerative lumbar scoliosis. METHODS: From August 2008 to August 2011, 28 patients of lumbar degenerative scoliosis were divided into one group (n = 14) undergoing modified transforaminal lumbar interbody fusion (TLIF) instrumented surgery and another group (n = 14) undergoing posterolateral fusion (PLF) instrumented surgery. There were 12 males and 16 females with a mean age of 66.2 years (range: 54-79). The operative durations and bleeding volumes of two groups were recorded. The post-operative efficacy was evaluated with VAS (visual analogue scale) for low back pain, ODI (Oswestry disability index), Cobb' angle and lumbar lordosis angle on plain film. RESULTS: The mean follow-up period was 25.9 months. The operative duration was 192.0 ± 44.7 min in modified TLIF group versus 163.0 ± 39.0 min in PLF group. The bleeding volume was 718.0 ± 197.2 ml in modified TLIF group versus 546.0 ± 226.6 ml in PLF group. All operated lumbar intervertebral achieved bony fusion in modified TLIF group by the last follow-up. Two cases had no bony fusion and there was one case of pseudarthrosis in PLF group. Significant differences existed between two groups in pre-operative and post-operative values of VAS, ODI, Cobb' s angle and lumbar lordosis angle (P < 0.05). There were significant differences between two groups in the values of pre-operative and post-operative VAS and lumbar lordosis angle (P < 0.05) but not in the values of pre-operative and post-operative ODI and Cobb' s angle (P > 0.05). CONCLUSION: As an alternative, safe and effective procedure, modified TLIF instrumented is superior to PLF instrumented in the treatment of lumbar degenerative scoliosis.

[Combination of bone marrow concentrate and PGA scaffolds enhancing bone marrow stimulation in the repair of rabbit articular cartilage].

OBJECTIVE: To observe the effects of bone marrow concentrate (BMC)-PGA scaffolds for bone marrow stimulation enhancement and repairing rabbit articular cartilage. METHODS: A rabbit model of articular cartilage defect was established for BMC-PGA stent implantation. After 8 weeks, the experimental animals were sacrificed. And the methods of hematoxylin and eosin stain, toluidine blue stain and immunohistochemistry were used to evaluate the effects of bone marrow stimulation enhancement and rabbit cartilage defect repairing. RESULTS: Visible new cartilage formation was evident after implantation. As compared with other groups, the repairing effect was better. CONCLUSION: The implantation of BMC-PGA scaffolds is both simple and effective in the repair of articular cartilage.

[Treatment of combination atlantoaxial fractures with posterior fusion plus atlantoaxial pedicle screw].

OBJECTIVE: To explore the clinical feasibility, approach, efficacy and indications of posterior fusion plus pedicle screw fixation in the treatment of combination atlantoaxial fractures. METHODS: A retrospective analysis was conducted for 26 cases of Jefferson and dens fracture treated with atlantoaxial pedicle screw from June 2008 to May 2010. RESULTS: The postoperative radiographs verified an excellent position of all screws with a satisfactory atlantoaxial reduction. Operative time was (126 ± 26) min, and blood loss was (350 ± 107) ml. During an average follow-up period of 14 months (range: 6 - 28), it showed no spine cord and vertebral artery injury or interfixation failure. Atlantoaxial alignment and stability were restored without any instrumentation-related complication. CONCLUSION: Posterior atlantoaxial pedicle screw and rod fixation may provide immediate three-dimensional rigid fixation of atlantoaxial joint. This technique is more effective than other previously reported approaches.

[Treatment of degenerative lumbar scoliosis via posterior decompression and fusion with internal fixation].

OBJECTIVE: To explore the pathogenic mechanism, operative techniques and therapeutic efficacy of lumbar degenerative scoliosis. METHODS: A retrospective analysis was performed for 32 patients (14 males and 18 females with a mean age of 67.4 years old) with degenerative lumbar scoliosis undergoing posterior decompression and fusion with pedicle screw system from January 2007 to March 2010. The post-operative outcomes were radiologically evaluated with Oswestry disability index (ODI), Cobb' angle and lumbar lordosis angle. RESULTS: All patients received a mean follow-up of 28.5 months (range: 12 - 50). All lumbar joints achieved bony fusion at Month 6 post-operation. No complications occurred due to instrumentation. There was no pseudoarthrosis. The significant differences of ODI existed between pre-operation and post-operation [(60 ± 11)% vs (21 ± 6)%, P < 0.01], Cobb' angle [(28 ± 9)° vs (13 ± 5)°, P < 0.01] and lumbar lordosis angle [(41 ± 12)° vs (20 ± 10)°, P < 0.01]. CONCLUSION: Individualized operations should be performed for degenerative lumbar scoliosis patients. Proper lumbar decompression and the reconstruction of coronal and sagittal balances may relieve symptoms and improve the quality of life.

[Pedicle screw implanting in the treatment of multi-level lumbar spine fractures].

OBJECTIVE: To review the clinical data of a group of patients with multi-level lumbar spine fractures treated with TSRH-3D pedicle screw system and to investigate the effect and post-operative efficacy of this technique. METHODS: Eighteen patients diagnosed as multi-level lumbar spine fracture were treated with TSRH-3D pedicle screw system. The operations were performed at 4 hours to 2 days after injury and the follow-up period was 15-32 months (mean: 23 months). The X-ray and CT scan were taken both preoperatively and postoperatively. RESULTS: (1) Twelve patients became injured from height falling (67%), 5 from traffic accident (28%) and 1 from crush (5%). (2) The improvement rate of spine motion, back pain and lower extremity pain was 67%, 94% and 78% respectively. (3) Compared with preoperative status, the height of anterior border of vertebra improved by 59.2%, lost by an average of 5.2% (P < 0.01) and the Cobb angle was corrected by 19.5 degrees , lost an average of 5.4 degrees . CONCLUSION: It is feasible to treat the multi-level lumbar spine fractures with TSRH-3D pedicle screw system. And the outcome of reduction and fixation is satisfactory. This technique is excellent at improving the height of injury vertebra, physiological curvature and neural function. It also can relieve lumbocrural pain with a definite postoperative efficacy.