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Objective To observe the effects of acetyl-l-carnitine (ALC) on autophagy, apoptosis and motor function after acute spinal cord injury (ASCI) in rats. Methods Thirty-six adult female Sprague-Dawley rats were randomly divided into sham operation group (Sham group, n=12), simple spinal cord injury group (SCI group, n=12), ALC treatment group (ALC group, n=12). Spinal cord injury model at the level of T10 segment was established using Allen's method. They were assessed with Basso-Beattle-Bresnahan (BBB) scale three days after injury. Then the rats were sacrificed, and the expression of microtubule associated protein 1 light chain 3 (LC3)-II in spinal cord was detect-ed with Western blotting and immunofluorescent labeling, and the number of apoptotic cells were assessed with TUNEL staining. Results The expression of LC3-II and the number of apoptotic cells increased in SCI group compared with those in Sham group (P<0.01), while the BBB score decreased (P<0.001). The expression of LC3-II increased and the number of apoptotic cells decreased in ALC group compared with those in SCI group (P<0.001), while the BBB score increased (P<0.01). Conclusion ALC may promote autophagy, and inhibit apopto-sis to improve the locomotor function after ASCI.
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Objective To investigate the effects of hydrogen sulfide on autophagy and the apoptosis after acute spinal cord injury in rats. Methods Thirty-six adult male SD rats (250-300 g) were randomly divided into three groups (n=12 for each group):sham operation group (Sham group), spinal cord injury group (Model group) and hydrogen sulfide pre-treatment group (H2S group). Allen’s method was used to establish the rat model of spinal cord injury. Rats of sham operation group re?ceived only laminectomy. Rats of H2S group received sodium hydrosulphide injection intraperitoneally (50μmol/kg) 1h after spinal cord injury, and Model group was given the same amount of saline solution. Rats in the three groups were sacrificed 24 h after spinal cord injury, then the spinal cord was removed. The expressions of LC3, p70S6K and Cleaved caspase-3 were detected by Western blot assay. The expression of LC3 was also detected by immunofluorescence. The cell apoptosis was as?sessed by TUNEL stain. Results Compared with Sham group, the expression levels of LC3Ⅱ/LC3Ⅰand Cleaved caspase-3 were increased in Model group, but the expression of p70S6K decreased and cell apoptosis increased in Model group (P<0.01). Compared with Model group, the expression levels of LC3Ⅱ/LC3Ⅰand Cleaved caspase-3 were decreased significant?ly, while the expression of p70S6K increased and cell apoptosis decreased significantly in H2S group (P < 0.01). Conclu?sion Hydrogen sulfide can inhibit autophagy and reduce cell apoptosis after acute spinal cord injury in rats.
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Objective To investigate the effects of edaravone (EDA) on cell apoptosis induced by endoplasmic reticu?lum stress (ESR) after spinal cord injury (SCI) in rats. Methods Thirty-six healthy adult SD rats were randomly divided in?to three groups (12 rats for each group):Sham group, SCI group and EDA group. The rat model of SCI was made by Allen’s method and the sham group was only received laminectomy and kept the spinal cord intact. Rats in sham group and SCI group accepted the same volume and frequency of saline injection as EDA group. The EDA group was given 10 mg/kg EDA once every 12 h intraperitoneally. Three days after injuring, the spinal cords were harvested, and the protein levels of C/EBP homologous protein (CHOP), Cleaved caspase-12 and Cleaved caspase-3 were detected by Western blot assay. Immunofluo?rescence staining was used to analyze the positive ratio of caspase-12 and CHOP in spinal cord of three groups. Meanwhile, TUNEL staining was used to identify cell apoptosis of spinal cord. Results Compared with sham group, the protein levels of CHOP, Cleaved caspase-12 and Cleaved caspase-3 were obviously higher in SCI group (P<0.01);the proportion of Cas?pase-12 and CHOP positive cells was significantly increased (P<0.01), and the apoptotic rates were also significantly in?creased in spinal cord (P<0.01). However, compared with SCI group, the protein levels of CHOP , Cleaved caspase-12 and Cleaved caspase-3 were significantly decreased in EDA group (P<0.01);the proportion of Caspase-12 and CHOP positive cells was significantly reduced (P<0.01), and the apoptotic rates were also significantly decreased in spinal cord (P<0.01). Conclusion EDA has neuroprotective potential to spinal cord injury. The mechanism of its neuroprotective effect may asso?ciate with its inhibitory effect to the cell apoptosis induced by endoplasmic reticulum stress after SCI.
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Objective To study the effect of Exendin-4 on oxidative stress and neural apoptosis following spinal cord injury (SCI). Methods Adult male SD rats, with weight between 200-250 g, were randomly divided into three groups (12 in each group):Sham group, SCI group and Exendin-4 group (Ex-4 group). Rats in Sham group achieved spinal cord exposure. SCI group and Ex-4 group were induced according to Allen′s test (using a weight-drop device). Rats in Ex-4 group were ad?ministrated with Exendin-4 (10 μg/rat) through intraperitoneal injection immediately after establishment of SCI models. Rats in Sham group and SCI group were given the same volume of normal saline solution instead. Level of malondialdehyde (MDA) and the activity of catalase (CAT) were assessed in spinal cord tissues 24 hour after drug administrations. Neural apoptosis was detected by TUNEL staining and the expression levels of caspase-9 and AIF were determined using Western blot. Results Compared with Sham group, the levels of MDA, caspase-9 and AIF as well as neuronal apoptosis rate in?creased obviously, while activity of CAT decreased markedly in SCI group(P<0.01). Compared with SCI group, the levels of MDA, caspase-9 and AIF as well as the neuronal apoptosis rate decreased obviously, while activity of CAT increased re?markably in Exendin-4 group(P < 0.01). Conclusion Exendin-4 restrain neural apoptosis following spinal cord injury through relieving oxidative damage.
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Objective To investigate the protective effect of Ebselen on mitochondrial damage and its influence to Cytochrome C expression and the neuronal apoptosis after spinal cord injury in rats. Methods Sixty adult SD rats were ran-domly divided into 5 groups (12 each group). Spinal cord injury model was made using Allen's method. Sham operation group received only laminectomy;SCI group received laminectomy and spinal trauma;Saline group received saline injection intraperitoneally (0.1%DMSO) after injury;methylprednisolone group received 30 mg/kg methylprednisolone injection intra-peritoneally, ebselen group received 10 mg/kg ebselen injection intraperitoneally. The malonaldehyde (MDA) and glutathi-one (GSH)level at the injured sites of the spinal cord were detected 24 hours after trauma, and the expression level of Cyto-chrome C was also observed. Finally, neuronal apoptosis was identified by TUNEL staining. Results MDA level in the Eb-selen group was significantly lower than that in the SCI group, and GSH level was significantly elevated in the Ebselen group compared with SCI group (P<0.01). Expression of Cytochrome C in Ebselen group was lower than that in SCI group shown by Western blot, and the neuronal apoptosis in Ebselen group reduced significantly too compared with SCI group (P<0.01). Conclusion Ebselen can alleviate peroxidation,prohibit expression of Cytochrome C and inhibit neuronal apoptosis,thus it shows a protective effect to experimental acute SCI.
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Objective To detect the effects of the selective mitochondrial fission inhibitor-Mdivi-1 on the malondi-alolehyde (MDA), glutathione (GSH) as well as cytochrome C (Cyt-C) in neuronal mitochondria and neuronal apoptosis. Methods Thirty-six adult female SD rats (250-300 g) were randomly divided into 3 groups (n=12):sham operation (Sham) group, single spinal cord injury (SCI) group and Mdivi-1 pretreatment (1.20 mg/kg, Mdivi-1) group. In sham group, the rats’ spinal cord was exposed, but no hit. The rat model of spinal cord injury was established by Allen’s method in SCI group and Mdivi-1 group. In Mdivi-1 group, rats were given Mdivi-1 through the tail vein 15 min before spinal cord injury, and SCI group received the same amount of dimethyl sulfoxide (DMSO). Rats in Sham group were sacrificed 8 h after exposing spinal cord. Rats in SCI group and Mdivi-1 group were sacrificed at 8 h after the spinal cord injury, then were removed the spinal cord T9-11. The contents of MDA and GSH in mitochondria of spinal cord tissues were detected with spectrophotometer. The expressions of Cyt-C protein in the mitochondria and cytoplasm were detected by Western blot assay. The neuronal apoptosis was assessed by TUNEL staining. Results Compared with Sham group, levels of Cyt-C and GSH in mitochondria were decreased significantly (P<0.01), while levels of MDA in mitochondria, Cyt-C in cytoplasm and the neuronal apopto-sis were increased significantly in SCI group (P<0.01). Compared with SCI group, Cyt-C and GSH levels in mitochondria were increased significantly in Mdivi-1 group (P<0.01), however, MDA in mitochondria,Cyt-C in cytoplasm and the neuro-nal apoptosis were significantly reduced (P<0.01). Conclusion Mdivi-1 can relieve neurons from mitochondrial oxidative damage, inhibit the release of cytochrome C and neuronal apoptosis after acute spinal cord injury, which plays a role in pro-moting the recovery of spinal cord function.
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This paper is aimed to investigate the repair of rabbit radial bone defect by the recombinant human bone morphogenetic protein 2/poly-lactideco-glycolic acid microsphere with fibrin sealant (rhBMP-2/PLGA/FS). The radial bone defect models were prepared using New Zealand white rabbits, which were randomly divided into 3 groups, experiment group which were injected with eMP-2/PLGA/FS at bone defect location, control group which were injected with FS at bone defect location, and blank control group without treatment. The ability of repairing bone defect was evaluated with X-ray radiograph. Bone mineral density in the defect regions was analysed using the level of ossification. The osteogenetic ability of repairing bone defect, the degradation of the material, the morphologic change and the bone formation were assessed by HE staining and Masson staining. The result showed that rhBMP-2/PLGA/FS had overwhelming superiority in the osteogenetic ability and quality of bone defect over the control group, and it could promote the repair of bone defect and could especially repair the radial bone defect of rabbit well. It may be a promising and efficient synthetic bone graft.
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Animals , Female , Male , Rabbits , Bone Morphogenetic Protein 2 , Therapeutic Uses , Bone Regeneration , Bone Substitutes , Therapeutic Uses , Fibrin Tissue Adhesive , Therapeutic Uses , Lactic Acid , Therapeutic Uses , Microspheres , Polyglycolic Acid , Therapeutic Uses , Radius Fractures , Therapeutics , Recombinant Proteins , Therapeutic Uses , Transforming Growth Factor beta , Therapeutic UsesABSTRACT
BACKGROUND: Traditional methods of repairing bone defect such as autograft and allograft have some disadvantages that are hard to deal with, gene treatment may be a new approach. OBJECTIVE: To investigate the biological properties of cultured human osteoblasts transfected with vascular endothelial growth factor (VEGF) gene. DESIGN, TIME AND SETTING: A controlled experiment based on cytology was carried out in the Scientific Experiment Centre of Liaoning Medical College from May 2005 to May 2006. MATERIALS: Human lilac born block was harvested from a cervical spondylosis patient who required lilac bone graft with his informed consent of this patient. Plasmid pCDI/VEGF_(121) was given as gift from Professor Ma, Peking Unviersity Human Disease Genomics Research Center. Competent Escherichia coU was given as gift from Professor Liu, Liaoning Medical College. METHODS: Human osteoblasts were isolated and cultured in vitro. There were a VEGF transfection group and a control group in the experiement. Using cation liposome, the pCDINEGF_(121) eukaryotic expression plasmis was induced into human osteoblasts.MAIN OUTCOME MEASURES: At 1, 3, 5, 7 days following passage culture, the expression of VEGF in human osteoblasts was detected. Its effects on the call proliferation, the secretion of osteocalcin and alkaline phosphatase were investigated.RESULTS: After the plasmid pCDI-VEGF_(121) was transferred into human osteoblasts 3 and 7 days, VEGF mRNA expression was detectable by RT-CPR method. The call number of transfection group was larger than that of control group (P < 0.05 or P < 0.01).When the cells were cultured for 3 days, the positive rate of alkaline phosphatase in the transfection group was increased compared with control group (P < 0.01 ); the secretion of osteocalcin in the transfection group was higher than that of control group (P < 0.05 or P < 0.01 ).CONCLUSION: VEGF gene transfection can improve the proliferation and biological function of human osteoblasts cultured in vitro.
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[Objective]To explore the technique and effect of anterior cervical decompression for the treatment of the mixed type of ossification of posterior longitudinal ligament. [Methods]Data on 37 patients(24 males and 13 females,with mean age of 54.3 years) who underwent resection or floating of posterior longitudinal ligament were reviewed.The occupying rate of OPLL ranged 32%~85% with an average of 51.4%.The Japanese Orthopaedic Association(JOA) scores ranged 4 ~14 points with an average of 7.9 points before operation.[Results]The Mean follow-up duration was 16 months(range,6~36 months).The JOA scores were 10.3 and 11.1 at 3 and 12 months after surgery.The results were excellent in 72.7% and good in 78.8%.[Conclusion]The resection of the longitude ligament and floating in anterior cervical decompression is a safe and effective treatment for the mixed type of ossification of posterior longitudinal ligament.