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Objective To investigate the osteogenic differentiation potency of ligament cells in thoracic ossification of the ligamentum flavum (TOLF) and analyze further by using transcriptome high-throughput sequencing.Methods Clinically,the patients with non-TOLF and TOLF (n=10 in each group) who underwent surgery in our hospital from October 2015 to April 2016 were included in this study.The primary ligament cells that derived from the two groups were separately cultured and induced osteogenesis with 15％ strength of cyclic mechanical stress for 12h and 24h using a device called Flexcell FX-4000.The ALP activity was determined to evaluate the osteogenesis using quantitative analysis and ALP staining assay.Real-time PCR and westernblotting were used to detect the mRNA and protein expression of osteogenic-related genes including ALP,BMP-2 and Osteocalcin.Then,three patients in each group were included in the study of transcriptome high-throughput sequencing and bioinformatics analysis using Illumina HiSeqTM 2500 sequencing platform to compare further.Results The morphology of the cells that derived from two groups was basically similar,all presented an elongate spindle-shape.To evaluate the ostogenesis,ALP activity assays including quantitative and staining assays were performed.Under microscope,the ALP staining in the TOLF group was higher than non-TOLF group and increased with the longer duration of stress induction.The result of semi-quantitative analysis showed the stained area and positive cells in TOLF group were more than non-TOLF group significantly at 0 h,and were increased with the induction.The results of quantitative analysis showed ALP activity in the TOLF group was significantly higher than non-TOLF group and were increased with the induction significantly all the time.But no significant change in ALP staining or quantitative analysis was found in non-TOLF.The results of real-time PCR indicated that the expression of ostegenic markers above in the TOLF group was more than non-TOLF group significantly except the expression of OCN at 0 h.The expression of the three ostegenic markers in TOLF group was increased with the stress induction for 12 h and 24 h significantly except the expression of BMP-2 and OCN at 12 h.The results of western-blotting indicated that the expression of the three ostegenic markers above in the TOLF group was more than non-TOLF group significantly except the expression of ALP at 0 h.The expression of the three ostegenic markers in TOLF group was increased with the stress induction for 12 h and 24 h,but only the expression of ALP at 24 h was significant.And no significant change in the expression of mRNA and protein was found in non-TOLF group.In the transcriptome analysis,671 genes of TOLF group were up-regulated and 314 genes were found to be down-regulated compared to the control group.In addition,22 significant GO terms associated with upregulated genes were found to be closely related to ossification.Conclusion TOLF ligament cells have high osteogenic differentiation potency,which could express obvious osteogenesis-related gene spectrum,and differentially expressed genes including L1RL1 、PTHLH、DKK1 、BMP6、SPP1 and FGF1 may be related with the osteogenic potency of ligament cells in thoracic ossification.
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Objective To employ intraoperative discography to determine the injured intervertebral disc segments that can not be identified on the preoperative MRI in patients with cervical spinal cord injury without fracture and dislocation for confirming the responsible segments needing surgical decompression and fusion.Methods The study involved 85 patients with cervical spinal cord injury without fracture and dislocation treated from January 2007 to December 2011,among which sixteen patients had not been identified with the responsible segments by preoperative MRI.The average preoperative Japanese Orthopedic Association (JOA) score was (9.1 ± 1.8) points.There was no obvious fracture or dislocation of the cervical spine on preoperative X-ray film,CT and MRI,but all patients displayed high intense signal in cervical spinal cord on MRI T2 weighted imaging.Besides,MRI revealed hemorrhagic swelling of anterior cervical soft tissue in nine patients and cervical intervertebral disk hernia in all patients.Annulus fibrosus rupture of cervical intervertebral disc with contrast leakage in intraoperative discography of suspected injury segments in all patients under direction of C-arm X-ray machine was set as the injury criterion.The patients with pure ruptured discs received cervical discectomy,interbody fusion and titanium plate fixation.The patients associated with multilevel cervical intervertebral disc hernia or ossification of posterior longitudinal ligament underwent anterior cervical corpectomy,bone graft with titanium cageand titanium plate fixation of ruptured discs.Results Nineteen injured discs were identified eventually by discography,including 2 discs at C3/4,4 at C4/5,8 at C5/6 and 5 at C6/7.Moreover,anterior annulus fibrosus rupture with intact anterior longitudinal ligament was found in 11 patients.The follow-up lasted for (24.4 ± 10.0) months.JOA scores were (13.3 ± 1.5) points and (14.5 ± 1.6) points at two weeks and three months after operation,and (15.1 ± 1.5) points at the last follow-up,indicating a relevant improvement rate of 53％,68％ and 76％ respectively.Mean operation time was 110 minutes and blood loss was 120 ml.Three patients had pain on shoulder and back and one patient had hoarse voice,but all the patients were relieved in two weeks after conservative treatments.No serious complications,such as deep infection,deterioration of neurological dysfunction,vertebral artery injury or internal fixation failure were noticed intra-or post-operatively.Conclusion For the intradiscal rupture that is hard to be determined by the conventional imaging methods,intraoperative discography can be used as an auxiliary method of imaging diagnosis in early surgical determination of responsible segments for cervical spinal cord injury without fracture and dislocation.
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Objective To evaluate the effect of en bloc open-door laminectomy and pedicle screw fixation for extremely severe cervical ossification of posterior longitudinal ligament (OPLL).Methods From January 2007 to December 2011,16 cases of serious cervical OPLL were treated,in which there were 13 males and 3 females,aged from 45 to 74 years (average,56.5 years).Spinal cord functional deterioration was related to minor activities of the neck,such as sneeze,cough or hyperextension of the neck,in 15 cases.One case suffered from aggravating neurological symptoms without a definite precipitating factor.The average preoperative Japanese Orthopaedic Score (JOA) was 7.1 ±1.8.Preoperative CT scans displayed the average stenotic rate (thickness of OPLL/sagittal diameter of the spinal canal) was 83.7％.All the patients received en bloc laminectomy and pedicle screw fixation of the cervical spine.Results All the patients were followed up for 3 months to 4 years (average,24.4±10 months).The JOA scores at 2 weeks,3 months,and the last follow-up postoperatively were 13.2±1.7,13.5 ±1.6 and 14.1 ±1.5,respectively; the improvement rates were 61.6％,64.6％ and 70.7％,respectively.The operative time ranged from 80 to 150 minutes (average,130 minutes),and blood loss ranged from 150 to 600 ml (range,300 ml).One case of postoperative wound hematoma,1 case of cerebrospinal fluid leakage and 3 cases of trapezius muscle pain occurred in this group.No serious complications,such as deep infection,deterioration of neurological dysfunction,vertebral artery injury and internal fixation failure,were noticed.Conclusion Owing to the minimum interference to the cervical spinal cord,stabilization of the decompressed segments and maintenance of cervical lordosis,en bloc open-door laminectomy and pedicle screw fixation is suitable for extremely severe cervical OPLL.
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Objective To study the onset,treatment,prognosis and possible causes of transient quadriplegia shortly after anterior cervical compression and fixation (within four hours postoperatively) in three patients with cervical spondylotic myelopathy who could function well for limbs after anesthesia awakening from the operation. Methods A retrospective study was carried out on medical data of three patients including two males and one females,at age of 41-61 years.Anterior cervical corpectomy,titanium mesh bone fusion and titanic plate fixation were performed under general anesthesia.The decompression segment was C5 in one patient,C6 in one and C5 plus C6/7intervertebral disc in one respectively.Paralysis occurred between 30 minutes and 4 hours postoperatively.Two patients were with complete paralysis and one with incomplete. All the patients received dehydration,neurotrophic drugs and high-dose methylprednisolone therapy immediately after paralysis. Meanwhile,emergent cervical MRI was performed,which showed spinal cord swelling,without obvious spinal cord compression by hematoma.Results The paralysis was alleviated completely in two patients within two hours after early medication without additional surgical compression.The other one patient was recovered incompletely at 24 hours after medicationand then underwent posterior cervical laminoplasty,when tremendous pressure was released from cervical spinal cord.But the spinal function had significant recovery after surgical compression and won complete recovery one week later. Conclusions Besides spinal cord ischemia-reperfusion injury,the transient paralysis after anterior cervical surgery may be associated with cervical spinal cord swelling and limited anterior decompression space.Early diagnosis and early intervention of paralysis may save the spinal cord function and attain a satisfactory prognosis.
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Objective To evaluate the mid-term clinical and radiographic results of artificial disc replacement (ADR) for discogenic low back pain.Methods From July 2004 to July 2007,21 patients with discogenic low back pain,aged from 26 to 67 years,underwent lumbar ADR with the Charité Ⅲ artificial disc at 22 levels and Activ L prosthesis at 2 levels:L4-5 5 cases,L5S1 13 cases,L4-5 and L5S1 3 cases.The diagnosis was proved by discography in all the patients.The clinical and radiographic results were evaluated and compared between pre-,post-operation and finial follow-up.Results All patients were followed up for 4to 7 years (average,5.1 years).There were statistical differences between pre-operation and final follow-up in Oswestry disability index and Visual Analogue Scale for back pain and leg pain.All the prostheses were mobile without dislocation,breakage,subsidence or spontaneous fusion,only a slight scoliosis was noted in one patient.At the ADR levels,the anterior and posterior intervertebral height increased averagely 6.3 mm and 1.9 mm respectively,the lordosis increased 2.9°,and the mean range of motion (ROM) was 4.6°.At the adjacent level proximal to ADR,the intervertebral height and lordosis decreased slightly,and the ROM decreased 2.5°.The total lumbar lordosis increased 7.8°,and the total lumbar ROM increased 2.4°.According to MRI and CT scans,the degeneration of proximal adjacent disc and facet were not evident,however the degeneration of facets at the ADR levels was evident.Conclusion The 5 years results of ADR for discogenic low back pain were satisfactory,with preserved motion at the ADR level,and the degeneration of adjacent level was not evident.However,there was obvious degeneration in facet joints of the ADR level.
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ObjectiveTo observe in vivo segmental lumbar motion in patients with lumbar disc herniation (LDH) during functional weight-bearing activities.MethodsFifteen patients with LDH at L4-5 were studied as experimental group.Ten healthy volunteers were recruited as control group.Three-dimension(3D) lumbar spine models of L3,L4 and L5 were reconstructed from thin section CT scans.Spine motions were then reproduced by matching lumbar spine models and images got from dual fluoroscopic imaging system (DFIS)under different motion state (standing,flexion-extension,left-right twisting and left-right bending).From local coordinate systems at the end plates,the motion of the cephalad vertebrae relative to the caudal vertebrae was calculated for vertebrae levels:L4-5 and L3-4.ResultsThe motion pattern at L4-5 was found to be altered.During flexion-extension,the migrations of the affected segments along the frontal axis,sagittal axis,vertical axis were similar with that of the control group,but the rotation angle along the frontal axis was significantly larger than that of the control group (P＜0.05).During left-right bending and left-right twisting,the migration and rotation angle along the frontal axis were significantly larger than those of control group.During flexionextension,the migrations of the neighboring segments (L3-4) along the three axes were larger than those of the control group,but there were no statistical significances.During left-right bending and left-right twisting,the migrations of the neighboring segments (L3-4) along the vertical axis were significantly larger than those of the control group (P＜0.05).ConclusionThe 3D lumbar motion pattern in LDH patient is different with that of normal people.For the affected segment,compared with the normal people,the range of flexion-extension motion and the translocation in left-right direction were significantly larger,but the rotation range along the vertical axis was smaller.
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Objective To investigate the effects of different cyclic tensile strain on the proliferation of human anulus fibrosus cells from degenerated discs.Methods Anulus fibrosus(AF) cells were isolated from a degenerated human IVD,expanded in monolayer,and cyclically strained for 3 hours,applying 0,5％,10％,15％ and 20％ strains at a frequency of 0.25 Hz with the use of the DioDynamic test instrument.The flow cytometry method was used to examine the Af cells proliferation at 24 hours following application of the cyclic tensile strains.Results The proliferative index (PI) increased with the magnitude value of cyclic tensile strain except 20％ group.The most significant increase of proliferation index were found in 15％ group.Conclusion There might be some corelationships between magnitude of cyclic tensile strain and the proliferation of the degenerative AF cells.