Clinical effectiveness of treatment of combined upper thoracic spinal stenosis and multilevel cervical spinal stenosis with different posterior decompression surgeries.

OBJECTIVE: To explore the clinical effectiveness of various posterior decompression surgeries in the treatment of upper thoracic spinal stenosis combined with multilevel cervical spinal stenosis. METHODS: From January 2010 to December 2015, 22 consecutive patients with combined upper thoracic spinal stenosis and multilevel cervical spinal stenosis were treated with two different approaches of posterior decompression surgeries. In group A with 10 patients, both cervical and thoracic spinal decompression surgeries were performed simultaneously (one-stage surgery); in group B with 8 patients, cervical and thoracic spinal decompression surgeries were performed separately within three months (two-stage surgery). Based on Japanese Orthopedic Association (JOA) scores, improvement rate and extent of neurological function were calculated and the difference was compared between the two groups. RESULTS: There was no significant difference in demographic data between the two groups. However, compared with those of group B, both short-term and long-term improvement rate of neurological function in group A was higher (P < 0.05). In addition, the hospitalization cost was also lower in group A. CONCLUSION: Both one-stage and two-stage posterior decompression surgeries were effective in treating patient with upper thoracic spinal stenosis combined with multilevel cervical spinal stenosis; however, one-stage combined surgery was superior to two-stage surgery.

Raloxifene microsphere-embedded collagen/chitosan/ß-tricalcium phosphate scaffold for effective bone tissue engineering.

Engineering novel scaffolds that can mimic the functional extracellular matrix (ECM) would be a great achievement in bone tissue engineering. This paper reports the fabrication of novel collagen/chitosan/ß-tricalcium phosphate (CCTP) based tissue engineering scaffold. In order to improve the regeneration ability of scaffold, we have embedded raloxifene (RLX)-loaded PLGA microsphere in the CCTP scaffold. The average pore of scaffold was in the range of 150-200µm with ideal mechanical strength and swelling/degradation characteristics. The release rate of RLX from the microsphere (MS) embedded scaffold was gradual and controlled. Also a significantly enhanced cell proliferation was observed in RLX-MS exposed cell group suggesting that microsphere/scaffold could be an ideal biomaterial for bone tissue engineering. Specifically, RLX-MS showed a significantly higher Alizarin red staining indicating the higher mineralization capacity of this group. Furthermore, a high alkaline phosphatase (ALP) activity for RLX-MS exposed group after 15days incubation indicates the bone regeneration capacity of MC3T3-E1 cells. Overall, present study showed that RLX-loaded microsphere embedded scaffold has the promising potential for bone tissue engineering applications.

A simple LC-ESI-MS/MS method for quantification of bacopaside I in rat plasma and its application to a pharmacokinetic study.

Bacopaside I is one of the main pseudojujubogenin glycosides isolated from Bacopa monniera. In the present study, a rapid and robust LC-ESI-MS/MS method was developed and validated to quantify bacopaside I in rat plasma. After plasma samples were deproteinized by methanol, the post-treatment samples were analyzed on a Zorbax Eclipse Plus C18 (2.1×50mm, 1.8µm) column using a mobile phase of acetonitrile and water (65:35, v/v). Detection was performed on a triple-quadrupole tandem mass spectrometer with selected reaction monitoring (SRM) mode via electrospray ionization source. This method covered a linearity range of 10-2000ng/mL with the lower limit of quantification of 10ng/mL. The intra- and inter-day precisions of analysis were less than 10.2%, and the accuracies were between -11.1% and 8.4% at the concentrations of 25, 150 and 1800ng/mL. The total run time was 6.0min. This method was successfully applied to the preclinical pharmacokinetic study of bacopaside I following intravenous or oral administration to rats.

Tumstatin induces apoptosis and stimulates phosphorylation of p65NF-κB in human osteoblastic osteosarcoma Saos-2 cells.

The present study was aimed to investigate the effect of tumstatin on inhibition of proliferation and induction of apoptosis in Saos-2 human osteosarcoma cells and to understand the mechanism involved. Inhibition of cell proliferation was analyzed by MTT assay and induction of apoptosis through nuclear fragmentation assay. Viability of Saos-2 cells was reduced to 19% on treatment with 25 µM concentration of tumstatin after 48 h. Presence of characteristic apoptotic nuclei, rounded cell shape and shrunken size were caused by tumstatin treatment at 25 µM concentration. The level of mRNA corresponding to PTEN, FasR and FasL was increased significantly in tumstatin treated Saos-2 cells compared to untreated control. Investigation of the mechanism revealed NF-κB activation by phosphorylation on serine 536. The activated NF-κB was translocated into the nucleus from the cytoplasm on treatment with tumstatin. Degradation of the IκBα by tumstatin was found to be much slower compared to that induced by treatment with TNF-α. Thus, tumstatin inhibits proliferation and induces apoptosis in Saos-2 cells through activation of NF-κB and its translocation to the nucleus. Therefore, tumstatin can play an important role in the treatment of osteosarcoma.

Thymosin Beta-4 Recombinant Adeno-associated Virus Enhances Human Nucleus Pulposus Cell Proliferation and Reduces Cell Apoptosis and Senescence.

BACKGROUND: Thymosin beta-4 (TB-4) is considered key roles in tissue development, maintenance and pathological processes. The study aimed to prove TB-4 positive biological function on nucleus pulposus (NP) cell apoptosis and slowing the process of cell aging while increasing the cell proliferation. METHODS: TB-4 recombinant adeno-associated virus (AAV) was constructed and induced to human NP cells. Cell of same group were cultured without gene modification as controlled group. Proliferation capacity and cell apoptosis were observed during 6 passages of the cells. Morphology and expression of the TB-4 gene were documented as parameter of cell activity during cell passage. RESULTS: NP cells with TB-4 transfection has normal TB-4 expression and exocytosis. NP cells with TB-4 transfection performed significantly higher cell activity than that at the control group in each generation. TB-4 recombinant AAV-transfected human NP cells also show slower cell aging, lower cell apoptosis and higher cell proliferation than control group. CONCLUSIONS: TB-4 can prevent NP cell apoptosis, slow NP cell aging and promote NP cell proliferation. AAV transfection technique was able to highly and stably express TB-4 in human NP cells, which may provide a new pathway for innovation in the treatment of intervertebral disc degenerative diseases.

Wogonoside induces cell cycle arrest and mitochondrial mediated apoptosis by modulation of Bcl-2 and Bax in osteosarcoma cancer cells.

Osteosarcoma (OS) is the most common bone cancer with a great tendency for local invasion and distant metastasis. Restricted by the severe toxicity of conventional drugs, the therapeutic challenge of osteosarcoma still remains unconquered. The objective of the present research work was to investigate the antiproliferative activity of wogonoside against human osteosarcoma (SaOS-2) cell line. Cell viability after wogonoside treatment was evaluated by MTT assay. Phase contrast microscopy was used to evaluate the change in cell morphology following drug treatment. The effect of wogonoside on cell cycle phase distribution and mitochondrial membrane potential was investigated by flow cytometry using propidium iodide (PI) and rhodamine-123 DNA-binding fluorescent dyes respectively. Western blotting was used to evaluate the effect of wogonoside on cell cycle-related proteins as well as on the expression levels of Bcl-2, Bax, cytosolic and mitochondrial cytochrome c and apoptotic protease activating factor-1 (Apaf-1). Wogonoside induced a dose-dependent as well as time-dependent growth inhibitory effects on cell proliferation of SaOS-2 cancer cells. Wogonoside induced G2/M cell cycle arrest as well as loss in mitochondrial membrane potential in these cells. Within 48 h of incubation, approximately 4.36%, 6.72%, 11.54%, 21.88% and 15.54% of the cells underwent early apoptosis after treatment with 0, 5, 10, 25 and 75 µM of wogonoside respectively. Wogonoside led to reduced Bcl-2 expression and increased Bax expression, while as it led to s decrease in the levels of mitochondrial cytochrome c and an increase in cytosolic fraction and expressions of cytosolic apoptotic protease activating factor-1 (Apaf-1).

Ultrasound assessment of injury to the posterior ligamentous complex in patients with mild thoracolumbar fractures.

OBJECTIVE: To assess the value of ultrasonographic imaging of the posterior ligamentous complex (PLC) to diagnose ligamentous injuries, in patients with mild thoracolumbar fractures. METHODS: Patients with thoracolumbar fractures were included in this prospective study. Patients underwent palpation of the midline of the back, and ultrasonography was performed over the entire thoracolumbar region by an experienced sonographer. A team that included a musculoskeletal radiologist, an orthopaedic surgeon and a sonographer assessed the ultrasound results. Ultrasonographic and magnetic resonance imaging (MRI) findings were jointly evaluated in a subgroup of patients who were able to fund MRI analysis. Conflicts regarding the results were resolved by a majority vote. RESULTS: A total of 21 patients participated in the study, all of whom exhibited abnormal ultrasonographic echogenicity on the supraspinous or interspinous ligaments. Three patients were diagnosed with a rupture of the supraspinous ligament. In 15/17 (88.2%) patients, interspinous ligament injuries were detected caudally to the injured vertebrae. CONCLUSIONS: Ultrasound examination is a reliable complementary diagnostic tool to identify PLC injuries in patients with mild thoracolumbar fractures.

[Assessment of correction effect in relation to fulcrum bending flexibility in adult idiopathic scoliosis patients].

OBJECTIVE: To study the relationship between the fulcrum bending flexibility (FBF) and correction rate (CR) of adult idiopathic scoliosis, and to explore the ability of FBF to assess the correction effect in relation to fulcrum bending flexibility. METHODS: 69 patients with adult idiopathic scoliosis with structural curves at thoracic or lumbarthoracic segments, 16 males and 53 females, aged 26.5 (19 - 53) were treated by pedicle screws instrumentation. Pre-operative standing and fulcrum bending films and postoperative standing X-ray film were taken. Cobb angle was measured. The data underwent regression analysis with the software SPSS 10.0. RESULTS: A regression formula was established: CR = 0.213 + 0.768 x FBF with P < 0.01. CONCLUSION: A definite linear relation exists between the FBF and CR of adult idiopathic scoliosis. By using the formula in proper samples, the effects of new instrumentation or correction technique can be objectively assessed.

[Association study of SIM2 gene polymorphisms with susceptibility to congenital scoliosis in a Chinese Han population].

OBJECTIVE: To investigate whether polymorphisms of SIM2 gene are associated with congenital scoliosis (CS) in a Chinese Han population. and explore the relationship of between polymorphisms of SIM2 and clinical phenotypes of CS. METHODS: A case-control design was employed in this study. A total of 127 patients (55 boys, 72 girls, mean age 12.90 y/o) diagnosed with CS admitted at Peking Union Medical College (PUMC) Hospital were enrolled between October 2005 and September 2007. The scoliosis-free control subjects (127 cases) at the same hospital during the same study period were frequency-matched to the cases on age (+/- 3 years) and gender. Genomic DNA was extracted by QIAamp DNA Blood Mini Kit from peripheral blood leukocytes of each subject who had signed informed consent. Based on genotype data from the International HapMap project, the main functional single nucleotide polymorphisms (SNPs) initially were selected. Case group were classified into different clinical phenotypes according to vertebral defect type, location of deformity, extent of developmental disruption, combined rib malformations and neural canal deformity. Genotying of all selected SNPs was done by SNPstream technology (Beckman Coulter SNPstream). All the data of SNPs with polymorphism were analyzed by association analysis based on a single SNP, the association analysis between phenotypes and SNPs. And pairwise linkage disequilibrium was calculated in the control population using Haploview 4.1 software. RESULTS: SNP1 (rs2073601), SNP2 (rs2073417) and SNP3 (rs2051397) of SIM2 are genotyped. SNP2 and SNP3 in linkage disequilibrium. No association (P > 0.05) is observed between SNP1, SNP2 and SNP3genotypes/allele polymorphisms and risk of CS and different clinical phenotypes. CONCLUSION: Genetic variants of SIM2 gene may not be associated with the susceptibility to CS and different clinical phenotypes of CS in Chinese Han population.

[Association of PAX1 gene polymorphisms with susceptibility to congenital scoliosis in Chinese Han population].

OBJECTIVE: To investigate the association of polymorphisms of PAX1 gene with congenital scoliosis (CS) in Chinese Han population and the relationship between the PAX1 gene polymorphisms and the clinical phenotypes of CS. METHODS: Peripheral blood samples were collected from 127 CS patients, 55 male and 72 female, aged (12.9 +/- 4.3) (2 - 23), and 127 sex- and age-matched controls. Based on genotype data from the International HapMap project, the tagging single nucleotide polymorphisms (tSNPs) were selected using Haploview 4.0 software. Hardy-Weinberg equilibrium was analyzed both in the control and case groups. The case group was classified into different clinical phenotypes according to vertebral defect type, location of deformity, extent of developmental disruption, combined rib malformations, and neural canal deformity. Genotyping of all selected SNPs was done by SNPstream technology. The association between phenotypes and SNP was analyzed. Pairwise linkage disequilibrium was calculated in the control population using Haploview 4.0 software. RESULTS: The sites: SNP1 (rs17861031) and SNP2 (rs6047590), of PAX1 gene were genotyped and both polymorphisms were distributed in line with the Hardy-Weinberg equilibrium in these 2 groups. There was no linkage disequilibrium between these 2 SNPs. The genotype frequencies of SNP1AA, SNP1AG, SNP1GG, SNP2AA, SNP2AT, and SNP2TT of the case group were 2%, 26%, 72%, 2%, 19%, and 80% respectively, all not significantly different from those of the control group (2%, 26%, 72%, 2%, 26%, and 82% respectively, all P > 0.05). The allele frequencies of SNP1A, SNP1G, SNP2A, and SNP2T of the case group were 15%, 85%, 11%, and 89% respectively, all not significantly different from those of the control group (15%, 85%, 10%, and 90% respectively, all P > 0.05). No positive sites were found in different clinical phenotypes. CONCLUSION: The genetic variants of PAX1 gene may not be associated with the susceptibility to CS and different clinical phenotypes of CS in Chinese Han population.

[Assessment of curve flexibility in adolescent idiopathic scoliosis before operation].

OBJECTIVE: To compare the effects in assessing the curve flexibility of the adolescent idiopathic scoliosis (AIS) and predicting the outcomes of operation among different radiological techniques: supine lateral bending (SB), traction (Tr), and fulcrum bending radiographs. METHODS: 68 consecutive AIS patients, all with the single-curve types Ia/Ib/Ic according to the PUMC classification, divided into 4 groups according to the magnitude of Cobb's angle: moderate thoracic curve (n = 19, 40 degrees < Cobb's angle < or = 60 degrees ), severe thoracic curve (n = 13, Cobb's angle > 60 degrees ), moderate lumbar curve (n = 28, 35 degrees < Cobb's angle < or = 60 degrees ), and severe lumbar curve(n = 8, Cobb's angle > 60 degrees ) who were treated surgically underwent preoperative radiological evaluation including standing anteroposterior and lateral Tr, SB, and fulcrum bending radiographs. COBB angle was measured and the flexibility ratio was determined on each radiograph. The amounts of correction obtained by all radiographic methods were compared with the amount of surgical correction. RESULTS: The post-operative Cobb's angle of the moderate thoracic curve group was 9 degrees , not significantly different from that by fulcrum bending radiograph (P = 0.076), but significantly different from those by the other methods (both P < 0.01). The post-operative COBB angle of the severe thoracic curve group was 40 degrees , significantly different from all the radiographs before operation (all P < 0.01). The post-operative Cobb's angle of the moderate lumbar curve group was 4 degrees , significantly different from those by fulcrum bending and Tr radiographs (both P < 0.01) and that by SB (P = 0.013). The post-operative Cobb's angle of the severe lumbar curve group was 24 degrees , significantly different from those of anteroposterior and Tr radiograph (both P < 0.01) and those of fulcrum-bending and SB radiographs (P = 0.021 and P = 0.011). In the moderate thoracic curve group the operation correction rate was not significantly different from the flexibility rate by fulcrum-bending radiograph (P = 0.111), and was significantly different from the flexibility rates by SB and Tr radiographs (P = 0.011 and P = 0.000). In the severe thoracic curve group the operation correction rate was significantly different from the flexibility rates by different kinds of radiograph (all P = 0.111). In the moderate lumbar curve group the operation correction rate was significantly different from the flexibility rates by different kinds of radiograph (P < 0.111 or P = 0.019). In the severe lumbar curve group the operation correction rate was significantly different from the flexibility rates by different kinds of radiograph (P < 0.01 or P = 0.017). CONCLUSION: Fulcrum-bending radiography can better assess the flexibility and correction rate of thoracic curves in AIS, however, it can only predict those in moderate thoracic curves. Fulcrum-bending radiograph and SB radiograph are similar in predicting the flexibility in lumbar curves.