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Objective: To investigate the effect of the non-PLC-dependent protein kinase C (PKC) pathway of parathyroid hormone (PTH) on the apoptosis and proliferation of osteoblast MC-3T3E1 cells., Methods: MC-3T3E1 cells were seeded in 96-well plates at the density of 1.5×10(4) cells/mL and incubated for 3 day. The cells were then exposed to 100 nmol/L of [Gly(1), Arg(19)]hPTH(1-28), 100 nmol/L of [Gly(1), Arg(19)]hPTH(1-34), 100 nmol/L of [Gly(1), Arg(19)]hPTH(1-34)+1 µmol/L Go6983, 1 µmol/L Go6983, or deionized water (control) for 1, 24 or 48 h. After the treatments, cell counting kit-8 (CCK-8) and Caspase-Glo® 3/7 Assay (Caspase-3) were used to examine the proliferation and apoptosis of MC3T3-E1 cells., Results: CCK-8 results showed that hPTH(1-34) increased the number of MC3T3-E1 cells compared with hPTH(1-34)+Go6983 at 1 h and 24 h, but this difference was not statistically different. At 48 h, treatment with hPTH(1-34), as compared with hPTH(1-28), significantly increased the number of MC3T3-E1 cells (P<0.05), and this effect was blocked by the PKC inhibitor Go6983 (P<0.05). hPTH(1-34) did not result in significant inhibition of MC3T3-E1 cell apoptosis at 1 h and 24 h as compared with hPTH(1-34)+Go6983, but significantly inhibited the cell apoptosis as compared with hPTH(1-28) (P<0.05); this inhibitory effect was blocked by Go6983 (P<0.05)., Conclusion: s A relatively long time (for 48 h) of exposure to PTH can inhibit apoptosis and promote the proliferation of MC3T3-E1cells through a non-PLC-dependent PKC pathway.

Objective: To evaluate the efficacy of conservative treatment with teriparatide for promoting bone fracture healing in patients with osteoporotic vertebral fracture., Methods: Twelve postmenopausal patients (aged 73±4.8 years) with osteoporotic spinal fracture confirmed by MRI or CT scanning received conservative treatment with teriparatidesc injection supplemented with calcium and analgesics for 6 months. At the beginning and at the end of the therapy, VAS score, Oswestry Disability Index (ODI), bone mass densitometry, and X-ray of the thoracic and lumbar spine, and serum P1NP and beta-CTX levels were measured. Six of the patients received a second MRI scan after the therapy to evaluate the bone healing., Results: All the 12 patients completed the treatment, during which no new fractures or adverse events occurred. At the end of the first month of treatment, analgesic was withdrawn for all the patients. The average VAS score decreased from 8±2 to 1±2 at 1 month during the therapy, and ODI was reduced from (76±12)% to (20±5)% at 1 month and further to (5±4)% at 6 month. After the 6-month therapy, the height of the fractured vertebrae (presented as the anterior to posterior wall height ratio) was insignificantly decreased from (75±20)% to (61±20)%, the BMD was increased by (20±5)%, P1NP increased significantly from 20.9±11.4 ng/mL to 80.0±41.2 ng/mL, and beta-CTX increased from 0.30±0.17 ng/mL to 0.51±0.3 ng/mL. The 6 patients re-examined with MRI demonstrated complete bone healing after the therapy., Conclusion: Teriparatide is effective for conservative treatment of osteoporotic spinal fracture and can promote bone fracture healing, improve the quality of life, and prevents vertebral collapse, and can be therefore an alternative treatment to PVP or BV.

Objective: To investigate the role of platelet-derived growth factor-BB (PDGF-BB) in the degeneration of the hypertrophied ligamentum flavum (LF) in the lumbar spine., Methods: Surgical specimens of degenerative hypertrophied LF were obtained from 11 patients with lumbar spinal stenosis (LSS, mean age 57.8 years), with those from 10 age-matched patients with lumbar disc herniation (LDH, mean age 53.5 years) as the normal controls. The thickness of the LF was measured preoperatively by axial T1-weighted magnetic resonance imaging (MRI) at the facet joint level. Immunohistochemistry was employed to determine the expression of PDGF β and PDGF-BB in the LF. The mRNA and protein expressions of PDGF-BB in the LF were detected using real-time PCR and Western blotting, respectively., Results: The thickness of the LF was 5.30±1.12 mm in the degenerative group and 2.80±1.53 mm in the control group, showing a significant difference between the two groups (P<0.001). PDGF-β and PDGF-BB were positive in the fibroblasts in hypertrophied LF. The mRNA and protein expressions of PDGF-BB were significantly higher in the degenerative group than in the control group (P=0.013 and 0.023, respectively)., Conclusion: The high expression of PDGF-BB in the hypertrophied LF suggests its important role in the development of hypertrophy of LF in lumbar spinal canal stenosis.

Objective: To study the changes in the biological behavior of bone marrow mesenchymal stem cells (BMSCs) transfected with red fluorescent protein by lentivirus (RFP-BMSCs) seeded on in poly-D, L-lactide acid (PDLLA) scaffolds with bioactive modification by ammonia plasma and Gly-Arg-Gly-Asp-Ser (GRGDS) in vitro., Methods: Circular sheets of PDLLA scaffolds (8 mm in diameter and 1 mm in thickness) were prepared and aminated with PDLLA (group A) or modified with the peptide conjugate A/PDLLA (group PA), with untreated PDLLA as the control (group P). The RFP-BMSCs were seeded on the scaffold materials and their proliferation and metabolic activity were detected using CyQuant NF and Alamar blue staining. The mineralization on the scaffolds was observed using calcein fluorescent dye under a fluorescent microscope. The adhesion and proliferation of RFP-BMSCs were observed by fluorescent microscope, and scanning electron microscope (SEM) was used to confirm the observed adhesion of the seed cells., Results: The RFP-BMSCs seeded on the 3 scaffolds all showed proliferative activity at different time points after cell seeding, and the cell numbers decreased significantly in the order of PA>A>P (P<0.001). The cell number was significantly greater in group PA than in group A at all the time points except for days 10 (P=0.077) and 12 (P=0.491), and gradually became similar with the passage of time. The metabolic changes of the cells follow a similar pattern of cell proliferation. RFP-BMSCs showed more active proliferation in group A and group PA than in group P. On days 14 and 21, the intensity of green fluorescence decreased in the order of group PA, A and P. The RFP-BMSCs showed better adhesion in group PA than in group A, and the cells in group P appeared more scattered under scanning electron microscope., Conclusion: Bioactive modification of PDLLA by ammonia treatment and conjugation with GRGDS peptides may promotes the adhesion, proliferation, metabolism and mineralization of RFP-BMSCs seeded on PDLLA scaffolds.

Objective: To compare the histological features of the thoracic vertebral body growth plates (VBGPs) of rats at different ages and assess their proliferative capability., Methods: The thoracic VBGPs obtained from rats aged 1 day and 1, 4, 8, 16 and 28 weeks were identified using safranin O-fast green staining, and the height of the hypertrophic zone, proliferative zone, and resting zone were measured. The chondrocytes were isolated from these VBGPs with a modified trypsin-collagenase type II digestion method for primary culture in vitro. The expressions of proliferating cell nuclear antigen (PCNA) mRNA and protein was detected by real time-PCR and Western blotting, respectively., Results: The 1-day- and 1-week-old rats showed significantly greater hypertrophic zone and proliferative zone in the VBGPs than older rats (P<0.01); the proliferative zone was significantly greater in rats aged 4 weeks than in those aged 28 weeks (P<0.05). The resting zone was obviously greater in rats aged 1 day and 1 week than in older rats (P<0.05), and also greater in rats aged 4 weeks than in those aged 16 and 28 weeks (P<0.05). Obvious ossification in the resting zone occurred at 16 weeks, and most of the resting zone became ossified at 28 weeks. The expression of PCNA decreased at both the mRNA and protein levels as the rats grew., Conclusion: The 3 zones of VBGPs are greater in rats aged 1 day and 1 week than in older ones. Ossification in the resting zone begins at 16 weeks, and till 28 weeks, most of the resting zone is ossified. The proliferation ability of VBGP chondrocytes decreases with the increase of age of the rats.

Objective: To introduce a rapid colorimetric method for assessing the viability of osteosarcoma cells after chemotherapy., Methods: Colorimetric assay and automatic microplate scanning spectrophotometer were used for assaying the viability of osteosarcoma cells., Results: Close correlation was found between the absorbance at 570 nm of the formazan products and the number of viable osteosarcoma cells., Conclusion: An effective, sensitive and convenient colorimetric assay has been established to assess the survival of osteosarcoma cells following chemotherapy.

Objective: To assess the feasibility of using MTT assay for detecting the apoptosis of osteosarcoma cells following chemotherapy., Methods: The osteosarcoma cells derived form surgical specimens were cultured in RPMI 1640 medium supplemented with 10% calf serum. Chemotherapeutic agents were administered in the cell culture, and MTT assay was used to observe the cell apoptosis under optical microscope., Results: MTT staining accurately identified apoptosis of the osteosarcoma cells, and the apoptotic cells were easily distinguished from normal cells and dead cells., Conclusion: MTT staining is convenient and practical for detecting osteosarcoma cell apoptosis.

Objective: To assess the effect of the anterior surgical approach for thoracolumbar spine tuberculosis and analyzed the causes of the surgical complications and the countermeasures., Methods: From Jan 1996 to Dec 2005, 120 patients with thoracolumbar spine tuberculosis underwent operations through the anterior approach either following the primary diagnosis (115 cases) or for recurrence (5 cases)., Results: Intraoperative pleural rupture occurred in 4 cases, and rupture of the external iliac vein occurred 1 case. Three patients had damages of the T12 dorsal ramus. One patient developed venous embolism of the lower extremity after the operation, two had paralytic ileus and 1 had false diabetes insipidus. Of the 5 recurrent patients, 1 died due to alcoholic cirrhosis and acute liver failure, 1 received a third operation for loosened internal fixation, and 1 had recurrence due to extensive drug resistance (XDR)., Conclusion: Standardized antituberculous therapy is fundamental for preventing the recurrence of tuberculosis, and individualized antituberculous therapy adjustment according to the results of bacterial culture and drug sensitivity tests can be the most effective means for preventing drug resistance and reducing tuberculosis recurrence.

Objective: To evaluate nacre powder-induced osteogensis in the femoral condyles of New Zealand rabbits and investigate the possible mechanism., Methods: Nacre powder was implanted into the femoral condyles of New Zealand rabbits and 4, 8, and 12 weeks after the implantation, radiographic examinations were carried out and the bone density was evaluated. The bone tissue specimens were sliced after fixation for histological observations. The osteogenesis area on the slice was estimated with Ponceau Red staining 8 and 12 weeks after the implantation and calculated with Imaga-Pro software., Results: X-ray after the operation did not reveal obvious evidence of angiogenesis in the femoral condyles, where the X-ray density underwent slight changes. The optical density decreased significantly after the implantation, and the quantity of the osteoid, woven and lamellar bone increased in the bone tissue with time. The osteogenesis area with Ponceau Red staining showed obvious bone formation, which was significantly different from the control group., Conclusion: T Obvious osteogenesis occurred in the femoral condyles after nacre powder implantation in New Zealand rabbits. Nacre powder has slow biodegradation in vivo and induces osteogenesis by osteoinduction.

Objective: To investigate the role of transforming growth factor (TGF)-beta1 in degeneration of the ligamentum flavum in the lumbar spine., Methods: The degenerative ligamentum flavum was obtained during surgery from 8 patients with lumbar spinal stenosis (mean age 58.6 years), and 8 young patients (mean age 24.2 years) with acute lumbar disc herniation were included as normal controls. The thickness of the ligamentum flavum was measured on preoperative magnetic resonance images, and the mRNA expressions of type-I collagen and TGF-beta1 in the ligamentum flavum were detected using reverse transcriptase-polymerase chain reaction. The protein expression and localization of TGF-beta1 were investigated by Western blotting and immunohistochemical staining, respectively., Results: The thickness of the ligamentum flavum were 4.70-/+0.40 mm in the degenerative group and 2.50-/+0.36 mm in the control group, showing significant difference between the two groups (P<0.001). The type-I collagen mRNA expression in the degenerative group was significantly higher than that in the control group (P=0.007). The mRNA and protein expressions of TGF-beta1 were significantly higher in the degenerative group than in the control group (P=0.008 and 0.004, respectively). Immunohistochemistry showed that TGF protein was localized in the fibroblasts within the ligamentum flavum., Conclusion: Degenerative ligamentum flavum shows hypertrophy and fibrosis, and TGF-beta1 overexpression may be associated with in the development and progression of ligamentum flavum degeneration in the lumbar spine.

Objective: To assess the improvements in the properties of nano-nacre artificial bone prepared on the basis of nacre/polylactide acid composite artificial bone and its potential for clinical use., Methods: The compound of nano-scale nacre powder and poly-D, L-lactide acid (PDLLA) was used to prepare the cylindrical hollow artificial bone, whose properties including raw material powder scale, pore size, porosity and biomechanical characteristics were compared with another artificial bone made of micron-scale nacre powder and PDLLA., Results: Scanning electron microscope showed that the average particle size of the nano-nacre powder was 50.4-/+12.4 nm, and the average pore size of the artificial bone prepared using nano-nacre powder was 215.7-/+77.5 microm, as compared with the particle size of the micron-scale nacre powder of 5.0-/+3.0 microm and the pore size of the resultant artificial bone of 205.1-/+72.0 microm. The porosities of nano-nacre artificial bone and the micron-nacre artificial bone were (65.4-/+2.9)% and (53.4-/+2.2)%, respectively, and the two artificial bones had comparable compressive strength and Young's modulus, but the flexural strength of the nano-nacre artificial bone was lower than that of the micro-nacre artificial bone., Conclusions: The nano-nacre artificial bone allows better biodegradability and possesses appropriate pore size, porosity and biomechanical properties for use as a promising material in bone tissue engineering.

Objective: To study the risk factors of secondary kyphotic angle increment after bone cement vertebroplasty for osteoporotic vertebral compression fractures., Methods: From October 2005 to May 2006, 32 (45 vertebrae) bone cement vertebroplasty procedures were performed. The operation time, injected cement volume, bone mineral density, visual analog scale (VAS) pain score, vertebral height, and kyphotic angle were recorded. The secondary increment of the kyphotic angle was calculated, and correlation analysis and linear regression analysis were performed., Results: The bone mineral density, the postoperative kyphotic angle and the vertebral midline height were significantly correlated to the secondary increment of the kyphotic angle., Conclusion: Large postoperative kyphotic angle, poor postoperative recovery of the vertebral midline height, and low bone mineral density are all risk factors of secondary increment of the kyphotic angle.

Objective: To express human platelet-derived growth factor (hPDGF) B chain mature peptide gene in a prokaryotic expression system and detect the bioactivity of the expressed protein., Methods: hPDGF B chain mature peptide gene was amplified and expressed in E. coli, and the recombinant protein, rhPDGF-BB, was purified and renatured in GSSG/GSS system. The bioactivity of rhPDGF-BB in vitro was evaluated with SD rat osteoblasts., Results: The full-length PDGF-B mature peptide gene was obtained and verified, and successfully expressed in E. coli. Bioactivity detection results showed that the expressed rhPDGF-BB obviously promoted the proliferation and DNA replication of SD rat osteoblasts in vitro (P<0.01)., Conclusion: he PDGF-B chain mature peptide cDNA has been successfully cloned and the PDGF-B precursor highly expressed in E. coli, and renatured rhPDGF-BB displays high bioactivity as shown by MTT assay and flow cytometry. This success provides the basis for production of functional PDGF-BB and facilitates further studies of its role in fracture healing and trauma reconstruction.

Objective: To provide insights into the mechanisms and pathways of osteogensis by observing the effects of water-soluble matrix of nacre (WSM) on bone morphogenetic protein-2 (BMP-2) and Cbfa1 gene expressions in rabbit marrow mesenchymal stem cells (BMSCs)., Methods: New Zealand rabbit BMSCs cultured in vitro were stimulated with different concentrations of WSM extracted at low temperature, and the activity of AKP in the cells was evaluated with the dose-effect curve generated. BMP-2 and Cbfa1 gene expressions in rabbit BMSCs exposed to WSM were assayed with one-step RT-PCR., Results: The activity of AKP in rabbit BMSCs increased after stimulation with different concentrations of WSM, and the effects were the most obvious with the WSM concentration ranging from 150 to 200 microg/ml. BMP-2 gene expression in the BMSCs increased after WSM exposure, but which did not result in obvious changes in Cbfa1 gene expression., Conclusion: WSM induces differentiation of rabbit BMSCs towards osteoblasts by increasing BMP-2 gene expression, in which process Cbfa1 gene does not seem to play a significant role.

Objective: To test the efficacy of kyphoplasty using an enhanced balloon expander in restoring the height of vertebral compression fractures (OVCF)., Methods: Fifteen lumbar vertebral bodies (L1-L5) were harvested from 3 young male fresh cadavers and separated into individual vertebral bodies with the bilateral pedicles of the vertebral arch removed. Before operation, plain X-ray films of all the vertebral bodies were obtained. All the vertebral bodies were compressed lengthwise to approximately 80% of their original heights using a universal material-testing machine to result in compression fractures. Post-compression vertebral bodies were then repaired using an enhanced balloon expander, and the delivery of the bone cement into the vertebral bodies was observed. The heights of the anterior and posterior borders of the vertebral bodied were measured before and after compression as well as after kyphoplasty., Results: The inflation of the balloon expander averaged 2.95-/+0.18 ml and the pressure was 122.67-/+27.89 psi (1 psi=6895 Pa). Kyphoplasty resulted in significant restoration of the vertebral body height lost due to the compression (P<0.01)., Conclusion: Kyphoplasty using an enhanced balloon expander may restore vertebral body height damaged by compression and correct the kyphotic deformity. The balloon expander can be a effective and economic choice for kyphoplasty for its relatively low cost.