Tumor necrosis factor-α inhibition restores matrix formation by human adipose-derived stem cells in the late stage of chondrogenic differentiation.

BACKGROUND: Cartilage tissue engineering is a promising strategy for treating cartilage damage. Matrix formation by adipose-derived stem cells (ADSCs), which are one type of seed cell used for cartilage tissue engineering, decreases in the late stage of induced chondrogenic differentiation in vitro, which seriously limits research on ADSCs and their application. AIM: To improve the chondrogenic differentiation efficiency of ADSCs in vitro, and optimize the existing chondrogenic induction protocol. METHODS: Tumor necrosis factor-alpha (TNF-α) inhibitor was added to chondrogenic culture medium, and then Western blotting, enzyme linked immunosorbent assay, immunofluorescence and toluidine blue staining were used to detect the cartilage matrix secretion and the expression of key proteins of nuclear factor kappa-B (NF-κB) signaling pathway. RESULTS: In this study, we found that the levels of TNF-α and matrix metalloproteinase 3 were increased during the chondrogenic differentiation of ADSCs. TNF-α then bound to its receptor and activated the NF-κB pathway, leading to a decrease in cartilage matrix synthesis and secretion. Blocking TNF-α with its inhibitors etanercept (1 µg/mL) or infliximab (10 µg/mL) significantly restored matrix formation. CONCLUSION: Therefore, this study developed a combination of ADSC therapy and targeted anti-inflammatory drugs to optimize the chondrogenesis of ADSCs, and this approach could be very beneficial for translating ADSC-based approaches to treat cartilage damage.

Association between Mean Corpuscular Hemoglobin and Platelet Distribution Width Ratio and Knee Osteoarthritis Severity.

BACKGROUND: The goal was to simply and efficiently predict the indicators of disease severity in knee osteoarthritis (KOA) patients. METHODS: One hundred eighty-four patients with KOA and 126 healthy subjects were included. WOMAC (Western Ontario and McMaster Universities Osteoarthritis Index) was used as a reference index for disease severity in KOA patients, in which WOMAC < 80 was classified as mild and WOMAC ≥ 80 as moderate and severe. Blood routine parameters of the KOA and the healthy groups were analyzed by the Mann Whitney U test. Receiver operating characteristic curves were used to analyze the sensitivity and specificity of mean corpuscular hemoglobin and platelet distribution width ratio (MPR) and monocyte and hemoglobin ratio (MHR) indicators. The correlation between MPR and MHR and disease severity of KOA was determined by bivariate regression analysis. Independent predictors of disease severity in patients with KOA were assessed by multivariate regression analysis. RESULTS: MPR, MHR, and WOMAC were significantly higher in the KOA group. The ROC curve indicated that the cutoff values of MPR and MHR were 2.09 and 0.0030, respectively, with sensitivity of 86.4% and 68.5% and specificity of 99.2% and 79.4%. Bivariate regression analysis found that MPR was better correlated with disease severity than MHR. The results of multivariate regression analysis demonstrated that the MPR values of moderate and severe patients were more than 19 times that of mild patients, and the OR values were 21.695 and 19.558, respectively. CONCLUSIONS: MPR and MHR demonstrated a good correlation with disease severity in patients with KOA. MPR is a potential independent predictor of disease severity in patients with KOA.

Development and validation of a pre-scoring system for nonspecific low back pain among general population in Guangzhou: a cross-sectional study.

BACKGROUND: Nonspecific Low Back Pain (NLBP) is a common disease with a low cure rate and significant impact on the population. This study aimed to develop and validate a pre-scoring system for identifying the risk of suffering from NLBP among the general population in Guangzhou. METHODS: A total of 1439 eligible subjects were surveyed in Guangzhou by stratified random sampling and was divided randomly into the development dataset (69.6%) and validation dataset (30.4%) subsequently. Based on the development dataset, potential associated factors (average exercise times weekly, the intensity of daily work, etc.) with NLBP were tested by the sequential logistic regression, and a pre-scoring system was formulated with Sullivan's method and graded afterward. The internal validity of the system was assessed by AUC and calibration plot, and the external validation was performed in the validation dataset. RESULTS: The prevalence rates of NLBP in the development dataset and the validation dataset were 12.97 and 13.27%, respectively. Age, BMI, average exercise times weekly, gender, educational level, the intensity of daily work, place of residence, monthly income, overall evaluation of health condition and physiology health were identified as significant factors. The total risk score ranged from 0 to 38, which was split into three risk grades: low risk (0 to 18), intermediate risk (19 to 22) and high risk (23 to 38). The pre-scoring system had an adequate calibration and a good discriminating ability with bootstrap-corrected AUC equaling 0.861 in the development dataset and 0.821 in the validation dataset. CONCLUSIONS: A pre-scoring system that could help clinicians to assess the risk of NLBP in the general population was validated. Further validation of the system in a new population or prospective cohort study is suggested.

[FGF-2/PELA/BMP-2 microcapsule scaffold promotes osteogenic differentiation of rat periosteum-derived stem cells in vitro].

OBJECTIVE: To observe the effect of a microencapsule scaffold capable of sustained release of fibroblast growth factor-2 (FGF-2) and bone morphogenetic protein-2 (BMP-2) in promoting the osteogenic differentiation of rat periosteum-derived stem cells (PDSCs) in vitro. METHODS: PDSCs from 4-week-old SD rats, after identification of the surface markers using flow cytometry, were induced to differentiate into osteoblast, chondroblast, and adipocyte lineages. The differentiated cells were verified by staining with Alizarin red, toluidine blue, alcian blue, oil red O and by immunofluorescence assay. FGF-2/PELA/BMP-2, FGF-2/PELA, PELA/BMP-2 and PELA microcapsules were prepared, examined for surface morphologies using scanning electron microscopy (SEM), and tested for controlled release of FGF-2 and BMP-2 using ELISA. The third passage of PDSCs were cultured in the presence of the aqueous extracts of one of the 4 materials, and alkaline phosphatase (AKP) activity in the culture media was detected at 7 and 14 days of culture; the expression levels of osteogenesis-related genes were quantified with quantitative real-time PCR (qRT-PCR). The osteogenic differentiation ability of the PDSCs cultured with the extracts was compared. RESULTS: The PDSCs, which expressed mesenchymal stem cell surface markers, were shown to have osteogenic, chondrogenic and adipogenic differentiation potentials. The cells cultured with the extract of FGF-2/PELA/BMP-2 microcapsules showed the highest AKP activity at 7 and 14 days of culture, and their expression levels of OCN and RunX-2 mRNA were the highest among the 4 groups; RunX-2 expression reached its peak level on day 14, and OCN mRNA expression level increased progressively as the culture time extended. CONCLUSION: FGF-2/PELA/BMP-2 biomimetic controlled release microcapsules preserve the cytokine activities and are capable of promoting the osteogenic differentiation of rat PDSCs.

[Analysis of clinical factors correlating with osteoporosis in patients with type 2 diabetes mellitus].

OBJECTIVE: To analyze the clinical factors that contribute to lowered bone mineral density (BMD) of the lumbar vertebrae in type 2 diabetic patients. METHODS: Forty type 2 diabetic patients with osteoporosis and 40 age- and gender-matched healthy individuals, selected using propensity score method, were examined for BMD of the L1 to L4 vertebrae and the left femur. Age, gender, course of the disease, body mass index (BMI), fasting blood glucose, serum calcium, serum phosphate, urinary calcium, and urinary phosphate were compared between the two groups. RESULTS: BMD of the L1 to L4 vertebras was significantly lower in the diabetic group than in the healthy individuals (P<0.05). In the diabetic patients, BMD showed an obvious difference between male and female patients and was negatively correlated with the course of the disease but positively with BMI, fasting blood glucose, urinary calcium and urinary phosphate; BMD was not correlated with age, serum calcium or serum phosphate in these patients. CONCLUSION: Osteoporosis in type 2 diabetic patients is closely related with gender, BMI, course of the disease and poorly controlled glucose level.

TGF-ß1, in association with the increased expression of connective tissue growth factor, induce the hypertrophy of the ligamentum flavum through the p38 MAPK pathway.

Hypertrophy of the ligamentum flavum (LF) is one of the key pathomechanisms of lumbar spinal stenosis (LSS). Transforming growth factor (TGF)-ß1 is abundantly expressed in hypertrophied degenerative LF tissues from LSS. However, the molecular mechanisms underling the association between TGF-ß1 and LF hypertrophy have not yet been fully elucidated. In this study, we investigated the important role of the mitogen-activated protein kinase (MAPK) pathway in the pathogenesis of LSS by analyzing the expression of connective tissue growth factor (CTGF) and extracellular matrix (ECM) components (collagen I and collagen III) in TGF-ß1-treated LF cells. Cell growth assay revealed that TGF-ß1, in association with CTGF, enhanced the the proliferation of LF cells, and we found that TGF-ß1 also elevated CTGF expression and subsequently enhanced the mRNA expression of collagen I and collagen III. The increased mRNA expression levels of CTGF, collagen I and collagen III were abolished by p38 inhibitors. Both immunofluorescence imaging and western blot analysis of p38 and p-p38 revealed the increased expression and phosphorylation of p38. Silencing the expression of p38 by siRNA in LF cells decreased the protein expression of p38, p-p38 and CTGF, as well as the mRNA expression of CTGF, collagen I and collagen III. Taken together, our findings indicate that TGF-ß1, in association with the increased expression of CTGF, contribute to the homeostasis of the ECM and to the hypertrophy of LF through the p38 MAPK pathway.

[Pullout test in expansive unilateral open-door laminoplasty of the cervical spine with OsteoMed M3 plate and screws: the screw orientation matters].

OBJECTIVE: To study the impact of screw orientation on the pullout strength of OsteoMed M3 titanium screws in expansive unilateral open-door laminoplasty of the cervical spine. METHODS: Six fresh human cervical spine specimens were randomly numbered and OsteoMed M3 plate and screws were used for an expansive unilateral open-door laminoplasty. The screws were inserted in the lateral mass at different extraversion angles (0°, 30° and 45°). The maximum pullout strength was tested on the ElectroForce material testing machine. RESULTS: The maximum pullout strength was 81.60∓7.33 N, 150.05∓15.57 N, and 160.08∓17.77 N in extraversion angle 0°, 30°, and 45° groups, respectively. The maximum pullout strength was significantly less in extraversion angle 0° group than in 30° and 45° groups (P<0.05), but similar in the latter two groups. CONCLUSION: The pullout strength of the screws inserted at an extraversion angle over 30° provides stronger fixation than an angle of 0° in the unilateral open-door laminoplasty using OsteoMed M3 titanium plate and screws.

Posterior cervical fixation following laminectomy: a stress analysis of three techniques.

The aim of this study was to compare the following three main fixation techniques: pedicle screw (PS) technique, lateral mass screw (LS) technique, and transarticular screw (TS) technique. A detailed, geometrically accurate, nonlinear C3-C7 FE model had been successfully developed and validated. Then three finite element (FE) models were reconstructed by different fixation techniques following C4-C6 level laminectomy. A compressive preload of 74 N combined with a pure moment of 1.8 Nm in flexion, extension, left-right lateral bending, and left-right axial rotation was applied to the models. The results showed that maximum von Mises stress on the fixation devices was much higher in the FE models of TS technique, compared with the models of PS and LS techniques. Furthermore, the screws inserted by TS technique had high stress concentration at the middle part of the screws. Screw inserted by PS and LS techniques had high stress concentration at the actual cap-rod-screw interface. The highest level of maximal stress was obtained with the fixation device of the TS technique. TS technique induces noticeable differences in the stress compared to the posterior cervical fixation technique, regarding the higher stress level on fixation devices.

[Placement of short iliac screw using Galveston technique in lumbosacral fusion].

OBJECTIVE: To evaluate the effect of placement of short iliac screw using Galveston technique in lumbosacral fusion. METHODS: From October 2003 to August 2007, 18 consecutive patients (mean age 46 years ranging from 25 to 62 years) received placement of short iliac screw in lumbosacral fusion. The patients were followed up for a mean of 18 months (12-23 months), and the effect of lumbosacral fusion was evaluated according to standing anterior-posterior and lateral plain films taken before and after the operation and at the follow-up and also on the basis of symptom relief. RESULTS: The mean time of surgery was 210 min (180-290 min). No complications occurred during and after the operation. According to the evaluation criteria of surgical treatment of low back pain formulated by the spine group of Chinese Orthopedic Association, excellent clinical outcome was achieved in 12 cases, good outcome in 3 cases, and tolerable outcome in 2 cases, with the excellent and good outcome rate of 83%. CONCLUSION: The Galveston technique for short iliac screw placement can obtain satisfactory outcome in the lumbosacral fusion.

[Effect of continuous passive motion on basic fibroblast growth factor expression during tendon-bone repair after surgical repair of acute rupture of the supraspinatus tendon in rabbits].

OBJECTIVE: To study the effect of continuous passive motion (CPM) on basic fibroblast growth factor (b-FGF) expression during tendon-bone repair in rabbits and explore the role of stress in the postoperative repair after acute rotator cuff injury. METHODS: Sixteen rabbits randomized into CPM group (n=8) and non-CPM group (n=8) were subjected to surgically induced acute rupture of the supraspinatus tendon and subsequent surgical repair, with another two rabbits serving as the control. Two weeks after the operation, the rabbits in CPM group underwent CPM training, and those in non-CPM group were normally fed only. At 2, 4, 6, and 8 weeks after the operation, 2 rabbits from each group were sacrificed and the tissue samples were obtained for detecting the changes in b-FGF expression. RESULTS: Two weeks after the operation, b-FGF expression was detected in both groups, and the CPM group showed slightly higher and more diffusive expression. At 4 weeks, b-FGF expression was significantly higher and distributed over a greater area in CPM group and in the non-CPM group. A large number of fibroblasts positive for b-FGF expression were identified in CPM group, aligning in parallel with the tendon membrane. At 6 weeks, b-FGF in the CPM group showed no obvious changes but that in the non-CPM group became lightened. At 8 weeks, b-FGF expression was reduced in both groups, which was more obvious in the non-CPM group. CONCLUSION: CPM can promote b-FGF expression to enhance type III collagen synthesis at the tendon-bone interface in early stage of tendon-bone repair following acute rupture of supraspinatus tendon in rabbits, thereby contributing to tendon-bone recovery after rotator cuff injury.

[Indication of fusion for degenerative lumbar spinal stenosis treated by "windows technique" laminoforaminotomy].

OBJECTIVE: To explore the indications of fusion for degenerative lumbar spinal stenosis treated by "windows technique". METHODS: From December 1999 to December 2005, 145 consecutive patients who were treated by primary decompression with "windows technique" laminoforaminotomy for degenerative lumbar spinal stenosis, a retrospective study, were divided into 3 groups (A and B and C) by preoperative lumbar conditions and surgical methods. In group A, 39 patients with spinal instability or degenerative lumbar spondylolisthesis or scoliosis underwent decompression and fusion; in group B, 31 patients with spinal instability or degenerative lumbar spondylolisthesis or scoliosis underwent decompression alone; In group C, 75 patients without spinal instability or degenerative lumbar spondylolisthesis or scoliosis were treated by decompression without fusion. On hospital medical records to review, they were followed up by telephone and out-patient referral. Statistics the duration of hospitalization, operative time, estimated blood loss; Observed recrudescence and reoperation and complication; and using Oswestry Disability Index and Visual Analog Scale and satisfaction rate for efficacy assessment, application SPSS 13.0 software. RESULTS: All 145 patients had at least a 3-year follow-up (ranging 37 to 108 months). In the group C, the duration of hospitalization less than in the group A or B (P < 0.05); In the group A, the operative time and estimated blood loss greater than in the group B or C (P < 0.05); The group B treated by decompression alone in the presence of instability or spondylolisthesis or scoliosis showed the worst results by the Oswestry Disability Index or Visual Analog Scale or ate of satisfaction (P < 0.05). The same good results can be obtained in the group A and C. There were not different about recrudescence or reoperation or complication in the three groups. CONCLUSIONS: Fusion should be performed on patients with instability or degenerative lumbar spondylolisthesis or scoliosis after primary decompression with "windows technique" laminoforaminotomy. The patient with simple lumbar spinal stenosis undergone primary surgery does not require fusion.

[Biomechanical evaluation of asymmetrical posterior internal fixation for transforaminal lumbar interbody fusion with transfacetopedicular screws].

OBJECTIVE: To assess the biomechanical stability of asymmetrical posterior internal fixation for transforaminal lumbar interbody fusion (TLIF) with transfacetopedicular screws (TFPS). METHODS: Range of motion (ROM) testing was performed in 7 fresh-frozen human cadaveric lumbar spine motion segments in flexion/extension, lateral bending, and axial rotation using 10.0 Nm torques at the L3-4 motion segment. The sequential test configurations included intact motion segment, TLIF and ipsilateral pedicle screw (PS), TLIF and ipsilateral PS plus contralateral TFPS according to Boucher technique, and TLIF and bilateral PS. The ROM was determined to assess the construct stability. RESULTS: In flexion/extension, lateral bending, and axial rotation, no measureable difference was found in the ROM between the standard bilateral pedicle screw and the novel asymmetric posterior internal fixation after TLIF. After TLIF, the ipsilateral PS construct provided less segment stability than the novel asymmetric construct with TFPS in flexion, left bending and left rotation. In rotation, the novel asymmetric construct allowed for significant off-axial rotation motions detrimental to the stability and fusion. CONCLUSION: Ipsilateral PS placement plus contralateral TFPS provides stability comparable to that of TLIF with bilateral PS, and can be an alternative in minimal invasive surgery.

[Surgery simulation with a percutaneous and transpedical interbody bone grafting apparatus].

OBJECTIVE: To develop a percutaneous and transpedical interbody bone grafting apparatus for vertebral bone defect reconstruction in thoracolumbar fracture correction via minimally invasive operation. METHODS: The percutaneous and transpedical interbody bone grafting apparatus was designed with CAD software, and the reduction effect, range of bone grafting and surgical complications of the apparatus were investigated in adult cadaveric thoracolumbar body and with computerized surgical simulation. RESULTS: The self-designed apparatus was convenient for percutaneous and transpedical interbody bone grafting that did not give rise to complications. CT showed large bone grafting area with increased density in the vertebral body corrected with this apparatus. CONCLUSION: The designed apparatus allows easy manipulation and efficient bone grafting and repositioning. Minimally invasive interbody bone grafting in thoracolumbar fracture can be easily performed with proper application of the apparatus.

[A digital model for lumbar motion segment reconstruction and three-dimensional visualization].

OBJECTIVE: To establish a digital model for reconstruction and three-dimensional visualization of lumbar motion segment from CT images. METHODS: Based on 64 row spiral CT continuous 2-dimensional images of L4-L5 segments, the lumbar structure and various soft tissues were reconstructed with the Mimics software, followed by validation of this model using finite element analysis software. RESULTS: The established three-dimensional digital model of lumbar motion segment included two vertebral bodies, cortical bone, cancellous bone, endplate, annulus fibrosus, nucleus pulposus and 6 types of ligaments. The output result of the digital model can be used to for research of computer aided design, rapid rototyping and finite element analysis. CONCLUSION: The application of thin-layer CT and Dicom standard ensures more accurate digital model establishment, and Mimics software allows more convenient reconstruction of the human skeleton and various soft tissues to facilitate further research.

[Development of an anti-infection nano-hydroxypatite drug delivery microsphere and its drug-release in vitro].

OBJECTIVE: To develop an anti-infection nano-hydroxypatite (nano-HA) microsphere for local drug delivery for treating osteomyelitis. METHODS: The nano-HA was used as the core carrier to load gentamicin (GM) and coated with poly(-hydroxybutyrate-co- hydroxyvalerate)/polyethylene glycol (PHBV/PEG), which was degradable and biocompatible, to prepare nano-HA-PHBV/PEG-GM microsphere. The surface structure and in vitro drug-release of the microsphere were studied. RESULTS: The microsphere had good drug delivery capability. The samples weighing 90 mg each were soaked in PBS and gentamicin release within the first day was 165.2 microg/ml, which maintained a low release rate in the following days. After 28 days, gentamicin release declined to 8.5 microg/ml, which was higher than the minimal inhibitory concentration of gentamicin (2 microg/ml). CONCLUSION: The local drug delivery system has good drug-release performance in vitro and may possess potential value in clinical management of osteomyelitis.

Three-dimensional porous poly-DL-lactide/basic fibroblast growth factor composites for bone defect repair: an experimental study.

OBJECTIVE: To investigate the osteoinductive ability of the composites consisting of basic fibroblast growth factor (bFGF) and porous poly-DL-lactide (PDLLA) for the development of a new absorbable osteosynthesis material. METHODS: Highly porous foams of PDLLA with the pore size ranging from 150 to 300 microm were prepared by a solvent-casting, particulate-leaching technique with NaCl as the leachable component. Animal models of radial diaphyseal defects of 1.0 cm with complete removal of the periosteum were induced in 45 rabbits, which were randomly divided into 3 groups to receive the defect repair with PDLLA and PDLLA/bFGF respectively, leaving one group untreated to serve as the control group. The implant specimens were harvested at 2, 4, 8, and 12 weeks respectively after the surgery and X-ray, histological and scanning electron microscopic (SEM) examinations were performed to evaluate the effectiveness of defect repair. At 8 and 12 weeks after implantation, biomechanical test (for three-point bending strength) was employed to study the quality of bone formation. RESULTS: PDLLA/bFGF composite stimulated more bone formation and had higher bending strength than PDLLA (P<0.05), and the bone formation induced by both materials was significantly more than that observed in the control group in every postoperative stage (P<0.05). CONCLUSION: PDLLA possesses good biocompatibility and absorbability, and when prepared into a porous material, it exhibits good osteoconductibility. As a good bFGF carrier, the foam of PDLLA with three- dimensional structure shows good osteoinductive ability with regard to the rapidity, quantity and quality of the bone formation.

Effect of pore size of D, L-polylactic acid as bone repair material on bone regeneration.

OBJECTIVE: To study the bone regeneration behavior in porous D,L-polylactic acid (D,L-PLA) with different pore sizes. METHOD: A particulate-leaching method was employed to prepare porous biodegradable D,L-PLA with different pore sizes (75, 250, 400, 750 micrometer) and with porosity of 75% as the materials to repair bone defects in rabbits. The materials were then implanted at random into 40 rabbits with bilateral radius bone defect, leaving another 10 rabbits without implantation as blank control. Gross observation and X-ray and histomorphological examination as well as assessment of the biomechanics of the implants were performed in 2, 4, 8 and 12 weeks respectively after the operation. RESULTS: New bone tissue occurred around the implanted materials with pore sizes of 250, 400 or 750 micrometer 12 weeks after the operation. In the control group and in the rabbits with implants with pore size of 75 micrometer, the bone defect was filled with connective tissues. The implants with 250-micrometer pores had the strongest biomechanical strengths of all the materials (P<0.01) at 8 weeks and 12 weeks after the operation. CONCLUSION: The pore size of the porous implants decides the behavior of bone regeneration, and D, L-PLA polymer with 250-micrometer pores produces the most desired effects.