USP7 promotes the osteoclast differentiation of CD14+ human peripheral blood monocytes in osteoporosis via HMGB1 deubiquitination.

Background: Abnormal osteoclast and osteoblast differentiation is an essential pathological process in osteoporosis. As an important deubiquitinase enzyme, ubiquitin-specific peptidase 7 (USP7) participates in various disease processes through posttranslational modification. However, the mechanism by which USP7 regulates osteoporosis remains unknown. Herein, we aimed to investigate whether USP7 regulates abnormal osteoclast differentiation in osteoporosis. Methods: The gene expression profiles of blood monocytes were preprocessed to analyze the differential expression of USP genes. CD14+ peripheral blood mononuclear cells (PBMCs) were isolated from whole blood collected from osteoporosis patients (OPs) and healthy donors (HDs), and the expression pattern of USP7 during the differentiation of CD14+ PBMCs into osteoclasts was detected by western blotting. The role of USP7 in the osteoclast differentiation of PBMCs treated with USP7 siRNA or exogenous rUSP7 was further investigated by the F-actin assay, TRAP staining and western blotting. Moreover, the interaction between high-mobility group protein 1 (HMGB1) and USP7 was investigated by coimmunoprecipitation, and the regulation of the USP7-HMGB1 axis in osteoclast differentiation was further verified. Osteoporosis in ovariectomized (OVX) mice was then studied using the USP7-specific inhibitor P5091 to identify the role of USP7 in osteoporosis. Results: The bioinformatic analyses and CD14+ PBMCs from osteoporosis patients confirmed that the upregulation of USP7 was associated with osteoporosis. USP7 positively regulates the osteoclast differentiation of CD14+ PBMCs in vitro. Mechanistically, USP7 promoted osteoclast formation by binding to and deubiquitination of HMGB1. In vivo, P5091 effectively attenuates bone loss in OVX mice. Conclusion: We demonstrate that USP7 promotes the differentiation of CD14+ PBMCs into osteoclasts via HMGB1 deubiquitination and that inhibition of USP7 effectively attenuates bone loss in osteoporosis in vivo.The translational potential of this article:The study reveals novel insights into the role of USP7 in the progression of osteoporosis and provides a new therapeutic target for the treatment of osteoporosis.

Comparison of Function- and Activity-Related Outcomes After Anterior Talofibular Ligament Repair With 1 Versus 2 Suture Anchors.

BACKGROUND: Few studies have compared the clinical outcomes of using 1 versus 2 suture anchors for anterior talofibular ligament (ATFL) repair. PURPOSE: To compare the function and activity-related outcomes of arthroscopic ATFL repair using 1 versus 2 suture anchors. STUDY DESIGN: Cohort study; Level of evidence, 3. METHODS: This retrospective study involved 46 patients (22 patients in the 1-anchor group, 24 patients in the 2-anchor group) who underwent ATFL repair between January 2015 and December 2017. American Orthopaedic Foot & Ankle Society score, Karlsson and Peterson score, and Tegner activity level were evaluated preoperatively and ≥2.5 years postoperatively. At follow-up, patients were also asked about time to return to sport as well as level and intensity of physical fitness. Satisfaction was evaluated with the Sefton grading system. RESULTS: After ≥2.5 years of follow-up (30 months in the 1-anchor group, 33 months in the 2-anchor group), patients in the 2-anchor group had a higher Tegner activity level than those in the 1-anchor group (mean ± SD, 4.75 ± 1.07 vs 4.05 ± 1.17; P = .039). As compared with patients in the 2-anchor group, fewer patients in the 1-anchor group returned to their preoperative activity level (54.2% vs 22.9%; P = .029); the rate of activity at the same or higher intensity as preinjury was also lower in the 1-anchor group (50% vs 79.2%; P = .038). However, there were no differences between the groups in terms of American Orthopaedic Foot & Ankle Society and Karlsson and Peterson scores, time to return to work/sport, duration of activity participation, level of physical fitness, or satisfaction according to Sefton grading. CONCLUSION: Arthroscopic ATFL repair appears to be an effective treatment regardless of whether 1 or 2 suture anchors are used. The techniques had similar functional outcome scores, but 1-anchor repair produced inferior activity-related outcomes.

Risk Factors and Scoring System of Cage Retropulsion after Posterior Lumbar Interbody Fusion: A Retrospective Observational Study.

OBJECTIVE: To investigate risk factors of cage retropulsion after posterior lumbar interbody fusion (PLIF) in China and to establish a scoring system of cage retropulsion. METHODS: The retrospective analysis was based on two hospital databases. The medical data records of posterior lumbar interbody fusion with cage retropulsion were selected from August 2009 to August 2019. Inclusion and exclusion criteria were set in advance. Risk factors including patients' baseline demographics (age, gender, operation diagnosis time difference), preoperative neurological symptoms, whether the fusion involves single or double segments, screw type, intraoperative compression, preoperative bone mineral density, whether there are neurological symptoms before surgery, whether there is urine dysfunction before surgery, disease type, complete removal of the endplate, and patient's education level. The research endpoint was the retropulsion of fusion cages. The Kaplan-Meier (K-M) method was used to analyze potential risk factors, and multivariate Cox regression was used to identify independent risk factors (P < 0.05). The Statistical Package for the Social Sciences (version 22.0; SPSS, IBM, Chicago, IL, USA) software was used for statistical analysis, and univariate analysis was used to screen out the factors related to cage retropulsion. All independent risk factors were included to predict the survival time of the retropulsion of cage. RESULTS: This study included a total of 32 patients with PLIF between 2009 to 2019. All patients were residents of China. Univariate analysis showed that there were 13 patients over 60 years old and 19 patients under 60 years old. There were 20 male patients and 12 female patients. The surgical diagnosis time was seven patients within 1 month, 17 patients within 1 to 3 months, and eight patients over 3 months. The disease type was 18 cases of lumbar disc herniation, 10 cases of lumbar spinal stenosis, four cases of lumbar spondylolisthesis. The fusion segment was 18 cases of single segment, 14 cases of double segment. The intraoperative compression was seven cases of compression, 25 cases of no compression. The preoperative bone mineral density was 10 cases of low density, 18 cases of normal, four cases of osteoporosis. The screw type was 27 cases of universal screw, five cases of one-way screw. Preoperative neurological symptoms were found in 25 cases and not in seven cases. Preoperative urination dysfunction occurred in 8 cases, whereas 24 cases did not have this dysfunction. The endplate was completely removed in 10 cases and not in 22 cases. Education level was nine cases of primary school education, 10 cases of secondary school, 13 cases of university level. Cox regression analysis showed that intraoperative pressure (hazard ratio [HR] = 4.604, P = 0.015) and complete removal of the endplate (HR = 0.205, P = 0.027) are associated with the time of cage retropulsion. According to the HR of each factor, the scoring rules were formulated, and the patients were divided into the low-risk group, moderate-risk group, and high-risk group according to the final score. The three median survival times of the three groups were 66 days in the low-risk group, 55 days in the moderate-risk group, and 45 days in the high-risk group, with statistical significance (P < 0.05). CONCLUSION: Intraoperative pressure and complete removal of the intraoperative endplate can be helpful to evaluating the expected time of cage retropulsion in patients with PLIF, and this clinical model guided the selection of postoperative prevention and follow-up treatment.

Comparison of Tri-Lock Bone Preservation Stem and the Conventional Standard Corail Stem in Primary Total Hip Arthroplasty.

OBJECTIVE: To compare the clinical and radiographic outcomes between the Tri-Lock Bone Preservation Stem (BPS) and the conventional standard Corail stem in primary total hip arthroplasty (THA). METHODS: From March 2012 to May 2014, we retrospectively reviewed 84 patients (104 hips) who received Tri-Lock (BPS) and 84 patients (115 hips) who received conventional standard Corail stem in THA. Their mean ages were 53.12 ± 2.32 years and 52.00 ± 2.11 years, respectively. The clinical outcomes were assessed by Western Ontario and McMaster University Osteoarthritis Index (WOMAC), Pain Visual Analogue Scale (VAS) and Harris Hip Score (HHS). The radiological outcomes were evaluated by the radiological examination. Accordingly, Intraoperative and postoperative complications were observed as well. RESULTS: The mean follow-up time was 48.23 ± 2.91 months in the Tri-Lock (BPS) group and 49.11 ± 2.11 months in the Corail group, respectively. The bleeding volumes in two groups were comparable (169.22 ± 58.11 mL vs 179.30 ± 59.14 mL, P = 0.003), with more bleeding volume in Corail group patients, while no statistically significance with respect to operation time was observed (65.41 ± 6.24 min vs 63.99 ± 6.33 min, P = 0.567). The rates of intraoperative fracture was 8% for the Corail group while 1% for the Tri-Lock (BPS) group (8% vs 1%, P = 0.030). At final follow-up, no statistical differences in regard to HHS, WOMAC, and Pain VAS were revealed between the two groups (P > 0.05). The rate of thigh pain was higher in Corail group than in Tri-lock (BPS) group (5% vs 0%, P = 0.043). However, incidence of stress shielding in grade 1 was higher in Tri-Lock (BPS) than in the Corail group (76% vs 23%, P < 0.01), while those in grade 2 and 3 were lower compared to the Corail stem (15% vs 28%, P < 0.01; 9% vs 16%, P = 0.008, respectively). Intriguingly, other assessments in relation to radiographic outcomes and postoperative complications were not comparable between the two groups. The Kaplan-Meier survival rate (revision surgery performed for any reason was defined as the end point) was similar between the two groups (P = 0.57), with 98.8% (95% confidence interval, 92.3%-100%) in Tri-lock (BPS) group and 97.6% (95% confidence interval, 94.6%-100%) in Corail group. CONCLUSIONS: The Tri-Lock (BPS) has similar clinic performances compared to the Corail stem. Furthermore, the Tri-lock (BPS) stem has some advantages in achieving lower incidence of thigh pain, stress shielding and intra-operative fracture. Therefore, we recommend the Tri-lock (BPS) stem as a good alternative in primary total hip arthroplasty, especially taking into account patient factors, including bone deficiency and convenience of extraction of the stem in hip revision.

All-Inside Arthroscopic Modified Broström Technique to Repair Anterior Talofibular Ligament Provides a Similar Outcome Compared With Open Broström-Gould Procedure.

PURPOSE: To introduce an all-inside modified Broström technique to suture the anterior talofibular ligament (ATFL) and inferior extensor retinaculum (IER) under arthroscopy and to compare its outcomes with those of the conventional open procedure. METHODS: All patients who underwent arthroscopic or open repair of the ATFL between June 2014 and December 2017 were included in this study. Visual analog scale (VAS), Karlsson and Peterson (K-P), American Orthopedic Foot and Ankle Society (AOFAS) ankle/hindfoot, and Tegner activity scores, as well as manual anterior drawer test (ADT), were used to evaluate the patients preoperatively and ≥2 years after surgery. The Sefton grading system was used to assess the level of satisfaction after surgery. Detailed surgical data and intraoperative findings were documented at the time of surgery. RESULTS: A total of 67 patients, 31 in the arthroscopic group and 36 in the open group, were included in this study (43 men and 24 women, mean body mass index 24.00, range 19.53 to 30.03). The surgical duration in the arthroscopic group (median, 34 minutes; range, 25 to 74) was significantly shorter than that in the open group (mean, 43.08 ± 8.11 minutes; 95% confidence interval [CI] 40.34 to 45.83) (P = .007). At the last follow-up, the subjective functional scores and ADT results improved significantly in both cohorts (P < .001). However, no significant difference was found in the VAS score (1.74 ± 1.24, 95% CI 1.29 to 2.2, in the open group versus 1.58 ± 1.2, 95% CI 1.18 to 1.99, in the arthroscopic group; P = .581), AOFAS score (91.71 ± 5.46, 95% CI 89.71 to 93.71, versus 90.67 ± 5.59, 95% CI 88.78 to 92.56; P = .444), K-P score (87.52 ± 7.59, 95% CI 84.73 to 90.3, versus 88.75 ± 5.56, 95% CI 86.87 to 90.63; P = .446), and ADT evaluation (normal: 96.77% versus 94.44%, P = .557) between the arthroscopic and open groups, respectively. In addition, 28 cases (90.32%) in the arthroscopic group and 32 (88.89%) in the open group achieved satisfactory results based on the Sefton grading system (P = .736). Seventeen patients (47.2%) in the open group and 18 patients (58.1%) in the arthroscopic group underwent Tegner evaluation after surgery, which showed no significant difference (5, interquartile range [IQR] 1 in the open group versus 5, IQR 3 in the arthroscopic group; P = .883). Complications were reported in 4 (11.1%) and 2 (6.5%) patients who underwent open and arthroscopic surgeries, respectively (P = .813). CONCLUSIONS: Both open and arthroscopic modified Broström surgeries generated favorable outcomes, with a significant improvement compared with the preoperative condition. Compared with the open Broström-Gould procedure, the all-inside arthroscopic modified Broström technique produced equivalent functional and clinical results at a minimum of 2 years after the operation, with a shorter surgical duration. Arthroscopic repair might be a safe and viable alternative to open surgery for lateral ankle stabilization. LEVEL OF EVIDENCE: III.

Recent advances of single-cell RNA sequencing technology in mesenchymal stem cell research.

Mesenchymal stem cells (MSCs) are multipotent stromal cells with great potential for clinical applications. However, little is known about their cell heterogeneity at a single-cell resolution, which severely impedes the development of MSC therapy. In this review, we focus on advances in the identification of novel surface markers and functional subpopulations of MSCs made by single-cell RNA sequencing and discuss their participation in the pathophysiology of stem cells and related diseases. The challenges and future directions of single-cell RNA sequencing in MSCs are also addressed in this review.

Long non-coding RNA: The functional regulator of mesenchymal stem cells.

Mesenchymal stem cells (MSCs) are a subset of multipotent stroma cells residing in various tissues of the body. Apart from supporting the hematopoietic stem cell niche, MSCs possess strong immunoregulatory ability and multiple differentiation potentials. These powerful capacities allow the extensive application of MSCs in clinical practice as an effective treatment for diseases. Therefore, illuminating the functional mechanism of MSCs will help to improve their curative effect and promote their clinical use. Long noncoding RNA (LncRNA) is a novel class of noncoding RNA longer than 200 nt. Recently, multiple studies have demonstrated that LncRNA is widely involved in growth and development through controlling the fate of cells, including MSCs. In this review, we highlight the role of LncRNA in regulating the functions of MSCs and discuss their participation in the pathogenesis of diseases and clinical use in diagnosis and treatment.

Neurotrophin-3 released from implant of tissue-engineered fibroin scaffolds inhibits inflammation, enhances nerve fiber regeneration, and improves motor function in canine spinal cord injury.

Spinal cord injury (SCI) normally results in cell death, scarring, cavitation, inhibitory molecules release, etc., which are regarded as a huge obstacle to reconnect the injured neuronal circuits because of the lack of effective stimulus. In this study, a functional gelatin sponge scaffold was used to inhibit local inflammation, enhance nerve fiber regeneration, and improve neural conduction in the canine. This scaffold had good porosity and modified with neurotrophin-3 (NT-3)/fibroin complex, which showed sustained release in vitro. After the scaffold was transplanted into canine spinal cord hemisection model, hindlimb movement, and neural conduction were improved evidently. Migrating host cells, newly formed neurons with associated synaptic structures together with functional blood vessels with intact endothelium in the regenerating tissue were identified. Taken together, the results demonstrated that using bioactive scaffold could establish effective microenvironment stimuli for endogenous regeneration, providing a potential and practical strategy for treatment of spinal cord injury. © 2018 The Authors Journal of Biomedical Materials Research Part A Published by Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 2158-2170, 2018.

DEPDC1 promotes cell proliferation and tumor growth via activation of E2F signaling in prostate cancer.

DEP domain containing 1 (DEPDC1) is recently reported to be overexpressed in several types of human cancer; however the role of DEPDC1 in prostate cancer remains to be investigated. Herein, we identified that the DEPDC1 mRNA and protein expression levels were dramatically increased in prostate cancer tissues and cell lines. Overexpression of DEPDC1 promoted, but depletion of DEPDC1 inhibited cell proliferation by regulating the G1-S phase cell cycle transition. Importantly, we found that DEPDC1 was essential for the tumor growth and formation of bone metastases of prostate cancer cells in vivo. Finally, we demonstrated that DEPDC1 interacted with E2F1 and increased its transcriptional activity, leading to hyper-activation of E2F signaling in prostate cancer cells. Our findings reveal an oncogenic role of DEPDC1 in prostate cancer progression via activation of E2F signaling, and suggest DEPDC1 might be a potential therapeutic target against the disease.

Intraoperative contrast-enhanced ultrasonography for microcirculatory evaluation in rhesus monkey with spinal cord injury.

This study tried to quantify spinal cord perfusion by using contrast-enhanced ultrasound (CEUS) in rhesus monkey models with acute spinal cord injury. Acute spinal cord perfusion after injury was detected by CEUS, coupling with conventional ultrasound (US) and Color Doppler US (CDFI). Time-intensity curves and perfusion parameters were obtained by autotracking contrast quantification (ACQ) software in the epicenter and adjacent regions of injury, respectively. Neurological and histological examinations were performed to confirm the severity of injury. US revealed spinal cords were hypoechoic and homogeneous, whereas dura maters, pia maters, and cerebral aqueducts were hyperechoic. After spinal cord contusion, the injured spinal cord was hyperechoic on US, and intramedullary vessels of adjacent region of injury were increased and dilated on CDFI. On CEUS hypoperfusion were found in the epicenter of injury, while hyperperfusion in its adjacent region. Quantitative analysis showed that peak intensity (PI) decreased in epicenters of injury but significantly increased in adjacent regions at all time points (p < 0.05). Functional evaluation demonstrated significant deterioration compared to pre-contusion (p < 0.05). Quantitative analysis with CEUS is a promising method for monitoring perfusion changes of spinal cord injury in overall views and real-time.

Transplantation of tissue engineering neural network and formation of neuronal relay into the transected rat spinal cord.

Severe spinal cord injury (SCI) causes loss of neural connectivity and permanent functional deficits. Re-establishment of new neuronal relay circuits after SCI is therefore of paramount importance. The present study tested our hypothesis if co-culture of neurotrophin-3 (NT-3) gene-modified Schwann cells (SCs, NT-3-SCs) and TrkC (NT-3 receptor) gene-modified neural stem cells (NSCs, TrkC-NSCs) in a gelatin sponge scaffold could construct a tissue engineering neural network for re-establishing an anatomical neuronal relay after rat spinal cord transection. Eight weeks after transplantation, the neural network created a favorable microenvironment for axonal regeneration and for survival and synaptogenesis of NSC-derived neurons. Biotin conjugates of cholera toxin B subunit (b-CTB, a transneuronal tracer) was injected into the crushed sciatic nerve to label spinal cord neurons. Remarkably, not only ascending and descending nerve fibers, but also propriospinal neurons, made contacts with b-CTB positive NSC-derived neurons. Moreover, b-CTB positive NSC-derived neurons extended their axons making contacts with the motor neurons located in areas caudal to the injury/graft site of spinal cord. Further study showed that NT-3/TrkC interactions activated the PI3K/AKT/mTOR pathway and PI3K/AKT/CREB pathway affecting synaptogenesis of NSC-derived neurons. Together, our findings suggest that NT-3-mediated TrkC signaling plays an essential role in constructing a tissue engineering neural network thus representing a promising avenue for effective exogenous neuronal relay-based treatment for SCI.

Graft of the NT-3 persistent delivery gelatin sponge scaffold promotes axon regeneration, attenuates inflammation, and induces cell migration in rat and canine with spinal cord injury.

Persistent neurotrophic factor delivery is crucial to create a microenvironment for cell survival and nerve regeneration in spinal cord injury (SCI). This study aimed to develop a NT-3/fibroin coated gelatin sponge scaffold (NF-GS) as a novel controlled artificial release therapy for SCI. In vitro, bone marrow-derived mesenchymal stem cells (MSCs) were planted into the NF-GS and release test showed that NF-GS was capable to generate a sustainable NT-3 release up to 28 days. MSCs in NF-GS had high cell activity with excellent cell distribution and phenotype. Then, the NF-GS was transplanted into the injury site of spinal cord of rat and canine in vivo, which exhibited strong biocompatibility during post-transplantation period. Four weeks following transplantation, the concentration of NT-3 was much higher than that in control groups. Cavity areas in the injury/graft site were significantly reduced due to tissue regeneration and axonal extensions associated with myelin sheath through the glial scar into the NF-GS. Additionally, the NF-GS decreased the inflammation by reducing the CD68 positive cells and TNF-α. A striking feature was the occurrence of some cells and myelin-like structure that appeared to traverse the NF-GS. The present results demonstrate that the NF-GS has the property to control the release of NT-3 from the NT-3/fibroin complex thus facilitating regeneration of injured spinal cord.

Regulation of human mesenchymal stem cell differentiation by TREM-2.

Activation of the triggering receptor expressed on myeloid cells 2 (TREM-2) regulates myeloid cell function in vitro. However, the failure to detect TREM-2 protein expression in vivo has hampered studies on immunological and other physiological TREM-2 functions. This study demonstrates that TREM-2 is expressed by human mesenchymal stem cells (h-MSCs) and responds to the toll-like receptor (TLR) ligand lipopolysaccharide (LPS). Knockdown of TREM-2 in h-MSCs using a small interfering RNA (siRNA) reduced the expression levels of TLR2, TLR4, and TLR6, inhibited osteogenic, chondrogenic, and adipogenic differentiation under specific induction conditions, and enhanced LPS-evoked inflammatory cytokine production. Thus, activation of TREM-2 may restrain h-MSC immune activation and promote differentiation for tissue repair.

Cholera Toxin B Subunit Shows Transneuronal Tracing after Injection in an Injured Sciatic Nerve.

Cholera toxin B subunit (CTB) has been extensively used in the past for monosynaptic mapping. For decades, it was thought to lack the ability of transneuronal tracing. In order to investigate whether biotin conjugates of CTB (b-CTB) would pass through transneurons in the rat spinal cord, it was injected into the crushed left sciatic nerve. For experimental control, the first order afferent neuronal projections were defined by retrograde transport of fluorogold (FG, a non-transneuronal labeling marker as an experimental control) injected into the crushed right sciatic nerve in the same rat. Neurons containing b-CTB or FG were observed in the dorsal root ganglia (DRG) at the L4-L6 levels ipsilateral to the tracer injection. In the spinal cord, b-CTB labeled neurons were distributed in all laminae ipsilaterally between C7 and S1 segments, but labeling of neurons at the cervical segment was abolished when the T10 segment was transected completely. The interneurons, distributed in the intermediate gray matter and identified as gamma-aminobutyric acid-ergic (GABAergic), were labeled by b-CTB. In contrast, FG labeling was confined to the ventral horn neurons at L4-L6 spinal segments ipsilateral to the injection. b-CTB immunoreactivity remained to be restricted to the soma of neurons and often appeared as irregular patches detected by light and electron microscopy. Detection of monosialoganglioside (GM1) in b-CTB labeled neurons suggests that GM1 ganglioside may specifically enhance the uptake and transneuronal passage of b-CTB, thus supporting the notion that it may be used as a novel transneuronal tracer.

Modified total en bloc spondylectomy for thoracolumbar spinal tumors via a single posterior approach.

PURPOSE: The objectives of this study were to describe our surgical management with a modified total en bloc spondylectomy (TES) and to evaluate the clinical effects in patients with thoracolumbar tumors. METHODS: Sixteen consecutive patients with thoracolumbar neoplasms underwent a modified TES via single posterior approach followed by dorsoventral reconstruction from December 2008 to July 2011. Details of the modified technique were described and the patients' clinical information was retrospectively reviewed and analyzed. RESULTS: Significant improvements in neurological function were achieved in most of the patients. Local pain or radicular leg pain was relieved postoperatively. The mean operation time was 7.2 h, with an average blood loss of 2,300 ml. No major complications, instrumentation failure or local recurrence was found at the final follow-up. Five patients died of the disease during mean 14-month (3.0-23) follow-up. CONCLUSIONS: The modified TES with a single posterior approach is feasible, safe and effective for thoracolumbar spine tumors.

Differentiation of human bone marrow precursor cells into neuronal-like cells after transplantation into canine spinal cord organotypic slice cultures.

BACKGROUND: Treatments to regenerate different tissue involving the transplantation of bone marrow derived mesenchymal precursor cells are anticipated. Using an alternative methods, in vitro organotypic slice culture method, would be useful to transplant cells and assessing the effects. This study was to determine the possibility of differentiating human bone marrow precursor cells into cells of the neuronal lineage by transplanting into canine spinal cord organotypic slice cultures. METHODS: Bone marrow aspirates were obtained from posterior superior iliac spine (PSIS) of patients that had undergone spinal fusion due to a degenerative spinal disorder. For cell imaging, mesenchymal precursor cells (MPCs) were pre-stained with PKH-26 just before transplantation to canine spinal cord slices. Canine spinal cord tissues were obtained from three adult beagle dogs. Spinal cords were cut into transverse slices of 1 mm using tissue chopper. Two slices were transferred into 6-well plate containing 3 ml DMEM with antibiotics. Prepared MPCs (1×10(4)) were transplanted into spinal cord slices. On days 0, 3, 7, 14, MPCs were observed for morphological changes and expression of neuronal markers through immunofluorescence and reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: The morphological study showed: spherical cells in the control and experiment groups on day 0; and on day 3, cells in the control group had one or two thick, short processes and ones in the experiment group had three or four thin, long processes. On day 7, these variously-sized processes contacted each other in the experiment group, but showed typical spindle-shaped cells in the control group. Immunofluorescence showed that PKH-26(+) MPCs stained positive for NeuN(+) and GFAP(+) in experimental group only. Also RT-PCR showed weak expression of ß-tubulin III and GFAP. CONCLUSIONS: Human bone marrow mesenchymal precursor cells (hMPCs) have the potential to differentiate into the neuronal like cells in this canine spinal cord organotypic slice culture model. Furthermore, these findings suggested the possibility that these cells can be utilized to treat patients with spinal cord injuries.

The clinical value of combined use of MR imaging and multi-slice spiral CT in limb salvage surgery for orthopaedic oncology patients: initial experience in nine patients.

BACKGROUND: The purpose of this prospective study was to evaluate the value of the combined use of MR imaging and multi-slice spiral CT for limb salvage surgery in orthopaedic oncology patients. PATIENTS AND METHODS: Nine consecutive patients with lower/upper limb malignant bone tumours (7 osteosarcomas and 2 chondrosarcomas) were treated with limb-salvaging procedures. Preoperative planning including determination of the osteotomy plane and diameters of the prosthesis was performed basing on the preoperative CT and MR images. The histopathology was performed as golden diagnostic criteria to evaluate the accuracy of CT and MR-based determination for tumour's boundary. RESULTS: The tumour extension measured on MRI was consistent with the actual extension (P>0.05, paired Student's t test), while the extension measured on CT imaging was less than the actual extension. The length, offset and alignment of the affected limb were reconstructed accurately after the operation. An excellent functional outcome was achieved in all patients. CONCLUSIONS: In the present study, MRI was found to be superior to CT for determining the tumour extension, combined use of MRI and CT measurement provided high precision for the fit of the prosthesis and excellent functional results.

Analysis of methylprednisolone-induced inhibition on the proliferation of neural progenitor cells in vitro by gene expression profiling.

Recently, it has been proved that methylprednisolone has inhibition effect on the proliferation of endogenous neural progenitor cells (NPCs) after spinal cord injury (SCI). Similar effect has also been found on NPCs cultured in vitro. However, the mechanism remains to be fully delineated. The purpose of this study is to investigate the potential molecular mechanism of this effect in NPCs cultured in vitro by gene expression profiling. Fetal mouse brain-derived NPCs were divided into 2 groups: NPCs incubated with methylprednisolone as a model of the methylprednisolone treatment after SCI, and without methylprednisolone as the control group. After the cell quantitative analysis and CCK-8 assay, the microarray analysis was carried out. Genes differentially expressed between NPCs treated with and without methylprednisolone were extracted. It was observed that the expression of 143 genes, including many members of distinct families, such as hypoxia inducible factors and neurotransmitter receptors, were significantly changed in response to the methylprednisolone treatment. Our results provide global molecular insights into the mechanisms of methylprednisolone-induced proliferation inhibition effect and suggest that EdnrB may play an important role in this effect.

Biomechanical stability of lower cervical spine immediately after discectomy with grafting.

OBJECTIVE: Anterior cervical discectomy is commonly used to treat radiculopathy and myelopathy. Although the size of the implanted graft may influence the clinical outcome of anterior reconstruction of the cervical spine, the ideal graft height remains arguable. The objective of the current study was to study the interrelations of graft height and immediate biomechanical stability in an anterior cervical discectomy model. METHODS: Six fresh-frozen human cadaver cervical spines (C1-T1) were tested in five sequential states. The first state tested was the "normal" state (specimens with intact discs). The other four states were tested after C5-C6 discectomy by the Smith-Robinson graft technique, using graft thicknesses of 100%, 120%, 140%, and 160% of the baseline height. The baseline height was defined as the intervertebral disc height of C5-C6 at the intact stage. Intervertebral segment flexion, extension, bending and rotation of C5-C6 were recorded using a 3D laser scanner and analyzed using Geomagic Studio 5.0 software. RESULTS: Bone grafting at 100% baseline height after discectomy provided the least stability and the greatest movement range. With increasing height of grafts, the movement range of the cervical spine declined. Immediate stability of the operated segments was significantly increased by grafting with 140% and 160% baseline heights compared to the baseline height condition. CONCLUSIONS: Strut-graft with appropriate distraction after Smith-Robinson anterior cervical discectomy plays an important role in the whole immediate biomechanical stability of the lower cervical spine. A graft height of 40% greater than baseline may be ideal after single discectomy in clinical practice.

Rosiglitazone enhances the proliferation of neural progenitor cells and inhibits inflammation response after spinal cord injury.

It has been previously shown that peroxisome proliferators-activated receptor gamma (PPAR-γ) is beneficial for nervous system injury. In present study, we examined the effect of rosiglitazone, a PPAR-γ agonist, on spinal cord injury (SCI) in rats. SCI was induced by dropping a 10g weight rod at a height of 25mm. The animals were randomly divided into vehicle group, rosiglitazone treated group, and G3335 treated group. Locomotor function recovery was evaluated by the Basso-Beattie-Bresnahan locomotor rating scale (BBB scale), NF-κB expression and endogenous neural progenitor cells (NPCs) proliferation and differentiation was assessed by flow cytometry and immunohistochemistry. Compared with the vehicle groups, we found that the rosiglitazone could significantly ameliorate locomotor recovery, reduce NF-κB expression, and increase the proliferation of endogenous NPCs. when the PPAR-γ antagonist was use, these effects were abolished. However, neurons differentiating from endogenous NPCs were inhibited when PPAR-γ was activated. Our results suggest that the activation of PPAR-γ may be a potential alternative treatment for spinal cord injury.