Short- versus long-segment posterior spinal fusion with vertebroplasty for osteoporotic vertebral collapse with neurological impairment in thoracolumbar spine: a multicenter study.

BACKGROUND: Vertebroplasty with posterior spinal fusion (VP + PSF) is one of the most widely accepted surgical techniques for treating osteoporotic vertebral collapse (OVC). Nevertheless, the effect of the extent of fusion on surgical outcomes remains to be established. This study aimed to evaluate the surgical outcomes of short- versus long-segment VP + PSF for OVC with neurological impairment in thoracolumbar spine. METHODS: We retrospectively collected data from 133 patients (median age, 77 years; 42 men and 91 women) from 27 university hospitals and their affiliated hospitals. We divided patients into two groups: a short-segment fusion group (S group) with 2- or 3-segment fusion (87 patients) and a long-segment fusion group (L group) with 4- through 6-segment fusion (46 patients). Surgical invasion, clinical outcomes, local kyphosis angle (LKA), and complications were evaluated. RESULTS: No significant differences between the two groups were observed in terms of neurological recovery, pain scale scores, and complications. Surgical time was shorter and blood loss was less in the S group, whereas LKA at the final follow-up and correction loss were superior in the L group. CONCLUSION: Although less invasiveness and validity of pain and neurological relief are secured by short-segment VP + PSF, surgeons should be cautious regarding correction loss.

Effect of bisphosphonates or teriparatide on mechanical complications after posterior instrumented fusion for osteoporotic vertebral fracture: a multi-center retrospective study.

BACKGROUND: The optimal treatment of osteoporosis after reconstruction surgery for osteoporotic vertebral fractures (OVF) remains unclear. In this multicentre retrospective study, we investigated the effects of typically used agents for osteoporosis, namely, bisphosphonates (BP) and teriparatide (TP), on surgical results in patients with osteoporotic vertebral fractures. METHODS: Retrospectively registered data were collected from 27 universities and affiliated hospitals in Japan. We compared the effects of BP vs TP on postoperative mechanical complication rates, implant-related reoperation rates, and clinical outcomes in patients who underwent posterior instrumented fusion for OVF. Data were analysed according to whether the osteoporosis was primary or glucocorticoid-induced. RESULTS: A total of 159 patients who underwent posterior instrumented fusion for OVF were included. The overall mechanical complication rate was significantly lower in the TP group than in the BP group (BP vs TP: 73.1% vs 58.2%, p = 0.045). The screw backout rate was significantly lower and the rates of new vertebral fractures and pseudoarthrosis tended to be lower in the TP group than in the BP group. However, there were no significant differences in lumbar functional scores and visual analogue scale pain scores or in implant-related reoperation rates between the two groups. The incidence of pseudoarthrosis was significantly higher in patients with glucocorticoid-induced osteoporosis (GIOP) than in those with primary osteoporosis; however, the pseudoarthrosis rate was reduced by using TP. The use of TP also tended to reduce the overall mechanical complication rate in both primary osteoporosis and GIOP. CONCLUSIONS: The overall mechanical complication rate was lower in patients who received TP than in those who received a BP postoperatively, regardless of type of osteoporosis. The incidence of pseudoarthrosis was significantly higher in patients with GIOP, but the use of TP reduced the rate of pseudoarthrosis in GIOP patients. The use of TP was effective to reduce postoperative complications for OVF patients treated with posterior fusion.

Surgical outcomes of spinal fusion for osteoporotic thoracolumbar vertebral fractures in patients with Parkinson's disease: what is the impact of Parkinson's disease on surgical outcome?

BACKGROUND: To date, there have been little published data on surgical outcomes for patients with PD with thoracolumbar OVF. We conducted a retrospective multicenter study of registry data to investigate the outcomes of fusion surgery for patients with Parkinson's disease (PD) with osteoporotic vertebral fracture (OVF) in the thoracolumbar junction. METHODS: Retrospectively registered data were collected from 27 universities and their affiliated hospitals in Japan. In total, 26 patients with PD (mean age, 76 years; 3 men and 23 women) with thoracolumbar OVF who underwent spinal fusion with a minimum of 2 years of follow-up were included (PD group). Surgical invasion, perioperative complications, radiographic sagittal alignment, mechanical failure (MF) related to instrumentation, and clinical outcomes were evaluated. A control group of 296 non-PD patients (non-PD group) matched for age, sex, distribution of surgical procedures, number of fused segments, and follow-up period were used for comparison. RESULTS: The PD group showed higher rates of perioperative complications (p < 0.01) and frequency of delirium than the non-PD group (p < 0.01). There were no significant differences in the degree of kyphosis correction, frequency of MF, visual analog scale of the symptoms, and improvement according to the Japanese Orthopaedic Association scoring system between the two groups. However, the PD group showed a higher proportion of non-ambulators and dependent ambulators with walkers at the final follow-up (p < 0.01). CONCLUSIONS: A similar surgical strategy can be applicable to patients with PD with OVF in the thoracolumbar junction. However, physicians should pay extra attention to intensive perioperative care to prevent various adverse events and implement a rehabilitation regimen to regain walking ability.

Surgical outcomes of spinal fusion for osteoporotic vertebral fracture in the thoracolumbar spine: Comprehensive evaluations of 5 typical surgical fusion techniques.

BACKGROUND: A consensus on the optimal surgical procedure for thoracolumbar OVF has yet to be reached due to the previous relatively small number of case series. The study was conducted to investigate surgical outcomes for osteoporotic vertebral fracture (OVF) in the thoracolumbar spine. METHODS: In total, 315 OVF patients (mean age, 74 years; 68 men and 247 women) with neurological symptoms who underwent spinal fusion with a minimum 2-year follow-up were included. The patients were divided into 5 groups by procedure: anterior spinal fusion alone (ASF group, n = 19), anterior/posterior combined fusion (APSF group, n = 27), posterior spinal fusion alone (PSF group, n = 40), PSF with 3-column osteotomy (3CO group, n = 92), and PSF with vertebroplasty (VP + PSF group, n = 137). RESULTS: Mean operation time was longer in the APSF group (p < 0.05), and intraoperative blood loss was lower in the VP + PSF group (p < 0.05). The amount of local kyphosis correction was greater in the APSF and 3CO groups (p < 0.05). Clinical outcomes were approximately equivalent among all groups. CONCLUSION: All 5 procedures resulted in acceptable neurological outcomes and functional improvement in walking ability. Moreover, they were similar with regard to complication rates, prevalence of mechanical failure related to the instrumentation, and subsequent vertebral fracture. Individual surgical techniques can be adapted to suit patient condition or severity of OVF.

Complications after spinal fixation surgery for osteoporotic vertebral collapse with neurological deficits: Japan Association of Spine Surgeons with ambition multicenter study.

BACKGROUND: There have been few reports on the incidence and risk factors of the complications after spinal fixation surgery for osteoporotic vertebral collapse (OVC) with neurological deficits. This study aimed to identify the incidence and risk factors of the complications after OVC surgery. METHODS: In this retrospective multicenter study, a total of 403 patients (314 women and 89 men; mean age 73.8 years) who underwent spinal fixation surgery for OVC with neurological deficits between 2005 and 2014 were enrolled. Data on patient demographics were collected, including age, sex, body mass index, smoking, steroid use, medical comorbidities, and surgical procedures. All postoperative complications that occurred within 6 weeks were recorded. Patients were classified into two groups, namely, complication group and no complication group, and risk factors for postoperative complications were investigated by univariate and multivariate analyses. RESULTS: Postoperative complications occurred in 57 patients (14.1%), and the most common complication was delirium (5.7%). In the univariate analysis, the complication group was found to be older (p = 0.039) and predominantly male (p = 0.049), with higher occurrence rate of liver disease (p = 0.001) and Parkinson's disease (p = 0.039) compared with the no-complication group. In the multivariate analysis, the significant independent risk factors were age (p = 0.021; odds ratio [OR] 1.051, 95% confidence interval [CI] 1.007-1.097), liver disease (p < 0.001; OR 8.993, 95% CI 2.882-28.065), and Parkinson's disease (p = 0.009; OR 3.636, 95% CI 1.378-9.599). CONCLUSIONS: Complications after spinal fixation surgery for OVC with neurological deficits occurred in 14.1%. Age, liver disease, and Parkinson's disease were demonstrated to be independent risk factors for postoperative complications.

Metatarsal Periosteal Plantar Ganglion Cyst Associated With Stress Fracture: A Case Report.

We encountered a rare case of a periosteal ganglion cyst at the plantar aspect of the metatarsal that induced a stress fracture in a 77-year-old female. The clinical manifestation of the plantar ganglion cyst of the foot was not evident because of its location deep beneath the plantar fascia. A pressure cortical indentation was detected at the metatarsal neck on the initial radiographs. Magnetic resonance imaging showed a stress fracture of the metatarsal with a ganglion cyst. The stress fracture was thought to have resulted from several factors, including structural weakness due to bony absorption from the ganglion cyst, osteoporosis that induced a fragility fracture, and a load-induced fatigue fracture. The fracture completely healed following complete resection of the ganglion cyst with the surrounding periosteum along with medication for osteoporosis. When confirmation of a stress fracture is necessary or when presentation of a stress fracture is atypical, magnetic resonance imaging should be considered to confirm or rule out any other associated pathologic features. Resection of the periosteal ganglion cyst with the surrounding periosteum is important to prevent recurrence.

Homing of vertebral-delivered mesenchymal stromal cells for degenerative intervertebral discs repair - an in vivo proof-of-concept study.

Introduction: Cell transplantation shows promising results for intervertebral disc (IVD) repair, however, contemporary strategies present concerns regarding needle puncture damage, cell retention, and straining the limited nutrient availability. Mesenchymal stromal cell (MSC) homing is a natural mechanism of long-distance cellular migration to sites of damage and regeneration. Previous ex vivo studies have confirmed the potential of MSC to migrate over the endplate and enhance IVD-matrix production. In this study, we aimed to exploit this mechanism to engender IVD repair in a rat disc degeneration model. Methods: Female Sprague Dawley rats were subjected to coccygeal disc degeneration through nucleus pulposus (NP) aspiration. In part 1; MSC or saline was transplanted into the vertebrae neighboring healthy or degenerative IVD subjected to irradiation or left untouched, and the ability to maintain the IVD integrity for 2 and 4 weeks was assessed by disc height index (DHI) and histology. For part 2, ubiquitously GFP expressing MSC were transplanted either intradiscally or vertebrally, and regenerative outcomes were compared at days 1, 5, and 14 post-transplantation. Moreover, the homing potential from vertebrae to IVD of the GFP+ MSC was assessed through cryosection mediated immunohistochemistry. Results: Part 1 of the study revealed significantly improved maintenance of DHI for IVD vertebrally receiving MSC. Moreover, histological observations revealed a trend of IVD integrity maintenance. Part 2 of the study highlighted the enhanced DHI and matrix integrity for discs receiving MSC vertebrally compared with intradiscal injection. Moreover, GFP rates highlighted MSC migration and integration in the IVD at similar rates as the intradiscally treated cohort. Conclusion: Vertebrally transplanted MSC had a beneficial effect on the degenerative cascade in their neighboring IVD, and thus potentially present an alternative administration strategy. Further investigation will be needed to determine the long-term effects, elucidate the role of cellular homing versus paracrine signaling, and validate our observations on a large animal model.

The Surgical Outcomes of Spinal Fusion for Osteoporotic Vertebral Fractures in the Lower Lumbar Spine with a Neurological Deficit.

INTRODUCTION: Osteoporotic vertebral fracture (OVF) is the most common osteoporotic fracture, and some patients require surgical intervention to improve their impaired activities of daily living with neurological deficits. However, many previous reports have focused on OVF around the thoracolumbar junction, and the surgical outcomes of lumbar OVF have not been thoroughly discussed. We aimed to investigate the surgical outcomes for lumbar OVF with a neurological deficit. METHODS: Patients who underwent fusion surgery for thoracolumbar OVF with a neurological deficit were enrolled at 28 institutions. Clinical information, comorbidities, perioperative complications, Japanese Orthopaedic Association scores, visual analog scale scores, and radiographic parameters were compared between patients with lower lumbar fracture (L3-5) and those with thoracolumbar junction fracture (T10-L2). Each patient with lower lumbar fracture (L group) was matched with to patients with thoracolumbar junction fracture (T group). RESULTS: A total 403 patients (89 males and 314 females, mean age: 73.8 ± 7.8 years, mean follow-up: 3.9 ± 1.7 years) were included in this study. Lower lumbar OVF was frequently found in patients with lower bone mineral density. After matching, mechanical failure was more frequent in the L group (L group: 64%, T group: 39%; p < 0.001). There was no difference between groups in the clinical and radiographical outcomes, although the rates of complication and revision surgery were still high in both groups. CONCLUSIONS: The surgical intervention for OVF is effective in patients with myelopathy or radiculopathy regardless of the surgical level, although further study is required to improve clinical and radiographical outcomes. LEVEL OF EVIDENCE: Level III.

Annulus fibrosus cell sheets limit disc degeneration in a rat annulus fibrosus injury model.

In recent years, studies have explored novel approaches for cell transplantation to enable annulus fibrosus (AF) regeneration of the intervertebral disc in particular for lumbar disc herniation. Nevertheless, successful engraftment of cells is structurally challenging, and no definitive method has yet been established. This study investigated the potential of cell sheet technology to facilitate cell engraftment for AF repair. AF injury was induced by a 1 × 1 mm defect in rat tails after which AF cell sheets were transplanted. Its regenerative effects were compared to a nondegenerated and degeneration only conditions. Degenerative changes of the entire intervertebral disc were examined by disc height measurements, histology, and immunohistochemistry for 4-, 8-, and 12-weeks post-transplantation. Cell engraftment was confirmed by tracing PKH26 fluorescent dyed AF cells. In the transplant group, disc degeneration was significantly suppressed after 4, 8, and 12 weeks when compared with the degenerative group, as indicated by histological scoring and DHI observations. At 2 and 4 weeks after transplant, PKH26 positive cells could be detected in defect region and surrounding AF. The results suggest cell engraftment into AF tissue could be established by the cell sheet technology without additional scaffolding or adhesives. In short, AF cell sheets appear to be an effective and accessible tool for AF repair and to support intervertebral disc regeneration.

Minimal Sustainability of Dedifferentiation by ROCK Inhibitor on Rat Nucleus Pulposus Cells In Vitro.

INTRODUCTION: Intervertebral disc degeneration is strongly associated with low back pain. Cell transplantation has been extensively studied as a treatment option for intervertebral disc degeneration. It is often necessary to perform cell culture prior to cell transplantation; however, during cell expansion, the cells tend to dedifferentiate and lose their potency. Although the ability to suppress dedifferentiation by ROCK inhibitor (ROCKi) has recently been reported for chondrocytes, its effects on nucleus pulposus cells are still largely unknown. METHODS: Rat nucleus pulposus cells were cultured with or without the addition of ROCKi (Y-27632), and cell proliferation; CD24 positivity; expression of SOX9, COL2A1, Aggrecan, and COL1A1; and cell redifferentiation ability in pellet culture were evaluated. RESULTS: Although the addition of ROCKi tended to slightly increase the cell proliferative capacity, no significant differences were observed between treated and untreated conditions. The addition of ROCKi showed a trend of minimally increased COL2A1, ACAN, and SOX9 expression. Increases in COL1A1 expression was slightly suppressed by ROCKi. In pellet culture, strong increase in type II collagen deposition was observed by the addition of ROCKi. The addition of ROCKi did not significantly change the levels of CD24 positivity. The supplementation of ROCKi did not significantly enhance nucleus pulposus cell marker expression during monolayer expansion. However, ROCKi addition did result in an increased type II collagen deposition in 3D pellet culture. CONCLUSIONS: Taken together, the results suggest a minimal effect by ROCKi on nucleus pulposus cell phenotype maintenance.

Changes in Spinal Alignment following eXtreme Lateral Interbody Fusion Alone in Patients with Adult Spinal Deformity using Computed Tomography.

This study examined the ability of the extreme lateral interbody fusion (XLIF) procedure to restore coronal and sagittal alignments for patients with adult spinal deformity (ASD) using computed tomography multiplanar reconstruction (CT-MPR). Thirty-eight patients with ASD undergoing correction and fixation with XLIF at 114 levels were studied. The coronal segmental Cobb angle, coronal regional Cobb angle (L1-5), sagittal segmental Cobb angle, sagittal regional Cobb angle (L1-5), intervertebral disc height and, vertebral body rotation (VBR) were measured before and after of XLIF surgery using CT-MPR. The mean sagittal segmental Cobb angle, the coronal segmental Cobb angle and VBR were corrected from 5.0° to 9.0°, from 6.3° to 4.3° and from 12.2° to 10.8°, respectively. The mean of the intervertebral disc heights increased significantly from 6.0 mm to 10.4 mm postoperatively. Although increases in coronal segmental Cobb, sagittal segmental Cobb, and intervertebral disc height at each level were significant, there were no significant differences in each parameter acquired by spine levels. The results also showed that it was difficult for L4/5 level to obtain the most postoperative coronal Cobb, sagittal Cobb and intervertebral disc height. This study evaluated the alignment improvement effect of stand-alone XLIF in ASD patients using CT-MPR. For the lower lumbar spine, it is difficult to obtain a lordosis more than 10 degrees with stand-alone XLIF for correcting ASD. Therefore, it is thought that correction such as osteotomy or compression technique to the posterior fusion may be necessary during the 2nd stage surgery.

Risk Factors for Proximal Junctional Fracture Following Fusion Surgery for Osteoporotic Vertebral Collapse with Delayed Neurological Deficits: A Retrospective Cohort Study of 403 Patients.

INTRODUCTION: Approximately 3% of osteoporotic vertebral fractures develop osteoporotic vertebral collapse (OVC) with neurological deficits, and such patients are recommended to be treated surgically. However, a proximal junctional fracture (PJFr) following surgery for OVC can be a serious concern. Therefore, the aim of this study is to identify the incidence and risk factors of PJFr following fusion surgery for OVC. METHODS: This study retrospectively analyzed registry data collected from facilities belonging to the Japan Association of Spine Surgeons with Ambition (JASA) in 2016. We retrospectively analyzed 403 patients who suffered neurological deficits due to OVC below T10 and underwent corrective surgery; only those followed up for　≥2 years were included. Potential risk factors related to the PJFr and their cut-off values were calculated using multivariate logistic regression analysis and receiver operating characteristic (ROC) analysis. RESULTS: Sixty-three patients (15.6%) suffered PJFr during the follow-up (mean 45.7 months). In multivariate analysis, the grade of osteoporosis (grade 2, 3: adjusted odds ratio (aOR) 2.92; p=0.001) and lower instrumented vertebra (LIV) level (sacrum: aOR 6.75; p=0.003) were independent factors. ROC analysis demonstrated that lumbar bone mineral density (BMD) was a predictive factor (area under curve: 0.72, p=0.035) with optimal cut-off value of 0.61 g/cm2 (sensitivity, 76.5%; specificity, 58.3%), but that of the hip was not (p=0.228). CONCLUSIONS: PJFr was found in 16% cases within 4 years after surgery; independent risk factors were severe osteoporosis and extended fusion to the sacrum. The lumbar BMD with cut-off value 0.61 g/cm2 may potentially predict PJFr. Our findings can help surgeons select perioperative adjuvant therapy, as well as a surgical strategy to prevent PJFr following surgery.

Surgical Treatment of Osteoporotic Vertebral Fracture with Neurological Deficit-A Nationwide Multicenter Study in Japan.

INTRODUCTION: The prevalence of patients with osteoporosis continues to increase in aging societies, including Japan. The first choice for managing osteoporotic vertebral compression fracture (OVF) is conservative treatment. Failure in conservative treatment for OVF may lead to non-union or vertebral collapse, resulting in neurological deficit and subsequently requiring surgical intervention. This multicenter nationwide study in Japan was conducted to comprehensively understand the outcomes of surgical treatments for OVF non-union. METHODS: This multicenter, retrospective study included 403 patients (89 males, 314 females, mean age 73.8 ± 7.8 years, mean follow-up 3.9 ± 1.7 years) with neurological deficit due to vertebral collapse or non-union after OVF at T10-L5 who underwent fusion surgery with a minimum 1-year follow-up. Radiological and clinical outcomes at baseline and at the final follow-up (FU) were evaluated. RESULTS: OVF was present at a thoracolumbar junction such as T12 (124 patients) and L1 (117 patients). A majority of OVF occurred after a minor trauma, such as falling down (55.3%) or lifting objects (8.4%). Short segment fusion, including affected vertebra, was conducted (mean 4.0 ± 2.0 vertebrae) with 256.8 minutes of surgery and 676.1 g of blood loss. A posterior approach was employed in 86.6% of the patients, followed by a combined anterior and posterior (8.7%), and an anterior (4.7%) approach. Perioperative complications and implant failures were observed in 18.1% and 41.2%, respectively. VAS scores of low back pain (74.7 to 30.8 mm) and leg pain (56.8 to 20.7 mm) improved significantly at FU. Preoperatively, 52.6% of the patients were unable to walk and the rate of non-ambulatory patients decreased to 7.5% at FU. CONCLUSIONS: This study demonstrated that substantial improvement in activity of daily living (ADL) was achieved by fusion surgery. Although there was a considerable rate of complications, fusion surgery is beneficial for elderly OVF patients with non-union.

Correlation analysis of sagittal alignment and skeletal muscle mass in patients with spinal degenerative disease.

We investigated how skeletal muscle mass (SMM) affects spinal sagittal balance (radiographic parameters) in symptomatic spinal patients. The first purpose of this study was to evaluate the body composition and the spinal sagittal alignment in symptomatic spinal patients. The second purpose of this study was to compare whether the body composition and the spinal sagittal alignment is different in patients with cervical spine disease and lumbar spine disease. We retrospectively evaluated 313 patients who were hospitalized for surgery to treat spinal degenerative disease, who were divided into cervical and lumbar spine disease groups. All patients underwent full-length standing whole-spine radiography and bioimpedance analysis (BIA) before surgery. We used standard measurements to assess the sagittal vertical axis (SVA), cervical lordosis (CL; C2-C7), lumbar lordosis (LL; T12-S1), thoracic kyphosis (TK; T5-12), pelvic incidence (PI), pelvic tilt (PT), and sacral slope (SS). We also analyzed radiological and body composition parameters, patient characteristics, and the correlation between SMM and each sagittal parameters. In the overall cohort, the mean age at the time of operation was 66.5 ± 15.3 years and 59.2% of the patients were men. The correlation coefficients (r) between SMM and PT were negative weak correlation (r = -0.343, P < 0.001). The correlation with SMM for other LL, PI, SS, and SVA was statistically significant, but the correlation was none. In addition, our results also suggested strong correlations (r > 0.5) between LL and SS (r = 0.744), between LL and SVA (r = -0.589), between PT and SS (r = -0.580), and LL and PT (r = -0.506). Fifty-seven patients (18.2%, cervical group) had cervical spine disease and 256 patients (81.8%, lumbar group) had lumbar spine disease. No significant differences in age, height, body weight, and body mass index were observed between the two groups. The SMM of patients with cervical and lumbar spine disease also did not differ significantly. In the lumbar group, correlations were found between SMM and PT (r = -0.288, P < 0.001), between SMM and LL (r = 0.179, P < 0.01), and between SMM and SS (r = 0.170, P < 0.01), while only PT (r = -0.480, P < 0.001) was negatively correlated with SMM in the cervical group. This analysis indicated that PT is the sagittal parameter most closely related to SMM in patients with the spinal degenerative disease. The SMM might be one of the important factors that influenced the posterior inclination of the pelvis in symptomatic spinal patients, especially in cervical spine disease.

Discogenic cell transplantation directly from a cryopreserved state in an induced intervertebral disc degeneration canine model.

A multitude of studies has indicated the potential of cell therapy as a method for intervertebral disc (IVD) regeneration. Transplantation of a variety of cells has been assessed and shown capable of deterring the rate of degeneration in animal models and in human clinical trials. In this study, a novel approach using human discogenic nucleus pulposus cells directly from their cryopreserved state was assessed. In an established canine disc degeneration model, the degeneration process was evaluated in IVDs receiving precultured discogenic cells, thawed-only discogenic cells, and a saline sham injection after induction of degeneration. Degeneration progression was followed over time by the evaluation of the disc height index (DHI). Finally, after 12 weeks, the manipulated and control discs were explanted, histologically stained, and scored. Treated discs demonstrated retained DHI values for all treatment options. Histologic evaluations demonstrated significant improvement of matrix features compared to the sham. Moreover, thawed-only cells function at least as well as precultured discogenic cells. In short, cell transplantation of human discogenic cells directly from their cryopreserved state can arrest disc height degeneration and maintain histological matrix features in a canine disc degeneration model. The presented work demonstrates the potential of an off-the-shelf cell therapy product to treat degenerative disc disease.

CD146 defines commitment of cultured annulus fibrosus cells to express a contractile phenotype.

Characterization of cells is important for facilitating cell-based therapies for degenerative diseases of intervertebral discs. For this purpose, we analyzed mouse annulus fibrosus cells by flowcytometory to detect phenotypic change in their primary cultures. After examination of sixteen cell surface proteins, we focused on CD146 that solely increased during culture expansion. CD146 is known to be a marker for mesenchymal stem cells and for their vascular smooth muscle commitment with expression of contractile phenotype enhanced by SM22α. We sorted CD146+ cells to elucidate their characteristics and the key factors that play a role in this change. Whole cell cultures showed the ability for tripotent differentiation toward mesenchymal lineages, whereas sorted CD146+ cells did not. Expression of CD146 was elevated by addition of transforming growth factor ß1, and sorted CD146+ cells expressed higher levels of mRNA for SM22α and Elastin than did CD146- cells. Morphologically, CD146+ cells more broadly deposited extracellular type I collagen than CD146- cells and showed filamentous actin bundles traversing their cytoplasm and cell-cell junctions. Moreover, CD146+ cells demonstrated significantly higher gel contraction properties than CD146- cells when they were embedded in collagen gels. Human annulus fibrosus CD146+ cells also showed higher contractility. Immunohistochemistry determined CD146+ cells localized to the outermost annulus layers of mouse intervertebral disc tissue with co-expression of SM22α. These results suggest that increment of CD146 expression indicates gradual change of cultured annulus fibrosus cells to express a contractile phenotype and that transforming growth factor ß1 enhances this cellular commitment. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:1361-1372, 2016.

Response to tumor necrosis factor-α mediated inflammation involving activation of prostaglandin E2 and Wnt signaling in nucleus pulposus cells.

The cyclooxygenase 2 (COX-2) product, prostaglandin E2 (PGE2 ), acts through a family of G protein-coupled receptors designated E-prostanoid (EP) receptors that mediate intracellular signaling by multiple pathways. However, it is not known whether crosstalk between tumor necrosis factor-α(TNF-α)-PGE2 -mediated signaling and Wnt signaling plays a role in the regulation of intervertebral disc (IVD) cells. In this study, we investigated the relationship between TNF-α-PGE2 signaling and Wnt signaling in IVD cells. TNF-α increased the expression of COX-2 in IVD cells. The EP receptors EP1, EP3, and EP4 were expressed in IVD cells, and TNF-α significantly increased PGE2 production. Stimulation with TNF-α also upregulated EP3 and EP4 mRNA and protein expression in IVD cells. The inductive effect of the EP3 and EP4 receptors on Topflash promoter activity was confirmed through gain- and loss-of-function studies using selective EP agonists and antagonists. PGE2 treatment activated Wnt-ß-catenin signaling through activation of EP3. We conclude that TNF-α-induced COX-2 and PGE2 stimulate Wnt signaling and activate Wnt target genes. Suppression of the EP3 receptor via TNF-α-PGE2 signaling seems to suppress IVD degeneration by controlling the activation of Wnt signaling. These findings may help identify the underlying mechanism and role of Wnt signaling in IVD degeneration.

A complex interaction between Wnt signaling and TNF-α in nucleus pulposus cells.

INTRODUCTION: Increased expression of the proinflammatory cytokine TNF-α in intervertebral discs (IVDs) leads to inflammation, which results in progressive IVD degeneration. We have previously reported that activation of Wnt-ß-catenin (hereafter called Wnt) signaling suppresses the proliferation of nucleus pulposus cells and induces cell senescence, suggesting that Wnt signaling triggers the process of degeneration of the IVD. However, it is not known whether cross talk between TNF-α and Wnt signaling plays a role in the regulation of nucleus pulposus cells. The goal of the present study was to examine the effect of the interaction between Wnt signaling and the proinflammatory cytokine TNF-α in nucleus pulposus cells. METHODS: Cells isolated from rat nucleus pulposus regions of IVDs were cultured in monolayers, and the expression and promoter activity of Wnt signaling and TNF-α were evaluated. We also examined whether the inhibition of Wnt signaling using cotransfection with Dickkopf (DKK) isoforms and Sclerostin (SOST) could block the effects of pathological TNF-α expression in nucleus pulposus cells. RESULTS: TNF-α stimulated the expression and promoter activity of Wnt signaling in nucleus pulposus cells. In addition, the activation of Wnt signaling by 6-bromoindirubin-3'-oxime (BIO), which is a selective inhibitor of glycogen synthase kinase 3 (GSK-3) activity that activates Wnt signaling, increased TNF-α expression and promoter activity. Conversely, the suppression of TNF-α promoter activity using a ß-catenin small interfering RNA was evident. Moreover, transfection with DKK-3, DKK-4, or SOST, which are inhibitors of Wnt signaling, blocked Wnt signaling-mediated TNF-α activation; these effects were not observed for DKK-1 or DKK-2. CONCLUSIONS: Here, we have demonstrated that Wnt signaling regulates TNF-α and that Wnt signaling and TNF-α form a positive-feedback loop in nucleus pulposus cells. The results of the present study provide in vitro evidence that activation of Wnt signaling upregulates the TNF-α expression and might cause the degeneration of nucleus pulposus cells. We speculate that blocking this pathway might protect nucleus pulposus cells against degeneration.