Objective: Our knowledge of human neural crest stem cells (NCSCs) is expanding, owing to recent advances in technologies utilizing human-induced pluripotent stem cells (hiPSCs) that generate NCSCs. However, the clinical application of these technologies requires the reduction of xeno-materials. To overcome this significant impediment, this study aimed to devise a novel method to induce NCSCs from hiPSCs without using a feeder cell layer.Materials and Methods: hiPSCs were cultured in feeder-free maintenance media containing the Rho-associated coiled-coil forming kinase inhibitor Y-27632. When the cells reached 50–70% confluence, differentiation was initiated by replacing the medium with knockout serum replacement (KSR) medium containing Noggin and SB431542. The KSR medium was then gradually replaced with increasing concentrations of Neurobasal medium from day 5 to 11.Results: Immunocytochemistry and flow cytometry were performed 12 days after induction of differentiation and revealed that the cells generated from hiPSCs expressed the NCSC markers p75 and HNK-1, but not the hiPSC marker SOX2.Conclusion: These findings demonstrate that hiPSCs were induced to differentiate into NCSCs in the absence of feeder cells.

STUDY DESIGN: A retrospective observational study was performed. PURPOSE: We investigated the prevalence of sarcopenia in dropped head syndrome (DHS), and the relationship between biochemical markers, including major advanced glycation end products (AGEs), pentosidine, and DHS in older women. OVERVIEW OF LITERATURE: AGEs have been implicated in the pathogenesis of sarcopenia. METHODS: We studied 13 elderly women with idiopathic DHS (mean age, 77.2 years) and 20 healthy volunteers (mean age, 74.8 years). We used a bioelectrical impedance analyzer to analyze body composition, including appendicular skeletal muscle mass index (SMI; appendicular lean mass [kg]/[height (m)]2). Cervical sagittal plane alignment, including C2–C7 sagittal vertical axis (C2–C7SVA), C2–C7 angle, and C2 slope (C2S), was measured. Biochemical markers, such as serum and urinary pentosidine, serum homocysteine, 1, 25-dihydroxyvitamin D, and 25-hydroxyvitamin D, were measured. The level of each variable was compared between DHS and controls. The relationship between biochemical markers and DHS was examined. RESULTS: Sarcopenia (SMI < 5.75) was observed at a high prevalence in participants with DHS (77% compared to 22% of healthy controls). Height, weight, femoral bone mineral density, appendicular lean mass, total lean mass, and SMI all had significantly lower values in the DHS group. Serum and urinary pentosidine, and serum homocysteine were significantly higher in the DHS group compared to controls. Analysis of cervical alignment revealed a significant positive correlation of serum pentosidine with C2–C7SVA and C2S. CONCLUSIONS: Sarcopenia was involved in DHS, and high serum pentosidine levels are associated with severity of DHS in older women.
Aged ; Biomarkers ; Body Composition ; Bone Density ; Electric Impedance ; Female ; Glycosylation End Products, Advanced ; Head ; Healthy Volunteers ; Homocysteine ; Humans ; Muscle, Skeletal ; Neck Muscles ; Observational Study ; Prevalence ; Retrospective Studies ; Sarcopenia

STUDY DESIGN: Retrospective observational study. PURPOSE: We considered the relationship between spinal alignment and skeletal muscle mass on clinical outcomes following a surgery for lumbar spinal stenosis (LSS). OVERVIEW OF LITERATURE: There are no reports of preoperative factors predicting residual low back pain following surgery for LSS. METHODS: Our target population included 34 women (mean age, 74.4 years) who underwent surgery for LSS. Prior to and 6 months after the surgery, systemic bone mineral density and lean soft tissue mass were measured using dual-energy X-ray absorptiometry. Skeletal muscle mass index (SMI) was calculated as the sum of the arm and leg lean mass in kilograms divided by height in meters squared. The spinal alignment was also measured. Clinical outcomes were evaluated using the Japanese Orthopedic Association scoring system, leg and low back pain Visual Analog Scale, and Roland–Morris Disability Questionnaire (RDQ). Additionally, we examined the bone mineral density, skeletal muscle mass, and spinal alignment before and after the surgery. We used the Spearman correlation coefficient to examine the associations among clinical outcomes, preoperative muscle mass, and spinal alignment. RESULTS: Sarcopenia (SMI <5.46) was observed in nine subjects (26.5%). Compared with normal subjects (SMI >6.12), RDQ was significantly higher in subjects with sarcopenia (p=0.04). RDQ was significantly negatively correlated with SMI (r=−0.42, p<0.05). There was a significant positive correlation between postoperative RDQ and pelvic tilt (PT; r=0.41, p<0.05). SMI and PT were significantly negatively correlated (r=−0.39, r<0.05). CONCLUSIONS: Good postoperative outcomes were negatively correlated with low preoperative appendicular muscle mass, suggesting that postoperative outcomes were inferior in cases of decreased appendicular muscle mass (sarcopenia). Posterior PT due to decreased limb muscle mass may contribute to postoperative back pain, showing that preoperatively reduced limb muscle mass and posterior PT are predictive factors in the persistence of postoperative low back pain.
Absorptiometry, Photon ; Arm ; Asian Continental Ancestry Group ; Back Pain ; Bone Density ; Extremities ; Female ; Health Services Needs and Demand ; Humans ; Leg ; Low Back Pain ; Muscle, Skeletal ; Observational Study ; Orthopedics ; Retrospective Studies ; Sarcopenia ; Spinal Stenosis ; Visual Analog Scale

OBJECTIVE: It is important to develop an easy means of diagnosing lumbar foraminal stenosis (LFS) in a general practice setting. We investigated the use of the Japanese Orthopaedic Association Back Pain Evaluation Questionnaire (JOABPEQ) to diagnose LFS in symptomatic patients. METHODS: Subjects included 13 cases (mean age, 72 years) with LFS, and 30 cases (mean age, 73 years) with lumbar spinal canal stenosis (LSCS) involving one intervertebral disc. The visual analogue scale score for low back pain and leg pain, the JOABPEQ were evaluated. RESULTS: Those with LFS had a significantly lower JOA score (p<0.001), while JOABPEQ scores (p<0.05) for lumbar dysfunction and social functioning impairment (p<0.01) were both significantly lower than the scores in LSCS. The following JOABPEQ questionnaire items (LFS vs. LSCS, p-value) for difficulties in: sleeping (53.8% vs. 16.6%, p<0.05), getting up from a chair (53.8% vs. 6.6%, p<0.001), turning over (76.9% vs. 40%, p<0.05), and putting on socks (76.9% vs. 26.6%, p<0.01) such as pain during rest, and signs of intermittent claudication more than 15 minutes (61.5% vs. 26.6%, p<0.05) were all significantly more common with LFS than LSCS. CONCLUSION: Results suggest that of the items in the JOABPEQ, if pain during rest or intermittent claudication is noted, LFS should be kept in mind as a cause during subsequent diagnosis and treatment. LFS may be easily diagnosed from LSCS using this established patient-based assessment method.
Asian Continental Ancestry Group* ; Back Pain* ; Constriction, Pathologic* ; Diagnosis ; General Practice ; Humans ; Intermittent Claudication ; Intervertebral Disc ; Leg ; Low Back Pain ; Methods ; Spinal Canal

STUDY DESIGN: Retrospective case series. PURPOSE: The purpose of this study was to examine changes in the ligamentum flavum thickness and remodeling of the spinal canal after anterior fusion during a 10-year follow-up. OVERVIEW OF LITERATURE: Extreme lateral interbody fusion provides minimally invasive treatment of the lumbar spine; this anterior fusion without direct posterior decompression, so-called indirect decompression, can achieve pain relief. Anterior fusion may restore disc height, stretch the flexure of the ligamentum flavum, and increase the spinal canal diameter. However, changes in the ligamentum flavum thickness and remodeling of the spinal canal after anterior fusion during a long follow-up have not yet been reported. METHODS: We evaluated 10 patients with L4 spondylolisthesis who underwent stand-alone anterior interbody fusion using the iliac crest bone. Magnetic resonance imaging was performed 10 years after surgery. The cross-sectional area (CSA) of the dural sac and the ligamentum flavum at L1–2 to L5–S1 was calculated using a Picture Archiving and Communication System. RESULTS: Spinal fusion with correction loss (average, 4.75 mm anterior slip) was achieved in all patients 10 years postsurgery. The average CSAs of the dural sac and the ligamentum flavum at L1–2 to L5–S1 were 150 mm² and 78 mm², respectively. The average CSA of the ligamentum flavum at L4–5 (30 mm²) (fusion level) was significantly less than that at L1–2 to L3–4 or L5–S1. Although patients had an average anterior slip of 4.75 mm, the average CSA of the dural sac at L4–5 was significantly larger than at the other levels. CONCLUSIONS: Spinal stability induced a lumbar ligamentum flavum change and a sustained remodeling of the spinal canal, which may explain the long-term pain relief after indirect decompression fusion surgery.
Decompression* ; Follow-Up Studies ; Humans ; Ligamentum Flavum* ; Magnetic Resonance Imaging ; Retrospective Studies ; Spinal Canal ; Spinal Fusion ; Spine ; Spondylolisthesis

STUDY DESIGN: Animal model study. PURPOSE: The purpose of this study was to evaluate the histological variation in the injured muscle and production of calcitonin gene-related peptide in rats over time. OVERVIEW OF LITERATURE: Vertebral surgery has been reported to cause atrophy of the back muscles, which may result in pain. However, few reports have described the time series histological variation in the injured muscle and changes in the dominant nerve. METHODS: We used 30 male, 8-week-old Sprague-Dawley rats. The right and left sides of the paravertebral muscle were considered as the injured and uninjured sides, respectively. A 115 g weight was dropped from a height of 1 m on the right paravertebral muscle. Hematoxylin and eosin (H&E) staining of the muscle was performed 1–3 weeks after injury for histological evaluation. Fluoro-Gold (FG) was injected into the paravertebral muscle. The L2 dorsal root ganglia on both sides were resected 1, 2, and 3 weeks after injury, and immunohistochemical staining for calcitonin gene-related peptide was performed. RESULTS: H&E staining of the paravertebral muscle showed infiltration of inflammatory cells and the presence of granulation tissue in the injured part on the ipsilateral side 1 week after injury. Muscle atrophy occurred 3 weeks after injury, but was repaired via spontaneous replacement of muscle cells/fibers. In contrast, compared with the uninjured side, the percentage of cells double-labeled with FG and calcitonin gene-related peptide in FG-positive cells in the dorsal root ganglia of the injured side was significantly increased at each time point throughout the study period. CONCLUSIONS: These results suggest that sensitization of the dominant nerve in the dorsal root ganglia, which may be caused by cicatrix formation, can protract injured muscle pain. This information may be helpful in elucidating the underlying mechanism of persistent pain after back muscle injury.
Animals ; Atrophy ; Back Muscles* ; Calcitonin Gene-Related Peptide ; Cicatrix ; Eosine Yellowish-(YS) ; Ganglia, Sensory ; Ganglia, Spinal ; Granulation Tissue ; Hematoxylin ; Humans ; Male ; Models, Animal ; Muscular Atrophy ; Myalgia ; Rats* ; Rats, Sprague-Dawley ; Spine

STUDY DESIGN: An experimental animal study. PURPOSE: To evaluate effects of anti-vascular endothelial growth factor (VEGF) on the content and distribution of the calcitonin gene-related peptide (CGRP) in the dorsal ganglia in a rat model. OVERVIEW OF LITERATURE: Increased expression of VEGF in degenerative disc disease increases the levels of inflammatory cytokines and nerve ingrowth into the damaged discs. In animal models, increased levels of VEGF can persist for up to 2 weeks after an injury. METHODS: Through abdominal surgery, the dorsal root ganglia (DRG) innervating L5/L6 intervertebral disc were labeled (FluoroGold neurotracer) in 24, 8-week old Sprague Dawley rats. The rats were randomly allocated to three groups of eight rats each. The anti-VEGF group underwent L5/6 intervertebral disc puncture using a 26-gauge needle, intradiscal injection of 33.3 µg of the pegaptanib sodium, a VEGF165 aptamer. The control-puncture group underwent disc puncture and intradiscal injection of 10 µL saline solution, and the sham-surgery group underwent labeling but no disc puncture. Two rats in each group were sacrificed on postoperative days 1, 7, 14, and 28 after surgery. L1–L6 DRGs were harvested, sectioned, and immunostained to detect the content and distribution of CGRP. RESULTS: Compared with the control, the percentage of CGRP-positive cells was lower in the anti-VEGF group (p<0.05; 40.6% and 58.1% on postoperative day 1, 44.3% and 55.4% on day 7, and 42.4% and 59.3% on day 14). The percentage was higher in the control group compared with that of the sham group (p<0.05; sham group, 34.1%, 40.7%, and 33.7% on postoperative days 1, 7, and 14, respectively). CONCLUSIONS: Decreasing CGRP-positive cells using anti-VEGF therapy provides fundamental evidence for a possible therapeutic role of anti-VEGF in patients with discogenic lower back pain.
Animals ; Back Pain ; Calcitonin Gene-Related Peptide ; Cytokines ; Diagnosis-Related Groups ; Endothelial Growth Factors ; Ganglia ; Ganglia, Spinal* ; Humans ; Intervertebral Disc* ; Low Back Pain ; Models, Animal ; Needles ; Neuropeptides* ; Punctures ; Rats* ; Rats, Sprague-Dawley ; Sodium ; Sodium Chloride ; Spinal Nerve Roots* ; Vascular Endothelial Growth Factor A*

STUDY DESIGN: Observational study. PURPOSE: To assess the correlation among inflammatory cytokine expression levels, degree of intervertebral disk (IVD) degeneration, and predominant clinical symptoms observed in degenerative disk disease (DDD). OVERVIEW OF LITERATURE: Low back pain (LBP) is associated with inflammatory cytokine expression levels, including those of tumor necrosis factor-alpha (TNF-α), interleukin 6 (IL-6), and nerve growth factor (NGF). However, the association between cytokine expression levels and the physiological mechanisms of disk degeneration and clinical pain remain controversial. METHODS: Using the enzyme-linked immunosorbent assay, TNF-α, IL-6, and NGF expression levels were analyzed in 58 IVD samples that were harvested from patients with lumbar DDD. Patient samples were grouped according to the degree of IVD degeneration using the Pfirrmann grading system and magnetic resonance imaging, and the correlations between the disease groups and each cytokine expression level were assessed. In addition, on the basis of their predominant preoperative symptoms, the patients were assigned to either an LBP or leg pain group to determine the correlation among these disease manifestations and individual cytokine expression levels. RESULTS: A gradual increase in TNF-α (R=0.391) and IL-6 (R=0.388) expression levels correlated with the degree of IVD degeneration, whereas NGF (R=0.164) expression levels exhibited a minimal decrease with disease progression. Regarding the predominant clinical manifestation, only the LBP group exhibited a significant increase in TNF-α expression levels (p=0.002). CONCLUSIONS: These results suggested that TNF-α and IL-6 play an important role in the pathophysiology of IVD degeneration at any stage, whereas NGF plays an important role during the early disease stages. Moreover, because TNF-α expression levels were significantly high in the LBP group, we propose that they are involved in LBP onset or progression.
Dichlorodiphenyldichloroethane ; Disease Progression ; Enzyme-Linked Immunosorbent Assay ; Humans ; Interleukin-6 ; Intervertebral Disc Degeneration* ; Intervertebral Disc* ; Leg ; Low Back Pain ; Magnetic Resonance Imaging ; Nerve Growth Factor ; Observational Study ; Tumor Necrosis Factor-alpha

No abstract available.
Spondylolisthesis*

STUDY DESIGN: Controlled laboratory study. PURPOSE: This study aimed to evaluate the efficacy of platelet-rich plasma (PRP) stored at room temperature (RT), frozen, or after freeze-drying. OVERVIEW OF LITERATURE: PRP enriches tissue repair and regeneration, and is a novel treatment option for musculoskeletal pathologies. However, whether biological activity is preserved during PRP storage remains uncertain. METHODS: PRP was prepared from blood of 12 healthy human volunteers (200 mL/person) and stored using three methods: PRP was stored at RT with shaking, PRP was frozen and stored at −80℃, or PRP was freeze-dried and stored at RT. Platelet counts and growth factor content were examined immediately after preparation, as well as 2, 4, and 8 weeks after storage. Platelet activation rate was quantified by flow cytometry. RESULTS: Platelet counts were impossible to determine in many RT samples after 2 weeks, but they remained at constant levels in frozen and freeze-dried samples, even after 8 weeks of storage. Flow cytometry showed approximately 80% activation of the platelets regardless of storage conditions. Almost no growth factors were detected in the RT samples after 8 weeks, while low but significant expression was detected in the frozen and freeze-dried PRP. Over time, the mean relative concentrations of various growth factors decreased significantly or disappeared in the RT group. In the frozen group, levels were maintained for 4 weeks, but decreased significantly by 8 weeks (p <0.05). The freeze-dried group maintained baseline levels of growth factors for the entire 8-week duration. CONCLUSIONS: Freeze-drying enables PRP storage while maintaining bioactivity and efficacy for extended periods.
Blood Preservation ; Flow Cytometry ; Freeze Drying ; Healthy Volunteers ; Humans* ; Intercellular Signaling Peptides and Proteins ; Pathology ; Platelet Activation ; Platelet Count ; Platelet-Rich Plasma* ; Regeneration