bluuebunny/crossref_metadata_2025_split huggingface dataset:
TGFbeta Cross-linked to Microcarriers as Injectable IVD Therapy in Dogs and Humans
Authors: Bertolo Alessandro , Steffen Frank , Arcolino Fanny , Capossela Simona , Taddei Anna Rita , Baur Martin , Poetzel Tobias , Stoyanov Jivko 
Published: April 2016

Introduction Studies designed to use large canine models and translate the results to human practice must take into account the ex vivo and in vitro differences in the bone marrow derived stem cells(MSCs) and their response to a covalently bound growth factor. Methods We analyze the activity of transforming growth factor-β1 (TGF-β1) covalently immobilized on microcarriers either by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide (EDC/NHS) or riboflavin/UV (RB/UV) light-mediated cross-linking. Collagen microcarriers cross-linked with TGF-β1 were used for chondrogenic differentiation of human or canine mesenchymal stem cells. Evaluation methods included cell viability test, chondrogenic marker expression (aggrecan and collagen type I and type II), histological detection of proteoglycans, and immunohistochemical analysis. Results Cross-linking strengthened the collagen structure of the microcarriers and reduced collagenase-mediated degradation. Chondrogenic differentiation of MSCs was induced by TGF-β1 cross-linked on microcarriers, promoting gene expression and protein accumulation of aggrecan and collagen type I and type II, as well as proteoglycans. Cross-linking by RB/UV enhanced chondrogenesis more than any other group. In addition, cross-linking reduced scaffold shrinkage exerted by MSCs during chondrogenesis, a desirable feature for microcarriers if used as tissue defect filler. Conclusion Cross-linking of TGF-β1 to collagen microcarriers supported chondrogenesis. Current veterinary clinical trial will show if such approach will lead a step closer to development of a cost-effective and locally acting device for cell-based therapy.
Compression Loading Induced Cellular Stress Response of Intervertebral Disc Cells in Organ Culture
Authors: Chooi Wai Hon , Chan Samantha C. W. , Gantenbein Benjamin , Chan Barbara P. 
Published: April 2016

Introduction Mechanical stress is often associated to interverterbal disc (IVD) degeneration and the effect of mechanical loading on IVD has been studied and reviewed.1,2 Previously, expression of heat shock proteins, HSP70 and HSP27 has been found in pathological discs.3 However, there is no direct evidence on whether IVD cells respond to the mechanical loading by expression of HSPs. The objective of this study is to investigate the stress response of IVD cells during compressive loading in an organ culture. Materials and Methods Fresh adult bovine caudal discs were cultured with compressive loading applied at physiological range. Effect of loading type (static and dynamic) and repeated loading (2 hours per day for 2 days) were studied. Nucleus pulposus (NP) and annulus fibrosus (AF) of the IVD were retrieved at different time points: right after loading and right after resting. Positive control discs were heat shocked (43°C). Cell activity was assessed and expression of stress response genes (HSP70 and HSF1) and matrix remodeling genes (ACAN, COL2, COL1, ADAMTS4, MMP3 and MMP13) were studied. Results Cell activity was maintained in all groups. Both NP and AF expressed high level of HSP70 in heat shock groups, confirming their expression in response to stress. In NP, expression of HSP70 was up-regulated after static loading and dynamic loading with higher fold change was observed after static loading. During repeated loading, HSP70 appeared to be upregulated right after loading and decreased after resting. Such trend was not observed in AF and HSF1 levels. Expressions of matrix remodeling genes did not change significantly with loading except ADAMTS4 decreased in AF during static loading. Conclusion This study demonstrated that NP cells upregulate expression of HSP70 in response to loading induced stress without changing cell activity and matrix remodeling significantly. Acknowledgments This project was funded by AO Spine (AOSPN) (grant number: SRN_2011_14) and a fellowship exchange award by AO Spine Scientific Research Network (SRN). References Chan SC, Ferguson SJ, Gantenbein-Ritter B. The effects of dynamic loading on the intervertebral disc. Eur Spine J 2011;20(11):1796–1812 Neidlinger-Wilke C, Galbusera F, Pratsinis H, et al. Mechanical loading of the intervertebral disc: from the macroscopic to the cellular level. Eur Spine J 2014;23(Suppl 3):S333–S343 Sharp CA, Roberts S, Evans H, Brown SJ. Disc cell clusters in pathological human intervertebral discs are associated with increased stress protein immunostaining. Eur Spine J 2009;18(11):1587–1594
Aberrant DNA Methylation Important Factor in Intervertebral Disc Pathophysiology
Authors: Chutkan Norman , Fulzele Sadanand 
Published: April 2016

Introduction Intervertebral disc (IVD) degeneration is one of the most important aging health problems and a leading cause of disability. However, little is known about pathophysiology of this condition. This study investigates the role of epigenetic factor in intervertebral disc gene regulation. Material and Methods IVD tissues were obtained from donors undergoing anterior lumbar discectomy procedures. The collected tissues were minced and immediately used for cell isolation by sequential enzyme digestion. The demethylation agent 5-azacytidine (5-AzaC) was used to inhibit DNA methylation in IVD cells. The cells were then analyzed for change in DNA methylation using Methyl-Profiler PCR array. We used Methyl-Profiler PCR array containing developmental and cell cycle regulatory genes. The Methyl-Profiler PCR arrays were performed as per manufacturer's instructions. Briefly, the cells were treated with and without 5-azaC for 4 days, and genomic DNA were isolated. Then the equal amounts of genomic DNA were overnight digested with methyl-specific restriction digestion enzyme, nonspecific restriction enzyme, a combination of methyl specific and non-specific, and a control mock digestion. Following digestion, real time PCR was performed using biorad-MyiQ cycler. The data was analyzed according to manufacturer's instructions. Results Methyl-Profiler PCR array showed novel genes hypo-methylated following 5-azacytidine treatment. We found that around 6 genes' promoters differentially methylated after experimental demethylation in IVD cells. The genes differentially methylated are CCND2, CDH1, HIC1, MGMT, PRDM2 and TP73. Real time PCR showed these genes were differentially regulated relative to control. Conclusion There are reports that these differentially methylated genes have a role in various age related degenerative diseases. We speculate that these genes may also play an important role in intervertebral disc degeneration. It would be interesting to investigate the expression of these genes and their methylation pattern in aging human degenerative IVD. In summary, our results suggest that demethylation of IVD cells may be one of the factors involved in IVD degeneration. Further studies are required to confirm our findings and to better understand the effects of aberrant DNA methylation on IVD biology and disease pathophysiology.
Mesenchymal Stem Cells Harvested from Vertebral Body and Iliac Crest Bone Marrow: Are They Equally Good? Comparison of Two Tissues and Two Harvesting Techniques
Authors: Fragkakis Evangelos M. , Jones Elena , Pountos Ippokratis , Giannoudis Peter 
Published: April 2016

Introduction/Objectives Clinical applications of Mesenchymal Stem Cells (MSCs) harvested from bone marrow (BM) are well established. Iliac crest (IC) is considered to be the gold standard. Vertebral body (VB) BM represents an alternative source, promoting bone fusion in spinal operations. Existing literature, comparing ICBM and VBBM MSCs, presents limitations. Density gradient centrifugation (lymphoprep, Lmp) for BM processing is used, despite the known effectiveness on ICBM, of the red cell lysis technique (ammonium chloride, AC). This study compared the yield and differentiation capacities of MSCs in VBBM and ICBM, and evaluated the effectiveness of two harvesting procedures on both BM sources. Patient/Methods Eighteen patients undergoing idiopathic scoliosis surgery were recruited (median age 15.4 years, range 13–17). BM was aspirated intraoperatively from IC (10ml) and 12th thoracic vertebra (10ml). Aspirates were processed using AC and Lmp. MSCs were enumerated using standard CFU-F assay. Colony areas, MSCs doubling rates and outcomes of the differentiation assays were statistically analyzed. Results AC yielded more leukocytes from ICBM (23x106cells/ml), compared with VBBM (14.4x106cells/ml, p = 0.013). Lmp yielded similar numbers of mononuclear cells from ICBM (7.5x106cells/ml) and VBBM (5x106cells/ml). More MSCs were harvested from 1ml of BM aspirate using AC technique, from both anatomical sites (IC: Lmp 199MSCs/ml, AC 367MSCs/ml, p = 0.012; VB: Lmp 296MSCs/ml, AC 440MSCs/ml, p = 0.043). There were no significant differences in MSC numbers/ml and average colony areas between the sites (IC: AC 14mm2, Lmp 15mm2; VB: AC 11mm2, Lmp 13mm2) or MSCs doubling rates in days (IC: AC 2, Lmp 1.9, VB: AC 1.9, Lmp 1.8). Calcium production, after 14 days of osteogenic induction, was higher in VBBM MSCs with both harvesting methods (AC: 15-fold, p = 0.003, Lmp: 6.6-fold). Adipogenic differentiation, measured by Nile-Red absorbance on day 21 post-induction, was not significantly different between two BM sources. Glycosaminoglycan (GAG) production, after 21 days of chondrogenic induction, was higher in VBBM MSCs (AC 2.1μg/ml, Lmp 3.7μg/ml) compared with ICBM (AC 1.2μg/ml, Lmp 2.9μg/ml), but not statistically significant. Conclusion AC results in more effective MSC isolation and does not affect MSCs’ proliferation and differentiation. Compared with ICBM, VBBM is richer in MSCs with osteo- and chondrogenic differentiation capacities and can therefore represent the first choice for MSC harvesting in spinal surgery.
Improvement of Bovine Nucleus Pulposus Cells Isolation Leads to Identification of Three Phenotypically Distinct Cell Subpopulations
Authors: Molinos Maria , Almeida Catarina , Gonçalves Raquel , Barbosa Mário 
Published: April 2016

Introduction Strategies to promote intervertebral disc (IVD) regeneration have been hindered by the lack of knowledge of IVD fundamental cellular/molecular components. One of the key points to be addressed is the characterization of nucleus pulposus (NP) cell population(s). This study establishes an improved method for bovine NP (bNP) cell isolation, whose procedure is still not consensual among the literature, allowing a thorough characterization of cell (sub)populations that exist in the young NP. Material and Methods bNP was digested with distinct enzymes (collagenase-type-I, collagenase-type-II, and collagenase- type-XI) at different concentrations (0.5, 1.0, and 2.0 mg/mL), for 4 and 19 hour. Cell yield, viability/apoptosis, and morphology were analyzed by flow cytometry and imaging flow cytometry. Identification of cell subpopulations within NP and its phenotype was investigated by assessing expression of CD29, CD44, CD45, CD34, CD146, and Brachyury. Results It was found that bNP cells present a similar morphology independently of the digestive enzyme used. However, cell yield was greatly improved by Coll-XI (2 mg/mL) treatment for a short digestion period. Interestingly, three subpopulations, with different sizes and auto-fluorescence, were consistently identified by flow cytometry. And crucially, differential expression of cell markers was found among these subpopulations. Conclusion This study demonstrated that collagenase-type-XI is an efficient enzyme that is used for digesting bNP. And most importantly, three phenotypically distinct subpopulations of cells where identified within the bNP. Such knowledge is key for a better understanding of NP cell biology and its potential endogenous regenerative capacity.
Modic Changes in Lumbar Spine: Prevalence and Genetic Association Analysis through Single Neucleotide Poymorphism Analysis of 71 Polymorphisms
Authors: Rajasekaran S. , Shetty Ajoy Prasad , Kanna Rishi 
Published: April 2016

Introduction A significant association between non-specific low back pain and Modic changes (MC) have been described. We hypothesied that a single nucleotide polymorphisms(SNP) may be involved in matrix and bone metabolism associated changes with modic changes. The present study purports to evaluate the prevalence of MC and the role of genetic polymorphisms in 40 candidate genes (including 71 SNPs). Methods The study was a cross sectional study performed on 809 patients irrespective of back pain status. Assessment was performed with MRI evaluation and genetic association analysis of 71 SNPs belonging to 40 genes. Inclusion criteria for patients were both sexes, age of 18 to 70 years, no evidence of other spinal diseases, previous surgery & trauma. The MRI included T1 and T2 sagittal sections of the lumbar spine. The presence and type of Modic changes & the genetic association analysis of 71 SNPs was made with reference to the presence or absence of Modic Changes. Genotyping of SNPs was performed using the Sequenome® platform. Results 809 individuals of mean age 36.7 ± 10.8 years were studied. Based on the presence of MC, the group was divided into 702 controls and 107 cases. 64 patients had single level MC (7.9%), 32 had double level (3.9%), 5 had three and four level MC (0.6%) and one had multi-level MC. Modic changes were identified in 251 of the 1070 endplates (EP). MC was more commonly situated in the lower EP (149, 59.4%) than in the upper EP (102, 40.6%). L4–5 EP were the most commonly affected level ( n = 77, 30.7%) followed by L5-S1 (26.3%), L3–4 (23.9%), L2–3 (12.4%) and L1–2 (6.8%). Type 2 MC was the most commonly observed pattern ( n = 206, 82%), followed by Type 1 ( n = 27, 10.8%) and Type 3 ( n = 18, 7.2%). Mirroring endplate MC was observed in 75 discs (29.8%) among the total 535 discs. SNPs of Vitamin D receptor gene (VDR) and Matrix Metallo proteinase (MMP20) ( p = 0.03) were significantly associated with MC. SNP of cyclo-oxygenase gene(COX2) and Insulin Growth Factor Receptor(IGF1R) ( p = 0.03) were significantly associated with Type 2 MC. Conclusion The study identifies genetic polymorphisms of VDR, MMP 20, COX 2 and IGF1R to be significantly associated with MC and have not been reported previously. Understanding the etiopathogenetic of Modic changes would help us to plan preventive and therapeutic strategies
Phenotype Standardization is the Urgent Need in Genetic Association Studies of Disc Degeneration - Results of Analysis of Genetic Association of 71 SNPs with Highly Specific Phenotypes in 809 Subjects
Authors: Rajasekaran S. , Kanna Rishi M. , Natesan Senthil , Muthurajan Raveendran , Ranjani Veera , Cheung Kenneth M. C. , Chan Danny , Kao Patrick Y. P. , Yee Anita , Prasad Ajoy 
Published: April 2016

Introduction Although the influence of genetics on etio-pathogenesis of disc degeneration is well recognized, there is a wide variation in the different genes observed to have association with disc degeneration. The clinical and radiographic features of disc degeneration used as phenotypes are quite variable in previously published studies. It is unclear whether the variations in genetic association studies depict variations in ethnicity or selection of phenotypes. Methods The purpose of the study was to evaluate the allelic diversity of 71 SNPs related to six different MRI markers of lumbar degenerative disc disease (annular tear, Pfirmann's grading, Schmorl's nodes, Modic changes, Total Endplate Damage score and disc bulge) in a single study population, and analyze how genetic associations can vary in the same study subjects with the choice of phenotype, based on age and sequence of selection of study subjects. Genotyping of cases and controls was performed on Genome wide SNP array to identify potentially associated disease loci. The results from the GWAS were then used to facilitate SNP selection and genotype validation was conducted using Sequenom based genotyping. Results The mean age of 809 subjects (M: 455, F:354) was 36.76 ± 10.8 years (range 10 – 80). Highly significant association ( p < 0.01) was observed with three SNPs of CILP for disc bulge and rs2249350 of ADAMTS5 and rs11247361IGF1R for annular tears. Significant association ( p < 0.05) was observed with polymorphisms of VDR and MMP20 for Modic changes, three SNPs of MMP20 for Schmorl's node and SNPs of CALM1 and FN1 for Pfirmann's grading. None of the SNPs had significant association with TEPS. Subgroup analysis based on age (<30, 30–40 and > 40 years) showed new set of genetic associations for all the six radiographic parameters. Similarly the population was divided into two groups based on numerical order and the association patterns completely differed as compared with the total group. Conclusions In the same study population with DDD, SNP associations completely change when phenotypes changed. Variations in age, sequence of study subjects, number of population apart from radiographic description of DDD significantly change the genetic association. Based on current results, it is difficult to consider one set of genes as responsible for disc degeneration considering these variations. Our study results demand an urgent need for standardizing the description of DDD, phenotype selection, and study criteria for genetic association studies.
Discectomy-Related Information on the Internet: Does the Quality Follow the Surge?
Authors: Sheridan Gerard , Elhassan Yahya , Mujtaba Nassiri , Mugtaba Sulman , Kiely Pat , Noel Jacques 
Published: April 2016

Introduction Over 2.5 billion users search the internet world wide. 70%- 80% look at health related information, yet no standards exist regarding the publication of medical literature on the internet. The quality of health information is of much concern and an emphasis on appraisal of internet Web sites is needed. We aim to look at the quality of health information on the internet relating to discectomy using various standardised tools and scoring systems. Material and Methods The three most commonly used search engines were identified and a search for “Discectomy” was performed on each. Two reviewers categorized the most popular Web sites according to their types and the quality of each was assessed using recognized scoring systems including the Journal of the American Medical Association (JAMA) benchmark, DISCERN and a novel Discectomy-specific content score. The quality of the information was also assessed according to the presence or absence of the Health on the Net (HON) code, a reported quality-assurance marker. Results Results confirmed that 53 websites were identified, and analyzed. Commercial Web sites were predominant at 24, 7 were governmental, 6 were produced by physicians, none were produced by allied health professionals, 3 were academic, 4 were public health information Web sites, 4 were attached to social media and discussion groups, 3 were media related, and 2 were unspecified. Using the 3 scoring systems mentioned above as a quality marker, internet sites with a 'Health on the Net’ (HON) code demonstrated significantly higher quality than those without the code ( p < 0.0001). Public health information Web sites attained the highest overall DISCERN and Discectomy-specific scores followed by the Governmental & Non-Profit Organizations Web sites. Conclusion The overall quality of information regarding discectomy remains poor and variable despite an exponential increase in the number of users and Websites, with a slight trend toward improvement. Only 20% to 30% are of good quality, compared with that 10 years ago (<10%). Presence of Health on the Net code is a very reliable marker for health information quality.
Analysis of Lumbar Foramina Size Change in Patients with Lumbar Spondylolisthesis using Kinetic Magnetic Resonance Imaging
Authors: Huang Hsiang-Ming , Yang Jui-Jung , Patel Neil , Wang Justin , Hsieh Patrick , Liu John , Buser Zorica , Wang Jeffrey C. 
Published: April 2016

Introduction The pathology of lumbar spondylolisthesis contributes to lumbar instability and significant clinical symptoms. However, the effect on intervertebral foraminal area (FA) during motion change is not well understood. Material and Methods In this study we used 107 patients diagnosed with single or multiple level lumbar spondylolisthesis at L3–L4, L4–L5 or L5–S1. All patients were initially divided into two groups: group A (sliding percentage 0–3%) and group B (sliding percentage ≥4%). Sliding percentage and FA size in neutral, flexion and extension were measured using kMRI. The FA change in different sliding groups at L5-S1 level was further analyzed. Results The number of patients in the group A (sliding percentage 1–3%) was 68 at L3–4, 38 at L4–5 and 32 at L5-S1 spine levels. The number of patients in the group B (sliding percentage ≥4%) was the following: 39 at L3–L4, 69 at L4–L5 and 75 at L5–S1. The mean FA in both groups changed with different spine positions. When considering all levels, the average FA increased ~17% from neutral to flexion and decreased ~21% from neutral to extension. Similarly, there was a significant difference in FA in flexion at L5-S1 ( p < 0.004) between the groups. For L5/S1 patients, 104 patients were divided into three sub-groups according to the sliding percentage (group I (0–9%), group II (10–19%) and group III (20–29%)). The FA was significantly increased in the flexion position and decreased in the extension position in all three groups. In group III FA size was significantly decreased from neutral to flexion when compared with Group I ( p < 0.022). The FA size was significantly increased from neutral to extension when group III was compared with groups I and II ( p < 0.000 and p < 0.017, respectively). Conclusion Our kMRI study revealed position-dependent changes of the intervertebral foramen. In patients with severe sagittal subluxation the change in FA from neutral to extension position demonstrated the largest decrease. This change may also correlate with changes in neuroforaminal stenosis and nerve root compression and its associated symptoms and should be taken in consideration when choosing the treatment option.