Gene Expression Data from Human Mesenchymal Stem Cells with and without TGF-B vs Human Annulus Disc Cells
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE17782 | GEO | 2009/08/25
SECONDARY ACCESSION(S): PRJNA117879
REPOSITORIES: GEO
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Project description:Autologous stem cell therapy has potential for biologic treatment of disc degeneration. Due to ease of harvest and abundance, adipose-derived mesenchymal stem cells (AD-MSC) are readily available. Our objectives were: 1) To develop/validate methods to harvest AD-MSC and direct them to a disc-like phenotype by three-dimensional (3D) culture and TGF-Ã3 exposure; 2) To perform gene expression profiling for human AD-MSC and annulus cells in 3D culture; 3) To test whether disc cell-AD-MSC co-culture could augment proteoglycan production. Stem cell plasticity offers potential for future biologic therapies for disc degeneration. Data indicated that human AD-MSC can successfully be manipulated in 3D culture to express gene products important in the disc ECM (types I and II collagen, chondroitin sulfate, keratin sulfate, decorin), and that co-culture of annulus cells with AD-MSC enhances proteoglycan production. Studies defined gene expression patterns of AD-MSC and human annulus cells in 3D culture, important as we explore the potential of MSC in biologic therapies for disc degeneration. Experiment Overall Design: AD-MSC were extracted from human adipose tissue, and characterized as stem cells using accepted criteria (direction into osteoblasts or chondrocytes; and cell surface marker criteria). Three AD-MSC cultures were grown in 3D with or without TGF-Ã3 for 2-3 weeks. Disc Tissue samples were obtained from surgical disc procedures performed on patients with herniated discs and degenerative disc disease. Seven disc cultures were also grown in 3-D for 2 weeks. RNA was harvested according to instructions with the Trizol isolation method, checked for quality using the 2100 Bioanalyzer (Agilent Technologies, Inc., Santa Clara, CA USA), reverse-transcribed to double-stranded cDNA, subjected to two rounds of transcription, and hybridized to the DNA microarray in the Affymetrix Fluidics Station 400. Affymetrix human U133 X3P arrays were used. Using Genesifter, gene expression in AD-MSC with and witout TGF-B3 was compared to annulus cells. Gene expression of stem cells with TGF-Ã3 was also compared to stem cells grown in the absence of TGF-Ã3.
2009-09-15 | E-GEOD-17782 | biostudies-arrayexpress
Project description:Asporin, also known as periodontal ligament-associated protein 1 (PLAP1), is a member of the family of small leucine-rich proteoglycan (SLRP) family. It is present within the cartilage extracellular matrix (ECM), and is reported have a genetic association with osteoarthritis. Its D14 allele has recently been found to be associated with lumbar disc degeneration in Asian subjects. There have been no studies, however, of this gene’s normal immunohistochemical localization within the human intervertebral disc, nor of expression levels in Caucasian individuals with disc degeneration. Studies were approved by our human subjects Institutional Review Board. Methods included immunohistochemical localization of asporin in the disc of humans and the sand rat (a small rodent with spontaneous age-related disc degeneration), and Affymetrix microarray analysis of asporin gene expression in vivo and in vitro. mmunohistochemical studies of human discs revealed that some, but not all, cells of the outer annulus expressed asporin. Fewer cells in the inner annulus contained asporin, and it was rarely present in cells in the nucleus pulposus. Similar patterns were found for the presence of asporin in lumbar discs of sand rats. Substantial relative gene expression levels were seen for asporin in both disc tissue and in annulus cells grown in three-dimensional culture. More degenerate human discs (Thompson grade 4) showed higher expression levels of asporin than did less degenerate (grade 1, 2 and 3) discs, p = 0.004. In the discs of Caucasian subjects studied here, and in the sand rat, greater immunolocalization levels were found in the outer compared to inner annulus. Localization was rare in the nucleus. Gene expression studies showed greatest expression of asporin in the more degenerate human discs in vivo.
2009-03-24 | GSE15227 | GEO
Project description:Asporin, also known as periodontal ligament-associated protein 1 (PLAP1), is a member of the family of small leucine-rich proteoglycan (SLRP) family. It is present within the cartilage extracellular matrix (ECM), and is reported have a genetic association with osteoarthritis. Its D14 allele has recently been found to be associated with lumbar disc degeneration in Asian subjects. There have been no studies, however, of this gene’s normal immunohistochemical localization within the human intervertebral disc, nor of expression levels in Caucasian individuals with disc degeneration. Studies were approved by our human subjects Institutional Review Board. Methods included immunohistochemical localization of asporin in the disc of humans and the sand rat (a small rodent with spontaneous age-related disc degeneration), and Affymetrix microarray analysis of asporin gene expression in vivo and in vitro. mmunohistochemical studies of human discs revealed that some, but not all, cells of the outer annulus expressed asporin. Fewer cells in the inner annulus contained asporin, and it was rarely present in cells in the nucleus pulposus. Similar patterns were found for the presence of asporin in lumbar discs of sand rats. Substantial relative gene expression levels were seen for asporin in both disc tissue and in annulus cells grown in three-dimensional culture. More degenerate human discs (Thompson grade 4) showed higher expression levels of asporin than did less degenerate (grade 1, 2 and 3) discs, p = 0.004. In the discs of Caucasian subjects studied here, and in the sand rat, greater immunolocalization levels were found in the outer compared to inner annulus. Localization was rare in the nucleus. Gene expression studies showed greatest expression of asporin in the more degenerate human discs in vivo. Disc Tissue samples were obtained via the National Cancer Institute Cooperative Tissue Network (CHTN) as well as surgical disc procedures performed on patients with herniated discs and degenerative disc disease. Tissue was fixed and paraffin embedded. Standard laser capture microdissection (LCM) techniques were used to collect the cells from which total RNA was isolated and analyzed via microarray. Based on the Thompson scouring system, unhealthy discs (grade 4) were compared to healthy discs (grades 2,3).
2009-03-24 | E-GEOD-15227 | biostudies-arrayexpress
Project description:Although the degenerating disc is considered to be the key source of pain in patients with low back pain, the relationship between disc cells, nerves, and pain production is poorly understood. Neurotrophins are signaling molecules involved in the survival, differentiation, migration, and neurite outgrowth of central and peripheral neurons. Neurotrophins are now known to be expressed in non-neuronal tissues, including the intervertebral disc. We hypothesize that the inflammatory cytokine interleukin-1 (IL-1beta), which is produced by disc cells during degeneration, is a key element in the cascade of events involving neurotrophins. To test this, we used an in vitro experimental design which challenged 3D-cultured human annulus cells with IL-1beta and utilized microarray analysis to evaluate neurotrophin- and nerve-related gene expression profiles in treated vs. control cells. Analysis of nerve growth factor levels in conditioned media was also performed. Findings presented here support the hypothesis that proinflammatory cytokines (which are produced by disc cells during degeneration) are involved with a significant increase in the expression of neurotrophins and other nerve-related genes. Findings expand previous data on expression of neurotrophins in the degenerating disc, and provide the first documentation of expression of neurotrophin 3 and neuropilin 2. These results have direct translational relevance because they address the primary clinical issue with disc degeneration (low back pain) and open the possibility of novel analgesic therapies based on development of specific small-molecular antagonists to neurotrophins. Human disc tissue samples were obtained from surgical disc procedures performed on patients with herniated discs and degenerative disc disease. Cultured annulus cells were grown in a 3D collagen construct with or without 10 2 pM IL-1 for a total of 14 days. Following homogenization in TRIzol reagent, total RNA was isolated and analyzed via mircoarray.
2012-06-26 | E-GEOD-27494 | biostudies-arrayexpress
Project description:The objective of this study was to determine how TNF-a, an important proinflammatory cytokine, affects gene expression in the human annulus. Cells were grown in a 3D collagen construct for 14 days with TNF-a. mRNA was isolated and subjected to microarray. Fold changes in gene expression were determined via GeneSifer software. Human disc tissue samples were obtained from surgical disc procedures performed on patients with herniated discs and degenerative disc disease. Cultured annulus cells were grown in a 3D collagen construct with or without 10e3 pM TNF-a for a total of 14 days. Following homogenization in TRIzol reagent, total RNA was isolated and analyzed via mircoarray.
2012-10-31 | E-GEOD-41883 | biostudies-arrayexpress
Project description:Transcriptional analysis of 6-month-old primary nucleus pulposus (NP) and annulus fibrosus (AF) mouse disc tissues from wild-type (WT) and N153S mice, which harbor a mutation wich constiuitively activates Sting. Mouse details available here: https://www.jax.org/strain/033543 The cGAS-STING pathway promotes the senescence-associated secretory phenotype (SASP), which is associated with intervertebral disc degeneration, and has had implications in inflammatory musculoskeletal disorders. We examined the role of STING in the disc by examining the transcriptomic profiles of nucleus pulposus and annulus fibrosus cells in WT and N153S mice using microarray.
2022-07-06 | GSE188909 | GEO
Project description:Low back pain is a major cause of disability especially for people between 20 and 50 years of age. As a costly healthcare problem, it imposes a serious socio-economic burden. Current surgical therapies have considerable drawbacks and fail to replace the normal disc in facilitating spinal movements and absorbing load. Therefore, the focus of regenerative medicine is on identifying biomarkers and signalling pathways to improve our understanding about the cascades of disc degeneration and allow for the design of specific therapies. We hypothesized that comparing microarray profiles from degenerative and non-degenerative discs will lead to the identification of dysregulated signalling and pathophysiological targets. Microarray data sets were generated from human annulus fibrosus cells and analysed using IPA ingenuity pathway analysis system. Gene expression values were validated by qRT-PCR, and respective proteins were identified by immunohistochemistry. Microarray analysis revealed 17 dysregulated molecular markers and various dysregulated cellular functions, including cell proliferation and inflammatory response, in the human degenerative annulus fibrosus. The most significant canonical pathway induced in degenerative annulus fibrosus was found to be the interferon signalling pathway. In conclusion, this study indicates interferon-alpha signalling pathway activation with IFIT3 and IGFBP3 up-regulation which may affect cellular function in human degenerative disc. 48 samples of intervertebral disc tissue - annulus fibrosus and nucleus pulposus - displaying varying degrees (grades) of degeneration
2015-10-23 | E-GEOD-70362 | biostudies-arrayexpress
Project description:Although it is well-recognized that apoptosis, senescence, and increased production of inflammatory cytokines and catabolic products are important factors in the degeneration of the human intervertebral disc, there is poor understanding of the underlying cause. The objective of the present study was to analyze gene expression patterns in the human annulus disc tissue.
2011-03-01 | GSE23130 | GEO
Project description:Description: The intervertebral disc (IVD) is a joint in the spine that is comprised of three major structures, the nucleus pulposus (NP), annulus fibrosus (AF) and cartilaginous endplate (EP). The NP is a proteoglycan-rich structure which contains a heterogenous population including the presence of progenitors such as the notochordal-like cells (NLCs) and NP cells, which are responsible for the synthesis and turnover of extracellular matrix in the NP. During aging and degeneration, there is a loss of cells, hydration, and changes in the ECM, that leads to the alterations in the biomechanical function of the IVD. The replenishment of cells, in particular, the progenitors are a potential therapy for IVD degeneration. However, a major challenge lies in obtaining sufficient numbers of healthy cells for treatment. This study used a 3 part protocol to differentiate pluripotent stem cells into NLCs in vitro.
2023-11-11 | GSE237546 | GEO
Project description:Although the degenerating disc is considered to be the key source of pain in patients with low back pain, the relationship between disc cells, nerves, and pain production is poorly understood. Neurotrophins are signaling molecules involved in the survival, differentiation, migration, and neurite outgrowth of central and peripheral neurons. Neurotrophins are now known to be expressed in non-neuronal tissues, including the intervertebral disc. We hypothesize that the inflammatory cytokine interleukin-1 (IL-1beta), which is produced by disc cells during degeneration, is a key element in the cascade of events involving neurotrophins. To test this, we used an in vitro experimental design which challenged 3D-cultured human annulus cells with IL-1beta and utilized microarray analysis to evaluate neurotrophin- and nerve-related gene expression profiles in treated vs. control cells. Analysis of nerve growth factor levels in conditioned media was also performed. Findings presented here support the hypothesis that proinflammatory cytokines (which are produced by disc cells during degeneration) are involved with a significant increase in the expression of neurotrophins and other nerve-related genes. Findings expand previous data on expression of neurotrophins in the degenerating disc, and provide the first documentation of expression of neurotrophin 3 and neuropilin 2. These results have direct translational relevance because they address the primary clinical issue with disc degeneration (low back pain) and open the possibility of novel analgesic therapies based on development of specific small-molecular antagonists to neurotrophins.
2012-06-27 | GSE27494 | GEO