Cartilage Endplate-derived Stem Cells Research Articles

Leptin Induces MMP-1 Expression Through the RhoA/ERK1/2/NF-κB Axis in Human Intervertebral Disc Cartilage Endplate-Derived Stem Cells.

Intervertebral disc (IVD) degeneration, associated with aging, may cause low back pain and disability, with obesity as a significant risk factor. In a prior study, we found a positive correlation between IVD degeneration and levels of matrix metalloproteinase-1 (MMP-1) and leptin. Yet, the interaction between MMP-1 and leptin in IVD degeneration is unclear. Our research seeks to explore leptin's influence on MMP-1 expression and the underlying mechanisms in human intervertebral disc cartilage endplate-derived stem cells, specifically SV40 cells. The mRNA and protein expression in leptin-stimulated SV40 cells were assessed using RT-real-time PCR and Western blotting or ELISA, respectively. We examined leptin-mediated RhoA activation through a GTP-bound RhoA pull-down assay. Furthermore, the phosphorylation levels of mitogen-activated protein kinases and AKT in leptin-stimulated SV40 cells were analyzed using Western blotting. The activation of NF-κB by leptin was investigated by assessing phosphorylation of IKKα/β, IκBα, and NF-κB p65, along with the nuclear translocation of NF-κB p65. To understand the underlying mechanism behind leptin-mediated MMP-1 expression, we employed specific inhibitors. Leptin triggered the mRNA and protein expression of MMP-1 in SV40 cells. In-depth mechanistic investigations uncovered that leptin heightened RhoA activity, promoted ERK1/2 phosphorylation, and increased NF-κB activity. However, leptin did not induce phosphorylation of JNK1/2, p38, or AKT. When we inhibited RhoA, ERK1/2, and NF-κB, it resulted in a decrease in MMP-1 expression. Conversely, inhibition of reactive oxygen species and NADPH oxidase did not yield the same outcome. Additionally, inhibiting RhoA or ERK1/2 led to a reduction in leptin-induced NF-κB activation. Moreover, inhibiting RhoA also decreased leptin-mediated ERK1/2 phosphorylation. These results indicated that leptin induced MMP-1 expression in SV40 cells through the RhoA/ERK1/2/NF-κB axis. This study provided the pathogenic role of leptin and suggested the potential therapeutic target for IVD degeneration.

Carbohydrate sulfotransferase 3 (CHST3) overexpression promotes cartilage endplate-derived stem cells (CESCs) to regulate molecular mechanisms related to repair of intervertebral disc degeneration by rat nucleus pulposus.

To investigate the regulatory effect of carbohydrate sulfotransferase 3 (CHST3) in cartilage endplate‐derived stem cells (CESCs) on the molecular mechanism of intervertebral disc degeneration after nucleus pulposus repair in rats. We performed GO and KEGG analysis of GSE15227 database to select the differential genes CHST3 and CSPG4 in grade Ⅱ, Ⅲ and Ⅳ intervertebral disc degeneration, IHC and WB to detect the protein profile of CHST3 and CSPG4, Co‐IP for the interaction between CHST3 and CSPG4. Then, immunofluorescence was applied to measure the level of CD90 and CD105, and flow cytometry indicated the level of CD73, CD90 and CD105 in CESCs. Next, Alizarin red staining, Alcian blue staining and TEM were performed to evaluate the effects of CESCs into osteoblasts and chondroblasts, respectively, CCK8 for the cell proliferation of osteoblasts and chondroblasts after induction for different times; cell cycle of osteoblasts or chondroblasts was measured by flow cytometry after induction, and WB for the measurement of specific biomarkers of OC and RUNX in osteoblasts and aggrecan, collagen II in chondroblasts. Finally, colony formation was applied to measure the cell proliferation of CESCs transfected with ov‐CHST3 or sh‐CHST3 when cocultured with bone marrow cells, WB for the protein expression of CHST3, CSPG4 and ELAVL1 in CSECs, transwell assay for the migration of CESCs to bone marrow cells, TEM image for the cellular characteristics of bone marrow cells, and WB for the protein profile of VCAN, VASP, NCAN and OFD1 in bone marrow cells. CHST3 and CSPG4 were differentially expressed and interacted in grade Ⅱ, Ⅲ and Ⅳ intervertebral disc degeneration; CD73, CD90 and CD105 were lowly expressed in CESCs, osteogenic or chondroblastic induction changed the characteristics, proliferation, cell cycle and specific biomarkers of osteoblasts and chondroblasts after 14 or 21 days,; CHST3 affected the cell proliferation, protein profile, migration and cellular features of cocultured CESCs or bone marrow cells. CHST3 overexpression promoted CESCs to regulate bone marrow cells through interaction with CSPG4 to repair the grade Ⅱ, Ⅲ and Ⅳ intervertebral disc degeneration.

BNIP3/Bcl-2-mediated apoptosis induced by cyclic tensile stretch in human cartilage endplate-derived stem cells.

The present study aimed to investigate the molecular mechanisms of cyclic stretch-induced apoptosis in human intervertebral disc cartilage endplate-derived stem cells (CESCs). CESCs were stretched by the Flexercell-4000™ Tension Plus system, the effect on cell viability was measured by a Cell Counting Kit-8 assay, while cell apoptosis was detected by flow cytometry. Western blot analysis was used to evaluate the expression of B-cell lymphoma 2 (Bcl-2)/adenovirus E1B 19 kDa interacting protein 3 (BNIP3), Bcl-2, Bcl-2 homologous antagonist killer (Bak), Bcl-2-associated X protein (Bax), Bcl extra large (Bcl-xl) and the activity of caspase-3, while Z-VAD-FMK was used to inhibit caspase-3. Compared with the control group, the cell viability decreased in a time-dependent manner after stretching. Furthermore, cell apoptosis and the activity of caspase-3 were increased in a time-dependent manner. The ratio of pro-death factor BNIP3 to anti-apoptotic protein Bcl-2 was significantly increased. When cells were stretched for 36 h, the apoptosis-associated proteins Bak and Bax were increased, while Bcl-xl was decreased. The viability and apoptotic ratio of stretched cells were significantly restored after caspase-3 was repressed. In conclusion, cyclic tensile stretch induced apoptosis of CESCs, which was probably due to upregulation of the expression of BNIP3.

The characteristics of stem cells in human degenerative intervertebral disc.

Background:The aim of this study is to identify which possessed the best stem-cell-like characteristics in 3 kinds of cell in human degenerative intervertebral disc: NPSCs (nucleus pulposus-derived stem cells), AFSCs (annulus fibrosus-derived stem cells), or CESCs (cartilage endplate-derived stem cells).Methods:We separated the disc samples obtained from 15 surgically treated patients with disc degenerative diseases into nucleus pulposus, annulus fibrosus, and cartilage endplate. After cultivating, we used the cell counting kit-8 to analysis the cell activity of 3 kinds of disc tissue-derived stem cell separately; different stem cells were defined with multilineage (osteogenic, chondrogenic, and adipogenic) differentiation. We extracted the total RNA and detected the expression of different lineage differentiation-related genes with the real-time polymerase chain reaction (RT-PCR).Results:Cell morphology of NPSCs, AFSCs, and CESCs did not show significant difference. Cell proliferation capacity of NPSCs and AFSCs was stronger than that of CESCs. The differentiation outcomes showed that osteocyte-like cells were stained red by Alizarin red S, chondrocyte-like cells blue by toluidine blue, and adipocyte-like red by oil red O. The RT-PCR reflected that the expression of different lineage differentiation-related genes of AFSCs was stronger than NPSCs and CESCs.Conclusion:In conclusion, we found that the cell morphology was not significantly different among NPSCs, AFSCs, and CESCs. Both differentiation and RT-PCR tests demonstrated that AFSCs had the best stem-cell-like characteristics in the human degenerative intervertebral disc.

A genome-wide analysis of the gene expression profiles and alternative splicing events during the hypoxia-regulated osteogenic differentiation of human cartilage endplate-derived stem cells.

It has been hypothesized that intervertebral disc degeneration is initiated by degeneration of the cartilage endplate (CEP), which is characterized by cartilage ossification. CEP-derived stem cells (CESCs), with the potential for chondro-osteogenic differentiation, may be responsible for the balance between chondrification and ossification in the CEP. The CEP remains in an avascular and hypoxic microenvironment; the present study observed that hypoxia was able to markedly inhibit the osteogenic differentiation of CESCs. This tissue-specific CESC differentiation in response to a hypoxic microenvironment was physiologically important for the prevention of ossification in the CEP. In order to study the hypoxia-regulated mechanisms underlying osteogenic differentiation of CESCs, a Human Transcriptome Array 2.0 was used to detect differentially expressed genes (DEGs) and alternatively spliced genes (ASGs) during the osteogenic differentiation of CESCs under hypoxia, compared with those induced under normoxia. High-throughput analysis of DEGs and ASGs demonstrated that genes in the complement pathway were enriched, which may be a potential mechanism underlying hypoxia inhibition of CESCs osteogenesis. The results of the present study may provide a basis for future mechanistic studies regarding gene expression levels and alternative splicing events during the hypoxia-regulated inhibition of osteogenesis, which may be helpful in identifying targets for CEP degeneration therapy.

The comparison of biological characteristics of three kinds of stem cells in human degenerative intervertebral disc

Objective To compare the biological characteristics of nucleus pulposus-derived stem cells (NPSCs), annulus fibrosus-derived stem cells (AFSCs) and cartilage endplate-derived stem cells (CESCs) from the same patient who had degenerative disc. Methods The human discs isolated from 7 patients with intervertebral disc herniation were separated carefully and digested and filtered before getting the original NPSCs, AFSCs and CESCs. When cultivated to third passage, the cell activity of stem cells was analyzed by using cell counting kit-8 (CCK-8); multilineage (adipogenic, osteogenic and chondrogenic) differentiation potential was analyzed to define different stem cells. The total RNA of third-passage-cells was extracted, and then reverse transcriptase-polymerase chain reaction (RT-PCR) was used to detect the relative expression of adipogenic, osteogenic and chondrogenic differentiation-related genes. Results Cell morphology of NPSCs, AFSCs and CESCs from the same patient showed no significant difference. The growth curve showed that proliferative ability of CESCs (A value at day 1, 3, 5, 7, 9, 11, 13, 15 was 0.35±0.01, 0.52±0.02, 0.64±0.08, 0.80±0.19, 0.79±0.2, 1.06±0.11, 1.37±0.09, 1.31±0.41) and AFSCs (0.38±0.01, 0.50±0.03, 0.72±0.06, 0.82±0.13, 0.87±0.08, 0.87±0.09, 1.20±0.05, 1.43±0.05)was more active than that of NPSCs (0.32±0.02, 0.42±0.04, 0.59±0.01, 0.64±0.17, 0.65±0.19, 0.68±0.21, 0.67±0.12, 0.59±0.12). Multilineage differentiation results showed adipocyte-like red by oil red O, osteocyte-like cells were stained red by Alizarin red, and chondrocyte-like cells blue by toluidine blue. The RT-PCR results showed that AFSCs was better than NPSCs and CESCs in the aspect of the expression of adipogenic [the relative expression of peroxisome proliferator-activated receptor gamma (PPARγ) gene was 6.85±0.32 vs. 6.63±0.50 and 7.21±0.21; the relative expression of lipoprotein lipase (LPL) gene was 13.72±0.37 vs. 8.41±0.42 and 10.62±0.37] and chondrogenic [the relative expression of type 2 collagen (COL-2) gene was 10.45±1.10 vs. 3.53±0.62 and 8.26±0.49; the relative expression of sex determining region Y-box 9(SOX-9) gene was 14.39±1.22 vs. 7.66±0.68 and 13.70±1.10] differentiation-related genes, whereas the expression of osteogenic differentiation-related genes was best in CESCs[the relative expression of runt related transcription factor-2(RUNX-2) gene was 7.30±1.1 vs. 2.61±0.06 and 6.20±0.92; the relative expression of COL-2 gene was 8.22±0.84 vs. 7.05±0.82 and 4.94±0.90] and the expression of adipogentic differentiation-related genes, particularly LPL gene was worse in NPSCs. Conclusion The cell morphology was similar among NPSCs, AFSCs and CESCs in vitro, but biological characteristics of those were different. Induced differentiation and RT-PCR results both showed that AFSCs had more advantages than NPSCs and CESCs in the field of intervertebral disc tissue engineering. Key words: Intervertebral disc degeneration; Nucleus pulposus stem cells; Annulus fibrosus stem cells; Cartilaginous endplate stem cells; Biological characteristics

A genome wide analysis of alternative splicing events during the osteogenic differentiation of human cartilage endplate-derived stem cells

Low back pain is a prevalent disease, which leads to suffering and disabilities in a vast number of individuals. Degenerative disc diseases are usually the underlying causes of low back pain. However, the pathogenesis of degenerative disc diseases is highly complex and difficult to determine. Current therapies for degenerative disc diseases are various. In particular, cell-based therapies have proven to be effective and promising. Our research group has previously isolated and identified the cartilage endplate‑derived stem cells. In addition, alternative splicing is a sophisticated regulatory mechanism, which greatly increases cellular complexity and phenotypic diversity of eukaryotic organisms. The present study continued to investigate alternative splicing events in osteogenic differentiation of cartilage endplate‑derived stem cells. An Affymetrix Human Transcriptome Array2.0was used to detect splicing changes between the control and differentiated samples. Additionally, molecular function and pathway analysis were also performed. Following rigorous bioinformatics analysis of the data,3,802alternatively spliced genes were identified, and10of these were selected for validation by reverse transcription‑polymerase chain reaction. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes pathway analysis also revealed numerous enriched GO terms and signaling pathways. To the best of our knowledge, the present study is the first to investigate alternative splicing mechanisms in osteogenic differentiation of stem cells on a genome‑wide scale. The illumination of molecular mechanisms of stem cell osteogenic differentiation may assist the development novel bioengineered methods to treat degenerative disc diseases.

Data for the gene expression profiling and alternative splicing events during the chondrogenic differentiation of human cartilage endplate-derived stem cells under hypoxia.

This article contains relevant data of the research article titled Global profiling of the gene expression and alternative splicing events during the hypoxia-regulated chondrogenic differentiation in human cartilage endplate-derived stem cells (Yao et al., 2016) [1]. The data show global profiling of the DEGs (Differentially expressed genes) and AS (Alternative splicing) events during the hypoxia-regulated chondrogenesis of CESCs (human cartilage endplate-derived stem cells) by using Affymetrix Human Transcriptome Array 2.0 (HTA 2.0) system. In addition, the enriched GO (Gene Ontology) functions and signaling pathways are listed. The information presented here includes the information of patients from which the clinical samples are obtained, the list of primers used for validation, the identification, GO and KEGG analysis of DEG and AS events.

Global profiling of the gene expression and alternative splicing events during hypoxia-regulated chondrogenic differentiation in human cartilage endplate-derived stem cells

The intervertebral disc (IVD) degeneration is initiated by cartilage endplate (CEP) degeneration and is characterised by reduced chondrification. Cartilage endplate-derived stem cells (CESCs) with chondrogenic differentiation abilities are responsible for the restoration of cartilage. CEP remains in an avascular and hypoxic microenvironment. In this study, we observed that the physiological hypoxia greatly promotes the chondrogenic differentiation of CESCs. This tissue specificity of the differentiation fate of CESCs in response to the hypoxic microenvironment was physiologically significant for the CEP to maintain the chondrification status. To investigate the mechanisms underlying the hypoxia-regulated chondrogenic differentiation of CESCs, we adopted a high-throughput scanning technology to detect the global profiling of gene expression and alternative splicing (AS) event changes during chondrogenic differentiation under hypoxia in CESCs compared to those induced under normoxia. An Affymetrix Human Transcriptome Array 2.0 was used to identify the differentially expressed genes (DEGs) and alternatively spliced genes (ASGs). After RT-PCR validation, GO and KEGG pathway analyses of both the DEGs and ASGs were performed. The enrichment of the GO functional terms and signalling pathways provided referential direction of the mechanism to study the gene expression and AS in the hypoxia-regulated chondrogenesis promotion, which could be helpful in understanding this physiological phenomenon, and it could also be instrumental in finding targets for CEP degeneration therapy.

Distinguishing characteristics of stem cells derived from different anatomical regions of human degenerated intervertebral discs.

Several types of stem cells have been successfully demonstrated to exist in the human degenerated intervertebral disc (IVD), which is composed of annulus fibrosus (AF), nucleus pulposus (NP) and cartilage endplate (CEP). However, the differences in the biological characteristics among these and bone marrow derived mesenchymal stem cells (BM-MSCs) remain unclear. To investigate this issue, cells were harvested from human AF, NP, CEP, and bone marrow, respectively; passage 2 cells were selected using the agarose suspension culture system to obtain stem cell clones. Following expansion in vitro, stem cells from different anatomical regions were compared regarding the morphology, proliferation ability, immunophenotypic expression, and multi-lineage differentiation capacity. In addition, stem cell-alginate bead compositions were constructed for the comparison of DNA and sGAG content. There were subtle differences regarding cell morphology, but no significant differences in proliferation ability among the four types of stem cells. For the immunophenotypic analysis, all stem cells basically fulfilled the criteria for mesenchymal stem cells (MSCs), which have been published by the International Society for Cellular Therapy (ISCT), with a significant difference in CD105 expression. A comparison of the osteogenic capacities indicated: cartilage endplate-derived stem cells (CESCs)>annulus fibrosus-derived stem cells (AFSCs)>BM-MSCs>nucleus pulposus-derived stem cells (NPSCs). The chondrogenesis difference was similar to osteogenesis. For adipogenesis: BM-MSCs>NPSCs>CESCs>AFSCs. In the stem cell/alginate composition, the CESCs consistently showed the superior chondrogenic potential among all those cell types. Our data indicated that all the four types of stem cells shared some similar biological properties (regarding shape, proliferation ability and immunophenotypic expression). CESCs, which had the strongest osteogenic and chondrogenic potentials, may serve as excellent seed cells for NP/cartilage or bone tissue engineering.

Global Gene Expression Profiling and Alternative Splicing Events during the Chondrogenic Differentiation of Human Cartilage Endplate-Derived Stem Cells.

Low back pain (LBP) is a very prevalent disease and degenerative disc diseases (DDDs) usually account for the LBP. However, the pathogenesis of DDDs is complicated and difficult to elucidate. Alternative splicing is a sophisticated regulatory process which greatly increases cellular complexity and phenotypic diversity of eukaryotic organisms. In addition, the cartilage endplate-derived stem cells have been discovered and identified by our research group. In this paper, we continue to investigate gene expression profiling and alternative splicing events during chondrogenic differentiation of cartilage endplate-derived stem cells. We adopted Affymetrix Human Transcriptome Array 2.0 (HTA 2.0) to compare the transcriptional and splicing changes between the control and differentiated samples. RT-PCR and quantitative PCR are used to validate the microarray results. The GO and KEGG pathway analysis was also performed. After bioinformatics analysis of the data, we detected 1953 differentially expressed genes. In terms of alternative splicing, the Splicing Index algorithm was used to select alternatively spliced genes. We detected 4411 alternatively spliced genes. GO and KEGG pathway analysis also revealed several functionally involved biological processes and signaling pathways. To our knowledge, this is the first study to investigate the alternative splicing mechanisms in chondrogenic differentiation of stem cells on a genome-wide scale.

Immortalization of Human Cartilage Endplate Stem Cell from Degenerated Human Intervertebral Disc

Introduction Low back pain is one of the most common reasons for seeking medical advice and is a major cause of work-related disabilities. Recently, cell-based therapies for regenerating or repairing the degenerated disc have become treatment options. Our group previously demonstrated that stem cells exit in human degenerated CEP and that cartilage endplate-derived stem cells (CESCs) are superior to BM-MSCs in terms of osteogenesis and chondrogenesis. CESCs may represent one of the most promising stem cells for several degenerative conditions due to their multipotency, immunoprivileged properties, and easy expansion in vitro. However, the limited life span of primary CESCs during in vitro expansion greatly hampers their use in basic research and clinical applications. Immortalization of CESCs will overcome this problem and may provide a very useful tool with which to study CESCs biology. Materials and Methods Cells isolated from CEP were cultured in a three-dimensional agarose suspension to acquire proliferative cell clusters. As previous method, we acquire primary CESCs. Then with these cells we showed that silencing p53 expression with lentivirus-mediated small interfering RNA and overexpressing human telomerase reverse transcriptase (hTERT). Results The effects of p53 knockdown and hTERT overexpression on CESCs, including proliferation, colony formation, and differentiation, were determined. The resultant immortal CESCs displayed similar surface antigen profile to primary CESCs and retained CESCs differentiation potential. Gene expression profile showed high similarity between immortalized CESCs and primary CESCs. In addition, immortalization-associated genes were also identified. Conclusion This study provides a new stem cell resources for basic studies of characteristics of CESCs and understanding the mechanism of disc degeneration. Disclosure of Interest None declared

Macrophage Migration Inhibitory Factor Inhibits the Migration of Cartilage End Plate-Derived Stem Cells by Reacting CD74

Introduction Macrophage migration inhibitory factor (MIF) is a multifunctional cytokine that regulates inflammatory reactions and the pathophysiology of many inflammatory diseases. Intervertebral disc (IVD) degeneration is characterized by an inflammatory reaction, but the potential role of MIF in IVD degeneration has not yet been determined. Recent studies have shown that MIF and its receptor, CD74, are involved in regulating the migration of human mesenchymal stem cells (MSCs). Thus, MIF might impair the ability of mesenchymal stem cells (MSCs) to home to injured tissues. Our previous studies indicated that cartilage endplate-derived stem cells (CESCs) as a type of MSCs exist in human degenerate IVDs. Here, we investigate the role of MIF in regulating the migration of CESCs. Material and Method CESCs were isolated and identified. Results We have shown that MIF was distributed in human degenerate IVD tissues and was subject to regulation by the proinflammatory cytokine tumor necrosis factor-α. Furthermore, in-vitro cell migration assays revealed that nucleus pulposus (NP) cells inhibited the migration of CESCs in a number-dependent manner, and enzyme-linked immunosorbent assays revealed that the amount of MIF in conditioned medium was significantly increased as a function of increasing cell number. In addition, recombinant human MIF inhibited the migration of CESCs in a dose-dependent manner. CESCs migration was restored when an antagonist of MIF, (S, R)-3(4-hydroxyphenyl)-4, 5-dihydro-5-isoxazole acetic acid methyl ester (ISO-1), was added. Finally, a CD74 activating antibody (CD74Ab) was used to examine the effect of CD74 on CESCs motility and inhibited the migration of CESCs in a dose-dependent manner. Conclusion We have identified and characterized a novel regulatory mechanism governing cell migration during IVD degeneration. The results will benefit understanding of another possible mechanism for IVD degeneration, and might provide a new method to repair degenerate IVD by enhancing CESCs migration to degenerated NP tissues to exert their regenerative effects. Disclosure of Interest None declared

Utilization of Stem Cells in Alginate for Nucleus Pulposus Tissue Engineering

Introduction In a general view of anatomy, intervertebral disc is composed of three parts: annulus fibrosus, nucleus pulposus (NP), and cartilage endplate. Recently, several types of stem cells were successfully isolated from these corresponding regions, but up to now, no research was performed about which kind of stem cells is the most efficient candidate for NP tissue engineering or for stem cell-based disc regeneration therapy. Materials and Methods In this study, we compared the regenerative potentials of the above-mentioned three kinds of disc-derived stem cells with that of the classic bone marrow (BM)-mesenchymal stem cells (MSCs) in a rabbit disc degeneration model. By magnetic resonance imaging (MRI), X-ray, histology, and so on. Results We found that cartilage endplate-derived stem cells (CESCs) showed superior capacity compared with the annulus fibrosus-derived stem cells (AFSCs), nucleus pulposus-derived stem cells (NPSCs), and BM-MSCs ( p< 0.05); in addition, when comparing the CESC group with the normal control group, there existed no statistical difference in X-ray ( p>0.05). Those results demonstrated that the CESC-seeded alginate construct performed the most powerful ability for NP regeneration, while AFSCs showed the most inferior potency, NPSCs and BM-MSCs had similar regenerative capacity and located in the middle. Conclusion Our study showed that CESCs might act as an efficient seed cell source for NP tissue engineering, which paved a new way for the biological solution of disc degeneration diseases. Disclosure of Interest None declared

Utilization of Stem Cells in Alginate for Nucleus Pulposus Tissue Engineering

In a general view of anatomy, intervertebral disc is composed of three parts: annulus fibrosus (AF), nucleus pulposus (NP), and cartilage endplate (CEP). Recently, several types of stem cells were successfully isolated from these corresponding regions, but up to now, no research was performed about which kind of stem cells is the most efficient candidate for NP tissue engineering or for stem cell-based disc regeneration therapy. In this study, we compared the regenerative potentials of the above-mentioned three kinds of disc-derived stem cells with that of the classic bone marrow (BM)-mesenchymal stem cells (MSCs) in a rabbit disc degeneration model. By magnetic resonance imaging (MRI), X-ray, histology, etc. evaluations, we found that cartilage endplate-derived stem cells (CESCs) showed superior capacity compared with the annulus fibrosus-derived stem cells (AFSCs), nucleus pulposus-derived stem cells (NPSCs), and BM-MSCs (p<0.05); additionally, when comparing the CESC group with the normal control group, there existed no statistical difference in X-ray (p>0.05). Those results demonstrated that the CESC-seeded alginate construct performed the most powerful ability for NP regeneration, while AFSCs showed the most inferior potency, NPSCs and BM-MSCs had similar regenerative capacity and located in the middle. All in all, our study showed that CESCs might act as an efficient seed cell source for NP tissue engineering, which paved a new way for the biological solution of disc degeneration diseases.

Correction: Macrophage Migration Inhibitory Factor Inhibits the Migration of Cartilage End Plate-Derived Stem Cells by Reacting with CD74

Correction: Macrophage Migration Inhibitory Factor Inhibits the Migration of Cartilage End Plate-Derived Stem Cells by Reacting with CD74

Macrophage migration inhibitory factor inhibits the migration of cartilage end plate-derived stem cells by reacting with CD74.

BackgroundMacrophage migration inhibitory factor (MIF) is a multifunctional cytokine that regulates inflammatory reactions and the pathophysiology of many inflammatory diseases. Intervertebral disc (IVD) degeneration is characterized by an inflammatory reaction, but the potential role of MIF in IVD degeneration has not been determined. Recent studies have shown that MIF and its receptor, CD74, are involved in regulating the migration of human mesenchymal stem cells (MSCs); Thus, MIF might impair the ability of mesenchymal stem cells (MSCs) to home to injured tissues. Our previous studies indicated that cartilage endplate (CEP)-derived stem cells (CESCs) as a type of MSCs exist in human degenerate IVDs. Here, we investigate the role of MIF in regulating the migration of CESCs.Methods and FindingsCESCs were isolated and identified. We have shown that MIF was distributed in human degenerate IVD tissues and was subject to regulation by the pro-inflammatory cytokine TNF-α. Furthermore, in vitro cell migration assays revealed that nucleus pulposus (NP) cells inhibited the migration of CESCs in a number-dependent manner, and ELISA assays revealed that the amount of MIF in conditioned medium (CM) was significantly increased as a function of increasing cell number. Additionally, recombinant human MIF (r-MIF) inhibited the migration of CESCs in a dose-dependent manner. CESCs migration was restored when an antagonist of MIF, (S, R)-3(4-hydroxyphenyl)-4, 5-dihydro-5-isoxazole acetic acid methyl ester (ISO-1), was added. Finally, a CD74 activating antibody (CD74Ab) was used to examine the effect of CD74 on CESCs motility and inhibited the migration of CESCs in a dose-dependent manner.ConclusionsWe have identified and characterized a novel regulatory mechanism governing cell migration during IVD degeneration. The results will benefit understanding of another possible mechanism for IVD degeneration, and might provide a new method to repair degenerate IVD by enhancing CESCs migration to degenerated NP tissues to exert their regenerative effects.