Single Cell RNA-Sequence Analyses Reveal Uniquely Expressed Genes and Heterogeneous Immune Cell Involvement in the Rat Model of Intervertebral Disc Degeneration
Intervertebral disc (IVD) degeneration is characterized by a loss of cellularity, and changes in cell-mediated activity that drives anatomic changes to IVD structure. In this study, we used single-cell RNA-sequencing analysis of degenerating tissues of the rat IVD following lumbar disc puncture. Two...
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MDPI AG
2022-08-01
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| Online Access: | https://www.mdpi.com/2076-3417/12/16/8244 |
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| author | Milad Rohanifar Sade W. Clayton Garrett W.D. Easson Deepanjali S. Patil Frank Lee Liufang Jing Marcos N. Barcellona Julie E. Speer Jordan J. Stivers Simon Y. Tang Lori A. Setton |
| author_facet | Milad Rohanifar Sade W. Clayton Garrett W.D. Easson Deepanjali S. Patil Frank Lee Liufang Jing Marcos N. Barcellona Julie E. Speer Jordan J. Stivers Simon Y. Tang Lori A. Setton |
| author_sort | Milad Rohanifar |
| collection | DOAJ |
| description | Intervertebral disc (IVD) degeneration is characterized by a loss of cellularity, and changes in cell-mediated activity that drives anatomic changes to IVD structure. In this study, we used single-cell RNA-sequencing analysis of degenerating tissues of the rat IVD following lumbar disc puncture. Two control, uninjured IVDs (L2-3, L3-4) and two degenerated, injured IVDs (L4-5, L5-6) from each animal were examined either at the two- or eight-week post-operative time points. The cells from these IVDs were extracted and transcriptionally profiled at the single-cell resolution. Unsupervised cluster analysis revealed the presence of four known cell types in both non-degenerative and degenerated IVDs based on previously established gene markers: IVD cells, endothelial cells, myeloid cells, and lymphoid cells. As a majority of cells were associated with the IVD cell cluster, sub-clustering was used to further identify the cell populations of the nucleus pulposus, inner and outer annulus fibrosus. The most notable difference between control and degenerated IVDs was the increase of myeloid and lymphoid cells in degenerated samples at two- and eight-weeks post-surgery. Differential gene expression analysis revealed multiple distinct cell types from the myeloid and lymphoid lineages, most notably macrophages and B lymphocytes, and demonstrated a high degree of immune specificity during degeneration. In addition to the heterogenous infiltrating immune cell populations in the degenerating IVD, the increased number of cells in the AF sub-cluster expressing <i>Ngf</i> and <i>Ngfr</i>, encoding for p75NTR, suggest that NGF signaling may be one of the key mediators of the IVD crosstalk between immune and neuronal cell populations. These findings provide the basis for future work to understand the involvement of select subsets of non-resident cells in IVD degeneration. |
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| institution | Directory Open Access Journal |
| issn | 2076-3417 |
| language | English |
| last_indexed | 2024-03-09T04:44:29Z |
| publishDate | 2022-08-01 |
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| spelling | doaj.art-f4770016de664b1696cc5ffc9054cc5f2023-12-03T13:18:03ZengMDPI AGApplied Sciences2076-34172022-08-011216824410.3390/app12168244Single Cell RNA-Sequence Analyses Reveal Uniquely Expressed Genes and Heterogeneous Immune Cell Involvement in the Rat Model of Intervertebral Disc DegenerationMilad Rohanifar0Sade W. Clayton1Garrett W.D. Easson2Deepanjali S. Patil3Frank Lee4Liufang Jing5Marcos N. Barcellona6Julie E. Speer7Jordan J. Stivers8Simon Y. Tang9Lori A. Setton10Department of Biomedical Engineering, Washington University in St. Louis, St. Louis, MO 63130, USADepartment of Orthopedic Surgery, Washington University School of Medicine, St. Louis, MO 63110, USADepartment of Orthopedic Surgery, Washington University School of Medicine, St. Louis, MO 63110, USADepartment of Biomedical Engineering, Washington University in St. Louis, St. Louis, MO 63130, USADepartment of Biomedical Engineering, Washington University in St. Louis, St. Louis, MO 63130, USADepartment of Biomedical Engineering, Washington University in St. Louis, St. Louis, MO 63130, USADepartment of Biomedical Engineering, Washington University in St. Louis, St. Louis, MO 63130, USADepartment of Biomedical Engineering, Washington University in St. Louis, St. Louis, MO 63130, USADepartment of Orthopedic Surgery, Washington University School of Medicine, St. Louis, MO 63110, USADepartment of Orthopedic Surgery, Washington University School of Medicine, St. Louis, MO 63110, USADepartment of Biomedical Engineering, Washington University in St. Louis, St. Louis, MO 63130, USAIntervertebral disc (IVD) degeneration is characterized by a loss of cellularity, and changes in cell-mediated activity that drives anatomic changes to IVD structure. In this study, we used single-cell RNA-sequencing analysis of degenerating tissues of the rat IVD following lumbar disc puncture. Two control, uninjured IVDs (L2-3, L3-4) and two degenerated, injured IVDs (L4-5, L5-6) from each animal were examined either at the two- or eight-week post-operative time points. The cells from these IVDs were extracted and transcriptionally profiled at the single-cell resolution. Unsupervised cluster analysis revealed the presence of four known cell types in both non-degenerative and degenerated IVDs based on previously established gene markers: IVD cells, endothelial cells, myeloid cells, and lymphoid cells. As a majority of cells were associated with the IVD cell cluster, sub-clustering was used to further identify the cell populations of the nucleus pulposus, inner and outer annulus fibrosus. The most notable difference between control and degenerated IVDs was the increase of myeloid and lymphoid cells in degenerated samples at two- and eight-weeks post-surgery. Differential gene expression analysis revealed multiple distinct cell types from the myeloid and lymphoid lineages, most notably macrophages and B lymphocytes, and demonstrated a high degree of immune specificity during degeneration. In addition to the heterogenous infiltrating immune cell populations in the degenerating IVD, the increased number of cells in the AF sub-cluster expressing <i>Ngf</i> and <i>Ngfr</i>, encoding for p75NTR, suggest that NGF signaling may be one of the key mediators of the IVD crosstalk between immune and neuronal cell populations. These findings provide the basis for future work to understand the involvement of select subsets of non-resident cells in IVD degeneration.https://www.mdpi.com/2076-3417/12/16/8244intervertebral disc degenerationsingle-cell RNA sequencingcell type |
| spellingShingle | Milad Rohanifar Sade W. Clayton Garrett W.D. Easson Deepanjali S. Patil Frank Lee Liufang Jing Marcos N. Barcellona Julie E. Speer Jordan J. Stivers Simon Y. Tang Lori A. Setton Single Cell RNA-Sequence Analyses Reveal Uniquely Expressed Genes and Heterogeneous Immune Cell Involvement in the Rat Model of Intervertebral Disc Degeneration Applied Sciences intervertebral disc degeneration single-cell RNA sequencing cell type |
| title | Single Cell RNA-Sequence Analyses Reveal Uniquely Expressed Genes and Heterogeneous Immune Cell Involvement in the Rat Model of Intervertebral Disc Degeneration |
| title_full | Single Cell RNA-Sequence Analyses Reveal Uniquely Expressed Genes and Heterogeneous Immune Cell Involvement in the Rat Model of Intervertebral Disc Degeneration |
| title_fullStr | Single Cell RNA-Sequence Analyses Reveal Uniquely Expressed Genes and Heterogeneous Immune Cell Involvement in the Rat Model of Intervertebral Disc Degeneration |
| title_full_unstemmed | Single Cell RNA-Sequence Analyses Reveal Uniquely Expressed Genes and Heterogeneous Immune Cell Involvement in the Rat Model of Intervertebral Disc Degeneration |
| title_short | Single Cell RNA-Sequence Analyses Reveal Uniquely Expressed Genes and Heterogeneous Immune Cell Involvement in the Rat Model of Intervertebral Disc Degeneration |
| title_sort | single cell rna sequence analyses reveal uniquely expressed genes and heterogeneous immune cell involvement in the rat model of intervertebral disc degeneration |
| topic | intervertebral disc degeneration single-cell RNA sequencing cell type |
| url | https://www.mdpi.com/2076-3417/12/16/8244 |
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