Bovine annulus fibrosus cell lines isolated from intervertebral discs

Kraus, Petra; Lufkin, Thomas

doi:10.1016/j.gdata.2016.09.012

Cited by 11 publications

(11 citation statements)

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“…In the IVD, AP‐RISH identified a significantly higher number of cells expressing Sox2 ( P < 0.001), Ptprc ( P < 0.001) and Zscan10 ( P < 0.001) in the NP tissue and significantly more Thy1 ‐expressing cells in the AF ( P = 0.002), whereas the proportion of cells expressing Esrrb, Nanog, Oct4 and Eng showed no significant difference (Table , Figs , ). This difference between AF and NP cell populations might be relevant for their therapeutic potential, however, we have previously demonstrated that cells can be isolated from all three tissues, outer AF, inner AF (TZ) and NP, and propagated in vitro in 2D monolayer culture under normal oxygen and zero‐applied pressure (Kraus & Lufkin, ; Kraus et al. ), which are common culture conditions but are unusual in vivo .…”

Section: Resultsmentioning

confidence: 99%

Potential biomarkers of the mature intervertebral disc identified at the single cell level

Kapper

Youngs

et al. 2018

Journal of Anatomy

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Intervertebral disc (IVD) degeneration and trauma is a major socio-economic burden and the focus of cell-based regenerative medicine approaches. Despite numerous ongoing clinical trials attempting to replace ailing IVD cells with mesenchymal stem cells, a solid understanding of the identity and nature of cells in a healthy mature IVD is still in need of refinement. Although anatomically simple, the IVD is comprised of heterogeneous cell populations. Therefore, methods involving cell pooling for RNA profiling could be misleading. Here, by using RNA in situ hybridization and z proportion test, we have identified potential novel biomarkers through single cell assessment. We quantified the proportion of RNA transcribing cells for 50 genetic loci in the outer annulus fibrosus (AF) and nucleus pulposus (NP) in coccygeal bovine discs isolated from tails of four skeletally mature animals. Our data reconfirm existing data and suggest 10 novel markers such as Lam1 and Thy1 in the outer AF and Gli1, Gli3, Noto, Scx, Ptprc, Sox2, Zscan10 and LOC101904175 in the NP, including pluripotency markers, that indicate stemness potential of IVD cells. These markers could be added to existing biomarker panels for cell type characterization. Furthermore, our data once more demonstrate heterogeneity in cells of the AF and NP, indicating the need for single cell assessment by methods such as RNA in situ hybridization. Our work refines the molecular identity of outer AF and NP cells, which can benefit future regenerative medicine and tissue engineering strategies in humans.

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Section: Resultsmentioning

confidence: 99%

Potential biomarkers of the mature intervertebral disc identified at the single cell level

Kapper

Youngs

et al. 2018

Journal of Anatomy

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“…We have previously identified positive and negative cells for T expression in cells of the AF and NP, however, with no significant difference in cell population proportions, indicating that assessment of heterogeneity within a cell population remains important to address the question of NP cell origin and heterogeneity [3], whether within their native environment or in culture. We have frequently isolated and propagated bovine cells from the outer AF and the NP through non-enzymatic explant outgrowth culture [20,77]. These cells exhibit some criteria established for mesenchymal stem cells, but continue to display heterogeneity when phenotyped by a selection of transcripts, by morphology or by cell movement [2,20,77,78,79].…”

Section: Discussionmentioning

confidence: 99%

“…We have frequently isolated and propagated bovine cells from the outer AF and the NP through non-enzymatic explant outgrowth culture [20,77]. These cells exhibit some criteria established for mesenchymal stem cells, but continue to display heterogeneity when phenotyped by a selection of transcripts, by morphology or by cell movement [2,20,77,78,79]. In this context others and we find morphological, molecular and “behavioral” differences in vivo and in vitro pointing to at least two cell populations within the outer AF and NP [2,3,61,68,73].…”

Section: Discussionmentioning

confidence: 99%

Quantitative Single-Cell Transcript Assessment of Biomarkers Supports Cellular Heterogeneity in the Bovine IVD

Kapper

Mondal

et al. 2019

Veterinary Sciences

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Severe and chronic low back pain is often associated with intervertebral disc (IVD) degeneration. While imposing a considerable socio-economic burden worldwide, IVD degeneration is also severely impacting on the quality of life of affected individuals. Cell-based regenerative medicine approaches have moved into clinical trials, yet IVD cell identities in the mature disc remain to be fully elucidated and tissue heterogeneity exists, requiring a better characterization of IVD cells. The bovine coccygeal IVD is an accepted research model to study IVD mechano-biology and disc homeostasis. Recently, we identified novel IVD biomarkers in the outer annulus fibrosus (AF) and nucleus pulposus (NP) of the mature bovine coccygeal IVD through RNA in situ hybridization (AP-RISH) and z-proportion test. Here we follow up on Lam1, Thy1, Gli1, Gli3, Noto, Ptprc, Scx, Sox2 and Zscan10 with fluorescent RNA in situ hybridization (FL-RISH) and confocal microscopy. This permits sub-cellular transcript localization and the addition of quantitative single-cell derived values of mRNA expression levels to our previous analysis. Lastly, we used a Gaussian mixture modeling approach for the exploratory analysis of IVD cells. This work complements our earlier cell population proportion-based study, confirms the previously proposed biomarkers and indicates even further heterogeneity of cells in the outer AF and NP of a mature IVD.

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“…The IVD is an unique non-vascularized fibrocartilage tissue consisting of a lamellar-structured circular AF surrounding the central NP core and incased above (superior) and below (inferior) by the vascularized cartilage endplates through which nutrients can diffuse to the AF and NP (Sivakamasundari and Lufkin, 2012; Kraus and Lufkin, 2016). The IVD is the largest non-vascularized tissue in the body.…”

Section: Developmental Biology Of the Intervertebral Discmentioning

confidence: 99%

Implications for a Stem Cell Regenerative Medicine Based Approach to Human Intervertebral Disk Degeneration

Kraus

Lufkin

2017

Front. Cell Dev. Biol.

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The human body develops from a single cell, the zygote, the product of the maternal oocyte and the paternal spermatozoon. That 1-cell zygote embryo will divide and eventually grow into an adult human which is comprised of ~3.7 × 1013 cells. The tens of trillions of cells in the adult human can be classified into approximately 200 different highly specialized cell types that make up all of the different tissues and organs of the human body. Regenerative medicine aims to replace or restore dysfunctional cells, tissues and organs with fully functional ones. One area receiving attention is regeneration of the intervertebral discs (IVDs), which are located between the vertebrae and function to give flexibility and support load to the spine. Degenerated discs are a major cause of lower back pain. Different stem cell based regenerative medicine approaches to cure disc degeneration are now available, including using autologous mesenchymal stem cells (MSCs), induced pluripotent stem cells (iPSCs) and even attempts at direct transdifferentiation of somatic cells. Here we discuss some of the recent advances, successes, drawbacks, and the failures of the above-mentioned approaches.

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Bovine annulus fibrosus cell lines isolated from intervertebral discs

Cited by 11 publications

References 11 publications

Potential biomarkers of the mature intervertebral disc identified at the single cell level

Potential biomarkers of the mature intervertebral disc identified at the single cell level

Quantitative Single-Cell Transcript Assessment of Biomarkers Supports Cellular Heterogeneity in the Bovine IVD

Implications for a Stem Cell Regenerative Medicine Based Approach to Human Intervertebral Disk Degeneration

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