Differential phenotypic behaviors of human degenerative nucleus pulposus cells under normoxic and hypoxic conditions: influence of oxygen concentration during isolation, expansion, and cultivation

Yang, Shuai; Hu, Ming-Hsiao; Sun, Yuan-Hui; Lin, Feng-Huei

doi:10.1016/j.spinee.2013.05.025

Cited by 22 publications

(25 citation statements)

References 31 publications

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“…Given that a lower O 2 tension has been shown to facilitate ECM deposition [ 23 ], the effect of human (s)Link-N on CD canine CLCs was also studied under hypoxia ( S3 Fig ). It appeared that the effects of human (s)Link-N were not affected by the O 2 tension: in both conditions, human (s)Link-N exerted a limited anabolic effect compared with controls.…”

Section: Resultsmentioning

confidence: 99%

“…It appeared that the effects of human (s)Link-N were not affected by the O 2 tension: in both conditions, human (s)Link-N exerted a limited anabolic effect compared with controls. Nevertheless, since the IVD is an avascular structure, hypoxia better mimics the in vivo situation and has been shown to better preserve the regenerative potential of CLCs [ 23 , 24 ], follow-up experiments (e.g. using canine (s)Link-N) were continued in hypoxia.…”

Section: Resultsmentioning

confidence: 99%

“…Follow-up culture experiments (i.e. using canine (s)Link-N) where performed at 5% O 2 , 5% CO 2 , 37°C, to improve the chondrogenic response of the CLCs [ 23 , 24 ].…”

Section: Methodsmentioning

confidence: 99%

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Link-N: The missing link towards intervertebral disc repair is species-specific

et al. 2017

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IntroductionDegeneration of the intervertebral disc (IVD) is a frequent cause for back pain in humans and dogs. Link-N stabilizes proteoglycan aggregates in cartilaginous tissues and exerts growth factor-like effects. The human variant of Link-N facilitates IVD regeneration in several species in vitro by inducing Smad1 signaling, but it is not clear whether this is species specific. Dogs with IVD disease could possibly benefit from Link-N treatment, but Link-N has not been tested on canine IVD cells. If Link-N appears to be effective in canines, this would facilitate translation of Link-N into the clinic using the dog as an in vivo large animal model for human IVD degeneration.Materials and methodsThis study’s objective was to determine the effect of the human and canine variant of Link-N and short (s) Link-N on canine chondrocyte-like cells (CLCs) and compare this to those on already studied species, i.e. human and bovine CLCs. Extracellular matrix (ECM) production was determined by measuring glycosaminoglycan (GAG) content and histological evaluation. Additionally, the micro-aggregates’ DNA content was measured. Phosphorylated (p) Smad1 and -2 levels were determined using ELISA.ResultsHuman (s)Link-N induced GAG deposition in human and bovine CLCs, as expected. In contrast, canine (s)Link-N did not affect ECM production in human CLCs, while it mainly induced collagen type I and II deposition in bovine CLCs. In canine CLCs, both canine and human (s)Link-N induced negligible GAG deposition. Surprisingly, human and canine (s)Link-N did not induce Smad signaling in human and bovine CLCs. Human and canine (s)Link-N only mildly increased pSmad1 and Smad2 levels in canine CLCs.ConclusionsHuman and canine (s)Link-N exerted species-specific effects on CLCs from early degenerated IVDs. Both variants, however, lacked the potency as canine IVD regeneration agent. While these studies demonstrate the challenges of translational studies in large animal models, (s)Link-N still holds a regenerative potential for humans.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

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Link-N: The missing link towards intervertebral disc repair is species-specific

et al. 2017

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“…Furthermore, the human CLC microaggregates were cultured under normoxic conditions. CLCs may respond more pronounced to the bioactive NCsecreted factors under hypoxic than normoxic conditions given the hypoxic nature of the NP (Feng et al, 2013;Yang et al, 2013). Therefore, further research under hypoxic conditions is necessary to confirm the obtained results with CLCs from several donors with different IVD degeneration grades.…”

Section: Ncs Of Different Species Regenerate Human Clcsmentioning

confidence: 91%

The species-specific regenerative effects of notochordal cell-conditioned medium on chondrocyte-like cells derived from degenerated human intervertebral discs

Bach

Vries

Krouwels

et al. 2015

eCM

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During intervertebral disc (IVD) maturation, the main cell type shifts from notochordal cells (NCs) to chondrocytelike cells (CLCs). NCs secrete factors with regenerative potential, making them an interesting focus for regenerative treatments. During initial development, these strategies preferably employ non-human donors due to easy availability of their NC-rich nucleus pulposus (NP) tissue. To increase the success of translating these strategies for clinical application, this study aimed to delineate whether NC-secreted factors of different species have a regenerative effect on human CLCs. Human, canine and porcine NC-rich NP tissue and NC-conditioned medium (NCCM) were analysed biochemically and histologically. Human CLC micro-aggregates from degenerated IVDs were cultured in human, canine or porcine NCCM. Collagen, glycosaminoglycan (GAG) and DNA content was determined and histology was performed. Canine and porcine NPs were richer in NCs than human NPs. Human NPs contained the highest collagen content, whereas the DNA and GAG content of canine NPs was significantly higher than that of human or porcine NPs. NCCM from all species significantly increased the DNA and GAG content of the human CLC micro-aggregates. Porcine and canine NCCM were significantly more potent than human NCCM in inducing GAG deposition, whereas only human NCCM induced collagen type II production. Secreted factors from human, canine and porcine NC-rich NPs exerted regenerative effects on human CLCs, indicating a cross-species effect. Bioactive compound(s) are present in NCCM of different species that may reverse human IVD degeneration, supporting further research into strategies based on NC-technology employing canine or porcine models for their translation into humans.

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“…IVD cells typically experience a dedifferentiation process when grown in monolayer culture [45]. Human NP cells cultivated under normoxia displayed poorer expressions of aggrecan and type II collagen while promoting expressions of MMPs [46]. Not only compromising the overall phenotypic behaviors, serial passages in monolayer and cultivation under normoxic environment also diminish the percentage positivity of CD24 in human NP cells.…”

Section: Discussionmentioning

confidence: 99%

CD24 expression indicates healthier phenotype and less tendency of cellular senescence in human nucleus pulposus cells

Yang

et al. 2019

Artificial Cells, Nanomedicine, and Biotechnology

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Identification of specific cell markers is crucial for recognizing functionally healthy nucleus pulposus (NP) cells. The objective of this study was to investigate the role of CD24 expression in adult human NP cells. Cells were retrieved from NP tissues of 20 patients (aged 17-44) operated on for lumbar disc herniation. Based on CD24 expression, NP cells were separated by sorting and then used to examine phenotypic behavior, the effects of culture conditions and cellular senescence pathway related proteins. CD24 expression was positive in 35.5 ± 3.7% (range 9.1-65.2%) of NP cells. Consistently, normoxic expansion and serial passages in monolayers decreased percentage positivity for CD24 in NP cells. CD24 -NP cells showed a markedly decreased GSK-3b activity and increased mitogen-activated protein kinase phosphorylation accompanying by an increased b-catenin expression. Higher levels of matrix metalloproteinases, as well as lower levels of ACAN and COL2 in CD24cells, indicated the breakdown and reduced the formation of key extracellular matrix components. CD24 þ NP cells presented a more favorable phenotype while CD24cells showed a more prominent cellular senescence fate. CD24 in NP cells may be a surrogate marker of healthy cells, in the cell-based therapeutic treatment of degenerative disc disorders. ARTICLE HISTORY

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Differential phenotypic behaviors of human degenerative nucleus pulposus cells under normoxic and hypoxic conditions: influence of oxygen concentration during isolation, expansion, and cultivation

Cited by 22 publications

References 31 publications

Link-N: The missing link towards intervertebral disc repair is species-specific

Link-N: The missing link towards intervertebral disc repair is species-specific

The species-specific regenerative effects of notochordal cell-conditioned medium on chondrocyte-like cells derived from degenerated human intervertebral discs

CD24 expression indicates healthier phenotype and less tendency of cellular senescence in human nucleus pulposus cells

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