Human anulus fibrosis and nucleus pulposus cells of the intervertebral disc -: Effect of degeneration and culture system on cell phenotype
Human anulus fibrosis and nucleus pulposus cells of the intervertebral disc -: Effect of degeneration and culture system on cell phenotype
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DOI：
10.1097/01.brs.0000192204.89160.6d
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发表时间：
2005-12-15
期刊：
影响因子：
3
通讯作者：
Gründer, T
中科院分区：
文献类型：
--
作者：
Kluba, T;Niemeyer, T;Gründer, T
研究设计。人腰椎间盘细胞取自青少年特发性脊柱侧凸(AIS)患者和退行性腰椎间盘疾病供者。从髓核(NP)中分离出纤维环(AF)，并在两种不同的细胞培养模型中分别培养细胞。目的：研究青少年特发性脊柱侧弯患者椎间盘细胞在体外扩增过程中基因表达的变化，并确定青少年特发性脊柱侧弯供者的细胞与退行性腰椎间盘疾病手术患者的细胞相比是否表现出不同的基因表达谱。在体外扩增过程中，细胞经历了一个去分化过程，其特征是基因表达的切换。人类椎间盘细胞分化和去分化状态的标志物尚不清楚。此外，目前尚不清楚在退变过程中是否会发生基因表达模式的变化。在控制和确定的条件下，用单层和藻酸盐培养法扩增AF和NP组织中的细胞。然后收集细胞，用实时定量聚合酶链式反应(PCR)进行表型分析。分析脊柱侧弯和退变的人椎间盘细胞中I、II、X型胶原、聚集素和白细胞介素1β的表达。在单层扩增过程中，两种细胞的II型和X型胶原及聚集素的基因表达均显著降低。在藻酸盐培养的细胞中观察到所有基因的重新表达。此外，与来自脊柱侧弯供者的NP细胞相比，来自退变组织的NP细胞的II型胶原水平显著降低。这些结果有助于更好地理解体外扩增对健康和退变的椎间盘细胞表型的影响。这可能对未来的组织工程用途有用。
Study Design. Human intervertebral disc cells were harvested from patients with adolescent idiopathic scoliosis (AIS) and from donors with degenerative disc disease. Anulus fibrosis (AF) was separated from nucleus pulposus (NP), and cells were cultured separately in two different cell culture models.Objectives. To investigate changes in gene expression of human disc cells during in vitro expansion and to determine whether cells from adolescent idiopathic scoliosis donors show different gene expression profiles compared with cells from patients operated for degenerative disc disease.Summary of Background Data. During in vitro expansion, cells undergo a dedifferentiation process, which is characterized by a switch in gene expression. Markers for the differentiation and dedifferentiation status of human disc cells are not yet known. Moreover, it is not known whether changes in the gene expression pattern occur during the degeneration process.Methods. Cells from AF and NP tissues were expanded in monolayer and alginate cultures under controlled and defined conditions. Cells were then harvested, and analysis of phenotype was performed using quantitative real-time polymerase chain reaction (PCR). The mRNA expression of Type I, II, and X collagen, aggrecan, and interleukin-1 beta in scoliosis and degenerative human intervertebral disc cells was analyzed.Results. The gene expression of Type II and X collagen and of aggrecan significantly decreased for both cell types during monolayer expansion. Reexpression of all genes was observed when cells were cultured in alginate. Additionally, NP cells from degenerative tissues displayed significant lower levels of Type II collagen compared with NP cells from scoliosis donors.Conclusions. These results provide a better understanding of how the phenotype of human healthy and degenerative disc cells is influenced by in vitro expansion. This may be useful for future tissue engineering purposes.