Adequate Serial Monolayer Passage Number of Human Intervertebral Disc Cells for Cell Therapy -Growth and Phenotype of Cells-

김용찬; Ki Bok Kim; Moon Soo Park; 김석우

Study Design: This is an in-vitro experimental study Objectives: We wanted to analyze the changes in the growth and phenotype of human degenerative intervertebral disc cells depending on the frequency of subculture in an in vitro monolayer culture system. Summary of the Literature Review: A subculture of disc cells is needed to obtain an adequate amount of disc cells for cell therapy,tissue engineering and analysis of the biological characteristics of degenerative disc cells Materials and Methods: The obtained intervertebral discs were divided into the nucleus pulposus (NP) and the annulus fibrosus (AF). The AF and NP cells were cultured in a monolayer manner, respectively. At each subculture time, we analyzed the morphological changes,the adhesion rate, the proliferation rate and the viability. The expressions of types I and II collagen and proteoglycan were analyzed at the mRNA gene level. Results: Both the AF and NP cells gradually showed a fibroblast-like spindle shape while undergoing subculture. The adhesion rate was higher at the second and third times of subculture. The cell proliferation was the highest at the second subculture time. The viability was markedly lower prior to the subculture. On RT-PCR, the type II collagen expression was gradually decreased in the NP cells. In the AF cells, Type II collagen was not expressed from the second time of subculture. The expression of proteoglycan was gradually decreased in both. Conclusions: Following the 3rd subculture, the degenerative disc cells had completely changed their original growth and phenotypic characteristics. Therefore, we believe that it is not desirable for us to do passage cultures more than three times for cell therapy.

Citation status

KCI Citation Counts (0)
REFERENCES (30)
열림/닫힘 버튼
* 2021년 이후 발행 논문의 참고문헌은 현재 구축 중입니다.

[journal] Urban JP / 2003 / Degeneration of the intervertebral disc / Arthritis Res Ther 5 : 120~130

[journal] Freemont AJ / 2002 / Current understanding of cellular and molecular events in intervertebral disc degeneration: implications for therapy / J Pathol 196 : 374~379

[journal] Hoppenfeld S / 1989 / Percutaneous removal of herniated lumbar discs. 50 cases with ten-year follow-up periods / Clin Orthop Relat Res 238 : 92~97

[journal] Yeung AT / 2002 / Posterolateral endoscopic excision for lumbar disc herniation: Surgical technique, outcome, and complications in 307 consecutive cases / Spine 27 : 722~731

[journal] White AH / 1987 / Lumbar laminectomy for herniated disc: a prospective controlled comparison with internal fixation fusion / Spine 12 : 305~307

[journal] Eie N / 1978 / Comparison of the results in patients operated upon for ruptured lumbar discs with and without spinal fusion / Acta Neurochir 41 : 107~113

[journal] Eie N / 1983 / The knee-elbow position in lumbar disc surgery: a review of complications / Spine 8 : 897~900

[journal] Freeman BJ / 2006 / Total disc replacement in the lumbar spine: a systematic review of the literature / Eur Spine J 15 : 439~447

[journal] An HS / 2003 / Biological repair of intervertebral disc / Spine 28 : 86~92

[journal] Ganey TM / 2002 / A potential role for cell-based therapeutics in the treatment of intervertebral disc herniation / Eur Spine J 11 : 206~214

[journal] Gan JC / 2003 / Intervertebral disc tissue engineering I: characterization of the nucleus pulposus / Clin Orthop Relat Res 411 : 305~314

[journal] Yoon ST / 2005 / Molecular therapy of the intervertebral disc / Spine J 5 : 280~286

[journal] Baums MH / 2006 / Autologous chondrocyte transplantation for treating cartilage defects of the talus / J Bone Joint Surg Am 88 : 303~308

[journal] Brittberg M / 1994 / Treatment of deep cartilage defects in the knee with autologous chondrocyte transplantation / N Engl J Med 331 : 889~895

[journal] Benya PD / 1982 / Dedifferentiated chondrocytes reexpress the differentiated collagen phenotype when cultured in agarose gels / Cell 30 : 215~224

[journal] Lemare F / 1998 / Dedifferentiated chondrocytes cultured in alginate beads: restoration of the differentiated phenotype and of the metabolic responses to interleukin-1beta / J Cell Physiol 176 : 303~313

[journal] Bayliss MT / 1988 / Volvo award in basic science. Proteoglycan synthesis in the human intervertebral disc. Variation with age, region and pathology / Spine 13 : 972~981

[journal] Chou AI / 2006 / The effect of serial monolayer passaging on the collagen expression profile of outer and inner anulus fibrosus cells / Spine 31 : 1875~1881

[journal] Kluba T / 2005 / Human anulus fibrosus and nucleus pulposus cells of the intervertebral disc: effect of degeneration and culture system on cell phenotype / Spine 30 : 2743~2748

[journal] Wang JY / 2001 / Intervertebral disc cells exhibit differences in gene expression in alginate and monolayer culture / Spine 26 : 1747~1751

[journal] Tullberg-Reinert H / 1999 / In situ measurement of collagen synthesis by human bone cells with a sirius red-based colorimetric microassay: effects of transforming growth factor beta2 and ascorbic acid 2-phosphate / Histochem Cell Biol 112 : 271~276

[journal] Farndale RW / 1986 / Improved quantitation and discrimination of sulphated glycosaminoglycans by use of dimethylmethylene blue / Biochim Biophys Acta 883 : 173~177

[journal] Loeser RF / 2000 / Integrin expression by primary and immortalized human chondrocytes: evidence of a differential role for alpha1beta1 and alpha2beta1 integrins in mediating chondrocyte adhesion to types II and VI collagen / Osteoarthritis Cartilage 8 : 96~105

[journal] Wang H / 2004 / Chondrocytes attach to hyaline or calcified cartilage and bone / Osteoarthritis Cartilage 12 : 56~64

[journal] Hong Y / 2005 / Collagen-coated polylactide microspheres as chondrocyte microcarriers / Biomaterials 26 : 6305~6313

[journal] Yashiki S / 2001 / Evaluation of attachment and growth of anchorage-dependent cells on culture surfaces with type I collagen coating / J Biosci Bioeng 92 : 385~388

[journal] Eyre DR / 1977 / Quantitative analysis of types I and II collagens in human intervertebral discs at various ages / Biochim Biophys Acta 492 : 29~42

[journal] Johnstone B / 1995 / The large proteoglycans of the human intervertebral disc. Changes in their biosynthesis and structure with age, topography, and pathology / Spine 20 : 674~684

[journal] Lee JY / 2001 / New use of a three-dimensional pellet culture system for human intervertebral disc cells: initial characterization and potential use for tissue engineering / Spine 26 : 2316~2322

[journal] Sato M / 2003 / An atelocollagen honeycomb-shaped scaffold with a membrane seal (ACHMS-scaffold) for the culture of annulus fibrosus cells from an intervertebral disc / J Biomed Mater Res A 64 : 248~256

KJC Korea
Journal Central

Statistics

Tools

Sharing

Issue List

Citation status