Optimization of chondrocyte isolation and phenotype characterization for cartilage tissue engineering
Current protocols for chondrocyte isolation are inconsistent, resulting in suboptimal cell yield and compromised cell quality. Thus, there is a need for an improved isolation protocol that is able to give a maximum yield with optimal cell viability while preserving the chondrocyte phenotype. In ligh...
| Main Authors: | , , , |
|---|---|
| Other Authors: | |
| Format: | Journal Article |
| Language: | English |
| Published: |
2015
|
| Subjects: | |
| Online Access: | https://hdl.handle.net/10356/106726 http://hdl.handle.net/10220/25104 |
| _version_ | 1811683695860383744 |
|---|---|
| author | Lau, Ting Ting Peck, Yvonne Huang, Weiliang Wang, Dong-An |
| author2 | School of Chemical and Biomedical Engineering |
| author_facet | School of Chemical and Biomedical Engineering Lau, Ting Ting Peck, Yvonne Huang, Weiliang Wang, Dong-An |
| author_sort | Lau, Ting Ting |
| collection | NTU |
| description | Current protocols for chondrocyte isolation are inconsistent, resulting in suboptimal cell yield and compromised cell quality. Thus, there is a need for an improved isolation protocol that is able to give a maximum yield with optimal cell viability while preserving the chondrocyte phenotype. In light of this, we developed an improved isolation protocol based on enzymatic digestion using 0.1% (w/v) collagenase II. Different from existing methods of digesting minced cartilage for a prolonged period (usually 14–16 h), we performed two additional digestions, with a 5- and 3-h interval in between. The results showed that this multiple digestion method was able to yield a total number of cells that are more than a fivefold increase as compared to any of the common isolation protocols. More importantly, a high percentage of the isolated cells remained viable. Furthermore, an evaluation of the effect of additional digestions on chondrocyte phenotype indicated that cells harvested from the second and third digestion showed a comparable or higher proliferative capacity than the first digestion and all the cells expressed chondrocyte-specific markers tested, with cells from the third digestion showing exceptionally high gene expression levels for collagen type II (Col II), aggrecan, and COMP. Additionally, their ability to produce collagen type II as well as their morphology were not affected by the two additional digestions. Taken together, the results suggested that the use of this isolation protocol resulted in a higher cell yield and the quality of the isolated cells was maintained. Hence, we recommend this isolation protocol to be employed for more efficient cell harvesting especially from limited biopsied cartilage tissue samples. |
| first_indexed | 2024-10-01T04:16:50Z |
| format | Journal Article |
| id | ntu-10356/106726 |
| institution | Nanyang Technological University |
| language | English |
| last_indexed | 2024-10-01T04:16:50Z |
| publishDate | 2015 |
| record_format | dspace |
| spelling | ntu-10356/1067262023-12-29T06:47:35Z Optimization of chondrocyte isolation and phenotype characterization for cartilage tissue engineering Lau, Ting Ting Peck, Yvonne Huang, Weiliang Wang, Dong-An School of Chemical and Biomedical Engineering DRNTU::Science::Medicine::Tissue engineering Current protocols for chondrocyte isolation are inconsistent, resulting in suboptimal cell yield and compromised cell quality. Thus, there is a need for an improved isolation protocol that is able to give a maximum yield with optimal cell viability while preserving the chondrocyte phenotype. In light of this, we developed an improved isolation protocol based on enzymatic digestion using 0.1% (w/v) collagenase II. Different from existing methods of digesting minced cartilage for a prolonged period (usually 14–16 h), we performed two additional digestions, with a 5- and 3-h interval in between. The results showed that this multiple digestion method was able to yield a total number of cells that are more than a fivefold increase as compared to any of the common isolation protocols. More importantly, a high percentage of the isolated cells remained viable. Furthermore, an evaluation of the effect of additional digestions on chondrocyte phenotype indicated that cells harvested from the second and third digestion showed a comparable or higher proliferative capacity than the first digestion and all the cells expressed chondrocyte-specific markers tested, with cells from the third digestion showing exceptionally high gene expression levels for collagen type II (Col II), aggrecan, and COMP. Additionally, their ability to produce collagen type II as well as their morphology were not affected by the two additional digestions. Taken together, the results suggested that the use of this isolation protocol resulted in a higher cell yield and the quality of the isolated cells was maintained. Hence, we recommend this isolation protocol to be employed for more efficient cell harvesting especially from limited biopsied cartilage tissue samples. Published version 2015-02-26T02:52:13Z 2019-12-06T22:17:02Z 2015-02-26T02:52:13Z 2019-12-06T22:17:02Z 2015 2015 Journal Article Lau, T. T., Peck, Y., Huang, W., & Wang, D.-A. (2015). Optimization of chondrocyte isolation and phenotype characterization for cartilage tissue engineering. Tissue engineering Part C : methods, 21(2), 105-111. 1937-3384 https://hdl.handle.net/10356/106726 http://hdl.handle.net/10220/25104 10.1089/ten.tec.2014.0159 en Tissue engineering Part C : methods © 2015 Mary Ann Liebert. This paper was published in Tissue Engineering Part C: Methods and is made available as an electronic reprint (preprint) with permission of Mary Ann Liebert. The paper can be found at the following official DOI: [http://dx.doi.org/10.1089/ten.tec.2014.0159]. One print or electronic copy may be made for personal use only. Systematic or multiple reproduction, distribution to multiple locations via electronic or other means, duplication of any material in this paper for a fee or for commercial purposes, or modification of the content of the paper is prohibited and is subject to penalties under law. 7 p. application/pdf |
| spellingShingle | DRNTU::Science::Medicine::Tissue engineering Lau, Ting Ting Peck, Yvonne Huang, Weiliang Wang, Dong-An Optimization of chondrocyte isolation and phenotype characterization for cartilage tissue engineering |
| title | Optimization of chondrocyte isolation and phenotype characterization for cartilage tissue engineering |
| title_full | Optimization of chondrocyte isolation and phenotype characterization for cartilage tissue engineering |
| title_fullStr | Optimization of chondrocyte isolation and phenotype characterization for cartilage tissue engineering |
| title_full_unstemmed | Optimization of chondrocyte isolation and phenotype characterization for cartilage tissue engineering |
| title_short | Optimization of chondrocyte isolation and phenotype characterization for cartilage tissue engineering |
| title_sort | optimization of chondrocyte isolation and phenotype characterization for cartilage tissue engineering |
| topic | DRNTU::Science::Medicine::Tissue engineering |
| url | https://hdl.handle.net/10356/106726 http://hdl.handle.net/10220/25104 |
| work_keys_str_mv | AT lautingting optimizationofchondrocyteisolationandphenotypecharacterizationforcartilagetissueengineering AT peckyvonne optimizationofchondrocyteisolationandphenotypecharacterizationforcartilagetissueengineering AT huangweiliang optimizationofchondrocyteisolationandphenotypecharacterizationforcartilagetissueengineering AT wangdongan optimizationofchondrocyteisolationandphenotypecharacterizationforcartilagetissueengineering |