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3D porous calcium-alginate scaffolds cell culture system improved human osteoblast cell clusters for cell therapy.

Chen CY, Ke CJ, Yen KC, Hsieh HC, Sun JS, Lin FH - Theranostics (2015)

Bottom Line: The Ca-Alginate scaffold facilitated the growth and differentiation of human bone cell clusters, and the functionally-closed process bioreactor system supplied the soluble nutrients and osteogenic signals required to maintain the cell viability.This system preserved the proliferative ability of cells and cell viability and up-regulated bone-related gene expression and biological apatite crystals formation.The described strategy could be used in therapeutic application and opens new avenues for surgical interventions to correct skeletal defects.

View Article: PubMed Central - PubMed

Affiliation: 1. Institute of Biomedical Engineering, College of Medicine and College of Engineering, National Taiwan University, Taipei, Taiwan (R.O.C.).

ABSTRACT
Age-related orthopedic disorders and bone defects have become a critical public health issue, and cell-based therapy is potentially a novel solution for issues surrounding bone tissue engineering and regenerative medicine. Long-term cultures of primary bone cells exhibit phenotypic and functional degeneration; therefore, culturing cells or tissues suitable for clinical use remain a challenge. A platform consisting of human osteoblasts (hOBs), calcium-alginate (Ca-Alginate) scaffolds, and a self-made bioreactor system was established for autologous transplantation of human osteoblast cell clusters. The Ca-Alginate scaffold facilitated the growth and differentiation of human bone cell clusters, and the functionally-closed process bioreactor system supplied the soluble nutrients and osteogenic signals required to maintain the cell viability. This system preserved the proliferative ability of cells and cell viability and up-regulated bone-related gene expression and biological apatite crystals formation. The bone-like tissue generated could be extracted by removal of calcium ions via ethylenediaminetetraacetic acid (EDTA) chelation, and exhibited a size suitable for injection. The described strategy could be used in therapeutic application and opens new avenues for surgical interventions to correct skeletal defects.

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Related in: MedlinePlus

Live/dead staining showed the difference between hOBs @Ca-Alginate scaffolds w/ and w/o perfusion. (A1, B1, C1, and D1) in green represented the live cells of bone-like tissues with calcein AM dye; (A2, B2, C2, and D2) in red (PI) indicated dead cells; (A3, B3, C3, and D3) were the merge images; (A4, B4, C4, and D4) displayed the percentage of live and dead cells.
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Figure 4: Live/dead staining showed the difference between hOBs @Ca-Alginate scaffolds w/ and w/o perfusion. (A1, B1, C1, and D1) in green represented the live cells of bone-like tissues with calcein AM dye; (A2, B2, C2, and D2) in red (PI) indicated dead cells; (A3, B3, C3, and D3) were the merge images; (A4, B4, C4, and D4) displayed the percentage of live and dead cells.

Mentions: Cell viability of bone-like tissues within Ca-Alginate scaffolds was determined by staining with two-color fluorescent dyes (Calcein AM/PI). Static conditions might cause death of human osteoblasts in 3D environment (Fig. 4A and 4B), whereas hOBs survived in Ca-Alginate scaffolds with dynamic fluids (Fig. 4C and 4D). In the static group, 21.2% of total cells were dead inside cell clusters at day 7 and 45.6% of total cells was stained with red fluorescence at day 14. Cell death was localized at the center of cell clusters, and all the 3D corresponding animated movies were provided in Additional File 2 (Video S2, S3, S4, and S5). Conversely, most cells in the dynamic group were stained in green and less than 1% of total cells died both at day 7 and day 14. Consequently, Ca-Alginate scaffolds could create an environment permissive for hOBs ingrowth and cell clusters formation and the perfusion system played a pivotal role in maintaining cell viability.


3D porous calcium-alginate scaffolds cell culture system improved human osteoblast cell clusters for cell therapy.

Chen CY, Ke CJ, Yen KC, Hsieh HC, Sun JS, Lin FH - Theranostics (2015)

Live/dead staining showed the difference between hOBs @Ca-Alginate scaffolds w/ and w/o perfusion. (A1, B1, C1, and D1) in green represented the live cells of bone-like tissues with calcein AM dye; (A2, B2, C2, and D2) in red (PI) indicated dead cells; (A3, B3, C3, and D3) were the merge images; (A4, B4, C4, and D4) displayed the percentage of live and dead cells.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4377732&req=5

Figure 4: Live/dead staining showed the difference between hOBs @Ca-Alginate scaffolds w/ and w/o perfusion. (A1, B1, C1, and D1) in green represented the live cells of bone-like tissues with calcein AM dye; (A2, B2, C2, and D2) in red (PI) indicated dead cells; (A3, B3, C3, and D3) were the merge images; (A4, B4, C4, and D4) displayed the percentage of live and dead cells.
Mentions: Cell viability of bone-like tissues within Ca-Alginate scaffolds was determined by staining with two-color fluorescent dyes (Calcein AM/PI). Static conditions might cause death of human osteoblasts in 3D environment (Fig. 4A and 4B), whereas hOBs survived in Ca-Alginate scaffolds with dynamic fluids (Fig. 4C and 4D). In the static group, 21.2% of total cells were dead inside cell clusters at day 7 and 45.6% of total cells was stained with red fluorescence at day 14. Cell death was localized at the center of cell clusters, and all the 3D corresponding animated movies were provided in Additional File 2 (Video S2, S3, S4, and S5). Conversely, most cells in the dynamic group were stained in green and less than 1% of total cells died both at day 7 and day 14. Consequently, Ca-Alginate scaffolds could create an environment permissive for hOBs ingrowth and cell clusters formation and the perfusion system played a pivotal role in maintaining cell viability.

Bottom Line: The Ca-Alginate scaffold facilitated the growth and differentiation of human bone cell clusters, and the functionally-closed process bioreactor system supplied the soluble nutrients and osteogenic signals required to maintain the cell viability.This system preserved the proliferative ability of cells and cell viability and up-regulated bone-related gene expression and biological apatite crystals formation.The described strategy could be used in therapeutic application and opens new avenues for surgical interventions to correct skeletal defects.

View Article: PubMed Central - PubMed

Affiliation: 1. Institute of Biomedical Engineering, College of Medicine and College of Engineering, National Taiwan University, Taipei, Taiwan (R.O.C.).

ABSTRACT
Age-related orthopedic disorders and bone defects have become a critical public health issue, and cell-based therapy is potentially a novel solution for issues surrounding bone tissue engineering and regenerative medicine. Long-term cultures of primary bone cells exhibit phenotypic and functional degeneration; therefore, culturing cells or tissues suitable for clinical use remain a challenge. A platform consisting of human osteoblasts (hOBs), calcium-alginate (Ca-Alginate) scaffolds, and a self-made bioreactor system was established for autologous transplantation of human osteoblast cell clusters. The Ca-Alginate scaffold facilitated the growth and differentiation of human bone cell clusters, and the functionally-closed process bioreactor system supplied the soluble nutrients and osteogenic signals required to maintain the cell viability. This system preserved the proliferative ability of cells and cell viability and up-regulated bone-related gene expression and biological apatite crystals formation. The bone-like tissue generated could be extracted by removal of calcium ions via ethylenediaminetetraacetic acid (EDTA) chelation, and exhibited a size suitable for injection. The described strategy could be used in therapeutic application and opens new avenues for surgical interventions to correct skeletal defects.

Show MeSH
Related in: MedlinePlus