Dental pulp stem cells immobilized in alginate microspheres for applications in bone tissue engineering

Authors

  • M. M. Kanafi,

    1. Manipal Institute of Regenerative Medicine, Manipal University, Bangalore, India
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    • These authors contributed equally to this work
  • A. Ramesh,

    1. Manipal Institute of Regenerative Medicine, Manipal University, Bangalore, India
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    • These authors contributed equally to this work
  • P. K. Gupta,

    1. Manipal Institute of Regenerative Medicine, Manipal University, Bangalore, India
    2. Stempeutics Research Pvt. Ltd., Bangalore, India
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  • R. R. Bhonde

    Corresponding author
    1. Manipal Institute of Regenerative Medicine, Manipal University, Bangalore, India
    • Correspondence: Ramesh Bhonde, Manipal Institute of Regenerative Medicine, GKVK Post, Bellary Road, Allalasandra, Yelahanka, Bangalore – 560065 India (e-mail: rr.bhonde@manipal.edu).

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Abstract

Aim

To immobilize dental pulp stem cells (DPSC) in alginate microspheres and to determine cell viability, proliferation, stem cell characteristics and osteogenic potential of the immobilized DPSCs.

Methodology

Human DPSCs isolated from the dental pulp were immobilized in 1% w/v alginate microspheres. Viability and proliferation of immobilized DPSCs were determined by trypan blue and MTT assay, respectively. Stem cell characteristics of DPSCs post immobilization were verified by labelling the cells with CD73 and CD90. Osteogenic potential of immobilized DPSCs was assessed by the presence of osteocalcin. Alizarin red staining and O-cresolphthalein complexone method confirmed and quantified calcium deposition. A final reverse transcriptase PCR evaluated the expression of osteogenic markers – ALP, Runx-2 and OCN.

Results

More than 80% of immobilized DPSCs were viable throughout the 3-week study. Proliferation appeared controlled and consistent unlike DPSCs in the control group. Presence of CD73 and CD90 markers confirmed the stem cell nature of immobilized DPSCs. The presence of osteocalcin, an osteoblastic marker, was confirmed in the microspheres on day 21. Mineralization assays showed high calcium deposition indicating elevated osteogenic potential of immobilized DPSCs. Osteogenic genes- ALP, Runx-2 and OCN were also upregulated in immobilized DPSCs. Surprisingly, immobilized DPSCs in the control group cultured in conventional stem cell media showed upregulation of osteogenic genes and expressed osteocalcin.

Conclusion

Dental pulp stem cells immobilized in alginate hydrogels exhibit enhanced osteogenic potential while maintaining high cell viability both of which are fundamental for bone tissue regeneration.

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