title: Dental pulp stem cells immobilized in alginate microspheres for applications in bone tissue engineering.
abstract: OBJECTIVE
  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.
  
  
  METHODS
  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.
  
  
  CONCLUSIONS
  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.
authors.affiliation: Manipal Institute of Regenerative Medicine, Manipal University, Bangalore, India.
authors.forename: M M
authors.initials: MM
authors.lastname: Kanafi
authors.forename: A
authors.initials: A
authors.lastname: Ramesh
authors.forename: P K
authors.initials: PK
authors.lastname: Gupta
authors.forename: R R
authors.initials: RR
authors.lastname: Bhonde
mesh_terms: D000464:Alginates / Q000737:chemistry*; D001846:Bone Development; D049109:Cell Proliferation; D002478:Cells, Cultured; D003782:Dental Pulp / Q000166:cytology*; D020723:Glucuronic Acid / Q000737:chemistry; D006603:Hexuronic Acids / Q000737:chemistry; D006801:Humans; D008863:Microspheres; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013234:Stem Cells / Q000166:cytology*; D023822:Tissue Engineering
chemical_list: D000464:Alginates; D006603:Hexuronic Acids; D020723:Glucuronic Acid
keywords: alginate hydrogels; bone tissue engineering; dental pulp stem cells; immobilization; microspheres