J Vet Sci.  2008 Dec;9(4):387-393. 10.4142/jvs.2008.9.4.387.

Implantation of canine umbilical cord blood-derived mesenchymal stem cells mixed with beta-tricalcium phosphate enhances osteogenesis in bone defect model dogs

• Jang BJ
• Byeon YE
• Lim JH
• Ryu HH
• Kim WH
• Koyama Y
• Kikuchi M
• Kang KS
• Kweon OK

• KMID: 1104914
• DOI: http://doi.org/10.4142/jvs.2008.9.4.387

Abstract

This study was performed to evaluate the osteogenic effect of allogenic canine umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs) mixed with beta-tricalcium phosphate (beta-TCP) in orthotopic implantation. Seven hundred milligrams of beta-TCP mixed with 1 x 10(6) UCB-MSCs diluted with 0.5 ml of saline (group CM) and mixed with the same volume of saline as control (group C) were implanted into a 1.5 cm diaphyseal defect and wrapped with PLGC membrane in the radius of Beagle dogs. Radiographs of the antebrachium were made after surgery. The implants were harvested 12 weeks after implantation and specimens were stained with H&E, toluidine blue and Villanueva-Goldner stains for histological examination and histomorphometric analysis of new bone formation. Additionally, UCB-MSCs were applied to a dog with non-union fracture. Radiographically, continuity between implant and host bone was evident at only one of six interfaces in group C by 12 weeks, but in three of six interfaces in group CM. Radiolucency was found only near the bone end in group C at 12 weeks after implantation, but in the entire graft in group CM. Histologically, bone formation was observed around beta-TCP in longitudinal sections of implant in both groups. Histomorphometric analysis revealed significantly increased new bone formation in group CM at 12 weeks after implantation (p < 0.05). When applied to the non-union fracture, fracture healing was identified by 6 weeks after injection of UCB-MSCs. The present study indicates that a mixture of UCB-MSCs and beta-TCP is a promising osteogenic material for repairing bone defects.

Keyword

beta-TCP; dog; mesenchymal stem cell; osteogenesis; umbilical cord blood

MeSH Terms

Animals
Biocompatible Materials/metabolism/therapeutic use
Bone Substitutes/*therapeutic use
Calcium Phosphates/*therapeutic use
Dogs
Fetal Blood/*cytology
Fracture Fixation/methods/veterinary
Mesenchymal Stem Cells/*physiology
Osteogenesis/*physiology
Tissue Engineering/methods
Wound Healing/physiology

Figure

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  Fig. 1 Radiographs of antebrachium in clinical application. (A) Preoperative: non-union fracture of the right distal radius. (B) Postoperative: plate & screw fixation with cortical allograft and cancellous bone autograft. (C) 9 weeks postoperative: Bone lysis, irregular margin and disuse atrophy of the bone were observed.

• 

  Fig. 2 Lateral radiographs of antebrachium after implantation and 2, 4, 8, 12 weeks. Radiolucency is found only near bone end in group C (implantation of β-TCP with saline), but entire graft in group CM (implantation of β-TCP with umbilical cord blood derived stem cells) at 12 weeks.

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  Fig. 3 Microphotographs of longitudinal sections of implants in the groups C (A) and CM (B, C) at 12 weeks after implantation of β-TCP. The proximal portion is at the top of the photomicrograph. The cranial aspect of bone (where the fixation plate had been) is at the left of photomicrograph. Bone appears blue or red, and ceramic appears black. (A, B) toluidine blue stain. (C) Villanueva-Goldner stain. ×10. Bone formation is observed around β-TCP throughout the implant. β-TCP remained in both side of the implant, especially under fixation plate and in the group C.

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  Fig. 4 Microphotographs of implants at 12 weeks after implantation in the group CM. New bone was in direct contact with the β-TCP granules. (A) H&E stain: Osteocyte is observed around the β-TCP. (B) Villanueva-Goldner stain (red-immature bone, green-mature bone). (C) Toluidine blue stain.

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  Fig. 5 Lateral radiographs of antebrachium at 0, 4, 8, and 12 weeks after first injection of canine umbilical cord blood derived stem cells.

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