The effects of platelet-rich plasma on bone marrow stromal cell transplants for tendon healing in vitro

Abstract

Purpose: In this study, we investigated the effect of platelet-rich plasma (PRP) and bone marrow-derived stromal cell (BMSC)-seeded interposition in an in vitro canine tendon repair model.

Methods: Bone marrow, peripheral blood, and tendons were harvested from mixed-breed dogs. Bone marrow-derived stromal cells were cultured and passaged from adherent cells of bone marrow suspension. Platelet-rich plasma was purified from peripheral blood using a commercial kit. A total of 192 flexor digitorum profundus tendons were used for the study. Tendons repaired with a simple suture were used as a control group. In treatment groups, a collagen gel patch was interposed at the tendon repair site before suture. There were 3 treatment groups, according to the type of collagen patch: a patch with PRP, a patch with BMSC, and a patch with PRP and BMSC. The repaired tendons were evaluated by biomechanical testing and by histological survey after 2 and 4 weeks in tissue culture. To evaluate viability, cells were labeled with PKH26 red fluorescent cell linker (Sigma, St. Louis, MO) and surveyed under confocal microscopy after culture.

Results: The maximum breaking strength and stiffness of the healing tendons with the BMSC-seeded PRP patch were significantly higher than those of the healing tendons without a patch or with a cell-seeded patch (p < .02). Viable BMSCs were present at both 2 and 4 weeks.

Conclusions: Platelet-rich plasma enhanced the effect of BMSC-seeded collagen gel interposition in this in vitro model. Based on these results, we now plan to investigate this effect in vivo.

Figure 1

Tissue culture of the repaired tendon on custom made wire mesh.

Figure 2

Healing tendon mounted on the micro-tester for mechanical testing.

Figure 3

(A) Appearance of the repaired tendon at mechanical testing. Both sutures were cut prior to testing. (B) Schema of the sutured tendon after suture cut. See the relation between suture, apposition site, and the site of suture cut.

Figure 4

Maximum strength of the repaired tendon. (mean ± SD. * =p< 0.01, ** = p < 0.02)

Figure 5

Stiffness of the repaired tendon. (mean ± SD. * =p< 0.01, ** = p < 0.02)

Figure 6

The labeled BMSC with PKH26 cell linker was observed under confocal microscopy with red fluorescence. (A) BMSC-seeded patch at 2 weeks, (B) BMSC-seeded PRP patch at 2 weeks, (C) BMSC-seeded patch at 4 weeks, (D) BMSC-seeded PRP patch at 4 weeks.

Figure 7

The repaired tendons stained with hematoxylin and eosin at 2 weeks in lower magnification (original photo taken at 90× magnification). (a) No patch group, (b) PRP patch group, (c) BMSC-seeded patch group, and (d) BMSC-seeded PRP patch group. Scale = 0.5 mm.

Figure 8

The repaired tendons stained with hematoxylin and eosin at 4 weeks in lower magnification (original photo taken at 90× magnification). (a) No patch group, (b) PRP patch group, (c) BMSC-seeded patch group, and (d) BMSC-seeded PRP patch group. Scale = 0.5 mm.

Figure 9

The repaired tendons stained with hematoxylin and eosin at 4 weeks in higher magnification (original photo taken at 450× magnification). Border of the collagen fibers within apposition site is partially indistinguishable in every group.(a) No patch group, (b) PRP patch group, (c) BMSC-seeded patch group, and (d) BMSC-seeded PRP patch group. Scale = 0.1 mm.