INTRODUCTION: Over years, the annulus fibrosus(AF) weakens and defects created in the AF causing disc herniation. An intervention increasing extracellular matrix(ECM) production and cell proliferation is needed to rejuvenate the AF tissue. Platelet rich plasma(PRP) is a useful delivery system for growth factors, enhancing fibroblast proliferation and ECM synthesis.
METHODS: Human blood aspirates were centrifuged producing PRP at 1/10 of the initial blood volume. Platelet‐released growth factors(PRGF) were produced by activation of the platelets in freeze‐thaw cycles. Bovine AF cells were harvested from fresh tail discs and cultured in Dulbecco's Modified Eagle's Medium(DMEM) with the following: 25% PRP, 50% PRP, 10% foetal calf serum, 50% PRGF, platelet poor plasma, and only DMEM. All samples were supplemented with ascorbic acid and ITS to encourage proliferation and collagen production. After 48 hours, DNA content, glycosaminoglycan(GAG) levels and mRNA expression of collagen I(Col‐I), Col‐II, aggrecan, matrix metalloproteinase 3(MMP3) and MMP13 were assessed. Finally, a cut was made in the AF of whole bovine discs and PRP was placed in the cut; after 7 days the discs underwent histological analysis.
RESULTS: DMEM containing 25% and 50% PRP gelled, creating a 3D structure in which the AF cells harboured. A significant increase in the DNA content was seen in the PRP groups. GAG production was significantly enhanced in the PRP groups. GAG/DNA levels were highest in the PRP and PRGF groups. Gene expression of Col‐I, aggrecan were up‐regulated and Col‐II was down‐regulated in the PRP groups. No change was seen in MMP3, MMP13 mRNA. In the whole disk, PRP caused an increase in GAG production and induced cell activation.
DISCUSSION: In vitro AF cells proliferate and produce increased ECM in the presence of PRP. Ex‐vivo PRP promoted GAG production in damaged intervertebral discs. Taken together, these results indicate that PRP might support AF regeneration in vivo.