Comparative evaluation of the effects of platelet-rich plasma formulations on extracellular matrix formation and the NF-κB signaling pathway in human articular chondrocytes

Concentrated leukocytes in leukocyte and platelet-rich plasma (L-PRP) may deliver increased levels of pro-inflammatory cytokines to activate the nuclear factor (NF)-κB signaling pathway, to counter or overwhelm the beneficial effects of growth factors on cartilage regeneration. However, to date, no relevant studies have substantiated this. In the present study, L-PRP and pure platelet-rich plasma (P-PRP) were prepared, and leukocytes, platelets, pro-inflammatory cytokines and growth factor concentrations were quantified; they were then used to treat human articular chondrocytes (HACs). Pyrrolidine dithiocarbamate (PDTC; 50 µM) was used to inhibit the activation of NF-κB. The nuclear translocation of NF-κB p65 and the protein expression of cartilaginous markers (collagen II, aggrecan and sex-determining region Y-box 9) were determined using western blot analysis. The mRNA expression of NF-κB-dependent inflammatory mediators, including inducible nitric oxide synthase and cyclooxygenase-2, and cartilaginous markers were determined using reverse transcription-quantitative polymerase chain reaction analysis. The production of prostaglandin E2, nitric oxide and glycosaminoglycan (GAG) were quantified using enzyme-linked immunosorbent assays, the Griess reaction and a 1,9-dimethylmethylene blue assay, respectively. The results demonstrated that L-PRP induced the nuclear translocation of NF-κB p65, upregulated the mRNA expression of NF-κB-dependent inflammatory mediators and upregulated the production of their products, whereas P-PRP, which had similar growth factor concentrations but significantly lower pro-inflammatory cytokine concentrations than L-PRP, did not. P-PRP promoted the mRNA and protein expression levels of cartilaginous markers and the production of GAG more effectively, compared with L-PRP. Furthermore, inhibition of the activation of NF-κB by PDTC enhanced the effects of L-PRP on extracellular matrix formation in the HACs to a level similar to that of P-PRP. These findings suggested that leukocytes in L-PRP activated the NF-κB signaling pathway via the delivery of interleukin-1β and tumor necrosis factor-α to counter the beneficial effects of growth factors on extracellular matrix formation in HACs. Therefore, P-PRP may be more suitable for the treatment of osteoarthritis.