Abstract
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.