Platelet-rich plasma protects rat chondrocytes from interleukin-1β-induced apoptosis

Abstract

Interleukin (IL)-1β-induced chondrocyte apoptosis is associated with the pathogenesis of arthritis. Platelet‑rich plasma (PRP), which is derived from the patient's own blood and contains numerous growth factors, has the potential for arthritis treatment. Therefore, the present study aimed to determine the effects of PRP on chondrocyte apoptosis, under IL‑1β‑induced pathological conditions. Chondrocytes isolated from the knee joint of Sprague Dawley rats were used in the present study. Cell viability was determined using the Cell Counting kit‑8 assay, cell apoptosis was evaluated by flow cytometry, and the expression of apoptosis‑, anabolism‑ and catabolism-associated genes were detected by quantitative polymerase chain reaction; protein expression was detected by western blot analysis. The results demonstrated that 10% PRP in the culture medium increased chondrocyte proliferation, whereas IL‑1β induced cell apoptosis. Treatment with PRP significantly attenuated cell apoptosis in IL‑1β‑treated chondrocytes, and altered apoptosis‑associated expression at the gene and protein level. Furthermore, treatment with PRP significantly reduced matrix metalloproteinase production and promoted anabolism of cartilage extracellular matrix under IL‑1β treatment. The present study demonstrated the protective effects of PRP on chondrocyte apoptosis and extracellular matrix anabolism, and provided scientific evidence to support the potential use of PRP as a promising therapeutic strategy for the treatment of arthritis.

Figure 1.

Characterization of rat chondrocytes. Phase contrast image shows the morphology of articular chondrocytes isolated from the knee joints of rat neonates. Following in vitro culture for two passages, cells were positive for collagen type II (COL II) as indicated by immunohistochemistry. Bar=50 µm.

Figure 2.

Interleukin (IL)-1β induces apoptosis of rat chondrocytes. (A) Annexin V staining was performed 24 h following treatment of chondrocytes with IL-1β. (B) Quantification of Annexin V-positive cells. The sum of gate Q2 and Q3 cells was used to determine the number of Annexin V-positive cells (N=3). FITC, fluorescein isothiocyanate; SSC, side scatter.

Figure 3.

Platelet-rich plasma (PRP) treatment reduced apoptosis, and the expression of apoptotic genes in rat chondrocytes. (A) Annexin V staining was conducted 24 h after incubation with interleukin (IL)-1β or IL-1β + PRP. (B) Quantification of Annexin V-positive cells. The sum of gate Q2 and Q3 cells was used to determine the number of Annexin V-positive cells (N=3). (C) Quantitative polymerase chain reaction (qPCR) was performed 24 h after treatment of chondrocytes (N=3). (D) Western blotting exhibited similar gene expression results as qPCR. (E) Semi-quantification of blots confirmed the expression of apoptotic proteins (N=3). *P<0.05, vs. the IL-1β group. Bcl-2, B-cell lymphoma 2; Bax, Bcl-2-associated X protein; PARP, poly (ADP-ribose) polymerase; GAPDH, glyceraldehyde 3-phosphate dehydrogenase.

Figure 4.

Platelet-rich plasma (PRP) inhibits the expression of catabolic genes induced by interleukin (IL)-1β. (A) Quantitative polymerase chain reaction was performed 24 h following treatment of chondrocytes (N=3). (B) Western blotting detected downregulation of matrix metalloproteinases (MMPs), and upregulation of tissue inhibitor of metalloproteinases (TIMP), SRY-box 9 (SOX9) and collagen type II (COL2) in the IL-1β + PRP group compared with the IL-1β group. (C) Semi-quantification of blots confirmed the expression of anabolic proteins was increased by PRP (N=3). *P<0.05, vs. the IL-1β group. GAPDH, glyceraldehyde 3-phosphate dehydrogenase.