Therapeutic Potential of Targeting Ferroptosis in Periprosthetic Osteolysis Induced by Ultra-High-Molecular-Weight Polyethylene Wear Debris

Abstract

Background/Objectives: Periprosthetic osteolysis is the primary cause of arthroplasty failure in the majority of patients. Mechanistically, wear debris released from the articulating surfaces of a prosthesis initiates local inflammation and several modes of regulated cell death programs, such as ferroptosis, which represents a promising therapeutic target in various chronic inflammatory diseases. Thus, the current study aimed at exploring the therapeutic potential of targeting ferroptosis in a polyethylene-wear-debris-induced osteolysis model. Methods: Inverted cell culture model was used for stimulating the cells with wear debris in vitro, and calvarial osteolysis model was used for evaluating the therapeutic effects of inhibitors in vivo. Results: The immunostaining of periprosthetic bone tissues demonstrated a number of osteocytes expressing ferroptosis markers. Likewise, the expressions of ferroptosis markers were confirmed in polyethylene-wear-debris-stimulated osteocyte-like cells and primary osteoblasts in a direct stimulation model but not in an indirect stimulation model. Furthermore, polyethylene wear debris was implanted onto calvarial bone and mice were treated with the ferroptosis inhibitors DFO and Fer-1. These treatments alleviated the inflammatory and pathological bone resorption induced by the wear debris implantation. Conclusions: Our data broaden the knowledge of the pathogenesis of periprosthetic osteolysis and highlight ferroptosis as a promising therapeutic target.

Keywords: ferroptosis inhibitors; periprosthetic osteolysis; therapeutics; wear debris.

Figure 1

Detection of GPX4- and NRF2-positive osteocytes in periprosthetic bone tissues. IHC staining of the bone tissues from patients who underwent revision surgery with specific antibodies. Arrows indicate positive reactions. Scale bars are 50 µm.

Figure 2

Expressions of ferroptosis markers in UHMWPE-wear-debris-simulated osteocytes and osteoblasts. Gene expressions of Gpx4, Chac1, and Nfe2l2. (A,B) Relative gene expressions of stimulated MLO-Y4-like osteocytes (A) and osteoblasts (B) in the direct stimulation model with UHMWPE debris. (C,D) Relative gene expressions of stimulated MLO-Y4-like osteocytes (C) and osteoblasts (D) in an indirect stimulation model using the UHMWPE-debris-stimulated macrophages. The results represent the means of 6 samples ± SEM. The significant difference between the two groups was determined by a two-tailed Student’s t-test. (E,F) Protein expression analysis of NRF2 and GPX4 in the stimulated MLO-Y4-like osteocytes. (E) Direct stimulation model. (F) Indirect stimulation model. * = p < 0.05; ** = p < 0.01; *** = p < 0.001; ns = not significant.

Figure 3

Evaluation of the beneficial effects of targeting ferroptosis in the inflammatory osteolysis model. The sham mice did not receive the debris implantation, while all the other groups received it. (A) Quantification of the bone resorption area in the calvarial bone tissues analyzed by micro-CT. The right panels are representative images for the micro-CT. (B) Quantification of the TRAP-stained areas in the calvarial bone sections. The right panels are representative images for the TRAP-stained sections. (C) Quantification of inflammatory infiltrates in the calvarial bone sections. The right panels are representative images for the HE-stained sections. The results represent the means ± SEM of 6 mice. Scale bars are 50 µm. The significant difference between the two groups was determined by one-way ANOVA, followed by Tukey’s multiple-comparison procedure. * = p < 0.05; ** = p < 0.01; *** = p < 0.001; **** = p < 0.0001.