Wear debris from implant interfaces is the major factor leading to periprosthetic osteolysis. Fibroblast-like synoviocytes (FLSs) populate the intimal lining of the synovium and are in direct contact with wear debris. This study aimed to elucidate the effect of Ti particles as wear debris on human FLSs and the mechanism by which they might participate in the bone remodeling process during periprosthetic osteolysis. FLSs were isolated from synovial tissue from patients, and the condition medium (CM) was collected after treating FLSs with sterilized Ti particles. The effect of CM was analyzed for the induction of osteoclastogenesis or any effect on osteogenesis and signaling pathways. The results demonstrated that Ti particles could induce activation of the NFκB signaling pathway and induction of COX-2 and inflammatory cytokines in FLSs. The amount of RANL in the conditioned medium collected from Ti particle-stimulated FLSs (Ti CM) showed the ability to stimulate osteoclast formation. The Ti CM also suppressed the osteogenic initial and terminal differentiation markers for osteoprogenitors, such as alkaline phosphate activity, matrix mineralization, collagen synthesis, and expression levels of Osterix, Runx2, collagen 1α, and bone sialoprotein. Inhibition of the WNT and BMP signaling pathways was observed in osteoprogenitors after the treatment with the Ti CM. In the presence of the Ti CM, exogenous stimulation by WNT and BMP signaling pathways failed to stimulate osteogenic activity in osteoprogenitors. Induced expression of sclerostin (SOST: an antagonist of WNT and BMP signaling) in Ti particletreated FLSs and secretion of SOST in the Ti CM were detected. Neutralization of SOST in the Ti CM partially restored the suppressed WNT and BMP signaling activity as well as the osteogenic activity in osteoprogenitors. Our results reveal that wear debris-stimulated FLSs might affect bone loss by not only stimulating osteoclastogenesis but also suppressing the bone-forming ability of osteoprogenitors. In the clinical setting, targeting FLSs for the secretion of antagonists like SOST might be a novel therapeutic approach for preventing bone loss during inflammatory osteolysis.

The development of a representative human, in vitro OA model could deepen understanding of disease mechanisms. Our research aimed to reprogram healthy and OA-derived synoviocytes to induced pluripotent stem cells (iPSCs), thereby generating a novel OA in vitro model. Comparison between the two models shall enable research into underlying processes with potential for clinical translation.

A meta-analysis of OA synovial biomarkers was conducted, identifying up to thirteen relevant pathophysiology-related factors, including, amongst others, IL-13, IL-10, IL-6, PIICP, and HA, with PIICP demonstrating the largest effect (SMD 6.11 [3.50, 8.72], p <0.00001). With these findings in mind, human fibroblast-like synoviocytes (HFLS) from healthy and OA patients were transduced using Sendai viral reprogramming. Two clones for each of the resulting iPSC lines were expanded and preliminarily analysed in triplicate by ICC and RT-qPCR for pluripotency characteristics.

Healthy HFLS-derived and OA-HFLS-derived iPSC (UoS-B and UoS-C lines, respectively) were generated, indicating successful reprogramming. Morphological observations demonstrated typical iPSC appearance, and ICC confirmed presence of pluripotency markers Tra-1-60, Oct3/4 and Nanog. Expression of Oct3/4, Nanog and Sox2 were confirmed by RT-qPCR with OA-iPSC lines expressing higher levels of all markers compared to non-OA iPSC. In particular, expression of Oct3/4 and Sox2 was 3.5 fold and 4.6 fold higher (p <0.001) in OA-iPSCs (UoS-C) vs. non-OA iPSCs (UoS-B), respectively. Sendai virus clearance was confirmed by passage 4.

The successfully obtained OA and non-OA iPSCs can be differentiated towards mesenchymal lineages, including chondrocyte and bone progenitor cells, enabling phenotypic comparison and biomarker analysis as identified in meta-analysis. Cell bank dissemination of these cell lines could deepen further in vitro OA research, with potential impact for clinical translation via the identification of novel cellular and molecular targets.