Recent evidence indicates the potential of gamma-irradiated (γi) Staphylococcus aureus to be used as an osteo-immunomodulator for bone regeneration. This study aims at characterizing the inflammatory milieu caused by the stimulation of γi S. aureus in immune cells and investigates its effects on MSC osteogenic differentiation. Furthermore, we aimed to recreate the immune-modulatory response exhibited by γi S. aureus by using a mixture of various synthetic pathogen recognition receptor (PRR) ligands consisting of TLR2, TLR8, TLR9, and NOD2 agonists. Human peripheral blood mononuclear cells (hPBMCs), isolated from healthy human donors, were exposed to γi S. aureus or seven different ligand mixtures. After 24 h, the conditioned medium (CM) from the hPBMCs was collected and its effects on hMSC osteogenic differentiation were investigated by assessing alkaline phosphatase (ALP) activity and matrix mineralization. The hPBMCs and their CM were also analyzed by bulk RNA sequencing and for cytokine secretion. CM from the γi S. aureus and the mixture consisting of Pam3CSK4, C-class CpG oligodeoxynucleotide (CpG ODN C), and murabutide targeting TLR2, TLR9, and NOD2 showed a fivefold increase in ALP and matrix mineralization in a donor-dependent manner. These effects were due to the upregulation of inflammatory signaling pathways, which led to an increase in cytokines and chemokines TNF, interleukin (IL)-6, IFN-γ, IL-1α, CXCL10, CCL18, CCL17, CXCL1, and CCL5. Upregulation of genes like BMP2R, BMP6R, BGLAP, and others contributed to the upregulation of osteogenic pathways in the hPBMCs stimulated with γi S. aureus and the aforementioned mix. Thus, formulations with mixtures of PRR ligands may serve as immune-modulatory osteogenesis-enhancing agents.

Background: Mesenchymal stromal/stem cells (MSCs) and MSC-derived extracellular vesicles (MSC-EVs) hold promise as a disease modifying treatment in osteoarthritis (OA). Obesity, and its associated inflammation, contribute to OA development and metabolic OA represents a specific and significant group of the OA patient population. Given their immunomodulatory properties, MSC and MSC-EVs are especially interesting for this group of patients as a therapeutic option. Here, we were the first to compare the therapeutic efficacy of MSCs and MSC-EVs in a mild OA model taking these metabolic aspects into consideration. Methods: Male Wistar-Han rats (Crl:WI(Han) (n = 36) were fed a high fat diet for 24 weeks, with unilateral induction of OA by groove surgery after 12 weeks. Eight days after surgery rats were randomized in three treatment groups receiving MSCs, MSC-EVs or vehicle injection. Pain-associated behavior, joint degeneration, and local and systemic inflammation were measured. Results: We demonstrated that despite not having a significant therapeutic effect, MSC-EV treatment results in lower cartilage degeneration, less pain behaviour, osteophytosis and joint inflammation, than MSC treatment. Suggesting that MSC-EVs could be a more promising therapeutic strategy than MSCs in this mild metabolic OA model. Conclusion: In summary, we find that MSC treatment has negative effects on the joint in metabolic mild OA. This is an essential finding for the significant group of patients with metabolic OA phenotype, and might help to understand why clinical translation of MSC treatment shows varying therapeutic efficacy thus far. Our results also suggest that MSC-EV-based treatment might be a promising option for these patients, however MSC-EV therapeutic efficacy will need improvement.

Introduction: Obesogenic diets aggravate osteoarthritis (OA) by inducing low-grade systemic inflammation, and diet composition may affect OA severity. Here, we investigated the effect of diet on joint damage and inflammation in an OA rat model. Methods: Wistar-Han rats (n = 24) were fed a chow, a high-fat (HF) diet, or a high-fat/high-sucrose (HFS) for 24 weeks. OA was induced unilaterally 12 weeks after the diet onset by groove surgery, and compared to sham surgery or no surgical intervention (contralateral limb). Knee OA severity was determined by OARSI histopathology scoring system. At several timepoints monocyte populations were measured using flow cytometry, and joint macrophage response was determined via CD68 immunohistochemistry staining. Results: Groove surgery combined with HF or HFS diet resulted in higher OARSI scores, and both HF and HFS diet showed increased circulating intermediate monocytes compared to chow fed rats. Additionally, in the HFS group, minimal damage by sham surgery resulted in an increased OARSI score. HFS diet resulted in the largest metabolic dysregulation, synovial inflammation and increased CD68 staining in tibia epiphysis bone marrow. Conclusion: Obesogenic diets resulted in aggravated OA development, even with very minimal joint damage when combined with the sucrose/fat-rich diet. We hypothesize that diet-induced low-grade inflammation primes monocytes and macrophages in the blood, bone marrow, and synovium, resulting in joint damage when triggered by groove OA inducing surgery. When the metabolic dysregulation is larger, as observed here for the HFS diet, the surgical trigger required to induce joint damage may be smaller, or even redundant.

Osteoarthritis (OA) is a common and debilitating joint disorder that leads to progressive joint breakdown and loss of articular cartilage. Accompanied by a state of low-grade inflammation, its etiology extends beyond that of a wear-and-tear disease, and the immune system might have a role in its initiation and progression. Obesity, which is directly associated with an increased incidence of OA, alters adipokine release, increases pro-inflammatory macrophage activity, and affects joint immune regulation. Studying inflammatory macrophage expression and strategies to inhibit inflammatory macrophage phenotype polarization might provide insights into disease pathogenesis and therapeutic applications. In pre-clinical studies, the detection of OA in its initial stages was shown to be possible using imaging techniques such as SPECT-CT, and advances are made to detect OA through blood-based biomarker analysis. In this review, obesity-induced osteoarthritis and its mechanisms in inducing joint degeneration are summarized, along with an analysis of the current developments in patient imaging and biomarker use for diagnostic and therapeutic strategies.

OBJECTIVE: Folate receptor beta (FR-β) has been used as a clinical marker and target in multiple inflammatory diseases, including osteoarthritis (OA) and rheumatoid arthritis (RA). However, the conditions under which FR-β+ macrophages arise remain unclear and could be affected by corticosteroids. Therefore, we studied FR-β expression in vitro in macrophage subtypes and determined their response to triamcinolone acetonide (TA), a clinically often-used corticosteroid. DESIGN: Human monocyte-derived macrophages were differentiated to the known M0, M1, or M2 macrophage phenotypes. The phenotype and FR-β expression and plasticity of the macrophage subtypes were determined using flow cytometry, reverse transcription-quantitative polymerase chain reaction (RT-qPCR), and enzyme-linked immunosorbent assay (ELISA). RESULTS: FR-β expression was low in granulocyte-macrophage colony-stimulating factor (GM-CSF)-generated (M1-like) macrophages and high in macrophage colony-stimulating factor (M-CSF)-generated (M0 and M2-like) macrophages. FR-β expression remained high once the M0 or M2 macrophages were stimulated with pro-inflammatory stimuli (interferon-γ plus lipopolysaccharide) to induce M1-like macrophages. On the contrary, anti-inflammatory TA treatment skewed GM-CSF macrophage differentiation toward an M2 and FR-β+ phenotype. CONCLUSIONS: As corticosteroids skewed monocytes toward an FR-β-expressing, anti-inflammatory phenotype, even in an M1 priming GM-CSF environment, FR-β has potential as a biomarker to monitor success of treatment with corticosteroids. Without corticosteroid treatment, M-CSF alone induces high FR-β expression which remains high under pro-inflammatory conditions. This explains why pro-inflammatory FR-β+ macrophages (exposed to M-CSF) are observed in arthritis patients and correlate with disease severity.