Stimulation of THP-1 Macrophages with LPS Increased the Production of Osteopontin-Encapsulating Exosome

Abstract

Osteopontin (OPN) mediates bone remodeling and tissue debridement. The OPN protein is cleaved, but it is unclear how full-length (FL)-OPN or its cleaved form perform their biological activities in target cells. We, therefore, performed the molecular characterization of OPN in exosomes (Exo). The Exo were isolated from lipopolysaccharide (LPS)-stimulated phorbol 12-myristate 13-acetate (PMA)-differentiated THP-1 macrophages. The Exo were also isolated from PMA-differentiated THP-1 macrophages. The Exo were identified using the qNano multiple analyzer (diameter 59–315 nm) and western blotting with a CD9 antibody. LPS-stimulated cells produced more particles than non-stimulated cells. The presence of the FL or the cleaved form of OPN was confirmed using western blot analysis. A mixture of FL and cleaved OPN was also measured using an ELISA system (Ud-OPN) and their presence in the Exo was confirmed. Ud/FL ratios became low after LPS stimulation, indicating the enhanced encapsulation of FL-OPN in the Exo by LPS. These findings suggest that LPS stimulation of human macrophages facilitates the synthesis of FL-OPN, which is cleaved in cells or the Exo after release. These findings indicate that Exo is a suitable vehicle to transfer OPN to the target cells.

High Levels of the Cleaved Form of Galectin-9 and Osteopontin in the Plasma Are Associated with Inflammatory Markers That Reflect the Severity of COVID-19 Pneumonia

“…Our results suggest that the moderate association of FL-OPN with CRP and sIL-2R, observed in the CV group was decreased in the CP group, which indicates that it might be rapidly cleaved in inflammatory environments. OPN was released in the form of exosome from a lipopolysaccharide-stimulated macrophage cell line and enhanced encapsulation of FL-OPN was reported, suggesting that the exosomes may be a suitable vehicle for transferring OPN to target cells [ 66 ]. We should determine whether OPN needs to be present in plasma in the form of exosomes to exert its biological functions.…”

Cell culture‐derived extracellular vesicles: Considerations for reporting cell culturing parameters

“…Other factors used to promote proliferation, differentiation, silencing, or priming of cells, and/or to otherwise manipulate cells and their functions, may influence the release of EVs (Bai et al., 2020; Emam et al., 2018; Martens‐Uzunova et al., 2021; Pecan et al., 2020; Savina et al., 2003; Segura et al., 2005; Soekmadji et al., 2017; Taylor et al., 2017; Wang et al., 2020) or their therapeutic potential (Akbar et al., 2017; Cossetti et al., 2014; De Jong et al., 2012; Hosseinkhani et al., 2017, 2020; Lopatina et al., 2014; Patel et al., 2019; Peltzer et al., 2020; Sutter et al., 2021; Varkouhi et al., 2019; Yang et al., 2020; Zhang et al., 2018, 2020).…”

Functional Properties of Cancer Epithelium and Stroma-Derived Exosomes in Head and Neck Squamous Cell Carcinoma

“…Interestingly, Xiao et al demonstrated that THP-1 treated with exosomes from OSCC cells exhibited an M1-like but not an M2-like phenotype in promoting migration [ 32 ], supporting the notion that TAMs are highly heterogeneous stromal cells whose crosstalk with cancer cells is mediated by multiple signals, a point that should be considered in antitumor therapy in OSCC based on the conversion of TAMs from the M2 to M1 phenotype. Of note, the exosomes secreted by THP-1 derived macrophages depends not only on PMA simulation but also the surrounding status including the lipopolysaccharide (LPS) [ 63 ] and IL-4 [ 64 , 65 ]. This suggests that the exosomes from THP-1 derived macrophages are highly sensitive to the exogenous stimulus, in part because the exosomes in different cancers exhibit heterogeneity and specificity.…”