Cross-linking of lipopolysaccharide (LPS) to CD14 on THP-1 cells mediated by LPS-binding protein.

Abstract

Recent work has established that bacterial endotoxin (LPS) binds to the plasma protein LPS-binding protein (LBP) forming high affinity complexes (LPS-LBP), that LBP is an opsonin for LPS-bearing particles, and that LPS-LBP complexes are potent agonists for monocytic cells (MO). mAb to the MO plasma membrane protein, CD14, inhibit LBP-dependent binding of LPS to MO, and LPS-LBP-dependent stimulation of cytokine release from MO. These data suggest that CD14 functions as a membrane receptor for LPS but do not demonstrate a direct association of LPS with CD14. Calcitriol was used to induce a high level of CD14 expression in the human monocyte-like cell line THP-1, resulting in enhanced responses of these cells to LPS-LBP complexes manifested by enhanced binding of LPS and a decrease in the amount of LPS needed to induce IL-8 release. An Re595 LPS derivative containing a radioiodinated, photoreactive, phenyl azide (125I-ASD-LPS) was used in cross-linking experiments to identify membrane proteins in calcitriol-treated THP-1 cells that interact with LPS. 125I-ASD-LPS was added to calcitriol-induced THP-1 cells in the presence or absence of LBP, the mixture photolyzed, and the resultant radioiodinated proteins analyzed by SDS-PAGE and autoradiography. We observed strong cross-linking of 125I-ASD-LPS to a 55-kDa membrane protein when LBP was present, but failed to observe radiolabeling of any other proteins with apparent molecular masses distinct from CD14. The cross-linked product was identified as CD14 by immunoprecipitation with anti-human CD14 mAb. Studies with human CD14 expressing transfectants of the murine B cell line 70Z/3 also revealed LBP-dependent cross-linking of 125I-ASD-LPS to CD14. These data document binding of LPS to a specific membrane protein that serves as an LPS receptor.

Research development on gut microbiota and vulnerable atherosclerotic plaque

“…In a healthy gut, a screen of the intestinal epithelium can prevent most LPS from activating the intestinal immune system, whereas unhealthy gut flora can cause increased intestinal permeability, the so-called "leaky gut," in which more LPS enters the blood circulation and triggers excessive inflammation of the vascular endothelium [ 95 ]. A large number of studies have shown that LPS can change the stability of plaques and make them more likely to rupture [ 19 , [96] , [97] , [98] , [99] , [100] , [101] ].…”

“…2 ). Oxygen radicals can lead to increased oxidative LDL formation downstream, and the atherosclerotic hypothesis currently recognized by major scholars is closely related to oxidative LDL, proving that LPS is a molecule that promotes atherosclerosis and may cause plaque rupture [ [97] , [98] , [99] , [100] ].

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