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Published on: September 9, 2014
Control of lipopolysaccharide (LPS) binding and LPS-induced tumor necrosis factor secretion in human peripheral blood
D Heumann1, P Gallay, C Barras
1Department of Internal Medicine, Centre Hospitalier Universitaire Vaudois, Lausanne, Switzerland.
Insights
LPS-binding protein (LBP) enhances lipopolysaccharide (LPS) binding to monocytes, primarily through CD14. This interaction significantly increases monocyte sensitivity to LPS, triggering tumor necrosis factor (TNF) secretion.
Area of Science:
- Immunology
- Cell Biology
Background:
- Lipopolysaccharide (LPS) is a potent immune activator found in Gram-negative bacteria.
- LPS-binding protein (LBP) is a key serum factor involved in LPS recognition by immune cells.
Purpose of the Study:
- To investigate the role of LPS-binding protein (LBP) in the binding of LPS to human monocytes.
- To elucidate the receptor-dependent mechanisms involved in LPS-monocyte interactions and subsequent TNF secretion.
Main Methods:
- Flow cytometry was used to quantify the binding of fluorescein-labeled LPS to human monocytes.
- Monocytes were treated with anti-CD14 monoclonal antibody (mAb) or subjected to enzymatic CD14 removal.
- LBP was depleted from serum using anti-LBP antibodies.
- Tumor necrosis factor (TNF) secretion was measured in response to LPS stimulation.
Main Results:
- Serum components, particularly LBP, significantly enhanced LPS binding to monocytes.
- CD14 was identified as the primary receptor mediating LPS binding and subsequent TNF secretion.
- Serum increased monocyte sensitivity to LPS-induced TNF secretion by 100- to 1000-fold.
- While CD14 was crucial, other receptors (Fc or complement) may also be involved in specific conditions.
Conclusions:
- LBP plays a critical role in facilitating LPS binding to monocytes, predominantly via CD14.
- The LBP-LPS complex significantly amplifies monocyte responses, leading to increased TNF secretion.
- Understanding these interactions is vital for comprehending innate immune responses to bacterial infections.
Abstract:
We used flow cytometry to determine how LPS-binding protein (LBP) effects the binding of fluorescein-labeled LPS to human monocytes via receptor-dependent mechanisms. The addition of human, rabbit, mouse, or FCS strikingly increased the binding of LPS to monocytes compared with controls incubated in serum-free medium. This binding was totally prevented by preincubation of monocytes with MY4, an anti-CD14 mAb, or by enzymatic removal of CD14 from monocytes. Depletion of LBP from rabbit serum with anti-LBP antibodies also produced a similar suppression. Solutions of albumin did not support the enhanced binding observed in serum but the addition of purified rabbit LBP to albumin solutions resulted in binding similar to that observed in serum-containing medium. When type-specific anti-LPS mAb was added to human serum, LPS binding to monocytes occurred but was only partly inhibited by anti-CD14 mAb, suggesting that receptors other than CD14 (presumably Fc or complement receptors) were involved. Serum increased by 100- to 1000-fold the sensitivity of monocytes to the triggering by LPS resulting in TNF secretion. TNF secretion was inhibited by anti-CD14 mAb up to 100 ng/ml of LPS and by anti-LPS mAb up to 1 to 10 ng/ml. The inhibition of TNF secretion by anti-LPS mAb appeared to be the result of directing LPS to monocyte receptors other than CD14. In contrast, in medium containing normal as well as acute serum and in the absence of anti-LPS antibodies, the binding of LPS to monocytes and the triggering of TNF secretion appeared to be mediated mainly by interactions between CD14 and LBP-LPS complexes.
