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Updated: Aug 13, 2026
Screening Assays to Characterize Novel Endothelial Regulators Involved in the Inflammatory Response
Published on: September 15, 2017
GPI-linked endothelial CD14 contributes to the detection of LPS
1Immunology Research Group, Department of Biophysics and Physiology, Institute of Infection, Immunity and Inflammation, University of Calgary, Calgary, Alberta T2N 4N1, Canada.
Abstract:
The inflammatory endothelial response to LPS is critical to the host's surviving a gram-negative bacterial infection. In this study we investigated whether human endothelial cells express the functional coreceptor for LPS, CD14, and most importantly whether it is glycosylphosphatidylinositol (GPI) linked. We also examined whether plasma proteins could reconstitute an LPS response in CD14-inhibited endothelium. RT-PCR- and CD14-specific MAbs demonstrated CD14 expression on primary human umbilical vein endothelial cells (HUVEC) but not passaged HUVEC. The amino acid sequence of endothelial CD14 was 99% homologous to CD14 on monocytes. Endothelium responded to relatively low levels of LPS in the absence of plasma, and this was entirely dependent on CD14. Removal of GPI-linked proteins with phosphatidylinositol-phospholipase C prevented LPS detection and subsequent protein synthesis (E-selectin expression). Endothelial CD14 was sufficient to initiate functional leukocyte recruitment, an event inhibited by blocking its LPS binding epitope and also by removing CD14 from the endothelial surface. Plasma proteins restored only approximately 30% of the LPS response in CD14-inhibited endothelium. In conclusion, our results strongly support an important role for endothelial membrane CD14 in the activation of endothelium for leukocyte recruitment.
Insights
Human endothelial cells express CD14, a key receptor for lipopolysaccharide (LPS), crucial for initiating inflammatory responses and leukocyte recruitment during bacterial infections. This membrane-bound CD14 is vital for endothelial cell activation.
Area of Science:
- Immunology
- Cell Biology
- Vascular Biology
Background:
- The inflammatory response of endothelial cells to lipopolysaccharide (LPS) is vital for survival during gram-negative bacterial infections.
- CD14 is a known coreceptor for LPS, primarily studied on monocytes.
Purpose of the Study:
- To investigate the expression and function of CD14 in human endothelial cells.
- To determine if endothelial CD14 is glycosylphosphatidylinositol (GPI)-linked.
- To assess the role of plasma proteins in reconstituting LPS response in CD14-inhibited endothelium.
Main Methods:
- RT-PCR and CD14-specific monoclonal antibodies (MAbs) were used to detect CD14 expression on primary human umbilical vein endothelial cells (HUVEC).
- Phosphatidylinositol-phospholipase C was employed to remove GPI-linked proteins.
- Functional assays measured leukocyte recruitment and E-selectin expression in response to LPS.
Main Results:
- CD14 was expressed on primary HUVEC but not on passaged HUVEC, with 99% amino acid homology to monocyte CD14.
- Endothelial cells responded to LPS independently of plasma, a response dependent on CD14.
- Removal of GPI-linked proteins abolished LPS detection and E-selectin expression; endothelial CD14 initiated leukocyte recruitment.
Conclusions:
- Endothelial membrane CD14 is expressed and functional, playing a critical role in LPS-induced endothelial activation.
- Endothelial CD14 is sufficient for initiating leukocyte recruitment, independent of plasma factors.
- GPI-linked endothelial CD14 is essential for detecting LPS and triggering downstream inflammatory signaling.
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