Cloning of differentially expressed genes following lipopolysaccharide stimulation in human umbilical vein endothelial cells.

Objective: To clone the differentially expressed genes in human umbilical vein endothelial cells (HUVEC) stimulated by lipopolysaccharide (LPS).

Methods: Two-directional (forward and backward) suppression subtractive hybridization (SSH) was performed on HUVEC cultured in either standard media or treated for 6 hours with LPS (100 ng/ml). To restrict the number of false-positive clones, colony dot hybridization was used to further verify the differentially expressed cDNA clones. Positive clones were sequenced.

Results: These analyses have identified both novel and known genes whose expression is influenced by LPS. The known genes include a group related to proinflammatory events, a group related to cellular apoptosis and proliferation, a group related to protein synthesis and cytoskeletal rearrangment, and a group related to energy metabolism and signal transduction.

Conclusions: SSH is a powerful technique of high sensitivity for the detection of differential gene expression in HUVEC stimulated by LPS.