Objective: To investigate whether the proinflammatory cytokines IL-1beta, TNF-alpha and lipopolysaccharide (LPS) enable to induce the expression of MCP-1 mRNA and protein in human umbilical vein endothelial cells (HUVECs).
Methods: After a four-hour exposure to 2 ng/ml IL-1beta, 20 ng/ml TNF-alpha or 100 ng/ml LPS, total RNA of HUVECs was extracted by single-step method. The expression of MCP-1 mRNA in HUVECs was examined by dot blot analysis using a probe of gamma-(32)P-end-labelled 35 mer oligonucleotide. Meanwhile, MCP-1 protein in the cytoplasm was detected by SABC immunostaining.
Results: Dot blot analysis showed that cultured HUVECs were able to express MCP-1 mRNA at a low level. Exposure to IL-1beta, TNF-alpha and LPS resulted in a 7.8-fold, 2.6-fold and 1.2-fold induction of MCP-1 mRNA expression in HUVECs, respectively. The cells on the coverglass in all groups revealed MCP-1 immunoreactivity. Densitometry scans showed that the mean absorbance (A) values of the cells in LPS, TNF-alpha and IL-1beta groups were 0.078 +/- 0.113, 0.102 +/- 0.005 and 0.117 +/- 0.010, respectively; whereas the absorbance values of the control group was 0.051 +/- 0.004. There were significant differences between all the experimental groups and the control group (F = 193.25, P < 0.01).
Conclusions: IL-1beta and TNF-alpha induce a strong expression of MCP-1 mRNA and protein in HUVECs. Both cytokines may be involved in the atherogenesis by inducing the liberation of MCP-1 by endothelial cells and increasing the recruitment of monocytes into the subendothelial space.