Interleukin-1 beta, tumor necrosis factor-alpha and lipopolysaccharide induce expression of monocyte chemoattractant protein-1 in calf aortic smooth muscle cells.
To investigate whether interleukin-1 beta(IL-1 beta), tumor necrosis factor-alpha (TNF-alpha) and lipopolysaccharide (LPS) induce expression of monocyte chemoattractant protein-1 (MCP-1) mRNA and protein in calf aortic smooth muscle cells(SMCs), calf aortic SMCs were cultured by a substrate-attached explant method. The cultured SMCs were used between the third to the fifth passage. After the cells became confluent, the SMCs were exposed to 2 ng/ml IL-1 beta, 20 ng/ml TNF-1 alpha and 100 ng/ml LPS respectively, and the total RNA of SMCs which were incubated for 4 h at 37 degrees C were extracted from the cells by using guanidinium isothiocyanate method. The expression of MCP-1 mRNA in SMCs was detected by using dot blotting analysis using a probe of gamma-32P-end-labelled 35-mer oligonucleotide. After a 24-h incubation, the media conditioned by the cultured SMCs were collected. The MCP-1 protein content in the conditioned media was determined by using sandwich ELISA. The results were as follows: Dot blotting analysis showed that the cultured SMCs could express MCP-1 mRNA. After a 4-h exposure to IL-1 beta, TNF-alpha and LPS, the MCP-1 mRNA expression in SMCs was increased (3.6-fold, 2.3-fold and 1.6-fold, respectively). ELISA showed that the levels of MCP-1 protein in the conditioned media were also increased (2.9-fold, 1.7-fold and 1.1-fold, respectively). The results suggest that calf aortic SMCs could express MCP-1 mRNA and protein. IL-1 beta and TNF-alpha can induce strong expression of MCP-1 mRNA and protein, and the former is more effective than the latter.
Meng
F
F
Department of Pathology, School of Basic Medical Sciences, Tongji Medical University, Wuhan 430030.
Deng
Z
Z
Ni
J
J
D000818:Animals; D001011:Aorta / Q000378:metabolism; D002417:Cattle; D018932:Chemokine CCL2 / Q000096:biosynthesis* / Q000235:genetics; D007375:Interleukin-1 / Q000494:pharmacology*; D008070:Lipopolysaccharides / Q000494:pharmacology*; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D018932:Chemokine CCL2; D007375:Interleukin-1; D008070:Lipopolysaccharides; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha