Fig. 5. Analysis of signal pathways in response to MCP-1. (A) cDNA microarray analysis of signal pathways induced by MCP-1. Data represent relative expression levels of specific genes normalized using a housekeeping gene control and compared with the group of untreated cells. Arrowheads indicate several representative genes whose levels increased in response to MCP-1. (B) Confluent mBMEC were pretreated with the following inhibitors: PD98059, SB 203580, LY294002, Ro-37840, Y27632, W7 and U7322 for 1 hour at 37°C. Recombinant mouse MCP-1 (100 nM) was then added for 2 hours. The cells were then fixed and processed for immunocytochemistry using anti-ZO-1 and Alexa 488 Phalloidin. The samples were viewed on a laser scanning Zeiss confocal microscope. Scale bar: 200 µm. (C) Changes in TEER during treatment with specific inhibitors of different signal pathways. Confluent mBMEC were grown on the filters in a Transwells chamber system and pretreated with the following inhibitors: PD98059, SB 203580, LY294002, Ro-37840, Y27632, W7 and U7322 for 1 hour at 37°C. After that the cells were exposed to MCP-1 (100 nM) in the presence of inhibitors. TEER was measured every 15 minutes over a time period of 2 hours. Results are presented as means±s.e.m., ^*P<0.01; ^**P<0.001.

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