Glucose Regulation of Thrombospondin and Its Role in the Modulation of Smooth Muscle Cell Proliferation
Figure 3
TSP-1 protein levels regulate the response of SMC to hyperglycemia. (a) SMCs expressing either the TSP-1 or a scrambled (Scr) RNAi construct were grown to confluency in DMEM containing 25 mM glucose and then incubated overnight in SFM with 25 mM. Equal amounts of lysates were either subject to immunoprecipitation (IP) followed by immunoblotting or immunoblotted (IB) directly with the antibody indicated. The graph shows the mean fold decrease of each protein in the RNAi cultures compared with the Scr control of three independent experiments, (). (b) SMC expressing either the TSP-1 or a scrambled (Scr) RNAi construct were grown to confluency in DMEM containing 25 mM glucose and then incubated overnight in SFM with 25 mM prior to exposure to IGF-I (50 ng/mL) for 5 minutes (). Equal amounts of lysates were then subject to immunoprecipitation (IP) followed by immunoblotting (IB) with the antibody indicated. The graph shows the mean fold increase in phosphorylation of each protein in response to IGF-I ( when the extent of phosphorylation in the absence of IGF-I is compared to the addition of IGF-I). (c) SMC expressing either the TSP-1 or a scrambled (Scr) RNAi construct were grown to confluency in DMEM containing 25 mM glucose and then incubated overnight in SFM with 25 mM prior to exposure to IGF-I (50 ng/mL) for 5 minutes (). Equal amounts of lysates were then subject to immunoblotting (IB) with either an antiphospho or total ERK antibody. The graph shows the mean fold increase in phosphorylation in response to IGF-I ( when the extent of phosphorylation in the absence of IGF-I is compared to the addition of IGF-I).
