FIGURE 5.
Hsp90α stabilizes MMP-2 and promotes the transmigration and tube formation of endothelial cells in vitro and in vivo. A, rHsp90α, rHsp90β, mouse IgG, Hsp90α mAb, or Hsp90β mAb (5 μg/ml) was added to the culture medium (without FCS) of HMECs. After 24 h of treatment, the CM was collected and concentrated by 10-fold. MMP-2 and its fragments were detected by anti-MMP-2 antibody. B, the transmigration of HMECs (transfected with MMP-2 shRNA or not) treated by Hsp90 recombinant proteins or antibodies (5 μg/ml) was examined by Matrigel transmigration assay in Millicell chambers. The transmigrated cells were stained by hematoxylin and eosin and counted. The error bars represent S.D. (n = 5). p value: Student's t test. *, p < 0.05; **, p < 0.01; #, p > 0.05. C, the tube formation of HMECs (transfected with MMP-2 shRNA or not) treated by Hsp90 recombinant proteins or antibodies (5 μg/ml) were examined by Matrigel model in vitro. The error bars represent S.D. (n = 5). p value: Student's t test. *, p < 0.05; **, p < 0.01; #, p > 0.05. D, the effects of Hsp90 recombinant proteins or antibodies (50 μg/ml) on tube formation in Matrigel plug containing lentivirus delivered MMP-2 shRNA or not were examined in nude mice. The tube formation was detected by immunofluorescence using CD31 staining. The density of the tube was calculated by NIS-Elements C, the software of Nikon A1 confocal. The error bars represent S.D. (n = 5). p value: Student's t test. *, p < 0.05; **, p < 0.01; #, p > 0.05. MW, molecular mass.