Figure 5.
Increased TF and PAI-1 in the IVC of animals with colon cancer xenografts. (A) IVC from mice shown in Figure 4A were harvested, divided for immunoblotting, and fixed for immunofluorescence microscopy. Of 8 mice per group, the lysates from 5 mice were resolved on sodium dodecyl sulfate–polyacrylamide gel electrophoresis gels due to sufficient quantity of protein in these samples. Equal amounts of samples were separately probed on different western blots with the indicated antibodies (owing to proximity of bands). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) served as a loading control. (B) The immunoblot bands of TF and PAI-1 shown in Figure 4A were normalized to GAPDH in the aforementioned gel. Normalized TF and PAI-1 from control and HT-29 mice are shown on box plots. Asterisks (* and **) compare the expression of TF and PAI-1, respectively, between the 2 groups. *P = .032 and **P = .0425. (C) Increased expression of TF in the IVC of animals with colon cancer xenografts. IVC from animals with a xenograft without IVC ligation were stained by using anti-TF and anti-CD31 antibodies. Alexa Fluor secondary antibodies were used. 4′,6-Diamidino-2-phenylindole (DAPI) was used for nuclear stain. Un-injected animals served as controls (CTL). Images are representative of IVCs from 8 mice in each group. Scale bar = 10 μm. (D) Three images per IVC were analyzed, and a region of interest was marked corresponding to the endothelial cells. Integrated density was normalized to the surface area of the region of interest measured in microns by using ImageJ. Data are presented as a box plot, and the Student t test was used to compare the 2 groups. **P = .002. (E) Increased expression of PAI-1 in the IVC of animals with colon cancer xenografts. IVCs from animals with xenografts without IVC ligation, as described earlier, were stained by using anti–PAI-1 and anti-CD31 antibodies. Images are representative from 8 mice in each group. Scale bar = 30 μm. (F) Integrated density for PAI-1 was performed as noted earlier. The data are presented as a box plot, and the Student t test was used to compare the 2 groups (**P = .009).

Increased TF and PAI-1 in the IVC of animals with colon cancer xenografts. (A) IVC from mice shown in Figure 4A were harvested, divided for immunoblotting, and fixed for immunofluorescence microscopy. Of 8 mice per group, the lysates from 5 mice were resolved on sodium dodecyl sulfate–polyacrylamide gel electrophoresis gels due to sufficient quantity of protein in these samples. Equal amounts of samples were separately probed on different western blots with the indicated antibodies (owing to proximity of bands). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) served as a loading control. (B) The immunoblot bands of TF and PAI-1 shown in Figure 4A were normalized to GAPDH in the aforementioned gel. Normalized TF and PAI-1 from control and HT-29 mice are shown on box plots. Asterisks (* and **) compare the expression of TF and PAI-1, respectively, between the 2 groups. *P = .032 and **P = .0425. (C) Increased expression of TF in the IVC of animals with colon cancer xenografts. IVC from animals with a xenograft without IVC ligation were stained by using anti-TF and anti-CD31 antibodies. Alexa Fluor secondary antibodies were used. 4′,6-Diamidino-2-phenylindole (DAPI) was used for nuclear stain. Un-injected animals served as controls (CTL). Images are representative of IVCs from 8 mice in each group. Scale bar = 10 μm. (D) Three images per IVC were analyzed, and a region of interest was marked corresponding to the endothelial cells. Integrated density was normalized to the surface area of the region of interest measured in microns by using ImageJ. Data are presented as a box plot, and the Student t test was used to compare the 2 groups. **P = .002. (E) Increased expression of PAI-1 in the IVC of animals with colon cancer xenografts. IVCs from animals with xenografts without IVC ligation, as described earlier, were stained by using anti–PAI-1 and anti-CD31 antibodies. Images are representative from 8 mice in each group. Scale bar = 30 μm. (F) Integrated density for PAI-1 was performed as noted earlier. The data are presented as a box plot, and the Student t test was used to compare the 2 groups (**P = .009).

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