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Figure 2 - Protein Z Exerts Pro-Angiogenic Effects and Upregulates CXCR4

Figure 2.  PZ in ischemia-induced neovascularization in vivo.
A, Representative pictures of thermal imaging of mice hindlimbs. B, Quantitative summary of pad temperature differences. On POD 21 PZ+/+ mice showed a significant reduction in temperature difference compared to immediately after induction of ischemia and significant lower temperature difference to PZ−/− mice, which offered no changes. Data are given as box plots indicating the median with the 25th and 75th percentiles. ANOVA on ranks repeated measures; * p<0.05 vs. pre OP, § p<0.05 vs. post OP, # p<0.05 vs. other genotype on POD 21, n = 11–16. C, After 21 days, PZ−/− mice treated with AdV-PZ showed significant reduction in temperature difference in the ischemic hindlimb. Data are given as box plots indicating the median with the 25th and 75th percentiles. ANOVA on ranks repeated measures; * p<0.05 vs. pre OP, § p<0.05 vs. post OP; n = 10. D, Representative pictures of M. gastrocnemius muscle tissue after immunofluorescent staining for CD31 (red) or cell nuclei (DAPI, blue). Intracardiac injection of an endothelial specific lectin confirmed functional perfusion of CD31/DAPI double positive capillaries (green). E, PZ+/+ mice exhibited significant higher density of CD31/DAPI-double positive capillaries per square millimeter of ischemic M. gastrocnemius tissue compared to the non-ischemic PZ+/+ muscle tissue. In muscle tissue of PZ−/− mice, ischemia did not result in remarkable neovascularization. The number of CD31/DAPI double-positive cells in ischemic PZ−/− mice M. gastrocnemius was even significantly lower compared to ischemic PZ+/+ mice. Data are given as box plots indicating the median with the 25th and 75th percentiles. ANOVA on ranks; & p<0.05 vs. non-ischemic, # p<0.05 vs. other genotype with ischemia; n = 11–16. F, After exposure of PZ−/− mice to AdV-PZ, the compromised angiogenic phenotype of PZ−/− mice after induction of ischemia was significantly reversed. Data are given as box plots indicating the median with the 25th and 75th percentiles. ANOVA on ranks; & p<0.05 vs. non-ischemic, # p<0.05 vs. AdV-GFP with ischemia, n = 10.
PZ in ischemia-induced neovascularization in vivo. A, Representative pictures of thermal imaging of mice hindlimbs. B, Quantitative summary of pad temperature differences. On POD 21 PZ+/+ mice showed a significant reduction in temperature difference compared to immediately after induction of ischemia and significant lower temperature difference to PZ−/− mice, which offered no changes. Data are given as box plots indicating the median with the 25th and 75th percentiles. ANOVA on ranks repeated measures; * p<0.05 vs. pre OP, § p<0.05 vs. post OP, # p<0.05 vs. other genotype on POD 21, n = 11–16. C, After 21 days, PZ−/− mice treated with AdV-PZ showed significant reduction in temperature difference in the ischemic hindlimb. Data are given as box plots indicating the median with the 25th and 75th percentiles. ANOVA on ranks repeated measures; * p<0.05 vs. pre OP, § p<0.05 vs. post OP; n = 10. D, Representative pictures of M. gastrocnemius muscle tissue after immunofluorescent staining for CD31 (red) or cell nuclei (DAPI, blue). Intracardiac injection of an endothelial specific lectin confirmed functional perfusion of CD31/DAPI double positive capillaries (green). E, PZ+/+ mice exhibited significant higher density of CD31/DAPI-double positive capillaries per square millimeter of ischemic M. gastrocnemius tissue compared to the non-ischemic PZ+/+ muscle tissue. In muscle tissue of PZ−/− mice, ischemia did not result in remarkable neovascularization. The number of CD31/DAPI double-positive cells in ischemic PZ−/− mice M. gastrocnemius was even significantly lower compared to ischemic PZ+/+ mice. Data are given as box plots indicating the median with the 25th and 75th percentiles. ANOVA on ranks; & p<0.05 vs. non-ischemic, # p<0.05 vs. other genotype with ischemia; n = 11–16. F, After exposure of PZ−/− mice to AdV-PZ, the compromised angiogenic phenotype of PZ−/− mice after induction of ischemia was significantly reversed. Data are given as box plots indicating the median with the 25th and 75th percentiles. ANOVA on ranks; & p<0.05 vs. non-ischemic, # p<0.05 vs. AdV-GFP with ischemia, n = 10.
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