![Figure 5. Thromboprotective mechanisms of Klkb1−/− mice are mediated by prostacyclin through Sirt1 and KLF4. (A) Plasma prostacyclin levels in Klkb1+/+ (n = 8), Klkb1−/− (n = 13), and Klkb1−/− mice + A779 (n = 6) as determined by 6-keto-PGF1α. (B) Carotid artery occlusion times in Klkb1+/+ and Klkb1−/− mice treated or untreated with nimesulide (Nimes) (n = 4-8 animals per group). (C) mRNA expression of Sirt1 and KLF4 in aortic tissues from Klkb1+/+ and Klkb1−/− mice in the absence or presence of nimesulide (n = 12 aortas per group). (D) The presence of Sirt1 and KLF4 antigen in aortic lysates. The gels on the left represent immunoblots of the murine aorta for Sirt1 or KLF4 antigen in Klkb1+/+ and Klkb1−/− mice. The graph on the right is a ratio of Sirt1 or KLF4 antigen compared with β-actin (n = 3-5 blots per condition). (E) The presence of the carotid artery Sirt1 antigen on immunofluorescence is shown on the left. The images were taken with a Nikon Eclipse TE2000-S microscope at 4× magnification. The graph on the right is the relative degree of immunofluorescence in the carotid arteries (n = 4-5 vessels per genotype). (F) The presence of carotid artery KLF4 antigen on immunofluorescence is shown on the left. The images were taken as before at 40×. The graph on the right is the relative degree of immunofluorescence in the carotid arteries (n = 10-11 vessels per genotype). (G) mRNA levels of Sirt1 and KLF4 in cultured MCECs after carbaprostacyclin (cPGI2) treatment. MCECs were treated with carbaprostacyclin (5 µg/mL) (n = 12) or vehicle (0.1% dimethyl sulfoxide [DMSO]) (n = 12) for 4 hours. (H-I) Immunoblot (H) and quantification (I) for protein expression of Sirt1 and KLF4 in untreated (UT) or cPGI2-treated (T) MCECs as in (G). (J) Carotid occlusion times in Klkb1+/+ and Klkb1−/− mice treated with splitomicin (Splito) or DMSO alone (n = 7-10 animals per group). Data are presented as mean ± SEM for all experiments. *Significant differences (P < .05) between the 2 groups on Student t test or 1-way ANOVA test when more than 2 groups of data are compared.](https://ash.silverchair-cdn.com/ash/content_public/journal/blood/125/4/10.1182_blood-2014-01-550285/4/710f5.jpeg?Expires=1735498793&Signature=hDEb7TWHu~ExYvD1on~EsVBJlmI8nzS7nGLCeOo3QQ69VIt3jTIQPiRsQ~~1bJKuy1E-USd6KeQXOmCweyrTzSo370BjcQvx1UMCgPCuoMcI7vVtC3Jp5V3K1Dn3CliTmhq-ktAXU8A6VDNSEwmovpgZmGkMaeGfPu9korvmIjRbnV~lBl24K6ON2VbQT1J-BQjqduhaZvdwxmZZjLg2rIib0koXFzEkF720M5KE0OzPWswgBHKFDVJq~eKwKEe-2HxZHuSn1VvuWDQGD2fIgW3emjXUfpss8Y8sm3qHz-MbM-pyXiQY-z3ZSTBcYrp2IKiV4bHg2zSQXTb2wxDDPA__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
Thromboprotective mechanisms of Klkb1−/− mice are mediated by prostacyclin through Sirt1 and KLF4. (A) Plasma prostacyclin levels in Klkb1+/+ (n = 8), Klkb1−/− (n = 13), and Klkb1−/− mice + A779 (n = 6) as determined by 6-keto-PGF1α. (B) Carotid artery occlusion times in Klkb1+/+ and Klkb1−/− mice treated or untreated with nimesulide (Nimes) (n = 4-8 animals per group). (C) mRNA expression of Sirt1 and KLF4 in aortic tissues from Klkb1+/+ and Klkb1−/− mice in the absence or presence of nimesulide (n = 12 aortas per group). (D) The presence of Sirt1 and KLF4 antigen in aortic lysates. The gels on the left represent immunoblots of the murine aorta for Sirt1 or KLF4 antigen in Klkb1+/+ and Klkb1−/− mice. The graph on the right is a ratio of Sirt1 or KLF4 antigen compared with β-actin (n = 3-5 blots per condition). (E) The presence of the carotid artery Sirt1 antigen on immunofluorescence is shown on the left. The images were taken with a Nikon Eclipse TE2000-S microscope at 4× magnification. The graph on the right is the relative degree of immunofluorescence in the carotid arteries (n = 4-5 vessels per genotype). (F) The presence of carotid artery KLF4 antigen on immunofluorescence is shown on the left. The images were taken as before at 40×. The graph on the right is the relative degree of immunofluorescence in the carotid arteries (n = 10-11 vessels per genotype). (G) mRNA levels of Sirt1 and KLF4 in cultured MCECs after carbaprostacyclin (cPGI2) treatment. MCECs were treated with carbaprostacyclin (5 µg/mL) (n = 12) or vehicle (0.1% dimethyl sulfoxide [DMSO]) (n = 12) for 4 hours. (H-I) Immunoblot (H) and quantification (I) for protein expression of Sirt1 and KLF4 in untreated (UT) or cPGI2-treated (T) MCECs as in (G). (J) Carotid occlusion times in Klkb1+/+ and Klkb1−/− mice treated with splitomicin (Splito) or DMSO alone (n = 7-10 animals per group). Data are presented as mean ± SEM for all experiments. *Significant differences (P < .05) between the 2 groups on Student t test or 1-way ANOVA test when more than 2 groups of data are compared.
