NTPDase1/CD39 Ectonucleotidase Is Necessary for Normal Arterial Diameter Adaptation to Flow

Figure 1. Impact of Entpd1 deficiency on contracting responses of conduit (ThAo) and resistance (MRA and LCA) arteries in response to pharmacological agonists. Aortic ((A); ThAo), mesenteric ((B); MRA) and coronary ((C); LCA) segments were isolated from male Entpd1+/+ and Entpd1−/− mice (n = 5 mice per group) and mounted on a wire-myograph. Phenylephrine (Phe) (ThAo and MRA) and serotonin (5HT) (LCA) induced concentration-dependent contractile responses. Means ± SEM are shown. Two-way ANOVA for repeated measurements Entpd1+/+ vs. Entpd1−/−, p values are shown on each graph such as Bonferroni post-test * p < 0.05 Entpd1+/+ vs. Entpd1−/−.

Figure 2. Impact of Entpd1 deficiency on vascular responses of conduit (ThAo) and resistance (MRA and LCA) arteries in response to ATP. Aortic ((A); ThAo), mesenteric ((B); MRA) and coronary ((C); LCA) segments were isolated from male Entpd1+/+ and Entpd1−/− mice (n = 5 mice per group) and mounted on a wire-myograph. Increasing cumulative concentrations of ATP on pre-contracted arteries induced a bimodal response resulting in a major relaxing response in Entpd1−/− mice. Means ± SEM are shown. Two-way ANOVA for repeated measurements Entpd1+/+ vs. Entpd1−/−, p values are shown on each graph such as Bonferroni post-test * p < 0.05 and ** p < 0.01, *** p < 0.001 Entpd1+/+ vs. Entpd1−/−.

Figure 3. Impact of Entpd1 deficiency on relaxing responses of conduit (ThAo) and resistance (MRA and LCA) arteries. Aortic (ThAo, left), mesenteric (MRA, middle) and coronary (LCA, right) segments were isolated from male Entpd1+/+ and Entpd1−/− mice (n = 5 mice per group) and mounted on a wire-myograph. (A) Endothelium-dependent relaxations were assessed in response to increasing cumulative concentrations of Ach on pre-contracted arteries. (B) NO-independent relaxations were measured in the presence of NOS antagonist LNAME (10−4 M) alone or with indomethacin (+INDO 10−5 M, in gray) for further COX-inhibition. (C) Endothelium-independent relaxation was assessed in response to the NO donor SNP. Means ± SEM are shown. Two-way ANOVA for repeated measurements Entpd1+/+ vs. Entpd1−/−, p values are shown on each graph such as Bonferroni post-test * p < 0.05 and ** p < 0.01 Entpd1+/+ vs. Entpd1−/−; # p < 0.05 Entpd1+/+ LNAME vs. Entpd1+/+ LNAME+INDO.

Figure 4. MRA vascular response to acute and chronic increase in intraluminal flow. (A) Flow-mediated dilation was measured in mesenteric resistance arteries isolated from male Entpd1+/+ and Entpd1−/− mice (n = 7 mice per group). (B) Schematic representation of the protocol used to induce the flow-mediated remodeling of mesenteric arteries in vivo (see Materials and Methods). Following ligation of some second-order mesenteric arteries downstream the first-order mesenteric arteries, first-order arteries with ligation are submitted chronically to high flow (HF) and compared with the first-order mesenteric arteries without ligation (normal flow, NF) and with ligation (low flow, LF). (C,D) 2 weeks after surgery, flow-mediated remodeling was evaluated by the measurement of passive diameter in mesenteric arteries isolated from WT Entpd1+/+ (C) and Entpd1−/− (D) mice in response to stepwise increases in intraluminal pressure (n = 6 per group). Means ± SEM are shown. two-way ANOVA for repeated measurements ((A): Entpd1−/− versus Entpd1+/+ mice; (C,D): HF and LF versus NF control; p values are shown on each graph) and Bonferroni post-test were performed * p < 0.05, ** p < 0.01, *** p < 0.001.

Figure 5. Impact of Entpd1 deletion on isolated perfused heart. (A) Coronary output, (B) heart rate and (C) left ventricular functions were evaluated on isolated pressurized and perfused hearts of Entpd1−/− (white circles) and Entpd1+/+ (black circles) littermate mice. Systolic function was evaluated through maximal pressure (Pmax) and maximal change in pressure over time (dP/dt max) and diastolic function through minimal (Pmin), end-diastolic pressures (Ped) and minimal change in pressure over time (dP/dt min). Measurements were performed either in resting conditions (Rest) or on paced working heart (Paced). Means ± SEM are shown. Regular two-way ANOVA ((A–C): Entpd1−/− versus Entpd1+/+ mice) and Bonferroni post-test were performed. * p < 0.05 Entpd1−/− vs. Entpd1+/+ mice.