Context-dependent effects of SOCS3 in angiotensin II-induced vascular dysfunction and hypertension in mice: mechanisms and role of bone marrow-derived cells

Abstract

Carotid artery disease is a major contributor to stroke and cognitive deficits. Angiotensin II (Ang II) promotes vascular dysfunction and disease through mechanisms that include the IL-6/STAT3 pathway. Here, we investigated the importance of suppressor of cytokine signaling 3 (SOCS3) in models of Ang II-induced vascular dysfunction. We examined direct effects of Ang II on carotid arteries from SOCS3-deficient (SOCS3(+/-)) mice and wild-type (WT) littermates using organ culture and then tested endothelial function with acetylcholine (ACh). A low concentration of Ang II (1 nmol/l) did not affect ACh-induced vasodilation in WT but reduced that of SOCS3(+/-) mice by ∼50% (P < 0.05). In relation to mechanisms, effects of Ang II in SOCS3(+/-) mice were prevented by inhibitors of STAT3, IL-6, NF-κB, or superoxide. Systemic Ang II (1.4 mg/kg per day for 14 days) also reduced vasodilation to ACh in WT. Surprisingly, SOCS3 deficiency prevented most of the endothelial dysfunction. To examine potential underlying mechanisms, we performed bone marrow transplantation. WT mice reconstituted with SOCS3(+/-) bone marrow were protected from Ang II-induced endothelial dysfunction, whereas reconstitution of SOCS3(+/-) mice with WT bone marrow exacerbated Ang II-induced effects. The SOCS3 genotype of bone marrow-derived cells did not influence direct effects of Ang II on vascular function. These data provide new mechanistic insight into the influence of SOCS3 on the vasculature, including divergent effects depending on the source of Ang II. Bone marrow-derived cells deficient in SOCS3 protect against systemic Ang II-induced vascular dysfunction.

Keywords: carotid artery disease; cerebral arteries; endothelial dysfunction; endothelium; suppressor of cytokine signaling 3.

Fig. 1.

Effects of angiotensin II (Ang II) on endothelial function are mediated by AT1 receptors. Vasodilation was induced by acetylcholine (A) and nitroprusside (B) in carotid arteries from C57BL/6 mice incubated with vehicle or Ang II (10 nM) in the presence and absence of losartan for 22 h; n = 6, *P < 0.05 vs. vehicle.

Fig. 2.

Effects of suppressor of cytokine signaling 3 (SOCS3) deficiency on local Ang II-induced endothelial dysfunction. A: SOCS3 and SOCS1 gene expression in aorta from SOCS3^+/+ and SOCS3^+/− mice (n = 6). *P < 0.05 vs. SOCS3^+/+. B: acetylcholine-induced relaxation in carotid arteries from SOCS3^+/+ and SOCS3^+/− mice incubated with vehicle or Ang II (1 nM) (n = 6). *P < 0.05 vs. vehicle.

Fig. 3.

Endothelium-independent relaxation in carotid arteries was not affected by Ang II or SOCS3 deficiency. Nitroprusside induced relaxation in arteries from SOCS3^+/+ and SOCS3^+/− mice incubated with vehicle or Ang II (1 nM); n = 6, *P < 0.05 vs. vehicle.

Fig. 4.

Ang II-induced endothelial dysfunction in SOCS3^+/− arteries was mediated by oxidative stress and inflammation. Acetylcholine induced relaxation in carotid arteries from SOCS3^+/+ and SOCS3^+/− mice incubated with vehicle or Ang II (1 nM) with or without a superoxide scavenger (tempol) (A), NF-κB essential modulator (NEMO)-binding domain (NBD), which is an inhibitory peptide of NF-κB (B), an IL-6-neutralizing antibody (anti-IL-6) (C), or a small-molecule inhibitor of STAT3 (S3I-201) (D); n = 6.

Fig. 5.

Endothelium-independent relaxation was not affected by Ang II or other treatments. Nitroprusside induced relaxation in carotid arteries from SOCS3^+/+ and SOCS3^+/− mice incubated with vehicle or Ang II in the presence and absence of tempol (1 mM) (A), an inhibitory peptide of NF-κB (NBD, 10 μM) (B), an IL-6-neutralizing antibody (anti-IL-6, 10 μg/ml) (C), or a small-molecule inhibitor of STAT3 (S3I-201, 10 μM) (D); n = 6.

Fig. 6.

Role of SOCS3 in Ang II-dependent hypertension. A: acetylcholine-induced relaxation in carotid arteries from SOCS3^+/+ and SOCS3^+/− mice systemically administrated with vehicle or Ang II (1.4 mg/kg per day) for 14 days. The role of oxidative stress was examined by acute administration of tempol; n = 6, *P < 0.05 vs. vehicle. B: vasodilation induced by acetylcholine in basilar arteries from SOCS3^+/+ and SOCS3^+/− mice systemically administrated with either vehicle or Ang II (1.4 mg/kg per day) for 14 days; n = 4, *P < 0.05 vs. vehicle. C: systolic blood pressure in SOCS3^+/+ and SOCS3^+/− mice systemically administrated with either vehicle or Ang II for 14 days. *P < 0.05 vs. vehicle.

Fig. 7.

Vascular function following systemic administration of Ang II. A and B: nitroprusside-induced relaxation (A) and U46619-induced contraction (B) in carotid arteries from SOCS3^+/+ and SOCS3^+/− mice treated with vehicle or Ang II (1.4 mg/kg per day) for 14 days; n = 6. C: nitroprusside-induced vasodilation in basilar arteries from SOCS3^+/+ and SOCS3^+/− mice treated with vehicle or Ang II (1.4 mg/kg per day) for 14 days; n = 4. D: systolic blood pressure measured using tail-cuff in SOCS3^+/+ and SOCS3^+/− mice systemically administrated with a nonpressor dose of Ang II for 14 days; n = 3. E and F: endothelium-dependent (E) and endothelium-independent (F) relaxation in carotid arteries from SOCS3^+/+ and SOCS3^+/− mice treated with the nonpressor dose of Ang II for 14 days; n = 3. The role of oxidative stress in each group was examined by acute administration of a superoxide scavenger tempol (1 mM).

Fig. 8.

Effects of SOCS3 deficiency on vasoconstriction. Ang II induced contraction in abdominal aorta (A), iliac arteries (B), and femoral arteries (C) from SOCS3^+/+ (n = 5) and SOCS3^+/− mice (n = 7). U46619 induced contraction in abdominal aorta (D), iliac arteries (E), and femoral arteries (F) from SOCS3^+/+ (n = 5) and SOCS3^+/− mice (n = 7).

Fig. 9.

Fluorescence-activated cell sorting. A: reconstitution of irradiated SOCS3^+/− mice with WT (CD45.1) bone marrow. X-axis represents WT (CD45.1) bone marrow. Y-axis represents SOCS3^+/− bone marrow. B: reconstitution of irradiated WT (CD45.1) mice with SOCS3^+/− bone marrow. X-axis represents WT (CD45.1) bone marrow. Y-axis represents SOCS3^+/− bone marrow. C: reconstitution of irradiated SOCS3^+/+ mice with WT (CD45.1) bone marrow. X-axis represents WT (CD45.1) bone marrow. Y-axis represents SOCS3^+/+ bone marrow.

Fig. 10.

Endothelial function and blood pressure of bone marrow chimeras in chronic Ang II-dependent hypertension. Acetylcholine induced relaxation in carotid arteries from WT (CD45.1) to SOCS3^+/+ (A), SOCS3^+/− to SOCS3^+/− (B), SOCS3^+/− to WT (CD45.1) (C), and WT (CD45.1) to SOCS3^+/− (D) bone marrow chimeras systemically treated with vehicle or Ang II (1.4 mg/kg per day) for 14 days. *P < 0.05 vs. vehicle. E: acetylcholine-induced maximum relaxation in carotid arteries from WT (CD45.1) to SOCS3^+/+, SOCS3^+/− to WT (CD45.1), SOCS3^+/− to SOCS3^+/−, and SOCS3^+/− to WT (CD45.1) bone marrow chimeras treated systemically with vehicle or Ang II (1.4 mg/kg per day) for 14 days; n = 6, *P < 0.05 vs. vehicle. F: blood pressure of WT (CD45.1) to SOCS3^+/−, SOCS3^+/− to WT (CD45.1) bone marrow chimeras was measured on day 14 of Ang II (1.4 mg/kg per day) infusion; n = 6, *P < 0.05 vs. vehicle.

Fig. 11.

Endothelial function in bone marrow chimeras in response to local effects of Ang II. Acetylcholine induced relaxation in carotid arteries from WT (CD45.1) to SOCS3^+/+ (A), SOCS3^+/− to SOCS3^+/− (B), SOCS3^+/− to WT (CD45.1) (C), and WT (CD45.1) to SOCS3^+/− (D) bone marrow chimeras incubated with either vehicle or Ang II; n = 3, *P < 0.05 vs. vehicle.