The angiotensin II AT1 receptor-associated protein Arap1 is involved in sepsis-induced hypotension

Abstract

Introduction: Hypotension in septic patients results from hypovolemia, vasodilatation and hyporeactivity to vasoconstrictors, such as angiotensin II. The AT1 receptor-associated protein 1 (Arap1) is expressed in vascular smooth muscle cells and increases the surface expression of the AT1-receptor in vitro. We hypothesized that dysregulation of Arap1 may contribute to vascular hyporeactivity to angiotensin II during endotoxemia.

Methods: Arap1-deficient mice were used to assess the role of Arap1 in sepsis-induced hypotension. The isolated perfused kidney was used as an in vitro model to determine the relevance of Arap1 for vascular resistance and sensitivity to angiotensin II.

Results: During endotoxemia, mean arterial blood pressure (MAP) decreased in both genotypes, with the time course of sepsis-induced hypotension being markedly accelerated in Arap1-/- compared to +/+ mice. However, baseline MAP was similar in Arap1-/- and wildtype mice (102 ± 2 vs.103 ± 2 mmHg; telemetry measurements; n = 10; P = 0.66). Following lipopolysaccharide (LPS) injections (3 mg/kg), Arap1 expression was successively down-regulated in the wildtype mice, reaching levels below 10% of baseline expression. The endotoxemia-related decline in Arap1 expression could be recapitulated in cultured mesangial cells by incubation with pro-inflammatory cytokines, such as tumor necrosis factor α and interferon γ. Plasma renin concentration was increased in Arap1-/- mice compared to wildtype mice (66 ± 6 vs. 41 ± 4 ng AngI/ml/h; n = 23; P = 0.001), presumably contributing to preserved MAP under baseline conditions. The sensitivity of the vasculature to angiotensin II was reduced in Arap1-/- compared to +/+ mice, as determined in the isolated perfused kidney.

Conclusions: Our data suggest that down-regulation of Arap1 expression during sepsis contributes to the development of hypotension by causing reduced vascular sensitivity to angiotensin II.

Figure 1

Mean arterial blood pressure (MAP), heart rate and activity score for angiotensin receptor-associated protein (Arap)1-/- and wildtype mice. MAP, heart rate and activity score were recorded by radio-telemetry for three consecutive days (n = 5 each). Shaded areas indicate light-off periods.

Figure 2

Mean arterial blood pressure (MAP) in wildtype and angiotensin receptor-associated protein (Arap)1 -/- mice during endotoxemia. MAP was measured by radio-telemetry following a single i.p. injection of lipopolysaccharide (LPS) (3 mg/kg) (n = 10 animals for each genotype).

Figure 3

Mean arterial blood pressure (MAP) during endotoxemia in angiotensin converting enzyme (ACE) inhibitor-pretreated mice. Wildtype and angiotensin receptor-associated protein (Arap)1 -/- mice were pretreated with the ACE inhibitor enalapril (10 mg/kg/d) (n = 5 animals for each genotype).

Figure 4

Angiotensin receptor-associated protein (Arap)1 mRNA expression after Lipopolysaccharide (LPS) injection (A) Renal Arap1 mRNA expression 0, 3, 6 and 12 hours after LPS injection determined by quantitative RT-PCR. Values are given as relative units normalized to β-actin expression (n = 6 for each genotype). (B) Relative Arap1 mRNA expression 0, 3, 6 and 12 hours after LPS injection in the heart, aorta, adrenal gland and lung compared to vehicle-injected animals (n = 6 for each group). (#P <0.05 compared to control; *P <0.01 compared to control).

Figure 5

Angiotensin receptor-associated protein (Arap)1 mRNA expression in cultured rat glomerular mesangial cells. Arap1 mRNA expression was measured following incubation with (A) a cytokine mix containing IL-1β (50 ng/mL), tumor necrosis factor (TNF)-α (100 ng/mL) and interferon (IFN)-γ (100 ng/mL) or (B) single cytokines (interleukin (IL)-1β (50 ng/mL), TNF-α (100 ng/mL), IFN-γ (100 ng/mL).

Figure 6

Isolated perfused kidney (A) Perfusate flow at constant perfusion pressure of 100 mmHg in isolated perfused kidneys from angiotensin receptor-associated protein (Arap)1-/- and wildtype mice (n = 5). After pre-relaxation with isoproterenol (10 nM), angiotensin II was added to the perfusate to a final concentration of 10, 30, 100, 300 and 100 pM. For each concentration, three values were determined over a period of three minutes. (B) Renin secretion from isolated kidneys perfused according to the same protocol (n = 5).

Figure 7

Plasma renin concentration in conscious angiotensin receptor-associated protein (Arap)1-/- and wildtype mice (n = 23 for each genotype).