Autoantibody activation of beta-adrenergic and muscarinic receptors contributes to an "autoimmune" orthostatic hypotension

Abstract

Background: Orthostatic hypotension (OH) is characterized by an abnormal autonomic response to upright posture. Activating autoantibodies to β1/2-adrenergic (AAβ1/2AR) and M2/3 muscarinic receptors (AAM2/3R) produce vasodilative changes in the vasculature that may contribute to OH.

Methods: Immunoglobulin (Ig)G from 6 patients with idiopathic OH harboring autoantibodies and from 10 healthy control subjects were examined for: 1) β1AR and M2R activity with a perfused Purkinje fiber assay and PKA assay in H9c2 cells and 2) vasodilator β2AR and M3R activity using a pressurized cremaster resistance arteriole assay. Changes in IgG activity with and without propranolol, atropine, and L-NAME were used to estimate AAβAR, AAM2R, and AAM3R activation of their respective functions.

Results: All six patients had elevated enzyme-linked immunosorbent assay titers to at least one of the receptors compared with controls. βAR-mediated contractility activity and M2R activity were increased in five of the six patients. IgG from all six patients produced a direct vasodilator effect on cremaster arterioles. βAR and nitric oxide synthase blockade led to near normalization of IgG-induced vasodilation.

Conclusion: AAβ1/2AR and AAM2/3R are present in some patients with idiopathic OH compatible with an in vivo effect. These autoantibodies and their cardiovascular effects provide new mechanistic insights into the pathophysiology of OH.

Figure 1. Effects of IgG from the 6 orthostatic hypotension patients on Purkinje contractility

A. Each value represents the mean ± SD of 15 consecutive contractions and was measured once the preparation had stabilized. The data are expressed as percent above basal levels of the buffer control. The IgG alone shows net activity. An increase in contractility over baseline with IgG plus muscarinic blocker atropine (Atr) represented the βAR effect of IgG (AAβAR). The difference in IgG effect on contractility between the presence and absence of atropine was a surrogate marker of the M2R inhibitory effect of IgG (AAM2R). The co-presence of atropine demonstrates that βAR activity was impaired partially by concurrent M2R inhibitory effects in 4 of the 6 patients. Patient 106 has marked M2R dominance while patient 159 is βAR dominant. B. The mean effect of IgG from the 6 patients on Purkinje contractility. The βAR activity of the IgG was measured as effect of IgG plus atropine (Atr) and M2R activity as IgG plus the non-selective β-blocker nadolol (Nad). The effect of IgG from 10 control subjects and the positive isoproterenol (ISO) control are also shown.

Figure 1. Effects of IgG from the 6 orthostatic hypotension patients on Purkinje contractility

A. Each value represents the mean ± SD of 15 consecutive contractions and was measured once the preparation had stabilized. The data are expressed as percent above basal levels of the buffer control. The IgG alone shows net activity. An increase in contractility over baseline with IgG plus muscarinic blocker atropine (Atr) represented the βAR effect of IgG (AAβAR). The difference in IgG effect on contractility between the presence and absence of atropine was a surrogate marker of the M2R inhibitory effect of IgG (AAM2R). The co-presence of atropine demonstrates that βAR activity was impaired partially by concurrent M2R inhibitory effects in 4 of the 6 patients. Patient 106 has marked M2R dominance while patient 159 is βAR dominant. B. The mean effect of IgG from the 6 patients on Purkinje contractility. The βAR activity of the IgG was measured as effect of IgG plus atropine (Atr) and M2R activity as IgG plus the non-selective β-blocker nadolol (Nad). The effect of IgG from 10 control subjects and the positive isoproterenol (ISO) control are also shown.

Figure 2. Effects of sera from the 6 orthostatic hypotension patients on PKA activity in H9c2 cells

The values are expressed as percent above basal levels of PKA in medium control. An increase in PKA activity with sera plus atropine (Atr) represented the βAR effect (AAβAR). The difference in sera effect between the presence and absence of atropine was a surrogate marker of the M2R inhibitory effect (AAM2R). This assay demonstrates a significant impact of muscarinic blockade on PKA activity in 5 of 6 subjects.

Figure 3. Effects of serum IgG from orthostatic hypotension patients on basal spontaneous myogenic tone of rat cremaster arterioles

Each IgG sample was perfused into the cremasteric arteriole after a stable baseline was achieved. A dosage response to buffer (control), 10, 100 and 300 μg/mL of IgG was recorded. The peak response for each IgG concentration is the percentage of the maximal Ca²⁺-free passive diameter (maximal dilation) for that arteriolar preparation. A 10% increase in dilation is considered significant. All of the samples caused a significant dilation compared to their tonic control value. Three control IgG samples and a dialyzed (to eliminate sodium azide preservative) pooled commercial IgG gave less than 5% change from baseline (data not shown).

Figure 4. The effects of βAR and eNOS blockade on IgG-mediated resistance arteriole dilatation

A. Representative tracing of the effect of sequential addition of the β-blocker propranolol (Prop 10⁻⁶ mol/L) and nitric oxide synthase inhibitor L-NAME (10⁻⁴ mol/L) on IgG-induced arteriolar dilation. B. Data for four of the subjects. Patients 118 and 236 had a return to baseline tone with combined blockade while 112 and 159 failed to return completely to control. No subject had residual vasoconstriction compared to baseline during combined blockade.

Figure 4. The effects of βAR and eNOS blockade on IgG-mediated resistance arteriole dilatation

A. Representative tracing of the effect of sequential addition of the β-blocker propranolol (Prop 10⁻⁶ mol/L) and nitric oxide synthase inhibitor L-NAME (10⁻⁴ mol/L) on IgG-induced arteriolar dilation. B. Data for four of the subjects. Patients 118 and 236 had a return to baseline tone with combined blockade while 112 and 159 failed to return completely to control. No subject had residual vasoconstriction compared to baseline during combined blockade.