A Unifying Hypothesis of the Pathophysiology of Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS): Recognitions from the finding of autoantibodies against ß2-adrenergic receptors.

CLASSIFICATION of receptor systems which are activated by sympathomimetic amines has engaged the attention of many investigators. Ahlquist1–3 has suggested the convenient designations of alpha (α) and beta (β) to distinguish major differences in the responses elicited in various organ systems. Our investigations have provided results which cannot be explained easily on the assumption that the β-type designates a single receptor population4–6. We now report additional studies which show that two distinct receptor types are at present included within this group.

Differentiation of receptor systems activated by sympathomimetic amines

Aims Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a complex multisystem disease. Evidence for disturbed vascular regulation comes from various studies showing cerebral hypoperfusion and orthostatic intolerance. The peripheral endothelial dysfunction (ED) has not been sufficiently investigated in patients with ME/CFS. The aim of the present study was to examine peripheral endothelial function in patients with ME/CFS. Methods and results Thirty-five patients [median age 40 (range 18-70) years, mean body mass index 23.8 ± 4.2 kg/m2 , 31% male] with ME/CFS were studied for peripheral endothelial function assessed by peripheral arterial tonometry (EndoPAT2000). Clinical diagnosis of ME/CFS was based on Canadian Criteria. Nine of these patients with elevated antibodies against β2-adrenergic receptor underwent immunoadsorption, and endothelial function was measured at baseline and 3, 6, and 12 months follow-up. ED was defined by reactive hyperaemia index ≤1.81. Twenty healthy subjects of similar age and body mass index were used as a control group. Peripheral ED was found in 18 of 35 patients (51%) with ME/CFS and in 4 healthy subjects (20%, P Conclusions Peripheral ED is frequent in patients with ME/CFS and associated with disease severity and severity of immune symptoms. As ED is a risk factor for cardiovascular disease, it is important to elucidate if peripheral ED is associated with increased cardiovascular morbidity and mortality in ME/CFS.

https://onlinelibrary.wiley.com/doi/pdf/10.1002/ehf2.12633

Peripheral endothelial dysfunction in myalgic encephalomyelitis/chronic fatigue syndrome

Chronic sustained stimulation of β-adrenoceptor is closely related to cardiac fibrosis which is bad for cardiac function. Growing evidence showed that the high prevalence of β1-adrenoceptor autoantibody (β1-AA) in the sera of patients with various types of cardiovascular diseases decreased cardiac function. In the current study, we demonstrated that β1-AA impaired the cardiac function evaluated by echocardiography and that β1-AA triggered cardiac fibrosis in terms of increased expression of α-smooth muscle actin as the marker of myofibroblast and collagen deposition in a passive β1-AA immunized mice model during 16 weeks. Further, we showed that β1-AA activated β1-AR/cAMP/PKA pathway and promoted proliferation in primary cardiac fibroblasts through specific binding to β1-AR but not to β2-AR. Moreover, β1-AA was also likely to promote proliferation in cardiac fibroblasts through activating p38MAPK and ERK1/2 as p38MAPK inhibitor SB203580 and ERK1/2 inhibitor PD98059 partially reversed the proliferative effect. The persistent activating signalling of PKA and P38MAPK in 1 h induced by β1-AA was associated with lacking agonist-induced desensitization phenomena. The conditioned medium from β1-AA-stimulated cardiac fibroblasts induced cardiomyocyte apoptosis, which indicated that β1-AA changed the secretion of cardiac fibroblasts contributing to cardiac injury. These findings will contribute to our understanding of the pathological mechanisms of β1-AA.

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Proliferation in cardiac fibroblasts induced by β1-adrenoceptor autoantibody and the underlying mechanisms

Thermostabilized G protein-coupled receptors used as antigens for in vivo immunization have resulted in the generation of functional agonistic anti-β1-adrenergic (β1AR) receptor monoclonal antibodies (mAbs). The focus of this study was to examine the pharmacology of these antibodies to evaluate their mechanistic activity at β1AR. Immunization with the β1AR stabilized receptor yielded five stable hybridoma clones, four of which expressed functional IgG, as determined in cell-based assays used to evaluate cAMP stimulation. The antibodies bind diverse epitopes associated with low nanomolar agonist activity at β1AR, and they appeared to show some degree of biased signaling as they were inactive in an assay measuring signaling through β-arrestin. In vitro characterization also verified different antibody-receptor interactions reflecting the different epitopes on the extracellular surface of β1AR to which the mAbs bind. The anti-β1AR mAbs only demonstrated agonist activity when in dimeric antibody format, but not as the monomeric Fab format, suggesting that agonist activation may be mediated through promoting receptor dimerization. Finally, we have also shown that at least one of these antibodies exhibits in vivo functional activity at a therapeutically-relevant dose producing an increase in heart rate consistent with β1AR agonism.

https://www.tandfonline.com/doi/pdf/10.4161/mabs.27226

Monoclonal anti-β1-adrenergic receptor antibodies activate G protein signaling in the absence of β-arrestin recruitment.

Antibodies against the second extracellular loop of β₁-adrenergic receptors induce endothelial dysfunction in conductance and resistance arteries of the Wistar rat.

Circulating autoantibodies directed against the 2nd extracellular loop (EL-2) of β(1)-adrenoceptors (β(1)-AABs) have been detected in the serum of patients with various cardiovascular pathologies. β(1)-AABs induce agonistic, positive inotropic effects via β(1)-adrenoceptors (β(1)ARs). In the mammalian heart, β(1)-AR can exist in 2 distinct activated configurations (the so-called high- and low-affinity states). The aim of the present study was to investigate whether the action of β(1)-AAB is dependent on the affinity state of β(1)AR in isolated ventricular cardiomyocytes of adult Wistar rats. Immunoglobulin G (IgG) containing β(1)-AAB obtained from animals immunized with a peptide corresponding to the EL-2 of human β(1)-AR, caused a dose-dependent increase in cell shortening. Isoproterenol-induced inotropy was significantly reduced in cardiomyocytes that had been preincubated with IgG containing β(1)-AAB and in cardiomyocytes isolated from immunized rats. The negative effects of preincubation with IgG containing β(1)-AAB on the response to isoproterenol was inhibited in the presence of bisoprolol. CGP 12177A and pindolol-induced inotropy was not affected by IgG preincubation or immunization. No detectable inotropic effect of cell shortening was obtained with IgG containing β(1)-AAB in the presence of propranolol and 3-isobutyl-1-methylxanthine. The present study demonstrates that β(1)-AABs have no agonist/antagonist-like effects upon low-affinity state β(1)-ARs. This result indicates that β(1)-AABs recognize and stabilize the high-affinity state, but are unable to stabilize and (or) induce the low-affinity state receptor.

Autoantibodies against cardiac β(1)-adrenoceptor do not affect the low-affinity state β(1)-adrenoceptor-mediated inotropy in rat cardiomyocytes.

In porcine coronary arteries (PCAs), celiprolol, a selective β(1)-adrenoceptors antagonist, induces vasodilatation by an endothelium- and nitric oxide (NO)-dependent pathway. However, the mechanisms of that vascular effect have not been precisely established. β(3)-Adrenoceptors have been shown to be involved in the relaxation per se of various vascular beds, including coronary vessels. Thus, we evaluated (i) the presence of β(3)-adrenoceptors in the PCA and (ii) their role in celiprolol-induced vasodilatation. PCA rings were placed in organ baths and preconstricted with KCl. All experiments were performed in the presence of nadolol (a β(1)/β(2)-adrenoceptor antagonist). Cumulative concentration-response curves to SR 58611A and ICI 215001 (2 β(3)-adrenoceptor agonists) and to celiprolol were constructed. We also used semiquantitative reverse transcription - polymerase chain reaction, which clearly showed the presence of β(3)-adrenoceptor transcripts. SR 58611A, ICI 215001, and celiprolol induced concentration-dependent relaxations in PCA rings. SR 58611A-induced relaxation was almost abolished after removal of endothelium or pretreatment with L-NAME (a NO synthase inhibitor). The vasorelaxations induced by SR 58611A and celiprolol were inhibited in the presence of SR 59230A and L-748337 (2 selective β(3)-adrenoceptor antagonists). We showed (i) that PCAs possess functional β(3)-adrenoceptors mediating endothelium- and NO-dependent relaxation, and (ii) that celiprolol exerts a β(3)-adrenoceptor agonistic activity in this vascular bed.

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Celiprolol induces β3-adrenoceptors-dependent relaxation in isolated porcine coronary arteries

Resume But de l’etude Analyser les modifications de la reactivite vasculaire en reponse a des protocoles d’immunisation par des antigenes correspondants a la seconde boucle extracellulaire des recepteurs adrenergiques (RA) –β3 ou –β1 et 3. Methodes Des rats Lewis ont ete immunises pendant 3 mois avec un peptide correspondant a la seconde boucle extracellulaire des RA–β3 ou –β1 et 3. Les anticorps anti-RA–β (AC–RA–β) produits ont ete titres par Elisa et purifies en utilisant le kit « Proteus Protein G ». Leur fonctionnalite a ensuite ete testee dans des cardiomyocytes ventriculaires isoles chez le lapin. Des anneaux d’aorte et d’artere mesenterique isoles ont ete montes dans des cuves a organes isoles et precontractes a la phenylephrine. Puis, des courbes de relaxation ont ete realisees. Resultats Le SR58611A (agoniste des RA–β3) (10 nM) et les AC–RA–β3 purifies (25 μg/mL) induisent une diminution du pourcentage de raccourcissement cellulaire (respectivement −39,6 ± 4,4 % [n = 11] et −18,5 ± 3,9 % [n = 10]) des cardiomyocytes isoles. Cette diminution est inhibee lorsque les cardiomyocytes sont preincubes avec le L–748337 (1 μM), un antagoniste selectif des RA–β3 (p  Conclusion Ces resultats montrent pour la premiere fois que les AC–RA–β3 exercent un effet agoniste β3– adrenergique. Ils n’auraient pas d’action pathogene vasculaire mais compenseraient la dysfonction endotheliale provoquee par les AC–RA–β1.

Effets cardiovasculaires des auto-anticorps anti-récepteurs béta1 et béta3-adrénergiques chez le rat Lewis

Resume Objectif Evaluer l’effet de l’immunisation active, a l’aide d’un peptide correspondant a la deuxieme boucle extracellulaire des recepteurs adrenergiques (RA) β 1 , sur la reactivite de l’aorte isolee de rat Wistar. Methodes Des rats Wistar âges de neuf semaines ont ete activement immunises pendant trois mois avec un peptide correspondant a la deuxieme boucle extracellulaire des RA β 1  humains. Les immunoglobulines G (IgG) specifiques ont ete caracterisees par Elisa et quantifiees par la methode BCA. Leur fonctionnalite a ensuite ete testee dans des cardiomyocytes ventriculaires isoles (CVI). Des anneaux d’aorte de rats temoins ou immunises ont ete montes dans des cuves a organes isoles. Ensuite, des courbes de contraction a la phenylephrine (1 nM a 300 μM) ou de relaxation a l’acetylcholine (1 nM a 100 μM) et l’isoprenaline (1 nM a 30 μM) ont ete etablies. Resultats Dans les CVI, les IgG anti-RA β 1 (10 ou 25 μg/mL) induisent une augmentation du raccourcissement cellulaire dependante de la concentration. L’effet inotrope positif induit par l’isoprenaline (10 nM et 1 μM) est fortement reduit dans les CVI incubes en presence de 25 μg/mL d’IgG pendant trois heures. Dans l’aorte de rats immunises, la contraction a la phenylephrine et la relaxation a l’acetylcholine sont significativement alterees. En revanche, la relaxation induite par l’isoprenaline est conservee. Conclusions Cette etude confirme que les anticorps diriges contre la deuxieme boucle extracellulaire des RA β 1  induisent un effet inotrope positif dans les cardiomyocytes isoles de rat adulte. Elle montre aussi, pour la premiere fois, que les rats immunises presentent une alteration de la fonction endotheliale, mais une conservation de la relaxation adrenergique β.

M.A. Abdelkrim

Papers

Antibodies against the second extracellular loop of β₁-adrenergic receptors induce endothelial dysfunction in conductance and resistance arteries of the Wistar rat.

Autoantibodies against cardiac β(1)-adrenoceptor do not affect the low-affinity state β(1)-adrenoceptor-mediated inotropy in rat cardiomyocytes.

Celiprolol induces β3-adrenoceptors-dependent relaxation in isolated porcine coronary arteries

Effets cardiovasculaires des auto-anticorps anti-récepteurs béta1 et béta3-adrénergiques chez le rat Lewis

[Antibodies against the second extracellular loop of beta1-adrenergic receptor induce aortic endothelial dysfunction in Wistar rat].