"Multisystem Inflammatory Syndrome in Children"-Like Disease after COVID-19 Vaccination (MIS-V) with Potential Significance of Functional Active Autoantibodies Targeting G-Protein-Coupled Receptors (GPCR-fAAb) for Pathophysiology and Therapy.

BACKGROUND: An infection with SARS-CoV-2 can trigger a systemic disorder by pathological autoimmune processes. A certain type of this dysregulation is known as Multisystemic inflammatory syndrome in children (MIS-C). However, similar symptoms may occur and have been described as Multisystemic inflammatory syndrome after SARS-CoV-2 Vaccination (MIS-V) following vaccination against SARS-CoV-2. We report the case of a 12-year-old boy who was identified with MIS-C symptoms without previous SARS-CoV-2 infection after receiving two doses of the Pfizer-BioNTech COVID-19 vaccine approximately one month prior to the onset of symptoms. He showed polyserositis, severe gastrointestinal symptoms and, consequently, a manifestation of a multiorgan failure. IgG antibodies against spike proteins of SARS-CoV-2 were detected, indicating a successful vaccination, while SARS-CoV-2 Nucleocapsid protein antibodies and SARS-CoV-2 PCR were not detected. Several functional, active autoantibodies against G-protein-coupled receptors (GPCR-fAAb), previously associated with Long COVID disease, were detected in a cardiomyocyte bioassay. Immunosuppression with steroids was initiated. Due to side effects, treatment with steroids and later interleukin 1 receptor antagonists had to be terminated. Instead, immunoadsorption was performed and continued with tacrolimus and mycophenolic acid therapy, leading to improvement and discharge after 79 days. GPCR-fAAb decreased during therapy and remained negative after clinical curing and under continued immunosuppressive therapy with tacrolimus and mycophenolic acid. Follow-up of the patient showed him in good condition after one year. CONCLUSIONS: Infection with SARS-CoV-2 shows a broad and severe variety of symptoms, partly due to autoimmune dysregulation, which, in some instances, can lead to multiorgan failure. Despite its rarity, post-vaccine MIS-C-like disease may develop into a serious condition triggered by autoimmune dysregulation. The evidence of circulating GPCR-fAAb and their disappearance after therapy suggests a link of GPCR-fAAb to the clinical manifestations. Thus, we hypothesize a potential role of GPCR-fAAb in pathophysiology and their potential importance for the therapy of MIS-C or MIS-V. However, this observation needs further investigation to prove a causative correlation.

Animals Experimentally Infected with SARS-CoV-2 Generate Functional Autoantibodies against G-Protein-Coupled Receptors.

(1) Background: SARS-CoV-2 infection has been linked to diverse clinical manifestations in humans, including cardiovascular complications. Functional autoantibodies targeting G-protein-coupled receptors have emerged as potential contributors to these effects. This study sought to investigate the production and activity of functional autoantibodies targeting G-protein-coupled receptors after SARS-CoV-2 infection of selected animal species. (2) Methods: The presence of functional autoantibodies such as 2-adrenoceptor, angiotensin II AT1 receptor, muscarinic M2 receptor, and angiotensin 1-7 MAS receptor was assessed in cattle and ferrets experimentally infected with SARS-CoV-2. Bioassays were conducted to evaluate the positive or negative chronotropic responses induced by these autoantibodies. Further experiments identified the extracellular domains to which the functional autoantibodies bind, and receptor antagonists were employed to block the induced responses. (3) Results: Only two out of six cattle that were inoculated with SARS-CoV-2 displayed viral replication and tested positive for functional autoantibodies against G-protein-coupled receptors. These functional autoantibodies specifically recognized ß2-adrenoceptor, angiotensin II AT1 receptor, muscarinic M2 receptor, and angiotensin 1-7 MAS receptor and induced distinct positive and negative chronotropic effects in the bioassay. Infected ferrets generated functional autoantibodies against ß2-adrenoceptor and muscarinic M2 receptor and presented bioactivity similar to that in cattle. (4) Conclusions: This study uncovers functional autoantibodies targeting G-protein-coupled receptors in cattle and ferrets post-SARS-CoV-2 infection, with implications for cardiovascular function.

B2 cells contribute to hypertension and natural killer cell activation possibly via AT1-AA in response to placental ischemia.

Preeclampsia, new onset hypertension during pregnancy, is associated with activated T helper cells (Th) and B cells secreting agonistic autoantibodies against the angiotensin II type 1 receptor (AT1-AA). The reduced uterine perfusion pressure (RUPP) model of placental ischemia recapitulates these characteristics. We have shown that Th-B cell communication contributes to AT1-AA and symptoms of preeclampsia in the RUPP rat. B2 cells are classical B cells that communicate with Th cells and are then transformed into memory B cells. We hypothesize that B2 cells cause hypertension, natural killer (NK) cell activation, and complement activation during pregnancy through the production of AT1-AA. To test this hypothesis, total splenic B cells and B2 cells were isolated from normal pregnant (NP) or RUPP rats on gestational day (GD)19 and adoptively transferred into GD12 NP rats. A group of recipient rats was treated with a specific inhibitor peptide of AT1-AA. On GD19, mean arterial pressure was measured, tissues were collected, activated NK cells were measured by flow cytometry, and AT1-AA was measured by cardiomyocyte assay. NP recipients of RUPP B cells or RUPP B2 cells had increased mean arterial pressure, AT1-AA, and circulating activated NK cells compared with recipients of NP B cells. Hypertension in NP recipients of RUPP B cells or RUPP B2 was attenuated with AT1-AA blockade. This study demonstrates that B cells and B2 cells from RUPP rats cause hypertension and increased AT1-AA and NK cell activation in response to placental ischemia during pregnancy.NEW & NOTEWORTHY This study demonstrates that placental ischemia-stimulated B2 cells induce hypertension and circulating natural killer cell activation and angiotensin II type 1 receptor production in normal pregnant rats.

Circulating angiotensin II type I receptor - autoantibodies in diabetic pregnancies.

Pregnant women with either pre-existing or gestational diabetes mellitus are at increased risk of preeclampsia as well as future cardiovascular disease. The renin-angiotensin system is dysregulated in both diabetes mellitus and preeclampsia. In preeclampsia, maternal levels of circulating agonistic autoantibodies against the angiotensin II Type I receptor (AT1-AAs) are increased. Circulating AT1-AAs are thought to contribute to both the pathophysiology of preeclampsia and the increased risk of future cardiovascular disease. Studies exploring AT1-AA in diabetes outside pregnancy suggest their potential for both metabolic and cardiovascular pathogenicity. No studies have investigated AT1-AAs in diabetic pregnancies. We hypothesized elevated maternal circulating AT1-AA levels in pregnancies complicated by any type of diabetes mellitus. Third-trimester maternal serum from 39 women (controls: n = 10; type 1 diabetes: n = 9; type 2 diabetes: n = 10; gestational diabetes=10) were analyzed for AT1-AA using an established bioassay method. Circulating AT1-AAs were present in 70% (7/10) of the controls and 83% (24/29) of the diabetes group (P = 0.399). Presence of AT1-AA was correlated to hsCRP levels (P = 0.036), but neither with maternal circulating angiogenic factors (soluble fms-like tyrosine kinase-1 and placental growth factor), nor with maternal or fetal characteristics indicative of metabolic disease or placental dysfunction. Our study is the first to demonstrate presence of circulating AT1-AAs in pregnant women with any type of diabetes. Our findings suggest AT1-AAs presence in pregnancy independently of placental dysfunction, nuancing the current view on their pathogenicity. Whether AT1-AAs per se contribute to increased risk of adverse pregnancy outcomes and future cardiovascular disease remains currently unanswered.

Post-COVID-19 Syndrome: Retinal Microcirculation as a Potential Marker for Chronic Fatigue.

Post-COVID-19 syndrome (PCS) is characterized by persisting sequelae after infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). PCS can affect patients with all COVID-19 disease severities. As previous studies have revealed impaired blood flow as a provoking factor triggering PCS, it was the aim of the present study to investigate the potential association between self-reported chronic fatigue and retinal microcirculation in patients with PCS, potentially indicating an objective biomarker. A prospective study was performed, including 201 subjects: 173 patients with PCS and 28 controls. Retinal microcirculation was visualized by OCT angiography (OCT-A) and quantified using the Erlangen-Angio-Tool as macula and peripapillary vessel density (VD). Chronic fatigue (CF) was assessed according to the variables of Bell's score, age and gender. VDs in the superficial vascular plexus (SVP), intermediate capillary plexus (ICP) and deep capillary plexus (DCP) were analyzed, considering the repetitions (12 times). Seropositivity for autoantibodies targeting G protein-coupled receptors (GPCR-AAbs) was determined by an established cardiomyocyte bioassay. Taking account of the repetitions, a mixed model was performed to detect possible differences in the least square means between the different groups included in the analysis. An age effect in relation to VD was observed between patients and controls (p < 0.0001). Gender analysis showed that women with PCS showed lower VD levels in the SVP compared to male patients (p = 0.0015). The PCS patients showed significantly lower VDs in the ICP as compared to the controls (p = 0.0001 (CI: 0.32; 1)). Moreover, considering PCS patients, the mixed model revealed a significant difference between those with chronic fatigue (CF) and those without CF with respect to VDs in the SVP (p = 0.0033 (CI: −4.5; −0.92)). The model included variables of age, gender and Bell's score, representing a subjective marker for CF. Consequently, retinal microcirculation might serve as an objective biomarker in subjectively reported chronic fatigue in patients with PCS.

Functional antibodies against G-protein coupled receptors in Beagle dogs infected with two different strains of Trypanosoma cruzi.

The interaction of the anti-beta1-adrenergic receptor autoantibodies (ß1ARAb) and the anti-muscarinic M2 receptor autoantibodies (M2RAb) with cardiac neurotransmitter receptors were identified in human chronic Chagas cardiomyopathy (CCC) related to the ECG and dysautonomia disturbances. Dogs are considered gold model to the study of Trypanosoma cruzi infection due the clinical similarities with CCC. This study aims to evaluate whether anti-ß1ARAb, anti-ß2ARAb, and anti-muscarinic M2RAb are generated in Beagle dogs infected by T. cruzi using Y and Berenice-78 strains of T. cruzi. Animals were infected with 4.0 x 103 bloodstream trypomastigotes/kg of body weight and, after 25 months of infection, blood sample was collected, and serum stored at -80°C. Dog serum was treated by ammonium sulphate precipitation and the IgG antibodies isolated and added to the beating neonatal rats' cardiomyocytes. All T. cruzi-infected dogs developed agonistic ß1ARAb, ß2ARAb, and M2RAb. Animals infected by Berenice strain presented less ß2ARAb and M2RAb activities than dogs infected by Y strain of the parasite. In cardiomyocytes culture, the antibodies recognized an epitope on the second extracellular loop of the receptors which were similar to findings in human Chagas disease. There was no detection of antibody against G protein-coupled receptor in serum from uninfected dogs. In conclusion, both Y and Berenice-78 strains of T. cruzi induced dog antibodies, whose targets located in the second extracellular loop of the adrenergic and muscarinic receptors were similar to those observed in individuals with CCC. Therefore, our findings highlight dogs as a promisor model to investigate pathogenic roles of functional Ab against G-protein coupled receptors.

Glaucoma and Alzheimer: Neurodegenerative disorders show an adrenergic dysbalance.

Glaucoma disease is characterized by an increased intraocular pressure (IOP), glaucomatous alterations of the optic disc and corresponding visual field defects. Even lowering the main risk factor IOP until an individual target level does not prevent this neurodegenerative disorder from proceeding. Several autoimmune mechanisms were discovered, partly showing a functionality. One of these autoimmune phenomena targets the ß2-adrenergic receptor (ß2-AR; i.e. agonistic autoantibodies; ß2-agAAb) and is linked to an elevated IOP and an impaired retinal microcirculation. As neurodegenerative disorder, Alzheimer's Disease (AD) is postulated to share a common molecular mechanism with glaucoma. In the present study we investigated autoimmune phenomena targeting the ß2-AR in patients with AD. Sera of the patients were analyzed in a rat cardiomyocyte bioassay for the presence of functional autoantibodies against ß2-AR. In addition, different species of amyloid beta (Aß) monomers were tested (Aß1-14, Aß10-25, Aß10-37 Aß1-40, Aß1-42, Aß28-40, and Aß-[Pyr]3-43). Our results demonstrate that none of the short-chain Aß (Aß1-14, Aß10-25, or Aß28-40) showed any agonistic or inhibitory effect on ß2-AR. Contrary, long-chain Aß-[Pyr]3-43, representing a major neurogenic plaque component, exerted an activation that after blocking by the ß2-AR antagonist ICI118.551, could be identified as that the effect was realized via the ß2-AR. Moreover, the long chain Aß1-40, Aß1-42, and Aß10-37, yet not the short-chain Aß peptides prevented the clenbuterol induced desensitization of the ß2-AR. In addition, we identified functional autoantibodies in the sera of AD patients, activating the ß2-AR, like the ß2-agAAb found in patients with glaucoma. As autoimmune mechanisms were reportedly involved in the pathogenesis of glaucoma and Alzheimer's Disease, we postulate that overstimulation of the ß2-AR pathway can induce an adrenergic overdrive, that may play an important role in the multifactorial interplay of neurodegenerative disorders.

Long COVID: Association of Functional Autoantibodies against G-Protein-Coupled Receptors with an Impaired Retinal Microcirculation.

Long COVID (LC) describes the clinical phenotype of symptoms after infection with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Diagnostic and therapeutic options are limited, as the pathomechanism of LC is elusive. As the number of acute SARS-CoV-2 infections was and is large, LC will be a challenge for the healthcare system. Previous studies revealed an impaired blood flow, the formation of microclots, and autoimmune mechanisms as potential factors in this complex interplay. Since functionally active autoantibodies against G-protein-coupled receptors (GPCR-AAbs) were observed in patients after SARS-CoV-2 infection, this study aimed to correlate the appearance of GPCR-AAbs with capillary microcirculation. The seropositivity of GPCR-AAbs was measured by an established cardiomyocyte bioassay in 42 patients with LC and 6 controls. Retinal microcirculation was measured by OCT-angiography and quantified as macula and peripapillary vessel density (VD) by the Erlangen-Angio Tool. A statistical analysis yielded impaired VD in patients with LC compared to the controls, which was accentuated in female persons. A significant decrease in macula and peripapillary VD for AAbs targeting adrenergic ß2-receptor, MAS-receptor angiotensin-II-type-1 receptor, and adrenergic α1-receptor were observed. The present study might suggest that a seropositivity of GPCR-AAbs can be linked to an impaired retinal capillary microcirculation, potentially mirroring the systemic microcirculation with consecutive clinical symptoms.

Induced antibodies directed to the angiotensin receptor type 1 provoke skin and lung inflammation, dermal fibrosis and act species overarching.

OBJECTIVE: To determine contributions and functions of autoantibodies (Abs) directed to the angiotensin receptor type 1 (AT1R), which are suggested to be involved in the pathogenesis of AT1R Abs-related diseases such as systemic sclerosis (SSc). METHODS: C57BL/6J mice were immunised with membrane-embedded human AT1R or empty membrane as control. Mice deficient for CD4+ or CD8+ T cells and B cells were immunised with membrane-embedded AT1R or an AT1R peptide proposed to be a dominant T cell epitope. A monoclonal (m)AT1R Ab was generated by hybridoma technique and transferred into C57BL/6J and AT1Ra/b knockout mice. The induced phenotype was examined by histology, immunohistochemistry, immunofluorescence, apoptosis assay and ELISA. In vitro, Abs responses towards AT1R were measured in cells of different origins and species. RESULTS: AT1R-immunised mice developed perivascular skin and lung inflammation, lymphocytic alveolitis, weak lung endothelial apoptosis and skin fibrosis accompanied by Smad2/3 signalling, not present in controls or mice deficient for CD4+ T and B cells. The AT1R peptide 149-172 provoked lung inflammation. Application of the mAT1R Ab induced skin and lung inflammation, not observed in AT1Ra/b knockout mice. In vitro, AT1R Abs activated rat cardiomyocytes and human monocytes, enhanced angiotensin II-mediated AT1R activation in AT1R-transfected HEK293 cells via AT1R binding and mAT1R Ab-activated monocytes mediated the induction of profibrotic markers in dermal fibroblasts. CONCLUSION: Our immunisation strategy successfully induced AT1R Abs, contributing to inflammation and, possibly, to fibrosis via activation of AT1R. Therefore, AT1R Abs are valuable targets for future therapies of SSc and other AT1R Ab-related diseases.

C-Reactive Protein (CRP) Blocks the Desensitization of Agonistic Stimulated G Protein Coupled Receptors (GPCRs) in Neonatal Rat Cardiomyocytes.

Recently, C-reactive protein (CRP) was shown to affect intracellular calcium signaling and blood pressure in vitro and in vivo, respectively. The aim of the present study was to further investigate if a direct effect on G-protein coupled receptor (GPCR) signaling by CRP can be observed by using CRP in combination with different GPCR agonists on spontaneously beating cultured neonatal rat cardiomyocytes. All used agonists (isoprenaline, clenbuterol, phenylephrine, angiotensin II and endothelin 1) affected the beat rate of cardiomyocytes significantly and after washing them out and re-stimulation the cells developed a pronounced desensitization of the corresponding receptors. CRP did not affect the basal beating-rate nor the initial increase/decrease in beat-rate triggered by different agonists. However, CRP co-incubated cells did not exhibit desensitization of the respective GPCRs after the stimulation with the different agonists. This lack of desensitization was independent of the GPCR type, but it was dependent on the CRP concentration. Therefore, CRP interferes with the desensitization of GPCRs and has to be considered as a novel regulator of adrenergic, angiotensin-1 and endothelin receptors.

Placental CD4+ T cells from preeclamptic patients cause autoantibodies to the angiotensin II type I receptor and hypertension in a pregnant rat model of preeclampsia.

Preeclampsia (PE) is a hypertensive disorder of pregnancy associated with activated CD4+ T cells and autoantibodies to angiotensin II type 1 receptor (AT1-AA). We have previously shown that CD4+ T cells isolated from women with PE cause hypertension, increased tumor necrosis factor alpha (TNF-α), endothelin-1, and soluble fms-like tyrosine kinase-1 (sFlt-1) when injected into pregnant nude-athymic rats compared to CD4+ T cells from normal pregnant (NP) women. However, the role of PE CD4+ T cells to cause AT1-AA as a mechanism of hypertension is not known. Aim: Our goal was to determine if PE CD4+ T cells stimulate AT1-AA in pregnant nude-athymic rats. CD4+ T cells were isolated from human NP and PE placentasand injected into nude-athymic rats on gestational day (GD) 12. In order to examine the role of the PE CD4+ T cells to stimulate B cell secretion of AT1-AA, a subset of the rats receiving PE CD4+ T cells were treated with rituximab on GD 14 or anti-CD40 ligand (anti-CD40L) on GD 12. On GD 19, mean arterial pressure (MAP) and tissues were obtained MAP [114 ± 1 mmHg (n = 9)] and AT1-AA [19.8 ± 0.9 beats per minute (bpm, n = 4)] were increased in NP nude + PE CD4+ T cells compared to NP nude + NP CD4+ T cells [98 ± 2 mmHg (n = 7, P < 0.05) and 1.3 ± 0.9 bpm (n = 5, P < 0.05)]. Rituximab (103 ± 2 mmHg, n = 3, P < 0.05) and anti-CD40L (102 ± 1 mmHg, n = 3, P < 0.05) lowered MAP compared to NP nude + PE CD4+ T cells. Circulating a proliferation-inducing ligand (APRIL) and placental angiotensin-converting enzyme 2 (ACE-2) activity was increased in response to PE CD4+ T cells. These results show that placental CD4+ T cells play an important role in the pathophysiology of PE, by activating B cells secreting AT1-AA to cause hypertension during pregnancy.

Case Report: Neutralization of Autoantibodies Targeting G-Protein-Coupled Receptors Improves Capillary Impairment and Fatigue Symptoms After COVID-19 Infection.

Clinical features of Coronavirus disease 2019 (COVID-19) are caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). Acute infection management is a substantial healthcare issue, and the development of long-Covid syndrome (LCS) is extremely challenging for patients and physicians. It is associated with a variety of characteristics as impaired capillary microcirculation, chronic fatigue syndrome (CFS), proinflammatory cytokines, and functional autoantibodies targeting G-protein-coupled receptors (GPCR-AAbs). Here, we present a case report of successful healing of LCS with BC 007 (Berlin Cures, Berlin, Germany), a DNA aptamer drug with a high affinity to GPCR-AAbs that neutralizes these AAbs. A patient with a documented history of glaucoma, recovered from mild COVID-19, but still suffered from CFS, loss of taste, and impaired capillary microcirculation in the macula and peripapillary region. He was positively tested for various targeting GPCR-AAbs. Within 48 h after a single BC 007 treatment, GPCR-AAbs were functionally inactivated and remained inactive during the observation period of 4 weeks. This observation was accompanied by constant improvement of the fatigue symptoms of the patient, taste, and retinal capillary microcirculation. Therefore, the removal of GPCR-AAb might ameliorate the characteristics of the LCD, such as capillary impairment, loss of taste, and CFS.

Maintenance therapy with subcutaneous immunoglobulin in a patient with immune-mediated neuropathic postural tachycardia syndrome.

AIMS: We describe the disease course of a 35-year-old female with an autoimmune mediated neuropathic postural tachycardia syndrome (PoTS), who responded to immunoglobulin therapy and stabilized on maintenance therapy with subcutaneous immunoglobulin (SCIg). METHODS: We provide longitudinal data of clinical scores, tilt-table results and antibody titers. RESULTS: Initial treatment with intravenous immunoglobulin caused infusion-related side-effects whereas SCIg was well tolerated and improved clinical symptoms and quality of life. Clinical improvement correlated with the reduction of serum antibody titers 22 months after first infusion. CONCLUSIONS: These findings suggest that autoimmune-mediated neuropathic PoTS can be treated sufficiently with IVIg whereas SCIg minimizes side-effects.

Inhibitory and Agonistic Autoantibodies Directed Against the ß2-Adrenergic Receptor in Pseudoexfoliation Syndrome and Glaucoma.

Pseudoexfoliation syndrome (PEXS) and glaucoma (PEXG) are assumed to be caused by a generalized elastosis leading to the accumulation of PEX material in ocular as well as in extraocular tissues. The exact pathophysiology of PEXS is still elusive. PEXG, the most common type of secondary open-angle glaucoma (OAG), is characterized by large peaks of intraocular pressure (IOP) with a progressive loss of the visual field. Agonistic autoantibodies (agAAbs) against the ß2-adrenergic receptor (AR) have been shown to be present in sera of patients with primary and secondary OAG and ocular hypertension and are seemingly linked to IOP. In the present study, we investigated the autoantibodies directed against the ß2-AR in sera of patients with PEXS and PEXG. We recruited 15, 10, and 15 patients with PEXG, PEXS, and primary OAG, respectively. Ten healthy individuals served as controls. All patients underwent standard ophthalmological examination with Octopus G1 perimetry. agAAbs prepared from serum samples were analyzed in a rat cardiomyocyte-based bioassay for the presence of agAAbs. We identified the interacting loop of the ß2-AR and the immunoglobulin G (IgG) subclasses using synthetic peptides corresponding to the extracellular loops of the receptors and enzyme-linked immunosorbent assay, respectively. None of the controls were ß2-agAAb-positive (0.2 ± 0.5 U). No ß2-agAAbs (0.2 ± 0.4 U), but inhibitory ß2-AAbs were observed in 80% of the patients that partially blocked the drug-induced ß2-adrenergic stimulation; 5.8 ± 1.7 U vs. 11.1 ± 0.9 U for clenbuterol in the absence and the presence of sera from patients with PEXS, respectively. Epitope analyses identified the third extracellular loop of the ß2-AR as the target of the inhibitory ß2-AAbs, being of IgG3 subtype in PEXS patients. In contrast, patients with PEXG showed ß2-agAAbs (5.6 ± 0.9 U), but no inhibitory ones. The ß2-agAAbs levels of patients with PEXG and primary OAG patients (3.9 ± 2.8 U; p > 0.05) were at a similar level. In two cases of PEXG, the ß2-agAAbs exert synergistic effects with clenbuterol. The activity increased from 11.5 ± 0.3 (clenbuterol only) to 16.3 ± 0.9 U. As autoimmune mechanisms were reportedly involved in the pathogenesis of glaucoma, agonistic and inhibitory ß2-AAbs seem to be a part of this multifactorial interplay.

Agonistic ß2-Adrenergic Receptor Autoantibodies Characterize the Aqueous Humor of Patients With Primary and Secondary Open-Angle Glaucoma.

Purpose: Agonistic ß2-adrenergic receptor autoantibodies (ß2-agAAbs) were recently observed in sera of patients with ocular hypertension (OHT), primary (POAG), and secondary open-angle glaucoma (SOAG), yet not in healthy controls (HCs). It was the aim of the present study to investigate the presence of ß2-agAAb in aqueous humor (AH) samples of OAG patients and to correlate these with the corresponding ß2-agAAb serum data. Material and Methods: Thirty-nine patients (21 male, 18 female) were recruited from the Department of Ophthalmology, University of Erlangen-Nürnberg: twenty-one POAG, 18 SOAG. Aqueous humor samples were collected during minimal invasive glaucoma surgery. Serum and AH samples were analyzed for ß2-agAAb by a bioassay quantifying the beating rate of cultured cardiomyocyte (cut-off: 2 U). Results: Thirty-six of 39 (92.3%) and 34 of 39 (87.2%) of OAG patients showed a ß2-agAAb in their sera and AH samples, respectively. All ß2-agAAb AH-positive OAG patients were also seropositive. We also observed a ß2-agAAb seropositivity in 95 and 89% of patients with POAG and SOAG, respectively. Beta2-agAAbs were seen in 86% (POAG) and 78% (SOAG) of AH samples. The ß2-agAAb adrenergic activity was increased in the AH of patients with POAG (6.5 ± 1.5 U) when compared with those with SOAG (4.1 ± 1.1 U; p = 0.004). Serum ß2-agAAb adrenergic activity did not differ between the cohorts [POAG (4.5 ± 1.5 U); SOAG (4.6 ± 2.1 U; p=0.458)]. No correlation of the beating rates were observed between serum and AH samples for group and subgroup analyses. Conclusion: The detection of ß2-agAAb in systemic and local circulations supports the hypothesis of a direct functional impact of these agAAbs on ocular G-protein coupled receptors. The high prevalence of ß2-agAAb in serum and AH samples of patients with POAG or SOAG suggests a common role of these AAbs in the etiopathogenesis of glaucoma, independent of open-angle glaucoma subtype.

Agonistic autoantibodies against ß2-adrenergic receptor influence retinal microcirculation in glaucoma suspects and patients.

PURPOSE: Agonistic ß2-adrenergic receptor autoantibodies (ß2-agAAb) have been observed in sera of patients with ocular hypertension and open-angle glaucoma (OAG). They target the ß2-receptors on trabecular meshwork, ciliary body and pericytes (Junemann et al. 2018; Hohberger et al. 2019). In addition to their influence on the intraocular pressure, an association to retinal microcirculation is discussed. This study aimed to investigate foveal avascular zone (FAZ) characteristics by en face OCT angiography (OCT-A) in glaucoma suspects and its relationship to ß2-agAAb status in patients with OAG. MATERIAL AND METHODS: Thirty-four patients (28 OAG, 6 glaucoma suspects) underwent standardized, clinical examination including sensory testing as white-on-white perimetry (Octopus G1, mean defect, MD) and structural measures as retinal nerve fibre layer (RNFL) thickness, neuroretinal rim width (BMO-MRW), retinal ganglion cell layer (RGCL) thickness, and inner nuclear layer (INL) thickness with high-resolution OCT. FAZ characteristics were measured by OCT-A scans of superficial vascular plexus (SVP), intermediate capillary plexus (ICP), and deep capillary plexus (DCP). FAZ-R was calculated (area FAZ (SVP)/area FAZ (ICP)). Using cardiomyocyte bioassays we analysed serum samples for the presence of ß2-agAAb. RESULTS: (I) Total mean FAZ area [mm2]: 0.34±0.16 (SVP), 0.24±0.12 (ICP), and 0.49±0.24 (DCP); mean FAZ-R 1.58±0.94. No correlation was seen for FAZ-R with MD, RNFL, BMO-MRW, RGCL thickness and INL thickness (p>0.05). (II) ß2-agAAb have been observed in 91% patients and showed no correlation with MD, RNFL, BMO-MRW, RGCL thickness and INL thickness (p>0.05). (III) FAZ-R correlated significantly with the ß2-agAAb-induced increase of the beat rate of cardiomyocyte (p = 0.028). CONCLUSION: FAZ characteristics did not correlate with any glaucoma associated functional and morphometric follow-up parameter in the present cohort. However, level of ß2-agAAb showed a significantly correlation with FAZ-ratio. We conclude that ß2-agAAb might be a novel biomarker in glaucoma pathogenesis showing association to FAZ-ratio with OCT-A.

Functional autoantibodies against G-protein coupled receptors in patients with persistent Long-COVID-19 symptoms.

Impairment of health after overcoming the acute phase of COVID-19 is being observed more and more frequently. Here different symptoms of neurological and/or cardiological origin have been reported. With symptoms, which are very similar to the ones reported but are not caused by SARS-CoV-2, the occurrence of functionally active autoantibodies (fAABs) targeting G-protein coupled receptors (GPCR-fAABs) has been discussed to be involved. We, therefore investigated, whether GPCR-fAABs are detectable in 31 patients suffering from different Long-COVID-19 symptoms after recovery from the acute phase of the disease. The spectrum of symptoms was mostly of neurological origin (29/31 patients), including post-COVID-19 fatigue, alopecia, attention deficit, tremor and others. Combined neurological and cardiovascular disorders were reported in 17 of the 31 patients. Two recovered COVID-19 patients were free of follow-up symptoms. All 31 former COVID-19 patients had between 2 and 7 different GPCR-fAABs that acted as receptor agonists. Some of those GPCR-fAABs activate their target receptors which cause a positive chronotropic effect in neonatal rat cardiomyocytes, the read-out in the test system for their detection (bioassay for GPCR-fAAB detection). Other GPCR-fAABs, in opposite, cause a negative chronotropic effect on those cells. The positive chronotropic GPCR-fAABs identified in the blood of Long-COVID patients targeted the ß2-adrenoceptor (ß2-fAAB), the α1-adrenoceptor (α1-fAAB), the angiotensin II AT1-receptor (AT1-fAAB), and the nociceptin-like opioid receptor (NOC-fAAB). The negative chronotropic GPCR-fAABs identified targeted the muscarinic M2-receptor (M2-fAAB), the MAS-receptor (MAS-fAAB), and the ETA-receptor (ETA-fAAB). It was analysed which of the extracellular receptor loops was targeted by the autoantibodies.

Activating autoantibodies against G protein-coupled receptors in narcolepsy type 1.

STUDY OBJECTIVES: Narcolepsy type 1 is a rare hypersomnia of central origin, which is caused by loss of hypothalamic neurons that produce the neuropeptides hypocretin-1 and -2. Hypocretin-containing nerve terminals are found in areas known to play a central role in autonomic control and in pain signaling. Cholinergic M2 receptors are found in brain areas involved with the occurrence of hallucinations and cataplexy. In addition to classical symptoms of narcolepsy, the patients suffer frequently from autonomic dysfunction, chronic pain, and hypnagogic/hypnopompic hallucinations. We aimed to test whether narcolepsy type 1 patients have autoantibodies against autonomic ß2 adrenergic receptor, M2 muscarinic receptors, or nociception receptors. METHODS: We tested the serum of ten narcolepsy type 1 patients (five female) for activating ß2 adrenergic receptor autoantibodies, M2 muscarinic receptor autoantibodies, and nociception receptor autoantibodies. RESULTS: Ten of ten patients were positive for muscarinic M2 receptor autoantibodies (P < 0.001), 9/10 were positive for autoantibodies against nociception receptors (P < 0.001), and 5/10 were positive for ß2 adrenergic receptor autoantibodies (P < 0.001). CONCLUSIONS: Narcolepsy type 1 patients harbored activating autoantibodies against M2 muscarinic receptors, nociception receptors, and ß2 adrenergic receptors. M2 receptor autoantibodies may be related to the occurrence of cataplexy and, moreover, hallucinations in narcolepsy since they are found in the same brain areas that are involved with these symptoms. The occurrence of nociception receptor autoantibodies strengthens the association between narcolepsy type 1 and pain. The connection between narcolepsy type 1, autonomic complaints, and the presumed cardiovascular morbidity might be associated with the occurrence of ß2 adrenergic receptor autoantibodies. On the other hand, the presence of the autoantibodies may be secondary to the destruction of the hypocretin pathways.

Autoantibodies directed against α1-adrenergic receptor and endothelin receptor A in patients with prostate cancer.

BACKGROUND: For prostate cancer, signaling pathways induced by over-boarding stimulation of G-protein coupled receptors (GPCR) such as the endothelin, α1- and ß-adrenergic, muscarinic and angiotensin 1 receptors were accused to support the carcinogenesis. However, excessive receptor stimulation by physiological receptor ligands is minimized by a control system that induces receptor sensitization and down-regulation. This system is missing when so-called "functional autoantibodies" bind to the GPCR (GPCR-AAB). If GPCR-AAB were found in patients with prostate cancer, uncontrolled GPCR stimulation could make these autoantibodies an additional supporter in prostate cancer. METHODS: Using the bioassay of spontaneously beating cultured rat neonatal cardiomyocytes, GPCR-AAB were identified, quantified and characterized in the serum of 25 patients (aged 56-78 years, median 70 years) with prostate cancer compared to 10 male patients (aged 48-82 years, median 64) with urinary stone disorders (controls). RESULTS: Of the cancer patients, 24 (96%) and 17 (68%), respectively, carried autoantibodies directed against the α1-adrenergic receptor (α1-AAB) and endothelin receptor A (ETA-AAB). No patient was negative for both GPCR-AAB. In contrast, ETA-AAB and α1-AAB were absent in all (100%) and 9 (90%) of the 10 control patients, respectively. While α1-AAB targeted a specific epitope of the first extracellular loop of the α1-adrenergic receptor subtype A, an epitope of the second extracellular loop of the ETA receptor was identified as a target of ETA-AAB. As demonstrated in vitro, the functional activity of both autoantibodies found in prostate cancer can be neutralized by the aptamer BC007. CONCLUSIONS: We hypothesized that α1-AAB and ETA-AAB, which are highly present in prostate cancer patients, could by their functional activity support carcinogenesis by excessive receptor stimulation. The in vitro demonstrated neutralization of α1- and ETA-AAB by the aptamer BC007 could open the door to complement the treatments already available for prostate cancer.