Suppression of β1-adrenoceptor autoantibodies is involved in the antiarrhythmic effects of omega-3 fatty acids in male and female hypertensive rats

The arrhythmogenic potential of β1-adrenoceptor autoantibodies (β1-AA), as well as antiarrhythmic properties of omega-3 in heart diseases, have been reported while underlying mechanisms are poorly understood. We aimed to test our hypothesis that omega-3 (eicosapentaenoic acid-EPA, docosahexaenoic acid-DHA) may inhibit matrix metalloproteinase (MMP-2) activity to prevent cleavage of β1-AR and formation of β1-AA resulting in attenuation of pro-arrhythmic connexin-43 (Cx43) and protein kinase C (PKC) signaling in the diseased heart. We have demonstrated that the appearance and increase of β1-AA in blood serum of male and female 12-month-old spontaneously hypertensive rats (SHR) was associated with an increase of inducible ventricular fibrillation (VF) comparing to normotensive controls. In contrast, supplementation of hypertensive rats with omega-3 for two months suppressed β1-AA levels and reduced incidence of VF. Suppression of β1-AA was accompanied by a decrease of elevated myocardial MMP-2 activity, preservation of cardiac cell membrane integrity and Cx43 topology. Moreover, omega-3 abrogated decline in expression of total Cx43 as well as its phosphorylated forms at serine 368 along with PKC-ε, while decreased pro-fibrotic PKC-δ levels in hypertensive rat heart regardless the sex. The implication of MMP-2 in the action of omega-3 was also demonstrated in cultured cardiomyocytes in which desensitization of β1-AR due to permanent activation of β1-AR with isoproterenol was prevented by MMP-2 inhibitor or EPA. Collectively, these data support the notion that omega-3 via suppression of β1-AA mechanistically controlled by MMP-2 may attenuate abnormal of Cx43 and PKC signaling; thus, abolish arrhythmia substrate and protect rats with an advanced stage of hypertension from malignant arrhythmias

Chronic Chagas disease: from basics to laboratory medicine

Chagas disease, caused by Trypanosoma cruzi infection, is ranked as the most serious parasitic disease in Latin America and has huge potential to become a worldwide problem, due to increasing migration, and international tourism, as well as infectant transfer by blood contact and transfusion, intrauterine transfer, and organ transplantation. Nearly 30% of chronically-infected patients become symptomatic, often with a latency of 10-30 years, developing life-threatening complications. Of those, nearly 90% develop Chagas heart disease, while the others manifest gastrointestinal disease and neuronal disorders. Besides interrupting the infection cycle and chemotherapeutic infectant elimination, starting therapy early in symptomatic patients is important for counteracting the disease. This would be essentially supported by optimized patient management, involving risk assessment, early diagnosis and monitoring of the disease and its treatment. From economic and logistic viewpoints, the tools of laboratory medicine should be especially able to guarantee this. After summarizing the basics of chronic Chagas disease, such as the epidemiological data, the pathogenetic mechanisms thought to drive symptomatic Chagas disease and also treatment options, we present tools of laboratory medicine that address patient diagnosis, risk assessment for becoming symptomatic and guidance, focusing on autoantibody estimation for risk assessment and heart marker measurement for patient guidance. In addition, increases in levels of inflammation and oxidative stress markers in chronic Chagas disease are discussed

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 ((f)AABs) targeting G-protein coupled receptors (GPCR-(f)AABs) has been discussed to be involved. We, therefore investigated, whether GPCR-(f)AABs 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-(f)AABs that acted as receptor agonists. Some of those GPCR-(f)AABs 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-(f)AAB detection). Other GPCR-(f)AABs, in opposite, cause a negative chronotropic effect on those cells. The positive chronotropic GPCR-(f)AABs identified in the blood of Long-COVID patients targeted the β(2)-adrenoceptor (β(2)-(f)AAB), the α1-adrenoceptor (α(1)-(f)AAB), the angiotensin II AT1-receptor (AT1-(f)AAB), and the nociceptin-like opioid receptor (NOC-(f)AAB). The negative chronotropic GPCR-(f)AABs identified targeted the muscarinic M(2)-receptor (M(2)-(f)AAB), the MAS-receptor (MAS-(f)AAB), and the ETA-receptor (ETA-(f)AAB). It was analysed which of the extracellular receptor loops was targeted by the autoantibodies

Rodenticide residues in non-target small mammal species and their occurrence in owl pellets

OBJECTIVES: Distinguishing the patterns of autoantibodies (AAB) against G-protein-coupled receptors in Chagas' cardiomyopathy and megacolon and the discovery of such a pattern in patients who are as yet asymptomatic could help to identify patients at high risk of developing the life-threatening complications of Chagas' disease. BACKGROUND: Such AAB against receptors as beta 1 (beta1-AAB), beta 2 (beta2-AAB), and muscarinergic 2 (M2-AAB) are thought to be involved in the pathogenesis of Chagas' cardiomyopathy and megacolon, the predominant manifestations of Chagas' disease, which is the most serious parasitic disease in Latin America. METHODS: Beta1-AAB, beta2-AAB, and M2-AAB were measured in the serum of asymptomatic Chagas' patients and in those with cardiomyopathy and/or megacolon. RESULTS: Nearly all Chagas' patients with cardiomyopathy and/or megacolon had AAB. Predominance of beta1-AAB combined with M2-AAB in Chagas' cardiomyopathy and beta2-AAB with M2-AAB in megacolon was found. Such patterns were also found in 34% of the asymptomatic patients, of whom 85% possessed a beta1-AAB level typical for Chagas' cardiomyopathy. CONCLUSIONS: The percentage of asymptomatic Chagas' patients who had a specific AAB pattern and had a beta1-AAB level above a defined cutoff point mirrors very well the epidemiological situation, which showed that clinical manifestations develop in nearly 30% of Chagas' patients and cardiomyopathy in nearly 90% of them. We hypothesize that beta1-, beta2-, and M2-AAB measurement might be a useful tool for risk assessment in the indeterminate state of Chagas' disease to select patients for earlier involvement in care programs. However, prospective studies are needed to further evaluate this hypothesis

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 (β1ARAb) 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-β1ARAb, anti-β2ARAb, 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 β1ARAb, β2ARAb, and M2RAb. Animals infected by Berenice strain presented less β2ARAb 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

Potential Relevance of α1-Adrenergic Receptor Autoantibodies in Refractory Hypertension

-AAB might have a mechanistic role and could represent a therapeutic target. in cardiomyocytes and induce mesentery artery segment contraction.-AAB in hypertensive patients, and the notion of immunity as a possible cause of hypertension

Agonistic β2-adrenergic receptor autoantibodies characterize the aqueous humor of patients with primary and secondary open-angle glaucoma

PURPOSE: Agonistic β2-adrenergic receptor autoantibodies (β2-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 β2-agAAb in aqueous humor (AH) samples of OAG patients and to correlate these with the corresponding β2-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 β2-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 β2-agAAb in their sera and AH samples, respectively. All β2-agAAb AH-positive OAG patients were also seropositive. We also observed a β2-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 β2-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 β2-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 β2-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 β2-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

RNA interference therapeutics targeting angiotensinogen ameliorate preeclamptic phenotype in rodent models

Preeclampsia, with the hallmark features of new-onset hypertension and proteinuria after 20 weeks of gestation, is a major cause of fetal and maternal morbidity and mortality. Studies have demonstrated a role for the renin-angiotensin system (RAS) in its pathogenesis; however, small-molecule RAS blockers are contraindicated because of fetal toxicity. We evaluated whether siRNA targeting maternal hepatic angiotensinogen (Agt) could ameliorate symptoms of preeclampsia without adverse placental or fetal effects in 2 rodent models. The first model used a cross of females expressing human Agt with males expressing human renin, resulting in upregulation of the circulating and uteroplacental RAS. The second model induced ischemia/reperfusion injury and subsequent local and systemic inflammation by surgically reducing placental blood flow midgestation (reduced uterine perfusion pressure [RUPP]). These models featured hypertension, proteinuria, and fetal growth restriction, with altered biomarkers. siRNA treatment ameliorated the preeclamptic phenotype in both models, reduced blood pressure, and improved intrauterine growth restriction, with no observed deleterious effects on the fetus. Treatment also improved the angiogenic balance and proteinuria in the transgenic model, and it reduced angiotensin receptor activating antibodies in both. Thus, an RNAi therapeutic targeting Agt ameliorated the clinical sequelae and improved fetal outcomes in 2 rodent models of preeclampsia

The quality control theory of aging

The quality control (QC) theory of aging is based on the concept that aging is the result of a reduction in QC of cellular systems designed to maintain lifelong homeostasis. Four QC systems associated with aging are 1) inadequate protein processing in a distressed endoplasmic reticulum (ER); 2) histone deacetylase (HDAC) processing of genomic histones and gene silencing; 3) suppressed AMPK nutrient sensing with inefficient energy utilization and excessive fat accumulation; and 4) beta-adrenergic receptor (BAR) signaling and environmental and emotional stress. Reprogramming these systems to maintain efficiency and prevent aging would be a rational strategy for increased lifespan and improved health. The QC theory can be tested with a pharmacological approach using three well-known and safe, FDA-approved drugs: 1) phenyl butyric acid, a chemical chaperone that enhances ER function and is also an HDAC inhibitor, 2) metformin, which activates AMPK and is used to treat type 2 diabetes, and 3) propranolol, a beta blocker which inhibits BAR signaling and is used to treat hypertension and anxiety. A critical aspect of the QC theory, then, is that aging is associated with multiple cellular systems that can be targeted with drug combinations more effectively than with single drugs. But more importantly, these drug combinations will effectively prevent, delay, or reverse chronic diseases of aging that impose such a tremendous health burden on our society

Blockade of CD40 ligand for intercellular communication reduces hypertension, placental oxidative stress, and AT1-AA in response to adoptive transfer of CD4(+) T lymphocytes from RUPP rats

Preeclampsia (PE) is associated with altered immune activation during pregnancy. We have previously shown that adoptive transfer of CD4+ T cells from the reduced uterine perfusion pressure (RUPP) rat model of PE increases blood pressure, oxidative stress (ROS), and inflammation in normal pregnant recipient rats. The objective of this study was to determine if blockade of communication via the CD40-CD40 ligand (CD40L) interaction between placental ischemia-induced CD4+ T cells with endogenous normal pregnant (NP) cells would improve pathophysiology that was previously observed in NP recipient rats of RUPP CD4+ T cells. Splenic CD4+ T lymphocytes were magnetically separated, incubated with 2.5 {mu}g/ml anti-CD40 ligand ({alpha}CD40L) overnight, and transferred into NP rats on day 12 of gestation (NP+RUPP CD4+ T+anti-CD40L). On day 19 of gestation, blood pressure (MAP), blood, and tissues were collected. MAP was 99 +/- 2 in NP (n = 13), 116 +/- 4 in NP+RUPP CD4+ T cells (n = 7; P < 0.01); MAP only increased to 104 +/- 2 in NP+RUPP CD4+ T cells+CD40L (n = 24) (P < 0.05 vs. NP+RUPP CD4+ T cells). Mechanisms of hypertension in response to RUPP CD4+ T cells include endothelin-1 (ET-1), ROS, and angiotensin II type I receptor (AT1-AA) were analyzed. Inhibition of CD40L binding reduced placental ET-1 to 2.3-fold above NP rats and normalized placental ROS from 318.6 +/- 89 in NP+RUPP CD4+ T cells (P < 0.05) to 118.7 +/- 24 in NP+RUPP CD4+ T+anti-CD40L (P < 0.05). AT1-AA was also normalized with inhibition of CD40L. These data suggest that placental ischemia-induced T-cell communication via the CD40L is one important mechanism leading to much of the pathophysiology of PE