Suli Zhang

Autoantibody against AT1 receptor from preeclamptic patients induces vasoconstriction through angiotensin receptor activation.

Preeclampsia is a serious pathologic complication during pregnancy, and its pathogenesis remains poorly understood. Recent studies have demonstrated that autoantibodies against angiotensin II type 1A receptor (AT1-AA) are present in women with preeclampsia. However, their role in the development of hypertension in preeclamptic patients has never been previously investigated. The present study was designed to determine whether AT1-AA isolated from the sera of preeclamptic patients causes vascular constriction and, if so, to further investigate the cellular receptors that mediate their vasoactivity. Blood samples were collected from 49 pregnant women (preeclampsia = 31, control = 18) and AT1-AA was detected using enzyme-linked immunosorbent assay. Vasoconstrictive effect of purified IgG from the sera of either preeclamptic patients or normal pregnant women was determined in isolated rat thoracic aorta, arteriae cerebri media and coronary artery. Compared with normal pregnant women, frequency of AT1-AA positive samples was markedly increased in preeclamptic patients (80.7 vs. 5.6%, P < 0.01). In isolated thoracic aortic rings, middle cerebral artery and coronary artery segments, AT1-AA induced vasoconstriction in a concentration-dependent fashion (P < 0.01). The vasoconstrictive effect of AT1-AA was completely blocked by losartan, an AT1-receptor antagonist. These data demonstrate that the AT1-AA causes significant vascular constriction in large conduit vessel as well as small resistant vessels though activation of the AT1 receptor. These results suggest that overproduction of AT1-AA is a novel risk factor in pregnant women and may play a causative role in the development of hypertension and vascular injury in preeclamptic patients.

Decreased Autophagy in Rat Heart Induced by Anti-β1-Adrenergic Receptor Autoantibodies Contributes to the Decline in Mitochondrial Membrane Potential

It has been recognized that changes in mitochondrial structure plays a key role in development of cardiac dysfunction, and autophagy has been shown to exert maintenance of mitochondrial homeostasis effects. Our previous study found that anti-β1-adrenergic receptor autoantibodies (β1-AABs) could lead to cardiac dysfunction along with abnormalities in mitochondrial structure. The present study tested the hypothesis that β1-AABs may induce the decline in mitochondrial membrane potential (ΔΨm) by suppression of cardiac autophagy, which contributed to cardiac dysfunction. Male adult rats were randomized to receive a vehicle or peptide corresponding to the second extracellular loop of the β1 adrenergic receptor (β1-AAB group, 0.4 μg/g every two weeks for 12 weeks) and treated with rapamycin (RAPA, an autophagy agonist) at 5 mg/kg/day for two days before detection. At the 4th week, 8th week and 12th week of active immunization, the rats were sacrificed and cardiac function and the levels of cardiac LC3 and Beclin-1 were detected. ΔΨm in cardiac myocytes was determined by myocardial radionuclide imaging technology and JC-1 staining. In the present study, β1-AABs caused cardiac dysfunction, reduced ΔΨm and decreased cardiac autophagy. Treatment with RAPA markedly attenuated β1-AABs-induced cardiac injury evidenced by recovered ΔΨm. Taken together, these results suggested that β1-AABs exerted significant decreased ΔΨm, which may contribute to cardiac dysfunction, most likely by decreasing cardiac autophagy in vivo. Moreover, myocardial radionuclide imaging technology may be needed to assess the risk in developing cardiac dysfunction for the people who have β1-AABs in their blood.

Angiotensin II type 1 receptor autoantibody as a novel regulator of aldosterone independent of preeclampsia.

Background: Preeclamptic women and their infants have significant morbidity and mortality worldwide. Abnormal aldosterone signaling is involved in the pathogenesis of preeclampsia, and the presence of agonistic autoantibodies against the angiotensin II type 1 receptor (AT1-AA) during disease has been observed. The role of AT1-AA in aldosterone generation with or without disease and the long-term impact of AT1-AA circulation in blood remain unclear. Method: We therefore assessed circulating AT1-AA and aldosterone levels in 76 patients with preeclampsia (35 severe and 41 mild), 26 patients with gestational hypertension, and 50 normotensive healthy pregnant women. Results: First, the correlation of AT1-AA levels was confirmed for preeclamptic patients. We report here that all AT1-AA-positive hypertensive pregnant women exhibited decreased aldosterone levels, and early-onset preeclampsia patients with high proteinuria showed an inverse correlation of aldosterone levels with AT1-AA. To study this effect in more detail, we confirmed that AT1-AA decreased aldosterone levels in pregnant rats and then demonstrated that aldosterone levels decreased in response to the chronic administration of AT1-AA into nonpregnant rats. Conclusion: These results suggested that AT1-AA regulates levels of aldosterone, which was tested with cell culture studies, revealing that activation of AT1 receptors by AT1-AA directly led to abnormal aldosterone generation in a time and dose-dependent manner. We present here a mechanism for regulation of aldosterone production: AT1-AA activates AT1 receptors on adrenocortical cells independent of pregnancy, in a time and dose-dependent manner.

Serum agonistic autoantibodies against type-1 angiotensin II receptor titer in patients with epithelial ovarian cancer: a potential role in tumor cell migration and angiogenesis

BackgroundAlthough agonistic autoantibodies against type-1 angiotensin-II receptor (AT1-AA) are frequently detected in women with preeclampsia, the clinical significance of AT1-AA in association with epithelial ovarian cancer (EOC) has not been identified.MethodsIn an attempt to clarify this issue, we measured serum AT1-AA titer from EOC patients (n = 89) and healthy normal subjects (n = 55), correlated AT1-AA titer with EOC stage and grade, and demonstrated the effects of purified AT1-AA on migration of ovarian cancer cells and angiogenesis of chick embryo chorioallantoic membrane.ResultsWe found that the AT1-AA titer was significantly higher in EOC patients compared with healthy control subjects (1.77 ± 0.28 vs. 0.35 ± 0.05, P < 0.01). The positive rate was averaged by 72.1±2.5% in EOC patients and 15.5 ±1.5% in control (P < 0.01). Increased AT1-AA titer in EOC patients was associated with advanced stages and grades of EOC, and positively correlated with level of vascular endothelial growth factor (r = 0.855, P < 0.01). Furthermore, AT1-AA directly stimulated migration of ovarian cancer cells and enhanced microvascular density of chick embryo chorioallantoic membrane. These AT1-AA-mediated effects were significantly blocked either by an autoantibody-neutralizing peptide or an angiotensin II type I receptor antagonist, losartan.ConclusionTaken together, we found that a higher serum AT1-AA titer may be associated with advanced progression of EOC in patients and play an important role in development of EOC by promoting cancer cell migration and angiogenesis. These findings implicate that AT1-AA might be selected as a detectable biomarker and potential therapeutic target in diagnosis and treatment of EOC patients.

Up-regulation of hypoxia-inducible factor-1α enhanced the cardioprotective effects of ischemic postconditioning in hyperlipidemic rats

Hyperlipidemia is an independent risk factor in the development of ischemic heart disease, which can increase myocardial susceptibility to ischemia/reperfusion (I/R) injury. Ischemic postconditioning (PostC) has now been demonstrated as a novel strategy to harness natures protection against myocardial I/R injury in normal conditions. However, the effect of PostC on hyperlipidemic animals remains elusive. It has been shown in our previous study that PostC reduces the myocardial I/R injury, and hypoxia-inducible factor-1α (HIF-1α) may play an important role in the cardioprotective mechanisms of PostC on normal rats. Here, we tested the hypothesis that the cardioprotection of PostC on hyperlipidemic rats is associated with the up-regulated HIF-1α expression. Male Wistar rats were fed with a high-fat diet for 8 weeks, and then randomly divided into five groups: sham, I/R, dimethyloxalylglycine (DMOG) + I/R, PostC, and DMOG + PostC group. The detrimental indices induced by I/R injury included infarct size, plasma creatine kinase (CK) activity and caspase-3 activity. The results showed that PostC could reduce the infarct size, when compared with the I/R group, which was consistent with the significant lower levels of plasma CK activity and caspase-3 activity, and that it increased the expression of HIF-1α in hyperlipidemic rats. When DMOG was given before PostC to up-regulate HIF-1α protein level, the degree of I/R injury was attenuated. In conclusion, these data suggested that the up-regulation of HIF-1α may be one of the cardioprotective mechanisms of PostC against I/R injury in hyperlipidemic rats.

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

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.

The Prognostic Role of Angiotensin II Type 1 Receptor Autoantibody in Non-Gravid Hypertension and Pre-eclampsia: A Meta-analysis and Our Studies.

AbstractAngiotensin II type 1 receptor autoantibody (AT1-AA) is found in patients with non-gravid hypertension or pre-eclampsia, but the relationship is uncertain.The aim of the present study was to assess the association between AT1-AA and high blood pressure using meta-analysis, and to evaluate the prognosis value of AT1-AA for hypertensive diseases.Literature search from PubMed, Embase, and Cochrane databases were conducted using keywords “hypertension” or “pre-eclampsia,” “angiotensin II receptor type 1 autoantibody,” and its aliases from April 1999 to December 2015.Studies evaluating the association between AT1-AA and non-gravid hypertension or pre-eclampsia were included in this analysis. The quality of the eligible studies was assessed based on the Newcastle–Ottawa Scale with some modifications.Two researchers then independently reviewed all included studies and extracted all relevant data. Association between AT1-AA and hypertension was tested with pooled odds ratios (ORs) and 95% confidence intervals (CIs). Finally, we evaluated whether AT1-AA predicted the prognosis of hypertension by using a summary receiver-operating characteristic (ROC) curve and sensitivity analysis.Ten studies were finally included in this meta-analysis. AT1-AA showed more significant association with pre-eclampsia than that with non-gravid hypertension (pooled OR 32.84, 95% CI 17.19–62.74; and pooled OR 4.18, 95% CI 2.20–7.98, respectively). Heterogeneity among studies was also detected probably due to different hypertensive subtypes and AT1-AA measuring methods. Area under summary ROC curve (AUC) of pre-eclampsia was 0.92 (sensitivity 0.76; specificity 0.86). Area under the ROC curve of overall hypertensive diseases or non-gravid hypertension was lower than that of pre-eclampsia (0.86 and 0.72, respectively) with lower sensitivities (0.46 and 0.26, respectively).The major limitation of this analysis was the publication bias due to lack of unpublished data and the language limitation during literature search. Prospective study with large simple size and specific measuring data collection are needed to enhance our findings in the future.Our analysis confirms that elevated AT1-AA in serum is significantly associated with hypertensive disorder, especially pre-eclampsia. AT1-AA may be a valuable indicator for poorer prognosis of patients with pre-eclampsia, and could be used in patients with hypertensive disease for risk evaluation and making individual treatment decision.

Decreased autophagy: a major factor for cardiomyocyte death induced by β1-adrenoceptor autoantibodies

Cardiomyocyte death is one major factor in the development of heart dysfunction, thus, understanding its mechanism may help with the prevention and treatment of this disease. Previously, we reported that anti-β1-adrenergic receptor autoantibodies (β1-AABs) decreased myocardial autophagy, but the role of these in cardiac function and cardiomyocyte death is unclear. We report that rapamycin, an mTOR inhibitor, restored cardiac function in a passively β1-AAB-immunized rat model with decreased cardiac function and myocardial autophagic flux. Next, after upregulating or inhibiting autophagy with Beclin-1 overexpression/rapamycin or RNA interference (RNAi)-mediated expression of Beclin-1/3-methyladenine, β1-AAB-induced autophagy was an initial protective stress response before apoptosis. Then, decreased autophagy contributed to cardiomyocyte death followed by decreases in cardiac function. In conclusion, proper regulation of autophagy may be important for treating patients with β1-AAB-positive heart dysfunction.

Decreased Autophagy Contributes to Myocardial Dysfunction in Rats Subjected to Nonlethal Mechanical Trauma

Autophagy is important in cells for removing damaged organelles, such as mitochondria. Insufficient autophagy plays a critical role in tissue injury and organ dysfunction under a variety of pathological conditions. However, the role of autophagy in nonlethal traumatic cardiac damage remains unclear. The aims of the present study were to investigate whether nonlethal mechanical trauma may result in the change of cardiomyocyte autophagy, and if so, to determine whether the changed myocardial autophagy may contribute to delayed cardiac dysfunction. Male adult rats were subjected to nonlethal traumatic injury, and cardiomyocyte autophagy, cardiac mitochondrial function, and cardiac function in isolated perfused hearts were detected. Direct mechanical traumatic injury was not observed in the heart within 24 h after trauma. However, cardiomyocyte autophagy gradually decreased and reached a minimal level 6 h after trauma. Cardiac mitochondrial dysfunction was observed by cardiac radionuclide imaging 6 h after trauma, and cardiac dysfunction was observed 24 h after trauma in the isolated perfused heart. These were reversed when autophagy was induced by administration of the autophagy inducer rapamycin 30 min before trauma. Our present study demonstrated for the first time that nonlethal traumatic injury caused decreased autophagy, and decreased autophagy may contribute to post-traumatic organ dysfunction. Though our study has some limitations, it strongly suggests that cardiac damage induced by nonlethal mechanical trauma can be detected by noninvasive radionuclide imaging, and induction of autophagy may be a novel strategy for reducing posttrauma multiple organ failure.

Autoantibody against angiotensin AT1 receptor from preeclamptic patients enhances collagen-induced human platelet aggregation

Hypercoagulability, platelet activation, and thrombocytopenia are the chief characteristics of preeclampsia, but their responsible underlying molecular mechanisms remain obscure. Recent studies have demonstrated that the autoantibody against angiotensin II type 1 receptor (AT1-AA) constitutes a novel risk factor for preeclampsia. However, the role of AT1-AA in platelet activation and hypercoagulability in preeclampsia has never been investigated. In the present study, we determined whether AT1-AA promotes platelet aggregation in vitro, and dissected the potential underlying mechanisms. AT1-AA was detected by enzyme-linked immunosorbent assay. After immunoglobulin G fractions purified from the preeclamptic patient positive sera were added to platelets isolated from healthy volunteers, platelet aggregation and intracellular Ca(2+) levels were detected. AT1-AA significantly enhanced in vitro collagen-induced platelet aggregation, an effect blocked by the AT1 receptor antagonist losartan. Additionally, AT1-AA increased and maintained collagen-induced cytosolic calcium concentration throughout the experiment. We demonstrated for the first time that AT1-AA significantly promotes collagen-induced platelet aggregation through angiotensin type 1 receptor activation in vitro, potentially via increased intracellular Ca(2+) concentration, supporting AT1-AA as a potential contributor to the hypercoagulable state of preeclampsia.