AT1 Expression Research Articles

High-cholesterol diet during pregnancy induces maternal vascular dysfunction in mice: potential role for oxidized LDL-induced LOX-1 and AT1 receptor activation.

The lectin-like oxidized low-density-lipoprotein (oxLDL) receptor-1 (LOX-1) has been shown to induce angiotensin II (AngII) type 1 receptor (AT1) activation, contributing to vascular dysfunction. Preeclampsia is a pregnancy complication characterized by vascular dysfunction and increased LOX-1 and AT1 activation; however, whether LOX-1 and AT1 activity contributes to vascular dysfunction in preeclampsia is unknown. We hypothesized that increased oxLDL levels during pregnancy lead to LOX-1 activation and subsequent AT1 activation, resulting in vascular dysfunction. Pregnant wild-type (WT) and transgenic LOX-1 overexpressing (LOX-1tg) mice were fed a control diet (CD) or high-cholesterol diet (HCD, to impair vascular function) between gestational day (GD) 13.5-GD18.5. On GD18.5, AngII-induced vasoconstriction and methylcholine (MCh)-induced endothelium-dependent vasodilation responses were assessed in aortas and uterine arteries. HCD decreased fetal weight and increased circulating oxLDL/cholesterol levels in WT, but not in LOX-1tg mice. HCD did not alter AngII responsiveness or AT1 expression in both vascular beds; however, AngII responsiveness and AT1 expression were lower in aortas from LOX-1tg compared with WT mice. In aortas from WT-CD mice, acute oxLDL exposure induced AT1-mediated vasoconstriction via LOX-1. HCD impaired endothelium-dependent vasodilation and increased superoxide levels in WT aortas, but not uterine arteries. Moreover, in WT-CD mice oxLDL decreased MCh sensitivity in both vascular beds, partially via LOX-1. In summary, HCD impaired pregnancy outcomes and vascular function, and oxLDL-induced LOX-1 activation may contribute to vascular dysfunction via AT1. Our study suggests that LOX-1 could be a potential target to prevent adverse outcomes associated with vascular dysfunction in preeclampsia.

Angiotensin AT1 and AT2 receptor heteromer expression in the hemilesioned rat model of Parkinson\u2019s disease that increases with levodopa-induced dyskinesia

Background/aimsThe renin-angiotensin system (RAS) is altered in Parkinson’s disease (PD), a disease due to substantia nigra neurodegeneration and whose dopamine-replacement therapy, using the precursor levodopa, leads to dyskinesias as the main side effect. Angiotensin AT1 and AT2 receptors, mainly known for their role in regulating water homeostasis and blood pressure and able to form heterodimers (AT1/2Hets), are present in the central nervous system. We assessed the functionality and expression of AT1/2Hets in Parkinson disease (PD).MethodsImmunocytochemistry was used to analyze the colocalization between angiotensin receptors; bioluminescence resonance energy transfer was used to detect AT1/2Hets. Calcium and cAMP determination, MAPK activation, and label-free assays were performed to characterize signaling in homologous and heterologous systems. Proximity ligation assays were used to quantify receptor expression in mouse primary cultures and in rat striatal sections.ResultsWe confirmed that AT1 and AT2 receptors form AT1/2Hets that are expressed in cells of the central nervous system. AT1/2Hets are novel functional units with particular signaling properties. Importantly, the coactivation of the two receptors in the heteromer reduces the signaling output of angiotensin. Remarkably, AT1/2Hets that are expressed in both striatal neurons and microglia make possible that candesartan, the antagonist of AT1, increases the effect of AT2 receptor agonists. In addition, the level of striatal expression increased in the unilateral 6-OH-dopamine lesioned rat PD model and was markedly higher in parkinsonian-like animals that did not become dyskinetic upon levodopa chronic administration if compared with expression in those that became dyskinetic.ConclusionThe results indicate that boosting the action of neuroprotective AT2 receptors using an AT1 receptor antagonist constitutes a promising therapeutic strategy in PD.

Effect of ischemic postconditioning on cell apoptosis and expression of relevant genes in non-culprit coronary arteries

This study was performed to determine the effect of ischemic postconditioning on cell apoptosis and angiotensin II receptor type 1 (AT1), connexin 43 (Cx43), and β-tubulin mRNA expression in non-culprit...

Cardioprotection Generated by Aerobic Exercise Training is Not Related to the Proliferation of Cardiomyocytes and Angiotensin-(1-7) Levels in the Hearts of Rats with Supravalvar Aortic Stenosis

The beneficial effect of aerobic exercise training (ET) on cardiac remodeling caused by supravalvar aortic stenosis (AS) has been demonstrated in experimental studies; however, the mechanisms responsible for improving cardiac function are not entirely understood. We evaluated whether ET-generated cardioprotection in pressure-overloaded rats is dependent on cardiomyocyte proliferation, increased angiotensin-(1-7) (Ang-1-7) levels, and its receptor in the myocardium. Eighteen weeks after ascending AS surgery, Wistar rats were randomly assigned to four groups: sedentary control (C-Sed), exercised control (C-Ex), sedentary aortic stenosis (AS-Sed) and exercised aortic stenosis (AS-Ex) groups. The moderate treadmill exercise protocol was performed for ten weeks. The functional capacity was assessed by treadmill exercise testing. Cardiac structure and function were evaluated by echocardiogram. Cardiomyocyte proliferation was evaluated by flow cytometry. Expression of cell cycle regulatory genes as CCND2, AURKB, CDK1, and MEIS1 was verified by RT-qPCR. Cardiac and plasma angiotensin I (Ang I), angiotensin II (Ang II), and Ang-(1-7) levels were analyzed by high-performance liquid chromatography (HPLC). The angiotensin-converting enzyme (ACE) activity was assessed by the fluorometric method and protein expression of AT1 and Mas receptors by Western blot. The AS-Ex group showed reduced left ventricular wall relative thickness and improved ejection fraction; also, it showed decreased gene expression of myocyte cell cycle regulators, ACE, Ang I, Ang II and Ang II/Ang-(1-7) ratio levels compared to AS-Sed group. However, ET did not induce alterations in Ang-(1-7) and cardiac Mas receptor expression and myocyte proliferation. Aerobic exercise training improves systolic function regardless of myocyte proliferation and Ang-(1-7)/Mas receptor levels. However, the ET negatively modulates the vasoconstrictor/hypertrophic axis (ACE/Ang II) and decreases the expression of negative regulatory genes of the cell cycle in cardiomyocytes of rats with supravalvular aortic stenosis.

All-trans retinoic acid attenuates the progression of Ang II-induced abdominal aortic aneurysms in ApoE\u2212/\u2212mice

BackgroundTo determine whether all-trans retinoic acid (ATRA) can influence the development of Angiotensin II (Ang II) induced experimental abdominal aortic aneurysms (AAAs).MethodsApolipoprotein E knock-out (ApoE−/−) mice were randomly assigned to 4 groups. Mice in the AAA and ATRA groups underwent continuous subcutaneous Ang II infusion for 28 days to induce AAA, while the Sham and Control groups were infused with saline. Systolic blood pressure was measured by the tail-cuff technique. The Control and ATRA groups received ATRA treatment. Aortic tissue samples were obtained at 28 days after surgery and evaluated by aortic diameter measurement, Western blotting, immunohistochemistry, and hematoxylin-eosin (H&E) and Verhoeff-Van Gieson (EVG) staining.ResultsThe abdominal aortic diameter was significantly reduced in the ATRA group compared with the AAA group (3 of 12 (25%) vs 9 of 12 (75%), P < 0.05), and the ATRA group exhibited reduced blood pressure on days 7, 14, and 28. Low expression of angiotensin II receptor type 1 (AT1), matrix metalloproteinase 2 (MMP2), and matrix metalloproteinase 9 (MMP9) and EVG staining revealed a significant reduction in the disruption of elastic fibers in the abdominal aortic tissue of the ATRA group compared to the AAA group. Western blot analysis indicated that protein levels of retinoic acid receptor α (RARα), MMP2, MMP9, and AT1 were dramatically affected by ATRA treatment.ConclusionsIn conclusion, ATRA attenuates the progression of Ang II-induced AAAs, possibly by downregulating MMP2, MMP9, and AT-1 expression.

Long-term effects of early overfeeding and food restriction during puberty on cardiac remodeling in adult rats.

Nutritional disorders during the perinatal period cause cardiometabolic dysfunction, which is observable in the early overfeeding (EO) experimental model. Therefore, severe caloric restriction has the potential of affecting homeostasis through the same epigenetic mechanisms, and its effects need elucidation. This work aims to determine the impact of food restriction (FR) during puberty in early overfed obese and non-obese animals in adult life. Three days after delivery (PN3), Wistar rats were separated into two groups: normal litter (NL; 9 pups) and small litter (SL; 3 pups). At PN30, some offspring were subjected to FR (50%) until PN60, or maintained with free access to standard chow. NL and SL animals submitted to food restriction (NLFR and SLFR groups) were kept in recovery with free access to standard chow from PN60 until PN120. Body weight and food intake were monitored throughout the experimental period. At PN120 cardiovascular parameters were analyzed and the animals were euthanized for sample collection. SLNF and SLFR offspring were overweight and had increased adiposity. Differences in blood pressure were observed only between obese and non-obese animals. Obese and FR animals have cardiac remodeling showing cardiomyocyte hypertrophy and the presence of interstitial and perivascular fibrosis. FR animals also show increased expression of AT1 and AT2 receptors and of total ERK and p-ERK. The present study showed that EO leads to the obese phenotype and cardiovascular disruptions. Interestingly, we demonstrated that severe FR during puberty leads to cardiac remodeling.

SAT-550 Changes in Albuminuria Precede Dietary Sodium-Dependent Changes in BP During Aging - a Longitudinal Study

Background: Hypertension (HT) is a well-established independent risk factor for adverse cardiovascular and renal (CVR) outcomes and a high salt (HS) diet is the main cause for high blood pressure (BP). Despite extensive research focusing on HT, surprisingly there are no longitudinal studies assessing the long-term effects of HS. Aims: This study aimed (1) to evaluate the timing of onset for changes in CVR health during long-term sodium loading and (2) to assess whether salt restriction can prevent these effects. Methods: C57BL/6 mice were randomized to HS, moderate (NS) or low (LS) salt diet and followed longitudinally for 50 weeks (wks). BP, urinary albumin/creatinine ratio (AC), plasma aldosterone (PA) and renin activity (PRA) were assessed monthly. At the end of the study, renal artery resistance and left ventricular (LV) parameters were measured by ultrasound and echocardiogram. Renal AT1 expression (Western Blot) and activity (IHQ) were quantified. Results: At the beginning of the study, there were no differences in BP and AC between the three dietary groups. Relative to wk 1, BP (mmHg) in the HS group was higher in wk 21 (131±1.7 vs. 115±3.0, p= 0.05). Sodium restriction delayed this increase: SBP was higher in wk 41 in the NS group compared to the wk 1 (128±3.4 vs. 115±6.4, p= 0.05) but did not reach significance in the LS group until the end of the study. Similarly, relative to wk 1, AC (µg/mg) only in the HS group reached significantly higher levels in wk 17 (44±4.2, p<0.05). Again, sodium restriction delayed the occurrence of renal damage. AC reached significance in wks 25 and 41 for NS and LS (35±1.1 and 42±2.6 respectively, p<0.05 vs. baseline). Interestingly, the changes in AC always preceded the changes in BP, irrespective of diet. PA and PRA were appropriately activated by dietary salt restriction and suppressed by aging. The aging-induced suppression appeared stronger for PA than for PRA in the HS group only. Long-term sodium loading (HS) induced increased renal resistance, which was prevented in the LS but not in the NS group. Relative to HS, the LV mass index and cardiac output were lower in the NS and LS groups (p<0.05). LV volume indices and ejection fraction did not differ between groups. Renal AT1 protein expression and activation status (IHQ) were decreased in the sodium restricted group. Conclusions: Our study showed that long-term exposure to HS induced a progressive increase in BP and AC in mice. Importantly, these changes were delayed by long-term reduction in sodium intake. Interestingly, changes in AC preceded those in BP, irrespective of diet. Cardiac parameters suggest a sodium-induced eccentric cardiac hypertrophy in the older age, which was prevented by sodium restriction. One possible mechanism behind these effects is the overactivation of the AT1 receptor pathway.

Role of Lectin-like Oxidized LDL Receptor-1 and Syncytiotrophoblast Extracellular Vesicles in the Vascular Reactivity of Mouse Uterine Arteries During Pregnancy

Vascular complications in pregnancy (e.g. preeclampsia) are a major source of maternal and foetal morbidity and mortality, and may be due to excessive release of placental syncytiotrophoblast-derived extracellular vesicles (STBEVs) into the maternal circulation. Increased activity of the multi-ligand scavenger receptor Lectin-like Oxidized LDL Receptor-1 (LOX-1) is associated with vascular dysfunction, and LOX-1 has been shown to interact with angiotensin II receptor type 1 (AT1). We hypothesized that STBEVs contribute to vascular dysfunction via LOX-1 and AT1 receptors during pregnancy. Uterine arteries from late pregnant wildtype and LOX-1 overexpressing mice were incubated overnight with or without STBEVs and vascular function was assessed using wire myography. STBEV-incubation decreased angiotensin II responsiveness only in wildtype mice, which coincided with decreased AT1 contribution and expression. Thus, STBEVs reduced angiotensin II responsiveness in normal pregnancy, but not in conditions of increased LOX-1 expression, suggesting that STBEVs (via LOX-1) play a role in normal adaptations to pregnancy. Oxidized LDL (a LOX-1 ligand) increased angiotensin II-induced vasoconstriction in STBEV-incubated arteries from both mouse strains, suggesting that the LOX-1 pathway may be involved in complicated pregnancies with elevated STBEVs and oxidized LDL levels (such as preeclampsia). These data increase our understanding of vascular complications during pregnancy.

Perinatal nicotine exposure alters miR‐181a‐mediated signaling leading to gender‐dependent development of cardiac ischemia‐sensitive phenotype in adult male offspring

Back groundCigarette smoking/nicotine exposure during pregnancy is one of the major perinatal insults resulting in an increased risk of cardiovascular diseases in offspring. However, the fetal nicotine exposure‐induced programming of adult cardiovascular dysfunction remains largely elusive. The present study tested the hypothesis that perinatal nicotine exposure caused gender‐dependent increase in heart vulnerability to ischemia‐reperfusion injury and cardiac dysfunction in adult male rat offspring through up‐regulation of miRNA‐181a (miR‐181a)‐mediated signaling pathway.MethodsNicotine or saline was administered to pregnant rats via subcutaneous osmotic minipumps from gestational day 4 until postnatal day 10. The adult (~ 3 month old) offspring were treated with miR‐181a antisense (LNA‐miR‐181a) (10 mg/kg × 2 every 5 day, i.p.) or saline. After 10 days of treatment, the rats underwent the cardiac ischemia/reperfusion (I/R) procedure and echocardiography analysis, and the left ventricle (LV) tissues were isolated for molecular biological studies.ResultsPerinatal nicotine exposure enhanced I/R‐induced cardiac infraction and dysfunction in adult male but not in female offspring. The miR‐181a levels in LV tissues were significantly higher in the nicotine‐treated group than the saline control group. In addition, nicotine exposure decreased global DNA methylation levels but enhanced AT1 and AT2 receptor protein expression, and ROS production in LV tissues in the male offspring. Furthermore, the nicotine exposure up‐regulated autophagy‐related proteins (Atg‐5, beclin‐1, LC3 II) expressions as compared to controls in the male offspring. The most importance, treatment with LNA‐miR‐181a reversed nicotine‐mediated I/R‐induced cardiac infarction and dysfunction in male offspring, which was associated with an elimination of the nicotine’s effects on miR‐181a expression, DNA methylation, AT1R/AT2R protein, ROS, and autophagy‐related proteins expressions in the male offspring.ConclusionOur data demonstrated that perinatal nicotine exposure could cause a gender‐dependent fetal programing of adult heart ischemia‐sensitive phenotype. Our findings further suggest that miR‐181a plays a casual role in nicotine‐mediated up‐regulation of oxidative stress, renin‐angiotensin system (RAS) and autophagic flux, which may contribute to the development of ischemia‐sensitive phenotype in adult male rats.Support or Funding InformationSupported in part by the NIH grants: R01HL135623, R03DA041492, and R01HD088039; TRDRP grants: 29IR0437 and T30FT0936

Spontaneous Pulmonary Hypertension Associated With Systemic Sclerosis in P-Selectin Glycoprotein Ligand 1-Deficient Mice.

ObjectivePulmonary arterial hypertension (PAH), one of the major complications of systemic sclerosis (SSc), is a rare disease with unknown etiopathogenesis and noncurative treatments. As mice deficient in P‐selectin glycoprotein ligand 1 (PSGL‐1) develop a spontaneous SSc‐like syndrome, we undertook this study to analyze whether they develop PAH and to examine the molecular mechanisms involved.MethodsDoppler echocardiography was used to estimate pulmonary pressure, immunohistochemistry was used to assess vascular remodeling, and myography of dissected pulmonary artery rings was used to analyze vascular reactivity. Angiotensin II (Ang II) levels were quantified by enzyme‐linked immunosorbent assay, and Western blotting was used to measure Ang II type 1 receptor (AT1R), AT2R, endothelial cell nitric oxide synthase (eNOS), and phosphorylated eNOS expression in lung lysates. Flow cytometry allowed us to determine cytokine production by immune cells and NO production by endothelial cells. In all cases, there were 4–8 mice per experimental group.ResultsPSGL‐1−/− mice showed lung vessel wall remodeling and a reduced mean ± SD expression of pulmonary AT2R (expression ratio [relative to β‐actin] in female mice age >18 months: wild‐type mice 0.799 ± 0.508 versus knockout mice 0.346 ± 0.229). With aging, female PSGL‐1−/− mice had impaired up‐regulation of estrogen receptor α (ERα) and developed lung vascular endothelial dysfunction coinciding with an increase in mean ± SEM pulmonary Ang II levels (wild‐type 48.70 ± 5.13 pg/gm lung tissue versus knockout 78.02 ± 28.09 pg/gm lung tissue) and a decrease in eNOS phosphorylation, leading to reduced endothelial NO production. These events led to a reduction in the pulmonary artery acceleration time:ejection time ratio in 33% of aged female PSGL‐1−/− mice, indicating pulmonary hypertension. Importantly, we found expanded populations of interferon‐γ–producing PSGL‐1−/− T cells and B cells and a reduced presence of regulatory T cells.ConclusionThe absence of PSGL‐1 induces a reduction in Treg cells, NO production, and ERα expression and causes an increase in Ang II in the lungs of female mice, favoring the development of PAH.

Baoyuan decoction ameliorates apoptosis via AT1-CARP signaling pathway in H9C2 cells and heart failure post-acute myocardial infarction rats

Ethnopharmacological relevancePrevious studies have approved that Baoyuan decoction (BYD) exerted remarkable cardioprotective effects on heart failure (HF) due to its anti-apoptotic properties. As a novel biomarker and target of HF, Cardiac ankyrin repeat protein (CARP) can exacerbate apoptosis via activation by angiotensin type 1 receptor (AT1) and subsequently deteriorate heart function. Transcriptome results in our previous study indicated BYD was beneficial to HF post-acute myocardial infarction (AMI) with a promising effect on CARP. However, the mechanism remains to be validated. Aim of the studyThis study aims to elucidate whether BYD ameliorates apoptosis to protect against HF via AT1-CARP signaling pathway. Materials and methodsLeft anterior descending ligation was applied to induce an HF rat model, Ang Ⅱ-stimulated H9C2 cells apoptotic model and overexpression of Ankrd1/CARP H9C2 cells were established to clarify the effects and potential mechanism of BYD. Ethanol extracts of BYD (0.64; 1.28; 2.57 g/kg) were orally administered for four weeks and Fosinopril (4.67 mg/kg) was selected as a positive group in vivo. In vitro, BYD (400, 600, 800 μg/ml) or RNH6270 (an inhibitor of AT1, 1 μM) was co-cultured with Ang Ⅱ stimulation for 48 h in H9C2 cells. Overexpression of Ankrd1/CARP was conducted by transient transfection with H9C2 cells to further confirm the exact mechanism. Finally, to define the active ingredients of anti-cardiomyocyte apoptosis in BYD, we furtherly used the Ang Ⅱ-induced cardiomyocyte apoptosis model to evaluate the effects. ResultsEchocardiography and TUNEL results showed that BYD in different doses remarkably improved heart function and inhibited apoptosis in vivo. Further study demonstrated that AT1 and CARP expressions in cardiac tissue were suppressed by BYD, accompanied with upregulation of B cell lymphoma-2 (Bcl-2) and downregulation of several pro-apoptotic molecules, including p53, Bcl-2 Associated X Protein (Bax) and Cleaved caspase 3. In parallel with the vivo experiment, in vitro research indicated BYD dramatically reduced the apoptotic cells and regulated expressions of critical apoptosis-related molecules mediated through downregulation of AT1 and CARP simultaneously which were consistent with the results in vivo experiment. Transiently transfected CARP over-expression further confirmed that BYD could suppress severe cardiomyocytes apoptosis induced by overexpression of CARP. Especially, the active ingredients of BYD including Astragaloside IV, Ginsenoside Rg3, Rb1, Rc and Re showed significantly anti-apoptosis effects. ConclusionBYD improves cardiac function and protects against cardiomyocytes injury by inhibiting apoptosis via regulating the AT1-CARP signaling pathway.

Angiotensin type 2 receptors: Role in aging and neuroinflammation in the substantia nigra

Overactivity of the angiotensin-type-1 receptor (AT1)/NADPH-oxidase axis enhances aging processes, neuroinflammation and neurodegeneration. The role of AT2 receptors in the above-mentioned AT1-related effects in the aged brain, particularly substantia nigra, was investigated in this study. In the nigra, we observed a progressive decrease in AT2 mRNA expression with aging, and AT2 deletion led to changes in spontaneous motor behavior, dopamine receptors, renin-angiotensin system, and pro-oxidative and pro-inflammatory markers similar to those observed in aged wild type (WT) mice. Both aged WT mice and young AT2 KO mice showed an increased AT1, decreased MAS receptor and increased angiotensinogen mRNA and/or protein expression, as well as upregulation of pro-oxidative and pro-inflammatory markers. In cultures of microglial cells, activation of AT2 receptors inhibited the LPS-induced increase in AT1 mRNA and protein expression and neuroinflammatory markers. Both in AT2 KO microglial cultures and microglia obtained from adult AT2 KO mice, an increase in AT1 mRNA expression was observed. In cultured dopaminergic neurons, AT2 activation down-regulated AT1 mRNA and protein, and dopaminergic neurons from adult AT2 KO mice showed upregulation of AT1 mRNA expression. Both in microglia and dopaminergic neurons the pathway AT2/nitric oxide/cyclic guanosine monophosphate mediates the regulation of the AT1 mRNA and protein expression through downregulation of the Sp1 transcription factor. MAS receptors are also involved in the regulation of AT1 mRNA and protein expression by AT2. The results suggest that an aging-related decrease in AT2 expression plays a major role in the aging-related AT1 overexpression and AT1-related pro-inflammatory pro-oxidative effects.

Ethanol Withdrawal Alters the Oxidative State of the Heart Through AT1-Dependent Mechanisms.

We investigated the cardiac effects of ethanol withdrawal and the possible role of AT1 receptors in such response. Male Wistar rats were treated with increasing doses of ethanol (3 to 9%, vol./vol.) for 21days. The cardiac effects of ethanol withdrawal were investigated 48h after abrupt discontinuation of ethanol. Some animals were orally treated with losartan (10mg/kg/day), a selective AT1 receptor antagonist. Ethanol withdrawal did not affect serum levels of creatine kinase (CK)-MB. Losartan prevented ethanol withdrawal-induced increase in superoxide anion (O2•-) production in the left ventricle (LV). However, ethanol withdrawal did no alter the levels of thiobarbituric acid reactive substances (TBARS) or the expression of Nox1, Nox2 or Nox4 were found in the LV. Ethanol withdrawal reduced the concentration of hydrogen peroxide (H2O2) in the LV and this response was prevented by losartan. Ethanol withdrawal increased catalase activity in the LV and losartan attenuated this response. No changes on superoxide dismutase (SOD) activity or expression were detected in the LV during ethanol withdrawal. The expression of AT1, AT2 or angiotensin converting enzyme (ACE) was not affected by ethanol withdrawal. Similarly, no changes on the expression of ERK1/2, SAPK/JNK, COX-1 or COX-2 were found in the LV during ethanol withdrawal. Ethanol withdrawal altered the cardiac oxidative state through AT1-dependent mechanisms. Our findings showed a role for angiotensin II/AT1 receptors in the initial steps of the cardiac effects induced by ethanol withdrawal.

Tanshinone IIA contributes to the pathogenesis of endometriosis via renin angiotensin system by regulating the dorsal root ganglion axon sprouting

AimsOur study was designed to explore the function and mechanism of Tanshinone IIA in alleviating pain syndrome caused by endometriosis (EMs). Main methodsFemale Sprague-Dawley rats went through autotransplantation operation to establish EMs model. The rats were randomly divided into five groups: sham, model, positive, Tanshinone IIA (L) (3 mg/kg/d) and Tanshinone IIA (H) (12 mg/kg/d) group. Volume of ectopic endometrium was measured after 21 days of continuous administration. Serum estradiol (E2) was detected by enzyme linked immunosorbent assay (Elisa). The protein expression of angiotensinogen (AGT), renin (REN), angiotensin converting enzyme (ACE), angiotensin II (ANGII) and angiotensin II type 2 receptor (AT2) in the dorsal root ganglion (DRG) neurons were measured by immunohistochemistry and Western Blotting. The mRNA expression levels of AGT and ANGII were measured by Real-time polymerase chain reaction (PCR). Key findingsTissue measurements showed that tanshinone IIA significantly inhibited the growth of ectopic endometrium. Tanshinone IIA could improve the paw withdrawal threshold thus reducing the mechanical hyperalgesia of EMs rats. Moreover, Tanshinone IIA regulated the DRG renin angiotensin system (RAS) by reducing the protein expression of AGT, REN, ACE, ANGII and AT2 in DRG neurons. Furthermore, Real-time PCR results also showed that the mRNA expression levels of AGT and ANGII in the DRG neurons were decreased. SignificanceThe Tanshinone IIA inhibitory effect on the EMs associated pain in EMs rats might occur through decreasing the expression of E2, ANGII and AT2, thus halting DRG sprouting and promoting hyperalgesia threshold.

Long-term triphenyltin exposure disrupts adrenal function in adult male rats.

Triphenyltin is an organotin, which is widely used as a fungicide in agriculture. Here, we reported the effects of triphenyltin on adrenal function in adult male rats. Adult male Sprague Dawley rats were daily gavaged with triphenyltin (0, 0.5, 1, and 2 mg/kg body weight) from postnatal day 56-86. Triphenyltin significantly decreased serum corticosterone levels at 1 and 2 mg/kg without affecting serum levels of aldosterone and adrenocorticotropic hormone. Triphenyltin increased thickness of zona glomerulosa without affecting that of zona fasciculata. Triphenyltin did not affect cell number in zona fasciculata and zona glomerulosa. Triphenyltin down-regulated the expression of Scarb1, Star, Cyp11a1, Hsd3b1, Cyp21, Cyp11b1, and Hsd11b1 at 1 and/or 2 mg/kg while it up-regulated the expression of At1, Nr4a2, and Hsd11b2 at 2 mg/kg. Triphenyltin activated the phosphorylation of AMPKα while suppressed the phosphorylation of AKT1 and SIRT1/PGC-1α in rat adrenals invivo and H295R cells invitro. Invitro, triphenyltin also induced ROS production in H295R cells at 100 nM, a concentration at which no apoptosis was induced. In conclusion, triphenyltin disrupts glucocorticoid synthesis in rat adrenal cortex via several mechanisms: 1) lowering AKT1 phosphorylation and SIRT1/PGC-1α levels; 2) activating AMPKα; and 3) possibly inducing ROS production.

Telmisartan ameliorates Aβ oligomer-induced inflammation via PPARγ/PTEN pathway in BV2 microglial cells

Telmisartan ameliorates inflammation in various brain disorders through angiotensin II type 1 receptor (AT1) blockade and peroxisome proliferator-activated receptor gamma (PPARγ) activation. Soluble β-amyloid oligomers (AβOs) play a causative role in neuronal dysfunction and memory loss in Alzheimer’s disease. In addition to directly targeting neurons, AβOs may also activate microglia to trigger toxic proinflammatory responses. Here, we investigated whether and how telmisartan ameliorates inflammatory responses in AβO-stimulated microglia. A mouse-derived BV2 microglial cell line lacking AT1 expression was selected as an in vitro model. Telmisartan not only inhibited AβO-induced proinflammatory interleukin (IL)-1β and tumor necrosis factor-α (TNF-α) expression, but also increased anti-inflammatory IL-10 expression, which was not affected by AβO stimulation. Telmisartan also inhibited AβO-induced nuclear factor (NF)-κB activity and phosphorylation of Akt and ERK, two upstream regulators of NF-κB activation. These anti-inflammatory effects were antagonized by PPARγ inhibitor GW9662. In addition, telmisartan increased the expression of PTEN (phosphate and tensin homolog deleted on chromosome 10), a lipid and protein phosphatase; PPARγ inhibitor GW9662 reversed this effect, indicating that telmisartan-induced PTEN expression is PPARγ dependent. The PTEN inhibitor blocked the effects of telmisartan on Akt and ERK phosphorylation, NF-κB transcriptional activity, and IL-1β and TNF-α production, but failed to reverse IL-10 expression. This data indicates that telmisartan-induced IL-10 expression is PPARγ-dependent but PTEN-independent. Altogether, telmisartan ameliorated AβO-induced microglial inflammation by inhibiting NF-κB-mediated proinflammatory cytokine expression via the PPARγ/PTEN pathways and by increasing PPARγ-mediated anti-inflammatory IL-10 expression. Telmisartan may present a promising therapy for the treatment of AβO pathology.

Stimulation of the ACE2/Ang-(1-7)/Mas axis in hypertensive pregnant rats attenuates cardiovascular dysfunction in adult male offspring.

The aim of this study was to investigate whether treatment with diminazene aceturate (DIZE), a putative ACE2 activator, or with angiotensin-(1-7) during pregnancy could attenuate the development of cardiovascular dysfunction in the adult offspring of spontaneously hypertensive rats (SHRs). For this, pregnant SHRs received DIZE or Ang-(1-7) throughout gestation. The systolic blood pressure (SBP) was measured in the male offspring from the 6th to16th weeks of age by tail-cuff plethysmography. Thereafter, the left ventricular contractile function and coronary reactivity were evaluated by the Langendorff technique. Samples of the left ventricles (LVs) and kidneys were collected for histology and western blot assay in another batch of adult rat offspring. Maternal treatment with DIZE or Ang-(1-7) during pregnancy attenuated the increase in SBP in adult offspring. In addition, both DIZE and Ang-(1-7) treatments reduced the cardiomyocyte diameter and fibrosis deposition in the LV, and treatment with Ang-(1-7) also reduced the fibrosis deposition in the kidneys. Maternal treatment with DIZE, as well as Ang-(1-7), improved the coronary vasodilation induced by bradykinin in isolated hearts from adult offspring. However, no difference was observed in the contractile function of the LVs of these animals. The expression levels of AT1 and Mas receptors, ACE, ACE2, SOD, and catalase in the LV were not modified by maternal treatment with Ang-(1-7), but this treatment elicited a reduction in AT2 expression. These data show that treatment with DIZE or Ang-(1-7) during gestation promoted beneficial effects of attenuating hypertension and cardiac remodeling in adult offspring.

Obesity and aging affects skeletal muscle renin-angiotensin system and myosin heavy chain proportions in pre-diabetic Zucker rats.

There is a gap in the knowledge regarding regulation of local renin-angiotensin system (RAS) in skeletal muscle during development of obesity and insulin resistance in vivo. This study evaluates the obesity- and age-related changes in the expression of local RAS components. Since RAS affects skeletal muscle remodelling, we also evaluated the muscle fibre type composition, defined by myosin heavy chain (MyHC) mRNAs and protein content. Gene expressions were determined by qPCR and/or Western blot analysis in musculus quadriceps of 3- and 8-month-old male obese Zucker rats and their lean controls. The enzymatic activity of aminopeptidase A (APA) was determined flourometrically. Activation of renin receptor (ReR)/promyelocytic leukaemia zinc finger (PLZF) negative feedback mechanism was observed in obesity. The expression of angiotensinogen and AT1 was downregulated by obesity, while neutral endopeptidase and AT2 expressions were upregulated in obese rats with aging. Skeletal muscle APA activity was decreased by obesity, which negatively correlated with the increased plasma APA activity and plasma cholesterol. The expression of angiotensin-converting enzyme (ACE) positively correlated with MyHC mRNAs characteristic for fast-twitch muscle fibres. The obesity- and age-related alterations in the expression of both classical and alternative RAS components suggest an onset of a new equilibrium between ACE/AngII/AT1 and ACE2/Ang1-7/Mas at lower level accompanied by increased renin/ReR/PLZF activation. Increased APA release from the skeletal muscle in obesity might contribute to increased plasma APA activity. There is a link between reduced ACE expression and altered muscle MyHC proportion in obesity and aging.

Angiotensin II Receptor 1 Blockage Limits Brain Damage and Improves Functional Outcome After Brain Injury in Aged Animals Despite Age-Dependent Reduction in AT1 Expression.

Traumatic brain injury (TBI) is a frequent pathology associated with poor neurological outcome in the aged population. We recently observed accelerated cerebral inflammation in aged mice in response to TBI. Candesartan is a potent specific inhibitor of angiotensin II receptor type 1 (AT1) which limits cerebral inflammation and brain damage in juvenile animals after experimental TBI. In the present study, we show significantly lower posttraumatic AT1 mRNA levels in aged (21 months) compared to young (2 months) mice. Despite low cerebral At1 expression, pharmacologic blockade by treatment with candesartan [daily, beginning 30 min after experimental TBI by controlled cortical impact (CCI)] was highly effective in both young and aged animals and reduced histological brain damage by −20% after 5 days. In young mice, neurological improvement was enhanced by AT1 inhibition 5 days after CCI. In older animals, candesartan treatment reduced functional impairment already on day 3 after TBI and post-traumatic body weight (BW) loss was attenuated. Candesartan reduced microglia activation (−40%) in young and aged animals, and neutrophil infiltration (−40% to 50%) in aged mice, whereas T-cell infiltration was not changed in either age group. In young animals, markers of anti-inflammatory microglia M2a polarization [arginase 1 (Arg1), chitinase3-like 3 (Ym1)] were increased by candesartan at days 1 and 5 after insult. In older mice 5 days after insult, expression of Arg1 was significantly higher independently of the treatment, whereas Ym1 gene expression was further enhanced by AT1 inhibition. Despite age-dependent posttraumatic differences in At1 expression levels, inhibition of AT1 was highly effective in a posttreatment paradigm. Targeting inflammation with candesartan is, therefore, a promising therapeutic strategy to limit secondary brain damage independent of the age.

Aerobic exercise training prevents obesity and insulin resistance independent of the renin angiotensin system modulation in the subcutaneous white adipose tissue.

We investigate the effects of aerobic exercise training (AET) on the thermogenic response, substrate metabolism and renin angiotensin system (RAS) in the subcutaneous white adipose tissue (SC-WAT) of mice fed cafeteria diet (CAF). Male C57BL/6J mice were assigned into groups CHOW-SED (chow diet, sedentary; n = 10), CHOW-TR (chow diet, trained; n = 10), CAF-SED (CAF, sedentary; n = 10) and CAF-TR (CAF, trained; n = 10). AET consisted in running sessions of 60 min at 60% of maximal speed, five days per week for eight weeks. The CAF-SED group showed higher body weight and adiposity, glucose intolerance and insulin resistance (IR), while AET prevented such damages in CAF-TR group. AET reduced the p-AKT/t-AKT ratio and increased ATGL expression in CHOW-TR and CAF-TR groups and increased t-HSL and p-HSL/t-HSL ratio in CAF-TR. AET prevented adipocyte hypertrophy in CAF-TR group and increased UCP-1 protein expression only in CHOW-TR. Serum ACE2 increased in CHOW-TR and CAF-TR groups, and Ang (1–7) increased in the CHOW-TR group. In the SC-WAT, CAF-TR group increased the expression of AT1, AT2 and Mas receptors, whereas CHOW-TR increased Ang (1–7) and Ang (1–7)/Ang II ratio in SC-WAT. No changes were observed in ACE and Ang II. Positive correlations were observed between UCP-1 and kITT (r = 0.6), between UCP-1 and Ang (1–7) concentration (r = 0.6), and between UCP-1 and Ang (1–7)/Ang II ratio (r = 0.7). In conclusion, the AET prevented obesity and IR, reduced insulin signaling proteins and increased lipolysis signaling proteins in the SC-WAT. In addition, the CAF diet precludes the AET-induced thermogenic response and the partial modulation of the RAS suggests that the protective effect of AET against obesity and IR could not be associated with SC-WAT RAS.