Adrenomedullin Improves Hypertension and Vascular Remodeling partly through the Receptor-Mediated AMPK Pathway in Rats with Obesity-Related Hypertension.

Adrenomedullin (ADM) is a novel cardiovascular peptide with anti-inflammatory and antioxidant properties. Chronic inflammation, oxidative stress and calcification play pivotal roles in the pathogenesis of vascular dysfunction in obesity-related hypertension (OH). Our study aimed to explore the effects of ADM on the vascular inflammation, oxidative stress and calcification in rats with OH. Eight-week-old Sprague Dawley male rats were fed with either a Control diet or a high fat diet (HFD) for 28 weeks. Next, the OH rats were randomly subdivided into two groups as follows: (1) HFD control group, and (2) HFD with ADM. A 4-week treatment with ADM (7.2 µg/kg/day, ip) not only improved hypertension and vascular remodeling, but also inhibited vascular inflammation, oxidative stress and calcification in aorta of rats with OH. In vitro experiments, ADM (10 nM) in A7r5 cells (rat thoracic aorta smooth muscle cells) attenuated palmitic acid (PA, 200 µM) or angiotensin II (Ang II, 10 nM) alone or their combination treatment-induced inflammation, oxidative stress and calcification, which were effectively inhibited by the ADM receptor antagonist ADM22-52 and AMP-activated protein kinase (AMPK) inhibitor Compound C, respectively. Moreover, ADM treatment significantly inhibited Ang II type 1 receptor (AT1R) protein expression in aorta of rats with OH or in PA-treated A7r5 cells. ADM improved hypertension, vascular remodeling and arterial stiffness, and attenuated inflammation, oxidative stress and calcification in OH state partially via receptor-mediated AMPK pathway. The results also raise the possibility that ADM will be considered for improving hypertension and vascular damage in patients with OH.

Adrenomedullin in paraventricular nucleus attenuates adipose afferent reflex and sympathoexcitation via receptors mediated nitric oxide-gamma-aminobutyric acid A type receptor pathway in rats with obesity-related hypertension.

BACKGROUND: Hypothalamic paraventricular nucleus (PVN) is an important central site for the control of the adipose afferent reflex (AAR) that increases sympathetic outflow and blood pressure in obesity-related hypertension (OH). METHOD: In this study, we investigated the effects of nitric oxide (NO) and cardiovascular bioactive polypeptide adrenomedullin (ADM) in the PVN on AAR and sympathetic nerve activity (SNA) in OH rats induced by a high-fat diet. RESULTS: The results showed that ADM, total neuronal NO synthase (nNOS) and phosphorylated-nNOS protein expression levels in the PVN of the OH rats were down-regulated compared to the control rats. The enhanced AAR in OH rats was attenuated by PVN acute application of NO donor sodium nitroprusside (SNP), but was strengthened by the nNOS inhibitor nNOS-I, guanylyl cyclase inhibitor (1H-[1,2,4]Oxadiazolo[4,3-a]quinoxalin-1-one, ODQ) and gamma-aminobutyric acid A type receptor (GABAA) antagonist Bicuculline. Moreover, PVN ADM microinjection not only decreased basal SNA but also attenuated the enhanced AAR in OH rats, which were effectively inhibited by ADM receptor antagonist ADM22-52, nNOS-I, ODQ or Bicuculline pretreatment. Bilateral PVN acute microinjection of ADM also caused greater increases in NO and cyclic guanosine monophosphate (cGMP) levels, and nNOS phosphorylation. Adeno-associated virus vectors encoding ADM (AAV-ADM) transfection in the PVN of OH rats not only decreased the elevated AAR, basal SNA and blood pressure (BP), but also increased the expression and activation of nNOS. Furthermore, AAV-ADM transfection improved vascular remodeling in OH rats. CONCLUSION: Taken together, our data highlight the roles of ADM in improving sympathetic overactivation, enhanced AAR and hypertension, and its related mechanisms associated with receptors mediated NO-cGMP-GABAA pathway in OH condition.

Adrenomedullin ameliorates palmitic acid-induced insulin resistance through PI3K/Akt pathway in adipocytes.

AIMS: White adipose tissue (WAT) dysfunction has been associated with adipose tissue low-grade inflammation and oxidative stress leading to insulin resistance (IR). Adrenomedullin (ADM), an endogenous active peptide considered as an adipokine, is associated with adipocytes function. METHODS: We evaluated the protective effects of ADM against IR in 3T3-L1 adipocytes treated by palmitic acid (PA) and in visceral white adipose tissue (vWAT) of obese rats fed with high-fat diet. RESULTS: We found that endogenous protein expressions of ADM and its receptor in PA-treated adipocytes were markedly increased. PA significantly induced impaired insulin signaling by affecting phosphatidylinositol 3-kinase (PI3K)-protein kinase B (Akt) axis and glucose transporter-4 (GLUT-4) levels, whereas ADM pretreatment enhanced insulin signaling PI3K/Akt and GLUT-4 membrane protein levels, decreased pro-inflammatory cytokines tumor necrosis factor α (TNFα), interleukin-1ß (IL-1ß) and IL-6 levels, and improved oxidative stress accompanied with reduced reactive oxygen species (ROS) levels and increased anti-oxidant enzymes manganese superoxide dismutase 2 (SOD2), glutathione peroxidase (GPx1) and catalase (CAT) protein expressions. Furthermore, ADM treatment not only improved IR in obese rats, but also effectively restored insulin signaling, and reduced inflammation and oxidative stress in vWAT of obese rats. CONCLUSIONS: This study demonstrates a prevention potential of ADM against obesity-related metabolic disorders, due to its protective effects against IR, inflammation and oxidative stress in adipocytes.

Hydrogen sulfide prevents arterial medial calcification in rats with diabetic nephropathy.

BACKGROUND: Arterial medial calcification (AMC) is associated with a high incidence of cardiovascular risk in patients with type 2 diabetes and chronic kidney disease. Here, we tested whether hydrogen sulfide (H2S) can prevent AMC in rats with diabetic nephropathy (DN). METHODS: DN was induced by a single injection of streptozotocin and high-fat diet (45% kcal as fat) containing 0.75% adenine in Sprague-Dawley rats for 8 weeks. RESULTS: Rats with DN displayed obvious calcification in aorta, and this was significantly alleviated by Sodium Hydrosulfide (NaHS, a H2S donor, 50 µmol/kg/day for 8 weeks) treatment through decreasing calcium and phosphorus content, ALP activity and calcium deposition in aorta. Interestingly, the main endogenous H2S generating enzyme activity and protein expression of cystathionine-γ-lyase (CSE) were largely reduced in the arterial wall of DN rats. Exogenous NaHS treatment restored CSE activity and its expression, inhibited aortic osteogenic transformation by upregulating phenotypic markers of smooth muscle cells SMα-actin and SM22α, and downregulating core binding factor α-1 (Cbfα-1, a key factor for bone formation), protein expressions in rats with DN when compared to the control group. NaHS administration also significantly reduced Stat3 activation, cathepsin S (CAS) activity and TGF-ß1 protein level, and improved aortic elastin expression. CONCLUSIONS: H2S may have a clinical significance for treating AMC in people with DN by reducing Stat3 activation, CAS activity, TGF-ß1 level and increasing local elastin level.

Circular RNA hsa_circ_0000700 promotes cell proliferation and migration in Esophageal Squamous Cell Carcinoma by sponging miR-1229.

Accumulating evidence has demonstrated that circular RNAs (circRNAs) are involved in the pathogenesis of cancer, including that of esophageal squamous cell carcinoma (ESCC). The current study aimed to investigate the role of hsa_circ_0000700 in ESCC. hsa_circ_0000700, miR-1229, and related functional gene expression was measured by RT-qPCR. To characterize the functions of hsa_circ_0000700 and miR-1229, ESCC cells were infected with hsa_circ_0000700-specific siRNA, miR-1229 mimics, and an inhibitor alone or in combination. Cell Counting Kit-8 (CCK8), colony formation, EdU, flow cytometry, and Transwell assays were employed to evaluate cell proliferation, apoptosis, and migration. Luciferase reporter and RNA immunoprecipitation assays were used to confirm the targeting relationship between hsa_circ_0000700 and miR-1229. Finally, a competing endogenous RNAs (ceRNA) network was built for hsa_circ_0000700, and miR-1229 targets were analyzed by bioinformatics. circ_0000700 expression was significantly upregulated in ESCC cell lines. Actinomycin D and RNase R treatment confirmed that circ_0000700 was more stable than its linear CDH9 mRNA form. Moreover, a cytoplasmic and nuclear fractionation assay suggested that circ_0000700 was mainly distributed in the cytoplasm of ECA-109 and TE-1 cells. In vitro, the proliferative and migratory capacities of ECA-109 and TE-1 cells were inhibited by knocking down circ_0000700 expression. Additionally, miR-1229 silencing reversed the circ_0000700-specific siRNA-induced attenuation of malignant phenotypes. Mechanistically, circ_0000700 was identified as a sponge of miR-1229 and could activate PRRG4, REEP5, and PSMB5 indirectly to promote ESCC progression. In summary, our results suggest that hsa_circ_0000700 functions as an oncogenic factor by sponging miR-1229 in ESCC.

Adrenomedullin Attenuates Inflammation in White Adipose Tissue of Obese Rats Through Receptor-Mediated PKA Pathway.

OBJECTIVE: Adrenomedullin (ADM) possesses therapeutic potential for inflammatory diseases. Consequently, the effects of ADM on inflammation in visceral white adipose tissue (vWAT) of obese rats or in adipocytes were explored in this study. METHODS: Male rats were fed a high-fat diet for 12 weeks to induce obesity, and obese rats were implanted with osmotic minipumps providing constant infusion of ADM (300 ng/kg per hour) and continued to be fed a high-fat diet for 4 weeks. RESULTS: When compared with the control group, endogenous protein expression of ADM and ADM receptors in vWAT and in lipopolysaccharide (LPS)-treated adipocytes was markedly increased. ADM significantly decreased the protein expression of the inflammatory mediators TNFα, IL-1ß, cyclooxygenase-2, and inducible nitric oxide synthase in vWAT of obese rats and in adipocytes stimulated by LPS. It also inhibited the activation of the inflammatory signaling pathways MAPK and NF-κB induced by LPS in adipocytes. These effects of ADM in adipocytes were inhibited by the administration of ADM receptor antagonist and cAMP-dependent protein kinase (PKA) activation inhibitor. CONCLUSIONS: ADM can inhibit inflammation in WAT in obesity, which may be mediated by the activation of ADM receptors and PKA.

Adrenomedullin 2 attenuates LPS-induced inflammation in microglia cells by receptor-mediated cAMP-PKA pathway.

Inflammation plays a critical role in the development of neurodegenerative diseases. Adrenomedullin 2 (AM2), a member of the calcitonin gene-related peptide family, has been known to have anti-inflammatory effects. Here, we evaluated the anti-inflammatory effects of AM2 in LPS-activated microglia and BV2 cells. The endogenous mRNA and protein expressions of AM2, calcitonin receptor-like receptor (CLR), receptor activity-modifying proteins (RAMPs) including RAMP1, RAMP2 and RAMP3 and the production of inflammatory mediators including tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) were detected by RT-PCR and Western blot. Our results revealed that LPS (1 µg/mL) significantly stimulated CLR, RAMP1, RAMP2 and RAMP3 protein expressions in BV2 microglia cells, but AM2 had a significant decrease. However, the mRNA levels of AM2, CLR, and RAMP1/2/3 were all markedly increased. LPS also induced obvious increases in mRNA and protein levels of the inflammatory mediators (TNF-α, IL-1ß, COX2 and iNOS). More importantly, AM2 (10 nM) administration effectively inhibited the mRNA and protein expressions of these mediators induced by LPS and increased the cAMP content in LPS-stimulated BV2 cells. Furthermore, the antagonism with AM2 receptor antagonist IMD17-47, adrenomedullin (AM) receptor antagonist by AM22-52 or the inhibition of protein kinase A (PKA) activation by P1195 effectively prevented the inhibitory role of AM2 in LPS-induced production of the above inflammatory mediators. In conclusion, AM2 inhibits LPS-induced inflammation in BV2 microglia cells that may be mainly through AM receptor-mediated cAMP-PKA pathway. Our results indicate AM2 plays an important protective role in microglia inflammation, suggesting therapeutic potential for AM2 in neuroinflammation diseases caused by activated microglia.

Adipose afferent reflex is enhanced by TNFα in paraventricular nucleus through NADPH oxidase-dependent ROS generation in obesity-related hypertensive rats.

BACKGROUND: The adipose afferent reflex (AAR), a sympatho-excitatory reflex, can promote the elevation of sympathetic nerve activity (SNA) and blood pressure (BP). Inflammation in the paraventricular nucleus (PVN) involves sympathetic abnormality in some cardiovascular diseases such as hypertension. This study was designed to explore the effects of tumor necrosis factor alpha (TNFα) in the PVN on the AAR and SNA in rats with obesity-related hypertension (OH) induced by a high-fat diet for 12 weeks. METHODS: Renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP) were continuously recorded in anesthetized rats, and their responses to capsaicin (CAP) stimulation of the right inguinal white adipose tissue were used to evaluate the AAR. RESULTS: Compared to the control rats, the systolic blood pressure (SBP), plasma norepinephrine (NE, indicating SNA) and TNFα levels, TNFα mRNA and protein levels, reactive oxygen species (ROS) content and NADPH oxidase activity in the PVN were significantly elevated in rats with OH. TNFα in the PVN markedly enhanced sympathoexcitation and AAR. Moreover, the enhancement of AAR caused by TNFα can be significantly strengthened by the pretreatment of diethyldithiocarbamate (DETC), a superoxide dismutase inhibitor, but attenuated by TNF-α receptor antagonist R-7050, superoxide scavenger PEG-SOD and NADPH oxidase inhibitor apocynin (Apo) in rats with OH. Acute microinjection of TNF-α into the PVN significantly increased the activity of NADPH oxidase and ROS levels in rats with OH, which were effectively blocked by R-7050. Furthermore, our results also showed that the increased levels of ROS, TNFα and NADPH oxidase subunits mRNA and protein in the PVN of rats with OH were significantly reversed by pentoxifylline (PTX, 30 mg/kg daily ip; in 10% ethanol) application, a cytokine blocker, for a period of 5 weeks. PTX administration also significantly decreased SBP, AAR and plasma NE levels in rats with OH. CONCLUSIONS: TNFα in the PVN modulates AAR and contributes to sympathoexcitation in OH possibly through NADPH oxidase-dependent ROS generation. TNFα blockade attenuates AAR and sympathoexcitation that unveils TNFα in the PVN may be a possible therapeutic target for the intervention of OH.

Prognostic factors affecting the all-cause death and sudden cardiac death rates of post myocardial infarction patients with low left ventricular ejection fraction.

BACKGROUND: Post myocardial infarction (post-MI) patients with low left ventricular ejection fraction (LVEF) have been candidates for an implantable cardioverter-defibrillator (ICD) since the Multicenter Automatic Defibrillator Implantation Trail II (MADIT II). However, due to the high costs of ICDs, widespread usage has not been accepted. Therefore, further risk stratification for post-MI patients with low LVEF may aid in the selection of patients that will benefit most from ICD treatment. METHODS: Four hundred and seventeen post-MI patients with low LVEF (< or = 35%) were enrolled in the study. All the patients received standard examination and proper treatment and were followed up to observe the all-cause death rate and sudden cardiac death (SCD) rate. Then COX proportional-hazards regression model was used to investigate the clinical factors which affect the all-cause death rate and SCD rate. RESULTS: Of 55 patients who died during (32 +/- 24) months of follow-up, 37 (67%) died suddenly. After adjusting for baseline clinical characteristics, multivariate COX proportional-hazards regression model identified the following variables associated with death from all causes: New York Heart Association (NYHA) heart failure class > or = III (Hazard ratio: 2.361), LVEF < or = 20% (Hazard ratio: 2.514), sustained ventricular tachycardia (Hazard ratio: 6.453), and age > or = 70 years (Hazard ratio: 3.116). The presence of sustained ventricular tachycardia (Hazard ratio: 6.491) and age > or = 70 years (Hazard ratio: 2.694) were specifically associated with SCD. CONCLUSIONS: In the post-MI patients with low LVEF, factors as LVEF < or = 20%, age > or = 70 years, presence of ventricular tachycardia, and NYHA heart failure class > or = III predict an adverse outcome. The presence of sustained ventricular tachycardia and age > or = 70 years was associated with occurrence of SCD in these patients.

Clinical study of 39 Chinese patients with arrhythmogenic right ventricular dysplasia/cardiomyopathy.

BACKGROUND: There are few studies on the clinical profile of Chinese patients with arrhythmogenic right ventricular dysplasia/cardiomyopathy (ARVD/C). The purpose of this study was to describe the clinical characteristics of ARVD/C patients from China, particularly to define the features of electrocardiograph and treatment outcomes. METHODS: Thirty-nine patients hospitalized in Fu Wai Cardiovascular Hospital from 1998 to 2006 were included. The data were obtained from the medical archive and the follow-up records. RESULTS: Of these patients 33 were male and 6 female (age at the first presentation was (34.9 +/- 9.8) years). The most common symptoms were palpitation (62%) and syncope (44%). Right precordial QRSd >or= 110 ms was detected in 69% of the patients, epsilon wave in 59%, and a ratio of QRSd in V(1) + V(2) + V(3)/V(4) + V(5) + V(6) >or= 1.2 in 82%. The most frequent features of electrocardiogram in patients without right bundle-branch block were T-wave inversions and S-wave upstroke in V(1)-V(3) >or= 55 ms (96% and 90% of 28 patients, respectively). Radiofrequency catheter ablation (RFCA) for ventricular tachycardia (VT) was successful in 15 (68%) of 22 patients. The recurrence rate of VT was 46% (7/15) during the follow-up of (16.7 +/- 11.2) months. Seven patients had cardioverter/defibrillator (ICD) implanted plus drug therapy and 17 patients took antiarrhythmic drugs alone. During the follow-up of (35.6 +/- 19.0) months, all patients with ICD implanted received at least one appropriate ICD shock. One patient died of ventricular fibrillation suddenly and one patient underwent heart transplantation for progressive biventricular heart failure during the drug therapy alone. CONCLUSIONS: This study demonstrated the clinical and ECG features of the 39 ARVD/C Chinese patients. ICD provided life-saving protection by effectively terminating malignant arrhythmias, and the high recurrence of VT was the major problem of RFCA therapy.

[Clinical and ECG features of arrhythmogenic right ventricular cardiomyopathy: a retrospective analysis of 31 cases].

OBJECTIVE: To retrospectively analyze the clinical and electrocardiographic features of patients with arrhythmogenic right ventricular cardiomyopathy (ARVC). METHODS: The clinical, electrocardiographic features and the efficacy of various therapies were analyzed in 31 patients (27 males) diagnosed as ARVC according to the criteria established by European Society of Cardiology. RESULTS: The averaged age when the ARVC was first diagnosed was (34.7 +/- 9.4) years (19 - 58 years), palpitation was present in 28 patients (90.3%) and syncope in 13 patients (41.9%), a family history of sudden death was present in 1 patient. Dilatated right ventricle was documented in 29 patients by echocardiography and (or) magnetic resonance imaging (MRI), 2 of them with dilated left ventricles. ECG changes included: T wave inversion, mostly seen in precordial leads (100%); epsilon (epsilon) wave (54.8%); QRS duration >or= 110 ms in V(1) to V(3) (83.9%); reduced extremity amplitude (41.9%); the first degree of AV block (22.6%); sustained VT (100%) including 15 monomorphic VT (48.4%) and 16 polymorphic VT (51.6%). The mean values of QRS duration in leads of V(1 - 3) [(120.8 +/- 13.7) ms] was significantly longer than that in V(4 - 6) [(99.4 +/- 13.7) ms, P < 0.05]. Fourteen patients underwent radiofrequency catheter ablation (RFCA) with an immediate success rate of 78.6% (11/14). During follow up (18.3 +/- 10.2) months, VT reoccurred in 6 patients (54.5%). The remaining 17 patients were treated with conventional medications, 7 of them were medicated under implanted cardioverter defibrillator (ICD). During the follow-up (35.6 +/- 19.0) months, VT reoccurred in 11 patients (64.7%) and one patient died suddenly. CONCLUSIONS: ARVC patients developed symptoms at mid-30s with significant ECG changes including appearance of an epsilon wave, T wave inversion and QRS duration >or= 110 ms in leads of V(1 - 3). The long term therapy efficacy was not satisfactory both for RFCA and conventional medications and ICD implantation should be recommended to patients with ARVC.