Genetic Deletion of ß-Arrestin 2 From the Subfornical Organ and Other Periventricular Nuclei in the Brain Alters Fluid Homeostasis and Blood Pressure.

BACKGROUND: ANG (angiotensin II) elicits dipsogenic and pressor responses via activation of the canonical Gαq (G-protein component of the AT1R [angiotensin type 1 receptor])-mediated AT1R in the subfornical organ. Recently, we demonstrated that ARRB2 (ß-arrestin 2) global knockout mice exhibit a higher preference for salt and exacerbated pressor response to deoxycorticosterone acetate salt. However, whether ARRB2 within selective neuroanatomical nuclei alters physiological responses to ANG is unknown. Therefore, we hypothesized that ARRB2, specifically in the subfornical organ, counterbalances maladaptive dipsogenic and pressor responses to the canonical AT1R signaling. METHODS: Male and female Arrb2FLOX mice received intracerebroventricular injection of either adeno-associated virus (AAV)-Cre-GFP (green fluorescent protein) to induce brain-specific deletion of ARRB2 (Arrb2ICV-Cre). Arrb2FLOX mice receiving ICV-AAV-GFP were used as control (Arrb2ICV-Control). Infection with ICV-AAV-Cre primarily targeted the subfornical organ with few off targets. Fluid intake was evaluated using the 2-bottle choice paradigm with 1 bottle containing water and 1 containing 0.15 mol/L NaCl. RESULTS: Arrb2ICV-Cre mice exhibited a greater pressor response to acute ICV-ANG infusion. At baseline conditions, Arrb2ICV-Cre mice exhibited a significant increase in saline intake compared with controls, resulting in a saline preference. Furthermore, when mice were subjected to water-deprived or sodium-depleted conditions, which would naturally increase endogenous ANG levels, Arrb2ICV-Cre mice exhibited elevated saline intake. CONCLUSIONS: Overall, these data indicate that ARRB2 in selective cardiovascular nuclei in the brain, including the subfornical organ, counterbalances canonical AT1R responses to both exogenous and endogenous ANG. Stimulation of the AT1R/ARRB axis in the brain may represent a novel strategy to treat hypertension.

MicroRNA-206 Contributes to the Progression of Preeclampsia by Suppressing the Viability and Mobility of Trophocytes via the Inhibition of AGTR1.

The development of preeclampsia (PE) is associated with the impaired trophoblast motility. MicroRNAs (miRs) contribute to the modulation of trophoblast invasion. In the current study, the role of miR-206/AGTR1 in the TNF-alpha-induced invasion defect of trophoblasts was explored. The levels of miR-206 and ATGR1 in clinical placenta tissues were investigated. Trophoblasts were treated with TNF-alpha, and the levels of miR-206 and ATGR1 were modulated. Changes in cell viability, invasion, and inflammation in trophoblasts were detected. The level of miR-206 was induced, while the level of AGTR1 was suppressed in placenta tissues. In in vitro assays, TNF-alpha suppressed viability, induced inflammatory response, inhibited invasion, upregulated miR-206, and down-regulated AGTR1. The inhibited expression of miR-206 or the overexpression of AGTR1 counteracted the effects of TNF-alpha, indicating the key role of the miR-206/AGTR1 in progression of PE. Collectively, miR-206 suppressed viability, induced inflammatory response, and decreased invasion of trophoblasts by inhibiting AGTR1.

Angiotensin receptor blockers retard the progression and fibrosis via inhibiting the viability of AGTR1+ CAFs in intrahepatic cholangiocarcinoma.

BACKGROUND: Intrahepatic cholangiocarcinoma (iCCA) is a highly lethal malignancy characterized by massive fibrosis and has ineffective adjuvant therapies. Here, we demonstrate the potential of angiotensin receptor blockers (ARBs) in targeting iCCA. METHODS: Masson's trichrome staining was used to assess the effect of ARBs in iCCA specimens, CCK8 and gel contraction assays in vitro and in xenograft models in vivo. RNA-seq and ATAC-seq were used for mechanistic investigations. RESULTS: Patients with iCCA who were administered ARBs had a better prognosis and a lower proportion of tumour stroma, indicating alleviated fibrosis. The presence of AGTR1, the ARBs receptor, is associated with a poor prognosis of iCCA and is highly expressed in tumour tissues and cancer-associated fibroblasts (CAFs). The ARBs strongly attenuated the viability of AGTR1+ CAFs in vitro and retarded tumour progression and fibrosis in xenograft models of co-cultured CAFs and iCCA cells. Still, they did not have a significant effect on AGTR1- CAFs. Moreover, ARBs decreased the secretion of AGTR1+ CAF-derived MFAP5 via the Hippo pathway, weakened the interaction between CAFs and iCCA cells, and impaired the aggressiveness of iCCA cells by attenuating the activation of the Notch1 pathway in iCCA cells. CONCLUSIONS: ARBs exhibit anti-fibrotic function by inhibiting the viability of AGTR1+ CAFs. These findings support using ARBs as a novel therapeutic option for targeting iCCA.

Genetic and phenotypic frequency distribution of ACE, ADRB1, AGTR1, CYP2C9*3, CYP2D6*10, CYP3A5*3, NPPA and factors associated with hypertension in Chinese Han hypertensive patients.

We analyzed the polymorphisms of 7 antihypertensive drugs-related genes and the factors associated with hypertension in hypertensive patients of Han ethnicity in Qingyang, China. A total of 354 hypertensive patients of Han ethnicity were enrolled from Qingyang, China. The ACE (I/D), ADRB1 (1165G > C), AGTR1 (1166A > C), CYP2C9*3, CYP2D6*10, CYP3A5*3 and NPPA (T2238C) polymorphisms were assessed. Clinical data of patients was also obtained. The influencing factors of hypertension were evaluated. The genotype frequencies of ACE, ADRB1, AGTR1, CYP2C9, CYP3A5 and NPPA loci were in Hardy-Weinberg equilibrium, with mutation frequencies of 39.27%, 74.29%, 6.21%, 4.80%, 72.46% and 0.71%, respectively. CYP2D6 locus was not in Hardy-Weinberg equilibrium. There was no statistical difference in allele frequencies between different genders (P > .05). There was significant difference in the frequencies of ACE (I/D) and NPPA (T2238C) loci among different regions of China (P < .05). Gender, ACE (I/D) and ADRB1 (1165G > C) gene polymorphism, smoking, homocysteine and HDL levels were associated hypertension. The mutation frequencies of ADRB1 (1165G > C) and CYP3A5*3 were high in hypertensive patients of Han ethnicity in Qingyang, suggesting these patients may be more sensitive to beta-blockers and calcium ion antagonists. Meanwhile, hypertension was associated with gender, ACE (I/D) and ADRB1 (1165G > C) gene polymorphisms, smoking, homocysteine and HDL levels.

[miR-152 inhibits the epithelial-mesenchymal transition and renin-angiotensin system of human hepatocellular carcinoma cells by down-regulating AGTR1].

Objective To investigate the effects of microRNA-152 (miR-152) targeting at angiotensin II type 1 receptor (AGTR1) on the epithelial mesenchymal transition (EMT) and renin-angiotensin system (RAS) of HCCLM3 human hepatocellular carcinoma cells. Methods The cultured HCCLM3 cells were divided into untransfected group (untreated), negative control group (transfection negative control sequence) and miR-152 group (transfected miR-152 mimic). The expressions of miR-152, angiotensin converting enzyme (ACE), angiotensin II (AngII) and angiotensin II type 1 receptor (AGTR1) mRNAs were detected by real-time fluorescence quantitative PCR. Cell invasion and migration were detected by TranswellTM assay. The expression of vimentin, N-cadherin, E-cadherin and AGTR1 were tested by western blot. The targeting relationship between miR-152 and AGTR1 were examined by double luciferase reporter assay. Results Compared with the untransfected group or the negative control group, the expression levels of miR-152 and E-cadherin protein in the miR-152 group significantly increased, while the expression levels of ACE, AngII, AGTR1 mRNA, the number of invaded cells, the number of migrating cells, and the protein expression levels of vimentin, N-cadherin, and AGTR1 decreased significantly. The results of double luciferase reporter gene assay confirmed that miR-152 can target binding with AGTR1. Conclusion miR-152 may inhibit EMT and RAS of HCCLM3 cells by targeting down-regulation of AGTR1 expression.

Angiotensin II receptor type 1 blocker candesartan improves morphine tolerance by reducing morphine­induced inflammatory response and cellular activation of BV2 cells via the PPARγ/AMPK signaling pathway.

Morphine is the most common drug of choice in clinical pain management; however, morphine tolerance presents a significant clinical challenge. The pathogenesis of morphine tolerance is known to be closely associated with angiotensin II receptor type 1 (AT1R) in microglia. As an AT1R antagonist, candesartan may serve an important role in regulating morphine tolerance. Therefore, the present study aimed to investigate the role of candesartan in morphine tolerance, and to explore the underlying mechanism. To meet this aim, BV2 microglial cells were treated with morphine or candesartan alone, or as a combination, and the expression levels of AT1R in BV2 cells were detected by reverse transcription­quantitative PCR (RT­qPCR) and western blotting. The levels of the inflammatory cytokines tumor necrosis factor­α, interleukin (IL)­1ß and IL­6 were subsequently detected by ELISA and western blotting. In addition, immunofluorescence analysis, western blotting and RT­qPCR were used to detect the expression levels of the BV2 cell activation marker, ionized calcium­binding adaptor molecule 1 (IBA­1). Western blotting was also used to detect the expression levels of peroxisome proliferator­activated receptor­Î³/AMP­activated protein kinase (PPARγ/AMPK) signaling pathway­associated proteins. Finally, the cells were treated with the PPARγ antagonist GW9662 and the AMPK inhibitor compound C to further explore the mechanism underlying the effects of candesartan on improving morphine tolerance. The expression levels of AT1R were revealed to be significantly increased following morphine induction; however, candesartan treatment inhibited the expression levels of AT1R, the levels of inflammatory cytokines and the protein expression levels of IBA­1 in morphine­induced BV2 cells in a dose­dependent manner. These processes may be associated with activation of the PPARγ/AMPK signaling pathway. Taken together, the present study revealed that treatment with candesartan reduced morphine­induced inflammatory response and cellular activation of BV2 cells via PPARγ/AMPK signaling.

Association of AGTR1 gene methylation and its genetic variant in Chinese farmer with hypertension: A case-control study.

The objective was to determine the potential associations of the angiotensin II receptor type 1 (AGTR1) gene polymorphism, methylation, and lipid metabolism in Chinese farmers with hypertension. A case-control study was conducted in Wuzhi county of Henan province in China in 2013 to 2014. A total of 1034 local residents (35-74 years, 386 hypertensive cases, and 648 normotensive subjects) were enrolled in this study. Triglyceride (TG), total cholesterol (TC), high-density lipoprotein, and low-density lipoprotein were measured using automatic chemistry analyzer. The AGTR1 gene promoter methylation level was measured using quantitative methylation-specific polymerase chain reaction method. The single nucleotide polymorphism rs275653 was genotyped with TaqMan probe assay at an applied biosystems platform. The gender, body mass index (BMI), TG, TC, and family history of hypertension in the hypertension group were significantly higher than those in control group (P < .05). No significant difference was observed in the distribution of AGTR1 rs275653 polymorphism in the hypertension and controls (P > .05). The AGTR1 gene methylation in subjects carrying different genotypes was not significantly observed (P > .05). The logistic regression analysis found the AGTR1 gene methylation level was negative correlation with hypertension in the present study (odds ratio, 0.946, 95% confidence interval, 0.896-0.999) through adjusting for age, gender, BMI, education, smoking, alcohol drinking, fruit and vegetable intake, pickles intake, and family history of hypertension. The association of AGTR1 gene hypomethylation and essential hypertension was observed in Chinese farmers; no significant difference was observed in the distribution of AGTR1 rs275653 polymorphism.

Central angiotensinergic mechanisms in female spontaneously hypertensive rats treated with estradiol.

Estrogens reduce 0.3 M NaCl intake and palatability in a widely used model of essential hypertension, the spontaneously hypertensive rats (SHRs). Here we investigated whether the inhibitory effects of ß-estradiol (E2, 10 µg/kg b.w. subcutaneously for 8 days) on water deprived partially-rehydrated (WD-PR) ovariectomized (OVX) adult female SHRs (fSHRs, n = 4-10/group) are related to interferences on brain angiotensin II AT1 receptors (AT1r). After WD-PR, E2 reduced 0.3 M NaCl intake (1.3 ± 0.6, vs. vehicle: 3.5 ± 1.2 ml/30 min), the number of hedonic responses to intraoral NaCl infusion (57 ± 11, vs. vehicle: 176 ± 32/min), and the relative angiotensin AT1r (Agtr1a) mRNA expression in the hypothalamus. Losartan (AT1r antagonist, 100 µg) intracerebroventricularly in OVX fSHRs treated with vehicle subcutaneously abolished 0.3 M NaCl intake (0.1 ± 0.1 ml/30 min) and only transiently reduced hedonic responses to intraoral NaCl. Losartan combined with E2 decreased the number of hedonic and increased the number of aversive responses to intraoral NaCl and abolished 0.3 M NaCl intake. E2 also reduced the pressor and dipsogenic responses to intracerebroventricular angiotensin II. The results suggest that AT1r activation increases palatability and induces NaCl intake in WD-PR fSHRs. E2 reduced hypothalamic Agtr1a mRNA expression, which may account for the effects of E2 on NaCl intake and palatability and intracerebroventricular angiotensin II-induced pressor and dipsogenic responses in OVX fSHRs. Future studies considering natural fluctuations in estrogen secretion might help to determine the degree of such interference in brain neuronal activity.

Type 1 Angiotensin Receptors on CD11c-Expressing Cells Protect Against Hypertension by Regulating Dendritic Cell-Mediated T Cell Activation.

BACKGROUND: Type 1 angiotensin (AT1) receptors are expressed on immune cells, and we previously found that bone marrow-derived AT1 receptors protect against Ang (angiotensin) II-induced hypertension. CD11c is expressed on myeloid cells derived from the bone marrow, including dendritic cells (DCs) that activate T lymphocytes. Here, we examined the role of AT1 receptors on CD11c+ cells in hypertension pathogenesis. METHODS: Mice lacking the dominant murine AT1 receptor isoform, AT1a, on CD11c+ cells (dendritic cell [DC] AT1aR knockout [KO]) and wild-type (WT) littermates were subjected to Ang II-induced hypertension. Blood pressures were measured by radiotelemetry. RESULTS: DC AT1aR KO mice had exaggerated hypertensive responses to chronic Ang II infusion with enhanced renal accumulation of effector memory T cells and CD40+ DCs. CCL5 (C-C motif chemokine ligand 5) recruits T cells into injured tissues, and CCR7 (C-C motif chemokine receptor 7) facilitates DC and T cell interactions in the kidney lymph node to allow T cell activation. DCs from the hypertensive DC AT1aR KO kidneys expressed higher levels of CCL5 and CCR7. mRNA expressions for CCR7 and tumor necrosis factor-α were increased in CD4+ T cells from the renal lymph nodes of DC AT1aR KO mice. During the second week of Ang II infusion when blood pressures between groups diverged, DC AT1aR KO mice excreted less sodium than WTs. Expressions for epithelial sodium channel subunits were increased in DC AT1aR KO kidneys. CONCLUSIONS: Following activation of the renin angiotensin system, AT1aR stimulation on DCs suppresses renal DC maturation and T cell activation with consequent protection from sodium retention and blood pressure elevation.

USF1-ATRAP-PBX3 Axis Promote Breast Cancer Glycolysis and Malignant Phenotype by Activating AKT/mTOR Signaling.

Angiotensin II type 1 receptor-associated protein (ATRAP) is widely expressed in different tissues and organs, although its mechanistic role in breast cancer remains unclear. Here, we show that ATRAP is highly expressed in breast cancer tissues. Its aberrant upregulation promotes breast cancer aggressiveness and is positively correlated with poor prognosis. Functional assays revealed that ATRAP participates in promoting cell growth, metastasis, and aerobic glycolysis, while microarray analysis showed that ATRAP can activate the AKT/mTOR signaling pathway in cancer progression. In addition, ATRAP was revealed to direct Ubiquitin-specific protease 14 (USP14)-mediated deubiquitination and stabilization of Pre-B cell leukemia homeobox 3 (PBX3). Importantly, ATRAP is a direct target of Upstream stimulatory factor 1 (USF1), and that ATRAP overexpression reverses the inhibitory effects of USF1 knockdown. Our study demonstrates the broad contribution of the USF1/ATRAP/PBX3 axis to breast cancer progression and provides a strong potential therapeutic target.

AGTR1, PLTP, and SCG2 associated with immune genes and immune cell infiltration in calcific aortic valve stenosis: analysis from integrated bioinformatics and machine learning.

Background: Calcific aortic valve stenosis (CAVS) is a crucial cardiovascular disease facing aging societies. Our research attempts to identify immune-related genes through bioinformatics and machine learning analysis. Two machine learning strategies include Least Absolute Shrinkage Selection Operator (LASSO) and Support Vector Machine Recursive Feature Elimination (SVM-RFE). In addition, we deeply explore the role of immune cell infiltration in CAVS, aiming to study the potential therapeutic targets of CAVS and explore possible drugs. Methods: Download three data sets related to CAVS from the Gene Expression Omnibus. Gene set variation analysis (GSVA) looks for potential mechanisms, determines differentially expressed immune-related genes (DEIRGs) by combining the ImmPort database with CAVS differential genes, and explores the functions and pathways of enrichment. Two machine learning methods, LASSO and SVM-RFE, screen key immune signals and validate them in external data sets. Single-sample GSEA (ssGSEA) and CIBERSORT analyze the subtypes of immune infiltrating cells and integrate the analysis with DEIRGs and key immune signals. Finally, the possible targeted drugs are analyzed through the Connectivity Map (CMap). Results: GSVA analysis of the gene set suggests that it is highly correlated with multiple immune pathways. 266 differential genes (DEGs) integrate with immune genes to obtain 71 DEIRGs. Enrichment analysis found that DEIRGs are related to oxidative stress, synaptic membrane components, receptor activity, and a variety of cardiovascular diseases and immune pathways. Angiotensin II Receptor Type 1(AGTR1), Phospholipid Transfer Protein (PLTP), Secretogranin II (SCG2) are identified as key immune signals of CAVS by machine learning. Immune infiltration found that B cells naï ve and Macrophages M2 are less in CAVS, while Macrophages M0 is more in CAVS. Simultaneously, AGTR1, PLTP, SCG2 are highly correlated with a variety of immune cell subtypes. CMap analysis found that isoliquiritigenin, parthenolide, and pyrrolidine-dithiocarbamate are the top three targeted drugs related to CAVS immunity. Conclusion: The key immune signals, immune infiltration and potential drugs obtained from the research play a vital role in the pathophysiological progress of CAVS.

Deficiency of the kidney tubular angiotensin II type1 receptor-associated protein ATRAP exacerbates streptozotocin-induced diabetic glomerular injury via reducing protective macrophage polarization.

Although activation of the renin-angiotensin system and of its glomerular components is implicated in the pathogenesis of diabetic nephropathy, the functional roles of the tubular renin-angiotensin system with AT1 receptor signaling in diabetic nephropathy are unclear. Tissue hyperactivity of the renin-angiotensin system is inhibited by the angiotensin II type 1 receptor-associated protein ATRAP, which negatively regulates receptor signaling. The highest expression of endogenous ATRAP occurs in the kidney, where it is mainly expressed by tubules but rarely in glomeruli. Here, we found that hyperactivation of angiotensin II type 1 receptor signaling in kidney tubules exacerbated diabetic glomerular injury in a mouse model of streptozotocin-induced diabetic nephropathy. These phenomena were accompanied by decreased expression of CD206, a marker of alternatively activated and tissue-reparative M2 macrophages, in the kidney tubulointerstitium. Additionally, adoptive transfer of M2- polarized macrophages into diabetic ATRAP-knockout mice ameliorated the glomerular injury. As a possible mechanism, the glomerular mRNA levels of tumor necrosis factor-α and oxidative stress components were increased in diabetic knockout mice compared to non-diabetic knockout mice, but these increases were ameliorated by adoptive transfer. Furthermore, proximal tubule-specific ATRAP downregulation reduced tubulointerstitial expression of CD206, the marker of M2 macrophages in diabetic mice. Thus, our findings indicate that tubular ATRAP-mediated functional modulation of angiotensin II type 1 receptor signaling modulates the accumulation of tubulointerstitial M2 macrophages, thus affecting glomerular manifestations of diabetic nephropathy via tubule-glomerular crosstalk.

Polymorphisms in ACE, ACE2, AGTR1 genes and severity of COVID-19 disease.

BACKGROUND: Infection by the SARS-Cov-2 virus produces in humans a disease of highly variable and unpredictable severity. The presence of frequent genetic single nucleotide polymorphisms (SNPs) in the population might lead to a greater susceptibility to infection or an exaggerated inflammatory response. SARS-CoV-2 requires the presence of the ACE2 protein to enter in the cell and ACE2 is a regulator of the renin-angiotensin system. Accordingly, we studied the associations between 8 SNPs from AGTR1, ACE2 and ACE genes and the severity of the disease produced by the SARS-Cov-2 virus. METHODS: 318 (aged 59.6±17.3 years, males 62.6%) COVID-19 patients were grouped based on the severity of symptoms: Outpatients (n = 104, 32.7%), hospitalized on the wards (n = 73, 23.0%), Intensive Care Unit (ICU) (n = 84, 26.4%) and deceased (n = 57, 17.9%). Comorbidity data (diabetes, hypertension, obesity, lung disease and cancer) were collected for adjustment. Genotype distribution of 8 selected SNPs among the severity groups was analyzed. RESULTS: Four SNPs in ACE2 were associated with the severity of disease. While rs2074192 andrs1978124showed a protector effectassuming an overdominant model of inheritance (G/A vs. GG-AA, OR = 0.32, 95%CI = 0.12-0.82; p = 0.016 and A/G vs. AA-GG, OR = 0.37, 95%CI: 0.14-0.96; p = 0.038, respectively); the SNPs rs2106809 and rs2285666were associated with an increased risk of being hospitalized and a severity course of the disease with recessive models of inheritance (C/C vs. T/C-T/T, OR = 11.41, 95% CI: 1.12-115.91; p = 0.012) and (A/A vs. GG-G/A, OR = 12.61, 95% CI: 1.26-125.87; p = 0.0081). As expected, an older age (OR = 1.47), male gender (OR = 1.98) and comorbidities (OR = 2.52) increased the risk of being admitted to ICU or death vs more benign outpatient course. Multivariable analysis demonstrated the role of the certain genotypes (ACE2) with the severity of COVID-19 (OR: 0.31, OR 0.37 for rs2074192 and rs1978124, and OR = 2.67, OR = 2.70 for rs2106809 and rs2285666, respectively). Hardy-Weinberg equilibrium in hospitalized group for I/D SNP in ACE was not showed (p<0.05), which might be due to the association with the disease. No association between COVID-19 disease and the different AGTR1 SNPs was evidenced on multivariable, nevertheless the A/A genotype for rs5183 showed an higher hospitalization risk in patients with comorbidities. CONCLUSIONS: Different genetic variants in ACE2 were associated with a severe clinical course and death groups of patients with COVID-19. ACE2 common SNPs in the population might modulate severity of COVID-19 infection independently of other known markers like gender, age and comorbidities.

The unfavorable clinical outcome of COVID-19 in smokers is mediated by H3K4me3, H3K9me3 and H3K27me3 histone marks.

Smoking could predispose individuals to a more severe COVID-19 by upregulating a particular gene known as mdig, which is mediated through a number of well-known histone modifications. Smoking might regulate the transcription-activating H3K4me3 mark, along with the transcription-repressing H3K9me3 and H3K27me3 marks, in a way to favor SARS-CoV-2 entry by enhancing the expression of ACE2, NRP1 and NRP2, AT1R, CTSD and CTSL, PGE2 receptors 2-4, SLC6A20 and IL-6, all of which interact either directly or indirectly with important receptors, facilitating viral entry in COVID-19.

Lay abstract The role of smoking in development of several respiratory diseases has been clearly established. A significant proportion of these deleterious effects is mediated through epigenetic mechanisms, particularly histone modifications. Recent evidence indicates that smoking induces the expression of a mediator known as mdig, which in turn alters the transcription of several key proteins that have been implicated in development of COVID-19.

Chronic oestrogen deficiency induced by ovariectomy may cause lung fibrosis through activation of the renin-angiotensin system in rats.

CONTEXT: Oestrogen deficiency is linked with pulmonary fibrosis. Additionally, it may lead to over-activation of the renin-angiotensin system (RAS), which worsens lung fibrosis. OBJECTIVE: The present study aims to investigate the role of RAS on lung fibrosis associated with oestrogen deficiency in ovariectomised rats. MATERIALS AND METHODS: Serum 17ß-oestradiol (E2), arterial blood gases, plasma angiotensin II levels, lung tissue hydroxyproline content, and transforming growth factor beta 1 (TGF-ß1) concentration, the mRNA expression of angiotensin type 1 receptor (AT1R), and angiotensin-converting enzyme (ACE1) were evaluated. Moreover, lung tissues were examined by histopathology and immunohistochemistry. RESULTS: Hydroxyproline content, TGF-ß1 concentration, plasma angiotensin II, the relative mRNA expression of ACE1, and AT1R is found to increase in ovariectomised rats. The mentioned changes can be largely rescued by administration of RAS blockers. CONCLUSION: Oestrogen deficiency activates RAS, which consequently increases the expression of pro-fibrotic factors and stimulates the fibrotic cascade causing lung fibrosis.

Angiotensin II receptor 1 controls profibrotic Wnt/ß-catenin signalling in experimental autoimmune myocarditis.

AIMS: Angiotensin (Ang) II signalling has been suggested to promote cardiac fibrosis in inflammatory heart diseases; however, the underlying mechanisms remain obscure. Using Agtr1a-/- mice with genetic deletion of angiotensin receptor type 1 (ATR1) and the experimental autoimmune myocarditis (EAM) model, we aimed to elucidate the role of Ang II-ATR1 pathway in development of heart-specific autoimmunity and post-inflammatory fibrosis. METHODS AND RESULTS: EAM was induced in wild-type (WT) and Agtr1a-/- mice by subcutaneous injections with alpha myosin heavy chain peptide emulsified in complete Freund's adjuvant. Agtr1a-/- mice developed myocarditis to a similar extent as WT controls at day 21 but showed reduced fibrosis and better systolic function at day 40. Crisscross bone marrow chimaera experiments proved that ATR1 signalling in the bone marrow compartment was critical for cardiac fibrosis. Heart infiltrating, bone-marrow-derived cells produced Ang II, but lack of ATR1 in these cells reduced transforming growth factor beta (TGF-ß)-mediated fibrotic responses. At the molecular level, Agtr1a-/- heart-inflammatory cells showed impaired TGF-ß-mediated phosphorylation of Smad2 and TAK1. In WT cells, TGF-ß induced formation of RhoA-GTP and RhoA-A-kinase anchoring protein-Lbc (AKAP-Lbc) complex. In Agtr1a-/- cells, stabilization of RhoA-GTP and interaction of RhoA with AKAP-Lbc were largely impaired. Furthermore, in contrast to WT cells, Agtr1a-/- cells stimulated with TGF-ß failed to activate canonical Wnt pathway indicated by suppressed activity of glycogen synthase kinase-3 (GSK-3)ß and nuclear ß-catenin translocation and showed reduced expression of Wnts. In line with these in vitro findings, ß-catenin was detected in inflammatory regions of hearts of WT, but not Agtr1a-/- mice and expression of canonical Wnt1 and Wnt10b were lower in Agtr1a-/- hearts. CONCLUSION: Ang II-ATR1 signalling is critical for development of post-inflammatory fibrotic remodelling and dilated cardiomyopathy. Our data underpin the importance of Ang II-ATR1 in effective TGF-ß downstream signalling response including activation of profibrotic Wnt/ß-catenin pathway.

Correlation of ACE2 with RAS components after Losartan treatment in light of COVID-19.

Angiotensin-converting enzyme 2 (ACE2) is an important factor in coronavirus disease (COVID-19) interactions. Losartan (LOS) belongs to the angiotensin receptor blocker (ARB) family. Additionally, the protective role of ACE2 restored by LOS has been suggested and clinically examined in the treatment of COVID-19 patients. Furthermore, clinical trials with LOS have been conducted. However, the mechanism through which LOS enhances ACE2 expression remains unclear. In addition, the response of ACE2 to LOS differs among patients. Our LOS-treated patient data revealed a correlated mechanism of ACE2 with components of the renin-angiotensinogen system. We observed a significant positive regulation of MAS1 and ACE2 expression. In the context of LOS treatment of COVID-19, ACE2 expression could depend on LOS regulated MAS1. Thus, MAS1 expression could predict the COVID-19 treatment response of LOS.

LINC00852 promotes the proliferation and invasion of ovarian cancer cells by competitively binding with miR-140-3p to regulate AGTR1 expression.

BACKGROUND: Dysregulation of long non-coding RNAs (lncRNAs) has been identified in ovarian cancer. However, the expression and biological functions of LINC00852 in ovarian cancer are not understood. METHODS: The expressions of LINC00852, miR-140-3p and AGTR1 mRNA in ovarian cancer tissues and cells were detected by quantitative reverse transcription polymerase chain reaction (qRT-PCR) assay. Gain- and loss-of-function assays were performed to explore the biological functions of LINC00852 and miR-140-3p in the progression of ovarian cancer in vitro. The bindings between LINC00852 and miR-140-3p were confirmed by luciferase reporter gene assay, RNA immunoprecipitation (RIP) assay and RNA pull-down assay. RESULTS: We found that LINC00852 expression was significantly up-regulated in ovarian cancer tissues and cells, whereas miR-140-3p expression was significantly down-regulated in ovarian cancer tissues. Functionally, LINC00852 knockdown inhibited the viability, proliferation and invasion of ovarian cancer cells, and promoted the apoptosis of ovarian cancer cells. Further investigation showed that LINC00852 interacted with miR-140-3p, and miR-140-3p overexpression suppressed the viability, proliferation and invasion of ovarian cancer cells. In addition, miR-140-3p interacted with AGTR1 and negatively regulated its level in ovarian cancer cells. Mechanistically, we found that LINC00852 acted as a ceRNA of miR-140-3p to promote AGTR1 expression and activate MEK/ERK/STAT3 pathway. Finally, LINC00852 knockdown inhibited the growth and invasion ovarian cancer in vivo. CONCLUSION: LINC00852/miR-140-3p/AGTR1 is an important pathway to promote the proliferation and invasion of ovarian cancer.

Effect of Angiotensin II on Chondrocyte Degeneration and Protection via Differential Usage of Angiotensin II Receptors.

The renin-angiotensin system (RAS) controls not only systemic functions, such as blood pressure, but also local tissue-specific events. Previous studies have shown that angiotensin II receptor type 1 (AT1R) and type 2 (AT2R), two RAS components, are expressed in chondrocytes. However, the angiotensin II (ANG II) effects exerted through these receptors on chondrocyte metabolism are not fully understood. In this study, we investigated the effects of ANG II and AT1R blockade on chondrocyte proliferation and differentiation. Firstly, we observed that ANG II significantly suppressed cell proliferation and glycosaminoglycan content in rat chondrocytic RCS cells. Additionally, ANG II decreased CCN2, which is an anabolic factor for chondrocytes, via increased MMP9. In Agtr1a-deficient RCS cells generated by the CRISPR-Cas9 system, Ccn2 and Aggrecan (Acan) expression increased. Losartan, an AT1R antagonist, blocked the ANG II-induced decrease in CCN2 production and Acan expression in RCS cells. These findings suggest that AT1R blockade reduces ANG II-induced chondrocyte degeneration. Interestingly, AT1R-positive cells, which were localized on the surface of the articular cartilage of 7-month-old mice expanded throughout the articular cartilage with aging. These findings suggest that ANG II regulates age-related cartilage degeneration through the ANG II-AT1R axis.

Stearoyl-CoA Desaturase (SCD) Induces Cardiac Dysfunction with Cardiac Lipid Overload and Angiotensin II AT1 Receptor Protein Up-Regulation.

Heart failure is a major cause of death worldwide with insufficient treatment options. In the search for pathomechanisms, we found up-regulation of an enzyme, stearoyl-CoA desaturase 1 (Scd1), in different experimental models of heart failure induced by advanced atherosclerosis, chronic pressure overload, and/or volume overload. Because the pathophysiological role of Scd1/SCD in heart failure is not clear, we investigated the impact of cardiac SCD upregulation through the generation of C57BL/6-Tg(MHCSCD)Sjaa mice with myocardium-specific expression of SCD. Echocardiographic examination showed that 4.9-fold-increased SCD levels triggered cardiac hypertrophy and symptoms of heart failure at an age of eight months. Tg-SCD mice had a significantly reduced left ventricular cardiac ejection fraction of 25.7 ± 2.9% compared to 54.3 ± 4.5% of non-transgenic B6 control mice. Whole-genome gene expression profiling identified up-regulated heart-failure-related genes such as resistin, adiponectin, and fatty acid synthase, and type 1 and 3 collagens. Tg-SCD mice were characterized by cardiac lipid accumulation with 1.6- and 1.7-fold-increased cardiac contents of saturated lipids, palmitate, and stearate, respectively. In contrast, unsaturated lipids were not changed. Together with saturated lipids, apoptosis-enhancing p53 protein contents were elevated. Imaging by autoradiography revealed that the heart-failure-promoting and membrane-spanning angiotensin II AT1 receptor protein of Tg-SCD hearts was significantly up-regulated. In transfected HEK cells, the expression of SCD increased the number of cell-surface angiotensin II AT1 receptor binding sites. In addition, increased AT1 receptor protein levels were detected by fluorescence spectroscopy of fluorescent protein-labeled AT1 receptor-Cerulean. Taken together, we found that SCD promotes cardiac dysfunction with overload of cardiotoxic saturated lipids and up-regulation of the heart-failure-promoting AT1 receptor protein.