Administration Of Angiotensin Research Articles

Fibroblast-derived extracellular vesicles as trackable efficient transporters of an experimental nanodrug with fibrotic heart and lung targeting.

The discovery of extracellular vesicles (EVs) as efficient exogenous biotransporters of therapeutic agents into cells across biological membranes is an exciting emerging field. Especially the potential of EVs as targeted delivery systems for diseases with selective treatments, such as fibrosis, whose treatment causes side effects in other organs not involved in the disease. Methods: In this study, we collected embryonic fibroblast-derived EVs from two different centrifugation fractions, 10 K g and 100 K g fractions from a NIH-3T3 cell line loaded with an experimental drug. Mice with fibrotic hearts and lungs were obtained by administration of angiotensin II. We generated fluorescent EVs and bioluminescent drug to observe their accumulation by colocalization of their signals in fibrotic heart and lung. The biodistribution of the drug in various organs was obtained by detecting the Au present in the drug nanostructure. Results: The drug-loaded EVs successfully reduced fibrosis in pathological fibroblasts in vitro, and modified the biodistribution of the experimental drug, enabling it to reach the target organs in vivo. We described the pre-analytical characteristics of EVs related to physical variables, culture and harvesting conditions, crucial for their in vivo application as nanotransporters using a previously validated protein-based antifibrotic drug. The results showed the colocalization of EVs and the experimental drug in vivo and ex vivo and the efficient reduction of fibrosis in vitro. This work demonstrates that 10K-EVs and 100K-EVs derived from fibroblasts can act as effective biotransporters for targeted drug delivery to profibrotic fibroblasts, lungs, or heart. Conclusion: We observed that fibroblast-derived 10K-EVs and 100K-EVs are useful biotransporters encapsulating a new generation drug leading to a reduction of fibrosis in profibrotic fibroblasts in vitro. In addition, drug containing EVs were shown to reach fibrotic heart and lungs in vivo, enhancing free drug biodistribution.

Effects of vericiguat on redox signaling in cardiomyocyte

Abstract Background Reactive oxygen species (ROS) inhibits the nitric oxide (NO)-soluble guanylate cyclase (sGC)-cyclic guanosine monophosphate (cGMP) pathway in the patients with advanced heart failure (HF) and contributes to the prognosis of HF. Vericiguat stimulates sGC independently of NO and reduced morbidity/mortality for HF patients with reduced ejection fraction. However, the effects of vericiguat on the cardiomyocyte in terms of ROS is unclear. Purpose We explored the cardioprotective effects of vericiguat focusing on the oxidative stress. Methods Cardiomyocytes isolated from the whole heart of neonatal rats were incubated with vericiguat (0μM as control, 0.1μM, 1.0μM, 10μM) for 72 hours. Administration of angiotensin II (1.0μM) was used to induce excessive ROS. Superoxide was quantified using luminometry, and whole ROS or mitochondria-derived superoxide was evaluated by using live-cell fluorescent probes. ERK 1/2 signaling pathway activity were evaluated with western blot analysis. Results Nicotinamide adenine dinucleotide phosphate (NADPH)-stimulated superoxide was significantly increased by the treatment of angiotensin II (40208 ± 6107 reactive light unit (RLU) / mg, p = 0.0009) and the angiotensin II-induced superoxide was significantly attenuated by vericiguat at doses of 0.1μM (28240 ± 6507 RLU / mg, p = 0.03) , 1.0μM (25221 ± 6496 RLU / mg, p = 0.007) and 10μM (8230 ± 2568 RLU / mg, p < 0.0001). Corrected total cell fluorescence value of live-cell fluorescent probes revealed that whole ROS and mitochondria-derived superoxide were significantly increased by angiotensin II treatment (525.3 ± 119.6, p = 0.004 and 945.3 ± 153.0, p <0.0001), which were attenuated by vericiguat (0.1μM, 431.2 ± 45.7, p = 0.48 and 548.1 ± 103.9, p = 0.0004; 1.0μM, 336.1 ± 109.0, p = 0.04 and 421.3 ± 111.9, p < 0.0001; 10μM, 270.0 ± 44.4, p = 0.005 and 157.7 ± 48.3, p < 0.0005) dose-dependently. Angiotensin II-induced phosphorylation of ERK 1/2 was suppressed by vericiguat at doses of 1.0μM (fold change 0.58, p = 0.04) and 10μM (fold change 0.45, p = 0.006) treatment. Conclusion Our results indicated that vericiguat could suppress the oxidative stress induced by angiotensin II and following activation of ERK 1/2 signaling. Suppressing the oxidative stress could lead to further activation of NO-sGC-cGMP pathway not only the stimulation by vericiguat and favorable cardio-protective cycle is expected.Quantification of superoxideQuantification of ROS or superoxide

SINGLE-CELL TRANSCRIPTOMIC PROFILING OF THE HYPERTENSION MOUSE BRAIN

Objective: Chronic hypertension has devastating effects on the brain, being the major cause of stroke and a leading cause of dementia. The alterations of cerebral blood vessels structure and vasoregulatory mechanisms leading by hypertension have been recognized for decades. However, the specific cellular and molecular mediators that drive these alterations remain unclear. Design and method: After 4 weeks of continuous administration of angiotensin II, we performed a single-cell transcriptomic analysis of brain cellulome. The brain cellular ecosystem in response to angiotensin II was analyzed, the network of cells that forms the mouse brain was profiled after a proinflammation stimulus that drives pathological brain vascular remodeling. Results: We analyzed the transcriptomes of 52,856 single cells (28,078 normal and 24,778 angiotensin II) derived from the brains of 10 normal and 12 angiotensin II mice, and compared all the cellular composition and transcriptomes of the whole brain cells. For all the major cell populations, we provide comprehensive datasets of genes and pathways whose transcriptional profiles change with hypertension. To reveal heterogeneity within each population, we grouped the cells into 14 classes based on the expression profile, lineage, function and anatomical organization. Of the 20718 total detected genes, 800 were significantly affected by hypertension in at least one cell type. Hypertensive- related cellular pathways and processes were also investigated, and we highlight changes in EC and EPC. Furthermore, the cellular responses to angiotensin II and the changes in intercellular communication were identified. Conclusions: These large-scale datasets provide a valuable resource for exploring the brain cellular landscape in health and after chronic cerebral vascular stress. These data provide insights into the cellular and molecular mechanisms that afford an additional discoveries of directed towards understanding the mechanisms by which hypertension disrupts cerebral blood vessels.

Macrophage scavenger receptor A1 antagonizes abdominal aortic aneurysm via upregulating IRG1

AimsAbdominal aortic aneurysm (AAA) is a common, usually asymptomatic disease with high mortality and limited therapeutic options. Extensive extracellular matrix (ECM) fragmentation and transmural inflammation act as major pathological processes of AAA. However, the underlying regulatory mechanisms remain incompletely understood. Herein, we aimed to investigate the role of scavenger receptor A1 (SR-A1), a key pattern recognition receptor modulating macrophage activity, in pathogenesis of AAA. Methods and resultsThe AAA model was generated by administration of angiotensin II (Ang II) into apolipoprotein E knockout mice or peri-arterial application of calcium phosphate in C57BJ/6L mice. We found that SR-A1 was markedly down-regulated in the macrophages isolated from murine AAA aortas. Global or myeloid-specific ablation of SR-A1 aggravated vascular inflammation, loss of vascular smooth muscle cells and degradation of the extracellular matrix. These effects of SR-A1 deficiency on AAA development were mediated by suppressed immunoresponsive gene 1 (IRG1) and increased inflammatory response in macrophages. Mechanically, binding of SR-A1 with Lyn led to STAT3 phosphorylation and translocation into the nucleus, in which STAT3 promoted IRG1 transcription through directly binding to its promoter. Restoration of macrophage SR-A1 in SR-A1-deficient mice by bone marrow transplantation or administration of 4-octyl itaconate, the derivate of IRG1 product itaconate, could relieve murine AAA. ConclusionOur study reveals a protective effect of macrophage SR-A1–STAT3–IRG1 axis against aortic aneurysm formation via inhibiting inflammation.

PS-BPB01-7: ANGIOTENSIN II INDUCES AORTIC RUPTURE AND DISSECTION IN OSTEOPROTEGERIN-DEFICIENT MICE

Background: The biological mechanism of action for osteoprotegerin (OPG), a soluble decoy receptor for the receptor activator of nuclear factor kappa-B ligand (RANKL) in the vascular structure, has not been elucidated. The study aim was to determine if OPG affects aortic structural integrity in angiotensin II-induced hypertension. Methods and Results: Mortality was higher (P < 0.0001 by Log-rank test) in 8-week-old male homozygotes of OPG gene knockout (OPG -/- mice given subcutaneous administration of angiotensin II (1,000 ng/kg/min) for 28 days, with an incidence of 21% fatal aortic rupture (Chi-square, 8.024, P = 0.005), and 23% aortic dissection (Chi-square, 4.285, P = 0.038), than in age-matched wild type mice. The absence of OPG was associated with decreased medial and adventitial thickness and increased numbers of elastin breaks as well as with 2.5-fold increased periostin expression and soluble RANKL concentrations. PEGylated human recombinant OPG administration decreased all-causes mortality (-22%, P < 0.001 by Log-rank test), the incidence of fatal aortic rupture (-13%, P = 0.08), and aortic dissection (-23%, P < 0.001) with decreasing numbers of elastin breaks (P = 0.0539) at the supra-renal aorta, periostin expressions (P = 0.0011), and soluble RANKL concentrations (P = 0.0012) in angiotensin II-infused OPG-/- mice. Conclusions: These data suggest that OPG protects against aortic rupture and dissection in Ang II-induced hypertension by inhibiting RANKL activity and periostin expression.

PS-BPB07-5: HYPERTENSIVE STIMULI PROMOTE NEUROINFLAMMATION AND COGNITIVE IMPAIRMENT IN A PRESSURE-DEPENDENT MANNER

Objective: Hypertension greatly increases the risk for cognitive impairment, and is associated with inflammation of the brain. However, it is unclear to what extent there is neuroinflammation during hypertension and whether this promotes cognitive impairment. This study aimed to quantify neuroinflammatory and cognitive changes caused by administration of angiotensin II, and determine whether this is blood pressure-dependent. Design and method: Male C57Bl/6 mice were administered vehicle (saline; n = 33) or angiotensin II (0.7 mg/kg/d, n = 34) for 14 d via an osmotic minipump. A subset of mice also received hydralazine hydrochloride (50 mg/kg; n = 38) in their drinking water for 14 d after minipump implantation. Systolic blood pressure (SBP) was measured using tail-cuff; inflammation was assessed using flow cytometry and RNA sequencing; and recognition memory was evaluated using the novel object recognition test. Results: Angiotensin II infusion increased SBP and promoted accumulation of neutrophils, monocytes, T cells and B cells in the brain compared to vehicle infusion which was blunted by co-administration of hydralazine (n = 7–8, P < 0.05). Angiotensin II infusion also impaired recognition memory and this effect could be prevented by co-treatment with hydralazine (n = 11–12, P < 0.05). Angiotensin II infusion promoted differential expression of 1194 genes in whole brain hemisphere and 634 genes in hippocampus compared to vehicle infusion. Out of these genes, 295 in whole brain hemisphere and 125 in hippocampus were inflammation/immune-related (n = 6, P-adjusted < 0.05). Angiotensin II caused downregulation of genes involved in cognitive function and memory consolidation (Dusp1, Arc) and upregulation of genes involved in immune cell function, regulation and migration (H2-Ab1, H2-Q6, Il17rc, Fcrlb, Ltb, Ccl8, Itga9) in whole brain hemisphere (n = 6, P-adjusted < 0.05). In the hippocampus, a key region in learning and memory, angiotensin II promoted upregulation of genes involved in immune cell proliferation and differentiation (Il12rb2, Acvr1b, Sema3c) and apoptosis (Plxna3, Anxa11, Hyal1, Axin1) (n = 6, P-adjusted < 0.05). Co-treatment with hydralazine altered expression of all of these genes suggesting possible reversal of the transcriptomic changes induced by angiotensin II infusion (n = 6, P-adjusted < 0.05). Conclusion: Angiotensin II promotes neuroinflammation and cognitive impairment in a blood pressure-dependent manner. Co-treatment with hydralazine may reverse some of the transcriptomic changes induced by angiotensin II. Chronic neuroinflammation may be a contributing factor to cognitive impairment during hypertension and could be modulated by blood pressure reduction.

Estimation of Central Systolic Blood Pressure from Peripheral Pressure Waves using a Novel Second Systolic Pressure-Based Method in Normal and Heritable Hypercholesterolemic Rabbits.

Central systolic blood pressure (cSBP) was closely related to hypertension-related organ damage rather than peripheral systolic blood pressure (pSBP). We aimed to estimate cSBP from pSBP without generalized transfer function in normal and Kurosawa and Kusanagi-hypercholesterolemic (KHC) rabbits aged 12 months. Two catheter-tip transducers were advanced into the ascending aorta (AA) and distal end of the right brachial artery (Br) through the right common carotid and right radial arteries, respectively, under pentobarbital anesthesia. Pressure waves in response to the intravenous administration of angiotensin II and sodium nitroprusside were simultaneously recorded in AA and Br under regular cardiac pacing. The first (pSBP) and second peaks (pSBP2) of the brachial blood pressure and their average (pSBPm) were significantly correlated with cSBP, despite Murgo's wave pattern of central pressure waves in both rabbit groups. In Bland-Altman plot and its modification as a function of the peripheral augmentation index (pAI) analyses, the differences between pSBP and cSBP decreased, and those between pSBP2 and cSBP increased significantly in their average- or pAI-dependent manner, with undeniable mean biases in both rabbit groups. When the same analyses for SBPm were performed instead, the mean bias was around zero, with reduced variance in the two rabbit groups. The observed pressure or pAI-dependent systematic biases for pSBP and pSBP2 disappeared, representing the precise feature of pSBPm as a cSBP estimate. We conclude that pSBPm could be more precise than pSBP2 as a cSBP estimate, irrespective of blood pressure levels, pAI, or the presence of atherosclerosis.

Combined Administration of Andrographolide and Angiotensin- (1-7) Synergically Increases the Muscle Function and Strength in Aged Mice

Sarcopenia is a progressive and generalized skeletal muscle disorder characterized by muscle weakness, loss of muscle mass, and decline in the capacity of force generation. Aging can cause sarcopenia. Several therapeutic strategies have been evaluated to prevent or alleviate this disorder. One of them is angiotensin 1-7 [Ang-(1-7)], an anti-atrophic peptide for skeletal muscles that regulates decreased muscle mass for several causes, including aging. Another regulator of muscle mass and function is andrographolide, a bicyclic diterpenoid lactone that decreases the nuclear factor kappa B (NF-κB) signaling and attenuates the severity of some muscle diseases. Evaluate the effect of combined administration of Ang-(1-7) with andrographolide on the physical performance, muscle strength, and fiber´s diameter in a murine model of sarcopenia by aging. Aged male mice of the C57BL/6J strain were treated with Andrographolide, Ang-(1-7), or combined for three months. The physical performance, muscle strength, and fiber´s diameter were measured. The results showed that aged mice (24 months old) treated with Ang-(1-7) or Andrographolide improved their performance on a treadmill test, muscle strength, and their fiber´s diameter compared to aged mice without treatment. The combined administration of Ang-(1-7) with andrographolide to aged mice has an enhanced synergically effect on physical performance, muscle strength, and fiber´s diameter. Our results indicated that in aged mice, the effects of andrographolide and Ang-(1-7) on muscle function, strength, and fiber´s diameter are potentiated.

Angiotensin-converting enzyme 2 improves hepatic insulin resistance by regulating GABAergic signaling in the liver

The angiotensin-converting enzyme 2 (ACE2)/angiotensin 1-7/MAS axis and the gamma-aminobutyric acid (GABA)ergic signaling system have both been shown to have the dual potential to improve insulin resistance (IR) and hepatic steatosis associated with obesity in the liver. Recent studies have demonstrated that ACE2 can regulate the GABA signal in various tissues. Notwithstanding this evidence, the functional relationship between ACE2 and GABA signal in the liver under IR remains elusive. Here, we used high-fat diet-induced models of IR in C57BL/6 mice as well as ACE2KO and adeno-associated virus-mediated ACE2 overexpression mouse models to address this knowledge gap. Our analysis showed that glutamate decarboxylase (GAD)67/GABA signaling was weakened in the liver during IR, whereas the expression of GAD67 and GABA decreased significantly in ACE2KO mice. Furthermore, exogenous administration of angiotensin 1-7 and adeno-associated virus- or lentivirus-mediated overexpression of ACE2 significantly increased hepatic GABA signaling in models of IR both invivo and invitro. We found that this treatment prevented lipid accumulation and promoted fatty acid β oxidation in hepatocytes as well as inhibited the expression of gluconeogenesis- and inflammation-related genes, which could be reversed by allylglycine, a specific GAD67 inhibitor. Collectively, our findings show that signaling via the ACE2/A1-7/MAS axis can improve hepatic IR by regulating hepatic GABA signaling. We propose that this research might indicate a potential strategy for the management of diabetes.

Role of endothelial dysfunction in the severity of COVID‑19 infection (Review).

COVID-19 patients with severe infection have been observed to have elevated auto-antibodies (AAs) against angiotensin II receptor type 1 (AT1R) and endothelin (ET) 1 receptor type A (ETAR), compared with healthy controls and patients with favorable (mild) infection. AT1R and ETAR are G protein-coupled receptors, located on vascular smooth muscle cells, fibroblasts, immune and endothelial cells, and are activated by angiotensin II (Ang II) and ET1 respectively. AAs that are specific for these receptors have a functional role similar to the natural ligands, but with a more prolonged vasoconstrictive effect. They also induce the production of fibroblast collagen, the release of reactive oxygen species and the secretion of proinflammatory cytokines (including IL-6, IL-8 and TNF-α) by immune cells. Despite the presence of AAs in severe COVID-19 infected patients, their contribution and implication in the severity of the disease is still not well understood and further studies are warranted. The present review described the major vascular homeostasis systems [ET and renin-angiotensin-aldosterone system (RAAS)], the vital regulative role of nitric oxide, the AAs, and finally the administration of angiotensin II receptor blockers (ARBs), so as to provide more insight into the interplay that exists among these components and their contribution to the severity, prognosis and possible treatment of COVID-19.

Angiotensin type 2 receptor antagonism as a new target to manage gout.

There is a growing search for therapeutic targets in the treatment of gout. The present study aimed to evaluate the analgesic and anti-inflammatory potential of angiotensin type 2 receptor (AT2R) antagonism in an acute gout attack mouse model. Male wild-type (WT) C57BL/6 mice either with the AT2R antagonist, PD123319 (10pmol/joint), or with vehicle injections, or AT2R KO mice, received intra-articular (IA) injection of monosodium urate (MSU) crystals (100µg/joint), that induce the acute gout attack, and were tested for mechanical allodynia, thermal hyperalgesia, spontaneous nociception and ankle edema development at several times after the injections. To test an involvement of AT2R in joint pain, mice received an IA administration of angiotensin II (0.05-5nmol/joint) with or without PD123319, and were also evaluated for pain and edema development. Ankle joint tissue samples from mice undergoing the above treatments were assessed for myeloperoxidase activity, IL-1β release, mRNA expression analyses and nitrite/nitrate levels, 4h after injections. AT2R antagonism has robust antinociceptive effects on mechanical allodynia (44% reduction) and spontaneous nociception (56%), as well as anti-inflammatory effects preventing edema formation (45%), reducing myeloperoxidase activity (54%) and IL-1β levels (32%). Additionally, Agtr2tm1a mutant mice have largely reduced painful signs of gout. Angiotensin II administration causes pain and inflammation, which was prevented by AT2R antagonism, as observed in mechanical allodynia 4h (100%), spontaneous nociception (46%), cold nociceptive response (54%), edema formation (83%), myeloperoxidase activity (48%), and IL-1β levels (89%). PD123319 treatment also reduces NO concentrations (74%) and AT2R mRNA levels in comparison with MSU untreated mice. Our findings show that AT2R activation contributes to acute pain in experimental mouse models of gout. Therefore, the antagonism of AT2R may be a potential therapeutic option to manage gout arthritis.

Effect of angiotensin II pathway inhibitors on post-surgical adhesion band formation: a potential repurposing of old drugs

BackgroundIn this study we investigated the therapeutic potential of angiotensin II pathway inhibitors in attenuating post-surgical adhesion band formation in tendon injury. MethodWe assigned 30 Wistar albino rats to 5 groups, including negative control, positive control, sham, Telmisartan- and Enalapril-treated groups (n=6). Telmisartan and Enalapril at a dose of 10 mg/kg were administered intraperitoneally for 21 days. Hematoxylin-Eosin, and Masson's trichrome staining were used to measure the inflammatory cell accumulation and collagen deposition in the Achilles tendon tissue sections. Oxidative stress markers were analyzed in tissue samples by spectrophotometric methods. Properties of Achilles tendon adhesions were compared based on Tang and Ishiyama scoring systems in the presence and absence of angiotensin II pathway inhibitors. ResultsTelmisartan and Enalapril reduced severity, length, and density of surgical-induced tendon adhesion at site of injury (***p < 0.001). Our results showed that administration of angiotensin II pathway inhibitors decreased infiltration of inflammatory cells to the injured area (*p < 0.05) and suppressed inflammation by regulating oxidative stress markers including MDA (***p < 0.001), total thiol (***p < 0.001), CAT (***p < 0.001), and SOD (***p < 0.001), in post-operative Achilles tendon tissues. Significant lower collagen deposition and formation of fibrotic tissues was seen in Telmisartan- and Enalapril-treated groups as detected by Masson's trichrome staining which correlated with a decrease in quantity (**p < 0.01) and grading of fibrosis score (***p < 0.001), in adhesive tissues. Moreover, inhibition of angiotensin II pathway could also ameliorate mechanical properties including ultimate load (***p < 0.001), and ultimate stress (*p < 0.05) in injured Tendons. ConclusionOur results showed that ssuppression of inflammation and fibrosis are two mechanisms by which Telmisartan and Enalapril elicit potent protective responses post Achilles tendon injuries.

Unveiling the Angiotensin-(1-7) Actions on the Urinary Bladder in Female Rats.

Angiotensin-(1–7) is a peptide produced by different pathways, and regardless of the route, the angiotensin-converting enzyme 2 (ACE-2) is involved in one of the steps of its synthesis. Angiotensin-(1–7) binds to Mas receptors localized in different cells throughout the body. Whether angiotensin-(1–7) exerts any action in the urinary bladder (UB) is still unknown. We investigated the effects of intravenous and topical (in situ) administration of angiotensin-(1–7) on intravesical pressure (IP) and cardiovascular variables. In addition, the Mas receptors and ACE-2 gene and protein expression were analyzed in the UB. Adult female Wistar rats were anesthetized with 2% isoflurane in 100% O2 and submitted to the catheterization of the femoral artery and vein for mean arterial pressure (MAP) and heart rate (HR) recordings, and infusion of drugs, respectively. The renal blood flow was acquired using a Doppler flow probe placed around the left renal artery and the renal conductance (RC) was calculated as a ratio of Doppler shift (kHz) and MAP. The cannulation of the UB was performed for IP recording. We observed that angiotensin-(1–7) either administered intravenously [115.8 ± 28.6% angiotensin-(1–7) vs. −2.9 ± 1.3% saline] or topically [147.4 ± 18.9% angiotensin-(1–7) vs. 3.2 ± 2.8% saline] onto the UB evoked a significant (p < 0.05) increase in IP compared to saline and yielded no changes in MAP, HR, and RC. The marked response of angiotensin-(1–7) on the UB was also investigated using quantitative real-time polymerase chain reaction and western blotting assay, which demonstrated the mRNA and protein expression of Mas receptors in the bladder, respectively. ACE-2 mRNA and protein expression was also observed in the bladder. Therefore, the findings demonstrate that angiotensin-(1–7) acts in the UB to increase the IP and suggest that this peptide can be also locally synthesized in the UB.

The Role of Angiotensin 1‐7 in Isolated Human Arterioles with SARS‐CoV‐2

IntroductionThe vascular endothelium plays a crucial role to regulate vascular tone. Recently, our laboratory reported severe acute respiratory syndrome coronavirus‐2 (SARS‐CoV‐2) infection significantly reduced endothelial‐dependent vasodilation months after exposed to SARS‐CoV‐2 in human microvessels. SARS‐CoV‐2 enters host cells through angiotensin‐converting enzyme (ACE)2 as a cell surface receptor. Downregulation of ACE2 protein expression by the virus leads to imbalance between angiotensin (ANG) II and ANG 1‐7, causing vasoconstriction, inflammation and increase oxidative stress. With this background, we intend to test our new hypothesis that administration of ANG 1‐7 will improve vasodilation in human arterioles from previously SARS‐CoV‐2 positive subjects.MethodsFresh human tissues were obtained as de‐identified surgical discarded specimens. Control subjects (n=9) were SARS‐CoV‐2 negative and without known virus exposure. We had 11 previously SARS‐CoV‐2 positive subjects or post‐COVID. For all post‐COVID patients, a negative COVID test was obtained prior to collection of surgical specimens. We isolated arterioles (100‐200 µm) and cannulated onto glass micropipettes under 60 mmHg and examined for diameter changes to the endothelial‐dependent vasodilator acetylcholine (ACh; 10−9 to 10−5 M) using videomicroscopy. Flow‐mediated dilation (FMD) was recorded at steady‐state during varying intraluminal pressure gradients (5‐100 cm H2O). All post‐COVID vessels were incubated in a culture media at 37°C for 12–15 hours with or without 1 nM of ANG 1‐7.ResultsDilation to ACh and FMD in arterioles from post‐COVID was significantly reduced (ACh max. dilation at 10−5 M: 74±6% vs. 93±4% in control, n=7‐9, P<0.001; FMD max. dilation at 100 cm H2O: 49±3% vs. 86±2% in control, n=6‐11, P<0.001) while overnight incubation of 1 nM ANG 1‐7 significantly improved the dilation (ACh max. dilation at 10−5 M: 74±6% in post‐COVID vs. 88±5% in post‐COVID + AND 1‐7, n=11, P<0.001; FMD max. dilation at 100 cm H2O: 49±3% in post‐COVID vs. 76±3% in post‐COVID + ANG 1‐7, n=11, P<0.001).ConclusionAdministration of ANG 1‐7 significantly improved vasodilation in isolated arterioles from post‐COVID subjects.

Angiotensin(1-7) Improves Islet Function in Diabetes Through Reducing JNK/Caspase-3 Signaling.

The aim of this study is to investigate whether Angiotensin (1-7), the physiological antagonist of Angiotensin II (AngII), has antidiabetic activity and the possible mechanism. Male Wistar rats were randomly divided into 3 groups: control group fed the normal diet, DM group fed high-fat diet and injected with STZ, and Angiotensin (1-7) group receiving injection of STZ followed by Angiotensin (1-7) treatment. Serum Ang II, fasting blood glucose, insulin, HOMA-IR, and HOMA-beta were determined in control, diabetes and Angiotensin (1-7) groups. The increased AngII and insulin resistance in diabetes group were accompanied by changes in islet histopathology. However, Angiotensin (1-7) improved the islet function and histopathology in diabetes without affecting the level of AngII. Western blot confirmed that Angiotensin (1-7) decreased the cleaved caspase 3 levels in pancreas of DM. The increased expression of JNK, Bax, and Bcl2 genes under diabetic conditions were partially reversed after Angiotensin (1-7) administration in pancreas. Immunofluorescence analysis showed that p-JNK was markedly increased in islet of DM rats, which was markedly alleviated after Angiotensin (1-7) treatment. Furthermore, Angiotensin (1-7) reversed high glucose(HG) induced mitochondrial apoptosis augments. Finally, Angiotensin (1-7) attenuated the apoptosis of INS-1 cells through reducing JNK activation in diabetes, which was blocked by anisomycin (a potent agonist of JNK). Our findings provide supporting evidence that Angiotensin (1-7) improved the islet beta-cells apoptosis by JNK-mediated mitochondrial dysfunction, which might be a novel target for the treatment and prevention of beta-cells dysfunction in DM.

Combinational Therapy with Aspirin and Ticagrelor Alleviates Vascular Inflammation and Angiotensin IIdriven Abdominal Aortic Aneurysm Formation in Mice

Background: Abdominal aortic aneurysm (AAA) is a severe form of blood vessel-related disease. Medial degeneration and inflammation are typical characteristic of AAA. Activated platelets release many pro-inflammatory cytokines and participate in the initial inflammatory response to various vascular diseases. Although there are some studies on the effects of APAs on AAA, it is yet unknown whether the new P2Y12 receptor inhibitor ticagrelor (T) can inhibit AAA. Herein, we explored the consequences of ticagrelor exposure on AAA progression and determined whether a combinational therapy, involving T and aspirin (A), exerts a stronger inhibitory effect in vivo. Methods: AAA was established in apolipoprotein E-deficient (ApoE-/- ) mice via a 28-day administration of angiotensin II (Ang II). Next, the mice were arbitrarily separated into 5 groups: saline infusion alone (sham), Ang II infusion alone, Ang II infusion plus oral A (10 mg•kg-1•d-1), Ang II infusion plus oral T (120 mg.kg-1.d-1), Ang II infusion plus combinational therapy with A (10 mg•kg-1•d-1) and T (120 mg.kg-1.d-1). Results: The combined treatment markedly suppressed the Ang II-driven elevation of maximal aortic diameter, aneurysm formation (26.7% decrease, P&lt;0.05), alterations in aortic expansion, elastic lamina destruction, platelet deposition, and inflammatory cytokine accumulation. In addition, it also diminished matrix metalloproteinase (MMP)-2 and MMP-9 production. Conclusions: A combinational therapy of A and T, but not individual drugs, inhibits Ang II-driven AAA generation in mice in vivo, and this process may be regulated by a suppressed inflammatory response.

Inhibition of XIST attenuates abdominal aortic aneurysm in mice by regulating apoptosis of vascular smooth muscle cells through miR-762/MAP2K4 axis.

Abdominal aortic aneurysm (AAA) is a common chronic aortic degenerative disease. Long non-coding RNA X-inactive specific transcript (XIST) is associated with the progression of AAA, while the underlying mechanism is still unclear. We investigated the functional role of XIST in AAA. AAA mouse model was established by administration of Angiotensin II (Ang II). Primary mouse vascular smooth muscle cells (VSMCs) were separated from the abdominal aorta of Ang II-induced AAA mice, and then treated with Ang II. XIST was highly expressed in Ang II-treated VSMCs. Cell proliferation ability was decreased and apoptosis was increased in VSMCs following Ang II treatment. XIST knockdown reversed the impact of Ang II on cell proliferation and apoptosis in VSMCs. XIST promoted mitogen-activated protein kinase kinase 4 (MAP2K4) expression by sponging miR-762. XIST overexpression suppressed cell proliferation and apoptosis of Ang II-treated VSMCs by regulating miR-762/MAP2K4 axis. Finally, Ang II-induced AAA mouse model was established to verify the function of XIST in AAA. Inhibition of XIST significantly attenuated the pathological changes of abdominal aorta tissues in Ang II-induced mice. The expression of miR-762 was inhibited, and MAP2K4 expression was enhanced by XIST knockdown in the abdominal aorta tissues of AAA mice. In conclusion, these data demonstrate that inhibition of XIST attenuates AAA in mice, which attributes to inhibit apoptosis of VSMCs by regulating miR-762/MAP2K4 axis. Thus, this study highlights a novel ceRNA circuitry involving key regulators in the pathogenesis of AAA.

Platelet protects angiotensin II-driven abdominal aortic aneurysm formation through inhibition of inflammation.

Inflammation is the primary cause of abdominal aortic aneurysm (AAA) formation and development. It has been reported that platelets protect against septic shock by inhibiting inflammation. However, it is unclear whether platelets protect AAA progress via suppressing inflammation. A mouse model of AAA was established by a daily administration of angiotensin II (Ang II, 1000ng/kg/min) for 28-day. The AAA mice received 1×109 platelets transfusion in normal saline every 3rd day for 1month. Hematoxylin and eosin, Masson's trichrome, and elastic van Gieson staining techniques were used to analyze the morphology of the abdominal aorta. Immunohistochemistry was used to detect any infiltration of inflammatory cells, inflammatory factors, and matrix metalloproteins (MMPs) in the aortic tissue. Western blot and enzyme-linked immunosorbent assay (ELISA) were used to examine the inflammatory factor proteins levels in the aortic wall and peripheral blood, respectively. Platelets infusion significantly suppressed the Ang II-driven elevation of aortic diameter, AAA formation (52.38% decrease, P<0.05), aortic expansion, elastic lamina destruction, and inflammatory response. In addition, MMP-2 and MMP-9 production were also reduced by platelets transfusion. For the first time, our study reported the beneficial effect of platelet transfusion in suppressing the Ang II-driven AAA progress in mice through the inhibition of inflammation.

Agave tequilana Counteracts Chronic Hypertension and Associated Vascular Damage.

Systemic arterial hypertension (SAH) is a health problem of great importance worldwide, and endothelial dysfunction underlies SAH development. This condition's main characteristics include vasoconstriction, inflammation, oxidative stress, and procoagulant and proliferative states. This study's objective was to evaluate the antihypertensive, anti-inflammatory, and antioxidant effects of the whole extract and fractions of Agave tequilana in a murine model of SAH. SAH was induced in male ICR or CD-1 (Strain obtained from animals from Charles River Laboratories, Massachusetts) mice by intraperitoneal administration of angiotensin II (AGII) (0.1 μg/kg) for 4 weeks, and then A. tequilana treatments were co-administered with AGII. At the end of the experiment, systolic and diastolic blood pressure were measured and the kidneys were dissected to quantify interleukin (IL)-1β, IL-6, tumor necrosis factor-alpha, IL-10, and malondialdehyde (MDA). The whole extract and the fractions of A. tequilana were chemically characterized using gas chromatography-mass spectrometry. The results indicate that the whole extract (At-W) and At-AcOEt fraction treatment are the most efficient in lowering blood pressure, although all the treatments had an immunomodulatory effect on the cytokines evaluated and an antioxidant effect on lipid peroxidation. Finally, the chromatographic profile shows that the integral extract and fractions of A. tequilana contained phytol (M)3,7,11,15-Tetramethyl-2-hexadecen-1-ol; 9,12-octadecadienoic acid; hentriacontane; 9,19-cyclolanost-24-en-3-ol,(3b); t-sitosterol; and stigmasta-3,5-dien-7-one.

IgA Nephropathy with Macroproteinuria and a GFR of 20-30 ml/min/1.73 m2 May Still Benefit from RAS Inhibition.

There has been controversy about renin-angiotensin system (RAS) inhibition in IgAN patients with advanced (stage 4) chronic kidney disease (CKD). Therefore, we investigated the effect of RAS blockade in these patients. Renal specimens of 50 IgAN patients who underwent renal biopsy during stage 4 CKD between 2010 and 2020, were stained using immunohistochemistry to detect the expression of RAS receptors (AT1R, AT2R, MasR, and MrgD). The primary endpoint was a composite of end-stage renal disease (ESRD) and death. Main baseline information and the administration of angiotensin-converting enzyme inhibitor (ACEI) or angiotensin II receptor blocker (ARB) were collected. During a median follow-up time of 25.5 months, 21 (42.0%) patients reached ESRD and none died. Six patients had a baseline eGFR of 15-20 ml/min/1.73m2, and reached ESRD with a median renal survival time of 7.0 (range 6.0-23.0) months. Among patients with a baseline eGFR of 20-30 ml/min/1.73m2, the percentage of patients using ACEI/ARB in progressive group was much lower than that in stable group (33.3% vs. 62.1%, P = 0.045), together with a shorter renal survival time in progressive group (26.0 vs. 30.5 months, P = 0.033). Macroproteinuria (24 h - UP ≥ 2.5 g) was also associated with a shorter renal survival time, as well as a significant decline in eGFR of stable group (24.4 vs. 26.4 ml/min/1.73 m2, P = 0.026). Lower eGFR [hazards ratio (HR), 0.829, 95% confidence interval (CI), 0.724-0.950; P = 0.007] and use of ACEI/ARB (HR, 0.356, 95% CI, 0.133-0.953; P = 0.040) were predictive of time to ESRD in this stage. No differences were found in the expression of AT1R, AT2R, MasR, and MrgD of renal tissues at the time of biopsy between stable and progressive groups. Contingent on monitoring serum creatinine and potassium levels, IgAN with macroproteinuria and a GFR of 20-30 ml/min/1.73m2 may still benefits from intrarenal RAS inhibition.