Adipocyte-Derived Serum Amyloid A Promotes Angiotensin II-Induced Abdominal Aortic Aneurysms in Obese C57BL/6J Mice.

BACKGROUND: Obesity increases the risk for human abdominal aortic aneurysms (AAAs) and enhances Ang II (angiotensin II)-induced AAA formation in C57BL/6J mice. Obesity is also associated with increases in perivascular fat that expresses proinflammatory markers including SAA (serum amyloid A). We previously reported that deficiency of SAA significantly reduces Ang II-induced inflammation and AAA in hyperlipidemic apoE-deficient mice. In this study. we investigated whether adipose tissue-derived SAA plays a role in Ang II-induced AAA in obese C57BL/6J mice. METHODS: The development of AAA was compared between male C57BL/6J mice (wild type), C57BL/6J mice lacking SAA1.1, SAA2.1, and SAA3 (TKO); and TKO mice harboring a doxycycline-inducible, adipocyte-specific SAA1.1 transgene (TKO-Tgfat; SAA expressed only in fat). All mice were fed an obesogenic diet and doxycycline to induce SAA transgene expression and infused with Ang II to induce AAA. RESULTS: In response to Ang II infusion, SAA expression was significantly increased in perivascular fat of obese C57BL/6J mice. Maximal luminal diameters of the abdominal aorta were determined by ultrasound before and after Ang II infusion, which indicated a significant increase in aortic luminal diameters in wild type and TKO-TGfat mice but not in TKO mice. Adipocyte-specific SAA expression was associated with MMP (matrix metalloproteinase) activity and macrophage infiltration in abdominal aortas of Ang II-infused obese mice. CONCLUSIONS: We demonstrate for the first time that SAA deficiency protects obese C57BL/6J mice from Ang II-induced AAA. SAA expression only in adipocytes is sufficient to cause AAA in obese mice infused with Ang II.

Therapeutic Assessment of Combination Therapy with a Neprilysin Inhibitor and Angiotensin Type 1 Receptor Antagonist on Angiotensin II-Induced Atherosclerosis, Abdominal Aortic Aneurysms, and Hypertension.

Combined neprilysin (NEP) inhibition (sacubitril) and angiotensin type 1 receptor (AT1R) antagonism (valsartan) is used in the treatment of congestive heart failure and is gaining interest for other angiotensin II (AngII)-related cardiovascular diseases. In addition to heart failure, AngII promotes hypertension, atherosclerosis, and abdominal aortic aneurysms (AAAs). Similarly, NEP substrates or products have broad effects on the cardiovascular system. In this study, we examined NEP inhibition (with sacubitril) and AT1R antagonism (with valsartan) alone or in combination on AngII-induced hypertension, atherosclerosis, or AAAs in male low-density lipoprotein receptor-deficient mice. Preliminary studies assessed drug delivery via osmotic minipumps for simultaneous release of sacubitril and/or valsartan with AngII over 28 days. Mice were infused with AngII (1000 ng/kg per minute) in the absence (vehicle) or presence of sacubitril (1, 6, or 9 mg/kg per day), valsartan (0.3, 0.5, 1, 6, or 20 mg/kg per day), or the combination thereof (1 and 0.3, or 9 or 0.5 mg/kg per day of sacubitril and valsartan, respectively). Plasma AngII and renin concentrations increased 4-fold at higher valsartan doses, indicative of removal of AngII negative feedback on renin. Sacubitril doubled plasma AngII concentrations at lower doses (1 mg/kg per day). Valsartan dose-dependently decreased systolic blood pressure, aortic atherosclerosis, and AAAs of AngII-infused mice, whereas sacubitril had no effect on atherosclerosis or AAAs but reduced blood pressure of AngII-infused mice. Combination therapy with sacubitril and valsartan did not provide additive benefits. These results suggest limited effects of combination therapy with NEP inhibition and AT1R antagonism against AngII-induced hypertension, atherosclerosis, or AAAs. SIGNIFICANCE STATEMENT: The combination of valsartan (angiotensin type 1 receptor antagonist) and sacubitril (neprilysin inhibitor) did not provide benefit above valsartan alone on AngII-induced hypertension, atherosclerosis, or abdominal aortic aneurysms in low-density lipoprotein receptor-deficient male mice. These results do not support this drug combination in therapy of these AngII-induced cardiovascular diseases.

Monosomy X in Female Mice Influences the Regional Formation and Augments the Severity of Angiotensin II-Induced Aortopathies.

OBJECTIVE: Turner syndrome women (monosomy X) have high risk of aortopathies consistent with a role for sex chromosomes in disease development. We demonstrated that sex chromosomes influence regional development of Ang II (angiotensin II)-induced aortopathies in mice. In this study, we determined if the number of X chromosomes regulates regional development of Ang II-induced aortopathies. Approach and Results: We used females with varying numbers of X chromosomes (XX female mice [XXF] or XO female mice [XOF]) on an C57BL/6J (ascending aortopathies) or low-density lipoprotein receptor deficient (Ldlr-/-) background (descending and abdominal aortopathies) compared with XY males (XYM). To induce aortopathies, mice were infused with Ang II. XOF (C57BL/6J) exhibited larger percent increases in ascending aortic lumen diameters than Ang II-infused XXF or XYM. Ang II-infused XOF (Ldlr-/-) exhibited similar incidences of thoracic (XOF, 50%; XYM, 71%) and abdominal aortopathies (XOF, 83%; XYM, 71%) as XYM, which were greater than XXF (XXF, 0%). Abdominal aortic lumen diameters and maximal external diameters were similar between XOF and XYM but greater than XXF, and these effects persisted with extended Ang II infusions. Larger aortic lumen diameters, abdominal aortopathy incidence (XXF, 20%; XOF, 75%), and maximal aneurysm diameters (XXF, 1.02±0.17; XOF, 1.96±0.32 mm; P=0.027) persisted in ovariectomized Ang II-infused XOF mice. Data from RNA-seq demonstrated that X chromosome genes that escape X-inactivation (histone lysine demethylases Kdm5c and Kdm6a) exhibited lower mRNA abundance in aortas of XOF than XXF (P=0.033 and 0.024, respectively). Conversely, DNA methylation was higher in aortas of XOF than XXF (P=0.038). CONCLUSIONS: The absence of a second X chromosome promotes diffuse Ang II-induced aortopathies in females.

Deletion of tetraspanin CD151 alters the Wnt oncogene-induced mammary tumorigenesis: A cell type-linked function and signaling.

Tetraspanin CD151 is increasingly implicated as a multifaceted mediator of cancer development and progression. Here we investigated the role of CD151 in breast cancer in the context of the Wnt oncogenic activation. Our data showed that removal of one or both of CD151 alleles in the MMTV-Wnt1 model significantly decreased the tumor-free survival of mice from 34 weeks on average to 22 weeks and 18 weeks, respectively. This effect coincided with an accelerated tumor growth and an increased number of Ki-67+ proliferative cells. Mechanistically, the CD151-deficient tumors were largely ER+, and exhibited hyperactivation of the Wnt pathway as reflected by a marked upregulation in ß-catenin and Cyclin D1, and their target genes. In addition, E-cadherin displayed a cytosolic distribution and transcription factor Snail was markedly upregulated. Collectively, this data implies that CD151 suppresses the Wnt1-driven tumorigenesis, at least in part, via counteracting the epithelial-mesenchymal transition (EMT)-like program in luminal epithelial cells. Meanwhile, the proportion of tumor cells expressing CK5 or p63, the biomarkers of myoepithelial/basal cells, markedly decreased in the absence of CD151. This change was accompanied by a decreased invasiveness of tumors and their incompetence to form a long-term cell culture. Consistent with this basal cell-linked role, the CD151 downregulation impairs mammosphere formation in MCF-10A cells and the defect was rescued by re-expression of intact CD151 ORF, but not its integrin binding-defective mutant. Overall, our study suggests that CD151 is a key player in the Wnt oncogene-driven tumorigenesis and impacts breast cancer malignancy in a cell type-dependent manner.

Adipocyte deficiency of ACE2 increases systolic blood pressures of obese female C57BL/6 mice.

BACKGROUND: Obesity increases the risk for hypertension in both sexes, but the prevalence of hypertension is lower in females than in males until menopause, despite a higher prevalence of obesity in females. We previously demonstrated that angiotensin-converting enzyme 2 (ACE2), which cleaves the vasoconstrictor, angiotensin II (AngII), to generate the vasodilator, angiotensin-(1-7) (Ang-(1-7)), contributes to sex differences in obesity-hypertension. ACE2 expression in adipose tissue was influenced by obesity in a sex-specific manner, with elevated ACE2 expression in obese female mice. Moreover, estrogen stimulated adipose ACE2 expression and reduced obesity-hypertension in females. In this study, we hypothesized that deficiency of adipocyte ACE2 contributes to obesity-hypertension of females. METHODS: We generated a mouse model of adipocyte ACE2 deficiency. Male and female mice with adipocyte ACE2 deficiency or littermate controls were fed a low (LF) or a high fat (HF) diet for 16 weeks and blood pressure was quantified by radiotelemetry. HF-fed mice of each sex and genotype were challenged by an acute AngII injection, and blood pressure response was quantified. To translate these findings to humans, we performed a proof-of-principle study in obese transwomen in which systemic angiotensin peptides and blood pressure were quantified prior to and after 12 weeks of gender-affirming 17ß-estradiol hormone therapy. RESULTS: Adipocyte ACE2 deficiency had no effect on the development of obesity in either sex. HF feeding increased systolic blood pressures (SBP) of wild-type male and female mice compared to LF-fed controls. Adipocyte ACE2 deficiency augmented obesity-induced elevations in SBP in females, but not in males. Obese female, but not obese male mice with adipocyte ACE2 deficiency, had an augmented SBP response to acute AngII challenge. In humans, plasma 17ß-estradiol concentrations increased in obese transwomen administered 17ß-estradiol and correlated positively with plasma Ang-(1-7)/AngII balance, and negatively to SBP after 12 weeks of 17ß-estradiol administration. CONCLUSIONS: Adipocyte ACE2 protects female mice from obesity-hypertension, and reduces the blood pressure response to systemic AngII. In obese transwomen undergoing gender-affirming hormone therapy, 17ß-estradiol administration may regulate blood pressure via the Ang-(1-7)/AngII balance.

Effects of Aryl Hydrocarbon Receptor Deficiency on PCB-77-Induced Impairment of Glucose Homeostasis during Weight Loss in Male and Female Obese Mice.

BACKGROUND: Lipophilic polychlorinated biphenyls (PCBs) accumulate with obesity, but during weight loss, liberated PCBs act as ligands of the aryl hydrocarbon receptor (AhR) to negatively influence health. Previous studies demonstrated that PCB-77 administration to obese male mice impaired glucose tolerance during weight loss. Recent studies indicate higher toxic equivalencies of dioxin-like PCBs in exposed females than males. OBJECTIVES: We compared effects of PCB-77 on weight gain or loss and glucose homeostasis in male vs. female mice. We defined effects of AhR deficiency during weight gain or loss in male and female mice exposed to PCB-77. METHODS: Study design was vehicle (VEH) or PCB-77 administration while fed a high-fat (HF) diet for 12 wk, followed by weight loss for 4 wk. The following groups were examined: male and female C57BL/6 mice administered VEH or PCB-77, female [Formula: see text] and [Formula: see text] mice administered VEH or PCB-77, and male [Formula: see text] and [Formula: see text] mice administered PCB-77. Glucose tolerance was quantified during weight gain (week 11) and loss (week 15); liver and adipose AhR and IRS2 (insulin receptor substrate 2) mRNA abundance, and PCB-77 concentrations were quantified at week 16. RESULTS: PCB-77 attenuated development of obesity in females but not males. During weight loss, PCB-77 impaired glucose tolerance of males. AhR-deficient females (VEH) were resistant to diet-induced obesity. Compared with VEH-treated mice, HF-fed [Formula: see text] females treated with PCB-77 has less weight gain, and [Formula: see text] females had greater weight gain. During weight loss, [Formula: see text] females but not [Formula: see text] males treated with PCB-77 exhibited impaired glucose tolerance. In [Formula: see text] females administered PCB-77, IRS2 mRNA abundance was lower in adipose tissue compared with VEH-treated mice. CONCLUSION: Male and female mice responded differently to PCB-77 and AhR deficiency in body weight (BW) regulation and glucose homeostasis. AhR deficiency reversed PCB-77-induced glucose impairment of obese males losing weight but augmented glucose intolerance of females. These results demonstrate sex differences in PCB-77-induced regulation of glucose homeostasis of mice. https://doi.org/10.1289/EHP4133.

XX sex chromosome complement promotes atherosclerosis in mice.

Men and women differ in circulating lipids and coronary artery disease (CAD). While sex hormones such as estrogens decrease CAD risk, hormone replacement therapy increases risk. Biological sex is determined by sex hormones and chromosomes, but effects of sex chromosomes on circulating lipids and atherosclerosis are unknown. Here, we use mouse models to separate effects of sex chromosomes and hormones on atherosclerosis, circulating lipids and intestinal fat metabolism. We assess atherosclerosis in multiple models and experimental paradigms that distinguish effects of sex chromosomes, and male or female gonads. Pro-atherogenic lipids and atherosclerosis are greater in XX than XY mice, indicating a primary effect of sex chromosomes. Small intestine expression of enzymes involved in lipid absorption and chylomicron assembly are greater in XX male and female mice with higher intestinal lipids. Together, our results show that an XX sex chromosome complement promotes the bioavailability of dietary fat to accelerate atherosclerosis.

Pseudomonas aeruginosa-derived pyocyanin reduces adipocyte differentiation, body weight, and fat mass as mechanisms contributing to septic cachexia.

Pseudomonas aeruginosa, a leading cause of sepsis, produces pyocyanin, a blue-pigmented virulence factor. Sepsis is associated with cachexia, but mechanisms are unknown and conventional nutrition approaches are not effective treatments. Pyocyanin has affinity for the aryl hydrocarbon receptor (AhR), which is expressed on adipocytes and regulates adipocyte differentiation. The purpose of this study was to define in vitro and in vivo effects of pyocyanin on adipocyte differentiation and body weight regulation as relates to septic cachexia. In 3T3-L1 preadipocytes, pyocyanin activated AhR and its downstream marker CYP1a1, and reduced differentiation. Administration of pyocyanin to male C57BL/6J mice acutely reduced body temperature with altered locomotion, but caused sustained weight loss. Chronic pyocyanin administration to male and female C57BL/6J mice resulted in sustained reductions in body weight and fat mass, with adipose-specific AhR activation. Pyocyanin-treated male mice had decreased energy expenditure and physical activity, and increased adipose explant lipolysis. In females, pyocyanin caused robust reductions in body weight, adipose-specific AhR activation, and increased expression of inflammatory cytokines in differentiated adipocytes. These results demonstrate that pyocyanin reduces adipocyte differentiation and decreases body weight and fat mass in male and female mice, suggesting that pyocyanin may play a role in septic cachexia.

Mas receptor deficiency augments angiotensin II-induced atherosclerosis and aortic aneurysm ruptures in hypercholesterolemic male mice.

OBJECTIVE: Previous studies demonstrated that deficiency of angiotensin-converting enzyme 2 (ACE2) augmented angiotensin II (AngII)-induced atherosclerosis and abdominal aortic aneurysm (AAA) formation in hypercholesterolemic mice. Effects of ACE2 deficiency could arise from increased concentrations of its substrate, AngII, or decreased concentrations of its product, angiotensin-(1-7) [Ang-(1-7)]. Infusion of Ang-(1-7), a Mas receptor (MasR) ligand, to hypercholesterolemic male mice reduced AngII-induced atherosclerosis, suggesting a protective role of the Ang-(1-7)/MasR axis. However, it is unclear whether endogenous Ang-(1-7) acts at MasR to influence AngII-induced vascular diseases. The purpose of this study was to define the role of MasR deficiency in AngII-induced atherosclerosis and AAA formation and severity in hypercholesterolemic male mice. METHODS: MasR+/+ and MasR-/- male mice on a low-density lipoprotein receptor-deficient (Ldlr-/-) or apolipoprotein E-deficient (Apoe-/-) background were infused with AngII at either 600 or 1000 ng/kg/min by osmotic minipump for 28 days. Atherosclerosis was quantified at study end point as percentage lesion surface area of the aortic arch in Ldlr-/- mice. Abdominal aortic internal diameters were quantified by ultrasound, and maximal external AAA diameters were quantified at study end point. Blood pressure was quantified by radiotelemetry and a tail cuff-based technique. Serum cholesterol concentrations and vascular tissue characterization were examined at study end point. RESULTS: MasR deficiency did not influence body weight, systolic blood pressure at baseline and during AngII infusion, or serum cholesterol concentrations in either Apoe-/- or Ldlr-/- mice. MasR deficiency increased AngII-induced atherosclerosis in aortic arches of Ldlr-/- mice (P < .05), associated with increased oxidative stress and apoptosis in aortic root sections (P < .05). MasR deficiency also augmented internal and external AAA diameters and increased aortic ruptures of both Ldlr-/- and Apoe-/- mice (P < .05). These effects were associated with increased elastin breaks and T-lymphocyte and macrophage accumulation into abdominal aortas of AngII-infused MasR-deficient mice (P < .05). CONCLUSIONS: These results demonstrate that MasR deficiency augmented AngII-induced atherosclerosis and AAA rupture through mechanisms involving increased oxidative stress, inflammation, and apoptosis, suggesting that MasR activation may provide therapeutic efficacy against vascular diseases.

IKKß is a ß-catenin kinase that regulates mesenchymal stem cell differentiation.

Mesenchymal stem cells (MSCs) can give rise to both adipocytes and osteoblasts, but the molecular mechanisms underlying MSC fate determination remain poorly understood. IκB kinase ß (IKKß), a central coordinator of inflammation and immune responses through activation of NF-κB, has been implicated as a critical molecular link between obesity and metabolic disorders. Here, we show that IKKß can reciprocally regulate adipocyte and osteoblast differentiation of murine and human MSCs through an NF-κB-independent mechanism. IKKß is a ß-catenin kinase that phosphorylates the conserved degron motif of ß-catenin to prime it for ß-TrCP-mediated ubiquitination and degradation, thereby increasing adipogenesis and inhibiting osteogenesis in MSCs. Animal studies demonstrated that deficiency of IKKß in BM mesenchymal stromal cells increased bone mass and decreased BM adipocyte formation in adult mice. In humans, IKKß expression in adipose tissue was also positively associated with increased adiposity and elevated ß-catenin phosphorylation. These findings suggest IKKß as a key molecular switch that regulates MSC fate, and they provide potentially novel mechanistic insights into the understanding of the cross-regulation between the evolutionarily conserved IKKß and Wnt/ß-catenin signaling pathways. The IKKß-Wnt axis we uncovered may also have important implications for development, homeostasis, and disease pathogenesis.

Sex Chromosome Complement Defines Diffuse Versus Focal Angiotensin II-Induced Aortic Pathology.

OBJECTIVE: Aortic pathologies exhibit sexual dimorphism, with aneurysms in both the thoracic and abdominal aorta (ie, abdominal aortic aneurysm [AAA]) exhibiting higher male prevalence. Women have lower prevalence of aneurysms, but when they occur, aneurysms progress rapidly. To define mechanisms for these sex differences, we determined the role of sex chromosome complement and testosterone on the location and progression of angiotensin II (AngII)-induced aortic pathologies. APPROACH AND RESULTS: We used transgenic male mice expressing Sry (sex-determining region Y) on an autosome to create Ldlr (low-density lipoprotein receptor)-deficient male mice with an XY or XX sex chromosome complement. Transcriptional profiling was performed on abdominal aortas from XY or XX males, demonstrating 1746 genes influenced by sex chromosomes or sex hormones. Males (XY or XX) were either sham-operated or orchiectomized before AngII infusions. Diffuse aortic aneurysm pathology developed in XY AngII-infused males, whereas XX males developed focal AAAs. Castration reduced all AngII-induced aortic pathologies in XY and XX males. Thoracic aortas from AngII-infused XY males exhibited adventitial thickening that was not present in XX males. We infused male XY and XX mice with either saline or AngII and quantified mRNA abundance of key genes in both thoracic and abdominal aortas. Regional differences in mRNA abundance existed before AngII infusions, which were differentially influenced by AngII between genotypes. Prolonged AngII infusions resulted in aortic wall thickening of AAAs from XY males, whereas XX males had dilated focal AAAs. CONCLUSIONS: An XY sex chromosome complement mediates diffuse aortic pathology, whereas an XX sex chromosome complement contributes to focal AngII-induced AAAs.

A Brief Introduction into the Renin-Angiotensin-Aldosterone System: New and Old Techniques.

The renin-angiotensin-aldosterone system (RAAS) is a complex system of enzymes, receptors, and peptides that help to control blood pressure and fluid homeostasis. Techniques in studying the RAAS can be difficult due to such factors as peptide/enzyme stability and receptor localization. This paper gives a brief account of the different components of the RAAS and current methods in measuring each component. There is also a discussion of different methods in measuring stem and immune cells by flow cytometry, hypertension, atherosclerosis, oxidative stress, energy balance, and other RAAS-activated phenotypes. While studies on the RAAS have been performed for over 100 years, new techniques have allowed scientists to come up with new insights into this system. These techniques are detailed in this Methods in Molecular Biology Series and give students new to studying the RAAS the proper controls and technical details needed to perform each procedure.

Measuring Blood Pressure Using a Noninvasive Tail Cuff Method in Mice.

The renin angiotensin system (RAS) is well known for its role in regulating blood pressure (BP). An activated RAS contributes to elevated blood pressure and is evident in both human and animal models of hypertension. Drugs that target the classic vasoconstrictive arm of the RAS (angiotensin II/AT1 receptor signaling) are potent anti-hypertensive agents in clinical setting. However, the newly discovered angiotensin-converting enzyme 2 (ACE2)/angiotensin-(1-7)/Mas receptor axis added new vitality to the hypertension field. Advances in genetic manipulation and the relative low cost made the mouse model as one of the most popular animal models to study hypertension. Since a reliable and accurate method for BP assessment is the key for such experiments, here we provide a protocol for BP measurement in mice using a noninvasive BP system. The CODA noninvasive BP system (a tail-cuff Method, Kent Scientific Corporation) enables blood pressure (BP) measurements in mice. This method uses a specialized volume pressure recording (VPR) sensor, and measures blood volume changes that are placed over the animal's tail. Mice do need to be restrained in specific holders and artificially heated to maintain normal BP.

Use of a Fluorescent Substrate to Measure ACE2 Activity in the Mouse Abdominal Aorta.

The use of fluorogenic substrates to measure enzymatic activity is widely used to understand function within different experimental models. ACE2 is important in understanding the balance between AngII and Ang-(1-7) and how this balance could then in turn influence hypertension or other disease outcomes. Here, we describe a method to measure ACE2 activity in abdominal aorta of hyperlipidemic mice under both saline and AngII infusion.

Blood Pressure Monitoring Using Radio Telemetry Method in Mice.

The TA11PA-C10 implantable transmitter (Data Sciences International, DSI) is designed to measure blood pressure (BP) and activity in freely moving laboratory mice. The fluid filled catheter is placed in the free flowing blood of the systemic artery (inserted into the left carotid artery and extended into the aorta), and the transmitter body is placed in a benign location for long-term biocompatibility. The transmitter can be used to monitor BP in mice (as small as 17 g) under normal physiological and unrestricted conditions 24 h a day while remaining free from stress associated with human interaction. Thus, telemetry is considered the gold standard for BP monitoring in small animals such as mice. However, this methodology does require a good understanding of the system as well as appropriate training to perform the delicate transmitter implantation surgery.

Differential effects of Mas receptor deficiency on cardiac function and blood pressure in obese male and female mice.

Angiotensin-(1-7) [ANG-(1-7)] acts at Mas receptors (MasR) to oppose effects of angiotensin II (ANG II). Previous studies demonstrated that protection of female mice from obesity-induced hypertension was associated with increased systemic ANG-(1-7), whereas male obese hypertensive mice exhibited increased systemic ANG II. We hypothesized that MasR deficiency (MasR-/- ) augments obesity-induced hypertension in males and abolishes protection of females. Male and female wild-type (MasR+/+ ) and MasR-/- mice were fed a low-fat (LF) or high-fat (HF) diet for 16 wk. MasR deficiency had no effect on obesity. At baseline, male and female MasR-/- mice had reduced ejection fraction (EF) and fractional shortening than MasR+/+ mice. Male, but not female, HF-fed MasR+/+ mice had increased systolic and diastolic (DBP) blood pressures compared with LF-fed controls. In HF-fed females, MasR deficiency increased DBP compared with LF-fed controls. In contrast, male HF-fed MasR-/- mice had lower DBP than MasR+/+ mice. We quantified cardiac function after 1 mo of HF feeding in males of each genotype. HF-fed MasR-/- mice had higher left ventricular (LV) wall thickness than MasR+/+ mice. Moreover, MasR+/+ , but not MasR-/- , mice displayed reductions in EF from HF feeding that were reversed by ANG-(1-7) infusion. LV fibrosis was reduced in HF-fed MasR+/+ but not MasR-/- ANG-(1-7)-infused mice. These results demonstrate that MasR deficiency promotes obesity-induced hypertension in females. In males, HF feeding reduced cardiac function, which was restored by ANG-(1-7) in MasR+/+ but not MasR-/- mice. MasR agonists may be effective therapies for obesity-associated cardiovascular conditions.NEW & NOTEWORTHY MasR deficiency abolishes protection of female mice from obesity-induced hypertension. Male MasR-deficient obese mice have reduced blood pressure and declines in cardiac function. ANG-(1-7) infusion restores obesity-induced cardiac dysfunction of wild-type, but not MasR-deficient, male mice. MasR agonists may be cardioprotective in obese males and females.

Reversal of Bone Marrow Mobilopathy and Enhanced Vascular Repair by Angiotensin-(1-7) in Diabetes.

The angiotensin (ANG)-(1-7)/Mas receptor (MasR) pathway activates vascular repair-relevant functions of bone marrow progenitor cells. We tested the effects of ANG-(1-7) on mobilization and vasoreparative functions of progenitor cells that are impaired in diabetes. The study was performed in streptozotocin-induced diabetic (db/db) mice. Diabetes resulted in a decreased number of Lineage-Sca-1+c-Kit+ (LSK) cells in the circulation, which was normalized by ANG-(1-7). Diabetes-induced depletion of LSK cells in the bone marrow was reversed by ANG-(1-7). ρ-Kinase (ROCK) activity was increased specifically in bone marrow LSK cells by ANG-(1-7) in diabetes, and the beneficial effects of ANG-(1-7) were prevented by fasudil. ANG-(1-7) increased Slit3 levels in the bone marrow supernatants, which activated ROCK in LSK cells and sensitized them for stromal-derived factor-1α (SDF)-induced migration. Diabetes prevented the mobilization of LSK cells in response to ischemia and impaired the recovery of blood flow, both of which were reversed by ANG-(1-7) in both models of diabetes. Genetic ablation of MasR prevented ischemia-induced mobilization of LSK cells and impaired blood flow recovery, which was associated with decreased proliferation and migration of LSK cells in response to SDF or vascular endothelial growth factor. These results suggest that MasR is a promising target for the treatment of diabetic bone marrow mobilopathy and vascular disease.

Female Mice With an XY Sex Chromosome Complement Develop Severe Angiotensin II-Induced Abdominal Aortic Aneurysms.

BACKGROUND: Abdominal aortic aneurysms (AAAs) are a deadly pathology with strong sexual dimorphism. Similar to humans, female mice exhibit far lower incidences of angiotensin II-induced AAAs than males. In addition to sex hormones, the X and Y sex chromosomes, and their unique complements of genes, may contribute to sexually dimorphic AAA pathology. Here, we defined the effect of female (XX) versus male (XY) sex chromosome complement on angiotensin II-induced AAA formation and rupture in phenotypically female mice. METHODS: Female low-density lipoprotein receptor (Ldlr) deficient mice with an XX or XY sex chromosome complement were infused with angiotensin II for 28 days to induce AAAs. Abdominal aortic lumen diameters were quantified by ultrasound, whereas AAA diameters were quantified at study end point. DNA microarrays were performed on abdominal aortas. To mimic males, female mice were administered a single dose of testosterone as neonates or as adults before angiotensin II infusions. RESULTS: Female Ldlr-/- deficient mice with an XX and XY sex chromosome complement had similar sex organ weights and low serum testosterone concentrations. Abdominal aortas from female XY mice selectively expressed Y chromosome genes, whereas genes known to escape X inactivation were higher in XX females. The majority of aortic gene differences in XY versus XX females fell within inflammatory pathways. AAA incidences doubled and aneurysms ruptured in XY females. AAAs from XY females exhibited inflammation, and plasma interleukin-1ß concentrations were increased in XY females. Moreover, aortas from XY females had augmented matrix metalloproteinase activity and increased oxidative stress. Last, testosterone exposure applied chronically, or as a single bolus at postnatal day 1, markedly worsened AAA outcomes in XY in comparison with XX adult females. CONCLUSIONS: An XY sex chromosome complement in phenotypic females profoundly influenced aortic gene expression profiles and promoted AAA severity. When XY females were exposed to testosterone, aneurysm rupture rates were striking. Mechanisms for augmented AAA severity in XY females include increased inflammation, augmented matrix metalloproteineases, and oxidative stress. Our results demonstrate that genes on the sex chromosomes regulate aortic vascular biology and contribute to sexual dimorphism of AAAs. Sex chromosome genes may serve as novel targets for sex-specific AAA therapeutics.

ACE2 deficiency reduces ß-cell mass and impairs ß-cell proliferation in obese C57BL/6 mice.

Drugs that inhibit the renin-angiotensin system (RAS) decrease the onset of type 2 diabetes (T2D). Pancreatic islets express RAS components, including angiotensin-converting enzyme 2 (ACE2), which cleaves angiotensin II (Ang II) to angiotensin-(1-7) [Ang-(1-7)]. Overexpression of ACE2 in pancreas of diabetic mice improved glucose homeostasis. The purpose of this study was to determine if deficiency of endogenous ACE2 contributes to islet dysfunction and T2D. We hypothesized that ACE2 deficiency potentiates the decline in ß-cell function and augments the development of diet-induced T2D. Male Ace2(+/y) or Ace2(-/y) mice were fed a low-fat (LF) or high-fat (HF) diet for 1 or 4 mo. A subset of 1-mo HF-fed mice were infused with Sal (Sal), losartan (Los), or Ang-(1-7). At 4 mo, while both genotypes of HF-fed mice developed a similar level of insulin resistance, adaptive hyperinsulinemia was reduced in Ace2(-/y) vs. Ace2(+/y) mice. Similarly, in vivo glucose-stimulated insulin secretion (GSIS) was reduced in 1-mo HF-fed Ace2(-/y) compared with Ace2(+/y) mice, resulting in augmented hyperglycemia. The average islet area was significantly smaller in both LF- and HF-fed Ace2(-/y) vs. Ace2(+/y) mice. Additionally, ß-cell mass and proliferation were reduced significantly in HF-fed Ace2(-/y) vs. Ace2(+/y) mice. Neither infusion of Los nor Ang-(1-7) was able to correct impaired in vivo GSIS of HF-fed ACE2-deficient mice. These results demonstrate a critical role for endogenous ACE2 in the adaptive ß-cell hyperinsulinemic response to HF feeding through regulation of ß-cell proliferation and growth.