Abdominal Aortic Aneurysm Lesions Research Articles

Long noncoding RNA Sngh20 promotes vascular smooth muscle cell senescence and abdominal aortic aneurysm growth by interacting with dead-box helicase 5

Abstract Background Development of non-surgical treatment of human abdominal aortic aneurysm (AAA) has clinical significance. Long noncoding RNAs (lncRNAs) are emerging as powerful mediators participated in cellular senescence process that a critical pathological alteration prompting AAA development. Purpose To investigate the role and underlying mechanism of lncRNA Small nucleolar RNA host gene-20 (Snhg20) in AAA formation. Methods Experimental AAA mice models were produced by peri-aortic CaPO4 injury in C57bl/6J mice or Subcutaneousangiotensin-II (Ang-II) infusion in Apoe–/– mice and gapmeR was used to knockdown Snhg20 expression. Adeno-associated virus and anti-IL-6 monoclonal antibody was used to knockdown dead-box helicase 5 (DDX5) and neutralize IL-6, respectively.Immunohistochemical and immunofluorescent staining, ELISA and real-time PCR were performed to determine the vascular structure and inflammation, cellular senescence and senescence-associated secretory phenotype (SASP) content in human and mouse AAA lesions. Human aortic SMC cell line was used to investigate the effect of Snhg20 knockdown on senescence and SASP in vitro. LncRNA pulldown and mass spectrometry were used to identify potential targets that interact with Snhg20. p53 DNA binding affinity was examined by EMSA. Results The expression of Snhg20 was downregulated in both human and mice AAA lesion. Snhg20 knockdown promoted SMC senescence and loss, vascular inflammation and AAA growth in both experimental AAA mice models. Mechanistically, Snhg20 interacted with DDX5, an important transcriptional coactivator of p53. The absence of snhg20 increased DDX5 and p53 interaction and thereby promoted p53 DNA binding affinity to transcriptionally initiate p21expression, resulting in increased cellular senescence and senescence-associated secretory phenotype (SASP) secretion, including IL-6. Moreover, DDX5 silencing or anti-IL-6 therapy rescued the SMC senescence and loss, vascular inflammation and AAA growth mediated by Snhg20 knockdown. Furthermore, human AAA lesions showed increased DDX5 co-localization with p53 and abundant senescent vascular SMC. Conclusion These findings identify an important role of Snhg20 in controlling vascular SMC senescence and SPAP expression and suggest that Snhg20 is a potential target for AAA treatment aimed at reducing VSMCs senescence.Schematic summary

NINJ1 Facilitates Abdominal Aortic Aneurysm Formation via Blocking TLR4-ANXA2 Interaction and Enhancing Macrophage Infiltration.

Abdominal aortic aneurysm (AAA) is a common and potentially life-threatening condition. Chronic aortic inflammation is closely associated with the pathogenesis of AAA. Nerve injury-induced protein 1 (NINJ1) is increasingly acknowledged as a significant regulator of the inflammatory process. However, the precise involvement of NINJ1 in AAA formation remains largely unexplored. The present study finds that the expression level of NINJ1 is elevated, along with the specific expression level in macrophages within human and angiotensin II (Ang II)-induced murine AAA lesions. Furthermore, Ninj1flox/flox and Ninj1flox/floxLyz2-Cre mice on an ApoE-/- background are generated, and macrophage NINJ1 deficiency inhibits AAA formation and reduces macrophage infiltration in mice infused with Ang II. Consistently, in vitro suppressing the expression level of NINJ1 in macrophages significantly restricts macrophage adhesion and migration, while attenuating macrophage pro-inflammatory responses. Bulk RNA-sequencing and pathway analysis uncover that NINJ1 can modulate macrophage infiltration through the TLR4/NF-κB/CCR2 signaling pathway. Protein-protein interaction analysis indicates that NINJ1 can activate TLR4 by competitively binding with ANXA2, an inhibitory interacting protein of TLR4. These findings reveal that NINJ1 can modulate AAA formation by promoting macrophage infiltration and pro-inflammatory responses, highlighting the potential of NINJ1 as a therapeutic target for AAA.

Colchicine Blocks Abdominal Aortic Aneurysm Development by Maintaining Vascular Smooth Muscle Cell Homeostasis.

Development of non-surgical treatment of human abdominal aortic aneurysm (AAA) has clinical significance. Colchicine emerges as an effective therapeutic regimen in cardiovascular diseases. Yet, whether colchicine slows AAA growth remain controversy. Here, we demonstrated that daily intragastric administration of low-dose colchicine blocked AAA formation, prevented vascular smooth muscle cell (SMC) phenotype switching and apoptosis, and vascular inflammation in both peri-aortic CaPO4 injury and subcutaneous angiotensin-II infusion induced experimental AAA mice models. Mechanistically, colchicine increased global mRNA stability by inhibiting the METTL14/YTHDC1-mediated m6A modification, resulting in increased sclerostin (SOST) expression and consequent inactivation of the WNT/β-catenin signaling pathway in vascular SMCs from mouse AAA lesions and in cultured human aortic SMCs. Moreover, human and mouse AAA lesions all showed increased m6A methylation, decreased SOST expression, and skewed synthetic SMC de-differentiation phenotype, compared to those without AAA. This study uncovers a novel mechanism of colchicine in slowing AAA development by using the METTL14/SOST/WNT/β-catenin axis to control vascular SMC homeostasis in mouse aortic vessels and in human aortic SMCs. Therefore, use of colchicine may benefit AAA patients in clinical practice.

Targeted PERK inhibition with biomimetic nanoclusters confers preventative and interventional benefits to elastase-induced abdominal aortic aneurysms.

Abdominal aortic aneurysm (AAA) is a progressive aortic dilatation, causing ∼80% mortality upon rupture. Currently, there is no approved drug therapy for AAA. Surgical repairs are invasive and risky and thus not recommended to patients with small AAAs which, however, account for ∼90% of the newly diagnosed cases. It is therefore a compelling unmet clinical need to discover effective non-invasive strategies to prevent or slow down AAA progression. We contend that the first AAA drug therapy will only arise through discoveries of both effective drug targets and innovative delivery methods. There is substantial evidence that degenerative smooth muscle cells (SMCs) orchestrate AAA pathogenesis and progression. In this study, we made an exciting finding that PERK, the endoplasmic reticulum (ER) stress Protein Kinase R-like ER Kinase, is a potent driver of SMC degeneration and hence a potential therapeutic target. Indeed, local knockdown of PERK in elastase-challenged aorta significantly attenuated AAA lesions in vivo. In parallel, we also conceived a biomimetic nanocluster (NC) design uniquely tailored to AAA-targeting drug delivery. This NC demonstrated excellent AAA homing via a platelet-derived biomembrane coating; and when loaded with a selective PERK inhibitor (PERKi, GSK2656157), the NC therapy conferred remarkable benefits in both preventing aneurysm development and halting the progression of pre-existing aneurysmal lesions in two distinct rodent models of AAA. In summary, our current study not only establishes a new intervention target for mitigating SMC degeneration and aneurysmal pathogenesis, but also provides a powerful tool to facilitate the development of effective drug therapy of AAA.

In Vivo Validation of Modulated Acoustic Radiation Force-Based Imaging in Murine Model of Abdominal Aortic Aneurysm Using VEGFR-2-Targeted Microbubbles.

The objective of this study is to validate the modulated acoustic radiation force (mARF)-based imaging method in the detection of abdominal aortic aneurysm (AAA) in murine models using vascular endothelial growth factor receptor 2 (VEGFR-2)-targeted microbubbles (MBs). The mouse AAA model was prepared using the subcutaneous angiotensin II (Ang II) infusion combined with the β-aminopropionitrile monofumarate solution dissolved in drinking water. The ultrasound imaging session was performed at 7 days, 14 days, 21 days, and 28 days after the osmotic pump implantation. For each imaging session, 10 C57BL/6 mice were implanted with Ang II-filled osmotic pumps, and 5 C57BL/6 mice received saline infusion only as the control group. Biotinylated lipid MBs conjugated to either anti-mouse VEGFR-2 antibody (targeted MBs) or isotype control antibody (control MBs) were prepared before each imaging session and were injected into mice via tail vein catheter. Two separate transducers were colocalized to image the AAA and apply ARF to translate MBs simultaneously. After each imaging session, tissue was harvested and the aortas were used for VEGFR-2 immunostaining analysis. From the collected ultrasound image data, the signal magnitude response of the adherent targeted MBs was analyzed, and a parameter, residual-to-saturation ratio ( Rres - sat ), was defined to measure the enhancement in the adherent targeted MBs signal after the cessation of ARF compared with the initial signal intensity. Statistical analysis was performed with the Welch t test and analysis of variance test. The Rres - sat of abdominal aortic segments from Ang II-challenged mice was significantly higher compared with that in the saline-infused control group ( P < 0.001) at all 4 time points after osmotic pump implantation (1 week to 4 weeks). In control mice, the Rres - sat values were 2.13%, 1.85%, 3.26%, and 4.85% at 1, 2, 3, and 4 weeks postimplantation, respectively. In stark contrast, the Rres - sat values for the mice with Ang II-induced AAA lesions were 9.20%, 20.6%, 22.7%, and 31.8%, respectively. It is worth noting that there was a significant difference between the Rres - sat for Ang II-infused mice at all 4 time points ( P < 0.005), a finding not present in the saline-infused mice. Immunostaining results revealed the VEGFR-2 expression was increased in the abdominal aortic segments of Ang II-infused mice compared with the control group. The mARF-based imaging technique was validated in vivo using a murine model of AAA and VEGFR-2-targeted MBs. Results in this study indicated that the mARF-based imaging technique has the ability to detect and assess AAA growth at early stages based on the signal intensity of adherent targeted MBs, which is correlated with the expression level of the desired molecular biomarker. The results may suggest, in very long term, a pathway toward eventual clinical implementation for an ultrasound molecular imaging-based approach to AAA risk assessment in asymptomatic patients.

PRDM16 deficiency in vascular smooth muscle cells aggravates abdominal aortic aneurysm.

Abdominal aortic aneurysm (AAA) is usually asymptomatic until life-threatening complications occur, predominantly involving aortic rupture. Currently, no drug-based treatments are available, primarily due to limited understanding of AAA pathogenesis. The transcriptional regulator PR domain-containing protein 16 (PRDM16) is highly expressed in the aorta, but its functions in the aorta are largely unknown. By RNA-seq analysis, we found that vascular smooth muscle cell-specific (VSMC-specific) Prdm16-knockout (Prdm16SMKO) mice already showed extensive changes in the expression of genes associated with extracellular matrix (ECM) remodeling and inflammation in the abdominal aorta under normal housing conditions without any pathological stimuli. Human AAA lesions displayed lower PRDM16 expression. Periadventitial elastase application to the suprarenal region of the abdominal aorta aggravated AAA formation in Prdm16SMKO mice. During AAA development, VSMCs undergo apoptosis because of both intrinsic and environmental changes, including inflammation and ECM remodeling. Prdm16 deficiency promoted inflammation and apoptosis in VSMCs. A disintegrin and metalloproteinase 12 (ADAM12) is a gelatinase that can degrade various ECMs. We found that ADAM12 is a target of transcriptional repression by PRDM16. Adam12 knockdown reversed VSMC apoptosis induced by Prdm16 deficiency. Our study demonstrated that PRDM16 deficiency in VSMCs promoted ADAM12 expression and aggravates AAA formation, which may provide potential targets for AAA treatment.

PRDM16 Deficiency in Vascular Smooth Muscle Cells Aggravates Abdominal Aortic Aneurysm

Abdominal aortic aneurysm (AAA) is usually asymptomatic until severe complications with high risk of lethality occur, predominantly involving aortic rupture. Currently, no drug-based treatments are available, primarily due to limited understanding of AAA pathogenesis. PR domain-containing protein 16 (PRDM16) is a transcriptional regulator that governs gene expression by cooperating with other transcription factors. It is highly expressed in the aorta, but its functions in the aortic tissue and AAA pathology are largely unknown. In this study, we investigated the roles of PRDM16 in vascular smooth muscle cells (VSMCs). By RNA-seq analysis, we found that VSMCs-specific Prdm16 knockout mice ( Prdm16 SMKO) already showed extensive changes in the expression of genes associated with extracellular matrix (ECM) remodeling and inflammation in the abdominal aorta under normal housing conditions without any pathological stimuli. We found that human AAA lesions displayed decreased PRDM16 expression in the aortic medial layer. Periadventitial elastase application to the suprarenal region of the abdominal aorta aggravated AAA formation in Prdm16 SMKO mice. During AAA development, VSMCs undergo apoptosis because of both intrinsic and environmental changes including inflammation and ECM remodeling. Prdm16 deficiency promoted inflammation and apoptosis in VSMCs. A disintegrin and metalloproteinase 12 (ADAM12) is a gelatinase which can degrade various ECM. We found that ADAM12 is a target of transcriptional repression by PRDM16. Adam12 knockdown reversed VSMC apoptosis induced by Prdm16 deficiency. Our study demonstrated that PRDM16 deficiency in VSMCs promoted ADAM12 expression and aggravates AAA formation, which may provide potential targets for AAA treatment. This work was partially supported by National Institutes of Health grants HL151524 (L. Chang) This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

DOCK2 Deficiency Attenuates Abdominal Aortic Aneurysm Formation-Brief Report.

Abdominal aortic aneurysm (AAA) is a potentially lethal disease that lacks pharmacological treatment. Degradation of extracellular matrix proteins, especially elastin laminae, is the hallmark for AAA development. DOCK2 (dedicator of cytokinesis 2) has shown proinflammatory effects in several inflammatory diseases and acts as a novel mediator for vascular remodeling. However, the role of DOCK2 in AAA formation remains unknown. Ang II (angiotensin II) infusion of ApoE-/- (apolipoprotein E deficient) mouse and topical elastase-induced AAA combined with DOCK2-/- (DOCK2 knockout) mouse models were used to study DOCK2 function in AAA formation/dissection. The relevance of DOCK2 to human AAA was examined using human aneurysm specimens. Elastin fragmentation in AAA lesion was observed by elastin staining. Elastin-degrading enzyme MMP (matrix metalloproteinase) activity was measured by in situ zymography. DOCK2 was robustly upregulated in AAA lesion of Ang II-infused ApoE-/- mice, elastase-treated mice, as well as human AAA lesions. DOCK2-/- significantly attenuated the Ang II-induced AAA formation/dissection or rupture in mice along with reduction of MCP-1 (monocyte chemoattractant protein-1) and MMP expression and activity. Accordingly, the elastin fragmentation observed in ApoE-/- mouse aorta infused with Ang II and elastase-treated aorta was significantly attenuated by DOCK2 deficiency. Moreover, DOCK2-/- decreased the prevalence and severity of aneurysm formation, as well as the elastin degradation observed in the topical elastase model. Our results indicate that DOCK2 is a novel regulator for AAA formation. DOCK2 regulates AAA development by promoting MCP-1 and MMP2 expression to incite vascular inflammation and elastin degradation.

Purinergic receptor P2X7 contributes to abdominal aortic aneurysm development via modulating macrophage pyroptosis and inflammation

The purinergic receptor P2X7 has been established as an important mediator of inflammation and participates in a variety of cardiovascular diseases including atherosclerosis, however, its role in abdominal aortic aneurysms (AAA) remains unclear. In this study, we demonstrate that P2X7 plays essential roles in AAA development via modulating macrophage pyroptosis and inflammation. P2X7 is highly expressed in human AAA specimen, as well as in experimental murine AAA lesions (both CaCl2- and Angiotensin II-induced AAA models), and it mainly confines in macrophages. Furthermore, P2X7 deficiency or pharmacological inhibition with its antagonist could significantly attenuate aneurysm formation in experimental murine AAA models, while P2X7 agonist could promote AAA development. The caspase-1 activity, matrix metalloproteinase (MMP) activity, reactive oxygen species (ROS) production and pro-inflammatory gene expression were significantly reduced in experimental AAA lesions in mice with P2X7 deficiency or inhibition. Mechanistically, macrophage P2X7 can mediate the activation of NLRP3 inflammasome and activate its downstream caspase-1 to initiate the pyroptosis pathway. After caspase-1 activation, it further cleaves pro-interleukin (IL)-1β and gasdermin D (GSDMD). Consequently, the N-terminal fragment of GSDMD forms pores on the cell membrane, leading to macrophage pyroptosis and release of the pro-inflammatory factor IL-1β. The resulting vascular inflammation further leads to the upregulation of MMP and ROS, thereby promoting AAA development. In summary, these data identify P2X7-mediated macrophage pyroptosis signaling pathway as a novel contributory mechanism of AAA formation.

ADAR1 Non-Editing Function in Macrophage Activation and Abdominal Aortic Aneurysm.

Macrophage activation plays a critical role in abdominal aortic aneurysm (AAA) development. However, molecular mechanisms controlling macrophage activation and vascular inflammation in AAA remain largely unknown. The objective of the study was to identify novel mechanisms underlying adenosine deaminase acting on RNA (ADAR1) function in macrophage activation and AAA formation. Aortic transplantation was conducted to determine the importance of nonvascular ADAR1 in AAA development/dissection. Ang II (Angiotensin II) infusion of ApoE-/- mouse model combined with macrophage-specific knockout of ADAR1 was used to study ADAR1 macrophage-specific role in AAA formation/dissection. The relevance of macrophage ADAR1 to human AAA was examined using human aneurysm specimens. Moreover, a novel humanized AAA model was established to test the role of human macrophages in aneurysm formation in human arteries. Allograft transplantation of wild-type abdominal aortas to ADAR1+/- recipient mice significantly attenuated AAA formation, suggesting that nonvascular ADAR1 is essential for AAA development. ADAR1 deficiency in hematopoietic cells decreased the prevalence and severity of AAA while inhibited macrophage infiltration and aorta wall inflammation. ADAR1 deletion blocked the classic macrophage activation, diminished NF-κB (nuclear factor kappa B) signaling, and enhanced the expression of a number of anti-inflammatory microRNAs. Mechanistically, ADAR1 interacted with Drosha to promote its degradation, which attenuated Drosha-DGCR8 (DiGeorge syndrome critical region 8) interaction, and consequently inhibited pri- to pre-microRNA processing of microRNAs targeting IKKβ, resulting in an increased IKKβ (inhibitor of nuclear factor kappa-B) expression and enhanced NF-κB signaling. Significantly, ADAR1 was induced in macrophages and interacted with Drosha in human AAA lesions. Reconstitution of ADAR1-deficient, but not the wild type, human monocytes to immunodeficient mice blocked the aneurysm formation in transplanted human arteries. Macrophage ADAR1 promotes aneurysm formation in both mouse and human arteries through a novel mechanism, that is, Drosha protein degradation, which inhibits the processing of microRNAs targeting NF-kB signaling and thus elicits macrophage-mediated vascular inflammation in AAA.

Vascular smooth muscle RhoA counteracts abdominal aortic aneurysm formation by modulating MAP4K4 activity

Whether a small GTPase RhoA plays a role in the pathology of abdominal aortic aneurysm (AAA) has not been determined. We show here that RhoA expression is reduced in human AAA lesions, compared with normal areas. Furthermore, incidence of AAA formation is increased in vascular smooth muscle cell (VSMC)-specific RhoA conditional knockout (cKO) mice. The contractility of the aortic rings and VSMCs from RhoA cKO mice is reduced, and expression of genes related to the VSMC contractility is attenuated by loss of RhoA. RhoA depletion activates the mitogen-activated protein (MAP) kinase signaling, including MAP4K4, in the aorta and VSMCs. Inhibition of MAP4K4 activity by DMX-5804 decreases AAA formation. Set, a binding protein to active RhoA, functions as an activator of MAP4K4 by sequestering PP2A, an inhibitor of MAP4K4, in the absence of RhoA. In conclusion, RhoA counteracts AAA formation through inhibition of MAP4K4 in cooperation with Set.

Abstract 350: Disruption Of Proprotein Convertase Subtilisin/Kexin 9 Suppresses Inflammation And Attenuates Abdominal Aortic Aneurysm Formation

Objective: Abdominal aortic aneurysm (AAA) is a degenerative inflammatory disease, with its rupture highly lethal. There have been no effective pharmacological medications. Proprotein convertase subtilisin/kexin 9 (PCSK9), which is known to modulate cholesterol metabolism, has recently been associated with AAA in human genome-wide association studies. However, its role in AAA pathogenesis is unknown. Approach and Results: Histology staining of mouse aortic aneurysmal tissue showed PCSK9 expression and its colocalization with macrophages. To investigate the role of PCSK9 in AAA formation, mice deficient in PCSK9 were crossed with ApoE SA/SA mice, which are hypercholesterolemic and inducible to develop angiotensin II-mediated hypertension upon doxycycline (Dox) administration. Deletion of PCSK9 reduced both incidence and severity of AAA and increased survival rate, without changing plasma cholesterol levels. This was associated with decreased aortic expression and activity of matrix metalloproteinase 9 (MMP9), less elastin breakdown, and reduced aortic expansion and aneurysm rupture. Additionally, PCSK9 deficiency suppressed systemic inflammatory response and macrophage infiltration into AAA lesions. In isolated macrophages stimulated with LPS, the upregulated expression of pro-inflammatory cytokines ( Tnfa, Il1b, Il6 ) was dramatically diminished in PCSK9-deficient macrophages. Furthermore, administration of PCSK9-specific siRNA in mice resulted in less macrophage accumulation in the aneurysmal aortic wall, decreased MMP9-mediated elastin degradation, and substantially constrained AAA formation. Conclusions: Endogenous PCSK9 promotes AAA formation, consistent with its novel action in mediating systemic and local inflammation, particularly, the macrophage-dependent MMP9 activity. Pharmacological inhibition of PCSK9 might be a promising strategy to curtail AAA. Key Words: aneurysm, PCSK9, elastin, inflammation, macrophages

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Abstract 14649: Chloride Channel Calcium-activated-1 Reduces Mouse Abdominal Aortic Aneurysm by Blocking Macrophage Migration Inhibitory Factor Activity in Macrophage Activation

Introduction: Human chloride channel calcium-activated-1 (CLCA1) protein is a secretory protein that goblet cells from trachea epithelium express abundantly where it participates in hypersecretory lung diseases by mucus overproduction. Studies from CLCA1-deficient mice or CLCA1 blocking agents have however yielded inconsistent conclusions. CLCA1 not only regulates mucin expression, but also participates in innate immune responses by binding to yet unidentified molecules on the inflammatory cells and regulating cytokine and chemokine production. Seeking CLCA1 binding proteins identified CLCA1 expression by endothelial cells (EC), smooth muscle cells (SMC), and macrophages in human and mouse abdominal aortic aneurysms (AAA). Ligation of CLCA1 mediated the actions of macrophage migration inhibitory factor (MIF). Hypothesis: CLCA1 controls AAA growth by regulating MIF activity. Methods and Results: Immunoblots revealed high levels of the 110 and 75-kDa CLCA1 fragments in human abdominal aneurysm lesions, but normal human aortas contained predominantly the 35-kDa carboxyl-terminal fragments. Cultured human EC and SMC contained only the 35-kDa fragment, but human monocyte-derived macrophages contained both the 75 and 35-kDa fragments. Confocal microscopy colocalized CLCA1 and MIF in macrophages from human AAA lesions. CLCA1 immunoprecipitation followed by MIF immunoblot showed that the 35-kDa CLCA1 fragment complexes with MIF in cultured macrophages. Adventitial CaPO 4 -induced AAA in male Clca1 -/- mice demonstrated significantly greater aneurysm diameter, and increased lesion neutrophil and eosinophil content, as well as neutrophil- and eosinophil-associated chemokine (CXCL1, 2, 5, CCR3, and eotaxin-2 and 3) expression than those in wild-type or sham-operated mice. Macrophages cultured from Clca1 -/- bone-marrow showed enhanced MIF activity compared to those from wild-type mice. Conclusions: The 35-kDa CLCA1 fragment reduces formation of experimental AAA by acting as a decoy receptor, sequestering MIF and thus limiting MIF-mediated macrophage activation.

Specific inhibition of SHP2 suppressed abdominal aortic aneurysm formation in mice by augmenting the immunosuppressive function of MDSCs

AimsTo address the roles of SHP2 in regulating angiotensin II (Ang II) induced abdominal aortic aneurysm (AAA) and the potential molecular mechanisms. Main methodsAAA model was established in apolipoprotein E-deficient (apoE−/−) mice infused with Ang II. Suprarenal aortic luminal diameters were ultrasonically measured to determine the presentation of AAA in mice. The inflammatory and immunosuppressive factors in serum were detected by ELISA. AAA lesion size, positive macrophages and elastic laminae degradation were examined by histological analysis. Myeloid-derived suppressor cells (MDSCs) were measured by flow cytometry after magnetic bead sorting. Bioinformatics analysis was applied to screen the crucial genes related the progression of AAA. Key findingsTreatment with PHPS1 (SHP2 inhibitor) significantly decreased the vascular diameter of AAA. Histological analysis showed that PHPS1 obviously reduced the Masson positive area, macrophages positive area, as well as the damage rate of elastic laminae. Moreover, PHPS1 suppressed the expression of INF-γ, TNF-α and MMPs, as well as elevated IL-10 and arginase-1 expression. Additionally, PHPS1 enhanced the expression of granulocytic MDSCs (G-MDSCs). By consulting with bioinformatics, STAT3 was selected. In G-MDSCs, PHPS1 stimulation obviously increased the phosphorylation level of STAT3, as well as elevated the protein expression of C/EBPβ and arginase-1. However, the above phenomena can be blocked after Stattic (STAT3 inhibitor) treatment. SignificanceSHP2 may affect the AAA progression by interfering with expansion and function of MDSCs to regulate the body immunity, which might afford a novel direction for the treatment of patients with AAA.

Eosinophils Protect Mice From Angiotensin-II Perfusion-Induced Abdominal Aortic Aneurysm.

Blood eosinophil count and ECP (eosinophil cationic protein) associate with human cardiovascular diseases. Yet, whether eosinophils play a role in cardiovascular disease remains untested. The current study detected eosinophil accumulation in human and murine abdominal aortic aneurysm (AAA) lesions, suggesting eosinophil participation in this aortic disease. To test whether and how eosinophils affect AAA growth. Population-based randomized clinically controlled screening trials revealed higher blood eosinophil count in 579 male patients with AAA than in 5063 non-AAA control (0.236±0.182 versus 0.211±0.154, 109/L, P<0.001). Univariate (odds ratio, 1.381, P<0.001) and multivariate (odds ratio, 1.237, P=0.031) logistic regression analyses indicated that increased blood eosinophil count in patients with AAA served as an independent risk factor of human AAA. Immunostaining and immunoblot analyses detected eosinophil accumulation and eosinophil cationic protein expression in human and murine AAA lesions. Results showed that eosinophil deficiency exacerbated AAA growth with increased lesion inflammatory cell contents, matrix-degrading protease activity, angiogenesis, cell proliferation and apoptosis, and smooth muscle cell loss using angiotensin-II perfusion-induced AAA in Apoe-/- and eosinophil-deficient Apoe-/-ΔdblGATA mice. Eosinophil deficiency increased lesion chemokine expression, muted lesion expression of IL (interleukin) 4 and eosinophil-associated-ribonuclease-1 (mEar1 [mouse EOS-associated-ribonuclease-1], human ECP homolog), and slanted M1 macrophage polarization. In cultured macrophages and monocytes, eosinophil-derived IL4 and mEar1 polarized M2 macrophages, suppressed CD11b+Ly6Chi monocytes, and increased CD11b+Ly6Clo monocytes. mEar1 treatment or adoptive transfer of eosinophil from wild-type and Il13-/- mice, but not eosinophil from Il4-/- mice, blocked AAA growth in Apoe-/-ΔdblGATA mice. Immunofluorescent staining and immunoblot analyses demonstrated a role for eosinophil IL4 and mEar1 in blocking NF-κB (nuclear factor-κB) activation in macrophages, smooth muscle cells, and endothelial cells. Eosinophils play a protective role in AAA by releasing IL4 and cationic proteins such as mEar1 to regulate macrophage and monocyte polarization and to block NF-κB activation in aortic inflammatory and vascular cells.

Hyperlipidemia does not affect development of elastase-induced abdominal aortic aneurysm in mice

Background and aimsHyperlipidemia is a suggested risk factor for abdominal aortic aneurysm (AAA). However, whether hyperlipidemia is causally involved in AAA progression remains elusive. Here, we tested the hypothesis that hyperlipidemia aggravates AAA formation in the widely used porcine pancreatic elastase (PPE) model of AAA in mice with varying levels of plasma lipids. MethodsPrior to PPE-surgery, 8-week-old male C57BL/6J mice (n = 32) received 1·1011 viral genomes of rAAV8-D377Y-mPcsk9 or control rAAV8 via the tail vein. Mice were fed either western type diet or regular chow. At baseline and during the 28 days following PPE-surgery, mice underwent weekly ultrasonic assessment of AAA progression. Experiments were repeated using Apolipoprotein E knockout (ApoE−/−) mice (n = 7) and wildtype C57BL/6J mice (n = 5). ResultsAt sacrifice, maximal intergroup plasma cholesterol and non-HDL/HDL ratio differences were >5-fold and >20-fold, respectively. AAA diameters expanded to 150% of baseline, but no intergroup differences were detected. This was verified in an independent experiment comparing 8-week-old male ApoE−/− mice with wildtype mice. Histological evaluation of experimental AAA lesions revealed accumulated lipid in neointimal and medial layers, and analysis of human AAA lesions (n = 5) obtained from open repair showed medial lipid deposition. ConclusionsIn summary, we find that lipid deposition in the aortic wall is a feature of PPE-induced AAA in mice as well as human AAA lesions. Despite, our data do not support the hypothesis that hyperlipidemia contributes to AAA progression.

Therapeutic potential of tricarboxylic acid cycle metabolite itaconate in cardiovascular diseases.

Therapeutic potential of tricarboxylic acid cycle metabolite itaconate in cardiovascular diseases.

BAF60a Deficiency in Vascular Smooth Muscle Cells Prevents Abdominal Aortic Aneurysm by Reducing Inflammation and Extracellular Matrix Degradation.

Currently, there are no approved drugs for abdominal aortic aneurysm (AAA) treatment, likely due to limited understanding of the primary molecular mechanisms underlying AAA development and progression. BAF60a-a unique subunit of the SWI/SNF (switch/sucrose nonfermentable) chromatin remodeling complex-is a novel regulator of metabolic homeostasis, yet little is known about its function in the vasculature and pathogenesis of AAA. In this study, we sought to investigate the role and underlying mechanisms of vascular smooth muscle cell (VSMC)-specific BAF60a in AAA formation. Approach and Results: BAF60a is upregulated in human and experimental murine AAA lesions. In vivo studies revealed that VSMC-specific knockout of BAF60a protected mice from both Ang II (angiotensin II)-induced and elastase-induced AAA formation with significant suppression of vascular inflammation, monocyte infiltration, and elastin fragmentation. Through RNA sequencing and pathway analysis, we found that the expression of inflammatory response genes in cultured human aortic smooth muscle cells was significantly downregulated by small interfering RNA-mediated BAF60a knockdown while upregulated upon adenovirus-mediated BAF60a overexpression. BAF60a regulates VSMC inflammation by recruiting BRG1 (Brahma-related gene-1)-a catalytic subunit of the SWI/SNF complex-to the promoter region of NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) target genes. Furthermore, loss of BAF60a in VSMCs prevented the upregulation of the proteolytic enzyme cysteine protease CTSS (cathepsin S), thus ameliorating ECM (extracellular matrix) degradation within the vascular wall in AAA. Our study demonstrated that BAF60a is required to recruit the SWI/SNF complex to facilitate the epigenetic regulation of VSMC inflammation, which may serve as a potential therapeutic target in preventing and treating AAA.

Reduced Nhe1 (Na+-H+ Exchanger-1) Function Protects ApoE-Deficient Mice From Ang II (Angiotensin II)-Induced Abdominal Aortic Aneurysms.

IgE-mediated activation of Nhe1 (Na+-H+ exchanger-1) induces aortic cell extracellular acidification and promotes cell apoptosis. A pH-sensitive probe pHrodo identified acidic regions at positions of macrophage accumulation, IgE expression, and cell apoptosis in human and mouse abdominal aortic aneurysm (AAA) lesions. Ang II (angiotensin II)-induced AAA in Nhe1-insufficient Apoe-/-Nhe1+/- mice and Apoe-/-Nhe1+/+ littermates tested Nhe1 activity in experimental AAA, because Nhe1-/- mice develop ataxia and epileptic-like seizures and die early. Nhe1 insufficiency reduced AAA incidence and size, lesion macrophage and T-cell accumulation, collagen deposition, elastin fragmentation, cell apoptosis, smooth muscle cell loss, and MMP (matrix metalloproteinase) activity. Nhe1 insufficiency also reduced blood pressure and the plasma apoptosis marker TCTP (translationally controlled tumor protein) but did not affect plasma IgE. While pHrodo localized the acidic regions to macrophage clusters, IgE expression, and cell apoptosis in AAA lesions from Apoe-/-Nhe1+/+ mice, such acidic areas were much smaller in lesions from Apoe-/-Nhe1+/- mice. Nhe1-FcεR1 colocalization in macrophages from AAA lesions support a role of IgE-mediated Nhe1 activation. Gelatin zymography, immunoblot, and real-time polymerase chain reaction analyses demonstrated that Nhe1 insufficiency reduced the MMP activity, cysteinyl cathepsin expression, IgE-induced apoptosis, and NF-κB activation in macrophages and blocked IgE-induced adhesion molecule expression in endothelial cells. A near-infrared fluorescent probe (LS662) together with fluorescence reflectance imaging of intact aortas showed reduced acidity in AAA lesions from Nhe-1-insufficient mice. This study revealed extracellular acidity at regions rich in macrophages, IgE expression, and cell apoptosis in human and mouse AAA lesions and established a direct role of Nhe1 in AAA pathogenesis.