Biology subjects

Lu, H. S.

Publications and source records attributed to Lu, H. S..

At least 19 recordsLinked to original sources

bioRxiv2026

Deletion of LRP1 in Smooth Muscle Cells Induces Infrarenal Aortic Aneurysms in Mice

Background: Low-density lipoprotein receptor-related protein 1 (LRP1) in smooth muscle cells (SMCs) is essential for maintaining aortic wall integrity. This study investigated whether deletion of LRP1 in SMCs induces region-specific aortic pathology and evaluated a potential therapeutic strategy. Methods: Lrp1-floxed mice were bred with Sm22-Cre+/0 mice to generate mice with SMC-LRP1 deletion (SMC-LRP1-/-) and wild-type littermates (SMC-LRP1+/+). Micro-computed tomography (micro-CT) was used to visualize the aorta, and aortic tissue sections were characterized histologically. To evaluate a potential therapeutic strategy, antisense oligonucleotides (ASOs) targeting hepatocyte-derived angiotensinogen (AGT) were administered to inhibit AGT synthesis. Results: Immunostaining and Western blotting confirmed LRP1 deletion in aortic SMCs. At 40 weeks of age, micro-CT revealed marked increases in aortic width and volume. Although dilatation occurred throughout the aorta, the infrarenal region exhibited the greatest expansion in both male and female SMC-LRP1-/- mice compared with SMC-LRP1+/+ littermates; subsequent analyses therefore focused on this region. LRP1 deletion in SMCs caused pronounced SMC loss and increased collagen deposition in the infrarenal aorta. Inhibition of hepatocyte-derived AGT with ASO attenuated infrarenal aortic dilatation and preserved aortic wall integrity in both male and female mice. Conclusions: Deletion of LRP1 in SMCs causes progressive aortic dilatation that includes the infrarenal aorta. Pharmacologic inhibition of hepatocyte-derived AGT attenuates infrarenal aortic dilatation.

pathology↗

bioRxiv2026

Angiotensin II Promotes Mitral Valve and Left Atrial Pathology in Fibrillin-1C1041G/+ Mice

Background: Activation of the renin-angiotensin system (RAS) contributes to aortic diseases caused by fibrillin-1 haploinsufficiency (Fbn1C1041G/+). However, whether RAS activation influences mitral valve disease associated with FBN1 variants remains undefined. This study determined whether inhibition or augmentation of the RAS affects mitral valve pathology in Fbn1C1041G/+ mice. Methods: Angiotensinogen (AGT) antisense oligonucleotides (ASOs) were administered for 12 weeks to inhibit the RAS in male and female Fbn1C1041G/+ mice. To augment the RAS, mice were infused with angiotensin II (AngII; 1,000 ng/kg/min) for 28 days. Mitral valve and heart function were monitored by ultrasound. At termination, hearts were dissected free and tissue sectioned to assess mitral valve leaflet thickness and pathology. Results: Mitral valve leaflet thickness was significantly increased in Fbn1C1041G/+ mice compared to Fbn1+/+ littermates of both sexes. ASO-mediated AGT inhibition did not alter this phenotype despite markedly reduced plasma AGT concentrations. AngII infusion increased mitral valve leaflet thickness in Fbn1C1041G/+ mice with increased proteoglycan and collagen content, while exerting minimal effects in Fbn1+/+ mice. Twenty-five percent of AngII-infused Fbn1C1041G/+ mice died of non-vascular causes. Ultrasound showed mitral valve dysfunction in AngII-infused Fbn1C1041G/+ mice. Grossly expanded left atria were common in AngII-infused Fbn1C1041G/+ mice. These markedly enlarged left atria contained increased fibrosis, macrophage accumulation, and platelet-rich masses. Conclusions: Mitral valves of Fbn1C1041G/+ mice did not respond to AGT inhibition but were markedly more susceptible to AngII infusion, with greater increases in thickness and compositional changes than mitral valves of Fbn1+/+ mice. In addition, AngII promoted mitral valve dysfunction that were associated with pronounced left atrial pathology.

pharmacology and toxicology↗

bioRxiv2026

Hepatocyte Angiotensinogen Deletion Protects Against Diet-induced Metabolic Disorders in Mice Under Thermoneutral Conditions

Angiotensinogen (AGT) deletion in hepatocytes reduces Western diet-induced adiposity and hepatic steatosis in mice maintained under conventional room-temperature (RT) housing. Given the high metabolic activity of mice, this temperature imposes adaptive metabolic responses in this species. Whether this metabolic protection persists independent of increased thermogenic demand remains unclear. In this study, we first determined whether thermoneutral housing (TN, 30 {degrees}C) alters Western diet-induced metabolic phenotypes compared with RT housing (20 {degrees}C) in wild-type mice. Although body weight did not differ significantly between housing conditions, Western diet-fed mice housed at TN exhibited brown adipose tissue whitening and more pronounced hepatic steatosis than mice housed at RT, confirming that thermoneutrality exacerbated diet-induced metabolic dysfunction. We then housed hepatocyte Agt deficient (hepAGT-/-) mice and wild-type (hepAGT+/+) littermates at TN and fed them Western diet for 12 weeks. Despite enhanced metabolic dysfunction under TN, hepatocyte AGT deletion resulted in reductions in diet-induced body weight gain, fat mass, liver weight, and hepatic triglyceride accumulation. Bulk RNA sequencing of liver revealed hepatocyte AGT deficiency-dependent alterations in lipid-metabolic pathways. Cross-temperature analysis of RT and TN housing identified 35 shared differentially expressed genes, including 27 concordantly downregulated genes enriched in lipid metabolism and transport. Extended Western diet feeding for 24 weeks confirmed sustained reductions in body weight gain, liver weight, and hepatic lipid accumulation in hepAGT-/- mice. These findings demonstrate that hepatocyte AGT deletion provides sustained protection against Western diet-induced metabolic dysfunction under thermoneutral housing, a condition that more closely recapitulates human basal metabolism. NEW & NOTEWORTHYThis study investigated hepatocyte angiotensinogen (AGT) biology during Western diet feeding in mice under thermoneutral housing, a condition relevant to human metabolism. By minimizing adaptive thermogenesis induced by standard room temperature housing, thermoneutrality more closely recapitulates human basal metabolic conditions. Under this condition, hepatocyte AGT deletion remains protective against adipo and hepatic lipid accumulation, despite exacerbated Western diet-induced metabolic dysfunction in wild-type mice, demonstrating that this protection persists in a human-relevant thermal environment. GRAPHIC ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=132 SRC="FIGDIR/small/742617v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@1d7285aorg.highwire.dtl.DTLVardef@9a401eorg.highwire.dtl.DTLVardef@e79ae9org.highwire.dtl.DTLVardef@16b5742_HPS_FORMAT_FIGEXP M_FIG C_FIG

pathology↗

bioRxiv2026

Elastogenesis by adventitial progenitors acquiring a smooth muscle cell phenotype following aortic dissection

Following aortic dissection (AD), there is a sustained risk of vascular complications, progressive false lumen aneurysm formation, and rupture. However, no effective therapy exists to prevent these complications, highlighting the need to elucidate the pathophysiology following AD. Elastic fibers are crucial for maintaining aortic wall integrity but are thought to have limited regenerative capacity once disrupted during AD. This study defined that elastic fibers were newly generated in the false lumen wall following AD in humans and mice. In human ADs, new elastic fibers were observed in the false lumen wall 6 months after onset. In mice with descending AD induced by {beta}-aminopropionitrile (BAPN), elastin mRNA was markedly upregulated in the chronic phase following AD, accompanied by elastic fiber formation. These fibers coincided with smooth muscle cell (SMC) markers within the false lumen wall. Of note, lineage tracing studies demonstrated that these cells were not derived from resident SMCs but adventitial progenitor cells. In vitro experiments further demonstrated that adventitial progenitor cells produced elastic fibers while expressing SMC markers. Collectively, these findings suggest that adventitial progenitor cells differentiate into elastogenic SMC-like cells, contributing to false lumen remodeling through de novo elastic fiber formation following AD.

pathology↗

bioRxiv2026

Epigenetic Programming of Macrophage Phenotypes by STING-IRF3 Drives Inflammation in Ascending Thoracic Aortic Dissection

BackgroundAscending thoracic aortic dissection (ATAD) is characterized by extensive macrophage (M{Phi}) accumulation and profound inflammation; however, the mechanisms sustaining pro-inflammatory M{Phi} activation remain incompletely defined. Emerging evidence indicates that epigenetically generated immune memory drives innate immune cells toward persistent inflammatory states. In this study, we investigated whether epigenetic reprogramming governs M{Phi} phenotypic fate and contributes to ATAD pathogenesis. MethodsWe performed single-cell RNA sequencing of human ascending aortic tissues from controls, patients with ascending thoracic aortic aneurysm (ATAA), and patients with acute ascending thoracic aortic dissection (ATAD). We also performed integrated single-cell RNA sequencing, single-cell ATAC sequencing, and spatial transcriptomics in an angiotensin II (Ang II)-infused mouse model. The role of the STING-IRF3 signaling axis in M{Phi} epigenetic programming was examined using M{Phi}-Sting -/- and M{Phi}-Irf3-/- mice. ResultsIn human and mouse aortic tissues, we identified multiple functional M{Phi} populations including pro-inflammatory, phagocytic/anti-inflammatory, proliferative, and reparative/healing M{Phi}s. Aortic M{Phi}s in both sporadic ATAD patients and Ang II-induced ATAD mice exhibited a pronounced pro-inflammatory bias with enhanced differentiation toward pro-inflammatory M{Phi}s and impaired differentiation toward phagocytic/anti-inflammatory states. Pro-inflammatory M{Phi}s were particularly abundant in dissection sites, whereas phagocytic M{Phi}s were enriched in discrete adventitial niches. Origin analyses revealed a substantial increase in CCR2 recruited M{Phi}s within the aortic wall, which preferentially differentiated into pro-inflammatory M{Phi}s. In contrast, LYVE1 resident M{Phi}s-- predominantly biased toward phagocytic phenotypes--were markedly depleted in ATAD. Single-cell ATAC sequencing identified coordinated chromatin remodeling with increased accessibility at pro-inflammatory gene loci and decreased accessibility at phagocytic gene loci. Among candidate transcriptional regulators identified, IRF family TFs, including IRF3 emerged as unique factors capable of simultaneously promoting pro-inflammatory gene programs while suppressing phagocytic gene expression. Mechanistically, STING-IRF3 signaling orchestrates this biased transcriptional state, likely through coordinated BRG1-dependent chromatin opening at pro-inflammatory gene loci and chromatin closing at phagocytic/anti-inflammatory gene loci. M{Phi} specific Sting -/- and Irf3-/- mice exhibited attenuated inflammatory reprogramming and reduced aortic destruction and dissection. ConclusionsThese findings identify STING-IRF3-mediated epigenetic programming of M{Phi}s as a fundamental mechanism driving aortic inflammation and ATAD development. Targeting M{Phi} epigenetic programming may represent a promising therapeutic strategy to prevent aortic dissection. Graphic Abstract O_FIG O_LINKSMALLFIG WIDTH=189 HEIGHT=200 SRC="FIGDIR/small/701198v1_ufig1.gif" ALT="Figure 1"> View larger version (29K): org.highwire.dtl.DTLVardef@c97bcdorg.highwire.dtl.DTLVardef@1df0ca8org.highwire.dtl.DTLVardef@b7fd04org.highwire.dtl.DTLVardef@1443e16_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2025

Angiotensin II Induces Abdominal Aortic Branch Aneurysms in Fibrillin-1C1041G/+ Mice

BackgroundMice harboring a missense variant (C1041G) of fibrillin-1 (Fbn1) have been used extensively for aortopathy research, but do not mimic all facets of the human disease. The role of increased angiotensin II (AngII) or blood pressure in determining the arterial phenotype of these mice remains incompletely defined. The purpose of this study was to determine whether AngII, either directly or indirectly through increased blood pressure, promotes pathology in the proximal thoracic aorta and beyond. MethodsFbn1+/+ and Fbn1C1041G/+ littermates were infused with either AngII or norepinephrine (NE) via subcutaneously implanted osmotic pumps. Micro computed tomography (microCT) was used to visualize vascular pathologies. Maximal arterial dimensions were measured using in situ or microCT images. ResultsAngII infusion dramatically augmented aortopathy in Fbn1C1041G/+ mice. Aortic dissection was visible within 3 days of AngII infusion. Over 50% of male Fbn1C1041G/+ mice died during AngII infusion, primarily due to aortic rupture in either the thoracic or abdominal regions. Surviving males had increased ascending and suprarenal aortic diameters and developed pathological lesions at the celiac and superior mesenteric branches of the abdominal aorta. Female mice had a much lower incidence of death but had increased ascending aortic and branch diameters. Although NE infusion also increased systolic blood pressure, it did not affect mortality or enlarge aortic or branch diameters in Fbn1C1041G/+ mice. MicroCT identified novel pathological changes during AngII infusion, including development of aortic branch aneurysms in the celiac and superior mesenteric arteries; however, the maximal diameters of the adjacent suprarenal aorta showed only modest increases in male Fbn1C1041G/+ mice. ConclusionAngII exacerbated aortic pathology in Fbn1C1041G/+ mice. It also promoted development of pathologies at aortic branch points, including the celiac and superior mesenteric arteries.

pathology↗

bioRxiv2025

Cardiac Hemorrhage Precedes Hypertension-induced Fibrosis inPlasminogen Activator Inhibitor-1 Deficient Mice

AimsPlasminogen activator inhibitor-1 (PAI-1) regulates plasmin-mediated proteolysis, thereby influencing vascular stability and tissue remodeling. Angiotensin II (AngII) induces an increase in PAI-1 during the development of ascending thoracic aortic aneurysm (ATAA). The initial purpose of this study was to determine whether deletion of PAI-1 influenced development of ATAA. Subsequently, this study aimed to define the early pathological events preceding cardiac fibrosis in PAI-1 deficiency and the structural domain responsible for its protective effect. Methods and resultsAngII was infused for 4 weeks in whole-body PAI-1 deficient (PAI-1-/-) mice and their wild-type littermates (PAI-1+/+) to examine the role of PAI-1 in ATAA. PAI-1 deficiency did not alter AngII-induced aortopathy but revealed a striking cardiac phenotype characterized by replacement fibrosis predominantly within the epicardium and posterior septum. Ferric iron, indicative of prior hemorrhage, was coincident with fibrosis. Similar phenotypes were observed in PAI-1-/- mice infused with norepinephrine for 4 weeks. To define the pathological events preceding cardiac fibrosis, either AngII or norepinephrine was infused for 1 week in PAI-1+/+ or -/- mice. Both infusions induced extensive epicardial hemorrhage and posterior septal fibrosis in PAI-1-/- mice. To explore the initiation of cardiac pathology, AngII was infused for approximately 1 day. PAI-1-/- mice developed diffuse hemorrhage and cardiomyocyte injury localized to the posterior septum, pathologic changes that preceded overt fibrosis. Finally, to determine the protective domain of PAI-1, saline or AngII was administered to mice harboring loss-of-function point mutations in the protease inhibitory (PAI-1Ala/Ala) or somatomedin B-binding domains (PAI-1AK/AK). Compared to saline infusion, 1 week of AngII induced hemorrhage and heterogeneous fibrosis in PAI-1Ala/Ala, but not PAI-1AK/AK mice. ConclusionsThese findings support that, under hemodynamic stress, PAI-1 deficiency promotes early cardiac hemorrhage and cardiomyocyte injury that lead to fibrosis. Mutational studies implicate dysregulated plasmin generation as an initiator of cardiac injury and fibrosis. TRANSLATIONAL PERSPECTIVECardiac fibrosis has been reported in a human population with PAI-1 deficiency and currently lacks targeted therapy. Our findings demonstrate that in animal models, PAI-1 deficiency confers susceptibility to cardiac injury in response to hemodynamic stress, which may accelerate fibrotic remodeling. Mutational disruption of the protease-inhibitory domain of PAI-1 induced similar pathology, supporting a protective role for this function. These observations suggest that interventions aimed at controlling hypertension, promoting endothelial integrity, or regulating plasmin activation could reduce fibrotic remodeling in this population.

pathology↗

bioRxiv2025

β-aminopropionitrile-induced thoracic aortopathy is refractory to cilostazol and sildenafil in mice

Thoracic aortopathies are life-threatening diseases including aneurysm, dissection, and rupture. Phosphodiesterases (PDEs) regulate intracellular cyclic nucleotide concentrations. Recent studies report the influences of cilostazol, a PDE3 inhibitor, and sildenafil, a PDE5 inhibitor, on abdominal aortic aneurysm formation. However, their impacts on thoracic aortopathy remain unknown. In this study, we investigated whether cilostazol and sildenafil affect thoracic aortopathy induced by {beta}-aminopropionitrile (BAPN) administration in mice. Bulk RNA sequencing analysis revealed that BAPN administration upregulated Pde3a transcription in the ascending aorta and Pde5a in both ascending and descending regions before thoracic aortopathy formation. Next, we tested the effects of cilostazol or sildenafil on BAPN-induced thoracic aortopathy. BAPN-administered mice were fed a diet supplemented with either cilostazol or sildenafil. Mass spectrometry measurements determined the presence of cilostazol or sildenafil in the plasma of mice fed drug-supplemented diets. However, neither drug altered BAPN-induced aortic rupture nor aneurysm formation and progression. These results provide evidence that cilostazol and sildenafil did not influence BAPN-induced thoracic aortopathy in mice.

pathology↗

bioRxiv2025

Smooth Muscle Cells and Fibroblasts in the Proximal Thoracic Aorta Exhibit Minor Differences Between Embryonic Origins in Angiotensin II-driven Transcriptional Alterations

BackgroundThoracic aortopathy is influenced by angiotensin II (AngII) and exhibits regional heterogeneity with the proximal region of the thoracic aorta being susceptible. Smooth muscle cells (SMCs) and selected fibroblasts in this region are derived from two embryonic origins: second heart field (SHF) and cardiac neural crest (CNC). While our previous study revealed a critical role of SHF-derived cells in AngII-mediated aortopathy formation, the contribution of CNC-derived cells remains unclear. MethodsMef2c-Cre R26RmT/mG mice were infused with AngII (1,000 ng/kg/min). Proximal thoracic aortas were harvested at baseline or after 3 days of infusion, representing the prepathological phase. Cells were sorted by origins using mGFP (SHF-derived) and mTomato (other origins, nSHF-derived) signals, respectively. After sorting cells by origin, single-cell RNA sequencing was performed and analyzed. ResultsShort-term AngII infusion induced significant transcriptomic changes in both SHF- and nSHF-derived SMCs, but differences between origins were modest. Fibroblast transcriptomes also underwent notable changes by AngII infusion, but differences between SHF and nSHF origins remained modest. Interestingly, AngII infusion resulted in the emergence of a new fibroblast sub-population. Several molecules related to the extracellular matrix, such as Eln and Col3a1, were downregulated in SHF-derived fibroblasts compared to nSHF-derived fibroblasts in the new subcluster. ConclusionFibroblasts in the new subcluster exhibited lineage-specific differences in extracellular matrix-related genes; however, overall transcriptomic differences between origins in SMCs and fibroblasts in response to AngII were modest in the pre-pathological phase of AngII-induced thoracic aortopathy. GRAPHIC ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=174 SRC="FIGDIR/small/610985v1_ufig1.gif" ALT="Figure 1"> View larger version (38K): org.highwire.dtl.DTLVardef@17c2e3corg.highwire.dtl.DTLVardef@1bf9517org.highwire.dtl.DTLVardef@d51aedorg.highwire.dtl.DTLVardef@dac951_HPS_FORMAT_FIGEXP M_FIG C_FIG

pathology↗

bioRxiv2024

Myeloid-Specific Thrombospondin-1 Deficiency Exacerbates Aortic Rupture via Broad Suppression of Extracellular Matrix Proteins

RationaleRupture of abdominal aortic aneurysms (AAA) is associated with high mortality. However, the precise molecular and cellular drivers of AAA rupture remain elusive. Our prior study showed that global and myeloid-specific deletion of matricellular protein thrombospondin-1 (TSP1) protects mice from aneurysm formation primarily by inhibiting vascular inflammation. ObjectiveTo investigate the cellular and molecular mechanisms that drive AAA rupture by testing how TSP1 deficiency in different cell populations affects the rupture event. Methods and ResultsWe deleted TSP1 in endothelial cells and macrophages --- the major TSP1-expressing cells in aneurysmal tissues ---- by crossbreeding Thbs1 flox/flox mice with VE-cadherin Cre and Lyz2-cre mice, respectively. Aortic aneurysm and rupture were induced by angiotensin II in mice with hypercholesterolemia. Myeloid-specific Thbs1 knockout, but not endothelial-specific knockout, increased the rate of lethal aortic rupture by more than 2 folds. Combined analyses of single-cell RNA sequencing and histology showed a unique cellular and molecular signature of the rupture-prone aorta that was characterized by a broad suppression in inflammation and extracellular matrix production. Visium spatial transcriptomic analysis on human AAA tissues showed a correlation between low TSP1 expression and aortic dissection. ConclusionsTSP1 expression by myeloid cells negatively regulates aneurysm rupture, likely through promoting the matrix repair phenotypes of vascular smooth muscle cells thereby increasing the strength of the vascular wall.

pathology↗

bioRxiv2024

Renal Proximal Tubule Cell-specific Megalin Deletion Induces Tubulointerstitial Nephritis in Mice Fed Western Diet

BackgroundPharmacological inhibition of megalin (also known as low-density lipoprotein receptor-related protein 2: LRP2) attenuates atherosclerosis in hypercholesterolemic mice. Since megalin is abundant in renal proximal tubule cells (PTCs), the purpose of this study was to determine whether PTC-specific deletion of megalin reduces hypercholesterolemia-induced atherosclerosis in mice. MethodsFemale Lrp2 f/f mice were bred with male Ndrg1-Cre ERT2 +/0 mice to develop PTC-LRP2 +/+ and -/- littermates. To study atherosclerosis, all mice were bred to an LDL receptor -/- background and fed a Western diet to induce atherosclerosis. ResultsPTC-specific megalin deletion did not attenuate atherosclerosis in LDL receptor -/- mice in either sex. Serendipitously, we discovered that PTC-specific megalin deletion led to interstitial infiltration of CD68+ cells and tubular atrophy. The pathology was only evident in male PTC-LRP2 -/- mice fed the Western diet, but not in mice fed a normal laboratory diet. Renal pathologies were also observed in male PTC-LRP2 -/- mice in an LDL receptor +/+ background fed the same Western diet, demonstrating that the renal pathologies were dependent on diet and not hypercholesterolemia. In contrast, female PTC-LRP2 -/- mice had no apparent renal pathologies. In vivo multiphoton microscopy demonstrated that PTC-specific megalin deletion dramatically diminished albumin accumulation in PTCs within 10 days of Western diet feeding. RNA sequencing analyses demonstrated the upregulation of inflammation-related pathways in kidney. ConclusionsPTC-specific megalin deletion does not affect atherosclerosis, but leads to tubulointerstitial nephritis in mice fed Western diet, with severe pathologies in male mice.

pathology↗

bioRxiv2023

β-aminopropionitrile Induces Distinct Pathologies in the Ascending and Descending Thoracic Aortic Regions of Young Mice

BACKGROUND{beta}-aminopropionitrile (BAPN) is a pharmacological inhibitor of lysyl oxidase and lysyl oxidase-like proteins. Administration of BAPN promotes aortopathies, although there is a paucity of data on experimental conditions to generate pathology. The objective of this study was to define experimental parameters and determine whether equivalent or variable aortopathies were generated throughout the aortic tree during BAPN administration in mice. METHODSBAPN was administered in drinking water for a period ranging from 1 to 12 weeks. The impacts of BAPN were first assessed with regard to dose, strain, age, and sex. BAPN-induced aortic pathological characterization was conducted using histology and immunostaining. To investigate the mechanistic basis of regional heterogeneity, ascending and descending thoracic aortas were harvested after one week of BAPN administration before the appearance of overt pathology. RESULTSBAPN-induced aortic rupture predominantly occurred or originated in the descending thoracic aorta in young C57BL/6J or N mice. No apparent differences were found between male and female mice. For mice surviving 12 weeks of BAPN administration, profound dilatation was consistently observed in the ascending region, while there were more heterogeneous changes in the descending thoracic region. Pathological features were distinct between the ascending and descending thoracic regions. Aortic pathology in the ascending region was characterized by luminal dilatation and elastic fiber disruption throughout the media. The descending thoracic region frequently had dissections with false lumen formation, collagen deposition, and remodeling of the wall surrounding the false lumen. Cells surrounding the false lumen were predominantly positive for -smooth muscle actin. One week of BAPN administration compromised contractile properties in both regions equivalently, and RNA sequencing did not show obvious differences between the two aortic regions in smooth muscle cell markers, cell proliferation markers, and extracellular components. CONCLUSIONSBAPN-induced pathologies show distinct, heterogeneous features within and between ascending and descending aortic regions in mice.

pathology↗

bioRxiv2023

Regional Heterogeneity of Aortic Histology and Transcriptomes in Young Cynomolgus Monkeys

BackgroundThe regional heterogeneity of vascular components and transcriptomes is an important determinant of aortic biology. This notion has been explored in multiple mouse studies. In the present study, we examined the regional heterogeneity of aortas in non-human primates. MethodsAortic samples were harvested from the ascending, descending, suprarenal, and infrarenal regions of young control monkeys and adult monkeys provided with high fructose for 3 years. The regional heterogeneity of aortic structure and transcriptomes was examined by histological and bulk RNA sequencing analyses. ResultsImmunostaining of CD31 and SMA revealed that endothelial and smooth muscle cells were distributed homogeneously across the aortic regions. In contrast, elastic fibers were less abundant and dispersed in the infrarenal aorta compared to other regions and associated with collagen deposition. Bulk RNA sequencing identified a distinct transcriptome related to the Notch signaling pathway in the infrarenal aorta with significantly increased NOTCH3 mRNA compared to other regions. Immunostaining revealed that NOTCH3 protein was increased in the media of the infrarenal aorta. The abundance of medial NOTCH3 was positively correlated with the dispersion of elastic fibers. Adult cynomolgus monkeys provided with high fructose displayed vascular wall remodeling, such as smooth muscle cell loss and elastic fiber disruption, predominantly in the infrarenal region. The correlation between NOTCH3 and elastic fiber dispersion was enhanced in these monkeys. ConclusionsAortas of young cynomolgus monkeys display regional heterogeneity of their transcriptome and the structure of elastin and collagens. Elastic fibers in the infrarenal aorta are dispersed along with upregulation of medial NOTCH3. HIGHLIGHTS- The present study determined the regional heterogeneity of aortas from cynomolgus monkeys. - Aortas of young cynomolgus monkeys displayed region-specific aortic structure and transcriptomes. - Elastic fibers were dispersed in the infrarenal aorta along with increased NOTCH3 abundance in the media. GRAPHIC ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=113 SRC="FIGDIR/small/530901v2_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@164c52corg.highwire.dtl.DTLVardef@ccfda8org.highwire.dtl.DTLVardef@213064org.highwire.dtl.DTLVardef@f007c3_HPS_FORMAT_FIGEXP M_FIG C_FIG

pathology↗

bioRxiv2022

Functional Exploration of Conserved Sequences in the Distal Face of Angiotensinogen

BackgroundAngiotensinogen (AGT) is an essential component in the renin-angiotensin system. AGT has highly conserved sequences in the loop and {beta}-sheet regions among species; however, their functions have not been studied. MethodsAdeno-associated viral vector (AAV) serotype 2/8 encoding mouse AGT with mutations of conserved sequences in the loop (AAV.loop-Mut), {beta}-sheet (AAV.{beta}sheet-Mut), or both regions (AAV.loop/{beta}sheet-Mut) were injected into male hepatocyte-specific AGT deficient (hepAGT-/-) mice in an LDL receptor -/- background. AAV containing mouse wild-type AGT (AAV.mAGT) or a null vector (AAV.null) were used as controls. Two weeks after AAV administration, all mice were fed a Western diet for 12 weeks. To determine how AGT secretion is regulated in hepatocytes, AAVs containing the above mutations were transducted into HepG2 cells. ResultsIn hepAGT-/- mice infected with AAV.loop-Mut or {beta}sheet-Mut, plasma AGT concentrations, systolic blood pressure, and atherosclerosis were comparable to those in AAV.mAGT-infected mice. Surprisingly, plasma AGT concentrations, systolic blood pressure, and atherosclerotic lesion size in hepAGT-/- mice infected with AAV.loop/{beta}sheet-Mut were not different from mice infected with AAV.null. In contrast, hepatic Agt mRNA abundance was elevated to a comparable magnitude as AAV.mAGT-infected mice. Immunostaining showed that AGT protein was accumulated trol and AAV containing wild-type mouse AGT as a positive control. We have demonstra ted consistently in this and previous studies tht located in the endoplasmic reticulum. ConclusionsThe conserved sequences in either the loop or {beta}-sheet region individually have no effect on AGT regulation, but the conserved sequences in both regions synergistically contribute to the secretion of AGT from hepatocytes. HIGHLIGHTSO_LIThe loop and {beta}-sheet regions in the distal face of angiotensinogen (AGT) have highly conserved sequences across species. C_LIO_LIMutations on either the loop or {beta}-sheet regions do not affect plasma AGT concentrations, blood pressure, and atherosclerosis in hypercholesterolemic mice. C_LIO_LIThe conserved sequences in the loop and {beta}-sheet regions regulate the secretion of AGT from hepatocytes synergistically in vivo and in cultured cells. C_LI

molecular biology↗

bioRxiv2021

Renal Angiotensinogen in Nonhuman Primates is Predominantly Liver-Derived

AGT (Angiotensinogen) is the unique substrate of the renin-angiotensin system. Liver is the primary source of circulating AGT. The present study determined whether hepatocyte-derived AGT regulates renal AGT accumulation by injecting ASO (antisense oligonucleotides) targeting hepatocyte-derived AGT (GalNAc AGT ASO) into female cynomolgus monkeys. Hepatocyte-specific inhibition of AGT led to profound reductions of plasma AGT concentrations. AGT protein in S1 and S2 of renal proximal tubules was greatly diminished by GalNAc AGT ASO. Given the similarity between nonhuman primates and human, our findings support the notion that renal AGT is predominantly derived from liver, and liver regulates renal angiotensin II production in humans. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=148 SRC="FIGDIR/small/444555v2_ufig1.gif" ALT="Figure 1"> View larger version (57K): org.highwire.dtl.DTLVardef@292f5aorg.highwire.dtl.DTLVardef@1e3e0aborg.highwire.dtl.DTLVardef@1b331e3org.highwire.dtl.DTLVardef@1904dba_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2021

GSDMD Deficiency Protects Against Aortic Rupture

ObjectiveAortic ruptures are fatal consequences of aortic aneurysms with macrophage accumulation being a hallmark at the site of ruptures. Pyroptosis is critical in macrophage-mediated inflammation. This study determined effects of pyroptosis on aortic dilation and rupture using GSDMD deficient mice. Approach and ResultsIn an initial study, male Gsdmd+/+ and Gsdmd-/- mice in C57BL/6J background (8 - 10 weeks old) were infected with adeno-associated viral vectors encoding mouse PCSK9D377Y gain-of-function mutation and fed a Western diet to induce hypercholesterolemia. After two weeks of AAV infection, angiotensin II (AngII, 1 {micro}g/kg/min) was infused. During the 4 weeks of AngII infusion, 5 of 13 Gsdmd+/+ mice died of aortic rupture, whereas no aortic rupture occurred in Gsdmd-/- mice. In surviving mice, no differences in either ascending or abdominal aortic dilation were observed between Gsdmd+/+ and Gsdmd-/- mice. To determine whether protection of GSDMD deficiency against aortic rupture is specific to AngII infusion, we subsequently examined aortic pathologies in mice administered beta-aminopropionitrile (BAPN). BAPN (0.5% wt/vol) was administered in drinking water to male Gsdmd+/+ and Gsdmd-/- mice (4 weeks old) for 4 weeks. Six of 13 Gsdmd+/+ mice died of aortic rupture, whereas no aortic rupture occurred in Gsdmd-/- mice. In mice survived, no differences of diameters in the ascending, arch, or abdominal aortic regions were observed between Gsdmd+/+ and Gsdmd-/- mice. ConclusionsGSDMD deficiency protects against AngII or BAPN-induced aortic ruptures in mice. HighlightsO_LIGSDMD deficiency protects against angiotensin II-induced aortic rupture in hypercholesterolemic mice. C_LIO_LIGSDMD deficiency protects against beta-aminopropionitrile (BAPN)-induced aortic dissection and rupture in C57BL/6J mice. C_LI

molecular biology↗

bioRxiv2020

Authentication of In Situ Measurements for Thoracic Aortic Aneurysms in Mice

Aortic diameter is a standard parameter for defining disease severity of thoracic aortic aneurysms. In mouse studies, aortic diameters can be measured in situ directly, but this approach has a potential confounder of underestimation due to the absence of physiological arterial blood pressure. In the present study, we developed an in situ approach for authentic aortic measurements. Thoracic aortic aneurysms were induced by beta-aminopropionitrile (BAPN, 0.5% wt/vol) administration in 4-week-old male C57BL/6J mice. Ultrasonography was performed to examine aortic dimensions, and mice with thoracic aortic dilatations were terminated subsequently. After saline perfusion through the left ventricle, periaortic tissues were removed to expose thoracic aortas. Optimal cutting temperature (OCT) compound was injected via the left ventricle to maintain aortic patency. In situ aortic images were captured pre- and post-OCT injection. In mice with severe thoracic aortic aneurysms, smaller aortic diameters were observed prior to OCT injection compared to ultrasound measurements, while aortic diameters in situ after OCT were comparable to diameters measured using ultrasound. A telemetry system demonstrated that maximal luminal pressures during 150 ul of OCT injection were 90 mmHg. Immunostaining for CD31 revealed that endothelial cells were preserved in the intima after OCT injection. These results indicate that OCT injection does not cause aortic damages due to excess pressures. In conclusion, in situ imaging with OCT injection provides authentic aortic measurements without overt aortic damage in mice with thoracic aortic aneurysms.

pathology↗

bioRxiv2020

Effects of Endogenous Angiotensin II on Abdominal Aortic Aneurysms and Atherosclerosis in Angiotensin II-infused Mice

Angiotensin II (AngII), a major effector of the renin-angiotensin system, exerts critical roles in regulating vascular function. AngII infusion induces abdominal aortic aneurysms (AAAs) and exacerbates atherosclerosis in hypercholesterolemic mice. We determined the effects of AngII infusion on endogenous AngII regulation and AngII-mediated AAAs and atherosclerosis. AngII infusion increased renal, but not plasma, AngII concentrations in male mice. AngI concentrations were decreased modestly in kidney, but more profoundly in plasma, during AngII infusion. Bovine AngII (DRVYVHPF) has one amino acid difference from mouse AngII (DRVYIHPF) that can be distinguished by LC-MS/MS. Therefore, we determined exogenous versus endogenous peptides in mice infused with bovine AngII. Bovine AngII infusion reduced endogenous renal AngII concentrations. To determine whether the residual endogenous AngII exerted an effect on AAAs and atherosclerosis in mice infused with AngII, aliskiren (a direct renin inhibitor) was administered to AngII-infused male LDL receptor deficient mice. Although aliskiren did not attenuate AAAs in AngII-infused mice, atherosclerotic lesion size was reduced. In conclusion, endogenous AngII concentrations are reduced during AngII infusion but still contribute to atherosclerosis, but not AAA, in AngII-infused hypercholesterolemic mice.

pathology↗