Inactivation of Group 1B Phospholipase A2 Enhances Disease Recovery and Reduces Experimental Colitis in Mice.

Phospholipase A2 (PLA2) enzymes influence inflammatory bowel disease in both positive and negative manners depending on the type of PLA2 that is expressed. This study explored the influence of the abundantly expressed Group 1B PLA2 (PLA2G1B) on ulcerative colitis. Wild-type C57BL/6J mice and Pla2g1b-/- mice were treated with dextran sulfate sodium (DSS) for 5 days to induce epithelial injury, followed by another 5 days without DSS for recovery. The Pla2g1b-/- mice displayed significantly less body weight loss, colitis pathology, and disease activity indexes compared to the wild-type mice. The differences in colitis were not due to differences in the colonic lysophospholipid levels, but higher numbers of stem and progenitor cells were found in the intestines of Pla2g1b-/- mice compared to the wild-type mice. The DSS-treated Pla2g1b-/- mice also showed higher expressions of genes that are responsible for epithelial repair and lower expressions of proinflammatory cytokine genes in the colon, as well as reduced inflammatory cytokine levels in the plasma. In vitro experiments revealed the PLA2G1B stimulation of inflammatory cytokine expression by myeloid cells. PLA2G1B inactivation protects against DSS-induced colitis in mice by increasing the intestinal stem cell reservoir for epithelial repair and reducing myeloid cell inflammation in the diseased colon. Thus, PLA2G1B may be a target for colitis management.

Sexual Dimorphism in Lipid Metabolism and Gut Microbiota in Mice Fed a High-Fat Diet.

The gut microbiome plays an essential role in regulating lipid metabolism. However, little is known about how gut microbiome modulates sex differences in lipid metabolism. The present study aims to determine whether gut microbiota modulates sexual dimorphism of lipid metabolism in mice fed a high-fat diet (HFD). Conventional and germ-free male and female mice were fed an HFD for four weeks, and lipid absorption, plasma lipid profiles, and apolipoprotein levels were then evaluated. The gut microbiota was analyzed by 16S rRNA gene sequencing. After 4-week HFD consumption, the females exhibited less body weight gain and body fat composition and significantly lower triglyceride levels in very-low-density lipoprotein (VLDL) and cholesterol levels in high-density lipoprotein (HDL) compared to male mice. The fecal microbiota analysis revealed that the male mice were associated with reduced gut microbial diversity. The female mice had considerably different microbiota composition compared to males, e.g., enriched growth of beneficial microbes (e.g., Akkermansia) and depleted growth of Adlercreutzia and Enterococcus. Correlation analyses suggested that the different compositions of the gut microbiota were associated with sexual dimorphism in body weight, fat mass, and lipid metabolism in mice fed an HFD. Our findings demonstrated significant sex differences in lipid metabolism and the microbiota composition at baseline (during LFD), along with sex-dependent responses to HFD. A comprehensive understanding of sexual dimorphism in lipid metabolism modulated by microbiota will help to develop more sex-specific effective treatment options for dyslipidemia and metabolic disorders in females.

Different sensitivity to diet-induced hyperinsulinemia and hyperglycemia between mice with global or bone marrow-specific apoE receptor-2 deficiency.

This study explored the role of apoE receptor-2 (apoER2), a unique member of the LDL receptor family proteins with a restricted tissue expression profile, in modulating diet-induced obesity and diabetes. Unlike wild-type mice and humans in which chronic feeding of a high-fat Western-type diet leads to obesity and the prediabetic state of hyperinsulinemia before hyperglycemia onset, the Lrp8-/- mice with global apoER2 deficiency displayed lower body weight and adiposity, slower development of hyperinsulinemia, but the accelerated onset of hyperglycemia. Despite their lower adiposity, adipose tissues in Western diet-fed Lrp8-/- mice were more inflamed compared with wild-type mice. Additional experiments revealed that the hyperglycemia observed in Western diet-fed Lrp8-/- mice was due to impaired glucose-induced insulin secretion, ultimately leading to hyperglycemia, adipocyte dysfunction, and inflammation upon chronic feeding of the Western diet. Interestingly, bone marrow-specific apoER2-deficient mice were not defective in insulin secretion, exhibiting increased adiposity and hyperinsulinemia compared with wild-type mice. Analysis of bone marrow-derived macrophages revealed that apoER2 deficiency impeded inflammation resolution with lower secretion of IFN-ß and IL-10 in response to LPS stimulation of IL-4 primed cells. The apoER2-deficient macrophages also showed an increased level of disabled-2 (Dab2) as well as increased cell surface TLR4, suggesting that apoER2 participates in Dab2 regulation of TLR4 signaling. Taken together, these results showed that apoER2 deficiency in macrophages sustains diet-induced tissue inflammation and accelerates obesity and diabetes onset while apoER2 deficiency in other cell types contributes to hyperglycemia and inflammation via defective insulin secretion.

Perivascular adipose tissue promotes vascular dysfunction in murine lupus.

Introduction: Patients with systemic lupus erythematosus (SLE) are at elevated risk for Q10 cardiovascular disease (CVD) due to accelerated atherosclerosis. Compared to heathy control subjects, lupus patients have higher volumes and densities of thoracic aortic perivascular adipose tissue (PVAT), which independently associates with vascular calcification, a marker of subclinical atherosclerosis. However, the biological and functional role of PVAT in SLE has not been directly investigated. Methods: Using mouse models of lupus, we studied the phenotype and function of PVAT, and the mechanisms linking PVAT and vascular dysfunction in lupus disease. Results and discussion: Lupus mice were hypermetabolic and exhibited partial lipodystrophy, with sparing of thoracic aortic PVAT. Using wire myography, we found that mice with active lupus exhibited impaired endothelium-dependent relaxation of thoracic aorta, which was further exacerbated in the presence of thoracic aortic PVAT. Interestingly, PVAT from lupus mice exhibited phenotypic switching, as evidenced by "whitening" and hypertrophy of perivascular adipocytes along with immune cell infiltration, in association with adventitial hyperplasia. In addition, expression of UCP1, a brown/beige adipose marker, was dramatically decreased, while CD45-positive leukocyte infiltration was increased, in PVAT from lupus mice. Furthermore, PVAT from lupus mice exhibited a marked decrease in adipogenic gene expression, concomitant with increased pro-inflammatory adipocytokine and leukocyte marker expression. Taken together, these results suggest that dysfunctional, inflamed PVAT may contribute to vascular disease in lupus.

Apolipoprotein E in Cardiometabolic and Neurological Health and Diseases.

A preponderance of evidence obtained from genetically modified mice and human population studies reveals the association of apolipoprotein E (apoE) deficiency and polymorphisms with pathogenesis of numerous chronic diseases, including atherosclerosis, obesity/diabetes, and Alzheimer's disease. The human APOE gene is polymorphic with three major alleles, ε2, ε3 and ε4, encoding apoE2, apoE3, and apoE4, respectively. The APOE gene is expressed in many cell types, including hepatocytes, adipocytes, immune cells of the myeloid lineage, vascular smooth muscle cells, and in the brain. ApoE is present in subclasses of plasma lipoproteins, and it mediates the clearance of atherogenic lipoproteins from plasma circulation via its interaction with LDL receptor family proteins and heparan sulfate proteoglycans. Extracellular apoE also interacts with cell surface receptors and confers signaling events for cell regulation, while apoE expressed endogenously in various cell types regulates cell functions via autocrine and paracrine mechanisms. This review article focuses on lipoprotein transport-dependent and -independent mechanisms by which apoE deficiency or polymorphisms contribute to cardiovascular disease, metabolic disease, and neurological disorders.

Receptor-independent fluid-phase macropinocytosis promotes arterial foam cell formation and atherosclerosis.

Accumulation of lipid-laden foam cells in the arterial wall plays a central role in atherosclerotic lesion development, plaque progression, and late-stage complications of atherosclerosis. However, there are still fundamental gaps in our knowledge of the underlying mechanisms leading to foam cell formation in atherosclerotic arteries. Here, we investigated the role of receptor-independent macropinocytosis in arterial lipid accumulation and pathogenesis of atherosclerosis. Genetic inhibition of fluid-phase macropinocytosis in myeloid cells (LysMCre+ Nhe1fl/fl) and repurposing of a Food and Drug Administration (FDA)-approved drug that inhibits macrophage macropinocytosis substantially decreased atherosclerotic lesion development in low-density lipoprotein (LDL) receptor-deficient and Apoe-/- mice. Stimulation of macropinocytosis using genetic (H-RASG12V) and physiologically relevant approaches promoted internalization of unmodified native (nLDL) and modified [e.g., acetylated (ac) and oxidized (ox) LDL] lipoproteins in both wild-type and scavenger receptor (SR) knockout (Cd36-/-/Sra-/-) macrophages. Pharmacological inhibition of macropinocytosis in hypercholesterolemic wild-type and Cd36-/-/Sra-/- mice identified an important role of macropinocytosis in LDL uptake by lesional macrophages and development of atherosclerosis. Furthermore, serial section high-resolution imaging, LDL immunolabeling, and three-dimensional (3D) reconstruction of subendothelial foam cells provide visual evidence of lipid macropinocytosis in both human and murine atherosclerotic arteries. Our findings complement the SR paradigm of atherosclerosis and identify a therapeutic strategy to counter the development of atherosclerosis and cardiovascular disease.

Chronic angiotensin receptor activation promotes hepatic triacylglycerol accumulation during an acute glucose challenge in obese-insulin-resistant OLETF rats.

PURPOSE: Angiotensin receptor blockers (ARBs) can ameliorate metabolic syndrome (MetS)-associated dyslipidemia, hepatic steatosis, and glucose intolerance, suggesting that angiotensin receptor (AT1) over-activation contributes to impaired lipid and glucose metabolism, which is characteristic of MetS. The aim of this study was to evaluate changes in the lipid profile and proteins of fatty acid uptake, triacylglycerol (TAG) synthesis, and ß-oxidation to better understand the links between AT1 overactivation and non-alcoholic fatty liver disease (NAFLD) during MetS. METHODS: Four groups of 25-week-old-rats were used: (1) untreated LETO, (2) untreated OLETF, (3) OLETF + angiotensin receptor blocker (ARB; 10 mg olmesartan/kg/d × 8 weeks) and (4) OLETF ± ARB (MINUS; 10 mg olmesartan/kg/d × 4 weeks, then removed until dissection). To investigate the dynamic shifts in metabolism, animals were dissected after an oral glucose challenge (fasting, 3 and 6 h post-glucose). RESULTS: Compared to OLETF, plasma total cholesterol and TAG remained unchanged in ARB. However, liver TAG was 55% lesser in ARB than OLETF, and remained lower throughout the challenge. Basal CD36 and ApoB were 28% and 29% lesser, respectively, in ARB than OLETF. PRDX6 abundance in ARB was 45% lesser than OLETF, and it negatively correlated with liver TAG in ARB. CONCLUSIONS: Chronic blockade of AT1 protects the liver from TAG accumulation during glucose overload. This may be achieved by modulating NEFA uptake and increasing TAG export via ApoB. Our study highlights the contributions of AT1 signaling to impaired hepatic substrate metabolism and the detriments of a high-glucose load and its potential contribution to steatosis during MetS.

Distinct pro-inflammatory properties of myeloid cell-derived apolipoprotein E2 and E4 in atherosclerosis promotion.

Polymorphisms in the apolipoprotein E (apoE) gene are risk factors for chronic inflammatory diseases including atherosclerosis. The gene product apoE is synthesized in many cell types and has both lipid transport-dependent and lipid transport-independent functions. Previous studies have shown that apoE expression in myeloid cells protects against atherogenesis in hypercholesterolemic ApoE-/- mice. However, the mechanism of this protection is still unclear. Using human APOE gene replacement mice as models, this study showed that apoE2 and apoE4 expressed endogenously in myeloid cells enhanced the inflammatory response via mechanisms independent of plasma lipoprotein transport. The data revealed that apoE2-expressing myeloid cells contained higher intracellular cholesterol levels because of impaired efflux, causing increasing inflammasome activation and myelopoiesis. In contrast, intracellular cholesterol levels were not elevated in apoE4-expressing myeloid cells, and its proinflammatory property was found to be independent of inflammasome signaling and related to enhanced oxidative stress. When ApoE-/- mice were reconstituted with bone marrow from various human APOE gene replacement mice, effective reduction of atherosclerosis was observed with marrow cells obtained from APOE3 but not APOE2 and APOE4 gene replacement mice. Taken together, these results documented that apoE2 and apoE4 expression in myeloid cells promotes inflammation via distinct mechanisms and promotes atherosclerosis in a plasma lipoprotein transport-independent manner.

Protein Phosphatase 2A Activation Via ApoER2 in Trophoblasts Drives Preeclampsia in a Mouse Model of the Antiphospholipid Syndrome.

[Figure: see text].

Apolipoprotein E receptor 2 deficiency decreases endothelial adhesion of monocytes and protects against autoimmune encephalomyelitis.

Under normal conditions, the blood-brain barrier effectively regulates the passage of immune cells into the central nervous system (CNS). However, under pathological conditions such as multiple sclerosis (MS), leukocytes, especially monocytes, infiltrate the CNS where they promote inflammatory demyelination, resulting in paralysis. Therapies targeting the immune cells directly and preventing leukocyte infiltration exist for MS but may compromise the immune system. Here, we explore how apolipoprotein E receptor 2 (ApoER2) regulates vascular adhesion and infiltration of monocytes during inflammation. We induced experimental autoimmune encephalitis in ApoER2 knockout mice and in mice carrying a loss-of-function mutation in the ApoER2 cytoplasmic domain. In both models, paralysis and neuroinflammation were largely abolished as a result of greatly diminished monocyte adherence due to reduced expression of adhesion molecules on the endothelial surface. Our findings expand our mechanistic understanding of the vascular barrier, the regulation of inflammation and vascular permeability, and the therapeutic potential of ApoER2-targeted therapies.

LDL receptor-related protein 1 and its interacting partners in tissue homeostasis.

PURPOSE OF REVIEW: LDL receptor-related protein 1 (LRP1) is a multifunctional protein with endocytic and signal transduction properties due to its interaction with numerous extracellular ligands and intracellular proteins. This brief review highlights key developments in identifying novel functions of LRP1 in liver, lung, and the central nervous system in disease pathogenesis. RECENT FINDINGS: In hepatocytes, LRP1 complexes with phosphatidylinositol 4-phosphate 5-kinase-1 and its related protein to maintain intracellular levels of phosphatidylinositol (4,5) bisphosphate and preserve lysosome and mitochondria integrity. In contrast, in smooth muscle cells, macrophages, and endothelial cells, LRP1 interacts with various different extracellular ligands and intracellular proteins in a tissue-dependent and microenvironment-dependent manner to either enhance or suppress inflammation, disease progression or resolution. Similarly, LRP1 expression in astrocytes and oligodendrocyte progenitor cells regulates cell differentiation and maturation in a developmental-dependent manner to modulate neurogenesis, gliogenesis, and white matter repair after injury. SUMMARY: LRP1 modulates metabolic disease manifestation, inflammation, and differentiation in a cell-dependent, time-dependent, and tissue-dependent manner. Whether LRP1 expression is protective or pathogenic is dependent on its interaction with specific ligands and intracellular proteins, which in turn is dependent on the cell type and the microenvironment where these cells reside.

Hepatic LDL receptor-related protein-1 deficiency alters mitochondrial dynamics through phosphatidylinositol 4,5-bisphosphate reduction.

The LDL receptor-related protein 1 (LRP1) is a multifunctional transmembrane protein with endocytosis and signal transduction functions. Previous studies have shown that hepatic LRP1 deficiency exacerbates diet-induced steatohepatitis and insulin resistance via mechanisms related to increased lysosome and mitochondria permeability and dysfunction. The current study examined the impact of LRP1 deficiency on mitochondrial function in the liver. Hepatocytes isolated from liver-specific LRP1 knockout (hLrp1-/-) mice showed reduced oxygen consumption compared with control mouse hepatocytes. The mitochondria in hLrp1-/- mouse livers have an abnormal morphology and their membranes contain significantly less anionic phospholipids, including lower levels of phosphatidylethanolamine and cardiolipin that increase mitochondrial fission and impair fusion. Additional studies showed that LRP1 complexes with phosphatidylinositol 4-phosphate 5-kinase like protein-1 (PIP5KL1) and phosphatidylinositol 4-phosphate 5-kinase-1ß (PIP5K1ß). The absence of LRP1 reduces the levels of both PIP5KL1 and PIP5K1ß in the plasma membrane and also lowers phosphatidylinositol(4,5) bisphosphate (PI(4,5)P2) levels in hepatocytes. These data indicate that LRP1 recruits PIP5KL1 and PIP5K1ß to the plasma membrane for PI(4,5)P2 biosynthesis. The lack of LRP1 reduces lipid kinase expression, leading to lower PI(4,5)P2 levels, thereby decreasing the availability of this lipid metabolite in the cardiolipin biosynthesis pathway to cause cardiolipin reduction and the impairment in mitochondria homeostasis. Taken together, the current study identifies another signaling mechanism by which LRP1 regulates cell functions: binding and recruitment of PIP5KL1 and PIP5K1ß to the membrane for PI(4,5)P2 synthesis. In addition, it highlights the importance of this mechanism for maintaining the integrity and functions of intracellular organelles.

Mutation in the distal NPxY motif of LRP1 alleviates dietary cholesterol-induced dyslipidemia and tissue inflammation.

The impairment of LDL receptor-related protein-1 (LRP1) in numerous cell types is associated with obesity, diabetes, and fatty liver disease. Here, we compared the metabolic phenotype of C57BL/6J wild-type and LRP1 knock-in mice carrying an inactivating mutation in the distal NPxY motif after feeding a low-fat diet or high-fat (HF) diet with cholesterol supplementation (HFHC) or HF diet without cholesterol supplementation. In response to HF feeding, both groups developed hyperglycemia, hyperinsulinemia, hyperlipidemia, increased adiposity, and adipose tissue inflammation and liver steatosis. However, LRP1 NPxY mutation prevents HFHC diet-induced hypercholesterolemia, reduces adipose tissue and brain inflammation, and limits liver progression to steatohepatitis. Nevertheless, this mutation does not protect against HFHC diet-induced insulin resistance. The selective metabolic improvement observed in HFHC diet-fed LRP1 NPxY mutant mice is due to an apparent increase of hepatic LDL receptor levels, leading to an elevated rate of plasma lipoprotein clearance and lower hepatic cholesterol levels. The unique metabolic phenotypes displayed by LRP1 NPxY mutant mice indicate an LRP1-cholesterol axis in modulating tissue inflammation. The LRP1 NPxY mutant mouse phenotype differs from phenotypes observed in mice with tissue-specific LRP1 inactivation, thus highlighting the importance of an integrative approach to evaluate how global LRP1 dysfunction contributes to metabolic disease development.

Mice lacking global Stap1 expression do not manifest hypercholesterolemia.

BACKGROUND: Autosomal dominant familial hypercholesterolemia (ADH; MIM#143890) is one of the most common monogenic disorders characterized by elevated circulatory LDL cholesterol. Initial studies in humans with ADH identified a potential relationship with variants of the gene encoding signal transducing adaptor family member protein 1 (STAP1; MIM#604298). However, subsequent studies have been contradictory. In this study, mice lacking global Stap1 expression (Stap1-/-) were characterized under standard chow and a 42% kcal western diet (WD). METHODS: Mice were studied for changes in different metabolic parameters before and after a 16-week WD regime. Growth curves, body fats, circulatory lipids, parameters of glucose homeostasis, and liver architecture were studied for comparisons. RESULTS: Surprisingly, Stap1-/- mice fed the 16-week WD demonstrated no marked differences in any of the metabolic parameters compared to Stap1+/+ mice. Furthermore, hepatic architecture and cholesterol content in FPLC-isolated lipoprotein fractions also remained comparable to wild-type mice. CONCLUSION: These results strongly suggest that STAP1 does not alter lipid levels, that a western diet did not exacerbate a lipid disorder in Stap1 deficient mice and support the contention that it is not causative for hyperlipidemia in ADH patients. These results support other published studies also questioning the role of this locus in human hypercholesterolemia.

Generating Multiscale Amorphous Molecular Structures Using Deep Learning: A Study in 2D.

Amorphous molecular assemblies appear in a vast array of systems: from living cells to chemical plants and from everyday items to new devices. The absence of long-range order in amorphous materials implies that precise knowledge of their underlying structures throughout is needed to rationalize and control their properties at the mesoscale. Standard computational simulations suffer from exponentially unfavorable scaling of the required compute with system size. We present a method based on deep learning that leverages the finite range of structural correlations for an autoregressive generation of disordered molecular aggregates up to arbitrary size from small-scale computational or experimental samples. We benchmark performance on self-assembled nanoparticle aggregates and proceed to simulate monolayer amorphous carbon with atomistic resolution. This method bridges the gap between the nanoscale and mesoscale simulations of amorphous molecular systems.

Distinct Influence of Hypercaloric Diets Predominant with Fat or Fat and Sucrose on Adipose Tissue and Liver Inflammation in Mice.

Overfeeding of a hypercaloric diet leads to obesity, diabetes, chronic inflammation, and fatty liver disease. Although limiting fat or carbohydrate intake is the cornerstone for obesity management, whether lowering fat or reducing carbohydrate intake is more effective for health management remains controversial. This study used murine models to determine how dietary fat and carbohydrates may influence metabolic disease manifestation. Age-matched C57BL/6J mice were fed 2 hypercaloric diets with similar caloric content, one with very high fat and low carbohydrate content (VHF) and the other with moderately high fat levels with high sucrose content (HFHS) for 12 weeks. Both groups gained more weight and displayed hypercholesterolemia, hyperglycemia, hyperinsulinemia, and liver steatosis compared to mice fed a normal low-fat (LF) diet. Interestingly, the VHF-fed mice showed a more robust adipose tissue inflammation compared to HFHS-fed mice, whereas HFHS-fed mice showed liver fibrosis and inflammation that was not observed in VHF-fed mice. Taken together, these results indicate macronutrient-specific tissue inflammation with excess dietary fat provoking adipose tissue inflammation, whereas moderately high dietary fat with extra sucrose is necessary and sufficient for hepatosteatosis advancement to steatohepatitis. Hence, liver and adipose tissues respond to dietary fat and sucrose in opposite manners, yet both macronutrients are contributing factors to metabolic diseases.

Hepatic HAX-1 inactivation prevents metabolic diseases by enhancing mitochondrial activity and bile salt export.

Increasing hepatic mitochondrial activity through pyruvate dehydrogenase and elevating enterohepatic bile acid recirculation are promising new approaches for metabolic disease therapy, but neither approach alone can completely ameliorate disease phenotype in high-fat diet-fed mice. This study showed that diet-induced hepatosteatosis, hyperlipidemia, and insulin resistance can be completely prevented in mice with liver-specific HCLS1-associated protein X-1 (HAX-1) inactivation. Mechanistically, we showed that HAX-1 interacts with inositol 1,4,5-trisphosphate receptor-1 (InsP3R1) in the liver, and its absence reduces InsP3R1 levels, thereby improving endoplasmic reticulum-mitochondria calcium homeostasis to prevent excess calcium overload and mitochondrial dysfunction. As a result, HAX-1 ablation activates pyruvate dehydrogenase and increases mitochondria utilization of glucose and fatty acids to prevent hepatosteatosis, hyperlipidemia, and insulin resistance. In contrast to the reduction of InsP3R1 levels, hepatic HAX-1 deficiency increases bile salt exporter protein levels, thereby promoting enterohepatic bile acid recirculation, leading to activation of bile acid-responsive genes in the intestinal ileum to augment insulin sensitivity and of cholesterol transport genes in the liver to suppress hyperlipidemia. The dual mechanisms of increased mitochondrial respiration and enterohepatic bile acid recirculation due to improvement of endoplasmic reticulum-mitochondria calcium homeostasis with hepatic HAX-1 inactivation suggest that this may be a potential therapeutic target for metabolic disease intervention.

Mycobacterium bovis Bacille-Calmette-Guérin Infection Aggravates Atherosclerosis.

Tuberculosis has been associated with increased risk of atherosclerotic cardiovascular disease. To examine whether mycobacterial infection exacerbates atherosclerosis development in experimental conditions, we infected low-density lipoprotein receptor knockout (Ldlr-/-) mice with Mycobacterium bovis Bacille-Calmette-Guérin (BCG), an attenuated strain of the Mycobacterium tuberculosis complex. Twelve-week old male Ldlr-/- mice were infected with BCG (0.3-3.0x106 colony-forming units) via the intranasal route. Mice were subsequently fed a western-type diet containing 21% fat and 0.2% cholesterol for up to 16 weeks. Age-matched uninfected Ldlr-/- mice fed with an identical diet served as controls. Atherosclerotic lesions in aorta were examined using Oil Red O staining. Changes induced by BCG infection on the immunophenotyping profile of circulating T lymphocytes and monocytes were assessed using flow cytometry. BCG infection increased atherosclerotic lesions in en face aorta after 8 weeks (plaque ratio; 0.021±0.01 vs. 0.013±0.01; p = 0.011) and 16 weeks (plaque ratio, 0.15±0.13 vs. 0.06±0.02; p = 0.003). No significant differences in plasma cholesterol or triglyceride levels were observed between infected and uninfected mice. Compared to uninfected mice, BCG infection increased systemic CD4/CD8 T cell ratio and the proportion of Ly6Clow non-classical monocytes at weeks 8 and 16. Aortic plaque ratios correlated with CD4/CD8 T cell ratios (Spearman's rho = 0.498; p = 0.001) and the proportion of Ly6Clow non-classical monocytes (Spearman's rho = 0.629; p < 0.001) at week 16. In conclusion, BCG infection expanded the proportion of CD4+ T cell and Ly6Clow monocytes, and aggravated atherosclerosis formation in the aortas of hyperlipidemic Ldlr-/- mice. Our results indicate that mycobacterial infection is capable of enhancing atherosclerosis development.

Prefrontal Cortex Regulates Chronic Stress-Induced Cardiovascular Susceptibility.

Background The medial prefrontal cortex is necessary for appropriate appraisal of stressful information, as well as coordinating visceral and behavioral processes. However, prolonged stress impairs medial prefrontal cortex function and prefrontal-dependent behaviors. Additionally, chronic stress induces sympathetic predominance, contributing to health detriments associated with autonomic imbalance. Previous studies identified a subregion of rodent prefrontal cortex, infralimbic cortex (IL), as a key regulator of neuroendocrine-autonomic integration after chronic stress, suggesting that IL output may prevent chronic stress-induced autonomic imbalance. In the current study, we tested the hypothesis that the IL regulates hemodynamic, vascular, and cardiac responses to chronic stress. Methods and Results A viral-packaged small interfering RNA construct was used to knockdown vesicular glutamate transporter 1 (vGluT1) and reduce glutamate packaging and release from IL projection neurons. Male rats were injected with a vGluT1 small interfering RNA-expressing construct or GFP (green fluorescent protein) control into the IL and then remained as unstressed controls or were exposed to chronic variable stress. IL vGluT1 knockdown increased heart rate and mean arterial pressure reactivity, while chronic variable stress increased chronic mean arterial pressure only in small interfering RNA-treated rats. In another cohort, chronic variable stress and vGluT1 knockdown interacted to impair both endothelial-dependent and endothelial-independent vasoreactivity ex vivo. Furthermore, vGluT1 knockdown and chronic variable stress increased histological markers of fibrosis and hypertrophy. Conclusions Knockdown of glutamate release from IL projection neurons indicates that these cells are necessary to prevent the enhanced physiological responses to stress that promote susceptibility to cardiovascular pathophysiology. Ultimately, these findings provide evidence for a neurobiological mechanism mediating the relationship between stress and poor cardiovascular health outcomes.

ApoER2 (Apolipoprotein E Receptor-2) Deficiency Accelerates Smooth Muscle Cell Senescence via Cytokinesis Impairment and Promotes Fibrotic Neointima After Vascular Injury.

OBJECTIVE: Genome-wide studies showed that mutation in apoER2 (apolipoprotein E receptor-2) is additive with ε4 polymorphism in the APOE gene on cardiovascular disease risk in humans. ApoE or apoER2 deficiency also accelerates atherosclerosis lesion necrosis in hypercholesterolemic mice and promotes neointima formation after vascular injury. This study tests the hypothesis that apoE and apoER2 modulate vascular occlusive diseases through distinct mechanisms. Approach and Results: Carotid endothelial denudation induced robust neointima formation in both apoE-/- and apoER2-deficient Lrp8-/- mice. The intima in apoE-/- mice was rich in smooth muscle cells, but the intima in Lrp8-/- mice was cell-poor and rich in extracellular matrix. Vascular smooth muscle cells isolated from apoE-/- mice were hyperplastic whereas Lrp8-/- smooth muscle cells showed reduced proliferation but responded robustly to TGF (transforming growth factor)-ß-induced fibronectin synthesis indicative of a senescence-associated secretory phenotype, which was confirmed by increased ß-galactosidase activity, p16INK4a immunofluorescence, and number of multinucleated cells. Western blot analysis of cell cycle-associated proteins showed that apoER2 deficiency promotes cell cycle arrest at the metaphase/anaphase. Coimmunoprecipitation experiments revealed that apoER2 interacts with the catalytic subunit of protein phosphatase 2A. In the absence of apoER2, PP2A-C (protein phosphatase 2A catalytic subunit) failed to interact with CDC20 (cell-division cycle protein 20) thus resulting in inactive anaphase-promoting complex and impaired cell cycle exit. CONCLUSIONS: This study showed that apoER2 participates in APC (anaphase-promoting complex)/CDC20 complex formation during mitosis, and its absence impedes cytokinesis abscission thereby accelerating premature cell senescence and vascular disease. This mechanism is distinct from apoE deficiency, which causes smooth muscle cell hyperplasia to accelerate vascular disease.