Lipoprotein Receptor Research Articles

LOX-1 in Cardiovascular Disease: A Comprehensive Molecular and Clinical Review.

LOX-1, ORL-1, or lectin-like oxidized low-density lipoprotein receptor 1 is a transmembrane glycoprotein that binds and internalizes ox-LDL in foam cells. LOX-1 is the main receptor for oxidized low-density lipoproteins (ox-LDL). The LDL comes from food intake and circulates through the bloodstream. LOX-1 belongs to scavenger receptors (SR), which are associated with various cardiovascular diseases. The most important and severe of these is the formation of atherosclerotic plaques in the intimal layer of the endothelium. These plaques can evolve into complicated thrombi with the participation of fibroblasts, activated platelets, apoptotic muscle cells, and macrophages transformed into foam cells. This process causes changes in vascular endothelial homeostasis, leading to partial or total obstruction in the lumen of blood vessels. This obstruction can result in oxygen deprivation to the heart. Recently, LOX-1 has been involved in other pathologies, such as obesity and diabetes mellitus. However, the development of atherosclerosis has been the most relevant due to its relationship with cerebrovascular accidents and heart attacks. In this review, we will summarize findings related to the physiologic and pathophysiological processes of LOX-1 to support the detection, diagnosis, and prevention of those diseases.

Radical Oxygen Species, Oxidized Low-Density Lipoproteins, and Lectin-like Oxidized Low-Density Lipoprotein Receptor 1: A Vicious Circle in Atherosclerotic Process.

Atherosclerosis is a complex condition that involves the accumulation of lipids and subsequent plaque formation in the arterial intima. There are various stimuli, cellular receptors, and pathways involved in this process, but oxidative modifications of low-density lipoprotein (ox-LDL) are particularly important in the onset and progression of atherosclerosis. Ox-LDLs promote foam-cell formation, activate proinflammatory pathways, and induce smooth-muscle-cell migration, apoptosis, and cell death. One of the major receptors for ox-LDL is LOX-1, which is upregulated in several cardiovascular diseases, including atherosclerosis. LOX-1 activation in endothelial cells promotes endothelial dysfunction and induces pro-atherogenic signaling, leading to plaque formation. The binding of ox-LDLs to LOX-1 increases the generation of reactive oxygen species (ROS), which can induce LOX-1 expression and oxidize LDLs, contributing to ox-LDL generation and further upregulating LOX-1 expression. This creates a vicious circle that is amplified in pathological conditions characterized by high plasma levels of LDLs. Although LOX-1 has harmful effects, the clinical significance of inhibiting this protein remains unclear. Further studies both in vitro and in vivo are needed to determine whether LOX-1 inhibition could be a potential therapeutic target to counteract the atherosclerotic process.

Spon1+ inflammatory monocytes promote collagen remodeling and lung cancer metastasis through lipoprotein receptor 8 signaling.

Lung cancer is the leading cause of cancer-related deaths in the world, and non-small cell lung cancer (NSCLC) is the most common subset. We previously found that infiltration of tumor inflammatory monocytes (TIMs) into lung squamous carcinoma (LUSC) tumors is associated with increased metastases and poor survival. To further understand how TIMs promote metastases, we compared RNA-Seq profiles of TIMs from several LUSC metastatic models with inflammatory monocytes (IMs) of non-tumor-bearing controls. We identified Spon1 as upregulated in TIMs and found that Spon1 expression in LUSC tumors corresponded with poor survival and enrichment of collagen extracellular matrix signatures. We observed SPON1+ TIMs mediate their effects directly through LRP8 on NSCLC cells, which resulted in TGF-β1 activation and robust production of fibrillar collagens. Using several orthogonal approaches, we demonstrated that SPON1+ TIMs were sufficient to promote NSCLC metastases. Additionally, we found that Spon1 loss in the host, or Lrp8 loss in cancer cells, resulted in a significant decrease of both high-density collagen matrices and metastases. Finally, we confirmed the relevance of the SPON1/LRP8/TGF-β1 axis with collagen production and survival in patients with NSCLC. Taken together, our study describes how SPON1+ TIMs promote collagen remodeling and NSCLC metastases through an LRP8/TGF-β1 signaling axis.

Cholesterol uptake in the intestine is regulated by the LASP1-AKT-NPC1L1 signaling pathway.

Cholesterol is essential for the stability and architecture of the plasma membrane and a precursor of bile acids and steroid hormones in mammals. Excess dietary cholesterol uptake leads to hypercholesterolemia and atherosclerosis and plays a role in cancer development. The role of actin-binding scaffolding protein LIM and SH3 protein 1 (LASP1) in cholesterol trafficking has not been investigated previously. Cholesterol levels, its uptake, and excretion were studied in mice deficient for low-density lipoprotein receptor and Lasp1 (Ldlr-/-Lasp1-/- mice) upon feeding a high-fat diet, and in LASP1-knockdown, differentiated human intestinal epithelial CaCo-2 cells. When compared with diet-fed Ldlr-/- control mice, Ldlr-/-Lasp1-/- mice displayed a reduction in serum cholesterol levels. Mechanistically, we identified a new role of LASP1 in controlling the translocation of the intestinal cholesterol transporter Niemann-Pick C1-like 1 (NPC1L1) to the apical cell surface, which was limited in LASP1-knockdown human CaCo-2 enterocytes and in the intestine of Ldlr-/- Lasp1-/- compared with Ldlr-/- mice, linked to LASP1-pAKT signaling but not CDC42 activation. In line, a reduction in cholesterol reabsorption was noted in LASP1-knockdown CaCo-2 cells in vitro, and an enhanced cholesterol excretion via the feces was observed in Ldlr-/- Lasp1-/- mice. These data uncover a novel function of Lasp1 in cholesterol trafficking, promoting cholesterol reabsorption in the intestine. Targeting LASP1 locally could thus represent a novel targeting strategy to ameliorate hypercholesterolemia and associated diseases.NEW & NOTEWORTHY We here uncovered LASP1 as a novel regulator of the shuttling of the sterol transporter NPC1L1 to the cell surface in enterocytes to control cholesterol absorption. Accordingly, LASP1-deficient mice displayed lowered serum cholesterol levels under dietary cholesterol supplementation.

Chandipura viral glycoprotein (CNV-G) promotes Gectosome generation and enables delivery of intracellular therapeutics

Overexpression of vesicular stomatitis virus G protein (VSV-G) elevates the secretion of EVs known as gectosomes, which contain VSV-G. Such vesicles can be engineered to deliver therapeutic macromolecules. We investigated viral glycoproteins from several viruses for their potential in gectosome production and intracellular cargo delivery. Expression of the viral glycoprotein (viral glycoprotein from the Chandipura virus [CNV-G]) from the human neurotropic pathogen Chandipura virus in 293T cells significantly augments the production of CNV-G-containing gectosomes. In comparison with VSV-G gectosomes, CNV-G gectosomes exhibit heightened selectivity toward specific cell types, including primary cells and tumor cell lines. Consistent with the differential tropism between CNV-G and VSV-G gectosomes, cellular entry of CNV-G gectosome is independent of the Low-density lipoprotein receptor, which is essential for VSV-G entry, and shows varying sensitivity to pharmacological modulators. CNV-G gectosomes efficiently deliver diverse intracellular cargos for genomic modification or responses to stimuli invitro and in the brain of mice invivo utilizing a split GFP and chemical-induced dimerization system. Pharmacokinetics and biodistribution analyses support CNV-G gectosomes as a versatile platform for delivering macromolecular therapeutics intracellularly.

Trem2 Agonist Reprograms Foamy Macrophages to Promote Atherosclerotic Plaque Stability-Brief Report.

Trem2 (triggering receptor on myeloid cells 2), a surface lipid receptor, is expressed on foamy macrophages within atherosclerotic lesions and regulates cell survival, proliferation, and anti-inflammatory responses. Studies examining the role of Trem2 in atherosclerosis have shown that deletion of Trem2 leads to impaired foamy macrophage lipid uptake, proliferation, survival, and cholesterol efflux. Thus, we tested the hypothesis that administration of a Trem2 agonist antibody (AL002a) to atherogenic mice would enhance macrophage survival and decrease necrotic core formation to improve plaque stability. To model a therapeutic intervention approach, atherosclerosis-prone mice (Ldlr [low-density lipoprotein receptor]-/-) were fed a high-fat diet for 8 weeks, then transitioned to treatment with AL002a or isotype control for an additional 8 weeks while continuing on a high-fat diet. AL002a-treated mice had increased lesion size in both the aortic root and whole mount aorta, which correlated with an expansion of plaque macrophage area. This expansion was due to increased macrophage survival and proliferation in plaques. Importantly, plaques from AL002a-treated mice showed improved features of plaque stability, including smaller necrotic cores, increased fibrous caps, and greater collagen deposition. Single-cell RNA sequencing of whole aorta suspensions from isotype- and AL002a-treated atherosclerotic mice revealed that Trem2 agonism dramatically altered foamy macrophage transcriptome. This included upregulation of oxidative phosphorylation and increased expression of collagen genes. In vitro studies validated that Trem2 agonism with AL002a promoted foamy macrophage oxidized low-density lipoprotein uptake, survival, and cholesterol efflux. Trem2 agonism expands atherosclerotic plaque macrophages by promoting cell survival and proliferation but improves features of plaque stability by rewiring foamy macrophage function to enhance cholesterol efflux and collagen deposition.

Cytotoxic sigma-2 ligands trigger cancer cell death via cholesterol-induced-ER-stress

Sigma-2-ligands (S2L) are characterized by high binding affinities to their cognate sigma-2 receptor, overexpressed in rapidly proliferating tumor cells. As such, S2L were developed as imaging probes (ISO1) or as cancer therapeutics, alone (SV119 [C6], SW43 [C10]) and as delivery vehicles for cytotoxic drug cargoes (C6-Erastin, C10-SMAC). However, the exact mechanism of S2L-induced cytotoxicity remains to be fully elucidated. A series of high-affinity S2L were evaluated regarding their cytotoxicity profiles across cancer cell lines. While C6 and C10 displayed distinct cytotoxicities, C0 and ISO1 were essentially non-toxic. Confocal microscopy and lipidomics analysis in cellular and mouse models revealed that C10 induced increases in intralysosomal free cholesterol and in cholesterol esters, suggestive of unaltered intracellular cholesterol trafficking. Cytotoxicity was caused by cholesterol excess, a phenomenon that contrasts the effects of NPC1 inhibition. RNA-sequencing revealed gene clusters involved in cholesterol homeostasis and ER stress response exclusively by cytotoxic S2L. ER stress markers were confirmed by qPCR and their targeted modulation inhibited or enhanced cytotoxicity of C10 in a predicted manner. Moreover, C10 increased sterol regulatory element-binding protein 2 (SREBP2) and low-density lipoprotein receptor (LDLR), both found to be pro-survival factors activated by ER stress. Furthermore, inhibition of downstream processes of the adaptive response to S2L with simvastatin resulted in synergistic treatment outcomes in combination with C10. Of note, the S2L conjugates retained the ER stress response of the parental ligands, indicative of cholesterol homeostasis being involved in the overall cytotoxicity of the drug conjugates. Based on these findings, we conclude that S2L-mediated cell death is due to free cholesterol accumulation that leads to ER stress. Consequently, the cytotoxic profiles of S2L drug conjugates are proposed to be enhanced via concurrent ER stress inducers or simvastatin, strategies that could be instrumental on the path toward tumor eradication.

The CD36 scavenger receptor Bez regulates lipid redistribution from fat body to ovaries in Drosophila.

Lipid distribution in an organism is mediated by the interplay between lipoprotein particles, lipoprotein receptors and class B scavenger receptors of the CD36 family. CD36 is a multifunctional protein mediating lipid uptake, mobilization and signaling at the plasma membrane and inside of the cell. The CD36 protein family has 14 members in Drosophila melanogaster, which allows for the differentiated analysis of their functions. Here, we unravel a role for the so far uncharacterized scavenger receptor Bez in lipid export from Drosophila adipocytes. Bez shares the lipid binding residue with CD36 and is expressed at the plasma membrane of the embryonic, larval and adult fat body. Bez loss of function lowers the organismal availability of storage lipids and blocks the maturation of egg chambers in ovaries. We demonstrate that Bez interacts with the APOB homolog Lipophorin at the plasma membrane of adipocytes and trace the Bez-dependent transfer of an alkyne-labeled fatty acid from adipocytes to Lipophorin. Our study demonstrates how lipids are distributed by scavenger receptor-lipoprotein interplay and contribute to the metabolic control of development.

Abstract 2004: Sex Differences In Bile Acid Metabolism Impact Cholesterol Homeostasis And Cardiovascular Disease

Cardiovascular disease (CVD) remains the leading cause of death in the United States for both men and women, despite cholesterol-lowering drugs such as statins. Diets rich in cholesterol are well-known to contribute to disease progression. The absorption of dietary cholesterol is facilitated by liver-synthesized detergents called bile acids. We hypothesized that manipulating bile acid metabolism to reduce cholesterol absorption would result in decreased plasma cholesterol levels and protect from atherosclerosis. To alter the bile acid pool, we used liver-directed AAV-CRISPR to disrupt Cyp7a1, which catalyzes the rate-limiting step of bile acid synthesis. Consequently, we show that loss of CYP7A1 in adult male mice reduces the size of the bile acid pool. To study atherosclerosis, we co-disrupted Cyp7a1 concurrently with Low Density Lipoprotein Receptor ( Ldlr ) to induce hypercholesterolemia on a Western diet for 20 weeks. Loss of hepatic CYP7A1 in males reduced atherosclerosis and reduced cholesterol absorption. In stark contrast to what we observed in male mice, the absence of this rate limiting enzyme in female mice resulted in only a modest reduction in the bile acid pool. This blunted reduction in bile acids in female Cyp7a1 CRISPR mice was not sufficient to decrease cholesterol absorption, resulting in accumulation of cholesterol, and consequently increasing cardiovascular risk. We hypothesize that there is a female-specific mechanism to preserve bile acid production that operates in the absence of CYP7A1. We are currently in the process of testing candidate female-specific enzymes for bile acid production. These studies indicate new avenues by which female bile acid and cholesterol metabolism regulation differs from than that of males. The resulting differences in cardiovascular risk highlight the essential need for male- and female-specific personalized medicine approaches for cardiovascular disease prevention.

Removal of endothelial surface-associated von villebrand factor suppresses accelerate datherosclerosis after myocardial infarction

BackgroundThromboinflammation involving platelet adhesion to endothelial surface-associated von Willebrand factor (VWF) has been implicated in the accelerated progression of non-culprit plaques after MI. The aim of this study was to use arterial endothelial molecular imaging to mechanistically evaluate endothelial-associated VWF as a therapeutic target for reducing remote plaque activation after myocardial infarction (MI).MethodsHyperlipidemic mice deficient for the low-density lipoprotein receptor and Apobec-1 underwent closed-chest MI and were treated chronically with either: (i) recombinant ADAMTS13 which is responsible for proteolytic removal of VWF from the endothelial surface, (ii) N-acetylcysteine (NAC) which removes VWF by disulfide bond reduction, (iii) function-blocking anti-factor XI (FXI) antibody, or (iv) no therapy. Non-ischemic controls were also studied. At day 3 and 21, ultrasound molecular imaging was performed with probes targeted to endothelial-associated VWF A1-domain, platelet GPIbα, P-selectin and vascular cell adhesion molecule-1 (VCAM-1) at lesion-prone sites of the aorta. Histology was performed at day 21.ResultsAortic signal for P-selectin, VCAM-1, VWF, and platelet-GPIbα were all increased several-fold (p < 0.01) in post-MI mice versus sham-treated animals at day 3 and 21. Treatment with NAC and ADAMTS13 significantly attenuated the post-MI increase for all four molecular targets by > 50% (p < 0.05 vs. non-treated at day 3 and 21). On aortic root histology, mice undergoing MI versus controls had 2–4 fold greater plaque size and macrophage content (p < 0.05), approximately 20-fold greater platelet adhesion (p < 0.05), and increased staining for markers of platelet transforming growth factor-β1 signaling. Accelerated plaque growth and inflammatory activation was almost entirely prevented by ADAMTS13 and NAC. Inhibition of FXI had no significant effect on molecular imaging signal or plaque morphology.ConclusionsPlaque inflammatory activation in remote arteries after MI is strongly influenced by VWF-mediated platelet adhesion to the endothelium. These findings support investigation into new secondary preventive therapies for reducing non-culprit artery events after MI.

STATINS PREVENT THE DELETERIOUS CONSEQUENCES OF PLACENTAL CHEMERIN UPREGULATION IN PREECLAMPSIA

Objective: Chemerin, an inflammatory adipokine, is upregulated in preeclampsia, and its placental overexpression results in preeclampsia-like symptoms in mice. Statins may lower chemerin. Design and method: Chemerin levels were determined in a prospective cohort study in women suspected of preeclampsia evaluating the predictive value of the soluble Fms-like tyrosine kinase-1/placental growth factor (sFlt-1/PlGF) ratio. Chemerin release was studied in perfused placentas and placental explants with or without the statins pravastatin and fluvastatin. We also addressed statin placental passage, and the effects of chemerin in chorionic plate and coronary arteries. Results: Serum chemerin was elevated in women with preeclampsia, and its addition to a predictive model yielded significant effects on top of the sFlt-1/PlGF ratio to predict preeclampsia and its fetal complications. Perfused placentas and explants of preeclamptic women released more chemerin and sFlt-1 and less PlGF than those of healthy pregnant women. Statins reversed this. Both statins entered the fetal compartment, and pravastatin's fetal/maternal concentration ratio was twice that of fluvastatin. Chemerin constricted plate arteries, and this was blocked by a chemerin receptor antagonist and pravastatin. Chemerin did not potentiate endothelin-1 in chorionic plate or coronary arteries. In explants, statins upregulated low-density lipoprotein receptor (LDLR) expression, which relies on the same transcription factor as chemerin, and NO release. Conclusions: Chemerin is a biomarker for preeclampsia, and statins both prevent its placental upregulation and effects, in an NO- and LDLR-dependent manner. Combined with their capacity to improve the sFlt-1/PlGF ratio, this offer an attractive mechanism by which statins may prevent or treat preeclampsia.

Abstract 3038: Platelet-specific Deletion Of Transforming Growth Factor-β1 Augments Angiotensin Ii-induced Abdominal Aortic Aneurysm In Mice

Background: Transforming growth factor beta (TGFβ) blockade via systemic antibody targeting has been shown to augment murine AAA progression in multiple independent studies. Despite these findings, the primary source of protective TGFβ within AAA remains unclear. Previous research conducted in our laboratory demonstrates deletion of platelet-specific TGFβ1 exacerbates aneurysms in the elastase mouse model of AAA. Our present study extends these results by showing a comparable phenotype in the angiotensin II (Ang II) murine model of AAA. Methods and Results: Low density lipoprotein receptor deficient ( Ldlr -/- ) mice with platelet-specific deletion of Tgfβ1 were created by breeding male mice expressing Cre recombinase under the control of a Pf4 promoter to female Tgfβ1 floxed mice (Jackson Labs). Male mice were fed high fat diet 1 week prior to and throughout subcutaneous infusion of AngII (1,000 ng/kg/min). After day 28 of infusion, ex vivo aortic diameter was significantly increased in Cre+ (n = 17) vs Cre- (n = 18) mice ( Cre- : 1.75 ± 0.17 mm; Cre+ : 2.43 ± 0.21 mm; P &lt; 0.05). AAA incidence, defined as aortic diameter ≥ 1.2mm, was 56% for Cre- mice vs. 93% in the Cre+ group. Histological analysis of picrosirius red staining revealed a marked reduction in type 1 abdominal collagen deposition as a proportion of total cellular area in aortas from Cre+ mice versus Cre- controls. OCT embedded sections of aneurysms from both cohorts were also probed for macrophage marker CD68 and analyzed using immunofluorescent microscopy. We found Cre+ aortas had significantly increased macrophage infiltration compared to Cre- aortas. Conclusions: This study of platelet-specific deletion of TGFβ1 in a secondary model of murine AAA supports the conclusion that TGFβ1 sourced form platelets plays a critically important role in the protection of AAA.

Abstract 2069: Impact Of Endothelial Stiffening And CD36-mediated Lipid Uptake On Atherosclerotic Lesion Formation

Objective: To determine the impact of endothelial stiffening induced by CD36 (cluster of differentiation 36) lipid uptake in the development of atherosclerosis. Approach and Results: Endothelial stiffening of intact en face monolayers, as assessed by atomic force microscopy, reveal the hyperlipidemic mouse model of atherosclerosis, LDLR -/- (low density lipoprotein receptor knockout) is significantly stiffer, especially in the atheroprone aortic arch, as compared to wild type controls (n=4 male mice with 8-15 tissue measurement sites per condition per mouse, p&lt;0.05). Furthermore, a 4-5 month high fat diet (HFD) feeding in LDLR -/- mice resulted in significantly enhanced stiffening compared to regular chow feeding, an effect that was observed in males but not females (n=6-8 mice, p&lt;0.05). There was a significantly greater plaque development, as analyzed using Oil O Red staining for lipids, in male HFD-fed LDLR -/- mice, as compared to age-matched females (n=5-6, p&lt;0.05). No significant difference between males and females was found in lipid panel parameters (n=4-6) or weight gain (n=10-14). Specific downregulation of CD36 in male hypercholesterolemic Cdh5.CreERT2CD36fl/fl LDLR -/- mice led to both a significant decrease in endothelial stiffening (n=5 mice, p&lt;0.05) and in the areas of atherosclerotic lesion formation (n=5-6 mice, p&lt;0.05). Mechanistically, we show that the saturated fatty acid, palmitic acid, increased endothelial stiffening in the en face monolayers of wild type mice, an effect which is prevented by the pre-treatment of the CD36 irreversible inhibitor, SSO (sulfo-N-succinimidyl oleate, n=5-6 male mice, p&lt;0.05). Conclusions: These results highlight that endothelial stiffness from intact aortas and atherosclerotic plaque formation following HFD feeding is strongly dependent on endothelial CD36.

Design and pilot results from the Million Veteran Program Return Of Actionable Results (MVP-ROAR) Study

As a mega-biobank linked to a national healthcare system, the Million Veteran Program (MVP) can directly improve the health care of participants. To determine the feasibility and outcomes of returning medically actionable genetic results to MVP participants, the program launched the MVP Return of Actionable Results (MVP-ROAR) Study, with familial hypercholesterolemia (FH) as an exemplar actionable condition. The MVP-ROAR Study consists of a completed single-arm pilot phase and an ongoing randomized clinical trial (RCT), in which MVP participants are recontacted and invited to receive clinical confirmatory gene sequencing testing and a telegenetic counseling intervention. The primary outcome of the RCT is 6-month change in low-density lipoprotein cholesterol (LDL-C) between participants receiving results at baseline and those receiving results after 6 months. The pilot developed processes to identify and recontact participants nationally with probable pathogenic variants in low-density lipoprotein receptor (LDLR) on the MVP genotype array, invite them to clinical confirmatory gene sequencing, and deliver a telegenetic counseling intervention. Among participants in the pilot phase, 8 (100%) had active statin prescriptions after 6 months. Results were shared with 16 first-degree family members. Six-month ΔLDL-C (low-density lipoprotein cholesterol) after the genetic counseling intervention was -37 mg/dL (95% CI: -12 to -61; P = .03). The ongoing RCT will determine between-arm differences in this primary outcome. While underscoring the importance of clinical confirmation of research results, the pilot phase of the MVP-ROAR Study marks a turning point in MVP and demonstrates the feasibility of returning genetic results to participants and their providers. The ongoing RCT will contribute to understanding how such a program might improve patient health care and outcomes. ClinicalTrials.gov ID NCT04178122.

A Sexually Dimorphic Hepatic Cycle of Very Low Density Lipoprotein Uptake and Assembly

The liver shows striking sexual dimorphism, which is reflected in differences between women and men in pathologies inherently associated with hepatic function. Recent single-cell transcriptomes revealed spatiotemporal programmes of liver function on the sublobular scale. However, how sexual dimorphism affects this space-time logic remains poorly understood. To address this, we performed single-cell RNA-seq in the mouse liver, which allowed us to model how lobular position, circadian time and sex shape the transcriptome. We found that sex, space and time markedly influence xenobiotic detoxification and lipid processing, including lipoprotein metabolism. Crucially, the very low density lipoprotein receptor (VLDLR) is restricted to the pericentral zone, with significantly higher mRNA and protein levels in female mice. Using human samples, we were able to recapitulate both VLDLR's pericentral expression as well as higher mRNA and protein levels in premenopausal women compared to similarly aged men. In humans, we found that VLDLR expression depends not just on age but also body mass index. Conversely, several genes involved in VLDL assembly are periportally biased, and we corroborated periportal VLDL generation with electron microscopy, optimized for detecting low density lipid particles. Together, this suggests a hepatic cycle of periportal formation and pericentral uptake of VLDL. Crucially, these processes provide a new window into the sexual dimorphism characterizing differences in VLDL kinetics and atherosclerotic cardiovascular disease. The work was supported by a Swiss National Science Foundation grant. The authors declare no conflicting interests. This is the full abstract presented at the American Physiology Summit 2024 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.

POLYMORPHISM OF LIPID METABOLISM GENES IN TATARSTAN-TYPE COWS

POLYMORPHISM OF LIPID METABOLISM GENES IN TATARSTAN-TYPE COWS

Abstract 1024: A Genome-wide CRISPR Screen Identifies The LDL Receptor As The Primary Mediator Of Lp(a) Uptake By HuH7 Cells

Elevated Lp(a) blood levels have been causally linked to the development of atherosclerotic and valvular heart disease, yet the identity of the specific receptor(s) that mediate hepatic clearance of Lp(a) remains controversial. Here, we report the development, execution, and validation of a genome-wide CRISPR screen for functional modifiers of Lp(a) uptake by HuH7 cells. We first optimized a method for fluorescently labeling purified human Lp(a). Sensitive and specific detection of fluorescent Lp(a) uptake by HuH7 cells was performed by flow cytometry. We then used this approach to isolate subpopulations of individual cells with aberrant Lp(a) uptake from a pool edited with a library of 123,411 gRNAs targeting 19,050 genes across the human genome. Strikingly, the top positive regulator of Lp(a) uptake in our screen was LDLR , which encodes the low-density lipoprotein receptor. MYLIP , which encodes a ubiquitin ligase that degrades LDLR, was the top negative regulator. We also observed a significant influence for several other established LDLR regulators, including those with a canonical role in SREBP signaling and those discovered in our analogous prior screen of LDL uptake. No other previously proposed Lp(a) receptor had a significant contribution to Lp(a) uptake in our screen. We subsequently validated our screen results by engineering HuH7 cell lines with targeted deletion or overexpression of LDLR and confirmed their corresponding impact on Lp(a) uptake. Lastly, we used biolayer interferometry to detect in vitro binding between purified Lp(a) and recombinant LDLR ectodomain. Together, our findings are consistent with a model in which hepatic clearance of Lp(a) is primarily mediated by the LDL receptor.

Myo-inositol supplementation in obese patients with non-alcoholic fatty liver disease: Assessment of sirtuin-1 pathway, atherogenic and hematological parameters

BackgroundNon-alcoholic fatty liver disease (NAFLD), the most common chronic liver disease, is closely associated with higher risks of atherosclerosis through a number of molecular cascades such as the sirtuin-1 (SIRT-1) pathway. Myo-inositol as a precursor isomer of cyclohexanehexols exerts anti-hyperlipidemic and anti-obesity features in metabolic disorders. The current study aimed to assess the effects of myo-inositol supplementation on genes expression of the SIRT-1 pathway, atherogenic indices, and hematological parameters in obese patients with NAFLD. MethodsThis double-blinded randomized clinical trial was conducted on 44 obese patients with ultrasound proven- NAFLD who received dietary recommendations and were randomly allocated into either myo-inositol (4 g/day) or placebo (maltodextrin 4 g/day) receiving groups for 8 weeks. At baseline and the end of the study mRNA expression levels of SIRT-1 pathway genes in peripheral blood mononuclear cells (PBMCs), complete blood cell count, lipid profile, and atherogenic indices were assessed. Inter-group differences were determined using analysis of covariance (by adjusting for the potential confounders) at the end of the trial. ResultsAt the end of the trial mRNA expression levels of low-density- lipoprotein receptor (LDLR) (only in myo-inositol group) and endothelial nitric oxide synthase (eNOS) (in both groups) significantly increased, compared with steady-state levels (p = 0.004 and p = 0.001, respectively). Moreover, myo-inositol supplementation significantly inhibited lectin-like oxidized- low-density lipoprotein receptor-1 (LOX-1) expression compared with the placebo group (p = 0.045). Slight improvements in Cholesterol-related atherogenic indices and hematological parameters were also observed in myo-inositol group, compared with baseline values (p < 0.05). ConclusionMyo-inositol supplementation could slightly activate SIRT-1 pathway and thereby exert marginal anti-atherosclerotic properties in obese patients with NAFLD.

Procyanidin B2 alleviates oxidized low-density lipoprotein-induced cell injury, inflammation, monocyte chemotaxis, and oxidative stress by inhibiting the nuclear factor kappa-B pathway in human umbilical vein endothelial cells

BackgroundOxidized low-density lipoprotein (ox-LDL) can initiate and affect almost all atherosclerotic events including endothelial dysfunction. In this text, the role and underlying molecular basis of procyanidin B2 (PCB2) with potential anti-oxidant and anti-inflammatory activities in ox-LDL-induced HUVEC injury were examined.MethodsHUVECs were treated with ox-LDL in the presence or absence of PCB2. Cell viability and apoptotic rate were examined by CCK-8 assay and flow cytometry, respectively. The mRNA and protein levels of genes were tested by RT-qPCR and western blot assays, respectively. Potential downstream targets and pathways of apple procyanidin oligomers were examined by bioinformatics analysis for the GSE9647 dataset. The effect of PCB2 on THP-1 cell migration was examined by recruitment assay. The effect of PCB2 on oxidative stress was assessed by reactive oxygen species (ROS) level, malondialdehyde (MDA) content, and mitochondrial membrane potential (MMP).Resultsox-LDL reduced cell viability, induced cell apoptosis, and facilitated the expression of oxidized low-density lipoprotein receptor 1 (LOX-1), C-C motif chemokine ligand 2 (MCP-1), vascular cell adhesion protein 1 (VCAM-1) in HUVECs. PCB2 alleviated ox-LDL-induced cell injury in HUVECs. Apple procyanidin oligomers triggered the differential expression of 592 genes in HUVECs (|log2fold-change| > 0.58 and adjusted p-value < 0.05). These dysregulated genes might be implicated in apoptosis, endothelial cell proliferation, inflammation, and monocyte chemotaxis. PCB2 inhibited C-X-C motif chemokine ligand 1/8 (CXCL1/8) expression and THP-1 cell recruitment in ox-LDL-stimulated HUVECs. PCB2 inhibited ox-LDL-induced oxidative stress and nuclear factor kappa-B (NF-κB) activation in HUVECs.ConclusionPCB2 weakened ox-LDL-induced cell injury, inflammation, monocyte recruitment, and oxidative stress by inhibiting the NF-κB pathway in HUVECs.

Gallic Acid Can Promote Low-Density Lipoprotein Uptake in HepG2 Cells via Increasing Low-Density Lipoprotein Receptor Accumulation.

Gallic acid (GA) is a type of polyphenolic compound that can be found in a range of fruits, vegetables, and tea. Although it has been confirmed it improves non-alcoholic fatty liver disease (NAFLD), it is still unknown whether GA can improve the occurrence of NAFLD by increasing the low-density lipoprotein receptor (LDLR) accumulation and alleviating cholesterol metabolism disorders. Therefore, the present study explored the effect of GA on LDLR and its mechanism of action. The findings indicated that the increase in LDLR accumulation in HepG2 cells induced by GA was associated with the stimulation of the epidermal growth factor receptor-extracellular regulated protein kinase (EGFR-ERK1/2) signaling pathway. When the pathway was inhibited by EGFR mab cetuximab, it was observed that the activation of the EGFR-ERK1/2 signaling pathway induced by GA was also blocked. At the same time, the accumulation of LDLR protein and the uptake of LDL were also suppressed. Additionally, GA can also promote the accumulation of forkhead box O3 (FOXO3) and suppress the accumulation of hepatocyte nuclear factor-1α (HNF1α), leading to the inhibition of proprotein convertase subtilisin/kexin 9 (PCSK9) mRNA expression and protein accumulation. This ultimately results in increased LDLR protein accumulation and enhanced uptake of LDL in cells. In summary, the present study revealed the potential mechanism of GA's role in ameliorating NAFLD, with a view of providing a theoretical basis for the dietary supplementation of GA.