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The dysfunction of the endothelial lining in lesion-prone areas of the arterial vasculature significantly contributes to the pathobiology of atherosclerotic cardiovascular disease. Recent studies suggested that UDP-glucose pyrophosphorylase 2 (UGP2) plays a role in cell proliferation and survival. This study investigates the anti-apoptotic and anti-atherogenic effects of UGP2 both in vitro and in vivo. We explored the effects and mechanisms of UGP2 on apoptosis in endothelial cells using flow cytometry and Western blot analysis. Additionally, we evaluate apoptosis levels in atherosclerotic lesions with ldlr -/- ugp2 +/- mice. Microarray analysis revealed reduced UGP2 expression in human atherosclerotic plaques. In vitro experiments demonstrated that TP53 interacts with the promoter region of the UGP2 gene, upregulating UGP2 expression. Enhanced UGP2 expression led to decreased reactive oxygen species (ROS) levels, reduced Cleaved caspase-3 expression, and lower apoptosis levels in endothelial cells. The anti-apoptotic effects of UGP2 were significantly diminished by H 2 O 2 . In vivo, UGP2 deficiency in ldlr -/- mice fed a Western high-fat diet promoted atherosclerosis, increased ROS levels, and elevated Cleaved caspase-3 expression and apoptosis in atherosclerotic lesions. Our findings identify UGP2 as a novel TP53 target gene that contributes to anti-apoptotic effects by regulating ROS homeostasis via a non-canonical pathway. UGP2 represents a potential therapeutic target for ameliorating atherosclerosis-related diseases., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2025 Elsevier Inc. All rights reserved.)

Background: Observational studies have identified a possible connection between lipid-lowering medications and respiratory illnesses. However, it remains unclear whether lipid-lowering drugs is causative for respiratory diseases, and we aimed to answer this question., Methods: We performed Mendelian randomization (MR) analyses by integrating data from genome-wide association studies (GWAS). Three statistical approaches were employed for MR analysis: inverse variance weighting (IVW), MR-Egger, and weighted median. The purpose was to evaluate the causal relationships between 10 drug targets that lower lipid levels and the likelihood of developing 7 respiratory diseases. Additional sensitivity analyses were conducted to ensure the robustness and validity of the results., Results: After adjusting for multiple testing, our MR analysis identified APOB (odd ratios [OR]: 0.86; 95% confidence interval [CI]: 0.77 to 0.97; P IVW  = 0.01) and PCSK9 (OR: 0.84; 95% CI: 0.72 to 0.97; P IVW  = 0.02) as significant risk targets for asthma. Additionally, LDLR was found to be a significant risk target for chronic obstructive pulmonary disease (OR: 0.81; 95% CI: 0.67 to 0.98; P IVW  = 0.03). The sensitivity analysis validated no proof of heterogeneity or pleiotropy amongst the mentioned results., Conclusions: Our findings suggest a likely causal relationship between respiratory diseases and lipid-lowering drug targets. Further mechanistic and clinical research is needed to confirm and validate these findings., Competing Interests: Declarations. Ethics approval and consent to participate: The ethical review and approval were not required for this study because all data used in this study are publicly available. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Proprotein convertase subtilisin/kexin type 9 (PCSK9) plays a crucial role in the degradation of the low-density lipoprotein receptor (LDLR), and PCSK9 inhibition emerges as an attractive strategy for atherosclerosis management. In this study, extracellular vesicles (EVs) were engineered to nanosponges, which could efficiently adsorb and deliver PCSK9 into lysosomes for degradation. Briefly, nanosponges were engineered by modifying EVs with EGF-A/PTGFRN fusion protein (PCSK9 binding domain EGF-A from the mutant LDLR with higher affinity was fused to the C terminus of prostaglandin F2 receptor negative regulator). The modification endowed the EVs with hundreds of EGF-As displayed on the surface, and thus the capacity to adsorb PCSK9 efficiently. The adsorbed PCSK9 would thus be delivered into lysosomes for degradation when the nanosponges were endocytosed by liver cells, thus releasing endogenous LDLR from degradation. In the ApoE -/- mouse model, tail vein-injected nanosponges were able to degrade PCSK9, increase LDLR expression, lower the LDL-C level, and thus alleviate atherosclerosis. In summary, here we not only develop a novel strategy for PCSK9 inhibition but we also propose a universal method for adsorption and degradation of circulating proteins for disease management., Competing Interests: Declaration of interests L.Y., G.Y., C.W., and M.W. are inventors on patents applied for by Fourth Military Medical University related to PCSK9 inhibition., (Copyright © 2024 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.)

Recent studies have demonstrated that chronic stress can enhance the development of multiple human diseases, including cancer. However, the role of chronic stress in esophageal carcinogenesis and its underlying molecular mechanisms remain unclear. This study uncovered that dysregulated cholesterol metabolism significantly promotes esophageal carcinogenesis under chronic stress conditions. Our findings indicate that the persistent elevation of glucocorticoids induced by chronic stress stimulates cholesterol uptake, contributing to esophageal carcinogenesis. The activated glucocorticoid receptor (GCR) enrichment at the promoter region of High Mobility Group Box 2 (HMGB2) facilitates its transcription. As a transcription coactivator, HMGB2 enhances Sterol Regulatory Element Binding Transcription Factor 1 (SREBF1) transcription and regulates cholesterol metabolism through LDL particle uptake into cells via Low Density Lipoprotein Receptor (LDLR). These results emphasize the significant impact of chronic stress on esophageal carcinogenesis and establish cholesterol metabolism disorder as a crucial link between chronic stress and the development of ESCC. The implications suggest that effectively managing chronic stress may serve as a viable strategy for preventing and treating ESCC., Competing Interests: Competing interests statement:The authors declare no competing interest.

Cholesterol is a vital component of cellular membranes and is an essential molecule in mammalian physiology. Yet dysregulation of hepatic cholesterol metabolism and an increase in plasma cholesterol is linked to development of atherosclerotic cardiovascular disease. Maintaining tight regulation of cholesterol homeostasis is therefore essential, elegantly highlighted by the control of hepatic low-density lipoprotein receptor (LDLR) abundance and associated lipoprotein clearance. The LDLR was discovered in the 1970's in the seminal work of Brown and Goldstein. This was followed by the development of statins, which promote hepatic clearance of LDL via the LDLR pathway. The discovery two decades ago of Proprotein Convertase Subtilisin-Kexin Type 9 (PCSK9), a secreted protein that binds to the LDLR ectodomain and promotes its degradation, and the clinical development of PCSK9 inhibitors has ushered an effort to uncover additional mechanisms that govern the function and abundance of the LDLR. In recent years this has led to the identification of novel post-transcriptional and post-translational mechanisms that govern the LDLR. This review focuses on these emerging regulatory mechanisms and specifically discusses: (1) Regulation of the LDLR mRNA by RNA-binding proteins and microRNAs, (2) Ubiquitin-dependent degradation of the LDLR protein by the E3 ubiquitin ligases inducible degrader of the LDLR (IDOL) and GOLIATH (RNF130), (3) Control of the LDLR pathway by the asialoglycoprotein receptor 1 (ASGR1), and (4) The role of LDLR ectodomain shedding mediated by membrane-type 1 matrix metalloprotease (MT1-MMP), Bone morphogenetic protein 1 (BMP1), and γ-secretase. Understanding the contribution of these emerging mechanisms to regulation of the LDLR is important for the development of novel LDLR-focused lipid-lowering strategies., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

We explore the interaction between estrogen and PCSK9 and their collective impact on lipid metabolism, especially concerning the regulation of low-density lipoprotein receptor levels. Utilizing both animal and cellular models, including ovariectomized mice and HepG2 cell lines, we demonstrate that estrogen deficiency leads to a disruption in lipid metabolism, characterized by elevated levels of total cholesterol and LDL-C. The study commences with mice undergoing ovariectomy, followed by a diet regimen comprising either high-fat diet or normal feed for a four-week duration. Key assessments include analyzing lipid metabolism, measuring PCSK9 levels in the bloodstream, and evaluating hepatic low-density lipoprotein receptor expression. We will also conduct correlation analyses to understand the relationship between PCSK9 and various lipid profiles. Further, a subset of ovariectomized mice on high-fat diet will undergo treatment with either estrogen or PCSK9 inhibitor for two weeks, with a subsequent re-evaluation of the earlier mentioned parameters. Our findings reveal that estrogen inhibits PCSK9-mediated degradation of low-density lipoprotein receptor, a process crucial for maintaining lipid homeostasis. Through a series of experiments, including immunohistochemistry and western blot analysis, we establish that PCSK9 is involved in lipid metabolism disorders caused by estrogen deficiency and that estrogen regulates PCSK9 and low-density lipoprotein receptor at post-transcriptional level. The study provides a mechanism for the involvement of PCSK9 in elucidating the disorders of lipid metabolism caused by estrogen deficiency due to perimenopause and ovarian decline., Competing Interests: Declarations. Animal Ethics: All animal experiments were approved by the Sichuan University academic ethics and ethics committee and were conducted in strict accordance with laboratory animal care and use program. Consent to Participate: Not applicable. Consent for Publication: All authors have read and approved to submit this article. Conflict of interest: The authors have no conflict of interests., (© 2024. The Author(s), under exclusive licence to Society for Reproductive Investigation.)

Background: Exome sequencing has been established as a fundamental tool in genetic diagnostics. It may also provide information about variants in genes unrelated to the primary purpose, so-called secondary findings. Especially, diagnoses of unnoticed inborn cardiac diseases are of high clinical relevance due to therapeutic options in context of prevention of sudden cardiac death., Methods: Exome data of 9962 individuals was analysed for relevant cardiogenetic findings. Genes were selected according to ACMG recommendations for secondary findings (v.3.1). First, a filter for (likely) pathogenic variants, published in the ClinVar database, was used. Second, exome data was screened for loss of function (LoF) variants in genes in which LoF is a known disease pathomechanism. All variants were evaluated by geneticists regarding their pathogenicity., Results: Pathogenic or likely pathogenic variants were identified in 136 different individuals (136/9962, 1.4%), with the Low-Density Lipoprotein Receptor gene (LDLR, 24/136, 17.6%) and the Titin gene (TTN, 24/136, 17.6%), being the most frequently affected ones. 31.6% (43/136) of the identified variants had been reported beforehand, while 47.1% (64/136) had not been reported. The remaining cases (29/136, 21.3%) were part of research projects with no written reports. In 26.5% (36/136), the finding would have been missed, if only index patients and not their parents had been screened for secondary findings in case of trio ES., Conclusion: As demonstrated in our study, at least one or two out of one hundred people are likely to carry a pathogenic cardiogenetic variant. Counselling geneticist and clinicians need to be aware of these findings in exome and genome sequencing. Informed consent of the patient regarding the report of secondary findings should absolutely be obtained beforehand., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Nanobodies (Nbs) hold great potential to replace conventional antibodies in various biomedical applications. However, conventional methods for their discovery can be time-consuming and expensive. We have developed a reliable protein selection strategy that combines magnetic activated cell sorting (MACS)-based screening of yeast surface display (YSD) libraries and functional ligand-binding identification by Tat-based recognition of associating proteins (FLI-TRAP) to isolate antigen-specific Nbs from synthetic libraries. This combined process enabled isolation of three unique Nb clones (NbT15, NbT21, and NbT22) that all bound specifically to a target antigen, namely proprotein convertase subtilisin/kexin type 9 (PCSK9) as well as a gain-of-function PCSK9 mutant (D374Y). All three clones bound to PCSK9 and blocked the interaction between the low-density lipoprotein receptor (LDLR) and either wild-type PCSK9 or the D374Y mutant. Overall, our combined protein selection method enables rapid and straightforward identification of potent antigen-specific Nbs in a manner that can be executed in a basic laboratory setting without the need for specialized equipment. We anticipate that our strategy will be a valuable addition to the protein engineering toolkit, allowing development of Nbs or virtually any other synthetic binding protein for a wide range of applications., Competing Interests: Declarations. Competing interests: M.P.D. has financial interests in Gauntlet, Inc., Glycobia, Inc., Resilience, Inc., UbiquiTX, Inc., and Versatope Therapeutics, Inc. M.P.D.‘s interests are reviewed and managed by Cornell University in accordance with their conflict-of-interest policies. All other authors declare no competing interests. A.C.K. is a cofounder and consultant for biotechnology companies Tectonic Therapeutic and Seismic Therapeutic, and also for the Institute for Protein Innovation, a nonprofit research institute. The other authors do not have any Competing-Interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results. Ethics statement: The study was approved by the Institutional Biosafety Committee of King Mongkut’s University of Technology (IBC-2020-031 and IBC-2023-004)., (© 2025. The Author(s).)

The Low Density Lipoprotein receptors (LDLRs) gene family includes 15 receptors: very low-density lipoprotein receptor (VLDLR), LDLR, Sorting-related receptor with A-type repeats (SORLA), and 12 LDL receptor-related proteins (LRPs): LRP1, LRP1B, LRP2, LRP3, LRP4, LRP5, LRP6, LRP8, LRP10, LRP11, LRP12, LRP13. Most of these are involved in the transduction of key signals during embryonic development and in the regulation of cholesterol homeostasis. In oviparous animals, the VLDL receptor is also known as VTGR since it facilitates the uptake of vitellogenin in ovary. In tetrapods, information concerning genes encoding these proteins is limited to a few taxa. Here, we report the characterization of VTGR in the amphibian Cynops orientalis. The secondary structure analyses and the expression profiles obtained from hepatic and gonadal tissues of C. orientalis supported the role of VTGR as vitellogenin oocyte membrane receptor in this species. Moreover, to get a holistic view of the evolutionary history of this gene superfamily, we extended our investigation to all 15 genes belonging to the LDLR superfamily analyzing through a phylogenetic analysis a total of 161 sequences belonging to 11 genera of vertebrates. The position of LRP8 in the tree and its expression findings in C. orientalis ovary allowed us to suggest that other proteins of the LDLR superfamily could act as receptors during vitellogenesis., Competing Interests: Declarations. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Positively charged residues are commonly located near the cytoplasm-membrane interface, which is known as the positive-inside rule in membrane topology. The mechanism underlying the function of these charged residues remains poorly understood. Herein, we studied the function of cytoplasmic residues in corin, a type II transmembrane serine protease in cardiovascular biology. We found that the positively charged residue at the cytoplasm-membrane interface of corin was not a primary determinant in membrane topology but probably served as a charge-repulsion mechanism in the endoplasmic reticulum (ER) to prevent interactions with proteins in the ER, including the signal peptidase. Substitution of the positively charged residue with a neutral or acidic residue resulted in corin secretion likely due to signal peptidase cleavage. In signal peptidase-deficient cells, the mutant corin proteins were not secreted but retained in the ER. Similar results were found in the low-density lipoprotein receptor and matriptase-2 that have positively charged residues at and near the cytoplasm-membrane interface. These results provide important insights into the role of the positively charged cytoplasmic residues in mammalian single-pass transmembrane proteins., Competing Interests: Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Objective: The impact of lipid-lowering medications on chronic kidney disease (CKD) remains a subject of debate. This Mendelian randomization (MR) study aims to elucidate the potential effects of lipid-lowering drug targets on CKD development., Methods: We extracted 11 genetic variants encoding targets of lipid-lowering drugs from published genome-wide association study (GWAS) summary statistics, encompassing LDLR, HMGCR, PCSK9, NPC1L1, APOB, ABCG5/ABCG8, LPL, APOC3, ANGPTL3, and PPARA. A Mendelian randomization analysis was conducted targeting these drug-related genes. CKD risk was designated as the primary outcome, while estimated glomerular filtration rate (eGFR) and blood urea nitrogen (BUN) were assessed as secondary outcomes. Additionally, mediation analysis was performed utilizing 731 immune cell phenotypes to identify potential mediators., Results: The meta-analysis revealed a significant association between ANGPTL3 inhibitors and a reduced risk of CKD (OR [95% CI] = 0.85 [0.75-0.96]). Conversely, LDLR agonists were significantly linked to an increased risk of CKD (OR [95% CI] = 1.11 [1.02-1.22]). Regarding secondary outcomes, lipid-lowering drugs did not significantly affect eGFR and BUN levels. Mediation analysis indicated that the reduction in CKD risk by ANGPTL3 inhibitors was mediated through modulation of the immune cell phenotype, specifically HLA-DR on CD14+ CD16+ monocytes (Mediated proportion: 4.69%; Mediated effect: -0.00899)., Conclusion: Through drug-targeted MR analysis, we identified a causal relationship between lipid-lowering drug targets and CKD. ANGPTL3 and LDLR may represent promising candidate drug targets for CKD treatment., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2025 Zhang, Ou, Peng, Pan, Zhang and Shi.)

Coronary artery disease (CAD) is a multigenic condition influenced by both nature and nurture (60% to 40%). Prognosis of CAD is based on familial patterns. This study examined and analyzed the susceptibility of CAD to genetic variants in various Pakistani families. A total of 50 families, 308 participants (79 affected and 229 unaffected were genotyped for NOS3 (rs1799983, rs2070744), PON1 (rs662), LPA-PLA2 (rs105193, rs1805017), APOE (rs429358, rs7412), PCSK9 (rs505151), MEF2A (rs325400), TNF (rs1800629) and LDLR (rs1122608, rs2228671) genes. The family-based association in CAD associated genes SNPs were NOS3 (rs1799983), PON1 (rs662), LPA-PLA2 (rs1805017), MEF2A (rs325400), and LDLR (rs1122608, rs222867) showed transmission within families p≤ 0.05 whereas NOS3 (rs2070744), APOE (rs429358, rs7412) and TNF (rs1800629) showed no association TDT asymptotic p-value >0.05. In DFAM and QFAM test NOS3 (rs1799983), PON1 (rs662), MEF2A (rs325400), and LDLR (rs1122608, rs222867) showed positive association p≤ 0.05 in both whereas NOS3 (rs2070744), APOE (rs429358, rs7412), LPA-PLA2 (rs1805017) and TNF (rs1800629) showed low risk of transmission asymptotic p-value >0.05 in DFAM but NOS3(rs2070744), APOE(rs7412), LPA-PLAG2(rs1805017) also showed association p≤ 0.05 whereas APOE (rs429358) and TNF (rs1800629) showed no association EMP1 p-value >0.05 in QFAM. In linkage analysis Chromosome 6 (Position 70.810): LOD = 3.16, Chromosome 7 (Position 107.190): LOD = 3.16, and chromosome 19 (Position 31.470): LOD = 3.90 also showed significant association with disease as p < 0.05. This discovery enhances the understanding about genetic variants of CAD and also facilitates early detection, targeted interventions, pattern of inheritance in population. This ultimately improving patient outcomes and guiding future research to highlight its significance as a potential diagnostic marker., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2025 Zahid et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Dietary fat drives the pathogenesis of atherosclerotic cardiovascular disease (ASCVD), particularly through circulating cholesterol and triglyceride-rich lipoprotein remnants. Industrially produced trans-unsaturated fatty acids (TFAs) incorporated into food supplies significantly promote ASCVD. However, the molecular trafficking of TFAs responsible for this association is not well understood. Here, we demonstrate that TFAs are preferentially incorporated into sphingolipids by serine palmitoyltransferase (SPT) and secreted from cells in vitro. Administering high-fat diets (HFDs) enriched in TFAs to Ldlr -/- mice accelerated hepatic very-low-density lipoprotein (VLDL) and sphingolipid secretion into circulation to promote atherogenesis compared with a cis-unsaturated fatty acid (CFA)-enriched HFD. SPT inhibition mitigated these phenotypes and reduced circulating atherogenic VLDL enriched in TFA-derived polyunsaturated sphingomyelin. Transcriptional analysis of human liver revealed distinct regulation of SPTLC2 versus SPTLC3 subunit expression, consistent with human genetic correlations in ASCVD, further establishing sphingolipid metabolism as a critical node mediating the progression of ASCVD in response to specific dietary fats., Competing Interests: Declaration of interests C.M.M. is a scientific founder and shareholder of Amprenta Therapeutics and a scientific advisor for Faeth Therapeutics., (Published by Elsevier Inc.)

Purpose of Review: Homozygous familial hypercholesterolaemia (HoFH) is characterized by marked elevation of low-density lipoprotein cholesterol (LDLC) and premature atherosclerotic cardiovascular disease. This is a review of novel pharmacological therapies to lower LDLC in patients with HoFH., Recent Findings: Novel therapies can be broadly divided by whether their efficacy is dependent or independent of residual low-density lipoprotein receptor (LDLR) function. Novel LDLR dependent therapies that reduce proprotein subtilisin kexin type 9 levels include monoclonal antibodies (alirocumab and evolocumab) and a small inhibitory RNA (inclisiran). LDLC reductions are highly variable and depend on residual LDLR function. Microsomal triglyceride inhibitors (lomitapide) and therapies that reduce angiopoietin like factor 3 (evinacumab and zodasiran) both reduce LDLC by approximately 50%, irrespective of residual LDLR function. Most patients with HoFH require multiple therapies to achieve LDLC targets. Better LDLC control with LDLR independent therapies is likely to improve the outlook for patients with HoFH while at the same time reducing the need for other therapies such as apheresis or hepatic transplantation., Competing Interests: Declarations. Competing Interests: DJ Blom DB reports personal fees from Amgen, Amryt, MSD, Novartis, Sanofi and Servier and outside the submitted work. A David Marais ADM reports no personal fees from the companies producing the agents discussed but had support for the performance of the study of atorvastatin and lomitapide by Warner Lambert Parke Davis and Aegerion, respectively. FJ Raal FJR reports personal fees from LIB Therapeutics, Amgen, Sanofi, Regeneron Pharmaceuticals, Inc., and Novartis outside the submitted work. Human and Animal Rights and Informed Consent: All reported studies/experiments with human or animal subjects performed by the authors have been previously published and complied with all applicable ethical standards (including the Helsinki declaration and its amendments, institutional/national research committee standards, and international/national/institutional guidelines)., (© 2024. The Author(s).)

Flaviviruses, which are transmitted by mosquitoes, are arthropod-borne infections that are pathogenic to both humans and animals, posing a significant global threat to public health. So far, various endocytic pathways have been reported for flaviviral entry; however, the role of cellular factors in viral replication and entry remains uncertain. Here in this study, we identified the role of Low-density lipoprotein receptor, which has long been established as a cholesterol carrier for neurons but remained unexplored as an essential host factor for JEV/WNV replication. To explore this, we utilized 10-day old BALB/c pups and two neuronal cell lines, NSC34 and HT22, both of different origin, as experimental models. Transient knockdown of LDLR gene in vitro using siRNA-mediated gene silencing drastically reduced viral specific transcripts and proteins upon viral incubation. Moreover, flaviviral binding and internalization were significantly compromised upon infection in LDLR-transfected cells when compared with non-specific eGFP-transfected cells. Antibody neutralization experiments using LDLR-specific polyclonal antibody significantly reduced viral entry in vitro, suggesting the role of LDLR as an important cell attachment factor for JEV and WNV uptake. Furthermore, ectopic expression of LDLR via plasmid transfection led to significant increase in virus replication in cells, indicating significant role of LDLR in flavivirus replication beside acting as an active attachment factor for JEV and WNV. Overall, our results indicate that LDLR act as novel host factor involved in both flaviviral entry and replication, thus serving as a suitable candidate for antiviral research., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Inc.)

Background: Pharmacological inhibition of megalin (also known as LRP2 [low-density lipoprotein receptor-related protein-2]) 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., Methods: Female Lrp2 f/f mice were bred with male Ndrg1 - Cre ERT2 +/0 mice to develop PTC-LRP2 +/+ and PTC-LRP2 -/- littermates. To study atherosclerosis, all mice were bred to an LDL (low-density lipoprotein) receptor -/- background and fed a Western diet to induce atherosclerosis., Results: PTC-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 a 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 on hypercholesterolemia. In contrast, female PTC-LRP2 -/- mice had no apparent renal pathologies. In vivo multiphoton microscopy demonstrated that PTC-specific megalin deletion dramatically diminished ALB (albumin) accumulation in PTCs within 10 days of Western diet feeding. RNA-sequencing analyses demonstrated the upregulation of inflammation-related pathways in the kidney., Conclusions: PTC-specific megalin deletion does not affect atherosclerosis but leads to tubulointerstitial nephritis in mice fed a Western diet, with severe pathologies in male mice., Competing Interests: A.E. Mullick is an employee of Ionis Pharmaceuticals, Inc. M. Yanagita has received research grants from Mitsubishi Tanabe Pharma and Boehringer Ingelheim. The other authors report no conflicts.

In non-small cell lung cancers, the non-squamous and squamous subtypes (nsqNSCLC and sqNSCLC) exhibit disparities in pathophysiology, tumor immunology, and potential genomic correlates affecting responses to immune checkpoint inhibitor (ICI)-based treatments. In our in-house training cohort (n=85), the presence of the LRP1B deleterious mutation (LRP1B-del) was associated with longer and shorter progression-free survival (PFS) on ICIs alone in nsqNSCLCs and sqNSCLCs, respectively (P interaction =0.008). These results were validated using a larger public ICI cohort (n=208, P interaction <0.001). Multiplex immunofluorescence staining revealed an association between LRP1B-del and increased and decreased numbers of tumor-infiltrating CD8 + T cells in nsqNSCLCs (P=0.040) and sqNSCLCs (P=0.014), respectively. In the POPLAR/OAK cohort, nsqNSCLCs with LRP1B-del demonstrated improved PFS benefits from atezolizumab over docetaxel (hazard ratio (HR) =0.70, P=0.046), whereas this benefit was negligible in those without LRP1B-del (HR=1.05, P=0.64). Conversely, sqNSCLCs without LRP1B-del benefited more from atezolizumab (HR=0.60, P=0.002) than those with LRP1B-del (HR=1.30, P=0.31). Consistent results were observed in the in-house CHOICE-01 cohort, in which nsqNSCLCs with LRP1B-del and sqNSCLCs without LRP1B-del benefited more from toripalimab plus chemotherapy than from chemotherapy alone (P interaction =0.008). This multi-cohort study delineates the antithetical impacts of LRP1B-del in nsqNSCLCs and sqNSCLCs on predicting the benefits from ICI alone or with chemotherapy over chemotherapy alone. Our findings highlight the distinct clinical utility of LRP1B-del in guiding treatment choices for nsqNSCLCs and sqNSCLCs, emphasizing the necessity for a detailed analysis based on pathological subtypes when investigating biomarkers for cancer therapeutics., Competing Interests: Compliance and ethics. The authors declare no potential conflicts of interest, except for the employment of Yu Xu, Qiaoxia Zhou, Guoqiang Wang, and Shangli Cai at Burning Rock Biotech. All human sample collection and usage procedures followed the principles of the Declaration of Helsinki and were approved by the Institutional Review Board of the Cancer Hospital of Chinese Academy of Medical Sciences and Peking Union Medical College. Written consent was obtained from all included patients., (© 2024. Science China Press.)

Background: Palmoplantar keratoderma (PPK) is a heterogeneous group of disorders characterized by abnormal thickening of the skin on the palms and soles. Striate palmoplantar keratoderma (SPPK) is commonly caused by heterozygous mutations in the desmoglein-1 (DSG1) gene., Objective: This study aimed to report a case of a 36-year-old Chinese female patient with SPPK caused by a novel DSG1 gene mutation, along with her family history, and explore its potential relationship with other genetic variants., Methods: Whole-exome sequencing was performed on the patient and their family members to identify the pathogenic mutation, which was validated by Sanger sequencing. Histological and electron microscopy analyses were conducted to examine the pathological characteristics of skin tissue.of skin tissue., Results: A frameshift mutation, c.1285del, in exon 10 of the DSG1 gene was identified, leading to a loss of protein function and resulting in SPPK. This mutation was also detected in two other family members with similar phenotypes. Additionally, a classical splicing variant, c.313+2dup, in the low-density lipoprotein receptor (LDLR) gene associated with hypercholesterolemia was identified in the patient; however, no direct association with SPPK was observed., Conclusion: This study was the first to report a novel mutation in the DSG1 gene associated with SPPK and suggested a potential role of the LDLR gene variant in SPPK patients, providing new insights for further research into the genetic mechanisms underlying SPPK., Competing Interests: Declarations. Conflict of interest: The authors declare no conflict of interest. Ethical approval: The study was approved by Xijing Hospital, Fourth Military Medical University Ethics Committee. Informed consent: The patient provided written informed consent for the publication of this report., (© 2024. The Author(s) under exclusive licence to The Genetics Society of Korea.)

Background: apoER2 (apolipoprotein E receptor-2) is a transmembrane receptor in the low-density lipoprotein receptor (LDLR) family with unique tissue expression. A single-nucleotide polymorphism that encodes the R952Q sequence variant has been associated with elevated plasma cholesterol levels and increased myocardial infarction risk in humans. The objective of this study was to delineate the mechanism underlying the association between the apoER2 variant with arginine-to-glutamine substitution at residue 952 (R952Q) and increased atherosclerosis risk., Methods: An apoER2 R952Q mouse model was generated and intercrossed with LDLR knockout mice, followed by feeding a Western-type high-fat high-cholesterol diet for 16 weeks. Atherosclerosis was investigated by immunohistology. Plasma lipids and lipid distributions among the various lipoprotein classes were analyzed by colorimetric assay. Tissue-specific effects of the R952Q sequence variant on atherosclerosis were analyzed by bone marrow transplant studies. sLRP1 (soluble low-density lipoprotein receptor-related protein 1) was measured in plasma and conditioned media from bone marrow-derived macrophages by ELISA and GST-RAP (glutathione S-transferase-receptor-associated protein) pull-down, respectively. P38 MAPK (mitogen-activated protein kinase) phosphorylation in VLDL (very-low-density lipoprotein)-treated macrophages was determined by Western blot analysis., Results: Consistent with observations in humans with this sequence variant, the apoER2 R952Q mutation exacerbated diet-induced hypercholesterolemia, via impediment of plasma triglyceride-rich lipoprotein clearance, to accelerate atherosclerosis in Western diet-fed LDLR knockout mice. Reciprocal bone marrow transplant experiments revealed that the apoER2 R952Q mutation in bone marrow-derived cells instead of non-bone marrow-derived cells was responsible for the increase in hypercholesterolemia and atherosclerosis. Additional data showed that the apoER2 R952Q mutation in macrophages promotes VLDL-induced LRP1 (low-density lipoprotein receptor-related protein 1) shedding in a p38 MAPK-dependent manner., Conclusions: The apoER2 R952Q mouse model recapitulates characteristics observed in human disease. The underlying mechanism is that the apoER2 R952Q mutation in macrophages exacerbates VLDL-stimulated sLRP1 production in a p38 MAPK-dependent manner, resulting in its competition with cell surface LRP1 to impede triglyceride-rich lipoprotein clearance, thereby resulting in increased hypercholesterolemia and accelerated atherosclerosis., Competing Interests: None.

The low-density lipoprotein receptor (LDLr) is the first member of a closely related transmembrane protein family. It is known for its involvement in various physiological processes, mainly in the regulation of lipid metabolism, especially in the brains of mammals and zebrafish. In zebrafish, two ldlr genes (ldlra and b) have been identified and their distribution in the brain is not well documented. Recently, the roles of ldlr and its inhibitor pcsk9 in regenerative process after telencephalic brain injury have been discussed. In this study, we explored the expression patterns of these genes during zebrafish development. We found that ldlra expression was detected at the end of the pharyngula period (48 hpf) and increased during the larval stage. Conversely, ldlrb expression was observed from zygotic to larval stages. Using techniques like in situ hybridization and taking advantage of transgenic fish, we demonstrated the widespread distribution of ldlra, ldlrb and pcsk9 in the brain of adult zebrafish. Specifically, these genes were expressed in neurons and neural stem cells and also at lower levels in endothelial cells. As expected, intraperitoneal injection of fluorescent-labelled LDLs resulted in their uptake by cerebral endothelial cells in a homeostatic context, whereas they diffused within the brain parenchyma after telencephalic injury. However, after intracerebroventricular injections into animals, LDL particles were not taken up by neural stem cells. In conclusion, our results provide additional evidence for LDLr expression in the brain of adult zebrafish. These results raise the question of the role of LDLr in the cholesterol/lipid imbalance in cerebral complications., (© 2024 The Author(s). European Journal of Neuroscience published by Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

The combination of alcohol and a low-carbohydrate, high-protein, high-fat atherogenic diet (AD) increases the risk of lethal arrhythmias in apolipoprotein E/low-density lipoprotein receptor double-knockout (AL) mice with metabolic dysfunction-associated steatotic liver disease (MASLD). This study investigates whether left ventricular (LV) myocardial interstitial fibrosis (MIF), formed during the progression of metabolic dysfunction-associated steatohepatitis (MASH), contributes to this increased risk. Male AL mice were fed an AD with or without ethanol for 16 weeks, while age-matched AL and wild-type mice served as controls. Liver and heart tissues were analyzed, and susceptibility to lethal arrhythmias was assessed through histopathology, fluorescence immunohistochemistry, RNA-Seq, RT-PCR, and lethal arrhythmia-evoked test. Ethanol combined with an AD significantly induced LV MIF in MASH-affected AL mice, as shown by increased fibrosis-related gene expression, Sirius-Red staining, and elevated collagen 1a1 and 3a1 mRNA levels, alongside a higher incidence of lethal arrhythmias. Cardiac myofibroblasts exhibited sympathetic activation and produced elevated levels of fibrosis-promoting factors. This study highlights the role of cardiac myofibroblasts in LV MIF, contributing to an increased incidence of lethal arrhythmias in MASH-affected AL mice fed ethanol and AD, even after the alcohol was fully metabolized on the day of consumption.

Background: Familial hypercholesterolemia (FH) is an inherited disorder mainly marked by increased low-density lipoprotein cholesterol (LDL-C) concentrations and a heightened risk of early-onset arteriosclerotic cardiovascular disease (ASCVD). This study seeks to characterize the genetic spectrum and genotype‒phenotype correlations of FH in Chinese pediatric individuals., Methods: Data were gathered from individuals diagnosed with FH either clinically or genetically at multiple hospitals across mainland China from January 2016 to June 2024., Results: In total, 140 children and adolescents (mean age of 6.00 years) with clinically and genetically diagnosed FH were enrolled in the study, with 87 distinct variants identified in the LDLR, APOB and PCSK9 genes. Among the variants, 11 variants were newly identified worldwide, with 9 classified as "pathogenic" or "likely pathogenic", and 2 classified as "variants of uncertain significance". Additionally, the 5 most common variants in the study were c.1448G > A (p.W483*), c.1879G > A (p.A627T), c.1216C > A (p.R406R), and c.1747C > T (p.H583Y) in the LDLR gene, as well as c.10579C > T (p.R3527W) in the APOB gene, accounting for 49.29% (69/140) of all patients. These variants are primarily observed in the Asian or Chinese population and are distinct from those present in Caucasian groups. In this cohort, 105 patients were diagnosed with heterozygous FH (HeFH), while 35 were diagnosed with homozygous FH (HoFH). Finally, only 28.57% of the patients (40/140) were using lipid-lowering medications with 33.33% of HoFH patients initiating treatment after the age of 8. Additionally, only 3 compound heterozygous patients (2.14%) underwent liver transplantation because of significantly high lipid levels., Conclusion: This study reveals the variable genotypes and phenotypes of children with FH in China and illustrates that the genotypes in the Chinese population differ from those in Caucasians, providing a valuable dataset for the clinical genetic screening of FH in China. Furthermore, the older age at diagnosis and treatment highlights the underdiagnosis and undertreatment of Chinese FH pediatric patients, suggesting that early identification should be improved through lipid or genetic screening, and that more timely and regular pharmacological treatments should be implemented., Competing Interests: Declarations. Ethics approval and consent to participate: The study received approval from the Ethics Committee of the Children’s Hospital of Zhejiang University School of Medicine. Written informed consent was obtained from all participants or their legal guardians prior to the performance of whole exome sequencing (WES). Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Semliki Forest virus (SFV) is a neuropathogenic alphavirus which is of interest both as a model neurotropic alphavirus and as an oncolytic virus with proven potency in preclinical cancer models. In laboratory mice, peripherally administered SFV infiltrates the central nervous system (CNS) and causes encephalitis of varying severity. The route of SFV CNS entrance is poorly understood but has been considered to occur through the blood-brain barrier. Here we show that neuroinvasion of intravenously administered SFV is strictly dependent on very-low-density-lipoprotein receptor (VLDLR) which acts as an entry receptor for SFV. Moreover, SFV primarily enters the CNS through the blood-cerebrospinal fluid (B-CSF) barrier via infecting choroid plexus epithelial cells which show distinctly high expression of VLDLR. This is the first indication of neurotropic alphavirus utilizing choroid plexus for CNS entry, and VLDLR playing a specific and crucial role for mediating SFV entry through this pathway., Competing Interests: Competing interests: The authors declare no competing interests. Ethical approval: The Swedish Work Environment Authority has approved the work with genetic modification of SFV (ID no. 202100-2932 v66a14 [laboratory] and v67a10 [mice]). All experiments regarding modified SFV were conducted under biosafety level 2. The local Animal Ethics Committee in Stockholm (13414/2020 and 04399/2023) approved the animal studies., (© 2024. The Author(s).)

Background: Familial hypercholesterolemia (FH) is a genetically inherited disorder caused by monogenic mutations or polygenic deleterious variants. Patients with FH innate with significantly elevated risks for coronary heart disease (CHD). FH prevalence based on genetic testing in Chinese CHD patients is missing. Whether classical index of coronary atherosclerosis severity can be used as indicators of FH needs to be explored. To investigate the FH prevalence in Chinese CHD patients and the association of SYNTAX I score with FH genotype., Methods: The monogenic and polygenic FH related genes were genotyped in 400 consecutively enrolled CHD patients. The clinical characteristics and SYNTAX I scores were analyzed in a retrospective nested case-control study., Results: The prevalence of genetically confirmed FH in our CHD cohort was 8.75%. The cLDL-C level, SYNTAX I scores and incidences of triple vessel lesions in FH patients were significantly higher, while cLDL-C and SYNTAX I scores were independent risk factors for FH. Furthermore, cLDL-C levels of polygenic FH were significantly lower than monogenic FH, while their severity of coronary atherosclerosis was comparable., Conclusions: Our study revealed that the SYNTAX I score was an independent risk factor for FH. Besides, polygenic origin of FH should be taken into consideration for CHD patients suspected of FH., Competing Interests: Declarations. Ethics approval and consent for participate: The study protocol was approved by the ethics committee of Beijing Chaoyang Hospital, Capital Medical University and performed in accordance with the ethical standers laid down in the 1964 Declaration of Helsinki and its later amendments. Written informed consents were obtained from all participants. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Sphingosine 1-phosphate (S1P) is a lysosphingolipid with antiatherogenic properties, but mechanisms underlying its effects remain unclear. We here investigated atherosclerosis development in cholesterol-rich diet-fed LDL receptor-deficient mice with high or low overexpression levels of S1P receptor 1 (S1P1) in macrophages. S1P1-overexpressing macrophages showed increased activity of transcription factors PU.1, interferon regulatory factor 8 (IRF8), and liver X receptor (LXR) and were skewed toward an M2-distinct phenotype characterized by enhanced production of IL-10, IL-1RA, and IL-5; increased ATP-binding cassette transporter A1- and G1-dependent cholesterol efflux; increased expression of MerTK and efferocytosis; and reduced apoptosis due to elevated B cell lymphoma 6 and Maf bZIP B. A similar macrophage phenotype was observed in mice administered S1P1-selective agonist KRP203. Mechanistically, the enhanced PU.1, IRF8, and LXR activity in S1P1-overexpressing macrophages led to downregulation of the cAMP-dependent PKA and activation of the signaling cascade encompassing protein kinases AKT and mTOR complex 1 as well as the late endosomal/lysosomal adaptor MAPK and mTOR activator 1. Atherosclerotic lesions in aortic roots and brachiocephalic arteries were profoundly or moderately reduced in mice with high and low S1P1 overexpression in macrophages, respectively. We conclude that S1P1 signaling polarizes macrophages toward an antiatherogenic functional phenotype and countervails the development of atherosclerosis in mice.

Cytokines are involved in all stages of atherosclerosis, generally contributing to disease progression. Previously, members of the Interleukin (IL)-6 cytokine family, such as IL-6, oncostatin M, and cardiotrophin-1, have been extensively studied in atherosclerosis. However, the role of leukemia inhibitory factor (LIF), member of the IL-6 family, and its receptor (LIFR), remains to be further elucidated. Therefore, the aim of this study is to provide insight in LIF receptor signalling in atherosclerosis development. Single-cell RNA sequencing analysis of human carotid artery plaques revealed that mast cells highly express LIF, whereas LIFR was specifically expressed on activated endothelial cells. A similar expression pattern of Lifr was observed in mouse atherosclerotic plaques. Next, female Western-type diet fed Ldlr -/- mice were treated with LIF receptor inhibitor EC359 (5 mg/kg s.c., n = 15) or control solvent (n = 15) three times per week for eight weeks. Stenosis grade was reduced in the aortic root of EC359 treated mice compared to control mice, but treatment did not affect plaque composition. Serum cholesterol levels were significantly reduced in EC359 treated mice, likely attributed to a reduction in VLDL cholesterol levels. Furthermore, LIF receptor inhibition reduced Pecam1 and Vcam1 expression in the aorta. Consequently, immune cell infiltration was reduced in aortic plaques of EC359 treated mice compared to control mice. Conclusively, we demonstrated that LIF receptor is a potential therapeutic target in atherosclerosis by reducing plaque size, attributed to lower serum cholesterol levels, reduced endothelial activation and less immune cell infiltration in the plaque., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Background and Aims: PCSK9 is a key regulator of LDL-cholesterol levels. PCSK9 gain of function variants (GOFVs) cause autosomal dominant hypercholesterolemia (ADH). The first described PCSK9-GOFV, p.Ser127Arg, almost exclusively reported in France, represents 67 % of the PCSK9 French GOFVs due to a founder effect. Few other carriers are reported in South Africa and Norway. This study aims to estimate when the common ancestor lived and to describe a cohort of p.Ser127Arg carriers., Methods: Eight families and 14 p.Ser127Arg carriers were genotyped and phenotyped. Haplotypes were constructed using 11 microsatellites around PCSK9 and 6 intragenic single nucleotide polymorphisms (SNPs). To add to the biological analysis, eight additional p.Ser127Arg carriers, 12 carriers of other PCSK9-GOFVs, 93 LDLR loss of function variant (LOFV) carriers and 49 non-carriers subjects were phenotyped., Results: The most common ancestor of p.Ser127Arg was estimated to have lived 775 years ago [95 % CI: 575-1075]. French Protestants exiled after the revocation of the Edict of Nantes in 1685 AD likely brought the variant to South Africa and Norway. As expected for ADH subjects, carriers of LDLR-LOFV, the p.Ser127Arg, or other PCSK9-GOFVs showed significantly higher LDL-C levels than that of the non-carriers. Interestingly, LDL-C levels are higher for LDLR-LOFVs and for the reduced secreted p.Ser127Arg than for secreted PCSK9-GOFVs, suggesting a greater effect of the p.Ser127Arg. Conversely, HDL-C was significantly lower for LDLR-LOFV and p.Ser127Arg carriers., Conclusions: This first report from a large cohort of PCSK9-p.Ser127Arg carriers provides observations suggesting a stronger hypercholesterolemic potential of the mutated pro-PCSK9 compared with the secreted mature protein. This work also provides additional data to support the association between PCSK9 and HDL metabolism, and molecular evidence that this variant appeared in France around 1248 AD (Graphical Abstract = Fig. 1)., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Objective: Aortic stenosis (AS) is characterized by narrowing of the aortic valve opening, resulting in peak blood flow velocity that induces high wall shear stress (WSS) across the valve. Severe AS leads to heart failure and death. There is no treatment available for AS other than valve replacement. Platelet-derived transforming growth factor beta 1 (TGF-β1) partially contributes to AS progression in mice, and WSS is a potent activator of latent TGF-β1. N-acetylcysteine (NAC) inhibits WSS-induced TGF-β1 activation in vitro . We hypothesize that NAC will inhibit AS progression by inhibiting WSS-induced TGF-β1 activation. Approach: We treated a cohort of Ldlr(-/-)Apob(100/100) low density lipoprotein receptor (LDLR) mice fed a high-fat diet with NAC (2% in drinking water) at different stages of disease progression and measured its effect on AS progression and TGF-β1 activation. Results: Short-term NAC treatment inhibited AS progression in mice with moderate and severe AS relative to controls, but not in LDLR mice lacking platelet-derived TGF-β1 (TGF-β1 platlet-KO -LDLR). NAC treatment reduced TGF-β signaling, p-Smad2 and collagen levels, and mesenchymal transition from isolectin B 4 and CD45-positive cells in LDLR mice. Mechanistically, NAC treatment resulted in plasma NAC concentrations ranging from 75.5 to 449.2 ng/mL, which were sufficient to block free thiol labeling of plasma proteins and reduce active TGF-β1 levels without substantially affecting reactive oxygen species-modified products in valvular cells. Conclusions: Short-term treatment with NAC inhibits AS progression by inhibiting WSS-induced TGF-β1 activation in the LDLR mouse model of AS, motivating a clinical trial of NAC and/or other thiol-reactive agent(s) as a potential therapy for AS.

VLDLR cerebellar hypoplasia is characterized by intellectual disability, non-progressive cerebellar ataxia, and seizures. The characteristic MRI findings include hypoplasia of the inferior portion of the cerebellar vermis and hemispheres, simplified cortical gyration, and a small brain stem. Biallelic VLDLR pathogenic variants cause loss-of-function of the encoded very low-density lipoprotein receptor. VLDLR exons 4 and 16 are alternatively spliced, resulting in the expression of four transcript variants, including two exon 4-lacking mRNAs expressed in the human brain. Previously reported VLDLR pathogenic variants affect all four transcript variants. Here we report on two sisters with facial dysmorphism, microcephaly, intellectual disability, and normal brain imaging. Exome sequencing in one patient identified the homozygous VLDLR nonsense variant c.376C>T; p.(Gln126*) in exon 4; her similarly affected sister also carried the homozygous variant and parents were heterozygous carriers. VLDLR transcript analysis identified mRNAs with and without exon 4 in patient fibroblasts, while exon 4-containing VLDLR mRNAs were predominantly detected in control fibroblasts. We found significantly reduced VLDLR mRNA levels in patient compared to control cells, likely caused by nonsense-mediated mRNA decay of exon 4-containing VLDLR transcripts. Expression of neuronal VLDLR isoforms produced from exon 4-lacking transcripts may have protected both patients from developing the cerebellar hypoplasia phenotype., Competing Interests: Competing interests: Seung Woo Ryu is an employee of 3billion, Inc. All other authors declare no conflicts of interest. Ethics: The parents of the two patients provided written informed consent for participation in the study, clinical data and specimen collection, genetic analysis, and publication of relevant findings., (© 2024. The Author(s).)

Hypertriglyceridemia (HTG) often accompanies diabetes and is considered a risk factor for diabetic vascular complications. However, inducing diabetic HTG typically requires high-fat diets in certain animal models. Leveraging our newly developed LDL receptor knockout hamster model, which exhibits features akin to human lipid metabolism, we sought to determine whether these animals would develop HTG without dietary manipulations in diabetes. Diabetes was induced via intraperitoneal injection of STZ in wild type and heterozygous LDL receptor deficient hamsters. Blood glucose, triglyceride, and cholesterol were measured over 60 days. Plasma TG clearance was determined via olive oil gavage. The effect of insulin on diabetic HTG was assessed on Day 60 post-diabetes induction. Blood glucose increased over threefold, while plasma insulin decreased to 30% of controls after STZ injection in both wild type and heterozygous hamsters by Day 7, remaining stable for 60 days. Plasma TG in wild-type hamsters remained unchanged at Day 7 post-STZ injection but increased slightly thereafter. Conversely, heterozygous hamsters exhibited severe HTG by Day 7 until the end of the study. Olive oil gavage revealed much slower plasma triglyceride clearance in heterozygous hamsters compared to WT animals, despite significantly reduced lipoprotein lipase activity in post-heparin plasma in both animals. Hyperglycemia and HTG in heterozygous hamsters were reversed to pre-diabetic levels following intraperitoneal insulin administration. In conclusion, severe HTG in diabetic heterozygous LDL receptor deficient hamsters developed spontaneously and was insulin-dependent. Thus, this hamster model holds promise for effectively studying the complications associated with human diabetic HTG., (© 2024 The Author(s). Animal Models and Experimental Medicine published by John Wiley & Sons Australia, Ltd on behalf of The Chinese Association for Laboratory Animal Sciences.)

Atherosclerosis caused major morbidity and mortality worldwide. Molecules possessing lipid-lowering and/or anti-inflammatory properties are potential druggable targets against atherosclerosis. We examined the anti-atherosclerotic effects of fluorescent gold nanoclusters (FANC), which were dihydrolipoic acid (DHLA)-capped 2-nm gold nanoparticles. We evaluated the 8-week effects of FANC in Western-type diet-fed ApoE-deficient mice by either continuous intraperitoneal delivery (20 μM, 50 μl weekly) or via drinking water (300 nM). FANC reduced aortic atheroma burden, serum total cholesterol, and oxidative stress markers malondialdehyde and 4-hydroxynonenal levels. FANC attenuated hepatic lipid deposit, with changed expression of lipid homeostasis-related genes HMGCR, SREBP, PCSK9, and LDLR in a pattern similar to mice treated with ezetimibe. FANC also inhibited intestinal cholesterol absorption, resembling the action of ezetimibe. The lipid-lowering and anti-atherosclerotic effects of FANC reappeared in Western-type diet-fed LDLr-deficient mice. FANC bound insulin receptor β (IRβ) via DHLA, leading to AKT activation. However, unlike insulin, which also bound IRβ to activate AKT to induce HO-1, activation of AKT by FANC was independent of HO-1 expression in human aortic endothelial cells (HAECs). Alternatively, FANC up-regulated NRF2, interfered the binding of KEAP1 to NRF2, and promoted KEAP1 degradation to free NRF2 for nuclear entry to induce HO-1 that suppressed the expression of ICAM-1 and VCAM-1. Consistently, FANC suppressed ox-LDL-induced enhanced attachment of THP-derived macrophages onto HAECs. In macrophages, FANC up-regulated ABCA1, and reversed ox-LDL-induced suppression of cholesterol efflux. FANC effected in vitro at nano moles. In conclusion, our findings showed novel actions and multiple mechanisms of FANC worked coherently against atherosclerosis., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Intracellular cholesterol metabolism is regulated by the SREBP-2 and LXR signaling pathways. The effects of inflammation on these molecular mechanisms remain poorly studied, especially at the blood-brain barrier (BBB) level. Tumor necrosis factor α (TNFα) is a proinflammatory cytokine associated with BBB dysfunction. Therefore, the aim of our study was to investigate the effects of TNFα on BBB cholesterol metabolism, focusing on its underlying signaling pathways. Using a human in vitro BBB model composed of human brain-like endothelial cells (hBLECs) and brain pericytes (HBPs), we observed that TNFα increases BBB permeability by degrading the tight junction protein CLAUDIN-5 and activating stress signaling pathways in both cell types. TNFα also promotes cholesterol release and decreases cholesterol accumulation and APOE secretion. In hBLECs, the expression of SREBP-2 targets (LDLR and HMGCR) is increased, while ABCA1 expression is decreased. In HBPs, only LDLR and ABCA1 expression is increased. TNFα treatment also induces 25-hydroxycholesterol (25-HC) production, a cholesterol metabolite involved in the immune response and intracellular cholesterol metabolism. 25-HC pretreatment attenuates TNFα-induced BBB leakage and partially alleviates the effects of TNFα on ABCA1, LDLR, and HMGCR expression. Overall, our results suggest that TNFα favors cholesterol efflux via an LXR/ABCA1-independent mechanism at the BBB, while it activates the SREBP-2 pathway. Treatment with 25-HC partially reversed the effect of TNFα on the LXR/SREBP-2 pathways. Our study provides novel perspectives for better understanding cerebrovascular signaling events linked to BBB dysfunction and cholesterol metabolism in neuroinflammatory diseases., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Objective: The gene encoding TOMM40 (Transporter of Outer Mitochondrial Membrane 40) is adjacent to that encoding APOE, which has a central role in lipid and lipoprotein metabolism. While human genetic variants near APOE and TOMM40 have been shown to be strongly associated with plasma lipid levels, a specific role for TOMM40 in lipid metabolism has not been established, and the present study was aimed at assessing this possibility., Methods: TOMM40 was knocked down by siRNA in human hepatoma HepG2 cells, and effects on mitochondrial function, lipid phenotypes, and crosstalk between mitochondria, ER, and lipid droplets were examined. Additionally, hepatic and plasma lipid levels were measured in mice following shRNA-induced knockdown of Tomm40 shRNA., Results: In HepG2 cells, TOMM40 knockdown upregulated expression of APOE and LDLR in part via activation of LXRB (NR1H2) by oxysterols, with consequent increased uptake of VLDL and LDL. This is in part due to disruption of mitochondria-endoplasmic reticulum contact sites, with resulting accrual of reactive oxygen species and non-enzymatically derived oxysterols. With TOMM40 knockdown, cellular triglyceride and lipid droplet content were increased, effects attributable in part to receptor-mediated VLDL uptake, since lipid staining was significantly reduced by concomitant suppression of either LDLR or APOE. In contrast, cellular cholesterol content was reduced due to LXRB-mediated upregulation of the ABCA1 transporter as well as increased production and secretion of oxysterol-derived cholic acid. Consistent with the findings in hepatoma cells, in vivo knockdown of TOMM40 in mice resulted in significant reductions of plasma triglyceride and cholesterol concentrations, reduced hepatic cholesterol and increased triglyceride content, and accumulation of lipid droplets leading to development of steatosis., Conclusions: These findings demonstrate a role for TOMM40 in regulating hepatic lipid and plasma lipoprotein levels and identify mechanisms linking mitochondrial function with lipid metabolism., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier GmbH.. All rights reserved.)

Toxoplasma gondii (T. gondii) is an obligate intracellular parasite that cannot biosynthesize cholesterol via the mevalonate pathway, it sources this lipid from its host. We discovered that T. gondii infection upregulated the expression of host cholesterol synthesis-related genes HMG-CoA reductase(HMGCR), squalene epoxidase (SQLE), and dehydrocholesterol reductase-7 (DHCR7), and increased the uptake pathway gene low-density lipoprotein receptor (LDLR). We found a protein, sterol regulatory element binding protein 2 (SREBP2), which is the key protein regulating the host cholesterol synthesis and uptake during T. gondii infection. T. gondii induced a dose-dependent nuclear translocation of SREBP2. Knockdown SREBP2 reduced T. gondii-induced cholesterol biosynthesis and uptake. Consequently, the parasite's ability to acquire cholesterol was significantly diminished, impairing its invasion, replication, and bradyzoites development. Interfering cholesterol metabolism using AY9944 effectively inhibited T. gondii replication. In summary, SREBP2 played an important role in T. gondii infection in vitro, serving as a potential target for regulating T. gondii-induced cholesterol metabolism, offering insights into the prevention and treatment of toxoplasmosis., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Oxidative stress plays a crucial role in the pathogenesis of coronary artery disease. In cardiovascular research using murine models, the generation and maintenance of models with robust coronary arterial atherosclerosis has been challenging., Methods: We characterized a new mouse model in which the last 3 amino acids of the carboxyl terminus of the HDL (high-density lipoprotein) receptor (SR-B1 [scavenger receptor, class B, type 1]) were deleted in a low-density lipoprotein receptor knockout (LDLR -/- ) mouse model (SR-B1ΔCT/LDLR -/- ) fed an atherogenic diet. We also tested the therapeutic effects of an oxidative stress-targeted nanoparticle in atherogenic diet-fed SR-B1ΔCT/LDLR -/- mice., Results: The SR-B1ΔCT/LDLR -/- mice fed an atherogenic diet had occlusive coronary artery atherosclerosis, impaired cardiac function, and a dramatically lower survival rate, compared with LDLR -/- mice fed the same diet. As SR-B1ΔCT/LDLR -/- mice do not exhibit female infertility or low pup yield, they are far easier and less costly to use than the previously described SR-B1-based models of coronary artery disease. We found that treatment with the targeted nanoparticles improved the cardiac functions and corrected hematologic abnormalities caused by the atherogenic diet in SR-B1ΔCT/LDLR -/- mice but did not alter the distinctive plasma lipid levels., Conclusions: The SR-B1ΔCT/LDLR -/- mice developed diet-inducible, fatal atherosclerotic coronary artery disease, which could be ameliorated by targeted nanoparticle therapy. Our study provides new tools for the development of cardiovascular therapies., Competing Interests: None.

Mitochondrial dysfunction and 12-lipoxygenase (ALOX12)-derived 12(S)-HETE production have been associated with vascular inflammation and the pathogenesis of atherosclerosis. However, the role of ALOX12 in regulating vascular energy metabolism in vascular inflammation has not been studied to date. Using mitochondrial and glycolysis functional profiling with the Seahorse extracellular flux analyzer, metabolipidomics, and proteomic analysis (LC-MS/MS), we characterized alterations in vascular energy metabolism in 2- and 6-month-old ApoE/LDLR -/- vs. control C57BL/6 mice. We identified that aorta of 6-month-old ApoE/LDLR -/- mice displayed compromised mitochondrial metabolism manifested by the reduced expression of mitochondrial enzymes, impaired mitochondrial respiration, and consequently diminished respiratory reserve capacity. An increased flux through the glycolysis/lactate shuttle, the hexosamine biosynthetic pathway (HBP), and the pentose phosphate pathway (PPP) was also recognized. Interestingly, ALOX12-12-HETE was the most upregulated axis in eicosanoid metabolism and histological examinations indicated that ApoE/LDLR -/- mice showed increased aortic expression of ALOX12, particularly in early atherosclerotic plaque areas. Remarkably, the joint blocking of ALOX12 and activation of AMPK, but not AMPK activation alone, resulted in the reprogramming of vascular metabolism, with improved mitochondrial respiration and suppressed auxiliary pathways (HBP, PPP, itaconate shunt). In conclusion, excessive activation of the ALOX12-12-HETE pathway in vascular inflammation in early atherosclerosis inhibits AMPK-dependent regulation of vascular metabolism. Consequently, ALOX12 may represent a novel target to boost impaired vascular mitochondrial function in pro-atherosclerotic vascular inflammation., Competing Interests: Declaration of Competing Interest No conflicts of interest relevant to this article were reported., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: Immune biomarkers for non-small-cell lung cancer (NSCLC) are programmed death ligand 1 (PD-L1) and tumor mutational burden (TMB). However, they cannot accurately predict the effectiveness of immunotherapy. Identifying appropriate biomarkers that can differentiate between beneficiary groups is imperative., Methods: We identified lipoprotein receptor-related protein 1B (LRP1B) mutation as a potential biomarker for immunotherapy by analyzing clinical data, combined with bioinformatics analysis. The effects of LRP1B on ferroptosis were assessed using qRT-PCR, Western blotting, CCK-8 assay, and flow cytometry. The potential mechanism underlying the regulation of ferroptosis by LRP1B was elucidated using qRT-PCR, Western blotting, ChIP, and dual-luciferase reporter gene assays., Results: Through the collection and analysis of clinical data, we had established that LRP1B mutations are closely associated with immunotherapy. Bioinformatics analysis revealed significant differences in the expression levels of PD-L1 and TMB between patients with LRP1B mutation and wild-type patients in lung adenocarcinoma (LUAD). Furthermore, we observed that patients with LRP1B mutation in LUAD had significantly higher levels of tumor-infiltrating lymphocytes (TILs) than wild-type patients. In addition, we found that patients with LRP1B mutation in LUAD had significantly prolonged progression-free survival (PFS) compared to wild-type patients. However, the differences of PD-L1 expression, TILs, and PFS were not observed in patients with LRP1B mutation in lung squamous cell carcinoma (LUSC). These findings provided strong evidence that LRP1B mutation was a potential biomarker for immunotherapy in LUAD. Moreover, our in vivo experiments indicated that knockdown of LRP1B enhanced the efficacy of mPD-1, and mechanistic studies revealed that LRP1B regulated the sensitivity of cells to ferroptosis by modulating the expression of SLC7A11 through altering the phosphorylation level of STAT3. Further analysis revealed that LRP1B knockdown promoted immunotherapy in vivo., Conclusions: Our results confirmed that LRP1B affected the efficacy of immunotherapy by modulating the sensitivity of NSCLC cells to ferroptosis. LRP1B mutations represent a highly promising immunotherapeutic biomarker for NSCLC., (© 2024 The Author(s). Cancer Medicine published by John Wiley & Sons Ltd.)

In multiple neurodegenerative diseases, including age-related macular degeneration, retinitis pigmentosa, and macular telangiectasia type 2 (MacTel), retinal pigment epithelial (RPE)-cells proliferate and migrate into the neuroretina, forming intraretinal pigment plaques. Though these pigmentary changes are hallmarks of disease progression, it is unknown if their presence is protective or detrimental.Here, we first evaluated the impact of pigment plaques on vascular changes and disease progression in MacTel. In a retrospective, longitudinal study, we analyzed multimodal retinal images of patients with MacTel and showed that pigment plaques were associated with decreased vascular leakage and stabilized neovascular growth. We then modeled the underlying pathomechanisms of pigment plaque formation in aberrant neovascular growth using the very-low-density lipoprotein receptor mutant (Vldlr -/- ) mouse. Our data indicated that during RPE-proliferation, migration and accumulation along neovessels RPE-cells underwent epithelial-mesenchymal transition (EMT). Pharmacologic inhibition of EMT in Vldlr -/- mice decreased pigment coverage, and exacerbated neovascular growth and vascular leakage.Our findings indicate that the proliferation, migration and perivascular accumulation of RPE-cells stabilize vascular proliferation and exudation, thereby exerting a protective effect on the diseased retina. We conclude that interfering with this "natural repair mechanism" may have detrimental effects on the course of the disease and should thus be avoided., Competing Interests: Declarations. Ethical approval: The studies were approved by the local ethics committees at each participating study site (Comité consultatif de protection des personnes dans la recherche biomédicale (CCPPRB); Human Research Ethics Committee (HREC); London City Road & Hampstead Research Ethics Committee; Sterling Institutional Review Board; Ethik-Kommission der Ärztekammer Westfalen-Lippe und der Medizinischen Fakultät der WWU Münster; The State of Israel Ministry of Health; The Chaim Sheba Medical Center; Ethik-Kommission - Medizinische Fakultät Bonn; University of Pennsylvania Institutional Review Board; Kantonale Ethikkommission Bern (KEK); University of Miami Human Subject Research Office), and were in adherence with the Declaration of Helsinki. All participants provided informed consent prior to participation. All animal experimental procedures were approved by The Scripps Research Institute Animal Care and Use Committee. Experiments were performed in accordance with the NIH Guide for the Care and Use of Laboratory Animals (National Academies Press, 2011). Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Background: Despite a proposed causal role for LDL-C (low-density lipoprotein cholesterol) in aortic stenosis (AS), randomized controlled trials of lipid-lowering therapy failed to prevent severe AS. We aimed to assess the impact on AS and peak velocity across the aortic valve conferred by lifelong alterations in LDL-C levels mediated by protein-disrupting variants in 3 clinically significant genes for LDL (low-density lipoprotein) metabolism ( LDLR , APOB , and PCSK9 )., Methods: We used sequencing data and electronic health records from UK Biobank (UKB) and All of Us and magnetic resonance imaging data from UKB. We identified predicted protein-disrupting variants with the Loss Of Function Transcript Effect Estimator (LOFTEE) and AlphaMissense algorithms and evaluated their associations with LDL-C and peak velocity across the aortic valve (UK Biobank), as well as diagnosed AS and aortic valve replacement (UK Biobank and All of Us)., Results: We included 421 049 unrelated participants (5621 with AS) in UKB and 195 519 unrelated participants (1087 with AS) in All of Us. Carriers of protein-disrupting variants in LDLR had higher mean LDL-C (UKB: +42.6 mg/dL; P =4.4e-237) and greater risk of AS (meta-analysis: odds ratio, 3.52 [95% CI, 2.39-5.20]; P =2.3e-10) and aortic valve replacement (meta-analysis: odds ratio, 3.78 [95% CI, 2.26-6.32]; P =4.0e-7). Carriers of protein-disrupting variants in APOB or PCSK9 had lower mean LDL-C (UKB: -32.3 mg/dL; P <5e-324) and lower risk of AS (meta-analysis: odds ratio, 0.49 [95% CI, 0.31-0.75]; P =0.001) and aortic valve replacement (meta-analysis: odds ratio, 0.54 [95% CI, 0.30-0.97]; P =0.04). Among 57 371 UKB imaging substudy participants, peak velocities across the aortic valve were greater in carriers of protein-disrupting variants in LDLR (+12.2 cm/s; P =1.6e-5) and lower in carriers of protein-disrupting variants in PCSK9 (-6.9 cm/s; P =0.022)., Conclusions: Rare genetic variants that confer lifelong higher or lower LDL-C levels are associated with substantially increased and decreased risk of AS, respectively. Early and sustained lipid-lowering therapy may slow or prevent AS development., Competing Interests: Dr Ellinor receives sponsored research support from Bayer AG, IBM Research, Bristol Myers Squibb, Pfizer, and Novo Nordisk; he has also served on advisory boards or consulted for Bayer AG. The other authors report no conflicts.

Aims: Diabetes leads to dysregulated macrophage immunometabolism, contributing to accelerated atherosclerosis progression. Identifying critical factors to restore metabolic alterations and promote resolution of inflammation remains an unmet goal. MicroRNAs orchestrate multiple signalling events in macrophages, yet their therapeutic potential in diabetes-associated atherosclerosis remains unclear., Methods and Results: miRNA profiling revealed significantly lower miR-369-3p expression in aortic intimal lesions from Ldlr-/- mice on a high-fat sucrose-containing (HFSC) diet for 12 weeks. miR-369-3p was also reduced in peripheral blood mononuclear cells from diabetic patients with coronary artery disease (CAD). Cell-type expression profiling showed miR-369-3p enrichment in aortic macrophages. In vitro, oxLDL treatment reduced miR-369-3p expression in mouse bone marrow-derived macrophages (BMDMs). Metabolic profiling in BMDMs revealed that miR-369-3p overexpression blocked the oxidized low density lipoprotein (oxLDL)-mediated increase in the cellular metabolite succinate and reduced mitochondrial respiration (OXPHOS) and inflammation [Interleukin (lL)-1β, TNF-α, and IL-6]. Mechanistically, miR-369-3p targeted the succinate receptor (GPR91) and alleviated the oxLDL-induced activation of inflammasome signalling pathways. Therapeutic administration of miR-369-3p mimics in HFSC-fed Ldlr-/- mice reduced GPR91 expression in lesional macrophages and diabetes-accelerated atherosclerosis, evident by a decrease in plaque size and pro-inflammatory Ly6Chi monocytes. RNA-Seq analyses showed more pro-resolving pathways in plaque macrophages from miR-369-3p-treated mice, consistent with an increase in macrophage efferocytosis in lesions. Finally, a GPR91 antagonist attenuated oxLDL-induced inflammation in primary monocytes from human subjects with diabetes., Conclusion: These findings establish a therapeutic role for miR-369-3p in halting diabetes-associated atherosclerosis by regulating GPR91 and macrophage succinate metabolism., Competing Interests: Conflict of interest: none declared., (© The Author(s) 2024. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Chronic activation of the adaptive immune system is a hallmark of atherosclerosis. As PI3Kδ is a key regulator of T and B cell differentiation and function, we hypothesized that alleviation of adaptive immunity by PI3Kδ inactivation may represent an attractive strategy counteracting atherogenesis. As expected, lack of hematopoietic PI3Kδ in atherosclerosis-prone Ldlr-/- mice resulted in lowered T and B cell numbers, CD4+ effector T cells, Th1 response, and immunoglobulin levels. However, despite markedly impaired peripheral pro-inflammatory Th1 cells and atheromatous CD4+ T cells, the unexpected net effect of hematopoietic PI3Kδ deficiency was aggravated vascular inflammation and atherosclerosis. Further analyses revealed that PI3Kδ deficiency impaired numbers, immunosuppressive functions, and stability of regulatory CD4+ T cells (Tregs), whereas macrophage biology remained largely unaffected. Adoptive transfer of wild-type Tregs fully restrained the atherosclerotic plaque burden in Ldlr-/- mice lacking hematopoietic PI3Kδ, whereas PI3Kδ-deficient Tregs failed to mitigate disease. Numbers of atheroprotective B-1 and pro-atherogenic B-2 cells as well as serum immunoglobulin levels remained unaffected by adoptively transferred wild-type Tregs. In conclusion, we demonstrate that hematopoietic PI3Kδ ablation promotes atherosclerosis. Mechanistically, we identified PI3Kδ signaling as a powerful driver of atheroprotective Treg responses, which outweigh PI3Kδ-driven pro-atherogenic effects of adaptive immune cells like Th1 cells.

Aim: Elevated levels of cholesterol in the bloodstream, also referred to as hypercholesterolemia, pose a significant risk for the onset of cardiovascular and cerebrovascular diseases. Oxysterols, cholesterol-derived oxidized compounds that form enzymatically or non-enzymatically, contribute to the development of atherosclerosis and coronary artery disease. This study aimed to examine the critical oxysterol levels in children and adolescents with hypercholesterolemia and explore the correlation between these levels, oxidative stress, and atherosclerosis progression., Materials and Methods: The study included 20 patients with familial hypercholesterolemia (FH) and 20 healthy individuals aged between 6 and 18 years. Participants were categorized into children (6-9 years) and adolescents (10-18 years). Pediatric and adolescent patients were selected from among subjects with LDL-C ≥ 130 mg/dL and diagnosed with heterozygous familial hypercholesterolemia (HeFH) based on the presence of mutations in the LDL receptor (LDL-R) gene. Patients with HeFH who were receiving regular atorvastatin therapy were included in the study., Results: There were no notable differences in catalase and paraoxonase (PON1) activities among the groups. However, the patient group displayed substantially higher levels of malondialdehyde (MDA) (P = 0.0108) and superoxide dismutase (SOD) activity (P = 0.0103). Compared to the healthy control group, serum chitotriosidase (CHITO) activity (P = 0.037) and chitinase 3-like protein 1 (YKL-40) levels (P = 0.0027) were significantly elevated in the patient group. Furthermore, the carotid intima-media thickness (CIMT) measurements of the patient group were significantly greater than those of the healthy group (**P < 0.0001****). The patient group exhibited significantly elevated levels of 5,6-α-epoxycholesterol, Cholestane-3β,5α,6β-triol (C-triol), and 7-ketocholesterol (7-KC), whereas 27-hydroxycholesterol (27-OHC) was significantly more abundant in the healthy group. On the other hand, while 27-OHC/Total cholesterol (Total-C) levels were significantly higher in healthy individuals, the C-Triol/Total-C ratio was significantly higher in patients. No significant differences were found between the groups in terms of 7-KC/Total-C and 5,6-α-epoxycholesterol/Total-C levels., Conclusion: This study highlights the key roles of oxysterols, oxidative stress, and macrophage activation in the development of atherosclerosis in pediatric and adolescent patients with FH. Elevated C-Triol levels in FH patients, alongside increased CIMT, point to early vascular changes despite atorvastatin therapy. In contrast, higher 27-OHC levels in healthy controls suggest differential oxysterol regulation due to cholesterol-lowering treatments in FH patients. C-Triol and 27-OHC/Total-C ratios showed potential as biomarkers to distinguish patients with FH. These findings emphasize the need for therapies targeting oxidative stress and macrophage activation in addition to cholesterol-lowering interventions., Competing Interests: Declarations Ethics approval and consent to participate This study was approved by the Ethics Committee of the Ege University Faculty of Medicine (Ethics Committee Approval No: 20-8T/31). Written informed consent was obtained from the legal guardian, and the child provided assent. All methods were performed in accordance with relevant guidelines and regulations. Consent for publication Not applicable. Competing interests The authors declare no competing interests., (© 2024. The Author(s).)

Background & Aims: Lipid metabolism disorders contribute to a range of human diseases, including liver-related pathologies. Rabbits, highly sensitive to dietary cholesterol, provide a model for understanding the development of liver disorders. Sterol regulatory element-binding protein isoform 2 (SREBP2) crucially regulates intracellular cholesterol pathways. Extra-virgin olive oil (EVOO) has shown reducing cholesterol levels and restoring liver parameters affected by HFD. The aim was to investigate the molecular impact of an HFD and supplemented with EVOO on rabbit liver cholesterol metabolism., Approach & Results: Male rabbits were assigned to dietary cohorts, including control, acute/chronic HFD, sequential HFD with EVOO, and EVOO. Parameters such as serum lipid profiles, hepatic enzymes, body weight, and molecular analyses. After 6 months of HFD, plasma and hepatic cholesterol increased with decreased SREBP2 and 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase (HMGCR) expression. Prolonged HFD increased cholesterol levels, upregulating SREBP2 mRNA and HMGCR protein. Combining this with EVOO lowered cholesterol, increased SREBP2 mRNA, and upregulated low-density lipoprotein receptor (LDLR) expression. HFD-induced metabolic dysfunction-associated fatty liver disease was mitigated by EVOO. In conclusion, the SREBP2 system responds to dietary changes., Conclusions: In rabbits, the SREBP2 system responds to dietary changes. Acute HFD hinders cholesterol synthesis, while prolonged HFD disrupts regulation, causing SREBP2 upregulation. EVOO intake prompts LDLR upregulation, potentially enhancing cholesterol clearance and restoring hepatic alterations., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Clonal hematopoiesis (CH) increases inflammasome-linked atherosclerosis, but the mechanisms by which CH mutant cells transmit inflammatory signals to nonmutant cells are largely unknown. To address this question, we transplanted 1.5% Jak2V617F (Jak2VF) bone marrow (BM) cells with 98.5% WT BM cells into hyperlipidemic Ldlr-/- mice. Low-allele-burden (LAB) mice showed accelerated atherosclerosis with increased features of plaque instability, decreased levels of the macrophage phagocytic receptors c-Mer tyrosine kinase (MERTK) and triggering receptor expressed on myeloid cells 2 (TREM2), and increased neutrophil extracellular traps (NETs). These changes were reversed when Jak2VF BM was transplanted with Il1r1-/- BM. LAB mice with noncleavable MERTK in WT BM showed improvements in necrotic core and fibrous cap formation and reduced NETs. An agonistic TREM2 antibody (4D9) markedly increased fibrous caps in both control and LAB mice, eliminating the difference between the groups. Mechanistically, 4D9 increased TREM2+PDGFB+ macrophages and PDGF receptor-α+ fibroblast-like cells in the cap region. TREM2 and PDGFB mRNA levels were positively correlated in human carotid plaques and coexpressed in macrophages. In summary, low frequencies of Jak2VF mutations promoted atherosclerosis via IL-1 signaling from Jak2VF to WT macrophages and neutrophils, promoting cleavage of phagocytic receptors and features of plaque instability. Therapeutic approaches that stabilize MERTK or TREM2 could promote plaque stabilization, especially in CH- and inflammasome-driven atherosclerosis.

Objective: Colorectal cancer is one of the most common malignant tumors in the world, which seriously threatens human health. It is essential for the search for new oncogene targets in colorectal cancer., Methods: Samples from 57 colorectal cancer patients were collected in this study. Next-Generation Sequencing (NGS) was performed to detect gene mutation, assess Microsatellite Instability (MSI), and evaluate Tumor Mutational Burden (TMB). RNA data from 528 CRC patients from the TCGA database were analyzed., Results: A total of 57 colon cancer patients were included in this study, including 30 males and 27 females, with a mean age of 56 years. In this study, the most common mutations were APC (79 %), TP53 (61 %), TTN (48 %), KRAS (42 %), SYNE1 (28 %), MUC16 (25 %), PIK3CA (25 %), FAT4 (22 %), RYR2 (19 %), OBSCN (18 %), and ZFHX4 (18 %). Subsequently, the authors analyzed gene mutations in colorectal cancer patients according to gender, age, and TMB status. Significant differences in immune cell infiltration were found between colorectal cancer tissues and normal tissues by CIBERSORT analysis. LRP1B may serve as a potential colorectal cancer therapeutic target, and its absence leads to changes in immune cell infiltration., Conclusion: The authors described the molecular characteristics of CRC. Loss of LRP1B leads to changes in immune cell infiltration and can be used as a therapeutic target for colorectal cancer., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2024. Published by Elsevier España, S.L.U.)

Familial hypercholesterolemia is an autosomal hereditary disease defined by an increased level of low-density lipoprotein cholesterol (LDL-C), which predisposes significant risks for premature cardiovascular disorders. We present a family trio study: proband, a 13-year-old Kazakh girl with homozygous familial hypercholesterolemia (HoFH) and her parents. HoFH is much more rare and severe than a heterozygous form of the disorder. HoFH patients generally present with LDL-C levels exceeding 13 mmol/L, resulting in early and life-threatening cardiovascular events within the first decades of life. In cases of neglected treatment, young patients have a risk of death from coronary diseases before the age of 30. The aim of this research was to identify genetic mutations in the affected patient and her parents. Genetic testing was necessary due to highly elevated LDL-C levels and the presence of multiple xanthomas. Targeted next-generation sequencing (NGS) was performed in this study using the Illumina TruSight cardio panel, which targets 174 genes related to cardiac disorders. The girl was diagnosed with HoFH based on the results of genetic testing. A biallelic mutation was observed in exon 3 of the low-density lipoprotein receptor ( LDLR ): c. 295 G>A (p.Glu99Lys). Sanger sequencing confirmed that the mutant gene was inherited from both parents. After confirming the genetic diagnosis of HoFH, the patient was treated with LDL apheresis and statins. This case report is the first study of HoFH in a pediatric patient from the Central Asian region. Globally, it emphasizes the need for increased clinical awareness among healthcare providers, as early detection and intervention are important for improving outcomes, particularly in pediatric patients with this rare genetic disorder.

In multiple myeloma (MM), while frequent mutations in driver genes are crucial for disease progression, they traditionally offer limited insights into patient prognosis. This study aims to enhance prognostic understanding in MM by analyzing pathway dysregulations in key cancer driver genes, thereby identifying actionable gene signatures. We conducted a detailed quantification of mutations and pathway dysregulations in 10 frequently mutated cancer driver genes in MM to characterize their comprehensive mutational impacts on the whole transcriptome. This was followed by a systematic survival analysis to identify significant gene signatures with enhanced prognostic value. Our systematic analysis highlighted 2 significant signatures, TP53 and LRP1B, which notably outperformed mere mutation status in prognostic predictions. These gene signatures remained prognostically valuable even when accounting for clinical factors, including cytogenetic abnormalities, the International Staging System (ISS), and its revised version (R-ISS). The LRP1B signature effectively distinguished high-risk patients within low/intermediate-risk categories and correlated with significant changes in the tumor immune microenvironment. Additionally, the LRP1B signature showed a strong association with proteasome inhibitor pathways, notably predicting patient responses to bortezomib and the progression from monoclonal gammopathy of unknown significance to MM. Through a rigorous analysis, this study underscores the potential of specific gene signatures in revolutionizing the prognostic landscape of MM, providing novel clinical insights that could influence future translational oncology research., (© The Author(s) 2024. Published by Oxford University Press.)

Background: Familial hypercholesterolemia (FH) is a frequently underdiagnosed genetic disorder characterized by elevated low-density lipoprotein (LDL) levels. Genetic testing of LDLR, APOB, and PCSK9 genes can identify variants in up to 80% of clinically diagnosed patients. However, limitations in time, scalability, and cost have hindered effective next-generation sequencing of these genes. Additionally, pharmacogenomic variants are associated with statin-induced adverse effects in FH patients. To address these challenges, we developed a multiplex primer-based amplicon sequencing approach for FH genetic testing., Methods: Multiplex primers were designed for the exons of the LDLR, APOB, and PCSK9 genes, as well as for pharmacogenomic variants rs4149056 (SLCO1B1:c.521T > A), rs2306283 (SLCO1B1:c.388A > G), and rs2231142 (ABCG2:c.421C > A). Analytical validation using samples with known pathogenic variants and clinical validation with 12 FH-suspected probands were conducted. Library preparation was based on a bead-based tagmentation method, and sequencing was conducted on the NovaSeq 6000 platform., Results: Our approach ensured no amplicon dropouts, with over 100× coverage on each amplicon. Known variants in 2 samples were successfully detected. Further, we identified one heterozygous LDLR (p.Glu228Ter) variant and 2 homozygous cases of LDLR (p.Lys294Ter) and LDLR (p.Ser177Leu) variants in patients. Pharmacogenomic analysis revealed that overall 3 patients may require reduced statin doses. Our approach offered reduced library preparation time (approximately 3 h), greater scalability, and lower costs (under $50) for FH genetic testing., Conclusions: Our method effectively sequences LDLR, APOB, and PCSK9 genes including pharmacogenomic variants that will guide appropriate screening and statin dosing, thus increasing both efficiency and affordability., (© Association for Diagnostics & Laboratory Medicine 2024. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Integrin α9β1 is known to stabilize leukocyte adhesion to the activated endothelium. We determined the role of myeloid cell α9β1 in early atherosclerosis in two models: α9Mye-KOApoe-/- or the Ldlr-/- mice transplanted with bone marrow (BM) from α9Mye-KO mice fed a high-fat "Western" diet for 4 wk. α9Mye-KOApoe-/- mice exhibited reduced early lesions in the aortae and aortic sinuses (P < 0.05 vs α9WT Apoe-/- mice). Similar results were obtained in α9Mye-KO BM→Ldlr-/- mice (P < 0.05 vs α9WT BM→Ldlr-/- mice). Reduced early atherosclerosis in α9Mye-KOApoe-/- mice was associated with decreased neutrophil and neutrophil extracellular traps (NETs) content in the aortic lesions (P < 0.05 vs α9WTApoe-/-). Vascular cell adhesion molecule-1-stimulated neutrophils from α9Mye-KO mice exhibited reduced adhesion, transmigration, and NETs formation (NETosis) (P < 0.05 vs α9WT neutrophils). Reduced NETosis was associated with decreased extracellular signal-regulated kinase phosphorylation, peptidyl arginine deiminase 4, and citrullinated histone H3 expression. In summary, genetic ablation of myeloid cell-specific α9 reduces early atherosclerosis, most likely by reducing neutrophil adhesion, transmigration, and NETosis., Competing Interests: Conflict of interest statement. None declared., (© The Author(s) 2024. Published by Oxford University Press on behalf of Society for Leukocyte Biology.)

Background: Familial hypercholesterolemia (FH) is a hereditary disease caused mainly by mutations in the gene encoding the low-density lipoprotein receptor (LDLR). This study aimed to confirm the pathogenicity of the LDLR c.97C>T (p.Gln33Ter) mutation through in vitro functional validation and determine whether this nonsense mutation induces nonsense-mediated mRNA decay (NMD)., Methods: The proband and his family were included in accordance with Chinese Expert Consensus on FH screening. The disease-causing mutations were fund using whole-exome sequencing and were confirmed using bidirectional Sanger sequencing. The pathogenicity of the mutation was predicted using in silico analysis. The LDLR c.97C>T (p.Gln33Ter) mutation was generated using site-directed mutagenesis and expressed in HEK293T cells lacking endogenous LDLR expression. The effects of this alteration on LDLR expression and LDL uptake were assessed using flow cytometry, quantitative polymerase chain analysis, western blotting, and confocal laser scanning microscopy., Results: The mutation that causes FH in this family was LDLR c.97C>T (p.Gln33Ter), and family members with this mutation exhibited elevated levels of low-density lipoprotein cholesterol (LDL-C). The cell experiment results showed that this mutation prevented the synthesis of LDLR protein and caused the cells to lose their LDL uptake ability., Conclusion: LDLR c.97C>T (p.Gln33Ter) is a pathogenic FH mutation. However, this nonsense mutation did not induce NMD., (© 2024 The Author(s). Molecular Genetics & Genomic Medicine published by Wiley Periodicals LLC.)