LDL Receptor-Related Protein 1B Polymorphisms Associated with Increased Risk of Lymph Node Metastasis in Oral Cancer Group with Diabetes Mellitus.

Oral cancer ranks fourth among malignancies among Taiwanese men and is the eighth most common cancer among men worldwide in terms of general diagnosis. The purpose of the current study was to investigate how low-density lipoprotein receptor-related protein 1B (LDL receptor related protein 1B; LRP1B) gene polymorphisms affect oral squamous cell carcinoma (OSCC) risk and progression in individuals with diabetes mellitus (DM). Three LRP1B single-nucleotide polymorphisms (SNPs), including rs10496915, rs431809, and rs6742944, were evaluated in 311 OSCC cases and 300 controls. Between the case and control groups, we found no evidence of a significant correlation between the risk of OSCC and any of the three specific SNPs. Nevertheless, in evaluating the clinicopathological criteria, individuals with DM who possess a minimum of one minor allele of rs10496915 (AC + CC; p = 0.046) were significantly associated with tumor size compared with those with homozygous major alleles (AA). Similarly, compared to genotypes homologous for the main allele (GG), rs6742944 genotypes (GA + AA; p = 0.010) were more likely to develop lymph node metastases. The tongue and the rs6742944 genotypes (GA + AA) exhibited higher rates of advanced clinical stages (p = 0.024) and lymph node metastases (p = 0.007) when compared to homozygous alleles (GG). LRP1B genetic polymorphisms appear to be prognostic and diagnostic markers for OSCC and DM, as well as contributing to genetic profiling research for personalized medicine.

Targeting PCSK9 to upregulate MHC-II on the surface of tumor cells in tumor immunotherapy.

BACKGROUND: Proprotein convertase subtilisin/kexin type 9 (PCSK9), the last member of the proprotein convertase family, functions as a classic regulator of low-density lipoprotein (LDL) by interacting with low-density lipoprotein receptor (LDLR). Recent studies have shown that PCSK9 can affect the occurrence and development of tumors and can be used as a novel therapeutic target. However, a comprehensive pan-cancer analysis of PCSK9 has yet to be conducted. METHODS: The potential oncogenic effects of PCSK9 in 33 types of tumors were explored based on the datasets of The Cancer Genome Atlas (TCGA) dataset. In addition, the immune regulatory role of PCSK9 inhibition was evaluated via in vitro cell coculture and the tumor-bearing mouse model. Finally, the antitumor efficacy of targeted PCSK9 combined with OVA-II vaccines was verified. RESULTS: Our results indicated that PCSK9 was highly expressed in most tumor types and was significantly correlated with late disease stage and poor prognosis. Additionally, PCSK9 may regulate the tumor immune matrix score, immune cell infiltration, immune checkpoint expression, and major histocompatibility complex expression. Notably, we first found that dendritic cell (DC) infiltration and major histocompatibility complex-II (MHC-II) expression could be upregulated by PCSK9 inhibition and improve CD8+ T cell activation in the tumor immune microenvironment, thereby achieving potent tumor control. Combining PCSK9 inhibitors could enhance the efficacies of OVA-II tumor vaccine monotherapy. CONCLUSIONS: Conclusively, our pan-cancer analysis provided a more comprehensive understanding of the oncogenic and immunoregulatory roles of PCSK9 and demonstrated that targeting PCSK9 could increase the efficacy of long peptide vaccines by upregulating DC infiltration and MHC-II expression on the surface of tumor cells. This study reveals the critical oncogenic and immunoregulatory roles of PCSK9 in various tumors and shows the promise of PCSK9 as a potent immunotherapy target.

LDLR c.415G > A causes familial hypercholesterolemia by weakening LDLR binding to LDL.

BACKGROUND: Familial hypercholesterolemia (FH) is a prevalent hereditary disease that can cause aberrant cholesterol metabolism. In this study, we confirmed that c.415G > A in low-density lipoprotein receptor (LDLR), an FH-related gene, is a pathogenic variant in FH by in silico analysis and functional experiments. METHODS: The proband and his family were evaluated using the diagnostic criteria of the Dutch Lipid Clinic Network. Whole-exome and Sanger sequencing were used to explore and validate FH-related variants. In silico analyses were used to evaluate the pathogenicity of the candidate variant and its impact on protein stability. Molecular and biochemical methods were performed to examine the effects of the LDLR c.415G > A variant in vitro. RESULTS: Four of six participants had a diagnosis of FH. It was estimated that the LDLR c.415G > A variant in this family was likely pathogenic. Western blotting and qPCR suggested that LDLR c.415G > A does not affect protein expression. Functional studies showed that this variant may lead to dyslipidemia by impairing the binding and absorption of LDLR to low-density lipoprotein ( LDL). CONCLUSION: LDLR c.415G > A is a pathogenic variant in FH; it causes a significant reduction in LDLR's capacity to bind LDL, resulting in impaired LDL uptake. These findings expand the spectrum of variants associated with FH.

Prior Exposure to Experimental Preeclampsia Increases Atherosclerotic Plaque Inflammation in Atherogenic Mice-Brief Report.

BACKGROUND: Women with a history of preeclampsia have evidence of premature atherosclerosis and increased risk of myocardial infarction and stroke compared with women who had a normotensive pregnancy. Whether this is due to common risk factors or a direct impact of prior preeclampsia exposure has never been tested in a mouse atherosclerosis model. METHODS: Pregnant LDLR-KO (low-density lipoprotein receptor knockout; n=35) female mice were randomized in midgestation to sFlt1 (soluble fms-like tyrosine kinase 1)-expressing adenovirus or identical control adenovirus. Postpartum, mice were fed high-fat diet for 8 weeks to induce atherogenesis. Comparison between the control and preeclampsia models was made for metabolic parameters, atherosclerosis burden and composition by histology, plaque inflammation by flow cytometry, and aortic cytokines and inflammatory markers using a cytokine array. RESULTS: In pregnant LDLR-KO mice, sFlt1 adenovirus significantly induced serum sFlt1, blood pressure, renal endotheliosis, and decreased pup viability. After 8 weeks of postpartum high fat feeding, body weight, fasting glucose, plasma cholesterol, HDL (high-density lipoprotein), and LDL (low-density lipoprotein) were not significantly different between groups with no change in aortic root plaque size, lipid content, or necrotic core area. Flow cytometry demonstrated significantly increased CD45+ aortic arch leukocytes and CD3+T cells and aortic lysate contained more CCL (CC motif chemokine ligand) 22 and fetuin A and decreased expression of IGFBP6 (insulin-like growth factor-binding protein 6) and CCL21 in preeclampsia-exposed mice compared with controls. CONCLUSIONS: In atherogenic LDLR-KO mice, exposure to sFlt1-induced preeclampsia during pregnancy increases future atherosclerotic plaque inflammation, supporting the concept that preeclampsia directly exacerbates atherosclerotic inflammation independent of preexisting risk factors. This mechanism may contribute to ischemic vascular disease in women after preeclampsia pregnancy.

In vivo chronic exposure to inorganic mercury worsens hypercholesterolemia, oxidative stress and atherosclerosis in the LDL receptor knockout mice.

Heavy metal exposure leads to multiple system dysfunctions. The mechanisms are likely multifactorial and involve inflammation and oxidative stress. The aim of this study was to evaluate markers and risk factors for atherosclerosis in the LDL receptor knockout mouse model chronically exposed to inorganic mercury (Hg) in the drinking water. Results revealed that Hg exposed mice present increased plasma levels of cholesterol, without alterations in glucose. As a major source and target of oxidants, we evaluated mitochondrial function. We found that liver mitochondria from Hg treated mice show worse respiratory control, lower oxidative phosphorylation efficiency and increased H2O2 release. In addition, Hg induced mitochondrial membrane permeability transition. Erythrocytes from Hg treated mice showed a 50% reduction in their ability to take up oxygen, lower levels of reduced glutathione (GSH) and of antioxidant enzymes (SOD, catalase and GPx). The Hg treatment disturbed immune system cells counting and function. While lymphocytes were reduced, monocytes, eosinophils and neutrophils were increased. Peritoneal macrophages from Hg treated mice showed increased phagocytic activity. Hg exposed mice tissues present metal impregnation and parenchymal architecture alterations. In agreement, increased systemic markers of liver and kidney dysfunction were observed. Plasma, liver and kidney oxidative damage indicators (MDA and carbonyl) were increased while GSH and thiol groups were diminished by Hg exposure. Importantly, atherosclerotic lesion size in the aorta root of Hg exposed mice were larger than in controls. In conclusion, in vivo chronic exposure to Hg worsens the hypercholesterolemia, impairs mitochondrial bioenergetics and redox function, alters immune cells profile and function, causes several tissues oxidative damage and accelerates atherosclerosis development.

CRISPR-Cas9-guided amplification-free genomic diagnosis for familial hypercholesterolemia using nanopore sequencing.

Familial hypercholesterolemia is an inherited disorder that remains underdiagnosed. Conventional genetic testing methods such as next-generation sequencing (NGS) or target PCR are based on the amplification process. Due to the efficiency limits of polymerase and ligase enzymes, these methods usually target short regions and do not detect large mutations straightforwardly. This study combined the long-read nanopore sequencing and CRISPR-Cas9 system to sequence the target DNA molecules without amplification. We originally designed and optimized the CRISPR-RNA panel to target the low-density lipoprotein receptor gene (LDLR) and proprotein convertase subtilisin/kexin type 9 gene (PCSK9) from human genomic DNA followed by nanopore sequencing. The average coverages for LDLR and PCSK9 were 106× and 420×, versus 1.2× for the background genome. Among them, continuous reads were 52x and 307x, respectively, and spanned the entire length of LDLR and PCSK9. We identified pathogenic mutations in both coding and splicing donor regions in LDLR. We also detected an 11,029 bp large deletion in another case. Furthermore, using continuous long reads generated from the benchmark experiment, we demonstrated how a false-positive 670 bp deletion caused by PCR amplification errors was easily eliminated.

[The diagnostic value of genetic testing in familial hypercholesterolemia in patients with premature myocardial infarction].

Objective: To evaluate the diagnostic value of gene testing in familial hypercholesterolemia (FH) in patients with premature myocardial infarction(PMI). Methods: This study was a single center cross-sectional study. A retrospective analysis was made on PMI patients who visited the People's Hospital of Peking University from May 1, 2015 to March 31, 2017. Clinical data of patients was collected and gene testing of FH related genes low density lipoprotein receptor (LDLR), proprotein convertase subtilisin/kexin type 9 (PCSK9), apolipoprotein B(APOB) and low density lipoprotein receptor adaptor protein 1(LDLRAP1) was carried out. Clinical diagnosis of FH patients was performed using Simon Broome criteria, DLCN criteria, and FH Chinese expert consensus. Results: There were 188 males (83.6%) among 225 PMI patients, and the age of the first myocardial infarction was (46.6±7.2) years old. Ten patients carried FH pathogenic or possibly pathogenic mutations (4.4%). Compared with Simon Broome standard, DLCN standard and FH Chinese expert consensus, gene testing increased the diagnostic rate of FH by 53.3%, 33.3% and 42.1% respectively. Conclusion: Gene testing is helpful to improve the diagnosis of FH, and it is important to start the standard treatment of FH as early as possible in patients with premature myocardial infarction.

Identification of a novel LDLR p.Glu179Met variant in Thai families with familial hypercholesterolemia and response to treatment with PCSK9 inhibitor.

Familial hypercholesterolemia (FH) is a genetic disease characterized by elevated LDL-C levels. In this study, two FH probands and 9 family members from two families from northeastern Thailand were tested for LDLR, APOB, and PCSK9 variants by whole-exome sequencing, PCR-HRM, and Sanger sequencing. In silico analysis of LDLR was performed to analyse its structure‒function relationship. A novel variant of LDLR (c.535_536delinsAT, p.Glu179Met) was detected in proband 1 and proband 2 in homozygous and heterozygous forms, respectively. A total of 6 of 9 family members were heterozygous for LDLR p.Glu179Met variant. Compared with proband 2, proband 1 had higher baseline TC and LDL-C levels and a poorer response to lipid-lowering therapy combined with a PCSK9 inhibitor. Multiple sequence alignment showed that LDLR p.Glu179Met was located in a fully conserved region. Homology modelling demonstrated that LDLR p.Glu179Met variant lost one H-bond and a negative charge. In conclusion, a novel LDLR p.Glu179Met variant was identified for the first time in Thai FH patients. This was also the first report of homozygous FH patient in Thailand. Our findings may expand the knowledge of FH-causing variants in Thai population, which is beneficial for cascade screening, genetic counselling, and FH management to prevent coronary artery disease.

Recapitulating familial hypercholesterolemia in a mouse model by knock-in patient-specific LDLR mutation.

Familial hypercholesterolemia (FH) is one of the most prevalent monogenetic disorders leading to cardiovascular disease (CVD) worldwide. Mutations in Ldlr, encoding a membrane-spanning protein, account for the majority of FH cases. No effective and safe clinical treatments are available for FH. Adenine base editor (ABE)-mediated molecular therapy is a promising therapeutic strategy to treat genetic diseases caused by point mutations, with evidence of successful treatment in mouse disease models. However, due to the differences in the genomes between mice and humans, ABE with specific sgRNA, a key gene correction component, cannot be directly used to treat FH patients. Thus, we generated a knock-in mouse model harboring the partial patient-specific fragment and including the Ldlr W490X mutation. LdlrW490X/W490X mice recapitulated cholesterol metabolic disorder and clinical manifestations of atherosclerosis associated with FH patients, including high plasma low-density lipoprotein cholesterol levels and lipid deposition in aortic vessels. Additionally, we showed that the mutant Ldlr gene could be repaired using ABE with the cellular model. Taken together, these results pave the way for ABE-mediated molecular therapy for FH.

High Prevalence of Familial Hypercholesterolemia Due to the Founder Effect of the LDLR c.2271del Variant in Communities of Oaxaca, Mexico.

INTRODUCTION: In Mexico, familial hypercholesterolemia (FH) is underdiagnosed, but population screening in small communities where at least one homozygous patient has already been detected results in a useful and inexpensive approach to reduce this problem. Considering that we previously reported nine homozygous cases from the state of Oaxaca, we decided to perform a population screening to identify patients with FH and to describe both their biochemical and genetic characteristics. METHODS: LDL cholesterol (LDLc) was quantified in 2,093 individuals from 11 communities in Oaxaca; either adults with LDLc levels ≥170 mg/dL or children with LDLc ≥130 mg/dL were classified as suggestive of FH and therefore included in the genetic study. LDLR and APOB (547bp fragment of exon 26) genes were screened by sequencing and MLPA analysis. RESULTS: Two hundred and five individuals had suggestive FH, with a mean LDLc of 223 ± 54 mg/dL (range: 131-383 mg/dL). Two pathogenic variants in the LDLR gene were detected in 149 individuals: c.-139_-130del (n = 1) and c.2271del (n = 148). All patients had a heterozygous genotype. With the cascade screening of their relatives (n = 177), 15 heterozygous individuals for the c.2271del variant were identified, presenting a mean LDLc of 133 ± 35 mg/dL (range: 60-168 mg/dL). CONCLUSIONS: The FH frequency in this study was 7.8% (164/2093), the highest reported worldwide. A founder effect combined with inbreeding could be responsible for the high percentage of patients with the LDLR c.2271del variant (99.4%), which allowed us to detect both significant biochemical heterogeneity and incomplete penetrance; hence, we assumed the presence of phenotype-modifying variants.

Familial hypercholesterolemia with special focus on Japan.

Familial hypercholesterolemia (FH) is an inherited disorder characterized by increased low-density lipoprotein LDL) cholesterol and atherosclerotic cardiovascular disease. Although initial genetic analysis linked FH to LDL receptor mutations, subsequent work demonstrated that a gain-of-function mutation in the proprotein convertase subtilisin/kexin type 9 (PCSK9), which causes LDL-R degradation, was shown to be the cause of FH. In this review, we describe the history of research on FH, its clinical phenotyping and genotyping and advances in treatment with special focus on Japan.

The Application of Peptide Nucleic Acids (PNA) in the Inhibition of Proprotein Convertase Subtilisin/Kexin 9 (PCSK9) Gene Expression in a Cell-Free Transcription/Translation System.

Proprotein convertase subtilisin/kexin 9 (PCSK9) is a protein that plays a key role in the metabolism of low-density lipoprotein (LDL) cholesterol. The gain-of-function mutations of the PCSK9 gene lead to a reduced number of surface LDL receptors by binding to them, eventually leading to endosomal degradation. This, in turn, is the culprit of hypercholesterolemia, resulting in accelerated atherogenesis. The modern treatment for hypercholesterolemia encompasses the use of biological drugs against PCSK9, like monoclonal antibodies and gene expression modulators such as inclisiran-a short, interfering RNA (siRNA). Peptide nucleic acid (PNA) is a synthetic analog of nucleic acid that possesses a synthetic peptide skeleton instead of a phosphate-sugar one. This different structure determines the unique properties of PNA (e.g., neutral charge, enzymatic resistance, and an enormously high affinity with complementary DNA and RNA). Therefore, it might be possible to use PNA against PCSK9 in the treatment of hypercholesterolemia. We sought to explore the impact of three selected PNA oligomers on PCSK9 gene expression. Using a cell-free transcription/translation system, we showed that one of the tested PNA strands was able to reduce the PCSK9 gene expression down to 74%, 64%, and 68%, as measured by RT-real-time PCR, Western blot, and HPLC, respectively. This preliminary study shows the high applicability of a cell-free enzymatic environment as an efficient tool in the initial evaluation of biologically active PNA molecules in the field of hypercholesterolemia research. This cell-free approach allows for the omission of the hurdles associated with transmembrane PNA transportation at the early stage of PNA selection.

LDLR gene rearrangements in Czech FH patients likely arise from one mutational event.

BACKGROUND: Large deletions and duplications within the low-density lipoprotein receptor (LDLR) gene make up approximately 10% of LDLR pathogenic variants found in Czech patients with familial hypercholesterolemia. The goal of this study was to test the hypothesis that all probands with each rearrangement share identical breakpoints inherited from a common ancestor and to determine the role of Alu repetitive elements in the generation of these rearrangements. METHODS: The breakpoint sequence was determined by PCR amplification and Sanger sequencing. To confirm the breakpoint position, an NGS analysis was performed. Haplotype analysis of common LDLR variants was performed using PCR and Sanger sequencing. RESULTS: The breakpoints of 8 rearrangements within the LDLR gene were analysed, including the four most common LDLR rearrangements in the Czech population (number of probands ranging from 8 to 28), and four less common rearrangements (1-4 probands). Probands with a specific rearrangement shared identical breakpoint positions and haplotypes associated with the rearrangement, suggesting a shared origin from a common ancestor. All breakpoints except for one were located inside an Alu element. In 6 out of 8 breakpoints, there was high homology (≥ 70%) between the two Alu repeats in which the break occurred. CONCLUSIONS: The most common rearrangements of the LDLR gene in the Czech population likely arose from one mutational event. Alu elements likely played a role in the generation of the majority of rearrangements inside the LDLR gene.

OptiMo-LDLr: An Integrated In Silico Model with Enhanced Predictive Power for LDL Receptor Variants, Unraveling Hot Spot Pathogenic Residues.

Familial hypercholesterolemia (FH) is an inherited metabolic disease affecting cholesterol metabolism, with 90% of cases caused by mutations in the LDL receptor gene (LDLR), primarily missense mutations. This study aims to integrate six commonly used predictive software to create a new model for predicting LDLR mutation pathogenicity and mapping hot spot residues. Six predictive-software are selected: Polyphen-2, SIFT, MutationTaster, REVEL, VARITY, and MLb-LDLr. Software accuracy is tested with the characterized variants annotated in ClinVar and, by bioinformatic and machine learning techniques all models are integrated into a more accurate one. The resulting optimized model presents a specificity of 96.71% and a sensitivity of 98.36%. Hot spot residues with high potential of pathogenicity appear across all domains except for the signal peptide and the O-linked domain. In addition, translating this information into 3D structure of the LDLr highlights potentially pathogenic clusters within the different domains, which may be related to specific biological function. The results of this work provide a powerful tool to classify LDLR pathogenic variants. Moreover, an open-access guide user interface (OptiMo-LDLr) is provided to the scientific community. This study shows that combination of several predictive software results in a more accurate prediction to help clinicians in FH diagnosis.

Novel LDLR variants affecting low density lipoprotein metabolism identified in familial hypercholesterolemia.

BACKGROUND: Familial hypercholesterolemia (FH) is an autosomal dominant disease of lipid metabolism mainly caused by mutations in the low-density lipoprotein receptor (LDLR) gene. Genetic detection of patients with FH help with precise diagnosis and treatment, thus reducing the risk of coronary heart disease (CHD) and other related diseases. The study aimed to identify the causative gene mutations in a Chinese FH family and reveal the pathogenicity and the mechanism of these mutations. METHODS AND RESULTS: Whole exome sequencing was performed in a patient with severe lipid metabolism dysfunction seeking fertility guidance from a Chinese FH family. Two LDLR variants c.1875 C > G (p.N625K; novel variant) and c.1448G > A (p.W483*) were identified in the family. Wildtype and mutant LDLR constructs were established by the site-direct mutagenesis technique. Functional studies were carried out by cell transfection to evaluate the impact of detected variants on LDLR activity. The two variants were proven to affect LDL uptake and binding, resulting in cholesterol clearance reduction to different degrees. According to The American College of Medical Genetics and Genomics (ACMG) Standards and Guidelines, the W483* variant was classified as "Pathogenic", while the N625K variant as "VUS". CONCLUSIONS: Our results provide novel experimental evidence of functional alteration by LDLR variants identified in our study and expand the mutational spectrum of LDLR mutation induced FH.

Finasteride delays atherosclerosis progression in mice and is associated with a reduction in plasma cholesterol in men.

Finasteride is commonly prescribed to treat benign prostate hyperplasia and male-pattern baldness in cis men and, more recently, trans individuals. However, the effect of finasteride on cardiovascular disease remains elusive. We evaluated the role of finasteride on atherosclerosis using low-density lipoprotein (LDL) receptor-deficient (Ldlr-/-) mice. Next, we examined the relevance to humans by analyzing the data deposited between 2009 and 2016 in the National Health and Nutrition Examination Survey. We show that finasteride reduces total plasma cholesterol and delays the development of atherosclerosis in Ldlr-/- mice. Finasteride reduced monocytosis, monocyte recruitment to the lesion, macrophage lesion content, and necrotic core area, the latter of which is an indicator of plaque vulnerability in humans. RNA sequencing analysis revealed a downregulation of inflammatory pathways and an upregulation of bile acid metabolism, oxidative phosphorylation, and cholesterol pathways in the liver of mice taking finasteride. Men reporting the use of finasteride showed lower plasma levels of cholesterol and LDL-cholesterol than those not taking the drug. Our data unveil finasteride as a potential treatment to delay cardiovascular disease in people by improving the plasma lipid profile.

Unlocking the mysteries of VLDL: exploring its production, intracellular trafficking, and metabolism as therapeutic targets.

Reducing circulating lipid levels is the centerpiece of strategies for preventing and treating atherosclerotic cardiovascular disease (ASCVD). Despite many available lipid-lowering medications, a substantial residual cardiovascular risk remains. Current clinical guidelines focus on plasma levels of low-density lipoprotein (LDL). Recent attention has been given to very low-density lipoprotein (VLDL), the precursor to LDL, and its role in the development of coronary atherosclerosis. Preclinical investigations have revealed that interventions targeting VLDL production or promoting VLDL metabolism, independent of the LDL receptor, can potentially decrease cholesterol levels and provide therapeutic benefits. Currently, methods, such as mipomersen, lomitapide, and ANGPTL3 inhibitors, are used to reduce plasma cholesterol and triglyceride levels by regulating the lipidation, secretion, and metabolism of VLDL. Targeting VLDL represents an avenue for new lipid-lowering strategies. Interventions aimed at reducing VLDL production or enhancing VLDL metabolism, independent of the LDL receptor, hold promise for lowering cholesterol levels and providing therapeutic benefits beyond LDL in the management of ASCVD.

Is It Ever Wise to Edit Wild-Type Alleles? Engineered CRISPR Alleles Versus Millions of Years of Human Evolution.

The tremendous burden of lipid metabolism diseases, coupled with recent developments in human somatic gene editing, has motivated researchers to propose population-wide somatic gene editing of PCSK9 (proprotein convertase subtilisin/kexin type 9) within the livers of otherwise healthy humans. The best-characterized molecular function of PCSK9 is its ability to regulate plasma LDL (low-density lipoprotein) levels through promoting LDL receptor degradation. Individuals with loss-of-function PCSK9 variants have lower levels of plasma LDL and reduced cardiovascular disease. Gain-of-function variants of PCSK9 are strongly associated with familial hypercholesterolemia. A new therapeutic strategy delivers CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats; CRISPR-associated protein 9) specifically to liver cells to edit the wild-type alleles of PCSK9 with the goal of producing a loss-of-function allele. This direct somatic gene editing approach is being pursued despite the availability of US Food and Drug Administration-approved PCSK9 inhibitors that lower plasma LDL levels. Here, we discuss other characterized functions of PCSK9 including its role in infection and host immunity. We explore important factors that may have contributed to the evolutionary selection of PCSK9 in several vertebrates, including humans. Until such time that more fully understand the multiple biological roles of PCSK9, the ethics of permanently editing the gene locus in healthy, wild-type populations remains highly questionable.

Deletion of Elovl5 leads to dyslipidemia and atherosclerosis in LDLR-deficient mice.

Atherosclerosis is a chronic inflammatory disease for which hepatic steatosis and atherogenic dyslipidemia are significant risk factors. We investigated the effects of endogenously generated very-long-chain polyunsaturated fatty acids (VL-PUFAs) on dyslipidemia and atherosclerosis development using mice that lack ELOVL5, a PUFA elongase that is required for the synthesis of arachidonic acid, EPA, and DHA from the essential fatty acids linoleic and linolenic acids, and the LDL receptor (LDLR). Elovl5-/-;Ldlr-/- mice manifest increased liver triglyceride and cholesterol concentrations due to the activation of sterol regulatory element binding protein-1, a transcription factor that activates enzymes required for de novo lipogenesis. Plasma levels of triglycerides and cholesterol in VLDL, IDL, and LDL were markedly elevated in Elovl5-/-;Ldlr-/- mice fed a chow and the mice exhibited marked aortic atherosclerotic plaques. Bone marrow-derived monocytes from wild-type (WT) and Elovl5-/- mice were polarized to M1 and M2 macrophages, and the effects of ELOVL5 on inflammatory activity were determined. There were no differences in most of the markers tested for M1 and M2 polarized cells between WT and Elovl5-/- cells, except for a slight increase in PGE2 secretion in Elovl5-/- cells, likely due to elevated Cox-2 expression. These results suggest that the deletion of Elovl5 leads to hepatic steatosis and dyslipidemia, which are the major factors in severe atherosclerosis in Elovl5-/-;Ldlr-/- mice.

Schisandrin A in Schisandra chinensis Upregulates the LDL Receptor by Inhibiting PCSK9 Protein Stabilization in Steatotic Model.

Schisandra chinensis extract (SCE) protects against hypocholesterolemia by inhibiting proprotein convertase subtilisin/kexin 9 (PCSK9) protein stabilization. We hypothesized that the hypocholesterolemic activity of SCE can be attributable to upregulation of the PCSK9 inhibition-associated low-density lipoprotein receptor (LDLR). Male mice were fed a low-fat diet or a Western diet (WD) containing SCE at 1% for 12 weeks. WD increased final body weight and blood LDL cholesterol levels as well as alanine transaminase and aspartate aminotransferase expression. However, SCE supplementation significantly attenuated the increase in blood markers caused by WD. SCE also attenuated WD-mediated increases in hepatic LDLR protein expression in the obese mice. In addition, SCE increased LDLR protein expression and attenuated cellular PCSK9 levels in HepG2 cells supplemented with delipidated serum (DLPS). Non-toxic concentrations of schisandrin A (SA), one of the active components of SCE, significantly increased LDLR expression and tended to decrease PCSK9 protein levels in DLPS-treated HepG2 cells. High levels of SA-mediated PCSK9 attenuation was not attributable to reduced PCSK9 gene expression, but was associated with free PCSK9 protein degradation in this cell model. Our findings show that PCSK9 secretion can be significantly reduced by SA treatment, contributing to reductions in free cholesterol levels.