Low Density Lipoprotein Receptor Gene Research Articles

Diosgenin and 4-Hydroxyisoleucine from Fenugreek Are Regulators of Genes Involved in Lipid Metabolism in The Human Colorectal Cancer Cell Line SW480.

Objective Diosignin and 4-hydroxy-L-isulosine (4-OH-Ile) are the two active ingredients of Fenugreek (Trigonella foenum- graecum). Thus, in this study, we examined the effects of hydroalcoholic extract of fenugreek seeds (HEFS), diosgenin and 4-OH-Ile on the expression of acetyl-CoA carboxylase (ACC), fatty acid synthase (FAS), peroxisome proliferator-activated receptor gamma (PPARγ) and low-density lipoprotein (LDL) receptor (LDLR) which are involved in lipid metabolism in SW480 cell line. Materials and Methods In this experimental study, SW480 cells were cultured in RPMI-1640 medium and treated with HEFS, diosignin, 4-OH-Ile or orlistat for 24 and 48 hours. Inhibitory concentration of 20% (IC20) was calculated using MTT method and cells were then pre-treated with the IC20 concentrations for 24 and 48 hours before RNA extraction and cDNA synthesis. Changes in the expression of ACC, FAS, PPARγ and LDLR genes were assayed by employing the real time-polymerase chain reaction (PCR) method. Results Our results showed a significant down-regulation in the expression of ACC (P<0.001 and P<0.001 after 24 and 48 hours, respectively) and FAS genes (P<0.001 and P<0.001 after 24 and 48 hours, respectively) in SW480 cells treated with HEFS, diosignin, 4-OH-Ile, or orlistat, but significant up-regulation in the expression of PPARγ (P<0.001 and P<0.001 after 24 and 48 hours, respectively) and LDLR (P=0.005 and P=0.001 after 24 and 48 hours, respectively). ConclusionAccording to the results of the present study, HEFS, diosgenin and 4-OH-Ile up or down-regulate the expression of some predominant genes involved in lipid metabolism pathway, similar to that observed for orlistat. These types of regulatory effects are presumably proper for the treatment of obesity and overweight.

The benign c.344G > A: p.(Arg115His) variant in the LDLR gene interpreted from a pedigree-based genetic analysis of familial hypercholesterolemia

BackgroundWe previously identified the c.344G > A: p.(Arg115His) variant in the low-density lipoprotein receptor (LDLR) gene, which was interpreted as “conflicting interpretations of pathogenicity” in ClinVar, based on a genetic analysis of patients with familial hypercholesterolemia (FH). However, whether this variant affects the pathophysiology of FH remains unclear. Therefore, our aim was to annotate the c.344G > A: p.(Arg115His) variant in the LDLR gene in FH. We present 2 families harboring the c.344G > A: p.(Arg115His) variant in the LDLR gene.MethodsGenetic analyses were performed for the coding regions and the exon-intron boundary sequence of the LDLR and proprotein convertase subtilisin/kexin type 9 (PCSK9) genes in 2 FH families. Next, the family without pathogenic variants in the LDLR and PCSK9 genes was screened by whole-exome sequencing. Detailed clinical and biochemical data were gathered from family members.ResultsIn one family, the index case had biallelic c.1567G > A: p.(Val523Met) and c.344G > A: p.(Arg115His) variants in the LDLR gene, while the sibling had only the c.1567G > A: p.(Val523Met) variant in the LDLR gene. There was no difference in the FH phenotype between the siblings. In another family, the index case and the sibling had no pathogenic variants in the LDLR, PCSK9, and apolipoprotein B (APOB) genes, but the sibling’s wife with nonFH had the c.344G > A: p.(Arg115His) variant in the LDLR gene. The sibling and his wife had 4 children, including an unaffected child and an affected child who had the c.344G > A: p.(Arg115His) variant in the LDLR gene. In addition, the allele frequency of the c.344G > A: p.(Arg115His) variant (0.0023–0.0043) in Japanese and East Asian populations is relatively high compared with that of the other LDLR pathogenic variants (0.0001–0.0008).ConclusionsThe c.344G > A: p.(Arg115His) variant in the LDLR gene is interpreted as benign in individuals with FH.

A Study to Explore the LDLR Gene Polymorphisms Contribute to Atorvastatin Response in a Sample of Iraqi Population with Atherosclerotic Coronary Artery Disease

Genetic factors will determine the higher variability, which found in response to lipid reduction treatment (statins). However, due to ethnicity the frequency and effect of single nucleotide polymorphisms (SNPs) may differ. The main aim of lipid lowering medical treatment is to eventually prevent the endogenous production of cholesterol through inhibition of HMG-CoA reductase, The resulting decrease in hepatocyte cholesterol concentration triggers up-regulation of low-density lipoprotein (LDL)-receptor expression by inducing sterol regulatory element-binding protein (SREBP) 2 cleavage, as SREBP-2 activates LDL receptor transcription. The aim of this study was to evaluate the effects of low density lipoprotein receptor (LDL-R) gene variants (rs200727689; rs72658860) in response to atorvastatin treatment in atherosclerotic coronary artery disease (ACAD) in a sample of Iraqi patients. The genetic polymorphisms of rs200727689 and rs72658860 were studied in patients undergoing coronary angiography (CAG). One hundred Iraqi patients include 52 patients treated with 20mg/day and 48 patients treated with 40mg/day, In addition, 100 apparently healthy subjects were genotype for these SNP by the thermal profile of allelic discrimination methods used Real Time PCR (RT-PCR) assay. A significant increase in A allele frequency (rs200727689) compared with controls for total ACAD patients (43% vs 23.5%; OR= 2.46; 95% C.I: 1.60-3.77; p=0.000). For (rs72658860) SNP, among total patients the A allele also increased the frequency substantially compared with the controls (40.5% vs 23.5%; P=0.0003; OR= 2.22; 95% C.I: 1.44 -3.41). In the patients who were treated with atorvastatin, SNP rs72658860 AA genotype significantly affected the response of atorvastatin in dose 20mg compared with 40mg. The result showed in AA genotype of (rs200727689) SNP a reduction in TC concentrations (277.55±108.38 vs 326.99±63.56) and LDL-C concentrations (198.33±100.0 vs 250.01±62.52) in sera compared with GG genotype in patients treated with 20 mg atorvastatin, although, none in all parameters statistically significant differences.

A catalog of the pathogenic mutations of LDL receptor gene in Japanese familial hypercholesterolemia

Little data exist on the pathogenic mutations of LDL receptor in Japanese familial hypercholesterolemia (FH). We aimed to catalog the pathogenic mutations of LDL receptor gene in the 2 major Japanese FH-care centers (Kanazawa University and National Cerebral and Cardiovascular Center Research Institute), where genetic testing of FH has been performed centrally on requests from institutes all over Japan during more than past 2 decades. 796 FH subjects from 472 families who had nonsynonymous mutations in LDL receptor gene were included in this study. Genetic mutations were analyzed for mutations by Sanger sequencing as well as by multiplex ligation probe dependent amplification technique for large rearrangements. Pathogenic mutations were defined either as 1) protein truncated variants, 2) registered as pathogenic in ClinVar, or Human Gene Mutation Database (HGMD), or meet the criteria of American College of Medical Genetics and Genomics guideline, or 3) CADD score >10. We found 138 different mutations. Among them, 132 mutations were considered as pathogenic, including 19 large rearrangement mutations. However, 6 missense mutations were classified as variants of unknown significance. A single mutation accounted for as much as 41% of the FH subjects recruited from Kanazawa University mainly due to founder gene effect, whereas many singleton mutations were found from National Cerebral and Cardiovascular Center Research Institute located in Osaka. We provided the largest catalog of pathogenic mutations of LDL receptor gene in Japanese FH. This could aid to determine the pathogenicity of the LDL receptor genetic mutations not only in Japanese but also in other ethnicities.

Clinical and serum lipid profiles and LDLR genetic analysis of xanthelasma palpebrarum with nonfamilial hypercholesterolemia.

XP is a flat xanthoma that usually presents as bilateral, symmetrical, soft, yellowish papules over the eyelids. The etiology of XP is unknown, but it may be related to complex heterozygous mutations. To investigate the lipid profiles, important clinical characteristics, and low-density lipoprotein receptor (LDLR) gene mutation in the patients suffering from xanthelasma palpebrarum (XP) with nonfamilial hypercholesterolemia. The prospective study included 25 individuals of XP with nonfamilial hypercholesterolemia, and 30 controls neither with XP nor familial hypercholesterolemia (FH). Each one underwent detailed clinical examination, serum lipid profile, and LDLR gene detection at the 400th exon and the 1246th exon. In our study, patients with XP were often not present with FH and family history. The mean serum cholesterol (CHOL) (5.20±1.82) and the prevalence of carotid atherosclerosis (4.34±0.78) were significantly higher in study group, while there was no statistically significant difference between two groups in terms of triglyceride (TG) (P=.38) and low-density lipoprotein (LDL) (P=.23). Unusually, the mean levels of high-density lipoprotein (HDL) (1.59±0.31) in the study group were much higher than the controls (1.31±0.30), and the LDLR gene mutation was not found. Clinical and serum lipid profiles indicated that XP was sporadic and underlying lipid abnormalities especially higher HDL. XP did not occur with mutations in the LDLR gene at the two exons. We suggested too high HDL level may be contributed to pathogenesis of XP and XP is often not associated with FH.

Familial Hypercholesterolemia and Atherosclerosis: Animal Models and Therapeutic Advances

Familial hypercholesterolemia (FH), mainly arising from loss-of-function mutation of the low-density lipoprotein receptor (LDLR), is a life-threatening inherited cardiometabolic disorder with limited therapies. In a recent study, Zhao et al. created a new model of FH and demonstrate that LDLR gene editing protects against both FH and atherosclerosis.

Interaction between Sex and LDLR rs688 Polymorphism on Hyperlipidemia among Taiwan Biobank Adult Participants.

Hyperlipidemia is one of the strong risk factors for ischemic heart disease. Using the Taiwan Biobank (TWB) database, we evaluated the risk of hyperlipidemia and its interaction with sex and rs688 polymorphism on the low-density lipoprotein receptor (LDLR) gene. Data collection in the biobank started in 2008 and is ongoing. Data analysis was performed on the participants’ data collected between 2008 and 2015. In general, 27.92% of the 9237 female participants and 32.65% of the 8690 male participants were identified with hyperlipidemia. Compared to the C/C genotype, C/T and T/T genotypes were not significant risk factors for hyperlipidemia (OR = 1.061, CI: 0.976–1.153 for C/T and OR = 1.052, CI: 0.845–1.309 for T/T genotype) in the general model. However, there was a significant interaction between sex and rs6888 on hyperlipidemia risk (p-interaction = 0.0321). With the male sex/CC genotype being the reference group, only the female sex/CT and T/T genotypes were closely associated with hyperlipidemia, with respective ORs of 1.153 (CI: 1.014–1.311) and 1.423 (CI: 1.056–1.917). Our data indicate that rs688 C/T and T/T genotypes may be associated with increased risk of hyperlipidemia in Taiwanese women. These findings may be relevant in lipid-modification therapy.

ABCG5 and ABCG8 genetic variants in familial hypercholesterolemia

Familial hypercholesterolemia (FH) is a common inherited disease characterized by elevated low-density lipoprotein cholesterol (LDL-C) plasma levels and increased cardiovascular disease risk. Most patients carry a mutation in the low-density lipoprotein receptor gene (LDLR). Common and rare variants in the genes encoding adenosine triphosphate-binding cassette transporters G5 and G8 (ABCG5 and ABCG8) have been shown to affect LDL-C levels. The objective of this study was to investigate whether and to which extent heterozygous variants in ABCG5 and ABCG8 are associated with the hypercholesterolemic phenotype. We sequenced ABCG5 and ABCG8 in a cohort of 3031 clinical FH patients and compared the prevalence of variants with a European reference population (gnomAD). Clinical characteristics of carriers of putative pathogenic variants in ABCG5 and/or ABCG8 were compared with heterozygous carriers of mutations in LDLR. Furthermore, we assessed the segregation of one ABCG5 and two ABCG8 variants with plasma lipid and sterol levels in three kindreds. The frequencies of (likely) pathogenic LDLR, APOB, PCSK9, ABCG5, and ABCG8 variants in our FH cohort were 11.42%, 2.84%, 0.69%, 1.48%, and 0.96%, respectively. We identified 191 ABCG5 and ABCG8 variants of which 53 were classified as pathogenic or likely pathogenic. Of these 53 variants, 51 were either absent from a reference population or more prevalent in our FH cohort than in the reference population. LDL-C levels were significantly lower in heterozygous carriers of a (likely) pathogenic ABCG5 or ABCG8 variant compared to LDLR mutation carriers (6.2±1.7 vs 7.2±1.7mmol/L, P<.001). The combination of both an ABCG5 or ABCG8 variant and a LDLR variant was found not to be associated with significant higher LDL-C levels (7.8±2.3 vs 7.2±1.7mmol/L, P=.259). Segregation analysis in three families (nine carriers, in addition to the index cases, and 16 noncarriers) did not show complete segregation of the ABCG5/G8 variants with high LDL-C levels, and LDL-C levels were not different (3.9±1.3 vs 3.5±0.6mmol/L in carriers and noncarriers, respectively, P=.295), while plasma plant sterol levels were higher in carriers compared to noncarriers (cholestanol: 10.2±1.7 vs 8.4±1.6μmol/L, P=.007; campesterol: 22.5±10.1 vs 13.4±3.5μmol/L, P=.008; sitosterol: 17.0±11.6 vs 8.2±2.6μmol/L, P=.024). 2.4% of subjects in our FH cohort carried putative pathogenic ABCG5 and ABCG8 variants but had lower LDL-C levels compared to FH patients who were heterozygous carriers of an LDLR variant. These results suggest a role for these genes in hypercholesterolemia in FH patients with less severely elevated LDL-C levels. We did not find evidence that these variants cause autosomal dominant FH.

CRISPR/Cas9-Mediated Vitellogenin Receptor Knockout Leads to Functional Deficiency in the Reproductive Development of Plutella xylostella.

The vitellogenin receptor (VgR) belongs to the low-density lipoprotein receptor (LDLR) gene superfamily and plays an indispensable role in Vg transport, yolk deposition, and oocyte development. For this reason, it has become a promising target for pest control. The involvement of VgR in Vg transport and reproductive functions remains unclear in diamondback moths, Plutella xylostella (L.), a destructive pest of cruciferous crops. Here, we cloned and identified the complete cDNA sequence of P. xylostella VgR, which encoded 1805 amino acid residues and contained four conserved domains of LDLR superfamily. PxVgR was mainly expressed in female adults, more specifically in the ovary. PxVgR protein also showed the similar expression profile with the PxVgR transcript. CRISPR/Cas9-mediated PxVgR knockout created a homozygous mutant of P. xylostella with 5-bp-nucleotide deletion in the PxVgR. The expression deficiency of PxVgR protein was detected in the ovaries and eggs of mutant individuals. Vg protein was still detected in the eggs of the mutant individuals, but with a decreased expression level. However, PxVg transcripts were not significantly affected by the PxVgR knockout. Knockout of PxVgR resulted in shorter ovarioles of newly emerged females. No significant difference was detected between wild and mutant individuals in terms of the number of eggs laid in the first 3 days after mating. The loss of PxVgR gene resulted in smaller and whiter eggs and lower egg hatching rate. This study represents the first report on the functions of VgR in Vg transport, ovary development, oviposition, and embryonic development of P. xylostella using CRISPR/Cas9 technology. This study lays the foundation for understanding molecular mechanisms of P. xylostella reproduction, and for making use of VgR as a potential genetic-based molecular target for better control of the P. xylostella.

High Serum Levels of Cholesterol Increase Antitumor Functions of Nature Killer Cells and Reduce Growth of Liver Tumors in Mice

The relationship between serum cholesterol level and development of hepatocellular carcinoma (HCC) remains unclear. We investigated the effects of serum cholesterol level on development of liver tumors in mice. We performed studies with C57BL/6J mice, mice with disruption of the low-density lipoprotein receptor gene (Ldlr-/-mice), and mice with conditional deletion of nature killer (NK) cells (NKdele mice). Some C57BL/6J and NKdele mice were given injections of diethylinitrosamine to induce liver tumor formation. Mice were placed on a normal diet (ND) or high-cholesterol diet (HCD) to induce high serum levels of cholesterol. We also studied mice with homozygous disruption of ApoE (ApoE-/- mice), which spontaneously develop high serum cholesterol. C57BL/6J and NKdele mice on the ND or HCD were implanted with Hep1-6 (mouse hepatoma) cells and growth of xenograft tumors and lung metastases were monitored. Blood samples were collected from mice and analyzed by biochemistry and flow cytometry; liver and tumor tissues were collected and analyzed by histology, immunohistochemistry, and RNA-sequencing analysis. NK cells were isolated from mice and analyzed for cholesterol content, lipid raft formation, immune signaling, and changes in functions. We obtained matched tumor tissues and blood samples from 30 patients with HCC and blood samples from 40 healthy volunteers; levels of cholesterol and cytotoxicity of NK cells were measured. C57BL/6J mice on HCD and ApoE-/- mice with high serum levels of cholesterol developed fewer and smaller liver tumors and lung metastases after diethylinitrosamine injection or implantation of Hep1-6 cells than mice on ND. Liver tumors from HCD-fed mice and ApoE-/- mice had increased numbers of NK cells compared to tumors from ND-fed mice. NKdele mice or mice with antibody-based depletion for NK cells showed similar tumor number and size in ND and HCD groups after diethylinitrosamine injection or implantation of Hep1-6 cells. NK cells isolated from C57BL/6J mice fed with HCD had increased expression of NK cell-activating receptors (natural cytotoxicity triggering receptor 1 and natural killer group 2, member D), markers of effector function (granzyme B and perforin), and cytokines and chemokines compared with NK cells from mice on ND; these NK cells also had enhanced cytotoxic activity against mouse hepatoma cells, accumulated cholesterol, increased lipid raft formation, and immune signaling activation. NK cells isolated from HCD-fed Ldlr-/- mice did not have increased cholesterol content or cytotoxic activity against mouse hepatoma cells compared with ND-fed Ldlr-/- mice. Serum levels of cholesterol correlated with number and activity of NK cells isolated from human HCCs. Mice with increased serum levels of cholesterol due to an HCD or genetic disruption of ApoE develop fewer and smaller tumors after injection of hepatoma cells or a chemical carcinogen. We found cholesterol to accumulate in NK cells and activate their effector functions against hepatoma cells. Strategies to increase cholesterol uptake by NK cells can be developed for treatment of HCC.

Construction of a Novel Fatty Liver Cell Model with Low Density Lipoprotein Receptor Gene Knockout Based On Cas9/Single Guide RNA Technique

Clustered regularly interspaced short palindromic repeats/Cas9 gene editing technique was used to construct the human liver Lo2 stable cell line with low density lipoprotein receptor gene deletion. According to the design principle of clustered regularly interspaced short palindromic repeats/Cas9 target, the single guide RNA specifically target low density lipoprotein receptor gene was designed and the recombinant plasmid lenticlustered regularly interspaced short palindromic repeats v2 was used to construct the recombinant plasmid which can express this single guide RNA and Cas9 protein. After sequencing identification, the recombinant plasmid was transferred into Human embryonic kidney 293T cells together with retroviral packaging plasmids VSVG and PAX2. 48 h after transfection, the virus supernatant was collected and directly infected with human liver Lo2 cell. Puromycin was used to screen low density lipoprotein receptor-deficient Lo2 cells, which was verified by polymerase chain reaction, western blotting, and immunofluorescence staining. Then polymerase chain reaction and western blotting were used to verify the effects of low density lipoprotein receptor knockout on the function, lipid metabolism, glucose metabolism pathway, and insulin resistance of the Lo2 cell line. In this paper, the clustered regularly interspaced short palindromic repeats/Cas9 plasmid targeting low density lipoprotein receptor was successfully constructed. The results of polymerase chain reaction and western blotting showed that the human liver Lo2 cell with a stable knockout of low density lipoprotein receptor was obtained. The results of immunofluorescence staining showed that compared with the control group, the content of prealbumin in low density lipoprotein receptor knockout cells decreased significantly, while the content of diacylglycerol increased. At the same time, low density lipoprotein receptor gene knockout can increase the expression level of fatty acid synthase, peroxisome proliferator-activated receptor-γ, sterol regulatory element binding protein 1c, and inhibit the expression of carnitine palmitoyltransferase 1a. In addition, compared with the control group, there was no significant difference in mRNA expression and protein level of mammalian target of rapamycin and phosphatidylinositol-3-kinase, and there was no significant change in epsilon isoform of protein kinase C and insulin receptor substrates 1 expression level. The Lo2 cell line with low density lipoprotein receptor gene knockout was constructed by clustered regularly interspaced short palindromic repeats/Cas9 technique, and the role of low density lipoprotein receptor in lipid metabolism and glucose metabolism was proved by Polymerase chain reaction and Western blotting. The low density lipoprotein receptor knockout polyclonal and monoclonal cells constructed in this study can provide a useful tool for future research.

Identification of Key Biomarkers and Potential Molecular Mechanisms in Oral Squamous Cell Carcinoma by Bioinformatics Analysis.

The aim of this study was to explore the key genes, microRNA (miRNA), and the pathogenesis of oral squamous cell carcinoma (OSCC) at the molecular level through the analysis of bioinformatics, which could provide a theoretical basis for the screening of drug targets. Data of OSCC were obtained from Gene Expression Omnibus (GEO) database. The differentially expressed genes (DEGs) were identified via GEO2R analysis. Next, protein-protein interaction (PPI) network of DEGs was constructed through Search Tool for the Retrieval of Interacting Gene and visualized via Cytoscape, whereas the hub genes were screened out with Cytoscape. Gene ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were performed by Database for Annotation, Visualization and Integrated Discovery. The miRNA, which might regulate hub genes, were screened out with TargetScan and GO and KEGG analysis of miRNA was performed by DNA Intelligent Analysis-miRPath. Survival analyses of DEGs were conducted via the Kaplan-Meier plotter. Finally, the relationships between gene products and tumors were analyzed by Comparative Toxicogenomics Database. A total of 121 differential genes were identified. One hundred thirty-five GO terms and 56 pathways were obtained, which were mainly related to PI3K-Akt signals pathway, FoxO signaling pathway, Wnt signaling pathway, cell cycle, p53 signaling pathway, cellular senescence, and other pathways; 10 genes were identified as hub genes through modules analyses in the PPI network. Finally, a survival analysis of 10 hub genes was conducted, which showed that the low expression of matrix metalloproteinase (MMP)1, MMP3, and C-X-C motif chemokine ligand (CXCL)1 and the high expression of CXCL9 and CXCL10 resulted in a significantly poor 5-year overall survival rate in patients with OSCC. In this study, the DEGs of OSCC was analyzed, which assists us in a systematic understanding of the pathogenicity underlying occurrence and development of OSCC. The MMP1, MMP3, CXCL1, CXCL9, and CXCL10 genes might be used as potential targets to improve diagnosis and as immunotherapy biomarkers for OSCC.

Preparation of a Functional Rat LDL Receptor Minigene

The majority of the low-density lipoprotein (LDL) receptors present in the body are expressed in the liver. Therefore, plasma LDL levels significantly correlate with changes in the activity of the hepatic LDL receptor. Based on this, there is a need to understand the regulatory mechanisms that control the hepatic expression of the low-density lipoprotein (LDL) receptor. Herein, we have prepared a functional rat LDL receptor minigene construct that can produce mRNA after splicing. Sequence analysis suggests that this construct has the potential to code for a truncated version of LDL receptor protein. This minigene could be used as a research tool to identify small molecules, natural products, and regulators of the LDL receptor gene that could be developed into LDL receptor-specific activators for therapeutic use.

UNIVERSAL THEORY OF ATHEROSCLEROSIS PATHOGENESIS

The paper shows that atherosclerosis results from disturbed cholesterol homeostasis in the body, the development of systemic stromal-vascular lipid-protein dystrophy (or lipidosis, or cholesterolosis) complicated by extracellular focal lipid deposits with a predominant cholesterol in the interstitial tissue of the inner arterial and aortic layer. These deposits are foreign to this body tissue. They induce the development of chronic productive granulomatous inflammation in it – granulomatosis around endogenous foreign bodies. Cholesterol deposition is promoted by a positive cholesterol balance in the body; increased permeability of the endothelium in hemodynamically vulnerable parts of the arteries, where blood components, including LP, infiltrate their wall; a lack of hydrolytic enzymes in the cell lysosomes that could destroy the steroid nucleus of the cholesterol molecule. Being essential for each specific organism (for building membranes, the formation of bile acids, the synthesis of steroid hormones, vitamin D3), cholesterol, if exceeded, can increase the probability of developing atherosclerosis. Genetic mechanisms are implicated in the disturbed lipid protein metabolism in the human body. Hyperlipoproteinemias (HLPs) are known to be the most common metabolic disorders. The dystrophy under consideration results from primary HLP types IIa, IIb, III, IV, V, as well as secondary HLPs in patients with various medical conditions associated with an increase in blood LDL and / or VLDL and/or a decrease in HDL. A reduced cholesterol efflux from peripheral tissues due to a decreased HDL content and the development of lipidosis are seen in diabetes, obesity, physical inactivity, stress, puberty, menopause, hypertriglyceridemia, cigarette smoking, uremia, treatment with anabolic steroids, beta-adrenergic blocking agents, gestagens, and the use of contraceptives. The most pronounced manifestations of dystrophy are characteristic of HLP types IIA, IIb, III, but its moderate development complicated by atherosclerosis also occurs in types IV and V, which are accompanied by increased blood VLDL. Mutations of LDL receptor genes, apoprotein genes lead to the development of stromal vascular dystrophy and atherosclerosis. A number of rare genetic disorders of sterol metabolism accompany impaired metabolism of cholesterol and its esters: hepatic lipase deficiency (with accumulation of VLDL and IDLs); a deficiency in lysosomal hydrolase of cholesterol esters with impaired LDL metabolism (Wolman disease); cholesterol ester accumulation disease; cerebrotendinous xanthomatosis; cerebral cholesterolosis; Toichlander and Hand Schüler Christian syndromes. The main factor contributing to the development of inflammation in the inner vascular wall of arteries and aorta in atherosclerosis is the cyclic hydrocarbonic structure of cholesterol, which cannot be cleaved in the lysosomes of MPs. The leading role of the cyclic hydrocarbonic structure of cholesterol, which is insoluble and indestructible by MPs, in the induction of atherosclerosis-related inflammation is confirmed by the fact of the atherogenic action of cholesterol derivatives having its structure. An important factor in inflammatory morphogenesis is the lipoprotein dyscolloidosis occurring in the arterial intima and the physical and chemical metamorphosis of cholesterol. A colloidal solution, solid crystals of free cholesterol and liquid crystals of cholesterol esters have a pronounced phlogogenic and sclerogenic effect on the interstitial tissue of the arterial intima.

Familial hypercholesterolemia: A complex genetic disease with variable phenotypes.

Familial hypercholesterolemia (FH) is the most frequent genetic disease and is characterized by elevation of LDL-cholesterol that accumulates in tissues leading to premature atherosclerosis and sometime tendon xanthomas. Main causes of FH are pathogenic variants in the genes encoding the LDL receptor (LDLR), its ligand - the apolipoprotein B (APOB) - or Proprotein Convertase Subtilisin/Kexin Type 9 (PCSK9). Rarer causes include variants in genes encoding apolipoprotein E (APOE) and the signal-transducing adaptor family member 1 (STAP1). Genetics of FH is extremely complicated by 1. high heterogeneity, 2. presence of variant clusters and 3. phenotypic variability. In fact, a great variability was observed among patients with the same genetic status: an overlap of LDL-cholesterol levels was observed between heterozygous patients (HeFH) and homozygous FH patients, as well as some HeFH showed a normal lipid profile. A correct pathogenicity evaluation is the first step to correctly define the genetic status helping to identify the variants which really cause the FH. Several phenotypic differences were observed among HeFH patients carrying different variant types (null or defective) or variants in different affected genes. Patients with a null variant in LDLR gene showed higher LDL-cholesterol levels and higher risk for coronary artery disease than patients with a defective variant. Pathogenic variants in several lipid-related genes causing different dyslipidemias were found among FH patients acting as both modifying factors (worsening the phenotype) and confounding factors (needing a differential diagnosis to be discriminated from FH). This review aims at depicting the complex genetic basis of FH.

DNA methylation at LRP1 gene locus mediates the association between maternal total cholesterol changes in pregnancy and cord blood leptin levels.

Placental lipids transfer is essential for optimal fetal development, and alterations of these mechanisms could lead to a higher risk of adverse birth outcomes. Low-density lipoprotein receptor (LDLR), LDL receptor-related protein 1 (LRP1), and scavenger receptor class B type 1 (SCARB1) genes are encoding lipoprotein receptors expressed in the placenta where they participate in cholesterol exchange from maternal to fetal circulation. The aim of this study was thus to investigate the association between maternal lipid changes occurring in pregnancy, placental DNA methylation (DNAm) variations at LDLR, LRP1, and SCARB1 gene loci, and newborn's anthropometric profile at birth. Sixty-nine normoglycemic women were followed from the first trimester of pregnancy until delivery. Placental DNAm was quantified at 43 Cytosine-phosphate-Guanines (CpGs) at LDLR, LRP1, and SCARB1 gene loci using pyrosequencing: 4 CpGs were retained for further analysis. Maternal clinical data were collected at each trimester of pregnancy. Newborns' data were collected from medical records. Statistical models included minimally newborn sex and gestational and maternal age. Maternal total cholesterol changes during pregnancy (ΔT3-T1) were correlated with DNAm variations at LDLR (r = -0.32, p = 0.01) and LRP1 (r = 0.34, p = 0.007). DNAm at these loci was also correlated with newborns' cord blood triglyceride and leptin levels. Mediation analysis supports a causal relationship between maternal cholesterol changes, DNAm levels at LRP1 locus, and cord blood leptin concentration (pmediation = 0.02). These results suggest that LRP1 DNAm link maternal blood cholesterol changes in pregnancy and offspring adiposity at birth, which provide support for a better follow-up of blood lipids in pregnancy.

Molecular Dynamics Simulation Reveals Exposed Residues in the Ligand-Binding Domain of the Low-Density Lipoprotein Receptor that Interacts with Vesicular Stomatitis Virus-G Envelope.

Familial hypercholesterolemia (FH) is an autosomal dominant disease most often caused by mutations in the low-density lipoprotein receptor (LDLR) gene, which consists of 18 exons spanning 45 kb and codes for a precursor protein of 860 amino acids. Mutations in the LDLR gene lead to a reduced hepatic clearance of LDL as well as a high risk of coronary artery disease (CAD) and sudden cardiac death (SCD). Recently, LDLR transgenes have generated interest as potential therapeutic agents. However, LDLR packaging using a lentiviral vector (LVV) system pseudotyped with a vesicular stomatitis virus (VSV)-G envelope is not efficient. In this study, we modified the LVV system to improve transduction efficiency and investigated the LDLR regions responsible for transduction inhibition. Transduction efficiency of 293T cells with a 5′-LDLReGFP-3′ fusion construct was only 1.55% compared to 42.32% for the eGFP construct. Moreover, co-expression of LDLR affected eGFP packaging. To determine the specific region of the LDLR protein responsible for packaging inhibition, we designed constructs with mutations or sequential deletions at the 3′ and 5′ ends of LDLR cDNA. All constructs except one without the ligand-binding domain (LBD) (pWoLBD–eGFP) resulted in low transduction efficiency, despite successful packaging of viral RNA in the VSV envelope, as confirmed through RT-PCR. When we evaluated a direct interaction between LDLR and the VSV envelope glycoprotein using MD simulation and protein–protein interactions, we uncovered Val119, Thr120, Thr67, and Thr118 as exposed residues in the LDLR receptor that interact with the VSV protein. Together, our results suggest that the LBD of LDLR interacts with the VSV-G protein during viral packaging, which significantly reduces transduction efficiency.

Cascade screening for familial hypercholesterolemia-identification of the C308Y mutation in multiple family members and relatives for the first time in mainland China

BackgroundFamilial hypercholesterolemia (FH), an autosomal dominant genetic disorder, is underdiagnosed and undertreated. The majority of FH cases are caused by low density lipoprotein receptor (LDL-R) gene mutations. The C308Y mutation in LDL-R results in approximately 70% loss of LDL-R activity, leading to the elevation of low density lipoprotein-cholesterol (LDL-C) and an increased risk of premature coronary heart disease (CHD). The aim of this study was to identify FH cases by cascade screening in family members and relatives of a 37-year old male with premature CHD and hypercholesterolemia.MethodsClinical exam, blood lipid profiling and genomic DNA sequencing of all exons of LDL-R were performed for the proband and his 14 family members and relatives. FH diagnosis was carried out using the Dutch Lipid Clinic Network (DLCN) criteria.ResultsLipid profiling showed that 9 individuals, including the proband, had hypercholesterolemia. All these 9 subjects had a G > A substitution at nucleotide 986 in exon 7 resulting in the C308Y mutation as determined by DNA sequencing, and all those carrying the mutation were diagnosed as having definite FH under the DLCN criteria. However, most (7/9) did not have suggestive clinical manifestations of CHD.ConclusionsThe C308Y mutation was discovered in multiple family members and relatives for the first time in mainland China. Cascade screening is key for the confirmatory diagnosis of FH. Our hypothesis that the C308Y is a common variant in the population of Southern China origin warrants further validation by screening for the C308Y mutation in a large population.

Molecular Features and Expression Patterns of Vitellogenin Receptor in Calliptamus italicus (Orthoptera: Acrididae).

Vitellogenin receptor (VgR) mediates the intake of vitellin via oocytes, thus exerting an important role in vitellogenesis. In this study, reverse transcription-polymerase chain reaction (RT-PCR) and rapid-amplification of cDNA ends techniques were adopted to clone the CiVgR gene, namely the VgR gene of Calliptamus italicus, i.e., Orthopteran. The full length of CiVgR was 5,589 bp, and the open reading frame was estimated to be 5,265 bp, which encoded 1,754 amino acids (aa). Sequence alignment analysis showed that CiVgR belonged to the superfamily of low-density lipoprotein receptor genes, which contained several conserved domains, including ligand-binding domains, epidermal growth factor precursor homology domains, transmembrane domains, and cytoplasmic domains. However, no O-linked sugar domain was identified. Phylogenetic analysis showed that CiVgR had the closest genetic relationship to Blattarias. RT-PCR showed that CiVgR was only specifically expressed in the ovarian tissue of females. quantitative real time polymerase chain reaction showed that the transcription of CiVgR already appeared in the fourth-instar nymph of C. italicus, which gradually increased after adult emergence, peaked at the previtellogenesis stage, and then started to decrease. The expression pattern of CiVgR was closely associated with vitellogenesis. The findings of this study further our understanding of the molecular mechanisms involved in the reproduction of C. italicus, and provide new ideas to control this insect.

The atherogenic role of immune cells in familial hypercholesterolemia.

Familial hypercholesterolemia (FH) is an autosomal dominant disorder of lipoprotein metabolism that mainly occurs due to mutations in the low-density lipoprotein receptor gene and is characterized by increased levels of low-density lipoprotein cholesterol, leading to accelerated atherogenesis and premature coronary heart disease. Both innate and adaptive immune responses, which mainly include monocytes, macrophages, neutrophils, T lymphocytes, and B lymphocytes, have been shown to play a key role for the initiation and progression of atherogenesis in the general population. In FH patients, these immune cells have been suggested to play specific pro-atherosclerotic activities, from the initial leukocyte recruitment to plaque rupture. In fact, the accumulation of cholesterol crystals and oxLDL in the vessels in FH patients is particularly high, with consequent abnormal mobilization of immune cells and secretion of various pro-inflammatory and chemokines. In addition, cholesterol accumulation in immune cells is exaggerated with chronic exposure to relevant pro-atherosclerotic triggers. The topics considered in this review may provide a more specific focus on the immune system alterations in FH and open new insights toward immune cells as potential therapeutic targets in FH.