Absence Of Low Density Lipoprotein Receptor Research Articles

The Expression of the Low-Density Lipoprotein Receptor Modulates the Effects of Hypogonadism on Mitochondrial Activity in Different Brain Domains

Background: Testosterone deficiency (TD), brain mitochondrial uncoupling protein 2 (UCP2), and the low-density lipoprotein receptor (LDLR) are independently linked to the triggering of several behavioral disorders. Here, we investigated how TD impacts mitochondrial activity and apoptosis in different brain domains in the presence and absence of LDLR. Materials and Methods: Sham-operated or surgically castrated C57BL/6 or LDLR-deficient (Ldlr−/−) mice were fed a western-type diet for 12 weeks. Then, mitochondrial UCP2 and cytochrome c (Cytc) relative protein levels were measured as markers of uncoupling and oxidative phosphorylation, respectively, in tissue specimens from different brain domains. The ratio of the relative cytoplasmic Cytc levels over the relative mitochondrial Cytc levels (CM ratio) was used as a biochemical marker of Cytc-dependent apoptosis. Results: In cerebral hemispheres and the cerebellum area of both C57BL/6 and Ldlr−/− mice, castration resulted in increased UCP2, suggesting protection from hypogonadism-induced oxidative stress. In the broader midbrain, castration reduced UCP2 expression in C57BL/6 mice but not in Ldlr−/− mice, suggesting that LDLR deficiency protects from oxidative stress in this region. CM ratio was ≤1 after castration in these three brain domains, suggesting increased protection from Cytc-dependent apoptosis. In diencephalon, castration had no effect on UCP2 and CM ratio in C57BL/6 mice but led to increased UCP2 expression and increased CM ratio in Ldlr−/− mice, suggesting an increase in apoptosis in the diencephalon of castrated Ldlr−/− mice. Conclusions: TD in Ldlr−/− mice fed a western-type diet is associated with protection from oxidative stress but could contribute to the emergence of behavioral or systemic impairments linked to apoptosis in the diencephalon.

Role of Lipoprotein Apheresis in the Management of Familial Hypercholesterolemia

Familial Hypercholesterolemia (FH) is an autosomal dominant disorder due to a congenital absence of low-density lipoprotein receptor (LDL-R) that transports cholesterol-carrying lipoprotein particles into cells. Absence of these receptors results in increased levels Non-HDL lipoproteins (like LDL, VLDL, Triglycerides, Lipoprotein a) which predisposes to premature atherosclerosis and cardiac complications. Treatment modalities for this condition include use of high intensity statins and diet control. However some of these patients with familial hypercholesterolemia do not respond to diet and drug therapy. Here Lipoprotein apheresis is believed to be a feasible and effective option. Lipoprotein apheresis is a relatively new technique in India used to treat familial hypercholesterolemia. Lipoprotein apheresis using cascade filter is effective in reducing non-HDL lipoprotein levels by 60-75% in a single procedure. However, the disadvantage of the procedure it is expensive and needs to be repeated every 2 weeks for lifetime. This case is being reported for rarity of this condition (1 case per million) and more so the rarity of the use of Lipoprotein apheresis procedure in our country.

3,5,3'-Triiodo-L-Thyronine- and 3,5-Diiodo-L-Thyronine- Affected Metabolic Pathways in Liver of LDL Receptor Deficient Mice.

3,5,3′-triiodo-L-thyronine (T3) and 3,5-diiodo-L-thyronine (T2), when administered to a model of familial hypercholesterolemia, i.e., low density lipoprotein receptor (LDLr)-knockout (Ldlr−/−) mice fed with a Western type diet (WTD), dramatically reduce circulating total and very low-density lipoprotein/LDL cholesterol with decreased liver apolipoprotein B (ApoB) production. The aim of the study was to highlight putative molecular mechanisms to manage cholesterol levels in the absence of LDLr. A comprehensive comparative profiling of changes in expression of soluble proteins in livers from Ldlr−/− mice treated with either T3 or T2 was performed. From a total proteome of 450 liver proteins, 25 identified proteins were affected by both T2 and T3, 18 only by T3 and 9 only by T2. Using in silico analyses, an overlap was observed with 11/14 pathways common to both iodothyronines, with T2 and T3 preferentially altering sub-networks centered around hepatocyte nuclear factor 4 α (HNF4α) and peroxisome proliferator-activated receptor α (PPARα), respectively. Both T2 and T3 administration significantly reduced nuclear HNF4α protein content, while T2, but not T3, decreased the expression levels of the HNFα transcriptional coactivator PGC-1α. Lower PPARα levels were found only following T3 treatment while both T3 and T2 lowered liver X receptor α (LXRα) nuclear content. Overall, this study, although it was not meant to investigate the use of T2 and T3 as a therapeutic agent, provides novel insights into the regulation of hepatic metabolic pathways involved in T3- and T2-driven cholesterol reduction in Ldlr−/− mice.

Human PCSK9 promotes hepatic lipogenesis and atherosclerosis development via apoE- and LDLR-mediated mechanisms.

Proprotein convertase subtilisin/kexin type 9 (PCSK9) promotes the degradation of hepatic low-density lipoprotein (LDL) receptors (LDLR), thereby, decreasing hepatocyte LDL-cholesterol (LDL-C) uptake. However, it is unknown whether PCSK9 has effects on atherogenesis that are independent of lipid changes. The present study investigated the effect of human (h) PCSK9 on plasma lipids, hepatic lipogenesis, and atherosclerotic lesion size and composition in transgenic mice expressing hPCSK9 (hPCSK9tg) on wild-type (WT), LDLR⁻/⁻, or apoE⁻/⁻ background. hPCSK9 expression significantly increased plasma cholesterol (+91%), triglycerides (+18%), and apoB (+57%) levels only in WT mice. The increase in plasma lipids was a consequence of both decreased hepatic LDLR and increased hepatic lipid production, mediated transcriptionally and post-transcriptionally by PCSK9 and dependent on both LDLR and apoE. Despite the lack of changes in plasma lipids in mice expressing hPCSK9 and lacking LDLR (the main target for PCSK9) or apoE (a canonical ligand for the LDLR), hPCSK9 expression increased aortic lesion size in the absence of apoE (268 655 ± 97 972 µm² in hPCSK9tg/apoE⁻/⁻ vs. 189 423 ± 65 700 µm(2) in apoE⁻/⁻) but not in the absence of LDLR. Additionally, hPCSK9 accumulated in the atheroma and increased lesion Ly6C(hi) monocytes (by 21%) in apoE⁻/⁻ mice, but not in LDLR⁻/⁻ mice. PCSK9 increases hepatic lipid and lipoprotein production via apoE- and LDLR-dependent mechanisms. However, hPCSK9 also accumulate in the artery wall and directly affects atherosclerosis lesion size and composition independently of such plasma lipid and lipoprotein changes. These effects of hPCSK9 are dependent on LDLR but are independent of apoE.

Host Cholesterol Synthesis Contributes to Growth of Intracellular Toxoplasma gondii in Macrophages

The intracellular protozoan Toxoplasma gondii lacks the ability to synthesize sterol and scavenges cholesterol from the low-density lipoprotein receptor (LDLR) pathway of its host to facilitate replication. Sterol biosynthesis inhibitors, however, have a demonstrated anti-Toxoplasma effect. In this study, we examined the host mevalonate pathway as a novel source of cholesterol for T. gondii and its effects on parasite growth in macrophages. Parasite growth did not significantly change in the absence of LDLR or when LDL was exogenously supplemented. Lovastatin and compactin, both inhibitors of hydroxymethylglutaryl-CoA (HMG-CoA) reductase in the mevalonate pathway, significantly inhibited T. gondii growth in both wild-type and LDLR-knockout macrophages. Parasite growth was also suppressed by squalestatin, an inhibitor of squalene synthase, despite mevalonate producing isoprenoid intermediates in host cells. The present study demonstrates that lovastatin, compactin and squalestatin have anti-Toxoplasma activities and that the host cholesterol synthesis may contribute to parasite growth in macrophages.

Plasma PCSK9 preferentially reduces liver LDL receptors in mice

Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a secreted protein that regulates the expression of LDL receptor (LDLR) protein. Gain-of-function mutations in PCSK9 cause hypercholesterolemia, and loss-of-function mutations result in lower plasma LDL-cholesterol. Here, we investigate the kinetics and metabolism of circulating PCSK9 relative to tissue levels of LDLRs. The administration of recombinant human PCSK9 (32 microg) to mice by a single injection reduced hepatic LDLRs by approximately 90% within 60 min, and the receptor levels returned to normal within 6 h. The half-life of the PCSK9 was estimated to be approximately 5 min. Continuous infusion of PCSK9 (32 microg/h) into wild-type mice caused a approximately 90% reduction in hepatic LDLRs within 2 h and no associated change in the level of LDLR in the adrenals. Parallel studies were performed using a catalytically inactive form of PCSK9, PCSK9(S386A), and similar results were obtained. Infusion of PCSK9(D374Y), a gain-of-function mutation, resulted in accelerated clearance of the mutant PCSK9 and a greater reduction in hepatic LDLRs. Combined, these data suggest that exogenously administrated PCSK9 in plasma preferentially reduces LDLR protein levels in liver at concentrations found in human plasma and that PCSK9's action on the LDLR is not dependent on catalytic activity in vivo.

The Proprotein Convertase PCSK9 Induces the Degradation of Low Density Lipoprotein Receptor (LDLR) and Its Closest Family Members VLDLR and ApoER2

The proprotein convertase PCSK9 gene is the third locus implicated in familial hypercholesterolemia, emphasizing its role in cardiovascular diseases. Loss of function mutations and gene disruption of PCSK9 resulted in a higher clearance of plasma low density lipoprotein cholesterol, likely due to a reduced degradation of the liver low density lipoprotein receptor (LDLR). In this study, we show that two of the closest family members to LDLR are also PCSK9 targets. These include the very low density lipoprotein receptor (VLDLR) and apolipoprotein E receptor 2 (ApoER2) implicated in neuronal development and lipid metabolism. Our results show that wild type PCSK9 and more so its natural gain of function mutant D374Y can efficiently degrade the LDLR, VLDLR, and ApoER2 either following cellular co-expression or re-internalization of secreted human PCSK9. Such PCSK9-induced degradation does not require its catalytic activity. Membrane-bound PCSK9 chimeras enhanced the intracellular targeting of PCSK9 to late endosomes/lysosomes and resulted in a much more efficient degradation of the three receptors. We also demonstrate that the activity of PCSK9 and its binding affinity on VLDLR and ApoER2 does not depend on the presence of LDLR. Finally, in situ hybridization show close localization of PCSK9 mRNA expression to that of VLDLR in mouse postnatal day 1 cerebellum. Thus, this study demonstrates a more general effect of PCSK9 on the degradation of the LDLR family that emphasizes its major role in cholesterol and lipid homeostasis as well as brain development.

Lipoprotein lipase-facilitated uptake of LDL is mediated by the LDL receptor

LPL mediates the uptake of lipoproteins into different cell types independent of its catalytic activity. The mechanism of this process and its physiological relevance are not clear. Taking into account the importance of the endothelial barrier for lipoprotein uptake, in vitro studies with primary aortic endothelial cells from wild-type and low density lipoprotein receptor (LDLR)-deficient (LDLR(-/-)) mice were performed. Addition of LPL almost doubled the uptake of LDL into wild-type cells. However, there was virtually no LPL-mediated change of LDL uptake into LDLR(-/-) cells. Upregulation of LDLR by lipoprotein-deficient serum/lovastatin in wild-type cells resulted in a 7-fold increase of LPL-mediated LDL uptake. Uptake of chylomicron remnants was not affected by LDLR expression. In proteoglycan-deficient cells, LPL did not increase the uptake of lipoproteins. The physiological relevance of this pathway was studied in mice that were both LDLR(-/-) and transgenic for catalytically inactive LPL in muscle. In the presence of LDLR, inactive LPL reduced LDL cholesterol significantly (13-24%). In the absence of LDLR, LDL cholesterol was not affected by transgenic LPL. Metabolic studies showed that in the presence of LDLR, LPL increased the muscular uptake of LDL by 77%. In the absence of LDLR, transgenic LPL did not augment LDL uptake. Chylomicron uptake was not affected by the LDLR genotype. We conclude that LPL-mediated cellular uptake of LDL, but not of chylomicrons, is dependent on the presence of both LDLR and proteoglycans.

Upregulation of liver VLDL receptor and FAT/CD36 expression in LDLR−/− apoB100/100 mice fed trans-10,cis-12 conjugated linoleic acid

This study explores the mechanisms responsible for the fatty liver setup in mice fed trans-10,cis-12 conjugated linoleic acid (t10c12 CLA), hypothesizing that an induction of low density lipoprotein receptor (LDLR) expression is associated with lipid accumulation. To this end, the effects of t10c12 CLA treatment on lipid parameters, serum lipoproteins, and expression of liver lipid receptors were measured in LDLR(-/-) apoB(100/100) mice as a model of human familial hypercholesterolemia itself depleted of LDLR. Mice were fed t10c12 CLA over 2 or 4 weeks. We first observed that the treatment induced liver steatosis, even in the absence of LDLR. Mice treated for 2 weeks exhibited hypertriglyceridemia with high levels of VLDL and HDL, whereas a 4 week treatment inversely induced a reduction of serum triglycerides (TGs), essentially through a decrease in VLDL levels. In the absence of LDLR, the mRNA levels of other proteins, such as VLDL receptor, lipoprotein lipase, and fatty acid translocase, usually not expressed in the liver, were upregulated, suggesting their involvement in the steatosis setup and lipoprotein clearance. The data also suggest that the TG-lowering effect induced by t10c12 CLA treatment was attributable to both the reduction of circulating free fatty acids in response to the severe lipoatrophy and the high capacity of liver to clear off plasma lipids.

Role of Low Density Lipoprotein Receptor in the Clearance of Coagulation Factor VIII In Vivo.

Low Density Lipoprotein Receptor (LDLR) is the archetype of the family of endocytic receptors that also includes Low-density lipoprotein Receptor-related Protein (LRP), Megalin, and Very Low Density Lipoprotein Receptor (VLDLR). While most of these receptors bind a variety of ligands, LDLR has restricted specifity. Its known ligands are apolipoprotein E (apoE) and apolipoprotein B100 (apoB100). Ligand binding to the LDL-receptor family is inhibited by the Receptor-Associated Protein (RAP). We have previously reported that RAP overexpression elevates plasma levels of factor VIII (FVIII) in both normal mice and mice with hepatic LRP deficiency [Bovenschen et al. (2003) Blood 101, 3933–3939]. This implies that LRP, but also another RAP-sensitive mechanism contributes to the regulation of FVIII in vivo. This study addresses the question whether LDLR, despite its restricted ligand specificity, binds FVIII and contributes to its clearance. In vitro binding was studied using a recombinant LDLR fragment spanning the extracellular domain of complement-type repeats 1–7. The purified fragment efficiently bound to immobilized ApoE and apoB-100 containing LDL. The immobilized LDLR fragment also bound human FVIII, with half-maximal binding at 156 nM FVIII, and binding was inhibited by RAP. Human von Willebrand Factor (VWF) or non-activated factor IX did not bind to the LDLR fragment. The relevance of the FVIII-LDLR interaction was assessed in vivo employing LDLR−/− mice, cre/loxP-mediated conditional LRP-deficient mice (LRP−), and mice with the combined deficiency. Plasma FVIII levels of controls, LDLR−/− and LRP− mice were 1.1, 0.9 and 1.8 U/ml, respectively. This suggests that LRP, but not LDLR regulates FVIII in plasma. Surprisingly, however, mice that combined LDLR deficiency with hepatic LRP deficiency displayed much higher FVIII levels (median value 4.6 U/ml) than mice lacking LRP alone. This suggests that LDLR does have the potential of regulating FVIII levels. LDLR−/− LRP− mice further displayed elevated levels of VWF (median value 3.3 U/ml), but not of factor V or factor IX. The possibility was considered that FVIII levels were elevated secondary to the profound changes in lipoprotein profiles. To this end, we also examined ApoE deficient mice, which have reduced LDL, and mice that overexpress ApoC1, which is associated with elevated levels of cholesterol- and triglyceride VLDL. ApoE−/− LDLR−/−LRP− mice had a median FVIII level of 4.2 U/ml, which is close to that of LDLR−/−LRP− mice. Mice that overexpressed human ApoC1 had elevated levels of cholesterol and triglycerides, but 0.5 U/ml FVIII. This demonstrates that elevated FVIII levels were independent of lipoprotein levels. The role of LDLR and LRP in FVIII clearance were further studied by analyzing the pharmacokinetics of human FVIII. In normal mice the Mean Residence Time (MRT) was 160 min [68% confidence intervals (CI) 117–218 min]. MRT was 200 [CI 154–259] min in LDLR−/− mice, and 263 [CI 206–336] min in LRP− mice. This confirms the previously described role of LRP in FVIII clearance. Strikingly, in LDLR−/−LRP− mice the MRT of FVIII was 760 [691–826] min, which is approximately 5-fold longer than in control mice. These data demonstrate that LRP and LDLR act in concert in regulating FVIII levels in plasma. In the absence of LDLR, LRP maintains normal FVIII levels, while hepatic LRP deficiency is largely compensated by LDLR. This regulatory role of LDLR represents a novel link between LDLR and the hemostatic system.

Multiple processes are involved in the uptake of chylomicron remnants by mouse peritoneal macrophages

The processes responsible for the uptake of chylomicron remnants by macrophages were investigated using freshly isolated cells from low density lipoprotein (LDL) receptor, very low density lipoprotein (VLDL) receptor and apolipoprotein E knockout mice. In peritoneal macrophages from normal mice, the metabolism of chylomicron remnants was inhibited 40% by anti-LDL receptor antibody and 60% by a high concentration of receptor-associated protein (RAP). Together they reduced the amount processed by 70%. Digestion of cell proteoglycans decreased remnant degradation by 20% while the addition of acetyl-LDL had no effect. When LDL receptors were absent, the absolute rates of metabolism were less than that of normal cells and were not inhibited by the anti-LDL receptor antibody; the rates, however, were reduced to less than half by RAP. These suggest that the LDL receptor-related protein (LRP) or another LDL receptor family member(s) contributes to chylomicron remnant uptake and becomes the major mechanism of uptake when LDL receptors are absent. In contrast, the VLDL receptor was not involved as its absence did not affect chylomicron remnant metabolism. Similarly, the absence of apoE production did not affect the amount of remnant uptake; however, the proportion that was sensitive to RAP was eliminated. The level of LRP expression was not altered in these cells whereas there was a decrease in LDL receptors. This suggests that the apoE content of chylomicron remnants is sufficient for its recognition by LDL receptors but additional apoE is required for its uptake by the LRP and that there is an up-regulation of a non-LDL receptor family mechanism in apoE deficiency. Together these studies suggest that even in the absence of LDL receptors or apoE secretion, chylomicron remnants could contribute to lipid accumulation in the artery wall during atherogenesis.