Low-density Lipoprotein Receptor Overexpression Research Articles

Overexpression of low-density lipoprotein receptor prevents neurotoxic polarization of astrocytes via inhibiting NLRP3 inflammasome activation in experimental ischemic stroke.

JOURNAL/nrgr/04.03/01300535-202502000-00027/figure1/v/2024-05-28T214302Z/r/image-tiff Neurotoxic astrocytes are a promising therapeutic target for the attenuation of cerebral ischemia/reperfusion injury. Low-density lipoprotein receptor, a classic cholesterol regulatory receptor, has been found to inhibit NLR family pyrin domain containing protein 3 (NLRP3) inflammasome activation in neurons following ischemic stroke and to suppress the activation of microglia and astrocytes in individuals with Alzheimer's disease. However, little is known about the effects of low-density lipoprotein receptor on astrocytic activation in ischemic stroke. To address this issue in the present study, we examined the mechanisms by which low-density lipoprotein receptor regulates astrocytic polarization in ischemic stroke models. First, we examined low-density lipoprotein receptor expression in astrocytes via immunofluorescence staining and western blotting analysis. We observed significant downregulation of low-density lipoprotein receptor following middle cerebral artery occlusion reperfusion and oxygen-glucose deprivation/reoxygenation. Second, we induced the astrocyte-specific overexpression of low-density lipoprotein receptor using astrocyte-specific adeno-associated virus. Low-density lipoprotein receptor overexpression in astrocytes improved neurological outcomes in middle cerebral artery occlusion mice and reversed neurotoxic astrocytes to create a neuroprotective phenotype. Finally, we found that the overexpression of low-density lipoprotein receptor inhibited NLRP3 inflammasome activation in oxygen-glucose deprivation/reoxygenation injured astrocytes and that the addition of nigericin, an NLRP3 agonist, restored the neurotoxic astrocyte phenotype. These findings suggest that low-density lipoprotein receptor could inhibit the NLRP3-meidiated neurotoxic polarization of astrocytes and that increasing low-density lipoprotein receptor in astrocytes might represent a novel strategy for treating cerebral ischemic stroke.

PAQR3 facilitates the ferroptosis of diffuse large B-cell lymphoma via the regulation of LDLR-mediated PI3K/AKT pathway.

Progesterone and adiponectin receptor 3 (PAQR3) has been found to regulate tumor progression by mediating cell ferroptosis. However, whether PAQR3 mediates ferroptosis in diffuse large B-cell lymphoma (DLBCL) needs further investigation. The mRNA and protein levels of PAQR3 and low-density lipoprotein receptor (LDLR) were assessed by qRT-PCR and WB assays. Cell proliferation was detected by MTT assay and EdU assay. Shrunken mitochondria was counted under transmission electron microscope. Cell ferroptosis was evaluated by measuring the levels of malondialdehyde, reactive oxygen species, glutathione, Fe2+ , and the protein expression of ferroptosis-related markers. PAQR3 and LDLR interaction was confirmed by RIP assay and pull-down assay. Our study showed that PAQR3 was underexpressed, while LDLR was overexpressed in DLBCL tissues and cells. Functionally, PAQR3 overexpression or LDLR knockdown restrained DLBCL cell proliferation and enhanced ferroptosis. Mechanistically, PAQR3 reduced LDLR expression by inhibiting its mRNA stability. Meanwhile, LDLR overexpression reversed PAQR3-mediated the promoting on DLBCL cell ferroptosis, and LY294002 (PI3K/AKT inhibitor) eliminated the inhibiting effects of LDLR overexpression on DLBCL cell ferroptosis. Additionally, excessive PAQR3 reduced DLBCL tumor growth by enhancing tumor cell ferroptosis through LDLR-mediated PI3K/AKT pathway. In conclusion, our data suggested that PAQR3 restrained DLBCL progression by aggravating ferroptosis, which was achieved by inhibiting LDLR expression to repress PI3K/AKT pathway.

Abstract LB066: Targeting low density lipoprotein receptors in the pancreatic adenocarcinoma

Abstract Introduction: Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive malignancy with a poor 5-year overall survival rate at all stages of disease (7-10%). The immunohistochemistry of pancreatic cancer tissue has shown a significantly higher level of positive LDLR (low-density lipoprotein receptor) staining in primary tumors compared to the normal adjacent pancreas. LDLR is overexpressed in the epithelial pancreatic adenocarcinoma, irrespectively of tumor size, stage, and aggressiveness. The objective of this study was to determine the LDLR-targeting properties of 68Ga-labeled peptide conjugate, 68Ga-RMX-VH, in pancreatic adenocarcinoma cell lines. Methods: RMX-VH peptide conjugate (25-50μg) was labeled with 68Ga (50-100mCi, ITM Germany) in mild conditions in the presence of scavenger, sodium ascorbate (8mg/ml). The cellular uptake and competition studies of the radiotracer (15μCi/well) were completed in multiple pancreatic adenocarcinoma cell lines and in vivo in PDAC xenografts generated in athymic nude mice. The PET/CT images were acquired using a G4 PET/X-ray camera (Sofie Biosciences; 10min/scan) at 1h, 2h post-injection. This study determined in vivo and in vitro time-dependent accumulation of this agent in LDLR-overexpressing pancreatic adenocarcinoma cells. Results: 68Ga-RMX-VH was synthesized with higher than 95% radiochemical purity. The radiotracer was incubated in pancreatic adenocarcinoma cell lines for 1h and has shown the highest uptake in HPAF-II (6.7%ID/mg) and BxPC-3 (5.3 %ID/mg) and MiaPaCa2 (4.75 %ID/mg), respectively. Western blot showed a variable level of LDLR overexpression in human pancreatic cancer cells. The microPET imaging studies confirmed rapid accumulation and retention of radiotracer in the tumor (1.18 %ID/g) and elimination through kidneys (4.5 %ID/g) and bladder (SUV ratio of tumor to kidneys: 0.21). These results correlate with previously reported retention of the agent in Pk4a pancreatic adenocarcinoma. Conclusions: RMX-VH has the potential to evaluate the role of LDL receptors in pancreatic adenocarcinoma. We have initiated the exploratory clinical study of this agent in LDLR-overexpressing solid tumors, including pancreatic adenocarcinoma. Citation Format: Leo Flores, Xuewei Qu, Cedric Malicet, Pascaline Lecorche, Ebrahim Delpassand, Jamal Temsamani, Izabela Tworowska. Targeting low density lipoprotein receptors in the pancreatic adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr LB066.

Abstract 11197: Nano-Triciribine Reduces SARS-CoV-2 Infection by Sequestering ACE2 and the Novel Host Factor LDLR

Introduction: People with previous CVD hospitalized for COVID-19 have elevated death rate. We reported that patients with diabetes and HF higher protein levels of the low density lipoprotein receptor (LDLR). We hypothesized that LDLR is a novel host factor for the SARS-CoV-2-Spike (S2S) protein that may be regulated by the Akt inhibitor Triciribine (TCN), a drug being tested in Phase III studies for breast cancer. We also hypothesized that nano-formulation of Triciribine (NanoTriciribine; NTCN) would enhance its efficacy and allow for intranasal delivery. Methods: Interactions between the recombinant proteins Spike-RBD (receptor binding domain), ACE2, LDLR and its ectodomains (EGFA-EFFB, C2-C5 and C2) were analyzed by binding assays and co-IP in HepG2, HK2, and 293T cells. Viral entry assays were performed with 2 S2S pseudoviruses using 293T cells + hACE2 and TMPRSS2 or Furin protease. The effect of NTCN or the LXR agonist GW-3965 on viral uptake (pseudotyped VSVΔG-GFP*S2S or chimera VSV-S2S-eGFP virus) was assessed. Akt, pAkt, ACE2, and LDLR levels were determined in 293T+hACE2 by flow cytometry. Assays were done in triplicates and 1-way-ANOVA with Tukey’s correction was used for statistics. Results: RBD protein binds modestly to the human LDLR (EC 50 :10μM) and its C2-C5 ectodomain (EC 50 :13.8μM). Co-IP revealed a novel and strong LDLR-ACE2 interaction in several human cell lines. LDLR overexpression in human cells increased the uptake of VSVΔG-GFP*S2S (FC=2.32;p<0.001) and chimera virus (FC=.33; p<.0001). NTCN and TCN each reduced pAkt/Akt ratio. 1μM TCN or NTCN reduced LDLR (7.2%;p<.01 & 15.6%;p<0.0001) and ACE2 (32%;p<0.05 & 44.7%;p<.01) cell surface expression, respectively. 1μM NTCN or GW-3965 reduced S2S viral entry by 64.2% (p<.0001) and 40.7% (p<.01), respectively, confirming a role for LDLR in S2S infection. In hACE2tg mice, chimera VSV-S2S caused significant lung infection as measured by qPCR, GFP expression in proximal and distal lung airway epithelial cells, and electron microscopy. Intranasal delivery of NTCN was well tolerated. Conclusions: LDLR enhanced S2S viral entry supporting the elevated COVID-19 susceptibility seen in patients with heart disease. NTCN is a promising candidate for prophylactic treatment against COVID-19.

Abstract P420: Nano-Triciribine Reduces SARS2-CoV-2 Infection By Sequestering ACE2 And The Novel Host Factor LDLR

Background: People with previous CVD hospitalized for COVID-19 have elevated death rate. We reported that patients with diabetes and HF higher protein levels of the low density lipoprotein receptor (LDLR). We hypothesized that LDLR is a novel host factor for the SARS-CoV-2-Spike (S2S) protein that may be regulated by the Akt inhibitor Triciribine (TCN), a drug being tested in Phase III studies for breast cancer. We also hypothesized that nano-formulation of Triciribine (NanoTriciribine; NTCN) would enhance its efficacy and allow for intranasal delivery. Methods: Interactions between the recombinant proteins Spike-RBD (receptor binding domain), ACE2, LDLR and its ectodomains (EGFA-EFFB, C2-C5 and C2) were analyzed by binding assays and co-IP in HepG2, HK2, and 293T cells. Viral entry assays were performed with 2 S2S pseudoviruses using 293T cells + hACE2 and TMPRSS2 or Furin protease. The effect of NTCN or the LXR agonist GW-3965 on viral uptake (pseudotyped VSVΔG-GFP*S2S or chimera VSV-S2S-eGFP virus) was assessed. Akt, pAkt, ACE2, and LDLR levels were determined in 293T+hACE2 by flow cytometry. Assays were done in triplicates and 1-way-ANOVA with Tukey’s correction was used for statistics. Results: RBD protein binds modestly to the human LDLR (EC 50 :10μM) and its C2-C5 ectodomain (EC 50 :13.8μM). Co-IP revealed a novel and strong LDLR-ACE2 interaction in several human cell lines. LDLR overexpression in human cells increased the uptake of VSVΔG-GFP*S2S (FC=2.32;p<0.001) and chimera virus (FC=.33; p<.0001). NTCN and TCN each reduced pAkt/Akt ratio. 1μM TCN or NTCN reduced LDLR (7.2%;p<.01 & 15.6%;p<0.0001) and ACE2 (32%;p<0.05 & 44.7%;p<.01) cell surface expression, respectively. 1μM NTCN or GW-3965 reduced S2S viral entry by 64.2% (p<.0001) and 40.7% (p<.01), respectively, confirming a role for LDLR in S2S infection. In hACE2tg mice, chimera VSV-S2S caused significant lung infection as measured by qPCR, GFP expression in proximal and distal lung airway epithelial cells, and electron microscopy. Intranasal delivery of NTCN was well tolerated. Conclusions: LDLR enhanced S2S viral entry supporting the elevated COVID-19 susceptibility seen in patients with heart disease. NTCN is a promising candidate for prophylactic treatment against COVID-19.

ApoE and immunity in Alzheimer’s disease and related tauopathies: Low-density lipoprotein receptor to the rescue

In this issue of Neuron, Shi etal. (2021) show a protective role for the low-density lipoprotein receptor (LDLR) in tau pathology. Brain overexpression of LDLR lowers apolipoprotein E (apoE), suppresses microglial activation, preserves myelin, and ameliorates neurodegeneration, pointing the way toward potential new therapies.

Overexpressing low-density lipoprotein receptor reduces tau-associated neurodegeneration in relation to apoE-linked mechanisms

APOE is the strongest genetic risk factor for late-onset Alzheimer's disease. ApoE exacerbates tau-associated neurodegeneration by driving microglial activation. However, how apoE regulates microglial activation and whether targeting apoE is therapeutically beneficial in tauopathy is unclear. Here, we show that overexpressing an apoE metabolic receptor, LDLR (low-density lipoprotein receptor), in P301S tauopathy mice markedly reduces brain apoE and ameliorates tau pathology and neurodegeneration. LDLR overexpression (OX) in microglia cell-autonomously downregulates microglial Apoe expression and is associated with suppressed microglial activation as in apoE-deficient microglia. ApoE deficiency and LDLR OX strongly drive microglial immunometabolism toward enhanced catabolism over anabolism, whereas LDLR-overexpressing microglia also uniquely upregulate specific ion channels and neurotransmitter receptors upon activation. ApoE-deficient and LDLR-overexpressing mice harbor enlarged pools of oligodendrocyte progenitor cells (OPCs) and show greater preservation of myelin integrity under neurodegenerative conditions. They also show less reactive astrocyte activation in the setting of tauopathy.

Long non-coding RNA CASC19 facilitates non-small cell lung cancer cell proliferation and metastasis by targeting the miR-301b-3p/LDLR axis.

Non-small cell lung cancer (NSCLC) is a lethal tumor resulting in a large number of cancer-related deaths globally. Long noncoding RNAs (lncRNAs) may modulate tumor initiation and metastasis. Although dysregulation of lncRNA cancer susceptibility 19 (CASC19) is validated in NSCLC, further exploration of the CASC19-regulated mechanism in NSCLC is still needed. CASC19 expression was examined in NSCLC cells by a quantitative reverse transcriptase-polymerase chain reaction. The specific role of CASC19 in NSCLC was analyzed by cell counting kit-8, EdU, Transwell and western blot assays. The interaction between miR-301b-3p and CASC19 or low-density lipoprotein receptor (LDLR) was confirmed by luciferase reporter and RNA immunoprecipitation assays. CASC19 is markedly overexpressed in NSCLC. Its deficiency impairs cell proliferation, as well as metastasis in NSCLC. Molecular mechanism experiments indicated that CASC19 negatively modulates the expression of miR-301b-3p and miR-301b-3p can bind with CASC19 in NSCLC. In addition, miR-301b-3p binds to LDLR to impair its expression in NSCLC. Finally, rescue experiments showed that miR-301b-3p inhibition or LDLR overexpression counteracted the CASC19 knockdown-mediated function on cell proliferation and metastasis in NSCLC. CASC19 facilitates NSCLC cell proliferation and metastasis by targeting the miR-301b-3p/LDLR axis, offering a possible strategy for lncRNA-targeted treatment in NSCLC.

Effects of human LDLR overexpression on apoE-related tau pathology and brain dysfunction

Abstract ApoE4 is the strongest genetic risk factor for late-onset Alzheimer Disease. It has been reported that apoE isoforms directly affect tauopathy and tau-mediated neurodegeneration in P301S tau transgenic mice expressing different human apoE isoforms, with apoE4 resulting in markedly increased tau-mediated neurodegeneration and the absence of apoE being marked protective against neurodegeneration. In the brain, low-density lipoprotein receptor (LDLR) is one of the main apoE receptors that regulates apoE levels, but LDLR has very few identified ligands compared to other apoE receptors. LDLR overexpression in the brain can dramatically lower apoE and Aβ levels, as well as decrease Aβ accumulation and deposition. Therefore, we propose to evaluate whether AAV-mediated overexpression of human LDLR (hLDLR) is an efficient way to reduce apoE levels, tau pathology, and neurodegeneration. To study the effect of LDLR overexpression on apoE4-related tau pathology and neurodegeneration, P301S Tau/ApoE4 (TE4) male mice received bilateral intracerebroventricular injection with AAV expressing hLDLR or GFP-control, starting before the onset of tau pathology development. Brains were isolated after 9 month, divided into hemispheres, and analyzed by histological and biochemical techniques. Widespread expression of hLDLR throughout the brain was observed and apoE levels were significantly decreased. Furthermore, mouse nest-building impairment was rescued after hLDLR overexpression. Strikingly, there is a significant decrease in brain atrophy, phosphorylated tau deposition in TE4 mice overexpressing hLDLR. In summary, these data suggest that LDLR may be a promising target to lower apoE levels and decrease neurodegeneration.

Overexpressing Low-Density Lipoprotein Receptor Reduces Tau-Associated Neurodegeneration Via ApoE-Dependent and Independent Mechanisms

APOE is the strongest genetic risk factor for late-onset Alzheimer’s disease. ApoE exacerbates tau-associated neurodegeneration by driving microglial activation. However, how apoE regulates microglial activation and whether targeting apoE is therapeutically beneficial in tauopathy is unclear. Here we show that overexpressing a low-density lipoprotein receptor (LDLR) transgene in P301S tau transgenic mice markedly reduces brain apoE and ameliorates tau pathology and neurodegeneration. ApoE specifically interacts with a high-molecular-weight tau species, and highly correlates with phospho-tau and insoluble tau levels. Microglial expression of the LDLR transgene reduces intracellular apoE and is associated with less microglial activation. snRNA-seq analysis of apoE-deficient or LDLR-overexpressing brains reveals that apoE deficiency drives microglial catabolism and increases the oligodendrocyte progenitor cell population. LDLR overexpression shares overlapping mechanisms, but uniquely upregulates microglial expression of specific ion channels and neurotransmitter receptors in tauopathy. A subset of disease-associated astrocytes with both neuroprotective and neurotoxic gene signatures is also identified.

Immunohistochemical Labeling of Low-Density Lipoprotein Receptor and Scavenger Receptor Class B Type 1 Are Increased in Canine Lymphoma.

Altered lipid metabolism is a well-documented hallmark of neoplastic transformation and impacts disease progression. Two major lipoprotein receptors, the low-density lipoprotein receptor (LDL-R) and scavenger receptor class B, type 1 (SR-BI) are overexpressed in a number of cancer types in people. These receptors serve to deliver cholesterol to the tumor cells and have been used to target drug therapies. In this study, we performed a retrospective analysis of LDL-R and SR-B1 expression in canine lymphoma using archived formalin-fixed tissue samples. Cases were immunophenotyped and classified according to World Health Organization (WHO) standards prior to immunostaining for the LDL_R and SR-B1. A total of 45 cases were evaluated; 21 high grade B (HGB), 11 low grade B (LGB), 7 high grade T (HGT), and 6 low grade T (LGT) lymphomas. One sided Wilcoxon rank sum tests were used to compare staining intensity between neoplastic and hyperplastic lymphoid tissue. The relationships between histological score and tumor grade and score and stage at presentation were assessed using non-parametric Kruskal-Wallis tests. Neoplastic lymphoid tissue expressed higher levels of both receptors compared to reactive lymph nodes. Median LDL-R score was 85.0 (interquartile range = 101.7), Median SR-B1 score was 209.0 (interquartile range 105.2). No relationship between LDL-R or SR-B1 staining score and tumor grade or phenotype was found. Serum cholesterol concentration was compared between dogs with high and low grade tumors using a two sample T-test, and correlations between cholesterol concentration and histological score, and between the score for the two receptors were determined using a Spearman correlation. The high expression level of these lipoprotein receptors on most of the tumors could underlie the lack of relationship between score and tumor grade. The overexpression of LDL-R and SR-B1 in canine lymphoma holds therapeutic potential particularly in dogs that overexpress one or both of these receptors, and this warrants further investigation.

The endonuclease APE1 processes miR-92b formation, thereby regulating expression of the tumor suppressor LDLR in cervical cancer cells

Background:The molecular mechanisms underlying cervical cancer require elucidation to identify novel therapeutic targets. Apurinic/apyrimidinic endodeoxyribonuclease 1 (APE1) is a multifunctional apurinic/apyrimidinic (AP) endonuclease that influences the transcription of many cancer-related genes via microRNome regulation. Herein, we examine the role of miR-92b-3p (hereinafter miR-92b), whose processing may be regulated by APE1, in cervical cancer progression.Methods:APE1’s processing of miR-92b from its pri-miR form was measured by a quantitative reverse transcription polymerase chain reaction (qRT-PCR)-based ratio. APE1’s endonuclease activity was measured with AP-site incision assays. APE1-DROSHA interaction was studied with immunofluorescence, confocal and proximity ligation analyses. The miR-92b’s targeting of low-density lipoprotein receptor (LDLR) was investigated with luciferase reporter assays. The miR-92b mimics and shRNA-based miR-92b silencing, as well as LDLR overexpression and short interfering RNA (siRNA)-based LDLR silencing, were employed in CaSki and SiHa cervical cancer cells. Cell proliferation and chemosensitivity to paclitaxel and cisplatin were assayed. Cell-cycle progression and apoptosis were assessed by flow cytometry. Tumor growth was studied in a murine xenograft model.Results:APE1’s endonuclease activity, via association with the DROSHA-processing complex, is necessary for processing mature miR-92b, thereby regulating expression of miR-92b’s direct target LDLR. The miR-92b promotes cell proliferation in vitro and in vivo, promotes cell-cycle progression, and reduces apoptosis and chemosensitivity. LDLR silencing recapitulated miR-92b’s transformative effects, while LDLR overexpression rescued these effects.Conclusions:APE1 enhances miR-92b processing, thereby suppressing LDLR expression and enhancing cervical carcinoma progression. Our identification of the novel APE1-miR-92b-LDLR axis improves our understanding of the complex pathogenesis of cervical carcinoma and reveals a novel therapeutic strategy for combating this disease.

Intestine-Specific Overexpression of LDLR Enhances Cholesterol Excretion and Induces Metabolic Changes in Male Mice.

Roux-en-Y gastric bypass (RYGB) surgery is one of the most effective treatment options for severe obesity and related comorbidities, including hyperlipidemia, a well-established risk factor of cardiovascular diseases. Elucidating the molecular mechanisms underlying the beneficial effects of RYGB may facilitate development of equally effective, but less invasive, treatments. Recent studies have revealed that RYGB increases low-density lipoprotein receptor (LDLR) expression in the intestine of rodents. Therefore, in this study we first examined the effects of RYGB on intestinal cholesterol metabolism in human patients, and we show that they also exhibit profound changes and increased LDLR expression. We then hypothesized that the upregulation of intestinal LDLR may be sufficient to decrease circulating cholesterol levels. To this end, we generated and studied mice that overexpress human LDLR specifically in the intestine. This perturbation significantly affected intestinal metabolism, augmented fecal cholesterol excretion, and induced a reciprocal suppression of the machinery related to luminal cholesterol absorption and bile acid synthesis. Circulating cholesterol levels were significantly decreased and, remarkably, several other metabolic effects were similar to those observed in RYGB-treated rodents and patients, including improved glucose metabolism. These data highlight the importance of intestinal cholesterol metabolism for the beneficial metabolic effects of RYGB and for the treatment of hyperlipidemia.

Combination Glioma Therapy Mediated by a Dual-Targeted Delivery System Constructed Using OMCN-PEG-Pep22/DOX.

Accumulating studies have investigated the efficacy of receptor-mediated delivery of hydrophobic drugs in glioma chemotherapy. Here, a delivery vehicle comprising polyethylene glycol (PEG) and oxidized nanocrystalline mesoporous carbon particles (OMCN) linked to the Pep22 polypeptide targeting the low-density lipoprotein receptor (LDLR) is designed to generate a novel drug-loaded system, designated as OMCN-PEG-Pep22/DOX (OPPD). This system effectively targets glioma cells and the blood-brain barrier and exerts therapeutic efficacy through both near-infrared (NIR) photothermal and chemotherapeutic effects of loaded doxycycline (DOX). Pathological tissue microarrays show an association of LDLR overexpression in human glioma tissue with patient survival.NIR irradiation treatment and magnetic resonance imaging results show that OPPD reaches the effective glioma-killing temperature in a glioma-bearing rat with a skull bone removal model and considerably reduces glioma sizes relative to the drug-loaded system without the Pep22 peptide modification and the control respectively. Thus, OPPD not only effectively targets LDLR-overexpressing glioma but also exerts a dual therapeutic effect by transporting DOX into the glioma and generating thermal effects with near-infrared irradiation to kill tumor cells. These collective findings support the utility of the novel OPPD drug-loaded system as a promising drug delivery vehicle for clinical application in glioma therapy.

Magneto low-density nanoemulsion (MLDE): A potential vehicle for combined hyperthermia and photodynamic therapy to treat cancer selectively

In this paper, we introduce a new drug delivery system (DDS) called magneto low-density nanoemulsion (MLDE), which can carry maghemite nanoparticles and Chlorin e6 as an active photosensitizer drug. This design can enhance tumor damage after minor heat dissipation and/or minimum visible light photosensitization doses by classical magnetic hyperthermia (MHT) and photodynamic therapy (PDT), respectively. We establish protocols to prepare the MLDE and to load the drug combination onto it. The MLDE prepared herein is nanometric (<200 nm), has high encapsulation efficiency, and is stable for at least 12 months in water dispersions. Flow cytometry results demonstrated that MLDE presents targeted selectivity toward the MCF-7 breast cancer cell line but not in NHI-3T3 mouse fibroblast cell lines, because the MCF-7 cancer cell surface contains overexpressed low density lipoprotein (LDL) receptors. Despite this targeted effect, MHT or PDT alone does not prompt significant antiproliferative outcomes. On the other hand, MHT and PDT in combination induce a strong and synergic action on MCF-7 cells and reduce the cell viability. In conclusion, the developed MLDE deserves further investigation because it is biocompatible, displays good encapsulation efficiency, and is highly stable. Moreover, it is selectively taken up by cancer cell surfaces with receptor recognition based on LDL receptor overexpression, which potentiates the action of combined MHT and PDT.

Overexpression of low-density lipoprotein receptors stimulated by vascular endothelial growth factor in fibroblasts from pterygium

The activation of subconjunctival fibroblasts is believed to be responsible for the pathogenesis of pterygium. Vascular endothelial growth factor (VEGF) appears to be the most potent stimulator of formation and progression of pterygium. Pterygium excision is a common procedure, although the recurrence rates remain high. Various postoperative adjuvant therapies are now attempted to lower the recurrence rate, with severe side effects. To offer a greater therapeutic effect and lower side effects, it’s necessary to discover a constant nanoparticle drug delivery targeting to subconjunctival fibroblasts in pterygium (PSFs). This study was designed to investigate the expression of low-density lipoprotein receptor (LDLr) stimulated by VEGF in PSFs. We found that after exposure to VEGF, mRNA and protein levels of LDLr were both increased significantly in PSFs, assessed using relative quantitative real-time RT-PCR and Western blot. Moreover, it’s demonstrated that the expression of LDLr were positively correlated with the cells proliferation. Uptake of DiI–LDL via live PSFs was increased with time, estimated by confocal microscopy. The protein expression of LDLr in pterygium subconjunctival tissues was significantly higher than in normal subconjunctival tissues. These results suggest that LDLr in the activated PSFs may become a novel target receptor for controlled drug delivery to lower postsurgical recurrence rate.

Lipid Accumulation in HepG2 Cells Is Attenuated by Strawberry Extract through AMPK Activation.

Regulation of lipid metabolism is essential for treatment and prevention of several chronic diseases such as obesity, diabetes, and cardiovascular diseases, which are responsible for most deaths worldwide. It has been demonstrated that the AMP-activated protein kinase (AMPK) has a direct impact on lipid metabolism by modulating several downstream-signaling components. The main objective of the present work was to evaluate the in vitro effect of a methanolic strawberry extract on AMPK and its possible repercussion on lipid metabolism in human hepatocellular carcinoma cells (HepG2). For such purpose, the lipid profile and the expression of proteins metabolically related to AMPK were determined on cells lysates. The results demonstrated that strawberry methanolic extract decreased total cholesterol, low-density lipoprotein (LDL)-cholesterol, and triglycerides levels (up to 0.50-, 0.30-, and 0.40-fold, respectively) while it stimulated the p-AMPK/AMPK expression (up to 3.06-fold), compared to the control. AMPK stimulation led to the phosphorylation and consequent inactivation of acetyl coenzyme A carboxylase (ACC) and inhibition of 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR), the major regulators of fatty acids and cholesterol synthesis, respectively. Strawberry treatment also entailed a 4.34-, 2.37-, and 2.47-fold overexpression of LDL receptor, sirtuin 1 (Sirt1), and the peroxisome proliferator activated receptor gamma coactivator 1-alpha (PGC-1α), respectively, compared to control. The observed results were counteracted by treatment with compound C, an AMPK pharmacological inhibitor, confirming that multiple effects of strawberries on lipid metabolism are mediated by the activation of this protein.

MRNA levels of low-density lipoprotein receptors are overexpressed in the foci of deep bowel endometriosis.

Is mRNA expression of LDL receptors altered in deep bowel endometriotic foci? SUMMARY ANSWER: mRNA expression of LDL receptors is up-regulated in deep bowel endometriotic foci of patients with endometriosis. Several studies have demonstrated the overexpression of low-density lipoprotein receptors in various tumour cell lines and endometriosis has similar aspects to cancer, mainly concerning the pathogenesis of both diseases. This is the first study we knowof to investigate lipoprotein receptors expression in deep endometriosis with bowel involvement. During 2014-2015, an exploratory case-control study was conducted with 39 patients, including 20 women with a histological diagnosis of deep endometriosis compromising the bowel and 19 women without endometriosis who underwent laparoscopic tubal ligation. Peripheral blood samples were collected on the day of surgery for lipid profile analysis, and samples of endometrial tissue and of bowel endometriotic lesions were also collected. The tissue samples were sent for histopathological analysis and for LDL-R and LRP-1 gene expression screening using quantitative real-time PCR. Patients with deep endometriosis had lower LDL-cholesterol than patients without the disease (119 ± 23 versus 156 ± 35; P = 0.001). Gene expression analysis of LDL receptors revealed that LDL-R was more highly expressed in endometriotic lesions when compared to the endometrium of the same patient but not more than in the endometrium of women without endometriosis (0.027 ± 0.022 versus 0.012 ± 0.009 versus 0.019 ± 0.01, respectively; P < 0.001). LRP-1 was more highly expressed in endometriotic lesions, both when compared with the endometrium of the same patient and when compared with the endometrium of patients without the disease (0.307 ± 0.207 versus 0.089 ± 0.076 and versus 0.126 ± 0.072, respectively; P < 0.001). The study also showed that LDL-R gene expression in the endometrium of women with endometriosis was higher during the secretory phase of the menstrual cycle (P = 0.001). LRP-1 gene expression was increased during the secretory phase in the endometrium of women without the disease (P = 0.008). In the endometriotic lesions, the presence of fibrosis is substantial, restricting access to the stromal and glandular components of the lesion. Despite that, we found that LDL receptor mRNA was overexpressed. Future studies may perform laser microdissection to isolate the area of interest in the target tissue, excluding fibrosis contamination. This study supports the feasibility of LDL-R targeted therapy in the treatment of deep endometriosis. This study was supported by Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP #2011/17245-0). The authors have no conflicts of interest to declare.

Release of Vesicular Stomatitis Virus Spike Protein G-Pseudotyped Lentivirus from the Host Cell Is Impaired upon Low-Density Lipoprotein Receptor Overexpression.

Production of a vesicular stomatitis virus spike protein G (VSVG)-pseudotyped lentiviral expression vector in HEK293 cells decreased on overexpression of low-density lipoprotein receptor (LDLR) but not that of ICAM1 or TfR1. Reverse transcription-quantitative PCR (RT-qPCR) revealed a reduction in vector RNA as a function of LDLR expression. Decreased syncytium formation suggested diminished surface expression of VSVG. Intracellular VSVG granules colocalized with LDLR, ER-Golgi intermediate compartment protein 53 (ERGIC53), LAMP2, and vimentin but not with GM130 or calnexin, suggesting that VSVG interacts with LDLR within the ERGIC, resulting in rerouting into the aggresome/autophagosome pathway.

Wnt directs the endosomal flux of LDL ‐derived cholesterol and lipid droplet homeostasis

The Wnt pathway, which controls crucial steps of the development and differentiation programs, has been proposed to influence lipid storage and homeostasis. In this paper, using an unbiased strategy based on high-content genome-wide RNAi screens that monitored lipid distribution and amounts, we find that Wnt3a regulates cellular cholesterol. We show that Wnt3a stimulates the production of lipid droplets and that this stimulation strictly depends on endocytosed, LDL-derived cholesterol and on functional early and late endosomes. We also show that Wnt signaling itself controls cholesterol endocytosis and flux along the endosomal pathway, which in turn modulates cellular lipid homeostasis. These results underscore the importance of endosome functions for LD formation and reveal a previously unknown regulatory mechanism of the cellular programs controlling lipid storage and endosome transport under the control of Wnt signaling.