Adverse effect of oxidized cholesterol exposure on colitis is mediated by modulation of gut microbiota.

Both cholesterol and oxidized cholesterol (OXC) are present in human diets. The incidence of inflammatory bowel diseases (IBDs) is increasing in the world. The present study was to investigate the mechanism by which OXC promotes colitis using C57BL/6 mice as a model. Results shown that more severe colitis was developed in OXC-treated mice with the administration of dextran sulfate sodium (DSS) in water. Direct effects of short-term OXC exposure on gut barrier or inflammation were not observed in healthy mice. However, OXC exposure could cause gut microbiota dysbiosis with a decrease in the relative abundance of short-train fatty acids (SCFAs)-producing bacteria (Lachnospiraceae_NK4A136_group and Blautia) and an increase in the abundance of some potential harmful bacteria (Bacteroides). OXC-induced symptoms of colitis were eliminated when mice were administered with antibiotic cocktails, indicating the promoting effect of OXC on DSS-induced colitis was mediated by its effect on gut microbiota. Moreover, bacteria-depleted mice colonized with gut microbiome from OXC-DSS-exposed mice exhibited a severe colitis, further proving the gut dysbiosis caused by OXC exposure was the culprit in exacerbating the colitis. It was concluded that dietary OXC exposure increased the susceptibility of colitis in mice by causing gut microbiota dysbiosis.

Statins repress needle-like carbon nanotube- or cholesterol crystal-stimulated IL-1ß production by inhibiting the uptake of crystals by macrophages.

Statins are 3-hydroxy-3-methylglutaryl-CoA reductase inhibitors that lower atherogenic LDL-cholesterol levels. Statins exert clinically relevant anti-inflammatory effects; however, the underlying molecular mechanism remains unclear. Studies have shown that endogenous and exogenous pathogenic crystals, such as cholesterol and monosodium urate (MSU), and needle-like nanomaterials, such as multi-wall carbon nanotubes (MWCNT), induce the production of IL-1ß and play a critical role in the development of crystal-associated sterile inflammatory pathologies. In this study, we evaluated the effect of statins on crystal-induced IL-1ß production in macrophages. We found that various statins, including pitavastatin, atorvastatin, fluvastatin, and lovastatin, but not squalene synthase inhibitor, repressed IL-1ß release upon MWCNT stimulation. In addition, IL-1ß production induced by cholesterol crystals and MSU crystals, but not by ATP or nigericin, was diminished. MWCNT-stimulated IL-1ß release was dependent on the expression of NLRP3, but not AIM2, NLRC4, or MEFV. Statin-induced repression was accompanied by reduced levels of mature caspase-1 and decreased uptake of MWCNT into cells. Supplementation of mevalonate, geranylgeranyl pyrophosphate, or farnesyl pyrophosphate prevented the reduction in IL-1ß release, suggesting a crucial role of protein prenylation, but not cholesterol synthesis. The statin-induced repression of MWCNT-elicited IL-1ß release was observed in THP-1-derived and mouse peritoneal macrophages, but not in bone marrow-derived macrophages where statins act in synergy with lipopolysaccharide to enhance the expression of IL-1ß precursor protein. In summary, we describe a novel anti-inflammatory mechanism through which statins repress mature IL-1ß release induced by pathogenic crystals and nanoneedles by inhibiting the internalization of crystals by macrophages.

Selenium nanoparticles inhibit the formation of atherosclerosis in apolipoprotein E deficient mice by alleviating hyperlipidemia and oxidative stress.

Atherosclerosis can cause severe cardiovascular diseases, which is the most common cause of death in the world. It's of great significance to study the prevention and treatment of atherosclerosis. Selenium nanoparticles (SeNPs) has drawn more and more attention due to high biological activity, high bioavailability, strong antioxidant capacity and low toxicity, exhibiting great potential in biomedical application. Thus, this study aimed at explore the anti-atherosclerotic effect of two kinds of SeNPs, bovine serum albumin (BSA) surface-decorated SeNPs and chitosan (CS) surface-decorated SeNPs (CS-SeNPs), in apolipoprotein E deficient (ApoE-/-) mice fed with a high-cholesterol and high-fat diet, and the possible mechanisms. The results demonstrated that both BSA-SeNPs (25, 50 and 100 µg Se/kg body weight/day) and CS-SeNPs (50 µg Se/kg body weight/day) could reduce atherosclerotic lesions in ApoE-/- mice after oral administration for 12 weeks. And these effects might mainly attributed to the ability of BSA-SeNPs and CS-SeNPs to inhibit hyperlipidemia by suppressing hepatic cholesterol and fatty acid metabolism, and alleviate oxidative stress by enhancing antioxidant activity. Moreover, the benefits of BSA-SeNPs were dose-dependent and the medium dose of BSA-SeNPs (50 µg Se/kg body weight/day) was optimal. Generally, BSA-SeNPs with mean size 38.5 nm and negative surface charge showed better anti-atherosclerotic effect than CS-SeNPs with mean size 65.8 nm and positive surface charge. These results suggested that SeNPs could significantly alleviate the formation of atherosclerosis in ApoE-/- mice, possibly by inhibiting hyperlipidemia and oxidative stress, exhibiting a potential to serve as an anti-atherosclerotic agent.

HMGB1/RAGE/TLR4 axis and glutamate as novel targets for PCSK9 inhibitor in high fat cholesterol diet induced cognitive impairment and amyloidosis.

AIMS: Alzheimer's disease (AD) is a leading health problem in which increased amyloid ß (Aß) accumulation may occur due to abnormal Aß precursor protein processing by ß-secretase 1 (BACE1) enzyme. Lately, neuro-inflammation was recognized as a significant contributor to its pathogenesis. Although the causes of AD are not yet well understood, much evidence has suggested that dyslipidemia has harmful effects on cognitive function and is inextricably involved in AD pathogenesis. Cholesterol is a vital molecule involved in neuronal development. Alteration in neuronal cholesterol levels affects Aß metabolism and results in neurodegeneration. Proprotein-convertase-subtilisin/kexin type-9 (PCSK9) was found to decrease neuronal cholesterol uptake by degradation of LDL-receptor related protein 1 (LRP-1) responsible for neuronal cholesterol uptake. Accordingly, this study was designed to evaluate the effect of PCSK9-inhibition by alirocumab (Aliro) in high-fat-cholesterol-diet (HFCD)-induced-AD-like condition. MAIN METHODS: Wistar Rats were divided into six groups; control; HFCD; HFCD and Memantine; HFCD and Aliro (4, 8 and 16 mg/kg/week) to test for ability of Aliro to modulate cognitive impairment, amyloidosis, brain cholesterol homeostasis and neuro-inflammation in HFCD-induced-AD-like condition. KEY FINDINGS: Our results demonstrated an association between PCSK9 inhibition by Aliro and amelioration of cognitive deficit, cholesterol hemostasis and reduction of neuro-inflammation. Aliro was able to alleviate hippocampal LRP-1expression levels and reduce brain cholesterol, hippocampal BACE1, Aß42, high-mobility-group-box-1 protein, receptor for advanced-glycation-end-products and toll like receptor-4 with subsequent decrease of different inflammatory mediators as nuclear-factor-kappa-B (NF-κB), tumor-necrosis-factor-alpha (TNF-α), interleukin-1beta (IL-1ß) and IL-6. SIGNIFICANCE: PCSK9-inhibition may represent a new therapeutic target in AD especially for HFCD-induced-AD-like condition.

Seven-Step Synthesis of All-Nitrogenated Sugar Derivatives Using Sequential Overman Rearrangements.

All-nitrogenated sugars (ANSs), in which all hydroxy groups in a carbohydrate are replaced with amino groups, are anticipated to be privileged structures with useful biological activities. However, ANS synthesis has been challenging due to the difficulty in the installation of multi-amino groups. We report herein the development of a concise synthetic route to peracetylated ANSs in seven steps from commercially available monosaccharides. The key to success is the use of the sequential Overman rearrangement, which enables formal simultaneous substitution of four or five hydroxy groups in monosaccharides with amino groups. A variety of ANSs are available through the same reaction sequence starting from different initial monosaccharides by chirality transfer of secondary alcohols. Transformations of the resulting peracetylated ANSs such as glycosylation and deacetylation are also demonstrated. Biological studies reveal that ANS-modified cholesterol show cytotoxicity against human cancer cell lines, whereas each ANS and cholesterol have no cytotoxicity.

Phenotypic Changes in Macrophage Activation in a Model of Nonalcoholic Fatty Liver Disease using Microminipigs.

AIM: Non-alcoholic fatty liver disease (NAFLD) is one of the most common chronic liver disorders associated with metabolic syndrome, and its prevalence has been on the rise. The pathogenesis of NAFLD has not yet been sufficiently elucidated due to the multifactorial nature of the disease, although the activation of macrophages/Kupffer cells is considered to be involved. We previously reported an animal model of NAFLD using MicrominipigsTM (µMPs) fed high-fat diets containing cholesterol with or without cholic acid. The aim of this study was to investigate the phenotypic changes of macrophages that occur during the development of NAFLD. METHODS: Immunohistochemistry of macrophages, lymphocytes, and stellate cells was performed using liver samples, and the density of positive cells was analyzed. RESULTS: The number of Iba-1-positive macrophages increased with increasing cholesterol content in the diet. The numbers of CD163-positive macrophages and CD204-positive macrophages also increased with increasing cholesterol content in the diet; however, the proportion of CD204-positive macrophages among Iba-1-positive macrophages was significantly reduced by cholic acid supplementation. CONCLUSION: The results suggest that lipid accumulation induced macrophage recruitment in swine livers, and that the number of M2-like macrophages increased at the early stage of NAFLD, while the number of M1-like macrophages increased at the late stage of NAFLD, resulting in a liver condition like non-alcoholic steatohepatitis. We provide evidence of the phenotypic changes that occur in macrophages during the development of NAFLD that has never been reported before using µMPs.

Cholesterol-Induced Phenotypic Modulation of Smooth Muscle Cells to Macrophage/Fibroblast-like Cells Is Driven by an Unfolded Protein Response.

OBJECTIVE: Vascular smooth muscle cells (SMCs) dedifferentiate and initiate expression of macrophage markers with cholesterol exposure. This phenotypic switching is dependent on the transcription factor Klf4 (Krüppel-like factor 4). We investigated the molecular pathway by which cholesterol induces SMC phenotypic switching. Approach and Results: With exposure to free cholesterol, SMCs decrease expression of contractile markers, activate Klf4, and upregulate a subset of macrophage and fibroblast markers characteristic of modulated SMCs that appear with atherosclerotic plaque formation. These phenotypic changes are associated with activation of all 3 pathways of the endoplasmic reticulum unfolded protein response (UPR), Perk (protein kinase RNA-like endoplasmic reticulum kinase), Ire (inositol-requiring enzyme) 1α, and Atf (activating transcription factor) 6. Blocking the movement of cholesterol from the plasma membrane to the endoplasmic reticulum prevents free cholesterol-induced UPR, Klf4 activation, and upregulation of the majority of macrophage and fibroblast markers. Cholesterol-induced phenotypic switching is also prevented by global UPR inhibition or specific inhibition of Perk signaling. Exposure to chemical UPR inducers, tunicamycin and thapsigargin, is sufficient to induce these same phenotypic transitions. Finally, analysis of published single-cell RNA sequencing data during atherosclerotic plaque formation in hyperlipidemic mice provides preliminary in vivo evidence of a role of UPR activation in modulated SMCs. CONCLUSIONS: Our data demonstrate that UPR is necessary and sufficient to drive phenotypic switching of SMCs to cells that resemble modulated SMCs found in atherosclerotic plaques. Preventing a UPR in hyperlipidemic mice diminishes atherosclerotic burden, and our data suggest that preventing SMC transition to dedifferentiated cells expressing macrophage and fibroblast markers contributes to this decreased plaque burden.

Glucose and cholesterol induce abnormal cell divisions via DAF-12 and MPK-1 in C. elegans.

People exposed to starvation have a high risk of developing cancer later in life, and prior studies have shown these individuals have high insulin and cholesterol levels and are sensitive to glucose. Using C. elegans as a model, we found that glucose and cholesterol can promote survival and cause starved L1 diapause worms to undergo abnormal neuronal cell divisions. Starvation has also been shown to promote long-term survival; however, we found that the functions of glucose and cholesterol in relation to these cell divisions are distinct from their effects on survival. We demonstrate that glucose functions in a DAF-16/FOXO-independent IIS pathway to activate the MAPK ontogenetic signaling to induce neuronal Q-cell divisions, and cholesterol works through DAF-12/steroidogenic pathways to promote these cell divisions. daf-12 and mpk-1/MAPK mutants suppress the function of glucose and cholesterol in these divisions, and a fully functioning dpMPK-1 requires the steroid hormone receptor DAF-12 for these divisions to occur. These afflictions also can be passed on to the immediate progeny. This work indicates a possible link between glucose and cholesterol in starved animals and an increased risk of cancer.

Dye-Loaded Quatsomes Exhibiting FRET as Nanoprobes for Bioimaging.

Fluorescent organic nanoparticles (FONs) are emerging as an attractive alternative to the well-established fluorescent inorganic nanoparticles or small organic dyes. Their proper design allows one to obtain biocompatible probes with superior brightness and high photostability, although usually affected by low colloidal stability. Herein, we present a type of FONs with outstanding photophysical and physicochemical properties in-line with the stringent requirements for biomedical applications. These FONs are based on quatsome (QS) nanovesicles containing a pair of fluorescent carbocyanine molecules that give rise to Förster resonance energy transfer (FRET). Structural homogeneity, high brightness, photostability, and high FRET efficiency make these FONs a promising class of optical bioprobes. Loaded QSs have been used for in vitro bioimaging, demonstrating the nanovesicle membrane integrity after cell internalization, and the possibility to monitor the intracellular vesicle fate. Taken together, the proposed QSs loaded with a FRET pair constitute a promising platform for bioimaging and theranostics.

The Protective Effect of Dabigatran and Rivaroxaban on DNA Oxidative Changes in a Model of Vascular Endothelial Damage with Oxidized Cholesterol.

BACKGROUND: Atherosclerotic plaques are unstable, and their release may result in thrombosis; therefore, currently, antiplatelet therapy with anticoagulants is recommended for the treatment of acute coronary syndrome. The aim of this study was to assess the effect of oxidized cholesterol on human umbilical vascular endothelial cells (HUVECs). The study also examines the protective and repairing effect of dabigatran and rivaroxaban in a model of vascular endothelial damage with 25-hydroxycholesterol (25-OHC). METHODS: HUVECs were treated with compounds induce DNA single-strand breaks (SSBs) using the comet assay. Oxidative DNA damage was detected using endonuclease III (Nth) or human 8 oxoguanine DNA glycosylase (hOOG1). Reactive oxygen species (ROS) formation was determined using flow cytometry. RESULTS: 25-hydroxycholesterol caused DNA SSBs, induced oxidative damage and increased ROS in the HUVECs; ROS level was lowered by dabigatran and rivaroxaban. Only dabigatran was able to completely repair the DNA SSBs induced by oxysterol. Dabigatran was able to reduce the level of oxidative damage of pyrimidines induced by oxysterol to the level of control cells. CONCLUSIONS: Observed changes strongly suggest that the tested anticoagulants induced indirect repair of DNA by inhibiting ROS production. Furthermore, dabigatran appears to have a higher antioxidant activity than rivaroxaban.

pH-Responsive nanoparticles based on cholesterol/imidazole modified oxidized-starch for targeted anticancer drug delivery.

Various nanoparticles have been developed for tumor-targeted drug delivery. However, nanoparticles with effective targeting and intelligent release capacity are still deficient. Herein, we present new pH-responsive and neutral charged nanoparticles for tumor-targeted anticancer drug delivery. Oxidized starch was synthesized and simultaneously modified by cholesterol and imidazole to obtain amphiphilic cholesterol/imidazole modified oxidized-starch (Cho-Imi-OS). Cho-Imi-OS easily self-assembled into nanoparticles by dialysis. Curcumin was selected as model drug to be encapsulated into the hydrophobic core of nanoparticles. The results showed that curcumin would effectively accumulate in cancer cells by encapsulating into the nanoparticles owing to the nano-sized structure and near neutral charged property of nanoparticles. Curcumin was released faster at pH 5.5 than that at pH 7.4 from the curcumin-loaded nanoparticles (Cur-NPs), indicating the pH-triggered release capacity of Cur-NPs after endocytosis by endosomes since the pH is low to 5.0∼6.0 in endosomes. Naturally, Cur-NPs showed significantly strong inhibitory effect on cancer cells.

Crystal Clots as Therapeutic Target in Cholesterol Crystal Embolism.

RATIONALE: Cholesterol crystal embolism can be a life-threatening complication of advanced atherosclerosis. Pathophysiology and molecular targets for treatment are largely unknown. OBJECTIVE: We aimed to develop a new animal model of cholesterol crystal embolism to dissect the molecular mechanisms of cholesterol crystal (CC)-driven arterial occlusion, tissue infarction, and organ failure. METHODS AND RESULTS: C57BL/6J mice were injected with CC into the left kidney artery. Primary end point was glomerular filtration rate (GFR). CC caused crystal clots occluding intrarenal arteries and a dose-dependent drop in GFR, followed by GFR recovery within 4 weeks, that is, acute kidney disease. In contrast, the extent of kidney infarction was more variable. Blocking necroptosis using mixed lineage kinase domain-like deficient mice or necrostatin-1s treatment protected from kidney infarction but not from GFR loss because arterial obstructions persisted, identifying crystal clots as a primary target to prevent organ failure. CC involved platelets, neutrophils, fibrin, and extracellular DNA. Neutrophil depletion or inhibition of the release of neutrophil extracellular traps had little effects, but platelet P2Y12 receptor antagonism with clopidogrel, fibrinolysis with urokinase, or DNA digestion with recombinant DNase I all prevented arterial occlusions, GFR loss, and kidney infarction. The window-of-opportunity was <3 hours after CC injection. However, combining Nec-1s (necrostatin-1s) prophylaxis given 1 hour before and DNase I 3 hours after CC injection completely prevented kidney failure and infarcts. In vitro, CC did not directly induce plasmatic coagulation but induced neutrophil extracellular trap formation and DNA release mainly from kidney endothelial cells, neutrophils, and few from platelets. CC induced ATP release from aggregating platelets, which increased fibrin formation in a DNase-dependent manner. CONCLUSIONS: CC embolism causes arterial obstructions and organ failure via the formation of crystal clots with fibrin, platelets, and extracellular DNA as critical components. Therefore, our model enables to unravel the pathogenesis of the CC embolism syndrome as a basis for both prophylaxis and targeted therapy.

Chemical Pathology of Homocysteine VII. Cholesterol, Thioretinaco Ozonide, Mitochondrial Dysfunction, and Prevention of Mortality.

The purpose of this review is to elucidate how low blood cholesterol promotes mitochondrial dysfunction and mortality by the loss of thioretinaco ozonide from opening of the mitochondrial permeability transition pore (mPTP). Mortality from infections and cancer are both inversely associated with blood cholesterol, as determined by multiple cohort studies from 10 to 30 years earlier. Moreover, low-density lipoprotein (LDL) is inversely related to all-cause and/or cardiovascular mortality, as determined by followup study of elderly cohorts. LDL adheres to and inactivates most microorganisms and their toxins, causing aggregation of LDL and homocysteinylated autoantibodies which obstruct vasa vasorum and produce intimal microabscesses, the vulnerable atherosclerotic plaques. The active site of mitochondrial oxidative phosphorylation and adenosine triphosphate (ATP) biosynthesis is proposed to consist of thioretinaco, a complex of two molecules of thioretinamide with cobalamin, oxidized to the disulfonium thioretinaco ozonide and complexed with oxygen, nicotinamide adenine dinucleotide (NAD+), phosphate, and ATP. Loss of the active site complex from mitochondria results from the opening of the mPTP and from decomposition of the disulfonium active site by electrophilic carcinogens, oncogenic viruses, microbes, and by reactive oxygen radicals from ionizing and non-ionizing radiation. Suppression of innate immunity is caused by the depletion of adenosyl methionine because of increased polyamine biosynthesis, resulting in inhibition of nitric oxide and peroxynitrite biosynthesis. Opening of the mPTP produces a loss of thioretinaco ozonide from mitochondria. This loss impairs ATP biosynthesis and causes the mitochondrial dysfunction observed in carcinogenesis, atherosclerosis, aging and dementia. Cholesterol inhibits the opening of the mPTP by preventing integration of the pro-apoptotic Bcl-2-associated X protein (BAX) in the outer mitochondrial membrane. This inhibition explains how elevated LDL reduces mitochondrial dysfunction by preventing loss of the active site of oxidative phosphorylation from mitochondria.

Lens cholesterol biosynthesis inhibition: A common mechanism of cataract formation in laboratory animals by pharmaceutical products.

CJ-12,918, a 5-lipoxygenase (5-LO) inhibitor, caused cataracts during a 1-month safety assessment studies in rats whereas the structurally similar ZD-2138 was without effect. For CJ-12,918 analogs, blocking different sites of metabolic liability reduced (CJ-13,454) and eliminated (CJ-13,610) cataract formation in both rats and dogs. Using this chemical series as a test set, models and mechanisms of toxicity were first explored by testing the utility of ex vivo rat lens explant cultures as a safety screen. This model overpredicted the cataractogenic potential of ZD-2138 due to appreciably high lens drug levels and was abandoned in favor of a mechanism-based screen. Perturbations in lens sterol content, from a decline in lathosterol content, preceded cataract formation suggesting CJ-12,918 inhibited lens cholesterol biosynthesis (LCB). A 2-day bioassay in rats using ex vivo LCB assessments showed that the level of LCB inhibition was correlated with incidence of cataract formation in animal studies by these 5-LO inhibitors. Thereafter, this 2-day bioassay was applied to other pharmaceutical programs (neuronal nitric oxide synthase, sorbitol dehydrogenase inhibitor, squalene synthetase inhibitor and stearoyl-CoA desaturase-1 inhibitors/D4 antagonists) that demonstrated cataract formation in either rats or dogs. LCB inhibition >40% was associated with a high incidence of cataract formation in both rats and dogs that was species specific. Bioassay sensitivity/specificity were further explored with positive (RGH-6201/ciglitazone/U18666A) and negative (tamoxifen/naphthalene/galactose) mechanistic controls. This body of work over two decades shows that LCB inhibition was a common mechanism of cataract formation by pharmaceutical agents and defined a level of inhibition >40% that was typically associated with causing cataracts in safety assessment studies typically ≥1 month.

Halting hyaluronidase activity with hyaluronan-based nanohydrogels: development of versatile injectable formulations.

Hyaluronan (HA) is among the most used biopolymers for viscosupplementation and dermocosmetics. However, the current injectable HA-based formulations present relevant limitations: I) unmodified HA is quickly degraded by endogenous hyaluronidases (HAase), resulting in short lasting properties; II) cross-linked HA, although shows enhanced stability against HAase, often contains toxic chemical cross-linkers. As such, herein, we present biocompatible self-assembled hyaluronan-cholesterol nanohydrogels (HA-CH NHs) able to bind to HAase and inhibit the enzyme activity in vitro, more efficiently than currently marketed HA-based cross-linked formulations (e.g. Jonexa™). HA-CH NHs inhibit HAase through a mixed mechanism, by which NHs bind to HAase with an affinity constant 7-fold higher than that of native HA. Similar NHs, based on gellan-CH, evidenced no binding to HAase, neither inhibition of the enzyme activity, suggesting this effect might be due to the specific binding of HA-CH to the active site of the enzyme. Therefore, HA-CH NHs were engineered into injectable hybrid HA mixtures or physical hydrogels, able to halt the enzymatic degradation of HA.

A novel method for conjugating the terminal amine of peptide ligands to cholesterol: synthesis iRGD-cholesterol.

AIM: Conventional conjugation reactions often involve the use of activated PEG as a linker, but concerns about PEG-mediated reduction in intracellular delivery and enhanced immunogenicity have generated interest in developing methods that eliminate the need for a PEG linker. MATERIALS & METHODS: Reaction conditions were identified that specifically couples the terminal amine of a cyclic iRGD peptide (CRGDRGPDC) to the hydroxyl moiety of cholesterol through a short carbamate linker. RESULTS & CONCLUSION: Using this method for synthesizing iRGD-cholesterol, peptide ligands can be incorporated into lipid-based delivery systems, thereby eliminating concerns about adverse reactions to PEG. Toxicity and stability data indicate low toxicity and adequate serum stability at low ligand levels.

Disease Progression and Pharmacological Intervention in a Nutrient-Deficient Rat Model of Nonalcoholic Steatohepatitis.

BACKGROUND: There is a marked need for improved animal models of nonalcoholic steatohepatitis (NASH) to facilitate the development of more efficacious drug therapies for the disease. METHODS: Here, we investigated the development of fibrotic NASH in male Wistar rats fed a choline-deficient L-amino acid-defined (CDAA) diet with or without cholesterol supplementation for subsequent assessment of drug treatment efficacy in NASH biopsy-confirmed rats. The metabolic profile and liver histopathology were evaluated after 4, 8, and 12 weeks of dieting. Subsequently, rats with biopsy-confirmed NASH were selected for pharmacological intervention with vehicle, elafibranor (30 mg/kg/day) or obeticholic acid (OCA, 30 mg/kg/day) for 5 weeks. RESULTS: The CDAA diet led to marked hepatomegaly and fibrosis already after 4 weeks of feeding, with further progression of collagen deposition and fibrogenesis-associated gene expression during the 12-week feeding period. Cholesterol supplementation enhanced the stimulatory effect of CDAA on gene transcripts associated with fibrogenesis without significantly increasing collagen deposition. Pharmacological intervention with elafibranor, but not OCA, significantly reduced steatohepatitis scores, and fibrosis-associated gene expression, however, was unable to prevent progression in fibrosis scores. CONCLUSION: CDAA-fed rats develop early-onset progressive NASH, which offers the opportunity to probe anti-NASH compounds with potential disease-modifying properties.

In Vivo MRI of Amyloid Plaques in a Cholesterol-Fed Rabbit Model of Alzheimer's Disease.

Hypercholesterolemia has been identified as a risk factor for Alzheimer's disease. In this study, rabbits were fed either a cholesterol diet or normal chow diet for 24 months. At endpoint, in vivo MRI was performed at the field strength of 3 Tesla using fast imaging employing steady state acquisition without (FIESTA) or with susceptibility-weighted post-processing (SWI-FIESTA) and susceptibility-weighted imaging with multi-echo acquisition (SWAN). This imaging revealed signal voids/hypointensities throughout the cortex, sub-cortex, and hippocampus of cholesterol-fed animals compared to control animals. Quantitative image analysis corroborated these qualitative findings and highlighted that SWI processing of FIESTA images significantly improved the detectability of plaques (p < 0.05). Aß immunostaining and Prussian blue staining for iron demonstrated that the voids in MR images corresponded to iron-laden Aß-positive plaques. This study demonstrates non-invasive in vivo visualization of Aß plaques in a diet-induced large animal model of Alzheimer's disease. This work lays the foundation for future work focusing on longitudinal monitoring of plaque formation in this model and the effects of diet or drug interventions.

Synthesis of lipid-black phosphorus quantum dot bilayer vesicles for near-infrared-controlled drug release.

Black phosphorus quantum dots are incorporated into liposomal bilayers to produce a drug delivery system with excellent near-infrared (NIR) photothermal properties and drug release capability controlled by light. In vitro experiments demonstrate its good biocompatibility and NIR-light-induced chemo-photothermal antitumor efficiency.

Role of dysfunctional adipocytes in cholesterol-induced nonobese metabolic syndrome.

Many studies have reported the causes of obese metabolic syndrome (MS); however, the causes of nonobese MS (NMS) remain unknown. In this study, we demonstrated that inflamed dysfunctional adipose tissue plays a crucial role in cholesterol-induced NMS. Control (C), high cholesterol (HC) and HC with 10% fructose in drinking water (HCF) diets were fed to Sprague-Dawley rats for 12 weeks. After 12 weeks, the body weights of the C- and HC-fed rats were comparable, but the weights of the HCF-fed rats were relatively low. Cholesterol caused metabolic problems such as high blood pressure, hypercholesterolemia and hypoinsulinemia. The HCF-fed rats exhibited whole-body insulin resistance with low circulating high-density lipoprotein levels. Increases in the tumor necrosis factor α level in the plasma, the number of CD68+ macrophages and the free nuclear factor-κB level in gonadal white adipose tissue (gWAT) resulted in local inflammation, which appeared as inflamed dysfunctional gWAT. Reduced superoxide dismutases (SODs) deteriorate natural antioxidant defense systems and induce reactive oxygen species in gWAT. Dysregulation of plasma levels of catecholamine, adipokines (leptin and adiponectin), hormone-sensitive lipase and perilipin in cholesterol-induced inflamed adipose tissue contributed to increased lipolysis and increased circulating nonesterified fatty acids. Cholesterol activated inflammation, lipolysis and cell death in 3T3-L1 adipocytes. Moreover, Chol-3T3-CM reduced the population of M2-type Raw264.7 macrophages, indicating that the macrophage polarization is mediated by cholesterol. Together, our findings indicate that inflamed dysfunctional adipocytes are critical in NMS, supporting the development of anti-inflammatory agents as potential therapeutic drugs for treating NMS.