A new sponge-type hydrogel based on hyaluronic acid and poly(methylvinylether-alt-maleic acid) as a 3D platform for tumor cell growth.

A new sponge-type hydrogel was obtained by cross-linking hyaluronic acid (HA) and poly(methylvinylether-alt-maleic acid) P(MVE-alt-MA) through a solvent-free thermal method. The sponge-type hydrogel was characterized and checked as a support for cell growth. The influence of concentration and weight ratio of polymers on the morphology and hydrogel stability was investigated. The total polymers concentration of 3% (w/w) and the weight ratio of 1:1 were optimal for the synthesis of a stable hydrogel (HA3P50) and to promote cell proliferation. The swelling measurements revealed a high-water absorption capacity of the hydrogel in basic medium. Diphenhydramine (DPH), lidocaine (Lid) and propranolol (Prop) were loaded within the hydrogel as a model drugs to investigate the ability of drug transport and release. In vitro studies revealed that HA3P50 hydrogel promoted the adhesion and proliferation of human hepatocellular carcinoma cell line HepG2, providing a good support for 3D cell culture to obtain surrogate tumor scaffold suitable for preclinical anti-cancer drug screening.

Hyperlipidemia Determines Dysfunctional HDL Production and Impedes Cholesterol Efflux in the Small Intestine: Alleviation by Ginger Extract.

SCOPE: To assess the impact of ginger extract (GIN) in stimulating the production of quality HDL and the cholesterol efflux in the small intestine (SI), key processes in the management of hyperlipidemia (HL)-induced hepatic steatosis, and atherosclerosis. METHODS AND RESULTS: Three groups of hamsters are used: (i) N, fed standard diet, (ii) HL, fed high-fat diet for 21 weeks, and (iii) HL-GIN, HL treated with GIN for the last 5 weeks of diet. Apolipoprotein A-I (apoA-I), malondialdehyde-apoA-I (MDA-apoA-I), paraoxonase1 (PON1), and myeloperoxidase (MPO) are measured in plasma and SI. ATP-binding cassette A1 transporter (ABCA1), ABCG5/G8, liver X receptor α/ß (LXRα/ß), peroxisome proliferator-activated receptor γ (PPARγ), and sirtuin1 (SIRT1) are assessed in the SI. Results show that in HL plasma, GIN decreases MDA-apoA-I, MPO/PON1 ratio and increases HDL-cholesterol/total cholesterol. In HL-SI, GIN decreases MDA-apoA-I and MPO, increases ApoA-I, PON1, and ABCA1, and restores cholesterol efflux disturbed by HL (SIRT1-LXRα/ß-PPARγ-ABCG8). GIN administration is associated with the reduction of the aortic valves lipid-deposits. CONCLUSION: In HL conditions, GIN stimulates the functional HDL production by restoring apoA-I quality and quantity through inhibition of the oxidative stress, and increases cholesterol efflux in the SI. These effects are associated with the restoration of SIRT1-LXRα/ß-PPARγ pathway.

Zingiber officinale extract administration diminishes steroyl-CoA desaturase gene expression and activity in hyperlipidemic hamster liver by reducing the oxidative and endoplasmic reticulum stress.

BACKGROUND: Stearoyl CoA desaturases (SCD) are enzymes that convert saturated to monounsaturated fatty acids and have increased activity in hepatic steatosis. PURPOSE: We aimed to investigate the potential of ginger extract (GIN) to modulate the liver SCD1 expression and activity in hyperlipidemic (HL) conditions, in order to lower lipid accumulation in the steatotic liver. STUDY DESIGN/METHODS: Male Golden Syrian hamsters were divided in three groups: (i) fed with standard chow (N), (ii) fed with standard chow plus 3% cholesterol and 15% butter for 21 weeks (HL), (iii) HL treated with GIN (800 µg/kg body weight/day) in the last 5 weeks of fat diet (HL-GIN). Cholesterol (C), triglycerides (TG), non-esterified fatty acids (NEFA), SCD1 estimated activity (C16:1n7/C16:0; C18:1n9/C18:0) and gene expression, acetyl-CoA carboxylase (ACC), thiobarbituric acid reactive substances (TBARS), paraoxonase1 (PON1) and myeloperoxidase (MPO) were determined in the plasma and liver of all hamsters. We measured protein expression of endoplasmic reticulum stress (ERS) markers, gene and protein expression of liver X receptor α/ß (LXRα/ß), peroxisome proliferator-activated receptor γ (PPARγ), ATP-binding cassette sub-family G member 5/8 (ABCG5/G8) and 7α-hydroxylase1 (CYP7A1) in all hamsters' livers. RESULTS: In plasma, in HL-GIN versus HL hamsters, SCD1 estimated activity was lower (27%; 15%, p < 0.05), NEFA levels decreased by 91%, p < 0.001, while C and TG levels did not vary; the oxidative stress expressed as MPO and TBARS levels decreased (15%; 11%, p < 0.01), while PON1 protein increased (75%, p < 0.05). In the liver of HL-GIN versus HL, C, TG, NEFA, MPO and TBARS levels decreased (8-40%, p < 0.05) and PON1 protein levels increased (30%, p < 0.05), SCD1 estimated activity decreased (8%; 9%, p < 0.05), in parallel with the reduced gene expression of SCD1 and ACC (70-80%, p < 0.05). The protein expression of the ERS sensors decreased (30-65%, p < 0.05), while that of ABCG5/G8, CYP7A1, LXRα/ß and PPARγ increased in HL-GIN (20-30%, p < 0.05) versus HL liver. CONCLUSION: GIN reduces SCD1 estimated activity and expression, as well as the lipids accumulated in the livers of HL hamsters. This is achieved through a mechanism involving the decrease of the oxidative and ERS, and the enhancement of cholesterol efflux.

Inhibition of miR-486 and miR-92a decreases liver and plasma cholesterol levels by modulating lipid-related genes in hyperlipidemic hamsters.

In the present study we aimed to evaluate the potential of in vivo inhibition of miR-486 and miR-92a to reverse hyperlipidemia, then to identify and validate their lipid metabolism-related target genes. Male Golden-Syrian hamsters fed a hyperlipidemic (HL) diet (standard chow plus 3% cholesterol and 15% butter, 10 weeks) were injected subcutaneously with lock-nucleic acid inhibitors for either miR-486 or miR-92a. Lipids and miRNAs levels in liver and plasma, and hepatic expression of miRNAs target genes were assessed in all HL hamsters. MiR-486 and miR-92a target genes were identified by miRWalk analysis and validated by 3'UTR cloning in pmirGLO vectors. HL hamsters had increased liver (2.8-fold) and plasma (twofold) miR-486 levels, and increased miR-92a (2.8-fold and 1.8-fold, respectively) compared to normolipidemic hamsters. After 2 weeks treatment, liver and plasma cholesterol levels decreased (23 and 17.5% for anti-miR-486, 16 and 22% for miR-92a inhibition). Hepatic triglycerides and non-esterified fatty acids content decreased also significantly. Bioinformatics analysis and 3'UTR cloning in pmirGLO vector showed that sterol O-acyltransferase-2 (SOAT2) and sterol-regulatory element binding transcription factor-1 (SREBF1) are targeted by miR-486, while ATP-binding cassette G4 (ABCG4) and Niemann-Pick C1 (NPC1) by miR-92a. In HL livers and in cultured HepG2 cells, miR-486 inhibition restored the levels of SOAT2 and SREBF1 expression, while anti-miR-92a restored ABCG4, NPC1 and SOAT2 expression compared to scrambled-treated HL hamsters or cultured cells. In vivo inhibition of miR-486 and miR-92a could be a useful and valuable new approach to correct lipid metabolism dysregulation.

Dysfunctional high-density lipoproteins have distinct composition, diminished anti-inflammatory potential and discriminate acute coronary syndrome from stable coronary artery disease patients.

There is a stringent need to find means for risk stratification of coronary artery diseases (CAD) patients. We aimed at identifying alterations of plasma high-density lipoproteins (HDL) components and their validation as dysfunctional HDL that could discriminate between acute coronary syndrome (ACS) and stable angina (SA) patients. HDL2 and HDL3 were isolated from CAD patients' plasma and healthy subjects. ApolipoproteinAI (apoAI), apoAII, apoCIII, malondialdehyde (MDA), myeloperoxidase (MPO), ceruloplasmin and paraoxonase1 (PON1) were assessed. The anti-inflammatory potential of HDL subfractions was tested by evaluating the secreted inflammatory molecules of tumor necrosis factor α-activated endothelial cells (EC) upon co-incubation with HDL2 or HDL3. We found in ACS versus SA patients: 40% increased MPO, MDA, apoCIII in HDL2 and HDL3, 35% augmented apoAII in HDL2, and in HDL3 increased ceruloplasmin, decreased apoAII (40%) and PON1 protein and activity (15% and 25%). Co-incubation of activated EC with HDL2 or HDL3 from CAD patients induced significantly increased levels of secreted inflammatory molecules, 15-20% more for ACS versus SA. In conclusion, the assessed panel of markers correlates with the reduced anti-inflammatory potential of HDL subfractions isolated from ACS and SA patients (mostly for HDL3 from ACS) and can discriminate between these two groups of CAD patients.

Hyperglycemia Determines Increased Specific MicroRNAs Levels in Sera and HDL of Acute Coronary Syndrome Patients and Stimulates MicroRNAs Production in Human Macrophages.

We aimed to determine the levels of microRNAs (miRNAs) in sera and HDL of acute coronary syndrome (ACS) compared to stable angina (SA) patients with/without hyperglycemia, and evaluate comparatively the functional effect of these sera on the processing machinery proteins (Drosha, DGCR8, Dicer) and miRNAs production in human macrophages. MiRNAs levels in sera and HDL from 35 SA and 72 ACS patients and 30 healthy subjects were measured by using microRNA TaqMan assays. MiR-223, miR-92a, miR-486, miR-122, miR-125a and miR-146a levels were higher in the hyperglycemic ACS compared to normoglycemic sera. MiR-223 and miR-486 prevailed in HDL2, while miR-92a predominated in HDL3, all three miRNAs discriminating between ACS and SA patients; their levels were increased in HDL from hyperglycemic ACS patients versus normoglycemic ones. The incubation of human macrophages with sera from ACS and SA patients showed that all patients' sera induced an increase of Drosha, DGCR8 and Dicer expressions and of selected miRNAs levels compared to control sera, the effect being higher in the case of hyperglycemic versus normoglycemic ACS sera. The addition of glucose to SA and ACS sera increased Drosha, DGCR8 and Dicer expression and miRNAs levels in the exposed macrophages. In conclusion, hyperglycemia is associated with increased miR-223, miR-92a, miR-486 levels in HDL, which discriminate between ACS and SA patients. Exposure of human macrophages to ACS compared to SA sera determines the upregulation of Drosha, DGCR8 and Dicer expression and the increase of selected miRNAs production, the effect being augmented by an increased glucose concentration.

MiR-486 and miR-92a Identified in Circulating HDL Discriminate between Stable and Vulnerable Coronary Artery Disease Patients.

Small non-coding microRNAs (miRNAs) are implicated in gene regulation, including those involved in coronary artery disease (CAD). Our aim was to identify whether specific serum miRNAs present in the circulating lipoproteins (Lp) are associated with stable or vulnerable CAD patients. A cardiovascular disease-focused screening array was used to assess miRNAs distribution in sera collected from 95 CAD patients: 30 with stable angina (SA), 39 with unstable angina (UA), 26 at one month after myocardial infarction (MI) and 16 healthy control subjects. We found that miR-486, miR-92a and miR-122 presented the highest expression in CAD sera. These miRNA together with miR-125a, miR-146a and miR-33a were further individually analyzed by TaqMan assays. The results were consistent with PCR-array screening data that all of these miRNAs were significantly increased in CAD patients compared to controls. Using a binary logistic regression model, we established that miR-486 and miR-92a in association with some high-density lipoprotein (HDL) components can designate vulnerable CAD patients. Further, all classes of Lp were isolated from sera by density gradient ultracentrifugation. Analysis of the selected miRNAs in each Lp class showed that they were associated mainly with HDL, miR-486 and miR-92a having the highest levels. In UA and MI patients, miR-486 prevailed in HDL2, while miR-92a prevailed in HDL3, and their levels discriminate between stable and vulnerable CAD patients. We identified two circulating miRNAs that in association with some lipid metabolism biomarkers can be used as an additional tool to designate vulnerable CAD patients.

Hyperlipidemia-induced hepatic and small intestine ER stress and decreased paraoxonase 1 expression and activity is associated with HDL dysfunction in Syrian hamsters.

SCOPE: We aimed at investigating the mechanisms linking hyperlipidemia (HL) with dysfunctional HDL and its main antioxidant enzyme, paraoxonase1 (PON1). PON1 expression and activity was determined in the small intestine, liver, and sera of normal and HL hamsters and associated with the ER stress (ERS) and the development of aortic valve lesions. METHODS AND RESULTS: Male Golden Syrian hamsters were fed standard chow (N) or standard diet with 3% cholesterol and 15% butter for 16 weeks. All hamsters on fat diet developed HL, 50% also hyperglycemia (HLHG) and a fourfold increased homeostasis model assessment of insuline resistance. PON1 expression was reduced in the small intestine and liver (N > HL > HLHG) along with the increased extent of ERS, oxidized lipids, and decreased expression of liver X receptors beta (LXRß) in the small intestine, peroxisome proliferator-activated receptor-γ (PPARγ) in the liver, and of the glucose transporter 4 in the myocardium. Serum PON1 levels decreased along with the increase of oxidized LDL and lesion areas of the aortic valves (N > HL > HLHG). CONCLUSION: The fat diet activates the ERS and oxidative stress, decreases LXRß, PPARγ, and PON1 in the small intestine, liver, and sera of all HL animals, in parallel with the appearance of atherosclerotic lesions in the aortic valves.