Radiolabeled multi-layered coated gold nanoparticles as potential biocompatible PET/SPECT tracers.

The demand for tailored, disease-adapted, and easily accessible radiopharmaceuticals is one of the most persistent challenges in nuclear imaging precision medicine. The aim of this work was to develop two multimodal radiotracers applicable for both SPECT and PET techniques, which consist of a gold nanoparticle core, a shell involved in radioisotope entrapment, peripherally placed targeting molecules, and biocompatibilizing polymeric sequences. Shell decoration with glucosamine units located in sterically hindered molecular environments is expected to result in nanoparticle accumulation in high-glucose-consuming areas. Gold cores were synthesized using the Turkevich method, followed by citrate substitution with linear PEG α,ω-functionalized with thiol and amine groups. The free amine groups facilitated the binding of branched polyethyleneimine through an epoxy ring-opening reaction by using PEG α,ω-diglycidyl ether as a linker. Afterwards, the glucose-PEG-epoxy prepolymer has been grafted onto the surface of AuPEG-PEI conjugates. Finally, the AuPEG-PEI-GA conjugates were radiolabeled with 99mTc or 68Ga. Instant thin-layer chromatography was used to evaluate the radiolabeling yield. The biocompatibility of non-labeled and 99mTc or 68Ga labeled nanoparticles was assessed on normal fibroblasts. The 99mTc complexes remained stable for over 22 hours, while the 68Ga containing ones revealed a slight decrease in stability after 1 hour.

Enhancing the Antioxidant Potential of Weissella confusa PP29 Probiotic Media through Incorporation of Hibiscus sabdariffa L. Anthocyanin Extract.

Lactic acid bacteria (LAB) produce important metabolites during fermentation processes, such as exopolysaccharides (EPS), which represent powerful natural antioxidants. On the other hand, H. sabdariffa L. anthocyanin extracts protect LAB and support their development. This study uncovers for the first time, the antioxidant profile of Weissella confusa PP29 probiotic media and focuses on elevating its impressive antioxidant attributes by synergistically integrating H. sabdariffa L. anthocyanin extract. The multifaceted potential of this innovative approach is explored and the results are remarkable, allowing us to understand the protective capacity of the fermented product on the intestinal mucosa. The total phenolic content was much lower at the end of the fermentation process compared to the initial amount, confirming their LAB processing. The DPPH radical scavenging and FRAP of the fermented products were higher compared to ascorbic acid and antioxidant extracts, while superoxide anion radical scavenging and lipid peroxidation inhibitory activity were comparable to that of ascorbic acid. The antioxidant properties of the fermented products were correlated with the initial inoculum and anthocyanin concentrations. All these properties were preserved for 6 months, demonstrating the promising efficacy of this enriched medium, underlining its potential as a complex functional food with enhanced health benefits.

Multifunctional Hydrogels Based on Cellulose and Modified Lignin for Advanced Wounds Management.

Considering the complex process of wound healing, it is expected that an optimal wound dressing should be able to overcome the multiple obstacles that can be encountered in the wound healing process. An ideal dressing should be biocompatible, biodegradable and able to maintain moisture, as well as allow the removal of exudate, have antibacterial properties, protect the wound from pathogens and promote wound healing. Starting from this desideratum, we intended to design a multifunctional hydrogel that would present good biocompatibility, the ability to provide a favorable environment for wound healing, antibacterial properties, and also, the capacity to release drugs in a controlled manner. In the preparation of hydrogels, two natural polymers were used, cellulose (C) and chemically modified lignin (LE), which were chemically cross-linked in the presence of epichlorohydrin. The structural and morphological characterization of CLE hydrogels was performed by ATR-FTIR spectroscopy and scanning electron microscopy (SEM), respectively. In addition, the degree of swelling of CLE hydrogels, the incorporation/release kinetics of procaine hydrochloride (PrHy), and their cytotoxicity and antibacterial properties were investigated. The rheological characterization, mechanical properties and mucoadhesion assessment completed the study of CLE hydrogels. The obtained results show that CLE hydrogels have an increased degree of swelling compared to cellulose-based hydrogel, a better capacity to encapsulate PrHy and to control the release of the drug, as well as antibacterial properties and improved mucoadhesion. All these characteristics highlight that the addition of LE to the cellulose matrix has a positive impact on the properties of CLE hydrogels, confirming that these hydrogels can be considered as potential candidates for applications as oral wound dressings.

Strategy Based on Michael Addition Reaction for the Development of Bioinspired Multilayered and Multiphasic 3D Constructs.

The high incidence of osteochondral defects has increased the interest in the development of improved repairing alternatives, with tissue engineering being considered a promising approach. The hierarchical, complex structure of osteochondral tissue requires the design of a biomimetic multilayered scaffold. Here, a multilayered and multiphasic 3D macroporous structure was achieved at subzero temperature by the Michael addition reaction of amino functionalities of collagen with acryloyl groups of a bifunctionalized poly(ε-caprolactone). This green approach has been successfully applied to crosslink layers of different composition, both for their efficient sequential formation and connection. Polyethylenimine functionalized nano-hydroxyapatite (nHApLPEI) was added to the bottom layer. The resulting hybrid cryogels were characterized by morphology, equilibrium swelling ratios, compressive strength analysis, and MTS assay. They presented good stability, integrity, and biocompatibility. The results revealed that the properties of the prepared constructs may be tuned by varying the composition, number, and thickness of the layers. The Young modulus values were between 3.5 ± 0.02 and 10.5 ± 0.6 kPa for the component layers, while for the multilayered structures they were more than 7.3 ± 0.2 kPa. The equilibrium swelling ratio varied between 4.6 and 14.2, with a value of ~10.5 for the trilayered structure, correlated with the mean pore sizes (74-230 µm).

New Smart Bioactive and Biomimetic Chitosan-Based Hydrogels for Wounds Care Management.

Wound management represents a continuous challenge for health systems worldwide, considering the growing incidence of wound-related comorbidities, such as diabetes, high blood pressure, obesity, and autoimmune diseases. In this context, hydrogels are considered viable options since they mimic the skin structure and promote autolysis and growth factor synthesis. Unfortunately, hydrogels are associated with several drawbacks, such as low mechanical strength and the potential toxicity of byproducts released after crosslinking reactions. To overcome these aspects, in this study new smart chitosan (CS)-based hydrogels were developed, using oxidized chitosan (oxCS) and hyaluronic acid (oxHA) as nontoxic crosslinkers. Three active product ingredients (APIs) (fusidic acid, allantoin, and coenzyme Q10), with proven biological effects, were considered for inclusion in the 3D polymer matrix. Therefore, six API-CS-oxCS/oxHA hydrogels were obtained. The presence of dynamic imino bonds in the hydrogels' structure, which supports their self-healing and self-adapting properties, was confirmed by spectral methods. The hydrogels were characterized by SEM, swelling degree, pH, and the internal organization of the 3D matrix was studied by rheological behavior. Moreover, the cytotoxicity degree and the antimicrobial effects were also investigated. In conclusion, the developed API-CS-oxCS/oxHA hydrogels have real potential as smart materials in wound management, based on their self-healing and self-adapting properties, as well as on the benefits of APIs.

Dextran Formulations as Effective Delivery Systems of Therapeutic Agents.

Dextran is by far one of the most interesting non-toxic, bio-compatible macromolecules, an exopolysaccharide biosynthesized by lactic acid bacteria. It has been extensively used as a major component in many types of drug-delivery systems (DDS), which can be submitted to the next in-vivo testing stages, and may be proposed for clinical trials or pharmaceutical use approval. An important aspect to consider in order to maintain high DDS' biocompatibility is the use of dextran obtained by fermentation processes and with a minimum chemical modification degree. By performing chemical modifications, artefacts can appear in the dextran spatial structure that can lead to decreased biocompatibility or even cytotoxicity. The present review aims to systematize DDS depending on the dextran type used and the biologically active compounds transported, in order to obtain desired therapeutic effects. So far, pure dextran and modified dextran such as acetalated, oxidised, carboxymethyl, diethylaminoethyl-dextran and dextran sulphate sodium, were used to develop several DDSs: microspheres, microparticles, nanoparticles, nanodroplets, liposomes, micelles and nanomicelles, hydrogels, films, nanowires, bio-conjugates, medical adhesives and others. The DDS are critically presented by structures, biocompatibility, drugs loaded and therapeutic points of view in order to highlight future therapeutic perspectives.

Enzymatically-Crosslinked Gelatin Hydrogels with Nanostructured Architecture and Self-Healing Performance for Potential Use as Wound Dressings.

Development of natural protein-based hydrogels with self-healing performance and tunable physical properties has attracted increased attention owing to their wide potential not only in the pharmaceutical field, but also in wounds management. This work reports the development of a versatile hydrogel based on enzymatically-crosslinked gelatin and nanogels loaded with amoxicillin (Amox), an antibiotic used in wound infections. The transglutaminase (TGase)-crosslinked hydrogels and encapsulating nanogels were formed rapidly through enzymatic crosslinking and self-assembly interactions in mild conditions. The nanogels formed through the self-assemble of maleoyl-chitosan (MAC5) and polyaspartic acid (PAS) may have positive influence on the self-healing capacity and drug distribution within the hydrogel network through the interactions established between gelatin and gel-like nanocarriers. The physicochemical properties of the enzymatically-crosslinked hydrogels, such as internal structure, swelling and degradation behavior, were studied. In addition, the Amox release studies indicated a rapid release when the pH of the medium decreased, which represents a favorable characteristic for use in the healing of infected wounds. It was further observed through the in vitro and in vivo biocompatibility assays that the optimized scaffolds have great potential to be used as wound dressings.

Carbonic Anhydrase inhibitors bearing organotelluride moieties as novel agents for antitumor therapy.

Solid tumors are mainly characterized by a specific hypoxic microenvironment which makes them particularly challenging to treat. The Carbonic Anhydrase IX (CA IX) is one of the major enzymes implicated in the regulation and maintaining of such conditions and therefore its targeting represents a winning approach in recent tumor targeted therapy. In our search for an innovative combination therapy, we attained the synthesis of selective CA IX inhibitors which are also used for cell specific delivery of cytotoxic organotellurium scaffolds. We investigated compounds 5b, 7b and 7c for their redox properties by means of radical species scavenging and lipid peroxidation inhibitory capacity, as well as intracellular (reactive oxygen species) ROS production in both normal and cancer cell lines. Subsequently, compounds were evaluated as possible free radical generators by ESR spectrometry showing to cause or promote the formation of free radicals. These results accounted for a novel, potent, and selective CA IX inhibitor (i.e. 7c, Ki = 32 nM) with high cytotoxic effect against malignant melanoma (MeWo) and hepatocellular carcinoma (HepG2) cells over normal fibroblasts (NHDF) through ROS-independent mechanisms. The preliminary data gives support to employ organotellurium moieties as useful pharmacological tools for further development in the oncological field.

Simultaneous Determination of Glibenclamide and Silymarin Released from Chitosan Microparticles by HPLC-ESI-MS Technique: Method Development and Validation.

The study aim was to develop and validate a high-performance liquid chromatography-electrospray ionization mass spectrometry (HPLC-ESI-MS) method to simultaneously determine glibenclamide (Gli) and silymarin (Sil) released from chitosan (CS) microparticles in aqueous solutions. The CS microparticles were synthesized using an ionic gelation method, and their morphology, swelling degree, encapsulation efficiency and active substance release were investigated. Gli and Sil were loaded in different concentrations, and their identification and quantification were performed using the HPLC-ESI-MS method, which was further validated. The drugs' characteristic m/z was found in the higher intensity of retention time (Rt) (Gli, 8.909 min; Sil A, 5.41 min; and Sil B, 5.66 min). The method selectivity and precision are very good, and the blank solution proved no interference. The linearity of the answer function is very good for Sil A (R2 = 1), Sil B (R2 = 0.9998) and Gli (R2 = 0.9991). For Gli, we obtained a limit of detection (LOD) = 0.038 mg/mL and limit of quantification (LOQ) = 1.275 mg/mL; for Sil A, a LOD = 0.285 mg/mL and LOQ = 0.95 mg/mL; and for Sil B, a LOD = 0.045 mg/mL and LOQ = 0.15 mg/mL. A high-resolution HPLC-ESI-MS method was developed and validated, which allowed the simultaneous determination of Gli and Sil loaded in CS microparticles, in a concentration range of 0.025-1 mg/mL.

One-Step Preparation of Carboxymethyl Cellulose-Phytic Acid Hydrogels with Potential for Biomedical Applications.

Hydrogels based on natural, biodegradable materials have gained considerable interest in the medical field due to their improved drug delivery profiles and tissue-mimicking architecture. In this regard, this study was devoted to the preparation and characterization of new physically crosslinked hydrogels based on carboxymethyl cellulose and an unconventional crosslinking agent, phytic acid. Phytic acid, in addition to its antioxidant and antibacterial effects, can improve the biological properties and stability of gels, without adding toxicity. Fourier transform infrared (FTIR) spectroscopy, rheological studies and thermal analysis confirmed the hydrogel formation. The influence of the ratio between the cellulose derivative and the crosslinker upon the morphological structure and water uptake was evidenced by scanning electron microscopy (SEM) and swelling measurements in simulated body fluids. Furthermore, procaine was entrapped within the hydrogels and used as a model drug for in vitro studies, which highlighted the dependence of the drug release on the phytic acid content of the matrix. The materials demonstrated antibacterial effects against Escherichia coli and Staphylococcus aureus bacteria. The biocompatibility was assessed on fibroblast cells, and according to our results, hydrogels can improve cell viability highlighting the potential of these systems as therapeutic scaffolds for skin tissue engineering.

Microvesicles and Microvesicle-Associated microRNAs Reflect Glioblastoma Regression: Microvesicle-Associated miR-625-5p Has Biomarker Potential.

Glioblastoma (GB) is the most aggressive and recurrent form of brain cancer in adults. We hypothesized that the identification of biomarkers such as certain microRNAs (miRNAs) and the circulating microvesicles (MVs) that transport them could be key to establishing GB progression, recurrence and therapeutic response. For this purpose, circulating MVs were isolated from the plasma of GB patients (before and after surgery) and of healthy subjects and characterized by flow cytometry. OpenArray profiling and the individual quantification of selected miRNAs in plasma and MVs was performed, followed by target genes' prediction and in silico survival analysis. It was found that MVs' parameters (number, EGFRvIII and EpCAM) decreased after the surgical resection of GB tumors, but the inter-patient variability was high. The expression of miR-106b-5p, miR-486-3p, miR-766-3p and miR-30d-5p in GB patients' MVs was restored to control-like levels after surgery: miR-106b-5p, miR-486-3p and miR-766-3p were upregulated, while miR-30d-5p levels were downregulated after surgical resection. MiR-625-5p was only identified in MVs isolated from GB patients before surgery and was not detected in plasma. Target prediction and pathway analysis showed that the selected miRNAs regulate genes involved in cancer pathways, including glioma. In conclusion, miR-625-5p shows potential as a biomarker for GB regression or recurrence, but further in-depth studies are needed.

Quantification of Low Amounts of Zoledronic Acid by HPLC-ESI-MS Analysis: Method Development and Validation.

Zoledronic acid (ZA) is used in the treatment of various bone pathologies, but it forms complexes with calcium ions present in body fluids, decreasing ZA bioavailability. Thereby, the study first describes the identification of ZA-calcium complexes that form in calcium-rich environments, in order to establish the bioavailable ZA concentration. Then, a new method for quantification of low ZA amounts in milieus that mimics in vivo conditions by using simulated body fluid and calcium sulfate hemihydrate was described. Almost all analytical methods of ZA quantification described in the literature require compound derivatization. At very low concentrations, derivatization is prone to analyte loss, therefore compromising the analytical results. In our study, we avoided ZA derivatization by using a high-performance liquid chromatography and electrospray ionization mass spectrometry (HPLC-ESI-MS) system, conducting the investigation based on the fragmentation mass extracted ion chromatograms specific to the ZA protonated form. The method was validated by selectivity, precision, accuracy, linearity, signal to noise ratio, and limit of detection and limit of quantification calculation. Experimentally, this method can detect ranges of 0.1-0.5 ng/mL and precisely quantify ZA concentrations as low as 0.1 ng/mL. This method could provide the basis for quantifying low amounts of ZA in the blood during long-term administration.

Design, preparation and in vitro characterization of biomimetic and bioactive chitosan/polyethylene oxide based nanofibers as wound dressings.

Chitosan-based nanofibers (CS-NFs) are excellent artificial extracellular matrices (ECMs) due to the resemblance of CS with the glycosaminoglycans of the natural ECMs. Despite this excellent feature, the poor electrospinnability and mechanical properties of CS are responsible for important limitations in respect to its biomedical applications. To improve the CS's physico-chemical properties, new bioactive and biomimetic CS-NFs were formulated with polyethylene oxide (PEO), having incorporated different active components (ACs) with important beneficial effects for healing. Manuka honey (trophic and antimicrobial effects), propolis (antimicrobial effects), Calendula officinalis infusion (antioxidant effect, reepithelialization stimulating agent), insulin (trophic effect), and L-arginine (angiogenic effect) were selected as ACs. SEM morphology analysis revealed well-alignment, unidirectional arrays, with small diameters, no beads, and smooth surfaces for developed CS_PEO-ACs NFs. The developed NFs showed good biodegradability (NFs mats lost up to 60% of their initial weight in PBS), increased hemocompatibility (hemolytic index less than 4%), and a reduced cytotoxicity degree (cell viability degree more than 90%). In addition, significant antioxidant and antimicrobial effects were noted for the developed NFs which make them suitable for chronic wounds, due to the role of oxidative stress and infection risk in delaying normal wound healing. The most suitable for wound healing applications seems to be CS_PEO@P_C which showed an improved hemolysis index (2.92 ± 0.16%), is non-toxic (cell viability degree more than 97%), and has also significant radical scavenging effect (DPPH inhibition more than 65%). In addition, CS_PEO@P_C presents increased antimicrobial effects, more noticeably for Staphylococcus aureus strain, which is a key feature in preventing wound infection and delaying the healing process. It can be concluded that the developed CS/PEO-ACs NFs are very promising biomaterials for wound care, especially CS_PEO@P_C.

The Multifaceted Role of Extracellular Vesicles in Glioblastoma: microRNA Nanocarriers for Disease Progression and Gene Therapy.

Glioblastoma (GB) is the most aggressive form of brain cancer in adults, characterized by poor survival rates and lack of effective therapies. MicroRNAs (miRNAs) are small, non-coding RNAs that regulate gene expression post-transcriptionally through specific pairing with target messenger RNAs (mRNAs). Extracellular vesicles (EVs), a heterogeneous group of cell-derived vesicles, transport miRNAs, mRNAs and intracellular proteins, and have been shown to promote horizontal malignancy into adjacent tissue, as well as resistance to conventional therapies. Furthermore, GB-derived EVs have distinct miRNA contents and are able to penetrate the blood-brain barrier. Numerous studies have attempted to identify EV-associated miRNA biomarkers in serum/plasma and cerebrospinal fluid, but their collective findings fail to identify reliable biomarkers that can be applied in clinical settings. However, EVs carrying specific miRNAs or miRNA inhibitors have great potential as therapeutic nanotools in GB, and several studies have investigated this possibility on in vitro and in vivo models. In this review, we discuss the role of EVs and their miRNA content in GB progression and resistance to therapy, with emphasis on their potential as diagnostic, prognostic and disease monitoring biomarkers and as nanocarriers for gene therapy.

New Insights on Hemp Oil Enriched in Cannabidiol: Decarboxylation, Antioxidant Properties and In Vitro Anticancer Effect.

This study aimed to obtain and characterize extracted hemp oil enriched in cannabidiol (CBD) by decarboxylation of cannabidiolic acid (CBDA) and to give new insights into its antioxidant and anticancer effects. Optimization of CBDA decarboxylation in hemp oil was performed, and CBD and CBDA contents and purities were determined by flash chromatography, 1H- and 13C-NMR. The antioxidant properties of CBD-enriched oil were investigated by Fe2+ chelating activity, Fe3+ reducing antioxidant power assay, O2●- scavenging activity, HO● scavenging ability and lipid peroxidation inhibitory assay, and its cytotoxicity, apoptosis- and oxidative stress-inducing effects on NHDF, MeWo, HeLa, HepG2 and HOS cells were determined. The CBD concentration in hemp oil was increased by CBDA soft decarboxylation optimized at 90 °C, for 1 h and the resulting oil was capable of reducing iron, scavenging free radicals and inhibiting lipid peroxidation in cell-free oxidative conditions. CBD-enriched oil promoted NHDF proliferation at up to 15 µg CBD/mL, while inducing apoptosis and ROS production and modulating antioxidant enzymes' gene expression in cancer cells, being selective for osteosarcoma cells, and induced apoptosis by p53- and ROS-independent mechanisms. CBD-enriched hemp oil demonstrated antioxidant properties in oxidative conditions and promoted normal fibroblasts' proliferation, while inducing apoptosis and ROS production in cancer cells.

New Hyaluronic Acid/Polyethylene Oxide-Based Electrospun Nanofibers: Design, Characterization and In Vitro Biological Evaluation.

Natural compounds have been used as wound-healing promoters and are also present in today's clinical proceedings. In this research, different natural active components such as propolis, Manuka honey, insulin, L-arginine, and Calendula officinalis infusion were included into hyaluronic acid/poly(ethylene)oxide-based electrospun nanofiber membranes to design innovative wound-dressing biomaterials. Morphology and average fiber diameter were analyzed by scanning electron microscopy. Chemical composition was proved by Fourier transform infrared spectroscopy, which indicated successful incorporation of the active components. The nanofiber membranes with propolis and Calendula officinalis showed best antioxidant activity, cytocompatibility, and antimicrobial properties against pathogen strains Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa and had an average diameter of 217 ± 19 nm with smooth surface aspect. Water vapor transmission rate was in agreement with the range suitable for preventing infections or wound dehydration (~5000 g/m2 24 h). Therefore, the developed hyaluronic acid/poly(ethylene)oxide nanofibers with additional natural components showed favorable features for clinical use as wound dressings.

SI-ATRP Decoration of Magnetic Nanoparticles with PHEMA and Post-Polymerization Modification with Folic Acid for Tumor Cells' Specific Targeting.

Targeted nanocarriers could reach new levels of drug delivery, bringing new tools for personalized medicine. It is known that cancer cells overexpress folate receptors on the cell surface compared to healthy cells, which could be used to create new nanocarriers with specific targeting moiety. In addition, magnetic nanoparticles can be guided under the influence of an external magnetic field in different areas of the body, allowing their precise localization. The main purpose of this paper was to decorate the surface of magnetic nanoparticles with poly(2-hydroxyethyl methacrylate) (PHEMA) by surface-initiated atomic transfer radical polymerization (SI-ATRP) followed by covalent bonding of folic acid to side groups of the polymer to create a high specificity magnetic nanocarrier with increased internalization capacity in tumor cells. The biocompatibility of the nanocarriers was demonstrated by testing them on the NHDF cell line and folate-dependent internalization capacity was tested on three tumor cell lines: MCF-7, HeLa and HepG2. It has also been shown that a higher concentration of folic acid covalently bound to the polymer leads to a higher internalization in tumor cells compared to healthy cells. Last but not least, magnetic resonance imaging was used to highlight the magnetic properties of the functionalized nanoparticles obtained.

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.

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.