Synthesis, Characterisation, and In Vitro Evaluation of Biocompatibility, Antibacterial and Antitumor Activity of Imidazolium Ionic Liquids.

In recent decades, ionic liquids (ILs) have garnered research interest for their noteworthy properties, such as thermal stability, low or no flammability, and negligible vapour pressure. Moreover, their tunability offers limitless opportunities to design ILs with properties suitable for applications in many industrial fields. This study aims to synthetise two series of methylimidazolium ILs bearing long alkyl chain in their cations (C9, C10, C12, C14, C16, C18, C20) and with tetrafluoroborate (BF4) and the 1,3-dimethyl-5-sulfoisophthalate (DMSIP) as counter ions. The ILs were characterised using 1H-NMR and MALDI-TOF, and their thermal behaviour was investigated through DSC and TGA. Additionally, the antimicrobial, anticancer, and cytotoxic activities of the ILs were analysed. Moreover, the most promising ILs were incorporated at different concentrations (0.5, 1, 5 wt%) into polyvinyl chloride (PVC) by solvent casting to obtain antimicrobial blend films. The thermal properties and stability of the resulting PVC/IL films, along with their hydrophobicity/hydrophilicity, IL surface distribution, and release, were studied using DSC and TGA, contact angle (CA), SEM, and UV-vis spectrometry, respectively. Furthermore, the antimicrobial and cytotoxic properties of blends were analysed. The in vitro results demonstrated that the antimicrobial and antitumor activities of pure ILs against t Listeria monocytogenes, Escherichia coli, Pseudomonas fluorescens strains, and the breast cancer cell line (MCF7), respectively, were mainly dependent on their structure. These activities were higher in the series containing the BF4 anion and increased with the increase in the methylimidazolium cation alkyl chain length. However, the elongation of the alkyl chain beyond C16 induced a decrease in antimicrobial activity, indicating a cut-off effect. A similar trend was also observed in terms of in vitro biocompatibility. The loading of both the series of ILs into the PVC matrix did not affect the thermal stability of PVC blend films. However, their Tonset decreased with increased IL concentration and alkyl chain length. Similarly, both the series of PVC/IL films became more hydrophilic with increasing IL concentration and alkyl chain. The loading of ILs at 5% concentration led to considerable IL accumulation on the blend film surfaces (as observed in SEM images) and, subsequently, their higher release. The biocompatibility assessment with healthy human dermal fibroblast (HDF) cells and the investigation of antitumoral properties unveiled promising pharmacological characteristics. These findings provide strong support for the potential utilisation of ILs in biomedical applications, especially in the context of cancer therapy and as antibacterial agents to address the challenge of antibiotic resistance. Furthermore, the unique properties of the PVC/IL films make them versatile materials for advancing healthcare technologies, from drug delivery to tissue engineering and antimicrobial coatings to diagnostic devices.

BDNF- and VEGF-Responsive Stimulus to an NGF Mimic Cyclic Peptide with Copper Ionophore Capability and Ctr1/CCS-Driven Signaling.

Neurotrophins are a family of growth factors that play a key role in the development and regulation of the functioning of the central nervous system. Their use as drugs is made difficult by their poor stability, cellular permeability, and side effects. Continuing our effort to use peptides that mimic the neurotrophic growth factor (NGF), the family model protein, and specifically the N-terminus of the protein, here we report on the spectroscopic characterization and resistance to hydrolysis of the 14-membered cyclic peptide reproducing the N-terminus sequence (SSSHPIFHRGEFSV (c-NGF(1-14)). Far-UV CD spectra and a computational study show that this peptide has a rigid conformation and left-handed chirality typical of polyproline II that favors its interaction with the D5 domain of the NGF receptor TrkA. c-NGF(1-14) is able to bind Cu2+ with good affinity; the resulting complexes have been characterized by potentiometric and spectroscopic measurements. Experiments on PC12 cells show that c-NGF(1-14) acts as an ionophore, influencing the degree and the localization of both the membrane transporter (Ctr1) and the copper intracellular transporter (CCS). c-NGF(1-14) induces PC12 differentiation, mimics the protein in TrkA phosphorylation, and activates the kinase cascade, inducing Erk1/2 phosphorylation. c-NGF(1-14) biological activities are enhanced when the peptide interacts with Cu2+ even with the submicromolar quantities present in the culture media as demonstrated by ICP-OES measurements. Finally, c-NGF(1-14) and Cu2+ concur to activate the cAMP response element-binding protein CREB that, in turn, induces the brain-derived neurotrophic factor (BDNF) and the vascular endothelial growth factor (VEGF) release.

W/O/W Microemulsions for Nasal Delivery of Hydrophilic Compounds: A Preliminary Study.

The administration of hydrophilic therapeutics has always been a great challenge because of their low bioavailability after administration. For this purpose, W/O/W microemulsion resulted to be a potential successful strategy for the delivery of hydrophilic compounds, interesting for the nasal mucosal therapy. Herein, an optimized biphasic W/O microemulsion was designed, through a preliminary screening, and it was inverted in a triphasic W/O/W microemulsion, intended for the nasal administration. In order to enhance the mucosal retention, surface modification of the biphasic W/O microemulsion was performed adding didodecyldimethylammonium bromide, and then converting the system into a cationic triphasic W/O/W microemulsion. The developed samples were characterized in terms of droplet size, polydispersity, zeta potential, pH and osmolality. The physical long-term stability was analyzed storing samples at accelerated conditions (40 ± 2 °C and 75 ± 5 % RH) for 6 months in a constant climate chamber, following ICH guidelines Q1A (R2). In order to verify the potential retention on the nasal mucosa, the two triphasic systems were analyzed in terms of mucoadhesive properties, measuring the in vitro interaction with mucin over time. Furthermore, fluorescein sodium salt was selected as a model hydrophilic drug to be encapsulated into the inner core of the two triphasic W/O/W microemulsions, and its release was analyzed compared to the free probe solution. The cytocompatibility of the two platforms was assessed on two cell lines, human fibroblasts HFF1 and Calu-3 cell lines, chosen as pre-clinical models for nasal and bronchial/tracheal airway epithelium.

The role of sperm-specific glyceraldehyde-3-phosphate dehydrogenase in the development of pathologies-from asthenozoospermia to carcinogenesis.

The review considers various aspects of the influence of the glycolytic enzyme, sperm-specific glyceraldehyde-3-phosphate dehydrogenase (GAPDS) on the energy metabolism of spermatozoa and on the occurrence of several pathologies both in spermatozoa and in other cells. GAPDS is a unique enzyme normally found only in mammalian spermatozoa. GAPDS provides movement of the sperm flagellum through the ATP formation in glycolytic reactions. Oxidation of cysteine residues in GAPDS results in inactivation of the enzyme and decreases sperm motility. In particular, reduced sperm motility in diabetes can be associated with GAPDS oxidation by superoxide anion produced during glycation reactions. Mutations in GAPDS gene lead in the loss of motility, and in some cases, disrupts the formation of the structural elements of the sperm flagellum, in which the enzyme incorporates during spermiogenesis. GAPDS activation can be used to increase the spermatozoa fertility, and inhibitors of this enzyme are being tried as contraceptives. A truncated GAPDS lacking the N-terminal fragment of 72 amino acids that attaches the enzyme to the sperm flagellum was found in melanoma cell lines and then in specimens of melanoma and other tumors. Simultaneous production of the somatic form of GAPDH and sperm-specific GAPDS in cancer cells leads to a reorganization of their energy metabolism, which is accompanied by a change in the efficiency of metastasis of certain forms of cancer. Issues related to the use of GAPDS for the diagnosis of cancer, as well as the possibility of regulating the activity of this enzyme to prevent metastasis, are discussed.

Copper(II) Complexes with Carnosine Conjugates of Hyaluronic Acids at Different Dipeptide Loading Percentages Behave as Multiple SOD Mimics and Stimulate Nrf2 Translocation and Antioxidant Response in In Vitro Inflammatory Model.

A series of copper(II) complexes with the formula [Cu2+Hy(x)Car%] varying the molecular weight (MW) of Hyaluronic acid (Hy, x = 200 or 700 kDa) conjugated with carnosine (Car) present at different loading were synthesized and characterized via different spectroscopic techniques. The metal complexes behaved as Cu, Zn-superoxide dismutase (SOD1) mimics and showed some of the most efficient reaction rate values produced using a synthetic and water-soluble copper(II)-based SOD mimic reported to date. The increase in the percentage of Car moieties parallels the enhancement of the I50 value determined via the indirect method of Fridovich. The presence of the non-functionalized Hy OH groups favors the scavenger activity of the copper(II) complexes with HyCar, recalling similar behavior previously found for the copper(II) complexes with Car conjugated using ß-cyclodextrin or trehalose. In keeping with the new abilities of SOD1 to activate protective agents against oxidative stress in rheumatoid arthritis and osteoarthritis diseases, Cu2+ interaction with HyCar promotes the nuclear translocation of erythroid 2-related factor that regulates the expressions of target genes, including Heme-Oxigenase-1, thus stimulating an antioxidant response in osteoblasts subjected to an inflammatory/oxidative insult.

In silico analysis of the solute carrier (SLC) family in cancer indicates a link among DNA methylation, metabolic adaptation, drug response, and immune reactivity.

Introduction: The oncogenic transformation is driven by genetic and epigenetic alterations influencing cancer cell fate. These alterations also result in metabolic reprogramming by modulating the expression of membrane Solute Carrier (SLC) transporters involved in biomolecules trafficking. SLCs act as tumor suppressors or promoters influencing cancer methylome, tumor growth, immune-escape, and chemoresistance. Methods: This in silico study aimed to identify the deregulated SLCs in various tumor types compared to normal tissues by analyzing the TCGA Target GTEx dataset. Furthermore, the relationship between SLCs expression and the most relevant tumor features was tackled along with their genetic regulation mediated by DNA methylation. Results: We identified 62 differentially expressed SLCs, including the downregulated SLC25A27 and SLC17A7, as well as the upregulated SLC27A2 and SLC12A8. Notably, SLC4A4 and SLC7A11 expression was associated with favorable and unfavorable outcome, respectively. Moreover, SLC6A14, SLC34A2, and SLC1A2 were linked to tumor immune responsiveness. Interestingly, SLC24A5 and SLC45A2 positively correlated with anti-MEK and anti-RAF sensitivity. The expression of relevant SLCs was correlated with hypo- and hyper-methylation of promoter and body region, showing an established DNA methylation pattern. Noteworthy, the positive association of cg06690548 (SLC7A11) methylation with cancer outcome suggests the independent predictive role of DNA methylation at a single nucleotide resolution. Discussion: Although our in silico overview revealed a wide heterogeneity depending on different SLCs functions and tumor types, we identified key SLCs and pointed out the role of DNA methylation as regulatory mechanism of their expression. Overall, these findings deserve further studies to identify novel cancer biomarkers and promising therapeutic targets.

Natural Compounds and Glutathione: Beyond Mere Antioxidants.

The tripeptide glutathione plays important roles in many cell processes, including differentiation, proliferation, and apoptosis; in fact, disorders in glutathione homeostasis are involved both in the etiology and in the progression of several human diseases, including cancer. Natural compounds have been found to modulate glutathione levels and function beyond their role as mere antioxidants. For example, certain compounds can upregulate the expression of glutathione-related enzymes, increase the availability of cysteine, the limiting amino acid for glutathione synthesis, or directly interact with glutathione and modulate its function. These compounds may have therapeutic potential in a variety of disease states where glutathione dysregulation is a contributing factor. On the other hand, flavonoids' potential to deplete glutathione levels could be significant for cancer treatment. Overall, while natural compounds may have potential therapeutic and/or preventive properties and may be able to increase glutathione levels, more research is needed to fully understand their mechanisms of action and their potential benefits for the prevention and treatment of several diseases. In this review, particular emphasis will be placed on phytochemical compounds belonging to the class of polyphenols, terpenoids, and glucosinolates that have an impact on glutathione-related processes, both in physiological and pathological conditions. These classes of secondary metabolites represent the most food-derived bioactive compounds that have been intensively explored and studied in the last few decades.

Prospects for the Use of Metal-Based Nanoparticles as Adjuvants for Local Cancer Immunotherapy.

Immunotherapy is among the most effective approaches for treating cancer. One of the key aspects for successful immunotherapy is to achieve a strong and stable antitumor immune response. Modern immune checkpoint therapy demonstrates that cancer can be defeated. However, it also points out the weaknesses of immunotherapy, as not all tumors respond to therapy and the co-administration of different immunomodulators may be severely limited due to their systemic toxicity. Nevertheless, there is an established way through which to increase the immunogenicity of immunotherapy-by the use of adjuvants. These enhance the immune response without inducing such severe adverse effects. One of the most well-known and studied adjuvant strategies to improve immunotherapy efficacy is the use of metal-based compounds, in more modern implementation-metal-based nanoparticles (MNPs), which are exogenous agents that act as danger signals. Adding innate immune activation to the main action of an immunomodulator makes it capable of eliciting a robust anti-cancer immune response. The use of an adjuvant has the peculiarity of a local administration of the drug, which positively affects its safety. In this review, we will consider the use of MNPs as low-toxicity adjuvants for cancer immunotherapy, which could provide an abscopal effect when administered locally.

Wild Artichoke (Cynara cardunculus subsp. sylvestris, Asteraceae) Leaf Extract: Phenolic Profile and Oxidative Stress Inhibitory Effects on HepG2 Cells.

Cynara cardunculus subsp. sylvestris (wild artichoke) is widespread in Sicily, where it has been used for food and medicinal purposes since ancient times; decoctions of the aerial parts of this plant have been traditionally employed as a remedy for different hepatic diseases. In this study, the phenolic profile and cell-free antioxidant properties of the leaf aqueous extract of wild artichokes grown in Sicily (Italy) were investigated. The crude extract was also tested in cells for its antioxidant characteristics and potential oxidative stress inhibitory effects. To resemble the features of the early stage of mild steatosis in humans, human HepG2 cells treated with free fatty acids at the concentration of 1.5 mM were used. HPLC-DAD analysis revealed the presence of several phenolic acids (caffeoylquinic acids) and flavonoids (luteolin and apigenin derivatives). At the same time, DPPH assay showed a promising antioxidant power (IC50 = 20.04 ± 2.52 µg/mL). Biological investigations showed the safety of the crude extract and its capacity to counteract the injury induced by FFA exposure by restoring cell viability and counteracting oxidative stress through inhibiting reactive oxygen species and lipid peroxidation and increasing thiol-group levels. In addition, the extract increased mRNA expression of some proteins implicated in the antioxidant defense (Nrf2, Gpx, and SOD1) and decreased mRNA levels of inflammatory cytokines (IL-6, TNF-α, and IL-1ß), which were modified by FFA treatment. Results suggest that the total phytocomplex contained in wild artichoke leaves effectively modulates FFA-induced hepatic oxidative stress.

Chemical, Antioxidant and Biological Studies of Brassica incana subsp. raimondoi (Brassicaceae) Leaf Extract.

Brassica incana subsp. raimondoi is an endemic taxon present in a restricted area located on steep limestone cliffs at an altitude of about 500 m a.s.l. in eastern Sicily. In this research, for the first time, studies on the phytochemical profile, the antioxidant properties in cell-free and cell-based systems, the cytotoxicity on normal and cancer cells by 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT) assay, and on Artemia salina Leach, were performed. The total phenolic, flavonoid, and condensed tannin contents of the leaf hydroalcoholic extract were spectrophotometrically determined. Ultra-performance liquid chromatography-tandem mass spectrometer (UPLC-MS/MS) analysis highlighted the presence of several phenolic acids, flavonoids, and carotenoids, while High-Performance Liquid Chromatography with Diode-Array Detection (HPLC-DAD) identified various kaempferol and isorhamnetin derivatives. The extract exhibited different antioxidant properties according to the five in vitro methods used. Cytotoxicity by MTT assay evidenced no impact on normal human fibroblasts (HFF-1) and prostate cancer cells (DU145), and cytotoxicity accompanied by necrotic cell death for colon cancer cells (CaCo-2) and hepatoma cells (HepG2), starting from 100 µg/mL and 500 µg/mL, respectively. No cytotoxic effects were detected by the A. salina lethality bioassay. In the H2O2-induced oxidative stress cell model, the extract counteracted cellular reactive oxygen species (ROS) production and preserved non-protein thiol groups (RSH) affected by H2O2 exposure in HepG2 cells. Results suggest the potential of B. incana subsp. raimondoi as a source of bioactive molecules.

DNA Damage and Apoptosis as In-Vitro Effect Biomarkers of Titanium Dioxide Nanoparticles (TiO2-NPs) and the Food Additive E171 Toxicity in Colon Cancer Cells: HCT-116 and Caco-2.

This study investigated the DNA damage and apoptosis in colon cancer cells HCT-116 and Caco-2 induced by engineered titanium dioxide nanoparticles (TiO2-NPs) (60 nm) and titanium dioxide food additive E171. MTT assays showed that both chemical forms significantly reduced cancer cell viability in a dose-dependent manner. In particular the food additive E171 induced a pronounced inhibitory effect on the growth of HCT-116 and Caco-2 cell lines (E171 IC50: 3.45 mg/L for HTC-116 and 1.88 mg/L Caco-2; TiO2-NPs 60 nm IC50: 41.1 mg/L for HTC-116 and 14.3 mg/L for Caco-2). A low level of genotoxicity was observed in Caco-2 cells, especially when treated with TiO2 60 nm. Western blot analysis showed that HCT116 and Caco-2 treated cells did not overexpress apoptotic markers such as cleaved Caspase 3 and cleaved Parp. Moreover, further analysis by quantitative real-time PCR (qRT-PCR) showed that TiO2-NPs and E171 did not promote the expression of Bax or downregulation of Bcl-2, nor did they increase the Bax/Bcl-2 ratio. The assay data provide clear evidence that TiO2 can cause DNA damage but does not induce apoptosis or decrease long-term cell proliferation. In addition, the results show that E171 has a slightly higher level of cytotoxicity and genotoxicity. This suggests that exposure to E171 may be hazardous to health and that further research on biological effects is needed to promote safer practices in the use of this compound.

Bioinformatic analysis of the LCN2-SLC22A17-MMP9 network in cancer: The role of DNA methylation in the modulation of tumor microenvironment.

Several features of cancer cells such as proliferation, invasion, metastatic spreading, and drug resistance are affected by their interaction with several tumor microenvironment (TME) components, including neutrophil gelatinase-associated lipocalin (NGAL), solute carrier family 22 member 17 (SLC22A17), and matrix metallopeptidase 9 (MMP9). These molecules play a key role in tumor growth, invasion, and iron-dependent metabolism of cancer cells. However, the precise epigenetic mechanisms underlying the gene regulation of Lipocalin 2 (LCN2), SLC22A17, and MMP9 in cancer still remain unclear. To this purpose, computational analysis was performed on TCGA and GTEx datasets to evaluate the expression and DNA methylation status of LCN2, SLC22A17, and MMP9 genes in different tumor types. Correlation analysis between gene/isoforms expression and DNA methylation levels of LCN2, SLC22A17, and MMP9 was performed to investigate the role of DNA methylation in the modulation of these genes. Protein network analysis was carried out using reverse phase protein arrays (RPPA) data to identify protein-protein interactions of the LCN2-SLC22A17-MMP9 network. Furthermore, survival analysis was performed according to gene expression and DNA methylation levels. Our results demonstrated that LCN2 and MMP9 were mainly upregulated in most tumor types, whereas SLC22A17 was largely downregulated, representing a specific hallmark signature for all gastrointestinal tumors. Notably, the expression of LCN2, SLC22A17, and MMP9 genes was negatively affected by promoter methylation. Conversely, intragenic hypermethylation was associated with the overexpression of SLC22A17 and MMP9 genes. Protein network analysis highlighted the role of the LCN2-SLC22A17-MMP9 network in TME by the interaction with fibronectin 1 and claudin 7, especially in rectal tumors. Moreover, the impact of expression and methylation status of LCN2, SLC22A17, and MMP9 on overall survival and progression free interval was tumor type-dependent. Overall, our analyses provide a detailed overview of the expression and methylation status of LCN2, SLC22A17, and MMP9 in all TCGA tumors, indicating that the LCN2-SLC22A17-MMP9 network was strictly regulated by DNA methylation within TME. Our findings pave the way for the identification of novel DNA methylation hotspots with diagnostic and prognostic values and suitable for epi-drug targeting.

Vitamin E and Non-Communicable Diseases: A Review.

Vitamin E, a nutrient found in several foods, comprises eight lipophilic vitamers, the α-, ß-, γ- and δ-tocopherols and the α-, ß-, γ- and δ-tocotrienols. This vitamin is capable of exerting antioxidant and anti-inflammatory activities, and acting as immunomodulators. Despite these well-known biological activities, the findings regarding the ability of vitamin E and its serum metabolites to prevent and/or control chronic disease are often conflicting and inconsistent. In this review, we have described the metabolism of vitamin E and its interaction with the gut microbiota, considering that these factors may be partially responsible for the divergent results obtained. In addition, we focused on the correlations between vitamin E serum levels, dietary intake and/or supplementation, and the main non-communicable diseases, including diabetes mellitus, asthma, cardiovascular diseases, and the four most common cancers (breast cancer, lung cancer, colorectal cancer, and prostate cancer) with the intention of providing an overview of its health effects in the non-communicable-diseases prevention.

Mangifera indica L. Leaves as a Potential Food Source of Phenolic Compounds with Biological Activity.

It is well recognized that functional foods rich in antioxidants and antiinflammation agents including polyphenols, probiotics/prebiotics, and bioactive compounds have been found to have positive effects on the aging process. In particular, fruits play an important role in regular diet, promoting good health and longevity. In this study, we investigated on biological properties of extract obtained from Mangifera indica L. leaves in preclinical in vitro models. Specifically, the profile and content of bioactive compounds, the antimicrobial potential toward food spoilage and pathogenic bacterial species, and the eventually protective effect in inflammation were examined. Our findings revealed that MLE was rich in polyphenols, showing a content exclusively in the subclass of benzophenone/xanthone metabolites, and these phytochemical compounds demonstrated the highest antioxidant capacity and greatest in vitro antibacterial activity toward different bacterial species such as Bacillus cereus, B. subtilis, Pseudomonas fluorescens, Staphylococcus aureus, and St. haemolyticus. Furthermore, our data showed an in vitro anti-inflammatory, antioxidant, and antifibrotic activity.

Co-Occurrence of Interleukin-6 Receptor Asp358Ala Variant and High Plasma Levels of IL-6: An Evidence of IL-6 Trans-Signaling Activation in Deep Vein Thrombosis (DVT) Patients.

Interleukin-6 (IL-6) is a pleiotropic cytokine involved in several mechanisms, and the alteration of IL-6 signaling leads to the overactivation of various processes including immunity, inflammation, and hemostasis. Although IL-6 increase has been documented in venous thromboembolic diseases, the exact involvement of IL-6 signaling in deep vein thrombosis (DVT) has not been fully understood. Consequently, we investigated the involvement of IL-6 trans-signaling in inflammatory events occurring in DVT, focusing on the role of the interleukin-6 receptor (IL6-R) Asp358Ala variant. The circulating levels of IL-6, soluble IL6-R (sIL6-R), and soluble glycoprotein 130, as well as the Asp358Ala genotyping, were assessed in a consecutive cohort of DVT patients and healthy controls. The results indicated that IL-6 was higher in DVT compared to controls. Moreover, sIL6-R levels were strongly correlated to Asp358Ala variant in both groups, showing a high frequency of this mutation across all samples. Interestingly, our results showed a high frequency of both Asp358Ala mutation and raised IL-6 levels in DVT patients (OR = 21.32; p ≤ 0.01), highlighting that this mutation could explain the association between IL-6 overactivation and DVT outcome. Overall, this study represents a proof of concept for the targeting of IL-6 trans-signaling as a new strategy for the DVT adjuvant therapy.

Orobanche crenata Forssk. Extract Affects Human Breast Cancer Cell MCF-7 Survival and Viral Replication.

BACKGROUND: Breast cancer (BC) is the leading cause of death worldwide. The severity of BC strictly depends on the molecular subtype. The less aggressive hormone-positive subtype is treated with adjuvant endocrine therapy (AET), which causes both physical and psychological side effects. This condition strongly impacts the adherence and persistence of AET among oncologic patients. Moreover, viral infections also constitute a serious problem for public health. Despite their efficacy, antiviral agents present several therapeutic limits. Accordingly, in the present work, we investigated the antitumor and antiviral activities of Orobanche crenata Forssk. (O. crenata), a parasitic plant, endemic to the Mediterranean basin, traditionally known for its beneficial properties for human health. METHODS: The MTT assay was carried out to evaluate the cytotoxic effect of O. crenata leaf extract (OCLE) on human breast cancer cells (MCF-7 and MDA-MB-231) and the primary HFF-1 cell line. The lactic dehydrogenase (LDH) assay was performed on MCF-7 cells to analyze necrotic cell death. The antioxidant effect of OCLE was evaluated by intracellular determination of the reactive oxygen species and thiol groups, by DPPH and ABTS assays. The antiviral activity of OCLE was determined against Poliovirus 1, Echovirus 9, Human respiratory syncytial virus, Adenovirus type 2 and type 5, Coxsackievirus B1 (CoxB1) and B3 (CoxB3), Herpes simplex type 1 (HSV-1) and type 2 (HSV-2), and ß-Coronavirus by the plaque reduction assay. RESULTS: The extract, after 24 h of incubation, did not affect MDA-MB-231 and HFF-1 cell viability. However, at the same time point, it showed a dose-dependent inhibitory effect on MCF-7 cells, with an increase in LDH release. OCLE exhibited free radical scavenging activity and significantly increased non-protein thiol levels in MCF-7 cells. OCLE effectively inhibited HSV-1, HSV-2, CoxB1, and CoxB3 replication. CONCLUSIONS: The overall results showed an interesting inhibitory effect of OCLE on both MCF-7 cell survival and viral replication.

Phytocomplex of a Standardized Extract from Red Orange (Citrus sinensis L. Osbeck) against Photoaging.

Excessive exposure to solar radiation is associated with several deleterious effects on human skin. These effects vary from the occasional simple sunburn to conditions resulting from chronic exposure such as skin aging and cancers. Secondary metabolites from the plant kingdom, including phenolic compounds, show relevant photoprotective activities. In this study, we evaluated the potential photoprotective activity of a phytocomplex derived from three varieties of red orange (Citrus sinensis (L.) Osbeck). We used an in vitro model of skin photoaging on two human cell lines, evaluating the protective effects of the phytocomplex in the pathways involved in the response to damage induced by UVA-B. The antioxidant capacity of the extract was determined at the same time as evaluating its influence on the cellular redox state (ROS levels and total thiol groups). In addition, the potential protective action against DNA damage induced by UVA-B and the effects on mRNA and protein expression of collagen, elastin, MMP1, and MMP9 were investigated, including some inflammatory markers (TNF-α, IL-6, and total and phospho NFkB) by ELISA. The obtained results highlight the capacity of the extract to protect cells both from oxidative stress­preserving RSH (p < 0.05) content and reducing ROS (p < 0.01) levels­and from UVA-B-induced DNA damage. Furthermore, the phytocomplex is able to counteract harmful effects through the significant downregulation of proinflammatory markers (p < 0.05) and MMPs (p < 0.05) and by promoting the remodeling of the extracellular matrix through collagen and elastin expression. This allows the conclusion that red orange extract, with its strong antioxidant and photoprotective properties, represents a safe and effective option to prevent photoaging caused by UVA-B exposure.

Overactivation of IL6 cis­signaling in leukocytes is an inflammatory hallmark of deep vein thrombosis.

Inflammation is a protective response of the body to various injuries, which is strictly regulated by a variety of factors, including immune cells and soluble mediators. However, dysfunction of this defensive mechanism often results in inflammation­driven diseases, such as deep vein thrombosis (DVT). The complex relationship between inflammatory cell activity and DVT has not been fully elucidated. The present study aimed to investigate the role of interleukin­6 (IL6) signaling transduction in DVT. To this aim, the expression levels of transmembrane isoforms of the IL6 receptor (IL6R) and the glycoprotein 130 responsible for the IL6 cis­signaling were evaluated in the peripheral blood mononuclear cells of patients with DVT and of healthy controls. The results indicated that leukocytes from patients with DVT exhibited overexpression of both IL6R and gp130 membrane isoforms and that these were strongly associated with the occurrence of DVT. Overall, the present findings indicated that IL6 cis­signaling may have a direct involvement in the leukocyte activation in DVT and may serve as a predictive biomarker of DVT development.

Protocatechuic Acid, a Simple Plant Secondary Metabolite, Induced Apoptosis by Promoting Oxidative Stress through HO-1 Downregulation and p21 Upregulation in Colon Cancer Cells.

Gastrointestinal cancers, particularly colorectal cancer, are mainly influenced by the dietary factor. A diet rich in fruits and vegetables can help to reduce the incidence of colorectal cancer thanks to the phenolic compounds, which possess antimutagenic and anticarcinogenic properties. Polyphenols, alongside their well-known antioxidant properties, also show a pro-oxidative potential, which makes it possible to sensitize tumor cells to oxidative stress. HO-1 combined with antioxidant activity, when overexpressed in cancer cells, is involved in tumor progression, and its inhibition is considered a feasible therapeutic strategy in cancer treatment. In this study, the effects of protocatechuic acid (PCA) on the viability of colon cancer cells (CaCo-2), annexin V, LDH release, reactive oxygen species levels, total thiol content, HO-1, γ-glutamylcysteine synthetase, and p21 expression were evaluated. PCA induced, in a dose-dependent manner, a significantly reduced cell viability of CaCo-2 by oxidative/antioxidant imbalance. The phenolic acid induced modifications in levels of HO-1, non-proteic thiol groups, γ-glutamylcysteine synthetase, reactive oxygen species, and p21. PCA induced a pro-oxidant effect in cancer cells, and the in vitro pro-apoptotic effect on CaCo-2 cells is mediated by the modulation of redox balance and the inhibition of the HO-1 system that led to the activation of p21. Our results suggest that PCA may represent a useful tool in prevention and/or therapy of colon cancer.

Novel Insights into Epigenetic Regulation of IL6 Pathway: In Silico Perspective on Inflammation and Cancer Relationship.

IL-6 pathway is abnormally hyperactivated in several cancers triggering tumor cell growth and immune system inhibition. Along with genomic mutation, the IL6 pathway gene expression can be affected by DNA methylation, microRNAs, and post-translational modifications. Computational analysis was performed on the Cancer Genome Atlas (TCGA) datasets to explore the role of IL6, IL6R, IL6ST, and IL6R transmembrane isoform expression and their epigenetic regulation in different cancer types. IL6 was significantly modulated in 70% of tumor types, revealing either up- or down-regulation in an approximately equal number of tumors. Furthermore, IL6R and IL6ST were downregulated in more than 10 tumors. Interestingly, the correlation analysis demonstrated that only the IL6R expression was negatively affected by the DNA methylation within the promoter region in most tumors. Meanwhile, only the IL6ST expression was extensively modulated by miRNAs including miR-182-5p, which also directly targeted all three genes. In addition, IL6 upregulated miR-181a-3p, mirR-214-3p, miR-18a-5p, and miR-938, which in turn inhibited the expression of IL6 receptors. Finally, the patients' survival rate was significantly affected by analyzed targets in some tumors. Our results suggest the relevance of epigenetic regulation of IL6 signaling and pave the way for further studies to validate these findings and to assess the prognostic and therapeutic predictive value of these epigenetic markers on the clinical outcome and survival of cancer patients.