Antiradical and Cytoprotective Properties of Allium nutans L. Honey Against CCL4-Induced Liver Damage in Rats

The aim of this study is determine the in vitro and in vivo antiradical properties and the cytoprotective activity of Allium nutans L. honey extract. The antiradical properties of the extracts were investigated in rabbit alveolar macrophages and human foreskin fibroblast (hFFs) cells in the presence of doxorubicin, a cytotoxic substance using DPPH and ABTS assays. The cytoprotective activities were determined using 18 Wistar rats divided into three different groups, a negative control, and two other groups with experimentally induced hepatotoxicity by a single intraperitoneal injection of 50% carbon tetrachloride (CCl4) oil solution. A positive control group, received drinking water only and an experimental group that was treated with Allium nutans L. honey extracts for 7 days. In vitro treatment with Allium nutans L. honey extracts resulted in 78% reduction in radical activity in DPPH and 91.6% inhibition using the ABTS. Also, honey extracts were able to preserve 100% of cell viability in the presence of the cytotoxic, doxorubicin. Furthermore, the treatment with honey extracts resulted in a significant reduction in damage to the structure of liver tissue, as well significant reduction in the levels of ALT and AST in the experimental group compared to the control group.

Prospects on the nano-plastic particles internalization and induction of cellular response in human keratinocytes

Background Today, cosmetic products are very popular with both men and women to improve their appearance and increase their social acceptability. Results In this study, nano-sized (30–300 nm) plastic particles were isolated from the commercial face-scrubs and treated on the human keratinocytes. The observed adherence of polyethylene nano-plastics (PENPs), polystyrene NPs (PSNPs), and face-scrubs isolated nano-plastics (NPs) on the keratin layer reveals a significant attachment of NPs from the cosmetics that are applied on the skin for a short duration. This attachment property could facilitate further adherence of protein molecules on NPs and the protein-corona formation. The protein-corona mimics protein aggregates, thereby triggers macropinocytosis, followed by the macropinolysosomal process in the cell. These internalized NPs induced the concentration-dependent cytotoxic, cytostatic and cytoprotective activity in keratinocytes. Both single dose and chronic long-term exposure of lethal and sub-lethal concentrations of NPs resulted in oxidative stress-mediated down-regulation of cell growth and proliferation inhibition. Autophagic structures and premature aging were also observed using an electron microscopy and a senescence marker, respectively in the NPs internalized HaCaT cells incubated in a fresh, NPs-free medium. Conclusion Though 2D culture models have many limitations, it produces significant conceptual advancements. This work provides an insight into the NPs concentration-dependent regulatory, cytoprotective, and cytotoxic effects in HaCaT cells. However, 3D model studies are required to identify the detailed mechanisms of NPs toxicity and cytoprotective events in cells at the molecular level. Graphic abstract

Human-Specific Regulation of Neurotrophic Factors MANF and CDNF by microRNAs

Mesencephalic astrocyte derived neurotrophic factor (MANF) and cerebral dopamine neurotrophic factor (CDNF) are novel evolutionary conserved trophic factors, which exhibit cytoprotective activity via negative regulation of unfolded protein response (UPR) and inflammation. Despite multiple reports demonstrating detrimental effect of MANF/CDNF downregulation, little is known about the control of their expression. miRNAs—small non-coding RNAs—are important regulators of gene expression. Their dysregulation was demonstrated in multiple pathological processes and their ability to modulate levels of other neurotrophic factors, glial cell line-derived neurotrophic factor (GDNF) and brain-derived neurotrophic factor (BDNF), was previously reported. Here, for the first time we demonstrated direct regulation of MANF and CDNF by miRNAs. Using bioinformatic tools, reporter assay and analysis of endogenous MANF and CDNF, we identified that miR-144 controls MANF expression, and miR-134 and miR-141 downregulate CDNF levels. We also demonstrated that this effect is human-specific and is executed via predicted binding sites of corresponding miRNAs. Finally, we found that miR-382 suppressed hCDNF expression indirectly. In conclusion, we demonstrate for the first time direct regulation of MANF and CDNF expression by specific miRNAs, despite the fact their binding sites are not strongly evolutionary conserved. Furthermore, we demonstrate a functional effect of miR-144 mediated regulation of MANF on ER stress response markers. These findings emphasize that (1) prediction of miRNA targets based on evolutionary conservation may miss biologically meaningful regulatory pairs; and (2) interpretation of miRNA regulatory effects in animal models should be cautiously validated.

In-vitro Cytoprotective Activity of Eichhornia crassipes Flowers Against Hydrogen Peroxideinduced Oxidative Stress in BRL 3A Rat Liver Cells

The aim of this study was to investigate the cytoprotective effect of ethanol extract of Eichhornia crassipes flowersand its fractions against hydrogen peroxide induced oxidative stress in BRL 3A liver cells. Powdered flowers of Eichhornia crassipeswere subjected to hot continuous extraction in soxhlet extractor using ethanol as solvent material. Initially, the solvent extracts were subjected to qualitative, quantitative analysis and assessed for in-vitro free radical scavenging activity and anti-oxidant activity. The ethanol extract was fractionated using benzene, chloroform and n-butanol. The crude ethanol extract and its fractions were evaluated for its potential cytoprotective effect against hydrogen peroxide (H2O2) induced oxidative stress in BRL 3A cell lines.Biochemical assays were carried out to determine the cytoprotective activity, including cell viability, lipid peroxidation by determining the formation of malondialdehyde, lactate dehydrogenase leakage into culture medium, the catalase activity and the content of reduced glutathione (GSH) in the cells. Exposure of BRL 3A to 2mM H2O2 reduced the cell viability, increased the malondialdehyde (MDA) level, increased the leakage of lactate dehydrogenase (LDH) and caused reduction in antioxidant activities. Pretreatment of cultured cells with crude ethanol extract of Eichhornia crassipes flowers and different solvent fractions at concentrations 0.01, 0.1, 1, 10, 100 μg/ml for 30 minutes before H2O2 exposure attenuated the oxidative injury in dose-dependent manner. It was observed that crude ethanol extract of Eichhornia crassipes flowers exhibited a strong cytoprotective by increasing cell viability, decreasing lipid peroxidation and LDH leakage. Further increase in catalase and reduced glutathione activity was noted in the cells pre-treated with ethanol extract of Eichhornia crassipes flowers. These findings suggest that ethanol extract of Eichhornia crassipes flowers has a strong cytoprotective activity against oxidative injury caused by reactive oxygen species.

Synthesis, antioxidant and cytoprotective activity evaluation of C-3 substituted indole derivatives

A series of fifteen indole derivatives substituted at the C-3 position were synthesized and characterized. The antioxidant activity of all derivatives was investigated by three in vitro antioxidant assays, and the derivative with pyrrolidinedithiocarbamate moiety was the most active as a radical scavenger and Fe3+-Fe2+ reducer. It can be stated that possible hydrogen and electron transfer mechanism is suggested for the quenching of the free radical. Moreover, the indolyl radical stabilization and the presence of unsubstituted indole nitrogen atom are mandatory for the observed antioxidant activity, which strongly depends on the type of the substituent directly connected to the methylene group at the C-3 position. Human red blood cells (RBC) have been used as a cell model to study derivatives interaction with the cell membrane. Haemolytic activity and RBC shape transformation were observed for certain derivatives in a concentration-dependent manner. However, most of the derivatives at sublytic concentration showed high cytoprotective activity against oxidative haemolysis induced by 2,2′-azobis(2-methylpropionamidine) dihydrochloride (AAPH). The cytoprotective properties of derivatives can be explained mostly due to their interactions with the RBC membrane components. Taking together, theoretical estimations and experimental data confirm the beneficial interactions between the selected C-3 substituted indole derivatives and the RBC membrane under oxidative stress conditions. These results encourage us to further structural optimization of C-3 substituted indole derivatives as potent antioxidant compounds.

Engineered EVs for Oxidative Stress Protection

Extracellular vesicles (EVs) are increasingly studied as vectors for drug delivery because they can transfer a variety of molecules across biological barriers. SerpinB3 is a serine protease inhibitor that has shown a protective anti-apoptotic function in a variety of stressful conditions. The aim of this study was to evaluate protection from oxidative stress-induced damage, using extracellular vesicles that overexpress SerpinB3 (EVs-SB3) in order to enhance the effect of extracellular vesicles on cellular homeostasis. EVs-SB3s were obtained from HepG2 cells engineered to overexpress SerpinB3 and they revealed significant proteomic changes, mostly characterized by a reduced expression of other proteins compared with EVs from non-engineered cells. These EV preparations showed a significantly higher protection from H2O2 induced oxidative stress in both the hepatoma cell line and in primary cardiomyocytes, compared to cells treated with naïve EVs or SerpinB3 alone, used at the same concentration. In conclusion, the induction of SerpinB3 transgene expression results in the secretion of EVs enriched with the protein product that exhibits enhanced cytoprotective activity, compared with naïve EVs or the nude SerpinB3 protein.

Personalized trimetazidine prescription as a cytoprotective agent in patients with coronary artery disease

Aim. To develop a personalized approach to the trimetazidine use in patients with coronary artery disease (CAD) based on the criteria for predicting the cytoprotective activity tested in vitro.Material and methods. We examined 30 patients with class I-III stable effort angina with concomitant hypertension and heart failure. The patients underwent echocardiography, complete blood count, biochemical tests with determination of the lipid profile, creatine phosphokinase (CPK), CPK-MB, renal and hepatic parameters. To determine the cytoprotective activity of trimetazidine, white blood cells (WBCs) of patients were examined in vitro using an Eclipse Ti-U inverted fluorescence microscope (Nikon, Japan). Living and dead cells were determined by staining WBCs with fluorescent dyes (Calcein AM, Ethidium bromide). Cell viability index (CVI) was calculated. The statistical processing was carried out. The criteria for predicting the trimetazidine cytoprotective effect were determined using Wald statistics.Results. When trimetazidine was injected into a WBC suspension sample, two types of cell viability changes were observed: in 60% of patients, CVI increased, on average, by 37% (from 23% to 60%, p<0,001) and in 40% of patients, CVI decreased, on average, by 30% (from 54% to 24%, p<0,05).A number of conditions of the patient initial status were identified for the manifestation of trimetazidine cytoprotective activity: grade 1 hypertension; right ventricular end diastolic dimension up to 30 mm according to echocardiography; normal lipid profile with a total cholesterol <5,3 mmol/L, very-low-density lipoproteins <1 mmol/L and an atherogenic coefficient up to 3 CU, myocyte and cardiomyocyte destruction (total CPK >100 U/L and CPK-MB >15 U/L), normal liver function (alanine aminotransferase <25 U/L), renal dysfunction (total protein <75 g/L, urea >8 mmol/L and blood creatinine >100 pmol/L), normal thrombopoiesis (immature platelet fraction <5%) and the state of functional adaptive system resistance (blood lymphocytes <30% and neutrophils >4x109/L).Conclusion. According to this in vitro analysis, the trimetazidine significantly increases (by an average of 37%) the cell (WBC) viability in 60% of patients with CAD. There are conditions of patient initial status, which specifies an individual pharmacodynamic target for the cytoprotective action of the drug.

In-vitro cytoprotective activity of Eichhornia crassipes ϐlowers against hydrogen peroxide induced oxidative stress in BRL 3A rat liver cells

The aim of this study was to see whether an ethanol extract of Eichhornia crassipes flowers and its fractions could protect BRL 3A liver cells from hydrogen peroxide-induced oxidative stress. Eichhornia crassipes powdered flowers were subjected to a hot continuous extraction in a soxhlet extractor using ethanol as the solvent material. The solvent extracts were first tested for in-vitro free radical scavenging and anti-oxidant activity using qualitative and quantitative methods. Benzene, chloroform, and n-butanol were used to fractionate the ethanol extract. In BRL 3A cell lines, the crude ethanol extract and its fractions were tested for their possible cytoprotective effect against hydrogen peroxide (H2O2) induced oxidative stress. Cell viability, lipid peroxidation by measuring the formation of malondialdehyde, lactate dehydrogenase leakage into culture medium, catalase activity, and the content of reduced glutathione (GSH) in the cells were all tested in biochemical assays to determine the cytoprotective activity. BRL 3A cells were exposed to 2mM H2O2, which decreased cell viability, increased malondialdehyde (MDA) levels, increased lactate dehydrogenase (LDH) leakage, and reduced antioxidant activities. Pretreatment of cultured cells for 30 minutes with crude ethanol extract of Eichhornia crassipes flowers and various solvent fractions at concentrations of 0.01, 0.1, 1, 10, 100 g/ml attenuated oxidative injury in a dose-dependent manner until H2O2 exposure. The crude ethanol extract of Eichhornia crassipes flowers was found to have a strong cytoprotective impact, raising cell viability while decreasing lipid peroxidation and LDH leakage. In the cells pre-treated with ethanol extract of Eichhornia crassipes flowers, there was a further increase in catalase and a decrease in glutathione activity. These results indicate that an ethanol extract of Eichhornia crassipes flowers has potent cytoprotective properties against reactive oxygen species-induced oxidative injury.

Cytoprotective Activity of p-Terphenyl Polyketides and Flavuside B from Marine-Derived Fungi against Oxidative Stress in Neuro-2a Cells

The influence of p-terphenyl polyketides 1–3 from Aspergillus candidus KMM 4676 and cerebroside flavuside B (4) from Penicillium islandicum (=Talaromyces islandicus) against the effect of neurotoxins, rotenone and paraquat, on Neuro-2a cell viability by MTT and LDH release assays and intracellular ROS level, as well as DPPH radical scavenging activity, was investigated. Pre-incubation with compounds significantly diminished the ROS level in rotenone- and paraquat-treated cells. It was shown that the investigated polyketides 1–3 significantly increased the viability of rotenone- and paraquat-treated cells in two of the used assays but they affected only the viability of paraquat-treated cells in the LDH release assay. Flavuside B statistically increased the viability of paraquat-treated cells in both MTT and LDH release assays, however, it increased the viability of rotenone-treated cells in the LDH release assay. Structure–activity relationships for p-terphenyl derivatives, as well as possible mechanisms of cytoprotective action of all studied compounds, were discussed.

Prospects on the nano-plastic particles internalization and induction of cellular response in human keratinocytes

Background Today, cosmetic usage has become customary in both men and women towards improving their appearance and increase societal visibility. In this study, the nano-sized (30 to 300 nm) plastic particles were isolated from the commercial face-scrubs and treated on the human keratinocytes. Results The adherence studies of polyethylene nano-plastics (PENPs), polystyrene NPs (PSNPs), and face-scrubs isolated nano-plastics (NPs) on the keratin layer revealed a rapid attachment of NPs on the skin from the cosmetics that have short exposure time. This attachment property could facilitate further adherence of protein molecules on NPs and the protein-corona formation. The protein-corona mimics protein aggregates, thereby triggers the macropinocytosis followed by the macropinolysosomal process in the cell. Then the internalized NPs induced the concentration-dependent cytotoxic, cytostatic and cytoprotective activity in keratinocytes. Both single dose and chronic long-term exposure of lethal and sub-lethal concentrations of NPs resulted in the oxidative stress-mediated down-regulation of cell growth and proliferation inhibition. Autophagy and premature aging were also observed in the NPs internalized HaCaT cells incubated in a fresh, NPs-free medium. Conclusion At the outset, this work provides insight into the NPs concentration-dependent regulatory, cytoprotective, and cytotoxic effects in HaCaT cells. Further studies are required to identify the detailed mechanisms of NPs toxicity and cytoprotective events in cells at the molecular level.