Cemtirestat, an aldose reductase inhibitor and antioxidant compound, induces ocular defense against oxidative and inflammatory stress in rat models for glycotoxicity.

Fructose, endogenously produced as a consequence of activation of the polyol pathway under hyperglycemic conditions, contribute to formation of advanced glycoxidation end products (AGEs) and carbonyl stress. Oxidative stress is increased in diabetes (DM) due to AGEs formation and the utilization of NADPH by aldo-keto reductase, AKR1B1(AR), the first enzyme in polyol pathway. Since inhibition of AR is an attractive approach for the management of diabetic eye diseases, we aimed to compare the effects of a novel AR inhibitor (ARI)/antioxidant (AO) compound cemtirestat on eye tissues with the effects of ARI drug epalrestat and AO agent stobadine in rat model for glycotoxicity. One group of rats was fed high fructose (10% drinking water; 14 weeks), while type-2 DM was induced in the other group of rats with fructose plus streptozotocin (40 mg/kg-bw/day). Diabetic (D) and nondiabetic fructose-fed rats (F) were either untreated or treated with two different doses of cemtirestat (2.5 and 7.5 mg/kg-bw/day), epalrestat (25 and 50 mg/kg-bw/day), or stobadine (25 and 50 mg/kg-bw/day) for 14 weeks. Cemtirestat, epalrestat, and stobadine elaviate the increase in TNF-α, IL-1ß, NF-ƙB, and caspase-3 in retina, lens, cornea, and sclera of F and D rats. Both glycotoxicity models resulted in a decrease in GSH to GSSG ratio and a change in glutathione S-transferase activity in eye tissues, but these alterations were improved especially with cemtirestat and stobadine. Lens D-sorbitol of D rats increased more than that of F rats, this increase was only attenuated by cemtirestat and epalrestat. Epalrestat was more effective than cemtirestat and stobadine in inhibiting the increase of vascular endothelial growth factor (VEGF) in the retina of F and D rats. Cemtirestat and stobadine but not epalrestat decreased high level of Nε-(carboxymethyl)lysine in the lens and retina of F and D rats. Cemtirestat is a potential therapeutic in protecting the rat eye against glycotoxicity insults.

A dual-acting aldose reductase inhibitor impedes oxidative and carbonyl stress in tissues of fructose- and streptozotocin-induced rats: comparison with antioxidant stobadine.

Inhibiting aldose reductase (ALR2, AR) as well as maintaining a concomitant antioxidant (AO) activity via dual-acting agents may be a rational approach to prevent cellular glucotoxicity and at least delay the progression of diabetes mellitus (DM). This study was aimed at evaluating the dual-acting AR inhibitor (ARI) cemtirestat (CMTI) on tissue oxidative stress (OS) and carbonyl stress (CS) biomarkers in rats exposed to fructose alone (F) or fructose plus streptozotocin (D; type-2 diabetic). D and F rats were either untreated or treated daily with low- or high-dose CMTI, ARI drug epalrestat (EPA) or antioxidant stobadine (STB) for 14 weeks. Malondialdehyde (MDA), glutathione S-transferase (GST), nitric oxide synthase (NOS), and catalase (CAT) were increased in the sciatic nerve of F and D. These increases were attenuated by low doses of CMTI and STB in D, but exacerbated by low-dose EPA and high-dose CMTI in F. STB and CMTI and to a lesser extent EPA improved MDA, protein-carbonyl, GST and CAT in the hearts and lungs of F and D. CMTI and STB were more effective than EPA in improving the increased MDA and protein-carbonyl levels in the kidneys of F and especially D. CMTI ameliorated renal GST inhibition in D. In the lungs, hearts, and kidneys of F and D, the GSH to GSSG ratio decreased and caspase-3 activity increased, but partially resolved with treatments. In conclusion, CMTI with ARI/AO activity may be advantageous in overcoming OS, CS, and their undesirable consequences, with low dose efficacy and limited toxicity, compared to ARI or antioxidant alone.

S-allylcysteine inhibits chondrocyte inflammation to reduce human osteoarthritis via targeting RAGE, TLR4, JNK, and Nrf2 signaling: comparison with colchicine.

The discovery of new pharmacological agents is needed to control the progression of osteoarthritis (OA), characterized by joint cartilage damage. Human OA chondrocyte (OAC) cultures were either applied to S-allylcysteine (SAC), a sulfur-containing amino acid derivative, or colchicine, an ancient anti-inflammatory therapeutic, for 24 h. SAC or colchicine did not change viability at 1 nM-10 µM but inhibited p-JNK/pan-JNK. While SAC seems to be more effective, both agents inhibited reactive oxygen species (ROS), 3-nitrotyrosine (3-NT), lipid hydroperoxides (LPO), advanced lipoxidation end-products (ALEs as 4-hydroxy-2-nonenal, HNE), advanced glycation end-products (AGEs), and increased glutathione peroxidase (GPx) and type-II-collagen (COL2). IL-1ß, IL-6, and osteopontin (OPN) were more strongly inhibited by SAC than by colchicine. In contrast, TNF-α was inhibited only by SAC, and COX2 was only inhibited by colchicine. Casp-1/ICE, GM-CSF, receptor for advanced glycation end-products (RAGE), and toll-like receptors (TLR4) were inhibited by both agents, but bone morphogenetic protein 7 (BMP7) was partially inhibited by SAC and induced by colchicine. Nuclear factor erythroid 2-related factor 2 (Nrf2) was induced by SAC; in contrast, it was inhibited by colchicine. Although they exert opposite effects on TNF-α, COX2, BMP7, and Nrf2, SAC and colchicine exhibit anti-osteoarthritic properties in OAC by modulating redox-sensitive inflammatory signaling.

Platinum nanoparticles Protect Against Lipopolysaccharide-Induced Inflammation in Microglial BV-2 Cells via Decreased Oxidative Damage and Increased Phagocytosis.

Neuroinflammation and oxidative stress cooperate to compromise the function of the central nervous system (CNS). Colloidal platinum nanoparticles (Pt NPs) are ideal candidates for reducing the deleterious effects of neuroinflammation since they act as free radical scavengers. Here we evaluated the effects of Pt NPs on several markers of lipopolysaccharide (LPS)-induced inflammation in cultured BV-2 microglial cells. BV-2 cells were treated with increased dilutions (1-100 ppm) of Colloidal Pt and/or LPS (1-10 µg/mL) at different exposure times. Three different protocols of exposure were used combining Pt NPs and LPS: (a) conditioning-protective effect (pre-post-treat), (b) therapeutic effect (co-treat) and (c) conditioning-therapeutic effect (pre-co-treat). After exposure to LPS for 24 h, cells were used for assessment of cell viability, reactive oxygen species (ROS) generation, lactate dehydrogenase (LDH) activity, apoptosis and caspase-3 levels, cell proliferation, mitochondrial membrane potential, inducible nitric oxide (iNOS) activity, pro-inflammatory cytokine (IL-1ß, TNF-α and IL-6) levels, and phagocytic activity. Low concentrations (below or equal to 10 ppm) of Colloidal Pt prevented or ameliorated the LPS-induced increase in ROS formation, loss of mitochondrial membrane potential, induction of apoptosis, increase in LDH release, increase in pro-inflammatory cytokines and iNOS, inhibition of phagocytosis linked to microglial persistence in the M1 phase phenotype, loss of cell adhesion, differentiation and/or proliferation, as well as loss of cell viability. These protective effects were evident when cells were preconditioned with Pt NPs prior to LPS treatment. Collectively, the findings demonstrate that at low concentrations, Pt NPs can regulate the function and phenotype of BV-2 cells, activating protective mechanisms to maintain the microglial homeostasis and reduce inflammatory events triggered by the inflammatory insults induced by LPS. These preventive/protective effects on the LPS pro-inflammatory model are linked to the antioxidant properties and phagocytic activity of these NPs.

Development of Novel Indole-Based Bifunctional Aldose Reductase Inhibitors/Antioxidants as Promising Drugs for the Treatment of Diabetic Complications.

Aldose reductase (AR, ALR2), the first enzyme of the polyol pathway, is implicated in the pathophysiology of diabetic complications. Aldose reductase inhibitors (ARIs) thus present a promising therapeutic approach to treat a wide array of diabetic complications. Moreover, a therapeutic potential of ARIs in the treatment of chronic inflammation-related pathologies and several genetic metabolic disorders has been recently indicated. Substituted indoles are an interesting group of compounds with a plethora of biological activities. This article reviews a series of indole-based bifunctional aldose reductase inhibitors/antioxidants (ARIs/AOs) developed during recent years. Experimental results obtained in in vitro, ex vivo, and in vivo models of diabetic complications are presented. Structure-activity relationships with respect to carboxymethyl pharmacophore regioisomerization and core scaffold modification are discussed along with the criteria of 'drug-likeness". Novel promising structures of putative multifunctional ARIs/AOs are designed.

Luteolin protects microglia against rotenone-induced toxicity in a hormetic manner through targeting oxidative stress response, genes associated with Parkinson's disease and inflammatory pathways.

Rotenone, an environmental toxin, triggers Parkinson's disease (PD)-like pathology through microglia-mediated neuronal death. The effects and molecular mechanisms of flavonoid luteolin against rotenone-induced toxicity was assessed in microglial BV2 cells. Cells were pretreated with luteolin (1-50 µM) for 12 h and then was co-treated with 20 µM of rotenone for an additional 12 h in the presence of luteolin. The viability (MTT), IL-1ß and TNF-α levels and lactate dehydrogenase (LDH) release (ELISA), and Park2, Lrrk2, Pink1, Nrf2 and Trx1 mRNA levels (qRT-PCR) were measured. In rotenone exposed microglia, luteolin increased viability significantly at lower concentrations (1-5 µM) compared to higher concentrations (25-50 µM). Rotenone increased LDH release and IL-1ß levels in a dose-dependent manner (1-20 µM). Luteolin inhibited rotenone-induced LDH release, however the activity decreased in concentration-dependent manner Neither rotenone nor luteolin altered TNF-α levels, but luteolin reduced IL-1ß levels in a concentration dependent manner in rotenone exposed cells. The mRNA levels of Nrf2 and Trx1, which are the master regulators of redox state, were increased by rotenone, as well as by luteolin, which exhibited an inverse relationship between its concentration and effect (1-20 µM). Park2 mRNA levels increased by luteolin, but decreased by rotenone. Pink1 mRNA levels was not altered by rotenone or luteolin. Lrrk2 mRNA levels reduced by luteolin, while it was increased by rotenone. Results suggest that luteolin have favorable effects on regulation of oxidative stress response, genes associated with PD and inflammatory pathways, hence protects microglia against rotenone toxicity in a hormetic manner.

3-Mercapto-5H-1,2,4-Triazino[5,6-b]Indole-5-Acetic Acid (Cemtirestat) Alleviates Symptoms of Peripheral Diabetic Neuropathy in Zucker Diabetic Fatty (ZDF) Rats: A Role of Aldose Reductase.

Peripheral neuropathy is the most prevalent chronic complication of diabetes mellitus. Good glycemic control can delay the appearance of neuropathic symptoms in diabetic patients but it is not sufficient to prevent or cure the disease. Therefore therapeutic approaches should focus on attenuation of pathogenetic mechanisms responsible for the nerve injury. Considering the role of polyol pathway in the etiology of diabetic neuropathy, we evaluated the effect of a novel efficient and selective aldose reductase inhibitor, 3-mercapto-5H-1,2,4-triazino[5,6-b]indole-5-acetic acid (cemtirestat), on symptoms of diabetic peripheral neuropathy in Zucker Diabetic Fatty (ZDF) rats. Since the age of 5 months, male ZDF rats were orally administered cemtirestat, 2.5 and 7.5 mg/kg/day, for two following months. Thermal hypoalgesia was evaluated by tail flick and hot plate tests. Tactile allodynia was determined by a von Frey flexible filament test. Two-month treatment of ZDF rats with cemtirestat (i) did not affect physical and glycemic status of the animals; (ii) partially inhibited sorbitol accumulation in red blood cells and the sciatic nerve; (iii) markedly decreased plasma levels of thiobarbituric acid reactive substances; (iv) normalized symptoms of peripheral neuropathy with high significance. The presented findings indicate that inhibition of aldose reductase by cemtirestat is not solely responsible for the recorded improvement of the behavioral responses. In future studies, potential effects of cemtirestat on consequences of diabetes that are not exclusively dependent on glucose metabolism via polyol pathway should be taken into consideration.

Quercetin and hydroxytyrosol attenuates xanthine/xanthine oxidase-induced toxicity in H9c2 cardiomyocytes by regulation of oxidative stress and stress-sensitive signaling pathways.

The increased activity of xanthine/xanthine oxidase (X/XO) has been suggested as a risk factor for heart disease and herbal polyphenols exhibits cardioprotection in vitro and in vivo. To understand the cardioprotective action mechanisms of polyphenol quercetin and hydroxytyrosol, the expression levels of stress-responsive proteins were studied in X/XO-induced toxicity model of H9c2 cardiomyocyocytes. Pretreatment with each polypenol (0.1-10 µg/ml; 24 h) enhanced viability (p < 0.01; MTT test) and inhibited reactive oxygen species (ROS) generation (p < 0.001; H2DCFDA assay) against 12 h exposure to a free radical generating system, X (0.5 mM) and XO (5 mU/ml). Western blotting experiments showed that X/XO increases the phosphorylation of downstream substrate of p38, MAPK-activated protein kinase 2 (MAPKAPK-2), p44/42-MAPK (Erk1/2) and cleaved caspase-3 (p < 0.001, vs. Control), however inhibits the levels of phosphorylated c-Jun and Hsp27 (p < 0.01, vs. Control). Pretreatment with quercetin or hydroxytyrosol attenuated the phosphorylation of MAPKAPK-2 and cleaved caspase-3 in X/XO-exposed cells (p < 0.01, vs. X/XO). Hydroxytyrosol enhanced the reduction of phosphorylation of a transcriptional target c-Jun and led to overphosphorylation in protective proteins, p44/42-MAPK and Hsp27 in X/XO-exposed cells (p < 0.01, vs. X/XO). Our data suggest that quercetin and hydroxytyrosol protects cardiomyocytes against X/XO-induced oxidative toxicity by diminishing intracellular ROS and the regulation of stress-sensitive protein kinase cascades and transcription factors.

Olive leaf extracts protect cardiomyocytes against 4-hydroxynonenal-induced toxicity in vitro: comparison with oleuropein, hydroxytyrosol, and quercetin.

Olive (Olea europaea) leaf, an important traditional herbal medicine, displays cardioprotection that may be related to the cellular redox modulating effects of its polyphenolic constituents. This study was undertaken to investigate the protective effect of the ethanolic and methanolic extracts of olive leaves compared to the effects of oleuropein, hydroxytyrosol, and quercetin as a positive standard in a carbonyl compound (4-hydroxynonenal)-induced model of oxidative damage to rat cardiomyocytes (H9c2). Cell viability was detected by the MTT assay; reactive oxygen species production was assessed by the 2',7'-dichlorodihydrofluorescein diacetate method, and the mitochondrial membrane potential was determined using a JC-1 dye kit. Phospho-Hsp27 (Ser82), phospho-MAPKAPK-2 (Thr334), phospho-c-Jun (Ser73), cleaved-caspase-3 (cl-CASP3) (Asp175), and phospho-SAPK/JNK (Thr183/Tyr185) were measured by Western blotting. The ethanolic and methanolic extracts of olive leaves inhibited 4-hydroxynonenal-induced apoptosis, characterized by increased reactive oxygen species production, impaired viability (LD50: 25 µM), mitochondrial dysfunction, and activation of pro-apoptotic cl-CASP3. The ethanolic and methanolic extracts of olive leaves also inhibited 4-hydroxynonenal-induced phosphorylation of stress-activated transcription factors, and the effects of extracts on p-SAPK/JNK, p-Hsp27, and p-MAPKAPK-2 were found to be concentration-dependent and comparable with oleuropein, hydroxytyrosol, and quercetin. While the methanolic extract downregulated 4-hydroxynonenal-induced p-MAPKAPK-2 and p-c-Jun more than the ethanolic extract, it exerted a less inhibitory effect than the ethanolic extract on 4-hydroxynonenal-induced p-SAPK/JNK and p-Hsp27. cl-CASP3 and p-Hsp27 were attenuated, especially by quercetin. Experiments showed a predominant reactive oxygen species inhibitory and mitochondrial protecting ability at a concentration of 1-10 µg/mL of each extract, oleuropein, hydroxytyrosol, and quercetin. The ethanolic extract of olive leaves, which contains larger amounts of oleuropein, hydroxytyrosol, verbascoside, luteolin, and quercetin (by HPLC) than the methanolic one, has more protecting ability on cardiomyocyte viability than the methanolic extract or each phenolic compound against 4-hydroxynonenal-induced carbonyl stress and toxicity.

Pomegranate Seed Oil Modulates Functions and Survival of BV-2 Microglial Cells in vitro.

Current evidence has demonstrated the immunomodulatory efficacy of omega-3 polyunsaturated fatty acids (PUFAs) in glial cells, suggesting their therapeutic potential for diseases in the central nervous system (CNS). However, conjugated omega-5 PUFAs have also attracted considerable attention because of their suggested anti-inflammatory effects. In the present study, the effect of pomegranate (Punica granatum L.) seed oil (PSEO) (a rich source of omega-5 PUFAs) on the activation of cultured BV-2 microglia was investigated within a 24-hour incubation period. PSEO (25 µg/ml) showed only a slightly smaller inhibitory effect on LPS-stimulated NO production (243 ± 12.5 % of control, p<0.001 vs. 437 ± 9.2 % in stimulated cells) and TNF-α release (87.1 ± 5.62 pg/ml vs. 229 ± 24.4 pg/ml in stimulated cells), as well as iNOS expression (7.36-fold of control, p < 0.01, vs. 17.5-fold increase in stimulated cells) compared to a standardized omega-3 PUFAs mixture (25 µg/ml) and the flavonoid quercetin (25 µmol/l). Unlike quercetin and stobadine, only the PUFA preparations effectively prevented apoptosis of microglia (as confirmed by the suppression of caspase 3 activation) exposed to the toxic concentration of LPS. The PUFA preparations did not provide a notable suppression of the intracellular oxidant generation and did not influence the intracellular distribution of cholesterol (as confirmed by filipin staining). However, they appeared to affect the morphology of activated cells. In conclusion, our data point to the first evidence of immunomodulation and cytoprotection of BV-2 microglia by the pomegranate seed oil, indicating that it may be (comparably to omega-3 PUFAs) efficient against microglia-mediated neuroinflammation while preventing the premature depletion of these immune effector cells in the brain.

Redox status related activation of endoplasmic reticulum stress and apoptosis caused by 4-hydroxynonenal exposure in INS-1 cells.

4-Hydroxynonenal (HNE), a diffusible aldehyde product of membrane lipid peroxidation, can be produced by oxidative stress and has been detected in several diseases such as diabetes. In this study, we investigated the effects of HNE exposure on cytotoxicity, intracellular redox status, endoplasmic reticulum (ER) stress and apoptosis in insulinoma cell line (INS-1). Short-term (1 h) incubation of INS-1 cells with 0-50 µM HNE decreased cell viability and caused depletion in reduced glutathione (GSH) levels and increased intracellular HNE-histidine adducts in a concentration-dependent manner. HNE activated the ER stress, leading to an increase in inositol-requiring enzyme-1a IRE1-α, phosphorylation of protein kinase-like ER kinase, phosphorylation of c-Jun N-terminal kinase (JNK) and increased the expression of CCAAT/enhancer binding protein (CHOP). Western blot analysis showed that HNE exposure induced dose-dependent activation of caspase 9 and caspase 3. These data indicate a potential role for HNE promoting deleterious effects toward pancreatic beta cell redox status and beta cell mass which may be important for the pathogenesis in diabetes.

Protection against α-Amanitin-induced liver toxicity: Efficacy of pomegranate seed oil and black cumin oil.

The consumption of mushrooms containing α-Amanitin (α-A) can lead to severe liver damage. In this study, toxicological experiments were conducted to confirm the protective effects of pomegranate seed oil (PSO) and black cumin oil (BCO) against α-A-induced hepatotoxicity. Rats exposed once to α-A (3 mg/kg bw, i.p.) or saline alone (0.1 ml, i.p.) were either left untreated or treated with PSO or BCO at a dose of 2 ml/kg bw/day by oral gavage on the same day, and the treatment was continued for 7 days. Serum aminotransferases (ALT and AST), alkaline phosphatase (ALP) and total protein levels were measured and the active caspase 3 (cl-caspase 3) was evaluated by western blotting in the liver. Serum ALT, AST and ALP levels tended to decrease in the α-A exposed group, but no statistically significant difference was found compared to the saline group (p > 0.05). PSO and BCO did not affect serum liver function tests in rats exposed to saline or α-A. α-A toxicity was demonstrated by a significant decrease in serum total protein level (p < 0.05), a significant increase in liver cl-caspase 3 expression (p < 0.05), and structural liver damage mainly characterized by mononuclear inflammation and steatosis. When α-A exposed rats were treated with BCO, the increase in cl-caspase 3 was not inhibited, on the contrary BCO increased cl-caspase 3 in healthy rats (p < 0.05). PSO significantly ameliorated α-A-induced cl-caspase 3 increase and inflammatory histopathology in the liver. Both PSO and BCO completely prevented α-A-induced protein degradation. The findings indicate that PSO and BCO may protect liver functions against α-A-induced hepatotoxicity, encouraging future comprehensive studies to test them at different doses and frequency.

S-allyl-cysteine triggers cytotoxic events in rat glioblastoma RG2 and C6 cells and improves the effect of temozolomide through the regulation of oxidative responses.

Glioblastoma (GBM) is an aggressive form of cancer affecting the Central Nervous System (CNS) of thousands of people every year. Redox alterations have been shown to play a key role in the development and progression of these tumors as Reactive Oxygen Species (ROS) formation is involved in the modulation of several signaling pathways, transcription factors, and cytokine formation. The second-generation oral alkylating agent temozolomide (TMZ) is the first-line chemotherapeutic drug used to treat of GBM, though patients often develop primary and secondary resistance, reducing its efficacy. Antioxidants represent promising and potential coadjutant agents as they can reduce excessive ROS formation derived from chemo- and radiotherapy, while decreasing pharmacological resistance. S-allyl-cysteine (SAC) has been shown to inhibit the proliferation of several types of cancer cells, though its precise antiproliferative mechanisms remain poorly investigated. To date, SAC effects have been poorly explored in GBM cells. Here, we investigated the effects of SAC in vitro, either alone or in combination with TMZ, on several toxic and modulatory endpoints-including oxidative stress markers and transcriptional regulation-in two glioblastoma cell lines from rats, RG2 and C6, to elucidate some of the biochemical and cellular mechanisms underlying its antiproliferative properties. SAC (1-750 µM) decreased cell viability in both cell lines in a concentration-dependent manner, although C6 cells were more resistant to SAC at several of the tested concentrations. TMZ also produced a concentration-dependent effect, decreasing cell viability of both cell lines. In combination, SAC (1 µM or 100 µM) and TMZ (500 µM) enhanced the effects of each other. SAC also augmented the lipoperoxidative effect of TMZ and reduced cell antioxidant resistance in both cell lines by decreasing the TMZ-induced increase in the GSH/GSSG ratio. In RG2 and C6 cells, SAC per se had no effect on Nrf2/ARE binding activity, while in RG2 cells TMZ and the combination of SAC + TMZ decreased this activity. Our results demonstrate that SAC, alone or in combination with TMZ, exerts antitumor effects mediated by regulatory mechanisms of redox activity responses. SAC is also a safe drug for testing in other models as it produces non-toxic effects in primary astrocytes. Combined, these effects suggest that SAC affords antioxidant properties and potential antitumor efficacy against GBM.

4,5-dianilinophtalimide protects neuroendocrine cells against serum deprivation-induced stress and apoptosis.

OBJECTIVE: The aim of this study was to reveal the effects of 4,5-dianilinophthalimide (DAPH), which inhibits amyloid ß fibrillization, against serum deprivation (SD)-induced apoptosis and the possible mechanisms in differentiated PC12 neuron cells. METHODS: Firstly, we evaluated whether DAPH protects cell viability exposed to SD by MTT assay. Next, we examined the changes of phospho-p38 MAPK (Thr180/Tyr182), phospho-HSP27 (Ser82), phospho-c-JUN (Ser73) and cleaved-CASP3 (Asp175) profiles by immunoblotting, in PC12 cells exposed to SD. Intracellular reactive oxygen species (ROS) level was also measured. RESULTS: SD induced apoptosis accompanied by up-regulation of phospho-p38 MAPK (Thr180/Tyr182), phospho-HSP27 (Ser82), phospho-c-JUN (Ser73), cleaved-CASP3 (Asp175) and intracellular ROS content. Co-treatment with non-toxic doses of DAPH prevented apoptosis by the attenuation of activated proteins and reduction of ROS level. These results suggest that serum deprivation-induced apoptosis inhibited by DAPH administration. CONCLUSION: We have provided for the first evidence that DAPH has a neuroprotective effect on SD-caused stress, probably via contributing the re-establishment of redox homeostasis.

Graphene- and MXene-based materials for neuroscience: diagnostic and therapeutic applications.

MXenes and graphene are two-dimensional materials that have gained increasing attention in neuroscience, particularly in sensing, theranostics, and biomedical engineering. Various composites of graphene and MXenes with fascinating thermal, optical, magnetic, mechanical, and electrical properties have been introduced to develop advanced nanosystems for diagnostic and therapeutic applications, as exemplified in the case of biosensors for neurotransmitter detection. These biosensors display high sensitivity, selectivity, and stability, making them promising tools for neuroscience research. MXenes have been employed to create high-resolution neural interfaces for neuroelectronic devices, develop neuro-receptor-mediated synapse devices, and stimulate the electrophysiological maturation of neural circuits. On the other hand, graphene/derivatives exhibit therapeutic applicability in neuroscience, as exemplified in the case of graphene oxide for targeted delivery of therapeutic agents to the brain. While MXenes and graphene have potential benefits in neuroscience, there are also challenges/limitations associated with their use, such as toxicity, environmental impacts, and limited understanding of their properties. In addition, large-scale production and commercialization as well as optimization of reaction/synthesis conditions and clinical translation studies are very important aspects. Thus, it is important to consider the use of these materials in neuroscience research and conduct further research to obtain an in-depth understanding of their properties and potential applications. By addressing issues related to biocompatibility, long-term stability, targeted delivery, electrical interfaces, scalability, and cost-effectiveness, MXenes and graphene have the potential to greatly advance the field of neuroscience and pave the way for innovative diagnostic and therapeutic approaches for neurological disorders. Herein, recent advances in therapeutic and diagnostic applications of graphene- and MXene-based materials in neuroscience are discussed, focusing on important challenges and future prospects.

Monoacylglycerol Lipase Inhibition Prevents Short-Term Mitochondrial Dysfunction and Oxidative Damage in Rat Brain Synaptosomal/Mitochondrial Fractions and Cortical Slices: Role of Cannabinoid Receptors.

Inhibition of enzymes responsible for endocannabinoid hydrolysis represents an invaluable emerging tool for the potential treatment of neurodegenerative disorders. Monoacylglycerol lipase (MAGL) is the enzyme responsible for degrading 2-arachydonoylglycerol (2-AG), the most abundant endocannabinoid in the central nervous system (CNS). Here, we tested the effects of the selective MAGL inhibitor JZL184 on the 3-nitropropinic acid (3-NP)-induced short-term loss of mitochondrial reductive capacity/viability and oxidative damage in rat brain synaptosomal/mitochondrial fractions and cortical slices. In synaptosomes, while 3-NP decreased mitochondrial function and increased lipid peroxidation, JZL184 attenuated both markers. The protective effects evoked by JZL184 on the 3-NP-induced mitochondrial dysfunction were primarily mediated by activation of cannabinoid receptor 2 (CB2R), as evidenced by their inhibition by the selective CB2R inverse agonist JTE907. The cannabinoid receptor 1 (CB1R) also participated in this effect in a lesser extent, as evidenced by the CB1R antagonist/inverse agonist AM281. In contrast, activation of CB1R, but not CB2R, was responsible for the protective effects of JZL184 on the 3-NP-iduced lipid peroxidation. Protective effects of JZL184 were confirmed in other toxic models involving excitotoxicity and oxidative damage as internal controls. In cortical slices, JZL184 ameliorated the 3-NP-induced loss of mitochondrial function, the increase in lipid peroxidation, and the inhibition of succinate dehydrogenase (mitochondrial complex II) activity, and these effects were independent on CB1R and CB2R, as evidenced by the lack of effects of AM281 and JTE907, respectively. Our novel results provide experimental evidence that the differential protective effects exerted by JZL184 on the early toxic effects induced by 3-NP in brain synaptosomes and cortical slices involve MAGL inhibition, and possibly the subsequent accumulation of 2-AG. These effects involve pro-energetic and redox modulatory mechanisms that may be either dependent or independent of cannabinoid receptors' activation.

Thallium Induces Antiproliferative and Cytotoxic Activity in Glioblastoma C6 and U373 Cell Cultures via Apoptosis and Changes in Cell Cycle.

Thallium (Tl+) is a heavy metal that causes toxicity in several organs, including the brain. Its cytotoxic profile, combined with its affinity for tumor cells when used as a radioligand for labeling these cells, suggests its potential use as antitumor therapy. In this study, glioblastoma cell lines C6 (from rat) and U373 (from human) were exposed to increased concentrations of thallium(I) acetate (5, 10, 50, 100, or 200 µM) and several toxic endpoints were evaluated, including loss of confluence and morphological changes, loss of cell viability, changes in cell cycle, and apoptosis. Tl+ was detected in cells exposed to thallium(I) acetate, demonstrating efficient uptake mechanism. Confluence in both cell lines decreased in a concentration-dependent manner (50-200 µM), while morphological changes (cell shrinkage and decreased cell volume) were more evident at exposures to higher Tl+ concentrations. For both parameters, the effects of Tl+ were more prominent in C6 cells compared to U373 cells. The same trend was observed for cell viability, with Tl+ affecting this parameter in C6 cells at low concentrations, whereas U373 cells showed greater resistance, with significant changes observed only at the higher concentrations. C6 and U373 cells treated with Tl+ also showed morphological characteristics corresponding to apoptosis. The cytotoxic effects of Tl+ were also assessed in neural and astrocytic primary cultures from the whole rat brain. Primary neural and astrocytic cultures were less sensitive than C6 and U373 cells, showing changes in cell viability at 50 and 100 µM concentrations, respectively. Cell cycle in both brain tumor cell lines was altered by Tl+ in G1/G2 and S phases. In addition, when combined with temozolamide (500 µM), Tl+ elicited cell cycle alterations, increasing SubG1 population. Combined, our novel results characterize and validate the cytotoxic and antiproliferative effects of Tl+ in glioblastoma cells.

Alpha-Mangostin Alleviates the Short-term 6-Hydroxydopamine-Induced Neurotoxicity and Oxidative Damage in Rat Cortical Slices and in Caenorhabditis elegans.

The development, at the experimental level, of therapeutic strategies based on natural products to attenuate neurological alterations in degenerative disorders has gained attention. Antioxidant molecules exhibit both anti-inflammatory and neuroprotective properties. Alpha-mangostin (α-Man) is a natural xanthonoid isolated from the mangosteen tree with demonstrated antioxidant and cytoprotective properties. In this study, we investigated the antioxidant and protective properties of α-Man, both ex vivo and in vivo. We assessed the mitochondrial reductant capacity and oxidative damage to lipids in rat cortical slices, and several endpoints characteristic of physiological stress in the nematode, Caenorhabditis elegans (C. elegans), upon exposure to the parkinsonian neurotoxin, 6-hydroxydopamine (6-OHDA). In rat cortical slices, α-Man (25 and 50 µM) reduced the 6-OHDA (100 µM)-induced oxidative damage to lipid levels, but failed to reverse loss in cell viability. In wild-type (N2) C. elegans, α-Man (5-100 µM) protected against 6-OHDA (25 mM)-induced decrease in survival when administered either as pre- or post-treatment. Protective effects of α-Man were also observed on survival in the VC1772 strain (skn-1 KO-) exposed to 6-OHDA, though the extent of the protection was lesser than in the wild-type N2 strain. However, α-Man (5-50 µM) failed to attenuate the 6-OHDA-induced motor alterations in the N2 strain. The loss of lifespan induced by 6-OHDA in the N2 strain was fully reversed by high concentrations of α-Man. In addition, while 6-OHDA decreased the expression of glutathione S-transferase in the CL2166 C. elegans strain, α-Man preserved and stimulated the expression of this protein. α-Man (25 µM) also prevented 6-OHDA-induced dopaminergic neurodegeneration in the BZ555 C. elegans strain. Altogether, our novel results suggest that α-Man affords partial protection against several, but not all, short-term toxic effects induced by 6-OHDA in cortical slices and in a skn-1-dependent manner in C. elegans.

Oxidant/antioxidant status in premenstrual syndrome.

PURPOSE: To investigate oxidant/antioxidant status in premenstrual syndrome (PMS). METHODS: Study group (n = 20) consisted of PMS and control group (n = 21) consisted of normal menstruating women. The serum oxidant status was assessed by the lipid hydroperoxide (LHP), malondialdehyde (MDA) and protein carbonyl (PC); the antioxidant status was assessed by the total thiol (T-SH) and total antioxidant capacity (TAC). RESULTS: The study and control groups revealed no statistical difference, in terms of day 3 LHP, MDA, PC, T-SH and TAC levels. There were no significant differences between groups in terms of day 21 MDA, PC and T-SH levels. However, day 21 LHP levels were increased and TAC levels were decreased in the study group compared with the control group. CONCLUSION: Increased oxidative stress and reduced antioxidant capacity may occur in PMS. It can be speculated that the imbalance of oxidant/antioxidant systems may be a cause or the consequence of the various stress symptoms in PMS.

Combatting Nitrosative Stress and Inflammation with Novel Substituted Triazinoindole Inhibitors of Aldose Reductase in PC12 Cells Exposed to 6-Hydroxydopamine Plus High Glucose.

Cellular redox dysregulation produced by aldose reductase (AR) in the presence of high blood sugar is a mechanism involved in neurodegeneration commonly observed in diabetes mellitus (DM) and Parkinson's disease (PD); therefore, AR is a key target for treatment of both diseases. The substituted triazinoindole derivatives 2-(3-thioxo-2H-[1,2,4]triazino[5,6-b]indol-5(3H)-yl) acetic acid (cemtirestat or CMTI) and 2-(3-oxo-2H-[1,2,4]triazino[5,6-b]indol-5(3H)-yl) acetic acid (COTI) are well-known AR inhibitors (ARIs). The neuroprotective properties of CMTI, COTI, the clinically used epalrestat (EPA), and the pyridoindole antioxidants stobadine and SMe1EC2 were all tested in the neurotoxic models produced by hyperglycemic glucotoxicity (HG, 75 mM D-glucose, 72 h), 6-hydroxydopamine (6-OHDA), and HG+6-OHDA models in PC12 cells. Cell viability decreased in all toxic models, increased by 1-5 µM EPA, and decreased by COTI at ≥ 2.5 µM. In the HG model alone, where compounds were present in the medium for 24 h after a continuous 24-h exposure to HG, cell viability was improved by 100 nM-5 µM EPA, 1-10 µM ARIs, and the antioxidants studied, but decreased by EPA at ≥ 10 µM. In the 6-OHDA model alone, where cells were treated with compounds for 24 h and further exposed to 100 µM 6-OHDA (8 h), only the antioxidants protected cell viability. In the HG+6-OHDA model, where cells were treated with all compounds (1 nM to 50 µM) for 48 h and exposed to 75 mM glucose for 24 h followed by incubation with 6-OHDA for 8 h, cell viability was protected by 100 nM-10 µM ARIs and 100-500 nM EPA, but not by antioxidants. All ARIs inhibited the HG+6-OHDA-induced increase in iNOS, IL-1ß, TNF-α, 3-NT, and total oxidant status at 1-50 µM, while increased SOD, CAT, GPx, and total antioxidant status at 1-10 µM. EPA and CMTI also reduced the HG+6-OHDA-induced increase in the cellular levels of nuclear factor kB (NF-KB). The neuroprotective potential of the novel ARIs and the pyridoindole antioxidants studied constitutes a promising tool for the development of therapeutic strategies against DM-induced and PD-related neurodegeneration.