The Efficacy of Combined Melatonin and Methylphenidate in Attenuating Pyroptosis and Cognitive Deficits Induced by Aluminum Chloride Administration in BALB/C Mice

l-Arginine/nitric oxide pathway and oxidative stress in adults with ADHD: Effects of methylphenidate treatment

IntroductionZearalenone (ZEN), a mycotoxin from Fusarium species, is widely present in contaminated grains and animal products. It exerts estrogen-like effects, disrupting hormonal balance and reproductive function, partly through oxidative stress-induced DNA damage. The ATM-Chk2-p53 pathway is a key mediator of the DNA damage response. Melatonin (MT), a natural antioxidant, supports ovarian function by regulating hormone secretion and reducing oxidative stress. This study explores whether MT alleviates ZEN-induced ovarian and granulosa cell damage via the ATM-Chk2-p53 pathway.MethodsFemale mice were exposed to ZEN (0.8mg/kg) with or without MT (10, 20, or 40 mg/kg) for 28 days. Ovarian morphology, hormone levels, oxidative stress markers, and DNA damage proteins were assessed. GRM02 cells were treated with ZEN (60 μM) and/or MT (100 μM). Apoptosis, cell cycle, oxidative stress, and DNA damage markers were evaluated. ATM-knockout and ATM-activated GRM02 models were used to examine pathway involvement.ResultsZEN caused ovarian atrophy, estrous disruption, reduced E2, FSH, and LH, elevated oxidative stress, and increased γH2AX, pATM, Chk2, and p53 expression. MT restored ovarian function, improved antioxidant capacity, and reduced DNA damage. In GRM02 cells, MT mitigated ZEN-induced G2/M arrest, apoptosis, and oxidative stress. ATM activation enhanced MT’s protective effect, while ATM knockout worsened ZEN toxicity.DiscussionMT protects against ZEN-induced ovarian and cellular damage by reducing oxidative stress and modulating the ATM-Chk2-p53 pathway. These findings highlight MT’s potential as a protective feed additive against mycotoxin-related reproductive toxicity.

ABSTRACTBackgroundTramadol (TM) abuse results in significant cognitive dysfunction. This study aimed to investigate the impact of an 8‐week high‐intensity interval training (HIIT) regimen and methylphenidate (MPH) administration (alone and in combination) on cognitive function and hippocampal oxidative stress markers following chronic TM administration.MethodsFifty‐six adult male rats (200–250 g) were divided into eight groups and received one of the following treatments: tramadol (50 mg/kg, intraperitoneally, 5 days/week for the first month and 3 days/week for the second month), methylphenidate (10 mg/kg, intraperitoneally, 3 times/week for 60 days), HIIT (five sessions/week for 8 weeks), or saline (1 mL, intraperitoneally, daily for 60 days). Learning and memory were assessed using the Morris water maze (MWM) and passive avoidance tests. Hippocampal malondialdehyde (MDA), glutathione peroxidase (GPx), and total antioxidant capacity (TAC) were measured using TBARS and FRAP methods, respectively. Hippocampal nitric oxide (NO) levels were determined with a commercial assay kit.ResultsTramadol induced significant impairments in learning and memory (p < 0.001). MPH, HIIT, and their combination attenuated these deficits. Tramadol and MPH increased MDA and NO levels (p < 0.001) and reduced TAC (p < 0.001). In contrast, HIIT reduced these parameters, even in the presence of MPH. In fact, HIIT reversed the adverse effects of tramadol and MPH by reducing MDA and NO (p < 0.001) and by increasing GPx (p < 0.05) and TAC (p < 0.001).ConclusionAlthough both MPH and HIIT interventions show promise in mitigating tramadol‐induced cognitive deficits, their mechanisms of action appear to differ significantly. HIIT likely exerts its effects by modulating oxidative stress, whereas MPH seems to disrupt it, suggesting distinct underlying mechanisms. Further studies are required to elucidate these mechanisms in greater detail.

The combination of temozolomide (TMZ) and paclitaxel (PTX) is the most commonly used chemotherapy regimen for glioblastoma, but there is no specific treatment for neuroblastoma due to the acquired multidrug resistance. Approximately half of treated glioblastoma patients develop resistance to TMZ and experience serious side effects. Melatonin (MEL), a multifunctional hormone long known for its antitumor effects, has a great advantage in combination cancer therapy thanks to its ability to affect tumors differently than normal cells. This study aims to evaluate the in vitro inhibitory effects of MEL in combination with TMZ on cancer cell viability and to elucidate the underlying mechanisms in the glioblastoma and neuroblastoma cell lines. C6 (Rattus norvegicus) and N1E-115 (Mus musculus) cancer cell lines and C8-D1A (mice) healthy cell lines were used. Cell proliferation was evaluated using the MTT test. IC50 values were determined by probit analysis. Two concentrations of TMZ (IC50 and 1/2 IC50) were used to induce cytotoxicity in the C6 and N1E-115 cell lines, both alone and in combination with PXT and MEL (all at IC50). The viable, dead, and apoptotic cells were determined by image-based cytometry using Annexin V/PI staining. The gene expression related to signaling pathways was assessed by the quantitative reverse transcription polymerase chain reaction (qRT-PCR), and key proteins were identified by the Western blot analysis. MTT assay showed that the combination of TMZ and MEL significantly reduces the viability of both glioblastoma and neuroblastoma cells compared to the vehicle-treated controls. Notably, MEL combined with 1/2 IC50 TMZ showed a significant death rate of cancer cells compared to controls and PTX. According to qRT-PCR data, the TMZ + MEL combination resulted in the upregulation of the genes of antioxidative enzymes (Sod1 and Sod2) and DNA repair genes (Mlh1, Exo1, and Rad18) in both cell lines. Moreover, the levels of Nfkb1 and Pik3cg were significantly reduced following the TMZ + MEL treatment. The combination of MEL with TMZ also enhanced the cell cycle arrest and increased the expression of p53 and pro-apoptotic proteins (Bax and caspase-3), while significantly decreasing the expression of anti-apoptotic protein Bcl-2. Our findings indicate that the combination of MEL with a low dose of TMZ may serve as an upstream inducer of apoptosis. This suggests the potential development of a novel selective therapeutic strategy as an alternative to TMZ for the treatment of both glioblastoma and neuroblastoma.

Aluminum, a widely occurring environmental metal, has been implicated as a neurotoxic agent and is associated with the development of several neurodegenerative conditions, including Alzheimer's disease (AD). Its neurotoxicity is largerly attributed to the induction of oxidative stress , which exacerbates neuroinflammation, leading to cognitive deficits and progressive neuronal dysfunction. Trans-anethole (TA) possesses diverse pharmacological activities, including anti-inflammatory, antioxidant, antifungal, and anticancer effects. However, its neuroprotective potential against AlCl₃-induced neurodegeneration, particularly in the context of memory impairment mediated by inflammation and oxidative stress, remains unknown. Therefore, this study was conducted to evaluate the potential role of TA in mitigating neurodegeneration. To establish an aluminum-induced neuroinflammation-associated neurodegenerative model, rats received oral administration of 150mg/kg AlCl₃ for 90 days. TA was administered at three different dosages between days 31 and 90: 40, 80, and 160mg/kg between days 31 and 90. Cognitive performance was assessed using the Morris water maze (MWM) and passive avoidance test (PAT). Neuroprotective effects were evaluated by analyzing acetylcholinesterase (AChE) activity, oxidative stress markers (catalase, glutathione, and malondialdehyde levels), and neuroinflammatory mediators (NLRP3 inflammasome, Interleukin-1β, and TNF-α) in AlCl₃-exposed rats. TA significantly mitigated aluminum-induced neuronal damage by restoring antioxidant defence, inhibiting AChE activity, and downregulating inflammatory proteins, including NLRP3, TNF-α, and IL-1β. Histopathological analysis further confirmed its neuroprotective role. The findings of this study suggests that TA holds therapeutic potential for neurodegenerative disorders by mitigating memory deficits, neuroinflammation, and oxidative stress.

Aging heightens susceptibility to ischemia-reperfusion (IR) injury, complicating liver transplantation, while the nucleotide-binding oligomerization domain (NOD)-like receptor pyrin domain containing 3 (NLRP3) inflammasome drives IR- and aging-induced inflammation. Although the effects of melatonin (MLT) on IR or aging have been studied separately, its impact on NLRP3 inflammasome activation in age- related IR injury remains unclear. This study investigates the impact of aging on hepatic IR injury, evaluating MLT therapeutic potential. for mitigating age-related damage. Aged and young male Wistar rats underwent 60min of ischemia followed by 6-24h of reperfusion. MLT (1mg/100g body weight) was injected 30min before ischemia, 10min before reperfusion, and 2h after reperfusion. Liver injury, oxidative stress and inflammatory responses, and activation of the NLRP3 inflammasome pathway were evaluated. Aged livers exhibited exacerbated IR injury, marked by elevated transaminases levels, severe histopathological damage, increased oxidative stress and heightened inflammatory responses compared to young IR-injured rats. MLT treatment significantly alleviated liver injury, reducing oxidative stress and inflammatory markers expression. Aging-associated IR injury correlated with increased NLRP3 inflammasome activation and pyroptosis, evidenced by the upregulation of apoptosis-associated speck-like protein containing a CARD (ASC-1), caspase-1 cleavage, interleukin (IL)-1β maturation and increased Il18 and Gsdmd gene expression; while MLT treatment suppressed this activation, downregulating these markers in aged IR-injured livers. These findings highlight the efficacy of MLT in mitigating IR-induced liver damage in aged rats by inhibiting the NLRP3 inflammasome activation, supporting its potential as a therapeutic strategy for age-related liver dysfunction.

Methylphenidate triggers retinal oxidative stress and mitochondrial dysfunction under physiological conditions but has beneficial effects in inflammatory settings.

Chronic stressful situations result in altered monoaminergic activity of neurotransmitters, resulting in various conditions characterized by deficits in learning, memory and attention. Stimulant effects can be visualized in terms of increased cognitive abilities through enhancement of dopamine (DA) release. This study examined cognitive responses and brain DA and 5-hydroxytryptamine (5HT) levels after prolonged methylphenidate (MPH) and modafinil administration, to demonstrate their effect on stress-induced cognitive deficits in rats. Effects on cognition were evaluated by passive avoidance and water maze tests. Furthermore brain levels of DA, homovanillic acid (HVA), dihydroxyphenylacetic acid (DOPAC), 5HT and 5-hydroxyindoleacetic acid (5HIAA) were analyzed by high-performance liquid chromatography coupled with electrochemical detection. We found that both MPH and modafinil improved cognition in both restrained and unrestrained rats, as examined through water maze and passive avoidance tests. Furthermore, these substance were associated with increased brain DA and 5-HT levels. Notabily, we observed decrease in DOPAC and HVA levels, while 5-HIAA levels exhibited a slight increase. The prevention of stress-induced cognitive deficits by MPH and modafinil could be elucidated through the interaction between 5HT and DA in regulating cognitive function.

Neuroprotection via IGF-1 neuronal signaling activation by melatonin and edaravone synergy in methylmercury-induced ALS-like neurotoxicity: Comprehensive analysis of brain regions, spinal cord, CSF, and blood plasma.

Melatonin improves brain function in a model of chronic Gulf War Illness with modulation of oxidative stress, NLRP3 inflammasomes, and BDNF-ERK-CREB pathway in the hippocampus

Chronic epigallocatechin-3-gallate ameliorates learning and memory deficits in diabetic rats via modulation of nitric oxide and oxidative stress

Chronic glaucoma is a multifactorial disease among which oxidative stress may play a major pathophysiological role. We conducted a systematic review and meta-analysis to evaluate the levels of oxidative and antioxidative stress markers in chronic glaucoma compared with a control group. The PubMed, Cochrane Library, Embase and Science Direct databases were searched for studies reporting oxidative and antioxidative stress markers in chronic glaucoma and in healthy controls using the following keywords: “oxidative stress” or “oxidant stress” or “nitrative stress” or “oxidative damage” or “nitrative damage” or “antioxidative stress” or “antioxidant stress” or “antinitrative stress” and “glaucoma”. We stratified our meta-analysis on the type of biomarkers, the type of glaucoma, and the origin of the sample (serum or aqueous humor). We included 22 case-control studies with a total of 2913 patients: 1614 with glaucoma and 1319 healthy controls. We included 12 studies in the meta-analysis on oxidative stress markers and 19 on antioxidative stress markers. We demonstrated an overall increase in oxidative stress markers in glaucoma (effect size = 1.64; 95%CI 1.20–2.09), ranging from an effect size of 1.29 in serum (95%CI 0.84–1.74) to 2.62 in aqueous humor (95%CI 1.60–3.65). Despite a decrease in antioxidative stress marker in serum (effect size = –0.41; 95%CI –0.72 to –0.11), some increased in aqueous humor (superoxide dismutase, effect size = 3.53; 95%CI 1.20–5.85 and glutathione peroxidase, effect size = 6.60; 95%CI 3.88–9.31). The differences in the serum levels of oxidative stress markers between glaucoma patients and controls were significantly higher in primary open angle glaucoma vs primary angle closed glaucoma (effect size = 12.7; 95%CI 8.78–16.6, P < 0.001), and higher in pseudo-exfoliative glaucoma vs primary angle closed glaucoma (effect size = 12.2; 95%CI 8.96–15.5, P < 0.001). In conclusion, oxidative stress increased in glaucoma, both in serum and aqueous humor. Malonyldialdehyde seemed the best biomarkers of oxidative stress in serum. The increase of some antioxidant markers could be a protective response of the eye against oxidative stress.

Sweet persimmon ‘Fuyu’ is highly sensitive to chilling injury (CI), which shortens its storage life. The aim of this study was to assess the effect of melatonin (MLT) dip treatment (100 µmol L −1 ), modified atmosphere packaging (MAP), and combined application of MLT and MAP on reduction of CI in ‘Fuyu’ persimmon fruit at 0 ± 1 °C up to 60 days. An untreated group of ‘Fuyu’ persimmon fruit was used as a control. The combined application of MLT and MAP significantly reduced CI incidence and index on both peel and flesh of persimmon fruit. This treatment also resulted in lower ethylene production, weight loss, soluble solids content (SSC), and better retention of texture properties including hardness, chewiness, gumminess, cohesiveness, and springiness. The combined treatment mitigated oxidative stress by reducing reactive oxygen species (ROS) production and decreasing lipid peroxidation by lowering hydrogen peroxide (H 2 O 2 ), malondialdehyde (MDA), and electrolyte leakage (EL), and reduced activities of lipoxygenase (LOX) and peroxidase (POD) enzymes. Both individual and combined MLT and MAP treatments preserved the antioxidant system by increasing the levels of flavonoids, total antioxidants, ascorbic acid, and glutathione content. These treatments also increased the activities of superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), monodehydroascorbate reductase (MDHAR), dehydroascorbate reductase (DHAR) and glutathione reductase (GR) enzymes. In conclusion, the combination of MLT and MAP effectively alleviated CI, reduced ethylene production, and oxidative stress which resulted in the maintenance of fruit quality of ‘Fuyu’ persimmons and prolonged storage life up to 60 days during cold storage. • ‘Fuyu’ persimmons are highly sensitive to CI during cold storage. • MLT followed by MAP application alleviates CI by reducing oxidative stress. • Ethylene production was downregulated by MLT followed by MAP treatment. • MLT followed by MAP synergistically maintains higher textural fruit quality. • MLT followed by MAP preserves higher antioxidants in persimmon fruit.

Aging is a complex and progressive process involving every organ in the body and it is the result of coordinated biology events that can span decades (1). At cardiovascular level, age-related degeneration and functional decline are quite heterogeneously, nevertheless oxidative stress is a well known pathological process involved. The aim of this study was to investigate the effects of a chronic and long-term treatment with melatonin on functional responses of small mesenteric arteries and on the expression of oxidative stress markers at aorta level of senescence-accelerated prone mice (SAMP8), a model of age-related vascular dysfunction and cognitive decline (2), respect relative controls, senescence-accelerated resistant mice (SAMR1). In the present study were investigated SAMP8 and SAMR1 mice orally treated or not treated for 10 months with melatonin (MelapureTM by Flamma S.p.A.). It was observed that the anticontractile effect of perivascular adipose tissue is impaired in untreated SAMP8, compared with SAMR1. On the contrary, the chronic treatment with melatonin decreased the contractile response to norepinephrine in mesenteric small arteries of SAMP8, restoring an anticontractile effect, probably through melatonin antioxidant mechanisms. In untreated SAMP8 mice was observed at aorta level also an overexpression of oxidative stress and inflammatory markers compared with controls; whereas the long-term treatment with melatonin in SAMP8 was able to increase the expression of some markers of vasculoprotection and to decrease oxidative stress and inflammation. A reduced expression of adiponectin and adiponectin receptor 1 was also observed at visceral fat level of untreated SAMP8 respect SAMR1, while a significant increase was observed after melatonin treatment. In conclusion, melatonin exhibited marked antioxidant and vasculoprotective effects, underlining its potential anti-aging properties at cardiovascular level. Sincere thanks to Flamma S.p.A.-Italy (www.flammagroup.com) for courteously providing the melatonin.

Maintenance of health depends on the ability to respond appropriately to environmental stressors via reciprocal interactions between the body and the brain. In this context, it is well recognized that the pineal hormone melatonin (MLT) plays an important role. T-helper cells bear G-protein-coupled MLT cell membrane receptors and, perhaps, MLT nuclear receptors. Activation of MLT receptors enhances the release of T-helper cell cytokines, such as gamma-interferon and interleukin-2 (IL-2), as well as activation of novel opioid cytokines which crossreact immunologically with both interleukin-4 and dynorphin B. MLT has been reported also to enhance the production of interleukin-1, interleukin-6 and interleukin-12 in human monocytes. These mediators may counteract secondary immunodeficiencies, protect mice against lethal viral and bacterial diseases, synergize with IL-2 against cancer and influence hematopoiesis. Hematopoiesis is influenced by MLT-induced-opioids (MIO) acting on kappa 1-opioid receptors present on bone marrow macrophages. Clinically, MLT could amplify the anti-tumoral activity of low dose IL-2, induce objective tumor regression, and prolong progression-free time and overall survival. MLT seems to be required for the effectiveness of low dose IL-2 in those neoplasias that are generally resistant to IL-2 alone. Similar findings were obtained in a study in which MLT was combined with gamma-interferon in metastatic renal cell carcinoma. In addition, MLT in combination with low-dose IL-2 was able to neutralize the surgery-induced lymphocytopenia in cancer patients. IL-2 treatment in patients results in activation of the immune system and creates the most suitable biological background for MLT. The finding that MLT stimulates IL-12 production from human monocytes only if incubated in presence of IL-2 further supports this concept. On the other hand, high concentrations of MLT have been found in human breast cancer tissue. The MLT concentration, which was 3 orders of magnitude higher than that present in the plasma, correlated positively with good prognostic markers such as estrogen receptor status and nuclear grade. Whether this relates to the immunoneuroendocrine action of MLT remains to be established. Clinical studies are needed on the effect of MLT in combination with IL-2 or other cytokines in cancer patients and viral diseases including HIV-infected patients.