Search Results (230)

Umbilical cord mesenchymal stem cells (UCMSCs)-derived small extracellular vehicles (sEVs) are reported to offer therapeutic effects in regenerative medicine, but they lack safety and biodistribution profiles to support smooth translation at the clinical stage and regulatory requirements. Our study aimed to determine the safety and biodistribution profile in a healthy animal model before application in the metabolic syndrome model. Method: Healthy male Sprague Dawley (SD) rats were given an intravenous (IV) injection of normal saline (control group) or pooled fetal UCMSCs-derived sEVs (treated group) every three weeks for 90 days. Morbidity and mortality observation (daily), physical measurements (weekly), selected serum biochemistry (every three weeks), and hematology (every three weeks) were performed for 90 days. Acute toxicity (on day 14) and sub-chronic toxicity (on day 90) were assessed for gross necropsy, relative organ weight, and histopathological assessment of lungs, liver, spleen, kidney, and lymph nodes. Separately, a biodistribution study was conducted with the sEVs preparations labeled with PKH26 fluorescent dye, given intravenously to the rats. The organs were harvested 24 h post-injection. There were no drastic changes in either group’s morbidity or mortality, physical, hematological, and biochemistry evaluation. The histopathological assessment concluded moderate (focal) inflammation in the treated group’s kidneys and signs of recovery from the inflammation and vascular congestion in the liver. A biodistribution study revealed a higher accumulation of sEVs in the spleen. Multiple IV injections of the pooled fetal UCMSCs-derived sEVs in healthy male SD rats were deemed safe. The sEVs were abundantly distributed in the spleen 24 h post-injection. Full article

Alzheimer’s disease (AD) is the most common cause of dementia in older adults and is becoming a major public health crisis as life expectancy increases worldwide. A major contributor to this disease is a deficiency in melatonin signaling. We have recently synthesised a series of melatonin analogs containing donepezil fragments. These compounds have been tested both in silico and in vitro. In this study, a particularly potent compound, 3a, was evaluated in a hybrid rat model of melatonin deficiency and AD. Rats underwent pinealectomy followed one week later by bilateral intracerebroventricular infusion of Aβ_1-42 (1 µg/µL). A 14-day subchronic treatment with compound 3a was started simultaneously with the neurotoxin infusion. Melatonin was used as a reference drug, while a matched sham group received vehicle treatment. One week after the Aβ_1-42 infusion, the rats’ cognitive functions were assessed using two Y-maze protocols, object recognition and object location tests. Markers of oxidative stress, including hippocampal glutathione, superoxide dismutase, and malondialdehyde, were assessed by ELISA. Compound 3a effectively prevented cognitive impairment induced by the AD model, and its effects were comparable to those of melatonin. In addition, this melatonin analogue with a donepezil fragment reduced AD-associated oxidative stress and suppressed model-associated increased Aβ_1-42 levels in the hippocampus. Our findings suggest that melatonin analogs containing donepezil fragments are promising therapeutic options for targeting oxidative stress associated with AD. Full article

This study explores the identification, characterization, and biological evaluation of angiotensin I-converting enzyme (ACE)-inhibitory peptides derived from enzymatic hydrolysates of crucian carp swim bladders. Following sequential purification by size-exclusion and reversed-phase chromatography, two bioactive peptides—Hyp-Gly-Ala-Arg (Hyp-GAR) and Gly-Ala-Hyp-Gly-Ala-Arg (GA-Hyp-GAR)—were identified using ultra-high-performance liquid chromatography coupled with linear ion trap–Orbitrap tandem mass spectrometry. The synthetic peptides demonstrated potent ACE-inhibitory activity in vitro, with IC₅₀ values of 12.2 μM (Hyp-GAR) and 4.00 μM (GA-Hyp-GAR). Molecular docking and enzyme kinetics confirmed competitive inhibition through key interactions with ACE active site residues and zinc coordination. In vivo antihypertensive activity was evaluated in spontaneously hypertensive rats, revealing that GA-Hyp-GAR significantly reduced systolic blood pressure in a dose-dependent manner. At a dose of 36 mg/kg, GA-Hyp-GAR reduced systolic blood pressure by 60 mmHg—an effect comparable in magnitude and timing to that of captopril. Mechanistically, GA-Hyp-GAR modulated levels of angiotensin II, bradykinin, endothelial nitric oxide synthase, and nitric oxide. A 90-day subchronic oral toxicity study in mice indicated no significant hematological, biochemical, or histopathological alterations, supporting the peptide’s safety profile. These findings suggest that GA-Hyp-GAR is a promising natural ACE inhibitor with potential application in functional foods or as a nutraceutical for hypertension management. Full article

PFASs are widely present and persistent in the environment, and exposure can occur via multiple pathways. Human and animal PFAS exposures have been associated with alterations in thyroid hormones, hepatotoxicity, and other adverse effects. This study evaluated the subchronic toxicities of four specific PFASs in 90-day oral rat studies. Studies were conducted in male and female Sprague–Dawley rats exposed to PFASs in corn oil via oral gavage. The PFASs studied were 1H,1H,9H-perfluorononyl acrylate (PFNAC), 1H,1H,2H,2H-perfluorohexyl iodide (PFHI), methyl heptafluoropropyl ketone (MHFPK), and 2-chloro-2,3,3,3-tetrafluoropropanoic acid (CTFPA). High doses were 10 mg/kg-day (male) and 30 mg/kg-day (female) for PFNAC, 200 mg/kg-day for PFHI, 300 mg/kg-day for MHFPK, and 30 (male) and 100 mg/kg-day (female) for CTFPA. The four lower doses for each PFAS were spaced at two- or threefold dose increments. The most consistent effect was dose-dependent increases in the relative and absolute liver weights for PFNAC, PFHI, and CTFPA but not for MHFPK. Increased liver weights were correlated with findings of hepatocellular hypertrophy. Increased kidney weights for PFNAC and PFHI were correlated with increased incidence of minimal tubule epithelial hypertrophy (PFNAC) or increased incidence and severity of chronic progressive nephropathy and hyaline droplet accumulation (PFHI). There were no compound-related effects on morbidity and mortality or overt signs of toxicity. Full article

Medium chain triglycerides (MCTs) are regarded as an important ingredient for functional foods and nutraceuticals. Cinnamomum camphora seed kernel oil (CCSKO) contains more than 95% medium chain fatty acids (MCFAs), which is a significantly higher level than palm kernel oil (62%) and coconut oil (55%). However, the safety assessment of CCSKO, as the only natural MCT oil rich in capric acid and lauric acid found so far in the world, has not been fully verified. The study aimed to investigate the 90-day sub-chronic oral toxicity and teratogenicity of CCSKO. In the sub-chronic oral toxicity study, no clinically significant adverse events occurred in male or female Sprague–Dawley (SD) rats with CCSKO daily administration for 13 weeks. Moreover, there were no dose–response relationships between CCSKO and body-weight gain, food intake and food utilization in male or female SD rats. No significant differences (p > 0.05) were found in the hematological properties or organ weights between the male and female SD rats. In the teratogenicity test, no toxicological signs were observed in either Wister pregnant rats or fetuses. The no-observed-adverse-effect level of CCSKO was determined to be more than 4 mL/kg body weight. These results suggested that CCSKO may be an excellent edible oil with high oral safety. Full article

Recently, compounds of plant origin have been the focus of increased scientific interest. Micromeria frivaldszkyana is a rare species endemic to Bulgaria, whose biological activity remains unknown. This article aims to evaluate the subchronic toxicity of Micromeria frivaldszkyana methanolic extract and its effect on cognition in rats. Following 90 days of oral administration, a histopathological evaluation of brain, kidney, and liver tissues was conducted. Additionally, serum levels of total bilirubin (TB), conjugated bilirubin (CB), alanine aminotransferase (ALT), aspartate aminotransferase (AST), creatinine (CR), uric acid (UA), and urea (U) were measured. Cognitive function was studied after 7 d of treatment using activity cage test, along with tests for active memory, passive memory, anxiety, spatial and working memory, and explorative activity. The experiments showed no toxic effects of the extract in subchronic application. No adverse effects on brain function were observed after 14 days of treatment. While the extract increased the motor activity of the animals, it did not significantly improve the learning and memory processes. In conclusion, the methanolic extract of Micromeria frivaldszkyana in doses 250 and 500 mg/kg bw did not induce toxicity after 90-day treatment in rats. These doses did not significantly affect central nervous system (CNS) functions, although increased motor activity was observed after 14 days of treatment with the extract. Full article

Arthrospira platensis, a filamentous photosynthetic cyanobacterium, is widely recognized for its high nutritional value, broad spectrum of bioactive compounds, and excellent safety profile, making it a promising natural source for health-promoting applications. This study aimed to profile the phenolic constituents of an ethanolic extract of A. platensis (EAP) using HPLC-DAD-ESI-MS and to investigate its pharmacological effects in attenuating acute and sub-chronic experimental colitis, as well as its antioxidant and antifungal properties. Colitis was induced in BALB/c mice by intrarectal administration of 2,4-dinitrobenzenesulfonic acid (DNBS), followed by oral administration of EAP at doses of 50, 100, and 200 mg/kg. Phenolic profiling revealed eight major compounds, with a cumulative content of 6.777 mg/g of extract, with Pyrogallol, Ferulic acid, and Chlorogenic acid being the most abundant. In vivo, EAP treatment significantly reduced the Disease Activity Index (DAI), alleviated macroscopic colonic damage, and preserved colonic mucosal integrity in both inflammatory phases. Biochemical analyses revealed significant reductions in myeloperoxidase (MPO) activity, nitric oxide (NO), and malondialdehyde (MDA) levels, accompanied by increased reduced glutathione (GSH) content and catalase activity. In vitro, EAP demonstrated notable antioxidant effects, including 56% DPPH and 47% ABTS radical scavenging activities, and an 81% ferrous ion-chelating capacity. Furthermore, it exhibited antifungal activity, with inhibition zones of 20 mm against Candida albicans and 15 mm against Aspergillus flavus, respectively. These findings highlight the multitarget bioactivity of EAP and support its potential as a natural agent for managing intestinal inflammation and oxidative stress across both acute and sub-chronic phases. Full article

Microcystin-leucine arginine (MC-LR) is a prominent water pollutant known for its potent hepatic toxicity. However, the effects of subchronic exposure to environmentally relevant concentrations of MC-LR on the fish liver remain poorly understood. This study aimed to systematically evaluate the impact of subchronic MC-LR exposure on the liver of darkbarbel catfish (Tachysurus vachelli). A total of 270 one-year-old fish were exposed to MC-LR (0, 2, and 5 μg/L) for 28 days and sampled on days 14 (D14) and 28 (D28). Histopathological analysis revealed marked hepatic inflammation in the MC-LR treatment groups, manifested as cellular degeneration, hyperemia, and inflammation. MC-LR exposure induced oxidative stress, evidenced by elevated malondialdehyde (MDA) levels and compensatory upregulation of superoxide dismutase (SOD) activity on D28. While hepatic lipid profiles were not altered by low-dose MC-LR, significant elevation of low-density lipoprotein cholesterol (LDL-C) specifically on D28 indicated incipient lipid metabolic disorder. Metabolomic analysis demonstrated a higher sensitivity, highlighting the stress response of the liver to low-dose MC-LR exposure. The results suggest MC-LR exposure disrupted hepatic phosphatidylcholine (PC) biosynthesis and inhibited lipoprotein formation, thereby impairing lipid transport and contributing to lipid metabolic disorders. In summary, subchronic exposure to environmentally relevant concentrations of MC-LR-induced hepatic tissue inflammation, oxidative stress, and lipid metabolic disorders in darkbarbel catfish. Full article

Background: Microplastics (MPs) can be inhaled by people. However, the relationships between long-term exposure to inhaled MPs, pulmonary fibrosis, and mitochondrial dysfunction are not completely clear. Methods: SD rats were exposed to a 0.0125, 0.125, 0.31, or 1.25 mg/day dosage of mixed polystyrene MPs (PS-MPs), with the particle sizes ranging from 500 nm to 4 µm, via intratracheal administration, for 7 to 35 consecutive days. Results: PS-MPs with particle sizes ranging from 1 µm to 4 µm were deposited in the lungs. The contents of NFκB-mediated proinflammatory cytokines were increased in the lungs of the rats after 7 days of PS-MP exposure. After exposure to PS-MPs, the degree of collagen deposition and the expression of TGF-β1/Smad increased significantly, and the levels of phosphorylated Akt (p-Akt) and nuclear β-catenin decreased significantly. The number of healthy mitochondria decreased, the expression of mitochondrial fission and fusion proteins increased, and the level of PINK1/Parkin-mediated mitophagy decreased in the lungs of the rats after 7 days of PS-MP exposure. A benchmark dose (BMD) of 0.151 mg/day and a benchmark dose lower confidence limit (BMDL) of 0.031 mg/day were identified on the basis of the subchronic effects of the intratracheal administration of the PS-MPs. Conclusions: Our study provides an in-depth understanding of the potential impacts of MP pollution on respiratory diseases. Full article

Sublancin, an S-linked antimicrobial (glycol) peptide produced by Bacillus subtilis, has emerged as a novel and promising veterinary drug due to its unique antibacterial mechanism, low risk of resistance, and properties that modulate the immune system, reduce inflammation, and promote gut health. This study comprehensively assessed the subchronic (90-day) and chronic (180-day) toxicity of Sprague–Dawley (SD) rats, following the guidelines issued by the Ministry of Agriculture of China. Rats were orally administered sublancin at doses of 2000, 10,000, or 50,000 mg/kg feed, representing 1666–5000 times the efficacious dose (1.0–1.2 mg/kg) reported in mice via the same administration route. Throughout this study, a wide range of physiological and behavioral parameters were monitored to access the toxicity of sublancin, including appetite, water intake, body weight gain, and organ weights. Hematological and biochemical analyses, as well as histopathological examinations of the major organs, were conducted at the end of each study period. The results indicated no adverse effects on any measured parameters at any dose level, with no significant differences observed between the sublancin-treated groups and the control group (p > 0.05). Notably, even the highest dose of 50,000 mg/kg did not induce growth inhibition or physiological dysfunction. A histopathological examination also revealed no tissue abnormalities in the major organs. The no-observed-effect level (NOEL) was determined to be 50,000 mg/kg for both study periods. These results demonstrate the long-term safety of sublancin in Sprague–Dawley rats, with no adverse effects during 180 days of oral administration at doses 1666–5000-fold the documented antimicrobially effective and immune-enhancing doses. Full article

Aflatoxin B1 (AFB1) and Ochratoxin A (OTA) are two of the most potent mycotoxins, recognized for their severe toxicity. In recent years, the consumption of bioactive substances has proven to be a valuable ally in combating their harmful effects on human health. For this purpose, this study evaluates the protective effects of fermented whey (FW) and pumpkin (P), as functional ingredients in bread, on duodenum tissue against sub-chronic toxicity induced by AFB1 and OTA. Nine groups of male and female Wistar rats (n = 5 per sex/group) were exposed to different combinations of AFB1, OTA, FW, and P for 28 days. The gene expression of apoptotic and antioxidant markers, including p53, Bax, Hmox1, NF-κB, and occludin, was measured by quantitative real-time PCR (RT-qPCR). AFB1 + OTA exposure led to an increased expression of p53 and NF-κB, with the downregulation of Bax and Hmox1. Occludin expression, which supports tight junction integrity, remained largely unaffected. Supplementation with FW and FW + P modulated gene expression favorably, offering protection against AFB1 and OTA toxicity. These bioactive components effectively mitigated oxidative stress and apoptosis in duodenal tissue. Notably, the results indicate that the protective effects of FW and P are not sex-dependent. These findings highlight the potential of FW and P as functional ingredients in combating the toxic effects of AFB1 and OTA in vivo. Full article

Diclofenac, a nonsteroidal anti-inflammatory drug widely used worldwide, has been detected in waterbodies at concentrations ranging from ng L⁻¹ to µg L⁻¹. Although diclofenac is not a persistent compound, aquatic organisms may be exposed to this drug for extended periods due to its incorporation into the environment by continuous release from hospitals and municipal discharges. This study aimed to evaluate the toxic effects of diclofenac on the microalga Pseudokirchneriella subcapitata, the cladoceran Daphnia curvirostris, and zebrafish embryos (Danio rerio). Toxicity bioassays for the microalga were performed according to the OECD 201 protocol with diclofenac concentrations of 0, 6.25, 12.5, 25, 50, 75, and 100 mg L⁻¹. For the determination of acute toxicity in the cladoceran (48 h), concentrations of 0, 10, 20, 30, 40, 50, and 60 mg L⁻¹ were tested; in subchronic bioassays, the effect of the drug on the reproductive parameters of D. curvirostris was determined for 21 days with sublethal concentrations of 10.3, 14.4, 17.2, and 21.3 mg L⁻¹. Toxicity bioassays on zebrafish embryos were performed according to the OECD 236 protocol, using concentrations of 0, 1, 2, 4, 6, 8, and 10 mg L⁻¹ of diclofenac. The results confirmed the toxic effects of the drug. The IC₅₀ for the microalga was 16.57 mg L⁻¹, while the LC₅₀ for D. curvirostris and D. rerio was 32.29 and 6.27 mg L⁻¹, respectively. In the microalga, chlorophyll-a and carotenoids increased at a concentration of 3.62 mg L⁻¹ of diclofenac; however, chlorophyll-b decreased at the highest drug concentration (13.51 mg L⁻¹). Protein and lipid concentrations in P. subcapitata exposed to all concentrations were higher than in the control. Chronic diclofenac exposure did not affect the survival of D. curvirostris; however, the cumulative progeny and number of clutches significantly decreased, and the age of first reproduction was delayed at all drug concentrations. Protein concentration in D. curvirostris hatchlings was higher at all diclofenac concentrations; in contrast, the amount of lipids and carbohydrates decreased significantly. In D. rerio, the hatching rate decreased by 40, 51.6, and 80% at concentrations of 6, 8, and 10 mg L⁻¹ diclofenac, respectively, and exposure to the drug caused lethal effects such as coagulation at 24 and 48 hpf; sublethal effects such as edema and curved tail were also observed at concentrations of 2 to 10 mg L⁻¹, and the effects increased with increasing concentration up to 144 hpf. The results demonstrate the vulnerability of aquatic organisms to the toxic effects of diclofenac, suggesting that discharging it into water bodies should be regulated to prevent potential ecological impacts on the various trophic levels of freshwater biota. Full article

Mycotoxins are common natural contaminants of crops and fruits, associated with negative effects on human and animal health. Currently, more than 300 mycotoxins have been identified, but data on their effects and their limits in feed and food are still inconsistent. The European Commission, by directive EC 574/2011, established regulations concerning the maximum limit allowed in farm animals’ feed for aflatoxins, but for all other mycotoxins there are only recommendations (EC 1319/2016) and there are no established limits. Considering their variety and toxic effects, but also the fact that not many details are yet known about the cumulative effects of co-contamination with various mycotoxins, it is necessary to monitor the evolution of their presence in animal feed. The aim of our study was to analyze for a four-year period (2021–2024) the concentrations of six mycotoxins (total aflatoxins-AFT, fumonisins-FB, deoxynivalenol-DON, zearalenone-ZEA, T2/HT2 and ochratoxin (A + B)-OTA), the most frequently encountered in the south area of Romania in poultry, piglets and pig’s complete feed. Our results showed that the maximum highest concentrations were 5.8 ppb for AFT, 4.7 ppm for FB, 1.9 ppm for DON, 62.8 ppb for ZEA, 32.1 ppb for T2/HT2 and 19.7 ppb for OTA irrespective of the type of feed. It should be noted that AFT and ZEA were identified in all samples during the entire monitored period, and the only mycotoxin that exceeded the guidance value was DON, for which the recommendation of 0.9 ppm for pig feed was exceeded. Recent studies demonstrated that sub-chronic and chronic exposure to low concentrations of mycotoxins and specially co-contamination is more common than acute exposure, being able to affect animal health over time by lowering the defense capacity, inducing inflammatory reactions and affecting intestinal health, which in the long term could have important economic consequences. Our survey study can provide important data showing the degree of contamination with mycotoxins in pig and poultry feed including the simultaneous presence of different mycotoxins in this complete feed. Full article

We investigated the cellular and neurophysiological mechanisms underlying the pro-cognitive effects of 5-HT4R activation in hippocampal–prefrontal pathways. Our findings show that, in addition to pyramidal neurons, 30–60% of parvalbumin+ interneurons in the CA1, CA3, and dentate gyrus (DG) of the hippocampus and the anterior cingulate (ACC), prelimbic (PL), and infralimbic (IL) regions of the prefrontal cortex co-express 5-HT4Rs. Additionally, 15% of somatostatin+ interneurons in CA1 and CA3 express 5-HT4Rs. Partial 5-HT4R agonist RS-67333 (1 mg/kg, i.p.) exerted anxiolytic effects and ameliorated short-term (3-min) and long-term (24-h) memory deficits in a mouse model of schizophrenia-like cognitive impairment induced by sub-chronic phencyclidine (sPCP) but did not enhance memory in healthy mice. At the neurophysiological level, RS-67333 normalized sPCP-induced disruptions in hippocampal–prefrontal neural dynamics while having no effect in healthy animals. Specifically, sPCP increased delta oscillations in CA1 and PL, leading to aberrant delta–high-frequency coupling in CA1 and delta–high-gamma coupling in PL. RS-67333 administration attenuated this abnormal delta synchronization without altering phase coherence or signal directionality within the circuit. Collectively, these results highlight the therapeutic potential of 5-HT4R activation in pyramidal, parvalbumin+, and somatostatin+ neurons of hippocampal–prefrontal pathways for mitigation of cognitive and negative symptoms associated with schizophrenia. Full article

Background: Many athletes experience sleep difficulties, and prior research within this cohort suggests that acute supplementation of alpha-lactalbumin (ALAC), a whey protein rich in the amino acid tryptophan, may improve sleep and performance. Therefore, this study investigated whether sub-chronic ALAC supplementation in the evening would improve sleep and physical performance within a poor-sleeping athletic population. Methods: In total, 24 athletically trained participants with sleep difficulties (Athlete Sleep Screening Questionnaire: 8.6 ± 2.2; Pittsburgh Sleep Quality Index: 10.0 ± 3.0) completed this double-blinded, randomised controlled, crossover trial. The participants were supplemented with 40 g of ALAC or control 2 h pre-sleep for seven consecutive nights within habitual settings, with sleep measured via actigraphy. Performance was assessed following the 1-week supplementation period, with the 30 s countermovement jump test, Yo-Yo Intermittent Recovery Test Level 1, and reaction time testing performed in a standardised sequence under controlled conditions. Results: During the ALAC condition, the objective number of awakenings increased (CON: 10.25 ± 5.28, ALAC: 11.01 ± 5.79; p = 0.031), the average jump height reduced (CON: 28.58 ± 5.53 cm, ALAC: 27.68 ± 5.14 cm; p = 0.037), the subjective physical and mental performance capabilities declined in the evening (p < 0.001), and evening negative emotional states (p = 0.001) were reduced. Conclusions: Seven days of ALAC supplementation may not improve the sleep and physical performance of an athletically trained population with mild–moderate sleep difficulties. Future research should recruit populations with more severe sleep difficulties and measure sleep architecture over an extended period to fully ascertain the effects, and potential benefits, of ALAC supplementation for athletes. Full article