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Platelet activation is closely related to thrombosis. Aspirin eugenol ester (AEE) is a novel medicinal compound synthesized by esterifying aspirin with eugenol using the pro-drug principle. Pharmacological and pharmacodynamic experiments showed that AEE has excellent anti-inflammatory, antioxidant, and inhibitory platelet activation effects, preventing thrombosis. However, the regulatory network and action target of AEE in inhibiting platelet activation remain unknown. This study aimed to investigate the effects of AEE on platelets of thrombosed rats to reveal its regulatory mechanism via a multi-omics approach. The platelet proteomic results showed that 348 DEPs were identified in the AEE group compared with the model group, of which 87 were up- and 261 down-regulated. The pathways in this result were different from previous results, including mTOR signaling and ADP signaling at P2Y purinoceptor 12. The metabolomics of heart and abdominal aortic tissue results showed that the differential metabolites were mainly involved in steroid biosynthesis, the citric acid cycle, phenylalanine metabolism, phenylalanine, tyrosine, and tryptophan biosynthesis, and glutathione metabolism. Molecular docking results showed that AEE had a better binding force to both the COX-1 and P2Y12 protein. AEE could effectively inhibit platelet activation by inhibiting COX-1 protein and P2Y12 protein activity, thereby inhibiting platelet aggregation. Therefore, AEE can have a positive effect on inhibiting platelet activation.

Aspirin eugenol ester (AEE) was a novel drug compound with aspirin and eugenol esterified. AEE had various pharmacological activities, such as anti-inflammatory, antipyretic, analgesic, anti-oxidative stress and so on. In this study, it was aimed to investigate the effect of AEE on the acute lung injury (ALI) induced by lipopolysaccharide (LPS) in rats. In vitro experiments evaluated the protective effect of AEE on the LPS-induced A549 cells. The tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and interleukin-1β (IL-1β) were measured in the cell supernatant. The Wistar rats were randomly divided into five groups ( n  = 8): control group, model group (LPS group), LPS + AEE group (AEE, 54 mg·kg -1 ), LPS + AEE group (AEE, 108 mg·kg -1 ), LPS + AEE group (AEE, 216 mg·kg -1 ). The lung wet-to-dry weight (W/D) ratio and immune organ index were calculated. WBCs were counted in bronchoalveolar lavage fluid (BALF) and total protein concentration was measured. Hematoxylin-Eosin (HE) staining of lung tissue was performed. Glutathione (GSH), glutathione peroxidase (GPx), catalase (CAT), antioxidant superoxide dismutase (SOD), total antioxidant capacity (T-AOC), lactate dehydrogenase (LDH), C-reactive protein (CRP), myeloperoxidase (MPO), malondialdehyde (MDA), macrophage mobility inhibitory factor (MIF), TNF-α, IL-6, and IL-1β activity were measured. The metabolomic analysis of rat serum was performed by UPLC-QTOF-MS/MS. From the results, compared with LPS group, AEE improved histopathological changes, reduced MDA, CRP, MPO, MDA, and MIF production, decreased WBC count and total protein content in BALF, pro-inflammatory cytokine levels, immune organ index and lung wet-dry weight (W/D), increased antioxidant enzyme activity, in a dose-dependent manner. The results of serum metabolomic analysis showed that the LPS-induced ALI caused metabolic disorders and oxidative stress in rats, while AEE could ameliorate it to some extent. Therefore, AEE could alleviate LPS-induced ALI in rats by regulating abnormal inflammatory responses, slowing down oxidative stress, and modulating energy metabolism., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Tao, Zhang, Qin, Liu, Li, Bai, Ge, Li and Yang.)

Aspirin eugenol ester (AEE) is a new pharmaceutical compound esterified by aspirin and eugenol, which has anti-inflammatory, antioxidant, and other pharmacological activities. The aim of this study was to investigate the protective effect of AEE on paraquat- (PQ-) induced cell damage of SH-SY5Y human neuroblastoma cells and its potential molecular mechanism. There was no significant change in cell viability when AEE was used alone. PQ treatment reduced cell viability in a concentration-dependent manner. However, AEE reduced the PQ-induced loss of cell viability. Flow cytometry, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), and 4'6-diamidino-2-phenylindole (DAPI) staining were used to evaluate cell apoptosis. Compared with the PQ group, AEE pretreatment could significantly inhibit PQ-induced cell damage. AEE pretreatment could reduce the cell damage of SH-SY5Y cells induced by PQ via reducing superoxide anion, intracellular reactive oxygen species (ROS), and mitochondrial ROS (mtROS) and increasing the levels of mitochondrial membrane potential (ΔΨ m ). At the same time, AEE could increase the activity of glutathione peroxidase (GSH-Px), catalase (CAT), and superoxide dismutase (SOD) and decrease the activity of malondialdehyde (MDA). The results showed that compared with the control group, the expression of p-PI3K, p-Akt, and Bcl-2 was significantly decreased, while the expression of caspase-3 and Bax was significantly increased in the PQ group. In the AEE group, AEE pretreatment could upregulate the expression of p-PI3K, p-Akt, and Bcl-2 and downregulate the expression of caspase-3 and Bax in SH-SY5Y cells. PI3K inhibitor LY294002 and the silencing of PI3K by shRNA could weaken the protective effect of AEE on PQ-induced SH-SY5Y cells. Therefore, AEE has a protective effect on PQ-induced SH-SY5Y cells by regulating the PI3K/Akt signal pathway to inhibit oxidative stress., Competing Interests: The authors declare no conflict of interest., (Copyright © 2021 Zhen-Dong Zhang et al.)

Aspirin eugenol ester (AEE) is a new pharmaceutical compound esterified by aspirin and eugenol, which has anti-inflammatory, antioxidant, and other pharmacological activities. This study is aimed at identifying the protective effect of AEE against H 2 O 2 -induced apoptosis in rat adrenal pheochromocytoma PC12 cells and the possible mechanisms. The results of cell viability assay showed that AEE could increase the viability of PC12 cells stimulated by H 2 O 2 , while AEE alone had no significant effect on the viability of PC12 cells. Compared with the control group, the activities of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px) were significantly decreased, and the content of malondialdehyde (MDA) was significantly increased in the H 2 O 2 group. By AEE pretreatment, the level of MDA was reduced and the levels of SOD, CAT, and GSH-Px were increased in H 2 O 2 -stimulated PC12 cells. In addition, AEE could reduce the apoptosis of PC12 cells induced by H 2 O 2 via reducing superoxide anion, intracellular ROS, and mitochondrial ROS (mtROS) and increasing the levels of mitochondrial membrane potential ( ΔΨ m). Furthermore, the results of western blotting showed that compared with the control group, the expression of p-PI3K, p-Akt, and Bcl-2 was significantly decreased, while the expression of Caspase-3 and Bax was significantly increased in the H 2 O 2 group. In the AEE group, AEE pretreatment could upregulate the expression of p-PI3K, p-Akt, and Bcl-2 and downregulate the expression of Caspase-3 and Bax in PC12 cells stimulated with H 2 O 2 . The silencing of PI3K with shRNA and its inhibitor-LY294002 could abrogate the protective effect of AEE in PC12 cells. Therefore, AEE has a protective effect on H 2 O 2 -induced PC12 cells by regulating the PI3K/Akt signal pathway to inhibit oxidative stress., Competing Interests: The authors declare no conflict of interest., (Copyright © 2021 Zhen-Dong Zhang et al.)

Paraquat (PQ) is an effective and commercially important herbicide that is widely used worldwide. However, PQ is highly toxic and can cause various complications and acute organ damage. Aspirin eugenol ester (AEE) is a potential new compound with anti-inflammatory and antioxidant stress pharmacological activity. The present study was to reveal the therapeutic effects and the protective effect of AEE against PQ-induced acute lung injury (ALI) with the help of PQ-induced oxidative damage in A549 cells and PQ-induced lung injury in rats. AEE might have no significant therapeutic effect on PQ-induced lung injury in rats. However, AEE had a significant protective effect on PQ-induced lung injury in rats. AEE pretreatment significantly reduced the stimulatory effect of PQ on malondialdehyde (MDA), the inhibitory effect of PQ on catalase (CAT) activity, superoxide dismutase (SOD) activity, glutathione peroxidase (GPx) activity, the ratio of GSH/GSSH, the activity of caspase-3 and the overexpression of p38 mitogen-activated protein kinase (MAPK) phosphorylation in vivo. In vitro, A549 cells were treated with 250 μM PQ for 24 h. Incubation of A549 cells with PQ led to apoptosis, and increased the level of superoxide anions, reactive oxygen species (ROS), malondialdehyde and the activity of caspase-3 and up-regulation of phosphorylated p38-MAPK, reduced mitochondrial membrane potential (ΔΨm) and the activity of SOD. However, after 24 h on AEE pretreatment of A549 cells, the above-mentioned adverse reactions caused by PQ were significantly alleviated. In addition, AEE pretreatment reduced p38-MAPK phosphorylation in PQ-treated A549 cells. SB203580, the specific p38-MAPK inhibitor, and p38-MAPK shRNA attenuated the activation of the p38-MAPK signaling pathway. N-acetylcysteine (NAC) reduced the level of phosphorylated p38-MAPK and the production of intracellular ROS and inhibited apoptosis. The results showed that AEE may inhibit PQ-induced cell damage through ROS/p38-MAPK-mediated mitochondrial apoptosis pathway., (Copyright © 2021 The Author(s). Published by Elsevier B.V. All rights reserved.)

The oxidative stress of vessel endothelium is a major risk factor of cardiovascular disorders. Antioxidative stress drugs are widely used in cardiovascular therapy. Aspirin eugenol ester (AEE) is a new pharmaceutical compound synthesized by esterification reaction of aspirin with eugenols and possesses antioxidative activity. The present study was designed to investigate the mechanism how AEE protects human umbilical vein endothelial cells (HUVECs) from H 2 O 2 -induced oxidative stress. H 2 O 2 was given to the HUVECs with or without AEE pretreatment. Changes in the oxidative stress-related factors, including those related to the mitochondria-lysosome axis, were determined with Western blotting, cellular immunofluorescence, and enzyme activity test. The results showed that, in the HUVECs, 300  μ M H 2 O 2 treatment significantly increased the apoptosis rate, MDA concentration, reactive oxygen species (ROS) production, mitochondrial membrane potential, expression of Bax and mature cathepsin D (CTSD), and activity of CTSD and Caspase3 (Cas3) but decreased the expression of Bcl2 and lysosomal membrane stability, while in the HUVECs pretreated with AEE, the above changes caused by either the stimulatory or the inhibitory effect of H 2 O 2 on the relevant factors were significantly reduced. AEE pretreatment significantly enhanced the activity of cellular superoxide dismutase and glutathione peroxidase in the HUVECs. Our findings suggest that AEE effectively reduced H 2 O 2 -induced oxidative stress in the HUVECs via mitochondria-lysosome axis., Competing Interests: The authors declare no conflict of interest., (Copyright © 2019 Mei-Zhou Huang et al.)

Aspirin eugenol ester (AEE) possesses anti-thrombotic, anti-atherosclerotic and anti-oxidative effects. The study aims to clarify the mechanism underlying the anti-atherosclerotic effects of AEE on vascular endothelial dysfunction. Both the high-fat diet (HFD)-induced atherosclerotic rat model and the H 2 O 2 -induced human umbilical vein endothelial cells (HUVECs) model were used to investigate the effects of AEE on vascular endothelial dysfunction. UPLC/QTOF-MS coupled with a multivariate data analysis method were used to profile the variations in the metabolites of HUVECs in response to different treatments. Pretreatment of HUVECs with AEE significantly ameliorated H 2 O 2 -induced apoptosis, the overexpression of E-selectin and VCAM-1, and the adhesion of THP-1 cells. Putative endogenous biomarkers associated with the inhibition of endothelial dysfunction were identified in HUVECs pretreated with AEE in the absence or presence of H 2 O 2 , and these biomarkers were involved in important metabolic pathways, including amino acid metabolism, carbohydrate metabolism, and glutathione metabolism. Moreover, in vivo, AEE also significantly reduced vascular endothelial dysfunction and decreased the overexpression of VCAM-1 and E-selectin. Based on our findings, the mechanism underlying the anti-atherosclerotic effects of AEE might be related to a reduction in vascular endothelial dysfunction mediated by ameliorating alterations in metabolism, inhibiting oxidative stress, and decreasing the expression of adhesion molecules.

Aspirin eugenol ester (AEE) was a promising drug candidate for treating inflammation, pain and fever and preventing cardiovascular diseases with fewer side effects than its precursors. Previous researches indicated that AEE could markedly inhibit agonist-induced platelet aggregation in vitro and ex vivo, however, the anti-platelet aggregation mechanisms of AEE remain to be defined. Here, AEE in vitro effects on agonist-induced granule-secretion, intercellular Ca 2+ mobilization and thromboxane A 2 (TXA 2 ) generation were examined. Vasodilator-stimulated phosphoprotein (VASP), mitogen-activated protein kinase (MAPK), Akt, Sirt 1 and CD40L expressions were also studied. In agonist-activated platelets in vitro, AEE markedly attenuated granule secretion markers (P-selectin expression and ATP release), intercellular Ca 2+ mobilization and thromboxane B 2 (TXB 2 ) formation. AEE also attenuated CD40L activation, suppressed extracellular-signal-regulated protein kinase 2 (ERK2), c-Jun N-terminal kinase 1 (JNK1) and Akt phosphorylation, and recovered Sirt1 expression, but the activation of p38, VASP Ser157 and VASP Ser239 , and the levels of cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) were not affected by AEE. Overall, this study demonstrates that AEE inhibits agonist-induced platelet aggregation in vitro by regulating PI3K/Akt, MAPK and Sirt 1/CD40L pathways., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Background and Purpose: Aspirin eugenol ester (AEE) is a new drug compound synthesized by combining aspirin with eugenol. It was reported to possess anti-thrombotic, anti-atherosclerotic, and anti-oxidative effects. However, its molecular mechanism against oxidative injury is unclear. This study investigated how AEE affected the oxidative injury of vascular endothelial cells in vivo and in vitro., Experimental Approach: A hamster model of atherosclerosis induced by a high fat diet (HFD) and an in vitro model of oxidative stress, H 2 O 2 -induced apoptosis of HUVECs, were used to investigate the anti-oxidative effects of AEE., Key Results: AEE significantly reduced the stimulatory effect of HFD on malondialdehyde, the inhibitory effect of HFD on SOD activity and GSH/GSSG ratio, and the overexpression of inducible NOS (iNOS) in the aorta. In vitro, incubation of HUVECs with H 2 O 2 led their apoptosis, dysfunctions of the NO systems (including increased iNOS activity, decreased endothelial NOS activity, and increased production of NO), an imbalance in calcium homeostasis and energy metabolism with an increase in intracellular free calcium and decrease in ATP, and a down-regulation of Nrf2. In contrast, in the HUVECs pretreated with 1 μM AEE for 24 hr, the above adverse effects induced by H 2 O 2 were significantly ameliorated. Moreover, the decrease in NO production and activity of iNOS induced by AEE was significantly attenuated in Nrf2-inhibited HUVECs., Conclusion and Implication: AEE protects vascular endothelial cells from oxidative injury by regulating NOS and Nrf2 signalling pathways. This suggests that AEE is a novel potential agent for the prevention of atherosclerosis., (© 2019 The British Pharmacological Society.)

Background: Hyperlipidemia, with an increasing of prevalence, has become one of the common metabolic diseases in companion animal clinic. Aspirin eugenol ester (AEE) is a novel compound that exhibits efficacious anti-hyperlipidemia activities. However, its mechanisms are still not completely known. The objective of present study was to investigate the intervention effects of AEE on cecal contents metabonomics profile and microbiota in hyperlipidemia rats., Results: Three groups of rats were fed with a control diet, or high fat diet (HFD) containing or not AEE. The results showed the beneficial effects of AEE in HFD-fed rats such as the reducing of aspartate aminotransferase (AST) and total cholesterol (TCH). Distinct changes in metabonomics profile of cecal contents were observed among control, model and AEE groups. HFD-induced alterations of eight metabolites in cecal contents mainly related with purine metabolism, linoleic acid metabolism, glycerophospholipid metabolism, sphingolipid metabolism and pyrimidine metabolism were reversed by AEE treatment. Principal coordinate analysis (PCoA) and cluster analysis of microbiota showed altered patterns with distinct differences in AEE group versus model group, indicating that AEE treatment improved the negative effects caused by HFD on cecal microbiota. In addition, the correction analysis revealed the possible link between the identified metabolites and cecal microbiota., Conclusions: This study showed regulation effects of AEE on cecal contents metabonomics profile and microbiota, which could provide information to reveal the possible underlying mechanism of AEE on hyperlipidemia treatment.

The main objective of this study was to investigate the ameliorative effects of aspirin eugenol ester (AEE) in hyperlipidemic rat. After five-week oral administration of AEE in high fat diet (HFD)-induced hyperlipidemic rats, the impact of AEE on plasma and urine metabonomics was investigated to explore the underlying mechanism by UPLC-Q-TOF/MS analysis. Blood lipid levels and histopathological changes of liver, stomach and duodenum were also evaluated after AEE treatment. Without obvious gastrointestinal (GI) side effects, AEE significantly relieved fatty degeneration of liver and reduced triglyceride (TG), low density lipoprotein (LDL) and total cholesterol (TCH) (P<0.01). Clear separations of metabolic profiles were observed among control, model and AEE groups by using principal component analysis (PCA) and orthogonal partial least-squares-discriminate analysis (OPLS-DA). 16 endogenous metabolites in plasma and 18 endogenous metabolites in urine involved in glycerophospholipid metabolism, fatty acid metabolism, fatty acid beta-oxidation, amino acid metabolism, TCA cycle, sphingolipid metabolism, gut microflora and pyrimidine metabolism were considered as potential biomarkers of hyperlipidemia and be regulated by AEE administration. It might be concluded that AEE was a promising drug candidate for hyperlipidemia treatment. These findings could contribute to the understanding of action mechanisms of AEE and provide evidence for further studies., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Background: Aspirin and eugenol were esterified to synthesize aspirin eugenol ester (AEE). As a pale yellow and odourless crystal, AEE reduced the gastrointestinal damage of aspirin and vulnerability of eugenol. The study was conducted to evaluate the preventive effects of AEE on blood lipids in rats with high fat diet (HFD)., Methods: Suspensions of AEE and simvastatin were prepared in 5% carboxymethyl cellulose sodium (CMC-Na). In order to observe the intervention effects, the drugs and HFD were administrated at the same time. Based on individual weekly body weight (BW), AEE was intragastrically administrated at the dosage of 18, 36 and 54 mg/kg. Simvastatin (10 mg/kg) and CMC-Na (20 mg/kg) were used as control drug. After 6 weeks of administration, the changes of BW and blood lipid indices including triglyceride (TG), low density lipoprotein (LDL), high density lipoprotein (HDL) and total cholesterol (TCH) were determined in the experiment., Results: The rat blood lipids profile in model group was remarkably different after feeding 6-weeks HFD. TG, TCH and LDL indexes in model group were increased significantly compared with those in control group (p < 0.01). AEE at the dosage of 54 mg/kg significantly decreased levels of TG, TCH and LDL (p < 0.01), and slowed the rate of BW gain in comparison with model group (p < 0.05). Moreover, high dose AEE showed better effects than simvastatin on reducing TCH level and similar effects on TG, HDL and LDL., Conclusion: AEE could remarkably reduce levels of TG, TCH and LDL in rats with high fat diet, and slow the rate of body weight gain. It was conducted that AEE was a potential candidate on reducing blood lipids level. The mechanism of action of AEE should be investigated in further studies.

Background: Based on the prodrug principle, aspirin and eugenol, as starting precursors, were esterified to synthesize aspirin eugenol ester (AEE). The aim of the present study was to evaluate the antithrombotic effect of AEE in an animal disease model. In order to compare the therapeutic effects of AEE and its precursors, aspirin, eugenol and a combination of aspirin and eugenol were designed at the same molar quantities as the AEE medium dose in the control group., Methods: After oral administration of AEE (dosed at 18, 36 and 72 mg/kg) for seven days, rats were treated with k-carrageenan to induce tail thrombosis. Following the same method, aspirin (20 mg/kg), eugenol (18 mg/kg) and 0.5 % CMC-Na (30 mg/kg) were administered as control drug. Different drug effects on platelet aggregation, hemorheology, TXB2/6-keto-PGF1α ratio and blood biochemistry were studied., Results: AEE significantly inhibited ADP and AA-induced platelet aggregation in vivo. AEE also significantly reduced blood and plasma viscosity. Moreover, AEE down-regulated TXB2 and up-regulated 6-keto-PGF1α, normalizing the TXB2/6-keto-PGF1α ratio and blood biochemical profile. In comparison with aspirin and eugenol, AEE produced more positive therapeutic effects than its precursors under the same molar quantity., Conclusion: It may be concluded that AEE was a good candidate for new antithrombotic and antiplatelet medicine. Additionally, this study may help to understand how AEE works on antithrombosis in different ways.

Background: Aspirin eugenol ester (AEE) is a promising drug candidate for treatment of inflammation, pain and fever and prevention of cardiovascular diseases with less side effects. The experiment will be conducted to investigate the efficacy of AEE on curing hyperlipidemia in Wistar rats. The rats were fed with high fat diet (HFD) for 8 weeks to induce hyperlipidemia., Results: Compared with the model group, the results showed that AEE at 54 mg/kg dosage could significantly decrease the hyperlipidemia indexes including triglyceride (TG), low density lipoprotein (LDL) and total cholesterol (TCH) (p < 0.01), increase high density lipoprotein (HDL) (p < 0.05) for five weeks drug administration. Meanwhile, simvastatin had same effect on hyperlipidemia indexes such as TG, LDL, TC, but no significant increase in HDL., Conclusion: AEE was effective against hyperlipidemia and had better anti-hyperlipidemic effect than its component, acetylsalicylic acid (Aspirin, ASA), eugenol and integration of ASA and eugenol. Under the experimental circumstance, the optimal dose of AEE to cure hyperlipidemia is 54 mg/kg for five weeks in Wistar rats.

Based on the prodrug principle, aspirin eugenol ester (AEE) was synthesized, which can reduce the side effects of aspirin and eugenol. As a good candidate for new antithrombotic and anti-inflammatory medicine, it is essential to evaluate its preventive effect on thrombosis. Preventive effect of AEE was investigated in κ-carrageenan-induced rat tail thrombosis model. AEE suspension liquids were prepared in 0.5% sodium carboxymethyl cellulose (CMC-Na). AEE was administrated at the dosage of 18, 36 and 72 mg/kg. Aspirin (20 mg/kg), eugenol (18 mg/kg) and 0.5% CMC-Na (30 mg/kg) were used as control drug. In order to compare the effects between AEE and its precursor, integration of aspirin and eugenol group (molar ratio 1:1) was also designed in the experiment. After drugs were administrated intragastrically for seven days, each rat was injected intraperitoneally with 20 mg/kg BW κ-carrageen dissolved in physiological saline to induce thrombosis. The length of tail-thrombosis was measured at 24 and 48 hours. The blank group just was given physiological saline for seven days without κ-carrageenan administrated. The results indicated that AEE significantly not only reduced the average length of thrombus, PT values and FIB concentration, but also reduced the red blood cell (RBC), hemoglobin (HGB), hematocrit (HCT) and platelet (PLT). The effects of AEE on platelet aggregation and anticoagulant in vitro showed that AEE could inhibit adenosine diphosphate (ADP)-induced platelet aggregation as dose-dependence but no notable effect on blood clotting. From these results, it was concluded that AEE possessed positive effect on thrombosis prevention in vivo through the reduction of FIB, PLT, inhibition of platelet aggregation and the change of TT and PT values.

Intestinal inflammation is a complex and recurrent inflammatory disease. Pharmacological and pharmacodynamic experiments showed that aspirin eugenol ester (AEE) has good anti-inflammatory, antipyretic, and analgesic effects. However, the role of AEE in regulating intestinal inflammation has not been explored. This study aimed to investigate whether AEE could have a protective effect on LPS-induced intestinal inflammation and thus help to alleviate the damage to the intestinal barrier. This was assessed with an inflammation model in Caco-2 cells and in rats induced with LPS. The expression of inflammatory mediators, intestinal epithelial barrier-related proteins, and redox-related signals was analyzed using an enzyme-linked immunosorbent assay (ELISA), Western blotting, immunofluorescence staining, and RT-qPCR. Intestinal damage was assessed by histopathological examination. Changes in rat gut microbiota and their functions were detected by the gut microbial metagenome. AEE significantly reduced LPS-induced pro-inflammatory cytokine levels (p < 0.05) and oxidative stress levels in Caco-2 cells and rats. Compared with the LPS group, AEE could increase the relative expression of Occludin, Claudin-1, and zonula occludens-1 (ZO-1) and decrease the relative expression of kappa-B (NF-κB) and matrix metalloproteinase-9. AEE could significantly improve weight loss, diarrhea, reduced intestinal muscle thickness, and intestinal villi damage in rats. Metagenome results showed that AEE could regulate the homeostasis of the gut flora and alter the relative abundance of Firmicutes and Bacteroidetes. Flora enrichment analysis indicated that the regulation of gut flora with AEE may be related to the regulation of glucose metabolism and energy metabolism. AEE could have positive effects on intestinal inflammation-related diseases. [ABSTRACT FROM AUTHOR]

Background: Aspirin eugenol ester (AEE) is a novel medicinal compound synthesized by esterification of aspirin with eugenol using the prodrug principle. AEE has the pharmacological activities of being anti-inflammatory, antipyretic, analgesic, anti-cardiovascular diseases, and anti-oxidative stress However, its oral bioavailability is poor, and its intestinal absorption and transport characteristics are still unknown. Objective: The purpose of this study was to investigate the uptake and transport mechanisms of AEE in Caco-2 cells. Methods: The effects of time, concentration, and temperature on the transport and uptake of AEE were studied. Results: The results showed that a higher concentration of salicylic acid (SA) was detected in the supernatant of cell lysates and cell culture medium, while AEE was not detected. Therefore, the content change of AEE was expressed as the content change of its metabolite SA. In the uptake experiment, when the factors of time, concentration, and temperature were examined, the uptake of SA reached the maximum level within 30 min, and there was concentration dependence. In addition, low temperature (4°C) could significantly reduce the uptake of SA in Caco-2 cells. In the transport experiment, under the consideration of time, concentration, and temperature, the transepithelial transport of SA from AP-BL and BL-AP sides was time-dependent. The amount of SA transported in Caco-2 cells increased with the increase of concentration, but the transmembrane transport rate had no correlation with the concentration. This phenomenon may be due to the saturation phenomenon of high concentration. The efflux ratio (ER) was less than 1, which indicated that their intestinal transport mechanism was passive transport. Moreover, the temperature had a significant effect on the transport of AEE. Conclusion: In summary, intestinal absorption of AEE through Caco-2 cell monolayers was related to passive transport. The uptake and transport of AEE were concentration-dependent, and temperature significantly affected their uptake and transport. The absorption and transport characteristics of AEE may contribute to the exploration of mechanisms of absorption and transport of chemosynthetic drugs in vitro. [ABSTRACT FROM AUTHOR]

Aspirin eugenol ester (AEE) is a new potential drug with anti-inflammatory and antioxidant stress pharmacological activity. Paraquat (PQ) is an effective and commercially important herbicide that is widely used worldwide. However, paraquat is highly toxic and can cause various complications and acute organ damage, such as liver, kidney and lung damage. The purpose of this study was to investigate whether AEE has a protective effect on hepatotoxicity induced by PQ in vivo and in vitro. Cell viability, apoptosis rate, mitochondrial function and intracellular oxidative stress were detected to evaluate the protective effect of AEE on PQ-induced BRL-3A (normal rat hepatocytes) cytotoxicity in vitro. In vivo , AEE pretreatment could attenuate oxidative stress and histopathological changes in rat liver induced by PQ. The results showed that AEE could reduce the hepatotoxicity induced by PQ in vivo and in vitro. AEE reduced PQ-induced hepatotoxicity by inhibitingoxidative stress and maintaining mitochondrial function. This study proved that AEE is an effective antioxidant and can reduce the hepatotoxicity of PQ. AEE reduced PQ-induced hepatotoxicity by inhibiting oxidative stress and maintaining mitochondrial function. [ABSTRACT FROM AUTHOR]

Aspirin eugenol ester (AEE) is a new potential pharmaceutical compound possessing anti-inflammatory, anti-cardiovascular disease, and antioxidative stress activity. The pharmacological activities of AEE are partly dependent on its regulation of cell apoptosis. However, it is still unclear how AEE inhibits cell apoptosis on the basis of its antioxidative stress effect. This study aimed to reveal the vascular antioxidative mechanism of AEE in response to H2O2-induced oxidative stress in HUVECs and paraquat-induced oxidative stress in rats. In the different intervention groups of HUVECs and rats, the expression of ASK1, ERK1/2, SAPK/JNK, and p38 and the phosphorylation levels of ERK1/2, SAPK/JNK, and p38 were measured. The effects of ASK1 and ERK1/2 on the anti-apoptotic activity of AEE in the oxidative stress model were probed using the corresponding inhibitors ASK1 and ERK1/2. The results showed that in the HUVECs, 200 μM H2O2 treatment significantly increased the phosphorylation of SAPK/JNK and the level of ASK1 but decreased the phosphorylation of ERK1/2, while in the HUVECs pretreated with AEE, the H2O2-induced changes were significantly ameliorated. The findings were observed in vitro and in vivo. Moreover, inhibition of ASK1 and ERK1/2 showed that ASK1 plays a vital role in the protective effect of AEE on H2O2-induced apoptosis. All findings suggested that AEE protects the vascular endothelium from oxidative injury by mediating the ASK1 pathway. [ABSTRACT FROM AUTHOR]

Exosomes play an essential role in maintaining normal brain function due to their ability to cross the blood-brain barrier. Aspirin eugenol ester (AEE) is a new medicinal compound synthesized by the esterification of aspirin with eugenol using the prodrug principle. Aspirin has been reported to have neuroprotective effects and may be effective against neurodegenerative diseases. This study wanted to investigate how AEE affected neurological diseases in vivo and in vitro. A multi-omics approach was used to explore the effects of AEE on the nervous system. Gene and protein expression changes of BDNF and NEFM in SY5Y cells after AEE treatment were detected using RT-qPCR and Western Blot. The multi-omics results showed that AEE could regulate neuronal synapses, neuronal axons, neuronal migration, and neuropeptide signaling by affecting transport, inflammatory response, and regulating apoptosis. Exosomes secreted by AEE-treated Caco-2 cells could promote the growth of neurofilaments in SY5Y cells and increased the expression of BDNF and NEFM proteins in SY5Y cells. miRNAs in the exosomes of AEE-treated Caco-2 cells may play an important role in the activation of SY5Y neuronal cells. In conclusion, AEE could play positive effects on neurological-related diseases. • The cardiac transcriptome results showed that AEE may prevent neurological diseases. • AEE could improve changes in liver gene expression and gut flora dysbiosis in ApoE-/- male mice caused by HFD. • Exosomes secreted by AEE-treated Caco-2 cells enable the growth of neurofilaments in SY5Y cells. [ABSTRACT FROM AUTHOR]