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. 2024 Oct 6;25(19):10747.
doi: 10.3390/ijms251910747.

Platelet Proteomics and Tissue Metabolomics Investigation for the Mechanism of Aspirin Eugenol Ester on Preventive Thrombosis Mechanism in a Rat Thrombosis Model

Qi Tao  1 , Li-Ping Fan  1 , Ji Feng  1 , Zhi-Jie Zhang  1 , Xi-Wang Liu  1 , Zhe Qin  1 , Jian-Yong Li  1 , Ya-Jun Yang  1
Affiliations

Platelet Proteomics and Tissue Metabolomics Investigation for the Mechanism of Aspirin Eugenol Ester on Preventive Thrombosis Mechanism in a Rat Thrombosis Model

Qi Tao et al. Int J Mol Sci. .

Abstract

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.

Keywords: aspirin eugenol ester (AEE); metabolomics; molecular docking; platelet; proteomics; thrombosis.

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Conflict of interest statement

The authors declare no conflicts of interest.

Figures

Figure 1
Figure 1
Changes in rat platelets. (A) Principal component analysis of normalized protein expression. (B) Histogram of DEP statistics. (C) Venn diagram of DEPs. (D) Volcano plot of DEPs between AEE and model groups. (E) Volcano plot of DEPs between AEE and control groups. (F) Volcano plot of DEPs between model and control groups. (G) Heat map of DEPs between AEE and model groups. (H) Heat map of DEPs between AEE and control groups. (I) Heat map of DEPs between model and control groups.
Figure 2
Figure 2
Reactome pathway analysis of DEPs. (A) AEE group vs. model group. (B) Model group vs. control group. (C) AEE group vs. control group.
Figure 3
Figure 3
GO enrichment analysis of DEPs. (A) AEE group vs. model group. (B) Model group vs. control group. (C) AEE group vs. control group.
Figure 4
Figure 4
KEGG pathway analysis of DEPs. (A) AEE group vs. model group. (B) Model group vs. control group. (C) AEE group vs. control group.
Figure 5
Figure 5
Interaction networks of the DEPs were analyzed using the STRING Database. (A) AEE group vs. control group. (B) AEE group vs. model group. (C) Model group vs. control group.
Figure 6
Figure 6
Effect of AEE on metabolomic profiles of hearts in thrombosed rats. The OPLS-DA score plots of different groups in ESI+ and ESI- modes. ESI+: electrospray ionization in positive ion mode; ESI-: electrospray ionization in negative ion mode. (A,E) The PCA score plots of the heart samples analyzed by the UPLC-Q-TOF/MS in ESI+ and ESI- modes. (B,F) The OPLS-DA score plots for AEE and model groups in ESI+ and ESI- models: ESI+: R2X = 0.842, R2Y = 1, Q2 = 0.949; ESI-: R2X = 0.789, R2Y = 1, Q2 = 0.968. (C,G) OPLS-DA score plots for model and control groups in ESI+ and ESI- models: ESI+: R2X = 0.605, R2Y = 1, Q2 = 0.851; ESI-: R2X = 0.791, R2Y = 0.997, Q2 = 0.797. (D,H) OPLS-DA score plots for AEE and control groups in ESI+ and ESI- models: ESI+: R2X = 0.452, R2Y = 0.996, Q2 = 0.931; ESI-: R2X = 0.723, R2Y = 1, Q2 = 0.949.
Figure 7
Figure 7
Effect of AEE on metabolomic profiles of abdominal aorta in thrombosed rats. The OPLS-DA score plots of different groups in ESI+ and ESI- modes. ESI+: electrospray ionization in positive ion mode; ESI-: electrospray ionization in negative ion mode. (A,E) PCA score plots of abdominal aorta samples analyzed by the UPLC-Q-TOF/MS in ESI+ and ESI- modes. (B,F) OPLS-DA score plots for AEE and model groups in ESI+ and ESI- models: ESI+: R2X = 0.750, R2Y = 1, Q2 = 0.973; ESI-: R2X = 0.889, R2Y = 1, Q2 = 0.953. (C,G) OPLS-DA score plots for model and control groups in ESI+ and ESI- models: ESI+: R2X = 0.765, R2Y = 1, Q2 = 0.944; ESI-: R2X = 0.883, R2Y = 0.1, Q2 = 0.875. (D,H) OPLS-DA score plots for AEE and control groups in ESI+ and ESI- models: ESI+: R2X = 0.681, R2Y = 1, Q2 = 0.959; ESI-: R2X = 0.871, R2Y = 1, Q2 = 0.986.
Figure 8
Figure 8
Heart metabolomics analysis results. (A) Metabolite enrichment analysis. (B) Metabolite pathway analysis. a: Nicotinate and nicotinamide metabolism; b: glycerophospholipid metabolism; and c: steroid biosynthesis.
Figure 9
Figure 9
Abdominal aorta metabolomic analysis results. (A) Metabolite enrichment analysis. (B) Metabolite pathway analysis. a: Phenylalanine metabolism; b: phenylalanine, tyrosine, and tryptophan biosynthesis; c: nicotinate and nicotinamide metabolism; d: glutathione metabolism; e: purine metabolism; and f: steroid hormone biosynthesis.
Figure 10
Figure 10
Docking pattern diagram. (A) AEE and protein P2Y12. (B) AEE and protein COX-1.
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