Acrylamide and bisphenol A: two plastic additives increase platelet activation, via oxidative stress

Abstract

Background: Since the mid-20th century, the widespread use of plastics has led to the buildup of harmful byproducts in the environment-most notably acrylamide (AA) and bisphenol A (BPA). These chemicals are now commonly detected in human tissues, raising concerns about their potential health effects. While their presence as environmental pollutants is well known, their specific impact on platelet function and the associated cardiovascular risks remains poorly understood.

Methods: To explore how AA and BPA affect platelet physiology, we performed in vitro assays to assess platelet activation and aggregation following exposure to these compounds. We also used bioinformatic tools to identify potential protein targets in human platelets and carried out molecular docking simulations to investigate how AA and BPA interact with key enzymes involved in platelet regulation.

Results: Both AA and BPA exposure led to a significant increase in platelet activation and aggregation, suggesting an elevated risk of thrombosis. Proteomic analysis identified around 1,230 potential protein targets, with 191 affected by AA and 429 by BPA. These proteins are primarily involved in oxidative stress, apoptosis, and signaling pathways regulated by protein kinase C (PKC), p38α-MAPK, and superoxide dismutase (SOD). Molecular modeling further revealed that AA and BPA form stable complexes with several of these enzymes, indicating direct interference with platelet function.

Discussion and conclusion: Our study shows that AA and BPA can enhance platelet reactivity and aggregation, which are key factors in the development of cardiovascular disease (CVD). By identifying specific molecular pathways and targets affected by these pollutants, we provide new insights into their potential role in promoting thrombotic conditions. These findings highlight the urgent need for greater public health awareness and stronger regulatory efforts to reduce human exposure to AA and BPA.

Keywords: acrylamide; bisphenol; cardiovascular diseases; microplastics; platelets; platelets and cardiovascular diseases.

FIGURE 1

Screening of targets related to Acrylamide (AA) and Bisphenol A (BPA). (A, B) Venn diagram displays 191 and 429 overlapping genes between AA and BPA respectively, and platelet-related target genes. (C–F) A Biological Process network of targets. C, all target protein of AA; D, platelet target protein of AA; E, all target protein of BPA; and F, platelet target protein of BPA.

FIGURE 2

Biological function of Acrylamide (AA) and AA-platelet target protein. (A, D) Gene Ontology (GO) second class enrichment assay of AA and AA-platelets targets. (B, E) Cnetplot depicts the linkages of targets and the top ten biological concepts. (C, F) The bubble chart shows GO terms’ top 10 biological processes (BP).

FIGURE 3

Biological function of Bisphenol A (BPA) and BPA-platelet target protein. (A, D) Gene Ontology (GO) second class enrichment assay of BPA and BPA-platelets targets. (B, E) Cnetplot depicts the linkages of targets and the top ten biological concepts. (C, F) The bubble chart shows GO terms’ top 10 biological processes (BP).

FIGURE 4

Pathways enrichment analysis of Acrylamide (AA) and Bisphenol A (BPA). (A, D) Bar chart showing the ten top AA and BPA pathways. (B, E) Cnetplot depicts the linkages of targets and the top ten pathways. (C, F) Bubble chart of top 10 signaling pathways linked to AA and BPA.

FIGURE 5

Platelet signaling pathways that trigger ROS production by (A) Acrylamide (AA) and (B) Bisphenol A (BPA). The diagram that shows the most important proteins that participate in the production of ROS by platelets also indicates the possible target proteins of AA and BPA. (C) Venn diagram displays overlapping platelet protein targets between three targets of AA and BPA. (D–G) Platelet protein association networks, these network interactions were generated with Cytoscape V3.4 (spring-embedded layout); the protein was represented by text (nodes), and the lines (edges) connecting the two texts signify an interaction between two proteins. (D), all target protein; (E), PKC-platelet target protein; (F), P38a-MAPK-platelet target protein; and (G), SOD-platelet target protein.

FIGURE 6

Interaction between acrylamide and bisphenol with kinases predicted by docking protein-ligand. Representative complexes were obtained by docking prediction using the structures of PKC C1A domain (A, B) and p38 MAPK (C, D). AA and BPA show orange and yellow spheres, respectively. The N-terminal is marked in blue and the C-terminal in red. The purple spheres in PKC correspond to Zn atoms. The graphs summarize the (E) docking score values reported by Glide, and (F) the ΔG_bind values calculated using the MM-GBSA method using Prime. All figures were created using PyMOL.

FIGURE 7

Interaction between acrylamide and bisphenol with Superoxide dismutase isoforms expressed on platelets predicted by docking protein-ligand. Representative complexes were obtained by docking prediction using the structures of SOD1 (A, B) and SOD2 (C, D). AA and BPA show orange and yellow spheres, respectively. The N-terminal is marked in blue and the C-terminal in red. The blue spheres in SOD1 correspond to Zn atoms while the orange spheres are Cu atoms. Meanwhile, the red spheres of SOD2 are Mn atoms. The graphs summarize the (E) docking score values reported by Glide, and (F) the ΔG_bind values calculated using the MM-GBSA method using Prime. All figures were created using PyMOL.

FIGURE 8

Molecular Dynamics Simulations of AA and BPA within a Phospholipid Bilayer. (A) Spatial Dynamics of AA within the Membrane Bilayer. The upper panel illustrates the interactions of AA with the external surface of the lipid membrane, revealing its association patterns. The lower panel showcases the translocation of AA as it traverses across the lipid bilayer. (B) Exploring Interactions of BPA within the Lipid Bilayer. This representation captures the dynamic displacements and insertion events of BPA within the phospholipid bilayer, shedding light on its behavior and influence on the membrane environment.

FIGURE 9

Cytotoxicity exposure induced by Acrylamide (AA) and Bisphenol A (BPA) in platelets. (A) Platelet viability using calcein-AM. The populations of calcein-negative platelets (anti-CD61) were non-viable cells. (B) LDH release from platelets was analyzed with the LDH cytotoxicity assay kit in the supernatant and measured at 490 nm in a microplate reader.

FIGURE 10

Effects on platelet aggregation mediated by Acrylamide (AA) and Bisphenol A (BPA) in platelets. (A–C) Upper panel: representative kinetic aggregation obtained for each condition. (D) the graph summarizes the percentage of platelet aggregation. The results were presented from six independent volunteers (each donor executed as single triplicates) and expressed as mean ± SEM.

FIGURE 11

Effects of Acrylamide (AA) and Bisphenol A (BPA) on platelets. (A, B) Intraplatelet reactive oxygen species (ROS) generation was measured using a DHE probe in a flow cytometer. The platelets were identified as the CD61⁺ population and were analyzed in terms of change in mean fluorescence intensity (ROS production). (C–D) representative kinetic aggregation obtained for each condition. (E) the graph summarizes the percentage of platelet aggregation; PMA was used as a platelet agonist. The results were presented from six independent volunteers (each donor executed as single triplicates) and expressed as mean ± SEM.