Tellimagrandin II Stimulates Inflammasomes by Causing an Accumulation of 3-Aminopropanal, Which Promotes Apoptosis of Endometriotic Cells While Inhibiting Invasion

Abstract

Background: Endometriosis is frequently treated with Paeoniae Radix. It contains Tellimagrandin II, which has the role of modulating immunity and anti-tumor. Therefore, we will explore the effects of Tellimagrandin II on the apoptosis and invasion/migration of ectopic endometrial cells (EECs).

Methods: Tellimagrandin II was used to treat EECs, and transcriptomics and bioinformatics techniques were used to identify its main pathways and targets of Tellimagrandin II. Western blotting was used to confirm the expression of essential targets. Flow cytometry was applied, the impact of tellimagrandin II on EECs apoptosis was identified. Transwell assays were conducted the effects of Tellimagrandin II on EECs invasion and migration. Finally, the binding of tellimagrandin II to key targets was confirmed using molecular docking techniques.

Results: Tellimagrandin II may inhibit pathways like beta-alanine metabolism and ECM-receptor interaction while activating JAK-STAT, NF-κB, and apoptotic pathways, according to transcriptomics and GSEA enrichment analysis. Tellimagrandin II can inhibit ALDH7A1 expression in EECs as well as increase SMOX expression, which may facilitate the accumulation of 3-Aminopropanal. This action becomes more pronounced as the dosage is increased. By upregulating the expression of NLRP3, TIMP-1, Caspase-3, BAX, and Caspase-1 in EECs while decreasing the expression of β-catenin and MMP2, tellimagrandin II can prevent EECs invasion and migration and encourage EECs apoptosis. Tellimagrandin II exhibited good docking with ALDH7A1 and SMOX, according to molecular docking.

Conclusion: Tellimagrandin II may stimulate inflammasomes by encouraging 3-aminopropanal accumulation within EECs. The increase in inflammasomes may promote EECs apoptosis and inhibit EECs invasion and migration. However, its in vivo inhibitory effects on endometriosis require further investigation.

Keywords: 3-aminopropanal; endometriosis apoptosis; inflammasome; invasion; tellimagrandin II.

Figure 1

CCK-8 assay. (Left) The effect of varying concentrations of tellimagrandin II on the survival rate of EECs; (Right) The effect of 50 μg/mL tellimagrandin II on the survival rate of EECs at different time points. ***:P <0.001; ****P <0.0001.

Figure 2

The figure shows the basic analysis of Tellimagrandin II acting on EECs: (a) depicts the expression details of all DEGs, with Orange dots indicating upregulation and blue dots indicating downregulation; (b) represents the KEGG enrichment analysis. The hue of the bubbles signifies the P-value, while their size represents the number of enriched DEGs. The redder the color, the more statistically significant the enrichment pathway.

Figure 3

Shows the GSEA enrichment analysis: (a) depicts the alterations in the pathway following Tellimagrandin II action on EECs, with red bars indicating activation and blue bars indicating inhibition; (b) depicts the 13 pathways where KEGG intersects with GSEA; (c) represents beta-alanine metabolism; (d) represents apoptosis; (e) represents ECM-receptor interaction; and (f) represents the IL-17 signaling pathway. On the c-f curves, an upward protrusion suggests activation, whereas a downward protrusion implies inhibition. ***:P <0.001; ****P <0.0001.

Figure 4

Western blot analysis of EECs treated with Tellimagrandin II: (a) protein immunoblot; (b) ALDH7A1; (c) SMOX; (d) NLRP3; (e) Caspase-1; (f) β-catenin; (g) cleaved Caspase-3; (h) BAX; (i) MMP2; (j) TIMP1. Each experiment was repeated three times. *:P <0.05; **:P <0.01; ***:P <0.001; ****P <0.0001.

Figure 5

Apoptosis flow cytometry analysis. (a–d) show flow cytometry results of EECs treated with Tellimagrandin II for 6 hours, while (e–h) show results after 18 hours of treatment. (a and e) represent the control group; (b and f) represent the low-dose Tellimagrandin II group; (c and g) represent the medium-dose group; (d and h) represent the high-dose group. The vertical axis represents PI (propidium iodide) staining, and the horizontal axis represents Annexin V staining.

Figure 6

Cell invasion and migration experiments. High, medium, and low doses of Tellimagrandin II can all inhibit the invasion and migration of EECs.

Figure 7

Molecular mechanism of Tellimagrandin II in the treatment of endometriosis. Tellimagrandin II (TII) may increase SMOX expression and decrease ALDH7 expression, leading to the accumulation of 3-aminopropanal in endometrial epithelial cells (EECs). The accumulated 3-aminopropanal can activate the inflammasome. Upon inflammasome activation, it may promote MMP2 expression via β-catenin and induce apoptosis by activating Caspase-1. However, whether TII inhibits invasion and promotes apoptosis in EECs through the inflammasome pathway requires further experimental verification.

Figure 8

Molecular docking diagram: (a) 3D representation of the docking between ALDH7A1 and Tellimagrandin II; (b) 2D representation of the docking between ALDH7A1 and Tellimagrandin II; (c) 3D representation of the docking between SMOX and Tellimagrandin II; (d) 2D representation of the docking between SMOX and Tellimagrandin II.