Supporting data for “Network pharmacology prediction and biological validation of the molecular mechanisms underlying the cardioprotection of botanicals against myocardial injury”

Acute myocardial infarction (AMI) is a leading cause of human death worldwide. Provided that timely medical interventions are in place, contemporary therapies could effectively reduce the rates of morbidity and mortality. However, new therapies are pressingly needed to prevent heart failure and improve the life quality of AMI patients. Botanicals are proven to be effective for treating myocardial injury and other heart diseases. Thus, the objective of this study was to characterize the cardioprotective properties of the well-documented botanicals and elucidate the underlying molecular mechanisms.

Firstly, we investigated the herbal medicine formula Baoyuan Decoction (BYD) for cardioprotective compounds and potential molecular targets. We validated that BYD not only protected cardiomyocyte H9C2 cells against oxygen glucose deprivation but also effectively reduced infarct size and preserved cardiac functions in a mouse model of myocardial ischemia reperfusion injury (MIRI). UPLC-MS/MS analysis identified 122 compounds from ethanolic BYD extract while network pharmacology approach predicted 37 MI-related target proteins in the regulation of inflammation and apoptosis. Pathway enrichment analysis revealed that BYD most likely targeted the tumor necrosis factor-alpha (TNF-α)/ nuclear factor-κB (NF-κB) pathway. Adenosine, ginsenoside Rh2, isoliquiritigenin, and licochalcone A were selected to form the four-compound mixture AGILe for targeting TNF-α/NF-κB pathway. Indeed, the formula AGILe effectively inhibited the TNF-α/NF-κB pathway and manifested the anti-inflammatory functions of BYD in the context of myocardial infarction.

Secondly, we previously discovered that C-glycosylated isoflavone puerarin preserved cardiac function against ischemia-reperfusion injury by upregulating SUMO2, a small ubiquitin-like modifier. We hypothesized that SUMO2 might activate endogenous cardioprotective mechanisms during myocardial infarction. We generated cardiac-specific SUMO2 knockout mice and evaluated their performance in myocardial infarction. As results, SUMO2 KO mice experienced severe myocardial injury and cardiac dysfunction, while SUMO2 overexpression markedly improved the performance of SUMO2 KO mice during ischemia reperfusion challenge. In vitro SUMO2 silencing increased ROS generation, disrupted the Bax/Bcl-2 signaling pathway, and exacerbated intrinsic cell apoptosis during oxygen glucose deprivation. These results confirmed the importance of SUMO2 in cardiac homeostasis and resistance. Ultimately, botanic drug puerarin may be a potential therapyvagainst myocardial injury for targeting SUMO2.

Thirdly, we prepared recombinant CRPPR-tagged human serum albumin (HSA) and puerarin-loaded HSA-CRPPR nanoparticles (HSA-CRPPR-PUE) for heart-targeting delivery of puerarin. Based on pharmacological evaluation in mouse model of MIRI, HSA-CRPPR-PUE showed better heart-targeting efficiency compared to free puerarin and HSA-PUE nanoparticles. Consistently, HSA-CRPPR-PUE more effectively protected the hearts against MIRI.

In conclusion, this thesis employed multiple state-of-art technologies to elucidate the molecular mechanisms underlying the cardioprotective effects of botanicals drugs. The key findings include: 1) A four-compound remedy AGILe was developed to manifest the potential of BYD for treating myocardial injury via regulating the TNFα/NF-Κb pathway; 2) Expression level of SUMO2 positively correlated with the health and functions of the hearts during myocardial infarction; 4) HSA-CRPPR-PUE nanoparticles supported heart-targeting delivery of puerarin for treating myocardial injury. Overall, these findings paved the way to develop botanical drugs for the therapy of myocardial infarction.