Investigating the Pro-Angiogenic Mechanism of Diammonium Glycyrrhizinate Injection in Zebrafish via Transcriptomic Analysis and RT-qPCR Validation

Abstract

Glycyrrhizic acid (GA) is the main metabolite of the traditional Chinese medicine Glycyrrhiza glabra. Many GA preparations in the pharmaceutical market have good efficacy in hepatoprotection and the treatment of liver diseases. However, the pro-angiogenic effects for GA-based agents have not been researched specifically. Angiogenesis is of vital importance for ischemic diseases, such as diabetic peripheral vascular disease, myocardial ischemia, and cerebral ischemia. The aim of this study was to investigate the pro-angiogenic activity of GA preparations commonly used in clinical practice and screen the representative angiogenesis promotor to further elucidate the mechanism of action. A zebrafish vascular injury model induced by PTK787 was used to assess the pro-angiogenic effect of GA preparations. The pro-angiogenic activity of diammonium glycyrrhizinate injection (DGI) was further systematically evaluated by detecting the growth of the intersegmental vessels (ISVs) and subintestinal vein vessels (SIVs) in zebrafish. The underlying mechanism of action for DGI was explored by transcriptomics analysis and RT-qPCR validation. The results showed that magnesium isoglycyrrhizinate injection (MII), DGI, compound glycyrrhizinate tablets (CGT), and diammonium glycyrrhizinate enteric-coated capsules (DGEC) demonstrated significantly pro-angiogenic effects. Among them, DGI exhibited the strongest activity. Transcriptomic analysis showed that the pro-angiogenic activity of DGI was closely associated with the mTOR signaling pathway. RT-qPCR results indicated that the expression levels of key genes of the mTOR/HIF-1 pathway were significantly upregulated in DGI-treated zebrafish relative to the PTK787 group. GA preparations have an obvious pro-angiogenic effect, in which DGI exerts particularly strong activity, appearing to be mediated mainly by activation of the mTOR/HIF-1 signaling pathway. This study broadens the clinical application of GA agents and provides new insights into the pharmacodynamics and potential mechanisms of DGI as a candidate agent for treating ischemic diseases.