Vasorelaxant Effects of the Crassirhizomae Rhizoma Water Extract via PI3K/Akt and NO/sGC/cGMP Pathways

다국어 초록 (Multilingual Abstract)

Vasorelaxant Effects of the Crassirhizomae Rhizoma Water Extract via PI3K/Akt and NO/sGC/cGMP Pathways
By Jihoon Kim Master of Korean Medicine Graduate School of Kyung Hee University Advised by Prof. Kyungjin Lee

Hypertension is a significant risk factor for cardiovascular disease. Hypertension can cause fatal complications, such as stroke, heart failure, and coronary artery disease; therefore, active management and treatment are required. In Korean medicine, various herbs are used in clinical practice to address hypertension-related issues, and accumulating evidence supports their efficacy and safety. Therefore, the possibility of treating hypertension using Korean medicines is worth exploring. Dryopteris crassirhizoma Nakai rhizome (DC) has long been used in traditional Korean medicine to treat parasitic infestations, colds, and hemorrhages. DC is also known for its antioxidant properties that protect vascular endothelial cells from oxidative stress, thus improving vascular health and supporting vascular relaxation. Vascular relaxation reduces blood flow resistance, facilitates smooth blood flow, and effectively lowers blood pressure. Therefore, the antioxidant activity of DC is expected to promote vascular relaxation and cause a decrease in blood pressure. However, studies on the vasorelaxant effects of DC are limited. This study aimed to investigate the vasorelaxant effects of DC. Before conducting vascular relaxation experiments with DC, we used network pharmacology to predict how DC induces vascular relaxation. The analysis suggested that DC could activate eNOS through the PI3K/Akt pathway leading to NO emission and vascular relaxation. Based on this, we focused our experiments on the mechanisms involving the PI3K/Akt pathway and eNOS. Thoracic aortas dissected from Sprague-Dawley rats were used to investigate vasorelaxant effects. Water extract of DC (DCW) (3, 10, 30, and 100 μg/mL) exhibited an endothelium- dependent vasorelaxant effect, which was found to involve the activation of the PI3K/Akt pathway and NO/sGC/cGMP pathway after phosphorylation of eNOS. Additionally, the DCW (100 μg/mL) demonstrated an inhibitory effect on aortic ring contractions induced by angiotensin II. The vascular relaxation effects of DCW were not related to vascular smooth muscle cell- associated mechanisms, such as K+ channels and Ca2+ channels. Considering its vasorelaxant effects, DCW has the potential for the prevention and treatment of hypertension. However, further studies are required to identify specific components responsible for its vasorelaxant effects and to determine whether DC exhibits vascular relaxation effects in vivo.

Keywords
Dryopteris crassirhizoma Nakai, Hypertension, Vasorelaxation, Network pharmacology, PI3K/Akt pathway

목차 (Table of Contents)

• Abstract v
• 1. Introduction 1
• 2. Materials and Methods 3
• 2.1. Network pharmacology analysis 3
• 2.2. Materials and chemicals 5
• 2.3. Sample preparation 5
• 2.4. Animals 5
• 2.5. Evaluation of vascular tension 6
• 2.6. Data analysis 9
• 3. Results 10
• 3.1. Effective compounds of DC and potential target proteins 10
• 3.2. KEGG pathway enrichment analysis and GO functional enrichment analysis of targets 13
• 3.3. Protein-protein interaction analysis of DC 17
• 3.4. Molecular docking of compounds and targets 21
• 3.5. Vasorelaxant effect of DCW on constriction induced by PE, KCl 24
• 3.6. Vasorelaxant effect of DCW on the rat aortic rings with intact or denuded endothelium 27
• 3.7. Effect of PI3K inhibitor pretreatment on vasorelaxation of DCW 30
• 3.8. Effect of nitric oxide synthase inhibitor, sGC inhibitor, cGMP inhibitor, COX inhibitor
• pretreatment on vasorelaxation of DCW 33
• 3.9. Effect of atropine pretreatment on vasorelaxation of DCW 36
• 3.10. Inhibitory effect of DCW on rat aortic rings constricted by extracellular Ca2+ 39
• 3.11. Vasorelaxant effect of DCW on aortic rings pretreated with K+ channel blockers .. 42
• 3.12. Inhibitory effect of DCW on constriction induced by Ang II 45
• 4. Discussion 48
• 5. Conclusions 54
• 6. References 55
• Abstract in Korean 60