Ji-Sheng Wang1, Xiao Li2, Zi-Long Chen3, Jun-Long Feng4, Bing-Hao Bao5, Sheng Deng6, Heng-Heng Dai7, Fan-Chao Meng8, Bin Wang9, Hai-Song Li10. 1. First Clinical Medical College, Beijing University of Chinese Medicine, Beijing, 100029, China; Andrology Department, Dongzhimen Hospital, Beijing University of Chinese Medicine, Beijing, 100700, China. Electronic address: houdejisheng@sina.com. 2. The First Affiliated Hospital, Henan University of Chinese Medicine, Zhengzhou, 450000, China. Electronic address: lixiao396396@sina.com. 3. First Clinical Medical College, Beijing University of Chinese Medicine, Beijing, 100029, China; Andrology Department, Dongzhimen Hospital, Beijing University of Chinese Medicine, Beijing, 100700, China. Electronic address: chenzilong369@sina.com. 4. First Clinical Medical College, Beijing University of Chinese Medicine, Beijing, 100029, China; Andrology Department, Dongzhimen Hospital, Beijing University of Chinese Medicine, Beijing, 100700, China. Electronic address: 707995037@qq.com. 5. First Clinical Medical College, Beijing University of Chinese Medicine, Beijing, 100029, China; Andrology Department, Dongzhimen Hospital, Beijing University of Chinese Medicine, Beijing, 100700, China. Electronic address: baobinghao369@sina.com. 6. First Clinical Medical College, Beijing University of Chinese Medicine, Beijing, 100029, China; Andrology Department, Dongzhimen Hospital, Beijing University of Chinese Medicine, Beijing, 100700, China. Electronic address: 2319076562@qq.com. 7. First Clinical Medical College, Beijing University of Chinese Medicine, Beijing, 100029, China; Andrology Department, Dongzhimen Hospital, Beijing University of Chinese Medicine, Beijing, 100700, China. Electronic address: daihengheng369@sina.com. 8. First Clinical Medical College, Beijing University of Chinese Medicine, Beijing, 100029, China; Andrology Department, Dongzhimen Hospital, Beijing University of Chinese Medicine, Beijing, 100700, China. Electronic address: mengfanchao396@sina.com. 9. Andrology Department, Dongzhimen Hospital, Beijing University of Chinese Medicine, Beijing, 100700, China. Electronic address: dayiwangbln@sina.com. 10. Andrology Department, Dongzhimen Hospital, Beijing University of Chinese Medicine, Beijing, 100700, China. Electronic address: lihs396@sina.com.
Abstract
ETHNOPHARMACOLOGICAL RELEVANCE: Leeches (pinyin name Shui Zhi; Latin scientific name Hirudo; Hirudinea; Hirudinidae) and centipedes (pinyin name Wu Gong; Latin scientific name Scolopendridae; Chilopoda; Scolopendridae) are traditional Chinese medicines, and they belong to the family entomology. A combination of leech and centipede is used as an effective medicine to promote blood circulation and remove blood stasis in traditional Chinese medicine, and "leech-centipede" medicine has been used in many prescriptions to treat diabetic vascular disease, including diabetic erectile dysfunction (DIED). However, its specific mechanism remains unclear and requires in-depth study. AIM OF THE STUDY: This study aimed to investigate the mechanism of "leech-centipede" medicine to improve erectile dysfunction-associated diabetes by detecting PKC pathway-related molecules. MATERIALS AND METHODS: The active ingredients of "leech-centipede" medicine were identified using high performance liquid chromatography (HPLC). Fifty male SPF rats were injected with streptozotocin to induce the DM model. Eight weeks later, the DMED model was validated with apomorphine. The DIED rats were divided into five groups-T,P,DD,DZ, and DG-and were separately treated with tadalafil, pathway inhibitor LY333531 and low-, medium-, and high-dose "leech-centipede" medicine for 8 weeks. After treatment, the blood glucose level was measured, erectile function with apomorphine was assessed, the LOX-1, sE-selectin, sICAM-1, SOD, and MDA in serum was evaluated by enzyme-linked immunosorbent assay, and flow cytometry was performed. After the collection of penile tissue, the related protein and mRNA expression was assessed by Western blotting and PCR, and the tissue and ultrastructure were analysed by HE staining, immunohistochemistry and scanning electron microscopy. RESULTS: After treatment, the erectile function of rats was significantly improved in the T,P,DD,DZ, and DG groups compared with that in the model group. Thus, "leech-centipede" medicine can significantly reduce the levels of LOX-1, sE-selectin, sICAM-1, EMPs and CD62P to protect vascular endothelial function and anti-platelet activation, improving DIED rat erectile function. Additionally, "leech-centipede" medicine can increase SOD expression and decrease MDA expression, reducing the possibility of oxidative stress injury in DIED rats and improving the antioxidant capacity. Moreover, "leech-centipede" therapy can dramatically reduce the protein and mRNA expression of DAG, PKCβ, NF-κB, and ICAM-1, improve vascular endothelial injury in DIED rats and inhibit abnormal platelet activation. CONCLUSION: "leech-centipede" medicine can improve erectile dysfunction by inhibiting the expression of PKC pathway-related molecules in DIED rats and protects endothelial function and anti-platelet activation.
ETHNOPHARMACOLOGICAL RELEVANCE: Leeches (pinyin name Shui Zhi; Latin scientific name Hirudo; Hirudinea; Hirudinidae) and centipedes (pinyin name Wu Gong; Latin scientific name Scolopendridae; Chilopoda; Scolopendridae) are traditional Chinese medicines, and they belong to the family entomology. A combination of leech and centipede is used as an effective medicine to promote blood circulation and remove blood stasis in traditional Chinese medicine, and "leech-centipede" medicine has been used in many prescriptions to treat diabetic vascular disease, including diabetic erectile dysfunction (DIED). However, its specific mechanism remains unclear and requires in-depth study. AIM OF THE STUDY: This study aimed to investigate the mechanism of "leech-centipede" medicine to improve erectile dysfunction-associated diabetes by detecting PKC pathway-related molecules. MATERIALS AND METHODS: The active ingredients of "leech-centipede" medicine were identified using high performance liquid chromatography (HPLC). Fifty male SPF rats were injected with streptozotocin to induce the DM model. Eight weeks later, theDMED model was validated with apomorphine. The DIED rats were divided into five groups-T,P,DD,DZ, and DG-and were separately treated with tadalafil, pathway inhibitor LY333531 and low-, medium-, and high-dose "leech-centipede" medicine for 8 weeks. After treatment, the blood glucose level was measured, erectile function with apomorphine was assessed, theLOX-1, sE-selectin, sICAM-1, SOD, and MDA in serum was evaluated by enzyme-linked immunosorbent assay, and flow cytometry was performed. After the collection of penile tissue, the related protein and mRNA expression was assessed by Western blotting and PCR, and the tissue and ultrastructure were analysed by HE staining, immunohistochemistry and scanning electron microscopy. RESULTS: After treatment, the erectile function of rats was significantly improved in the T,P,DD,DZ, and DG groups compared with that in the model group. Thus, "leech-centipede" medicine can significantly reduce the levels of LOX-1, sE-selectin, sICAM-1, EMPs and CD62P to protect vascular endothelial function and anti-platelet activation, improving DIED rat erectile function. Additionally, "leech-centipede" medicine can increase SOD expression and decrease MDA expression, reducing the possibility of oxidative stress injury in DIED rats and improving the antioxidant capacity. Moreover, "leech-centipede" therapy can dramatically reduce the protein and mRNA expression of DAG, PKCβ, NF-κB, and ICAM-1, improve vascular endothelial injury in DIED rats and inhibit abnormal platelet activation. CONCLUSION:"leech-centipede" medicine can improve erectile dysfunction by inhibiting the expression of PKC pathway-related molecules in DIED rats and protects endothelial function and anti-platelet activation.