本研究旨在探讨补阳还五汤通过血小板源性生长因子(PDGF)信号通路对小鼠后肢缺血后血流恢复及动脉生成的影响。将40只C57BL/6小鼠随机分为模型(清水,10mL·kg~(-1)·d~(-1)),贝前列素钠(阳性对照,18μg·kg~(-1)·d~(-1)),和低,medium-,和高剂量(10、20和40g·kg~(-1)·d~(-1),分别)补阳还五汤组(n=8)。采用股动脉结扎法建立后肢缺血模型。结扎后,每天通过管饲法给小鼠施用相应的药物,持续14天。对于激光多普勒灌注成像,将小鼠麻醉并在PeriscanPSI成像仪下测量。使用冷冻组织切片的免疫荧光染色测量缺血腓肠肌中毛细血管和动脉的密度。Westernblot检测PDGF亚基B(PDGFB)的表达,磷酸化丝裂原细胞外激酶(p-MEK),MEK,磷酸化细胞外信号调节激酶(p-ERK),和ERK.采用实时PCR测定PDGFB的mRNA水平。用补阳还五汤含药血清分别以10%和20%的剂量治疗缺氧血管平滑肌细胞(VSMC)。体外评估VSMC的增殖和迁移。结果表明,与模型组相比,贝前列素钠和补阳还五汤促进血流恢复,增加毛细血管和小动脉密度,并上调PDGFB的蛋白质水平,p-MEK,p-ERK,和PDGFB的mRNA水平,以中剂量补阳还五汤效果最显著。10%补阳还五汤含药血清可增强VSMCs的增殖和迁移。我们的发现表明补阳还五汤上调PDGFB转录并激活PDGF信号通路,从而促进缺血腓肠肌动脉生成和血流恢复。
This study aims to investigate the effect of Buyang Huanwu Decoction on blood flow recovery and
arteriogenesis after hindlimb ischemia in mice via the platelet-derived growth factor(PDGF) signaling pathway. Forty C57BL/6 mice were randomized into model(clean water, 10 mL·kg~(-1)·d~(-1)), beraprost sodium(positive control, 18 μg·kg~(-1)·d~(-1)), and low-, medium-, and high-dose(10, 20, and 40 g·kg~(-1)·d~(-1), respectively) Buyang Huanwu Decoction groups(n=8). The hindlimb ischemia model was established by femoral artery ligation. The mice were administrated with corresponding agents by gavage daily for 14 days after ligation. For laser Doppler perfusion imaging, the mice were anesthetized and measured under a Periscan PSI imager. The density of capillary and arterio-le in the ischemic gastrocnemius was measured using immunofluorescence staining of the frozen tissue sections. Western blot was employed to determine the expression of PDGF subunit B(PDGFB), phosphorylated mitogen extracellular kinase(p-MEK), MEK, phosphorylated extracellular signal-regulated kinase(p-ERK), and ERK. Real-time PCR was employed to determine the mRNA level of PDGFB. The Buyang Huanwu Decoction-containing serum was used to treat the vascular smooth muscle cells(VSMCs) in hypoxia at doses of 10% and 20%. The proliferation and migration of VSMCs was assessed in vitro. The results showed that compared with the model group, beraprost sodium and Buyang Huanwu Decoction enhanced the blood flow recovery, increased the capillary and arteriole density, and up-regulated the protein levels of PDGFB, p-MEK, p-ERK, and mRNA levels of PDGFB, with the medium-dose Buyang Huanwu Decoction demonstrating the most significant effect. The 10% Buyang Huanwu Decoction-containing serum enhanced the proliferation and migration of VSMCs. Our findings demonstrate that Buyang Huanwu Decoction up-regulates PDGFB transcription and activates PDGF signaling pathway to promote
arteriogenesis and blood flow recovery in ischemic gastrocnemius.