摘要
目的 探讨三突变型缺氧诱导因子-1α(HIF-1α)对老年小鼠缺血后肢的促血管生成作用。方法 24只老年小鼠结扎一侧后肢股动脉制备后肢缺血模型并随机等分为3组,分别经肌肉注射腺病毒包裹的β-半乳糖苷酶基因(Ad-LacZ),腺病毒包裹的Pro402、Pro564双位点突变型HIF-1α基因(Ad-DM),腺病毒包裹的Pro402、Pro564、Asn803三位点突变型HIF-1α基因(Ad-TM)。术前,术后0、7、14、21、28 d分别对小鼠双侧后肢骨骼肌行对比超声灌注成像评价缺血后肢骨骼肌血流情况(用缺血后肢血流量/对侧非缺血后肢血流量表示)。术后28 d利用运动评分评价缺血后肢的运动功能。术后28 d处死小鼠取缺血后肢骨骼肌分别行Bandeiraea Simplicifolia-1凝集素、α-平滑肌肌动蛋白免疫荧光染色标记微血管和小动脉,评价血管发生和小动脉生成。结果 对比超声灌注成像显示:术后0 d, 3组的血流均下降至15%(P=0.811)。随时间的推移,3组的血流均有所恢复,其中,Ad-TM组的血流恢复快于其他两组。术后28 d, Ad-TM组中缺血后肢的血流量达到对侧非缺血后肢血流量的(67.4±8.1)%,较Ad-LacZ组的(33.2±5.2)%和Ad-DM组的(48.4±5.3)%显著增加(P<0.001)。术后28 d, Ad-TM组的运动评分(3.4±0.7)分显著高于Ad-LacZ组的(1.8±0.6)分和Ad-DM组的(2.5±0.5)分,差异有统计学意义(P=0.001)。免疫荧光染色显示:Ad-TM组的微血管密度(3.1±0.6)显著高于Ad-LacZ组的(1.6±0.4)和Ad-DM组的(2.0±0.5),差异有统计学意义(P<0.001);Ad-TM组的小动脉密度(10.6±2.2)显著高于Ad-LacZ组的(5.7±1.0)和Ad-DM组的(6.7±1.5),差异有统计学意义(P<0.001)。结论 三突变型HIF-1α可通过血管发生和小动脉生成的方式有效促进老年小鼠缺血后肢的血管生成,改善缺血后肢血流灌注和运动功能。
Objective To investigate the neovascularizational effects of hypoxia-inducible factor-1α(HIF-1α) with triple-point mutant on ischemic hindlimbs in aged mice.Methods Twenty four aged mice were ligated the femoral artery of one hindlimb to prepare the hindlimb ischemia model and were randomly divided into three groups, which were injected with adenovirus encapsulated β-galactosidase gene(Ad-LacZ),adenovirus encapsulated Pro402,Pro564 double site mutant HIF-1α gene(Ad-DM),adenovirus encapsulated three site mutant HIF-1α gene of Pro402,Pro564 and Asn803(Ad-TM).The bilateral hindlimb skeletal muscles of mice were compared through serial contrast-enhanced ultrasonic perfusion imaging before and immediately after surgery, and postoperative day 7,14,21 and 28.Movement score was used to assess functional recovery of the limbs on postoperative day 28. Postoperative day 28,the ischemic hindlimb muscle specimens were harvested, then the density of the capillaries and arterioles were determined by immunofluorescence analysis using antibodies against Bandeiraea Simplicifolia-1 lectin or smooth muscle α-actin.Results Immediately after femoral artery ligation, blood flow(as a percentage of that in the non-ischemia hindlimb) was reduced to 15% in all groups(P=0.811). Although blood flow gradually recovered over time in all groups, recovery was persistently better in the Ad-TM group as compared with that of the other groups.On postoperative day 28,the blood flow ratio was significantly increase in the Ad-TM group(67.4±8.1)% as compared with that of the Ad-LacZ group(33.2±5.2)% and the Ad-DM group(48.4±5.3)%(P<0.001). The movement score for the Ad-TM group was(3.4±0.7),much superior to that of the Ad-LacZ group(1.8±0.6) and the Ad-DM group(2.5±0.5),and the difference was statistically significant(P=0.001).Immunofluorescence staining showed that the microvessel density of Ad-TM group(3.1±0.6) was significantly higher than that of Ad-LacZ group(1.6±0.4) and Ad-DM group(2.0±0.5),and the difference was statistically significant(P<0.001).The arteriolar density in Ad-TM group(10.6±2.2)was significantly higher than that in Ad-LacZ group(5.7±1.0) and Ad-TM group(6.7±1.5),and the difference was statistically significant(P<0.001).Conclusion Triple-point mutant HIF-1α can effectively promote the angiogenesis of the ischemic hindlimb of aged mice through angiogenesis and arteriogenesis, and improve the blood perfusion and motor function of the ischemic hindlimb.
作者
谢佳佳
廖文君
吴碧君
唐其东
李瑜辉
XIE Jiajia;LIAO Wenjun;WU Bijun;TANG Qidong;LI Yuhui(Department of Cardiology,Guangdong Second Provincial General Hospital,Guangzhou 510317,Guangdong,China;Department of Ultrasound,Guangdong Second Provincial General Hospital,Guangzhou 510317,Guangdong,China)
出处
《心血管病学进展》
CAS
2022年第9期841-845,共5页
Advances in Cardiovascular Diseases
基金
国家自然科学基金(81401426)。
关键词
下肢缺血
缺氧诱导因子-1Α
对比超声
血管生成
Lower limb ischemia
Hypoxia-inducible factor-1α
Contrast ultrasound
Angiogenesis
