血管外支架预防猪大隐静脉移植血管再狭窄的实验研究

目的以猪大隐静脉-颈总动脉旁路移植动物模型为基础,观察涤纶血管外支架支持预防静脉移植血管内、中膜增生的作用。方法将10只25~30kg普通长白猪行双侧大隐静脉-颈总动脉旁路移植术(端侧吻合),一侧静脉移植血管放置涤纶外支架(实验组),另一侧作为对照(对照组)。术后35d取出移植血管进行组织学和免疫组织化学检测。结果对照组静脉移植血管内膜增生较实验组明显增加(0.4872±0.0706mm vs.0.2259±0.0553mm,P<0.01);对照组中膜增生亦较实验组增加(0.6246±0.0859mm vs.0.4201±0.0615mm,P<0.01);对照组内膜面积是实验组的2倍,中膜面积是实验组的近1.5倍。实验组内膜及中膜内侧区域增殖细胞核抗原(PCNA)、血小板源性生长因子(PDGF)阳性细胞显著减少,PCNA从7.98%±4.06%减少至3.35%±0.95%(P<0.01),PDGF从9.47%±5.35%减少至2.67%±0.97%(P<0.01)。结论非限制性涤纶血管外支架可以显著抑制大隐静脉移植血管新内膜及中膜增生,可能预防大隐静脉移植血管的再狭窄。     

转染单纯疱疹病毒胸苷激酶基因抑制移植静脉内膜增生

目的探讨腺病毒载体转染单纯疱疹病毒胸苷激酶（herpes simplex virus thymidine kinase，HSVtk）基因对移植静脉内膜增生的作用。方法用Wistar大鼠建立自体静脉移植模型。以载有HsVtk（4&#215;10^9 plaque forming units）基因的腺病毒孵育颈静脉，将静脉倒置移植于肾下腹主动脉。从移植术后第3d开始腹腔注射丙氧鸟苷[ganciclovir，GCV，60mg／（kg&#183;d），2次／d]共21d，取出移植静脉标本后用PCR法检测HSVtk基因的表达，用原位杂交法检测基因的转录；进行弹力纤维染色及增殖细胞核抗原（PCNA）免疫组化染色，观察内膜增生及平滑肌细胞增殖；用TUNEL法观察移植静脉平滑肌细胞凋亡情况。结果移植静脉内膜进行性增厚，新生内膜中有大量增殖状态的平滑肌细胞。移植后第3d，转染HSVtk基因的静脉经PCR扩增出590bp阳性条带，HSVtk基因开始表达mRNA。HSVtk／GCV对静脉内膜增生有明显抑制作用，而单独予以HSVtk对内膜增生无影响[（17．2&#177;3．2）μm vs（31.1&#177;2．5）μm，P〈0．05]。HSVtk／GCV使静脉中平滑肌细胞增殖指数下降为（9．1&#177;2．3）％，凋亡细胞指数升高到（28．7&#177;3．6）％，分别于空白对照组[（38．7&#177;5．6）％，（18．5&#177;2．6）％]相比差异均有统计学意义（P〈0．05）。结论HSVtk／GCV系统通过抑制平滑肌细胞增殖及促进细胞凋亡抑制移植静脉内膜增生。     

特异亲合转录因子E2F的寡脱氧核苷酸对移植静脉内膜增生的影响

目的　观察E2F亲合性寡脱氧核苷酸对大鼠自体移植静脉内膜增生的影响。方法以Wistar大鼠建立自体静脉移植模型 ,术中将含有E2F亲合性双链寡脱氧核苷酸的医用生物胶涂沫于移植静脉的外膜。术后 14d取移植静脉 ,观察细胞周期基因c myc ,核增殖抗原基因 (PCNA)的表达指数和内膜增生程度。以空白胶和错配双链寡脱氧核苷酸作为对照。结果　对照组移植静脉中有大量平滑肌细胞表达c myc和PCNA ,内膜明显增生。错配双链寡脱氧核苷酸对c myc ,PC NA的表达和内膜增生无抑制作用。当静脉外膜施加 10 0 μgE2F亲合性寡脱氧核苷酸时 ,PCNA和c myc的表达率分别是 ( 2 0 .41± 4.15 ) %、( 10 .83± 2 .87) % ,比对照静脉分别下降 5 0 %、34% ,内膜厚度为 ( 18.2± 2 .8) μm ,比对照组减轻 44% ( P <0 .0 5 )。 2 0 0 μg的E2F亲合性双链寡脱氧核苷酸对PCNA、c myc和内膜增生的抑制更为显著。 结论　E2F亲合性寡核苷酸可能通过抑制c myc、PCNA的表达抑制内膜增生 ,存在剂量效应关系。     

腹主动脉人工血管置换术后放疗对人工血管内膜的影响

目的:观察腹主动脉人工血管置换术后35Gy体外分割放疗对人工血管通畅率、内膜增生及血管内皮细胞(VEC)覆盖的影响。方法:对20只犬行腹主动脉膨体聚四氟乙烯（ePTFE）人工血管置换术。术后动物分成对照组和放疗组，每组10只。两组均于术后4，8周采集标本，行HE染色，增殖细胞核抗原（PCNA）及分化抗原簇34（CD34）免疫组化检测。结果:两组术后各有1例人工血管内血栓形成，通畅率均为90.0％。术后4周，放疗组和对照组人工血管内膜形成完整，内膜厚度和PCNA表达差异无统计学意义（均P>0.05），VEC覆盖均不完全；术后8周，放疗组内膜厚度较对照组明显变薄（P<0.05），对照组人工血管近端、中间、远端3处内膜厚度分别为(72.30±9.15)μm，(40.46±7.75)μm和(98.06±6.90)μm，放疗组人工血管近端、中间、远端3处内膜厚度分别为(37.67±6.77)μm， (21.16±4.98)μm和(56.64±5.13)μm，PCNA表达减少（P<0.05）；VEC均覆盖完整。结论:腹主动脉人工血管术置换后35Gy体外分割放疗不影响移植人工血管通畅率，对内膜VEC的覆盖亦无明显影响，但可抑制人工血管内膜增生和PCNA的表达。     

重组arresten蛋白对自体移植静脉内膜增生的抑制作用

目的 观察原核表达人arresten重组蛋白对自体移植静脉内膜增生的抑制作用.方法 用pRSET原核表达系统表达并纯化人arresten重组蛋白.将Wistar大鼠颈外静脉移植于腹主动脉,建立大鼠自体静脉移植模型,实验分3组:假手术组、移植对照组和移植实验组.自术后第3天起,皮下注射给予arresten重组蛋白(每日4 mg/kg体重)处理.4周后取移植静脉组织标本,进行病理组织学观察与免疫组织化学染色分析.结果 移植组移植静脉均呈现典型的内膜增生、肥厚,导致血管管腔狭窄;新生内膜主要有过度增殖的α-SMA染色阳性平滑肌细胞组成.移植实验组移植静脉内膜增生受到明显抑制,新生内膜面积(0.12±0.07)mm2及新生内膜/中膜面积比(0.373±0.085)均显著低于对照组[(0.38±0.11)mm2,1.621±0.086,P<0.01];并且实验组移植静脉新生内膜细胞PCNA标记指数显著低于对照组[(15.62±3.97)%比(56.36±3.49)%,P<0.01].结论 重组arresten蛋白通过抑制新生内膜平滑肌细胞的增殖能有效抑制自体移植静脉内膜增生的发生发展,在防治血管重建术后再狭窄方面显示出良好的应用前景.     

实验性自体静脉移植平滑肌细胞增殖周期动力学研究

为观察移植静脉平滑肌细胞增植周期变化，取１２０只大鼠建立自体颈静脉移植于腹主动脉动物模型，分别于移植早期（２ｈ，６ｈ，２４ｈ），移植中期（１ｗ，２ｗ，４ｗ），以透射电镜观察其超微结构，流式细胞仪检测整体移植血管增殖活性，利用ＰＣＮＡ免疫组化观察平滑肌细胞增殖活性及增殖平滑肌细胞在血管中分布。结果表明：（１）自体静脉移植早期虽然组织学未见平滑肌细胞数量明显增加，但平滑肌细胞增殖业已启动；移植中期，平滑肌细胞大量增殖移行；（２）在增殖细胞的分布上，移植早期，局限于血管中膜；移植中期，内膜及中膜均表现出较高增殖率。因此，移植静脉平滑肌增殖、移行的控制应着力于移植术后２周内。     

脂质体与腺病毒转染EDRz抑制自体移植静脉内膜增生的比较研究

目的探讨以脂质体和腺病毒为载体,早期生长反应基因-1 DNA酶(Egr-1 DNA enzyme,EDRz)局部外用对自体移植静脉血管平滑肌(VSMC)增殖和内膜增生的抑制作用。方法大鼠建立自体静脉移植模型后随机分为脂质体组、腺病毒组和对照组,两实验组分别用脂质体-EDRz和腺病毒-EDRz涂抹移植静脉行在体转染。于术后1,2,6,24 h及3,7,14,28 d取材,荧光显微镜检测移植静脉的转染情况;采用原位杂交方法检测Egr-1 mRNA的表达;用免疫组织化学方法检测Egr-1蛋白表达,同时进行组织形态学观察。结果术后1 h,EDRz主要位于移植静脉的外膜、中膜和部分内皮细胞。脂质体组荧光灰度值为70.3±13.5,腺病毒组荧光灰度值为60.5±11.2。术后2～24 h,EDRz主要位于移植静脉的中膜;移植7 d,EDRz主要位于移植静脉的内膜。移植早期Egr-1蛋白表达主要位于中膜的血管平滑肌细胞(VSMC)和部分单核细胞、内皮细胞;移植后期未检测到Egr-1 DNA酶的表达。移植后期在中膜和新生内膜的VSMC均未检测到Egr-1蛋白的表达。移植2 h阳性表达率,脂质体组为(15.3±4.2)%,腺病...     

大鼠颈静脉分支-颈动脉间置模型新生内膜细胞来源研究

目的探讨大鼠颈静脉分支-颈动脉间置模型新生内膜细胞来源。方法建立SD大鼠颈静脉分支-颈动脉间置模型,分别于术后1、3、7、14和28d取静脉移植血管,定量分析新生内膜厚度,并进行α-SM—actin和CD34免疫组织化学分析。结果新生内膜增生在28d时最明显,血管吻合部位狭窄最严重,增生厚度近端（65．2±4．6）μm,远端（64．7±5．3）μm,中段（63．5±5．6）μm。新生内膜细胞主要来源于内皮细胞、相邻动脉血管平滑肌细胞或循环祖细胞,以新生内膜腔面为著。结论静脉移植血管新生内膜细胞主要来源于静脉移植血管本身内皮细胞、相邻动脉血管平滑肌细胞或循环祖细胞,提示可于血管吻合完成后进行局部干预或术后尽早全身用药,以防止移植血管狭窄。     

血管外雷帕霉素缓释涂层抑制移植静脉再狭窄

目的观察血管周围局部应用雷帕霉素对兔自体移植静脉段内膜过度增殖的抑制作用,并初步探讨其作用机制。方法大耳白兔30只,采用自身对照设计建立模型。任取一侧静脉旁路为干预组,外涂含0.3 mg雷帕霉素的Pluronic凝胶;另一侧为对照组,外涂空凝胶。4周后测旁路静脉内膜厚度、细胞的增殖指数及PCNA、P27~(kipl)免疫组化阳性细胞数。并于术后1、2、4、8周检测血管壁中雷帕霉素的含量。结果干预组旁路静脉内膜厚度(63.72±14.00)μm与对照组(77.76±14.90)μm相比,增生明显减少(P<0.05)。增殖指数对照组(29.30±7.15)明显高于干预组(20.10±9.48)(P<0.05)。在干预组和对照组均有阳性PCNA免疫组化染色表达细胞,正常静脉未见阳性表达,对照组较干预组细胞的阳性表达更强(P<0.05)。P27~(kipl)免疫组化染色干预组见阳性表达细胞,对照组表达阴性(P<0.05)。干预组血管壁术后1～8周均可测到雷帕霉素,含量呈逐步递减趋势。结论血管周围局部应用雷帕霉素具有抑制兔移植静脉内膜过度增殖的作用;这种作用和P27~(kipl)的上调表达有关。     

肝素对自体移植静脉内膜增殖的影响

目的 探讨肝素对移植静脉内膜增殖的影响。方法 建立大鼠自体静脉移植模型;将颈静脉内侧支移植于髂总动脉,应用肝素,每日２次,皮下注射５天。移植后２８天,应用病理形态学及ＰＣＮＡ免疫组织化学,通过计算机图像定量分析,观察其对移植静脉内膜平滑肌细胞增殖的影响。结果 肝素治疗组和空白对照组移植静脉段内膜增厚程度,ＰＣＮＡ阳性细胞数无差异。结论 肝素对移植静脉内膜增殖无作用,对受损的动脉内膜平滑肌细胞增殖有     

Vein to artery grafts: a morphological and histochemical study of the histogenesis of intimal hyperplasia.

Failure of vein to artery grafts has been associated with intimal thickening (hyperplasia) and atherosclerosis. Current theories of intimal development, derived from arterial studies, show that smooth muscle cells migrate from the media to the intima after endothelial damage, where they proliferate and produce intimal hyperplasia. However, little is known of the histogenesis of these lesions in vein grafts. Experimental ilio-lumbar vein to iliac artery autografts were placed in 52 rats and analysed by light microscopy and histochemistry from 2 to 140 days after surgery. On day 2 the grafts and adjacent artery were severely damaged. Regeneration of damaged arterial tissue occurred by day 5, and thickening was already evident in the arterial intima. The intimal cells had histochemical characteristics of smooth muscle. By day 15, this hyperplastic intima was continuous across the anastomosis from the artery into the graft. After day 28 a wedge of densely packed cells was present in the vein graft intima for approximately 2 mm into the graft. By day 140, all the grafts were fully re-endothelialized. Intimal hyperplasia was present in all grafts and varied in thickness from 3 to 20 cells. Histochemical staining of these cells showed them to be of smooth muscle origin.     

VEIN TO ARTERY GRAFTS: A MORPHOLOGICAL AND HISTOCHEMICAL STUDY OF THE HISTOGENESIS OF INTIMAL HYPERPLASIA

Failure of vein to artery grafts has been associated with intimal thickening (hyperplasia) and atherosclerosis. Current theories of intimal development, derived from arterial studies, show that smooth muscle cells migrate from the media to the intima after endothelial damage, where they proliferate and produce intimal hyperplasia. However, little is known of the histogenesis of these lesions in vein grafts. Experimental ilio-lumbar vein to iliac artery autografts were placed in 52 rats and analysed by light microscopy and histochemistry from 2 to 140 days after surgery. On day 2 the grafts and adjacent artery were severely damaged. Regeneration of damaged arterial tissue occurred by day 5, and thickening was already evident in the arterial intima. The intimal cells had histochemical characteristics of smooth muscle. By day 15, this hyperplastic intima was continuous across the anastomosis from the artery into the graft. After day 28 a wedge of densely packed cells was present in the vein graft intima for approximately 2 mm into the graft. By day 140, all the grafts were fully re-endothelialized. Intimal hyperplasia was present in all grafts and varied in thickness from 3 to 20 cells. Histochemical staining of these cells showed them to be of smooth muscle origin.     

Modulation of phosphatidylinositol 3-kinase signaling reduces intimal hyperplasia in aortocoronary saphenous vein grafts

OBJECTIVES: Fifty percent of human aortocoronary saphenous vein grafts are occluded after 10 years. Intimal hyperplasia is an initial step in graft occlusion and consists of vascular smooth muscle cell proliferation. Phosphatidylinositol 3-kinase and its downstream regulator, the inositol 3-phosphatase PTEN (phosphatase and tensin homolog deleted on chromosome 10), are important regulators of vascular smooth muscle cell proliferation, migration, and cell death. This study tests whether overexpression of PTEN in aortocoronary saphenous vein grafts can reduce intimal hyperplasia. METHODS: Adult dogs underwent aortocoronary bypass grafting to the left anterior descending artery by using the autologous saphenous vein. Saphenous vein grafts were treated with phosphate-buffered saline (n = 9), empty adenovirus (n = 8), or adenovirus encoding for PTEN (n = 8). Arteriography at 30 and 90 days assessed saphenous vein graft patency. A subset received saphenous vein grafts treated with a marker transgene (beta-galactosidase, n = 3), empty adenovirus (n = 4), or adenovirus encoding for PTEN (n = 4) and were killed on postoperative day 3 to confirm expression. Vascular smooth muscle cells were isolated from canine saphenous vein infected with adenovirus encoding for PTEN, and immunoblotting and proliferation assays were performed. RESULTS: Saphenous vein graft transgene expression was confirmed by means of immunohistochemistry, immunoblotting, and polymerase chain reaction. Arteriograms revealed all saphenous vein grafts to be patent. Saphenous vein grafts treated with adenovirus encoding for PTEN demonstrated reduced intimal area compared with those treated with empty adenovirus and phosphate-buffered saline (1.39 +/- 0.11 vs 2.35 +/- 0.3 and 2.57 +/- 0.4 mm 2 , P < .05), and the intima/media ratio was lower in saphenous vein grafts treated with adenovirus encoding for PTEN (0.50 +/- 0.05 vs 1.43 +/- 0.18 and 1.11 +/- 0.14, P < .005). PTEN overexpression in vascular smooth muscle cells inhibited platelet-derived growth factor-induced phosphorylation of Akt, a downstream effector of phosphatidylinositol 3-kinase. PTEN-treated vascular smooth muscle cells demonstrated decreased basal, platelet-derived growth factor-stimulated, and serum-stimulated proliferation. CONCLUSION: This study demonstrates that PTEN overexpression in aortocoronary saphenous vein grafts reduces intimal hyperplasia. The mechanism of this antiproliferative effect in vascular smooth muscle cells is likely due to inhibition of phosphatidylinositol 3-kinase signaling through Akt, with resultant decreases in vascular smooth muscle cell growth and survival. Therefore modulation of the phosphatidylinositol 3-kinase pathway through PTEN overexpression might represent a novel therapy to prevent saphenous vein graft intimal hyperplasia after coronary artery bypass grafting.     

Neointimal hyperplasia rapidly reaches steady state in a novel murine vein graft model

OBJECTIVE: Neointimal hyperplasia remains a principal cause of vein graft failure. Genetic contributions to vein graft neointimal hyperplasia could be well studied in the mouse; however, surgical approaches to vein bypass surgery in the mouse have yet to replicate approaches commonly employed in human patients. Consequently, the goal of this study was to develop a murine interposition vein graft model that reproduces characteristics of human vein graft disease. METHOD: Using C57BL/6J mice, we excised inferior venae cavae (IVCs) from donor mice and grafted them, with end-to-side anastomosis, into the carotid circulation of recipients. IVC grafts were harvested from 3 to 56 days postoperatively, and analyzed for the development of neointima and media. RESULTS: Thickening of both the vein graft neointima and media progressed rapidly between postoperative weeks 1 and 4, and reached steady state levels by approximately week four, with a graft-wall thickness of 91 +/- 4 microm (14 cell layers), a lumen area of 0.56 mm(2), an average neointima-media ratio of 0.4 to 0.6, and a predominance of alpha-smooth muscle actin-staining cells. Comprising predominately smooth muscle actin-expressing cells, the neointima was 50% thicker in the proximal than in the distal third of the grafts (P <.001), but proximal and distal vein graft anastomoses were widely patent. CONCLUSIONS: In syngeneic murine carotid interposition IVC grafts implanted with end-to-side anastomoses, moderate, nonocclusive neointimal hyperplasia reaches steady state after the fourth postoperative week. This neointimal hyperplasia, like that of human grafts, predominates near vein graft anastomoses. This vein graft model should facilitate genetic analyses of the pathogenesis of neointimal hyperplasia.     

三七总皂苷对移植静脉再狭窄的影响

目的 观察三七总皂苷(PNS)对大鼠移植静脉再狭窄的影响.方法 建立SD大鼠颈外静脉颈总动脉旁路移植模型,随机分为假手术组、模型组和PNS组,造模后第2天开始灌胃给药,PNS组给予PNS(100 mg/kg),假手术组、模型组灌胃等量蒸馏水(10 ml/kg).给药14 d后,取静脉血管桥固定,观察血管内膜增生,免疫组织化学测定增殖细胞核抗原(PCNA)表达及TUNEL法检测平滑肌细胞凋亡.结果 上述3组的内膜厚度分别为(4.05±0.85)、(26.30±1.15)、(10.80±0.98)μm;内膜/中膜分别为(0.22±0.09)、(0.76±0.27)、(0.45±0.23),PNS组新内膜厚度明显低于模型组(P＜0.05);模型组的PCNA表达量(A值)为(23.6±4.3),明显高于PNS组(11.5±3.6,P＜0.05);模型组的凋亡细胞百分数为(4.3±1.9),明显低于PNS组(20.3±3.5,P＜0.05).结论 PNS可以抑制静脉移植血管桥新生内膜的增生,对预防移植血管再狭窄有一定作用.

Abstract：

Objective To observate the effects of panax notoginseng saponin (PNS) on restenosis of rat vein graft.Methods Jugular vein-to-artery bypass grafting was performed on 30 Sprague-Dawley rats.The rats were divided into three groups: sham group, model group and PNS group.Drugs were administered at the second day after the operation for 14 days.The grafts were harvested for histochemical staining to observe the hyperplasia of neointima, and proliferating cell nuclear antigen (PCNA) expression,and apoptosis of smooth muscle cells were detected by TUNEL.Results In sham group, model group and PNS group, the endometrial thickness was (4.05 ±0.85), (26.30 ± 1.15), ( 10.80 ±0.98) μm; the intima/media was (0.22 0.09), (0.76 0.27), (0.45 0.23), respectively.The neointimal thickness in PNS group was significantly less than in model group ( P ＜ 0.05 ).The PCNA expression in model group (A value) was ( 23.6 ± 4.3 ), significantly higher than in PNS group ( 11.5 ± 3.6 ) ( P ＜ 0.05 ).The percentage of apoptotic cells in model group was (4.3 ± 1.9), significantly lower than in PNS group (20.3 ± 3.5 ) (P ＜ 0.05 ).Conclusion PNS can inhibit the neointimal hyperplais of vein graft, which can prevente restenosis after bypass grafts.     

Biologic degeneration of vein grafts after thrombotic occlusion: thrombectomy within 3 days results in better indices of viability

OBJECTIVES: To clarify the mechanism for poor patency of vein grafts after thrombectomy and the time limit for successful salvage operation, we investigated the time course of biologic degenerative changes in thrombosed vein grafts.Materials and methods The right femoral artery was replaced with a femoral vein graft in 25 mongrel dogs. After 3 months, grafts were explanted in 5 dogs (control grafts), and the remaining 20 dogs underwent femoral artery ligation to create a thrombosed graft. Of the 20 grafts, 5 were explanted at 3 days after ligation (group I-3) and 5 were explanted at 5 days after ligation (group I-5). Of the remaining 10 grafts, 5 underwent thrombectomy at 3 days after ligation (group II-3) and 5 underwent thrombectomy at 5 days after ligation, and were reimplanted into the left femoral artery, then explanted 28 days after reimplantation. The grafts were assessed with immunohistochemistry and prostaglandin (PG) I(2) assay (6-keto-PGI(1alpha)). RESULTS: Of the 25 grafts, occlusion recurred in 3 in group II-5 within 28 days after reimplantation. There were significant differences between group I-5 and group I-3 or control grafts for percentage of areas positive for alpha-actin, total number of cells per field, and proliferating cell nuclear antigen (PCNA)-positive cells in layer of thickened intima and atrophied media (I/M), and for total cell and PCNA- positive cell numbers per field in the adventitia. Mean 6-ketoPGF(1alpha) was 40 +/- 14.1 pg/mg/min in control dogs, 84 +/- 18.9 pg/mg/min in group I-3, and 15.4 +/- 7.7 pg/mg/min in group I-5, demonstrating a significant reduction in group I-5 (P =.009). CONCLUSION: Graft wall cell viability and PGI(2) production in thrombosed vein grafts are well preserved for up to 3 days. Therefore graft salvage operations no later than 3 days after thrombotic occlusion may provide acceptable long-term patency of salvaged grafts.     

Hydrophilic statin suppresses vein graft intimal hyperplasia via endothelial cell-tropic Rho-kinase inhibition

BACKGROUND: Recent studies suggest that statins can protect the vasculature in a manner that is independent of their lipid-lowering activity through inhibition of the small guanosine triphosphate-binding protein, Rho, and Rho-associated kinase. Little information is available on the inhibitory effect of statins on vein graft intimal hyperplasia, the main cause of late graft failure after bypass grafting. We therefore examined the effects of a hydrophilic statin on vein graft intimal hyperplasia in vivo and Rho-kinase activity in vitro. METHODS: In the first experiment, rabbits were randomized to a control group (n = 7) that was fed regular rabbit chow or to a pravastatin group (n = 7) that was fed regular rabbit chow supplemented with 10 mg/kg pravastatin sodium. The branches of the jugular vein were ligated and an approximately 3-cm segment of the jugular vein was taken for an autologous reversed-vein graft. The carotid artery was cut and replaced with the harvested autologous jugular vein. At 2 and 4 weeks after the operation, vein grafts in both groups were harvested, and intimal hyperplasia of the vein grafts was assessed. In the second experiment, human umbilical vein endothelial cells and vascular smooth muscle cells were cultured and then treated with 1 micromol/L and 30 micromol/L pravastatin for 24 hours and harvested. Immunoblotting was performed on the resulting precipitates. Quantitative evaluation of phosphorylated myosin binding subunit and endothelial nitric oxide synthase was performed by densitometric analysis. RESULTS: We demonstrated that oral administration of the hydrophilic statin pravastatin to normocholesterolemic rabbits inhibited intimal hyperplasia of carotid interposition-reversed jugular vein grafts 4 weeks after implantation (pravastatin group, 39.5 +/- 3.5 microm vs control group, 64.0 +/- 7.1 microm; n = 7; P < .05) and suppressed cell proliferation and apoptosis in the neointima 2 weeks after implantation. In addition, we found that pravastatin inhibited Rho-kinase activity and accelerated endothelial nitric oxide synthase expression in human umbilical vein endothelial cells but did not inhibit Rho-kinase activity in vascular smooth muscle cells. CONCLUSIONS: These novel findings clearly demonstrate that a hydrophilic statin can suppress intimal hyperplasia of the vein graft in vivo and also show endothelial cell-tropic inhibition of Rho-kinase in vitro. Furthermore, these results strongly support the clinical use of hydrophilic statins to prevent intimal hyperplasia of the vein graft after bypass grafting. CLINICAL RELEVANCE: Late graft failure caused by neointimal hyperplasia limits the efficacy of vein grafting. Various treatments were examined to reduce neointimal hyperplasia, but a standard clinical treatment has not yet been established. We report here the inhibitory effect of pravastatin on the development of vein graft intimal hyperplasia. In addition, we demonstrate that pravastatin showed endothelial cell-tropic benefits through both the inhibition of Rho-kinase activity and acceleration of eNOS expression in vitro. Because the clinical benefits and safety of pravastatin have been established to a certain extent through long-term clinical usage, pravastatin may soon become standard treatment after vein bypass grafting.     

Development and regression of intimal thickening of arterially transplanted autologous vein grafts in dogs

To investigate the influence of hemodynamic conditions on intimal thickening of arterially transplanted autologous vein grafts, two experimental models of canine femoral arteries were prepared. In group I, in which an autologous vein graft was transplanted under abnormal flow conditions (distal poor runoff), intimal thickening gradually developed and reached 358 +/- 33 microns at 8 months, whereas no thickening was observed under normal flow conditions at any time throughout the observation period. In group II the thickened intima, which developed under abnormal flow conditions for 1 month, was reimplanted into the contralateral leg under normal flow conditions. The thickness of the intima markedly regressed to about 66% at 1 month, 50% at 3 months, and 25% at 8 months, respectively, whereas no regression of the thickened intima was observed under continued abnormal flow conditions. Electron microscopic studies revealed that the thickened intima in group I was composed of proliferation of transformed smooth muscle cells with a marked increase in the mitochondria, the rough endoplasmic reticulum, and an abundant fibrous matrix, whereas with the regressed thickness of the intima of group II, the smooth muscle cells were spindle-shaped with distinct myofibrillae. These results provide pertinent data on the process involved in the intimal thickening in cases of graft placement.     

Reduction of intimal hyperplasia and enhanced reactivity of experimental vein bypass grafts with verapamil treatment.

Recent studies have shown that calcium antagonists exert an antiatherogenic effect in animals fed cholesterol. Vein graft intimal hyperplasia is believed to be an early event in atherosclerotic lesion formation, which is a significant cause of graft failure. Altered vasoreactivity has also been postulated in the etiology of vein graft failure. Therefore this study examined the effect of verapamil treatment on the development of intimal hyperplasia and the vasoreactivity of experimental vein bypass grafts. The right external jugular vein was grafted into the right carotid artery of 30 male New Zealand white rabbits fed normal rabbit chow. The left external jugular vein was used as the control vein. Fifteen animals received verapamil (1.25 mg/day for 28 days) via the femoral vein by means of an osmotic pump. In 15 control animals the pump contained saline. Plasma verapamil concentration was 50.9 +/- 13.2 ng/mL (x +/- SEM), a dose that showed no effect on either blood pressure, total serum cholesterol, or in vitro platelet aggregation to ADP. Fourteen of fifteen grafts were patent in each group, for a patency rate of 93%. Histologic examination using computer morphometry showed significant reduction of intimal hyperplasia at the proximal, middle, and distal graft segments (p less than 0.05). In addition in vitro isometric tension studies of the vein grafts and control veins showed that verapamil causes enhanced reactivity of both vein grafts and control veins in response to norepinephrine and histamine (p less than 0.05). Reactivity of vein grafts to serotonin was unaltered. While none of the normal veins in the control group responded to serotonin, normal veins treated with verapamil contracted readily in response to serotonin. Endothelial-dependent relaxation to acetylcholine was absent in both control and verapamil-treated vein grafts, while normal veins from both groups responded to the same extent to acetylcholine. Because we could not demonstrate any difference in platelet or endothelium function between untreated and verapamil-treated animals, we examined the direct effect of verapamil on smooth muscle. Verapamil significantly inhibited [3H]-thymidine incorporation into DNA in vascular smooth muscle cells in culture in a dose-dependent manner. Verapamil treatment significantly reduces intimal hyperplasia in experimental vein grafts and inhibits smooth muscle cell proliferation in culture. Furthermore the enhanced reactivity to norepinephrine and histamine in the verapamil-treated vessels has no detrimental effect on the patency rate at 4 weeks. Thus by inhibiting intimal hyperplasia, calcium antagonists may improve the long-term patency of vein bypass grafts.     

聚对二氧环己酮血管外支架抑制移植静脉内膜增生的机制

目的 构建可降解性聚对二氧环己酮(PDS)血管外支架抑制移植静脉内膜增生的动物模型,探讨PDS外支架抑制移植静脉内膜增生的作用及其机制。方法 建立兔颈外静脉移植模型,24只新西兰大白兔分为单纯移植组(n=12)和PDS外支架组(n=12)。术后4周及12周取出移植静脉,测量移植静脉内膜、中层面积及厚度,免疫组织化学法和实时荧光定量逆转录-聚合酶链反应(RT-PCR)法检测增殖细胞核抗原(PCNA)、转化生长因子-β1(TGF-β1)和肾素-血管紧张素Ⅱ受体1( AT1 R)表达。结果 24只兔均存活,移植血管全部通畅。外支架组中膜面积、内膜面积、中膜厚度、内膜厚度各值皆小于单纯移植组,差异有统计学意义(P＜0.05)。免疫组织化学与RT-PCR检测表明血管外支架组TGF-β1在外膜中过度表达,而中膜和内膜表达减少。术后4周,外支架组AT1R表达水平低于对照组,12周时,两组AT1R表达水平都降低,差异无统计学意义(P＞0.05)。结论 大孔隙、非限制性PDS血管外支架通过形成新生外膜、调节细胞因子再分布等机制有效地抑制内膜和中膜的增生,其用于抑制移植静脉内膜增生是可行的。