活体荧光显微镜技术在观察肝硬化门静脉高压大鼠肝脏微循环结构变化中的应用

目的探讨活体荧光显微镜技术观察肝硬化门静脉高压模型大鼠的肝脏微循环结构变化的应用效果。方法取雄性SD大鼠70只,将其中40只SD大鼠采用随机数字表法分为假手术组、胆总管结扎（BDL）模型2周组、BDL模型4周组及BDL模型6周组,每组10只,应用活体荧光显微镜技术观察大鼠肝脏微循环结构变化。将余下的30只sD大鼠于BDL模型建立4周后采用随机数字表法分为3组,动态观察给予生理盐水（生理盐水组）、内皮素-1（ET-1,ET-1组）、NO供体亚硝基半胱氨酸（GSNO,GSNO组）后BDL模型大鼠肝脏微循环的急性改变。组间比较采用单因素方差分析,组内比较采用配对样本t检验。结果BDL模型大鼠共死亡9只,存活率为85．0％（51／60）。假手术组大鼠全部存活。随着BDL造模时间延长,肝脏肝窦直径逐渐变小,BDL模型2周组、BDL模型4周组及BDL模型6周组肝窦直径分别为（13．6±1．0）μm、（8．8±0．7）μm和（8．0±0．5）μm,与假手术组的（17．4±1．0）μm比较,差异有统计学意义（t=5．86,18．24,15．57,P〈0．05）。随着BDL造模时间延长,肝窦开放数量逐渐减少,BDL模型2周组、BDL模型4周组及BDL模型6周组肝窦开放数量分别为（6．8±0．8）个／200μm、（4．3±1．8）个／200μm、（4．0±1．2）个／200μm,与假手术组的（8．8±0．5）／200μm比较,差异有统计学意义（t=3．25,2．77,2．12,P〈0．05）。注射生理盐水15min后,生理盐水组肝窦直径为（7．2±1．2）μm,与注射前的（6．9±0．5）Dm比较,差异无统计学意义（t=0．89,P〉0．05）;肝窦开放数量为（6．8±1．4）＋／200μm,与注射前的（6．6±0．4）个／200μm比较,差异无统计学意义（t=1．12,P〉0．05）。注射ET-115min后,ET-1组肝窦直径为（5．4±0．5）μm,与注射前的（7．9±0．6）μm比较,差异有统计学意义（t=7．39,P〈0．05）;肝窦开放数量为（5．4±1．8）＋／200μm,与注射前的（5．8±1．2）个／200μm比较,差异无统计学意义（t=0．84,P〉0．05）。注射GSNO15rain后,GSNO组肝窦直径为（11．4±1．3）μm,与注射前的（7．5±1．7）μm比较,差异有统计学意义（t=5．95,P〈0．05）;肝窦开放数量为（6．4±1．6）＋／200μm,与注射前的（5．6±0．8）个／200μm比较,差异无统计学意义（t=0．54,P〉0．05）。结论活体荧光显微镜技术可以通过观察肝脏微循环结构变化在一定程度上评价肝纤维化的程度,并且能够通过动态捕捉肝窦直径和肝窦开放数量在药物作用影响下发生的改变。     

肢体缺血再灌注后微循环变化及其发生机制的实验研究 1．兔足背肌腱表面微循环活体观察法的建立

我们对兔后肢趾长伸肌腱的血供系统进行了解剖观察，发现其环韧带以远接近跖趾关节的一段是制作微循环活体观察标本的理想部位。该处肌腱表面有丰富的微血管，且血管的背景为白色的肌腱，反衬明显；肌腱上方仅有皮肤和少量疏松组织覆盖，因此容易解剖。在此基础上，我们成功地创立了足背肌腱表面微循环活体观察法，为利用兔后肢缺血模型研究肢体缺血再灌注后的活体微循环变化开辟了一条新途径。     

降钙素基因相关肽对急性肝损伤小鼠肝脏微循环的干预

目的：研究降钙素基因相关肽（CGRP）对刀豆蛋白A（ConA）诱导的小鼠肝损伤模型肝脏微循环的影响。方法：昆明种小鼠60只,随机分为ConA损伤组、CGRP干预组和空白对照组（n=20）,空白对照组注射生理盐水,ConA损伤组用ConA诱导建立小鼠急性肝损伤模型,CGRP干预组在用ConA诱导建模前体外注射CGRP,各组中10只于处理后用激光多普勒血流仪测肝脏的平均血流灌注量,另10只活体观察肝脏微循环流速,最后处死,全部HE染色观察肝脏病理变化。结果：CGRP干预组肝脏的平均血流灌注量和血液流速较ConA损伤组明显增加（P〈0.01）,病理学改变明显减轻。2组的血细胞浓度差异无统计学意义（P〉0.05）。结论：CGRP能够通过改变肝脏组织的灌注影响急性肝损伤时肝脏微循环障碍的程度和病理改变。     

肢体缺血再灌注后微循环变化及其发生机制的实验研究——1.兔足背肌腱表面微循环活体观察法的建立

我们对兔后肢趾长伸肌腱的血供系统进行了解剖观察,发现其环韧带以远接近跖趾关节的一段是制作微循环活体观察标本的理想部位。该处肌腱表面有丰富的微血管,且血管的背景为白色的肌腱,反衬明显;肌腱上方仅有皮肤和少量疏松组织覆盖,因此容易解剖。在此基础上,我们成功地创立了足背肌腱表面微循环活体观察法,为利用兔后肢缺血模型研究肢体缺血再灌注后的活体微循环变化开辟了一条新途径。     

肢体缺血再灌注后微循环变化及其发生机制的实验研究：1.兔…

我们对兔后肢趾长伸肌腱的供系统进行了解剖观察，发现其环韧带以远接近跖趾关节的一段是制作微循环活体观察标本的理想部位。该处肌键表面有丰富的微血管，且血管的背景为白色的肌腱，反衬明显；肌腱上方仅有皮肤和少量疏松组织覆盖，因此容易解剖。在此基础上，我们成功地创立了足背肌键表面微循环活体观察法，为利用兔后肢缺血模型研究肢体缺血再灌注后的活体微循环变化开辟了一条新途径。     

黄腐酸钠对大白鼠肝缺血/再灌注微循环作用的实验研究

目的　探讨通过肝缺血 /再灌期 ( ischem ia and reperfusion I/ R)给予药物预处理动物模型来探索不同药物、不同给药途径防治肝脏 I/ R期损伤的有效方法。方法　 60只 Wistar大鼠分成 5组 :A组 (正常对照组 )、B组 ( I/ R期无药物干预模型组 )、C( 10 % L- arg静注干预组 )、D组 ( 1% SF静注干预组 )、E组 ( 2 % SF实验前 3 d口服干预组 ) ,动脉夹阻断肝门 4 5 min后去除动脉夹再灌注 1h。于阻断肝门前、阻断后 4 5 m in、再灌注 1h测定肝脏微循环血流量 ,检测血清 AL T、AST值 ,同时取测定点处肝脏组织病理进行光镜、电镜观察分析。分别用统计学 t检验及单因素方差析的方法比较 5组间的差异和意义。结果　阻断肝门前 E组肝脏微循环血流量明显高于 A、B、C、D组 ( P<0 .0 1) ,阻断 4 5 m in时 B、C、D、E组微循环血流量均呈一致下降趋势 ,E组再灌注 1h后回升至 2 8.3 9± 0 .83 ( V) ,与 B组 2 4 .71± 0 .4 5 ( V)比较明显增高 ( P<0 .0 5 )。病理分析证实 I/ R期肝组织损伤主要发生在再灌注初期肝小叶的中央静脉区及肝腺泡 区 ,光镜半定量分析显示 E组肝细胞坏死数目明显减少 ( P<0 .0 5 )。结论　阻断肝门后肝脏微循环明显下降 ,再灌注早期可回升 ,但不能达到正常值。黄腐酸钠 ( SF)可提高正常肝脏     

活体肝脏移植猪动物实验训练模型的建立

目的建立活体肝脏移植猪动物实验训练模型,让外科手术人员能够学习和掌握活体肝移植所必需的技巧。方法施行6组供、受体共12头猪活体肝移植模拟手术,每组猪性别不限。结果5头受体猪存活至术后第7天,并进行无痛处死;1头受体猪于术后第2天死于创面广泛出血。4头受体猪发生了胆瘘,均没有发生显著的胆汁性腹膜炎。结论本实验模型的各个步骤,基本整合了进行活体肝移植所需要的所有外科操作技巧;作为一种模拟活体肝移植动物实验模型,能够有效地应用于对移植人员进行人活体肝移植手术技巧的训练。     

碱性成纤维细胞生长因子促进缺血皮瓣微循环重建的实验研究

目的探讨碱性成纤维细胞生长因子对缺血皮瓣微循环重建的影响。方法以Ｗｉｓｔａｒ大鼠为实验动物，在其背部形成７ｃｍ×１ｃｍ随意皮瓣，通过测定皮瓣存活面积、病理切片、电镜观察、组织化学染色、图像分析技术等手段进行观察。结果给药组以毛细血管增生为特征，微血管断面面积及皮瓣存活率均显著高于对照组。结论碱性成纤维细胞生长因子可促进缺血皮瓣微循环重建。     

碱性成纤维细胞生长因子促进缺血皮瓣微循环重建的实验研究

目的探讨碱性成纤维细胞生长因子对缺血皮瓣微循环重建的影响。方法以 Wistar 大鼠为实验动物,在其背部形成7cm×1cm 随意皮瓣,通过测定皮瓣存活面积、病理切片、电镜观察、组织化学染色、图像分析技术等手段进行观察。结果给药组以毛细血管增生为特征,微血管断面面积及皮瓣存活率均显著高于对照组。结论碱性成纤维细胞生长因子可促进缺血皮瓣微循环重建。     

Analysis of the microcirculation during xenogeneic liver perfusion in the guinea pig - rat model. The contribution of leukocytes to the rejection process

Abstract Since the main feature of hyperacute rejection is a disturbance of the xenograft's microcirculation, we analyzed microhemodynamic parameters during xenogeneic hemoperfusion of the guinea pig (GP) liver and investigated the contribution of leukocytes to the rejection process using intravital fluorescence microscopy. Isolated GP livers were hemoperfused via the portal vein in a recirculating system with a constant flow of 1 ml/min per g liver. In contrast to isogeneic perfusion with heparinized GP blood, a disturbance in the microcirculation was observed during xenogeneic perfusion using heparinized rat blood, with significantly higher values of perfusion pressure, reduced sinusoidal perfusion rates, and a larger number of stagnant leukocytes. A complete breakdown of the microcirculation, with the highest values of perfusion pressure and the smallest perfusion index as associated with 100% accumulated leukocytes when rat blood was anticoagulated with sodium citrate. Almost isogeneic perfusion values were obtained when fucoidin, which inhibits L-selectin-dependent cell interaction, was added to heparinized rat blood. These data indicate that leukocyte-endothelial cell interaction contributes to xengeneic rejection.     

Role of hypotension in brain-death associated impairment of liver microcirculation and viability

Hypotension in brain-dead organ donors is considered a determinant factor of graft viability. The aim of this study was to elucidate the role of hypotension in brain-death associated impairment of hepatic microcirculation and function. Male Sprague-Dawley rats with an intracranial balloon were used. Group I (n = 7) served as sham controls. In group II (n = 7) brain death was induced through inflation of an intracranial balloon. In group III (n = 7) hypotension without brain death was induced by means of pentobarbital. In group II, a steep rise of arterial pressure was followed by a fall to a lower level (P < 0.01, vs. group I). Also in group III arterial pressure was lower (P < 0.01, vs. group I). In group II, bile production was diminished (P < 0.05). Impaired sinusoidal perfusion (P < 0.01) and enhanced leukocyte endothelium interaction (P < 0.05) were documented in hepatic microvasculature. Electron microscopic analysis revealed vacuolization of hepatocytes; these changes were not observed in group III. Brain death induces specific changes of liver microcirculation, function and histomorphology. Independent of associated hypotension, brain death per se impairs donor liver graft quality. Received: 28 September 1999 Revised: 5 May 2000 Accepted: 13 September 2000     

Influence of anti-inflammatory and vasoactive drugs on microcirculation and angiogenesis after burn in mice

Objective

The treatment of burns remains a challenge. Besides the administration of physiological saline, local disinfection and symptomatic medications, no causal therapy is known to accelerate angiogenesis and wound healing.The aim of this study was to investigate the influences of dilatative and anti-inflammatory acting drugs on microcirculation, angiogenesis and leukocyte behavior, which had shown positive effects in former burn studies.

Methods

The ears of male hairless mice (n = 47) were inflicted with full thickness burns using a hot air jet. Then the affects of five intraperitoneal injections of either acetylsalicylic acid (ASA), isosorbide dinitrate, prostaglandin E1 (PGE1) or sodium chloride (each administered to one of four corresponding study groups), on microcirculation, leukocyte-endothelial interaction and angiogenesis were investigated over a 12 day period using intravital fluorescent microscopy.

Results

Angiogenesis was slightly improved by PGE1 (0.3 vs. 1.3% non-perfused area in other groups on day 12, p = 0.029). Additionally, blood flow increased and rolling leukocytes decreased compared to other groups. The ASA-group showed best functional vessel density and lowest leukocyte-adhesion. The often described posttraumatic expansion of tissue damage could not be observed in either group.

Conclusion

Prostaglandin E1 improved angiogenesis, increased the blood flow and reduced the number of rolling leukocytes. ASA had positive influences on functional vessel density, edema formation and the number of sticking leukocytes. However, it reduced the blood flow.Overall, out of all the drugs tested, prostaglandin seems to have the greatest positive impact on microcirculation and angiogenesis in burns.     

Effects of warm Carolina rinse on microvascular reperfusion injury in rat liver transplantation

Abstract Recently, it has been demonstrated that the use of both cold Carolina rinse (CR, 4°C) as well as warm Ringer's lactate (RL, 37°C) attenuates microvascular perfusion failure and leukocyte (WBC) accumulation in liver grafts. The aim of this study was to analyse in vivo whether warming of CR can also lead to a reduction in microvascular reperfusion injury in rat liver transplantation. Syngeneic orthotopic liver transplantation, including arterial reconstruction, was performed in male Lewis rats (180–300 g). Livers were stored in University of Wisconsin (UW) solution for 24 h and rinsed with 15 ml CR which was either cold 4 °C ( n = 7) or warm 37 °C ( n = 8) prior to reperfusion. Hepatic microcirculation and WBC accumulation were assessed by intra-vital fluorescence microscopy, and graft function was determined by analysis of bile flow during the 90-min reperfusion period. Warm CR yielded significantly ( P < 0.01) improved sinusoidal perfusion when compared with cold CR; however, the extent of WBC adherence in both sinusoids and postsinusoidal venules did not vary between the groups. In addition, bile flow was slightly increased after warm CR. We conclude that after 24 h of cold storage in UW solution, warming of CR may offer additional benefit in the prevention of microcirculatory reperfusion injury without affecting WBC accumulation.