改良原位胶原酶循环肝灌注法分离猪肝细胞

目的:建立改良原位胶原酶循环肝灌注猪肝细胞分离方法方法:猪门静脉插管,分别用D-Hanks液和胶原酶循环原位肝灌注分离猪肝细胞,分离后的肝细胞以5×108/L-1培养,观察分离和培养7d的肝细胞产量、活率和蛋白质、葡萄糖合成功能、G-6Pase活性、安定转化功能和LDH漏出量.结果:分离获得的肝细胞总量为5.2×107/g肝组织,肝细胞活率98.7%,培养7Dk保持稳定.安定转化功能在培养3d最强(安定浓度3d时为68±6pgcell-1);葡萄糖合成功能从1d时1.05±0.15nmolcell-1下降到3d时0.74±009nmolcell-1G-6-Pase活性从0d时1871±282zmolcell1下降到1d时1218±218zmolcell-1,然后维持在较低水平;LDH漏出量在3d较高.结论:改良原位胶原酶循环肝灌注法分离猪肝细胞.     

毛毕吸虫成虫分离方法的研究初报

目的为提高毛毕吸虫成虫的收集率。方法对家鸭行离体或在体肝静脉逆行灌注收集肝内和门静脉内成虫;同时对家鸭灌注降主动脉,以收集肝内、门静脉内和肠系膜静脉的成虫。结果应用上述方法可收集到大量活的无损伤的成虫。结论灌注法优于解剖法,且简便、快捷,实用性强。     

Survivin在原发性肝细胞癌中的表达及意义

目的　Survivin是凋亡抑制蛋白中的一种 ,选择性地表达于恶性肿瘤组织。该文研究Sur vivin基因在原发性肝细胞癌中的表达及生物学意义。方法　收集 2株肝细胞癌细胞株 ,4 0例原发性肝癌组织标本及相应的癌旁组织 ,以Westernblotting法检测Survivin蛋白表达 ;半定量RT PCR法检测SurvivinmRNA表达 ;肝癌细胞凋亡指数采用原位末端标记法检测。结果　 2株肝癌细胞株和 85 % (34例 )的肝癌组织表达Survivin蛋白和mRNA ,而癌旁组织内无一例阳性表达。Survivin蛋白表达的阳性率在肝内转移组为 93.5 % ,显著高于肝内无转移组 (5 5 .6 % ,P <0 .0 5 ) ;在门静脉癌栓浸润组为 92 .8% ,显著高于无门静脉癌栓浸润组 (6 6 .7% ,P <0 .0 5 )。RT PCR显示 ,2株肝癌细胞株和 85 .0 % (34例 )的肝癌组织表达SurvivinmRNA ,与Westernblotting的结果一致 ,SurvivinmRNA的表达水平在肝内转移组 (1.10 5± 0 .396 )和门静脉癌栓浸润组 (1.137± 0 .4 0 4 )中 ,显著高于肝内无转移组 (0 .5 72± 0 .0 82 )和无门静脉癌栓浸润组 (0 .6 2 7± 0 .12 2 ,P <0 .0 5 )。所有肝癌组织标本中均可检测到凋亡细胞 ,但Survivin表达阳性组的凋亡指数 (1.15 2 %± 0 .32 6 % )显著低于Survivin表达阴性组 (4.5 0 2 %± 0 .830 % ,P <0 .0 5     

经颈静脉肝内门体分流术治疗食道静脉曲张破裂出血

采用经颈静脉肝内门体分流术(TIPSS)治疗5例肝硬化门静脉高压症(CPH)食管静脉反复破裂出血病人，平均门静脉压力由术前3．6±0．7kPa，降至术后1.73±0．35kPa，术后24小时全部病例出血均停止，门静脉血流阻力系数降低，曲张静脉消失或减轻。TIPSS治疗门脉高压食管静脉反复性出血具有损伤小、合并症少，近期疗效确切等特点，是治疗门脉高压食道静脉破裂出血的首选方法。     

肾上腺髓质素在肝硬化大鼠肝脏、门静脉的表达及意义

目的　探测肾上腺髓质素(ADM)在肝硬化大鼠门静脉血浆浓度及在肝脏、门静脉的表达。方法　以CCL4诱导肝硬化大鼠模型,正常大鼠作对照。造模完成后于门静脉采血,同时取门静脉及肝组织。以放射免疫法测定大鼠门静脉血浆ADM浓度。免疫组化染色(SABC法)检测ADM在大鼠肝脏及门静脉的表达,并以Ridit分析作半定量分析。结果　肝硬化大鼠门静脉血浆ADM浓度明显高于正常大鼠(5 3 .61±18.2 7对3 6.84±12 .67,t=3 .2 1,P <0 .0 1)。正常大鼠除肝窦外,肝细胞内也有ADM表达,肝硬化时肝脏和门静脉表达均明显增加(U肝脏=2 .69,P <0 .0 5 ;U门静脉=2 .2 7,P <0 .0 5 )。结论　肝硬化时ADM在肝细胞、肝窦内皮细胞,门静脉血管的表达及门静脉血浆浓度均较正常大鼠升高。肝细胞、肝窦内皮细胞,门静脉表达ADM增加可能是循环ADM浓度增高的一个因素。肝硬化时ADM可能在门静脉压力的调节中发挥重要作用。     

内皮素受体拮抗剂BMS-182874对大鼠心肌缺血再灌注损伤的保护作用

为观察选择性内皮素A受体拮抗剂BMS 182 874对离体灌注心脏缺血再灌损伤的影响 ,并进一步探讨其可能的作用机制。按照Langendorff方式灌注离体大鼠心脏 ,全心缺血 30min后再灌注 30min诱导急性心肌缺血模型。给予BMS 182 874 (10 μmol/L或 5 0 μmol/L)持续灌注离体心脏 ,连续记录心功能 ,检测冠状动脉流出液中的肌酸激酶和心肌组织中的肿瘤坏死因子α含量。结果发现 ,与对照组相比 ,缺血再灌注组心功能显著下降 ,左心室内压和左心室内压最大变化率 (±dp/dtmax)、冠状动脉流量明显降低 (P <0 .0 1) ,肌酸激酶 (2 3.9± 2 .2比 2 17.5± 14 .1,P<0 .0 1)以及心肌组织中肿瘤坏死因子α含量明显增加 (6 4 5± 4 5比 192 6± 14 1,P <0 .0 1) ,而 10 μmol/L和 5 0 μmol/L的BMS 182 874显著减轻缺血后心功能的损害 ,升高左心室内压和左心室内压最大变化率 (±dp/dtmax) (P <0 .0 1) ,同时降低肌酸激酶 (185± 14和 14 3± 11,与缺血再灌注组相比P <0 .0 5和P <0 .0 1)和心肌组织中的肿瘤坏死因子α含量 (12 2 2± 6 7和 110 8± 5 7,与缺血再灌注组相比P <0 .0 1)。结果提示 ,内皮素A受体拮抗剂能显著减轻心肌缺血再灌注损伤 ,其保护作用可能与抑制心肌组织中肿瘤坏死因子α生成有关。     

血清铁、铁蛋白和脂肪肝关系的研究

目的　研究血清铁、血清铁蛋白与酒精性、非酒精性脂肪肝的关系。方法　采用 1秒钟快速肝穿刺 ,对 97例脂肪肝患者取肝组织标本 ,行HE和铁染色 ,分别用原子吸收光谱法和放射免疫法检测患者的血清铁和血清铁蛋白。结果　中重度酒精性脂肪肝 (AFL)血清铁、血清铁蛋白测定值 [(2 0 .9± 9.3) μmol/L ,(2 17.6± 71.8)ng/ml;(2 9.1± 6 .5 ) μmol/L ,(2 84 .7± 77.9)ng/ml]与对照组 [(10 .5± 5 .7) μmol/L ,(14 3.3± 71.9)ng/ml]比较明显升高 (P <0 .0 1)。重度非酒精性脂肪肝 (NAFL)的血清铁和血清铁蛋白 [(2 1.5± 11.1) μmol/L ,(199.3±72 .1)ng/ml]和对照组 [(10 .5± 5 .7) μmol/L ,(14 3.3± 71.9)ng/ml]比较亦显著升高 (P <0 .0 1) ,而且AFL多出现肝细胞灶性坏死 (6 7% )和肝铁过载(87% )。结论　重度NAFL及中重度AFL多出现血清铁和铁蛋白增高 ,AFL多合并肝铁过载 ,血清铁、铁蛋白可以作为肝铁过载的重要指标。     

成人脂肪肝整肝肝细胞分离及大量冻存的实验研究

目的建立成人脂肪肝整肝肝细胞的分离以及人肝细胞大量冻存技术,为生物人工肝提供稳定的人肝细胞来源。方法采用胶原酶经肝静脉逆行灌注的方法分离成人重度脂肪肝的整肝肝细胞,并比较常规冻存和程序冻存后肝细胞在细胞活性、贴壁率、LDH漏出量及白蛋白合成能力的差异。结果采用添加N-乙酰半胱氨酸(NAC)的胶原酶灌注液分离肝细胞的产量为(7.4±0.5)×10~6 cells/g肝组织,活性为(81.4±3.4)%,而未添加NAC组的肝细胞产量为(5.6±0.8)×10~6 cells/g肝组织和活性为(67.3±5.0)%,差异均有统计学意义(P0.05)。分离的人肝细胞采用程序冻存后在细胞活性、贴壁率及白蛋白合成能力方面均相应高于常规冻存组(P0.05),LDH漏出量低于常规冻存组(P0.05)。结论应用添加NAC的胶原酶灌注液经肝静脉逆行灌注可提高脂肪肝整肝肝细胞分离的活性及产量,采用程序冻存的方法可以提高冻存人肝细胞的活性,满足生物人工肝对肝细胞的需要。     

逆灌注对移植肝缺血再灌注损伤的影响

目的:探讨原位肝移植中经下腔静脉逆行灌注对移植肝缺血再灌注损伤的影响.方法:36例大鼠肝移植随机分为3组,每组12例.门静脉组即经门静脉顺行灌注,肝动脉+门静脉组即同时开放肝动脉及门静脉顺行灌注,下腔静脉组即先吻合下腔静脉后开放逆行灌注,然后吻合门静脉及肝动脉.分别检测术后1、6及24 h的血清转氨酶、移植肝病理变化及...     

血管内皮细胞生长因子基因涂布支架预防门腔静脉分流支架狭窄的实验研究

目的　通过转染人血管内皮细胞生长因子 (phVEGF)基因促使血管内支架处内皮细胞生长以预防支架内再狭窄的发生。方法　将 phVEGF基因局部涂布于由多聚赖氨酸包被的血管内支架上 ,采用介入方法经颈静脉插管至 6只狗的右肝静脉 ,同时以置入裸支架为对照组。结果　支架置入后第 1周 ,在转染 phVEGF基因组局部血管组织内 ,RT PCR法检测到 phVEGF的表达 ;荧光显微镜下观察到部分血管内皮细胞发出黄绿色荧光 ;扫描电镜显示支架腔内皮分化程度较对照组高。置入后第 8周 ,转染 phVEGF基因组 (n =6 )血管造影显示支架通畅 ,而对照组 (n =5 )则均发生再狭窄 ;支架端肝静脉血管平均新生内膜厚度 ,平均新生内膜面积 ,百分狭窄面积均明显小于对照组 [(0 .16 7± 0 .10 3)mm比 (1.96 5± 0 .72 5 )mm ,(2 .5 6 8± 1.5 2 6 )mm2 比 (17.5 96± 7.939)mm2 ,(11.46 2± 5 .42 3) %比 (6 8.5 0 5± 2 4.0 0 3) %,P <0 .0 5 ];免疫组化显示平滑肌细胞增殖程度 (PLI)也显著高于对照组 [(0 .0 30± 0 .0 0 2 )比 (0 .0 42± 0 .0 0 3) ,P <0 .0 5 ) ]。结论　phVEGF基因涂布支架可加速支架内皮化的形成 ,进而抑制血管内支架置入术后血栓形成和内膜增殖引起的再狭窄发生。     

Magnetic resonance assessment of myocardial perfusion via catheter-based ventricle-coronary vein bypass in porcine myocardial infarction model.

OBJECTIVE: The goal of this study was to investigate the efficacy of VPASS with physiological measurements, magnetic resonance imaging (MRI), and histology in a porcine model of myocardial infarction. BACKGROUND: A catheter-based ventricle-to-coronary vein bypass (VPASS) has been proposed as a potential treatment strategy for refractory coronary artery disease patients. METHODS: In an acute setting, the VPASS implant was deployed percutaneously in three swine. The partial pressure of oxygen (PO(2)) in the anterior interventricular vein (AIV) and left ventricle (LV) were measured before and after VPASS implant with various combinations of balloon occlusion in the AIV and left anterior descending artery (LAD). In a separate chronic study, the VPASS procedure was completed on three swine with a mid-LAD occlusion. Thirty days post-VPASS procedure, angiography, contrast-enhanced MRI, and histology were performed to assess myocardial viability. Perfusion was analyzed using the average percent signal intensity change (APSIC) in the anterior walls (AW) and inferior walls (IW). RESULTS: The VPASS implant was performed without complication. Post-VPASS implantation, the distal AIV PO(2) increased up to the LV PO(2) level during simultaneous AIV and LAD blockage (432 +/- 24 mmHg). At day 30, quantitative perfusion analysis demonstrated no difference in APSIC between AW and IW (125 +/- 26% vs. 137 +/- 38%, P = 0.46). Delayed enhancement and histology showed focal subendomyocardial infarction. CONCLUSIONS: VPASS implant with simultaneous AIV and LAD occlusion allows perfusion of oxygenated blood to the distal AIV, which in the setting of an acute myocardial infarction model was capable of rescuing most of the myocardium at risk.     

Coronary angioplasty via saphenous vein grafts

Percutaneous transluminal angioplasty of coronary stenoses distal to anastomosis of a venous graft was attempted through the graft 22 times in 19 patients. Ten patients had stable angina, seven unstable angina and two patients acute myocardial infarction. The mean interval between bypass surgery and angioplasty was 6.5 years (range 1-15). Fifteen lesions were dilated in the left anterior descending artery, five in the right coronary artery, and two in the circumflex artery. Three procedures were for double lesions. In two cases, a stenosed vein graft was also dilated. All grafts were cannulated with an El Gamal guiding catheter. The procedure failed in two cases. The remaining 20 lesions were successfully dilated. Early and late occlusion of the graft occurred in one patient, and coronary arterial stenosis recurred in two patients. All three patients underwent successful redilatation. The 17 patients undergoing successful dilatation were asymptomatic, with a normal exercise test and/or maintained angiographic result at follow-up of 14 months mean duration (range 2-48). Angioplasty of coronary stenosis through a vein graft is feasible, safe and effective. This therapeutic approach avoids the need for repeat bypass surgery and, as judged by long-term follow-up, has a favourable clinical outcome.     

选择性上肺静脉造影显示下肺静脉开口位置的研究

目的探讨心房颤动(简称房颤)射频消融治疗中通过选择性上肺静脉造影显示下肺静脉开口位置的可行性与效果。方法97例房颤患者,取左前斜位(LAO)50°和右前斜位(RAO)50°两个体位进行左上肺静脉造影。取LAO50°及RAO30°两个体位进行右上肺静脉造影。结果97例(100%)均可通过选择性左上肺静脉造影清晰显示左下肺静脉开口的下缘,其中78例(80.4%)可以清晰显示左下肺静脉的开口前缘。79例(81.4%)患者可以清晰显示左下肺静脉开口的后缘。选择性右上肺静脉造影时,86例(88.7%)能够清晰识别右下肺静脉开口下缘,76例(78.4%)能清晰显示右下肺静脉开口前缘,81例(83.5%)能清晰显示右下肺静脉开口后缘。结论选择性上肺静脉造影不仅能够显示上肺静脉的开口位置,而且在大部分患者中亦能较清晰显示下肺静脉的开口解剖。     

经腋静脉途径置入心脏电极导线的临床研究

目的探讨经皮腋静脉穿刺置入冠状窦电极或心内膜电极导线的可行性与安全性。方法入选216例具有电生理检查指征、起搏器和ICD置入指征的患者,按简单随机法分为实验组与对照组,分别经腋静脉途径和经锁骨下静脉途径置入冠状窦电极或心内膜电极导线。比较两种途径置入冠状窦电极或心内膜电极导线的穿刺成功率和并发症。结果实验组与对照组穿刺成功率分别为97.3%(109/112)和95.2%(99/104);两组并发症发生率分别为9.8%(11/112)和8.7%(9/104),两组患者穿刺成功率和并发症发生率均无统计学差异(P均0.05)。其中,对照组发生气胸3例,实验组无一例发生气胸(P0.05)。结论经腋静脉途径置入冠状窦电极或心内膜电极导线安全可行。     

Simultaneous urokinase perfusion in renal artery and vein in a case of renal vein thrombosis]

We present he case of a young man with nephrotic syndrome, caused by membranous glomerulonephritis, who developed renal vein thrombosis with extension to the inferior vena cava is presented. Renal vein thrombosis was diagnosed by echo Doppler and confirmed by angio-CT scan. At the hospitalization the patient presented a severe left flank pain, edema of the lower limbs and painful left testicular tumefaction. The treatment consisted of: 1) systemic anticoagulation with sodic heparin, 2) placement of temporary vena cava filter through the right jugular vein, 3) direct thrombolysis into endocaval thrombus with early lysis of thrombus, and 4) renal thrombolysis with selective simultaneous renal artery and renal vein infusion of urokinase. Angiography performed after 24 hours of loco-regional thrombolysis showed complete lysis of renal thrombus; clinically there was a regression of left flank pain. We conclude that, face to renal vein thrombosis, thrombolytic treatment with simultaneous renal artery and renal vein perfusion is mandatory. Furthermore it is very important, in presence of caval extension of renal thrombus, to place a temporary vena cava filter before starting thrombolysis, considering the high risk of pulmonary embolism related to this pathology.