急性肝损伤后猪自体骨髓间充质干细胞肝内移植的MR活体示踪实验研究

目的 探讨应用临床型1.5T磁共振活体示踪磁标记猪骨髓间充质干细胞(MSCs)自体移植肝脏的可行性.方法 获取、分离、培养、扩增猪骨髓MSCs.应用菲立磁标记细胞,普鲁士蓝染色鉴定,建立急性肝损伤模型,分为标记细胞组(n=6)和未标记细胞组(n=4)经门静脉行肝内移植,分别于移植前,移植后6 h、3 d、7 d、14 d行磁共振FFE(T_2WI)序列成像,14 d后行组织切片普鲁士蓝染色.结果 普鲁士蓝染色表明MSCs的标记率达100%,磁标记MSCs肝内移植后行磁共振T_2WI序列呈明显低信号改变,并持续至细胞移植后14 d,组织切片普鲁士蓝染色显示14 d后仍有磁标记细胞存在于肝实质及肝窦中.结论 利用菲立磁可以在体外成功标记猪骨髓间充质干细胞,磁共振成像可以对经门静脉移植到肝内的磁标记MSCs进行活体示踪.     

磁标记猪骨髓间充质干细胞自体肝内移植后MR活体示踪研究

目的探讨利用磁共振成像对移植肝脏的磁标记猪骨髓间充质干细胞进行活体示踪的可行性。方法获取猪自体骨髓间充质干细胞,分离、培养、扩增。应用菲立磁（Feridex）标记细胞,普鲁士蓝染色鉴定,标记细胞组（n=6）和未标记细胞组（n=4）行经门静脉行肝内移植,分别于移植前,移植后6h、3d、7d行磁共振T1WI,T2WI,GRE序列成像,7d后行组织切片普鲁士蓝染色。结果：普鲁士蓝染色表明MSCs的标记率达接近100％,磁标记MSCs肝内移植后行磁共振T2＊WI序列呈明显低信号改变,并持续至细胞移植后7d,组织切片普鲁士蓝染色显示7d后肝内仍有移植的磁标记细胞存在于肝实质及肝窦中。结论利用SPIO可以在体外成功标记猪骨髓间充质干细胞,磁共振成像可以对经门静脉移植到肝内的磁标记MSCs进行活体示踪。     

骨髓间充质干细胞经脾移植治疗大鼠急性肝功能衰竭的实验研究

目的 探讨大鼠骨髓间充质干细胞（MSCs）经脾移植治疗急性肝功能衰竭的疗效，并观察MSCs在体内的迁徙情况。方法 收集1只SD雄性大鼠胫骨及股骨的骨髓，采用密度梯度离心联合贴壁培养法分离、纯化及扩增雄性SD大鼠的骨髓MSCs，再行免疫组化染色以观察第4代骨髓MSCs的表面标志物。联合应用D-氨基半乳糖和肿瘤坏死因子-α （TNF-α）建立24只雌性大鼠急性肝功能衰竭模型，将其随机分为2组：实验组（n=12）大鼠于造模后24 h行骨髓MSCs脾内移植；空白对照组（n=12）仅于脾内注射0.5 mL生理盐水。2组大鼠于移植后均取血检测丙氨酸氨基转移酶（ALT）、总胆红素（TBIL）及白蛋白（ALB）水平，采用PCR法检测大鼠肝脏组织中Y性别决定区基因（SRY基因）的表达，并行HE染色观察肝脏组织的病理学改变。结果 第4代MSCs表达CD44和CD29，但不表达CD34。MSCs移植72 h及以后，实验组存活5只大鼠（41.7%），空白对照组存活3只大鼠（25.0%），2组大鼠的存活率比较差异有统计学意义（P＜0.05），实验组较高。将雄性大鼠的骨髓MSCs移植于雌性大鼠的脾内后，在雌性大鼠肝脏中能检测到SRY基因的表达；且HE染色结果显示，实验组大鼠的肝功能在移植后4周明显改善。移植后与空白对照组比较，实验组各时点的ALT和TBIL水平均较低（P＜0.05）；移植后1周和2周，实验组的ALB水平均高于空白对照组（P＜0.05）。结论 骨髓MSCs经脾内移植后迁徙并定居于受损的肝脏内，可替代肝细胞的功能。     

静脉注射骨髓间质干细胞对脊髓损伤修复作用的实验研究

目的探讨静脉注射大鼠骨髓间质干细胞(MSCs)对脊髓损伤后神经修复和功能恢复的影响。方法32只SD大鼠,体重约300g,雌雄不限。体外分离、培养、纯化MSCs,应用流式细胞技术检测MSCs表面细胞标志CD34、CD45、CD29、CD90。根据改良Allen法制备大鼠脊髓损伤模型。暴露T10段脊髓,将一直径为3mm的圆形薄铜垫片置于T10段脊髓表面,以重量为10g的砝码,从5cm高度自由坠落打击该垫片,造成T10段脊髓冲击伤,损伤组24只,假手术组8只。模型建立后24h,损伤组随机分为实验组14只,对照组10只。实验组及假手术组经尾静脉注射Brdu标记的MSCs,对照组经静脉注射PBS。损伤后24h、注射MSCs后1、3、5周评价各组大鼠的神经功能状况,并检测MSCs在体内迁移、存活以及分化情况。结果细胞CD34、CD45阴性表达,CD29、CD90阳性表达。实验组运动功能改善,BBB评分高于对照组(P<0.05)。注射的MSCs在宿主损伤脊髓中聚集并存活,注射MSCs3～5周后部分细胞表达微管相关蛋白2(MAP2)、神经元特异性烯醇化酶(NSE)染色的Brdu阳性细胞。结论MSCs经静脉注射后可向脊髓损伤处聚集并存活,促进神经修复及神经功能的恢复。     

磁性荧光双标人骨髓间充质干细胞归巢至大鼠急性损伤肝组织示踪显像

目的观察磁粒子和荧光染料双标人骨髓间充质干细胞归巢至大鼠急性损伤肝组织的示踪。方法以超顺磁性氧化铁(SPIO)和氯甲基苯甲酰氨(CM-Dil)标记永生化人骨髓间充质干细胞(UE7T-13)。普鲁士蓝染色检测细胞内铁,荧光显微镜和流式细胞仪检测CM-Dil标记阳性率。建立12只急性肝损伤大鼠模型,分为实验组(n=6)和对照组(n=6),将标记细胞经尾静脉移植入实验组大鼠,未标记细胞移植入对照组大鼠。分别于移植前3h、移植后3h、3天、5天、7天应用MR T2*W、T2*map对大鼠活体成像,获得肝脏R2*值。并与肝脏组织切片普鲁士蓝染色和CM-Dil荧光表达情况对照。结果双标细胞的SPIO-PLL标记率接近100%,流式细胞仪检测CM-Dil标记阳性率达99.97%。实验组细胞移植后3天、5天肝脏R2*值均较移植前明显升高(t=7.282、7.608,P均0.01),对照组细胞移植后各时间点与移植前比较,R2*值差异均无统计学意义(F=0.99,P0.05)。普鲁士蓝阳性细胞主要分布于肝小叶中央静脉周围病变区,荧光显微镜下观察红色荧光阳性细胞与普鲁士蓝阳性细胞分布基本一致。结论 SPIO和CM-Dil可有效标记人永生化骨髓间充质干细胞(UE7T-13),不影响细胞增殖能力,临床应用型1.5T MR可对磁粒子标记的UE7T-13进行肝归巢活体示踪,CM-Dil有助于证实存活干细胞的肝内定植。     

补阳还五汤对骨髓间质干细胞移植治疗大鼠脊髓损伤的影响

目的:观察补阳还五汤联合骨髓间质干细胞(MSCs)移植对大鼠脊髓损伤后神经功能恢复以及移植的MSCs迁移情况的影响,并探讨其作用机制。方法:SD大鼠80只,其中70只用改良Allen法制备大鼠T10脊髓损伤模型,并随机分为中药 MSCs组20只、MSCs组20只、假手术组(无脊髓损伤)10只、中药组20只、空白对照组(无治疗)10只。中药 MSCs组、MSCs组、假手术组行大鼠尾静脉移植带Brdu标记的MSCs。各组大鼠于术后1、3、5周观察神经功能恢复、免疫组化检测带标记的MSCs迁移情况。结果:与空白对照组相比,治疗组神经功能测定在术后1、3、5周时均明显高于对照组(P<0·01)。术后1周移植组的脊髓组织内即可见Brdu标记阳性细胞(假手术组除外),术后5周中药 MSCs组Brdu阳性细胞计数较MSCs组有显著性差异(P<1.05)。结论:静脉注射移植的MSCs能够迁移到脊髓损伤组织,并促进神经功能的恢复。补阳还五汤能促进移植的MSCs迁移,同时有利于脊髓功能的恢复。     

骨髓间充质干细胞移植促进脊髓损伤大鼠神经功能的恢复

目的 探讨骨髓间充质干细胞(MSCs)移植对脊髓损伤大鼠神经功能恢复的影响极其可能机制.方法 采集SD大鼠骨髓,分离出MSCs,并培养、纯化,取传至第6代的MSCs,移植前1 d以5溴2脱氧尿嘧啶核苷(Brdu)标记,然后用显微注射器将其缓慢注射到脊髓损伤大鼠模型(成年SD大鼠)的受损脊髓中心,以不进行移植的脊髓损伤大鼠模型(损伤组)和假手术组为对照.术后进行运动功能评分以判断神经功能恢复情况,并切取移植区域脊髓组织,应用荧光免疫化学染色检测脊髓损伤灶边周的神经元凋亡情况(TUNEI.与NeuN双标记)以及移植的MSCs的分布和向神经元分化情况(Brdu与NeuN双标记),应用逆转录聚合酶链反应检测损伤灶区域脑源性神经营养因子(BDNF)mRNA的表达.结果 移植后第3天开始,损伤组和移植组大鼠的BBB运动功能评分明显上升,第14天后进入平台期,但移植组各时间点的.BBB运动功能评分均明显高于损伤组(P<0.05).移植后3 d,在移植组的脊髓组织中可见Brdu标记阳性细胞,少部分Brdu标记阳性细胞同时表达神经元特异性标志物NeuN;移植后第3、7天,损伤组和移植组脊髓组织中的凋亡神经元均显著多于假手术组(P<0.01),凋亡的神经元多存在于损伤周边区,但移植组的凋亡神经元显著少于损伤组(P<0.01);移植后第3、7天,假手术组未检测到BDNF mRNA的表达,损伤组和移植组均可检测到BDNF mRNA的表达,移植组的表达量分别较损伤组高28.6%和39.2%(P<0.05).结论 移植的骨髓MSCs可促进脊髓损伤大鼠神经功能的恢复,其机制除MSCs直接分化为神经元进行修复外,还可能与其改善脊髓损伤区的微环境、上调BDNF、mRNA表达、减少神经细胞凋亡有关.     

定量RT-PCR检测肝脏损伤与移植干细胞定居量的关系

目的探索经不同途径移植到同种异体大鼠的大鼠骨髓间质干细胞（MSCs），在受体大鼠肝脏的定居情况。方法以绿色荧光蛋白标记从大鼠骨髓中分离培养的MSCs，体外扩增后，分别从不同途径移植到同种异体正常和肝脏损伤大鼠体内，于移植后的第3、7天，通过荧光定量PCR检测移植的MSCs在大鼠体肝内的表达情况。结果不同途径移植的同种异体MSCs，均可在受体大鼠肝脏内定居，定居于肝脏受损大鼠的细胞量大于非肝脏受损大鼠，差异显著（P〈0.05）。在肝脏受损大鼠，移植途径与MSCs在肝脏的定居量无明显的相关性（P〉0.05）。非肝脏受损大鼠，移植途径与MSCs在肝脏的定居量明显相关（P〈0.05），并与移植后时间有关。结论标记的MSCs可以定居于受体大鼠肝脏，其定居时间与细胞数量，与肝脏是否受损伤关系密切，与移植途径关系不密切；标记的MSCs可定居于肝脏未受损伤的大鼠肝脏，定植的细胞量与移植途径和移植后时间有关。     

一种可恢复性门静脉高压症大鼠模型

目的:探讨建立可恢复性门静脉高压症大鼠模型的可行性。方法:Wistar大鼠60只,分为实验组 A、实验组B和对照组。实验组A、组B通过手术将20-gauge钝头注射针头及外径1.5mm标志管与门静脉主干平行适度结扎以缩窄门静脉,然后抽出针头,对照组开腹后只游离门静脉主干。3周后,实验组A开腹拔除门静脉旁标志管前后分别行门体静脉测压并门静脉系统造影;对照组行门静脉压力测定和造影。实验组B开腹拔除门静脉旁标志管,4周后测量门静脉压力。各组检测血清ALT,AST,BIL, 处死后肝脏常规病理检查。结果:3周后实验组A、B大鼠食管胃底静脉迂曲扩张,门体间侧支开放;实验组A大鼠拔管前、后门静脉压分别为(15.6±3.1)mmHg、(13．4±2．3)mmHg,与对照组门静脉压(7．7± 1．7)mmHg差别有统计学意义(P<0．05)。实验组B拔管4周后门静脉压为(8.4±2.7)mmHg,与对照组差别无统计学意义(P>0．05),与实验组A差别有统计学意义(P<0.05)。各组大鼠肝细胞形态结构正常,血清ALT,AST,BIL值各组无显著性差异。结论:可复性肝前性门静脉高压症大鼠模型是成功的,可用于肝硬化肝移植后血流动力学研究。     

骨髓间充质干细胞移植重建大鼠缺血心肌的实验研究

目的　探讨大鼠骨髓间充质干细胞 (MSCs)移植于缺血心肌后的增殖分化情况和对缺血心肌细胞的修复重建能力及心功能改善情况。　方法　实验组为将体外培养SD大鼠的MSCs经溴氮胞苷 (BrdU)标记后显微注射于结扎冠状动脉后的大鼠缺血心肌内 ,并以无血清培养基注射动物为对照组。 4周后观察移植细胞的分化情况 ,并通过超声多普勒、心肌核素显像、免疫组化和新生血管形成情况来检测心功能变化。　结果　实验组MSCs移植 4周后 ,在缺血心肌区内可发现不同分化阶段的心肌样细胞。超声检查发现实验组的左室射血分数 (LVEF)的改善明显好于对照组 (P <0 0 5 ) ;SPECT显示实验组心肌核素摄取显著高于对照组 (P <0 0 1) ;在促新生血管形成方面 ,实验组也明显好于对照组(P <0 0 5 )。　结论　骨髓间充质干细胞移植于缺血心肌后可重建缺血心肌 ,增加心肌灌注 ,显著改善心功能。     

自体骨髓细胞诱导肝移植大鼠长期存活的机制探讨

目的 研究自体骨髓细胞诱导肝移植大鼠长期存活的可能机制.方法 雌性受体大鼠随机分成空白对照组(A组)、D-hanks液组(B组)、全骨髓细胞组(C组)、间充质干细胞组(D组).观察大鼠的中位生存时间(median survival time,MST)、肝功能、病理变化、Sry基因原位杂交和甲胎蛋白、白蛋白免疫组化双标检测观察自体骨髓细胞的分化情况.结果 C组、D组MST均>180 d(P<0.01);血肝功能指标C组、D组降低明显,有显著差异(P<0.01);C组、D组之间比较无显著差异(P>0.05);移植术后60 d C、D组均无明显的急性排斥反应;C组、D组Sry基因原位杂交和甲胎蛋白、白蛋白免疫组化双标检测呈阳性.结论 自体骨髓细胞能减少排斥反应、诱导大鼠肝移植术后长期存活,其中间充质干细胞在移植肝内诱导分化为肝细胞发挥肝细胞功能,是其中可能的机制.     

肝移植缺血再灌注对肺损伤的影响

目的 研究原位肝移植缺血再灌注对肺部病理生理变化的影响,探讨肺损伤的发生机制.方法 选择南京军区福州总医院施行的23例原位肝移植患者.分别于手术开始进腹后5 min(Ta)、门静脉开放前5 min(Tb)和新肝期3 h(Tc)各切取右下肺一小块组织行病理检查,并作IL-1β和TNF-a的免疫组化实验.分别于麻醉后手术前(T1)、门静脉开放前5 min(T2)、门静脉开放后10 min(T3)、新肝期60 min(T4)、新肝期3 h(T5)以及术后12 h(T6)各时间点采集外周血检测血浆中细胞因子IL-1β和TNF-a含量水平.结果 外周血TNF-a和IL-1β于门静脉开放前升高不明显,但门静脉开放后显著升高.T1和T2分别与T3、T4和T5比较,差异均有显著统计学意义(P<0.01).光镜下和电镜下,Ta时间点的肺组织结构正常,Tb时间点变化不明显,Tc时间点出现显著异常变化.Tc时间点TNF-a和IL-1β表达的阳性平均积分明显高于Ta和Tb时间点,差异具有统计学意义(P<0.01).结论 移植术中移植肝缺血再灌注可使机体产生严重的全身炎症反应综合征,并导致早期急性肺损伤的发生.TNF-a和IL-1β参与了急性肺损伤的发生过程.

Abstract：

Objective To study the effects of ischemia-reperfusion in liver transplantation on the pathophysiological changes of the lung and mechanisms of lung injury. Methods We studied 23 patients who received liver transplantation at Fuzhou General Hospital of PLA. We cut a small piece of the right lung for pathological study and for L-1β and TNF-a immunohistochemistry studies at 5 minutes after the beginning of operation (Ta), 5 minutes before the portal vein was opened (Tb) and three hours after the new liver was transplanted (Tc). We also collected peripheral blood to study the concentration of IL-1β and TNF-a in the plasma at the beginning of operation (T1), the portal vein 5 minutes before opening, the portal vein (T2) ten minutes after the opening (T3) , and one hour after the new liver was transplanted (T4), three hours after the new liver was transplanted (T5), and 12 hours after operation (T6). Results The cytokines TNF-a and IL-1β in peripheral blood were not obviously increased in the portal vein before it was opened, but were significantly increased after the portal vein was opened. Comparison of T1 and T2 separately with T3, T4 and T5 showed significant differences (P<0. 01). In light and electron microscopy, the structures of the lung tissues were normal at Ta and did not change significantly at Tb. There were significant abnormalities at Tc. The average positive points of TNF-a and IL-1β expressions in the lung tissues at Tc were significantly higher than Ta and Tb(P<0. 01). Conclusion Ischemia-reperfusion in liver transplantation led to a serious systemic inflammatory syndrome,and acute lung injury. TNF-a and IL-1β were involved in acute lung injury.     

大鼠脊髓损伤后不同时期移植异体骨髓间充质干细胞的局部分布

目的 观察大鼠脊髓损伤后不同时间点经尾静脉注射移植异体骨髓间充质干细胞(mes-enchymal stem cells,MSCs)后,MSCs在损伤局部的聚集情况.方法 取成年雄性大鼠48只,建立脊髓半横断模型,术后即刻、1 d、1周、2周、3周、4周、5周、6周,经尾静脉注射移植Hoechst预标记的同种异体MSCs(5×106个/只),每个时间点6只.另设立未损伤组(空白对照)及假损伤组(实验对照)作为对照,每组6只.细胞移植后2周,处死实验动物.损伤脊髓部位连续水平纵行切片,在荧光显微镜下观察,计数单张切片平均阳性细胞数.结果 行方差分析.结果 对照组脊髓内仅见零星标记细胞[末损伤组(14+2)个,假损伤组(11±3)个],损伤后即刻至损伤4周进行细胞移植组脊髓损伤部位可见大量标记细胞聚集,其中损伤1周组最多达(2197±14)个,损伤5周组标记细胞数明显减少至(259±66)个,损伤6周组标记细胞仪为(43±15)个.标记的移植细胞集中在距离损伤部位0.5 cm区域内,主要分布于白质.结论 大鼠脊髓损伤后1个月内经静脉移植同种异体骨髓MSCs,移植细胞可向脊髓损伤部位聚集.     

骨髓间充质干细胞移植途径对胰岛移植物免疫排斥反应的影响

目的:探讨骨髓间充质干细胞(MSC)与胰岛共移植对诱导胰岛移植物免疫耐受的作用,并比较MSC不同途径移植的效果。方法:SD大鼠和Lewis大鼠分别作为供、受体。取SD大鼠股骨,贴壁培养法分离和扩增MSC,胶原酶V分离胰岛。应用链脲佐菌素制备Lewis大鼠糖尿病模型后,将其随机均分为A组(将BrdU标记的MSC与胰岛经门静脉混合输入),B组(将胰岛经门静脉输入,BrdU标记的MSC经尾静脉输入),C组(胰岛经门静脉输入,联合应用环孢素A)和D组(单纯胰岛门静脉移植)。观察各组术后血糖变化,比较各组胰岛移植物存活时间。术后第7天切取各组部分存活大鼠肝脏、胸腺、脾脏行免疫组化染色观察MSC归巢位置。结果:A,B两组大鼠术后正常血糖维持时间最长,C组次之,D组最短;各组胰岛存活时间A组为(12.1±2.3)d,B组为(8.6±1.4)d,C组为(13.2±1.9)d,D组为(2.2±0.6)d;MSC归巢部位观察显示,A组BrdU阳性的MSC主要分布于肝脏,并在植入胰岛周围形成"类微囊化效应",B组BrdU阳性的MSC主要分布于胸腺、脾脏。结论:MSC与胰岛共移植能诱导胰岛移植物免疫耐受,且MSC和胰岛混合经门静脉移植效果优于胰岛门静脉移植联合MSC外周静脉移植。     

Portal hyperperfusion causes disturbance of microcirculation and increased rate of hepatocellular apoptosis: investigations in heterotopic rat liver transplantation with portal vein arterialization

Clinical results of portal vein arterialization (PVA) in liver transplantation are controversial. One reason for this is the lack of a standardized flow regulation. Our experiments in rats compared PVA with blood-flow regulation to PVA with hyperperfusion in heterotopic auxiliary liver transplantation (HALT). In group I (n = 19), the graft's portal vein was completely arterialized via the right renal artery in-stent technique, using a 0.3-mm stent, leading to a physiological average portal blood flow. In group II (n = 19), a 0.5-mm stent was used. In group II, the average portal blood flow after reperfusion was significantly elevated (group II: 6.4 +/- 1.5; group I: 1.7 +/- 0.4 mL/min/g of liver weight; P < .001). The sinusoidal diameter after reperfusion was significantly greater in group II (9.8 +/- 0.5 microm) than in group I (5.5 +/- 0.2 microm; P < .001). Red blood cell velocity in the dilated sinusoids was significantly lower in group II (171 +/- 18 microm/s) than in group I (252 +/- 13 microm/s). Stasis of erythrocytes occurred; consequently, the functional sinusoidal density was significantly reduced in group II (38 +/- 7%) compared with group I (50 +/- 3%; P < .01). Two hours after reperfusion of the portal vein, the number of apoptotic hepatocytes was significantly higher in group II than in group I (I: 0 +/- 0 vs II: 7 +/- 9 M30-positive hepatocytes/10 high-power fields). The 6-week survival rate was 9 of 11 in both groups. In group II, 6 of 9 grafts showed massive hepatocellular necroses after 6 weeks, whereas in group I, only 1 of 9 presented a slight hepatocellular necrosis. Finally, our results demonstrate negative effects of portal hyperperfusion in transplanted livers, which are correctable by adequate flow regulation.     

Portal flow diversion is essential for graft survival in canine auxiliary partial orthotopic liver transplantation

After auxiliary partial orthotopic liver transplantation for inborn errors of metabolism, finding a balance in portal blood flow distribution between native liver and graft is complicated. We investigated the correction of hypoallantoinuria in the Dalmatian dog with a reduced-size Beagle orthotopic auxiliary liver graft, depending on intra-operative intervention in the portal flow. There were three groups: a ligation group, where the host portal vein was tied off, a free-flow group with random flow to both livers and a banding group, where the host portal vein was banded with an adjustable strapband. Metabolic correction was initially seen in all groups, but ligation led to portal hypertension and early mortality. In the free-flow group, correction was lost after 7 days, while banding preserved correction until 6 weeks. We conclude that acute ligation can lead to portal hypertension and free-flow leads to hypoperfusion and early loss of metabolic correction. Banding divided the portal blood flow between host liver and graft and prolonged metabolic correction.     

骨髓间充质干细胞自体移植时间对兔急性肾功能衰竭治疗效果的影响

目的 观察骨髓间充质干细胞(MSCs)自体移植对兔急性肾功能衰竭的治疗作用并探讨不同移植时间对其治疗效果的影响.方法 骨髓穿刺抽取新西兰大耳白兔骨髓,分离、培养、扩增MSCs.30只兔通过夹闭双肾动脉90 min后再灌注制作急性肾功能衰竭模型,随机分为移植A组、移植B组和未治疗C组,每组10只.移植A组在恢复血流即刻,移植B组在恢复血流后72 h将BrdU标记的自体MSCs经颈静脉回输移植,未治疗C组仅制作急性肾功能衰竭模型.于造模后21 d处死兔,比较各组的存活率、肾功能及肾组织形态学改变.结果 与未治疗组比较,移植组肾功能较快恢复(P<0.05),肾组织损伤明显改善(P<0.05).移植A组的效果优于移植B组(P<0.05).结论 骨髓间充质干细胞自体移植能有效治疗缺血再灌注损伤引起的急性肾功能衰竭,早期移植的效果好.     

Importance of portal flow diversion in experimental auxiliary partial orthotopic liver transplantation

BACKGROUND: Auxiliary partial orthotopic liver transplantation (APOLT) has successfully been performed in patients with noncirrhotic metabolic diseases. It remains, however, unclear if intervention in the portal venous inflow is necessary to ensure adequate portal blood flow to graft and host liver. In this experimental study we evaluate the hepatic flow during APOLT. METHODS: Left lateral/medial segmental grafts were transplanted from beagle to dalmatian dogs. Vascular structures were anastomosed end-to-end. The effect of diversion of the portal flow was studied in three groups: in the ligation group (n=3) the host portal vein was tied off, the free flow group (n=6) had random flow to both livers. In the banding group (n=11) the host portal vein was banded with a adjustable strapband to restore the pretransplantation flow distribution. RESULTS: After reperfusion the blood flow through the common portal vein decreased from 49 to 36 ml/kg/min (P<0.03) in all animals. Flow through the left portal vein decreased from 26 to 5 ml/kg/min (P<0.0001). Banding restored the flow in the left portal vein to 12 ml/kg/min, although the flow in the free-flow group remained 4 ml/kg/min. In the ligation group the total portal flow was forced toward the graft leading to the highest perfusion: 24 ml/kg/min (P<0.005). Adverse effect of this ligation was the development of portal hypertension. CONCLUSIONS: This experimental study confirms that diversion of the portal flow is necessary for adequate graft perfusion in APOLT. Banding can restore the pretransplantation flow distribution, without compromising the flow in the common portal vein.     

Hepatocyte transplantation through the hepatic vein: a new route of cell transplantation to the liver

The efficiency of hepatocyte transplantation into the liver varies with the method of administration. This study investigated whether retrograde infusion via the hepatic vein provides a sufficient number of donor cells for the liver. Donor hepatocytes were isolated from dipeptidyl peptidase IV (DPPIV(+)) rats and transplanted into DPPIV(-) rat livers either by antegrade portal vein infusion or retrograde hepatic vein infusion. Hepatocyte engraftment ratios and localization were evaluated by histological DPPIV enzymatic staining at 1 week and 8 weeks after the transplantation. No significant differences in engraftment efficiency were observed at either 1 week or 8 weeks after transplantation by either route. However, the localization of the transplanted hepatocytes differed with the administration route. Portal vein infusion resulted in predominantly periportal engraftment, whereas hepatic vein infusion led to pericentral zone engraftment. Immunohistochemical analysis showed that the transplanted hepatocytes engrafted in the pericentral zone after retrograde infusion displayed intense CYP2E1 staining similar to the surrounding native hepatocytes. CYP2E1 staining was further enhanced by administration of isosafrole, an inducing agent for various cytochrome P450 enzymes, including CYP2E1. This study demonstrates a novel approach of transplanting hepatocytes into the liver through retrograde hepatic vein infusion as the means to target cell implantation to the pericentral zone.     

内皮祖细胞移植对大鼠肝硬化作用的研究

目的 探讨内皮祖细胞(endothelial progenitor cells,EPCs)移植对四氯化碳(carbon tetrachloride,CCl4)诱导的大鼠肝硬化的作用.方法 本组38只SD大鼠中8只大鼠为正常对照,其余30只采用25%的CCl4/橄榄油灌胃制备肝硬化模型.再将肝硬化大鼠分为3组,每组10只.12周后直接处死的为肝硬化模型组,门静脉输入大鼠EPCs为EPCs移植组,经门脉输入生理盐水为移植对照组.移植4周后检测移植组和移植对照组大鼠肝组织胶原Ⅲ(collagen Ⅲ,COL Ⅲ)、平滑肌动蛋白(smooth muscle actin α,α-SMA)和Ki67的表达,检测外周血肝功能和血凝分析.结果 肝硬化模型组大鼠肝脏体积增至正常时的2倍.EPCs移植组大鼠与肝硬化模型组比较,肝组织学活动指数(histological activity index,HAI)(F=75.062,P<0.01),丙氨酸氨基转移酶(alanine aminotransferase,ALT)(F=29.942,P<0.05),门冬氨酸氨基转移酶(aspartate aminotransferase,AST)(F=16.618,P<0.05)和总胆红素(total bilirubin,TBIL)(F=9.911,P<0.05)水平降低,白蛋白(albumin,Alb)(F=4.944,P<0.05)和Ki67(F=45.966,P<0.01)水平升高,纤维化面积(F=25.025,P<0.05)减少,α-SMA(F=7.86,P<0.05)和COL Ⅲ(F=135.787,P<0.01)表达降低;与正常肝脏相比,移植对照组HAI,ALT,AST和TBIL水平升高,Alb和Ki67水平降低,纤维化面积增加,α-SMA和COL Ⅲ表达升高(P<0.05).移植对照组大鼠凝血酶原时间延长,差异有统计学意义(P<0.05). 结论移植EPCs可以促进大鼠肝硬化的肝细胞增生,减轻肝纤维化程度.