非酒精性脂肪肝大鼠部分肝切除术后肝再生功能的变化

目的 探讨大鼠非酒精性脂肪肝（NAFLD）部分肝脏切除术后肝再生功能的变化。方法 80只Wistar大鼠，随机分为正常对照组（C组，35只）与NAFLD组（F组，45只），C组给予正常饮食喂养，F组给予高脂饲料喂养。在喂养至第12周时行70％肝切除术，两组动物分别于术后0、1、12、24、36h处死，取出残肝，计算再生肝重比；光镜下计数核分裂肝细胞；透射电镜观察术后肝细胞超微结构的变化；免疫组织化学染色法检测增殖细胞核抗原阳性表达率；半定量逆转录聚合酶链反应检测细胞周期蛋白D1的表达变化。结果 光镜和电镜观察显示F组肝窦狭窄迂曲，细胞质内大量脂滴沉积，细胞核小，细胞器少，能量代谢及细胞增殖均不活跃。F组术后12、24、36h核分裂相计数明显低于C组同时相点（P〈0．01）；F组术后再生肝重比、S期细胞分数及增殖指数也较C组下降，差异有统计学意义（P〈0．01）；F组增殖细胞核抗原阳性率、细胞周期蛋白D1的mRNA表达在术后12、24、36h均明显低于C组同时相点（P〈0．01）。结论 中至重度NAFLD大鼠部分肝切除术后DNA合成高峰滞后，肝再生延迟，再生进程主要被阻滞在细胞周期的G1／S期调控点。     

Liver regeneration in response to partial hepatectomy in rats treated with retrorsine: a kinetic study

BACKGROUND/AIM: We have designed an experimental model in which transplantation of normal hepatocytes into rats previously treated with retrorsine (a naturally-occurring pyrrolizidine alkaloid) results in near-complete replacement of the recipient liver by donor-derived cells. Two/thirds partial hepatectomy was found to be essential for this process to occur. To probe this finding, in the present study we describe the kinetics of liver regeneration in response to partial hepatectomy in rats given retrorsine. METHODS: Six-weeks-old male Fisher 344 rats received retrorsine (2 injections of 30 mg/kg each, i.p., 2 weeks apart), or the vehicle. Four weeks after the last injection, partial hepatectomy was performed and rats were killed at 1, 2, 3, 6, and 15 days thereafter. RESULTS: At time zero, i.e. prior to partial hepatectomy, liver weight and total liver DNA content were significantly lower in retrorsine-treated animals compared to controls (DNA content: 19.2+/-1.7 vs. 25.7+/-1.1 mg/liver). Diffuse megalocytosis (enlarged hepatocytes) was present in the group exposed to retrorsine. By day 3 post-partial hepatectomy liver DNA content in control animals had more than doubled compared to day 1 values (20.2+/-1.5 vs. 8.8+/-1.2), while very little increase was seen in retrorsine-treated rats at the same time points (7.6+/-0.4 vs. 6.1+/-0.2). At 2 weeks after partial hepatectomy, total DNA content returned close to normal levels in the control group (26.9+/-1.0 mg/liver); however, the value was still very low in animals receiving retrorsine (9.1+/-0.7). Data on BrdU labeling were consistent with this pattern and indicated that DNA synthesis following partial hepatectomy was largely inhibited in the retrorsine group. Similarly, no mitotic response was observed in hepatocytes following partial hepatectomy in animals exposed to retrorsine. CONCLUSIONS: These results clearly indicate that retrorsine exerts a strong and persistent cell cycle block on hepatocyte proliferation. Further, these results are in agreement with the hypothesis that selective proliferation of transplanted hepatocytes in retrorsine-treated animals is dependent, at least in part, on the persistent cell cycle block imposed by the alkaloid on endogenous parenchymal cells.     

Gene expression profile in the regenerating rat liver after partial hepatectomy

BACKGROUND/AIMS: When a loss of hepatic mass occurs, the expression of a large number of genes is either induced or altered, accompanying hepatocyte proliferation. In the present study, we made an in-house cDNA microarray containing 4608 elements (Liver chip), and analyzed extensively gene expression profiles of the regenerating liver after 70% partial hepatectomy (PHx) in rats. METHODS: RNAs were prepared from three rat livers at each time point (taken at 0, 6, 12, 18, 24, 48, 72 h, and 1 week after PHx). Using the liver chip, we performed large-scale analysis of gene expression during liver regeneration. Elements either up- or down-regulated more than twofold at one or more time points were selected. RESULTS: Among the 4608, 382 were identified. Using cluster analysis, we found great similarity between gene-expression profiles at 12 and 18 h after PHx as well as between 48 and 72 h after PHx. We also found that there are at least six distinct temporal patterns of gene expression in the regenerating rat liver after PHx. CONCLUSIONS: These results indicated that microarray analysis is a powerful approach for monitoring molecular events in the regenerating liver.     

Immediate onset of DNA synthesis in remnant rat liver after 90% hepatectomy by an administration of follistatin

Background/Aim: Follistatin is an antagonist of activins and is effective in promoting liver regeneration after 70% hepatectomy. To examine its efficacy under more critical conditions, we studied the effect of follistatin on liver regeneration in 90% hepatectomized rat.Methods: Human recombinant follistatin was infused into the portal vein immediately after 90% hepatectomy in 24-h-starved rats, and changes in the liver regeneration rate and nuclear bromodeoxyuridine labeling were measured.Results: In control rats, nuclear labeling was first detected at 11 h of hepatectomy. In follistatin-treated rats, nuclear labeling was markedly increased at 3 h, and was significantly higher than that in control rats at 24, 72, 96, 120 and 144 h. The liver regeneration rate was significantly higher in follistatin-treated rats at 48, 72, 96, 120, 144 and 168 h. To determine the reason for the accelerated growth in starved rats, we compared the expression of mRNA for c-myc, p53, p21^C1P1, p15^INK4B, p27^KIP1, and subunits of activins in fed and starved rats. mRNA for p21^CIP1 and p15^INK4B, but not p27^KIP1 were decreased in 24 h-starved rats compared to the fed rats. mRNA for β_A subunit of activin was not detected in either fed ro 24-h-starved rats, whereas that for β_C subunit was increased in starved rats.Conclusion: These results indicate that follistatin induces immediate onset of DNA synthesis in 90% hepatectomized rats and is quite effective in promoting liver regeneration.     

Liver failure following partial hepatectomy

While major liver resections have become increasingly safe due to better understanding of anatomy and refinement of operative techniques, liver failure following partial hepatectomy still occurs from time to time and remains incompletely understood. Observationally, certain high-risk circumstances exist, namely, massive resection with small liver remnants, preexisting liver disease, and advancing age, where liver failure is more likely to happen. Upon review of available clinical and experimental studies, an interplay of factors such as impaired regeneration, oxidative stress, preferential triggering of apoptotic pathways, decreased oxygen availability, heightened energy-dependent metabolic demands, and energy-consuming inflammatory stimuli work to produce failing hepatocellular functions.     

Liver sinusoidal endothelial and biliary cell repopulation following irradiation and partial hepatectomy

AIM: To investigate whether irradiation (IR) and partial hepatectomy (PH) may prepare the host liver for nonparenchymal cell (NPC) transplantation.METHODS: Livers of dipeptidyl peptidase(DPP)-deficient rats were pre-conditioned with external beam IR (25 Gy) delivered to two-thirds of the right liver lobules followed by a one-third PH of the untreated lob-ule. DPP-positive liver cells (NPC preparations enriched for liver sinusoidal endothelial cells (LSECs) and hepatocytes) were transplanted via the spleen i...     

Acute renal injury after partial hepatectomy

Currently, partial hepatectomy is the treatment of choice for a wide variety of liver and biliary conditions. Among the possible complications of partial hepatectomy, acute kidney injury(AKI) should be considered as an important cause of increased morbidity and postoperative mortality. Difficulties in the data analysis related to postoperative AKI after liver resections are mainly due to the multiplicity of factors to be considered in the surgical patients, moreover, there is no consensus of the exact definition of AKI after liver resection in the literature, which hampers comparison and analysis of the scarce data published on the subject. Despite this multiplicity of risk factors for postoperative AKI after partial hepatectomy, there are main factors that clearly contribute to its occurrence. First factor relates to large blood losses with renal hypoperfusion during the operation, second factor relates to the occurrence of post-hepatectomy liver failure with consequent distributive circulatory changes and hepatorenal syndrome. Eventually, patients can have more than one factor contributing to post-operative AKI, and frequently these combinations of acute insults can be aggravated by sepsis or exposure to nephrotoxic drugs.     

熊脱氧胆酸促进肝脏部分切除后肝细胞再生

目的 探讨熊脱氧胆酸（ｕｒｓｏｄｅｏｘｙｃｈｏｌｉｃ ａｃｉｄ，ＵＤＣＡ）对胆道梗阻肝脏部分切除（ＰＨ）后肝细胞再生的影响。方法Ｗｉｓｔａｒ大鼠随机分为正常７０％肝部分切除组（Ｎ－ＰＨ）、胆道梗阻２周７０％ＰＨ组（ＢＤＯ－ＰＨ）、ＢＤＯ—ＰＨ ＵＤＣＡ治疗组及ＢＤＯ—ＰＨ生理盐水治疗组。观察肝组织学改变，检测７０％ＰＨ后肝细胞ＢｒｄＵ标记、肝内肝细胞生长因子（ＨＧＦ）及其受体Ｍｅｔ ｍＲＮＡ表达。结果 ＵＤＣＡ治疗能促进胆道梗阻后肝功能好转并减轻肝组织学病变；ＵＤＣＡ治疗组大鼠７０％ＰＨ后肝内ＨＧＦ／Ｍｅｔ ｍＲＮＡ高峰表达值均高于ＢＤＯ—ＰＨ组（Ｐ ＜ ０．０５），肝细胞 ＢｒｄＵ高峰标记指数（５９．３９±１０．８２）％高于 ＢＤＯ—ＰＨ组肝细胞 ＢｒｄＵ高峰标记指数（３６．２２±８．３７％（ｔ＝４．１４９，Ｐ＜０．０１），而与Ｎ－ＰＮ组肝细胞ＢｒｄＵ高峰标记指数（６８．６４±１１．２６％）％相比差异无显著性（ｔ＝１．４５１，Ｐ＞０．０５）。结论 ＵＤＣＡ通过缓解胆道梗阻后肝组织损害并上调７０％ＰＨ后肝内ＨＧＦ／Ｍｅｔ ｍＲＮＡ表达，从而促进胆道梗阻肝脏部分切除后肝细胞再生。     

Randomized study of chemoembolization as an adjuvant therapy for primary liver carcinoma after hepatectomy

From April 1990 to December 1993, 140 patients were recruited to a randomized study to evaluate transcatheter hepatic arterial chemoembolization (TACE) as an adjuvant therapy for primary liver carcinoma after hepatectomy. This study investigated the principle, techniques and results of TACE. The results showed that the intrahepatic recurrence rate was 48.9% in the patients who underwent radical resection only, but only 21.3% in the patients who also underwent TACE 3–4 weeks after hepatectomy (P<0.01). The 1-, 2-, 3-, and 4-year survival rates were 72.3%, 52.7%, 35.1%, and 35.1% respectively for the patients who underwent radical resection only, and were 97.9%, 85.5%, 69.5%, and 56.9% for the patients who also underwent TACE 3–4 weeks after radical resectionP<0.001). The 1-, 2-, 3-, and 4-year survival rates were 38.9%, 0%, 0%, and 0% for the patients who underwent palliative resection only, and were 68.3%, 32.3%, 21.5%, and 21.5% respectively for the patients undergoing TACE 3–4 weeks after palliative hepatectomyP<0.001).Abbreviations TACE transcatheter hepatic arterial chemoembolization - PLC primary liver carcinoma     

生长激素对鼠部分肝切除术后肝再生影响

目的　探讨生长激素对 70 %肝切除后肝再生的影响。方法　 60只SD大鼠随机分为对照组及生长激素组 ,按Higgins方法行 70 %肝切除术 ,术后给药并分批于术后 6、2 4、48、72、96h处死 ,作如下比较 :①残肝肝重 ;②增殖细胞核抗原 (PCNA)标记指数 ;③图像定量分析法测量PCNA阳性产物面积及灰度值。结果　与对照组比较 ,生长激素组残肝肝重、PCNA标记指数、PCNA阳性产物面积在术后均显著增高 (P <0 .0 5 ) ,而灰度值则显著降低 (P <0 .0 5 )。结论　生长激素具有强烈促进肝细胞增殖和刺激肝再生的作用     

Interplay of neuropilin-1 and semaphorin 3A after partial hepatectomy in rats

AIM:To elucidate the role of neuropilin-1(Nrp-1) and semaphorin 3A(Sema3A) in sinusoidal remodeling during liver regeneration in rats.METHODS:Male Wistar/ST rats at 7 wk of age,weighing about 200 g,were used for all animal experiments.In vivo,at 24,48,72,96,144 and 192 h after twothirds partial hepatectomy(PHx),the remnant livers were removed.Liver tissues were immunohistochemically stained for Nrp-1,Sema3A and SE-1,a liver sinusoidal endothelial cell(SEC) marker.Total RNA of the liver tissue was extracted and reversely transcribed into cDNA.The mRNA expression of Sema3A was analyzed by quantitative real-time polymerase chain reaction and normalized to that of ribosomal protein S18.In vitro,SECs were isolated from rat liver and cultured in endothelial growth medium containing 20 ng/mL vascular endothelial cell growth factor.Migration of SECs in primary culture was assessed by cell transwell assay with or without recombinant Sema3A.Apoptotic cells were determined by a terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling method.RESULTS:In vitro,immunohistochemistry study revealed that Sema3A and Nrp-1 were constitutively expressed in hepatocytes and SECs,respectively,in normal rat liver tissues.Nrp-1 expression in SECs was quantified by the percentage of immunostained area with antiNrp-1 antibody in relation to the area stained with SE-1.Between 24 h and 96 h following resection of liver,Nrp-1 expression in SECs was transiently increased.Compared with the baseline(5.2% ± 0.1%),Nrp-1 expression in SECs significantly increased at 24 h(17.3% ± 0.7%,P 0.05),48 h(39.1% ± 0.6%,P 0.01),72 h(46.9% ± 4.5%,P 0.01) and 96 h(29.9% ± 3.8%,P 0.01) after PHx,then returned to the basal level at termination of liver regeneration.Interestingly,the expression of Sema3A was inversely associated with that of Nrp-1 in liver after PHx.Sema3A mRNA expression was significantly reduced by about 75% over the period 24-144 h after PHx(P 0.05),and returned to basal levels at 192 h after PHx.In vitro,SECs isolated from rats after PHx(PHx-SECs) were observed to migrate to the lower chamber of the cell transwell system after incubation for 24 h,but not cells from normal rats(CONT-SECs),indicating that mobility of PHx-SECs increases as compared with that of CONT-SECs.Moreover,recombinant Sema3A significantly attenuated migration in PHx-SECs in primary culture(vehicle-treated 100% ± 7.9% vs Sema3A-treated 42.6% ± 5.4%,P 0.01),but not in CONT-SECs.Compared with CONTSECs,the apoptotic rate of PHx-SECs decreased by 78.3%(P 0.05).There was no difference in apoptosis between CONT-SECs that were treated with vehicle and Sema3A.However,in PHx-SECs,apoptosis was induced by the presence of 5 nmol Sema3A for 24 h(vehicle-treated 21.7% ± 7.6% vs Sema3A-treated 104.3% ± 8.9%,P 0.05).In addition,immunohistochemistry confirmed the increased expression of Nrp-1 in PHx-SECs,while it was noted to a lesser extent in CONT-SECs.CONCLUSION:The interplay of Nrp-1 and Sema3A shown in our results may lead to a better understanding of interaction between sinusoidal remodeling and SECs during liver regeneration.     

小鼠部分肝切除术的现状与应用

部分肝切除术创建于20世纪30年代,他为哺乳动物的肝再生和肝疾病等研究提供了重要的实验工具,现有的手术方法主要以大鼠和小鼠为对象.小鼠的部分肝切除手术与大鼠的手术方法有较大的相似性,但因其体质差异不能完全复制大鼠的手术方法.本文结合文献报道和实践操作的经验,除了对部分肝切除术的原理和主要方法进行介绍外,也对小鼠部分肝切...     

Liver regeneration after partial hepatectomy in rats with defective bilirubin conjugation or biliary excretion

The role of conjugated bilirubin in liver regeneration after partial hepatectomy (PH) was studied in Gunn rats, in transport-mutant (TR^–) rats, and in rats with extrahepatic biliary obstruction. Ornithine decarboxylase (ODC) and thymidine kinase (TK) activities in liver homogenates and immunohistochemistry ofin vivo bromodeoxyuridine (BrdU) incorporation in hepatic DNA were followed as regeneration parameters at 24 and 48 hr after PH. The results relative to TK activity and BrdU incorporation were consistent with significantly delayed hepatic DNA synthesis in Gunn rats in comparison to control Wistar and TR^– rats. This delay in DNA synthesis was not reflected in the hepatic ODC activity. After one week of complete common bile duct obstruction (CBO), an increased TK activity and BrdU incorporation was seen. PH following CBO resulted in a further increase in ODC activity and BrdU incorporation. TK activity did not change, however. These data relative to the regulation of hepatic DNA synthesis after PH in Gunn rats and in rats with extrahepatic biliary obstruction suggest a possible stimulatory role for conjugated bilirubin in hepatic regeneration; however, the normal hepatic DNA synthesis in TR^– rats studied before PH and the subnormal DNA synthesis seen 24hr after PH in TR^– rats and in rats with CBO indicate that conjugated bilirubin does not stimulate hepatic DNA synthesis.Presented at the Proceedings of the International Meeting on Normal and Neoplastic Growth in Hepatology, Bari, Italy, June 1989.     

Differentiation of rat bone marrow stem cells in liver after partial hepatectomy

AIM: To investigate the differentiation of rat bone marrow stem cells in liver after partial hepatectomy. METHODS: Bone marrow cells were collected from the tibia of rat with partial hepatectomy, the medial and left hepatic lobes were excised. The bone marrow stem cells (Thy CD3-CD45RA- cells) were enriched from the bone marrow cells by depleting red cells and fluorescence-activated cell sorting. The sorted bone marrow stem cells were labeled by PKH26-GL in vitro and autotransplanted by portal vein injection. After 2 wk, the transplanted bone marrow stem cells in liver were examined by the immunohistochemistry of albumin (hepatocyte-specific marker). RESULTS: The bone marrow stem cells (Thy CD3-CD45RA- cells) accounted for 2.8% of bone marrow cells without red cells. The labeling rate of 10μM PKH26-GL on sorted bone marrow stem cells was about 95%. There were sporadic PKH26-GL-labeled cells among he-patocytes in liver tissue section, and some of the cells expressed albumin. CONCLUSION: Rat bone marrow stem cells can differentiate into hepatocytes in regenerative environment and may participate in liver regeneration after partial hepatectomy.     

白细胞介素-10与减体积大鼠肝移植后肝再生的关系

目的 探讨白细胞介素-10(IL-10)与减体积大鼠肝移植术后移植肝再生的关系。方法 建立减体积大鼠肝移植模型,实验分为:肝切除组、全肝移植组和减体积肝移植组,分别于术后1、2、4、7d取肝组织,免疫组织化学检测各组IL-10的表达,流式细胞仪检测移植肝的增殖活性。结果 肝切除组、全肝移植组和减体积肝移植组肝细胞增生活跃,术后4d增殖高峰分别为26.3±0.9、35.8±2.2、32.4±1.8。IL-10与移植后肝再生呈负相关(r=-0.58,P<0.01)。结论 减体积肝移植和全肝移植术后肝脏具有同样的增殖活性,但增殖峰值较肝切除延迟。IL-10对移植肝肝再生具有明显的调控作用,同时受免疫系统产生的其它细胞因子和激素的影响。     

肝硬化大鼠膈肌功能改变的实验研究

目的研究肝硬化大鼠膈肌损伤的作用机制。方法SD大鼠16只被随机分成对照组(n=8)和实验组(n=8)。实验组大鼠皮下注射40?l4制备肝硬化动物模型,共成模8只。应用体外灌流大鼠膈肌条的方法,分别测量其单收缩力(Pt),最大强直张力(Po),峰值收缩时间(CT),半舒张时间(1/2RT),张力最大上升速率( dT/dtmax),张力最大下降速率(-dT/dtmax),并测量在10、20、40、60、100Hz的张力,绘制力—频率曲线,同时测定膈肌组织中SOD、SDH、NOS、Na -K -ATPase、Ca2 -Mg2 ATPase的活性及MDA含量,并观察膈肌的形态学变化。结果肝硬化大鼠膈肌Pt,Po, dT/dtmax,-dT/dtmax均低于对照组(P<0.01);CT,1/2RT高于对照组(P<0.05);给予10、20、40、60、100Hz的电压刺激膈肌时,肝硬化大鼠膈肌张力明显低于对照组(P<0.05);肝硬化大鼠膈肌组织SOD、SDH、Na -K -ATPase,Ca2 -Mg2 ATPase活性明显低于对照组(P<0.05),而MDA含量和NOS活性显著高于对照组(P<0.05);肝硬化大鼠膈肌肌纤维断裂,溶解。结论肝硬化大鼠膈肌肌纤维结构受到破坏,收缩功能下降,可能与SOD,SDH,Na -K -ATPase,Ca2 -Mg2 ATPase活性降低,NOS活性和MDA含量增高有关。     

Expressed genes in regenerating rat liver after partial hepatectomy

AIM: To reveal the liver regeneration (LR) and its control as well as the occurrence of liver disease and to study the gene expression profiles of 551 genes after partial hepatectomy (PH) in regenerating rat livers. METHODS: Five hundred and fifty-one expressed sequence tags screened by suppression subtractive hybridization were made into an in-house cDNA microarray, and the expressive genes and their expressive profiles in regenerating rat livers were analyzed by microarray and bioinformatics. RESULTS: Three hundred of the analyzed 551 genes were up- or downregulated more than twofolds at one or more time points during LR. Most of the genes were up- or downregulated 2-5 folds, but the highest reached 90 folds of the control. One hundred and thirty-nine of them showed upregulation, 135 displayed downregulation, and up or down expression of 26 genes revealed a dependence on regenerating livers. The genes expressed in 24-h regenerating livers were much more than those in the others. Cluster analysis and generalization analysis showed that there were at least six distinct temporal patterns of gene expression in the. regenerating livers, that is, genes were expressed in the immediate early phase, early phase, intermediate phase, early-late phase, late phase, terminal phase. CONCLUSION: In LR, the number of down-regulated genes was almost similar to that of the upregulated genes; the successively altered genes were more than the rapidly transient genes. The temporal patterns of gene expression were similar 2 and 4 h, 12 and 16 h, 48 and 96 h, 72 and 144 h after PH. Microarray combined with suppressive subtractive hybridization can effectively identify the genes related to LR.