Hepatocyte regeneration after partial liver irradiation in rats

Liver has a strong potential for regeneration after physical, biological or chemical injury, but no data have been reported so far on liver regeneration in response to irradiation injury. The present experiment in rats was designed to clarify whether partial liver irradiation could induce and stimulate the unirradiated part of liver to regenerate. The left-half of rat liver was irradiated with a single dose of 25 Gy. Liver tissues from the irradiated and unirradiated liver, and blood sample were collected at different time points after irradiation. Radiation injury was evaluated by serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities, histopathologic features and trichrome stain. The hepatic regeneration was assessed by serum hepatic growth factor (HGF), mitotic index and proliferation cell nuclear antigen (PCNA) immunohistochemical stain. Expression of transforming growth factor-β1 (TGF-β1) by immunohistochemistry assay was also performed. Results showed that 25 Gy of single-dose irradiation produced severe hepatic injury in the irradiated liver, and the unirradiated liver had been stimulated to regenerate, demonstrated by significant increases of serum HGF 30 days after irradiation, and increase of mitotic index and the number of PCNA-positive hepatocytes 60 days after irradiation. TGF-β1 was strongly and uniformly expressed in the irradiated liver 90-day-post-irradiation, and it was also expressed slightly in unirradiated liver region. In summary, partial liver irradiation could stimulate the unirradiated liver to regenerate, and the role of TGF-β1 in hepatic injury and proliferation needs further investigation.     

Changes in the liver of rats at late stages of regeneration

Summary The author carried out a histological study of the regenerating liver of rats at remote periods (2, 5, 11 and 17 months) after the operation.Development of dystrophic processes was observed 17 months after the excision of 2/3 of the organ by Higgins and Anderson's method. The weight of the liver decreased in experimental animals and was much lower than in controls. A large number of binuclear cells with small nuclei appeared; mononuclear hepatic cells decreased in size; proliferation and thickening of reticular fibers was detected by Gomori's staining.(Presented by Active Member of the Akad. Med. Nauk SSSR N. N. Zhukov-Verezhnikov) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 51, No. 4, pp. 114–118, April, 1961     

大鼠下颌下腺切除对肝再生的影响

目的：探讨大鼠下颌下腺切除后对肝再生的影响。方法：将21只雄性SD大鼠分成三组：3只正常大鼠作为对照组(N组)；9只行肝大部切除术(H组)；9只行肝大部切除术和双侧下颌下腺切除术(E十H组)。应用增殖细胞核抗原(PCNA)免疫组化和核仁组成区嗜银蛋白(AgNOR)染色方法，对N组，H组，E H组的肝组织进行检测和计量。结果：术后第1天及第3天E H组的PCNA指数、AgNOR数及残余肝脏重量均明显低于H组。结论：结果表明大鼠下颌下腺切除后，肝细胞增殖延迟，肝再行过程延缓，提示下颌下腺分泌的EGF对肝再生起重要调节作用。     

索拉菲尼对肝硬化大鼠部分肝切除术后肝脏再生的影响

目的: 在大鼠肝硬化模型的基础上行肝脏部分切除(PH),研究索拉菲尼(sorafenib)对大鼠肝脏再生的影响。方法: 使用二乙基亚硝胺(DEN)诱导Wistar大鼠肝硬化,成功建立30只肝硬化大鼠PH模型后,随机分2组,每组15只。术后第1 d开始,分别给予实验组索拉菲尼(30 mg·kg^-1·d^-1)、对照组生理盐水灌胃10 d后处死。留取PH后及实验结束后的血液及肝脏标本,检测2组肝脏再生率(LRR),增殖细胞核抗原(PCNA),生化指标: 丙氨酸转移酶(ALT)和血清白蛋白(ALB)、血清总胆红素(TBIL)和血清直接胆红素(DBIL)的变化,血管生成相关因子:血管内皮生长因子(VEGF)、血管内皮生长因子受体2(VEGFR-2)、血小板源性生长因子受体β(PDGFR-β),以及肝脏微血管密度(MVD)的变化。结果: (1)LRR在实验组及对照组分别为45.43%±3.36%和44.21%±2.77%,无显著差异(P>0.05);(2)免疫组织化学(IHC)没有检测到PCNA;(3)2组的生化指标无显著差异(P>0.05);(4)实验组VEGFR-2和PDGFR-β的表达受到抑制,MVD降低,并且实验组与对照组差异有统计学意义(P<0.01)。结论: 索拉菲尼虽然对肝硬化血管再生相关因子有抑制作用,但是对肝细胞再生和肝功能没有明显影响。     

The effect of functional loading on liver regeneration in rats

Summary Ninety albino rats with the large left liver lobe extirpated were sustained on various diets (control, sparing and choleretic). In rats fed a choleretic diet regencration of the liver was most complete. In one week after the operation the initial relative weight of the liver was already restored, while in the other groups restoration took place only after two weeks.Restoration of the weight of the liver was due to an increase in the number of liver cells.Thus, an enhanced functional load of the liver stimulates its regeneration.Presented by Prof. N. N. Zhukov-Verezhnikov. Active Member of the USSR Academy of Medical Sciences     

大鼠肝硬化与肝再生的基因表达谱比较分析

目的 了解肝硬化(LC)与肝再生(LR)的相关性.方法 取成年SD雄性大鼠24只,每组6只,用CCl4诱导方法 建立大鼠LC模型;将114只SD雄性大鼠随机分为19组,包括9个部分肝切除组,9个假手术组和1个正常对照组,用2/3肝切除手术建立LR模型.用大鼠全基因组表达谱芯片Genome 230 2.0检测大鼠LC发生与大鼠LR中肝组织的基因表达谱,用生物信息学和系统生物学等方法 分析基因表达谱预示的生理活动的异同,并用定量PCR证明芯片检测结果 的可靠性.结果 LC发生中304个基因和LR中948个基因发生了有意义表达变化,其中121个基因为两者共有,183个基因为LC特有,827个基因为LR特有.H-clustering分析表明,LC和LR的生理活动未显示时间上的相关性.K-means聚类分析显示,LC和LR的C1～C5组基因表达趋势相似,C6组相反,但LR的变化更为丰富.应用基因本体论(GO)分类和功能聚类分析发现,在LC和LR中,免疫反应、炎症反应、细胞迁移、细胞黏附等生理活动增强,各种物质代谢活动减弱.其中,LC的刺激反应在C2强于肝再生,在C6弱于肝再生,而DNA修复、细胞增殖、脂类代谢、内环境稳态和氧化应激等均弱于肝再生.结论 LC与LR的基因表达变化和生理活动有共同方面,也有差异之处.     

大鼠肝再生过程中毛细胆管增生的形态计量研究

Morphometric analysis of the liver during regeneration after partial hepatectomy showed that the mean area and number of bile canaliculi increased, while the volume density of microvilli related to the lumina of the canaliculi decreased. These changes were observed 12 hours after the operation, which gradually returned to normal on the third day after operation. It indicated that the decrease of volume density of microvilli was the result of enlargement of bile canaliculi, which was different from the decrease of microvilli due to passive widening of the canaliculi such as in cholestasis from loss of microvilli. All these phenomena suggest that enlargement of the area of bile canaliculi during liver regeneration may be due to the increase of membranous protein synthesis. Moreover, the increase of the number of bile canaliculi suggests that the proliferation of hepatocytes is associated with formation of new branches of bile canaliculi.     

肝脏再生医学的研究进展

基于机体本身具有的组织自然愈合能力的再生医学(regenerative medicine)研究,近年来在皮肤、血管、肺和肝等组织的研究领域取得了飞速发展。通过在培养液中加入3T3条件培养基和人血清的方法成功地获得了体外人肝细胞的增殖培养。研究表明,小型肝细胞较实质肝细胞有很强的增殖能力。这为利用肝细胞进行的肝细胞移植及人工肝脏的临床应用开辟了广阔的前景。     

大鼠肝大部切除后肝再生过程中肝小叶结构的重建

目的:观察正常大鼠肝大部切除后肝再生过程中肝小叶大小的动态变化,有助于探明大鼠肝再生过程中肝小叶结构的重建过程,并提供组织学依据.方法:正常雄性Wistar大鼠随机分为正常对照组、假手术组和部分肝切除组.采用Higgins和Anderson方法行大鼠70％肝切除,术中即刻称取切除肝叶的湿重.分别于术后12 h、24h、72 h、120 h、1周、2周处死大鼠,留取全部肝组织称湿重,统一留取肝右叶肝组织中性甲醛固定,石蜡包埋连续切片进行H-E染色,显微镜下观察再生肝肝小叶大小变化.结果:正常大鼠部分肝切除后,12h可观察到肝小叶面积开始增大,120 h达高峰,从1周开始可见汇管区门静脉终末支的增大并与邻近中央静脉相联系,肝小叶结构逐步重建,2周时肝小叶面积接近正常,小叶数目增加.正常对照组和假手术组大鼠未见上述动态变化.结论:正常大鼠部分肝切除后,肝再生通过早期肝小叶面积增大,后期肝小叶数目增多而恢复.     

Facilitation of liver regeneration after partial hepatectomy by ventromedial hypothalamic lesions in rats

Whether or not the hypothalamus is involved in initiating hepatic DNA synthesis after partial hepatectomy is unclear. To determine the role of the ventromedial hypothalamic nuclei in liver regeneration after partial hepatectomy, we studied hepatic DNA synthesis during liver regeneration in rats with bilateral lesions of these nuclei. Lesions of the ventromedial hypothalamus accelerated the increase in hepatic DNA synthesis and raised the peak level of thymidine incorporation after partial hepatectomy. These effects of hypothalamic lesions were completely inhibited by hepatic vagotomy. Thus, lesions of the ventromedial hypothalamus appear to promote hepatic regeneration by increasing vagal stimulation of the liver.     

The effect of preconditioning on liver regeneration after hepatic resection in cirrhotic rats

Background/Aims

Ischemic preconditioning (IP) decreases severity of liver necrosis and has anti-apoptotic effects in previous studies using liver regeneration in normal rats. This study assessed the effect of IP on liver regeneration after hepatic resection in cirrhotic rats.

Methods

To induce liver cirrhosis, thioacetamide (300 mg/kg) was injected intraperitoneally into Sprague-Dawley rats twice per week for 16 weeks. Animals were divided into four groups: non-clamping (NC), total clamping (TC), IP, and intermittent clamping (IC). Ischemic injury was induced by clamping the left portal pedicle including the portal vein and hepatic artery. Liver enzymes alanine transaminase (ALT) and aspartate aminotransferase (AST) were measured to assess liver damage. Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) staining for apoptosis and proliferating cell nuclear antigen (PCNA) staining for cell replication were also performed.

Results

Day-1 ALT and AST were highest in IP, however, levels in NC and IC were comparably low on days 1-7. There was no significant correlation of AST or ALT with experimental groups (P=0.615 and P=0.186). On TUNEL, numbers of apoptotic cells at 100× magnification (cells/field) were 31.8±24.2 in NC, 69.0±72.3 in TC, 80.2±63.1 in IP, and 21.2±20.8 in IC (P<0.05). When regeneration capacity was assessed by PCNA staining, PCNA-positive cells (cells/field) at 400× were 3.4±6.0 in NC, 16.9±69 in TC, 17.0±7.8 in IP and 7.4±7.6 in IC (P<0.05).

Conclusions

Although regeneration capacity in IP is higher than IC, the liver is vulnerable to ischemic damage in cirrhotic rats. Careful consideration is needed in applying IP in the clinical setting.     

法尼酯X受体在肝硬化大鼠肝切除术后肝再生中的作用

目的 探讨法尼酯X受体(FXR)在肝硬化大鼠肝切除术后肝再生中的作用.方法 30只肝硬化大鼠肝切除术后按完全随机法分为对照组、胆酸组和消胆胺组饲养1周,每组10只.对照组给以标准饲料饲养,胆酸组给以含0.2%胆酸的饲料饲养,消胆胺组以含2%消胆胺的饲料饲养.1周后检测各组大鼠胆汁分泌速度及总胆汁酸(TBA)含量,检测肝功能、有丝分裂指数(MI)、增殖细胞核抗原(PCNA)标记指数和细胞核DNA含量,并应用RT-PCR和Western免疫印迹分别检测肝组织FXR、胆固醇7α羟化酶(CYP7A1)两者mRNA和蛋白的表达.结果 胆酸组胆汁分泌速度、TBA含量、血清白蛋白(ALB)含量、MI、PCNA标记指数、细胞核DNA含量均高于对照组(P<0.05),血清谷草转氨酶(AST)、谷丙转氨酶(ALT)均低于对照组(P<0.05).而消胆胺组胆汁分泌速度、胆汁TBA含量、血清ALB含量、MI、PCNA标记指数、细胞核DNA含量均低于对照组和胆酸组(P<0.05),血清谷草转氨酶(AST)、谷丙转氨酶(ALT)均高于对照组和胆酸组(P<0.05).胆酸组肝组织FXR mRNA和蛋白表达均高于对照组(mRNA:0.671±0.027比0.528±0.017,蛋白:0.702±0.039比0.566±0.020,P<0.05).肝组织CYP7A1mRNA和蛋白表达含量均低于对照组(mRNA:0.237±0.020比0.325±0.076,蛋白:0.264±0.015比0.325±0.084,P<0.05).而消胆胺组肝组织FXR mRNA(0.468±0.023)和蛋白表达(0.502±0.021)均低于对照组和胆酸组(P<0.05),肝组织CYP7A1 mRNA(0.411±0.021)和蛋白表达(0.476±0.018)则高于对照组和胆酸组(P<0.05).结论 胆汁酸与其受体FXR结合激活细胞内信号转导对肝硬化大鼠肝切除术后肝再生起重要作用,升高胆汁酸水平促进肝再生,降低胆汁酸水平则抑制肝再生.     

Effect of cytidine and uridine on regeneration of the liver in rats poisoned with carbon tetrachloride

The effect of uridine and cytidine on the course of repair processes in the liver of rats with experimental hepatitis due to CCl₄ was studied. Injection of uridine or cytidine simultaneously with CCl₄ over a period of 7 days did not prevent damage to the liver by the poison. Further treatment with the nucleosides (up to 15 and 20 days) accelerated, although to different degrees, the course of repair processes after discontinuation of CCl₄. Cytidine, for instance, caused marked hypertrophy of regenerating hepatocytes, combined with proliferation of mesenchymal cells, which, however, was not accompanied by restoration of the conjugating and excretory functions of the liver. Unlike cytidine, uridine led to more rapid normalization of the abovementioned functions, although restoration of the structure of the organ in this case was less complete.Metabolic Regulation Division, Academy of Sciences of the Belorussian SSR, Medical Institute, Grodno. (Presented by Academician of the Academy of Medical Sciences of the USSR A. P. Avtsyn.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 88, No. 12, pp. 722–723, December, 1979.     

Experimental models of hepatectomy and liver regeneration using newborn and weaning rats

OBJECTIVES: Liver regeneration is a complex process that has not been completely elucidated. The model most frequently used to study this phenomenon is 70% hepatectomy in adult rats; however, no papers have examined this effect in developing animals. The aims of the present study were: 1) to standardize two models of partial hepatectomy and liver regeneration in newborn suckling and weaning rats, and 2) to study the evolution of remnant liver weight and histological changes of hepatic parenchyma on the days that follow partial hepatectomy. METHODS: Fifty newborn and forty-four weaning rats underwent 70% hepatectomy. After a midline incision, compression on both sides of the upper abdomen was performed to exteriorize the right medial, left medial and left lateral hepatic lobes, which were tied inferiorly and resected en bloc. The animals were sacrificed on days 0 (just after hepatectomy), 1, 2, 3, 4 and 7 after the operation. Body and liver weight were determined, and hepatic parenchyma was submitted to histological analysis. RESULTS: Mortality rates of the newborn and weaning groups were 30% and 0%, respectively. There was a significant decrease in liver mass soon after partial hepatectomy, which completely recovered on the seventh day in both groups. Newborn rat regenerating liver showed marked steatosis on the second day. In the weaning rat liver, mitotic figures were observed earlier, and their amount was greater than in the newborn. CONCLUSIONS: Suckling and weaning rat models of partial hepatectomy are feasible and can be used for studies of liver regeneration. Although similar, the process of hepatic regeneration in developing animals is different from adults.     

Intracellular distribution of hyaluronidase in acute and chronic hepatitis and reparative regeneration of the liver in rats

The dynamics of the subcellular distribution of hyaluronidase was studied in the rat liver after acute and chronic hepatitis induced by CCl₄. In the early period of recovery an increase was found in the relative specific activity of the enzyme in the supernatant fraction but a decrease in the fraction of light mitochondria. Later this index increased in the fraction of heavy and light mitochondria, evidence of active processes of phagocytosis and digestion. After acute hepatitis the process of recovery at the subcellular level lasted much longer than at the morphological level.Central Research Laboratory, Novosibirsk Medical Institute. (Presented by Academician of the Academy of Medical Sciences of the USSR D. D. Yablokov.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 81, No. 3, pp. 295–297, March, 1976.