Superoxide dismutase activity of normal murine liver, regenerating liver, and H6 hepatoma.

By means of both direct assay and gel electrophoresis, normal A/J mouse liver was shown to possess both Cu-Zn and Mn superoxide dismutase (SD) activity. H6 hepatoma cells contained Cu-Zn SD activity, but no Mn SD activity was detectable. Isolated mitochondria from normal liver contained both forms of the enzyme, but isolated mitochondria from H6 hepatoma cells contained no SD activity. To ascertain whether this loss of Mn SD activity was characteristic of these tumor cells or was simply a property of rapidly dividing cells, SD activity was measured in regenerating liver. Mn SD activity was present in the regenerating liver at all times after surgery. Hence loss of the Mn SD activity seemed to be a characteristic of some tumor cells but not of corresponding rapidly dividing normal cells.     

小鼠肝癌原位移植模型的建立及其研究意义

目的:建立小鼠的肝癌原位移植模型,为研究肝癌的侵袭转移机制奠定基础.方法:体外培养小鼠肝癌细胞株H22细胞并进行传代,调整细胞浓度为1×107/ml,接种到小鼠腹腔,进行腹腔传代培养,腹腔传代3次后,收集腹水瘤细胞,用生理盐水洗涤2遍,调整细胞浓度为2×107/ml,用50 μl的微量注射器无菌开腹直视下向小鼠肝脏左叶注入50 μl的H22细胞,术后常规饲养,14天后人道主义处死小鼠,解剖观察小鼠肝脏肿瘤形成情况,病理组织学方法鉴定肿瘤组织.结果:肉眼可见肿瘤结节形成良好,成瘤率高,病理鉴定为肝癌.结论:成功建立了小鼠的肝癌原位移植模型,为后续研究肝癌的免疫逃逸和侵袭转移提供了实验基础.     

小鼠肝癌H22细胞LDL受体活性的研究

目的 :观察小鼠肝癌H2 2 细胞LDLR活性。方法 :对小鼠肝癌H2 2 细胞和正常肝细胞进行受体结合的Scatchard分析和单点分析。结果 :①H2 2 细胞对LDL的亲和性与正常肝细胞相同 ,但H2 2 细胞Bmax明显增多 ;②H2 2 细胞对LDL的非特异性结合、内移和降解能力与正常肝细胞相同。结论 :小鼠肝癌H2 2 细胞LDLR活性增高 ,其对LDL的内移和降解功能未变。     

Differences in GTPase-activating protein activity between liver tumors and normal liver tissue in mice.

The intrinsic GTPase activity of the cellular protein p21ras is strongly increased by two cytosolic proteins, the GTPase-activating protein (GAP) produced by the neurofibromatosis type 1 gene (NF1-GAP) and a GAP of 120 kDa molecular mass (p120-GAP). The GAP-mediated stimulation of p21ras GTPase activity was measured in cytosol obtained from carcinogen-induced liver tumors and normal liver tissues of mice of two strains, namely C3H/He and C57BL/6J. For this purpose, cytosolic extracts were incubated with recombinant human p21ras complexed to [gamma-32P]GTP and the time-dependent decrease in p21ras bound radioactivity was measured. Liver cytosolic extracts mediated an increase in the GTPase activity of wild-type p21ras. There were great differences between tumor and normal tissues in the maximal velocity (Vmax) and in the apparent Michaelis constant (KM) of the p21ras GTPase reaction. Both Vmax and apparent KM were decreased in the liver tumors. Cytosolic extracts isolated from liver tumors that harbored point mutations in codon 61 of the c-H-ras gene did not differ in their activity from extracts obtained from non-mutated liver tumors. Since both GAP proteins are important cellular regulators of the ras signaling pathway and probably also effectors of p21ras, the observed differences in GAP activity may be of relevance for the tumorigenic process in mouse liver.     

诱导小鼠肝硬化对结肠癌肝转移的抵抗作用

目的:研究诱导小鼠肝硬化对结肠癌肝转移的抵抗作用及其可能的机制。方法:诱导小鼠肝硬化及结肠癌肝转移模型,比较正常肝与肝硬化小鼠肝转移率;免疫组化法测定层粘连蛋白(laminin,LN)、纤维连接蛋白(fibronectin,FN)及E-选择素(E-selectin)在小鼠肝脏的表达。结果:①肝硬化小鼠肝转移率(4/16,25%)低于正常肝小鼠(14/16,87.5%)(P=0.001);②肝硬化后移植瘤小鼠LN、FN表达高于正常肝移植瘤小鼠,E-selectin表达低于正常肝移植瘤小鼠。结论:动物实验证实肝硬化对结肠癌肝转移有抵抗作用;LN、FN及E-selectin在此过程中可能起一定作用。     

Metabolism of nucleic acids in normal, regenerating and neoplastic tissues of the liver]

Under study was the kinetic growth of three hepatomas-22A, 60 and 46, characterized by a various degree of differentiation. There was found a correlation between the degree of differentiation and parameters of the hepatomas growth. For the hepatomas a correlation between the DNA content and rate of growth was observed, for hepatoma 46 a value of the DNA content proved to be near to that of normal liver. Moreover, in a regenerating liver the DNA content is the same as normal, that indicates the difference between normal and neoplastic actively proliferating tissues. The processes of DNA synthesis, studied by thymidine-C14 incorporation, showed a linear correlation with the rate of growth in regenerating, normal and neoplastic tissues. The ratio RNA/DNA in tumors is regularly decreased with respect to normal liver (a linear correlation between RNA/DNA and the maximum rate of growth). In a regenerating liver the ratio is high. Variations in the ratio RNA/DNA reflect changes in the functional activity and related disorders in the metabolism of nucleic acids in hepatomas. There was found a suppressed DNA decomposition in autolysis of homogenates from hepatoma tissues. It is essential to note that in a minimum deviated hepatoma some inhibition of the DNA decomposition is the only observed disorder in the nucleic acids metabolism. Spin-labelled DNA preparations of tissues under study were obtained. The presence of structural differences between separate DNA samples has been demonstrated.     

Characterization of nucleolar antigens of normal rat liver, regenerating rat liver, and Novikoff ascites hepatoma cells following in vitro translation of polyadenylic acid-containing messenger RNA

Nucleolar antigens of normal rat liver, regenerating liver, and Novikoff ascites hepatoma cells were transplanted in vitro from polyadenylic acid-containing messenger RNAs isolated from the respective tissues and immunoprecipitated with specific antinucleolar antibodies and Protein A. By two-dimensional gel electrophoresis of the translation products, five antigens were detected in normal rat liver. The antigens detected in 18-hr regenerating rat liver were the same as those of normal rat liver when immunoprecipitated with the anti-liver nucleolar antibodies. In the Novikoff tumor, 11 antigens were detected with anti-Novikoff nucleolar antibodies. Of these, two were not found in either normal or regenerating liver. Four major antigens were detectable in both Novikoff hepatoma and regenerating liver messenger RNA translation products with anti-Novikoff nucleolar antibodies. Two antigens were found in normal and regenerating liver that were not found in Novikoff hepatoma. In agreement with previous results, these immunoprecipitation analyses provide further evidence that some nucleolar antigens are present in Novikoff hepatoma that are not found in either normal or regenerating rat liver.     

Effects of N-acetoxy-2-acetylaminofluorene and N'-nitro-N-nitrosoguanidine on nuclear DNA synthesis in rat liver.

A system for the study of DNA synthesis in isolated nuclei is described for sham and regenerating rat liver. The system has been characterized with respect to nuclear purity, conditions for optimum incorporation of [5-methyl-3H]thymidine triphosphate, time course of incorporation, product analysis by neutral and alkaline sucrose gradients, and the effect of exogenously added DNA. No difference in the basal level of activity was detected between nuclei prepared from normal or regenerating liver when isolated 24 hr after operation. However, exogenous activated DNA preferentially stimulated [5-methyl-3H]thymidine triphosphate incorporation in nuclei from regenerating liver. Activated DNA caused to react with the carcinogen N-acetoxy-2-acetylaminofluorene was a less effective primer-template in this system and decreased in a dose-dependent fashion the incorporation of [5-methyl-3H]thymidine triphosphate to below basal levels in nuclei from both normal and regenerating liver. The carcinogen N-methyl-N'-nitro-N-nitrosoguanidine had no inhibitory effect when assayed in this fashion.     

重组质粒pIRES-GM-CSF-IL-21的构建及其治疗小鼠肝癌移植瘤的疗效

目的 构建重组质粒plRES-GM-CSF-IL-21,观察其在小鼠体内的抑瘤作用。方法 将体外培养的肝癌H22细胞接种于小鼠肝左叶,成瘤后,将小鼠分为5组,每组10只,分别尾静脉注射重组质粒plRES-GM-CSF-IL-21、plRES-GM-CSF、plRES-IL-21、plRES和PBS,观察各组荷瘤小鼠的肿瘤生长情况以及GM-CSF和IL-21的抗肿瘤效应。采用酶联免疫吸附法检测小鼠血清γ干扰素(lFN-γ)和白细胞介素2(IL-2)的含量。采用四甲基偶氮唑蓝法检测小鼠脾脏自然杀伤细胞(NK)和细胞毒T淋巴细胞(CTL)的活性。结果 H22组、H-22/Neo组、H22/GM-CSF组、H22/IL-21组和H22/GM-CSFIL-21组小鼠的肿瘤重量分别为(1.591±0.280)g、(1.489±0.155)g、(0.603±0.223)g、(0.583±0.290)g和(0.303±0.323)g,H22/GM-CSF-IL-21组、H22/IL-21组和H22/GM-CSF组小鼠的肿瘤重量明显低于H22组和H22/Neo组(P＜0.01),H22/GM-CSF-IL-21组小鼠的肿瘤重量明显低于H22/GM-CSF组和H22/IL-21组(P＜0.05)。与H22/GM-CSF组和H22/IL-21组比较,H22/GM-CSF-IL-21组小鼠血清中IFN-γ和IL-2的浓度显著升高(P＜0.01),而H22组和H22/Neo组小鼠血清中IFN-Y和IL-2的浓度显著降低(P＜0.01)。与H22/GM-CSF组和H22/IL-21组比较,H22/GM-CSF-IL-21组小鼠脾脏CTL和NK细胞的杀伤活性显著增强(P＜0.01),而H22组和H22/Neo组小鼠脾脏CTL和NK细胞的杀伤活性显著降低(P＜0.01)。结论 重组质粒plRES-GM-CSF-IL-21对小鼠肝癌移植瘤具有明显的抗肿瘤效应,并能促进小鼠体内IFN-γ和IL-2的分泌,增强脾脏中NK和CTL细胞的活性,其效果优于单—GM-CSF或II-21基因治疗。     

内源性一氧化氮在5-FU联合L-Arg治疗裸鼠人肝癌移植瘤中的作用

目的:观察内源性一氧化氮(NO)在5-氟尿嘧啶(5-FU)联合L-精氨酸(L-Arg)治疗裸鼠人肝癌移植瘤中的作用。方法:采用BEL-7402细胞株建立裸鼠人肝癌移植瘤模型,分别给裸鼠腹腔注射生理盐水、5-FU和(5-FU L-Arg),观察各组药物对肿瘤的抑制作用,病理学观察移植瘤的坏死程度和范围,TUNEL法检测细胞凋亡,免疫组化法检测诱导型一氧化氮合酶(inducible nitric oxidesyn-thesase,iNOS)、p16和Bax的表达,化学比色法检测iNOS的活性,硝酸还原酶法检测瘤组织内的NO浓度,对免疫组化结果采用BI2000免疫组化图像分析系统进行光密度测定,统计分析采用SPSS11.0软件进行One-Way ANOVA、Bonferroni、等级资料的秩和检验(Kruskal-Wallis H)及Pearson相关分析。结果:5-FU联合L-Arg能明显抑制移植瘤的生长,移植瘤组织的坏死范围明显增大,肿瘤细胞的凋亡指数增加,移植瘤组织内的iNOS表达与活性明显增加,NO含量增高,NO浓度与Bax和p16的表达有明显相关性。结论:5-FU联合L-Arg对裸鼠人肝癌移植瘤有明显的抑制作用,其机制可能与诱导iNOS表达和活性增强、内源性NO生成增加、NO诱导抑癌基因和凋亡相关基因表达有关。     

低密度脂蛋白- 阿克拉霉素复合物抗肿瘤作用的研究

目的：观察低密度脂蛋白－ 阿克拉霉素（ Ｌ Ｄ Ｌ Ａ Ｃ Ｍ） 复合物对小鼠肝癌 Ｈ２２ 细胞的细胞毒作用。方法： Ｍ Ｔ Ｔ 法及蛋白质合成抑制实验。结果： Ｍ Ｔ Ｔ 实验显示,与正常肝细胞组相比, Ｌ Ｄ Ｌ Ａ Ｃ Ｍ 复合物对 Ｈ２２ 细胞有更显著的生长抑制作用。蛋白质合成抑制实验除显示了 Ｌ Ｄ Ｌ Ａ Ｃ Ｍ 复合物的生长抑制作用外,还反映出该复合物作用的饱和性。结论： Ｌ Ｄ Ｌ Ａ Ｃ Ｍ 复合物对小鼠肝癌 Ｈ２２ 细胞有靶向杀伤作用。     

The effect of liver cytosol on hepatic regeneration and tumor growth

This report further evaluates the concept that the interaction of factors that originate within the liver can contribute, regulate or even initiate the actual development of hepatic regeneration after liver cell necrosis or partial hepatectomy. The effect of liver cytosol (100,000 g supernatant), both from intact adult rat liver (NLC) and from adult rat liver remnants that had been regenerating for 24 hours after 70% partial hepatectomy (PH) in posthepatectomy liver regeneration in the rat was studied. The specificity of the growth-controlling properties in liver cytosol was determined using tumor cells. The intraperitoneal administration of NLC after PH resulted in approximately 70-80% inhibition of the peak 3H-DNA specific activity seen in controls at 18 and 24 hours post-PH, with a significant increase in DNA synthesis at 31-40 hours post-PH. The intraperitoneal administration of RLC after PH, augmented the hepatic regenerative response normally produced. Autoradiographic determination of hepatic nuclear labeling confirmed the inhibitory and stimulatory properties of NLC and RLC respectively. Syngeneic NLC or RLC at six and 24 days after subcutaneous tumor inoculation resulted in significant inhibition of tumor growth for both a methylcholanthrene-induced bladder carcinoma (FBCa) and an HTC-hepatoma. The retardation of FBCa growth could be enhanced by administering NLC or RLC every three or seven days. Syngeneic and xenogeneic liver cytosol resulted in dose-dependent inhibition of P815 mastocytoma cell proliferation in vitro. It is apparent from these studies that both stimulatory and inhibitory factors can be extracted from liver tissue that not only influence liver cell regeneration, but also affect tumor growth. Further isolation and characterization of these factors may lead to an understanding of more fundamental problems such as the control of normal and malignant cell growth.     

熊果酸对小鼠H22 移植瘤的抑制作用研究

目的： 观察熊果酸 (ursolic acid,UA)对小鼠H22移植瘤的抑制作用并对其机制进行探讨。方法：将75只昆明种小鼠于左前腋下皮下接种H22肝癌细胞,24 h后随机分为5组,每组15只,模型组、环磷酰胺 (cyclophosphamide,CP)对照组[25 mg/ (kg·d)] 和UA低、中、高剂量药物组[50、100、150 mg/ (kg·d)]。UA各剂量组和CP对照组分别以相应剂量UA和CP灌胃,模型组以等体积花生油灌胃,每天灌胃1次,连续给药2周。末次给药24 h后,称小鼠体质量,摘除眼球取血后处死。无菌条件下完整剥离小鼠肿瘤及肝、肾、脾组织,计算抑瘤率及肝、肾、脾指数;HE染色观察肿瘤组织病理学改变;分离脾淋巴细胞,CCK8法测定小鼠脾淋巴细胞增殖活性;免疫组化法检测肿瘤组织中CD4+、CD8+ T细胞亚群的分布情况,计数CD4+ T细胞和CD8+ T细胞个数。并采用酶联免疫吸附法 (ELISA)检测血清中白细胞介素12 (interleukin-12,IL-12)的浓度。结果：UA各剂量组小鼠H22肿瘤质量增长均较缓慢,其中UA中、高剂量组肿瘤质量明显减小,与模型组比较,差异均具有统计学意义 (P<0.05);UA中、高剂量组抑瘤率分别为30.15%和39.80%。UA各剂量组肝、肾指数与模型组比较差异无统计学意义 (P>0.05),而脾指数则随UA剂量增加逐渐升高,其中,UA中、高剂量组脾指数与模型组比较差异具有统计学意义 (P<0.05)。HE染色病理观察结果显示,模型组肿瘤细胞生长旺盛,细胞体积较大,胞质丰富,少见坏死区域;UA干预组肿瘤组织中瘤细胞生长增殖明显受到抑制,肿瘤细胞数量减少,密度降低,排列稀疏,细胞核固缩深染或破裂,细胞间质较多,核浆比例减少,坏死区域明显增多。CCK8实验结果显示,随着UA剂量增加,脾淋巴细胞增殖活性逐渐升高,其中,UA中、高剂量组脾淋巴细胞增殖活性显著升高,与模型组间的差异具有统计学意义 (P<0.05)。免疫组化结果显示,UA中、高剂量组肿瘤组织中CD4+、CD8+ T细胞个数与模型组比较均明显增加,差异具有统计学意义 (P<0.05)。UA高、中、低剂量组血清中IL-12的含量均较模型组升高,差异均有统计学意义 (P<0.05);UA高剂量组与CP对照组相比亦显著升高 (P<0.05)。 结论：一定剂量UA可抑制H22荷瘤小鼠肿瘤生长,其作用机制可能与UA促进机体免疫器官发育和淋巴细胞增殖,增加CD4+、CD8+ T细胞向肿瘤组织浸润,并提高血清中IL-12等抗肿瘤活性细胞因子的浓度诱导细胞免疫有关。     

Asialoglycoprotein receptors in rat liver nodules

The asialoglycoprotein (ASGP) receptor, binding and internalizingglycoproteins exposing terminal galactose or N-acetylgalactosamineresidues, was compared in normal liver, regenerating liver andliver nodules. The total cellular content of ASGP binding siteswas reduced to 50 and 40% in regenerating liver and liver nodulesrespectively, compared to the level in normal liver. The ASGPreceptor was heterogeneously distributed in subcellular fractions,with the highest enrichment (16-fold) found in a low-densitymembrane fraction (LDMF), enriched in endosomes and Golgi complexmembranes. The subcellular distribution pattern was similarin the three different liver tissues, except that the relativeenrichment in LDMF was less pronounced in regenerating liver(13-fold) and even less in liver nodules (5-fold). Scatchardanalysis of the binding data indicated that the receptor populationsin all liver tissues were homogeneous with dissociation constantsin the range of 0.12-0.47 nM. The difference in ASGP receptorbinding activity was not found to be the result of an increasedoccupancy with endogenous ligand. In vivo endocytosis of [¹²⁵I]asialo-orosomucoid([¹²⁵I]ASOR) showed a reduction in the amount of internalizedligand in liver nodules, well correlated with the reduced numberof binding sites compared to normal liver. However, a slowerthan normal intracelhilar metabolism of internalized ligandin the nodules was indicated. Bifunctional cross-linking experimentsshowed [¹²⁵I)ASOR-receptor complexes of M_r 250 000, 110 000and 85 000 in normal and regenerating liver, whereas in livernodules only the M_r 85 000 was seen. It is concluded that ASGPbinding activity is reduced in regenerating liver and in livernodules. This reduction was manifested predominantly in membranesderived from the Golgi complex, in endocytic vesicles and atthe cell surface. The slower than normal rate of decay of theinternalized ligand in liver nodules is suggested to be theresult of alterations in ligand dissociation and receptor recyclingprocesses. Furthermore, the abscence of high molecular cross-linkable[¹²⁵I]ASOR-receptor complexes in liver nodules may reflect analteration in receptor oligomerization.     

Estrogen receptors in human gastric, hepatocellular, and gallbladder carcinomas and normal liver tissues

Steroid binding assay using the dextran coated charcoal (DCC) method was applied to human tissues including tumors of the digestive organs, and the results were compared with those of enzymeimmunoassay (EIA) and immunocytochemical assay (ICA) with monoclonal antibody against human estrogen receptor of MCF-7 breast cancer cells. Using the DCC method, estrogen receptor activity was detected in 6 of 26 cases (23.1%) with gastric carcinoma, 3 of 16 hepatocellular carcinoma cases (18.8%), 1 of 3 gallbladder carcinoma cases (33.3%), and both of the 2 cases (100%) with normal liver tissue. However, using EIA, no ER activity was detected in any case. Moreover, ER positive cells were not found by immunohistochemical staining in the gastric carcinoma cases or in normal liver tissue, both of which showed ER activity by the DCC method. These results suggest that the estrogen receptor like material exists in cytosol of the human digestive tumors and normal liver tissue, but that the specificity of the antibodies against estrogen receptor molecules in these tumors may be different from that of the breast tumors.     

射频治疗对荷H22肝癌小鼠脾淋巴细胞免疫功能的影响

背景与目的:射频治疗(radiofrequencytherapy)不同于手术之处在于治疗后的肿瘤组织仍留于体内,这种差别对机体细胞免疫功能的影响有待深入研究。本研究目的是评价射频治疗荷H22肝癌小鼠后脾淋巴细胞增殖活性、细胞毒作用及Th1/Th2细胞因子漂移状况的变化。方法:24只BALB/c小鼠随机分为射频治疗组、手术治疗组、荷瘤对照组及正常对照组4组。MTT法检测小鼠脾淋巴细胞增殖活性,双色流式细胞技术检测脾淋巴细胞细胞毒作用,双抗体夹心ELISA法测定脾细胞培养上清中IL-2、IFN-γ、IL-4及IL-10细胞因子浓度,半定量RT-PCR测定脾细胞IL-2、IFN-γ、IL-4和IL-10mRNA相对含量。结果:射频治疗组小鼠脾淋巴细胞增殖活性和细胞毒作用显著高于正常对照组、手术治疗组及荷瘤对照组(P<0.05)。射频治疗组IL-2和IFN-γ浓度及mRNA表达水平显著高于其他3组(P<0.05),IL-4和IL-10浓度及mRNA表达水平显著低于其他3组(P<0.05)。手术治疗组IL-2、IFN-γ、IL-4和IL-10浓度及mRNA表达水平与正常对照组相比无显著性差异(P>0.05)。结论:射频治疗能够促进荷H22肝癌小鼠脾淋巴细胞活化、增殖,增强对同源肿瘤细胞的杀伤作用。射频治疗可以促进Th1类细胞因子IL-2和IFN-γ的表达。     

Suppression of nuclear ADP-ribosyltransferase activity in regenerating rat liver by 5-azacytidine and its relevance to the nuclear methylating activities

A single administration of 5-azacytidine (5-ACR) to partially hepatectomized rats 24 h following operation resulted in a dose-dependent reduction of nuclear ADP-ribosyltransferase (ADPRT) activity in the liver, when assayed after the nuclei were isolated 22 h after injection. No such a suppression by 5-ACR was observed in the liver of intact rats. Cytidine, a known agent which prevents the incorporation of 5-ACR into DNA, abolished the suppression of ADPRT, when it was given in combination with 5-ACR. The 5-ACR suppressed nuclei from regenerating liver showed no decreased DNA methylating activity, as estimated from the rate of radiolabel transfer from [methyl-3H]SAM to the bulk DNA. The methylation of nuclear RNA and protein was markedly reduced. These results suggest that the incorporation of 5-ACR into nucleic acids inactivates chromatin-bound ADPRT without inhibition of DNA methylation.     

Comparison study of the expressions of myristoylated alanine-rich C kinase substrate in hepatocellular carcinoma, liver cirrhosis, chronic hepatitis, and normal liver

The myristoylated alanine-rich C kinase substrate (MARCKS) is a prominent substrate for protein kinase C (PKC) in a variety of cells. The aim of this study was not only to evaluate the expression and localization of MARCKS in various pathological liver tissues, including HCC, but also to analyze the difference in MARCKS expression between hepatitis virus-induced HCC and cirrhosis. The level of MARCKS and its phosphorylated proteins, as well as its localization, were determined using Western blot and/or immunohistochemistry in HCC and other pathological liver tissues. We also analyzed the change of MARCKS localization on the influence of MARCKS phosphorylation in the HLF cancer cell line by phosphorylation study. In addition, the relationship between MARCKS expression and proliferative activity was studied in HCC. In the immunohistochemical study, a very small amount of MARCKS protein was found along the contour of the hepatocellular membrane in normal liver and in cases of chronic hepatitis. MARCKS was up-regulated in liver cirrhosis tissue and was localized in the cytoplasm of hepatocytes. The expression of MARCKS was down-regulated in HCC tissues, as compared with non-tumorous liver cirrhosis tissues from the same patients. Furthermore, MARCKS was serine-phosphorylated in liver cirrhosis and HCC, and phosphorylated MARCKS was detected in a cytosolic fraction of these tissues. In a phosphorylation study using the HLF HCC cell line, MARCKS was displaced from the plasma membrane to the cytosol following the activation of protein kinase C (PKC) by phorbol 12-myristrate 13-acetate (PMA). Furthermore, the activity of cyclin D1 and cyclin E kinases was found to be higher in HCCs with low MARCKS expression than in HCCs with high MARCKS expression. These results suggest that up-regulation of MARCKS might be essential in the generation of cirrhotic nodules through chronic hepatitis from normal liver, and that the phosphorylation and/or down-regulation of MARCKS might play an important role in the development and progression of HCC from liver cirrhosis.     

Transient inhibition of liver regeneration in mice by transforming growth factor-beta 1 encapsulated in liposomes

We determined whether transforming growth factor-beta 1 (TGF-beta 1) encapsulated in phosphatidylcholine and phosphatidylserine liposomes could inhibit the proliferative response of mouse liver subsequent to partial hepatectomy. C57BL/6 mice were hepatectomized (60%) or underwent laparotomy (control). Peak mitotic activity occurred 48 hr after hepatectomy, as did incorporation of [125I]IdUrd into dividing cells of the liver. The number of Kupffer cells in the regenerating liver was constant relative to liver size, and the uptake of circulating liposomes correlated with liver size and the number of Kupffer cells. Liposomes containing saline (control) or TGF-beta 1 were injected i.v. into hepatectomized mice 24 and 36 hr after the operation. Liposome-TGF-beta 1, but not control liposomes, significantly inhibited the uptake of [125I]IdUrd by liver cells when measured 48 hr after hepatectomy. However, by 72 hr after hepatectomy, the uptake of [125I]IdUrd in livers of mice treated with liposome-TGF-beta 1 was similar to or exceeded that found for hepatectomized mice injected with control liposomes. We conclude that liposomes containing TGF-beta 1 can postpone the proliferative response of regenerating mouse liver cells and are therefore suitable carriers for the in vivo use of TGF-beta 1.