肝脏星形细胞的生物学特性研究进展

肝脏星形细胞(HSCs)是肝脏的非实质细胞之一,具有重要的生理功能,在病理情况下,HSCs被激活,具有许多重要的生物学特性,在肝纤维化与门静脉高压的形成中发挥重要作用.对影响HSCs激活的因素、HSCs激活及凋亡机制的研究成为近年来肝脏细胞学研究的热点之一,并取得了很大进展.     

肝窦内皮细胞生理功能及病理过程的分子机制

肝窦内皮细胞（liver sinusoidal endothelial cell,LSEC）是肝非实质细胞的主要细胞群,具有物质转运、吞噬、抗原提呈、免疫耐受等功能. 肝在遭到多种病原侵袭时,肝窦内皮细胞窗孔逐渐减少或消失,内皮下基膜形成,产生类似于连续型毛细血管的结构,这一过程称为肝窦毛细血管化. 它由多种因素引起,其过程极复杂,在多种肝病的发病前期阶段均有出现,近年来受到广泛关注. 而目前关于肝窦内皮细胞的生理功能及病理机制研究方面的系统总结仍少有报道. 本文对肝窦内皮细胞的生理功能及肝窦病理机制作一较为全面的综述. 除了阐述肝窦毛细血管化自身分子机制的研究进展外,还重点介绍了肝窦毛细血管化参与肝多种疾病发病过程的作用机制. 此外,对肝窦内皮细胞相关的研究方法也作了详细的介绍,为全面了解肝窦内皮细胞生理功能及肝窦毛细血管化的分子机理提供参考.     

肝星形细胞在肝纤维化发生及治疗中的作用

肝纤维化是指在修复肝脏损害和炎症的过程中,肝脏细胞外基质过度增多和过度沉积的病理过程。目前认为,肝星形细胞在肝纤维化形成的过程中起着非常重要的作用。细胞外基质主要来自肝星形细胞,肝实质中降解细胞外基质的基质金属蛋白酶也来自肝星形细胞。肝星形细胞已成为肝纤维化治疗的新靶点。     

肝脏细胞分化和成熟的分子调控机制

肝脏是重要的代谢调控和药物解毒器官,执行体内多种生理功能。肝脏疾病已经越来越严重地影响着人体健康和生存质量。考虑到临床研究和转化医学的迫切需求,人们必须深入研究肝脏内各种细胞特别是肝实质细胞和胆管细胞的分化成熟过程及分子调控机制。该文概述了肝脏内起源于内胚层的肝实质细胞和胆管分化成熟的发育过程,总结了调控此过程的信号通路和转录因子,并简要介绍了最新技术对于肝脏发育研究的推动作用。这些结果对于人们在体外高效地诱导得到或建立更成熟、结构功能更完善的肝脏样细胞或肝脏类器官以及肝脏疾病的研究与治疗有重要意义。     

小胶质细胞在脑中的生理功能

小胶质细胞是脑中的巨噬细胞,也是脑实质中唯一的一种免疫细胞,因而被看作是中枢神经系统抵御病原入侵的第一道防线。在其他非感染病理状态下,如脑损伤及神经退行性疾病等,小胶质细胞也发挥着保护和毒性损伤的双重作用。相比较其病理功能,人们对小胶质细胞的生理功能长期以来很少关注。然而,近几年关于小胶质细胞生理功能的研究在多个方面都有突破。这些研究结果揭示,小胶质细胞在发育的神经系统中起着调控神经元存活和修饰突触的作用,并且在成熟的健康脑中具有探测和调控神经元活动的功能。将着重对近几年关于小胶质细胞生理功能的相关研究做一综述。     

肝纤维化发生发展及逆转过程中肝巨噬细胞亚群分类

肝纤维化是由持续性损伤修复反应引起的,导致肝组织内细胞外基质异常沉积,进一步引发肝脏结构和肝功能异常改变的一种病理过程.已有大量研究表明,肝纤维化在去除损伤因素后是可以逆转的.肝星状细胞作为主要的效应细胞,合成和分泌各种胶原和细胞外基质,一直被认为是肝纤维化发生发展的中心环节.最近的研究发现,巨噬细胞作为主要的调节细胞,能同时调节肝星状细胞的功能和基质胶原的降解,促进肝纤维化的形成.而在肝纤维化逆转过程中,促使活化的肝星状细胞凋亡和纤维胶原的降解,促进肝纤维化的逆转.目前已有研究表明,巨噬细胞亚群在肝纤维化发生发展及逆转中具有双向调控作用,但是对于动物模型体内还没有系统的研究巨噬细胞亚群的分类.本文对巨噬细胞亚群的分类研究做一个全面的综述,对肝脏巨噬细胞在肝纤维化中分子机制的进一步研究具有一定的参考价值和借鉴意义.     

肝细胞极化的结构功能特征及在肝脏疾病时的病理表现

肝细胞极化的形成和维持是肝细胞发挥正常功能的保证。与简单极化上皮细胞不同,肝细胞在肝脏血管与胆小管间形成多个极化膜面,并由紧密连接分隔。极化肝细胞膜及细胞内骨架结构与功能复杂并有序,其分子组成及物质转运机制近年来已被逐渐认识。由于肝细胞极化与肝脏生理功能及多种肝脏疾病时的病理改变有着密切关系,该文就目前肝细胞极化分子和细胞水平研究现状进行综述,并探讨此领域研究发展方向。     

肝星状细胞中表皮形态发生素表达调控机制及其作用研究

肝星状细胞是肝脏中重要的间质细胞,是肝细胞外基质的主要来源.表皮形态发生素(epimorphin、EPM、syntaxin2)在肝脏发育、再生及癌变过程中发挥了重要的作用,目前其表达变化的调控机制及对肝星状细胞的作用还未有报道.通过对肝组织标本进行检测,发现肝纤维化过程中肝星状细胞表达EPM上调.从表观遗传学的角度对EPM表达变化调控机制进行研究,发现DNA去甲基化促进了EPM的表达.为了研究EPM对肝星状细胞的可能的调节作用,将EPM表达质粒转染肝星状细胞,之后检测了EPM对肝星状细胞增殖及迁移能力的变化.结果证明EPM能够促进肝星状细胞的增殖与迁移.本研究发现,激活的肝星状细胞高表达EPM可能是由于DNA去甲基化引起的,同时,高表达的EPM能够促进肝星状细胞的增殖与迁移,进而促进肝纤维化进展.     

细胞相容性物质的生理功能及其作用机制

以生物界广泛存在的细胞相容性物质甜菜碱为代表,介绍了其生理功能的研究进展,同时对植物中其它细胞相容性物质的一些生理功能,以及这些相容性物质在保护生物大分子结构与功能中的可能作用机制也作了评述。     

Stimulation of growth of primary cultured adult rat hepatocytes without growth factors by coculture with nonparenchymal liver cells

DNA synthesis of adult rat parenchymal hepatocytes alone in primary culture can be stimulated only by the addition of humoral growth factors to the culture medium. However, when parenchymal hepatocytes were cocultured with nonparenchymal liver cells from adult rats, their DNA synthesis was markedly stimulated in the absence of added growth factors or calf serum. DNA synthesis of parenchymal hepatocytes was not stimulated by conditioned medium from nonparenchymal liver cells and was greatest when the parenchymal cells were plated on 24-h cultures of nonparenchymal liver cells. A dead feeder layer of nonparenchymal cells was almost as effective as a feeder layer of viable nonparenchymal cells. These results suggest that the stimulation of DNA synthesis in parenchymal hepatocytes was not due to some soluble factors secreted by nonparenchymal liver cells but to an insoluble material(s) produced by the nonparenchymal liver cells. This insoluble material(s) was collagenase- and acid-sensitive, suggesting that it was a protein containing collagen. The effect of nonparenchymal liver cells was specific: coculture with hepatoma cells, liver epithelial cells, or Swiss 3T3 cells did not stimulate DNA synthesis in parenchymal hepatocytes. Added insulin and epidermal growth factor showed additive effects with nonparenchymal cells in the cocultures. These results suggest that DNA synthesis in parenchymal hepatocytes is stimulated not only by various humoral growth factors but also by cell-cell interaction between parenchymal and nonparenchymal hepatocytes, possibly endothelial cells. This cell-cell interaction may be important in repair of liver damage and liver regeneration.     

Uptake and 25-hydroxylation of vitamin D3 by isolated rat liver cells

The physiological roles played by hepatocytes and nonparenchymal cells of rat liver in the metabolism of vitamin D3 have been investigated. Tritium-labeled vitamin D3 dissolved in ethanol was administered intravenously to two rats. Isolation of the liver cells 30 and 70 min after the injection showed that vitamin D3 had been taken up both by the hepatocytes and by the nonparenchymal liver cells. The relative proportion of vitamin D3 that accumulated in the nonparenchymal cells increased with time. Perfusion of the isolated rat liver with [3H] vitamin D3 added to the perfusate confirmed the ability of both cell types to efficiently take up vitamin D3 from the circulation. By a method based on high pressure liquid chromatography and isotope dilution-mass fragmentography it was found that isolated liver cells in suspension had a considerable capacity to take up vitamin D3 from the medium. About 2.5 fmol of vitamin D3 were found to be associated with each hepatocyte or nonparenchymal cell after 1 h of incubation. 25-Hydroxylation in vitro was found to be carried out only by the hepatocytes. The rate of hydroxylation was about the same whether the cells were isolated from normal or rachitic rats (3.5 and 4 pmol of 25-hydroxyvitamin D3 formed per h per 10(6) cells, respectively). The possibility that the nonparenchymal cells might serve as a storage site for vitamin D3 in the liver is discussed.     

Effect of cell-cell interactions in preservation of cellular phenotype: cocultivation of hepatocytes and nonparenchymal cells.

Heterotypic cell interaction between parenchymal cells and nonparenchymal neighbors has been reported to modulate cell growth, migration, and/or differentiation. In both the developing and adult liver, cell-cell interactions are imperative for coordinated organ function. In vitro, cocultivation of hepatocytes and nonparenchymal cells has been used to preserve and modulate the hepatocyte phenotype. We summarize previous studies in this area as well as recent advances in microfabrication that have allowed for more precise control over cell-cell interactions through 'cellular patterning' or 'micropatterning'. Although the precise mechanisms by which nonparenchymal cells modulate the hepatocyte phenotype remain unelucidated, some new insights on the modes of cell signaling, the extent of cell-cell interaction, and the ratio of cell populations are noted. Proposed clinical applications of hepatocyte cocultures, typically extracorporeal bioartificial liver support systems, are reviewed in the context of these new findings. Continued advances in microfabrication and cell culture will allow further study of the role of cell communication in physiological and pathophysiological processes as well as in the development of functional tissue constructs for medical applications.     

Distribution and some properties of NADPH and NADH oxidase in parenchymal and nonparenchymal liver cells

The activities of NADPH and NADH oxidase were determined in homogenates of isolated pure parenchymal and nonparenchymal rat liver cells at neutral (7.4) and acid (5.5) pH. The NADPH oxidase at pH 7.4 is about equally active in parenchymal and nonparenchymal cells and in both cell types is rather insensitive to KCN (1 mm) inhibition. By lowering the pH to 5.5, the NADPH oxidase of the nonparenchymal cells is stimulated (twofold) while the activity in parenchymal cells is decreased. The NADH consumption at neutral pH in parenchymal cells is 75% inhibited by KCN, while this activity in nonparenchymal cells is relatively insensitive to KCN. The NADH oxidase in both parenchymal and nonparenchymal liver cells is less active when the pH is lowered from 7.4 to 5.5. The distribution of NAD(P)H oxidases between parenchymal and nonparenchymal liver cells and the effect of pH on their activities suggest that in the nonparenchymal cells, the NADPH oxidase might play a role in the synthesis of H₂O₂ within the phagocytic vacuole. A scheme is proposed which describes the metabolic events involved in H₂O₂ formation and catabolism of endo(phago)cytosed particles in nonparenchymal liver cells.     

Uptake of yeast invertase by rat liver cells in vivo and in vitro.

Yeast invertase injected intravenously in rats is rapidly taken up by the liver, reaching levels in that organ of 20% or more of the injected dose in about 12 h. At early time points, the bulk of the liver invertase appears in the sedimentable homogenates but, with time, there is a progressive increase in the fraction in the soluble phase, which remains at a constant proportion as the total hepatic invertase declines. The uptake of polyvinylpyrrolidone by the liver is much slower, as is its redistribution to the soluble fraction of homogenates. Separation of cell types from livers containing the markers revealed that the invertase was almost exclusively in the nonparenchymal cell population, while polyvinylpyrrolidone was distributed relatively indiscriminately between parenchymal and nonparenchymal cells. Measurements of uptake of invertase by liver cell preparations in vitro confirmed that nonparenchymal cells were much more active than parenchymal cells in this regard. Furthermore, the process was saturable with the former cell types and inhibitable by α-methylmannoside. Thus, it may be concluded that the uptake of invertase is via fluid pinocytosis in parenchymal cells and adsorptive pinocytosis in the nonparenchymal cells.     

Some kinetic properties of Mucor rouxii phosphofructokinase. Effect of cyclic adenosine 3', 5'-monophosphate.

Intact and pure parenchymal and nonparenchymal cells were isolated from rat liver. The activities of Superoxide dismutase in these cell types were determined by two different methods. With both methods the specific activity of this enzyme is 1.5 times higher in parenchymal than in nonparenchymal liver cells. It can be calculated that about 7% of the total rat liver Superoxide dismutase activity is localized in the nonparenchymal liver cells. Electrophoresis on polyacrylamide gels indicates that the isolated parenchymal cells contain both cytosolic and mitochondrial isoenzymes, whereas with nonparenchymal cells only the cytosolic enzyme could be detected. The mitochondrial band observed in isolated parenchymal cells is absent in the original total liver homogenate. This isoenzyme seems to be activated during the parenchymal cell isolation procedure. Isoelectrofocusing indicates that the cytosolic Superoxide dismutase consists in four different isoelectric forms in both parenchymal and nonparenchymal cells. With the mitochondrial isoenzyme two bands are obtained. The possibility that O₂^? is an important intermediate in H₂O₂ formation in nonparenchymal liver cells is discussed. In this respect, Superoxide dismutase might not only protect the cell against a toxic reagent as O₂^t-, but might also help to regulate the level of the important antimicrobial agent, H₂O₂.     

Evidence that the nonparenchymal cells of the liver are the principal source of cholesteryl ester transfer protein in primates

Previous studies showed that 90% or more of the cholesteryl ester transfer protein (CETP) mRNA is contained in the liver of cynomolgus monkeys. The purpose of this study was to determine if the parenchymal cells (hepatocytes) were the hepatic cell type that contained that mRNA. The parenchymal and nonparenchymal cells were separated by standard methods, and the CETP, apoA-I, apoB, and apoE mRNA content of the preparation determined at each step in the purification process. ApoA-I and apoB are produced only in the parenchymal cells; apoE is produced by both cell types. The mRNA measurements showed that the CETP mRNA: apoA-I mRNA and the CETP mRNA: apoB mRNA ratios were more than 2500-fold greater in the nonparenchymal cell preparation than in the starting material, and that the purified parenchymal cell fraction was virtually devoid of CETP mRNA. In situ hybridization studies showed that, whereas the apoA-I mRNA signal was evenly distributed over the tissue section, the CETP mRNA signal was associated with the hepatic sinusoids, suggesting that it was the hepatic sinusoidal cells that were principally responsible for the high CETP mRNA levels in the liver. We conclude that the nonparenchymal cells are the principal source of CETP in the cynomolgus monkey.     

Cell of origin of distinct cultured rat liver epithelial cells, as typed by cytokeratin and surface component selective expression

The cell of origin of the nonparenchymal epithelioid cells that emerge in liver cell cultures is unknown. Cultures of rat hepatocytes and several types of nonparenchymal cells obtained by selective tissue dispersion procedures were typed with monoclonal antibodies to rat liver cytokeratin and vimentin, polyvalent antibodies to cow hoof cytokeratins and porcine lens vimentin, and monoclonal antibodies to surface membrane components of ductular oval cells and hepatocytes. Immunoblot analysis revealed that, in cultured rat liver nonparenchymal epithelial cells, the anti-rat hepatocyte cytokeratin antibody recognized a cytokeratin of relative mass (Mr) 55,000 and the anti-cow hoof cytokeratin antibody reacted with a cytokeratin of Mr 52,000, while the anti-vimentin antibodies detected vimentin in both cultured rat fibroblasts and nonparenchymal epithelial cells. Analyses on the specificity of anti-cytokeratin and anti-vimentin antibodies toward the various cellular structures of liver by double immunofluorescence staining of frozen tissue sections revealed unique reactivity patterns. For example, hepatocytes were only stained with anti-Mr 55,000 cytokeratin antibody, while the sinusoidal cells reacted only with the anti-vimentin antibodies. In contrast, epithelial cells of the bile ductular structures and mesothelial cells of the Glisson capsula reacted with all the anti-cytokeratin and anti-vimentin antibodies. It should be stressed, however, that the reaction of the anti-vimentin antibodies on bile ductular cells was weak. The same analysis on tissue sections using the anti-ductular oval cell antibody revealed that it reacted with bile duct structures but not with the Glisson capsula. The anti-hepatocyte antibody reacted only with the parenchymal cells. The differential reactivity of the anti-cytokeratin and anti-vimentin antibodies with the various liver cell compartments was confirmed in primary cultures of hepatocytes, sinusoidal cells, and bile ductular cells, indicating that the present panel of antibodies to intermediate filament constituants allowed a clear-cut distinction between cultured nonparenchymal epithelial cells, hepatocytes, and sinusoidal cells. Indirect immunofluorescence microscopy on nonfixed and paraformaldehyde-fixed cultured hepatocytes and bile ductular cells further confirmed that both anti-hepatocyte and anti-ductular oval cell antibodies recognized surface-exposed components on the respective cell types.(ABSTRACT TRUNCATED AT 400 WORDS)     

Uptake of beta-hexosaminidase by nonparenchymal liver cells and peritoneal macrophages

The uptake of beta-hexosaminidase (EC 3.2.1.30) in nonparenchymal liver cells (i.e. endothelial and Kupffer's cells) and peritoneal macrophages has been determined by an enzymatic assay. A considerable uptake was noted in nonparenchymal liver cells, whereas no measurable uptake was seen in peritoneal macrophages. The endothelial cells were more active in the uptake of beta-hexosaminidase than were the Kupffer's cells. The uptake of beta-hexosaminidase by nonparenchymal liver cells showed saturation kinetics and was competitively inhibited by mannan. These findings support the concept that a cell surface receptor on nonparenchymal liver cells mediates uptake of beta-hexosaminidase and suggests a difference in the receptor mechanisms on liver and peritoneal macrophages.     

Altered distribution of hexokinase and glucokinase between parenchymal and nonparenchymal cells of rat liver after methapyrilene intoxication

The cell number as well as the hexokinase and glucokinase activity of liver parenchymal and nonparenchymal cells were studied in methapyrilene treated rats. The number of nonparenchymal cells was doubled after treatment with methapyrilene for two weeks while that of hepatocytes remained constant. The hexokinase activity was increased fourfold in the nonparenchymal cell fraction while it was unchanged in the parenchymal cells. The glucokinase activity was decreased in the hepatocytes to one third. Hence, the increased hexokinase activity was due to a proliferation of nonparenchymal cells rather than to a toxic dedifferentiation of hepatocytes.