生物人工肝中肝细胞来源及培养的新进展

肝细胞的活性是生物人工肝成功的关键，而良好的细胞来源及培养是肝细胞活性的保证。近年来，生物型人工肝中肝细胞来源及培养有了很大的进展。其中。肝细胞来源的研究主要表现在原代肝细胞，人肝癌细胞，传代及转化的肝细胞等方面，肝细胞培养的研究集中在培养方式，培养条件，培养时间及所需肝细胞数量等方面。这些研究解决了许多理论和实践上的问题，促进了生物人工肝的发展。     

肝细胞凋亡与自身免疫的关系及影响因素

就应答不同刺激而言，细胞凋亡或程序性细胞死亡是一种正常细胞反应。凋亡细胞能在健康成年人生长发育组织中被检测到，例如肝。在许多慢性炎症和自身免疫性疾病中，受到影响的组织中的凋亡细胞数呈现递增。本文旨在探讨肝细胞凋亡与自身免疫的相关性及相关条件，以揭示肝细胞凋亡的机制、肝病防治对肝细胞凋亡的影响。     

细胞角蛋白与肝脏疾病

细胞角蛋白在肝细胞和胆管细胞的不同表型与细胞功能密切相关。通过胚胎肝组织发生过程中角蛋白表型的变化和肿瘤性肝细胞不同成份角蛋白表达的异质性研究，提示成年人肝脏可能贮备有多潜能干细胞。此外，细胞角蛋白作为细胞系标记物，其病理学的改变是肝脏疾病的参考指标。     

肝细胞癌细胞外基质的分布形式及其临床意义

目的：了解细胞外基质在肝细胞癌的分布形式；探讨其与肝细胞癌恶性程度的关系。方法：用５种细胞外基质抗体对４０例肝细胞癌作免疫组化染色。结果：显示４种分布形式：连续梁周腺泡型、断续梁周腺泡周型、血管间质型、胞膜胞浆型。前３型与肿瘤生长方式、细胞分化和增殖密切相关。结论：提示细胞外基质在评价肝细胞癌的恶性程度是有用的标志物。     

肝组织工程种子细胞来源的研究进展

肝脏组织工程是组织工程领域中重要的学科之一，其研究主要采用2条基本途径治疗各种终末期肝病：一是通过细胞体内移植的方法，将种子细胞植入体内修复替代肝组织缺损；二是将种子细胞接种到支架材料上，在体外构建”肝脏组织”，替代肝脏功能。理想的种子细胞是其关键组成部分，包括人或动物的成体肝细胞、各种肝干细胞、永生化肝细胞系等。但目前尚未确定一种理想的细胞来源，临床广泛应用还存在许多问题。就肝组织工程种子细胞来源的研究进展综述如下。     

肝星状细胞研究进展

肝星状细胞(hepatic stellate cells，HSCs)是肝脏的非实质细胞之一，以前也称为贮脂细胞(fat-storing cells)、Ito细胞(Ito cells)、脂细胞(1ipocytes)、间质细胞(interstitial cells)、窦周细胞(peri-sinusoidal cells)或维生素A储存细胞(vitamin A-storing cells)等，其位于Disse间隙内，形态不规则，胞体呈卵圆形，其核周部分包埋在邻近肝细胞的隐窝内，     

细胞角蛋白7和19在肝细胞癌细胞系中的异常表达及其机制

正常肝脏中，肝细胞的中间丝细胞骨架．仅由细胞角蛋白（ｃｙｌｏｋｅｒａｔｉｎ，ＣＫ）８和 ＣＫ１８构成；而胆管上皮细胞除含有这两种ＣＫ外，还表达ＣＫ７和ＣＫ１９。这两种细胞类型的ＣＫ组型，曾被认为能在各种生理、病理状态下保持不变，因此，ＣＫ８和ＣＫ１８被称为“肝细胞型”ＣＫ，ＣＫ７和ＣＫ１９被称为“胆管型”ＣＫ。最近研究表明，肝细胞的ＣＫ组型在某些肝损伤和肝脏疾病状态下会发生一定的改变，肝细胞和肝细胞癌可额外地表达“胆管型”ＣＫ（ＣＫ１９ 和／或ＣＫ７），称为肝细胞ＣＫ的异常表达［１－５］，其机制尚…     

生物人工肝中肝细胞来源及培养的新进展

肝细胞的活性是生物人工肝成功的关键 ,而良好的细胞来源及培养是肝细胞活性的保证。近年来 ,生物型人工肝中肝细胞来源及培养有了很大的进展。其中 ,肝细胞来源的研究主要表现在原代肝细胞、人肝癌细胞、传代及转化的肝细胞等方面。肝细胞培养的研究集中在培养方式、培养条件、培养时间及所需肝细胞数量等方面。这些研究解决了许多理论和实践上的问题 ,促进了生物人工肝的发展     

大鼠睾丸支持细胞对体外共微囊化肝细胞的双重保护作用

目的 观察大鼠睾丸支持细胞(Sertoli cell)对混合共微囊化肝细胞的营养支持和免疫豁免(immune privilege)的双重保护作用.方法 气流法制备含不同比例肝细胞与Sertoli细胞混合的微囊进行体外培养,测定培养上清中白蛋白、尿素的含量,结合形态学判断肝细胞的功能和活性,并选择细胞最佳混合比例.CCK-8法测定Sertoli细胞对肝细胞引起的脾细胞增殖的影响.结果 与纯肝细胞微囊组相比,一定比例的Sertoli细胞可促进共微囊化肝细胞的白蛋白和尿素合成(P＜0.01),肝细胞存活时间也相应延长.其中,肝细胞和Sertoli细胞以20:1混合时即可达最佳效果.在脾细胞增殖反应中,与对照组相比,微囊化肝细胞仍可引起明显的脾细胞增殖(P＜0.01),而Sertoli细胞可显著抑制这一反应.结论 Sertoli细胞与肝细胞混合共微囊化能明显改善囊内肝细胞的形态、功能与寿命,并使后者得到一定的免疫豁免保护作用.     

慢性丙型肝炎时贮脂细胞的定量研究

应用免疫组化等方法，对慢性迁延性丙型肝炎肝组织中呈结蛋白阳性的贮脂细胞进行了定量的研究结果；结蛋白阳性的细胞数量与同阶段的乙型肝炎相比显著减少；电镜观察可见肝窦中有较多的枯否细胞，肝细胞间隙及窦周胶原纤维的沉积。我们认为，在丙型肝炎早期纤维化中，贮脂细胞可能并非主要导致纤维化的效应细胞，这和乙型肝炎有所不同。     

肝癌细胞侵袭行为与其流变特性相关

以原代肝细胞、肝细胞癌（Ｈｅｐａｔｏｃｅｌｌｕｌａｒ ｃａｒｃｉｎｏｍａ,ＨＣＣ）细胞及对人工基底膜Ｍａｔｒｉｇｅｌ具有侵袭能力的ＨＣＣ细胞（ＨＣＣ－Ｉｎｖ细胞）为研究对象,研究了三种细胞的粘弹特性、与胶原蛋白Ⅰ的粘附力学特性及与肝血窦内皮细胞（Ｌｉｖｅｒ ｓｉｏｎｓｏｉｄａｌ ｅｎｄｏｔｈｅｌｉａｌ ｃｅｌｌ,ＬＥＣ）的粘附力学特性。结果发现,ＨＣＣ细胞的上述流变特性较之于肝细胞发生明显的变化,     

Interactive effects of O3, cytochalasin D, and vinblastine on transepithelial transport and cytoskeleton in rat airways

Cytoskeletal perturbations and associated changes in transepithelial transport in rat airways were analyzed after in vivo treatment with cytochalasin D or vinblastine or exposure to ozone (O3). Exposure of O3 or cytochalasin D, but not vinblastine, increased permeability in the bronchoalveolar region. Combined treatment with cytochalasin D and O3 did not increase the effect seen with each agent alone. However, treatment with vinblastine plus 0.8 ppm O3 resulted in a slight enhancement of permeability over that seen with O3 alone. This enhancement was not seen with 2 ppm O3. When cytochalasin and vinblastine treatment were combined, a synergistic effect on bronchoalveolar permeability was seen, suggesting participation of both microfilamentous and microtubular cytoskeletal elements in maintaining the integrity of the bronchoalveolar epithelium. Potentially harmful effects of O3 on cytoskeletal elements were confirmed in rat lung epithelial cells in culture. O3 exposure produced reversible changes in microfilamentous structures comparable to those produced by cytochalasin D. The results of these studies support the hypotheses that the cytoskeleton has a central role in maintenance of respiratory epithelial integrity and that a target for O3 toxicity may be the components of cytoskeleton. These results, however, do not rule out the possibility that treatment with cytoskeleton destabilizing drugs leads to the release of mediators, which in turn contribute to the airway epithelial dysfunction and increased permeability.     

Morphometric study of hepatocytes containing hepatitis B surface antigen

The development of hepatocellular carcinoma (HCC) is probably related to infection with hepatitis B virus (HBV). Hepatocytes in livers of patients with HCC have been reported to show putative preneoplastic changes such as hyperplasia, dysplasia, or adenomatous regeneration. To determine quantitatively whether these morphologic changes are associated with HBV-infected cells, the authors performed morphometry of hepatitis B surface antigen (HBsAg)-positive hepatocytes in the nontumorous portion of 10 livers with HCC and in 10 livers without HCC. The diameter of nuclei and cytoplasm of HBsAg-positive hepatocytes was measured after demonstration of HBsAg by the peroxidase-antiperoxidase method. As controls, HBsAg-negative hepatocytes in the same liver sections were measured as well as hepatocytes of 20 age-matched HBsAg-negative patients with normal liver or alcoholic cirrhosis. HBsAg-positive hepatocytes exhibited significantly larger nuclei and a higher nucleocytoplasmic ratio than control hepatocytes. In addition, HBsAg-positive cells were often arranged in foci that consisted of two cell populations: hypertrophic (enlarged nuclei and nucleocytoplasmic ratio) and hyperplastic (two-cell-thick plates of small cells with a high nucleocytoplasmic ratio). While precancerous cells have been difficult to identify, these morphologic changes are frequently associated with the development of malignant neoplasia.     

Role of the cytoskeleton in Ca2+ release from the intracellular Ca store of rat peritoneal mast cells

In order to study the role of the cytoskeleton in histamine release from mast cells, the effects of cytochalasin D, cholchicine and vinblastine on Ca²⁺ release from the intracellular Ca store induced by compound 48/80 were investigated by means of a video-intensified microscopy system. When the quin 2-loaded mast cells were stimulated by 0.35 g/ml of compound 48/80, a rapid increase in intracellular Ca²⁺ was observed. At concentrations higher than 10^–6 M, both colchicine and vinblastine pretreatments significantly inhibited the increase in intracellular Ca²⁺ concentrations caused by compound 48/80, although cytochalasin D had no effect. When permeabilized mast cells were exposed to potassium-antimonate solution, microtubules became attached to the endoplasmic reticulum, where many dots of Ca-antimonate were observed; in some areas, the microtubules interconnected the endoplasmic reticulum and granules in the mast cells. From the results of the present study, it was assumed that microtubules play some important role in the processes leading to Ca²⁺ release from the intracellular Ca store.     

Influence of Inherent Mechanophenotype on Competitive Cellular Adherence

Understanding the role of mechanophenotype in competitive adherence of cells to other cells versus underlying substrates can inform such processes as tissue development, cancer progression, and wound healing. This study investigated how mechanophenotype, defined by whole-cell, elastic/viscoelastic properties for the perinuclear region, and cellular assembly are intertwined through the mechanosensing process. Atomic force microscopy was used to characterize the temporal elastic/viscoelastic properties of individual and assembled fibroblasts grown on substrates with elastic moduli above, below, or similar to whole-cell mechanophenotypes measured for three, genetically modified cell lines. All cells were at their most compliant immediately after plating but transitioned to distinct, stiffer mechanophenotypes by Day 1 after acclimation. This mechanical state, and cellular assembly/morphology, did not change significantly over the following three days of testing, regardless of substrate compliance or cellular organization (multi-cell nodules/plaques or single cells). Interestingly, cells formed 3D nodules when attached to substrates with elastic moduli less than their own but spread readily on substrates with moduli equal to or greater than their own, suggesting a preference to adhere to the stiffest surface sensed (substrate or cell). This suggests that inherent mechanophenotype plays a role as a competing surface during microenvironment mechanosensing and subsequent cell–cell-substrate organization.     

Human T-lymphocyte rosette formation: inhibition by cytochalasin B.

The effects of cytochalasin B, colchicine and vinblastine on the rosette formation of human T lymphocytes with neuraminidase-treated human erythrocytes (nHRBC) and with sheep red blood cells (SRBC) have been studied. Pretreatment of the lymphocytes with cytochalasin B which affects microfilament action reversibly inhibits both nHRBC and SRBC rosette formation. Colchicine and vinblastine known to interact with microtubules causes no major reduction in rosette-forming cells. The results suggest that normally functioning microfilaments are necessary for nHRBC and SRBC rosetting, whereas microtubules are not essential for the blinding of the erythrocytes to the lymphocytes.     

Vinblastine-induced autophagocytosis: the effect of disorganization of microfilaments by cytochalasin B

The effects of disorganization of cellular microfilaments by cytochalasin B on vinblastine-induced autophagocytosis was studied in Ehrlich ascites tumor cells in vitro. Incubation with vinblastine induced a formation of autophagic vacuoles in the cytoplasm. The disorganization of microfilaments by cytochalasin B failed to inhibit vinblastine-induced autophagocytosis. Incubation with cytochalasin B alone induced a rapid formation of blebs on the cell surface. These contained cytoplasmic organelles and were connected by a narrow shaft to the main part of the cell. Thin subcortical microfilaments seen in the control cell cytoplasm were apparently relocated after cytochalasin B treatment and formed amorphous masses deeper in the cytoplasm. Vinblastine did not affect the formation of blebs after cytochalasin B treatment.     

Characterization of hyperplastic foci observed in surgical specimens of hepatocellular carcinoma

By reviewing previous surgical specimens of hepatocellular carcinoma, 17 cases with hyperplastic foci (HPF) characterized by discernible increase in nuclear densities, could be histologically selected. Nuclear densities of HPF and control hepatic parenchyma were assessed quantitatively by counting the nuclear number of hepatic cells, and proliferating cell nuclear antigen labeling index was measured. HPF occurred multifocally, confined within a lobular unit, smoothly merging into surrounding hepatic parenchyma. Nuclear densities of HPF were 1.71 times greater than those of control hepatic parenchyma. The hepatocytes of HPF also showed significantly higher proliferative activities than those of control parenchyma. In addition, noticeable structural distortions, such as focal trabecular thickening or microacinar formation of hepatocytes, were sometimes observed in HPF. However, these HPF seemed to be distinguished from minute de novo hepatocellular carcinoma (HCC) or intrahepatic HCC metastasis, because of paucity of distinctive atypical changes, and intimate correlation with neighboring hepatocytes. Several adjacent HPF were aggregated to form a much larger unit of a hyperplastic area with loss of fibrous septa of liver cirrhosis. It was suggested that grossly detectable large regenerative nodules are produced via fusion of several adjacent HPF.     

Lectin binding patterns in normal liver, chronic active hepatitis, liver cirrhosis, and hepatocellular carcinoma. An immunohistochemical and immunoelectron microscopic study.

We studied the binding patterns of 14 lectins in human normal and cirrhotic liver (LC) tissues and hepatocellular carcinomas (HCC) using the ABC method. Lectins were divided into 4 groups according to their binding patterns in normal tissues: (A) PHA, MPA, LcH, RCA-I, and WGA, which bound to hepatocytes and all three types of sinusoidal cells; (B) BPA, GS-I, PNA, and SBA, which bound to Kupffer cells and endothelia of interlobular arteries and veins and bile duct epithelia in the portal tract, but not to hepatocytes; (C) UEA-I, which bound only to endothelia of interlobular arteries and veins and bile duct epithelia in the portal tract; (D) LBA, Lotus, LPA, and SJA, which showed no binding. Thus group B lectins may be useful markers of Kupffer cells. Only electron microscopic examination revealed the precise binding sites of lectins in sinusoidal cells and hepatocytes. Hepatocyte cell surface polarities demonstrated by lectin binding in LC and HCC were different from those in the normal liver. The binding pattern of PHA to LC hepatocytes changed from a membranous to both a membranous and a cytoplasmic pattern, and that of LcH to HCC cells changed to dot-like staining in the cytoplasm. These changes of polarities in LC and HCC might be caused by changes in the distribution of lectin-binding carbohydrates or by the altered glycosylation of glycoconjugates.     

Optical tracking of local surface wave for skin viscoelasticity

Rapid and effective determination of biomechanical properties is important in examining and diagnosing skin thermal injury. Among the methods used, viscoelasticity quantification is one of the most effective methods in determining such properties. This study aims to rapidly determine skin viscoelasticity by optically tracking the local surface wave. New elastic and viscous coefficients were proposed to indicate skin viscoelasticity based on a single impulse response of the skin. Experiments were performed using fresh porcine skin samples. Surface wave was generated in a single impulse using a vibrator with a ball-tipped device and was detected using a laser Doppler vibrometer. The motions along the depth direction were monitored using an ultrasound system. The ultrasound monitoring results indicated the multi-layered viscoelasticity of the epidermis and dermis. The viscoelastic coefficients from four healthy samples show a potential viscoelasticity variation of porcine skin. In one sample, the two coefficients were evidently higher than those in a healthy area if the skin was slightly burned. These results indicate that the proposed method is sensitive, effective, and quick in determining skin viscoelasticity.