肝卵圆细胞的研究进展

肝卵圆细胞（OCs）是肝脏的干细胞,能够分化为肝细胞、胆管上皮细胞及其他细胞,亦与肝再生和肝脏疾病密切相关。本文简要地总结了近几年有关肝卵圆细胞的来源、定位、增殖、分化、凋亡、移植及在肝再生和肝脏疾病中作用等研究进展。     

肝损伤时诱导肝星状细胞中小窝蛋白-1表达升高

目的 探究肝损伤介导肝星形细胞小窝蛋白-1的表达的影响.方法 通过肝总管结扎手术诱导大鼠肝损伤模型和体外培养肝星形细胞LX-2,检测肝组织和肝星形细胞小窝蛋白-1的表达水平.结果 ①与假手术组相比,胆总管结扎手术可显著诱导肝脏损伤,手术后7d,手术组大鼠体重显著减轻,肝脏重量和肝指数显著增加,以及血清谷丙转氨酶和谷草转氨酶水平显著增加;②与假手术组相比,肝损伤可显著诱导小窝蛋白-1在mRNA和蛋白水平显著增加;③与胆总管结扎组相比,LPS处理可显著增加肝星形细胞小窝蛋白-1的表达水平.结果 肝损伤可显著诱导肝星形细胞小窝蛋白-1的表达.     

大鼠肝纤维化过程中肝内成纤维细胞的免疫组化研究

目的肝脏内存在有两种成纤维细胞,一种是肝星形细胞(hepatic stellate cell,HSC),一种是门脉区域的成纤维细胞。用免疫组化方法对这两种细胞群在肝纤维化(hepatic fibrosis,HF)中活化后的细胞形态进行组织学鉴别。方法Wistar系雄性鼠用于实验动物模型制作:CCL4腹腔注射和胆总管结扎(bile duct ligation,BDL)诱导肝纤维化动物模型,用SABC法来检测。一次抗体分别为GFAP、α-SM、Desmin、Vinculin。结果CCl4腹腔注射后48h中央静脉周围的星形细胞在GFAP、α-SMA、Desmin、Vinculin染色呈强阳性;另外,BDL96h后门脉区域成纤维细胞在α-SMA、Vinculin染色中显示阳性,而在GFAP、Desmin染色中未发现有变化。结论α-SMA、Vinculin在肝星形细胞和门脉区域成纤维细胞有相同的阳性结果,GFAP、Desmin被证明能够用于区别有活性的肝星形细胞和门脉区域的成纤维细胞。     

JNK信号通路调控大鼠再生肝8种细胞的增殖和凋亡

目的 从基因转录水平了解JNK信号通路在大鼠再生肝8种细胞中的作用。方法 用密度梯度离心和免疫磁珠等方法分离肝细胞（HC）、胆管上皮细胞（BEC）、卵圆细胞（OC）、肝星形细胞（HSC）、窦内皮细胞（SEC）、库普弗细胞（KC）、陷窝细胞（PC）、树突状细胞（DC）等8种肝脏细胞,用大鼠Genome 230 2.0芯片检测大鼠再生肝8种细胞的基因表达谱,用生物信息学和系统生物学等方法分析基因表达变化预示的JNK信号通路在调控大鼠再生肝8种细胞增殖、凋亡中的作用。用实时荧光定量PCR方法验证了芯片结果的可靠性。结果 JNK信号通路涉及240个基因和42条途径,其中,225个基因与大鼠肝再生相关。基因协同作用分析显示,在大鼠肝再生启动阶段,JNK信号通路启动HC和KC增殖,促进OC凋亡,启动部分PC和SEC增殖和促进部分PC和SEC凋亡;在进展阶段,JNK信号通路促进HC、BEC、KC和DC增殖,促进部分PC增殖、部分PC凋亡。在终止阶段,JNK信号通路促进HC、OC和PC凋亡,促进部分KC增殖、部分KC凋亡。结论 大鼠肝再生中JNK信号通路的42条途径和225个基因参与调控大鼠再生肝8种细胞的增殖和凋亡。     

大鼠肝卵圆细胞的诱导、分离及鉴定

目的建立大鼠肝卵圆细胞的增殖模型,并探索其分离及鉴定方法。方法雄性Wistar大鼠每天1次连续灌胃给予不同剂量二乙酰氨基芴(2-AAF熏5、10、15、20、25mg/kgBW),第5天行标准的2/3肝切除术,术后按各自剂量继续给予11天,不同时间取肝脏组织,行甲胎蛋白、细胞角蛋白18及19染色并观察。以确定的2-AAF最佳剂量制备大鼠肝干细胞增殖模型,Seglen胶原酶原位灌注结合Percoll密度梯度离心分离纯化大鼠肝卵圆细胞,光镜、电镜下观察细胞特点,并进行上述细胞表型标志免疫组化染色。结果2-AAF15mg/kgBW能建立较理想的肝卵圆细胞增殖模型。HE染色可见汇管区及中央静脉周围大量增殖的嗜碱性小细胞,电镜下观察此种细胞具有卵圆形细胞核、细胞质少而淡、核/浆比例较大等特点,免疫组化染色证实甲胎蛋白、细胞角蛋白18和19染色阳性,白蛋白及白细胞共同抗原(LCA)染色阴性。分离所得底层细胞,光镜下表现大小不等、不规则圆形细胞,体积较小,细胞核/浆比例较大,电镜下细胞表面可见少量短而小的微绒毛状突起,余同增殖细胞特点,免疫组化染色与增殖细胞表现相同细胞表型特点。结论本方法可成功诱导、分离、纯化大鼠肝卵圆细胞,符合肝卵圆细胞的形态特点、超微结构及细胞表型标志特点。     

肝卵圆细胞的研究进展

肝卵圆细胞(hepaticovalcells,HOC)是存在于肝脏中的一类肝干细胞,目前已证实了它在人及 多种动物中存在。HOC来源并定位于肝内的胆管系统中,具有多向分化潜能,正常时处于静止状态,只有当 肝脏发生严重损伤且肝细胞再生障碍时才受到激活并大量增殖,可分化为肝细胞和胆管细胞,以修复重建肝 脏组织、恢复生化功能。本文就HOC的来源、定位、生物学特性、研究方法及应用方面的进展情况作一综述。     

肝纤维化形成过程中Ⅰ型血管紧张素Ⅱ受体表达的研究

目的 :探讨Ⅰ型血管紧张素Ⅱ受体 (AngiotensinⅡType1Receptor,AT1R)在人肝脏组织中的表达情况。方法 :运用免疫组织化学SABC法和间接免疫荧光标记法 ,对 12例正常肝组织及 18例纤维化肝组织进行检测。结果 :AT1R阳性表达主要分布在肝小叶周边及肝窦区星形细胞 (HepaticStellateCell,HSC)的胞浆内。 12例正常肝组织中 8例呈阳性表达 ,18例纤维化肝组织均表达AT1R ,且纤维化组阳性细胞数明显多于正常组 (P <0 0 0 1)。结论 :AT1R在肝星形细胞胞浆中的表达随肝纤维化的发展明显增强 ,并且可能在肝纤维化的发生发展中发挥着一定作用。     

解耦联蛋白2对大鼠肝纤维化形成中星形细胞活化的影响

目的探讨解耦联蛋白2(UCP2)在肝纤维化形成过程中的作用及其发生机制。方法体内实验采用四氯化碳(CCl4)诱导肝纤维化模型,取肝脏观察病理变化,用Western blotting、免疫组织化学和Real-time PCR等方法检测UCP2和p38丝裂素活化蛋白激酶(p38MAPK)的表达水平;体外实验采用UCP2特异性抑制剂京尼平和CCl4刺激星形细胞,检测UCP2和p38MAPK相关蛋白的表达情况。结果与正常组相比,模型组大鼠肝脏α-平滑肌肌动蛋白(α-SMA)和UCP2表达增高(P0.05,n=10);加入CCl4刺激细胞后,星形细胞α-SMA表达增加,p38MAPK及其磷酸化水平增高(P0.05,n=6);而在加入京尼平后,α-SMA表达增加,p38 MAPK及其磷酸化水平明显降低(P0.05,n=6)。结论 UCP2参与了肝纤维化的发生,可能促进了星形细胞的活化及增殖过程。     

细胞外间质与肝纤维化

肝纤维化(liver fibrosis)是指各种致病因子所致的肝脏内纤维结缔组织增生,其特点为肝脏中细胞外间质,特别是胶原的过度沉积,纤维化发展的后果是形成肝硬化。细胞生物学和分子生物学研究表明,肝纤维化不是简单的固定不变的组织学改变,而是一种主动的动态病理过程。细胞外间质亦不仅仅起支架作用,它对细胞的结构和功能有重要影响。本文结合这些新概念对肝纤维化的发生机理及诊断治疗进展作一综述。     

肝源性肝干细胞

肝干细胞是指能够参与肝细胞再生和损伤修复的同时具有干细胞特质的细胞的总称。当肝脏或肝功能受到严重损伤,肝干细胞被激活并增殖,分化为肝实质细胞和胆管上皮细胞,使得肝功能得以重建。根据起源不同,肝干细胞可分为肝源性肝干细胞和非肝源性肝干细胞。肝干细胞是具有重要临床价值的细胞,在自我更新、高度增殖和多向分化潜能方面都具有巨大的潜能。近年有实验表明,肝干细胞亦可能是肝肿瘤细胞的起源,它与肝肿瘤的发生发展密切相关,提示肝干细胞的研究对于了解肝损伤修复机理,肝肿瘤的发生发展机制具有重要意义。本文对肝干细胞的分类、来源及特性,肝干细胞的表面标志的主要差别,肝干细胞的活化、分离和培养,肝干细胞分化增殖的调节,肝干细胞与原发性肝癌的关系以及肝干细胞的发展前景的相关研究进展作一阐述。     

JNK1 and JNK2 regulate α‐SMA in hepatic stellate cells during CCl4‐induced fibrosis in the rat liver

Following liver injuries, hepatic stellate cells (HSCs) express α‐SMA. Mitogen activated protein kinase (MAPK) signaling pathways mediate α‐SMA expression in distinct cell types. However, the regulation of α‐SMA expression by MAPKs in HSCs has been rarely studied. We aimed to study the role of MAPKs in the activation of HSCs during liver fibrosis. Liver fibrosis of rats was induced by carbon tetrachloride. HSC‐T6 cells, murine embryonic fibroblasts, JNK1^?/? and JNK2^?/? cells were used for in vitro studies. Immunohistochemistry and immunoblot analysis were used. We have found that the expression of JNK and α‐SMA co‐localized in HSCs during liver fibrosis, but ERK and p38 expressed in macrophages. The expression of α‐SMA was up‐regulated by JNK1 and JNK2 in non‐stress condition. Under TGF‐β stimulation, however, the level α‐SMA expression was increased by only JNK1, but not significantly changed by JNK2. We suggest that JNKs are responsible for α‐SMA regulation, and especially JNK1 has a major role in up‐regulation of α‐SMA expression in HSCs under stress condition induced by TGF‐β during liver fibrosis.     

Innervation of the sinusoidal wall: regulation of the sinusoidal diameter

In the livers of humans, cats, guinea pigs, and tupaia, nerve endings are distributed all over the hepatic lobules. Nerve endings in the intralobular spaces are localized mainly in the Disse spaces and are oriented toward the hepatic stellate cells (HSCs), sinusoidal endothelial cells, and hepatocytes. They are especially closely related to HSCs. Various neurotransmitters such as substance P exist in the nerve endings. In addition, HSCs possess endothelin (ET) and adrenergic receptors and contract in response to the corresponding agonists. In contrast, nitric oxide (NO) inhibits the contraction of HSCs. HSCs thus appear to be involved in the regulation of hepatic sinusoidal microcirculation by contraction and relaxation. In the cirrhotic liver, intralobular innervation is decreased, but ET, ET receptors, and NO are overexpressed in the HSCs. These findings indicate that HSCs in cirrhotic liver may play an important role in the sinusoidal microcirculation through agents such as ET or NO rather than through intralobular innervation.     

Growth inhibition and apoptosis in liver myofibroblasts promoted by hepatocyte growth factor leads to resolution from liver cirrhosis

Liver cirrhosis is characterized by hepatic dysfunction with extensive accumulation of fibrous tissue in the liver. In response to chronic hepatic injury, hepatic portal myofibroblasts and activated hepatic stellate cells (HSCs) play a role in liver fibrosis. Although administration or gene expression of hepatocyte growth factor (HGF) leads to improvement in hepatic fibrosis/cirrhosis, the related mechanisms are not fully understood. We investigated mechanisms involved in resolution from liver cirrhosis by HGF, focusing on growth regulation and apoptosis in portal myofibroblasts. Cultured rat HSCs could not proliferate, were withdrawn after passage, and were replaced by proliferating portal myofibroblasts during the passages. In quiescent HSCs, c-Met receptor expression was undetected whereas c-Met receptor expression was detected in activated HSCs and liver myofibroblasts expressing alpha-smooth muscle actin (alpha-SMA), suggesting that activated HSCs and portal myofibroblasts are targets of HGF. For cultured rat portal myofibroblasts, HGF counteracted phosphorylation of extracellular signal-regulated kinase (Erk) 1/2 and mitogenic stimulus induced by platelet-derived growth factor, induced c-jun N-terminal kinase (JNK) 1 phosphorylation, and promoted apoptotic cell death. In the dimethylnitrosamine rat model of liver cirrhosis, administration of HGF suppressed proliferation while promoting apoptosis of alpha-SMA-positive cells in the liver, events that were associated with reduced hepatic expressions of alpha-SMA and histological resolution from liver cirrhosis. Growth inhibition and enhanced apoptosis in portal myofibroblasts by HGF are newly identified mechanisms aiding resolution from liver fibrosis/cirrhosis by HGF.     

大鼠肝纤维化过程中4种肝脏细胞对纤溶系统的调节

目的:探讨在大鼠肝纤维化发展过程中,肝细胞、肝星状细胞(HSCs)、Kupffer细胞和内皮细胞(ECs)对纤溶系统的调节作用。方法:采用1周2次皮下注射50%CCl4、持续12周的方法制备大鼠慢性肝纤维化模型;将肝纤维化不同时期的4种肝脏细胞进行分离;采用Northern印迹法及Western印迹法分别测定大鼠各类肝脏细胞中尿激酶型纤溶酶原激活物(uPA)、以及它的抑制物(PAI-1)和受体(uPAR)的mRNA和蛋白表达。结果:Northern印迹法及Western印迹法分析均显示,在大鼠肝纤维化过程中,HSCs和ECs均为PAI-1、uPAR的主要产生细胞,相比之下,HSCs产生更多。结论:在大鼠肝纤维化发生发展过程中,HSCs和ECs对纤溶系统成分的产生起着重要的调节作用,而HSCs则是PAI-1、uPAR最主要的产生细胞。     

Hepatic stellate cells—the pericytes in the liver

Hepatic stellate cells (HSCs) are pericytes of liver in the space between parenchymal cells and sinusoidal endothelial cells of the hepatic lobule. HSCs comprise specialized functions such as vitamin A storage, hemodynamic functions, support of liver regeneration, and immunoregulation. In pathological conditions, HSCs transform to an activated myofibroblasts-like phenotype, start to proliferate, and de novo express several proinflammatory and profibrogenic genes. These processes are particularly important in the development of cirrhosis, portal hypertension, and hepatocellular cancer. This review highlights recent findings in understanding the biology of HSCs and discusses the physiological functions of HSCs and the role of activated HSCs in pathophysiology and disease.     

Isolation of a unique hepatic stellate cell population expressing integrin α8 from embryonic mouse livers

Results: Using antibodies against ALCAM and PDPN, we separated mesothelial cells (MCs) and HSCs from developing livers and identified integrin α8 (ITGA8) as a marker for embryonic desmin+ HSCs that are preferentially localized near the developing liver surface and α‐smooth muscle actin+ perivascular mesenchymal cells around the vein. A cell lineage–tracing study revealed that upon differentiation, MC‐derived HSCs or perivascular mesenchymal cells express ITGA8 during liver development. Using anti‐ITGA8 antibodies, we succeeded in isolating MC‐derived HSCs and perivascular mesenchymal cells from embryonic livers. In direct co‐culture, ITGA8+ mesenchymal cells promoted the expression of hepatocyte and cholangiocyte markers in hepatoblasts. In the normal adult liver, expression of ITGA8 was restricted to portal fibroblasts in the portal triad. Upon liver injury, myofibroblasts increased the expression of ITGA8. 相似文献   

Complement 5a stimulates hepatic stellate cells in vitro,and is increased in the plasma of patients with chronic hepatitis B

Complement 5a (C5a) is a critical modulator of liver immunity. In this study, we investigated the role of C5a and its receptor in liver fibrosis in patients with hepatitis B virus infection. We found that plasma C5a concentration was significantly increased in patients with chronic hepatitis B, particularly in those patients with higher grade and stage scores. Further analysis indicated that the increased C5a concentration was positively correlated with clinical parameters reflecting liver fibrosis severity, including type IV collagen and procollagen type III N‐terminal peptide. Our in vitro data indicated that the C5a receptor is highly expressed in hepatic stellate cells (HSCs). Addition of C5a significantly activated HSCs and up‐regulated α‐smooth muscle actin, hyaluronic acid and type IV collagen expression. Also, addition of C5a could inhibit the spontaneous and soluble tumour necrosis factor‐related apoptosis‐inducing ligand‐induced apoptosis of HSCs. These findings highlight the potential role of C5a in the regulation of liver fibrosis.     

Pleiotrophin/heparin-binding growth-associated molecule as a mitogen of rat hepatocytes and its role in regeneration and development of liver

Previously pleiotrophin (PTN) was identified among proteins secreted by Swiss 3T3 cells as a mitogen for cultured adult rat hepatocytes. The present study showed that the growth of rat hepatocytes was enhanced when cultured with rat hepatic stellate cells (HSCs). HSCs expressed PTN mRNA and secreted its protein in the co-cultures. Recombinant PTN enhanced the growth of hepatocytes in culture, suggesting that HSCs stimulate the growth of hepatocytes through the action of PTN. To know the biological role of PTN in the growth of hepatocytes in vivo, we examined the expression of PTN in four regeneration models of adult liver and embryonic liver of rat. The expression of PTN mRNA in the liver was markedly up-regulated by the treatment with D-galactosamine (GalN) or with acetylaminofluorene followed by partial hepatectomy. HSCs expressed PTN mRNA in response to GalN treatment and its protein was found on hepatocytes. The mRNA expression of N-syndecan, a PTN receptor, was up-regulated in GalN-treated hepatocytes. The mesenchymal cells in the septum transversum enclosing the embryonic liver, but not embryonic HSCs, expressed PTN mRNA. We suggest that PTN is secreted from activated adult HSCs and embryonic mesenchymal cells as a mitogen of parenchymal cells in adult and embryonic liver, respectively.     

骨髓造血干细胞龛

造血干细胞(hematopoietic stem cell,HSC)位于骨髓的造血微环境即龛(niche)中,它们与龛内特定的细胞相互作用以调节其自我更新和定向分化。研究发现,骨髓中的成骨细胞和内皮细胞与造血干细胞关系密切,分别构成了HSC龛中的成骨龛和血管龛,其中成骨龛维持静态的HSC微环境,而血管龛调控HSC的增殖、分化和动员等行为。对骨髓HSC龛的研究为将来临床治疗血液系统相关疾病提供了新的思路。     

Increased SSeCKS Expression in Rat Hepatic Stellate Cells Upon Activation In Vitro and In Vivo

Recent reports suggest that src suppressed c kinase substrates (SSeCKS) are early inflammatory response protein. However, there is only scarce knowledge on the functional role of SSeCKS in liver under conditions of acute inflammation. In the present study, we investigated SSeCKS expression in liver after administration of carbon tetrachloride (CCl₄) in rats and in isolated primary hepatic stellate cells (HSCs) upon activation on a plastic dish. We found that SSeCKS mRNA was hardly detectable in healthy liver tissue and further increased in carbon tetrachloride-mediated acute liver failure. SSeCKS protein expression was mainly found in hepatic stellate cells. In vitro, SSeCKS expression in activated rat HSCs was dramatically increased. The upregulation of SSeCKS protein expression in rat HSCs during activation in vitro and in vivo suggested the possibility of SSeCKS, an important part of function of the activated HSCs, perhaps through modulation of liver regeneration or formation of liver fibrosis after various injuries.