Hepatic progenitor cells in hepatocellular adenomas.

Hepatocellular adenoma is a benign tumor of the liver that has a small but not negligible risk of malignant transformation into hepatocellular carcinoma. In analogy with the established role of oval cells in hepatocarcinogenesis in rodent models, human hepatic progenitor cells may have a function in the development of liver tumors. To investigate this issue, we performed immunohistochemistry on biopsies of 10 consecutively resected hepatocellular adenomas using markers for hepatic progenitor cells. Sections of paraffin-embedded and frozen biopsies were stained using antibodies against cytokeratins 7, 8, 18, and 19, chromogranin-A, OV-6, and neural cell adhesion molecule. Hepatic progenitor cells were observed in five of 10 hepatocellular adenomas. These five tumors also contained cells with an immunohistochemical phenotype intermediate between hepatic progenitor cells and hepatocytes. Hepatic progenitor cells and intermediate hepatocyte-like cells were scattered throughout the tumors with a density that varied from area to area. Ultrastructural examination confirmed the presence of hepatic progenitor cells. Our study shows that hepatic progenitor cells are present in a considerable proportion of hepatocellular adenomas, supporting the hypothesis that human hepatic progenitor cells can play a role in the development of hepatocellular tumors.     

Kupffer细胞在肝细胞癌肿瘤免疫中的作用及其机制

作为人体内最大的巨噬细胞群和肝内最重要的免疫细胞,Kupffer细胞在肝脏甚至整个人体的感染、炎症、脂质代谢和移植免疫方面扮演着重要的角色.然而,如此重要的故有免疫细胞,在肝脏肿瘤发生、发展方面的作用机制仍然是学术界的肓点.本文通过搜集和总结为数不多的文献并结合我们的研究成果,从巨噬细胞吞噬、分泌和抗原呈递三个方面,对Kupffer细胞在肝脏肿瘤免疫中发挥的作用及其机制作一简单综述.我们认为,在肝脏病变早期,Kupffer细胞可通过上述三种途径,对于肝细胞癌的发生起到了监视与抑制作用;然而在肝脏炎症长期存在的情况下,Kupffer细胞则可能通过分泌机制,诱导肝细胞癌的发生.     

Kupffer cells are responsible for producing inflammatory cytokines and hepatocellular dysfunction during early sepsis.

BACKGROUND: Although hepatocellular dysfunction occurs early after the onset of sepsis, the mechanism responsible for this remains unknown. We tested the hypothesis that the reduction in Kupffer cell (KC) numbers prior to the onset of sepsis prevents the occurrence of hepatocellular dysfunction and reduces levels of the proinflammatory cytokines IL-1beta and IL-6 during the early stage of polymicrobial sepsis. MATERIALS AND METHODS: The number of KC in male adult rats was reduced in vivo by intravenous injection of gadolinium chloride 48 h before the induction of sepsis. KC-reduced and KC-normal rats were then subjected to cecal ligation and puncture (CLP, i.e., a model of polymicrobial sepsis) or sham operation followed by administration of normal saline solution. At 5 h after CLP (the early stage of sepsis), hepatocellular function [i.e., the maximal velocity of clearance (Vmax) and efficiency of active transport (Km) of indocyanine green] was measured using a fiber-optic catheter and in vivo hemoreflectometer. Whole blood was drawn to measure plasma levels of IL-1beta and IL-6 using enzyme-linked immunosorbent assays. RESULTS: Hepatocellular function was depressed and the circulating levels of IL-1beta and IL-6 were increased significantly at 5 h after CLP. KC reduction by prior administration of gadolinium chloride, however, prevented the occurrence of hepatocellular dysfunction and the upregulation of IL-1beta and IL-6. CONCLUSIONS: The KC-derived proinflammatory cytokines IL-1beta and IL-6 appear to be directly or indirectly responsible for producing hepatocellular dysfunction during early sepsis. Thus, pharmacologic agents that downregulate the production of one or both of these proinflammatory cytokines in the liver may be useful for maintaining hepatocellular function during the early stage of sepsis.     

Prostanoid production by lipopolysaccharide-stimulated Kupffer cells

Although some data suggest that macrophages in the reticuloendothelial system (RES) are important sources of thromboxane A2 (TxA2) and prostacyclin (PGI2) during endotoxic shock, we are unaware of data documenting the ability of hepatic macrophages (Kupffer cells) to release either TxA2 or PGI2 when exposed to lipopolysaccharide (endotoxin, LPS). In this study, Kupffer cells were examined for their ability to release prostaglandin E2 (PGE2), TxA2, and PGI2 following stimulation with 0, 1.0, 50.0, and 100.0 micrograms/ml of Escherichia coli LPS. Kupffer cells were obtained from rat livers by enzymatic digestion with 0.05% collagenase followed by enrichment of the macrophage population on the basis of differences in density and adherence among the various cell populations isolated. Based on several criteria (phagocytosis of opsonized sheep erythrocytes, positive staining for esterase and peroxidase, failure to replicate), 95% of adherent cells were Kupffer cells. After 4 days of incubation, cells were stimulated with various doses of LPS for 4 and 8 hr. Prostanoid concentrations in culture supernatants were determined by radioimmunoassay. Increasing doses of LPS significantly (P less than 0.001) increased the concentration of immunoreactive PGE2 (iPGE2) and iTxB2 (the stable metabolite of TxA2). The concentration of i6-keto-PFG1 alpha (stable metabolite of PGI2) increased following stimulation with 1.0 microgram/ml of LPS, but declined as the dose of LPS was increased. The results provide evidence that endotoxin-activated Kupffer cells, like other macrophage populations, release several metabolites of arachidonic acid. Kupffer cell-derived prostanoids, particularly TxA2, may be important mediators of some of the pathophysiologic manifestations of acute endotoxemia.     

Activation of human Kupffer cells by thymostimulin (TP-1) to produce cytotoxicity against human hepatocellular cancer

Background: In a small pilot study, thymostimulin (TP-1) produced tumor regression in almost 50% of patients with hepatocellular cancer (HCC) who were treated with TP-1 alone. However, the mechanism of the TP-1-mediated antitumor effect against HCC is unknown. Methods: Human hepatocytes and Kupffer cells were isolated from liver biopsy specimens by collagenase infusion and counterflow elutriation. Hepatocytes and Kupffer cells were incubated in vitro with clinically relevant doses of TP-1. Cell-free supernatants were collected at various time points after incubation. Hepatocyte and Kupffer cell supernatant levels for a panel of growth factors and monokines were determined by enzyme-linked immunosorbent assay. The cytotoxic activity of TP-1 alone and of TP-1-stimulated hepatocyte and Kupffer cell supernatants against Hep G2 and Hep 3B human HCC cells in vitro was measured by MTT assay. Results: Doses of TP-1 up to 100 µg/ml produced no cytotoxicity against Hep G2 or Hep 3B cells. Furthermore, supernatants from TP-1-treated hepatocytes produced no cytotoxicity against Hep G2 or Hep 3B cells, and TP-1 did not stimulate the release of transforming growth factor (TGF)-, TGF-, or hepatocyte growth factor. TP-1-treated Kupffer cell supernatants produced significant cytotoxicity against Hep G2 cells but produced no cytotoxicity against Hep 3B cells. Kupffer cells stimulated by TP-1 released significant amounts of tumor necrosis factor-alpha (TNF-), interleukin (IL)-1, and IL-6 compared with control Kupffer cells (p<0.01). The activity of TP-1-treated Kupffer cell supernatants against Hep G2 cells was blocked by anti-TNF- antibodies, whereas neither anti-IL-1 nor anti-IL-6 antibodies blocked cytotoxicity. Conclusions: These results demonstrate that TP-1 cytotoxicity against human HCC cells is not mediated directly or through hepatocytes, but occurs through activation of Kupffer cells and release of TNF-. Understanding the mechanism of TP-1 cytotoxicity against human HCC has been used to plan a phase I trial of TP-1 combined with regional infusion of doxorubicin to treat unresectable HCC.Presented at the 49th Annual Cancer Symposium of The Society of Surgical Oncology, Atlanta, Georgia, March 21–24, 1996.     

脂质体包裹氯膦酸二钠剔除大鼠肝脏枯否细胞作用的研究

目的探讨脂质体包裹氯膦酸二钠剔除大鼠肝脏枯否细胞的作用。方法实验组大鼠给予脂质体包裹的氯膦酸二钠,对照组给予等量生理盐水。给药后不同时间点计数ED1、ED2阳性细胞数目;尾静脉注射印度墨汁判断枯否细胞吞噬碳素颗粒的情况;RT-PCR检测枯否细胞受体mRNA的表达情况。结果给药后2d大鼠肝脏吞噬碳素颗粒的枯否细胞消失,ED1、ED2阳性细胞基本消失,PCR检测不到肝脏枯否细胞受体mRNA的表达。给药后第8天ED1阳性细胞开始明显增多,至第11天其数目基本恢复正常。ED2阳性细胞至第11天开始增加,但平均每中倍视野仍仅有(4.8±1.7)个细胞,明显少于对照组的(17.7±2.1)个细胞,差异具有统计学意义(P0.01)。结论静脉单次注射脂质体包裹氯膦酸二钠至少在1周内能有效剔除肝脏枯否细胞。     

肝细胞腺瘤的临床诊治及文献复习

目的分析肝细胞腺瘤的临床特点及诊治经验。方法回顾性分析我院2000年1月至2012年12月外科切除并经病理诊断为肝细胞腺瘤的16例患者临床资料。结果肝细胞腺瘤一般无明显症状,邵分表现为右上腹隐痛不适等非特异性症状。1例合并乙肝,l例合并乙肝及丙肝,所有患者AFP均为阴性。术前诊断符合率为43．75％（7／16）。均接受手术治疗,随访10～160个月,l例术后130个月复发,再次手术后健康存活,其余患者均健康生存。结论肝细胞腺瘤好发于年轻女性,超重可能与发病有关,其临床表现多不典型,常规影像学方法确诊率较低,CT、MRI增强扫描有一定诊断价值。手术切除是肝细胞腺瘤主要的治疗方法。治疗后可复发,需定期复查。     

Hepatic allograft-derived Kupffer cells regulate T cell response in rats

In liver transplantation, the development of tolerance is associated with an increased rate of apoptosis of T lymphocytes in the portal inflammatory infiltrate and the presence of an intragraft Th2-like T cell population. Underlying mechanisms are poorly understood. Kupffer cells (KC), which reside in the hepatic sinosoids, can directly interact with circulating T lymphocytes and thus are uniquely positioned to play a role in immunomodulation. In this study, the immunoregulatory effects of KC were investigated. We show that KC can significantly suppress T cell proliferation in mixed leukocyte reaction (MLR). Furthermore, KC express functional Fas ligand (FasL) and can induce apoptosis of Fas+ cells. This process can be blocked by addition of neutralizing anti-FasL antibody. Moreover, using an allogeneic liver transplant model we have determined that 1. KC recovered from chronically accepted hepatic allografts have increased FasL messenger RNA (mRNA) and protein expression and a greater ability to induce apoptosis of alloreactive T cells compared with KC recovered from an acute rejection model; 2. KC not only induce apoptosis of T cells, but also regulate cytokine production and Th2/Th3-like cytokine (interleukin [IL]-10 / transforming growth factor [TGF]-β) mRNA expression in allogeneic MLR in vitro; and 3. administration of KC derived from chronically accepted liver allografts significantly prolongs the survival of hepatic allografts in an acute rejection model in an alloantigen-specific manner. In conclusion, these data implicate the possible role of KC-mediated regulation of T cell response in the induction of immune tolerance in liver allografts. (Liver Transpl 2003;9:489-497.)     

Kupffer cells participate in rejection following liver transplantation in the rat

Abstract Recognition of foreign antigens involves macrophages which release mediators such as immunoactive interleukins, and in the liver, the resident macrophages (Kupffer cells) are activated following transplantation. Therefore, we evaluated the hypothesis that Kupffer cells participate in the rejection reaction following transplantation. Orthotopic liver transplantation was performed between different syngenic rat strains. Livers from Lewis rats were stored in lactated Ringer's solution for 1 h to minimize cold ischemic injury and transplanted into PVG recipients. At 24 h postoperatively, transaminases (AST) were elevated to values around 2000 U/l, total bilirubin was increased to values around 20 μmol/l, and five of six rats died within 3 days. Macroscopic and histological examination showed large areas of necrosis without cellular infiltration, characteristic of rejection. When donor rats were treated with gadolinium chloride (GdCl₃, 10 mg/kg i.v. 24 h before storage of the liver) to inactivate the Kupffer cells, AST levels only rose to around 700 U/l, and the total bilirubin level was in the normal range (<4 μmol/l). Survival was improved significantly by GdCl₃, with five of seven rats surviving more than 1 month ( P < 0.05) and four of seven rats surviving for at least 100 days without immunosuppressive drug therapy. Rejection was not totally prevented, however, since the surviving rats had elevated AST and bilirubin levels, and cellular infiltration in portal areas along with proliferation of bile canaliculi was observed. These data are consistent with the hypothesis that Kupffer cells participate in mechanisms of early rejection following liver transplantation.     

Human Kupffer cells are cytotoxic against human colon adenocarcinoma

Colorectal liver metastases are a common clinical problem and require more effective therapy. Kupffer cells (KC) perform many important homeostatic functions within the liver and may also possess the ability to mediate tumor cytotoxicity. We investigated the ability of human KC to mediate cytotoxicity against human colon adenocarcinoma targets (HT 29) in vitro. Unstimulated human KC were cytotoxic against the HT 29 targets at all effector/target ratios tested. This cytotoxicity was increased significantly (p less than 0.05) when the KC were stimulated with interferon-gamma and lipopolysaccharide. Human KC produced tumor necrosis factor (TNF), and KC stimulation significantly (p less than 0.05) increased secretion of this monokine. The addition of anti-TNF antibody to the KC-HT 29 cocultures completely neutralized all of the available TNF yet cytotoxicity was not affected, suggesting the participation of a membrane-bound form of TNF or other mechanisms.