Macrophage requirement for host defence against experimental hepatic amoebiasis in the hamster

The role of macrophages in hepatic amoebiasis in hamsters has been investigated by means of their treatment with bacille Calmette-Guérin (BCG) for activation, and with silica for elimination of these cells. Silica-treated animals inoculated intrahepatically with 1 x 10(5) trophozoites of Entamoeba histolytica developed amoebic abscesses in the liver and more metastases to other organs than control animals, and this effect was silica-dose-dependent. In contrast, BCG-treated animals developed significantly smaller abscesses in the liver and fewer metastatic foci. These data suggest that macrophages are involved in host defence against the establishment of amoebic liver abscess and metastatic dissemination of amoebae.     

Absence of CD38 delays arrival of neutrophils to the liver and innate immune response development during hepatic amoebiasis by Entamoeba histolytica

To define the role of CD38 in the migration of neutrophils to the liver and consequently in the induction of an innate immune response during murine hepatic amoebiasis by Entamoeba histolytica, we examined amoebic liver abscess development (ALA), presence of amoebae and neutrophils, and expression levels of cytokines and other inflammation mediators mRNA, in infected wild-type and CD38 Knockout (CD38KO) C57BL/6J mice. Results showed that CD38KO mice undergo a delay in ALA development in comparison with the wild-type strain. The presence of amoebae lasted longer in CD38(-/-), and although neutrophils arrived to the liver in both strains, there was a clear difference in the time between the two strains; whereas in the wild-type strain, neutrophils arrived at early times (6-12 h), in the CD38KO strain, neutrophils arrived later (48-72 h). Cytokines profile during the innate immune response development (TNF-α, IL-1β, IL-6) was, for WT mice concomitant with, and preceded, for CD38KO mice, the time in which neutrophils were present in the liver lesion. In conclusion, CD38 is important for neutrophils migration during hepatic amoebiasis, and in turn, these cells play an important role in the innate immune response.     

Hepatic ultrasound in a population with high incidence of invasive amoebiasis: evidence for subclinical,self-limited amoebic liver abscesses

About 10% of successfully treated amoebic liver abscesses (ALA) do not completely resolve and can be detected by ultrasound as typical residual liver lesions. The frequency of these residues should be an indicator for the prevalence of ALA in a given population, and may help to solve the question whether non-clinical, self-healing ALAs occur. We have performed hepatic ultrasound in 1036 adult individuals living in a high-risk area for ALA in Central Vietnam and identified typical ALA residual lesions in about 1.2% of the subjects. As expected, these lesions were associated with positive amoeba serology and were found in 11.9% of individuals with a previous ALA history. However, more than 50% of the residues were identified in individuals who had never developed symptoms suspected for ALA and who never received any ALA specific treatment, suggesting that subclinical, self-limited hepatic amoeba abscesses truly exist.     

Reconstruction of hepatic organoid by hepatic stem cells

Recent advances in culture methods, stem cell research, and tissue engineering provide clues for making tissues in vitro that are functionally and structurally similar to hepatic tissues. To reconstruct hepatic organoids, two approaches to establish the methods have been proposed: the use of cells and the combination of cells and a scaffold (called tissue engineering). Recently, the coculture of hepatic cells (mature hepatocytes, small hepatocytes, hepatoblasts) and hepatic nonparenchymal cells has been reported to form hepatic organoids that possess differentiated hepatic functions. On the other hand, hepatocytes in a roller bottle were shown to form specific structures, consisting of biliary epithelial cells, connective tissue, mature hepatocytes, and endothelial cells. In this review, the studies of hepatic tissue formation in vitro will be summarized.     

Regulation of the inflammatory immune response by the cytokine/chemokine network in amoebiasis

Amoebiasis is caused by the protozoa Entamoeba histolytica and persists as one of the leading parasitic diseases affecting millions worldwide. This parasite invades the intestinal mucosa, causing amoebic colitis and ulcers. It may also spread to other organs, mainly the liver, causing amoebic liver abscess (ALA). Current research efforts have focused on the development of specific diagnostic tests and animal models searching for a better understanding of the complex physiopathology of this disease. Analysis of the inflammatory immune response during intestinal amoebiasis in both human disease and animal murine models has revealed an important regulatory role for chemokines and cytokines. Recruitment and activation of inflammatory cells can also be modulated by specific protease-mediated cleavage of cytokines and by secreted amoebic factors such as amoebapores and monocyte locomotion inhibitory factor (MLIF). Unlike intestinal amoebiasis, analysis of the immune response in ALA has mainly been done in the hamster model. This has limited our information regarding the immune response during this phase of the disease. However, even with these limitations, several Th1/2 cytokines, such as IL-6 and IL-4, and regulatory cytokines, like IL-10 and TGFbeta, have been associated to the development of this disease.     

Kupffer cell functions during intestinal amoebiasis in guinea pigs

Kupffer cells play an important role in all alimentary tract infections. Their role in intestinal amoebiasis is not clear. Hence, the present study examined Kupffer cell functions--phagocytic capacity and levels of a key lysosomal enzyme, N-acetyl-beta-glucosaminidase -- in guinea pigs infected with Entamoeba histolytica intracaecally. Animals were sacrificed on days 0, 3, 7, 14 and 21 post-infection. During intestinal amoebiasis the phagocytic capacity in Kupffer cells was depressed, whereas lysosomal enzyme levels were highly elevated. Maximum alteration in Kupffer cell functions was observed on the 14th post-infection day (P less than 0.01). Animals which survived until 21 days post-infection did not show any significant (P greater than 0.05) change in their Kupffer cell functions compared with controls and day zero values. Phagocytic capacity was inversely correlated with severity of caecal lesions. In contrast, enzyme levels were directly correlated with degree of caecal lesions. Similar trends were observed in blood monocytes. The depressed phagocytic capacity of mononuclear phagocytes and the increased enzyme release by these cells accompanying intestinal amoebic infection may contribute to the establishment of the infection and the tissue damage that accompanies it.     

Protection against experimental echinococcosis by non-specifically stimulated peritoneal cells

Summary Infection of cotton rats with Echinococcus multilocularis or vaccination with BCG, or its cell walls, activates peritoneal cells to kill the protoscolices of the parasite in vitro and protects laboratory animals against the cestode. To determine whether other 'non-specific' stimuli would also protect against the parasite, cotton rat peritoneal cells were activated in vivo with PHA and transferred to recipients 3 days later. The recipients, controls and PHA-treated animals were then inoculated with the parasite; 3 days after inoculation other untreated infected animals received cells activated in vivo with PHA. PHA-activated cells, the PHA treatment itself and immunization with a homogenate of the parasite stimulated a leucocytosis and protected against infection by E. multilocularis; carrageenan abrogated protection in PHA-treated animals. The results of this study confirm that protection against echinococcosis can be induced non-specifically; these results suggest that immunity in hydatid disease may have an important component in the inflammatory reaction.     

Dendritic cells and immunity against cancer

T cells can reject established tumours when adoptively transferred into patients, thereby demonstrating the power of the immune system for cancer therapy. However, it has proven difficult to maintain adoptively transferred T cells in the long term. Vaccines have the potential to induce tumour-specific effector and memory T cells. However, clinical efficacy of current vaccines is limited, possibly because tumours skew the immune system by means of myeloid-derived suppressor cells, inflammatory type 2 T cells and regulatory T cells (Tregs), all of which prevent the generation of effector cells. To improve the clinical efficacy of cancer vaccines in patients with metastatic disease, we need to design novel and improved strategies that can boost adaptive immunity to cancer, help overcome Tregs and allow the breakdown of the immunosuppressive tumour microenvironment. This can be achieved by exploiting the fast increasing knowledge about the dendritic cell (DC) system, including the existence of distinct DC subsets that respond differentially to distinct activation signals, (functional plasticity), both contributing to the generation of unique adaptive immune responses. We foresee that these novel cancer vaccines will be used as monotherapy in patients with resected disease and in combination with drugs targeting regulatory/suppressor pathways in patients with metastatic disease.     

Modulation of ischemia-reperfusion-induced hepatic injury by kupffer cells

To elucidate the role of Kupffer cells in ischemia-reperfusion-induced hepatic injury, hepatic injury induced by ischemia-reperfusion was analyzed after modulation of Kupffer cell function. Ischemia of the liver was performed by occlusion of both the portal vein and hepatic artery, which enter into the left lateral and median lobes of the liver. Blood flow in the ischemic lobe was reduced, in contrast to an increased blood flow in the nonischemic lobe during occlusion of the veins. Although hepatocyte damage was not demonstrated by ischemia for <60 min, hepatic injury was found after reperfusion of the liver, and activation of Kupffer cells was morphologically demonstrated by electron microscopies. Suppression of Kupffer cells, induced by previous administration of gadolinium chloride or latex particles, reduced the grade of hepatic injury induced by ischemia-reperfusion. On the other hand, stimulation of Kupffer cell phagocytosis, induced by administration of latex particles at the time of reperfusion, aggravated the ischemia-reperfusion-induced hepatotoxicity, which was then reduced by simultaneous administration of superoxide dismutase. Kupffer cells, isolated from the rats treated with the ischemia-reperfusion procedure, have been found to release increased amounts of oxygen radical intermediates. These results suggest that hepatic injury induced by ischemia-reperfusion is modulated by the function of Kupffer cells and that superoxide anion released from Kupffer cells cound play an important role in ischemia-reperfusion hepatic injury.     

Inhibition of hepatic stellate cells proliferation by mesenchymal stem cells and the possible mechanisms

Aim: During fibrosis, hepatic stellate cells (HSCs) undergo a complex activation process characterized by increased proliferation and extracellular matrix deposition. Previous studies have suggested that mesenchymal stem cells (MSCs) may ameliorate fibrogenesis and represent a promising strategy for cell therapy. However, the underlying mechanisms are not fully understood. Methods: Hepatic stellate cells were treated with or without MSCs. Then cell proliferation and cell cycle were analyzed. Production of soluble factors by MSCs and its relation with cell proliferation suppression was evaluated by transwell co‐culture and RNA interference. Effects of MSCs on the gene expression of collagen were also evaluated. Results: MSCs induced G₀/G₁ arrest of HSCs growth partly through secreting soluble factors TGF‐β3 and HGF, which resulted in up‐regulation of p21^Cip1 and p27^Kip1 expression and down‐regulation of cyclinD1. MSCs inhibited the phosphorylation of extracellular signal‐regulated kinase (ERK) 1/2 and reduced gene expression of collagen type I and III. MSCs did not reverse the proliferation and collagen type I gene expression of HSCs provoked by PDGF. Conclusions: The growth inhibition of HSCs induced by MSCs through an arrest in the G₀/G₁ phase of the cell cycle is partially mediated by secretion of TGF‐β3 and HGF. MSCs inhibit HSCs activation through decreasing phosphorylation of extracellular signal‐regulated kinase (ERK) 1/2. These results further support MSCs may be used as a novel therapy for treating fibrotic diseases in human.     

Relationship between hepatic progenitor cell‐mediated liver regeneration and non‐parenchymal cells

Hepatic progenitor cells (HPCs) are thought to reside in the canals of Hering and can be activated and contribute to liver regeneration in response to liver injury by proliferating and differentiating towards both hepatocytes and biliary epithelial cells. In this setting, several cytokines, chemokines, and growth factors related to liver inflammation and other liver cells comprising the HPC niche, namely hepatic stellate cells (HSCs), play crucial roles in HPC activation and differentiation. In response to several types of liver injury, tumor necrosis factor‐like weak inducer of apoptosis (TWEAK) is secreted by several inflammatory cells, including monocytes, T lymphocytes, and macrophages, and acts as an initiator of the HPC niche and HSC activation. Following TWEAK‐induced activation of the HPC niche, fibroblast growth factor 7 and hepatocyte growth factor released from activated HSC play central roles in maintaining HPC proliferation. In contrast, HGF‐MET and Wnt3a‐β‐catenin signals are the predominant mediators of the hepatocyte differentiation of HPC, whereas epidermal growth factor receptor–NOTCH signaling controls HPC differentiation towards biliary epithelial cells. These signals are maintained exclusively by activated HSC and inflammatory cells surrounding HPC. Together, HSC and inflammatory cells surrounding HPC are responsible for the precise control of HPC proliferation and differentiation fate. In this review, we discuss recent progress in understanding of interactions between HPC and other liver cells in HPC‐mediated liver regeneration in the setting of liver inflammation.     

骨髓来源干细胞门静脉移植治疗肝纤维化的实验研究

目的研究骨髓间充质千细胞（MSCs）诱导分化的肝样细胞对大鼠肝纤维化的治疗作用。方法在体外分离培养MSCs中加入细胞因子,诱导其分化成为肝样细胞,通过门静脉移植入肝纤维化模型大鼠体内后,观察病理组织学变化、评价纤维化分期并监测肝功能的变化。结果肝纤维化模型大鼠经门静脉移植诱导肝样细胞4周后,其肝脏纤维化明显改善。结论肝样细胞移植对肝纤维化模型大鼠的纤维化有明显的治疗作用。     

Increased macrophage migration inhibition factor production in hamsters sensitized by amoebic antigen and glucan

Well defined cell-mediated immune responses were detectable following experimental immunization of hamsters with Entamoeba histolytica antigen, using glucan as an adjuvant. Peritoneal cells from amoeba antigen-glucan sensitized animals, upon incubation with specific antigen in vitro, were found to release into the supernatant a macrophage migration inhibition factor (MIF). Such supernatant fluids inhibited the migration of macrophages from non-sensitized hamsters. The production of MIF was found to be greatly increased if glucan is added to amoeba antigen when sensitizing animals. The optimal concentration for maximum inhibition was recorded at 10(-8) dilution of the supernatant.     

Fenretinide stimulates the apoptosis of hepatic stellate cells and ameliorates hepatic fibrosis in mice

Aim: To investigate whether fenretinide, a clinically proved apoptosis‐inducing chemopreventive agent in tumor cells, can induce apoptosis in hepatic stellate cells (HSCs) and resolve hepatic fibrosis. Methods: CCl₄‐induced liver fibrosis in mice and rat activated hepatic stellate cells (HSC‐T6) as well as hepatocytes (BRL‐3A) were studied. Results: The duplex staining of proliferating cell nuclear antigen and α‐ smooth muscle actin or terminal deoxynucleotidyl transferase‐mediated deoxyuridine triphosphate nick‐end labeling and α‐ smooth muscle actin demonstrated that fenretinide executed its anti‐fibrosis effect in liver by inducing apoptosis rather than inhibiting proliferation of HSCs, while it had no apparently apoptotic effect on hepatocytes. Fenretinide could elicit apoptosis of HSC‐T6 in vitro at the concentration range from 0.5 to 5 µM, but at higher concentrations ≥5 µM was required to induce apoptosis in hepatocytes (BRL‐3A). Conclusion: Further studies using malondialdehyde measurement, Western blot, antioxidant, inhibitors for p53, caspase 8 and 9 – as well as anti‐Fas neutralizing antibody – have shown that in HSC‐T6, fenretinide‐induced apoptosis involves a reactive oxygen species (ROS)‐generated, P53‐independent, mitochondria‐associated intrinsic pathway, whereas in hepatocytes (BRL‐3A), a ROS‐generated, P53‐dependent, Fas‐related extrinsic pathway is triggered only at high concentration.     

Protection of mice against intestinal amoebiasis with BCG, Corynebacterium parvum and Listeria monocytogenes

Treatment with Corynebacterium parvum or BCG, or infection with live Listeria monocytogenes was found to protect mice against subsequent infection with Entamoeba histolytica. Complete protection was obtained in mice treated with 10(7) (colony forming units) of BCG but not with 10(5). Partial protection was achieved with 10(6) of BCG. These data provide evidence that non-specific immunity plays an important role in the host defense against amoebic infection.     

Role of cellular immunity in protection against Trypanosoma cruzi in mice

The importance of antibodies in the maintenance of immunity against Trypanosoma cruzi in mice is emphasized by the failure of either immunization or transfer of immune T cells to afford B cell suppressed mice complete protection against a lethal infection. Both procedures did, however, significantly prolong survival indicating a contributory role for T cells other than simply as helper cells in antibody production. The complete protection afforded intact mice following transfer of immune T cells can be attributed to a significant T cell-mediated augmentation of IgG antibody production.