ABSTRACT
Chemotactic cytokines (chemokines) play an important role in the recruitment of lymphocytes to tissue by regulating cellular adhesion and transendothelial migration. This study examined the expression and function of CXC (human monokine induced by gamma-interferon [HuMig], interleukin-8 [IL-8], and interferon-inducible protein-10 [IP-10]) and CC (macrophage inflammatory protein-1alpha [MIP-1alpha], MIP-1beta, regulated upon activation normal T lymphocyte expressed and secreted (RANTES), and macrophage chemoattractant protein-1 [MCP-1]) chemokines and their respective receptors on lymphocytes infiltrating human liver tumors. Chemokine and chemokine receptor expression was assessed by immunohistochemistry, flow cytometry, in situ hybridization and ribonuclease (RNAse) protection assays and function by in vitro chemotaxis of tumor-derived lymphocytes to purified chemokines and to HepG2 tumor cell culture supernatants. Tumor-derived lymphocytes showed strong chemotactic responses to both CC and CXC chemokines in vitro and expressed high levels of CXCR3 (HuMig and IP-10 receptor) and CCR5 (RANTES, MIP-1alpha, and MIP-1beta receptor). Expansion of tumor-derived lymphocytes in recombinant IL-2 increased expression of CXCR3. The corresponding chemokines were detected on vascular endothelium (HuMig, IL-8, MIP-1alpha, and MIP-1beta) and sinusoidal endothelium (HuMig, MIP-1alpha, MIP-1beta) in hepatocellular carcinoma. In vitro, HepG2 cells secreted functional chemotactic factors for tumor-derived lymphocytes that could be inhibited using anti-CCR5 or anti-CXCR3 monoclonal antibodies (MoAbs). Thus, lymphocytes infiltrating human liver tumors express receptors for and respond to both CXC and CC chemokines. The relevant chemokine ligands are expressed in hepatocellular carcinoma (HCC), particularly HuMig, which was strongly expressed by tumor endothelium, suggesting that they play a role in lymphocyte recruitment to these tumors in vivo. The ability of HepG2 cells to secrete lymphocyte chemotactic factors in vitro suggests that the tumor contributes to lymphocyte recruitment in vivo.
Subject(s)
Carcinoma, Hepatocellular/immunology , Chemokines, CC/genetics , Chemokines, CXC/genetics , Gene Expression Regulation, Neoplastic/immunology , Interferon-gamma/immunology , Liver Neoplasms/immunology , Liver/immunology , Lymphocytes, Tumor-Infiltrating/immunology , Aged , Antibodies, Monoclonal , Carcinoma, Hepatocellular/genetics , Carcinoma, Hepatocellular/pathology , Carcinoma, Hepatocellular/surgery , Chemokines, CC/analysis , Chemokines, CXC/analysis , Female , Humans , Immunohistochemistry , Liver/pathology , Liver Neoplasms/genetics , Liver Neoplasms/pathology , Liver Neoplasms/surgery , Lymphocytes, Tumor-Infiltrating/pathology , Male , Middle Aged , RNA, Messenger/analysis , Receptors, Chemokine/analysis , Receptors, Chemokine/genetics , Transcription, GeneticABSTRACT
This study demonstrates a novel role for the Fas pathway in the promotion of local tumor growth by inducing apoptotic cell death in normal hepatocytes at the tumor margin in colorectal hepatic metastases. Our results show that >85% of lymphocytes infiltrating colorectal liver cancer express high levels of Fas-ligand (Fas-L) by flow cytometry. Using immunohistochemistry of tumor tissue we showed strong Fas expression in noninvolved hepatocytes, whereas Fas-L expression was restricted to tumor cells and infiltrating lymphocytes at the tumor margin. Apoptosis was observed in 45 +/- 13% of the Fas(high) hepatocytes at the tumor margin whereas only 7 +/- 3% tumor cells were apoptotic (n = 10). In vitro, primary human hepatocytes expressed Fas receptor and crosslinking with anti-Fas antibody induced apoptosis in 44 +/- 5% of the cells compared with 4. 6 +/- 1.0% in untreated controls (P = 0.004). Both tumor-infiltrating lymphocytes (TIL) and human metastatic colon cancer cells cells are able to induce Fas-mediated apoptosis of primary human hepatocytes in coculture cytotoxic assays. TIL induced apoptosis in 47 +/- 9% hepatocytes compared with control 4.3 +/- 1. 0% (P = 0.009) and this effect was reduced by anti-human Fas-L mAb (18.7 +/- 1.3%, P = 0.009). SW620 cells induced apoptosis in 26 +/- 2% hepatocytes compared with control 5.6 +/- 1.7% (P = 0.004) and this was reduced to 11.2 +/- 1.8% (P = 0.004) in the presence of anti-human Fas-L mAb. These data suggest that the inflammatory response at the margin of colorectal liver metastases induces Fas expression in surrounding hepatocytes, allowing them to be killed by Fas-L-bearing TIL or tumor cells and facilitating the invasion of the tumor into surrounding liver tissue.
Subject(s)
Colorectal Neoplasms/pathology , Liver Neoplasms/immunology , Liver Neoplasms/secondary , Membrane Glycoproteins/immunology , fas Receptor/immunology , Aged , Cells, Cultured , Colonic Neoplasms/pathology , Colonic Neoplasms/physiopathology , Fas Ligand Protein , Female , Humans , Liver/cytology , Liver/pathology , Liver/physiology , Lymphocytes, Tumor-Infiltrating/physiology , Male , Middle Aged , Neoplasm Invasiveness/physiopathology , Reference Values , fas Receptor/physiologyABSTRACT
Repeat orthotopic liver transplantation (ReOLT) is controversial because of limited donor organ availability and increasing health care costs. The purpose of this study is to analyse and compare the outcome of reOLT in the 1990s and the 1980s. Prospective data of 1077 adult OLT from the Liver Unit database were used for the study. The log-rank test was used for statistical analysis. Between January 1982 and December 1996, a total of 1077 adult OLTs were performed including 107 reOLTs. The proportion of retransplants decreased from 13% in the 1980s to 9% in the 1990s. There was a significant improvement in outcome; the overall 1-year graft and patient survival for reOLT was 60% and 74% in the 1990s compared to 29% (P < 0.0001) and 51% (P < 0.0001) in the 1980s. In the second half of the study between January 1990 and December 1996, 732 adult OLTs were undertaken including 70 (9%) reOLTs which consisted of 62 second, 7 third and 1 fourth grafts. The main indications for retransplantation were chronic rejection (31%), hepatic artery thrombosis (30%), primary non-function (16%), ischaemic injury (11%), recurrent disease (6%) and biliary complications (6%). During this period, the 1-year graft survival for all reOLTs was significantly lower than for primary OLTs (67% vs 78%, P < 0.001). The timing of reOLT was found to be associated with graft survival; 1-year graft survival for early reOLT (< 30 days) was 50% compared to 73% for late reOLT (P < 0.001). The worse outcome associated with early reOLT is explained by the poor preoperative medical condition of patients who were retransplanted from intensive care. Subgroup analysis of indications for reOLT revealed 1-year graft survival of 81% for late vascular complications, 75% for early vascular complications, 69% for chronic rejection and 30% for primary non-function. One-year graft survival rates for third and fourth grafts were 42% and 0%, respectively. Graft survival and resource utilisation in patients who received a late regraft for the first time is now comparable to that for primary OLT. The favourable overall results should not preclude this group of patients from consideration for reOLT.
Subject(s)
Liver Transplantation , Adult , Graft Survival , Humans , Multivariate Analysis , Prospective Studies , Reoperation , Survival Rate , Treatment OutcomeABSTRACT
Metastatic colorectal cancer is usually progressive despite infiltration of the tumours by T lymphocytes, suggesting that these tumour-infiltrating lymphocytes (TILs) are functionally deficient. Recently, TILs from other tumours have been shown to express reduced levels of the T-cell receptor signal-transducing CD3-zeta chain. We were interested to determine whether a similar abnormality existed in TILs from human colorectal hepatic metastasis (CHM) and, if so, whether correcting the abnormality in vitro would restore anti-tumour activity and provide support for the development of immunotherapy for colorectal hepatic metastases. Twelve of 19 TILs from colorectal hepatic metastases were successfully expanded in vitro in high-dose recombinant interleukin 2 (rlL-2) and their specific anti-tumour cytolytic activity was determined. CD3-positive (CD3+) TILs were HLA-Drhigh and CD69high, suggesting that they had been activated by exposure to antigen but expressed low levels of CD25, CD71 and the nuclear proliferation antigen Ki-67. Furthermore, they showed reduced expression of CD3-zeta compared with autologous peripheral blood T cells (PBTs) and failed to proliferate in the absence of high-dose rIL-2. Expansion of TILs in rIL-2 resulted in restoration of CD3-zeta expression and the ability to lyse K562 and Daudi cells but not autologous tumour cells. The absence of autologous tumour-specific cytolytic T-cell (CTL) activity may be due to the poor immunogenicity of colorectal tumour cells, which we found expressed only low levels of MHC I antigens and CD54 and failed to express MHC II antigens or the co-stimulatory molecules CD80, CD86 or CD106. The inability of rIL-2 to generate tumour-specific CTLs despite restoration of CD3-zeta expression and the presence of an intact lytic mechanism suggests that successful immunotherapy may require the development of strategies to increase the immunogenicity of this tumour.
Subject(s)
CD3 Complex/metabolism , Colorectal Neoplasms/immunology , Interleukin-2/metabolism , Lung Neoplasms/immunology , Lymphocyte Subsets/physiology , Lymphocytes, Tumor-Infiltrating/physiology , Neoplasm Proteins/metabolism , Adult , Aged , Colorectal Neoplasms/pathology , Female , Flow Cytometry , Humans , Immunotherapy , Lung Neoplasms/secondary , Lung Neoplasms/therapy , Lymphocyte Activation , Lymphocytes, Tumor-Infiltrating/immunology , Male , Middle AgedABSTRACT
T cell-mediated mechanisms are important in the defense against solid organ tumors. Why some tumors are more heavily infiltrated by T cells than others is poorly understood but is likely to depend upon adhesive interactions between circulating lymphocytes and tumor endothelium. In support of this hypothesis, the present study shows that primary human hepatocellular carcinomas (HCC) are more heavily infiltrated with T cells than colorectal hepatic metastases (CHM), and that their tumor vessels express high levels of several adhesion molecules. In HCC, an intense T cell infiltrate is observed within the tumor associated with strong expression of ICAM-1 and vascular adhesion protein-1 (VAP-1) on tumor endothelium. In contrast, fewer T cells infiltrated CHM and these tumors have little ICAM-1 and no detectable VAP-1 or VCAM-1 on tumor endothelium. T cells infiltrating both tumors are LFA-1 and very late Ag (VLA)-4 high. In vitro tissue-binding studies demonstrated that T cells bound readily to tumor endothelium in HCC, and Abs to ICAM-1, VAP-1, and to a lesser extent VCAM-1 could inhibit this binding. VAP-1 supported sialic acid-dependent adhesion under shear stress, suggesting that VAP-1 and ICAM-1 mediate, respectively, tethering and firm adhesion. In contrast, very few T cells bound to tumor vessels in CHM. Thus our data suggest that the VAP-1/VAP-1 receptor and ICAM-1/LFA-1 pathways are important in the recruitment of T cells to HCC. The strong expression of VAP-1 on tumor endothelium distinguishes HCC from CHM and supports our previous hypothesis that VAP-1 is an important hepatic endothelial adhesion molecule.