北山 丈二

A unique subset of T cells that co-express NKR-P1, which is a lectin type of NK receptor and is thought to have a major role in triggering NK activity, has been identified. In mice, NK1.1 (mouse NKR-P1C)(+) T cells, called NKT cells, preferentially accumulate in the liver and bone marrow. They predominantly use invariant V alpha 14 chain VCR and phenotypically are CD4(+)CD8(-) or CD4(-)CD8(-) T cells. In this study, we analyzed, phenotypically and functionally, the NKR-P1A (analogue of murine NKR-P1C)(+) T cells resident: in the human liver. Here, we show that in complete contrast to the NKT cells in the mouse liver, the majority of NKR-P1A(+) T cells in the human liver are CD8(+) and their TCR repertoire is not skewed to V alpha 24 TCR, the homologue of murine V alpha 14 TCR. Almost all of the NKR-P1A(+) T cells in the human liver expressed CD69, suggesting that they were activated. Furthermore, the NKR-P1A(+) T cells in the human liver exhibited strong cytotoxicity against a variety of tumor cell lines including K562, Molt4 and some colonic adenocarcinoma cell lines.

Integrins play an important role in various lymphocyte functions. In this study, we isolated lamina propria lymphocytes (LPL) and tumor-infiltrating lymphocytes (Tn) from normal and malignant tissues in patients with colorectal cancer, and examined the expression of beta 1 and beta 2 integrins on these lymphocytes quantitatively with two-color flow cytometry, Both LPL and Tn expressed a lower level of common beta 1 chain (CD29) in CD4 and CD8 subpopulations than did peripheral blood lymphocytes (PBL). Among the associated alpha chains, the expression levels of alpha 1 (CD49a) and alpha 2 (CD49b) were slightly higher, whereas those of alpha 4 (CD49d) and alpha 6 (CD49f) were markedly reduced in LPL and Tn. No significant differences were observed in expressions of any beta 1 integrin chains between these two lymphocytes populations. Similarly, both alpha L (CD11a) and beta 2 (CD18) were down-regulated in TIL and LPL with CD8(+) cytotoxic phenotype, but not in these with CD4 phenotype. CD8(+) TIL expressed a slightly but significantly higher level of alpha L beta 2 than did CD8 LPL. CD8(+) LPL and CD8(+) TIL consistently showed significantly decreased binding to purified ICAM-1, VCAM-1 and HT29 colon cancer cells as compared with CD8(+) PBL. Although CD8(+) TIL showed a slightly higher level of adhesion to these substrates than did CD8(+) LPL, the level was much lower than that in PBL. The expression pattern and functional down-regulation of these integrins may be one of the reasons why TIL cannot eradicate the cancer cells in colorectal cancer.

Background: p21Waf1/Cip1 (p21), p27Kip1 (p27), p53, and Rb play critical roles in cell cycle regulation and may influence the clinical behavior of tumors. We examined whether their expression is useful to predict survival of patients with esophageal squamous cell carcinoma (ESC). Methods: Expression of p21, p27, p53, and Rb was studied by the immunohistochemical method in specimens from 62 patients with curatively resected ESC tumors and scored by a computerized image analysis system. Results: The median expression scores of p21, p27, p53, and Rb (14, 12, 27, and 50, respectively) were used as cut-off points to define low and high expression groups for each protein. The 5-year survival rate for the high p21 expression group was 68%; that for the low expression group was 31% (P = .0062). p27, p53, and Rb were not correlated with overall survival. When patients were categorized into four groups based on p21 expression level and lymph node involvement (pN), the survival curves were significantly different (P = .0017). Thus, patients without lymph node involvement but with low p21 expression had survival similar to that of patients with lymph node involvement and high p21 expression. Multivariate analysis showed that age (P = .0102), lymph node involvement (P = .0076), and p21 (P = .0276) were independent prognostic factors, Conclusions: Expression of p21 is an independent prognostic factor in curatively resected ESC. Definition of new subgroups of patients based on p21 expression may help to enhance the stratification of stage.

Integrins play an important role in various lymphocyte functions. In this study, tumor-infiltrating lymphocytes (TIL) were isolated from colorectal cancer tissues and the expression of beta 1 and beta 2 integrins on the TIL was quantitatively examined with two-color flow cytometry. In comparison with peripheral blood lymphocytes (PBL), TIL expressed a lower level of common beta 1 chain (CD29) in both CD4 and CD8 subpopulations. Among the associated a chains, the expressions of alpha 1 (CD49a) and alpha 2 (CD49b) were slightly higher in TIL than in PBL, whereas alpha 4 (CD-49d) and alpha 6 (CD49f) were markedly downregulated in TIL. Both alpha L (CD11a) and beta 2 (CD18) were reduced in CD8(+) TIL but not in CD4(+) TIL, TIL with the CD8(+) cytotoxic phenotype showed significantly decreased binding to purified intracellular adhesion molecules (ICAM)-1, and vascular adhesion cell molecule (VCAM)-1, and HT29 colon cancer cells, compared with the in counterparts in PBL, The peculiar expression pattern and functional down regulation of these integrins may explain why TIL in colorectal cancer cannot eradicate the malignant cells.

Background: The frequency of lymph node metastasis in mucosal gastric cancers 2-4 cm in diameter was low (three (1.3 per cent) of 234) in patients treated in this unit between 1966 and 1995. This study was a prospective report on local resection with lymphadenectomy for early gastric cancer. Methods: Eight patients with a single early gastric cancer underwent local resection with lymphadenectomy. The tumour was excised with a non-cancerous rim of approximately 2 cm. The extent of lymphadenectomy depended on tumour location. Intraoperative endoscopic examination and frozen-section analysis of the dissected nodes were used to determine the resection line and evaluate nodal status. Results: Mean operating time, blood loss and number of dissected nodes were 171 min, 87 ml and 8 respectively. There were no operative complications. Cancer invasion was confined to the mucosa in six tumours but two patients had minute submucosal invasion. The maximum diameter of the resected specimens was 10 cm and no nodal involvement was detected. No patient developed postgastrectomy syndrome. Conclusion: For selected patients with early gastric cancer, local resection with lymphadenectomy can provide a good quality of life without compromising cure rate.

It has been established that lymphocytes obtained from tumor-draining lymph nodes (DLN) are sensitized to the tumor antigen in vivo. Moreover, after being activated in vitro, these cells can be utilized for adoptive immunotherapy. In the present study, DLN cells, obtained from C57BL/6 mice with fibrosarcoma (MC-1), were activated and expanded with anti-CD3 monoclonal antibody followed by culture with recombinant interleukin-2 (rIL-2), These CD4(-) CD8(+) CD25(+) CD44(+) T-cells shelved specific antitumor efficacy to the pulmonary micrometastases of an autologous tumor, against which lymphokine-activated killer cells were ineffective; however, they did not show cytolytic activity in vitro. The supernatant, obtained by coculturing the activated DLN cells with MC-1 cells, exhibited the specific production of interferon-gamma (IFN-gamma) which was enhanced by rIL-2, The therapeutic effect of the activated DLN cells correlated with the specific IFN-gamma production better than with the cytolytic activity.

A technique, by which the root of the right gastric artery can be clearly exposed and the area of the right gastric artery can thus be completely dissected, is presented herein, In conventional procedures, the approach to the right gastric artery is performed from the lessor omentum, and a lymphadenectomy along the common hepatic artery is carried out after the division of the right gastric artery. Using the present technique, the common hepatic artery is exposed before the exposure of the right gastric artery and lymph node dissection is thus continued peripherally to the proper hepatic artery, During these procedures, the root of the right gastric artery arising from the hepatic artery can be easily identified. The lymphatic flow from the area of the right gastric artery to that of the common hepatic artery is transected using the conventional procedure, but not with the present technique. In addition, an en bloc dissection of the regional lymph nodes is also possible when using this procedure.

BACKGROUND. To evaluate the immunologic activity of regional lymph nodes, the phenotype of lymphocytes and the functional expression of cell adhesion molecules (CAMs) on lymph node lymphocytes (LNL-: uninvolved, LNL+: involved) were investigated in patients with gastrointestinal carcinoma. METHODS. The lymphocyte subpopulation and the expression of CD11a, CD44, and CD29 on CD4+ and CD8+ cells in peripheral blood lymphocytes (PBL), LNL-, and LNL+ derived from 37 patients with gastrointestinal carcinoma were studied. In addition, the adherence of CD8+ cells to ICAM-1 which reflects the adhesive function of CD11a, was examined, and changes in this adherence were studied by experimental coculture with cancer cells (DLD-1). RESULTS. Although there were no differences in the overall proportion of T cells between the groups, CD8+ cells and CD16+ cells were considerably diminished in LNL+. The expression of CD11a and CD29 on CD4+ and CD8+ cells was significantly lower in LNL than in PBL, whereas the expression of CD44 showed no significant differences. The expression levels of these CAMs were almost the same in LNL- and LNL+. Only CD11a expression on CD8+ cells in LNL+ was significantly lower than that in LNL- (P < 0.005). The adherence of CD8+ cells in LNL+ to ICAM-1 was lower than that in PBL and LNL-, and was extremely enhanced by experimental coculture with cancer cells (DLD-1). CONCLUSIONS. These data indicate that the functional expression of CD11a (LFA-1) on CD8+ T cells is suppressed in cancer-involved regional lymph nodes in patients with gastrointestinal carcinoma. (C) 1996 American Cancer Society.

The CC chemokines regulated on activation normal T expressed and secreted (RANTES) and monocyte chemotactic protein 3 (MCP-3), and the anaphylatoxin C5a, induce activation, degranulation, chemotaxis, and transendothelial migration of eosinophils. Adhesion assays on purified ligands showed differential regulation of beta 1 and beta 2 integrin avidity in eosinophils. Adhesiveness of VLA-4 (alpha 4 beta 1, CD29/CD49d) for vascular cell adhesion molecule 1 or fibronectin was rapidly increased but subsequently reduced by RANTES, MCP-3, or C5a. The deactivation of VLA-4 lead to cell detachment, whereas phorbol 12-myristate 13-acetate in duced sustained activation of VLA-4. In contrast, chemoattractants stimulated a prolonged increase in the adhesiveness of Mac-1 (alpha M beta 2, CD11b/CD18) for intercellular adhesion molecule 1, Inhibition by pertussis toxin confirmed signaling via G protein-coupled receptors, Chemoattractants induced transient, while phorbol 12-myristate 13-acetate induced sustained actin polymerization, Disruption of actin filaments by cytochalasins inhibited increases in avidity of VLA-4 but not of Mac-1, Chemoattractants did not upregulate a Mn2+-inducible beta 1 neoepitope defined by the mAb 9EG7, but induced prolonged expression of a Mac-1 activation epitope recognized by the mAb CBRM1/5. This mAb inhibited chemoattractant stimulated adhesion of eosinophils to intercellular adhesion molecule 1, Thus, regulation of VLA-4 was dependent on the actin cytoskeleton, whereas conformational changes appeared to be crucial for activation of Mac-1, To our knowledge, this is the first demonstration that physiological agonists, such as chemoattractants, can differentially regulate the avidity of a beta 1 and a beta 2 integrin expressed on the same leukocyte.

The effect of basic fibroblast growth factor (b-FGF), one of the commonest angiogenic factors in various cancer types, on lymphocyte adhesion and transmigration across the endothelial cell monolayer was investigated using human umbilical vein-derived endothelial cells (HUVEC) and type I collagen gel. Forty-eight h exposure of HUVEC with 2 ng/ml b-FGF significantly decreased the basal adhesion of lymphocytes to endothelial cells. The decrease ratio is further enhanced by the addition of shear stress in this assay system. When HUVEC was stimulated for the last 24 h with optimal conditions of recombinant interleukin 1 beta, the percentages of transmigration as well as adhesion were also decreased significantly by the presence of b-FGF. The expression of intercellular adhesion molecule 1 and vascular cell adhesion molecule 1 was downregulated by b-FGF exposure in both resting and activated conditions by recombinant interleukin 1 beta, supposedly the main reason for this phenomenon. The migrating cells across b-FGF-stimulated HUVEC contained a markedly lower percentage of CD4(+) T-cells than those across nontreated HUVEC, although the 4B4(+)/2H4(+) ratio in CD4(+) T-cell populations did not differ significantly. These facts suggest that the presence of b-FGF in the angiogenic area suppresses lymphocyte emigration, especially that of CD4(c) T-cells, and thus causes insufficient helper function in local immune response. This effect of b-FGF was possibly one of the critical mechanisms by which cancer cells escape from the host immune reactions in the angiogenic stage of tumor development.

We describe a 56-year-old woman with Stewart-Treves syndrome who had severe dyspnea from a pleural effusion caused by metastatic angiosarcoma in the right lung. Tumorinfiltrating lymphocytes (TIL) in the pleural effusion were cultured and expanded in vitro in the continuous presence of recombinant interleukin 2 with periodic stimulation by CD3 antibody. The expanded TIL were administered intrapleurally seven times at 1- to 4-week intervals in combination with intravenous infusion of recombinant interleukin 2. A panel of T-cell clones was also obtained from TIL. Immunotherapy dramatically improved the patient's dyspnea and pleural effusion. A CD4(+) T-cell clone and a CD8(+) T-cell clone established from TIL had specific cytotoxicity to the tumor cells.

The mechanisms of lysis of endothelial cells derived from human umbilical vein (HUVEC) by autologous lymphokine-activated killer (LAK) cells, generated from cord blood lymphocytes of the same donor, were investigated. Freshly isolated HUVEC as well as HUVEC cultured for several passages were efficiently lysed by autologous LAK cells, and their susceptibility to the LAK cells was almost the some as that of allogenic HUVEC. Complement-depletion experiments revealed that the lysis was mainly dependent on CD16(+) natural killer (NK) LAK cells. Pretreatment of HUVEC with recombinant interferon gamma (rIFN gamma) for 24 h made them resistant to lysis by autologous LAK cells, while pretreatment with either rIL-1 beta. rTNF alpha or acidic or basic fibroblast growth factor did not alter the lyric sensitivity of HUVEC. The resistance of rIFN gamma-treated HUVEC was specific to lysis by CD16(+) NK LAK cells, and their lysis by CD3(+) T-LAK cells was not significantly altered. Moreover, in comparison with control HUVEC or rIL-1 beta-treated HUVEC, rlFN beta-treated HUVEC had a significantly less potent inhibitory effect on the lysis of untreated HUVEC, when used as an unlabeled target. This suggests that rIFN gamma treatment may down-regulate the recognition of some molecules on HUVEC by rIL-2-activated NK cells. These data suggest that damage of the endothelium during LAK therapy is mainly dependent on LAK cells with a NK phenotype that can specifically recognize a certain molecule on autologous endothelial cells.

In six patients with advanced pancreatic carcinoma, TIL and tumour-draining lymphocytes (TDL) were isolated from primary pancreatic tumour and regional lymph nodes. In comparison with TDL and peripheral blood lymphocytes (PBL), TIL contained a comparatively higher percentage of TCR gammadelta+ cells, although they were still a small fraction. By 2 weeks culture with rIL-2 and immobilized OKT-3 antibody, the TCR gammadelta + cells in TIL were preferentially expanded at the early culture periods, although it was temporary. In four cases, the TCR gammadelta+ and CD8+ TCR alphabeta+ TIL were separated by negative sorting using flowcytometry. All the TCR gammadelta+ TIL were CD4-, CD8- (double negative), and one of the TIL lines was mostly composed of deltaTCS1+ cells, while the others were deltaTCS1-. In comparison with CD8+ TCR alphabeta+ TIL, all the TCR gammadelta+ TIL exhibited much stronger lytic activity against freshly isolated autologous pancreatic cancer cells. However, all the gammadelta+ TIL also exhibited a strong non-MHC-restricted cytotoxicity, and there was no correlation between the lytic pattern and the percentage of deltaTCS1+ cells. These data suggest that the TCR gammadelta+ T cells can proliferate vigorously in a certain condition, and if successfully expanded in vitro they might be helpful material for effective adoptive immunotherapy.