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Biomedical subjects

A Stoppacciaro

Publications and source records attributed to A Stoppacciaro.

At least 55 records · Page 3Linked to original sources
PubMed1997

Genetic modification of a carcinoma with the IL-4 gene increases the influx of dendritic cells relative to other cytokines.

Tumor cells genetically modified with certain cytokine genes gain immunogenic properties that allow the development of systemic anti-tumor immunity. Whether different cytokines may influence infiltration of transduced tumors by dendritic cells (DC) has not been investigated. Therefore, we analyzed the C26 murine colon carcinoma genetically modified to release interleukin (IL)-2, IL-4, IL-12, granulocyte colony-stimulating-factor (CSF) or granulocyte-macrophage (GM)-CSF for immunostaining with the monoclonal antibody NDLC145 recognizing the DEC205 determinant which, on tumor sections, is virtually restricted to DC. Infiltrating leukocytes were also characterized for expression of co-stimulatory molecules like CD54, CD86 and major histocompatibility complex class II. The intratumoral DC content was dependent on the type of transduced cytokines with C26/IL-4 being the most abundant in DEC205+ cells. The effect of IL-4 in recruiting DC did not depend on the type of tumor since it was confirmed in the TSA mammary carcinoma. In comparison with C26/GM-CSF, C26/IL-4 had more B7.2+ cells but less Ia+ cells. Furthermore, the hypertrophic skin overlaying tumors producing GM-CSF showed numerous Langerhans cells stained by NDLC145 and the draining lymph nodes showed abundance and paucity of DC in C26/GM-CSF and C26/IL-4, respectively. When injected into the ear pinna, C26/GM-CSF stimulated, whereas C26/IL-4 inhibited DC-mediated priming of delayed-type hypersensitivity reaction by 2,4-dinitro-1-fluorobenzene. These findings prove that transduced cytokines differently influence DC recruitment at the tumor site and DC function in nearby tissues. Along with the other leukocytes and their secondary produced cytokines, DC create an environment in which T cells can be differently modulated. Such a phenomenon may have implications on genetic modification of tumor cells to be used as cancer vaccine.

Animals↗
PubMed1997

Kaposi's sarcoma cells express the macrophage-associated antigen mannose receptor and develop in peripheral blood cultures of Kaposi's sarcoma patients.

The mannose receptor (MR) is a surface 175-kd C-type lectin expressed by macrophages and dendritic cells. MR is involved in removal of effete cells, phagocytosis of mannose-coated particles, pinocytosis, and antigen presentation. Expression of MR was investigated in 17 biopsies of Kaposi's sarcoma (3 AIDS KS, 13 classical KS, and 1 transplant-associated KS) using three anti-MR monoclonal antibodies (3.29, D547, and PAM1). Immunostaining for MR was detected in 94 +/- 7% KS cells with spindle morphology. In normal tissues, MR was expressed by sinus-lining cells of spleen and lymph nodes, but it was not detected in endothelial cells lining normal hematic and lymphatic vessels, hemangioma, hemangioendothelioma, and lymphangioma. Expression of MR in KS cells prompted us to investigate the possibility that they derive from a circulating precursor cell. Peripheral blood mononuclear cells from 16 patients with KS (10 classical, 1 transplanted, and 5 AIDS) were cultured in PHA-conditioned medium for 10 to 14 days. Confluent monolayers of adherent spindle cells were detected in 8 of 11 classical KS, in 5 of 5 AIDS KS patients, and in 0 of 34 control patients. Peripheral-blood-derived KS-like cells were characterized by co-expression of macrophage and endothelial antigens being positive for CD45 (60%), CD68 (98%), MR (70%), CD14 (25%), VE-cadherin (70%), and von Willebrand factor (10%). When the immunophenotype of peripheral-blood-derived adherent cells was compared with that of KS spindle cells of tissue biopsies, it was found that both cell types are VE-cadherin+/MR+/CD68+, that peripheral-blood-derived spindle cells are CD34- and are less frequently stained for CD31 and von Willebrand factor, and that lesional KS cells do not express the leukocyte markers CD45 and CD18. Our findings are consistent with the possibility that KS lesions derive from tissue accumulation and local proliferation of a special subset of macrophages with endothelial features the normal counterpart of which are the sinus-lining cells of spleen and lymph nodes.

Acquired Immunodeficiency Syndrome↗
PubMed1997

Characterization of MEC 14.7, a new monoclonal antibody recognizing mouse CD34: a useful reage for identifying and characterizing blood vessels and hematopoietic precursors.

Endothelial cell-specific molecules are potential targets for new therapeutic strategies in the control of inflammatory reactions, immune responses and neoangiogenesis. We describe the production and characterization of MEC 14.7, a monoclonal antibody directed to murine endothelial cells recognizing a glycosylated protein with an apparent molecular mass of about 100 kDa in cultured endothelioma cell lysate and about 80 kDa in lung lysate. MEC 14.7 antigen was selectively expressed by the endothelium in vivo, particularly in small vessels and neoformed capillaries and by developing vascular structures in embryonal bodies. Deglycosylation of the molecule with neuraminidase, O- and N-glycanase showed that the MEC 14.7 epitope is neuraminidase-sensitive. MEC 14.7 antigen was purified from lung lysates by chromatographic techniques, and sequenced internal peptides indicated it was identical with murine CD34. Thus the apparent molecular mass of CD34 is heterogeneous, depending on the glycosylation state in the different cell types. Immunomagnetic isolation and culture of MEC 14.7-positive bone marrow cells showed that this antibody recognizes hematopoietic progenitors (particularly myelomonocytic) and can be used in murine models of bone marrow reconstitution.

Amino Acid Sequence↗
PubMed1996

Amount of interleukin 12 available at the tumor site is critical for tumor regression.

The C26 colon carcinoma is resistant to systemic recombinant interleukin 12 (rIL-12) therapy. Transduction of C26 with genes encoding the two subunits of murine IL-12 to release 30-80 pg/ml resulted in delayed tumor onset after injection of 5 x 10(4) cells into syngeneic BALB/c mice and in 40% tumor regression after injection into CD4-depleted mice. Here, we analyzed the activity of rIL-12 (1 microgram/day) against C26 grown into CD4-depleted mice. Like in mice given injections of interleukin 12 (IL-12) gene-transduced C26 cells, depletion of CD4+ cells led to tumor regression in 6 of 14 mice, and immumocytochemical characterization of tumor-infiltrating leukocytes showed abundant infiltration by CD8+ T cells and asialoGM1+ natural killer cells, which were scanty in tumors from nondepleted mice. On the basis of the percentage of tumor regression and leukocyte infiltration we can conclude that, in the C26 system, systemic rIL-12 (1 pmicrogramg/day) produces the same results as 30-80 pg/ml IL-12 released at the tumor site. A new polycistronic retroviral vector was then used to increase the amount of IL-12 produced by C26-transduced cells. C26 cells releasing 5 ng/ml IL-12, nearly 100 times more than the above-mentioned transduced cells, were tumorigenic in less than 50% of the mice given injections of 5 x 10(4) cells. In mice given injections of 5 x 10(5) cells, an initial tumor take of 100% followed by a complete tumor regression. Tumor regression was associated with infiltration of CD8+ and asialoGM1+ cells, and mice that remained tumor free were immune to a rechallenge of nontransduced C26 cells. The results indicate that the amount of IL-12 made available at the tumor site may determine both the type and number of infiltrating leukocytes and the events leading to tumor regression as well as it may overcame host immunosuppression.

Adenocarcinoma↗
PubMed1996

HOXB7 constitutively activates basic fibroblast growth factor in melanomas.

Homeobox (HOX) genes control axial specification during mammalian development and also regulate skin morphogenesis. Although selected HOX genes are variably expressed in leukemias and kidney and colon cancer cell lines, their relationship with the neoplastic phenotype remains unclear. In both normal development and neoplastic transformation, HOX target genes are largely unknown. We investigated the expression and function of HOXB cluster genes in human melanoma. The HOXB7 gene was constitutively expressed in all 25 melanoma cell lines and analyzed under both normal and serum-starved conditions, as well as in in vivo primary and metastatic melanoma cells; conversely, HOXB7 was expressed in proliferating but not quiescent normal melanocytes. Treatment of melanoma cell lines with antisense oligomers targeting HOXB7 mRNA markedly inhibited cell proliferation and specifically abolished expression of basic fibroblast growth factor (bFGF) mRNA. Band shift and cotransfection experiments showed that HOXB7 directly transactivates the hFGF gene through one out of five putative homeodomain binding sites present in its promoter. These novel findings indicate a key role for constitutive HOXB7 expression in melanoma cell proliferation via bFGF. The results also raise the possibility that growth factor genes are critical HOX target genes in other developmental and/or neoplastic cell systems.

Antisense Elements (Genetics)↗
PubMed1996

Hypoxic tumor cell death and modulation of endothelial adhesion molecules in the regression of granulocyte colony-stimulating factor-transduced tumors.

C-26 colon adenocarcinoma cells transduced with the granulocyte colony-stimulating factor (G-CSF) gene form large tumors when injected into sublethally irradiated mice. These tumors regress when leukocyte function is reconstituted. Electron microscopy and immunocytochemical analysis of regressing C-26/G-CSF nodules indicates that tumor destruction is due mainly to hypoxia resulting from the functional loss of tumor vasculature and is only marginally due to direct cytolysis. Desegregation of basal lamina, cell swelling, and loss of junctions characterized the vessels within regressing tumors. Tumor cells were necrotic or filled with lipid vacuoles regardless of the distance from nearby vessels. Damage of tumor vasculature was dependent on the infiltrating leukocytes and the cytotoxic cytokines they produced. Locally produced interleukin-1 and tumor necrosis factor-alpha (TNF-alpha) induced vascular cellular adhesion molecule-1 (VCAM-1) and E-selectin on tumor vessels. Treatment with monoclonal antibodies to interferon-gamma (IFN-gamma) or TNF-alpha blocked tumor regression by inhibiting VCAM-1 and E-selectin expression on tumor-associated endothelial cells resulting in a reduced number of infiltrating leukocytes. Thus, C-26/G-CSF tumor regression presents features typical of hemorrhagic necrosis that occurs through the cytokines produced by infiltrating leukocytes in response to G-CSF.

Adenocarcinoma↗
PubMed1995

TCL1 oncogene activation in preleukemic T cells from a case of ataxia-telangiectasia.

The TCL1 oncogene on human chromosome 14q32.1 is involved in chromosome translocations [t(14;14)(q11;q32.1) and t(7;14)(q35;q32.1)] and inversions [inv14(q11;q32.1)] with TCR alpha/beta loci in T-cell leukemias, such as T-prolymphocytic (T-PLL). It is also involved in T-acute and -chronic leukemias arising in cases of ataxia-telangiectasia (AT), an immunodeficiency syndrome. Similar chromosomal rearrangements occur also in the clonally expanded T cells in AT patients before the appearance of the overt leukemia. We have analyzed the expression of TCL1 mRNA and protein in peripheral blood lymphocytes (PBLs) from four AT cases and from healthy controls. We found that the TCL1 gene was overexpressed in the PBLs of an AT patient with a large clonal T-cell population exhibiting the t(14;14) translocation but not in the lymphocytes of the other cases. Fluorescence in situ hybridization of the TCL1 genomic locus to lymphocyte metaphases from the AT patient with the T-cell clonal expansion showed that the breakpoint of the t(14;14) translocation lies within the TCL1 locus and is accompanied by an inverted duplication of the distal part of chromosome 14. These data indicate that TCL1 is activated in preleukemic clonal cells as a consequence of chromosome translocation involving sequences from the TCR locus at 14q11. Deregulation of TCL1 is the first event in the initiation of malignancy in these types of leukemias and represents a potential tool for clinical evaluation.

Adolescent↗
PubMed1995

An in vivo model to compare human leukocyte infiltration in carcinoma xenografts producing different chemokines.

In this study we tested whether the pattern of cytokines expressed by human carcinomas could account for a different in vivo recruitment of leukocyte subpopulations as a part of the anti-tumor immune response. Two carcinoma cell lines, SK-OV-3 ovary carcinoma and CALU-3 lung carcinoma, were analyzed by reverse transcriptase-polymerase chain reaction (RT-PCR), immunofluorescence and ELISA for the expression and in vitro production of cytokines with chemotactic, proinflammatory and growth-stimulating activity. Although both cell lines displayed a constitutive expression of granulocyte colony-stimulating factor (G-CSF), granulocyte macrophage-CSF (GM-CSF), M-CSF, interleukin (IL-) 1 alpha and IL-8, only CALU-3 cell line expressed IL-10, RANTES (Regulated upon Activation, Normal T Expressed and Secreted) and monocyte-activating protein (MCP)-1. MCP-1 and IL-8 were detected by immunohistochemistry on sections from tumors xenografted in nude mice. To analyze whether the tumor-released cytokines modulate leukocytes in tumor infiltration, we studied the distribution of human peripheral blood leukocytes injected in the proximity of SK-OV-3 and of CALU-3 tumor xenografts. While SK-OV-3 was unable to recruit human leukocytes and appeared to be barely infiltrated by murine CD45+ cells, CALU-3 appeared to be rapidly and heavily infiltrated by human leukocytes which induced tumor necrosis within 18-24 hr.

Adenocarcinoma↗
PubMed1995

CD4 T cells inhibit in vivo the CD8-mediated immune response against murine colon carcinoma cells transduced with interleukin-12 genes.

Retroviral-mediated cytokine gene transfer into tumor cells is a highly effective way of inducing tumor inhibition and immunity. We analyzed the tumorigenicity of C-26 murine colon carcinoma cells transduced with genes encoding the two subunits of murine interleukin-12 (IL-12) in a polycistronic retroviral vector and selected for resistance to G418 and for IL-12 production (30-80 pg/ml). BALB/c mice injected s.c., i.v. and intrasplenically with C-26/IL-12 cells from three different IL-12-producing clones showed delayed tumor onset as compared with mice injected with control NeoR-transduced or parental tumor cells. Although C-26/IL-12 tumor-bearing mice eventually died of lung metastasis, their survival time was twice as long as that of mice injected with control cells. In experiments with mice selectively depleted of natural killer (NK) cells before tumor cell injection, the time of tumor onset and survival of mice injected with C-26/IL-12 s.c. and i.v., respectively, was reduced. CD8+ T cell depletion had no effect on latency or survival, whereas removal of CD4+ T cells led to C-26/IL-12 tumor regression in about 40% of mice. Histological and immunocytochemical characterization of leukocytes infiltrating C-26/IL-12 tumors showed only slight infiltration with few T cells in non-depleted mice but abundant infiltration by CD8+ T cells and asialo-GM1+ NK cells in tumors of mice depleted of CD4+ T cells. The lack of CD8+ T cell infiltration is not due to a CD4-mediated suppression of their activation because irradiated C-26/IL-12 cells primed for the induction of a strong cytotoxic T lymphocyte response against C-26 parental cells and induced CD8+ effector cells that protected against C-26/IL-12 in a Winn assay. Rather, the results suggest that, although C-26/IL-12 cells injected in vivo stimulate both NK and CD8+ T cells, tumor infiltration by the latter is inhibited by CD4+ T cells.

Animals↗
PubMed1995

The lymphoepithelial organization of the tonsil: an immunohistochemical study in chronic recurrent tonsillitis.

Interactions between leukocytes and crypt epithelium were extensively investigated in 12 cases of chronic recurrent tonsillitis, using immunohistochemistry and cytofluorimetric analysis of cell suspensions. Intraepithelial leukocytes are a mixed cell population composed of 50 per cent CD20-positive B lymphocytes, 40 per cent T lymphocytes with a 2.7 CD4/CD8 ratio, and 10 per cent CD68-positive macrophages. About 4 per cent of intraepithelial leukocytes are proliferating cells, as indicated by Ki-67 staining. Leukocyte infiltration is associated with expression on epithelial cells of the adhesion molecules ICAM-1 and VCAM-1. Crypt epithelium is supported by a basement membrane showing frequent interruptions and connected with the reticular stroma of the lymphoid tissue, which was stained for fibronectin, tenascin, collagen, and laminin. Extracellular matrix (ECM) distribution was correlated with integrin expression on B and T lymphocytes. It was found that the ECM was arranged differently in the follicles and in the extrafollicular area and that B and T lymphocytes exhibited different patterns of integrin expression.

Adolescent↗
PubMed1995

Local immunotherapy of superficial bladder cancer by intravesical instillation of recombinant interleukin-2.

Local immunotherapy of superficial bladder cancer by endovesical administration of recombinant interleukin-2 (rIL-2) was investigated in a phase I-II study. Twenty-five patients with Ta-T1, N0, M0, G1-G2 transitional cell carcinoma of the bladder received an induction course of rIL-2 (10 daily instillations) with the tumor in place using an interpatient dose escalation scheme from 3 to 18 x 10(6) IU/day. Seven to fourteen days after the end of the induction course, the objective clinical response was evaluated and transurethral resection of the bladder tumor was carried out. Four maintenance courses (10 daily instillations) were started 1 month after surgery and carried out every 4 months at a dose of 6 x 10(6) IU/day in all patients. Follow-up visits were scheduled bimonthly during treatment and then every 6 months. No evidence of laboratory, local or systemic toxicity was observed even at the highest rIL-2 dosages. Induction of a local inflammatory response could be demonstrated at the tumor site after intravesical rIL-2 treatment. The significant reduction in tumor diameters observed in some patients may be interpreted as a sign of the biological activity of this rIL-2 regimen. Further exploratory work is required to evaluate the clinical efficacy of this immunotherapeutic approach.

Administration, Intravesical↗
PubMed1994

Inhibition of the metastatic spread and growth of B16-BL6 murine melanoma by a synthetic matrix metalloproteinase inhibitor.

The synthetic matrix metalloproteinase inhibitor batimastat was tested for its ability to inhibit growth and metastatic spread of the B16-BL6 murine melanoma in syngeneic C57BL/6N mice. Intraperitoneal administration of batimastat resulted in a significant inhibition in the number of lung colonies produced by B16-BL6 cells injected i.v. The effect of batimastat on spontaneous metastases was examined in mice inoculated in the hind footpad with B16-BL6 melanoma. The primary tumor was removed surgically after 26-28 days. Batimastat was administered twice a day from day 14 to day 28 (pre-surgery) or from day 26 to day 44 (post-surgery). With both protocols, the median number of lung metastases was not significantly affected, but there was a significant reduction in the weight of the metastases. Finally, the effect of batimastat was examined on s.c. growth of B16-BL6 melanoma. Batimastat administered daily, starting at day of tumor transplantation, resulted in a significant growth delay, whereas treatment starting at advanced stage tumor only reduced tumor growth marginally. Our results indicate that a matrix metalloproteinase inhibitor can not only prevent the colonization of secondary organs by B16-BL6 cells but also limit the growth of solid tumors.

Animals↗
PubMed1994

Co-expression of endothelial cell and macrophage antigens in Kaposi's sarcoma cells.

The histopathogenesis of Kaposi's sarcoma (KS) was investigated using immunocytochemistry in 12 skin biopsies obtained from two AIDS patients, nine patients with the classic form, and one organ-transplant patient. KS cells revealed a peculiar antigenic profile, being characterized by co-expression of endothelial and macrophage markers. KS cells were stained for von Willebrand factor (vWF); for vascular endothelial (VE) cadherin, an endothelial specific adhesion molecule; and for PECAM/CD31. In addition, they expressed the macrophage antigens PAM-1, CD68, and CD14, and were positive for vitronectin receptor and alpha-1,5,6/beta-1 integrins. KS cells were weakly stained for ICAM-1 in 6 of 12 cases and were negative for VCAM-1 and E-selectin.

Acquired Immunodeficiency Syndrome↗
PubMed1994

Expression of intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 in undifferentiated nasopharyngeal carcinoma (lymphoepithelioma) and in malignant epithelial tumors.

Immunoreactivity for intercellular adhesion molecule-1 (ICAM-1) and for vascular cell adhesion molecule-1 (VCAM-1), two adhesion molecules of the immunoglobulin (Ig) superfamily, was tested and measured on tissue sections from 16 undifferentiated nasopharyngeal carcinomas (U-NPC), 12 keratinizing squamous cell carcinomas (SCCs) of the head and neck region, and 54 malignant epithelial tumors of various origin. Neoplastic cells of all cases of U-NPC were diffusely and intensely stained for ICAM-1 and VCAM-1. Moreover, ICAM-1 messenger RNA (mRNA) and VCAM-1 mRNA were detected by Northern blot analysis of RNA extracts from two tumors. In the other epithelial tumors focal or diffuse staining for ICAM-1 was observed in 40 cases (66%), whereas reactivity for VCAM-1 was detected in a single case of metastatic undifferentiated carcinoma of unknown origin. The biopsy specimens of U-NPC showed variable infiltration by leukocytes, which were positive for the integrins lymphocyte function antigen-1 (LFA-1) and alpha-4/beta-1, the corresponding ligands for ICAM-1 and VCAM-1. The possibility that ICAM-1 and VCAM-1 on neoplastic cells may favor the intratumoral recruitment of leukocytes in a way similar to that occurring in crypt epithelium of the palatine tonsil is discussed.

Adolescent↗
PubMed1994

Different tumours, transduced with different cytokine genes as G-CSF and IL-2, show inhibition of tumour take through neutrophil activation but differ in T cell functions.

The immune effectors and the cellular mechanisms responsible for tumour rejection of two different tumours transduced with different cytokines have been characterized by immunocytochemistry and in situ hybridization. A colon (C-26) and a mammary (TS/A) adenocarcinoma engineered to release, respectively, 90 pg/ml of G-CSF (C-26/G-CSF) and 30 U or 6000 U of IL-2 (low B1.30 and high B4.6000 level of IL-2, respectively) were compared for the type of infiltrating leucocytes and for the repertoire of secondary cytokines produced by the leucocytes recruited at the tumour site. The results indicate that in both systems tumour rejection is associated with prominent infiltration of CD45+/RB6-8C5+ polymorphonuclear (PMN) leucocytes expressing mRNA for IL-1 alpha, IL-1 beta and TNF alpha. TS/A B1.30 and B4.6000 also showed a small proportion of infiltrating T lymphocytes, expressing IFN gamma and IL-4 mRNA, which were virtually absent in the C-26/G-CSF tumour. In mice injected with C-26/G-CSF cells after 600 rad irradiation, the tumours grew to about 1.5 cm and then regressed completely. During the regression phase, T lymphocytes were recruited within C-26/G-CSF, and the infiltrating leucocytes were similar, in terms of PMN/macrophages/T lymphocytes ratio, to those found during the memory response elicited by injection of a challenging dose of parental TS/A into mice pre-immunized with B1.30 IL-2-producing cells. The memory response was characterized by a CD4/CD8 ratio of 0.4 and by IFN-gamma and IL-4 mRNA expression, whereas the T lymphocytes present within regressing C-26/G-CSF were mostly CD4 (CD4/CD8 ratio of 2.1) and expressed IFN gamma mRNA only. The gene transfers of cytokines as different as G-CSF and IL-2 are able to inhibit tumour take through a similar anti-tumour immune response mostly due to non-specific effectors (PMN), thus resembling acute inflammation phenomena. Regression of C-26/G-CSF initially established in irradiated mice as well as rejection of TS/A tumour injected into B1.30 immunized mice are similar to a chronic infection but the immune reaction elicited by C-26/G-CSF has an impaired T-lymphocyte function.

Adenocarcinoma↗