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A Stoppacciaro

Publications and source records attributed to A Stoppacciaro.

At least 37 records · Page 2Linked to original sources
PubMed1999

Heat shock protein 70 induced during tumor cell killing induces Th1 cytokines and targets immature dendritic cell precursors to enhance antigen uptake.

Previously, we reported that killing tumor cells in vivo with the HSV thymidine kinase/ganciclovir system generates potent antitumor immunity, determined in part by the mechanism by which the cells die and by the levels of inducible heat shock protein (hsp) expression induced during the process of cell death. Here, we show that induction of hsp70 expression induces an infiltrate of T cells, macrophages, and predominantly dendritic cells (DCs) into the tumors as well as an intratumoral profile of Th1 cytokine expression (IFN-gamma, TNF-alpha, and IL-12) and enhances immunogenicity via a T cell-mediated mechanism. In addition, the protection conferred by hsp70 is both tumor and cell specific. We also demonstrate that hsp70 targets immature APC to make them significantly more able to capture Ags. This is likely to optimize cross-priming of the infiltrating APC with tumor Ags, which are simultaneously being released by the dying cells. In addition, using an Myc epitope-tagged hsp70 expression vector, we present evidence that hsp70 released from dying tumor cells is taken up directly into DCs and may, therefore, be involved in direct chaperoning of Ags into DCs. Taken together, our data suggest that hsp70 induction serves to signal the immune system of the presence of an immunologically relevant (dangerous) situation against which an immune reaction should be raised.

Animals↗
PubMed1999

Dendritic cells infiltrating tumors cotransduced with granulocyte/macrophage colony-stimulating factor (GM-CSF) and CD40 ligand genes take up and present endogenous tumor-associated antigens, and prime naive mice for a cytotoxic T lymphocyte response.

We transduced BALB/c-derived C-26 colon carcinoma cells with granulocyte/macrophage colony-stimulating factor (GM-CSF) and CD40 ligand (CD40L) genes to favor interaction of these cells with host dendritic cells (DCs) and, therefore, cross-priming. Cotransduced cells showed reduced tumorigenicity, and tumor take was followed by regression in some mice. In vivo tumors were heavily infiltrated with DCs that were isolated, phenotyped, and tested in vitro for stimulation of tumor-specific cytotoxic T lymphocytes (CTLs). BALB/c C-26 carcinoma cells express the endogenous murine leukemia virus (MuLV) env gene as a tumor-associated antigen. This antigen is shared among solid tumors of BALB/c and C57BL/6 mice and contains two epitopes, AH-1 and KSP, recognized in the context of major histocompatibility complex class I molecules H-2Ld and H-2K(b), respectively. DCs isolated from C-26/GM/CD40L tumors grown in (BALB/c x C57BL/6)F1 mice (H-2d x b) stimulated interferon gamma production by both anti-AH-1 and KSP CTLs, whereas tumor-infiltrating DCs (TIDCs) of BALB/c mice stimulated only anti-AH-1 CTLs. Furthermore, TIDCs primed naive mice for CTL activity as early as 2 d after injection into the footpad, whereas double-transduced tumor cells required at least 5 d for priming; this difference may reflect direct DC priming versus indirect tumor cell priming. Immunohistochemical staining indicated colocalization of DCs and apoptotic bodies in the tumors. These data indicate that DCs infiltrating tumors that produce GM-CSF and CD40L can capture cellular antigens, likely through uptake of apoptotic bodies, and mature in situ to a stage suitable for antigen presentation. Thus, tumor cell-based vaccines engineered to favor the interaction with host DCs can be considered.

Animals↗
PubMed1999

Thrombospondin-1 is a mediator of the neurotypic differentiation induced by EGF in thymic epithelial cells.

Thymic epithelial cell component originates from cranial neural crest as well as from endoderm and ectoderm of the third pharyngeal pouch and branchial cleft. Epidermal growth factor (EGF) has been previously shown to play a crucial role in directing thymic epithelial cells toward a neural-oriented cell fate. To identify genes that are involved in the EGF-induced neurotypic differentiation of the thymic stroma-derived TC-1S cell line, we studied EGF-treated and untreated cells by RNA fingerprinting PCR-based differential screening. We obtained 23 distinct sequences including 18 known genes and 5 sequences previously unreported, which are currently under characterization. Here, we describe the involvement of one of the isolated genes, the thrombospondin-1, as a mediator of the neurotypic differentiation induced by EGF in TC-1S cells. We show that thrombospondin-1 mRNA and protein levels are increased by EGF. Moreover, exogenous thrombospondin-1 is able to enhance the outgrowth of neurite-like processes as well as the expression of neurofilaments and neural cell adhesion molecule in TC-1S cells. These observations suggest that the up-regulation of thrombospondin-1 synthesis induced by EGF contributes to the differentiation choice of thymic epithelial cells toward a neural fate, reminiscent of their neural crest origin.

Animals↗
PubMed1999

Fas/Fas ligand-driven T cell apoptosis as a consequence of ineffective thyroid immunoprivilege in Hashimoto's thyroiditis.

Hashimoto's thyroiditis (HT) is a chronic autoimmune disease resulting from Fas-mediated thyrocyte destruction. Although autocrine/paracrine Fas-Fas ligand (FasL) interaction is responsible for thyrocyte cell death during the active phases of HT, the role of infiltrating T lymphocytes (ITL) in this process is still unknown. Therefore, we investigated the expression and function of Fas and FasL in ITL. All ITL expressed high levels of Fas and CD69, an early marker of T cell activation associated with functional Fas expression in T cells in vivo. In contrast to thyrocytes that were found to produce high levels of FasL, ITL did not express significant amounts of FasL, suggesting that ITL are not directly involved in thyrocyte destruction. The analysis of ITL purified from HT thyroids showed that ITL were massively killed by Fas crosslinking and that a considerable number (24-36%) underwent spontaneous apoptosis within 36 h of culture. Accordingly, in situ TUNEL (terminal deoxynucleotidyl transferase dUTP nick end labeling) staining revealed that a significant number (10-15%) of ITL in proximity to FasL-producing thyroid follicles were apoptotic. Moreover, virtually all ITL in proximity to thyroid follicles were preapoptotic, as they expressed high levels of GD3 ganglioside, a killer glycolipid responsible for the generation of irreversible apoptotic signals that accumulate in hematopoietic cells shortly after Fas crosslinking. These data demonstrate that ITL are not directly involved in thyrocyte cell death during HT, suggesting that autocrine/paracrine Fas-FasL interaction is a major mechanism in autoimmune thyrocyte destruction.

Adult↗
PubMed1999

Expression of EDA/EDB isoforms of fibronectin in papillary carcinoma of the thyroid.

Cellular fibronectins containing the extracellular domain A or B (EDA and EDB) are particularly abundant in fetal and neoplastic tissues. The presence of EDA and EDB was investigated in 28 cases of papillary carcinoma of the thyroid using IST-9 and BC-1 monoclonal antibodies. Immunostaining for EDA and EDB was detected in tumour stroma, in tumour basement membranes, and in tumour blood vessels. EDA was present in 27 of the 28 cases, in 20 of which more than 75 per cent of the tumour stroma was stained. Immunostaining for EDB was detected in 23 of the 28 cases and was less pronounced than that for EDA, being present in less than 25 per cent of the tumour stroma in most cases. Reactivity for EDA/EDB was not observed in the adjacent normal thyroid in any of the cases investigated. In a group of 20 non-papillary tumours, immunostaining for EDA was present in the stroma of three follicular carcinomas (one minimally and two widely invasive), one medullary carcinoma, and 5 of 16 follicular adenomas; expression of EDB was more restricted, being present in only the two cases of widely invasive follicular carcinoma. The presence of EDA and EDB was not correlated with the extent of fibrosis or the degree of tumour cell differentiation. Immunoreactivity was already present in microcarcinomas. These observations raise the possibility that the production of oncofetal fibronectins is an important step in papillary carcinoma tumourigenesis, perhaps facilitating adhesion and spreading of tumour cells.

Carcinoma, Papillary↗
PubMed1999

Hepatocyte growth factor (HGF) stimulates tumour invasiveness in papillary carcinoma of the thyroid.

The present study has investigated the functional role of the Met receptor in primary cultures of 20 papillary carcinomas and of normal thyroid cells obtained from the same patients. Normal and tumour cells grew as adherent cells, formed a confluent monolayer after 10-20 days, had epithelial morphology, and were immunoreactive for cytokeratin, vimentin, and thyroglobulin. The potential effect of hepatocyte growth factor (HGF) on cell invasiveness was investigated in Boyden chambers, using a nucleopore filter coated with Matrigel as the barrier and HGF as the chemoattractant. Tumour cells of five out of seven cases of papillary carcinoma were more responsive to HGF than the corresponding normal cells in terms of the number of migrated cells per mm(2). Involvement of the Met receptor in the HGF-induced migratory response was suggested by the observation that the agonistic anti-Met monoclonal antibody (MAb) DO-24 was equally effective. HGF did not affect the proliferative activity of thyroid cells. Under the same experimental conditions, 10 per cent fetal bovine serum (FBS) induced a two-fold increase in [(3)H]thymidine incorporation into normal cells and tumour cells. These findings are consistent with the possibility that HGF plays a crucial role in determining the invasiveness of tumour cells in papillary carcinoma of the thyroid.

Antibodies, Monoclonal↗
PubMed1998

The chemokine receptor CCR8 is preferentially expressed in Th2 but not Th1 cells.

In this paper we report on the cloning and characterization of mouse CCR8. Like its human homologue, it is predominantly expressed in the thymus. In the periphery, murine CCR8 mRNA was found most abundantly expressed in activated Th2-polarized cells and in NK1.1+ CD4+ T cells. Human CCR8 is also preferentially expressed in human Th2-polarized cells and clones. This pattern of expression suggests that CCR8 is part of a Th2-specific gene expression program. The CCR8 ligands I-309 and its mouse homologue T cell activation gene 3 (TCA-3) are potent chemoattractants for Th2-polarized cells. Taken together, these observations strongly suggest that CCR8 plays a role in the control of Th2 responses, and may represent a potential target for treatment of allergic diseases.

Amino Acid Sequence↗
PubMed1998

Junctional adhesion molecule, a novel member of the immunoglobulin superfamily that distributes at intercellular junctions and modulates monocyte transmigration.

Tight junctions are the most apical components of endothelial and epithelial intercellular cleft. In the endothelium these structures play an important role in the control of paracellular permeability to circulating cells and solutes. The only known integral membrane protein localized at sites of membrane-membrane interaction of tight junctions is occludin, which is linked inside the cells to a complex network of cytoskeletal and signaling proteins. We report here the identification of a novel protein (junctional adhesion molecule [JAM]) that is selectively concentrated at intercellular junctions of endothelial and epithelial cells of different origins. Confocal and immunoelectron microscopy shows that JAM codistributes with tight junction components at the apical region of the intercellular cleft. A cDNA clone encoding JAM defines a novel immunoglobulin gene superfamily member that consists of two V-type Ig domains. An mAb directed to JAM (BV11) was found to inhibit spontaneous and chemokine-induced monocyte transmigration through an endothelial cell monolayer in vitro. Systemic treatment of mice with BV11 mAb blocked monocyte infiltration upon chemokine administration in subcutaneous air pouches. Thus, JAM is a new component of endothelial and epithelial junctions that play a role in regulating monocyte transmigration.

Amino Acid Sequence↗
PubMed1998

Interferon gamma-independent rejection of interleukin 12-transduced carcinoma cells requires CD4+ T cells and Granulocyte/Macrophage colony-stimulating factor.

We analyzed the ability of interferon (IFN)-gamma knockout mice (GKO) to reject a colon carcinoma transduced with interleukin (IL)-12 genes (C26/IL-12). Although the absence of IFN-gamma impaired the early response and reduced the time to tumor onset in GKO mice, the overall tumor take rate was similar to that of BALB/c mice. In GKO mice, C26/IL-12 tumors had a reduced number of infiltrating leukocytes, especially CD8 and natural killer cells. Analysis of the tumor site, draining nodes, and spleens of GKO mice revealed reduced expression of IFN- inducible protein 10 and monokine induced by gamma-IFN. Despite these defects, GKO mice that rejected C26/IL-12 tumor, and mice that were primed in vivo with irradiated C26/IL-12 cells, showed the same cytotoxic T lymphocyte activity but higher production of granulocyte/macrophage colony-stimulating factor (GM-CSF) as compared with control BALB/c mice. Treatment with monoclonal antibodies against GM-CSF abrogated tumor regression in GKO but not in BALB/c mice. CD4 T lymphocytes, which proved unnecessary or suppressive during rejection of C26/IL-12 cells in BALB/c mice, were required for tumor rejection in GKO mice. CD4 T cell depletion was coupled with a decline in GM-CSF expression by lymphocytes infiltrating the tumors or in the draining nodes, and with the reduction and disappearance of granulocytes and CD8 T cells, respectively, in tumor nodules. These results suggest that GM-CSF can substitute for IFN-gamma in maintaining the CD8-polymorphonuclear leukocyte cross-talk that is a hallmark of tumor rejection.

Animals↗
PubMed1998

Reduced tumorigenicity and augmented leukocyte infiltration after monocyte chemotactic protein-3 (MCP-3) gene transfer: perivascular accumulation of dendritic cells in peritumoral tissue and neutrophil recruitment within the tumor.

Monocyte chemotactic protein-3 (MCP-3) is a C-C chemokine that interacts with the CCR1, CCR2, and CCR3 receptors and has a spectrum of action encompassing T cells, NK cells, eosinophils, and dendritic cells (DC), in addition to mononuclear phagocytes. This broad spectrum of action prompted the present study aimed at assessing the antitumor activity of MCP-3 in a gene transfer approach and at providing information as to the actual in vivo leukocyte recruiting capacity of MCP-3. P815 mastocytoma cells transfected with the gene coding MCP-3 (P815/MCP-3) grew in syngeneic hosts and underwent rejection. Rejection was associated with profound alterations of leukocyte infiltration and resistance to subsequent challenge with P815 cells. Tumor-associated macrophages, already present in copious numbers, T cells, eosinophils, and neutrophils, increased in tumor tissues after gene transfer. DC, identified as DEC205+, high MHC class II+, CD11c+ cells, did not increase substantially in the tumor mass. However, in peritumoral tissues, DC accumulated in perivascular areas. P815/MCP-3-transfected tumor cells grew normally in nude mice. Increased accumulation of macrophages and polymorphonuclear neutrophils was evident also in nude mice. mAb against CD4, CD8, and IFN-gamma, but not against IL-4, inhibited rejection of MCP-3-producing cells. An anti-polymorphonuclear mAb caused only a retardation of MCP-3-elicited tumor rejection. Thus, MCP-3 gene transfer elicits tumor rejection by activating type I T cell-dependent immunity. It is tempting to speculate that altered trafficking of APCs, which express receptors and respond to MCP-3, together with recruitment of activated T cells, underlies activation of specific immunity by MCP-3-transfected cells.

Animals↗
PubMed1998

Interleukin 12 gene therapy of MHC-negative murine melanoma metastases.

Immunological gene therapy of cancer relies heavily on the activation of T cells, but tumors with defects in MHC gene expression are not recognized by MHC-restricted T cells. To investigate the potential of cytokine genes for the therapy of MHC-negative tumors, we transduced B78H1, a class I-negative murine melanoma clone, with a polycistronic vector carrying murine interleukin (IL)-12 genes. The clones studied produced 400-25,000 pg/ml IL-12; their in vitro growth properties were similar to those of parental cells. A complete inhibition of growth was observed in vivo both after s.c. and i.v. administration of all IL-12 clones. IL-12-transduced cells were also used as a therapeutic vaccine in mice bearing micrometastases by nontransduced parental cells. A significant (80-90%) reduction in the number of lung nodules was obtained. Immunohistochemical analysis and studies in immunocompromised hosts showed that T cells and natural killer cells had a significant role in the elimination of IL-12-releasing cells. In situ hybridization with cytokine probes detected a strong increase in the proportion of leukocytes positive for IFN-gamma, tumor necrosis factor alpha, IL-1beta, and IFN-inducible protein 10 at the site of rejection of IL-12-engineered tumor cells. However, it was clear that the loss of in vivo growth was also due to T-cell- and natural killer cell-independent factors, possibly related to the antiangiogenic properties of IL-12. In conclusion, tumor therapy based on IL-12 gene transduction was effective on a MHC-negative metastatic tumor, suggesting a possible application to MHC-defective human neoplasms.

Animals↗
PubMed1998

Expression of Met protein and urokinase-type plasminogen activator receptor (uPA-R) in papillary carcinoma of the thyroid.

Met protein encoded by MET oncogene is the high affinity receptor for hepatocyte growth factor (HGF)/scatter factor (SF). HGF/SF has to be cleaved in its heterodimeric form by the urokinase-type plasminogen activator (uPA) to become active as a ligand for Met receptor. The expression of Met protein and of the high affinity receptor for uPA (uPA-R) was investigated in 39 samples of papillary carcinoma using immunohistochemistry. Reactivity for Met protein was present in 33 of 34 tumours, mostly with a diffuse pattern of staining. Reactivity for uPA-R was present in 78 per cent of papillary tumours and exhibited a pattern of staining similar to that of Met protein. Staining for uPA-R was present in 23 of 25 cases (92 per cent) of papillary carcinoma with prominent sclerosis, and in only 1 of 7 cases (14 per cent) without sclerosis. Peritumoural normal thyroid, follicular adenomas, and follicular carcinomas were negative for Met protein and for uPA-R. Hyperfunctioning tall thyroid cells showed weak membrane reactivity for uPA-R and for Met protein. The findings of immunohistochemistry were confirmed at the mRNA level using in situ hybridization, since the signal for uPA-R and Met RNAs was detected in most tumour cells of five cases of papillary carcinoma.

Adult↗
PubMed1998

IL-10 prevents the differentiation of monocytes to dendritic cells but promotes their maturation to macrophages.

Human monocytes cultured with granulocyte-macrophage colony-stimulating factor (GM-CSF) and IL-13 for 7 days differentiate into cells with the morphology and function of dendritic cells (DC). We have investigated the effect of IL-10 on this differentiation pathway. In the presence of IL-10 cells did not develop DC morphology, did not express CD1a and had lower levels of MHC class II. IL-10 promoted the differentiation of large cells with the morphology, cytochemistry and membrane phenotype of macrophages, including staining for nonspecific esterase and high levels of CD14, CD16 and CD68. The effect of IL-10 was dose dependent and was best appreciated when the cytokine was added at the initiation of the culture, as addition on day 3 was less inhibitory. When added to already differentiated DC on day 6, IL-10 caused only a modest reduction of MHC class II and CD1a expression, and no acquisition of the macrophage markers CD14, CD16 and CD68. Prolonged incubation up to 5 days with IL-10 did not induce a shift of differentiated DC to macrophages. On the other hand, the macrophages obtained by culturing for 7 days with GM-CSF+IL-13+IL-10 did not shift to DC upon removal of IL-10 for up to 3 days. Thus, the effect of IL-10 on monocyte differentiation, occurs only at the precursor level and confers an irreversible phenotype. From a functional point of view, cells cultured in the presence of IL-10 were poor stimulators of allogeneic cord blood T cells in mixed lymphocyte reaction (MLR) and presented tetanus toxin (TT) to specific T cell lines with much less efficiency than control DC. In contrast, IL-10-cultured DC showed 7 times greater endocytosis of FITC-dextran. This increased endocytosis was mostly mediated via the mannose receptor, as demonstrated by blocking with unlabeled mannose. In conclusion, IL-10 inhibits DC differentiation from monocytes and, in a substantial proportion of the cells, promotes the differentiation to mature macrophages. Intriguingly, IL-10 inhibits antigen presentation while it stimulates endocytic activity.

Antigen Presentation↗
PubMed1998

Monocyte chemotactic protein-1 in the inflammatory pseudotumour of the lung.

Inflammatory pseudotumour of the lung is a lesion mainly composed of histiocytes. Histiocyte accumulation may arise from local proliferation of migratory cells, from cytokine induced recruitment of monocytes from the systemic circulation, or both. Cell proliferation was investigated with Ki-67 immunostaining and cytokine production with reverse transcriptase-polymerase chain reaction in two cases of inflammatory pseudotumour of the lung. It was found that the two lesions were composed mainly of non-proliferating (Ki-67 non-binding) macrophages that stained positive for CD68, CD14, CD4, and mannose receptor. Both cases contained mRNA transcripts for monocyte chemotactic protein-1 (MCP-1), a monocyte chemoattractant, and for interleukin 6 (IL-6), an inducer of plasma cell differentiation. One of the two cases also contained mRNA transcripts for IL-8, a neutrophil chemoattractant. These findings are consistent with the possibility that accumulation of non-proliferating histiocytes induced by MCP-1 is one of the pathogenic events occurring in inflammatory pseudotumour of the lung.

Adult↗
PubMed1998

The monocyte chemotactic protein a (MCP-1) and interleukin 8 (IL-8) in Hodgkin's disease and in solid tumours.

AIMS: Monocyte chemotactic protein 1 (MCP-1) and interleukin 8 (IL-8) are small, inducible proteins with chemotactic activity for specific subsets of leucocytes. The possibility that MCP-1 and IL-8 are produced in tissues involved by Hodgkin's disease, thus contributing to the inflammatory-type background of the lesion, was investigated. METHODS: The presence of RNA transcripts for MCP-1 and IL-8 was investigated in biopsy samples of 24 cases of Hodgkin's disease, 17 non-Hodgkin's malignant lymphomas, 30 solid tumours, and 30 histologically normal tissues by means of reverse transcription-polymerase chain reaction (RT-PCR)/Southern blot analysis. RESULTS: MCP-1 expression was detected in 23 of 24 cases of Hodgkin's disease, in seven of 17 cases of B cell non-Hodgkin's lymphoma, and in seven of 14 cases of reactive lymphoid hyperplasia. IL-8 was present in six of 14 cases of Hodgkin's disease, and was seen only rarely in B cell non-Hodgkin's lymphoma and in reactive lymphoid tissues. MCP-1 and IL-8 RNA transcripts were detected in 13 of 25 carcinomas originating from the lung, breast, thyroid, and ovary. CONCLUSIONS: These findings are consistent with the possibility that MCP-1 and IL-8 are two additional cytokines involved in the pathogenesis of Hodgkin's disease.

Blotting, Southern↗
PubMed1997

TCL1 is overexpressed in patients affected by adult T-cell leukemias.

Among mature postthymic T-cell leukemias, adult T-cell leukemia (ATL) has characteristic clinicopathological entities. The association with the human T-cell leukemia/lymphotropic virus type I is one of the distinctive etiopathogenetic features of this disease. However, unlike other acute transforming retroviruses, the human T-cell leukemia/lymphotropic virus type I lacks an oncogene within its genome. Other human postthymic leukemias, such as T-prolymphocytic leukemias, involve mostly the CD4 cellular subset and share many similarities to ATLs (aggressive course, cutaneous involvement, CD4+, CD29+, CD45RA- phenotype, and alpha-naphthyl-acetate esterase positivity). A chromosomal rearrangement at 14q32.1, involved in translocations or inversions with either the alpha/delta locus [t(14;14)(q11;q32.1), inv14(q11;q32.1)], or the beta-chain locus of the T-cell receptor [t(7;14)(q35;q32.1)] is found. These rearrangements disregulate a gene, TCL1, located at the 14q32.1 region, that we show is physiologically expressed in CD4/CD8 double-negative thymocyte cells, but not in more differentiated CD4+ and CD8+ subpopulations. Here, using molecular and immunocytochemical analysis, we report that TCL1 is also overexpressed in 10 of 10 ATL specimens, indicating that this gene may play an important role in the pathogenesis of this disease.

Adult↗