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Biomedical subjects

E Di Carlo

Publications and source records attributed to E Di Carlo.

At least 37 records · Page 2Linked to original sources
PubMed2000

p185(neu) protein is required for tumor and anchorage-independent growth, not for cell proliferation of transgenic mammary carcinoma.

Transgenic FVB-NeuN mice (N202) bearing the rat neu protooncogene driven by the mouse mammary tumor virus promoter/enhancer develop focal mammary carcinomas overexpressing the neu-encoded p185(neu) protein. In vitro expression of p185(neu) among mammary carcinoma cultures was heterogeneous, and we could establish some cell lines and clones displaying a complete loss of p185(neu) expression, along with others with very high p185(neu) protein level. Upon in vivo injection, p185(neu)-positive cells gave rise to fast-growing tumors with a short latency, while p185(neu)-negative cells required a very long latency time, and the resulting tumors were invariably p185(neu)-positive. The lower growth ability of p185(neu)-negative cells in vivo was also confirmed in athymic nude mice. In vitro, analysis of anchorage-independent growth in soft agar revealed colony formation from p185(neu)-positive but not p185(neu)-negative cells. The direct involvement of p185(neu) in clonogenicity was demonstrated by the inhibition of p185(neu)-positive colony growth in soft agar in the presence of an anti-p185(neu) monoclonal antibody. By contrast, a higher level of anchorage-dependent clonogenic growth and proliferation was observed in p185(neu)-negative cells as compared to p185(neu)-positive cells, thus explaining the relative ease with which p185(neu)-negative cell lines and clones were established in vitro. Together, the results indicate that p185(neu) expression can lead to tumor formation and metastasis through the modification of intrinsic properties of cells related to anchorage-independent growth ability rather than to proliferation or host-dependent mechanisms.

Animals↗
PubMed2000

Tumor rejection and immune memory elicited by locally released LEC chemokine are associated with an impressive recruitment of APCs, lymphocytes, and granulocytes.

The human beta chemokine known as LEC (also called NCC-4, HCC-4, or LMC) displays chemotactic activity for monocytes and dendritic cells. The possibility that its local presence increases tumor immunogenicity is addressed in this paper. TSA parental cells (TSA-pc) are poorly immunogenic adenocarcinoma cells that grow progressively, kill both nu/nu and syngeneic BALB/c mice, and give rise to lung metastases. TSA cells engineered to release LEC (TSA-LEC) are still able to grow in nu/nu mice, but are promptly rejected and display a marginal metastatic phenotype in BALB/c mice. Rejection is associated with a marked T lymphocyte and granulocyte infiltration, along with extensive macrophage and dendritic cell recruitment. NK cells and CD4+ T lymphocytes are uninfluential in TSA-LEC cell rejection, whereas both CD8+ lymphocytes and polymorphonuclear leukocytes play a major role. An antitumor immune memory is established very quickly after rejection, since 6 days later 75% of BALB/c mice were already resistant to a TSA-pc challenge. Spleen cells from rejecting mice display specific cytotoxic activity against TSA-pc and secrete IFN-gamma and IL-2 when restimulated by TSA-pc. The ability of LEC to markedly improve recognition of poorly immunogenic cells by promoting APC-T cell cross-talk suggests that it could be an effective component of antitumor vaccines.

Adenocarcinoma↗
PubMed2000

Ability of systemic interleukin-12 to hamper progressive stages of mammary carcinogenesis in HER2/neu transgenic mice.

Previous studies in mice have shown that chronic administration of recombinant interleukin-12 (IL-12) hampers the progression of both chemical- and oncogene-dependent carcinogenesis. This suggests that a new preventive strategy may be envisaged for individuals with a genetic risk of cancer or carrying preneoplastic lesions. Starting at progressive stages of mammary carcinogenesis, female BALB/c and FVB mice carrying the activated rat HER2/neu oncogene (BALB-neuT) or the proto-oncogene (FVB-neuN) under the mouse mammary tumor virus promoter received multiple 5-day courses of different doses of IL-12. The times of tumor appearance, multiplicity, and histopathological features of the neoplastic lesions were evaluated. In both BALB-neuT and FVB-neuN mice, 5-day i.p. courses of 50/100 ng of IL-12/day inhibited mammary carcinogenesis when they coincided with the progression of early preneoplastic lesions. Inhibition appears to depend primarily on the ability of IL-12 to interfere with early tumor angiogenesis. Later treatments are much less effective, and daily doses of 10 and 2 ng are useless. The efficacy of early IL-12 courses suggests that they could be used to prevent mammary tumors in individuals at risk, whereas their lower efficacy in later stages of carcinogenesis and the dose range required pose some constraints on their use in the management of overt preneoplastic lesions. Precise understanding of tumor progression means that effective treatments can be commenced relatively late in the life of individuals at risk and that no lifetime administration is required.

Animals↗
PubMed2000

Dissimilar anti-tumour reactions induced by tumour cells engineered with the interleukin-2 or interleukin-15 gene in nude mice.

Interleukin (IL)-15 shares immuno-stimulatory properties with IL-2 and is a potent inducer of natural killer (NK) cell function. The major histocompatibility complex (MHC) class I-negative human small cell lung cancer (SCLC) cell line N592, engineered to express a modified IL-15 cDNA (N592/IL-15), secreted biologically active IL-15 (300-500 pg/ml), capable of boosting T-cell proliferation and NK activity 'in vitro'. The effect of IL-15 gene transfer on natural immunity 'in vivo' was assessed by xenotransplants in nude mice and compared with that of the IL-2 gene. N592 cells engineered with IL-2 (N592/IL-2) were promptly rejected, while N592/IL-15 displayed a significant delay in tumour growth and a slightly reduced take rate. However, in NK-depleted nude mice, N592/IL-15 displayed the same growth kinetics as unmodified N592 cells, and N592/IL-2 grew with slightly reduced kinetics. An impressive reactive cell infiltration, consisting mainly of macrophages and granulocytes, was associated with N592/IL-2 tumour rejection, while a more evident recruitment of NK cells was found in N592/IL-15 tumours. In both N592 transfected tumours, we found expression of chemoattractant molecules, such as granulocyte macrophage-colony stimulating factor (GM-CSF) and monocyte chemoattractant protein (MCP)-1, while macrophage inflammatory protein (MIP)-2 was produced by endothelial cells only in N592/IL-2 tumours. In this tumour, very few and severely damaged microvessels were found, while microvessels were numerous in N592/IL-15 tumours. The potent recruitment of NK cells mediated by IL-15 gene transfer suggests its possible therapeutic use in tumours lacking MHC class I.

Animals↗
PubMed1999

A "stealth effect": adenocarcinoma cells engineered to express TRAIL elude tumor-specific and allogeneic T cell reactions.

BALB/c mammary adenocarcinoma cells engineered to express TNF-related apoptosis-inducing ligand (TRAIL)/APO-2 ligand (APO-2L) on their membrane (TSA-TRAIL) grow with kinetics similar to that of parental cells (TSA-pc) in vitro and in nu/nu mice. In contrast, TSA-TRAIL cells grow faster than TSA-pc in normal BALB/c mice. In DBA/2 mice, which differ from BALB/c mice at minor histocompatibility Ags, they also grow faster and display a higher percentage of tumor takes than TSA-pc. In fully histoincompatible C57BL/6 (B6) mice, TSA-TRAIL cells form evident tumors that are slowly rejected by most mice, but outgrow in a few. In contrast, TSA-pc cells are rejected at once by B6 mice. Since TRAIL/APO-2L induces apoptosis by interacting with a variety of specific receptors, this rapid growth in both syngeneic and allogeneic mice may be the result of an immunosuppressive mechanism. The following evidence supports this hypothesis: 1) TSA-TRAIL cells overcome the strong immunity against TSA-pc cells elicited in BALB/c mice by preimmunization with TSA cells engineered to release IL-4; 2) their rejection by B6 mice does not prime a CTL-mediated memory; 3) thymidine uptake by T lymphocytes unstimulated or stimulated by allogeneic cells is inhibited when TSA-TRAIL cells are added as third party cells; 4) CTL kill TSA-pc but not TSA-TRAIL cells in 48-h assays; and 5) activated lymphocytes interacting with TSA-TRAIL cells in vivo and in vitro undergo apoptosis.

Adenocarcinoma↗
PubMed1999

Immune events associated with the cure of established tumors and spontaneous metastases by local and systemic interleukin 12.

The antitumor activity of recombinant murine interleukin-12 (rIL-12) is documented by a large set of data from numerous mouse models. Because the cellular and molecular mechanisms by which rIL-12 impairs tumor growth are still not fully defined, we compared the effects of local and systemic rIL-12 administration in mice harboring an invasive 7-day-old moderately differentiated and spontaneously metastasizing mammary adenocarcinoma (TSA). Whereas the immune events elicited via the two routes of rIL-12 administration seem to be the same, systemic rIL-12 is markedly more effective; tumor destruction is dependent on a prompt antitumor response resulting from the cooperation of several subsets of reactive cells. The reactions that seem to play a key role are: (a) indirect inhibition of angiogenesis by secondary cytokines (mainly IFN-gamma) and third-level chemokines (inducible protein 10 and monokine induced by IFN-gamma); (b) systemic activation of leukocyte subsets capable of producing proinflammatory cytokines, CTLs, and antitumor antibodies; and (c) destruction of tumor vessels by polymorphonuclear cells. The markedly higher efficacy of systemic rIL-12 seems to rest on its ability to recruit these systemic reactions more quickly and efficiently than local rIL-12.

Animals↗
PubMed1999

Interferon (IFN)-beta gene transfer into TS/A adenocarcinoma cells and comparison with IFN-alpha: differential effects on tumorigenicity and host response.

Our group had previously shown that transfer of the mouse interferon (IFN)-alpha1 gene into the metastasizing TS/A mammary adenocarcinoma resulted in T-cell-mediated tumor rejection and development of antitumor immunity. Moreover, we had shown that the metastatic ability of TS/A tumor cells producing IFN-alpha was strongly impaired, whereas IFN-gamma expression did not influence or augmented metastasis formation by TS/A cells. In this study, we have analyzed the in vitro and in vivo behavior of various TS/A tumor cell clones isolated after the transduction with a recombinant retroviral vector carrying the mouse IFN-beta gene. We have also compared the tumorigenicity of these clones with that of TS/A cells expressing IFN-alpha1. BALB/c mice were inoculated subcutaneously with parental TS/A cells, transduction control TS/A cells, or TS/A cells producing IFN-alpha or IFN-beta. Tumor growth was evaluated by the measurement of tumor masses and analysis of survival. The features of tumor growth and rejection were examined by histological and immunohistochemical analyses. The metastatic ability of parental TS/A cells, transduction control TS/A cells, or TS/A cells producing IFN-alpha, IFN-beta, or IFN-gamma was evaluated after intravenous injection of the tumor cells into BALB/c mice by counting of the lung metastatic nodules and analysis of survival. A strong inhibition of tumorigenicity and development of tumor immunity were observed upon subcutaneous injection of syngeneic mice with TS/A tumor cells producing high amounts of IFN-beta, but not with clones expressing low levels of the cytokine, as observed for cells expressing IFN-alpha. IFN-alpha secretion by TS/A cells at the site of tumor growth induced a stronger inflammatory response as compared with IFN-beta, which appeared to be more active in the inhibition of tumor-induced angiogenesis. Notably, the metastatic ability of IFN-beta-producing TS/A cells after intravenous injection was either not affected or only slightly impaired as compared with parental TS/A tumor cells. In contrast, even cells producing low levels of IFN-alpha proved to be poorly metastatic. These findings represent the first comparison of the effectiveness of IFN-alpha versus IFN-beta produced by genetically modified cells on their tumorigenic behavior and suggest the existence of some notable differences in the capabilities of these two cytokines to induce a host antitumor reactivity in mice.

Adenocarcinoma↗
PubMed1999

Analysis of mammary carcinoma onset and progression in HER-2/neu oncogene transgenic mice reveals a lobular origin.

Morphologic examinations of mammary neoplasias arising in BALB/c (H-2d) mice carrying the activated rat HER-2/neu oncogene (BALB-NeuT), and in FVB (H-2q) mice bearing the wild-type proto-oncogene (FVB-NeuN), indicate that both conditions result in a very human-like lobular carcinoma of alveolar type, whose histotype is the result of the preferential expression of HER-2/neu products in the epithelium of lobular ducts and lobules. Detailed analysis of tumor progression indicates that transition from lobular hyperplasia to overt carcinoma is associated with a high epithelial proliferation rate, as assessed by anti-proliferating cell nuclear antigen immunostaining, and coincides with the activation and maximal extension of tumor angiogenic process as assessed by microvessel count (anti-CD31), anti-beta3 integrin, and anti-laminin immunostaining. Neovascularization is accompanied by vascular endothelial cell growth factor and basic fibroblast growth factor production by hyperplastic epithelial cells. By contrast with the BALB-NeuT tumors, E-cadherin expression is almost nonexistent in those arising in FVB-NeuN mice and this may explain their high metastatic potential. Despite their different kinetics, however, the lung metastases observed in both strains are histologically similar and resemble the primary tumor. Both strains can thus be proposed as models for "in vivo" investigation of the origin and progression of the alveolar type of lobular mammary carcinoma and the testing of new therapeutic approaches.

Animals↗
PubMed1998

Coexpression of IL-6 and soluble IL-6R causes nodular regenerative hyperplasia and adenomas of the liver.

Studies with tumor necrosis factor p55 receptor- and interleukin-6 (IL-6)-deficient mice have shown that IL-6 is required for hepatocyte proliferation and reconstitution of the liver mass after partial hepatectomy. The biological activities of IL-6 are potentiated when this cytokine binds soluble forms of its specific receptor subunit (sIL-6R) and the resulting complex interacts with the transmembrane signaling chain gp130. We show here that double transgenic mice expressing high levels of both human IL-6 and sIL-6R under the control of liver-specific promoters spontaneously develop nodules of hepatocellular hyperplasia around periportal spaces and present signs of sustained hepatocyte proliferation. The resulting picture is identical to that of human nodular regenerative hyperplasia, a condition frequently associated with immunological and myeloproliferative disorders. In high expressors, hyperplastic lesions progress with time into discrete liver adenomas. These data strongly suggest that the IL-6/sIL-6R complex is both a primary stimulus to hepatocyte proliferation and a pathogenic factor of hepatocellular transformation.

Adenoma↗
PubMed1998

Down regulation of major histocompatibility complex class I expression in mammary carcinoma of HER-2/neu transgenic mice.

Transgenic mice carrying the HER-2/neu proto-oncogene under tissue-specific transcriptional control of a mammary tumor virus long terminal repeat (Tg-MMTVneu mice) spontaneously develop mammary carcinomas. HER-2/neu is a tumor antigen that can be recognized by cytotoxic T lymphocytes if tumor cells present the appropriate major histocompatibility complex (MHC) class I glycoproteins. The purpose of this work was to assess whether mammary carcinomas arising in Tg-MMTVneu mice correctly expressed MHC (H-2q) class I gene products. We analyzed by flow cytometry 51 primary tumors from 19 transgenic mice. About one-half of the tumors showed a reduced expression of class I antigens. All tumors were highly positive for membrane neu. Some mice had multiple mammary carcinomas with widely different MHC expression levels, and most mice had at least one tumor with a low expression. Treatment with gamma-interferon of carcinoma cells cultured in vitro induced a strong reexpression of H-2q antigens. Our results suggest that the immune response activated in vivo by HER-2/neu-positive tumors can lead to the emergence of escape variants characterized by a down-regulation of MHC class I products.

Animals↗
PubMed1998

Interleukin 12-mediated prevention of spontaneous mammary adenocarcinomas in two lines of Her-2/neu transgenic mice.

The ability of interleukin (IL)-12 to prevent tumors when administered to individuals with a genetic risk of cancer was studied in two lines of transgenic mice expressing rat HER-2/neu oncogene in the mammary gland. Female BALB/c (H-2(d)) mice carrying the activated HER-2/ neu oncogene show no morphological abnormalities of the mammary gland until 3 wk of age. They then progress through atypical hyperplasia to in situ lobular carcinoma and at 33 wk of age all 10 mammary glands display invasive carcinomas. Adult FVB mice (H-2(q)) carrying the HER-2/neu protooncogene develop mammary carcinomas with a longer latency (38-49 wk) and a lower multiplicity (mean of 2.6 tumors/mice). Treatment with IL-12 (5 daily intraperitoneal injections, 1 wk on, 3 wk off; the first course with 50 ng IL-12/day, the second with 100 ng IL-12/day) begun at 2 wk of age in BALB/c mice and at 21 wk of age in FVB mice markedly delayed tumor onset and reduced tumor multiplicity. Analogous results were obtained in immunocompetent and permanently CD8(+) T lymphocyte-depleted mice. In both transgenic lines, tumor inhibition was associated with mammary infiltration of reactive cells, production of cytokines and inducible nitric oxide synthase, and reduction in microvessel number, in combination with a high degree of hemorrhagic necrosis.

Animals↗
PubMed1998

The immune response elicited by mammary adenocarcinoma cells transduced with interferon-gamma and cytosine deaminase genes cures lung metastases by parental cells.

The parental cells of the TSA murine mammary adenocarcinoma (TSA-pc) were transfected with both the interferon-gamma (IFN-y) gene and the cytosine deaminase (CD) suicide gene to obtain a therapeutic vaccine active against TSA-pc lung metastases. Even in the absence of treatment with the prodrug 5-fluorocytosine (5-FC), the local growth of double transfectants (CD-y clones) was inhibited by a marked recruitment of granulocytes and macrophages. In mice harboring TSA-pc micrometastases, therapeutic vaccination with either IFN-gamma or CD single transfectants reduced the number of lung nodules, whereas CD-gamma double transfectants abrogated metastasis growth in up to 80% of mice. Treatment of mice with 5-FC did not alter the curative efficacy of CD-gamma double-transfectant cells. By contrast, in mice vaccinated with CD single-transfectant cells, 5-FC treatment caused a significant loss of their curative activity. Host T cells played an active role in the cure of lung metastases, because vaccination of nude mice with CD-gamma cells was uneffective.

Adenocarcinoma↗
PubMed1998

Interaction between endothelial cells and the secreted cytokine drives the fate of an IL4- or an IL5-transduced tumour.

Injection of interleukin-4 (IL4) gene-transduced tumour cells into syngeneic immunocompetent mice resulted in tumour rejection in which a key role for eosinophils was suggested. To evaluate whether IL5 inhibits tumour growth by selectively inducing eosinophil recruitment and activation, a poorly differentiated mammary adenocarcinoma cell line (TSA) was transfected with the IL5 gene and the cells secreting IL5 (TSA-IL5) were injected subcutaneously (s.c.) in syngeneic mice. The oncogenicity of TSA-IL5 was compared with that exhibited by TSA cells transfected with the IL4 gene (TSA-IL4) and with the neomycin resistance gene only (TSA-neo). At progressive times after subcutaneous challenge, tumour growth areas were studied histologically, ultrastructurally, and immunohistochemically to identify the reactive cells, visualize tumour vessels, and detect the cytokines and chemokines involved in the anti-tumour reaction. Both the morphological and the functional data showed that TSA-IL5, despite the large eosinophil infiltrate, grew progressively like TSA-neo, suggesting that eosinophils per se do not play a crucial role in TSA tumour rejection. Furthermore, our data indicate that the rejection of TSA-IL4 depends on the IL4-induced expression of VCAM-1 and MCP-1 by endothelial cells. MCP-1 together with VCAM-1 results in recruitment and activation of basophils, mast cells, and macrophages, and hence a pro-inflammatory cytokine cascade that initially favours the influx and activation of neutrophils and finally tumour rejection. In this context, the rejection of TSA-IL4 seems to involve a variety of reactive cells and rests on a continuous cross-talk between basophils, mast cells, macrophages, CD8-positive lymphocytes, and granulocyte subsets, mostly neutrophils.

Animals↗
PubMed1998

Local release of interleukin-10 by transfected mouse adenocarcinoma cells exhibits pro- and anti-inflammatory activity and results in a delayed tumor rejection.

Tumor cells engineered to release cytokines are a valuable tool for investigating biological activities elicited by local cytokines. The parental cells of a mouse mammary adenocarcinoma (TSA-pc) were transduced with the cDNA coding for mouse interleukin-10 (IL-10). In vitro, transduced TSA cells secrete about 200 ng of IL-10/10(5) seeded cells in 48 hours (TSA-IL-10). When injected subcutaneously into syngeneic BALB/c mice, TSA-IL-10 cells gave rise to a tumor that grew progressively during the first 7-10 days and then rapidly and completely regressed. To study the events associated with this growth and disappearance, histological, immunohistochemical and ultrastructural analyses of the tumor area were performed at progressive times after challenge. A slow, but progressive and massive recruitment of leukocytes (mainly macrophages and neutrophils) into the tumor was evident. Several CD8+, CD4+ lymphocytes and a few NK cells were present. Marked inhibition of neoangiogenesis was also observed. On day 9, the microvascular network in the growth area had almost vanished, while vascular damage was present in the surrounding stromal tissue. From day 4, down-modulation of VEGF expression in the tumor area and inhibition of tumor necrosis factor-alpha (TNF-alpha) and IL-6 production by reactive leukocytes were evident. The few vessels present in the tumor area displayed poor expression of monocyte chemotactic protein-1 (MCP-1), moderate expression of VCAM-1, and strong expression of ELAM-1, three molecules that result in adhesion of inflammatory cells to the endothelium. A few tumor-infiltrating macrophages were moderately stained with anti-iNOS antibodies. These findings suggest that the collapse of established TSA-IL-10 tumors is the result of the pro- and anti-inflammatory activity of IL-10, which: a) is a signal for the local recruitment of leukocytes; b) leads to vascular damage; c) suppresses cytokine production. The coexistence of both a direct stimulatory activity on endothelial cells and an anti-angiogenic activity is evidence of the ambivalence of the local effects of IL-10.

Adenocarcinoma↗
PubMed1997

Interleukin 6 gene-transfected mouse mammary adenocarcinoma: tumour cell growth and metastatic potential.

Cells from the spontaneous metastatic TSA mammary adenocarcinoma of BALB/C mouse were transfected with the murine (interleukin-6) IL6 gene. The clone (TSA-IL6) secreting the largest amount of IL6 displayed an in vitro increased growth rate compared with that of TSA cells transfected with the neomycin resistance gene only (TSA-neo). TSA-IL6 cell colonies consisted mainly of fusiform cells and TSA-neo colonies of polygonal cells. When subcutaneously (s.c.) injected in syngeneic mice, TSA-IL6 cells gave rise to tumours that grew significantly slower than TSA-neo cell tumours. Microscopically, TSA-IL6 tumours displayed a fascicular pattern of growth, associated with a very scanty macrophage infiltrate. S.c. TSA-IL6 tumours were significantly less metastatic than TSA-neo tumours. By contrast, following intravenous (i.v.) challenge, TSA-IL6 cells produced 5-7 times more lung metastases than TSA-neo cells. The i.v. TSA-IL6 cell lung metastases showed a marked macrophage infiltrate and a rich vascularization. The high in vitro TSA-IL6 cell growth rate is attributable to the IL6-induced production of growth factors, some of which possess heparin-binding properties, such as amphiregulin. The differences in vascularization and macrophage infiltrate may underlie the observed differences between s.c. TSA-IL6 tumour growth with low spontaneous metastatic potential and the widespread growth of i.v. metastasis.

Adenocarcinoma↗
PubMed1988

Calcium-acidic phospholipid-phosphate complexes in human hydroxyapatite-containing pathologic deposits.

The deposition of calcium-containing crystals in tissues is due to a combination of factors: elevation in the concentrations of precipitating ions, formation of specific nucleators, and removal of macromolecules that inhibit crystal deposition. This study tested the hypothesis that calcium acidic phospholipid phosphate complexes, which promote hydroxyapatite deposition both in vitro and in vivo, are associated only with hydroxyapatite deposits, and furthermore, that the presence of these complexes is associated with all such hydroxyapatite deposits. Lipid analysis of 76 surgical specimens containing evidence of pathologic calcification (35 hydroxyapatite, 35 calcium pyrophosphate dihydrate, and 6 containing other crystalline materials) had mean complexed acidic phospholipid contents of 8.7, 1, and 0.012, (microgram/mg demineralized dry weight) respectively. Tissues that contained larger, more perfect hydroxyapatite crystals based on x-ray diffraction analyses, had a higher complexed acidic phospholipid content (7.5 +/- 4 micrograms/mg demineralized dry weight, N = 16) than tissues with poorly crystallized hydroxyapatite (3.9 +/- 2 micrograms/mg, N = 11). Histologically, tissues containing larger crystals were characterized by cell or tissue necrosis. Poorly crystalline deposits were found in tissues showing little or no evidence of cell necrosis or tissue degeneration.

Calcinosis↗