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Biomedical subjects

G Forni

Publications and source records attributed to G Forni.

At least 91 records · Page 5Linked to original sources
PubMed1994

Transduction of the IL2 gene into human acute leukemia cells: induction of tumor rejection without modifying cell proliferation and IL2 receptor expression.

BACKGROUND: Previous studies have suggested that some of the limitations associated with the administration of high-dose exogenous interleukin 2 (IL2) may be overcome, at least partly, by cytokine gene transfer modalities. These findings have prompted investigations into whether human tumor cells may be transduced with the IL2 gene and whether tumor cell lines could be engineered to release IL2. PURPOSE: The purpose of this study was to evaluate the possibility of inducing a productive transfer of the IL2 gene into human acute leukemia cells and to assess the phenotypic and proliferative changes generated in the engineered cells, as well as their tumorigenic potential in nude mice. METHODS: Three retroviral vectors (DC/TK/IL2, DC/AD/R/IL2, and N2/CMV/IL2) carrying the IL2 gene were used to transduce three human leukemic cell lines: K562 and U937 (myeloid) and ST4 (lymphoid). Messenger RNA expression of the IL2 gene was analyzed by reverse transcriptase-polymerase chain reaction (RT-PCR) and productive IL2 release using a human IL2 assay and an enzyme-linked immunosorbent assay kit. The expression of the p55 (alpha) and p75 (beta) chains of the IL2 receptor were determined by RT-PCR and indirect immunofluorescence. The kinetics of in vitro growth and proliferation of parental and engineered cells were also measured. Parental and IL2 gene-transduced ST4 lymphoblasts were injected into immunosuppressed nude mice that had their tumors measured twice weekly. RESULTS: The productive insertion of the IL2 gene was achieved in all three cell lines studied. The amounts of IL2 constitutively released by the engineered neoplastic cells ranged between 1 and 11 U/mL of IL2 produced from 10(6) cells in 72 hours. A fivefold increase in IL2 production was obtained in ST4 cells by further limiting dilution cloning of the bulk-infected cells. The stable integration of the IL2 gene did not modify the phenotype of the leukemic cells, the expression of the IL2 receptor alpha and beta chains and of several cytokine genes, or the kinetics of in vitro growth and proliferation. In nude mice injected with various IL2-producing ST4 clones, tumor growth associated inversely with the amounts of IL2 secreted by the leukemic cells. CONCLUSIONS: The results of this study demonstrate that the IL2 gene can be productively transduced into human myeloid and lymphoid leukemic cells without modifying their phenotypic and proliferative properties and that this transduction leads to a reduced or abrogated in vivo tumorigenic potential.

Acute Disease↗
PubMed1994

Environmental signals influencing expression of the IFN-gamma receptor on human T cells control whether IFN-gamma promotes proliferation or apoptosis.

IFN-gamma R expression is subject to contrasting modulation on human T cells. IFN-gamma R constitutive expression is low on three human malignant T cells (ST4, PF382, and Jurkat) growing in medium supplemented with serum. The addition of IFN-gamma down-modulates IFN-gamma R expression and increases both proliferation and MHC class I Ag expression. By contrast, when malignant T cells are cultured in medium without serum, IFN-gamma R expression dramatically increases and the cells undergo a slow apoptotic death. The addition of IFN-gamma enhances apoptosis and inhibits cell rescue in serum-supplemented medium. This opposite ability of IFN-gamma to stimulate malignant T cell proliferation or death correlates with the intensity of IFN-gamma R cell expression, high expression being a marker for cell apoptosis. IFN-gamma R up-modulation also occurs on malignant T cells undergoing apoptosis after treatment with dexamethasone, on irradiated normal CD3+ PBL, and on cultured normal CD3+ thymocytes. Moreover, the ability of IFN-gamma to augment apoptosis of highly IFN-gamma R-positive thymocytes suggests that its role in promoting T cell apoptosis is also important in physiologic conditions.

Apoptosis↗
PubMed1994

Interleukin-2 prevention of apoptosis in human neutrophils.

Evidence is presented that interleukin (IL)-2 maintains viability of human polymorphonuclear cells (PMN) in culture by preventing these cells from undergoing programmed cell death (PCD) and induces the synthesis of new RNA and protein. Our laboratory has recently discovered that human PMN constitutively express IL-2 beta receptor and more importantly, PMN are able to respond functionally to IL-2 by enhanced growth inhibitory activity against an opportunistic fungal pathogen, Candida albicans. We now report that IL-2 was able to interfere with the PCD process and reduce the number of apoptotic PMN to < 40% in 72-h culture. Freshly isolated PMN usually underwent a time-dependent aging process and > 80% of PMN cultured in medium alone for 72 h showed morphologic features of PCD as depicted by hematoxylin and eosin staining as well as by electron microscopy. During the PCD process, untreated PMN not only exhibited condensed nuclear structure and decrease in cell size, but also displayed DNA fragmentation. DNA fragmentation in PMN was prevented by IL-2. Prevention of PCD by IL-2 was associated with an increase in new RNA and protein synthesis in PMN, which may reflect cytokine induction, such as tumor necrosis factor, as we have recently shown. Thus, our data expands our current understanding of PMN in that they may be an active component of the immune system, with a longer life-span when activated than expected.

Apoptosis↗
PubMed1994

LPS-inducible nuclear factor in human monocytes that binds the negative regulatory element of the HIV LTR.

We studied the constitutive and lipopolysaccharide (LPS)-induced expression of nuclear protein binding to the negative regulatory element (NRE) of the human immunodeficiency virus type 1 (HIV-1) long terminal repeat (LTR) in fresh human monocytes. We demonstrated the existence of a constitutive factor binding to the NRE 73-bp HpaII/HpaII fragment (-216 to -143) whose expression is up-regulated by LPS treatment. Competition experiments with overlapping oligonucleotides covering the HpaII/HpaII fragment and with mutated oligonucleotides mapped the binding within the TTTCATCAC region (-171 to -163). This binding pattern is unique to human monocytes.

Base Sequence↗
PubMed1994

Temporary regression of recurrent squamous cell carcinoma of the head and neck is achieved with a low but not with a high dose of recombinant interleukin 2 injected perilymphatically.

The efficacy of ten daily injections of 500 or 500,000 U of recombinant interleukin 2 (IL-2) day-1 given 1.5 cm from the insertion of the sternocleidomastoid muscle on the mastoid was evaluated in 31 patients with recurrent head and neck squamous cell carcinoma. No toxic effects were noted. One complete response (CR) and three partial responses (PRs) were observed in the 16 patients who received 500 U of IL-2, whereas the higher dose was not effective. The CR was recorded in one of the seven patients with a oropharyngeal recurrence. Partial responses were obtained in 1/5 patients with hypopharyngeal recurrences, in 1/5 patients with oral cavity recurrences and 1/7 patients with laryngeal recurrences. The duration of the responses was 3-5 months and additional courses of ten injections of IL-2 had no further effect.

Carcinoma, Squamous Cell↗
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↗
PubMed1994

[Usefulness of ultrasonographic techniques in catheterization of the internal jugular vein in patients with chronic heart failure].

BACKGROUND: The right internal jugular vein as a route for right heart catheterization and continuous infusion of drugs is increasingly used in patients with heart failure. Although this approach has several advantages, a small but definite number of unsuccessful vein punctures and/or of complications have been reported. This prospective study was designed to evaluate the usefulness of ultrasound techniques for cannulating internal jugular vein in a series of 310 consecutive patients with chronic heart failure. METHODS: In all patients a duplex scanning of internal jugular veins was performed before the cannulation. A subgroup of 62 patients was selected for having a "difficult" cannulation according to the following criteria: previous failure of cannulating the vein (3 unsuccessful needle advances); neck abnormalities; severe emphysema and respiratory insufficiency. In these patients a Doppler-guidance system, which consisted of a miniature ultrasound Doppler transducer inserted in a standard 19 gauge needle, was used. The needle was advanced under the skin following the maximal audio signal of the venous flow. The following variables were considered: success rate, number of needle advances to cannulate the vein, time elapsed from local anesthesia and the insertion of the catheter, minor and major complications. RESULTS: Duplex scanning showed that in 14 patients (4.5%) the right internal jugular vein was occluded or severely narrowed. In all patients the left internal jugular vein, which showed a compensatory dilatation, was successfully cannulated. In 294 of the remaining 296 patients the position of the vein was anterior to the carotid artery at a depth of 4-27 mm below the skin. With respect of the triangle formed by the two heads of the sternocleidomastoid muscle, the vein was central in 35%, medial in 15% and lateral in 60% of cases. Based on duplex scanning ultrasound data, 285 patients underwent internal jugular vein cannulation, which was achieved at the first attempt in 74% and within 3 attempts in 87% of cases. Minor and major complications occurred in 4 (1.4%) and in 1 (0.3%) respectively. By the Doppler guidance system, the cannulation was successfully achieved in 79% of patients at the first attempt and in 98% of patients (61/62) within 3 attempts. In one patient an uneventful puncture of the carotid artery occurred. The time to perform the cannulation was not significantly different using the two approaches (conventional approach: 4.4 +/- 3 minutes; Doppler guidance system: 4.2 +/- 2). Overall the internal jugular vein was successfully cannulated in 307/310 patients (99%). CONCLUSIONS: Ultrasound techniques provide useful information which facilitates the cannulation of the internal jugular vein in patients with heart failure. The Doppler guidance method allows a rapid and safe cannulation of the vein even in cases that are difficult using the conventional approach.

Adolescent↗
PubMed1994

[Loco-regional treatment with low and high doses of interleukin-2 of head and neck squamous cell carcinoma recurrences].

The present study reports the effects of repeated administration of two different doses of recombinant (r) Interleukin-2 (IL-2) around tumor-draining lymph nodes in 22 patients with recurrent squamous cell carcinoma of the head and neck (HNSCC). Each patient was randomly assigned a dose of 500 or 500,000 IU/r-IL-2/day. Inoculation was made at a depth of 15 mm in the anterior margin of the sternocleidomastoid muscle at 15 mm from its insertion into the mastoid. The injections were made on the same side as the recurrent tumor when draining lymph nodes were still present and contralaterally when only contralateral lymph nodes were present. The 10 daily injection were followed by an interval without treatment until day 40. A further three courses of 10 injections were made at thirty-day intervals in the absence of a CR. There was no toxicity. One complete response (CR) and 2 partial responses (PR) were observed in the 11 patients who received 500 U of IL-2, whereas the higher dose was not effective. The CR was recorded in the 4 patients with an oropharyngeal recurrence. One PR was obtained in the 4 patients with hypopharyngeal recurrences and one in 3 with oral cavity recurrences. Clinical responses were nearly always preceded by improvement of the functional impairment caused by tumour infiltrations of the upper respiratory-digestive organs. The responses lasted 3-5 months and additional courses of 10 injections of IL-2 had no further effects.

Adolescent↗
PubMed1993

Protective and curative potential of vaccination with interleukin-2-gene-transfected cells from a spontaneous mouse mammary adenocarcinoma.

The potential of interleukin 2-gene-transfected tumor cells to prevent tumor growth and cure established tumors was evaluated using cells from a spontaneous, invasive, and metastasizing mouse mammary adenocarcinoma. Tumor cells engineered to secrete interleukin 2 initially trigger a local inflammatory reaction that leads to inhibition of established parental adenocarcinomas, as well as an antigenically unrelated fibrosarcoma. The ensuing systemic immunity selectively inhibits subsequent parental cell challenges and cures established parental adenocarcinomas and their lung metastases, although less effectively as the neoplastic mass increases. Multiple injections of interleukin 2-gene-transfected tumor cells may thus be considered a new form of vaccination in the management of minimal residual disease and incipient metastases.

Adenocarcinoma↗
PubMed1993

Ultrastructural evidence of the mechanisms responsible for interleukin-4-activated rejection of a spontaneous murine adenocarcinoma.

The ultrastructural pattern of the anti-tumor response elicited by interleukin-4 (IL-4) was investigated by using a spontaneous mammary adenocarcinoma (TS/A) unable to elicit protective immunity in syngeneic BALB/c mice as suggested by a variety of preimmunization-challenge experiments. A subcutaneous lethal challenge of TS/A tumor cells was inhibited in a significant number of BALB/c mice receiving recombinant murine IL-4 injected daily for 10 days around the tumor-draining lymph node. Tumor rejection was mainly the result of direct membrane and cytoplasmic damage to tumor cells by eosinophils, neutrophils and macrophages that deeply penetrated the proliferating tumor mass. Lymphocytes and fibroblasts participated in the reaction by interacting with tumor cells, granulocytes and each other. The most frequent cell interactions in the peri- and intra-tumoral areas and in the tumor-draining lymph nodes are illustrated. The efficiency with which the IL-4-activated reaction leads to tumor inhibition and induction of a T-lymphocyte-dependent tumor-specific immune memory appears to depend on interactions between distinct leukocytes.

Adenocarcinoma↗
PubMed1993

Selective transformation of host lymphocytes in vivo by retrovirus-producing macrophages.

Macrophage-mediated retroviral transformation of host cells was studied in vivo utilizing the cloned murine macrophage-line GG2EE, generated by in vitro infection of bone marrow cells from C3H/HeJ mice (H-2k) with the acute transforming retrovirus J2 bearing the v-myc and v-raf oncogenes. Because GG2EE macrophages produce the J2 retrovirus, the development of secondary, J2 virus-induced tumors after the injection of the cell line into several strains of mice was evaluated. GG2EE cells proliferated and gave rise to histiocytic tumors in syngeneic mice and in allogeneic athymic Swiss mice. The inoculum of GG2EE cells in allogeneic DBA/2 mice (H-2d) and, to a lesser extent, in BALB/c (H-2d) and BALB/k (H-2k) mice gave rise to a small, solid mass at the injection site. Although the initial tumor was slowly rejected, secondary lymphomas belonging to the B or T cell lineage developed, leading to mouse death. Extensive phenotypic, functional, and chromosomal analyses proved that lymphomas were derived from host T and B cell transformation. Southern and Northern blot studies showed that J2 virus was integrated and expressed in lymphoma cells, demonstrating that the virus was transmitted to the host lymphocytes and suggesting that it was causal in lymphoma development. The existence of close and protracted interactions between GG2EE macrophages and allogeneic host lymphocytes and the presence of viral particles in the area of macrophage-lymphocyte contact were demonstrated by histologic and ultrastructural analysis. Rejection of J2 virus-infected lymphocytes in allogeneic mice suggested that host lymphocyte transformation was dependent upon the macrophage cell type. These results demonstrate that macrophage-derived J2 retrovirus transforms host lymphocytes in vivo in allogeneic mice and that a condition of host alloreactivity is critical for such event.

Animals↗
PubMed1993

Modulation of interferon-gamma receptor during human T lymphocyte alloactivation.

Previous work has shown that neutralization of physiologically secreted interferon(IFN)-gamma or blockade of its receptor during T lymphocyte activation inhibits both proliferation and cytotoxic T lymphocyte generation, suggesting that IFN-gamma plays a crucial role in T lymphocyte induction and differentiation. In this study, the kinetics of the surface expression of the 90-kDa IFN-gamma receptor (IFN-gamma R) was followed during human mixed lymphocyte reaction (MLR) to alloantigens. IFN-gamma R mRNA is constitutively expressed on resting peripheral blood lymphocytes emerging from nylon wood column (NW-PBL) and its expression increases two- to threefold on alloactivated NW-PBL. IFN-gamma R protein is poorly expressed on the membrane of resting CD3+ cells, but up-modulates after 3-day MLR and sharply down-modulates at day 6. Both the p55 and the p75 chains of interleukin-2 receptor (IL-2R) were shown to up-modulate in parallel with IFN-gamma R, whereas they were still highly expressed at day 6. After alloactivation, IFN-gamma and IL-2 secretion starts at 24 h, peaks at day 3 and decreases just when IFN-gamma R and IL-2R begin to up-modulate. Proliferation peaks at day 6. Lastly, stimulation with distinct cell populations showed that the intensity of lymphocyte proliferation, IFN-gamma R membrane up-modulation, and IFN-gamma and IL-2 secretion are regulated in a parallel manner, thus suggesting that they are interrelated. Taken as whole these results demonstrate that increased expression of IFN-gamma R on T lymphocytes can be a critical event during their activation, and strongly support the hypothesis that IFN-gamma/IFN-gamma R interaction provides a signal for its progression.

Cell Line↗
PubMed1993

Strategies for cytokine utilisation in tumor therapy.

The state of the art with regard to the employment of various cytokine-based tumor immunotherapy strategies and their mechanisms of action are critically reviewed. As matters now stand, adoptive transfer of LAK cells or tumor infiltrating lymphocytes together with high doses of IL-2 constitutes the only immunologic way to hinder tumor growth in advanced stages of cancer. On the other hand, many experimental data show that the local presence of cytokines, either injected repeatedly at tumor site or released by cytokine-gene engineered tumor cells, arouses immunogenicity in apparently nonimmunogenic spontaneous tumors. By strengthening the notion that most tumors are potentially immunogenic, these findings offer substantial evidence to stress the potential use of cytokines as a component of new tumor vaccines.

Animals↗
PubMed1993

Cytokine-induced tumor immunogenicity: from exogenous cytokines to gene therapy.

The local presence of cytokines can drastically alter tumor host immune relations and activate a nonspecific reaction that in some cases leads to induction of specific responses to otherwise nonimmunogenic tumors. The employment of cytokines in the creation of new antitumor vaccines is thus a tempting prospect. Analogous effects have been obtained with cytokines inoculated locally and cytokines released from tumor cells engineered to produce them. An account is given of some mechanisms whereby this cytokine-induced reaction results in increased tumor immunogenicity. However, the real value of this potential form of vaccine in inducing the regression of incipient or established tumors remains to be established.

Animals↗
PubMed1993

Quantitative ultrasonic analysis of myocardium in patients with thalassemia major and iron overload.

BACKGROUND: Patients with beta-thalassemia major present with severe anemia and need continuous transfusion therapy. The consequent iron overload leads to hemochromatosis. Initial cardiac dysfunction has been documented even in thalassemics without clinical manifestations of heart failure as well as by conventional echocardiographic-Doppler techniques. The purpose of this study was to assess the acoustic quantitative properties of myocardium in patients with iron overload. METHODS AND RESULTS: Thirty-eight patients with beta-thalassemia major, without clinical signs of cardiac failure, and 20 age- and sex-matched young controls were studied by echocardiography. An on-line analysis of the ultrasonic radiofrequency signal was performed to obtain quantitative operator-independent measurements of the integrated backscatter (IB) signal of the ventricular septum and the posterior wall. The integrated values of the radiofrequency signal were normalized for the pericardial interface and expressed in percent (IB%). Thalassemic patients had been receiving transfusion therapy for 16 +/- 5 years and had received 313 +/- 138 transfusion units; they all had received chelation treatment (desferroxiamine) for 9 +/- 2 years. Patients and controls showed comparable values of echocardiographically assessed percent fractional shortening (32 +/- 3% versus 36 +/- 4%, p = NS), whereas thalassemics showed higher values of left ventricular mass index (118 +/- 30 versus 98 +/- 15 g/m2, p < 0.05). The IB% values were higher in patients with thalassemia major than in controls for both septum (35 +/- 14% versus 21 +/- 6%, p < 0.001) and posterior wall (16 +/- 6% versus 11 +/- 3%, p < 0.001). In thalassemic patients, no significant correlation was found between the septum IB% value and hematological parameters, such as the total number of transfusions (r = 0.2, p = NS) or the mean ferritin value (r = 0.1, p = NS). No significant correlation was also found between the septum IB% value and the echocardiographically assessed left ventricular mass index (r = 0.2, p = NS). CONCLUSIONS: These data demonstrate that myocardial reflectivity is abnormally increased in patients with thalassemia major under transfusion treatment, probably due to myocardial iron deposits and/or secondary structural changes. These quantitatively assessed abnormalities in regional reflectivity can be detected when conventional echocardiographic parameters of systolic left ventricular function are undistinguishable from normal controls.

Adolescent↗