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D Fradelizi

Publications and source records attributed to D Fradelizi.

At least 73 records · Page 4Linked to original sources
PubMed1987

Differential effects of human and porcine interleukin 2 on natural killing (NK) activity of newborn piglets and adult pigs lymphocytes.

The in vitro effects of human or porcine Interleukin 2 (IL2) on Natural Killing (NK) activity were studied with blood lymphocytes of newborn piglets and of adult pigs. Large volumes of porcine IL2 were prepared by PHA stimulation of irradiated mesenteric lymph node cells and, following purification by gel-filtration chromatography, the apparent molecular weight of IL2 was 15,000 Da. Purified human and porcine IL2 as well as recombinant human IL2 were found to increase markedly NK activity of lymphocytes derived from adult animals. However, although newborn piglets-derived lymphocytes are sensitive to crude porcine IL2 supernatants (and to Interferon alpha as previously shown) their low NK activity is unaffected by purified IL2. These data suggest therefore the existence of differences in the ontogenic development of the lymphokines responsiveness of porcine NK cells.

Animals↗
PubMed1986

Purification to homogeneity and NH2-terminal amino acid sequence of a novel interleukin 1 species derived from a human B cell line.

A subclone, referred to as 3B6, derived from a DR-negative EBV-transformed B cell line, has been found to spontaneously produce IL 1. 3B6-IL 1 displays a pI of 5 on FPLC chromatofocusing. It has been purified to homogeneity by a sequence of ion-exchange chromatography and affinity chromatography on procion red agarose. The homogeneous material migrated with an apparent m.w. of 13,500 on SDS-PAGE. The overall recovery of IL 1 activity was estimated at 57%. The final material had a specific activity of 7.8 X 10(6) half-maximal units/mg and represented a 50,000-fold purification. A partial NH2-terminal amino acid sequence has been obtained that is different from those reported from monocytic IL 1. However, this molecule can formally be identified as IL 1 on its spectrum of biologic activities. In addition to inducing the proliferation of murine thymocytes in the co-stimulator assay. 3B6-IL 1 is active on both human T and B cells, respectively, in inducing IL 2 synthesis by cells from a subcloned HSB 2 line and promoting the proliferation of anti-IgM-stimulated human peripheral blood B lymphocytes. Furthermore, 3B6-IL 1 acts as a growth factor for normal human fibroblasts and for the 3B6 line itself. However, 3B6-IL 1 is not pyrogenic in rabbits. Thus, the 3B6 cell line was shown to produce a new molecular species of IL 1, with respect to its NH2-terminal sequence, which shared all of the studied biologic activities of monocytic IL 1 except for pyrogenicity.

Amino Acid Sequence↗
PubMed1986

Interleukin 2 (IL 2) up-regulates its own receptor on a subset of human unprimed peripheral blood lymphocytes and triggers their proliferation.

Several reports indicate that human peripheral blood lymphocytes (PBL) seeded in culture with purified or recombinant interleukin 2 (IL 2) immediately after separation from the blood display a substantial level of proliferation at day 5 or 6, even in the absence of any activating signal. The spontaneously IL 2 proliferating cells are large lymphocytes, and they co-purify on a Percoll gradient in the large granular lymphocytes (third (LGL) fraction) together with the natural killer (NK) activity. When LGL were separated into NKH1 (an NK-specific surface marker)-positive and NKH1-negative cells by fluorescence-activated cell sorting (FACS), proliferating cells were mainly found in the NKH1-negative fraction. On the contrary, when cells from Percoll fraction 3 were separated into OKT3-negative and positive cells, the majority of the proliferating cell was found in the OKT3-positive cells. These results indicate that spontaneously IL 2 proliferating (SIP) cells most probably belong to the T cell lineage, but are distinct from NK cells. Surprisingly, cells from this Percoll fraction examined immediately after separation from the blood do not express detectable amounts of IL 2 receptors as assessed by three different techniques: binding of [3H]IL 2, binding of [125I]anti-Tac antibodies, and FACS analysis with the use of anti-Tac antibodies. However, after 18 hr of culture in IL 2-supplemented medium, 5 to 7% of these cells became Tac-positive by FACS analysis. Additional analysis of IL 2 receptor induced in culture with IL 2 was performed by [125I]anti-TAC binding and by [3H]IL 2 binding. Scatchard analysis of [3H]IL 2 binding, in the range of concentrations leading to the detection of high-affinity binding sites, showed an affinity constant similar to that of conventional phytohemagglutinin blasts. The role of IL 2/IL 2 receptor interaction in the proliferation process was confirmed by the fact that proliferation, in contrast with NK activation, was clearly inhibited by anti-Tac antibodies. When LGL activated with IL 2 for 60 hr were sorted into Tac+ and Tac- cells, equal levels of NK activity was found in the two fractions. Proliferation, however, was only observed in the Tac+ population. We interpret these results to indicate that SIP cells are preactivated cells circulating in the blood. They are large cells and represent a very small proportion of circulating lymphocytes (0.3%). They express a subliminal amount of IL 2 receptor. Cultivated in the presence of IL 2, IL 2 receptor expression is enhanced to a detectable level, and the SIP cells begin to proliferate. These SIP cells could be activated T cells present in every normal individual.

Antibodies, Monoclonal↗
PubMed1986

Differentiation factors for human specific B cell response.

Nonspecific T cell factors produced by lectin-activated human peripheral blood lymphocytes (PBL) were used to restore the T-dependent B cell response to trinitrophenyl-polyacrylamide (TNP-PAA). Preincubation experiments with the particulate antigen TNP-PAA and/or a soluble TNP-protein conjugate show that a first specific signal provided by the antigen and nonspecific lymphokines sequentially acts on B cells. By gel filtration the T cell-replacing factor (TRF) activity is present in the 30-15-kDa fraction of T cell supernatants and is associated to interleukin 2 (IL2). However, absorption of IL2 does not abolish the TRF activity. Moreover, chromatofocusing of this 30-15-kDa material allows the obtaining of an IL2-free fraction containing a differentiation factor (with an isoelectric point of 5.7 +/- 0.2). The ability of this fraction to restore the anti-TNP response is manifest in the presence of a 50-kDa B cell growth factor. This latter, prepared by a combination of absorption on concanavalin A-Sepharose and gel filtration, was IL2 free and unable to support the anti-TNP response. We thus directly demonstrate that in the absence of IL2 three separate signals (the antigen, T cell-derived growth and differentiation factors) are involved in human-specific B cell response.

B-Lymphocytes↗
PubMed1986

Modulation of c-fos and c-myc mRNA levels in normal human lymphocytes by calcium ionophore A23187 and phorbol ester.

A transient expression of the proto-oncogenes c-fos and c-myc is rapidly induced by growth factors or mitogens in different cell types including lectin-stimulated lymphocytes. To elucidate the early events of lymphocyte activation, we determined (by Northern blot analysis) the steady-state levels of c-fos and c-myc mRNA in normal human peripheral blood lymphocytes (PBL) stimulated with the Ca2+ ionophore A23187 and/or 12-O-tetradecanoylphorbol-13-acetate (TPA), whose biological activities are well defined. We report that ionophore A23187 (0.5 microM) or, to a significantly lesser extent, TPA (0.5 ng/ml), neither of which is mitogenic alone at these concentrations, nevertheless can induce a transient accumulation of the proto-oncogene transcripts. These results indicate that a significant accumulation of c-fos and c-myc mRNA can occur without subsequent lymphocyte proliferation. The combination of these two mitogens increases the induced levels of both types of c-onc mRNAs. The level of mRNA accumulation correlates with the degree of proliferation induced by mitogenic combinations of ionophore A23187 and TPA, as measured by [3H]thymidine incorporation. Thus, these compounds act synergistically both with respect to c-fos and c-myc mRNA accumulation and to mitogenicity in human PBL. We also studied the level of c-fos transcripts following stimulation of the T lymphocyte T3/Ti receptor complex, using monoclonal anti-T3 antibodies. We observed that mitogenic doses of anti-T3 also induce an accumulation of c-fos mRNA, whose induction also is synergized by TPA. These results suggest that transient accumulation of c-fos; as well as c-myc, mRNA is a normal event of the immune response. They confirm that Ca2+ ionophore combined with TPA can mimic some molecular features of the early steps of normal human PBL activation.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1986

Natural killer susceptibility of human cells may be regulated by genes in the HLA region on chromosome 6.

Natural killer (NK) cells exist in each individual in the absence of any intentional immunization. They are able to kill a wide range of targets from tumoral as well as from normal origin. However, their exact physiologic role is not clearly understood. In this study we report results about a human Epstein-Barr virus-transformed B-cell line from which variants perturbed in the expression of HLA molecules have been derived. Our results indicate that in these cell lines an inverse relationship exists between expression of HLA antigens and susceptibility to NK lysis. The original cell line is highly resistant to NK lysis. On the contrary, the variant perturbed in class I antigen expression is highly susceptible. Variant perturbed in class II antigen expression is intermediate in susceptibility. Interferon, which induces HLA class I expression and NK resistance in the unrelated classical K-562 target cells, does not change either HLA expression or NK susceptibility in the variant cell lines. The difference between the original cell line and the variants does not reside in the ability to be bound by NK effectors. Our results suggest a different role for HLA molecules. By some unknown mechanism discussed here, the presence of HLA molecules at the surface of a cell would prevent this cell from being killed by NK cells. The loss of this "good health" signal would lead to the elimination of the cell through NK lysis.

Antigens, Surface↗
PubMed1986

[Production of IL2 and induction of IL2 receptor: two independent and dissociable events during allogenic stimulation in vitro in man].

Donors from HLA genotyped families are extremely useful for immunogenetic studies. Analysis of the in vitro allogenic response of lymphocytes from HLA recombinant siblings can dissect the respective role of HLA class I and class II antigens for the lymphocyte activation. Using lymphocytes from such donors, we have demonstrated that IL2 production is triggered by HLA class II disparities (but not by HLA class I), HLA class I disparities can prime the cytotoxic T lymphocytes precursors inducing the IL2 receptor on the specific clones which will proliferate in presence of IL2 either from endogenous origin (in the case of HLA class II disparities) or from experimentally added IL2 in the culture.

Cytotoxicity, Immunologic↗
PubMed1985

Accessory function of human leukemic cell lines: properties of B and B-K562 hybrid cell lines.

The accessory function (AF) of various B cell lines has been investigated by studying their ability to replace monocytes inducing the proliferation of highly purified T lymphocytes in the presence of phytohemagglutinin. AF could be exerted by B cells from different origin, including Epstein-Barr virus (EBV) transformed lymphoblastoid cell lines (LCL), Burkitt's lymphoma (BL)-derived lines, either EBV-positive or not, pre-B and B leukemias and lymphomas. The EBV-converted BJAB-B95 BL cells could exert an AF as efficient as BJAB cells, their EBV-negative counterparts, which demonstrates that EBV itself does not play any role in this function. The HLA-DR antigen-negative BL-K562 hybrids PUTKO and DUTKO, T51.4.1.6 cells which derived from an HLA-DR-negative variant of the T51 LCL and 721.84.5 cells from a mutagenically dissected HLA-DR-negative clone of the 721 LCL, also exerted very efficient AF. 721 LCL, PUTKO and DUTKO hybrids could produce interleukin 1 activity when triggered with 12-O-tetradecanoyl-phorbol 13-acetate (TPA). Furthermore, addition of anti-DR monoclonal antibodies which blocked completely the AF of the B cell lines in mixed lymphocyte-tumor cell reaction did not affect the mitogen-triggered AF exerted by the same B cell lines. The induction of cellular differentiation with TPA markedly reduced the AF exerted by all B cell lines studied as well as by the hybrid cells DUTKO. By contrast, the PUTKO hybrid was unsensitive to sodium butyrate and TPA treatments, which were unable to abrogate its AF. Taken together, these data indicate that the AF in mitogen-induced T lymphocyte responses is dependent upon some precise maturational stages of accessory cells and HLA-DR antigen expression is not required for AF in mitogen-induced proliferation assays.

Animals↗
PubMed1985

T lymphocyte function during experimental Chagas' disease: production of and response to interleukin 2.

Infection of mice with Trypanosoma cruzi results in a severe immunosuppression, accompanied by the appearance of autoimmune symptoms. We have previously shown that proliferation and interleukin 2 production by concanavalin A-stimulated T cells from infected mice is severely depressed. In this study we show that at least two phenomena are responsible for this depression. First, mixing experiments showed the existence, in spleens of infected animals, of adherent, Thy-1-negative and radioresistant suppressor cells. Second, studies of enriched T cell populations and analysis of precursors by limiting dilution showed that the T cell compartment itself was impaired in infected animals: responses of enriched T cells, even when reconstituted with normal accessory cells, reached only 40% of those obtained with normal uninfected mice.

Animals↗
PubMed1985

IL 2 receptor induction on human T lymphocytes: role for IL 2 and monocytes.

In this report we studied the requirements for the activation and proliferation of highly purified human T lymphocytes. Purified T cells incubated for 3 days with PHA neither proliferate nor express IL 2 receptors as detected by FACS analysis with the use of anti-Tac antibodies. However, purified T cells incubated with Con A or anti-T3 moAb do not proliferate, albeit 30 to 35% T cells express Tac epitopes. The addition of IL 2, either natural purified or recombinant, resulted in both the appearance of Tac antigen and the proliferation of PHA-activated T cells. Much to our surprise, IL 2 did not induce proliferation of Tac-positive T cells activated by Con A or soluble anti-T3 unless monocytes were added to the cultures. These data suggested that two classes of IL 2 receptors might exist on T cells, one of which was not functionally involved in T cell proliferation. In keeping with this interpretation, we have been able to demonstrate, using a radiolabeled IL 2 binding assay, that anti-T3 moAb induced almost exclusively IL 2 receptors of low affinity (Kd = 30 to 70 X 10(-9) M) and that additional signals, provided by monocytes, are required for the acquisition of high affinity receptors. IL 2 itself can induce high affinity receptors on PHA-stimulated T cells but not on cells activated by Con A or anti-T3. In this latter case the physical presence of monocytes is required and cannot be substituted by IL 1, thus indicating a previously unreported role for monocytes. It is postulated that the contact of monocytes with T cells induces a switch from an inactive low affinity conformation of the IL 2 receptor to a functional high affinity one.

Adult↗
PubMed1985

Functional properties of two human B cell growth factor species separated by lectin affinity column.

The supernatants from PHA-activated normal human T cells (conditioned medium) were fractionated by differential adsorption on Con A-Sepharose columns. The effluent contained both IL 2 activity (tested on the CTL-L2 cell line) and BCGF activity (tested on anti-mu-activated normal B-enriched cells). Absorption of such effluent on PHA-activated T cell blasts removed the IL 2 activity without affecting the BCGF activity. The eluate also displayed BCGF activity (tested on anti-mu-activated B-enriched cells) without detectable IL 2 activity. The two BCGF species isolated by this criteria synergize with semipurified IL 1 to support the anti-mu-induced proliferation of highly monocyte-depleted B cells. Both BCGF species required the presence of anti-mu Ab to support the proliferation of small B cells (at the G0 stage). However, in contrast to crude conditioned medium and to the BCGF present in the effluent, the BCGF present in the eluate was unable to induce the proliferation of unfractionated or large (at the G1 stage) B cells in the absence of anti-mu Ab. These results support the existence of two functionally different human BCGF species, differing at least in their relative sugar content and distinct from IL 2.

Adsorption↗
PubMed1985

Mechanism of accessory cell requirement in inducing IL 2 responsiveness by human T4 lymphocytes that generate colonies under PHA stimulation.

PHA-driven monoclonal colony formation by low concentrations of resting T4 lymphocytes in agar culture requires the presence of interleukin 2 (IL 2) and accessory cells. Using recombinant IL 2 and anti-Tac monoclonal antibody as a probe for the IL 2 receptor, we demonstrate that the requirement of accessory cells (here an irradiated B cell line) in inducing IL 2 responsiveness relies on their enhancing effect in functional IL 2 receptor expression by the T colony progenitors. Furthermore, it is shown that cell to cell interaction between accessory cells and colony progenitors results in IL 2 response, i.e., colony formation, when the IL 2 receptor density reaches a critical threshold. The asynchronism in IL 2 responsiveness expression by the T colony progenitors upon activation and the short-lived T cell-accessory cell interaction, due to accessory cell death, determine the 10% colony efficiency of the culture system. In addition, we demonstrate that the accessory function in IL 2 receptor and IL 2 responsiveness expression by the T colony progenitors can be supported by irradiated T lymphocytes as well as B cells. The absence of lineage restriction of the signal delivered by accessory cells, and the requirement of physical interaction between T colony progenitors and accessory cells, emphasize the necessity of cross-linking the activation-signal receptors in inducing IL 2 responsiveness by resting T4 cells.

Antibodies, Monoclonal↗
PubMed1985

[Abnormal regulation in the production of IL2 in Hodgkin's disease].

Patients with active Hodgkin's disease demonstrate a significant depression of cellular immunity. The present study, performed with lymphocytes from 16 untreated patients with active Hodgkin's disease and 13 healthy control volunteers, demonstrate an equivalent IL 2 production in vitro in both groups. Our results, however, demonstrate an abnormal regulation of IL 2 production in patients. A negative control of IL 2 production involving monocytes producing PGE 2 able to induce radiosensitive suppressor T lymphocytes, has been identified previously with cells from healthy donors. In the present study we demonstrate that this negative control is hyper functioning with cells from Hodgkin's disease patients.

Hodgkin Disease↗
PubMed1984

Heterogeneous accessory cell requirement for human peripheral blood T lymphocyte activation by PHA into IL-2-responsive colony-forming cells.

Mitogen-driven T cell proliferation in liquid culture requires accessory cells that cooperate in interleukin 2 production. We have investigated the accessory cell requirement for human lymphocyte colony formation under PHA stimulation. Semisolid medium limits cell-to-cell contact emphasizing the role of cooperating cells both in growth factor production and in triggering events. Culturing at high T cell density demonstrates that accessory cells can be substituted for colony formation by exogenous IL-2. Culturing at low T cell density in the presence of IL-2 also demonstrates that accessory cells are required for activation of a subset of progenitors into IL-2 responsive colony-forming cells. Consequently, T colony progenitors, contained in the E-rosetting cell fraction of peripheral blood, are heterogeneous in their triggering signals: a minor subset is directly inducible by PHA, and a major subset is inducible by PHA in the presence of accessory cells. We found that monocytes and some leukemic B cells support effective accessory function in both colony growth factor production and colony progenitor sensitization.

B-Lymphocytes↗
PubMed1984

The mechanisms of inhibition of human IL 2 production. II. PGE2 induction of suppressor T lymphocytes.

In vitro and in vivo experiments indicate that the production of interleukin 2 (IL 2) by T lymphocytes is critical for the development of the effector phase of immunity. Complex cellular interactions are involved for the induction of IL 2 production. We have shown in a previous study that in humans monocytes can transmit opposite signals to the IL 2-producing cells. In addition to the positive signal delivered through the release of interleukin 1, human monocytes can deliver a negative signal through the release of prostaglandin E2 (PGE2). This monokine, known to activate suppressor mechanisms in several systems, was shown to inhibit IL 2 production. The data presented in this paper show that this PGE2-dependent inhibition is strictly dependent upon the presence of radiosensitive T cells in the culture, suggesting that PGE2 induces the activation of suppressor T cells modulating IL 2 production. Kinetics experiments indicate that these suppressor cells are radiosensitive during their induction phase but become radioresistant after 18 hr of incubation in the presence of PGE2. Successful in vitro induction of suppressor cells by incubation of enriched T cells with PGE2 was decisive for the analysis of the phenomenon. The induced suppressors were capable of inhibiting IL 2 production by fresh autologous T cells as well as inhibiting PHA proliferative response by these cells. A quantitative evaluation of IL 2 receptors on PGE2-treated cells has indicated that this absorption capacity was similar to the capacity of PBL known to express a low number of IL 2 receptors, thus excluding a suppression via absorption or competition for IL 2. No detectable killing of IL 2-producing cells by PGE2-induced suppressors was observed. The OKT4 and OKT8 phenotype of suppressor cells was examined. T cells were purified at two stages of differentiation before or after induction by PGE2 in vitro treatment. We conclude from these experiments that PGE2 activates suppressor cells among precursors segregating predominantly with the OKT8 subset and fewer cells with the OKT4 subset. After differentiation, however, the suppressor cells segregate with the OKT8 subset only. Such results were obtained by using positive selection (cellular affinity columns) and negative selection (monoclonal antibodies plus complement).

Absorption↗
PubMed1984

Accessory function and interleukin 1 production by human leukemic cell lines.

Highly purified T lymphocytes do not proliferate in response to mitogens, unless adherent HLA-DR-positive monocytes are added to the culture. This accessory function (AF) of monocytes requires the release of interleukin 1 (IL 1). Cells from three human leukemic cell lines, K562, HL60, and U937, could very efficiently replace monocytes in a 72-hr mitogen-induced T cell proliferation assay. The AF was clearly related to precise maturational stages of these cells; the hematopoietic precursor K562 cells spontaneously exerted high AF, but lost this property when treated with differentiation inducers. On the contrary, the promyelocytic HL60 cells and the "histiocytic" U937 cells exhibited no spontaneous AF, but acquired this property when induced to differentiate along the granulocytic and/or monocytic pathway. Three leukemic cells could not only stimulate T cells to proliferate and produce IL 2 in the presence of mitogens, but also under appropriate culture conditions these cells could produce IL 1, which could not be distinguished from normal human monocyte derived IL 1 by gel filtration and isoelectric focusing. Moreover, analysis of phenotypic markers revealed that AF and production of IL 1 could be demonstrated in different cell types and therefore are not restricted to the monocytic lineage. No HLA-DR antigen could be detected on K562 and HL60 cells. Thus, the expression of the DR antigens is not required for AF and IL 1 production in response to mitogens. These three human leukemic cell lines will provide convenient sources of human IL 1.

Cell Line↗
PubMed1984

[Interleukin 2 and rheumatoid polyarthritis].

The dysimmune state associated with rheumatoid arthritis was approached by the study of a T lymphocyte function, the production of helper factor: Interleukin 2 (I12). This production was measured in 11 patients in comparison with 9 controls. The I12 activity is significantly higher in the patient group. Several mechanisms may be involved in this over-production. The radiosensitive T suppressor function, which was also investigated in this study, was the same in the two groups, so can not explain this overproduction. However, the radiosensitive suppressor function is very variable in the patients, suggesting a clinical correlation which could not be confirmed in this small group of subjects.

Arthritis, Rheumatoid↗