Biomedical subjects

M Piccoli

Publications and source records attributed to M Piccoli.

At least 73 records · Page 4Linked to original sources

PubMed1992

Differential expression of granzyme A and granzyme B proteases and their secretion by fresh rat natural killer cells (NK) and lymphokine-activated killer cells with NK phenotype (LAK-NK).

Granzymes A-G are a family of serine proteases localized in the cytoplasmic granules of cytotoxic T lymphocytes (Masson, D. et al. Cell 1987. 49: 679) and granzyme A is secreted by T lymphocytes in response to antigenic stimulation. Granzyme A is also expressed by natural killer (NK) and lymphokine-activated killer (LAK) cells. Here we show that fresh rat NK cells constitutively express granzyme B and that granzyme A and granzyme B are differentially regulated in unstimulated NK cells vs. LAK cells with NK phenotype (LAK-NK cells). We also show that both granzymes A and B are secreted in a calcium-dependent manner, by NK and LAK-NK cells in response to stimuli which trigger NK cell functions.

Animals↗

PubMed1992

[Imaging diagnosis, echography and CAT in mechanical jaundice].

In a review of 70 cases of extra-hepatic cholestasis of different origin, the great diagnostic value of imaging techniques, echography and computed tomography, was demonstrated in distinguishing the type of jaundice, and in defining the cause and site of obstruction.

Adult↗

PubMed1991

Human natural killer cells express VLA-4 and VLA-5, which mediate their adhesion to fibronectin.

Very late Ag (VLA)-3, VLA-4, and VLA-5, belonging to the beta-1 subfamily of integrins, have been recently identified as receptors for different binding regions of fibronectin (FN). We have detected VLA-4 and VLA-5, but not VLA-3, on fresh CD3-, CD16+, CD56+ human NK cells by flow cytometry and immunochemical analyses using mAb directed against beta-1, alpha-3, alpha-4, and alpha-5 subunits. Binding assays, performed on FN-coated plates, showed that NK cells specifically adhere to FN and their binding capacity is increased by MgCl2 but not by CaCl2. Using as inhibitory probes a polyclonal antibody against the beta-1 chain of the human FN receptor, the synthetic peptide GRGDSP, which is able to inhibit cellular adhesion mediated by VLA-5, the CS1 fragment, which contains the principal adhesion site in the IIICS domain recognized by VLA-4, and functional mAb directed against alpha-4 or alpha-5 subunits, we show that both VLA-4 and VLA-5 mediate the adhesion of human NK cells to FN. The expression of these integrin receptors may be relevant for NK interaction with extracellular matrix components and other cell types.

Amino Acid Sequence↗

PubMed1991

Inhibition of NK cell generation by Corynebacterium parvum.

Treatment of mice with Corynebacterium parvum (Cp) resulted in a substantial decrease of splenic NK activity associated with a reduced number of LGL. Cp also inhibited in vitro augmentation of NK cytotoxicity by IFN or IL-2 as well as generation of LAK activity. Localization experiments by using radiolabelled LGL indicated that the lower number of LGL in the spleen was not attributable to a Cp-induced alteration of LGL homing. Finally Cp was found to affect the ability of bone marrow cells to reconstitute NK activity in lethally irradiated mice, indicating that it can interfere with development of NK cells from bone marrow progenitors.

Animals↗

PubMed1990

Characterization of Corynebacterium parvum-induced suppressor cells of mouse NK and ADCC activity.

Treatment of mice with Corynebacterium parvum (Cp) resulted in a substantial decrease in natural killer activity in the spleen at 10 days. The decrease in cytotoxicity was associated with the presence of splenic nonadherent (NA) suppressor cells, capable of inhibiting natural as well as antibody-dependent cellular cytotoxicity (ADCC). The nonadherent suppressor cells appeared to be null cells, lacking detectable expression of Thy 1, L3T4 (CD4), Lyt 2 (CD8), or asialo-GM1 and could be physically separated from cells with NK activity by centrifugation on Percoll discontinuous density gradients. Our results indicate that Cp can negatively modulate cytolytic functions of NK cells by inhibiting the effector phase of cytotoxicity.

Animals↗

PubMed1990

Augmentation of mouse natural killer (NK) activity by GM-1/P, a processed form of monosialoganglioside GM-1.

We describe the immunomodulatory activity of GM-1/P a processed form of GM-1 (monosialoganglioside) extracted from ox brain, purified and physically modified. We examined the effect of in vivo and in vitro treatment of GM-1/P on natural (NK) activity and its ability to induce the production of interleukin-2 (IL-2) in the mouse. In vivo treatment with GM-1/P (1 mg/Kg, i.v., day-1) resulted in a marked increase and in a change of distribution of NK activity, which was associated with lower density Percoll fractions. Marked increase was already observed at 18 hrs and then declined by day 4. In vitro treatment with GM-1/P (2 micrograms/ml) enhanced NK activity of B6 spleen cells, already after 6 hours of incubation, remaining at plateau levels within 18 hours. A role of IL-2 in this enhancement was suggested by the ability of an anti-IL-2 rabbit antiserum to abolish in vitro increased cytotoxicity. The presence of IL-2 in the supernatants of splenocytes from GM-1/P (1mg/Kg, i.v., ,day-1) treated mice stimulated with Con A or Con A plus TPA for 48 hrs was evaluated by proliferation of an IL-2 dependent CTLL cell line. GM-1/P by itself was unable to stimulate IL-2 production; however it markedly increased IL-2 production induced by Con A or Con A plus TPA.

Adjuvants, Immunologic↗

PubMed1990

Enhancement of lymphocyte proliferation and IL-2 receptor expression by a processed form (GM-1/P) of monosialoganglioside GM-1.

In this study we investigated the ability of GM-1/P, a calcium mediated processed form of monosialoganglioside GM-1, of in vivo augmenting mouse T and B-lymphocyte blastogenesis induced by mitogens. We have also determined its effect on IL-2 responsiveness by analyzing the induction of the expression of IL-2 receptor (IL-2r) on mouse spleen cells. Lymphocyte blastogenesis was evaluated by 3H-TdR incorporation of spleen cells from untreated or GM-1/P (1mg/Kg, i.v., day-1) treated mice cultured in the presence of T (PHA, ConA) B (LPS) cell specific mitogens. The stimulatory effects appeared to be due to a direct action on T and B lymphocytes, since proliferative response was not abolished by removal of macrophages. Splenocytes from GM-1/P treated mice showed increased proliferation in response to various concentrations of HrIL-2; moreover under these conditions an increased generation of LAK activity was found. A direct evidence for enhanced expression of IL-2r was obtained by immunofluorescence and FACS analysis using a monoclonal antibody (PC.61) directed against the p55 subunit of murine IL-2r. 29% PC.61+ cells were found in IL-2 cultures from treated spleen cells.

Animals↗

PubMed1990

Local/regional recombinant interleukin 2 (rIL-2) immunotherapy of tumors. Intra-arterial continuous infusion of rIL-2 in bladder cancer patients: a phase I study.

We have investigated clinical and immunological effects of two 5-day cycle continuous infusion of four escalating doses (3,000; 30,000; 300,000; 3,000,000 U/m2/day) of intra-arterial (i.a.) rIL-2, in ten patients with superficial (Ta-T1, N0, M0) bladder TCC. Tumor TUR was performed four days after the end of the second IL-2 cycle. No clinical or laboratory toxicity was detected in any but one patient, who developed grade III hypotension and grade III neurological toxicity. Two CR and two PR were observed in patients treated with 30,000 and 300,000 U/m2/day of rIL-2, respectively. Clinical responses lasted 8+, 8+, 4+ and 4+ months, respectively. Two out of ten patients developed recurrences two months after tumor resection. In the peripheral blood, no changes in the percentage of T (CD3+) lymphocytes were observed, while a significant increase of CD3+ CD16+ T cells was found in 6/9 patients. In contrast, NK (CD3- CD16+) cells were augmented only in 2/9 patients. An increase of B lymphocytes (CD19+ cells) and monocytes (CD14+ cells) was also observed in 3/9 and 7/9 patients, respectively. Lymphocyte activation marker expression (CD25, CD71, HLA-DR) increased mainly on T lymphocytes. NK and LAK activities were enhanced in 4/10 patients, while ADCC activity augmented in 2/10 patients. No detectable increased levels of plasmatic lymphokines (gamma-IFN and alpha-TNF) were found. Enhanced CD8+ and CD4+ tumor infiltrating lymphocytes were demonstrated after IL-2 treatment. Moreover, at the tumor site, lymphocytes expressing CD25 and HLA-DR antigens were noted. Tumor infiltrating macrophages were positive for IL-1 alpha, IL-1 beta and alpha-TNF.

Adult↗

PubMed1989

Rat natural killer cells synthesize fibronectin. Possible involvement in the cytotoxic function.

The ability of NK cells to synthesize and secrete fibronectin (FN), an extracellular matrix glycoprotein which plays a key role in many biologic processes including cellular adhesion, morphology, cytoskeletal organization, cell migration, and invasiveness, was studied. By using affinity-purified polyclonal antibodies directed against human cellular or plasma FN, the presence of FN was evidentiated on Percoll-purified rat large granular lymphocyte or on a large granular lymphocyte tumor cell line (CRC) by flow cytometry and immunoelectron microscopy. Its expression increased after NK cell activation by poly I:C administration. Biochemical analysis by immunoprecipitation and SDS-PAGE indicated that FN was associated to cell surface and secreted in the supernatant in a molecular form similar to that of FN from L929 fibroblasts. In an attempt to understand the role of FN in the NK cell function, we found that an antibody against human plasma FN and its F(ab')2 fragment inhibited NK cytotoxicity against YAC-1 target at the effector cell level. Inhibition occurred at the postbinding level, because F(ab')2 anti-FN inhibited induction of phosphatidylinositol hydrolysis by YAC-1 target cells, whereas binding to target cells was not affected. The possible role of FN in the NK cytotoxic function is suggested.

Animals↗

PubMed1989

Granzyme A expression by normal rat natural killer (NK) cells in vivo and by interleukin 2-activated NK cells in vitro.

Granzyme A (TSP-1) is a serine esterase expressed in cytotoxic T and natural killer (NK) cell lines. Its presence in in vivo primed T cells is disputed at present. Here we show that normal rat NK cells contain granzyme A in vivo and that its expression is augmented by their short-term in vitro treatment with IL 2. Granzyme A expression in a NK cell population with LAK activity has also been examined.

Animals↗

PubMed1989

Effects of protein kinase C (PK-C) activators and inhibitors on human large granular lymphocytes (LGL): role of PK-C on natural killer (NK) activity.

The role of protein kinase C (PK-C) in the early metabolic events involved in human natural killer (NK) cell activation has been studied through the action of PK-C-specific activators and inhibitors. Highly purified human large granular lymphocytes (LGL) were treated for 1 hr with the diacylglycerol analog 1-oleoyl-2-acetyl glycerol (OAG) (10(-4)-10(-5) g/ml) or with 12-O-tetradecanoylphorbol-13-acetate (TPA) (10(-8)-10(-10) g/ml), both specific activators of PK-C. Both these agents consistently increased NK activity against K562 target cells. Suboptimal doses of either OAG or TPA also synergized with Ca2+ ionophores to augment spontaneous cytotoxic activity. Pretreatment of LGL with 1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrocloride (H7) (5-40 microM), a potent PK-C inhibitor, greatly reduced NK activity in a time- and dose-dependent fashion. By contrast, N-(2-guanidinoethyl)-5-isoquinolinesulfonamide hydrochloride (HA 1004), a potent cAMP- and cGMP-dependent PK inhibitor with almost no effect on PK-C, marginally reduced NK activity. Moreover, almost complete NK activity inhibition was observed when H7 (10 microM), but not HA 1004 (50 microM), was present in the NK assay. Finally, 48 hr stimulation of LGL with TPA (10(-6) g/ml), a treatment able to inactivate most of the PK-C cellular pool, almost completely abrogated NK activity. This functional evidence was supported by phosphorylation of several endogenous substrates which occurs within 5 min in TPA-treated LGL. Two proteins of 70 and 56 kDa have been identified as major PK-C substrates, together with other phosphorylated proteins with MW ranging from 177 to 43 kDa. H7, but not HA 1004, almost completely inhibited the TPA-induced phosphorylation of all of these proteins in the NK cells. These data strongly suggest that selective activation of PK-C plays an essential role in the mechanisms of NK cell activation.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

PubMed1989

Laminin synthesis by NK cells and modulation of its expression by TPA (12-O-tetradecanoylphorbol-13-acetate).

Natural killer (NK) cells have been suggested to play a major role in resistance against metastatic spread of tumors. This study was aimed at understanding whether laminin (LM), a component of the extracellular matrix involved in the mechanism of tumor invasion and cell interaction, is expressed by NK cells. The results indicate that NK cells can synthesize and display on the cell surface LM and that TPA can modulate its expression. Our findings suggest that the presence of LM on NK cells could be relevant in the control of tumor invasion by NK cells.

Animals↗

PubMed1989

Overnight incubation of mouse spleen cells in recombinant IL-2 generates cytotoxic cells with NK characteristics from precursors enriched with or devoid of LGL.

Interleukin-2 (IL-2) augments natural killer (NK) activity as well as generating effector cells named lymphokine activated killer cells (LAK) which are capable of lysing a wide spectrum of target cells. A large body of evidence has been accumulated to evaluate the relationship between NK and LAK cells and conflicting results have been reported. Our study was addressed to further analyse this relationship and in particular to investigate whether in a short incubation IL-2 is merely capable of augmenting the activity of pre-existing killer cells, or whether it can also promote the differentiation of precursor cells. Eighteen-hour culture of mouse spleen cells in human recombinant IL-2 induced a DNA-synthesis-independent generation of cytotoxic cells bearing an NK phenotype (aGM-1+, Thy1.2+/-, CD8-, CD4-). These were generated from precursor cells also bearing an NK phenotype, recovered either from low density Percoll fractions enriched in lytic cells with LGL morphology as well as from high density fractions devoid of LGL and cytotoxic activity.

Animals↗

PubMed1988

Proliferative effects of 12-O-tetradecanoylphorbol-13-acetate (TPA) and calcium ionophores on human large granular lymphocytes (LGL).

The proliferative responses of natural killer (NK) cells to 12-O-tetradecanoylphorbol-13-acetate (TPA), which directly activates protein kinase c(PKC), and to the Ca2+ ionophores A23817 and ionomycin, known to enhance the intracellular calcium, have been investigated. Highly purified large granular lymphocytes (LGL) were cultured for 12-30 hr in the presence of TPA, ionomycin, or A23817. TPA alone (1-20 ng/ml) triggered rapid LGL proliferation, whereas the calcium ionophores were ineffective. The addition of either calcium ionophore to suboptimal doses or TPA (0.1-0.5 ng/ml) resulted in a synergistic effect on LGL proliferation. Under these conditions high levels of IL-2 activity were released by the LGL. Phenotypic analysis revealed the rapid loss of the Fc gamma receptors (CD16) on LGL and the induction of the expression of IL-2 (CD25) and transferrin receptors and of HLA-DR, but not of CD3. Removal of extracellular Ca2+ by addition of EGTA at the beginning of the culture greatly depressed LGL proliferation and IL-2 production, and blocked phenotypic changes, such as the expression of Tac antigen. Finally, progression to the proliferative phase of LGL, activated by TPA alone or with ionomycin, was completely abrogated by a hyperimmune anti-IL-2 antiserum.

Antigens, Surface↗

PubMed1986

Sequential metabolic events and morphological changes during in vivo large granular lymphocyte activation and proliferation.

Interferon (IFN) and IFN inducers are known to boost natural killer (NK) activity in vivo and in vitro. In vivo enhancement of NK activity results from activation of preexisting NK cells as well as from an increased number of large granular lymphocytes (LGL), with a portion of them undergoing cell division. Our study was addressed to analyze the sequence of metabolic events occurring within the LGL population of Fischer rats treated with poly(I:C), as an IFN inducer. The increase in cytotoxic activity and LGL number in the peripheral blood already reached maximal levels by 12 hr after poly(I:C) injection, remained on a plateau 24 to 48 hr later, then slightly decreased on Day 4, and returned to control levels by Day 6. A similar kinetics was observed for RNA synthesis. In contrast DNA synthesis first increased at 24 hr, peaked at 48 hr, then decreased on Day 4, and was not detectable on Day 6. Percoll fractionation resulted in 92-97% of LGL in fraction 1, and cells in this fraction accounted for the increase of cytotoxicity as well as for newly synthesized RNA and DNA. However, LGL recovered on Day 1 or 2 after poly(I:C) stimulation displayed quite heterogeneous morphology, and a number of mitotic configurations were seen on Day 2 within the LGL population. Our results indicate that the boosting of NK activity by poly(I:C) is always associated with an increase in LGL numbers, the enhanced lytic capacity is associated in vivo with new RNA synthesis by the NK cells, and only in a later phase NK cell proliferation may account for the increase in LGL numbers.

Animals↗

PubMed1986

Evidence for disregulation in the control of Epstein-Barr virus latency in patients with AIDS-related complex.

Patients affected by AIDS-related complex (ARC) show several immunological abnormalities which may lead to a disregulation of immunosurveillance against viral latent infections. In this paper evidence for Epstein-Barr virus (EBV) reactivation in seven out of eight affected by persistent generalized lymphadenopathy is given. These patients showed either elevated levels of circulating EBNA-positive transformed cells or depressed EBV-specific T cell cytotoxicity, as assessed by the regression assay, or both. A direct involvement of EBV in the pathogenesis of ARC is thus suggested. Natural killer cell activity was found decreased, correlating to the evidence of circulating EBV-infected cells and of impaired EBV-specific immune control. These data provide evidence that, when specific immune mechanisms lose control on ubiquitous latent viruses, the risk for reactivation becomes higher. In the case of EBV, direct evidence of this event is provided by the emergence in the peripheral blood of clones of infected cells with unlimited growth potential.

AIDS-Related Complex↗

PubMed1985

Changes in number and density of large granular lymphocytes upon in vivo augmentation of mouse natural killer activity.

NK activity of mice as well as humans and rats has been clearly associated with large granular lymphocytes (LGL). To better understand the effects of interferon (IFN) and IFN inducers on natural killer (NK) cells, we have compared the LGL in the spleens of normal and boosted mice. Cells were fractionated by centrifugation on discontinuous Percoll density gradients, and each fraction was tested for NK activity against YAC-1 targets and for the presence of LGL. In vivo treatment with C. parvum (0.7 mg/mouse, i.p., day-3), MVE-2 (25 mg/kg, i.p., day-3), poly I:C (4 mg/kg, i.p., day-3), or IFN (10(5) U/mouse, i.p., day-1) resulted in a marked augmentation and a change of distribution of cytotoxic activity. Most of the NK activity of boosted spleen cells was associated with lower density fractions 1 and 2, whereas active normal spleen cells had somewhat higher density (fractions 2 and 3). In parallel to their increased reactivity, the boosted spleens had a marked increase in the percentage of LGL, particularly in fractions 1 and 2. The augmented activity appeared to be mediated by the LGL, because treatment with anti-asialo GM1 or anti-Thy-1.2 plus complement reduced NK as well as the number of LGL. These results indicate that IFN-mediated boosting of NK activity in the spleen is due to an increase in the lower density LGL, as well as to an increase in the function of preexisting NK cells.

Adjuvants, Immunologic↗

PubMed1984

Bimodal effects of MVE-2 on cytotoxic activity of natural killer cell and macrophage tumoricidal activities.

Maleic anhydride divinyl ether of molecular weight 15,500 (MVE-2) increased tumoricidal activity of NK cells and M phi in a dose-dependent manner with the peak response of both effector cells occurring 3 days following treatment. Both effector cell responses were sustained for over 7 days following one injection. However, repeated injections with MVE-2 led to a hyporesponsiveness of NK cells whereas M phi activity remained high. Poly ICLC, but not C. parvum, reconstituted NK cell activity in mice hyporesponsive to MVE-2. Significant antitumor response was achieved when tumor cells were inoculated at the peak time of effector cell response, indicating the in vivo role of NK and/or M phi in exerting a tumoricidal effect.

Adjuvants, Immunologic↗