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Biomedical subjects

I Nicoletti

Publications and source records attributed to I Nicoletti.

At least 55 records · Page 3Linked to original sources
PubMed1993

The acute promyelocytic leukemia-specific PML-RAR alpha fusion protein inhibits differentiation and promotes survival of myeloid precursor cells.

Acute promyelocytic leukemia is a clonal expansion of hematopoietic precursors blocked at the promyelocytic stage. The differentiation block can be reversed by retinoic acid, which induces blast maturation both in vitro and in vivo. Acute promyelocytic leukemia is characterized by a 15;17 chromosome translocation with breakpoints within the retinoic acid alpha receptor (RAR alpha) gene on 17 and the PML gene, which encodes a putative transcription factor, on 15. A PML-RAR alpha fusion protein is formed as a consequence of the translocation. We expressed the PML-RAR alpha protein in U937 myeloid precursor cells and showed that they lost the capacity to differentiate under the action of different stimuli (vitamin D3 and transforming growth factor beta 1), acquired enhanced sensitivity to retinoic acid, and exhibited a higher growth rate consequent to diminished apoptotic cell death. These results provide evidence of biological activity of PML-RAR alpha and recapitulate critical features of the promyelocytic leukemia phenotype.

Animals↗
PubMed1993

Interleukin-4 protects double-negative and CD4 single-positive thymocytes from dexamethasone-induced apoptosis.

Glucocorticoid hormones (GCH) and anti-CD3 monoclonal antibodies (MoAbs) induce in mouse thymocytes and T-cell tumor lines an active process of cell death called apoptosis. Interleukins (IL), including IL-1 and IL-2, have been reported to inhibit such apoptosis. In this study we show that IL-4 also reduced the DNA fragmentation characteristic of dexamethasone (DEX)-induced apoptosis in thymocytes. This effect, studied in both time-course and dose-response experiments, was also detected at low IL-4 concentrations (1 U/mL) and against high DEX levels (10(-7) mol/L). The effect of IL-4 was blocked by an anti-IL-4 but not by an anti-IL-1 alpha MoAb, and was thus both specific and direct. Phenotypic analysis showed that IL-4 protects predominantly CD4-CD8- and CD4+CD8- cells. Our findings suggest that intrathymic T-cell development may be influenced by IL-4.

Animals↗
PubMed1993

Interleukin-2 induces apoptosis in mouse thymocytes.

Interleukins play a role in the process of T-cell development and, like other cytokines, seem able to modulate apoptosis. Interleukin-2 has been reported to inhibit apoptotic cell death of thymocytes induced in vitro by either activation of CD3/TCR complex or treatment with glucocorticoid hormone. We demonstrate here that IL-2 can provoke DNA fragmentation and cell death of CD4+ CD8+ mouse thymocytes by activating an endogenous apoptotic pathway. Thymocytes, incubated with high IL-2 concentrations in vitro, showed the morphological characteristics of apoptotic cells, including reduction in nuclear size, derangement in chromatin structure, and DNA fragmentation in oligonucleosomal subunits. Inhibition of mRNA and protein synthesis and addition of the PKC-inhibitor H-7, Zn2+ ions, and IL-4 counteracted the IL-2 effect. These data suggest that high IL-2 concentrations may induce an active process of cell death on mouse thymocytes in vitro.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1993

Cellular stress and glucocorticoid hormones protect L929 mouse fibroblasts from tumor necrosis factor alpha cytotoxicity.

Adaptive responses to the environment depend on the induction of the "stress response" in less differentiated organisms and cultured cells and the activation of the hypothalamic-pituitary-adrenal axis in animals and humans. This indicates that adrenal steroids and stress proteins play an important role in regulating cell survival in response to noxious stimuli. In an in vitro model, we analyzed the effects of either dexamethasone (DEX) treatment or environmental changes which can elicit a stress response, on the survival of cultured L-929 mouse fibroblasts exposed to the cytotoxic cytokine tumor necrosis factor alpha (TNF-alpha). DEX treatment produced a significant reduction in the apoptotic death of L-929 cells produced by TNF-alpha. Abrogation of the protective effect of DEX by actinomycin D and cycloheximide demonstrated that protection against TNF-alpha requires de novo synthesis of mRNA and proteins. The results were similar when L-929 cells were exposed to metabolic (serum starvation) or thermal (heat shock) stresses before TNF-alpha treatment. In both cases the stress process afforded significant protection against TNF-alpha cytotoxicity. Inhibition of mRNA and protein synthesis abrogated the protection exerted by stress (serum starvation) or produced massive death during the stress event (heat shock). The similarities in the protective activities of DEX and stress response and the reported interactions between heat shock proteins and glucocorticoid hormones suggest that stress proteins and glucocorticoids both belong to an ancient evolutionary pathway which controls cell survival.

Animals↗
PubMed1993

The S-100 protein causes an increase of intracellular calcium and death of PC12 cells.

The S-100 protein-PC12 cell interaction has been studied as a model system of the possible physiological role played by S-100 protein in the nervous system. The data reported demonstrate that S-100 exerts a cytotoxic action which eventually leads to PC12 cell death, regardless of the cell cycle phase. The effect is specific for the S-100 isoforms, which are made up of two identical subunits and is abolished by a monoclonal antibody directed against the same isoforms. Other isoforms and/or calcium-binding proteins, such as troponin or calmodulin, do not induce the same effects. The action of S-100 on cell viability is not detectable in other cell lines of different embryological origin, such as 3T3, L1210, GH3. S-100 causes a rapid and considerable increase (two- to three-fold) of intracellular Ca2+ concentration in PC12 cells accompanied by cytostatic and cytotoxic action. It is postulated that this action also occurs in vivo, as part of the physiological action of this protein.

Animals↗
PubMed1993

Genistein inhibits tumour cell growth in vitro but enhances mitochondrial reduction of tetrazolium salts: a further pitfall in the use of the MTT assay for evaluating cell growth and survival.

The natural isoflavone genistein inhibits the growth of a number of tumour cell lines in vitro. During investigations on the antiproliferative effects of genistein we observed that, with respect to direct cell counting, a tetrazolium (MTT) colorimetric assay consistently underestimated the growth inhibitory activity of the substance. Cell proliferation was markedly inhibited by genistein in three tumour cell lines (MCF-7, human breast tumour; Jurkat cells, human T-cell leukaemia; L-929, mouse transformed fibroblasts) when cell number was evaluated by direct counting, whereas a 72-h MTT assay failed to reveal any growth-inhibitory effect. Cell cycle analysis by propidium iodide staining and flow-cytometry revealed a G2/M cell cycle arrest after genistein treatment. Genistein-treated cells displayed an increase in cell volume and in mitochondrial number and/or activity, as revealed by enhanced formazan generation and increased uptake of the vital mitochondrial dye rhodamine 123. These results suggest that alterations in cell cycle phase redistribution of tumour cells by genistein may significantly influence mitochondrial number and/or function and, consequently, MTT reduction to formazan. This may constitute an important bias in analysing the effects of genistein, and possibly other drugs that block the G2/M transition, on growth and viability of cancer cells in vitro by MTT assay.

Animals↗
PubMed1993

Cytostatic and cytotoxic effects of tumor necrosis factor alpha on MCF-7 human breast tumor cells are differently inhibited by glucocorticoid hormones.

To investigate the mechanisms of growth inhibition exerted by TNF-alpha on tumor cells in vitro, we analyzed the cytokine effects on growth and cell-cycle parameters of cultured MCF-7 human breast cancer cells. TNF-alpha exerted a dose-dependent inhibition of MCF-7 cell growth, which reached its maximum at 1000 U/ml TNF-alpha concentrations. Flow-cytometric analysis of cell nuclei revealed two main components in TNF-alpha activity: an earlier cytostatic effect (G1/S block), was followed by nuclear shrinkage and cytolysis. The 55-60-kDa TNF-alpha receptor is involved in the growth inhibitory activity of the cytokine, since the H398 anti-55-kDa receptor antibody significantly counteracted the cytostatic and cytotoxic effects of TNF-alpha while an antibody (htr-9) with agonistic activity on the same receptor produced both cytostasis and cytolysis. Culture conditions strongly influenced the MCF-7 cell response to TNF-alpha. Serum deprivation of log-growing (i.e., high S phase percentage) cultures potentiated the cytotoxic effect, while reduction in S phase cell percentage by preculture in serum-free medium resulted in a significant inhibition of TNF-alpha action. Mitogenic hormones, such as insulin and 17 beta-estradiol+insulin, restored the sensitivity of MCF-7 cells precultured in serum-free medium to both the cytostatic and cytolytic effects of TNF-alpha. The synthetic glucocorticoid hormone dexamethasone, at micromolar concentrations, counteracted the TNF-alpha effect on MCF-7 cell growth. Flow-cytometric analysis showed that dexamethasone did not antagonize the cytostatic activity of either TNF-alpha or htr-9 agonistic antibody, but only the subsequent cytolysis.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies↗
PubMed1993

Hyaluronic acid modulates growth, morphology and cytoskeleton in embryonic chick skin fibroblasts.

The action that hyaluronic acid (HA) exerts on cell proliferation was investigated in embryonic chick skin fibroblasts at different ages (7-14 days) and in different cell-cycle stages evaluated by flow cytometry (cells maintained with and without serum). Proliferation was estimated by 3H-thymidine incorporation and cell counting. The results demonstrated hyaluronic acid inhibits cell multiplication in all different environmental conditions examined. The inhibitory effect of HA is more evident in 14-day than 7-day old fibroblasts. The ability of HA to modulate 3H-thymidine incorporation did not involve a change in the time required for cells entering the S phase of the replicating cycle, but is due to a smaller number of cells entering in this phase. As the relationships between components of the extracellular matrix (ECM) and the cytoskeleton are known, parallel studies were carried out on some cytoskeleton proteins. Furthermore, by modifying the capacity of cells to adhere to the substrate, HA induced alterations in cell shape and in cytoskeleton components involved in these processes. We may hypothesize that HA, binding specific membrane receptors, affects cell adhesion and morphology inducing less receptivity of fibroblasts to mitogenic stimuli by transmembrane interactions with cytoskeleton.

Actinin↗
PubMed1992

A novel transforming protein (SHC) with an SH2 domain is implicated in mitogenic signal transduction.

A new SH2-containing sequence, SHC, was isolated by screening cDNA libraries with SH2 representative DNA probes. The SHC cDNA is predicted to encode overlapping proteins of 46.8 and 51.7 kd that contain a single C-terminal SH2 domain, and an adjacent glycine/proline-rich motif with regions of homology with the alpha 1 chain of collagen, but no identifiable catalytic domain. Anti-SHC antibodies recognized three proteins of 46, 52, and 66 kd in a wide range of mammalian cell lines. These SHC proteins complexed with and were phosphorylated by activated epidermal growth factor receptor. The physical association of SHC proteins with activated receptors was recreated in vitro by using a bacterially expressed SHC SH2 domain. NIH 3T3 mouse fibroblasts that constitutively overexpressed SHC acquired a transformed phenotype in culture and formed tumors in nude mice. These results suggest that the SHC gene products couple activated growth factor receptors to a signaling pathway that regulates the proliferation of mammalian cells.

3T3 Cells↗
PubMed1992

Interleukins modulate glucocorticoid-induced thymocyte apoptosis.

Glucocorticoid hormones, calcium ionophores and anti-CD3 monoclonal antibodies induce apoptosis in mouse thymocytes. This type of cell death, which is characterized by an extensive DNA fragmentation into oligonucleosomal subunits, occurs in the intrathymic process of negative selection, and is involved in the deletion of autoreactive T-cells during thymic maturation. A number of cytokines are able to modulate apoptosis, and interleukins, including interleukin-1, interleukin-2, and interleukin-4, play a crucial role in thymic maturation and T-cell development. We tested the effects of several cytokines on the glucocorticoid hormone-induced apoptosis of mouse thymocytes in vitro, and demonstrated that interleukin-1 alpha, interleukin-2, and interleukin-4 inhibit the apoptosis induced by dexamethasone, but that interleukin-3 and interleukin-6 exert no noteworthy effect. Dose-response experiments indicated that interleukin-4 is more potent than interleukin-1 alpha and interleukin-2 in inhibiting dexamethasone-induced apoptosis. Furthermore, interleukin-4 fully inhibited the DNA fragmentation induced by the protein kinase-C activator 12-O-tetradecanoylphorbol-13-acetate, but was ineffective against apoptosis induced by the calcium ionophore A23187. These results suggest that interleukins regulate the thymic selection process by acting as modulators of the negative selection process.

Animals↗
PubMed1992

Heat shock induces apoptosis in mouse thymocytes and protects them from glucocorticoid-induced cell death.

Thymocyte death is a complex phenomenon under the control of different signals and stimuli. We evaluated the effect of elevated temperature (heat shock, HS) on mouse thymocyte apoptosis. Incubation of thymocytes at 43 degrees C for 20 min induced DNA fragmentation and cell death, but it was also able to decrease the apoptosis induced by dexamethasone (DEX), TPA or Ca2+ ionophore. The anti-apoptotic effect was correlated with induction of heat shock proteins (HSPs) and abolished by protein synthesis inhibition. On the other hand, HS-induced unlike DEX-induced apoptosis was not inhibited by protein synthesis and mRNA transcription inhibitors, the PKC inhibitors H-7 and staurosporine, or interleukin-4 (IL-4), but only by Zn2+. These results suggest that HS interferes in thymocyte death by either inducing or inhibiting thymocyte apoptosis and that the induction process mechanisms are different from those of GCH.

Animals↗
PubMed1992

IL-2-dependent generation of natural killer cells from bone marrow: role of MAC-1-, NK1-1- precursors.

We have previously shown that interleukin-2 (IL-2) is able to induce the generation of natural killer (NK) activity in bone marrow (BM) cell cultures from mice pretreated with 5-fluorouracil (5-FU). Cell fractionation experiments to analyze the nature of BM precursors indicate that MAC-1-, NK1-1- noncytotoxic precursors are induced by IL-2 to proliferate and generate cytolytic NK cells. These data demonstrate that the phenotype and functional characteristics of the IL-2-responsive cells in the FUBM are different from those of mature NK cells in that they are MAC-1+, NK1.1+, CD3- and susceptible to boosting by IFN-alpha.

Animals↗
PubMed1992

Clinical and metabolic effects of a triphasic pill containing gestodene.

The clinical and metabolic effects of a low-dose triphasic oral contraceptive containing gestodene were investigated in 42 healthy women during 6 months of treatment. No pregnancies and no severe side effects occurred during the study. The pill exerted good cycle control and the incidence of irregular bleeding was very low. As for the coagulatory system, there was an increase in prothrombin activity and in fibrinopeptide A plasma levels, and a decrease in activated partial thromboplastin time. Antithrombin III activity, fibrinogen concentration and platelets count did not change during pill intake. No significant modifications in plasma total-cholesterol, low density lipoprotein-cholesterol or in the subfraction high density lipoprotein2-cholesterol (HDL2-CH) were observed. Serum triglycerides, HDL-CH and HDL3-CH levels were significantly higher at the end of treatment. The pill did not alter fasting insulin and glucose levels or their response to an oral glucose tolerance test. These minimal effects on metabolism, combined with its high efficacy and acceptability, may suggest that this triphasic formulation with gestodene seems to be a safe and reliable contraceptive agent.

Adult↗
PubMed1992

Glucocorticoid-induced DNA fragmentation: role of protein-kinase-C activity.

Glucocorticoid hormones (GCH) and IL-2 induce apoptotic cell death by a PKC-dependent mechanism. IL-4 counteracts apoptosis by inhibiting PKC activity. GCH and IL-2 show antagonistic effects on apoptosis when administered together. These data indicate that PKC activation in response to different stimuli can both enhance or reduce thymocyte survival.

Animals↗
PubMed1992

Intestinal pseudoobstruction secondary to hypothyroidism. Importance of small bowel manometry.

Hypothyroidism may lead to secondary pseudoobstruction. We report a patient with intestinal symptoms from hypothyroidism in which previous conventional examinations were negative. Gastrointestinal manometry disclosed features of pseudoobstruction, and we discuss the importance of performing functional studies in selected cases, in as much as symptoms seemed to resolve on replacement therapy.

Aged↗