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M Piacentini

Publications and source records attributed to M Piacentini.

At least 73 records · Page 4Linked to original sources
PubMed1997

The role of proliferating cell nuclear antigen (PCNA) in differentiating idiopathic orbital inflammatory disease and lymphoid proliferations.

The purpose of the present study was to determine whether proliferating cell nuclear antigen (PCNA), an immunohistochemical marker for a nuclear protein abundant in actively proliferating (dividing) cells, is useful as an aid in differentiating idiopathic orbital inflammatory syndrome (IOIS) from lymphoproliferative lesions (LLs). Records of all patients with IOIS and LLs were studied retrospectively. Tissue biopsy specimens from four patients with IOIS and nine patients with LLs were examined. The diagnosis in each case was based on presenting signs and symptoms, orbital computed tomography (CT) and/or magnetic resonance (MR) scans, histopathologic criteria, and follow-up data consistent with the entity. These findings were correlated with the percentage of B- and T-cells in the lesions as well as with the number of cells that demonstrated staining for PCNA in formalin-fixed tissue. PCNA activity was markedly increased in the higher grade (HG) lymphoma group as compared to that in the low grade (LG) lymphoma and idiopathic inflammatory group. Lymphoma cases showed a significantly increased B-/T-cell ratio compared to IOIS lesions. PCNA activity in conjunction with the ratio of B-/T-cells may be a helpful immunohistologic adjunct for differentiating purely inflammatory lesions of the orbit from lymphoid tumors. Further studies are necessary to compare PCNA activity in fresh frozen tissue with that in formalin-fixed tissue.

B-Lymphocytes↗
PubMed1997

Tissue transglutaminase-dependent posttranslational modification of the retinoblastoma gene product in promonocytic cells undergoing apoptosis.

The retinoblastoma gene product (pRB) plays an important role in controlling both cell release from the G1 phase and apoptosis. We show here that in the early phases of apoptosis, pRB is posttranslationally modified by a tissue transglutaminase (tTG)-catalyzed reaction. In fact, by employing a novel haptenized lysis synthetic substrate which allows the isolation of glutaminyl-tTG substrates in vivo, we identified pRB as a potential tTG substrate in U937 cells undergoing apoptosis. In keeping with this finding, we showed that apoptosis of U937 cells is characterized by the rapid disappearance of the 105,000- to 110,000-molecular-weight pRB forms concomitantly with the appearance of a smear of immunoreactive products with a molecular weight of greater than 250,000. The shift in pRB molecular weight was reproduced by adding exogenous purified tTG to extracts obtained from viable U937 cells and was prevented by dansylcadaverine, a potent enzyme inhibitor. The effect of the pRB posttranslational modification during apoptosis was investigated by determining the E2F-1 levels and by isolating and characterizing pRB-null clones from U937 cells. Notably, the lack of pRB in these U937-derived clones renders these p53-null cells highly resistant to apoptosis induced by serum withdrawal, calphostin C, and ceramide. Taken together, these data suggest that tTG, acting on the pRB protein, might play an important role in the cell progression through the death program.

Apoptosis↗
PubMed1996

Apoptosis of L929 cells by etoposide: a quantitative and kinetic approach.

Exponentially growing L929 cells were continuously exposed to 1 or 10 microM etoposide (VP-16). The effects of such treatment on cell growth, cycle distribution, morphology, and selected biochemical events were examined. DNA synthesis rates were markedly decreased and the protein/DNA ratio increased (unbalanced growth). Growth was blocked, with most cells being cycle arrested by 24 h in (late S-)G2-M. An asynchronous process of cell death then developed. Cells initially shrank into eosinophilic, trypan blue-excluding bodies, which were then released into the medium, and eventually became permeable to trypan blue. Transmission electron microscopy confirmed that dying cells acquired an apoptotic morphotype, with compaction and margination of chromatin, loss of microvilli, and shrinkage of cytoplasm and nucleus. Tissue transglutaminase activity and intensity of immunostaining rapidly increased in treated cultures. Internucleosomal DNA fragmentation could not be detected by agarose gel electrophoresis, yet flow cytometry revealed that the apoptotic bodies had a very low DNA fluorescence (< or = 10% of the 2n value). In agreement with the microscopic findings, this suggested that extensive DNA degradation had occurred in dead cells. While rates of cell loss from the monolayer amounted to 21 and 57% day(-1) (1 and 10 microM VP-16, respectively), apoptotic indexes largely underestimated the extent of the process. These indexes only measured the accumulation of apoptotic bodies, i.e., the balance between their generation and disposal. The latter occurred by mechanisms similar to those that operate in tissues: "secondary necrosis" or phagocytosis by viable homotypic cells in the monolayer ("homophagy").

Animals↗
PubMed1996

Induction of "tissue" transglutaminase in HIV pathogenesis: evidence for high rate of apoptosis of CD4+ T lymphocytes and accessory cells in lymphoid tissues.

Peripheral blood mononuclear cells and lymphoid tissues from HIV-infected individuals display high levels of "tissue" transglutaminase (tTG) with respect to seronegative persons. In asymptomatic individuals, > 80% of the circulating CD4+ T cells synthesize tTG protein and the number of these cells matches the level of apoptosis detected in the peripheral blood mononuclear cells from the same patients. In HIV-infected lymph nodes tTG protein is localized in large number of cells (macrophages, follicular dendritic cells, and endothelial cells), showing distinctive morphological and biochemical features of apoptosis as well as in lymphocytes and syncytia. These findings demonstrate that during the course of HIV infection, high levels of apoptosis also occur in the accessory cells of lymphoid organs. The increased concentration of epsilon(gamma-glutamyl)lysine isodipeptide, the degradation product of tTG cross-linked proteins, observed in the blood of HIV-infected individuals demonstrates that the enzyme accumulated in the dying cells actively cross-links intracellular proteins. The enhanced levels of epsilon(gamma-glutamyl)lysine in the blood parallels the progression of HIV disease, suggesting that the isodipeptide determination might be a useful method to monitor the in vivo rate of apoptosis.

Apoptosis↗
PubMed1996

Differential growth of N- and S-type human neuroblastoma cells xenografted into scid mice. correlation with apoptosis.

This study concerns the role of apoptosis in the growth of human neuroblastomas transplanted into immunodeficient SCID mice. Human neuroblastoma cell lines may consist of one or more distinct phenotypes including the neural 'N-type' and flat substrate-adherent 'S-type'. A differential phenotype-specific proliferation was apparent among S- and N-type cell clones transplanted into SCID mice when compared with the wild-type SK-N-BE(2) cell line. This differential growth capacity of the tumours was correlated with spontaneous apoptosis. Another SK-N-BE(2)-derived cell line (TGA), displaying high levels of apoptosis upon stable transfection with the full length 'tissue' transglutaminase (tTG) cDNA, was unable to induce tumour development when xenografted into SCID mice. To support these observations, the expression of apoptosis-related genes (i.e., bcl-2, p53, and tTG) in the various neuroblastomas was also investigated.

Animals↗
PubMed1995

Tamoxifen and somatostatin affect tumours by inducing apoptosis.

Tamoxifen is a commonly used chemotherapeutic agent in human breast cancer, although some tumours develop resistance. Somatostatin is also being introduced as an anti-tumour agent. Here we show that the action of these drugs is, at least partly, due to their induction of apoptosis. Both 50 nM somatostatin, and 60 nM tamoxifen significantly enhanced the percentage of cells undergoing apoptosis, when compared to untreated or oestrogen treated control cells. This effect was observed in SK-N-BE(2) human neuroblastoma cells and in MCF-7G human breast cancer cells but not in their drug-resistant counterpart MCF-7A which showed a very low rate of spontaneous programmed cell death. Finally, we propose a simple test of the sensitivity and resistance of individual tumours to these agents by assessing their ability to induce apoptosis in vitro as measured by flow cytometry.

Antineoplastic Agents↗
PubMed1994

HIV-1 gp120-dependent induction of apoptosis in antigen-specific human T cell clones is characterized by 'tissue' transglutaminase expression and prevented by cyclosporin A.

We investigated the effect of cyclosporin (CsA) on HIV-gp120-dependent induction of cell death by apoptosis of human T cell clones specific for influenza virus haemagglutinin and restricted by HLA-DR1. Preincubation of the clones with gp120 induced a large inhibition of their proliferation which was paralleled by the induction of apoptosis. Exposure to the specific antigen alone was able to trigger apoptosis in a significant fraction of cells, this effect was potentiated by pretreatment with gp120. Apoptosis was characterized by the typical morphological changes and by the expression of 'tissue' Transglutaminase (tTG), one of the few characterized effector elements of programmed cell death. Interestingly, the tTG protein induction was detectable within the first 24 hours following the gp120 treatment and preceded the appearance of the typical apoptotic phenotype. Noteworthy, CsA treatment prevented the gp120-dependent induction of apoptosis by blocking the activation of the Ca(2+)-dependent effector elements such as tTG.

Apoptosis↗
PubMed1994

Apoptosis in human skin development: morphogenesis, periderm, and stem cells.

During human skin development, embryonic- and fetal-specific periderm cells and incompletely keratinized cells are replaced by keratinocytes that differentiate while stratifying to form the fully functional epidermis. Proliferating basal cells of fetal skin also develop into epidermal appendages such as hair follicles and glands. We demonstrate that programmed cell death, not emphasized in conventional epidermal biology, has an important function in establishing the final architecture of the human epidermis and its appendages. Immunohistochemical localization of transglutaminases in fetal periderm, intermediate epidermal cells, and within appendages coincides with DNA fragmentation indicating that apoptosis is involved in deletion of these stage-specific cells and remodeling of appendages. The data also suggest that terminal differentiation of epidermal cells might be a specialized form of apoptosis. The pattern of expression of bcl-2, a gene associated with survival of some cells, is exclusive of the distribution patterns of markers of the cell death pathway. Bcl-2 protein is correlated with specific morphogenetic events in hair follicles and eccrine sweat glands, and its presence in single cells of the hair follicle bulge suggests that Bcl-2 may be a stem cell marker.

Adult↗
PubMed1994

Macrophage-colony stimulating factor (M-CSF) stimulation induces cell death in HIV-infected human monocytes.

We show here that HIV-infected monocyte-macrophages stimulated by macrophage-colony stimulating factor (M-CSF) undergo massive syncytia formation and die. The M-CSF-stimulated HIV-infected monocyte-macrophages (M/M) destroy themselves by blebbing out particles (resembling apoptotic bodies) which may contain condensed and marginated chromatin. The death of monocyte-macrophages is also characterized by the expression of "Tissue" Transglutaminase (tTG) which is one of the genes specifically expressed and activated in apoptising cells. Noteworthy, when the syncytia formation and consequently death is prevented, infected monocyte-macrophages remain viable and produce large amounts of virus for an extended period. The concentrations of M-CSF (1000 U/ml) used in this work are similar to those that stimulate macrophages in vivo. This suggests that HIV killing of M/M in the presence of M-CSF could lead, in vivo, to a greater than expected loss of immune cells and may contribute to explain the complex derangement of the immune function observed in HIV-infected patients.

Cell Death↗
PubMed1994

Immunohistochemical localization of tissue transglutaminase and Bcl-2 in rat uterine tissues during embryo implantation and post-partum involution.

Rat pregnancy is characterized by a progressive continuous induction of apoptosis in the maternal tissues lining the conceptus. Different cell types (decidual cells, giant cells, epithelial cells, etc.) undergo apoptosis at the different stages of development. We demonstrated here that, independently from their origin and stage of pregnancy, the dying cells were invariably characterized by the presence of 'tissue' transglutaminase (tTG) and negligible amounts of Bcl-2 proteins. This finding confirms the antithetic roles played by these two gene products within the apoptotic pathway. It is noteworthy that Bcl-2 was detected in decidual cells just after the implant; however, starting from day 9, its positive detection was reduced in the maternal tissues and appeared in the embryonal ones. The apoptotic nature of death was also suggested by the presence of a high phagocytic activity in the maternal involuting regions and by morphological observations showing that the tTG positive cell remnants displayed typical phenotypic features of apoptosis. The possibility that the cells lining the conceptus that die by apoptosis could be envisaged as an inert barrier between the maternal and fetal tissues is discussed.

Animals↗
PubMed1994

Abnormal Bcl-2 and "tissue" transglutaminase expression in psoriatic skin.

Cell death by apoptosis plays a key role in skin development and homeostasis. Previous studies have shown that increased apoptosis characterizes several pathologic conditions affecting human skin. Thus, the pathogenesis of cutaneous diseases may involve an imbalance in the homeostatic mechanisms determining whether the death of keratinocytes will occur by terminal differentiation or apoptosis. We investigated the involvement of apoptosis in psoriasis. For this purpose, we assessed, in addition to morphology and DNA fragmentation, the expression of two putative apoptotic genes, bcl-2 and "tissue" transglutaminase, in normal and psoriatic skin. A large number of keratinocytes showing biochemical and morphologic features of cells undergoing apoptosis was observed in all the suprabasal layers of the psoriatic epidermis. The plaques from all patients analyzed showed a dramatic reduction in the number of bcl-2-positive cells localized in the basal cell compartment. In contrast, the psoriatic lesions presented a marked induction in "tissue" transglutaminase, which was localized specifically to the cytoplasm of apoptotic keratinocytes. "Tissue" transglutaminase protein staining was undetectable in normal epidermis. The bcl-2 and "tissue" transglutaminase staining pattern observed in psoriasis also was found in the skin of patients affected by lichen planus. These findings indicate that these two genes are regulated in an opposite fashion in psoriatic keratinocytes undergoing apoptosis, thus confirming their antithetic role in the cascade of events leading to the establishment of the mature apoptotic phenotype.

Apoptosis↗
PubMed1994

Tissue transglutaminase and apoptosis: sense and antisense transfection studies with human neuroblastoma cells.

In this report, we show that the overexpression of tissue transglutaminase (tTG) in the human neuroblastoma cell line SK-N-BE(2) renders these neural crest-derived cells highly susceptible to death by apoptosis. Cells transfected with a full-length tTG cDNA, under the control of a constitutive promoter, show a drastic reduction in proliferative capacity paralleled by a large increase in cell death rate. The dying tTG-transfected cells exhibit both cytoplasmic and nuclear changes characteristic of cells undergoing apoptosis. The tTG-transfected cells express high Bcl-2 protein levels as well as phenotypic neural cell adhesion molecule markers (NCAM and neurofilaments) of cells differentiating along the neuronal pathway. In keeping with these findings, transfection of neuroblastoma cells with an expression vector containing segments of the human tTG cDNA in antisense orientation resulted in a pronounced decrease of both spontaneous and retinoic acid (RA)-induced apoptosis. We also present evidence that (i) the apoptotic program of these neuroectodermal cells is strictly regulated by RA and (ii) cell death by apoptosis in the human neuroblastoma SK-N-BE(2) cells preferentially occurs in the substrate-adherent phenotype. For the first time, we report here a direct effect of tTG in the phenotypic maturation toward apoptosis. These results indicate that the tTG-dependent irreversible cross-linking of intracellular protein represents an important biochemical event in the induction of the structural changes featuring cells dying by apoptosis.

Apoptosis↗
PubMed1994

c-myb down regulation is associated with apoptosis in human neuroblastoma cells.

The c-myb transcriptional regulator plays a crucial role in the control of several proliferative/differentiative processes in haematopoietic cells. Its expression and function is not lineage-restricted, since c-myb is also expressed in solid tumours such as neuroblastomas, where its transcription is decreased by retinoic acid. In response to retinoic acid, neuroblastomas differentiate either towards a neuronal phenotype or undergo apoptosis. The temporal relationship between reduction in c-myb mRNA levels and the differentiative/proliferative/apoptotic processes suggests that c-myb may play a key role in the control of growth of these neuroectodermal tumours. Transfection of neuroblastoma cells with expression vectors containing segments of human c-myb cDNA in antisense orientation yielded fewer transfectant clones, with a far slower proliferation rate, than transfection with the corresponding sense construct. The dramatic growth arrest and reduction in cell number in the antisense transfectants is due to the induction of apoptosis. Apoptosis in the c-myb antisense-transfected cells is further increased in reduced serum conditions.

Journal Article↗
PubMed1993

Multiple cell cycle access to the apoptotic death programme in human neuroblastoma cells.

We report the induction of apoptosis in a human neuroblastoma cell line SK-N-BE(2) by cisplatin or retinoic acid, and its relation to cell cycle. Apoptosis was monitored by counting apoptotic bodies and evaluating the activity of 'tissue' transglutaminase (EC 2.3.2.13), one of the genes specifically expressed in apoptotic cells. Data indicate that both agents enhance apoptosis, even though cells arrest at different cell cycle phases. In fact, retinoic acid causes accumulation in G1, whilst cisplatin induces accumulation of cells in the G2/M phase. This evidence suggests the presence of multiple start points for the apoptotic death programme within the cell cycle of human neuroblastoma cells.

Apoptosis↗