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Biomedical subjects

A Faggioni

Publications and source records attributed to A Faggioni.

At least 37 records · Page 2Linked to original sources
PubMed1998

Upregulation of Epstein-Barr virus-encoded latent membrane protein by human herpesvirus 6 superinfection of EBV-carrying Burkitt lymphoma cells.

The effect of HHV-6 strain A infection on the expression of Epstein-Barr virus- (EBV-) encoded growth transformation-associated genes in two EBV-positive Burkitt lymphoma cell lines, Akata and P3HR-3, was investigated. The results indicate that HHV-6A upregulates the expression of the latent membrane protein LMP-1 in both cell lines. Expression of EBNA-2 was also upregulated in Akata cells following HHV-6A infection. Transfection of reporter constructs carrying the LMP-1 regulatory sequences (LRS; -634/+40) or its 5' deleted derivatives in Akata and in a T-lymphoblastoid cell line, J-Jhan, confirmed the presence of positive and negative regulatory elements responsive to HHV-6A infection in LMP-1 regulatory sequence (LRS). The majority of LRS constructs were under the influence of dominant negative factors. HHV-6A was able to override the effect of such factors acting on reporter plasmids containing the -634/-54, -324/-54, -214/-54, and -106/-54 parts of LRS. The plasmid that carried only the -54/+40 LRS region was constitutively active in both Akata and J-Jhan cells; in Akata, its activity was influenced by HHV-6A. The finding that HHV-6A infection may activate LMP-1 and EBNA-2 expression, which is essential for the immortalization of B-lymphocytes by EBV, shows a novel aspect of the interaction between these two herpesviruses.

Antibodies, Monoclonal↗
PubMed1997

Transcription of latent and replicative Epstein-Barr-virus genes in bone-marrow and peripheral-blood mononuclear cells of healthy donors.

Reverse-transcriptase polymerase chain reaction has been used to analyze the expression of 2 latent genes (EBNA-1 and LMP-1) and one replicative gene (BZLF-1) of Epstein-Barr virus in mononuclear cells from bone marrow and peripheral blood of healthy donors. EBV-gene transcription was detected in 8 out of 15 bone-marrow samples. Among these, 5 allowed the detection of latency-associated transcripts in the absence of BZLF-1 expression. Only one sample showed positivity for expression of both latent and lytic genes. In 2 cases, BZLF-1 was the only transcript detected. In peripheral blood, 4 out of 7 samples showed evidence of EBNA-1 transcription; LMP-1 was expressed in 5 samples, and in 2 cases concomitant expression of EBNA-1 and BZLF-1 was detected. These results provide a direct demonstration by RT-PCR of EBV-gene transcription in bone-marrow-resident viral infected cells and suggest, in contrast to previous studies on peripheral blood, that LMP-1 and BZLF-1 are frequently transcribed also in absence of EBV-related disease. The heterogeneous viral gene expression found makes it difficult to define a pattern of viral latency in vivo which coincides with that described for lymphoblastoid or Burkitt's-lymphoma cell lines at different stages of differentiation.

Animals↗
PubMed1997

Neuramide inhibits Epstein-Barr virus-induced transformation activity and proliferation of transformed B lymphocytes.

Neuramide, in Italy, is a prescription drug and contains various polypeptides, with molecular weights ranging between 10,000 and 1,000. It is commonly used in Italy for the clinical therapy of varicella-zoster and other viral herpetic diseases. In the present study, we investigated its effect on the in vitro transformation activity and transformed status of the Epstein-Barr virus (EBV), ab herpesvirus closely associated with infectious mononucleosis (IM), nasopharyngeal carcinoma (NPC) and Burkitt's lymphoma (BL). Antiviral effects of neuramide were evaluated on viral-induced immortalization, transformed status and DNA synthesis. Our results show that simultaneous and/or post-treatment of EBV-infected lymphocytes with neuramide, at a concentration of 2 inhibiting units (IU)/ml, blocks EBV-mediated transformation and related events, whereas pretreatment does not show any inhibiting activity against EBV. Neuramide was also found to be a potent inhibitor of the proliferation of EBV-transformed B lymphocytes in vitro. These results suggest that the molecular mechanisms of action of the drug should be investigated more closely and also support the necessity to further purify the active polypeptides, in order to improve its possible in vivo efficacy in EBV-associated diseases, such as infectious mononucleosis and other lymphoproliferative diseases.

Antimicrobial Cationic Peptides↗
PubMed1996

Expression of Egr-1 correlates with the transformed phenotype and the type of viral latency in EBV genome positive lymphoid cell lines.

In this paper we have investigated the role of Egr-1 in B cell growth regulation by examining the gene expression in a panel of B cell lines, including both EBV genome negative and EBV carrying cell lines. Egr-1 expression correlates with the cellular phenotype and the specific pattern of viral latency established within the individual cell lines. Thus, constitutive activation of Egr-1 gene is invariably associated with unrestricted expression of viral latent genes in all group III EBV genome carrying cell lines. In contrast, Egr-1 expression is abrogated in group I Burkitt tumor cells, irrespective of the EBV genome carrying status. Activated viral gene expression associated with phenotypic conversion of group I cell lines in to group II or III restores the Egr-1 gene expression. Several forms of EGR-1 protein are found within the different groups of cell lines, and the binding activity to DNA consensus sequences was investigated. Finally, time course analysis of Egr-1 expression during the early steps of EBV infection in vitro demonstrated that Egr-1 is upregulated within minutes from the initial interaction with the B lymphocyte.

B-Lymphocytes↗
PubMed1996

Infection of human T lymphoid cells by human herpesvirus 6 is blocked by two unrelated protein tyrosine kinase inhibitors, biochanin A and herbimycin.

Human herpesvirus 6 is a T lymphotropic herpesvirus that causes exanthem subitum in infants and is considered a potential cofactor in AIDS etiopathogenesis and progression owing to its in vivo and in vitro interactions with human immunodeficiency virus. We report that no differences in phosphorylation on tyrosine residues of cellular proteins were detectable at early times following HHV-6 infection in comparison to uninfected cells. On the contrary, several cellular proteins appeared phosphorylated on tyrosine at 24-48 hr postinfection. In addition, when tyrosine phosphorylation induced by HHV-6 infection was inhibited by the tyrosine kinase inhibitor biochanin A, the infection of HSB-2 cells was also coordinately reduced, as judged by inhibition of cytopathic effect and by inhibition of early and late viral antigen expression. Similar results were obtained with a second unrelated tyrosine kinase inhibitor, herbimycin. The inhibitors seem to act at a late stage of the viral infectious cycle, since neither viral binding nor internalization were affected. Thus, our results indicate that HHV-6 infection leads to the phosphorylation of protein tyrosine kinases, which may play a role in the course of viral infection, probably by participating in the cytopathic effect induced by the virus.

Benzoquinones↗
PubMed1996

EAES ductal stone study. Preliminary findings of multi-center prospective randomized trial comparing two-stage vs single-stage management.

BACKGROUND: The current management of patients with ductal calculi and gallstone disease consists of endoscopic stone extraction (ESE) followed by laparoscopic cholecystectomy (LC). The advent of techniques of laparoscopic ductal stone clearance has introduced an alternative single stage laparoscopic treatment for these patients. The EAES ductal stone trial was set up to compare the relative efficacy and outcome of these two management options. METHODS: The study consists of a prospective randomized controlled clinical trial comparing two management options of patients undergoing LC and suspected of harbouring common duct stones. Patients registered into the trial are randomized to one of two arms: (i) Group A-preoperative ERC with ESE followed by LC during the same hospital admission. (ii) Group B-single stage laparoscopic management consisting of LC and laparoscopic stone extraction either by the trans-cystic duct route or by direct supraduodenal common duct exploration. RESULTS: This preliminary analysis was carried out on 207 randomized patients with comparisons being made on the intention to treat principle. The two groups (A = 106, B = 101) were comparable with respect to clinical features. ASA grade, serum biochemistry and ultrasound findings. CONCLUSIONS: These preliminary findings indicate equivalent success rates and patient morbidity between the two management options but a shorter hospital stay (cost benefit) with the single stage laparoscopic treatment. Trans-cystic duct extraction is a more benign procedure than laparoscopic supraduodenal CBD exploration and is accompanied by a significantly shorter hospital stay. The higher incidence of conversion in the single stage laparoscopic group compared to the two-stage arm is due to the preference for open common duct exploration when the laparoscopic attempt failed by the majority of participating surgeons. The results to-date suggest that in fit patients, single stage laparoscopic treatment is the better option and the role of ESE should change to selective use in those patients in whom laparoscopic ductal stone extraction has failed.

Adolescent↗
PubMed1996

Paraphenylene diamine, a contact allergen, induces oxidative stress in normal human keratinocytes in culture.

During the course of evaluating the interaction between allergens and keratinocytes in the pre-immunological phase of contact sensitization, we have studied the effects of paraphenylene diamine (pPD) on membrane lipid peroxidation and on intracellular antioxidant levels in cultured human keratinocytes. pPD is an aromatic amine which undergoes spontaneous oxidation in culture medium, generating short-lived free radical species including oxyradicals. Following exposure to non-toxic concentrations of pPD (0.5-10 micrograms/ml), we have evaluated the fatty acid pattern of membrane phospholipids as a target of peroxidative damage, and the intracellular level of reduced glutathione (GSH), the activity of superoxide dismutase (SOD), and that of catalase (CAT) as parameters of the antioxidant system. Depending on pPD concentration and the period of exposure, peroxidative damage with a significant decrease in membrane polyunsaturated fatty acids, was detected. Concentrations between 0.5 and 2 micrograms/ml produced an initial increase and then a decrease in both SOD and CAT activities, and in the oxidation of GSH, up to 12 h. After 24 h, when all the pPD had decomposed, recovery of the initial levels of the antioxidants was detected. Concentrations over 5 micrograms/ml induced a progressive decrease in both the enzymatic activities and the GSH concentrations. These results are consistent with the view that oxidative stress can be an essential event in the pre-immunological phase of contact sensitization.

Allergens↗
PubMed1995

Cross-linking of at least three binding sites mediates signal transduction in a CR2-positive Burkitt lymphoma derived cell line (Raji).

In this study we demonstrate that Raji cells, a CR2-positive Burkitt lymphoma-derived cell line, during cell growth, need the cross-linking of multiple OKB7 binding sites or C3d determinants to mediate signal transduction. The loss of one of these affects the cellular response. Moreover, OKB7, the anti-CR2 MoAb, recognizes C3d determinants on the cell surface and inhibits signal transduction induced by anti-C3d polyclonal antibody. Since Raji cells are always CR2 positive during cell growth, we suppose that at least another protein, along with CR2, may be involved in setting up a cell surface complex able to receive and transduce the signal triggered by OKB7. In our experimental system the protein that offers a third binding site to OKB7, may be represented by a 33 kDa protein bearing C3d determinants.

Binding Sites↗
PubMed1995

Epstein-Barr virus DNA is present both in CD10/CD77 positive and negative subsets of human tonsillar lymphocytes.

We report the fractionation of freshly isolated subsets of tonsillar lymphocytes according to cell density and double sorting for the differential expression of CD10 and CD77, and their analysis for the presence of Epstein Barr virus genome by nested PCR. All the subsets of tonsillar lymphocytes gave unequivocal evidence for the presence of EBV DNA, when obtained from EBV seropositive individuals. Confirmation of all cases examined demonstrates that B lymphocytes from the germinal centers of tonsils are also involved in carrying the EBV infection in vivo.

Adolescent↗
PubMed1995

Human herpesvirus 6 variant A, but not variant B, infects EBV-positive B lymphoid cells, activating the latent EBV genome through a BZLF-1-dependent mechanism.

Human herpesvirus 6, a predominantly T lymphotropic virus, has been recently shown to infect some EBV-positive B cell lines, and to induce in them the activation of the EBV lytic cycle. Here we have confirmed and extended such observations, showing that (1) this phenomenon is restricted to the variant A of HHV-6: in fact two isolates belonging to the HHV-6 variant B (BA92 and Z29) were neither able to infect any B cell line, independently of the EBV status, nor to induce the EBV genome expression. The only exception is represented by the P3HR1 cells, in which, however, the infection by the variant B does not determine induction of EBV antigens; (2) the presence of the EBV genome contributes to the susceptibility of the B cell lines to HHV-6 infection, increasing the binding sites and the percentage of infectable cells, as detected by immunoelectron microscopy; and (3) HHV-6 infected T cells, transfected with plasmids bearing the promoter regions of the EBV early genes BZLF1 and BMRF1, show a strong transactivation of these promoters.

B-Lymphocytes↗
PubMed1994

HHV-6 infection in Italy: characterization of an endemic isolate and seroepidemiologic analysis.

A biologic, immunologic and molecular characterization of an HHV-6 isolate (BA92) rescued by the peripheral blood mononuclear cells of a child affected by Exanthem subitum is reported. The comparison with the known HHV-6 prototype strains showed that BA92 is indistinguishable from the Z29 isolate, and can be included in the variant B group of HHV-6. A seroepidemiologic analysis of the antibody response to BA92 of normal individuals as well as patients affected by diseases potentially associated to HHV-6 infection has shown an overall seroprevalence of 81%, and that no variations in seroprevalence or in antibody geometric mean titer are observed assaying the sera also against G.S., U1102, or Z29 infected cells, respectively. These findings indicate: (1) HHV-6 infection is widely diffuse in Italy; (2) it is not possible to discriminate between the viral variants by the currently available IF assays, and (3) no conclusions can be drawn on the potential association of HHV-6 with any of the diseases examined.

Journal Article↗
PubMed1994

Characterization of cultured nail matrix cells.

BACKGROUND: Cultures of epidermal cells are commonly used to study skin biology and differentiation. Recently a method to culture nail matrix cells has been established. OBJECTIVE: We report the biologic characteristics of nail matrix cells in vitro compared with those of epidermal keratinocytes. METHODS: Human nail matrix cells were isolated and cultured in defined medium. Electron-microscopic examination, growth rate, integrin expression and keratin synthesis pattern were evaluated. In addition, the cells were cultured in serum-containing medium. RESULTS: Nail matrix cells appear to be larger than human epidermal keratinocytes and, at the ultrastructural level, they contain a higher euchromatin/heterochromatin ratio and a lower nucleus/cytoplasm ratio and have a higher growth rate. The synthesis of "hard" keratins was detected at all calcium concentrations. Immunofluorescence analyses showed the expression of alpha 2, alpha 3, and alpha 6 integrin subunits. When cultured in serum-containing medium, nail matrix cells produced an outgrowth of epithelium and a spontaneous migration phenomenon associated with a tendency to stratify in a semilunar area that resembles the architecture of the nail matrix. The pluristratified epithelium showed characteristic markers of nail differentiation. CONCLUSION: Culture of nail matrix cells may represent a useful model to study the biologic properties of nail structure, alterations in some nail diseases and the effects of drugs.

Cell Division↗
PubMed1994

Human herpesvirus 6 envelope glycoproteins B and H-L complex are undetectable on the plasma membrane of infected lymphocytes.

Membrane immunofluorescence analysis of cells infected with either variant (A or B) of human herpesvirus 6 revealed a typical punctate staining, after labeling with several HHV-6-positive human sera or with two monoclonal antibodies directed to gB and gH. Immunoprecipitation studies showed a sharp difference in glycoprotein content in whole-cell extracts versus on the cell surface, suggesting the occurrence of gB in the extracellular virions juxtaposed to plasma membranes. By immunoelectron microscopy, the extracellular virions still attached to the cell surface appeared consistently and specifically labeled, whereas the plasma membrane was always unlabeled, independent of viral variant, antibody, or target cell used. These findings may reflect an atypical maturation pathway of HHV-6, and could have important implications in the control of cellular immune response to HHV-6-infected lymphocytes.

Cell Line↗
PubMed1994

Events related to Epstein-Barr virus binding and superinfection of Raji cells.

Raji cells, a CR2-positive Burkitt lymphoma cell line, incubated in normal human serum, activate C3 and fix C3-derived fragments. The presence of these molecules on the cell surface does not affect subsequent Epstein-Barr virus (EBV) binding but it prevents superinfection. On the other hand, EBV superinfection is enhanced if Raji cells were incubated with heat-inactivated serum whose C3 fragments may bind only through receptor-binding sites. These results indicate that the region on cell surface offering the covalent site to C3 fragments would be essential for EBV superinfection. Incubation of Raji cells for 1 min with EBV results in the phosphorylation of CR2 and of a high-molecular-weight protein followed by their dephosphorylation, completed already after 20 min. This finding ascribes to EBV a prompt action through its receptor, different from that of other compounds causing a prolonged CR2 phosphorylation. Our data suggest that at least two binding sites are required for EBV superinfection of Raji cells or that specific patterns of CR2 phosphorylation may modulate Raji superinfection by EBV.

Binding Sites↗