High-field plasma acceleration in a high-ionization-potential gas.

Plasma accelerators driven by particle beams are a very promising future accelerator technology as they can sustain high accelerating fields over long distances with high energy efficiency. They rely on the excitation of a plasma wave in the wake of a drive beam. To generate the plasma, a neutral gas can be field-ionized by the head of the drive beam, in which case the distance of acceleration and energy gain can be strongly limited by head erosion. Here we overcome this limit and demonstrate that electrons in the tail of a drive beam can be accelerated by up to 27 GeV in a high-ionization-potential gas (argon), boosting their initial 20.35 GeV energy by 130%. Particle-in-cell simulations show that the argon plasma is sustaining very high electric fields, of ∼150 GV m(-1), over ∼20 cm. The results open new possibilities for the design of particle beam drivers and plasma sources.

JC human polyomavirus is associated to chromosomal instability in peripheral blood lymphocytes of Hodgkin's lymphoma patients and poor clinical outcome.

BACKGROUND: B cells are potential sites for latency and reactivation of the human neurotropic JC polyomavirus (JCV). We investigated JCV and Epstein-Barr virus (EBV) status in peripheral blood lymphocytes (PBL) from 74 Hodgkin's lymphoma (HL) and 91 B-cell non-Hodgkin's lymphoma (B-NHL) patients. PATIENTS AND METHODS: JCV and EBV DNA were assessed by PCR, and FISH technique was used to localize viral infection and to estimate chromosomal instability (rogue cells, 'chromosomal aberrations') throughout evolution. The influence of viral infection and chromosomal instability on freedom from progression (FFP) was investigated in HL patients. RESULTS: PCR product sequencing of PBL identified JCV in 42 (57%) circulating lymphocytes of HL patients. FISH analysis revealed that the presence of cells with a high JCV genome copy number--associated to the presence of rogue cells and 'higher frequency of chromosomal aberrations'--increased from 15% before treatment to 52% (P < 10(-5)) after. The co-activation of JCV and EBV was independent of known prognostic parameters and associated with a shorter FFP (JCV and EBV co-activation P < 0.001, rogue cells P < 0.002). CONCLUSION: In HL, JCV activation and chromosomal instability have been identified in PBL and associated with a poorer prognosis, especially in EBV+.

Blastoid mantle cell lymphoma: evidence for nonrandom cytogenetic abnormalities additional to t(11;14) and generation of a mouse model.

Mantle cell lymphoma (MCL) is characterized by the t(11;14)(q13;q32), which is associated with cyclin D1 hyperexpression and a poor prognosis. MCL cases have been shown to progress to a more aggressive disease but the molecular events responsible of this phenomenon have not been determined. We have established two cell lines from the pleural effusions of two patients with MCL that we have used for further cytogenetic characterization to better define the incidence and nature of secondary chromosome abnormalities using multicolor fluorescence in situ hybridization, whole chromosome paint, and specific probes. Both cell lines grew independently without growth factors. Using CCND1/IGH-specific probes, patient UPN1 was found to have a masked t(11;14). Numerous and complex chromosomal abnormalities were found in both cell lines affecting chromosomes 2, 8, 13, 18, 22, X, and Y. These abnormalities included 8p losses, suggesting the presence of an anti-oncogene in this region, rearrangements of 8q24, MYC gene, and translocations involving 8, X, and Y chromosomes, which might be significant in the pathogenesis of MCL progression. The use of the cell lines (UPN1) allowed us to generate a mouse model of human MCL, mimicking a disseminated lymphoma and leading to the death of the animals in 4 weeks. This blastoid MCL model could be of major interest to determine molecular events involved in MCL progression, allowing isolation of involved genes and their functional characterization, and to study the effects of new chemotherapy regimens in mouse models.

In vivo elimination of acentric double minutes containing amplified MYCN from neuroblastoma tumor cells through the formation of micronuclei.

Neuroblastoma, the most common solid extracranial neoplasm in children, shows an appreciable variability in clinical evolution. Amplification of the MYCN oncogene in this tumor is detected in 25 to 30% of cases and is associated with poor clinical outcome. In this study, quantitative polymerase chain reaction and fluorescence in situ hybridization were used to determine MYCN amplification status in 46 neuroblastoma tumors. MYCN amplification was detected in tumors from 11 patients. Fluorescence in situ hybridization revealed the presence of micronuclei containing amplified MYCN sequences in 8 of the 11 tumors. Micronuclei are indicative of spontaneous elimination or loss of amplified sequences by tumor cells. Because the elimination of amplified sequences can be enhanced in vitro by specific drugs such as hydroxyurea, our observations suggest a new therapeutic strategy specifically targeted to cells with amplified genes.

Evidence of LMP1-TRAF3 interactions in glycosphingolipid-rich complexes of lymphoblastoid and nasopharyngeal carcinoma cells.

Latent membrane protein 1 (LMP1) is an Epstein-Barr virus (EBV) protein expressed in EBV-transformed B lymphocytes and in approximately 50% of nasopharyngeal carcinomas (NPCs). LMP1 signaling involves several cellular signaling intermediates, especially TNF receptor-associated factors (TRAFs). We have shown previously that LMP1 is highly concentrated in a cell fraction called glycosphingolipid-rich membrane complexes (GSL complexes). We report here that parallel accumulation of LMP1 and TRAF3, but not TRAF1 or TRADD, was observed in GSL complexes from lymphoblastoid and LMP1-positive NPC cells. In contrast, TRAF3 was not concentrated in GSL complexes from LMP1-negative cells. Binding of LMP1 and TRAF3 in GSL complexes was demonstrated in lymphoblastoid and NPC cells, by co-immunoprecipitation with both anti-LMP1 and anti-TRAF3 antibodies.

Control of apoptosis in Epstein Barr virus-positive nasopharyngeal carcinoma cells: opposite effects of CD95 and CD40 stimulation.

The expression and function of CD95 and CD40 were investigated in malignant cells from EBV-positive undifferentiated nasopharyngeal carcinomas (NPCs). Large amounts of CD95 and CD40 expression were detected in 15 of 16 EBV-positive NPC specimens. In contrast, CD95 was not detected in two biopsies from patients with EBV-negative differentiated NPCs. We tested whether the CD95 apoptotic pathway was functional in NPC cells by treating two EBV-positive NPC tumor lines in vitro with a CD95 agonist. In both cases, NPC cells were extremely susceptible to CD95-mediated apoptosis, despite strong constitutive expression of Bcl-x. Combined CD40 and CD95 stimulation was used to investigate the possible anti-apoptotic activity mediated by CD40. The CD40 receptor was activated by incubating NPC cells with murine L cells producing CD154, the CD40 ligand. This treatment resulted in a strong inhibition of CD95-related cytotoxicity. Such an anti-apoptotic effect of CD40 is well known for B lymphocytes, but has not previously been reported for epithelial cells. These data suggest that NPC tumor-infiltrating lymphocytes, which often produce the CD40 ligand in situ, may increase the survival of malignant cells, thereby enhancing tumor growth in patients.

High concentration of the EBV latent membrane protein 1 in glycosphingolipid-rich complexes from both epithelial and lymphoid cells.

Latent Membrane Protein 1 (LMP1) is an EBV-transforming protein which is detected both in lymphoblastoid cell lines-resulting from EBV-immortalization in vitro- and in undifferentiated nasopharyngeal carcinoma (NPC), an EBV-associated malignancy of epithelial origin. To better define LMP1 subcellular targets, LMP1 distribution was analyzed in cellular glycosphingolipid-rich complexes (GSL-complexes) derived from epithelial and lymphoid cells. These complexes are obtained by extraction of glycosphingolipid-rich membrane domains (GSL-domains), which are clustering sites for heterotrimeric G-proteins and G-protein-associated receptors. LMP1 concentration was enriched 50-fold in GSL-complexes extracted from a NPC tumor line, C15. High concentrations of LMP1 were also observed in GSL-complexes derived from cultured lymphoid and epithelial cells. These data suggest that association with GSL-domains is an important step in LMP1 trafficking and is probably required for some aspects of its biological activity.

Heterogeneity among human nasopharyngeal carcinoma cell lines for inflammatory cytokines mRNA expression levels.

Using polymerase chain reaction (PCR), we confirmed the expression of interleukin-1 alpha (IL-1 alpha) by the human nasopharyngeal carcinoma (NPC) cell line C15 without contribution of either human IL-1 beta or mouse IL-1 alpha in the biological activity previously found in C15. However we showed that IL-1 alpha was not expressed in all NPCs. IL-1 beta and/or tumor necrosis factor (TNF)-alpha genes could also be activated, independently from the number of Epstein Barr Virus (EBV) copies harbored by the cells. Interestingly, the primary tumor C15 showed a profile of TNF-sensitive tumor while C17, C18 and C19 which were derived from metastasis have a typical profile of TNF-resistant cells. Furthermore, the inflammatory cytokines whose genes are classically induced by IL-1 and TNF were found expressed only in C17 and C19 suggesting another level of heterogeneity among NPCs.

Elevated expression of ICAM1 (CD54) and minimal expression of LFA3 (CD58) in Epstein-Barr-virus-positive nasopharyngeal carcinoma cells.

Undifferentiated nasopharyngeal carcinoma (NPC) is a remarkable entity among human tumors because of its constant association with the Epstein-Barr virus (EBV). Malignant epithelial cells harbor the EBV genome and often express at least 2 species of latent EBV protein (EBNA1 and LMP1). Despite the massive presence of tumor-infiltrating lymphocytes, NPC cells obviously escape immune surveillance directed to EBV antigens. Previous investigations carried out on EBV-positive Burkitt lymphoma (BL) cells have shown that this fact may be partially accounted for by a lack of expression of ICAM1 (CD54) and LFA3 (CD58). ICAM1 and LFA3 have therefore been investigated in fresh NPC biopsies and transplanted NPCs. With only 1 exception out of 9 cases, NPC cells appear to express high levels of ICAM1 and low levels of LFA3. This is a complete inversion of the pattern observed in normal epithelial cells in vivo. Additional investigations will be required to determine to what extent these characteristics affect T-cell interactions with NPC cells, specially in the process of EBV-antigen recognition.

Detection of anti-Epstein-Barr-virus transactivator (ZEBRA) antibodies in sera from patients with nasopharyngeal carcinoma.

The Epstein-Barr virus (EBV) is a ubiquitous Herpes virus which causes infectious mononucleosis and is associated with such different neoplasms as Burkitt's lymphoma and nasopharyngeal carcinoma. EBV latently infects its target cells; nevertheless, evidence of viral replication in NPC tumours has been uncovered. Among the EBV transactivators, the ZEBRA protein plays a crucial role in switching the virus from a latent to a productive mode. ZEBRA protein was produced using a eukaryotic expression vector: the open reading frame containing the BZFLI cDNA has previously been inserted down-stream from the adenovirus major late promoter leading to expression of a 38-kDa nuclear protein. We performed serological studies by employing ZEBRA protein expressed in human cells for immunofluorescence and Western-blot assays. We were able to detect IgG anti-ZEBRA antibodies (IgG/ZEBRA) in 87% of NPC patients. These antibodies were absent in control sera; IgG/ZEBRA antibodies can be proposed as a useful marker for diagnosis of NPC tumors.

Increased cytotoxicity of nanoparticle-carried Adriamycin in vitro and potentiation by verapamil and amiodarone.

The cytotoxicities of free cyanoacrylic nanoparticles, free Adriamycin, Adriamycin-loaded nanoparticles and a mixture of Adriamycin and nanoparticles are compared in cancer cell cultures. Increased cytotoxicity was observed in the sensitive (DC3F) subline when Adriamycin was in particulate form rather than free. In the derived pleiotropic resistant subline (DC3F AD/AZA), sensitivity to Adriamycin was completely restored with the conjugate. Addition of verapamil or amiodarone allowed an enhancement of efficiency of tenfold for free Adriamycin and between two- and fourfold for its conjugate form. Vectorization by nanoparticles and pharmacological modulation of cell membrane can act in synergy in synergy to overcome the resistance to Adriamycin in vitro.

Establishment and characterization of three transplantable EBV-containing nasopharyngeal carcinomas.

Three transplantable nasopharyngeal carcinoma (NPC) tumors, designated C15, C17 and C18, have been obtained and characterized. C15, derived from a primary NPC tumor, has been propagated in nude mice for 30 passages. C17 and C18, derived from metastatic NPC tissue, have been passaged 10 times. Desmosomes, present in every case, provided confirmation of the epithelial origin of all 3 tumors. The Epstein-Barr virus (EBV) genome is contained in C15, C18 and C17 tumor cells with 30, 12 and 3 copies, respectively. The Epstein-Barr virus nuclear antigen (EBNA) was stained by the classical anti-complement immunofluorescence (ACIF) technique. Fluorescence intensity was strong in C15, moderate in C18, and hardly detectable in C17 cells. No expression of the EA and VCA antigens was detected. Flow cytometry analysis performed on monocellular suspensions showed the absence of detectable CR2 molecules (the EBV receptor on B lymphocytes) in all 3 tumors, and the constitutive expression of HLA class-II antigens in C15 and C17 cells. IL-1 activity was demonstrated in the supernatant of C15 and C17 cells cultivated in vitro for 3 days. These data confirm that the constitutive synthesis of MHC class-II molecules and the release of IL-1-like activities are frequent features of NPC cells. These characteristics could be of importance in relation with the T-cell infiltrate found in NPC primary tumors.

Constitutive expression of HLA class II antigens on EBV positive malignant cells from nasopharyngeal carcinoma: possible involvement in T cell infiltration.

Undifferentiated nasopharyngeal carcinoma (NPC) was originally referred to as "lymphoepithelial carcinoma." Two main cell types are constantly associated within these tumors: the malignant epithelial cells which harbor the Epstein-Barr virus (EBV) genome, and the nonmalignant lymphocytes mostly of the T lineage. Three characteristic features of malignant NPC cells might explain T cell migration within NPC tumor tissue: HLA class II molecule expression, IL-1 production, and presence of EBV antigens. Homogeneous suspensions of malignant NPC cells were derived from a nude-mouse transplantable tumor in order to specify the status of HLA class II molecules in these tumors. These suspensions were stained for HLA class II antigens and analyzed by flow cytometry in comparison with other carcinoma cell lines. Three characteristics of transplantable malignant NPC cells were demonstrated: constitutive and coordinate expression of DR, DP, and DQ molecules, and concomitant expression with the CDw40 antigen ("B lymphocyte carcinoma cross-reacting antigen").

[Non-Hodgkin's malignant lymphoma: reliability of "typing" using cyto-enzymatic markers. Comparison with immunological markers (author's transl)]. / Lymphomes malins non hodgkiniens: fiabilité du "typage" par les marqueurs cyto-enzymatiques. Comparaison avec les marqueurs immunologiques.

Comparison between membrane markers and enzyme markers was made in 74 cases of non-Hodgkin's malignant lymphomas and a good correlation appears between both methods in order to distinct lymphomas into T and B origin. Enzyme markers are reliable and provide quickly made and easily interpretable documents. As far as T-lymphomas are concerned, three hydrolases namely acid phosphates e, acid esterase and B-glucuronidase give the same good results. As for B-lymphomas, a specific enzyme marker has to be found. Furthermore, typing of malignant lymphomas by enzymatic and/or immunologic methods appears to be quite better than from morphologic features such as convoluted or cleaved nuclei for example.

Subsets of malignant lymphomas in children related to the cell phenotype.

We studied the lymphomatous cells of 39 children presenting with the classical features of malignant lymphoma. Twenty-two had T lymphoblasts. We could classify these patients into three subsets: The T lymphoblasts from children group 1 displayed antigen(s) shared by a thymocyte subpopulation, had terminal deoxynucleotidyl transferase (TDT), but no affinity for peanut agglutinin (PNA). The T lymphoblasts from children group 2 lacked the thymocyte antigen(s), had no TDT, but showed affinity for PNA. The T lymphoblasts from children group 3 displayed mature T-cell antigens, had no TDT, and no affinity for PNA. Children from the three groups were similar in terms of clinical presentation, age and sex distribution, and cell morphology; however patients from the three groups might have a different prognosis. Fourteen children had B lymphoblasts that, in half of the cases, had affinity for Helix pomatia agglutinin. Three patients had lymphoblasts lacking specific marker. Two of them had cells displaying an antigen found on common acute lymphoblastic leukemia cells and had TDT.