Biomedical subjects
Publications and source records attributed to J Dumont.
The YDp plasmids (Yeast Disruption plasmids) are pUC9 vectors bearing a set of yeast gene disruption cassettes, all uniform in structure and differing only in the selectable marker used (HIS3, LEU2, LYS2, TRP1 or URA3). The markers, surrounded by translational termination codons, are embedded in the slightly modified sequence of the pUC9 multiple cloning sites.
We investigated the correlations between the biological effects of interferon-alpha (IFN-alpha) and clinical responsiveness in low-grade non-Hodgkin's lymphomas (NHL). In this disease, 40-50% of cases respond to IFN-alpha therapy. Patients with nodular NHL were selected for a phase II trial in which they were treated daily with 9 x 10(6) U of IFN-alpha 2a. Binding experiments with [125I]IFN-alpha 2a showed the presence of IFN-alpha receptors on tumor B-cells isolated from lymph nodes before therapy in 9 out of 10 patients. Receptor levels were not related to the subsequent clinical responses. However, no specific binding was detected in one patient who turned out to be unresponsive to IFN-alpha treatment. Single injections of IFN-alpha 2a before beginning the therapeutic protocol resulted in down-regulation of IFN-alpha receptors without change in their affinity in peripheral blood leukocytes from only patients who subsequently responded to therapy (4/10). In 4/5 non-responders and one patient displaying a minor response, receptor numbers did not decrease but Kd values rose markedly in all six cases. These results indicate that lack of in vivo IFN-alpha receptor down-regulation and reduced receptor affinity, as detected before therapy, may be correlated with failure of IFN-alpha therapy in nodular NHL.
A hybrid gene comprising the bovine thyroglobulin gene promoter and the coding region for the simian virus-40 large T- and small t-antigens was used to generate 30 transgenic mice by microinjection into the pronuclei of single cell embryos. All animals except three developed, as single primitive pathology, a dramatic enlargement of the thyroid gland. Compression of trachea and esophagus, accompanied by dyspnea, inspiratory stridor, and dysphagia, led to a progressive cachexia and premature death attributed to respiratory failure. Despite the large thyroid volume, T4 levels were abnormally low, and the progression of the syndrome could be delayed by a substitutive treatment with thyroid hormones. The rapid evolution of the disease, leading to the death of most founder transgenic animals before the breeding age, prevented transmission of the transgene to their offspring. Only two transgenic lines are presently surviving. Immunohistochemical analysis of the tissues revealed a specific expression of the simian virus-40 antigens in the thyroid cells. Hyperplasia was already obvious at birth. Older animals displayed moderately to poorly differentiated thyroid adenocarcinomas. Electron microscopy revealed, however, the persistence of cell polarity and the presence of microfollicles between the densely packed cells. Cell lines derived from these large T-expressing thyroids were shown to have lost expression of both thyroglobulin and thyroperoxidase, while expressing low levels of TSH receptors. These transgenic mice could constitute an interesting model of aggressive adenocarcinoma, sharing phenotypical similarities with the anaplastic type of human thyroid tumors.
An analysis of R-banded PHA-stimulated lymphocytes from 13 patients with secondary acute non-lymphocytic leukemia (S-ANLL) following breast cancer or lymphoma, and treatment by alkylating agents and/or radiotherapy, is reported. We found that chromosomes 5, 7, 11 and 17 are over-involved in structural rearrangements. These anomalies are similar to those observed in the same categories of patients without S-ANLL, and after in vitro treatment of normal lymphocytes by the alkylating agent melphalan. These anomalies are thus likely to be induced by treatment, independently of S-ANLL. However, the same chromosomes (5, 7, 11 and 17) are recurrently deficient in leukemic S-ANLL clones. In spite of these similarities, it remains unlikely that the deficiencies observed in leukemic clones were directly induced at the time of treatment. Probably, treatment of primary cancers induces nonrandom mutations of recessive genes located on these chromosomes as also indicated by chromosomal lesions. Various rearrangements including deletions of the homologous normal counterparts may then occur, unmasking mutated recessive genes. The latter stage would be concomitant with the leukemogenic process.
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Selenium status was determined in an endemic-goiter area and in a control area of Zaire. Compared with the reference values of a noniodine-deficient area, serum selenium in subjects living in the core of the northern Zaire endemic-goiter belt (Karawa villages) was seven times lower in 52 school-children and similarly low in 23 cretins; erythrocyte glutathione peroxidase (RBC-GPX) was five times lower in schoolchildren and still two times lower in cretins (P = 0.004). In a less severely iodine-deficient city of the same endemia (Businga), selenium status was moderately altered. RBC-GPX activity was linearly associated with serum selenium concentration up to a value of 1140 nmol/L and leveled off at approximately 15 U/g Hb at greater selenium concentration. At Karawa villages, selenium supplementation normalized both the serum selenium and the RBC-GPX. This combined iodine and selenium deficiency could be associated with the elevated frequency of endemic myxedematous cretinism in Central Africa.
RDC8 (correction of RCD8), a recently cloned new putative member of the G protein-coupled receptor family protein, is exclusively present in the medium-sized neurons of the striatum. This restricted localisation mimics the major striatal dopamine D1 receptor localisation and is of major importance for the understanding of basal ganglia physiology and degenerative diseases pathogeny such as Huntington's and Parkinson's disease. RDC7, another putative G protein-coupled receptor chemically closely related to RDC8 (correction of RCD8), is mainly distributed in pyramidal neurons of the cerebral cortex, the hippocampus and the claustrum, and in the amygdala, and may represent the minor extra-striatal variant of the D1 receptor.
A cytogenetic study of 14 patients with secondary acute nonlymphocytic leukemia (S-ANLL) with prior treatment for breast cancer is reported. The chromosomes recurrently involved in numerical or structural anomalies are chromosomes 7, 5, 17, and 11, in decreasing order of frequency. The distribution of the anomalies detected in this sample of patients is similar to that observed in published cases with prior breast or other solid tumors, though anomalies of chromosome 11 were not pointed out, but it significantly differs from that of the S-ANLL with prior hematologic malignancies. This difference is principally due to a higher involvement of chromosome 7 in patients with prior hematologic malignancies and of chromosomes 11 and 17 in patients with prior solid tumors. A genetic determinism involving abnormal recessive alleles located on chromosomes 5, 7, 11, and 17 uncovered by deletions of the normal homologs may be a cause of S-ANLL. The difference between patients with prior hematologic malignancies or solid tumors may be explained by different constitutional mutations of recessive genes in the two groups of patients.
Bovine leukemia virus (BLV) p24 gene was expressed in Saccharomyces cerevisiae under the control of the PHO5 (encoding repressible acid phosphatase, rAPase) promoter. Yeast cells were transformed by a yeast-E. coli shuttle vector carrying the PHO5 promoter, the p24 gene and the CYC1 transcription terminator. After low inorganic phosphate (Pi) induction of the PHO5 promoter, p24 accumulated in the producing cells up to a concentration representing 10% of total soluble proteins. The expression level of p24 gene was not increased by insertion of the positive regulatory gene PHO4 on the p24 expression vector. The p24 produced in this system and incubated in crude yeast extract showed a remarkably high resistance to proteolytic degradation, a feature that presumably correlates with the compact globular conformation of the protein combined to the stabilizing effect of the N-terminal residue.
The nucleotide sequence coding for bovine leukemia virus (BLV) envelope glycoprotein gp51 was inserted into a yeast-Escherichia coli shuttle vector carrying the promoter and secretion signal sequence of PHO5 (the yeast gene coding for repressible acid phosphatase) and the CYC1 transcriptional terminator. Yeast cells transformed by this construction synthesized gp51 after PHO5 induction by inorganic phosphate deprivation. The yeast-expressed gp51 was partially glycosylated into heterodisperse protein molecules ranging from 40 to 48 kDa. No gp51 was excreted in the culture medium. The amount of protein accumulated in yeast cells was estimated to reach 0.06% of soluble proteins. This modest level of expression seemed to be due to the toxicity of gp51 to the yeast cell. The yeast-expressed gp51 products were used in enzyme-linked immunosorbent assays for the detection of antibodies in sera from BLV-infected animals; they were also screened for the presence of well-defined biological epitopes. In both studies poor reactivity was observed. Rabbits immunized with the recombinant gp51 showed high antibody titers to native BLV gp51. However, these antibodies did not neutralize BLV in vitro.
The antibacterial activity of five commercially available amalgams (Cupromuc, Dispersalloy, Fluoralloy, Predalloy, and Synalloy) against two bacterial species frequently isolated from dental plaque and caries (Actinomyces viscosus and Streptococcus mutans) was studied by use of a modification of the method of Orstavik (1985). All amalgams displayed some antibacterial properties. This antibacterial activity was amalgam- and bacteria-dependent. Cupromuc and Fluoralloy were the most active, and the non-gamma 2 amalgams showed intermediate activity. Combined with mechanical and biological properties, these differences should be taken into account when one is choosing an amalgam for clinical use.
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A simple, rapid SDS-Page method for protein profiles analysis of Actinomyces species was developed. Eighteen strains (12 reference strains and 6 fresh isolates) representing four species were used in this study. Eight detergents were tested for protein extraction. Cell extracts were obtained by shaking the bacteria suspended in a solution containing a detergent and glass beads. After electrophoresis, according to the Laemmli's technique, two protein stain procedure were tested (Coomassie blue and silver stain). Best results were obtained with 0.5% Triton X-100 for 4 h and the silver stain procedure of Oakley. Protein profiles analysis showed that the method is reproducible and distinguishes not only species but sometimes also subspecies.
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A finding of large amounts of extracellular amorphous material is rare in malignant lymphomas. We report four such cases of follicular lymphoma that were investigated by electron microscopy and immunohistochemistry using monoclonal antibodies. This extracellular material was clearly different from the sclero-hyalinosis commonly found in malignant lymphomas. Our observations revealed that the material is ultrastructurally composed of membranous structures, membrane-bound vesicles, and electron-dense bodies. Immunohistochemistry confirmed its neoplastic cell origin and its identity with cell membrane antigens (Dako-LC+, anti-B antibodies positive). In one case investigated using frozen sections, it showed the same immunophenotype as the neoplastic cells: SB4+ (CD19), SB2+ (CD21), TO15+ (CD22), SB3+ (CD37), IgML+, and Calla+. The mechanism for accumulation of this extracellular amorphous material is probably a function of a deregulation of cell wall synthesis and an exocytosis of cell membranes.
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Seventeen patients with Hodgkin's disease (HD) were treated with high-dose chemotherapy followed by autologous bone marrow transplantation (ABMT). Eleven patients were resistant to initial therapy. Three patients had relapsed and were still responders to second or third line therapy. Three patients had relapsed but were progressing under second or third line therapy. Pre-ABMT chemotherapy included high dose cyclophosphamide in all patients (50 mg Kg-1 day-1 bolus for 4 days), most often associated with BCNU or CCNU, aracytine and 6 thioguanine. Four patients received additional TBI (10 Gy). In 9 patients complete remission (CR) was achieved, 4 failed to respond and 4 cases were not evaluable due to early death. Among CR patients, 2 died from late toxicity, 4 relapsed between the 2nd and 5th months, but 3 patients remain in CR, off therapy at 25+, 43+, and 66+ months, including 1/11 initially resistant and 2/6 who had relapsed. There were 9 treatment related deaths: 6 due to infection, 1 cardiac failure and 2 multiorgan failure. The high complete response rate in these heavily pretreated patients suggests that there may be an indication for high dose therapy earlier in resistant HD. Moreover under such conditions, treatment related morbidity would be expected to be lower.