Analysis by in situ hybridization of cytokine mRNAS expression in thymic nurse cells.
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Animals↗
Biomedical subjects
J Boniver
Publications and source records attributed to J Boniver.
TNF-alpha is involved in the mechanism of murine thymic lymphoma prevention by bone marrow grafting.
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Animals↗
Analysis by in situ hybridization of cytokine mRNA expression in the murine developing thymus.
We have used in situ hybridization to investigate the expression of IL-1, IL-2, IL-4, IL-6 and IFN-gamma genes by thymic cells during fetal development in mice. Two waves of mRNAs were detected in thymic cells for IL-1 at days 16 and 19 of gestation, for IL-2 at days 14 and 18, and for IL-4 at days 14 and 16. Three peaks for IL-6 were observed at days 13, 17 and around birth. Finally, only one peak of cells positive for IFN-gamma was detected. Whereas cells positive for IL-1 were generally grouped and more often localized in the external area of the thymus, the other positive cells were isolated and evenly distributed in the thymus. Our results illustrated the presence of cytokine transcripts in the developing thymus following a developmentally controlled sequence and support the hypothesis that cytokines could play a role in T cell development.
Animals↗
Population dynamics of CD4+ T cells lacking Thy-1 in murine retrovirus-induced immunodeficiency syndrome (MAIDS).
Increased numbers of CD4+ Thy-1- cells have been described in the spleen (SP) of mice with retrovirus-induced immunodeficiency (MAIDS). Since this phenotypic abnormality might have considerable functional importance, the expansion of the CD4+ Thy-1- subset in MAIDS was characterized further. CD4+ Thy-1- and Thy-1+ T-cells from infected mice expressed similar densities of CD3 and TCR alpha/beta. In contrast, the Thy-1- subset was uniformly CD44hi, even early in the disease when part of Thy-1+ cells were still CD44lo. The emergence of CD4+ Thy-1- cells occurred first in SP and lymph nodes and was observed later in thymus. The important fraction of CD4+ cells lacking Thy-1 normally present in Peyer's patches was only weakly modified. Despite the major expansion of the CD4+ Thy-1- phenotype, the proliferating fraction was not higher in this subset than in CD4+ Thy-1+ cells from infected mice. Persistence after hydroxyurea administration was identical in both subsets, indicating similar mean cell lifespans. Taken together, these results show that the major expansion of CD4+ Thy-1- T-cells in MAIDS is not ascribable solely to increased proliferation within this subset. Phenotypic analysis suggests that CD4+ Thy-1- cells result from the differentiation of Thy-1+ cells induced by activation signals related to retroviral infection.
Animals↗
A RadLV-induced gamma delta T cell lymphoma displaying an antitumoral cytotoxicity.
We described previously the induction by RadLV infection of a lymphoma (NS8) expressing a cytolytic activity against an MCA-induced fibrosarcoma. We report here that the cytolytic activity of these immortalized CD3+, CD8+ T cells is non-MHC-restricted. We then determined the structure and expression of the TCR chains expressed by these cells. Only partial rearrangement of the beta chain associated to an abnormally short transcript was detected in NS8 cells, whereas the gamma chain is rearranged and normally transcribed. On the opposite, rearrangement and expression of these genes were found in the other RadLV-induced lymphomas analysed. Moreover, gamma delta TCR proteins were detected on the cell surface of NS8 cells only, whereas the alpha beta complex, presents on the other T cell lines, was not expressed by NS8 cells. The ability of NS8 cells or of cells obtained from activated lymph nodes (harvested from mice grafted with the T2 sarcoma used to induce the NS8 line) to lyse the T2 sarcoma cell line was analysed. With both types of lymphocytes, the cytotoxicity was partially inhibited by a preincubation of the effector cells with anti-gamma delta antibodies. These results demonstrate that gamma delta lymphocytes can mediate anti-tumour cytotoxicity and NS8 lymphoma line may be representative of the TCR gamma delta CD8+ T cell subpopulation expressing non MHC-restricted cytotoxicity and displaying antitumoral activity.
Animals↗
Thymus involvement in murine acquired immunodeficiency (MAIDS).
Due to self-renewal of the peripheral pool of T-cells, adult thymectomy has normally little influence on immunocompetence. However, thymus might play a more important role in the setting of viral-induced cytopathic effects on T-cells in the periphery. Therefore, thymus weight, cell numbers, and subset distribution were sequentially analysed after infection with RadLV-Rs, a viral mixture known to induce murine retrovirus induced immunodeficiency (MAIDS). Infection induced thymic atrophy (concerning organ weight as well as total cell number) which culminated seven weeks after inoculation. The atrophic process mostly reflected the depletion of double positive CD4+ CD8+ cells since their proportion sharply decreased around week 6. Single positive T-cells were less affected by the process. The proportion of B-cells progressively increased. Surprisingly, there was a strong correlation between the extent of atrophy and the frequency of B-cells in the thymus. Finally, an abnormal CD4+ T-cell subset lacking Thy-1 and previously described in the periphery also appeared in the thymus and its frequency was strongly correlated with the expansion of B-cells in this organ.
Animals↗
Lymphocyte lifespan in murine retrovirus-induced immunodeficiency.
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Animals↗
[Lethal varicella. Immunohistochemistry and in situ hybridization].
We report a case of lethal varicella developed in an immunocompromised man in his seventies. A study by immunohistochemistry and in situ hybridization was conducted revealing the presence of the glycoprotein gpI specific for VZV and its corresponding nucleic acids found in multiple foci of most tissues and organs, except in the brain and paravertebral sympathetic ganglia.
Aged↗
[Rupture of a superficial femoral aneurysm in the context of neurofibromatosis. Report of a case].
The authors observed the spontaneous rupture of a dysplastic superficial femoral artery in a 47 years old patient with Von Recklinghausen disease. The patient presents a gigantism of his right lower limb. On angiography, the femoropopliteal axis is polyaneurysmal and there is evidence of rupture at mid-thigh. The diseased artery is excluded by interposition of a dacron prosthesis, with reimplantation of the deep femoral artery. Histology shows fibrohyalin thickening of the intima. Dysplasia of large limb arteries is excessively rare in Von Recklinghausen disease.
Aneurysm, Ruptured↗
Detection of human papillomaviruses in paraffin-embedded biopsies of cervical intraepithelial lesions: analysis by immunohistochemistry, in situ hybridization, and the polymerase chain reaction.
One hundred and forty biopsies with an initial diagnosis of cervical intraepithelial lesion (CIL) were tested for the presence of human papillomavirus (HPV) by immunohistochemistry and in situ hybridization using commercial biotinylated probes (Vira-Type in situ assay; Digene Diagnostics, Silver Spring, MD) or probes labeled with digoxigenin by the random primer technique. Immunohistochemistry was more inferior to the in situ hybridization method, with a detection rate of 14% (20/140) compared to 61% (86/140) for the in situ assay with the digoxigenin-labeled probes. Biotinylated probes proved to be slightly less sensitive than digoxigenin-labeled probes, with a detection rate of 53% (74/140). Although less sensitive in our series taken as a whole, immunohistochemistry was positive in a few cases of CILs negative by in situ hybridization, so that the association of these techniques gave the highest detection rate (66%; 92/140). The CILs that remained negative with these methods (34%; 48/140) were investigated by the polymerase chain reaction (PCR) using consensus primers to determine definitively the presence of HPV in these lesions and were reviewed histologically to assess the diagnosis of CILs. The PCR method increased the detection rate of HPV in our series to 76% (107/140). The diagnosis of CILs was confirmed for all the biopsy specimens positive by PCR (15/15; 100%) and for all the HPV negative tissues with histological features of a high-grade lesions (7/7; 100%).(ABSTRACT TRUNCATED AT 250 WORDS)
Female↗
Flow cytometry in prostate cancer: histological and clinical correlation.
DNA histograms were obtained by flow cytometry for 39 human prostate carcinomas (27 total prostatectomy specimens, 5 biopsies and 7 transuretral resections). The study was performed on formalin-fixed and paraffin-embedded material. In this report, ploidy index did not seem to be a good marker of prognosis as no significant variation in ploidy was found neither among the different stages nor among the different Gleason categories. Proliferative index of the tumors seemed to be a more sensitive parameter; a significant relation was observed between proliferative index and stage of the tumor. The authors discuss these results under the light of previously reported observations.
Aged↗
Misdiagnosed malignancy in transplanted organs.
The case reports of three patients who received cancer-bearing organs at this institution are presented. A fourth recipient, who was to be transplanted with a cancerous kidney, was spared this disastrous complication. The relevant data regarding the donors is also alluded to, with special reference to the type and site of the primary malignancy. Following these case reports, the implications of these issues, their possible prevention, and further management are discussed.
Adult↗
Detection of Epstein-Barr virus in a case of undifferentiated nasopharyngeal carcinoma by in situ hybridization with digoxigenin-labelled PCR-generated probes.
A case of nasopharyngeal carcinoma is presented. Epstein-Barr viral genome was identified in the neoplastic cells by in situ hybridization with digoxigenin-labelled polymerase chain reaction-generated probes. We report the development of this technique in paraffin-embedded sections and propose that such identification may prove valuable for the diagnosis of this tumour in routine material.
Adult↗
Phenotypical and functional analyses of mononuclear cells during rejection of a transplanted murine fibrosarcoma.
Repeated injections of mitomycin C-treated T2 fibrosarcoma cells into tumor-sensitized mice cause regression of a secondary tumor graft and more than 90% of the mice are cured. In the data presented here, an enhancement of the cytolytic cell-mediated activities measured in vitro against the specific T2 targets is shown in lymph nodes draining the tumor and in the spleen during the process of tumor rejection. Histopathologic studies revealed a rapid and marked accumulation of mononuclear cells mostly at the periphery of the rejected tumor tissue. A significant increase of CD8-positive, asialo GM1-positive and acid phosphatase-positive cells was observed in the rejected tumors whereas CD4-positive cells were similarly detected in both progressing and rejected tumor tissue. As macrophages seemed to be the population presenting the most persistent variation after immunization, the production of TNF-alpha was studied within the tumor site and in the lymphoid tissues during the regression process. Firstly, the presence of TNF-alpha within the cytoplasm of most of the adherent cell fractions isolated from the spleen and the tumor of immune mice was demonstrated by immunocytochemistry. Next, TNF-alpha mRNA-containing cells were determined by in situ hybridization of frozen tumor sections and identified essentially as tumor infiltrating macrophages. Finally, the macrophage populations isolated from tumors and from the spleen of immune mice were able to produce in vitro large quantities of TNF-alpha without exogenous stimulation. These findings support the role of TNF-alpha in the effector mechanisms contributing to the tumor regression process.
Animals↗
Efficient immunoselection of cytolytic effectors with a magnetic cell sorter.
This paper describes a rapid and efficient method for the sorting of in vitro activated cytolytic effectors cells. For cytotoxic assays, a large number of cells with conserved function must be rapidly obtained. Immunomagnetic sorting was chosen because it is faster than flow cytometry sorting. The MACS system requires the use of paramagnetic beads of small diameter (100-150 nm), reputed to interfere minimally with cell function. In order to generate the cytolytic effectors, peripheral blood lymphocytes were cultivated in the presence of interleukin-2 (50 U/ml) and anti-CD3 monoclonal antibody (BMA030, 100 ng/ml) for 4 days. Cell separation was based on the membrane expression of the CD3 complex. The purity obtained for positive (CD3+) cell sorting with the MACS was higher than 95%. The purity of negative (CD3-) cell fraction was more variable, but further purification by flow cytometry rapidly yielded purity higher than 95%. Cytotoxic assays were performed against four target cell lines (K562, Daudi, HL60 and U937) and proliferation assays showed that both negatively and positively selected populations had conserved their function acquired during culture in the presence of anti-CD3 mAb and IL2.
Antibodies, Monoclonal↗
Detection of cytomegalovirus, Pneumocystis carinii, and aspergillus species in bronchoalveolar lavage fluid. A comparison of techniques.
Cytomegalovirus (CMV), Pneumocystis carinii, and Aspergillus species are common causes of fatal pulmonary infections in immunocompromised hosts. Therefore, rapid and reliable methods of establishing the diagnosis of these types of pneumonia are essential. Bronchoalveolar lavage (BAL) has proved to be a rapid and safe procedure for procuring large numbers of bronchial and alveolar cells from the lung. To assess various methods of detecting CMV, P. carinii, and aspergillosis in lavage specimens, 47 BAL samples from adults at high risk for these infections were evaluated. The visualization of these agents was performed by cytologic examination and in situ hybridization for CMV; cytologic examination, Gomori's methenamine silver (GMS) stain, and immunofluorescence for P. carinii; and cytologic examination, GMS stain, and immunocytologic studies for Aspergillus species. Cytomegalovirus was detected in 2 of 47 specimens (4%) by cytologic examination and 7 of 47 (15%) by in situ hybridization. Cells with nuclear and/or cytoplasmic inclusions invariably were labeled with the CMV DNA probe. The weak diagnostic value of the cytologic examination resulted from the absence of characteristic inclusions in many specimens with positive results by in situ hybridization. P. carinii was the most frequent pathogen isolated from BAL fluid (9 of 47 cases; 19%). It was found in 1 of 47 specimens (2%) by the cytologic examination of Papanicolaou-stained smears, 4 of 47 (8.5%) by the GMS stain, and 8 of 47 (17%) by immunofluorescence. Most P. carinii-positive cases (five of nine cases) were detected by immunofluorescence only. Aspergillus species was diagnosed in 2 of 47 specimens (4%) by cytologic examination and GMS staining. Immunocytologic studies had positive results in these specimens and detected one additional case of Aspergillus infection (3 of 47 cases; 6%). These data show that techniques using CMV DNA probes and anti-P. carinii or anti-Aspergillus antibodies are rapid and more sensitive than conventional diagnostic procedures.
Aspergillus↗
Comparison of bone marrow sections, smears and immunohistological staining for immunoglobulin light chains in the diagnosis of benign and malignant plasma cell proliferations.
Bone marrow specimens from 118 patients with clinical suspicion of multiple myeloma were studied to assess the diagnostic reliability of histological and cytological criteria. Plasma cell clonality was assessed by demonstrating light chain restriction. In most cases of multiple myeloma, the classical cytological and histological criteria were found. In eight cases in which the marrow contained less than 10% of plasma cells, there was discordance between the light microscopic and immunohistochemical findings. In six cases without evidence of multiple myeloma and with polyclonal plasma cells in bone marrow, abnormal plasma cells resembling a malignant proliferation were found. These findings indicate that comparison of histological and cytological results with immunohistological studies for immunoglobulin light chains in bone marrow biopsy sections can be helpful in the evaluation of patients with a suspicion of multiple myeloma and when the marrow contains less than 10% of plasma cells.
Biopsy↗
Evidence that insulin-like growth factor 2 (IGF2) is the dominant thymic peptide of the insulin superfamily.
The central T-cell tolerance of neuroendocrine functions has been proposed to be primarily induced by the thymic repertoire of neuroendocrine self antigens. The present study aimed at characterizing the human thymic insulin-related autoantigen able to represent the pancreatic B-cell function in face of the developing T-cells. Immunofluorescence studies were performed on human and rat thymic sections, as well as on the rat IT-45R1 thymic epithelial cell line using several antibodies to epitopes of the insulin peptide superfamily. These studies identify beyond any doubt that insulin-like growth factor 2 (IGF2) is the dominant thymic peptide of the insulin family. The sequence of an insulin-derived autoantigen is proposed. This autoantigen is a nonamer and has a hydrophobic residue leucine (L) at position 9. In the human species, this autoantigen would primarily be tolerogenic for the pancreatic B-cell endocrine function during fetal development.
Amino Acid Sequence↗