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Biomedical subjects

J G Fournier

Publications and source records attributed to J G Fournier.

At least 37 records · Page 2Linked to original sources
PubMed1992

Ultrastructural localization of HIV-1 RNA and core proteins. Simultaneous visualization using double immunogold labelling after in situ hybridization and immunocytochemistry.

The analysis of human immunodeficiency virus type 1 (HIV-1) RNA sequences in CEM and Jurkat lymphoid cells infected with the virus has been performed at the subcellular level. Using a biotinylated DNA probe specific for HIV-1, virus RNA sequences were detected on Lowicryl thin sections after immunogold cytochemistry. The labelling observed on the cytoplasm was localized near the plasma membrane connected with extracellular cluster of virions. On free immature and nascent form of the virus the detection of HIV-1 RNA was associated with the peripheral electron-dense structure, whereas on mature form the labelling was concentrated on the central nucleoid known to be the site of the HIV-1 genomic RNA. The identification of virus RNA was also performed simultaneously with the detection of HIV-1 core protein p24 or p17 using a double immunogold labelling. Whereas the HIV-1 RNA showed again a cytoplasmic and virions localization, the structural protein was only observed on viral formations. The cytoplasmic localization of virus RNA, at the time of virus production, suggests that they are of genomic origin destined to be packaged in virions once the assembly of virus structural proteins has taken place in the plasma membrane at the viral budding site. The present molecular investigation conducted at the subcellular level provides insight into the cell periphery distribution of HIV-1 RNA observed at the light microscope as corresponding to the detection of HIV-1 infected lymphoid cells actually releasing virions.

Biotin↗
PubMed1992

Localization by immunogold labelling of HIV-1 structural proteins on Lowicryl embedded HIV-1 infected cell ultrathin sections.

Using several HIV-1-specific antibodies and the immunogold labelling technique, we have detected and localized distinct viral proteins on ultrathin sections of HIV-1 infected cells embedded in the Lowicryl K4M resin. Monoclonal antibodies (MAbs) against p24, p17 and gp160/gp41 showed a preferential labelling on viral formations still attached to the cell membrane (budding process) or free in the extracellular space. The anti-p24 and the anti-p17 MAb yielded a gold labelling not only on mature but also on immature virions where the gold particles were associated with the ring-like electron-dense material. The three HIV-1-specific antibodies against p24, p17 and p55 yielded a cross-reaction with HIV-2 in agreement with the conservation of the internal antigenic determinants of both viruses. In all instances, there was no specific immunogold labelling over the cells, suggesting that once the virus structural proteins were synthesized, they were promptly utilized for virus assembly at the plasma membrane level.

Acrylic Resins↗
PubMed1991

Skeletal muscle involvement in human immunodeficiency virus infection.

In addition to muscle changes due to peripheral nervous system involvement, primary myopathic changes associated with the human immunodeficiency virus (HIV) have also been described. We studied seven cases: two had developed an acquired immunodeficiency syndrome (AIDS), four had seroconverted to HIV but were otherwise asymptomatic, one was HIV seronegative when the biopsy was performed and one was biopsied twice. Besides the HIV no other infectious agent was detected. Muscle biopsies showed: (a) muscle fiber necrosis and regeneration; (b) inflammatory changes with moderate perivascular infiltration; and (c) unusual myofibrillary disorganization. Immunocytochemical techniques using anti-HIV monoclonal antibodies showed the presence of the virus in one biopsy. HIV-RNA was detected by in situ hybridization in the same biopsy. With both techniques the HIV was detected in isolated mononuclear cells in the muscle endomysium and not within the muscle fibers. Muscle involvement associated with HIV infection may be related, at least in some cases, to the presence of the virus in interstitial cells.

Acquired Immunodeficiency Syndrome↗
PubMed1991

Do measles early giant cells result from fusion of non-infected cells? An immunohistochemical and in situ hybridization study in a case of morbillous appendicitis.

A case of acute appendicitis with features of measles inflammatory reaction is studied. Two types of lymphoid polykaryons are seen: Warthin Finkeldey cells inside germinal centres (LN1 ++, LN2 +, L26 +, MB1 +, MB2 +/-) and multinucleate plasma cells in the lamina propria (mu +, alpha ++, kappa +, lambda +). Both types of polykaryon are devoid of inclusions. The search for viral genetic information by in situ hybridization was negative in these cells. A positive signal was observed in interfollicular mononuclear cells and rare enterocytes. A possible mechanism of fusion from without, acting at the beginning of the disease to induce the appearance of polykaryons, is discussed.

Acute Disease↗
PubMed1991

Analysis of human mitochondrial transcripts using electron microscopic in situ hybridization.

Human mitochondrial transcripts have been examined at the ultrastructural level. After contact with ultrathin sections of a human lymphoid cell line (CEM) embedded in Lowicryl K4M, biotinylated mitochondrial probes yield specific hybrids identified by a colloidal gold immunocytochemistry marker that visualizes rRNA and mRNA coding for respiratory chain polypeptides CO II, CO III and ATPase-6. The mitochondrial transcripts are preferentially located close to the inner membrane, particularly the cristae, suggesting that intra-organelle protein synthesis is intimately associated with the mitochondrial membrane system. Quantitative analysis indicates that the mitochondria concentrate the labeling with intensities that vary with the type of RNA and that the nucleus induces a light hybridization signal with each mitochondrial probe. The visualization of human mitochondrial DNA expression in correlation with the fine anatomy of the mitochondria constitutes a new approach for fundamental research on the organelle and for analyzing its behaviour in human mitochondrial diseases.

Adenosine Triphosphatases↗
PubMed1991

Ultrastructural detection of cellular and viral RNA with biotinylated DNA probes.

We describe a procedure for detecting and localizing cellular rRNA and HIV virus RNA, based on in situ hybridization at the ultrastructural level. After contact with ultrathin sections of cells embedded in Lowicry K4M, biotinylated DNA probes yield specific hybrids identified by a colloidal gold immunocytochemistry marker. The cellular transcripts are preferentially located in the ribosome structures of the cytoplasme and in the dense fibrillar component of the nucleolus. With the HIV probe, viral RNA molecules, were observed over infected cell and some extracellular viral particles. In the cytoplasm the hybridization signal is detected frequently at the periphery close to the plasma membrane, whereas the gold signal associated with virions is localized mainly over the center (core) of the particle.

Biotin↗
PubMed1990

[The detection of viral antigens and ribosomal RNA in cells by using low-temperature media].

A post-embedding technique for immunocytochemical analysis at the ultrastructural level was used to detect and localize HIV antigens on ultrathin sections of Lowicryl-embedded HIV-infected cells. A genomic probe containing ribosomal sequences and labeled with biotin was used to hybridize rRNA molecules in sections of animal cells embedded in Lowicryl. The method presently described offers the possibility to detect rapidly and precisely ribosomal gene expression and viral proteins at the ultrastructural level.

Acrylic Resins↗
PubMed1989

Presence of a constitutive paracrine beta-interferon in v-mos-bearing nonmalignant reverted cells.

A stable nonmalignant revertant cell line was derived from Moloney murine sarcoma virus-transformed BALB/c cells after long-term cultivation in the presence of murine type I interferon (IFN). These cells gradually established resistance to exogenous IFN and were also seen to contain IFN-dependent proteins. The presence of an endogenous IFN was confirmed by the results of Northern blot analysis and in situ hybridization with an IFN-beta probe, showing that only mRNA specific for IFN-beta- could be found in the uninduced reverted cells. The latter synthesized only a small amount of IFN-beta protein and exhibited few IFN-specific membrane receptors, which bound recombinant IFN-beta with a high affinity. After treatment with IFN antibody, the overexpression of H-2 major histocompatibility antigen genes was significantly down-regulated. These findings strongly suggest the existence in this reverted cell line of a constitutive IFN which, acting through an autocrine and/or paracrine mechanism, might play a role in maintaining the reverted state.

Animals↗
PubMed1989

Detection by immunogold techniques of HIV antigens in Lowicryl ultrathin sections of infected cells.

A post-embedding technique for immunocytochemical analysis at the ultrastructural level was used to detect and localize HIV antigens on ultrathin sections of Lowicryl K4M-embedded HIV-infected cells. With serum from an AIDS patient, specific immunogold labelling was obtained exclusively on mature viral extracellular structures. The more intense reactivity was obtained with core antigens. The present immunoelectron microscopy method provides several advantages - high sensitivity of immunodetection, good preservation of cellular morphology, easy preparation procedure - which could lead to the use of this method for HIV-infected human tissues.

Acquired Immunodeficiency Syndrome↗
PubMed1989

Detection by in situ hybridization of HIV I RNA in spleens of HIV I sero-positive patients with thrombocytopenic purpura.

In situ hybridization with a 3H labelled probe on cryosections from 6 spleens of HIV I sero-positive patients with thrombocytopenic purpura showed the presence of HIV RNA in 4 of the 6 spleens at the follicular hyperplasia stage. Two patterns of hybridization were observed: first, a diffuse autoradiographic signal, displaying an irregular network, detected in 1 or 2 germinal centres (GC) per section (17%); secondly, the presence of very few distinct radioactive cells in the labelled GC. A similar pattern was observed in an ARC (Acquired immunodeficiency syndrome-Related Complex) lymph node, but with a more intense and frequent hybridization signal. These results indicate that the spleens, like the lymph nodes, are involved in the course of HIV infection but with a less intense tissue-virus interaction, which may explain the minor morphological changes observed in the spleens. In addition, a careful examination of the lymph node tissue indicated that lymphocytes are the predominant cell type infected with the virus. As for the follicular dendritic cells (FDC), a similarity of the hybridization signal observed in the GC and in vitro HIV infected cells suggests that the FDC could also be sensitive to the virus.

AIDS Serodiagnosis↗
PubMed1989

[Ultrastructural detection of ribosomal RNA by in situ hybridization using a biotinylated probe on ultrathin sections of cultured animal cells].

A genomic probe containing ribosomal sequences and labelled with biotin was used to hybridize rRNA molecules in ultrathin sections of animal cells embedded in Lowicryl K4M. After detection with streptavidin conjugated with 10 nm gold particles, ribosomal target sequences were localized preferentially in the dense fibrillar component of the nucleolus and in the polyribosome structures of the cytoplasm. The method presently described offers the possibility to detect rapidly and precisely ribosomal gene expression at the ultrastructural level, particularly under different physiological and pathological conditions.

Animals↗
PubMed1988

Detection of mononuclear cells expressing hepatitis B virus in peripheral blood from HBsAg positive and negative patients by in situ hybridisation.

In situ hybridisation was used to detect RNA of Hepatitis B virus (HBV) in peripheral mononuclear blood cells. Presence of HBV-RNA was detected in 10/17 HBsAg positive HBeAg positive patients, in 8/18 HBsAg positive HBeAg negative patients, in 1/3 anti-HBs positive subjects. Four control subjects were negative. The frequency of labelled cells varied from 1 to 10% of mononuclear cells. These results indicate that the DNA detected so far in mononuclear blood cells may be transcriptionally active.

Adolescent↗
PubMed1988

Detection of vesicular stomatitis virus (VSV) RNA in the central nervous system of infected mice by in situ hybridization.

Using in situ hybridization with a cloned DNA probe specific for the VSV G protein, viral RNA was detected and localized in CNS tissue of mice infected i.c. with either wild or ts G 31 VSV mutant. In both cases, brain and spinal cord neurons were the only cells seen to contain viral RNA. Virus-positive neurons were observed enclosed in spongious areas induced by the ts VSV mutant. These results suggest that the VSV shows a strong tropism for the neuronal cell and indicate that the vacuole formation might be associated with the expression of the VSV G protein gene in infected neurons.

Animals↗
PubMed1988

Peritubular cells are the site of erythropoietin synthesis in the murine hypoxic kidney.

Erythropoietin (Epo)-producing cells were identified in the murine hypoxic kidney by in situ hybridization. Profound anemia was induced in order to greatly increase Epo production. This resulted in high levels of Epo mRNA in the kidney. 35S-labeled DNA fragments of the murine Epo gene were used as probes for in situ hybridization. Control experiments conducted in parallel included kidneys of nonanemic mice, RNase-treated hypoxic kidney sections, and 35S-labeled non-Epo-related DNA. The Epo probe gave a specific hybridization signal in the hypoxic kidney in the cortex and to a lesser extent in the outer medulla. Glomerular and tubular cells were not labeled. All positive cells were identified as peritubular cells. Using immunofluorescence, we showed that cells with the same topography contained Factor VIII-related antigen. These data demonstrated that peritubular cells, most likely endothelial cells, constitute the major site of Epo production in the murine hypoxic kidney.

Animals↗
PubMed1988

Detection and localization of renin messenger RNA in human pathologic tissues using in situ hybridization.

In order to investigate the synthesis of renin in human pathologic tissues, the authors used in situ hybridization to detect and localize renin messenger RNA (mRNA). The probe was a 35S-radiolabeled 1.1-kb length complementary DNA of human renal renin. To compare the synthesis with the presence and the storage of renin, renin antigen was assessed by immunohistochemistry in the same tissues. The human pathologic tissues were as follows: two ischemic kidneys related to renovascular hypertension; two renal juxtaglomerular cell tumors; one extrarenal renin-secreting epithelioid sarcoma of soft tissues. In ischemic kidneys, the cells containing both renin mRNA and renin protein were found in numerous juxtaglomerular apparatus and in the wall of arterioles, shown by combined in situ hybridization and immunohistochemistry. Most of the tumor cells in the juxtaglomerular cell tumors and scarce tumor cells in the epithelioid sarcoma of soft tissues were positive by in situ hybridization and immunohistochemistry. These findings demonstrate that the presence of renin in these tissues is associated with local cellular production of renin. In particular, smooth muscle cells of the wall of arterioles are definitely capable of synthesizing renin. Moreover, in these tissues, gene expression (renin synthesis) and renin storage are concordant.

Adult↗