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Biomedical subjects

E Bertrand

Publications and source records attributed to E Bertrand.

At least 37 records · Page 2Linked to original sources
PubMed2000

Alkaline fixation drastically improves the signal of in situ hybridization.

In situ hybridization (ISH) is widely used to detect DNA and RNA sequences within the cell and tissue sections. The important step in performing this technique is tissue fixation. We investigated the influence of the pH of the fixative on the outcome of ISH. Our studies indicate that alkaline formaldehyde dramatically increases the ISH signal with RNA probes. The increase in signal was observed for detection of low as well as for high abundance messages. The sensitivity of the method was increased 5- to 6-fold.

Cell Cycle Proteins↗
PubMed2000

Dynamics of wetting layer formation

We study the formation and growth of wetting layers in the binary liquid mixture cyclohexane-methanol. By progressively deuterating the methanol we can tune the equilibrium wetting layer thickness. Hysteresis of the transition is observed for large thicknesses and is absent for thinner ones. This can be understood by calculating the activation energy for wetting layer nucleation as a function of the film thickness. We also show that the late-stage growth of the wetting layer after the nucleation process follows a power law in time, in agreement with a diffusion-limited growth mechanism proposed theoretically.

Journal Article↗
PubMed2000

Expressing active ribozymes in cells.

Artificially engineered ribozymes can be used to specifically regulate expression of target genes. Such ribozymes can be synthesized chemically and delivered into the cell exogeneously. Alternatively, ribozymes can be produced by the cell endogenously, after introduction of the artificial gene into the cellular genome. In the latter case, the design of the artificial gene defines the ribozyme properties, such as: expression level, intracellular localization, folding and association with proteins. Generally speaking, design of the expression vector is critical to obtain active ribozyme molecules. This paper first describes factors that are known or predicted to affect ribozyme activity in the cell, then reviews various expression systems that have been specifically developed for ribozymes. Lastly, a recently developed ribozyme system termed snorbozymes (small nucleolar RNA:ribozyme hybrids) will be discussed. This powerful test system has generated several important observations that are likely to affect the future development of ribozyme technology.

Animals↗
PubMed2000

[Conduction disorders in chronic parietal endocarditis or endomyocardial fibrosis. 170 cases at the Cardiology Institute of Abidjan].

The authors have conducted research on conduction disturbances in the endomyocardial fibrosis, synonymous with chronic parietal endocarditis, about 170 cases at the Institute of Cardiology in Abidjan, from January 1977 to June 1991. The anatomical and/or angiographic examination have permitted to describe 64 cases of right fibrosis, 24 cases of left fibrosis and 82 cases of bilateral fibrosis. Conduction anomalies have been observed among 42.9% of the patients. Among 92 anomalies recorded, the most frequent has been the first degree heart block (43.5%) and the incomplete right bundle branch block (30.4%). The old age of the patients and the right localization were the factors associated with conduction disturbances in endomyocardial fibrosis (difference not significant). Yet, fibrosis surgery, especially the decortication of the fibrous endocardium of the right ventricle, have generated one or many conduction anomalies among most of our operated patients. The right branch of the fasciculus of the His has been the most injured by the fragmented techniques of METRAS who had, therefore, the merit to have minimized the incidence of the complete post-operative heart blocks still high in European and Brazilian series. In term of prognosis, no conduction disturbance has directly caused a patient's death, even if those anomalies cannot be totally ruled out in the 16 cases of sudden death.

Adolescent↗
PubMed1999

A small nucleolar RNA:ribozyme hybrid cleaves a nucleolar RNA target in vivo with near-perfect efficiency.

A hammerhead ribozyme has been localized to the yeast nucleolus by using the U3 small nucleolar RNA as a carrier. The hybrid small nucleolar RNA:ribozyme, designated a "snorbozyme," is metabolically stable and cleaves a target U3 RNA with nearly 100% efficiency in vivo. This is the most efficient in vivo cleavage reported for a trans-acting ribozyme. A key advantage of the model substrate featured is that a stable, trimmed cleavage product accumulates. This property allows accurate kinetic measurements of authentic cleavage in vivo. The system offers new avenues for developing effective ribozymes for research and therapeutic applications.

Base Sequence↗
PubMed1999

[Severe hemorrhage from rupture of an intra-oral arteriovenous malformation].

We report the case of a 47-year-old woman with a sudden, spontaneous exsanguinating haemorrhage from an arteriovenous malformation of the mandible. The diagnosis was obtained with a contrast enhanced CT-scan and a digital subtraction angiography. As arterial embolisation was not available the lesion was treated surgically.

Angiography↗
PubMed1999

mRNA localization signals can enhance the intracellular effectiveness of hammerhead ribozymes.

Subcellular localization signals for several mRNAs are positioned in their 3' untranslated regions (UTR). We have utilized the human alpha- and beta-actin 3' UTRs as signals for colocalizing hammerhead ribozymes with a lacZtarget mRNA. Ribozyme and target genes containing matched or unmatched 3' UTRs were cotransfected into 12-day-old chicken embryonic myoblast and fibroblast (CEMF) cultures and assayed by in situ hybridization (ISH) using a dual label, antibody sandwich procedure, and dual fluorescence microscopy to monitor intracellular colocalization. Beta-galactosidase localization in transfectants was visualized by incubation with X-gal and also quantitated by an o-nitrophenyl beta-D-galactopyranoside (ONPG) assay. We found that the percentage of colocalization using the matched alpha- or beta-actin 3' UTR (alpha-alpha or beta-beta) was enhanced approximately threefold relative to unmatched 3' UTRs. The increase in ribozyme-mediated inhibition of beta-galactosidase activity observed when matched 3' UTRs were used was consistent with the observed percentage of colocalization. These results represent the first direct demonstration that mRNA localization signals (zipcodes) can be utilized to enhance intracellular ribozyme efficacy.

3' Untranslated Regions↗
PubMed1999

Central nervous system infection caused by Borrelia burgdorferi. Clinico-pathological correlation of three post-mortem cases.

The spirochete Borrelia burgdorferi (B. burgdorferi) may cause severe meningoencephalomyelitis as the sole manifestation of Lyme borreliosis. We would like to present three such cases, where definite neuroborreliosis was clinically diagnosed in two cases and possible neuroborreliosis was recognized in one case. Alive spirochetes were isolated and cultured from blood and cerebrospinal fluid (CSF) in both definite cases. B. burgdorferi as the causative agent of the infection was confirmed in CSF by polymerase chain reaction (PCR) in one definite case. In the possible case spirochetes were cultured from blood and CSF. Alive spirochetes were not isolated, however anti-B. burgdorferi antibody value in serum was significantly elevated. On necropsy gross examination brain edema without focal changes was detected in two cases. Cerebral atrophy was seen in Case 3. Microscopically, lymphocytic infiltrates, microglial diffuse and nodular activation, spongiform changes, diffuse demyelination of the cerebral and cerebellar white matter, and diffuse astrocytosis, were characteristic pathological features in all presented cases. Multifocal, perivascular degenerative changes in the cerebral and cerebellar white matter were observed in the first case. Inflammatory changes in the nuclei and roots of cranial nerves were present in the third case.

Adult↗
PubMed1999

The significance of immunocytochemical markers, synaptophysin and neurofilaments in diagnosis of ganglioglioma.

Ganglioglioma is a tumor composed of neoplastic neurons and neoplastic glial cells mixed in different proportions. Astrocytes are the essential glial component. The tumor proliferates mostly in the temporal lobe cortex, scarcely in other areas of the brain. In ganglioglioma the population of ganglionic neurons is very difficult to diagnose. In the last ten years, immunocytochemical reaction with synaptophysin and neurofilament protein (NFP) triad considered as neoplastic neuron markers seemed to differentiate neoplastic nerve cells from normal neurons occurring incidentally within neoplastic proliferation area. In our study, the reaction with synaptophysin and NFP was performed on fragments of normal frontal lobe, pons and cerebellum as well as on fragments of post-operation tumor diagnosed as ganglioglioma. A positive synaptophysin reaction was obtained on the surface of neoplastic neurons in gangliogliomas, as well as on the surface of normal neurons in the encephalon, cerebellum and pons. Both neoplastic and normal neurons and their processes showed the expression of neurofilaments protein triad. Thus, a positive reaction of neurons with synaptophysin and NFP seems to be nonpathognomic in the diagnosis of the neoplastic neuron population in gangliogliomas.

Adult↗
PubMed1999

Hallervorden-Spatz disease in an adult patient.

Hallervorden-Spatz disease (HSD) is an extremely rare degenerative process. The familial studies point to inherited, autosomal recessive neurodegenerative disorder. Quite recently this disease gene has been identified to chromosome 20p12.3-p13. Clinical manifestations of HSD leading to death after several years of illness are most frequently observed in childhood. HSD in adults is very scarce. The case reported concerns a woman who at the age of 26 years began to suffer from slowly progressing psycho-organic syndrome with muscular rigidity, involuntary movements and dysarthria. The patient was hospitalized several times with successive diagnoses of multiple sclerosis, amyotrophic lateral sclerosis and Huntington's disease. Shortly before death magnetic resonance imaging (MRI) scan showed a decreased signal in both basal ganglia. The patient died at the age of 34 years after an eight-year illness. In the brain autopsy symmetric hyperpigmentation of globus pallidus (GP) and reticular part of substantia nigra (SN) was found. The microscopic observation revealed abundant deposits of brown pigment mostly in GP and SN. In addition, numerous spheroids disseminated in the basal ganglia, mesencephalon and medulla oblongata, as well as Lewy bodies in SN were noted. Pigment deposits expressed intensive iron positive reaction by Perls' Prussian-blue method. Based on the described neuropathological changes occurring mostly in GP and SN, Hallervorden-Spatz disease was diagnosed.

Adult↗
PubMed1999

Microglia and neuritic plaques in familial Alzheimer's disease induced by a new mutation of presenilin-1 gene. An ultrastructural study.

The results of the ultrastructural study of the brains of two sisters with familial Alzheimer's disease (AD) induced by a new mutation of presenilin-1 (PS-1) gene who died at the young age (35 and 37 years) are presented. In both cases, the changes typical of AD with particularly large number of neuritic plaques (NPs) were found. Microglial cells were located between amyloid core and neurites. At the ultrastructural level, the content of microglial cytoplasm was differentiated (amyloid fibrils or/and phagocytic bodies). This may suggest that microglial cells participate in forming of amyloid fibrils and/or phagocytosis of amyloid.

Adult↗
PubMed1999

Border zone neovascularization in cerebral ischemic infarct.

Immunocytochemical and quantitative studies on vascular reaction (angiogenesis) in cortical border zone of infarct were undertaken. Intensity and temporal profile of angiogenesis was assessed in 60 patients aged between 48 and 69 (younger group), and between 70 and 92 (older group), with cerebral infarct in the area of middle cerebral artery vascularization, who died during the first six weeks following the stroke. We have found that angiogenesis was a multistage process in which four stages were distinguished: phase of primary activation of endothelial cells, two consecutive phases of active angiogenesis and final phase of only sporadic proliferation of vessels. The distinction of phases in a multiphase angiogenic cascade helped us to evaluate the correlation with survival time and the age of patients. The most pronounced intensification of angiogenesis and increased density of CD 31 positive capillaries in penumbra were observed in the second phase, especially in younger patients. The duration of the penumbral neovascularization decreased in the older age patient. Our results indicate that sprouting angiogenesis is a quantitatively significant source of vessels in the cerebral infarct border zone. However, non-therapeutically stimulated angiogenesis developed only 3-4 days after the stroke, that is beyond the period of reversible changes in ischemic penumbra recognized as a "therapeutical window" in the human brain. The angiogenic therapy opens a new way towards the revascularization of ischemic brain infarct.

Aged↗
PubMed1998

Nucleolar localization of early tRNA processing.

There is little information as to the location of early tRNA biosynthesis. Using fluorescent in situ hybridization in the budding yeast, Saccharomyces cerevisiae, examples of nuclear pre-tRNAs are shown to reside primarily in the nucleoli. We also probed the RNA subunit of RNase P. The majority of the signal from RNase P probes was nucleolar, with less intense signals in the nucleoplasm. These results demonstrate that a major portion of the tRNA processing pathway is compartmentalized in nucleoli with rRNA synthesis and ribosomal assembly. The spatial juxtaposition suggests the possibility of direct coordination between tRNA and ribosome biosynthesis.

Base Sequence↗
PubMed1998

The snoRNA box C/D motif directs nucleolar targeting and also couples snoRNA synthesis and localization.

Most small nucleolar RNAs (snoRNAs) fall into two families, known as the box C/D and box H/ACA snoRNAs. The various box elements are essential for snoRNA production and for snoRNA-directed modification of rRNA nucleotides. In the case of the box C/D snoRNAs, boxes C and D and an adjoining stem form a vital structure, known as the box C/D motif. Here, we examined expression of natural and artificial box C/D snoRNAs in yeast and mammalian cells, to assess the role of the box C/D motif in snoRNA localization. The results demonstrate that the motif is necessary and sufficient for nucleolar targeting, both in yeast and mammals. Moreover, in mammalian cells, RNA is targeted to coiled bodies as well. Thus, the box C/D motif is the first intranuclear RNA trafficking signal identified for an RNA family. Remarkably, it also couples snoRNA localization with synthesis and, most likely, function. The distribution of snoRNA precursors in mammalian cells suggests that this coupling is provided by a specific protein(s) which binds the box C/D motif during or rapidly after snoRNA transcription. The conserved nature of the box C/D motif indicates that its role in coupling production and localization of snoRNAs is of ancient evolutionary origin.

Animals↗
PubMed1998

Localization of ASH1 mRNA particles in living yeast.

ASH1 mRNA localizes to the bud tip in Saccharomyces cerevisiae to establish asymmetry of HO expression, important for mating type switching. To visualize real time localization of the mRNA in living yeast cells, green fluorescent protein (GFP) was fused to the RNA-binding protein MS2 to follow a reporter mRNA containing MS2-binding sites. Formation and localization of a GFP particle in the bud required ASH1 3'UTR (untranslated region) sequences. The SHE mutants disrupt RNA and particle localization and SHE 2 and 3 mutants inhibit particle formation as well. Both She3myc and She1myc colocalized with the particle. Video microscopy demonstrated that She1p/Myo4p moved particles to the bud tip at 200-440 nm/sec. Therefore, the ASH1 3'UTR-dependent particle serves as a marker for RNA transport and localization.

3' Untranslated Regions↗
PubMed1998

3'-End modification of the adenoviral VA1 gene affects its expression in human cells: consequences for the design of chimeric VA1 RNA ribozymes.

Polymerase III (pol III)-dependent genes, like the adenoviral VA1 gene, are of particular interest for expressing small therapeutic RNAs into cells. A new VA1 RNA carrier molecule was generated through the deletion of the VA1 RNA central domain to give rise to the VAdeltaIV RNA vector that was devoid of undesirable physiologic activity (i.e., inhibition of the interferon-induced protein kinase, PKR). This vector was used to express in human cells hammerhead ribozymes targeted against the human immunodeficiency virus (HIV). Eight anti-HIV ribozymes were inserted at the 3'-end of this vector immediately before the four T-residues that serve as a transcription termination signal. Although the constructs were active in vitro, they failed to inhibit HIV replication in transient assays. Analysis of the intracellular ribozyme expression in cells revealed several anomalies. First, using mutant derivatives, we showed that the presence of two or three consecutive T-residues in the ribozyme portion was sufficient to promote the release of anomalous short transcripts. Second, when the ribozyme did not contain T-rich sequence, full-length transcripts were produced, but these transcripts were very unstable and were retained in the cell nucleus. In contrast, insertion of the ribozyme in place of the central domain of VA1 RNA led to production of full-length transcripts that were stable and located in the cytoplasm but that were not found to be active in vitro. Taken together, these results have important consequences for the future design of active intracellular ribozymes based on the use of pol III-transcribed genes.

3' Untranslated Regions↗
PubMed1998

Monitoring retroviral RNA dimerization in vivo via hammerhead ribozyme cleavage.

We have used a strategy for colocalization of Psi (Psi)-tethered ribozymes and targets to demonstrate that Psi sequences are capable of specific interaction in the cytoplasm of both packaging and nonpackaging cells. These results indicate that current in vitro dimerization models may have in vivo counterparts. The methodology used may be applied to further genetic analyses on Psi domain interactions in vivo.

3T3 Cells↗