Biomedical subjects

G Chevalier

Publications and source records attributed to G Chevalier.

At least 19 recordsLinked to original sources

PubMed2004

PCR-RFLP using a SNP on the mitochondrial Lsu-rDNA as an easy method to differentiate Tuber melanosporum (Perigord truffle) and other truffle species in cans.

Canned truffle products labeled Tuber melanosporum, the famous Perigord truffle, may contain other less tasty and cheaper truffle species. To protect consumers from fraud, a PCR DNA-based method was used to unequivocally identify the nature of the product. Several rapid and simple cell lysis procedures, used in conjunction with a commercially available DNA purification kit, were evaluated for their effectiveness in recovering DNA from canned truffle. In parallel, a marker for T. melanosporum was tested on the mitochondrial rDNA. These two techniques were then combined to differentiate T. melanosporum from other truffle species like T. aestivum, T. brumale or T. indicum up to the legal threshold in canned products. These findings not only allow a comparison of the effectiveness of the different DNA extraction methods but also provide a preliminary indication of the specificity and sensitivity of the detection with the mitochondrial marker that might be attainable for truffle species in a quantitative PCR-based analysis method.

Ascomycota↗

PubMed2004

Rapid molecular typing of Tuber melanosporum, T. brumale and T. indicum from tree seedlings and canned truffles.

We have developed new DNA extraction and purification procedures for investigation of mycorrhized seedlings and canned truffles. Use of these procedures on approximately 100 mg initial material enabled good sample representation. For mycorrhized seedlings, Taq polymerase inhibitors were discarded irrespective of tree species. In routine analysis we systematically used consensus primers ITS1/ITS4 to check the absence of Taq polymerase inhibitors and the presence of fungus DNA. Positive response with ITS validates other positive or negative PCR results. Absence of amplification with ITS prevents validation of other results. For canned truffles, DNA harvested from ascocarps sterilized for one and a half hours at 115 degrees C was amplified with specific primers. We have developed consensus primers, named R12/F12, to check for the presence of amplifiable fungus DNA and the absence of Taq polymerase inhibitors. Here also, positive response with consensus R12/F12 validates other positive or negative PCR results. We have developed one primer pair specific for T. brumale and another specific for T. melanosporum. We can then characterize these two taxa, which enables the use of "truffle or truffled" French designations. We can also characterize T. indicum, the Asiatic black truffle that might fraudulently be sold as T. melanosporum and T. brumale. These three specific primer pairs were used independently of DNA extraction from tree seedlings or canned truffles. Our process is specific, sensitive, convenient, and quick.

DNA, Fungal↗

PubMed2001

Honeycomb-like structure of the intermediate layers of the rat superior colliculus: afferent and efferent connections.

There is increasing evidence that acetylcholinesterase is organised in a lattice-like fashion in the intermediate layers of the mammalian superior colliculus. In a recent study, we described this organisation in rat by showing that it comprises a well formed honeycomb-like lattice with about 100 cylindrical compartments or modules occupying both the intermediate collicular layers. Considering this enzyme domain as a reference marker for comparing the organisation of collicular input-output systems, the present study investigates whether the principal sensori-motor systems in intermediate layers also have honeycomb-like arrangements. In 33 animals, the distributions of afferents (visual from extrastriate cortex; somatic from the primary somatosensory cortex, the trigeminal nucleus and the cervical spinal cord) and efferents (cells of origin of the crossed descending bulbospinal tract and uncrossed pathway to the pontine gray, the ascending system to the medial dorsal thalamus) were examined in a tangential plane following applications of horseradish peroxidase-wheatgerm agglutinin conjugate (used as an anterograde and retrograde tracer). In 22 of the 33 rats, axonal tracing was made within single tangential sections also stained for cholinesterasic activity in order to compare the neuron profiles with the cholinesterasic lattice.The results show that these afferent and efferent systems are also organised in honeycomb-like networks. Moreover, those related to the cortical, trigeminal and some of the spinal afferents are aligned with the cholinesterasic lattice. Likewise most of colliculo-pontine, colliculo-bulbospinal and half of colliculo-diencephalic projecting cells also tend to be in spatial register with the enzyme lattice. This indicates that the honeycomb-like arrangement is a basic architectural plan in the superior colliculus for the organisation of both acetylcholinesterase and major sensori-motor systems for orientation.

Acetylcholinesterase↗

PubMed2001

The fine organization of nigro-collicular channels with additional observations of their relationships with acetylcholinesterase in the rat.

The nigro-collicular pathway that links the basal ganglia to the sensorimotor layers of superior colliculus plays a crucial role in promoting orienting behaviors. This connection originating in the pars reticulata and lateralis of the substantia nigra has been shown in rat and cat to be topographically organized. In rat, a functional compartmentalization of the substantia nigra has also been shown reflecting that of the striatum. In light of this, we reinvestigated the topographical arrangement of the nigro-collicular pathway by examining the innervation of each nigral functional zone. We performed small injections of either biocytin or wheatgerm agglutinin conjugated with horseradish peroxidase restricted to identified somatic, visual and auditory nigral zones. Frontally cut sections showed that innervations provided by the three main nigral zones form a mosaic of complementary domains stratified from the stratum opticum to the ventral part of the intermediate collicular layers, with the somatic afferents sandwiched between the visual and the auditory ones. When reconstructed from semi-horizontal sections, nigral innervations organized in the form of a honeycomb-like array composed of 100 cylindrical modules covering three-quarters of the collicular surface. Such a modular architecture is reminiscent of the acetylcholinesterase lattice we previously described in rat intermediate collicular layers. In the enzyme lattice, the surroundings of the cylindrical modules are composed of a mosaic of dense and diffuse enzyme subdomains. Thus, we compared the distribution of the overall nigral projection and of its constituent channels with the acetylcholinesterase lattice. The procedure combined axonal labelling with histochemistry on single sections for acetylcholinesterase activity. The results demonstrate that the overall nigral projection overlaps the acetylcholinesterase lattice and its constituent channels converge with either the dense or the diffuse enzyme subdomains. The stereometric arrangement of the nigro-collicular pathway is suggestive of an architecture promoting the selection of collicular motor programs for different classes of orienting behavior.

Acetylcholine↗

PubMed2001

[Rapid determination of sex in Myocastor coypus embryos in the first stage of gestation].

The early knowledge of the sex may be crucial for the understanding of many features of ecological and evolutive biology, including offspring sex-ratio adjustment and evolution of breeding systems. In coypu (Myocastor coypus), significant variation in birth sex-ratios can be observed and selective abortion of entire litters is one of the cited mechanisms. In order to determine the sex of coypu embryos in the earlier stages of gestation (second week), we developed a molecular technique based on PCR amplification of a region of the Sry gene. These method used the combination of two sets of primers: one specific of the Y-chromosome; the other one, autosomal, is a positive control for amplification. Because of the direct amplification of embryo lysate without DNA extraction, the present sexing technique is rapid, relatively simple and inexpensive, and presents numerous advantages for the study at population scale.

Animals↗

PubMed2000

Honeycomb-like structure of the intermediate layers of the rat superior colliculus, with additional observations in several other mammals: AChE patterning.

The aim of the present study was to reinvestigate the stereometric pattern of acetylcholinesterase (AChE) activity staining in the intermediate layers of the superior colliculus in several mammalian species. A pioneering study in the cat and the monkey by Graybiel (1978) stressed the regular arrangement of AChE staining in the deep collicular layers. According to her description, made in the frontal plane, the enzyme was arranged in a mediolateral series of patches, the cores of which tended to line up in the longitudinal axis of the structure, so they formed roughly parallel bands. As exhaustive a description as possible of the AChE distribution was undertaken in the rat by compiling observations in the frontal, sagittal, and tangential planes. It emerged that AChE-positive elements are organized in the form of a conspicuous honeycomb-like network that is divided into about 100 rounded compartments, over virtually the full extent of the intermediate layers. The generality of the rat model was then tested in other rodents such as mouse and hamster and also in cat and monkey. For these species we resorted to a single tangential cutting plane, which proved to be more appropriate for disclosing such a modular arrangement. The data revealed that in all species AChE staining followed the same architectural plan and identified the striking similarity in the number of compartments that compose the various honeycomb-like lattices. In conclusion, the present findings support a unified model of the AChE arrangement within the intermediate layers of the mammalian colliculus; the model comprehensively incorporates the classical description of the patchy and stripy features of the enzyme distribution. We hypothesize here that the modular AChE arrangement might be the anatomical basis for collicular vectorial encoding of orienting movements.

Acetylcholinesterase↗

PubMed2000

Adult corneal epithelium basal cells possess the capacity to activate epidermal, pilosebaceous and sweat gland genetic programs in response to embryonic dermal stimuli.

Recent work has shown remarkable plasticity between neural and hematopoeitic, as well as between hematopoeitic and muscle stem cells, depending on environmental stimuli (Fuchs, E. and Segre, J. A. (2000) Cell 100, 143-155). Stem cells give rise to a proliferative transient amplifying population (TA), which is generally considered to be irreversibly committed. Corneal epithelium provides a particularly useful system for studying the ability of TA cells to activate different genetic programs in response to a change in their fibroblast environment. Indeed, corneal stem and TA cells occupy different localities - stem cells at the periphery, and TA cells more central (Lehrer, M. S., Sun, T. T. and Lavker, R. M. (1998) J. Cell Sci. 111, 2867-2875) - and thus can be discretely dissected from each other. It is well known that pluristratified epithelia of cornea and skin display distinct programs of differentiation: corneal keratinocytes express keratin pair K3/K12 and epidermal keratinocytes keratin pair K1-2/K10; moreover, the epidermis forms cutaneous appendages, which express their own set of keratins. In our experiments, central adult rabbit corneal epithelium was thus associated either with a mouse embryonic dorsal, upper-lip or plantar dermis before grafting onto nude mice. Complementary experiments were performed using adult mouse corneal epithelium from the Rosa 26 strain. The origin of the differentiated structures were identified in the first case by Hoechst staining and in the second by the detection of beta-galactosidase activity. The results show that adult central corneal cells are able to respond to specific information originating from embryonic dermis. They give rise first to a new basal stratum, which does not express anymore corneal-type keratins, then to pilosebaceous units, or sweat glands, depending of the dermis, and finally to upper layers expressing epidermal-type keratins. Our results provide the first evidence that a distinct TA cell population can be reprogrammed.

Animals↗

PubMed1999

Pilot study on the relationship between personality traits and skin conductivity of specific surface points as measured by Motoyama's apparatus.

This pilot study used Motoyama's electrodermal response-measuring apparatus to test resting skin conductivity from 28 extremity digit surface coordinates of 33 subjects who also were administered the Eysenck Personality Inventory on an alternatively assigned basis for the dimensions of extraversion and neuroticism. Correlations show significant relationships between neuroticism and extraversion raw scores on two of the 14 bilateral finger surface coordinates.

Adult↗

PubMed1998

Vitamin D receptor stable transfection restores the susceptibility to 1,25-dihydroxyvitamin D3 cytotoxicity in a rat glioma resistant clone.

Recently, 1,25-dihydroxyvitamin D3 (1,25-D3) and less hypercalcemic analogs were shown to exert a delayed cytotoxic effect on rat C6 glioma cells. 1,25-D3 induces in these cells a programmed cell death, accompanied by the induction of c-myc, p53 and gadd 45 genes. The involvement of the intracellular vitamin D receptor (VDR) remained to be determined. In this lethal process, we have investigated its role in a subclone of C6 cells, which was isolated on the basis of its resistance to 1,25-D3, and in which VDR expression was not detected either at the mRNA or protein levels. The stable transfection of a rat VDR cDNA into this clone restored its susceptibility to the cytotoxic effects of 1,25-D3. This phenomenon was accompanied by a dramatic upregulation of c-myc mRNA expression, as already described in a C6-sensitive clone. These results provide the first evidence that VDR expression, if not sufficient, is necessary to mediate 1,25-D3 cytotoxic effect in C6 glioma cells. Since VDR mRNA expression has been already reported in human brain tumors, our data imply that the identification of VDR expression could become a prerequisite in any strategy of glioma treatment with vitamin D analogs.

Animals↗

PubMed1998

Epigenetic control of programmed cell death: inhibition by 5-azacytidine of 1,25-dihydroxyvitamin D3-induced programmed cell death in C6.9 glioma cells.

In mammalian DNA cytosine methylation occurs specifically at CpG dinucleotide. Although the full array of function of DNA methylation is yet to be elucidated, it is well established that DNA methylation is an important mechanism involved in gene expression, DNA replication and cancer. Rat glioma C6.9 cells undergo programmed cell death (PCD) after treatment with 1,25-dihydroxyvitamin D3 (1,25-D3). Hence, these cells were used to study whether DNA methylation was involved in the control of PCD. We found that 1,25-D3-mediated PCD of C6.9 cells was suppressed by exposure of the cells to the DNA demethylating agents 5-azacytidine (5-AzaC) and 5-aza-2'-deoxycytidine. This effect remains detectable several cell divisions following removal of 5-AzaC and, therefore, involves DNA methylation as an epigenetic regulatory mechanism of PCD. Accordingly, internucleosomal fragmentation, a feature of apoptosis that is detected in 1,25-D3-treated cells, is no longer observable after treatment of these cells with 5-AzaC. However, 5-AzaC does not totally suppress the responsiveness of C6.9 cells to 1,25-D3 since the induction of the c-myc gene remains unaffected. These results suggest that a change in DNA methylation pattern could suppress 1,25-D3-mediated PCD through the expression of previously hypermethylated genes such as proto-oncogenes with death-repressor activity, endogenous virus sequences or even genes inducing change in the differentiated state of these cells.

Animals↗

PubMed1998

Both retinoic acid receptors alpha (RARalpha) and gamma (RARgamma) are able to initiate mouse upper-lip skin glandular metaplasia.

Embryonic mouse upper-lip skin explants treated with 16.7 microM all-trans retinoic acid (tRA) give rise to a glandular metaplasia of hair vibrissa follicles; however, at this concentration, tRA can activate not only the three retinoic acid receptors (RARalpha, beta, and gamma), but also the retinoid X receptors (RXRalpha, beta, and gamma) as a consequence of its isomerization to 9-cis retinoic acid. We therefore studied the respective roles of the RXR and RAR by treating RARalpha(-/-), beta(-/-), and gamma(-/-) skin explants with tRA and wild-type explants with synthetic retinoids specific for RXR or for each of the RAR. The null mutation of the RARalpha, RARbeta, and RARgamma genes did not prevent tRA-induced hair glandular metaplasia, but RARgamma inactivation dramatically reduced its ratio. As demonstrated by treating explants with a RAR- or a RXR-specific panagonist (CD367 and Ro25-7386, respectively), RAR are primarily responsible for this metaplasia. The use of two retinoids (Ro40-6055, 8 x 10(-3) microM, or CD437, 7.7 x 10(-2) microM) that are believed to act, respectively, as a RARalpha- or a RARgamma-specific agonist showed that both these receptors can initiate a metaplasia. In contrast, BMS453, a RARbeta-specific agonist, was unable to give rise to any metaplasia. Nevertheless, the highest degrees and ratios of metaplasia were only obtained after treatment with the CD367 RAR panagonist, or with either Ro40-6055 or CD437 at a concentration sufficient to allow the activation of the three RAR, suggesting that RARbeta activation is required for a metaplasia of all vibrissae.

Animals↗

PubMed1998

novH: differential expression in developing kidney and Wilm's tumors.

We previously established that the expression of the human nov gene (novH) was altered in Wilms' tumors and that levels of novH and WT1 mRNA were inversely correlated in individual Wilms' tumors. Insofar as novH has been shown to be a target for WT1 regulation, novH might play an important role during normal nephrogenesis and in the development of Wilms' tumors. We now show that during normal nephrogenesis novH protein is tightly associated with differentiation of glomerular podocytes. NovH expression is not restricted to renal differentiation but is also detected in endothelium and neural tissue of the kidney. Our results establish that alteration of novH expression in sporadic and heritable Wilms' tumors is associated with dysregulated expression of both novH mRNA and protein. In general, the highest novH expression was noted in the Wilms' tumor, genitourinary anomalies, aniridia, and mental retardation (WAGR)-associated Wilms' tumors. Expression in the Denys-Drash syndrome (DDS)-associated Wilms' tumors fell within the variable spectrum observed in sporadic Wilms' tumor cases. As in developing kidney podocytes, novH protein was also prominent in the abnormal hypoplastic podocytes from DDS cases and in kidney podocytes adjoining Wilms' tumors. In Wilms' tumors exhibiting heterotypic differentiation, novH protein was expressed at high levels in tumor-derived striated muscle and at lower levels in tumor-derived cartilage. These observations taken together indicate that novH may represent both a marker of podocytic differentiation in kidney and a marker of heterotypic mesenchymal differentiation in Wilms' tumors. In addition, absence or very low levels of WT1 are correlated with higher novH expression, and its variable expression in cases with mutant WT1 (sporadic and DDS) suggests that the potential activation and repression transcriptional functions possessed by WT1 are likely dependent on the specific mutation incurred.

Animals↗

PubMed1997

Noradrenaline inhibits the programmed cell death induced by 1,25-dihydroxyvitamin D3 in glioma.

The rat glioma cell line C6.9 has been recently reported to respond to 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) by the induction of a programmed cell death. Since, in vivo, glial cells are thought to be exposed to several neurotransmitters, we investigated the possibility of a neurotransmitter-mediated inhibition of this active cell death process. Noradrenaline and the beta-adrenoceptor agonist isoproterenol showed significant inhibition of the 1,25(OH)2D3-induced programmed cell death. The beta-adrenoceptor antagonist propanolol reversed this inhibition, while the alpha-adrenoceptor antagonist yohimbin was devoid of any effect. This suggests that the efficiency of antiproliferative vitamin D-related therapies could be influenced by endogenous levels of noradrenaline.

Adrenergic beta-Agonists↗

PubMed1997

Was the formation of 1,25-dihydroxyvitamin D3 initially a catabolic pathway?

Following solar ultraviolet radiation, epidermal 7-dehydrocholesterol is converted to previtamin D3, which then undergoes a thermal isomerization into vitamin D3. The metabolism of vitamin D3, which is usually considered as an inactive compound, gives rise to the active hormone 1,25-dihydroxyvitamin D3, following two hydroxylation steps occurring in liver and kidney. Here, we propose that this anabolic pathway can also be interpreted as a catabolic one leading to the degradation of the photoproducts of 7-dehydrocholesterol, for which a specific biological role in the skin is proposed.

Calcitriol↗

PubMed1997

Typing Tuber ectomycorrhizae by polymerase chain amplification of the internal transcribed spacer of rDNA and the sequence characterized amplified region markers.

Identification of some economically important Tuber species using classical morphological characteristics is sometimes difficult. We report here the molecular characterization of a species coming from China, Tuber indicum, mistaken with Tuber melanosporum species. Using restriction analysis of the amplified internal transcribed spacer (ITS) of rDNA, ITS sequence analysis, and sequence characterized amplified region markers, with DNA from fruit bodies or mycorrhizae, genetic variation was found between these two species, allowing to differentiate and characterize them.

Ascomycota↗

PubMed1997

[Genetic alterations associated with pathologic differentiation of Wilms' tumors].

Although several cytogenetic alterations have been associated with development of Wilms' tumor, a multigenic neoplasia, molecular mechanisms of its induction, development and maintenance remain to be elucided. In order to characterize these different steps we have developed a unique animal model of Wilms' tumor constituted by the MAV-1(N) induced avian nephroblastoma. This animal model led to the discovery in our laboratory of a new gene now (nephroblastoma overexpressed gene) which is overexpressed in all avian nephroblastoma. Expression of the human nov gene (novH), which is down-regulated by WT1, is also deregulated in Wilms' tumors. Nov characteristics suggest that it would play a role in the control of cellular proliferation and differentiation. Our observations also indicate that nov could be involved in the development of Wilms' tumors, and represent a marker of their differentiation state.

Animals↗

PubMed1996

1,25-Dihydroxyvitamin D3 induces programmed cell death in a rat glioma cell line.

1,25-Dihydroxyvitamin D3 (1,25(OH)2D3), a seco-steroid hormone with potential antitumoral activities, has been recently reported to exert cytotoxic effects on C6 glioma cells. However, the molecular mechanisms which trigger this cell death remain unknown. We show here that this 1,25(OH)2D3-induced cell death is dependent upon protein synthesis and is accompanied by the expression of c-myc, p53, and gadd45 genes. Two other genes, coding for interleukin-6 and vaso-endothelial growth factor, are also upregulated after addition of 1,25(OH)2D3. This programmed cell death can be suppressed when cells are treated with forskolin, a drug which increases intracellular cAMP concentration, or with genistein, an inhibitor of tyrosine protein kinases. However, in spite of the demonstration of fragmented DNA in 1,25(OH)2D3-treated cells, the C6.9 cells used in this study do not show the classical morphological features of apoptosis. These results provide the first evidence for the existence of a programmed cell death triggered by 1,25(OH)2D3 in glioma cells and may provide a basis for the development of new therapeutic strategies. In addition, these data also suggest that the treatment of C6.9 cells with 1,25(OH)2D3 may be a useful model to study the molecular mechanisms involved in the programmed cell death of a cell of glial origin.

Animals↗

PubMed1996

Regulation of nov by WT1: a potential role for nov in nephrogenesis.

The nov gene encodes a putative Insulin-like-Growth Factor-Binding-Protein (IGFBP) of a novel type which is structurally related to a family of growth-factors likely to play a role in the control of cell proliferation. In the kidney, nov is expressed essentially at the embryonic stage and alterations of nov expression, relative to the normal kidney, have been detected in both avian nephroblastomas and human Wilms' tumors. The levels of human nov (novH) and WT1 mRNA in individual Wilms' tumors have been shown to be inversely correlated, suggesting that the expression of novH could be under the negative control of WT1. We have now established the nucleotide sequence of the 5' flanking region and identified two transcription start sites by RNase protection assays and primer extension. We report that in transient cotransfection experiments the transcription activity of novH promoter constructs was repressed by two isoforms of WT1 proteins (WT1 and WT1+KTS). Repression of the novH promoter required both intact zinc finger regions and the NH2 transcription repression domain of WT1. Inasmuch as the minimal region of novH promoter required to mediate WT1 repression in vivo failed to bine recombinant WT1 protein in in vitro footprinting assays this repression may be mediated by either (i) low affinity sites cooperative interactions or (ii) indirectly via protein-protein interactions with another factor(s). Furthermore, constitutive expression of wild type WT1 into 293 cells resulted in a decrease of endogenous NOVH protein levels, suggesting that novH may be a physiological target for WT1. The downregulation of novH expression by WT1 might represent a key element in normal and tumoral nephrogenesis.

3T3 Cells↗