Biomedical subjects

M Charlier

Publications and source records attributed to M Charlier.

At least 19 recordsLinked to original sources

PubMed2004

Interferon-tau upregulates prolactin receptor mRNA in the ovine endometrium during the peri-implantation period.

Our objective was to determine the effect of ovine interferon-tau (IFN-tau) on prolactin receptor (PRL-R) gene expression in the ovine endometrium. IFN-tau is an embryonic cytokine which, via its paracrine anti-luteolytic activity, plays a critical role in maternal recognition of pregnancy in ruminants. Using ribonuclease protection assay procedures, we compared endometrial PRL-R mRNA levels in ewes that were intrauterine injected with either 2 mg bovine serum albumin or 2 mg recombinant ovine IFN-tau on day 10 of the oestrous cycle (day 0 = day of oestrus). IFN treatment significantly increased the abundance of both the long and short forms of PRL-R mRNA in the ovine uterus, but had no effect on the long:short form ratio. In situ hybridization experiments revealed that the increase in abundance of PRL-R mRNA in the uterus was localized to the glandular compartment of the endometrium. In pregnant ewes, a similar increase in PRL-R mRNA abundance was found to occur in ovine endometrium on days 14-15 post conception. Collectively, these data provided strong evidence that IFN-tau modulates the level of lactogenic hormone receptor mRNA in the ovine uterus. Whether the effect of IFN-tau on PRL-R expression is mediated directly or influenced, at least in part, by progesterone remains to be elucidated.

Animals↗

PubMed2003

DNA radiolysis in DNA-protein complexes: a stochastic simulation of attack by hydroxyl radicals.

PURPOSE: To propose an improved version of RADACK, a stochastic simulation of radiolytic attack on DNA, that takes into account the reactivity of each amino acid of a specifically bound protein with hydroxyl radicals. To apply it to the natural lactose operator-repressor complex taking advantage of recently reported structures. To compare the obtained probabilities of DNA strand break induction with those calculated with the previous versions and with an experimental pattern of strand break probabilities. MATERIALS AND METHODS: Models of complexes close to the natural ones, derived from crystallography- and NMR-based structures recently available in the PDB databank, were used. The specific chemical reactivity of each amino acid was introduced in the new version of RADACK (the reactivity model). The probabilities of strand break induction by the irradiation of the complex were calculated with this new version as well as with previous ones. RESULTS: The patterns of probabilities of strand break induction calculated with the improved version of RADACK were partially different from those obtained with previous versions. The patterns obtained for both, using putative models of natural complexes, were consistent with the experimental results, but some discrepancies were suggestive of slight differences between these structures and the real natural system. The crystallographic structure agreed best with the experimental results. CONCLUSIONS: A new version of RADACK was validated that took into account the reactivity of atoms in both DNA and protein. The putative modelled structures of a natural lactose operator-repressor complex were discussed.

Amino Acid Sequence↗

PubMed2001

RADACK, a stochastic simulation of hydroxyl radical attack to DNA.

RADACK was conceived to simulate the radiation-induced attack to different DNA forms and complexes. It allows to separately calculate the probability of attack to each reactive atom of the sugar and of the base and takes into account the sequence-dependent structure of DNA as known from crystallographic or NMR studies or resulting from molecular modelling. The calculations are aimed to assess sequence-, structure- and ligand-dependent modulation of damages of sugar and bases, leading to single strand breaks (frank strand breaks, FSB) and alkali-labile base modifications (alkali-revealed breaks, ARB), respectively. The modelling procedure and the results of simulations for some representative structures (B, Z and quadruplex forms) are here described and discussed. The calculated relative probabilities of OH* radical attack to all reaction sites are compared to experimental FSB and ARB values. By a fitting procedure, the relative efficiencies of conversion of the C4' and C5'-centred radicals into FSB, epsilon (C4'): epsilon (C5'), and the relative efficiencies of base radicals- to- ARB conversion, epsilon(T) : epsilon(A) : epsilon(C) : epsilon(G), are then deduced for each DNA form. The ability of the model to account for the distribution of damages in DNA-ligand complexes is proven by its successful application to two DNA-protein systems : the lac repressor-lac operator complex and the nuclcosome core.

Bacterial Proteins↗

PubMed2001

Radiosensitivity of DNA in a specific protein-DNA complex: the lac repressor-lac operator complex.

PURPOSE: To calculate the probability of radiation-induced frank strand breakage (FSB) at each nucleotide in the Escherichia coli lac repressor-lac operator system using a simulation procedure. To compare calculated and experimental results. To asses the contribution of DNA conformational changes and of the masking by the protein to DNA protection by the repressor. MATERIALS AND METHODS: Two structures of the complex were extracted from the PDB databank: crystallography- and NMR-based structures. Calculations were made of the accessibility of the atoms mainly involved in strand breakage (H4' and H5') to O&Hdot; and of the FSB probabilities, along: (1) DNA in the complex; (2) DNA in the complex depleted of the repressor; and (3) a linear DNA having the same sequence. An 80bp fragment bearing the operator was irradiated alone or in presence of the repressor. The relative probabilities of FSB at each nucleotide were determined using sequencing gel electrophoresis. RESULTS: Calculations predict modulation of the accessibility of H4' and H5' atoms and of the probabilities of FSB along the DNA fragments of complexes. This is due to the protein-induced conformational change and to masking by bound protein. The best agreement with the experimental FSB was observed for calculations that use the crystallography-based structure. CONCLUSIONS: For specific DNA-protein complexes, our calculations can predict the protein radiolytic footprints on DNA. They show the significant contribution of the protein-induced DNA conformational change to DNA protection.

Bacterial Proteins↗

PubMed2001

Visuoperceptual impairment in MS patients: nature and possible neural origins.

Failures on visuoperceptual neuropsychological tasks (on neuropsychological tests of visuo-spatial perception or on tests concerning semantic properties of visual objects), may indicate focal deficits of visuoperceptual function, or could be the result of (an)other (peripheral) visual deficit(s), or be the effect of a more general cognitive decline. In multiple sclerosis (MS) patients exhibiting sufficient visual acuity and not showing severe cognitive deterioration, impairment on a comprehensive set of 31 visuoperceptual neuropsychological tasks was compared with spatial resolution deficits (SRD), temporal resolution deficits (TRD) for visual stimuli, abnormal pattern shift visual evoked potential (PSVEP) responses, and failing scores on neuropsychological tasks other than visuoperceptual tasks. Impairment on the visuoperceptual neuropsychological tasks was highly independent from the other abnormal visual and cognitive neurological impairments examined, suggesting that it mostly represented focal deficits. Only TRD in both eyes related to this impairment and this relationship was rather weak. Thus in some MS patients a slowed visual information processing may be one of the combined deficits underlying visuoperceptual neuropsychological task impairment. Given that SRD and TRD were not related to another stage of MS and reflect disturbances of a P (parvocellular channel and ventral stream projections) and M (magnocellular channel and dorsal stream projections) visual-system function respectively, demyelination of a certain M pathway may become a co-determinant of visuoperceptual neuropsychological task impairment more rapidly than damage to a certain P pathway.

Adult↗

PubMed2001

Radiation disrupts protein-DNA complexes through damage to the protein. The lac repressor-operator system.

Eon, S., Culard, F., Sy, D., Charlier, M. and Spotheim-Maurizot, M. Radiation Disrupts Protein-DNA Complexes through Damage to the Protein. The lac Repressor-Operator System. Radiat. Res. 156, 110-117 (2001). Binding of a protein to its cognate DNA sequence is a key step in the regulation of gene expression. If radiation damage interferes with protein-DNA recognition, the entire regulation process may be perturbed. We have studied the effect of gamma rays on a model regulatory system, the E. coli lactose repressor-operator complex. We have observed the disruption of the complex upon irradiation in aerated solution. The complex is completely restored by the addition of nonirradiated repressor, but not by the addition of nonirradiated DNA. Thus radiation disrupts the DNA-protein complex by affecting the binding ability of the protein. This interpretation is supported by the dramatic loss of binding ability of a free irradiated repressor toward nonirradiated DNA. Interestingly, the dose necessary for the disruption of the irradiated complex is higher than that for inducing the complete loss of the binding ability of the free irradiated repressor. This may be due to the protection of key amino acids by the bound DNA. As seen from calculations of the accessibility of amino acids to radiolytic OH(.), the protection is due to both masking and conformational effects.

Amino Acids↗

PubMed2001

Enhanced strand break induction of DNA by resonant metal-innershell photoabsorption.

We determined the number of single and double strand breaks (ssb and dsb) in a DNA-chloroterpyridine platinum complex induced by resonant photoabsorption in the L(III) innershell of a platinum atom. The number of ssb and dsb were measured in supercoiled plasmids (AG30) versus the chloroterpyridine platinum concentration, i.e., the ratio of intercalated molecules to the number of phosphate sites in DNA. A significant increase in the number of ssb and dsb was observed when the DNA contained intercalated molecules. This technique is an efficient way to induce ssb and dsb triggered by the atomic Auger effect.

Algorithms↗

PubMed2000

Increase in prolactin receptor (PRL-R) mRNA level in the mammary gland after hormonal induction of lactation in virgin ewes.

In order to examine the hormonal regulation of the prolactin-receptor (PRL-R) gene expression during mammary gland development, ewes were treated to induce lactation via an estrogen-progesterone-hydrocortisone and ovine growth hormone treatment. In situ hybridization analysis was used and revealed that sex steroids increased PRL-R mRNA levels in the mammary gland. Using RNase protection assay we showed that the estradiol + progesterone treatment increased both the levels of the long and the short forms of PRL-R mRNA. Addition of hydrocortisone increased the level of alphaS1-casein transcripts and the level of the ratio of the long to the short form of the PRL-R mRNA. This ratio can be further enhanced by addition of ovine growth hormone to the latter treatment. This suggests a role of hydrocortisone and ovine growth hormone in the alternative splicing that leads to the preferential expression of the long form of the PRL-R mRNA. In conclusion, the present experiments suggest that estrogen, progesterone and hydrocortisone are the major regulators of the PRL-R gene expression during pregnancy and prepare the mammary gland for its differentiation.

Alternative Splicing↗

PubMed2000

Effect of ethidium bromide intercalation on DNA radiosensitivity.

PURPOSE: To assess the influence of the intercalating drug ethidium bromide (EtBr) on the yields of single strand breaks (ssb) induced by fast neutrons in supercoiled pBR322 plasmid and in a linear DNA restriction fragment. MATERIALS AND METHODS: The yield of ssb in the plasmid was measured by agarose gel electrophoresis. The proportion of fragments bearing one ssb and the probability of breakage at each nucleotide site was determined using sequencing gel electrophoresis. The volume variations due to the intercalation of EtBr were calculated. The expected radio-modifying effect at each nucleotide site of the linear fragment was evaluated using a reported simulation procedure. RESULTS: The ssb yield in the plasmid increased for concentrations up to 0.04 drug/bp and fell back in the range 0.04-0.1 drug/bp. For the linear DNA, only a slight protective effect was observed over the whole concentration range. The effect was almost the same at all nucleotide sites. CONCLUSION: For the linear DNA fragment, radioprotection was mainly due to scavenging of OH* radicals by the intercalated drug. For the plasmid, the radio-modifying effect results mainly from the variation of its effective volume, due to the modification of superhelicity.

DNA↗

PubMed2000

Radiolysis of nucleosome core DNA: a modelling approach.

PURPOSE: To calculate the expected pattern of frank strand breaks (FSB) induced in the DNA of a nucleosome core particle (NCP) by the attack of radiolytic OH* radicals and to compare this with the experimental pattern of FSB induced by the in vitro irradiation of chicken NCP. MATERIALS AND METHODS: The structure of NCP was obtained from the PDB databank. Using molecular modelling, the structure of a linear DNA fragment with the central 60 bp sequence of NCP-DNA was determined. The accessibility of the sugar attack sites to OH* radicals at each nucleotide of the linear fragment or of the NCP-DNA was calculated. The probability of reaction of OH* with each sugar reactive site was calculated using a Monte-Carlo method-based stochastic model. RESULTS: The accessibility of attack sites in the NCP-DNA and the calculated probabilities of sugar attack are mainly modulated through masking by histones, and only slightly through bending. The periodicity of the calculated FSB probabilities is identical to that of the experimental FSB probabilities in chicken NCP (period 10.4 +/- 0.1 bp). CONCLUSIONS: The good agreement between the calculated and experimental results recommends the presented modelling procedure as a tool in predicting the radiosensitivity of DNA in DNA-ligand complexes of known structure.

Animals↗

PubMed2000

Visuoperceptual impairment in multiple sclerosis patients diagnosed with neuropsychological tasks.

A comprehensive set of 31 binocular neuropsychological tasks assessing a series of spatial and non-spatial visuoperceptual abilities was used to study visuoperceptual impairment in a representative group of 49 MS-clinic patients exhibiting neither diagnosed ophthalmological afflictions nor major psychiatric diagnoses. Among these patients, true frequency rate of visuoperceptual impairment, i.e. of subjects failing four or more tasks, was estimated at 26%. The pattern of visuoperceptual impairment was non-uniform, non-selective, restricted and idiosyncratic. Only four tasks yielded significant rates of impairment. They concerned colour discrimination, the perception of the Müller-Lyer illusion and object recognition in two separate conditions. Each of the four factors identified by factor analysis had an important representative (with factor loading >0.35) among these four tasks. Failures on these tasks correlated poorly. Together, the four tasks satisfactorily predicted visuoperceptual impairment as defined by the comprehensive set of tasks (sensitivity 86.7%; specificity 81.3%), but with regard to an uncontaminated criterion, their aggregate sensitivity and specificity was only 75 and 56% respectively. Visuoperceptual neuropsychological task performance related significantly but weakly to cognitive status, physical disability and to pyramidal, cerebellar and brain stem neurological signs, and did not correlate with other clinical neurological signs, disease duration, type of MS, a history of optic neuritis, depression or medication status Multiple Sclerosis (2000) 6 241 - 254

Adult↗

PubMed2000

A questionnaire assessment of unmet needs for rehabilitation services and resources for people with multiple sclerosis: results of a pilot survey in five European countries. Needs Task group of MARCH (Multiple Sclerosis and Rehabilitation, Care and Health Services Research in Europe).

OBJECTIVE: To develop an international services and needs assessment instrument (SUN) for people with multiple sclerosis and their carers and to pilot this in different countries of the European Community. DESIGN: Interview study of people with multiple sclerosis, their carers and nominated key professionals examining the unmet needs of patients and carers. SETTING: Belgium, Estonia, Greece, Italy and the United Kingdom. MAIN OUTCOME MEASURES: Needs assessment questionnaire. RESULTS: The study comprised 137 people with multiple sclerosis, 125 carers and 111 professionals. Patients reported on average 2.9 unmet needs for themselves; their carers and professionals reported on average 2.4. Needs were categorized into seven broad categories. Due to difficulties experienced by the local researchers in distinguishing between needs and objectives a large proportion of needs had to be assigned to the 'other' category. CONCLUSIONS: The SUN is a valuable and practicable tool for the identification of unmet needs for people with multiple sclerosis and their carers. Formal validation and reliability testing of the different language versions is recommended.

Europe↗

PubMed2000

Strand break induction by photoabsorption in DNA-bound molecules.

Dried samples of a DNA-chloroterpyridine platinum complex were irradiated with monochromatic X rays tuned to the photoabsorption resonance of the L(III) inner shell of the platinum atom. The number of single- and double-strand breaks (SSBs and DSBs) triggered by the Auger effect in supercoiled DNA plasmids was measured by the production of circular nicked and linear forms. To probe the specific contribution of the L(III) inner-shell excitation of the platinum atom, photon wavelengths were tuned on the resonance energy (on peak) and below (off peak). The quantum yields of the resonance radiation were typically found to be 11 for the SSBs and 1 for the DSBs. The DSB-to-SSB ratio increased by 20% when switching from off-resonance to on-resonance irradiation.

DNA↗

PubMed1999

Cellular localization and evolution of prolactin receptor mRNA in ovine endometrium during pregnancy.

In this study, we have investigated the expression of the prolactin receptor gene in ovine endometrium during oestrus cycle and pregnancy. Using reverse transcription-PCR analysis, we provided evidence that the prolactin receptor gene is specifically transcribed in this tissue. As shown by Northern blot analysis, the level of the prolactin receptor transcripts increased dramatically during late pregnancy. In situ hybridization experiments revealed that prolactin receptor mRNA was specifically expressed in the glandular compartment and confirmed the dramatic increase of its expression that occurs at the end of pregnancy. Taken together, these findings are consistent with a putative role of prolactin and/or related molecules in the regulation of the proliferation of the glandular compartment and/or in the control of the secretory activity of the endometrium.

Animals↗

PubMed1999

Temporal resolution deficits in the visual fields of MS patients.

We assessed the relationship between temporal resolution and MS-induced neuropathy. A diagnostic strategy comprising assessments of temporal resolution at 16 points in the extra-foveal visual field up to 12 degrees from the fovea was first compared with foveal temporal resolution and with a standard VEP procedure in the same MS patients. At the group level, foveal temporal resolution was less sensitive to demyelination than the 16-point diagnostic strategy, the detection rate of which was comparable to that of the VEP procedure. Cross-sensitivity of the VEP and the 16-point diagnostic procedure was low. Subsequently, the average severity of MS-induced temporal resolution deficits was studied at three retinal loci of the same size but different eccentricities. Foveal deficits were not significantly greater than more peripheral deficits within the central 12 degrees.

Adult↗

PubMed1999

Radioprotection of DNA by spermine: a molecular modelling approach.

PURPOSE: To observe and explain the sequence-dependence of DNA radioprotection by spermine. MATERIALS AND METHODS: Sequencing gel electrophoresis was used to analyse the probability of frank strand break (FSB) induction at each nucleotide site. Molecular modelling of complexes of DNA with spermine molecules and of a curved electrically null DNA has been performed. RESULTS: The effect of spermine on radiation-induced strand breakage varied significantly along the studied fragment. At low spermine concentration, some sequences were protected while others were unprotected. Molecular modelling calculations show that the most electro-negative sites are located in the minor or in the major groove of DNA. The positively charged spermine (Z=+4) should preferentially bind to such sites. When bound in the minor groove, spermine triggers a reduction of the accessibility of radiolytic attack sites to OH* radicals. This is due to induced structural modifications and to the masking of attack sites. In the case of major groove binding, no reduction of accessibility occurs. This type of binding can explain the lack of protection of sequences with electro-negative sites in the major groove. At high spermine concentration, the fragment is strongly protected. A nucleosome-like pattern of breakage with periodically distributed regions of protection was observed. Molecular modelling calculations show that the accessibility of the attack sites in a curved electrically null DNA is also periodically reduced. CONCLUSIONS: Molecular modelling of DNA-spermine complexes that takes into account the electrostatic properties of DNA, allows an explanation of the experimentally observed effects of spermine on DNA radiosensitivity.

Base Sequence↗

PubMed1999

Monte Carlo simulation of radiolytic attack to 5'-d[T4G4]4 sequence in a unimolecular quadruplex.

PURPOSE: To extend to a quadruplex the stochastic model of radiolytic attack previously applied to a quasi-random duplex DNA. MATERIALS AND METHODS: The quadruplex structure is obtained from the PDB databank (first structure from 201D entry). The probabilities of OH* radical attack at all sugar and base reaction sites are calculated using a stochastic model based on the Monte Carlo method. RESULTS: Good agreement between the calculated and experimental frank strand break (FSB) probabilities is obtained using the relative efficiencies of conversion of the C4' and C5'-centred radicals into FSB determined for the quasi-random duplex (2.8:1 respectively). Efficiencies of base radicals-to-alkali-revealed breaks (ARB) conversion are determined by fitting the calculated probabilities of base attack to the previously reported experimental probabilities for ARB. The efficiency of conversion of thymine radical into an ARB is 3.4+/-0.7 times higher than for guanine radical. CONCLUSIONS: This paper supports the calculation method and allows evaluation of the relative efficiencies of thymine and guanine radicals-to-ARB conversion.

Base Sequence↗

PubMed1998

Radiosensitivity as support for the structure of a unimolecular DNA quadruplex.

PURPOSE: To search for possible variations of radiosensitivity along a unimolecular DNA quadruplex and to use them as support for choosing one of the two possible structures proposed by NMR studies. MATERIALS AND METHODS: The probability of radiation-induced frank strand breaks (FSB) and of alkali-revealed breaks (ARB) at each nucleotide site along the oligomer AATTCGTGGAGCACC (T4G4)4, was determined by sequencing gel electrophoresis as has been previously described in the literature. The oligomer was irradiated as a unimolecular quadruplex (QS) or paired to its complementary oligonucleotide in a double-stranded B-helix (DS). RESULTS: The oligomer globally presents the same radiosensitivity in the quadruplex or in the double-stranded helix. FSB occur with the same probability at each nucleotide site in both QS- and DS-DNA. More ARB are induced at T- and less at G-sites in QS-DNA. The probability of ARB induction in the second T-run is lower than in the first and the third one. CONCLUSION: Intrastrand radiosensitivity variations support the folding of the studied oligomer into a quadruplex with two lateral loops and one central loop of T-runs spanning the diagonal of the terminal G-quartet.

Circular Dichroism↗