Xq27 FRAXA locus is a strong candidate for dyslexia: evidence from a genome-wide scan in French families.

Dyslexia is a frequent neurodevelopmental learning disorder. To date, nine susceptibility loci have been identified, one of them being DYX9, located in Xq27. We performed the first French SNP linkage study followed by candidate gene investigation in dyslexia by studying 12 multiplex families (58 subjects) with at least two children affected, according to categorical restrictive criteria for phenotype definition. Significant results emerged on Xq27.3 within DYX9. The maximum multipoint LOD score reached 3,884 between rs12558359 and rs454992. Within this region, seven candidate genes were investigated for mutations in exonic sequences (CXORF1, CXORF51, SLITRK2, FMR1, FMR2, ASFMR1, FMR1NB), all having a role during brain development. We further looked for 5'UTR trinucleotide repeats in FMR1 and FMR2 genes. No mutation or polymorphism co-segregating with dyslexia was found. This finding in French families with Dyslexia showed significant linkage on Xq27.3 enclosing FRAXA, and consequently confirmed the DYX9 region as a robust susceptibility locus. We reduced the previously described interval from 6.8 (DXS1227-DXS8091) to 4 Mb also disclosing a higher LOD score.

Genome-wide scan for genes involved in bipolar affective disorder in 70 European families ascertained through a bipolar type I early-onset proband: supportive evidence for linkage at 3p14.

Preliminary studies suggested that age at onset (AAO) may help to define homogeneous bipolar affective disorder (BPAD) subtypes. This candidate symptom approach might be useful to identify vulnerability genes. Thus, the probability of detecting major disease-causing genes might be increased by focusing on families with early-onset BPAD type I probands. This study was conducted as part of the European Collaborative Study of Early Onset BPAD (France, Germany, Ireland, Scotland, Switzerland, England, Slovenia). We performed a genome-wide search with 384 microsatellite markers using non-parametric linkage analysis in 87 sib-pairs ascertained through an early-onset BPAD type I proband (AAO of 21 years or below). Non-parametric multipoint analysis suggested eight regions of linkage with P-values<0.01 (2p21, 2q14.3, 3p14, 5q33, 7q36, 10q23, 16q23 and 20p12). The 3p14 region showed the most significant linkage (genome-wide P-value estimated over 10 000 simulated replicates of 0.015 [0.01-0.02]). After genome-wide search analysis, we performed additional linkage analyses with increased marker density using markers in four regions suggestive for linkage and having an information contents lower than 75% (3p14, 10q23, 16q23 and 20p12). For these regions, the information content improved by about 10%. In chromosome 3, the non-parametric linkage score increased from 3.51 to 3.83. This study is the first to use early-onset bipolar type I probands in an attempt to increase sample homogeneity. These preliminary findings require confirmation in independent panels of families.

Short-term treatment with mycophenolic acid and tacrolimus is tolerogenic for INS-1 cell clone transplantation and the deleterious effects of the drugs are limited: in vivo and in vitro studies.

One of the major requirements for a successful and life-lasting organ transplant is the access to safe, least toxic and permanent tolerance-inducing drugs. In this study we wished to evaluate the effects of tolerogenic doses of the immunosuppressive drugs mycophenolic acid (MPA) and tacrolimus (Tac) on clonal beta-cell lines, both in vivo and in vitro. Here we demonstrate that combined administration of low-dose MPA and Tac for 23 days induced permanent tolerance in an allogeneic beta-cell line transplant in Wistar rat liver through the portal vein. This short-term treatment of tolerogenic doses of the two drugs was deleterious to the survival of the transplanted cells but a small percentage of the cells could resist the effect and become fully active when the drugs were removed. The surviving cells, retrieved from growth in vivo, did not exhibit increased resistance in comparison to the original cells when tested in vitro at two glucose concentrations, 10 and 20 mM. The presence of a small percentage of resistant cells at the two glucose concentrations was also detected in the in vitro study after a continuous 8-day treatment demonstrating that the in vivo resistance was not related to micro-environmental protection but possibly to a phenotypic cell state that is yet to be determined.

Medicago truncatula plants overexpressing the early nodulin gene enod40 exhibit accelerated mycorrhizal colonization and enhanced formation of arbuscules.

The mutualistic symbiosis between flowering plants and arbuscular mycorrhizal fungi is extremely abundant in terrestrial ecosystems. In this symbiosis, obligately biotrophic fungi colonize the root of the host plants, which can benefit from these fungi by enhanced access to mineral nutrients in the soil, especially phosphorus. One of the main goals of research on this symbiosis is to find plant genes that control fungal development in the host plant. In this work, we show that mycorrhizal colonization is regulated by enod40, an early nodulin gene known to be involved in the nodule symbiosis of legumes with nitrogen-fixing bacteria. Medicago truncatula plants overexpressing enod40 exhibited stimulated mycorrhizal colonization in comparison with control plants. Overexpression of enod40 promoted fungal growth in the root cortex and increased the frequency of arbuscule formation. Transgenic lines with suppressed levels of enod40 transcripts, likely via a cosuppression phenomenon induced by the transgene, exhibited reduced mycorrhizal colonization. Hence, enod40 might be a plant regulatory gene involved in the control of the mycorrhizal symbiosis.

Translational and structural requirements of the early nodulin gene enod40, a short-open reading frame-containing RNA, for elicitation of a cell-specific growth response in the alfalfa root cortex.

A diversity of mRNAs containing only short open reading frames (sORF-RNAs; encoding less than 30 amino acids) have been shown to be induced in growth and differentiation processes. The early nodulin gene enod40, coding for a 0.7-kb sORF-RNA, is expressed in the nodule primordium developing in the root cortex of leguminous plants after infection by symbiotic bacteria. Ballistic microtargeting of this gene into Medicago roots induced division of cortical cells. Translation of two sORFs (I and II, 13 and 27 amino acids, respectively) present in the conserved 5' and 3' regions of enod40 was required for this biological activity. These sORFs may be translated in roots via a reinitiation mechanism. In vitro translation products starting from the ATG of sORF I were detectable by mutating enod40 to yield peptides larger than 38 amino acids. Deletion of a Medicago truncatula enod40 region between the sORFs, spanning a predicted RNA structure, did not affect their translation but resulted in significantly decreased biological activity. Our data reveal a complex regulation of enod40 action, pointing to a role of sORF-encoded peptides and structured RNA signals in developmental processes involving sORF-RNAs.

Reduction of noise-induced hearing loss using L-NAC and salicylate in the chinchilla.

The effects of a combination of two antioxidant compounds were studied in a chinchilla model of noise-induced hearing loss. After obtaining baseline hearing thresholds using inferior colliculus evoked potentials, chinchillas were exposed for 6 h to octave band noise centered at 4 kHz (105 dB SPL). Post-noise thresholds were obtained 1 h after the noise exposure, and then animals received either saline or salicylate and N-L-acetylcysteine combination. Another group received antioxidant treatment 1 h prior to noise. Hearing was tested at 1, 2 and 3 weeks post-noise. Subsequently, the cochleae were harvested, and cytocochleograms were prepared. There was a 20-40 dB SPL threshold shift at 3 weeks for tested controls. Permanent threshold shifts (PTS) were significantly reduced (P<0.05) to approximately 10 dB for the pre-treatment group at week 3. The PTS for the post-treatment group at week 3 was similar to the pre-treatment group at 1 and 2 kHz (0-10 dB) but was intermediate between the control and pre-treatment groups at 4 and 8 kHz (23 dB). Animals pre-treated with antioxidant had a significant reduction in hair cell loss but those post-treated with antioxidant had no protection from hair cell loss. These findings demonstrate the feasibility of reduction of noise-induced hearing loss using clinically available antioxidant compounds.

Temporal and spatial order of events during the induction of cortical cell divisions in white clover by Rhizobium leguminosarum bv. trifolii inoculation or localized cytokinin addition.

We examined the timing and location of several early root responses to Rhizobium leguminosarum bv. trifolii infection, compared with a localized addition of cytokinin in white clover, to study the role of cytokinin in early signaling during nodule initiation. Induction of ENOD40 expression by either rhizobia or cytokinin was similar in timing and location and occurred in nodule progenitor cells in the inner cortex. Inoculation of rhizobia in the mature root failed to induce ENOD40 expression and cortical cell divisions (ccd). Nitrate addition at levels repressing nodule formation inhibited ENOD40 induction by rhizobia but not by cytokinin. ENOD40 expression was not induced by auxin, an auxin transport inhibitor, or an ethylene precursor. In contrast to rhizobia, cytokinin addition was not sufficient to induce a modulation of the auxin flow, the induction of specific chalcone synthase genes, and the accumulation of fluorescent compounds associated with nodule initiation. However, cytokinin addition was sufficient for the localized induction of auxin-induced GH3 gene expression and the initiation of ccd. Our results suggest that rhizobia induce cytokinin-mediated events in parallel to changes in auxin-related responses during nodule initiation and support a role of ENOD40 in regulating ccd. We propose a model for the interactions of cytokinin with auxin, ENOD40, flavonoids, and nitrate during nodulation.

Analysis of Medicago truncatula nodule expressed sequence tags.

Systematic sequencing of expressed sequence tags (ESTs) can give a global picture of the assembly of genes involved in the development and function of organs. Indeterminate nodules representing different stages of the developmental program are especially suited to the study of organogenesis. With the vector lambdaHybriZAP, a cDNA library was constructed from emerging nodules of Medicago truncatula induced by Sinorhizobium meliloti. The 5' ends of 389 cDNA clones were sequenced, then these ESTs were analyzed both by sequence homology search and by studying their expression in roots and nodules. Two hundred fifty-six ESTs exhibited significant similarities to characterized data base entries and 40 of them represented 26 nodulin genes, while 133 had no similarity to sequences with known function. Only 60 out of the 389 cDNA clones corresponded to previously submitted M. truncatula EST sequences. For 117 cDNAs, reverse Northern (RNA) hybridization with root and nodule RNA probes revealed enhanced expression in the nodule, 48 clones are likely to code for novel nodulins, 33 cDNAs are clones of already known nodulin genes, and 36 clones exhibit similarity to other characterized genes. Thus, systematic analysis of the EST sequences and their expression patterns is a powerful way to identify nodule-specific and nodulation-related genes.

Alteration of enod40 expression modifies medicago truncatula root nodule development induced by sinorhizobium meliloti

Molecular mechanisms involved in the control of root nodule organogenesis in the plant host are poorly understood. One of the nodulin genes associated with the earliest phases of this developmental program is enod40. We show here that transgenic Medicago truncatula plants overexpressing enod40 exhibit accelerated nodulation induced by Sinorhizobium meliloti. This resulted from increased initiation of primordia, which was accompanied by a proliferation response of the region close to the root tip and enhanced root length. The root cortex of the enod40-transformed plants showed increased sensitivity to nodulation signals. T(1) and T(2) descendants of two transgenic lines with reduced amounts of enod40 transcripts (probably from cosuppression) formed only a few and modified nodulelike structures. Our results suggest that induction of enod40 is a limiting step in primordium formation, and its function is required for appropriate nodule development.

Giant juvenile xanthogranuloma of the tongue.

Juvenile xanthogranulomas (JXGs) are rare, benign, fibrohistiocytic lesions. They usually appear as one or more cutaneous papules on the head, neck, or trunk in infants. Twelve cases of oral JXGs have been reported, four of which involved the tongue. We present a 6-year-old girl with a large tongue mass diagnosed as JXG after an excisional biopsy. Histological and immunohistochemical staining results are presented. This is the first reported case of a giant oral JXG. A review of the literature on these unusual lesions is presented, along with discussion of their differential diagnosis and key aspects of the patient's evaluation, management, and pathological diagnosis.

Evidence for the presence of several phosphodiesterase isoforms in brown adipose tissue of Zucker rats: modulation of PDE2 by the fa gene expression.

The present study was undertaken to characterise the phosphodiesterases (PDEs) present in brown adipose tissue (BAT) of Zucker rat pups and to determine whether the capacity for degradation of cyclic nucleotides was affected by the fatty genotype. Regardless of the genotype, PDE2-4 contributed to total PDE activity, the PDE3 activity equalling the sum of PDE2 and 4 activities. In fa/fa compared to Fa/fa rats, (a) PDE2 activity was significantly increased, (b) Western blot analysis of PDE2 revealed two signals at 71 and 105 kDa, with changes in protein being in good parallelism with changes in activity, (c) the PDE2 mRNA concentration was also significantly increased. In good agreement, the cGMP concentration was decreased in BAT from fa/fa pups.

enod40 induces dedifferentiation and division of root cortical cells in legumes.

Under nitrogen-limiting conditions Rhizobium meliloti can establish symbiosis with Medicago plants to form nitrogen-fixing root nodules. Nodule organogenesis starts with the dedifferentiation and division of root cortical cells. In these cells the early nodulin gene enod40, which encodes an unusually small peptide (12 or 13 amino acids), is induced from the beginning of this process. Herein we show that enod40 expression evokes root nodule initiation. (i) Nitrogen-deprived transgenic Medicago truncatula plants overexpressing enod40 exhibit extensive cortical cell division in their roots in the absence of Rhizobium. (ii) Bombardment of Medicago roots with an enod40-expressing DNA cassette induces dedifferentiation and division of cortical cells and the expression of another early nodulin gene, Msenod12A. Moreover, transient expression of either the enod40 region spanning the oligopeptide sequence or only the downstream region without this sequence induces these responses. Our results suggest that the cell-specific growth response elicited by enod40 is involved in the initiation of root nodule organogenesis.

Down-regulation of beta3-adrenergic receptor expression in rat adipose tissue during the fasted/fed transition: evidence for a role of insulin.

The beta3-adrenergic receptor (beta3-AR) exerts a central role in the transduction of catecholamine effects in white and brown adipose tissue (WAT and BAT). A recent report has documented that insulin strongly down-regulates beta3-AR expression and catecholamine responsiveness in 3T3-F442A adipocytes [Fève, El Hadri, Quignard-Boulangé and Pairault (1994) Proc. Natl. Acad. Sci. U.S.A. 91, 5677-5681]. In the present report we show that the rise in plasma insulin levels elicited by the fasted/fed transition is associated with a reduction in beta3-AR mRNA levels and beta-adrenergic responsiveness in WAT and BAT. beta3-AR transcripts are also decreased in adipose tissue from animals subjected for 6 h to euglycaemic hyperinsulinaemic glucose clamps. Moreover, insulin acts directly on cultured rat white and brown adipocytes to decrease beta3-AR gene expression and adenylate cyclase activity in response to beta3-AR-selective agonists. These results suggest that there is a close relationship between food intake, plasma insulin levels and beta3-AR expression.

Early alterations in the brown adipose tissue adenylate cyclase system of pre-obese Zucker rat fa/fa pups: decreased G-proteins and beta 3-adrenoceptor activities.

This study was undertaken to determine whether receptor and non-receptor components of the adenylate cyclase (AC) cascade were altered in brown adipose tissue (BAT) of 14-day-old pre-obese (fa/fa) rats, before endocrine status is strongly modified by fa gene expression. Activity of the AC catalytic subunit did not differ between the two genotypes. In fa/fa rats compared with control Fa/fa rats, there was a 50% decrease in the activity of alpha Gs (stimulated by NaF or guanosine 5'-[gamma-thio]triphosphate) but no change in protein content (Western blotting). alpha Gi function, assessed by the inhibitory action of low concentrations of guanosine 5'-[beta gamma-imido]triphosphate upon 10(-4) M forskolin-stimulated AC activity, was equally low in both genotypes. Analysis of dose-response curves for different beta-agonists revealed that (i) both the basal and the maximally stimulated activity of AC were 2-fold lower in fa/fa rats than in Fa/fa rats; (ii) BRL37344 and CGP12177 (beta 3 agonists) were less potent in fa/fa than in Fa/fa rats (Kact. multiplied by 2); (iii) noradrenaline and isoprenaline (Iso), at the low-affinity site (beta 3-AR), were less potent in fa/fa than in Fa/fa pups (Kact. increased by 30 and 20% respectively). At the high-affinity site (mainly beta 1) these two agonists were more potent in fa/fa than in Fa/fa rats (Kact. decreased by 40 and 80% respectively). In good agreement with the latter result, the beta 1-adrenergic receptor (beta 1-AR)-selective antagonist CGP20712A had more effect on the Iso-stimulated AC activity in pre-obese than in lean pups (2-fold decreased in IC50). Binding experiments with [3H]CGP12177 show that in BAT of suckling rats, beta 3-ARs represent 80% of the total beta-ARs. Bmax values for the two sites were not affected by the genotype, although the beta 3-AR mRNA concentration in BAT (quantitative reverse-transcriptase PCR) was 3-fold lower in fa/fa rats than in Fa/fa pups. In conclusion, these results provide evidence for alterations in beta 1- and beta 3-AR signalling in BAT of 14-day-old suckling pre-obese Zucker rats with a decreased activity of alpha Gs. The impaired AC responsiveness to catecholamines might be a primary contributor to the development of this genetic obesity.

Effect of the beta-adrenoceptor agonist BRL-35135 on development of obesity in suckling Zucker (fa/fa) rats.

This study was undertaken to determine whether administration of a thermogenic beta-agonist drug to Zucker fatty rats could correct some of the earliest metabolic defects detectable in brown adipose tissue (BAT). Fa/fa and fa/fa littermates were given oral administration of BRL-35135 from 8 to 16 days of age. In fa/fa rats, the lipid content of white and brown adipose tissues was significantly reduced. In the BAT of fa/fa rats, thermogenic capacity was restored to the level observed in Fa/fa rats, whereas hyperactivity of fatty acid synthetase was abolished, and a deficit in lipoprotein lipase (activity and mRNA) was partly corrected. Hyperinsulinemia in fa/fa pups was significantly reduced. The decreased content of GLUT-4 mRNA that characterized BAT of fa/fa pups was also restored to normal. At variance with observations in preobese rats, BRL had very little or no effect on lean Fa/fa rats. The present study reveals that chronic administration of a beta-agonist drug early in life prevents emergence of most of the metabolic abnormalities that characterize fa/fa rats at the onset of obesity. This suggests that impaired sympathetic activity may play a role in the development of this genetic obesity.