Genetic isolates in Corsica (France): linkage disequilibrium extension analysis on the Xq13 region

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Reversible Disruption of Pre-Pulse Inhibition in Hypomorphic-Inducible and Reversible CB1-/- Mice

Although several genes are implicated in the pathogenesis of schizophrenia, in animal models for such a severe mental illness only some aspects of the pathology can be represented (endophenotypes). Genetically modified mice are currently being used to obtain or characterize such endophenotypes. Since its cloning and characterization CB1 receptor has increasingly become of significant physiological, pharmacological and clinical interest. Recently, its involvement in schizophrenia has been reported. Among the different approaches employed, gene targeting permits to study the multiple roles of the endocannabinoid system using knockout (-/-) mice represent a powerful model but with some limitations due to compensation. To overcome such a limitation, we have generated an inducible and reversible tet-off dependent tissue-specific CB1-/- mice where the CB1R is re-expressed exclusively in the forebrain at a hypomorphic level due to a mutation (IRh-CB1-/-) only in absence of doxycycline (Dox). In such mice, under Dox+ or vehicle, as well as in wild-type (WT) and CB1-/-, two endophenotypes motor activity (increased in animal models of schizophrenia) and pre-pulse inhibition (PPI) of startle reflex (disrupted in schizophrenia) were analyzed. Both CB1-/- and IRh-CB1-/- showed increased motor activity when compared to WT animals. The PPI response, unaltered in WT and CB1-/- animals, was on the contrary highly and significantly disrupted only in Dox+ IRh-CB1-/- mice. Such a response was easily reverted after either withdrawal from Dox or haloperidol treatment. This is the first Inducible and Reversible CB1-/- mice model to be described in the literature. It is noteworthy that the PPI disruption is not present either in classical full CB1-/- mice or following acute administration of rimonabant. Such a hypomorphic model may provide a new tool for additional in vivo and in vitro studies of the physiological and pathological roles of cannabinoid system in schizophrenia and in other psychiatric disorders

The role of the –50 region of the human γ-globin gene in switching

During the switch from human γ- (fetal) to β- (adult) globin gene expression, the γ and β genes are expressed competitively by an alternating transcription mechanism. The –50 region of the γ gene promoter has been proposed to be responsible for the early competitive advantage of the γ genes and to act as a stage selector element (SSE) in hemoglobin switching. We analyzed the effect of mutating the –50 region of the γ gene in the presence of a competing β gene in transgenic mice. This shows that the –50 region does not affect silencing of the β gene in early development and does not act as a stage selector. However, it affects the ratio of γ versus β gene expression in the early, but not later, stages of fetal development. Interestingly, both the wild-type and mutant minilocus constructs show a higher frequency of alternate transcription than observed in the complete locus, suggesting that sequences normally present between the γ and β genes facilitate the interaction of the locus control region (LCR) and β-globin gene in the complete locus

Delta-globin gene expression improves sickle cell disease in a humanised mouse model

Sickle cell disease (SCD) is a widespread genetic disease associated with severe disability and multi-organ damage, resulting in a reduced life expectancy. None of the existing clinical treatments provide a solution for all patients. Gene therapy and fetal haemoglobin (HbF) reactivation through genetic approaches have obtained promising, but early, results in patients. Furthermore, the search for active molecules to increase HbF is still ongoing. The delta-globin gene produces the delta-globin of haemoglobin A2 (HbA2). Although expressed at a low level, HbA2 is fully functional and could be a valid anti-sickling agent in SCD. To evaluate the therapeutic potential of a strategy aimed to over-express the delta-globin gene in vivo, we crossed transgenic mice carrying a single copy of the delta-globin gene, genetically modified to be expressed at a higher level (activated), with a humanised mouse model of SCD. The activated delta-globin gene gives rise to a consistent production of HbA2, effectively improving the SCD phenotype. For the first time in vivo, these results demonstrate the therapeutic potential of delta-globin, which could lead to novel approaches to the cure of SCD

Startle response and percentage of PPI in IR-hCB1^-/-, CB1^-/- and WT mice: effect of Dox treatment.

<p>The startle amplitude (panel A) and % of PPI (panel B) are the mean±S.E.M. of WT (n = 13), CB1^-/- (n = 13), IR-hCB1^-/- (n = 15) and IR-hCB1^-/- Dox (n = 14) mice. For PPI the data are expressed as the average PPI response over the 3 prepulse intensities (see supplemental files for the figure illustrating PPI by prepulse intensity). ***P<0.0001 IR-hCB1^-/- Dox vs all other groups (Newman-Keuls Multiple Comparison test).</p

Real time-PCR CB1 mRNA expression in both WT and IR-hCB1^-/- mice either Dox treated or untreated.

<p>The mRNA expression detected by real time-PCR was measured in different brain areas from WT, untreated and Dox-treated IR-hCB1^-/- mice (n = 3 for each group). *P<0.001.</p

Percentage of PPI in IR-hCB1^-/- mice under different conditions.

<p>All values represent the mean±S.E.M. Panel A: reversibility of Dox-induced disruption of PPI. Two groups of IR-hCB1^-/- mice have been randomly assigned to vehicle group (dotted color, n = 6) or Dox group (plain color, n = 7) and then tested for PPI (Exp. 1). Dox administration lasted 12 days, and extra 12 days served for the washout, after which the schedule was repeated by switching the treatments (i.e. Dox given to previous No-Dox treated mice, and vice versa) (Exp. 2). Panel B: Dox-induced disruption of PPI is antagonized by haloperidol (1 mg/kg i.p., 30 min before PPI session).</p

Schematic representation of the construct for generating the IR-CB1-/-.

<p>Tetracycline-controlled transactivator (tTA) expression was driven by the CaMKII promoter. In its active form, tTA binds to the tetO operator sequence that controls two promoters in opposite direction and initiates expression of the CB1 and Lac Z transgenes. Expression can be inhibited by Dox as it binds to tTA, making it inactive.</p