Molecular understanding of a rat model with schizophrenia-related features. Gene-dosage imbalance of the gamma-secretase component Aph-1b in APO-SUS and -UNSUS rats

Contains fulltext : 30145.pdf (publisher's version ) (Open Access)Wistar rats pharmacogenetically selected for a high susceptibility to the dopamine agonist apomorphine (APO-SUS rats) display many differences with their phenotypic counterpart (APO-UNSUS rats), which are remarkably similar to those observed in schizophrenic patients. In this thesis we tried to understand the molecular mechanism underlying the APO-SUS/-UNSUS complex phenotype, which might aid in finding the cause of schizophrenia in human. Microarray expression analyses revealed in APO-SUS rats, relative to -UNSUS rats, a reduced expression of Aph-1b, a component of the gamma-secretase enzyme complex that is involved in multiple (neuro)developmental signalling pathways. These differences were caused by different gene copy numbers; APO-SUS rats harbour only one or two gene copies of Aph-1b, whereas APO-UNSUS rats contain three copies. This gene-dosage imbalance was caused by an unequal crossing-over event involving a segmental duplication within the Aph-1b locus. Although the expression levels of the other gamma-secretase components were not different, the gamma-secretase cleavage activity towards several substrates was affected and the Aph-1b genotypes segregated with a number of behavioural phenotypes. Through crossing and genetic re-selection, new rat lines were generated with one, two or three Aph-1b gene copies against otherwise similar genetic backgrounds. These rats revealed, next to affected Aph-1b expression levels throughout development, temporal, tissue- and substrate-specific changes in gamma-secretase cleavage activity. Preliminary studies on transgenic Xenopus laevis expressing Aph-1a under control of the proopiomelanocortin promoter, imply that the manipulation of Aph-1 expression in Xenopus melanotrope cells leads to a modified sensitivity of the dopaminergic system. In conclusion, these studies suggest that a subtle change in the expression of a (neuro)developmentally important protein may spatio-temporally affect diverse signalling pathways, ultimately resulting in a complex phenotype that is generally thought to be caused by multiple affected genes. Furthermore, we implicate the possible involvement of the gamma-secretase enzyme in complex disorders, like schizophrenia.RU Radboud Universiteit Nijmegen, 22 december 2006Promotores : Martens, G.J.M., Cools, A.R. Co-promotor : Ellenbroek, A.A.160 p

Reduced Aph-1b expression causes tissue- and substrate-specific changes in gamma-secretase activity in rats with a complex phenotype.

Contains fulltext : 32917.pdf (publisher's version ) (Open Access

Comparison of methyl-DNA immunoprecipitation (MeDIP) and methyl-CpG binding domain (MBD) protein capture for genome-wide DNA methylation analysis reveal CpG sequence coverage bias

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Dietary methyl donor deficiency during pregnancy in rats shapes learning and anxiety in offspring

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Dynamics of bivalent chromatin domains upon drug induced reactivation and resilencing in cancer cells.

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Culture of keratinocytes for transplantation without the need of feeder layer cells

Patients with large burn wounds have a limited amount of healthy donor skin. An alternative for the autologous skin graft is transplantation with autologous keratinocytes. Conventionally, the keratinocytes are cultured with mouse feeder layer cells in medium containing fetal calf serum (FCS) to obtain sufficient numbers of cells. These xenobiotic materials can be a potential risk for the patient. The aim of the present study was to investigate if keratinocytes could be expanded in culture without the need of a feeder layer and FCS. Keratinocytes were cultured on tissue culture plastic with or without collagen type IV coating in medium containing Ultroser G (serum substitute) and keratinocyte growth factor (KGF). An in vitro skin equivalent model was used to examine the capacity of these cells to form an epidermis. Keratinocytes in different passages (P2, P4, and P6) and freshly isolated cells were studied. Keratinocytes grown on collagen type IV were able to form an epidermis at higher passage numbers than cells grown in the absence of collagen type IV (P4 and P2, respectively). In both cases the reconstructed epidermis showed an increased expression of Ki-67, SKALP, involucrin, and keratin 17 compared to normal skin. Only 50,000 keratinocytes grown on collagen type IV in P4 were needed to form 1 cm2 epidermis, whereas 150,000 of freshly isolated keratinocytes were necessary. Using this culture technique sufficient numbers of keratinocytes, isolated from 1 cm2 skin, were obtained to cover 400 cm2 of wound surface in 2 weeks. The results show that keratinocytes can be cultured without the need of a fibroblast feeder layer and FCS and that these cells are still able to create a fully differentiated epidermis. This culture technique can be a valuable tool for the treatment of burn wounds and further development of tissue engineered skin

Insufficient slow-flow suppression mimicking aneurysm wall enhancement in magnetic resonance vessel wall imaging: a phantom study

OBJECTIVE: MR vessel wall imaging (VWI) is increasingly performed in clinical settings to support treatment decision-making regarding intracranial aneurysms. Aneurysm wall enhancement after contrast agent injection is expected to be related to aneurysm instability and rupture status. However, the authors hypothesize that slow-flow artifacts mimic aneurysm wall enhancement. Therefore, in this phantom study they assess the effect of slow flow on wall-like enhancement by using different MR VWI techniques. METHODS: The authors developed an MR-compatible aneurysm phantom model, which was connected to a pump to enable pulsatile inflow conditions. For VWI, 3D turbo spin echo sequences-both with and without motion-sensitized driven equilibrium (MSDE) and delay alternating with nutation for tailored excitation (DANTE) preparation pulses-were performed using a 3-T MR scanner. VWI was acquired both before and after Gd contrast agent administration by using two different pulsatile inflow conditions (2.5 ml/sec peak flow at 77 and 48 beats per minute). The intraluminal signal intensity along the aneurysm wall was analyzed to assess the performance of slow-flow suppression. RESULTS: The authors observed wall-like enhancement after contrast agent injection, especially in low pump rate settings. Preparation pulses, in particular the DANTE technique, improved the performance of slow-flow suppression. CONCLUSIONS: Near-wall slow flow mimics wall enhancement in VWI protocols. Therefore, VWI should be carefully interpreted. Preparation pulses improve slow-flow suppression, and therefore the authors encourage further development and clinical implementation of these techniques

Noradrenergic activation of the basolateral amygdala maintains hippocampus-dependent accuracy of remote memory

Item does not contain fulltextEmotional enhancement of memory by noradrenergic mechanisms is well-described, but the long-term consequences of such enhancement are poorly understood. Over time, memory traces are thought to undergo a neural reorganization, that is, a systems consolidation, during which they are, at least partly, transferred from the hippocampus to neocortical networks. This transfer is accompanied by a decrease in episodic detailedness. Here we investigated whether norepinephrine (NE) administration into the basolateral amygdala after training on an inhibitory avoidance discrimination task, comprising two distinct training contexts, alters systems consolidation dynamics to maintain episodic-like accuracy and hippocampus dependency of remote memory. At a 2-d retention test, both saline- and NE-treated rats accurately discriminated the training context in which they had received footshock. Hippocampal inactivation with muscimol before retention testing disrupted discrimination of the shock context in both treatment groups. At 28 d, saline-treated rats showed hippocampus-independent retrieval and lack of discrimination. In contrast, NE-treated rats continued to display accurate memory of the shock-context association. Hippocampal inactivation at this remote retention test blocked episodic-like accuracy and induced a general memory impairment. These findings suggest that the NE treatment altered systems consolidation dynamics by maintaining hippocampal involvement in the memory. This shift in systems consolidation was paralleled by time-regulated DNA methylation and transcriptional changes of memory-related genes, namely Reln and Pkmzeta, in the hippocampus and neocortex. The findings provide evidence suggesting that consolidation of emotional memories by noradrenergic mechanisms alters systems consolidation dynamics and, as a consequence, influences the maintenance of long-term episodic-like accuracy of memory

Differential contribution of the three Aph1 proteins to g-secretase activity in vivo

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