Sonic hedgehog (Shh) and its main receptor Patched (Ptc) are implicated in both neural development and tumorigenesis1, 2. Beside the classic morphogen activity of Shh, Shh is also a survival factor3, 4. Along this line, Ptc has been shown to function as a dependence receptor, inducing apoptosis in the absence of Shh, while its pro-apoptotic activity is blocked in Shh presence5. Here we show that, in the absence of its ligand, Ptc interacts with the adaptor protein DRAL/FHL2. DRAL/FHL2 is required for the pro-apoptotic activity of Ptc both in immortalized cells and during neural tube development in chick embryo. We demonstrate that, in the absence of Shh, Ptc recruits a protein complex that includes DRAL, the CARD containing domain proteins TUCAN or NALP1 and the apical caspase-9. Ptc triggers caspase-9 activation and enhances cell death via a caspase-9-dependent mechanism. Thus, we propose that, upon absence of its ligand Shh, the dependence receptor Ptc serves as the anchor for a caspase-activating complex that includes DRAL, a CARD domain containing protein and caspase-9.

Numerous studies have demonstrated oxidative damage in the central nervous system in subjects with Alzheimer disease and in animal models of this dementing disorder. In the current study, we show that transgenic mice modeling Alzheimer disease-PDAPP mice with Swedish and Indiana mutations in human amyloid precursor protein (APP)-develop oxidative damage in brain, including elevated levels of protein oxidation (indexed by protein carbonyls and 3-nitrotyrosine) and lipid peroxidation (indexed by protein-bound 4-hydroxy-2-nonenal). This oxidative damage requires the presence of a single methionine residue at position 35 of the amyloid β-peptide (Aβ), since all indices of oxidative damage in brain were completely prevented in genetically and age-matched PDAPP mice with a M631L mutation in APP. No significant differences in levels of APP, Aβ(1-42), Aβ (1-40), or the ratio Aβ(1-42)/Aβ(1-40) were found, suggesting that the loss of oxidative stress in vivo in brain of PDAPP(M631L) mice results solely from the mutation of the Met35 residue to Leu in the Aβ peptide. However, a marked reduction in Aβ-immunoreactive plaques was observed in the M631L mice, which instead displayed small punctate areas of non-plaque immunoreactivity and a microglial response. In contrast to the requirement for Met at residue 35 of the Aβ sequence (M631 of APP) for oxidative damage, indices of spatial learning and memory were not significantly improved by the M631L substitution. Furthermore, a genetically matched line with a different mutation-PDAPP(D664A)-showed the reverse: no reduction in oxidative damage but marked improvement in memory. This is the first in vivo study to demonstrate the requirement for Aβ residue Met35 for oxidative stress in brain of a mammalian model of Alzheimer disease. However, in this specific transgenic mouse model of AD, oxidative stress is neither required nor sufficient for memory abnormalities.

The β-amyloid precursor protein (APP) is an orphan transmembrane receptor whose physiological role is largely unknown. APP is cleaved by proteases generating amyloid-β (Aβ) peptide, the main component of the amyloid plaques that are associated with Alzheimer’s disease. Here, we show that APP binds netrin-1, a multifunctional guidance and trophic factor. Netrin-1 binding modulates APP signaling triggering APP intracellular domain (AICD)-dependent gene transcription. Furthermore, netrin-1 binding suppresses Aβ peptide production in brain slices from Alzheimer model transgenic mice. In this mouse model, decreased netrin-1 expression is associated with increased Aβ concentration, thus supporting netrin-1 as a key regulator of Aβ production. Finally, we show that netrin-1 brain administration in Alzheimer model transgenic mice may be associated with an amelioration of the Alzheimer’s phenotype.

In the adult mammalian brain, neural stem cells persist in the subventricular zone (SVZ) where dopamine D 3 receptors are expressed. Here, we demonstrate that addition of 1 lM apomorphine increases cell numbers in post-natal SVZ cell cultures. This effect was prevented by a co-treatment with haloperidol, sulpiride or U-99194A, a D 3 -preferring antagonist, and mimicked by the dopamine D 3 receptor selective agonist 7-hydroxy-dipropylaminotetralin (7-OH-DPAT). EC 50 values were 4.04 ± 1.54 nM for apomorphine and 0.63 ± 0.13 nM for 7-OH-DPAT, which fits the pharmacological profile of the D 3 receptor. D 3 receptors were detected in SVZ cells by RT-PCR and immunocytochemistry. D 3 receptors were expressed in numerous b-III tubulin immunopositive cells. The fraction of apoptotic nuclei remained unchanged following apomorphine treatment, thus ruling out any possible effect on cell survival. In contrast, proliferation was increased as both the proportion of nuclei incorporating bromo-deoxyuridine and the expression of the cell division marker cyclin D 1 were enhanced. These findings provide support for a regulatory role of dopamine over cellular dynamics in post-natal SVZ.

Background:The small stress heat shock protein 27 (Hsp27) has recently turned as a promising target for cancer treatment. Hsp27 upregulation is associated with tumour growth and resistance to chemo- and radio-therapeutic treatments, and several ongoing drugs inhibiting Hsp27 expression are under clinical trial. Hsp27 is now well described to counteract apoptosis and its elevated expression is associated with increased aggressiveness of several primary tumours. However, its role in the later stage of tumour progression and, more specifically, in the later and most deadly stage of tumour metastasis is still unclear.Methods/results:In the present study, we showed by qRT–PCR that Hsp27 gene is overexpressed in a large fraction of the metastatic breast cancer area in 53 patients. We further analysed the role of this protein in mice during bone metastasis invasion and establishment by using Hsp27 genetically depleted MDA-MB231/B02 human breast cancer cell line as a model. We demonstrate that Hsp27 silencing led to reduced cell migration and invasion in vitro and that in vivo it correlated with a decreased ability of breast cancer cells to metastasise and grow in the skeleton.Conclusion:Altogether, these data characterised Hsp27 as a potent therapeutic target in breast cancer bone metastasis and skeletal tumour growth.

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Objective-Selective neuronal vulnerability in neurodegenerative diseases is poorly understood. In Alzheimer's disease, the basal forebrain cholinergic neurons are selectively vulnerable, putatively because of their expression of the cell death mediator p75 NTR (the common neurotrophin receptor), and its interaction with proapoptotic ligands pro-nerve growth factor and amyloid-β peptide. However, the relation between amyloid precursor protein (APP) and p75 NTR has not been described previously.Methods-APP and p75 NTR were assayed for interaction by coimmunoprecipitation in vitro and in vivo, yeast two-hybrid assay, bioluminescence resonance energy transfer, and confocal microscopy. Effects on APP processing and signaling were studied using immunoblotting, enzyme-linked immunosorbent assays, and luciferase reporter assays.Results-The results of this study are as follows: (1) p75 NTR and APP interact directly; (2) this interaction is modified by ligands nerve growth factor and β-amyloid; (3) APP and p75 NTR colocalization in vivo is modified in Alzheimer's model transgenic mice; (4) APP processing is altered by p75 NTR , and to a lesser extent, p75 NTR processing is altered by the presence of APP; (5) APP-dependent transcription mediated by Fe65 is blocked by p75 NTR ; and (6) coexpression of APP and p75 NTR triggers cell death.Interpretation-These results provide new insight into the emerging signaling network that mediates the Alzheimer's phenotype and into the mechanism of basal forebrain cholinergic neuronal selective vulnerability. In addition, the results argue that the interaction between APP and p75 NTR may represent a therapeutic target in Alzheimer's disease.The common neurotrophin receptor, p75 NTR , has been implicated as a potential mediator of Alzheimer's disease pathogenesis in several different ways. First, p75 NTR expression is highly restricted in the adult nervous system, and its main site of expression, the basal forebrain cholinergic neurons, represents a selectively vulnerable region in Alzheimer's disease. [1][2][3] Second, p75 NTR mediates programmed cell death, 4-7 and the expression of p75 NTR sensitizes cells to β-amyloid (Aβ) toxicity. 8 Third, Aβ has been shown to interact directly with p75 NTR,9 and this interaction may lead to apoptosis induction. 9 These studies suggest that one mechanism by which p75 NTR may participate in Alzheimer's disease pathogenesis is by binding Aβ peptide and triggering programmed cell death. However, recent studies suggest that Aβ peptide also binds to its parent, APP (β-amyloid precursor protein), multimerizing APP, inducing caspase cleavage of APP intracytoplasmically at Asp664, and inducing programmed cell death. 14,15 Furthermore, the Aβ-induced cleavage of APP at Asp664 may play a key role in nonapoptotic features of the Alzheimer's phenotype, such as hippocampal synapse loss, dentate gyral atrophy, and reduction in excitatory postsynaptic potentials at CA1 after stimulation of the Schaeffer collaterals of CA3 neurons, because the addition of a...

The netrin-1/DCC ligand/receptor pair has key roles in central nervous system (CNS) development, mediating axonal, and neuronal navigation. Although expression of netrin-1 and DCC is maintained in the adult brain, little is known about their role in mature neurons. Notably, netrin-1 is highly expressed in the adult substantia nigra, leading us to investigate a role of the netrin-1/DCC pair in adult nigral neuron fate. Here, we show that silencing netrin-1 in the adult substantia nigra of mice induces DCC cleavage and a significant loss of dopamine neurons, resulting in motor deficits. Because loss of adult dopamine neurons and motor impairments are features of Parkinson's disease (PD), we studied the potential impact of netrin-1 in different animal models of PD. We demonstrate that both overexpression of netrin-1 and brain administration of recombinant netrin-1 are neuroprotective and neurorestorative in mouse and rat models of PD. Of interest, we observed that netrin-1 levels are significantly reduced in PD patient brain samples. These results highlight the key role of netrin-1 in adult dopamine neuron fate, and the therapeutic potential of targeting netrin-1 signaling in PD.