Methylation and Expression of Mutant FUS in Motor Neurons Differentiated From Induced Pluripotent Stem Cells From ALS Patients.

Amyotrophic lateral sclerosis (ALS) is a rapidly progressive disease leading to degeneration of motor neurons (MNs). Epigenetic modification of gene expression is increasingly recognized as potential disease mechanism. In the present study we generated motor neurons from induced pluripotent stem cells from ALS patients carrying a mutation in the fused in sarcoma gene (FUS) and analyzed expression and promoter methylation of the FUS gene and expression of DNA methyltransferases (DNMTs) compared to healthy control cell lines. While mutant FUS neural progenitor cells (NPCs) did not show a difference in FUS and DNMT expression compared to healthy controls, differentiated mutant FUS motor neurons showed significantly lower FUS expression, higher DNMT expression and higher methylation of the proximal FUS gene promoter. Immunofluorescence revealed perceived proximity of cytoplasmic FUS aggregates in ALS MNs together with 5-methylcytosin (5-mC). Targeting disturbed methylation in ALS may therefore restore transcriptional alterations and represent a novel therapeutic strategy.

Species-specific vesicular monoamine transporter 2 (VMAT2) expression in mammalian pancreatic beta cells: implications for optimising radioligand-based human beta cell mass (BCM) imaging in animal models.

AIMS/HYPOTHESIS: Imaging of beta cell mass (BCM) is a major challenge in diabetes research. The vesicular monoamine transporter 2 (VMAT2) is abundantly expressed in human beta cells. Radiolabelled analogues of tetrabenazine (TBZ; a low-molecular-weight, cell-permeant VMAT2-selective ligand) have been employed for pancreatic islet imaging in humans. Since reports on TBZ-based VMAT2 imaging in rodent pancreas have been fraught with confusion, we compared VMAT2 gene expression patterns in the mouse, rat, pig and human pancreas, to identify appropriate animal models with which to further validate and optimise TBZ imaging in humans. METHODS: We used a panel of highly sensitive VMAT2 antibodies developed against equivalently antigenic regions of the transporter from each species in combination with immunostaining for insulin and species-specific in situ hybridisation probes. Individual pancreatic islets were obtained by laser-capture microdissection and subjected to analysis of mRNA expression of VMAT2. RESULTS: The VMAT2 protein was not expressed in beta cells in the adult pancreas of common mouse or rat laboratory strains, in contrast to its expression in beta cells (but not other pancreatic endocrine cell types) in the pancreas of pigs and humans. VMAT2- and tyrosine hydroxylase co-positive (catecholaminergic) innervation was less abundant in humans than in rodents. VMAT2-positive mast cells were identified in the pancreas of all species. CONCLUSIONS/INTERPRETATION: Primates and pigs are suitable models for TBZ imaging of beta cells. Rodents, because of a complete lack of VMAT2 expression in the endocrine pancreas, are a 'null' model for assessing interference with BCM measurements by VMAT2-positive mast cells and sympathetic innervation in the pancreas.