ABSTRACT
Epidemiological studies suggest that low dietary folate increases risk for intestinal neoplasia. We recently developed a unique tumor model in which mice fed low dietary folate developed DNA damage and intestinal tumors. To identify additional mechanisms by which reduced folate intake contributes to tumor formation in this model, we characterized gene expression signatures in tumors. A total of 175 probe sets had altered expression, with the majority (173) showing increased expression compared with normal intestine. Functional categorization revealed that most genes were involved in cancer (51 genes) or cell death (37 genes); 31 genes appeared in both categories. Because apoptosis resistance is a hallmark of neoplasia, we assessed apoptosis in normal intestine of mice fed control (CD) and low-folate diets (FD); apoptosis was reduced in FD normal intestine compared with CD intestine by active caspase-3 expression (P = 0.027) and caspase-3/7 activities (P = 0.059). We selected candidate genes with antiapoptotic properties that had increased expression in tumor microarrays, CD44, and gelsolin and confirmed these increases at the mRNA and protein levels. CD44 and gelsolin protein also increased in mice fed the FD compared with the CD, normal intestine. Bcl-2-like 1:Bcl-2-antagonist/killer 1 mRNA ratios tended to be greater in FD than in CD normal intestine (P = 0.056). In conclusion, tumors induced by low dietary folate exhibited gene expression profiles that are characteristic of disrupted apoptosis. Folate depletion in normal intestine may trigger neoplasia through increased DNA damage and defective apoptosis; upregulation of CD44 and gelsolin, and the mitochondrial apoptotic pathway are implicated.
Subject(s)
Apoptosis , Folic Acid Deficiency/complications , Gene Expression Regulation, Neoplastic/drug effects , Intestinal Neoplasms/metabolism , Neoplasm Proteins/metabolism , Animals , Apoptosis/drug effects , Folic Acid/pharmacology , Gelsolin/genetics , Gelsolin/metabolism , Gene Expression Profiling , Hyaluronan Receptors/genetics , Hyaluronan Receptors/metabolism , Intestinal Neoplasms/etiology , Intestinal Neoplasms/pathology , Mice , Mice, Inbred BALB C , Microarray Analysis , Mutation , Neoplasm Proteins/geneticsABSTRACT
The expression of nine genes was analyzed by real-time RT-PCR in the central nervous system in order to investigate the molecular pathogenesis of natural scrapie. An up-regulation of genes related to glial activation (GFAP) and apoptosis (CASP3) was detected in obex and cerebrum, indicating a reactive glia. Another glial activation-related gene (CTSS) was slightly up-regulated in obex, whereas constitutive expression was detected for SOD1, YWHAZ, PRNP, and the apoptosis-related genes BCL2, MCL1, and BAX. This differential gene expression might reflect a spatial-temporal and tissue-specific molecular pathogenesis of scrapie.
Subject(s)
Central Nervous System/metabolism , Gene Expression Regulation/physiology , Gene Expression/physiology , Scrapie/metabolism , Amyloid/genetics , Amyloid/metabolism , Animals , Caspase 3 , Caspases/genetics , Caspases/metabolism , Central Nervous System/pathology , Fibroblast Growth Factor 9/genetics , Fibroblast Growth Factor 9/metabolism , Glial Fibrillary Acidic Protein/genetics , Glial Fibrillary Acidic Protein/metabolism , Molecular Chaperones/genetics , Molecular Chaperones/metabolism , Prions , Protein Precursors/genetics , Protein Precursors/metabolism , RNA, Messenger/metabolism , Reverse Transcriptase Polymerase Chain Reaction/methods , Sheep , Statistics, Nonparametric , Superoxide Dismutase/genetics , Superoxide Dismutase/metabolism , Superoxide Dismutase-1ABSTRACT
BACKGROUND: Cellular prion protein expression is essential for the development of transmissible spongiform encephalopathies (TSEs), and in sheep, genetic susceptibility to scrapie has been associated to PrP gene polymorphisms. To test the hypothetical linkage between PrP gene expression and genetic susceptibility, PrP mRNA levels were measured by real-time RT-PCR in six ovine tissues of animals with different genotypes. RESULTS: Previous to the PrP gene expression analysis the stability of several housekeeping (HK) genes was assessed in order to select the best ones for relative quantification. The normalisation of gene expression was carried out using a minimum of three HK genes in order to detect small expression differences more accurately than using a single control gene. The expression stability analysis of six HK genes showed a large tissue-associated variation reflecting the existence of tissue-specific factors. Thereby, a specific set of HK genes was required for an accurate normalisation of the PrP gene expression within each tissue. Statistical differences in the normalised PrP mRNA levels were found among the tissues, obtaining the highest expression level in obex, followed by ileum, lymph node, spleen, cerebellum and cerebrum. A tendency towards increased PrP mRNA levels and genetic susceptibility was observed in central nervous system. However, the results did not support the hypothesis that PrP mRNA levels vary between genotypes. CONCLUSION: The results on PrP gene expression presented here provide valuable baseline data for future studies on scrapie pathogenesis. On the other hand, the results on stability data of several HK genes reported in this study could prove very useful in other gene expression studies carried out in these relevant ovine tissues.
ABSTRACT
The expression of brain derived neurotrophic factor (BDNF) and its preferred receptor (TrkB) in rat retinal ganglion cells (RGCs) have been determined in the present study. To identify RGCs retrograde labelling was performed with fluorogold (FG). Subsequently, retinas were immunostained with antibodies to BDNF and TrkB. We found that all RGCs labelled with FG express both BDNF and its preferred receptor, TrkB. Moreover, displaced amacrine cells were also found to be immunolabelled by both antibodies. Thus BDNF/TrkB signalling in RGCs probably involves endogenous BDNF produced by the RGCs themselves.
