DNA Repair Deficiency in Huntington\u27s Disease Fibroblasts and Induced Pluripotent Stem Cells

Mutant huntingtin protein (mhtt)– the protein responsible for cellular dysfunction in Huntington’s disease (HD) –is a product of an expanded trinucleotide repeat (TNR) cytosine-adenine-guanine (CAG) sequence in exon 1 of the huntingtin (HTT) gene. The pathology of HD has been extensively researched; however, the mechanism by which the disease-causing TNR expansions occur in somatic cells remains elusive. Interestingly, HD has often been referred to a ‘DNA repair disease’, even though DNA repair dysfunction in situ has not been identified. We hypothesized that presence of the mhtt protein affects the expression of DNA repair genes used to address DNA repair, ultimately affecting genome stability, thus providing a possible mechanism for TRN instability. Using quantitative polymerase chain reaction (qPCR) gene arrays for 84 DNA repair genes, we identified 18 DNA repair genes with decreased fold changes between 2- and 3- fold, as well as 11 genes down regulated greater than 3- fold in one HD fibroblast sample relative to a wild-type sample. To ensure our results were not limited to the samples tested, we then increased our number of HD samples and investigated gene expression of APEX1, BRCA1, RPA1, and RPA3 using sensitive TaqMan Gene Expression assays. Further, immunocytochemistry (ICC) analysis validated expression deficiencies at the protein level. These data identify down-regulated genes necessary to maintain stability in the genome of multiple HD affected fibroblast lines. Our data infers that the presence of the toxic mutant huntingtin (mHtt) protein is involved in the DNA repair gene inhibition. The mutant huntingtin protein (mHtt) produced in HD exhibits a partial gain-of-function in that the hydrophobic, expanded polyglutamine region at the N-termini aggregates to unintended targets, such as transcription factors and histone modifiers. To identify the broad pathway regulating gene expression down-regulation, we investigated epigenetic regulatory mechanisms. Rapid revival of selected DNA repair expression was observed in response to pharmacological hypomethylation treatment, but not to histone modification treatments. This identifies differential methylation patterns occur as a result of mHtt presence. Using capillary electrophoresis fragment analysis to characterize HTT TNR gene expansions, our data reveals that intermittent 5-azacytidine treatments induced HTT gene stability over 4 population doublings, elucidating methylation patterning involvement in TNR instability. Furthermore, induced pluripotent stem cells (iPSCs) undergo global epigenetic changes relative to its native cell type that include methylation patterning changes. Upon reprogramming of HD-affected fibroblasts into a pluripotent state we revealed that gene expression was recovered to wild-type levels and was maintained through 20 population doublings. As well, iPSC-HD lines show contraction-biased instability, opposite to expansion-biased instability in native fibroblast cell types. Differentiation of iPSC-HD lines into mesenchymal-like cells (MLCs) further revealed that APEX1 expression remained static, while others retreated to pre-iPSC expression levels. Interestingly, HD-iPSC derived MLCs showed that TNR regions maintained stability in the HTT gene pathogenic region, showing no changes in (CAG) repeats. These findings demonstrate that DNA repair gene expression in HD fibroblasts is altered, thus providing insight into the mechanism in which TNR instability persists, ultimately leading to genetic anticipation. This also identifies possible biomarkers that can be used to monitor disease progression and therapeutic treatment success. This study also provides evidence that TNR instability is pharmacologically alterable. This is the first evidence that a DNA repair gene deficiency is present in cells affected by Huntington’s disease. More so, our data suggests that mechanisms involved in pluripotency has a protective effect on the pathogenic TNR region of the HTT gene

Introduction to carbon dioxide sequestration–based cementitious construction materials

This chapter briefly reviews important issues that justify the importance of carbon dioxide sequestration. It includes carbon dioxide emissions and projections, global warming consequences namely extreme weather events, ocean acidification, sea level rise, economic losses, the increase of vector-borne illnesses, or even the revival of long dormant deadly bacteria and viruses. Comments about the Conference of parties are made with a special attention for the China situation. Comments are also made about the shortcomings of market-based instruments to try to reduce carbon dioxide emissions. A book outline is also included.(undefined)info:eu-repo/semantics/publishedVersio

The Impact of Auctions on Residential Sale Prices : Australian Evidence

This study re-examines the variation in selling prices between the auction and private treaty method of sales. Using sales data from five major Australian capital cities over a four year period, we estimate a hedonic pricing model. Results indicate that for house sales, auctions lead to greater selling prices across all cities examined. However, results for unit sales reveal that this auction premium is only evident in two cities where auctions are less prevalent. Further analysis reveals that self-selection (where a particular method of sale is selected to maximise the selling price) is evident across the sample. After controlling for this self-selection bias using a two-stage model, houses sold via auction generally command a higher price. This suggests that the auction method of selling provides a price premium over the private treaty method of sale