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Huntington disease iPSCs show early molecular changes in intracellular signaling, the expression of oxidative stress proteins and the p53 pathway.

Szlachcic WJ, Switonski PM, Krzyzosiak WJ, Figlerowicz M, Figiel M - Dis Model Mech (2015)

Bottom Line: Several molecular and developmental effects of HD have been identified using neural stem cells (NSCs) and differentiated cells, such as neurons and astrocytes.Surprisingly, we found that a number of changes affecting cellular processes in HD were also present in undifferentiated pluripotent HD iPSCs, including the dysregulation of the MAPK and Wnt signaling pathways and the dysregulation of the expression of genes related to oxidative stress, such as Sod1.Interestingly, a common protein interactor of the huntingtin protein and the proteins in the above pathways is p53, and the expression of p53 was dysregulated in HD YAC128 iPSCs and human HD iPSCs.

View Article: PubMed Central - PubMed

Affiliation: Institute of Bioorganic Chemistry, Polish Academy of Sciences, Noskowskiego 12/14, Poznań 61-704, Poland.

No MeSH data available.


Related in: MedlinePlus

The expression of mutant huntingtin increased upon the differentiation of the YAC128 iPSCs. Protein lysates for western blotting analysis were prepared on days 4 and 8 of the formation of non-adherent embryoid bodies (EB4 and EB8 lanes, respectively), on days 7 and 14 of adherent differentiation (after 4 days of EB formation) in either FBS-containing medium (−) or N2B27 neuronal medium (+). The relative level of expression of mutant huntingtin increased several fold during adherent differentiation, whereas the increase in mouse wild-type (WT) huntingtin was not significant. The level of increase in mutant huntingtin was significant under neuronal-cell culture conditions. *P<0.01 in Bonferroni post-test.
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DMM019406F2: The expression of mutant huntingtin increased upon the differentiation of the YAC128 iPSCs. Protein lysates for western blotting analysis were prepared on days 4 and 8 of the formation of non-adherent embryoid bodies (EB4 and EB8 lanes, respectively), on days 7 and 14 of adherent differentiation (after 4 days of EB formation) in either FBS-containing medium (−) or N2B27 neuronal medium (+). The relative level of expression of mutant huntingtin increased several fold during adherent differentiation, whereas the increase in mouse wild-type (WT) huntingtin was not significant. The level of increase in mutant huntingtin was significant under neuronal-cell culture conditions. *P<0.01 in Bonferroni post-test.

Mentions: The YAC128 and YAC128/Oct-eGFP iPSC lines contained a human HTT transgene (supplementary material Fig. S4A) and expressed both mutant human huntingtin mRNA (supplementary material Fig. S4B) and protein (supplementary material Fig. S4C). In immunoblots, the mutant huntingtin protein was represented by bands of noticeably lower intensity compared with those of mouse huntingtin. Similarly, bands of mutant huntingtin of lower intensity than that of the bands of normal huntingtin were observed in samples of human 4281 HD fibroblasts. We investigated the levels of the expression of the mutant and normal huntingtin proteins upon iPSC differentiation. We found that, in cells that differentiated toward neurons, the level of mutant huntingtin was increased several fold, whereas the degree of increase in mouse WT huntingtin was not significant (Fig. 2; time of differentiation×allele interaction, P=0.0192; main effect of time, P=0.0002; two-way ANOVA; Bonferroni post-test, *P<0.01). Our results indicated that mutant huntingtin had accumulated in neuronal cells at an early stage of differentiation.Fig. 2.


Huntington disease iPSCs show early molecular changes in intracellular signaling, the expression of oxidative stress proteins and the p53 pathway.

Szlachcic WJ, Switonski PM, Krzyzosiak WJ, Figlerowicz M, Figiel M - Dis Model Mech (2015)

The expression of mutant huntingtin increased upon the differentiation of the YAC128 iPSCs. Protein lysates for western blotting analysis were prepared on days 4 and 8 of the formation of non-adherent embryoid bodies (EB4 and EB8 lanes, respectively), on days 7 and 14 of adherent differentiation (after 4 days of EB formation) in either FBS-containing medium (−) or N2B27 neuronal medium (+). The relative level of expression of mutant huntingtin increased several fold during adherent differentiation, whereas the increase in mouse wild-type (WT) huntingtin was not significant. The level of increase in mutant huntingtin was significant under neuronal-cell culture conditions. *P<0.01 in Bonferroni post-test.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4582098&req=5

DMM019406F2: The expression of mutant huntingtin increased upon the differentiation of the YAC128 iPSCs. Protein lysates for western blotting analysis were prepared on days 4 and 8 of the formation of non-adherent embryoid bodies (EB4 and EB8 lanes, respectively), on days 7 and 14 of adherent differentiation (after 4 days of EB formation) in either FBS-containing medium (−) or N2B27 neuronal medium (+). The relative level of expression of mutant huntingtin increased several fold during adherent differentiation, whereas the increase in mouse wild-type (WT) huntingtin was not significant. The level of increase in mutant huntingtin was significant under neuronal-cell culture conditions. *P<0.01 in Bonferroni post-test.
Mentions: The YAC128 and YAC128/Oct-eGFP iPSC lines contained a human HTT transgene (supplementary material Fig. S4A) and expressed both mutant human huntingtin mRNA (supplementary material Fig. S4B) and protein (supplementary material Fig. S4C). In immunoblots, the mutant huntingtin protein was represented by bands of noticeably lower intensity compared with those of mouse huntingtin. Similarly, bands of mutant huntingtin of lower intensity than that of the bands of normal huntingtin were observed in samples of human 4281 HD fibroblasts. We investigated the levels of the expression of the mutant and normal huntingtin proteins upon iPSC differentiation. We found that, in cells that differentiated toward neurons, the level of mutant huntingtin was increased several fold, whereas the degree of increase in mouse WT huntingtin was not significant (Fig. 2; time of differentiation×allele interaction, P=0.0192; main effect of time, P=0.0002; two-way ANOVA; Bonferroni post-test, *P<0.01). Our results indicated that mutant huntingtin had accumulated in neuronal cells at an early stage of differentiation.Fig. 2.

Bottom Line: Several molecular and developmental effects of HD have been identified using neural stem cells (NSCs) and differentiated cells, such as neurons and astrocytes.Surprisingly, we found that a number of changes affecting cellular processes in HD were also present in undifferentiated pluripotent HD iPSCs, including the dysregulation of the MAPK and Wnt signaling pathways and the dysregulation of the expression of genes related to oxidative stress, such as Sod1.Interestingly, a common protein interactor of the huntingtin protein and the proteins in the above pathways is p53, and the expression of p53 was dysregulated in HD YAC128 iPSCs and human HD iPSCs.

View Article: PubMed Central - PubMed

Affiliation: Institute of Bioorganic Chemistry, Polish Academy of Sciences, Noskowskiego 12/14, Poznań 61-704, Poland.

No MeSH data available.


Related in: MedlinePlus