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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

Human HD iPSCs show dysregulation of HD markers. (A) HD 109 CAG and HD 71 CAG cell lines contained both normal and mutant huntingtin. Western blots demonstrate decreased ERK1/2 phosphorylation, increased SOD1 expression, and unchanged PRDX1 and β-catenin protein levels. Ctrl, control. (B) A twofold decrease in the phosphorylation of ERK1/2 in the juvenile HD 109 CAG line and a milder 1.4-fold decrease in the HD 71 CAG line were detected after quantification. (C) The levels of total and 33/37 phosphorylated β-catenin remained unchanged. (D) SOD1 expression was dysregulated in human HD iPSCs, showing an increase of 1.88- and 1.6-fold in the juvenile HD 109 CAG cells and HD 71 CAG lines, respectively (1-way ANOVA P=0.0182; P<0.05, Bonferroni post-test).
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DMM019406F6: Human HD iPSCs show dysregulation of HD markers. (A) HD 109 CAG and HD 71 CAG cell lines contained both normal and mutant huntingtin. Western blots demonstrate decreased ERK1/2 phosphorylation, increased SOD1 expression, and unchanged PRDX1 and β-catenin protein levels. Ctrl, control. (B) A twofold decrease in the phosphorylation of ERK1/2 in the juvenile HD 109 CAG line and a milder 1.4-fold decrease in the HD 71 CAG line were detected after quantification. (C) The levels of total and 33/37 phosphorylated β-catenin remained unchanged. (D) SOD1 expression was dysregulated in human HD iPSCs, showing an increase of 1.88- and 1.6-fold in the juvenile HD 109 CAG cells and HD 71 CAG lines, respectively (1-way ANOVA P=0.0182; P<0.05, Bonferroni post-test).

Mentions: We next tested whether the phenotypes observed in mouse iPSCs could also be detected in human HD patient-derived iPSCs. Several HD iPSC lines were acquired from the NINDS repository. Two lines contained 71 CAG repeats in HTT and originated from a 20-year-old HD patient, and another two lines contained 109 CAG repeats and originated from a 9-year-old juvenile HD patient. All HD lines contained both normal and mutant huntingtin, whereas the control lines contained only normal huntingtin (Fig. 6A). The total protein was isolated from human iPSCs, and the expression levels of phosphorylated and total ERK1/2, phosphorylated and total β-catenin, and SOD1 were examined by immunoblotting (Fig. 6A). In our studies (two lines of each genotype; three technical replicates), we observed a twofold decrease in the phosphorylation of ERK1/2 in the juvenile HD 109 CAG line and a milder 1.4-fold decrease in the HD 71 CAG line when compared with control lines (Fig. 6B). Notably, human pluripotent stem cells are dependent on an active ERK1/2 pathway and Essential 8 medium contains bFGF. Therefore, relatively high levels of ERK1/2 phosphorylation were observed in western blots without performing additional bFGF stimulation, which was inevitable for ERK assay in mouse iPSCs. Additionally, the SOD1 expression was dysregulated in human HD iPSCs, showing an increase of 1.88- and 1.6-fold in juvenile HD 109 CAG lines and HD 71 CAG lines, respectively (Fig. 6D). This increase in SOD1 expression reached statistical significance (1-way ANOVA, P=0.0182; P<0.05, Bonferroni post-test). The levels of PRDX1 protein (Fig. 6D) and β-catenin (Fig. 6C) remained unchanged. In conclusion, the protein expression patterns observed in human HD and juvenile-HD iPSCs were similar to the changes detected in YAC128 iPSCs.Fig. 6.


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)

Human HD iPSCs show dysregulation of HD markers. (A) HD 109 CAG and HD 71 CAG cell lines contained both normal and mutant huntingtin. Western blots demonstrate decreased ERK1/2 phosphorylation, increased SOD1 expression, and unchanged PRDX1 and β-catenin protein levels. Ctrl, control. (B) A twofold decrease in the phosphorylation of ERK1/2 in the juvenile HD 109 CAG line and a milder 1.4-fold decrease in the HD 71 CAG line were detected after quantification. (C) The levels of total and 33/37 phosphorylated β-catenin remained unchanged. (D) SOD1 expression was dysregulated in human HD iPSCs, showing an increase of 1.88- and 1.6-fold in the juvenile HD 109 CAG cells and HD 71 CAG lines, respectively (1-way ANOVA P=0.0182; P<0.05, Bonferroni post-test).
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DMM019406F6: Human HD iPSCs show dysregulation of HD markers. (A) HD 109 CAG and HD 71 CAG cell lines contained both normal and mutant huntingtin. Western blots demonstrate decreased ERK1/2 phosphorylation, increased SOD1 expression, and unchanged PRDX1 and β-catenin protein levels. Ctrl, control. (B) A twofold decrease in the phosphorylation of ERK1/2 in the juvenile HD 109 CAG line and a milder 1.4-fold decrease in the HD 71 CAG line were detected after quantification. (C) The levels of total and 33/37 phosphorylated β-catenin remained unchanged. (D) SOD1 expression was dysregulated in human HD iPSCs, showing an increase of 1.88- and 1.6-fold in the juvenile HD 109 CAG cells and HD 71 CAG lines, respectively (1-way ANOVA P=0.0182; P<0.05, Bonferroni post-test).
Mentions: We next tested whether the phenotypes observed in mouse iPSCs could also be detected in human HD patient-derived iPSCs. Several HD iPSC lines were acquired from the NINDS repository. Two lines contained 71 CAG repeats in HTT and originated from a 20-year-old HD patient, and another two lines contained 109 CAG repeats and originated from a 9-year-old juvenile HD patient. All HD lines contained both normal and mutant huntingtin, whereas the control lines contained only normal huntingtin (Fig. 6A). The total protein was isolated from human iPSCs, and the expression levels of phosphorylated and total ERK1/2, phosphorylated and total β-catenin, and SOD1 were examined by immunoblotting (Fig. 6A). In our studies (two lines of each genotype; three technical replicates), we observed a twofold decrease in the phosphorylation of ERK1/2 in the juvenile HD 109 CAG line and a milder 1.4-fold decrease in the HD 71 CAG line when compared with control lines (Fig. 6B). Notably, human pluripotent stem cells are dependent on an active ERK1/2 pathway and Essential 8 medium contains bFGF. Therefore, relatively high levels of ERK1/2 phosphorylation were observed in western blots without performing additional bFGF stimulation, which was inevitable for ERK assay in mouse iPSCs. Additionally, the SOD1 expression was dysregulated in human HD iPSCs, showing an increase of 1.88- and 1.6-fold in juvenile HD 109 CAG lines and HD 71 CAG lines, respectively (Fig. 6D). This increase in SOD1 expression reached statistical significance (1-way ANOVA, P=0.0182; P<0.05, Bonferroni post-test). The levels of PRDX1 protein (Fig. 6D) and β-catenin (Fig. 6C) remained unchanged. In conclusion, the protein expression patterns observed in human HD and juvenile-HD iPSCs were similar to the changes detected in YAC128 iPSCs.Fig. 6.

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