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Decreased expression of cell proliferation-related genes in clonally derived skin fibroblasts from children with Silver-Russell syndrome is independent of the degree of 11p15 ICR1 hypomethylation.

Heckmann D, Urban C, Weber K, Kannenberg K, Binder G - Clin Epigenetics (2015)

Bottom Line: Cell proliferation was diminished in SRShypo compared to Cnormo (p = 0.035).The analysis of severely ICR1 hypomethylated clonal fibroblasts did not reveal functional differences compared to normomethylated clones with respect to IGF2 and H19 expression.A difference compared to the clones from healthy individuals was present in the form of a lower proliferation rate, presumably due to impaired cell cycle progression.

View Article: PubMed Central - PubMed

Affiliation: Pediatric Endocrinology, University Children's Hospital Tuebingen, Hoppe-Seyler-Straße 1, 72076 Tuebingen, Germany.

ABSTRACT

Background: The in vitro analysis of the hypomethylation of imprinting control region 1 (ICR1) within the IGF2/H19 locus is challenged by the mosaic distribution of the epimutation in tissues from children with Silver-Russell syndrome (SRS). To exclude mosaicism, clonal cultures of skin fibroblasts from four children with SRS and three controls were analyzed. Cell proliferation, IGF-II secretion, and IGF2 and H19 expression were measured, and a microarray expression analysis was performed.

Results: Single-cell expansion established severely ICR1 hypomethylated clones (SRShypo) and normomethylated clones (SRSnormo) from the patients and controls (Cnormo). IGF2 expression was below the detection limit of the quantitative real-time PCR (qRT-PCR) assay, whereas H19 expression was detectable, without differences between fibroblast clones. Cell count-related IGF-II release was comparable in SRShypo and Cnormo supernatants. Cell proliferation was diminished in SRShypo compared to Cnormo (p = 0.035). The microarray analysis revealed gene expression changes in SRS clones, predicting a decrease in cell proliferation and a delay in mitosis.

Conclusions: The analysis of severely ICR1 hypomethylated clonal fibroblasts did not reveal functional differences compared to normomethylated clones with respect to IGF2 and H19 expression. A difference compared to the clones from healthy individuals was present in the form of a lower proliferation rate, presumably due to impaired cell cycle progression.

No MeSH data available.


Related in: MedlinePlus

IGF-II secretion by SRShypo und Cnormo clones. SRShypo clones and Cnormo clones were cultured for 14 days (d) in standard medium. Cell count-based IGF-II secretion into the supernatant was measured on days 1, 7, and 14 using IGF-II RIA.
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Fig3: IGF-II secretion by SRShypo und Cnormo clones. SRShypo clones and Cnormo clones were cultured for 14 days (d) in standard medium. Cell count-based IGF-II secretion into the supernatant was measured on days 1, 7, and 14 using IGF-II RIA.

Mentions: The cell number-normalized amount of the IGF-II protein measured in the supernatants after the incubation of 25,000 cells for 1, 7, and 14 days was not different in the SRShypo (S1sh_12, S5sh_11) and Cnormo clones (K3li_3, K3re_12) (Figure 3).Figure 3


Decreased expression of cell proliferation-related genes in clonally derived skin fibroblasts from children with Silver-Russell syndrome is independent of the degree of 11p15 ICR1 hypomethylation.

Heckmann D, Urban C, Weber K, Kannenberg K, Binder G - Clin Epigenetics (2015)

IGF-II secretion by SRShypo und Cnormo clones. SRShypo clones and Cnormo clones were cultured for 14 days (d) in standard medium. Cell count-based IGF-II secretion into the supernatant was measured on days 1, 7, and 14 using IGF-II RIA.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4318184&req=5

Fig3: IGF-II secretion by SRShypo und Cnormo clones. SRShypo clones and Cnormo clones were cultured for 14 days (d) in standard medium. Cell count-based IGF-II secretion into the supernatant was measured on days 1, 7, and 14 using IGF-II RIA.
Mentions: The cell number-normalized amount of the IGF-II protein measured in the supernatants after the incubation of 25,000 cells for 1, 7, and 14 days was not different in the SRShypo (S1sh_12, S5sh_11) and Cnormo clones (K3li_3, K3re_12) (Figure 3).Figure 3

Bottom Line: Cell proliferation was diminished in SRShypo compared to Cnormo (p = 0.035).The analysis of severely ICR1 hypomethylated clonal fibroblasts did not reveal functional differences compared to normomethylated clones with respect to IGF2 and H19 expression.A difference compared to the clones from healthy individuals was present in the form of a lower proliferation rate, presumably due to impaired cell cycle progression.

View Article: PubMed Central - PubMed

Affiliation: Pediatric Endocrinology, University Children's Hospital Tuebingen, Hoppe-Seyler-Straße 1, 72076 Tuebingen, Germany.

ABSTRACT

Background: The in vitro analysis of the hypomethylation of imprinting control region 1 (ICR1) within the IGF2/H19 locus is challenged by the mosaic distribution of the epimutation in tissues from children with Silver-Russell syndrome (SRS). To exclude mosaicism, clonal cultures of skin fibroblasts from four children with SRS and three controls were analyzed. Cell proliferation, IGF-II secretion, and IGF2 and H19 expression were measured, and a microarray expression analysis was performed.

Results: Single-cell expansion established severely ICR1 hypomethylated clones (SRShypo) and normomethylated clones (SRSnormo) from the patients and controls (Cnormo). IGF2 expression was below the detection limit of the quantitative real-time PCR (qRT-PCR) assay, whereas H19 expression was detectable, without differences between fibroblast clones. Cell count-related IGF-II release was comparable in SRShypo and Cnormo supernatants. Cell proliferation was diminished in SRShypo compared to Cnormo (p = 0.035). The microarray analysis revealed gene expression changes in SRS clones, predicting a decrease in cell proliferation and a delay in mitosis.

Conclusions: The analysis of severely ICR1 hypomethylated clonal fibroblasts did not reveal functional differences compared to normomethylated clones with respect to IGF2 and H19 expression. A difference compared to the clones from healthy individuals was present in the form of a lower proliferation rate, presumably due to impaired cell cycle progression.

No MeSH data available.


Related in: MedlinePlus