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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 microarray analysis revealed gene expression changes in SRS clones, predicting a decrease in cell proliferation and a delay in mitosis.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 microarray analysis revealed gene expression changes in SRS clones, predicting a decrease in cell proliferation and a delay in mitosis.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