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Dicer1 ablation in the mouse epididymis causes dedifferentiation of the epithelium and imbalance in sex steroid signaling.

Björkgren I, Saastamoinen L, Krutskikh A, Huhtaniemi I, Poutanen M, Sipilä P - PLoS ONE (2012)

Bottom Line: The postnatal development of the epididymis is a complex process that results in a highly differentiated epithelium, divided into several segments.The dedifferentiated epithelium also showed an increase in estrogen receptor 1 expression while the expression of androgen receptor and its target genes; glutathione peroxidase 5, lipocalin 5 and cysteine-rich secretory protein 1 was downregulated, indicating imbalanced sex steroid signaling.At the time of the final epididymal development, Dicer1 acts as a regulator of signaling pathways essential for maintaining epithelial cell differentiation.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Institute of Biomedicine, University of Turku, Turku, Finland.

ABSTRACT

Background: The postnatal development of the epididymis is a complex process that results in a highly differentiated epithelium, divided into several segments. Recent studies indicate a role for RNA interference (RNAi) in the development of the epididymis, however, the actual requirement for RNAi has remained elusive. Here, we present the first evidence of a direct need for RNAi in the differentiation of the epididymal epithelium.

Methodology/principal findings: By utilizing the Cre-LoxP system we have generated a conditional knock-out of Dicer1 in the two most proximal segments of the mouse epididymis. Recombination of Dicer1, catalyzed by Defb41(iCre/wt), took place before puberty, starting from 12 days postpartum. Shortly thereafter, downregulation of the expression of two genes specific for the most proximal epididymis (lipocalin 8 and cystatin 8) was observed. Following this, segment development continued until week 5 at which age the epithelium started to regress back to an undifferentiated state. The dedifferentiated epithelium also showed an increase in estrogen receptor 1 expression while the expression of androgen receptor and its target genes; glutathione peroxidase 5, lipocalin 5 and cysteine-rich secretory protein 1 was downregulated, indicating imbalanced sex steroid signaling.

Conclusions/significance: At the time of the final epididymal development, Dicer1 acts as a regulator of signaling pathways essential for maintaining epithelial cell differentiation.

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Related in: MedlinePlus

Cell proliferation and apoptosis.Ki67 immunostaining; (A) Control; (B) Dicer1fl/fl; Defb41iCre/wt mouse. TUNEL labeling; (D) Control; (E) Dicer1fl/fl; Defb41iCre/wt mouse. Arrows mark apoptotic cells. (C, F) Comparison of number of proliferating cells and apoptotic cells in the initial segment (IS) and caput (CAP) of control (white bars) and Dicer1fl/fl; Defb41iCre/wt mice (grey bars). The number of proliferating and apoptotic cells were calculated from ten tubular cross sections of 5 control and 5 (Ki67 stained) and 8 (TUNEL labeled) Dicer1fl/fl;Defb41iCre/wt mice epididymides and the total cell number was divided by the circumference of the tubular cross sections (mm). Statistical significance of changes, calculated using the unpaired t-test, is indicated as follows: ns, not significant; *, P≤0.05; ***, P≤0.001. Scale bars 100 µm.
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pone-0038457-g005: Cell proliferation and apoptosis.Ki67 immunostaining; (A) Control; (B) Dicer1fl/fl; Defb41iCre/wt mouse. TUNEL labeling; (D) Control; (E) Dicer1fl/fl; Defb41iCre/wt mouse. Arrows mark apoptotic cells. (C, F) Comparison of number of proliferating cells and apoptotic cells in the initial segment (IS) and caput (CAP) of control (white bars) and Dicer1fl/fl; Defb41iCre/wt mice (grey bars). The number of proliferating and apoptotic cells were calculated from ten tubular cross sections of 5 control and 5 (Ki67 stained) and 8 (TUNEL labeled) Dicer1fl/fl;Defb41iCre/wt mice epididymides and the total cell number was divided by the circumference of the tubular cross sections (mm). Statistical significance of changes, calculated using the unpaired t-test, is indicated as follows: ns, not significant; *, P≤0.05; ***, P≤0.001. Scale bars 100 µm.

Mentions: Immunohistochemical studies showed a marked increase in cell proliferation throughout the proximal epididymis of 2 month-old Dicer1fl/fl; Defb41iCre/wt mice (Figure 5B). The number of Ki-67 positive cells was on average 2 times higher in Dicer1 cKO IS (15.2±3.0 cells/mm, control: 6.4±0.6 cells/mm, P≤0.05) and almost 6 times higher in Dicer1 cKO CAP (11.1±1.7 cells/mm, control: 1.9±0.5 cells/mm, P≤0.001) compared with those of control mice epididymides (Figure 5C). Although the epithelium was highly proliferative, it did not lead to a marked increase in the size of the Dicer1fl/fl; Defb41iCre/wt mouse epididymides. On the contrary, at the age of 2 months the Dicer1 cKO epididymis was significantly smaller than that of the control mouse. Furthermore, the number of apoptotic cells was increased in Dicer1 cKO IS and CAP (Dicer1 cKO IS: 0.89±0.39 cells/mm, control IS: 0.07±0.07 cells/mm; Dicer1 cKO CAP: 1.12±0.34 cells/mm, control CAP: 0.07±0.07 cells/mm, P≤0.05) (Figure 5D–F). The difference seen in IS was not statistically significant owing to high variation between the individual mice.


Dicer1 ablation in the mouse epididymis causes dedifferentiation of the epithelium and imbalance in sex steroid signaling.

Björkgren I, Saastamoinen L, Krutskikh A, Huhtaniemi I, Poutanen M, Sipilä P - PLoS ONE (2012)

Cell proliferation and apoptosis.Ki67 immunostaining; (A) Control; (B) Dicer1fl/fl; Defb41iCre/wt mouse. TUNEL labeling; (D) Control; (E) Dicer1fl/fl; Defb41iCre/wt mouse. Arrows mark apoptotic cells. (C, F) Comparison of number of proliferating cells and apoptotic cells in the initial segment (IS) and caput (CAP) of control (white bars) and Dicer1fl/fl; Defb41iCre/wt mice (grey bars). The number of proliferating and apoptotic cells were calculated from ten tubular cross sections of 5 control and 5 (Ki67 stained) and 8 (TUNEL labeled) Dicer1fl/fl;Defb41iCre/wt mice epididymides and the total cell number was divided by the circumference of the tubular cross sections (mm). Statistical significance of changes, calculated using the unpaired t-test, is indicated as follows: ns, not significant; *, P≤0.05; ***, P≤0.001. Scale bars 100 µm.
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Related In: Results  -  Collection

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pone-0038457-g005: Cell proliferation and apoptosis.Ki67 immunostaining; (A) Control; (B) Dicer1fl/fl; Defb41iCre/wt mouse. TUNEL labeling; (D) Control; (E) Dicer1fl/fl; Defb41iCre/wt mouse. Arrows mark apoptotic cells. (C, F) Comparison of number of proliferating cells and apoptotic cells in the initial segment (IS) and caput (CAP) of control (white bars) and Dicer1fl/fl; Defb41iCre/wt mice (grey bars). The number of proliferating and apoptotic cells were calculated from ten tubular cross sections of 5 control and 5 (Ki67 stained) and 8 (TUNEL labeled) Dicer1fl/fl;Defb41iCre/wt mice epididymides and the total cell number was divided by the circumference of the tubular cross sections (mm). Statistical significance of changes, calculated using the unpaired t-test, is indicated as follows: ns, not significant; *, P≤0.05; ***, P≤0.001. Scale bars 100 µm.
Mentions: Immunohistochemical studies showed a marked increase in cell proliferation throughout the proximal epididymis of 2 month-old Dicer1fl/fl; Defb41iCre/wt mice (Figure 5B). The number of Ki-67 positive cells was on average 2 times higher in Dicer1 cKO IS (15.2±3.0 cells/mm, control: 6.4±0.6 cells/mm, P≤0.05) and almost 6 times higher in Dicer1 cKO CAP (11.1±1.7 cells/mm, control: 1.9±0.5 cells/mm, P≤0.001) compared with those of control mice epididymides (Figure 5C). Although the epithelium was highly proliferative, it did not lead to a marked increase in the size of the Dicer1fl/fl; Defb41iCre/wt mouse epididymides. On the contrary, at the age of 2 months the Dicer1 cKO epididymis was significantly smaller than that of the control mouse. Furthermore, the number of apoptotic cells was increased in Dicer1 cKO IS and CAP (Dicer1 cKO IS: 0.89±0.39 cells/mm, control IS: 0.07±0.07 cells/mm; Dicer1 cKO CAP: 1.12±0.34 cells/mm, control CAP: 0.07±0.07 cells/mm, P≤0.05) (Figure 5D–F). The difference seen in IS was not statistically significant owing to high variation between the individual mice.

Bottom Line: The postnatal development of the epididymis is a complex process that results in a highly differentiated epithelium, divided into several segments.The dedifferentiated epithelium also showed an increase in estrogen receptor 1 expression while the expression of androgen receptor and its target genes; glutathione peroxidase 5, lipocalin 5 and cysteine-rich secretory protein 1 was downregulated, indicating imbalanced sex steroid signaling.At the time of the final epididymal development, Dicer1 acts as a regulator of signaling pathways essential for maintaining epithelial cell differentiation.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Institute of Biomedicine, University of Turku, Turku, Finland.

ABSTRACT

Background: The postnatal development of the epididymis is a complex process that results in a highly differentiated epithelium, divided into several segments. Recent studies indicate a role for RNA interference (RNAi) in the development of the epididymis, however, the actual requirement for RNAi has remained elusive. Here, we present the first evidence of a direct need for RNAi in the differentiation of the epididymal epithelium.

Methodology/principal findings: By utilizing the Cre-LoxP system we have generated a conditional knock-out of Dicer1 in the two most proximal segments of the mouse epididymis. Recombination of Dicer1, catalyzed by Defb41(iCre/wt), took place before puberty, starting from 12 days postpartum. Shortly thereafter, downregulation of the expression of two genes specific for the most proximal epididymis (lipocalin 8 and cystatin 8) was observed. Following this, segment development continued until week 5 at which age the epithelium started to regress back to an undifferentiated state. The dedifferentiated epithelium also showed an increase in estrogen receptor 1 expression while the expression of androgen receptor and its target genes; glutathione peroxidase 5, lipocalin 5 and cysteine-rich secretory protein 1 was downregulated, indicating imbalanced sex steroid signaling.

Conclusions/significance: At the time of the final epididymal development, Dicer1 acts as a regulator of signaling pathways essential for maintaining epithelial cell differentiation.

Show MeSH
Related in: MedlinePlus