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Ornithine Decarboxylase Activity Is Required for Prostatic Budding in the Developing Mouse Prostate.

Gamat M, Malinowski RL, Parkhurst LJ, Steinke LM, Marker PC - PLoS ONE (2015)

Bottom Line: Inhibiting ornithine decarboxylase using DFMO in UGS organ culture blocked the induction of prostatic buds by androgens, and significantly decreased expression of key prostate transcription factor, Nkx3.1, by androgens.DFMO also significantly decreased the expression of developmental regulatory gene Notch1.Together these results indicate that Odc1 and polyamines are required for androgens to exert their effect in mediating prostatic bud induction, and are required for the expression of a subset of prostatic developmental regulatory genes including Notch1 and Nkx3.1.

View Article: PubMed Central - PubMed

Affiliation: School of Pharmacy, University of Wisconsin-Madison, 777 Highland Avenue, Madison, WI, United States of America.

ABSTRACT
The prostate is a male accessory sex gland that produces secretions in seminal fluid to facilitate fertilization. Prostate secretory function is dependent on androgens, although the mechanism by which androgens exert their effects is still unclear. Polyamines are small cationic molecules that play pivotal roles in DNA transcription, translation and gene regulation. The rate-limiting enzyme in polyamine biosynthesis is ornithine decarboxylase, which is encoded by the gene Odc1. Ornithine decarboxylase mRNA decreases in the prostate upon castration and increases upon administration of androgens. Furthermore, testosterone administered to castrated male mice restores prostate secretory activity, whereas administering testosterone and the ornithine decarboxylase inhibitor D,L-α-difluromethylornithine (DFMO) to castrated males does not restore prostate secretory activity, suggesting that polyamines are required for androgens to exert their effects. To date, no one has examined polyamines in prostate development, which is also androgen dependent. In this study, we showed that ornithine decarboxylase protein was expressed in the epithelium of the ventral, dorsolateral and anterior lobes of the adult mouse prostate. Ornithine decarboxylase protein was also expressed in the urogenital sinus (UGS) epithelium of the male and female embryo prior to prostate development, and expression continued in prostatic epithelial buds as they emerged from the UGS. Inhibiting ornithine decarboxylase using DFMO in UGS organ culture blocked the induction of prostatic buds by androgens, and significantly decreased expression of key prostate transcription factor, Nkx3.1, by androgens. DFMO also significantly decreased the expression of developmental regulatory gene Notch1. Other genes implicated in prostatic development including Sox9, Wif1 and Srd5a2 were unaffected by DFMO. Together these results indicate that Odc1 and polyamines are required for androgens to exert their effect in mediating prostatic bud induction, and are required for the expression of a subset of prostatic developmental regulatory genes including Notch1 and Nkx3.1.

No MeSH data available.


Related in: MedlinePlus

E16 UGS cultured in the presence or absence of testosterone and DFMO.The tissues were fixed in 4% PFA then stained for E-Cadherin to visualize the urogenital epithelium. In the no steroid control media, the urogenital epithelium remained smooth with no evidence of prostatic bud formation (A). After six days of treatment with testosterone, the UGS underwent prolific bud formation (B). Culturing the UGS with testosterone and DFMO decreased the number of prostatic buds compared to the testosterone treatment, and comparable to the no steroid control UGS (C). For each treatment, the numbers of buds were counted. Testosterone induced significantly more buds compared to no steroid treatment, and treating UGS tissues with DFMO and testosterone significantly decreased the number of buds compared to testosterone alone (D). Culturing the UGS for six days did not adversely affect the histology of cells regardless of whether they were cultured in NS media (E), T (F) or T+DFMO (G) media. In all three treatments, the nuclei were large and healthy looking with no evidence of pyknosis. To assess proliferation, we assessed Ki67 expression in our UGS culture. Ki67 was expressed in specific epithelial cells and mesenchymal cells in the NS control UGS (H). In the T treated UGS, Ki67 was expressed in cell clusters in prostatic buds, as well as specific cells within the mesenchyme (I). In T+DFMO treated UGS, Ki67 was expressed in certain cells in the epithelium as well as the mesenchyme. Abbreviations: NS no steroid control, T testosterone, DFMO difluoromethylornithine. The dotted line demarcates the border between epithelium and mesenchyme. Images E-J were taken at 100X magnification.
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pone.0139522.g004: E16 UGS cultured in the presence or absence of testosterone and DFMO.The tissues were fixed in 4% PFA then stained for E-Cadherin to visualize the urogenital epithelium. In the no steroid control media, the urogenital epithelium remained smooth with no evidence of prostatic bud formation (A). After six days of treatment with testosterone, the UGS underwent prolific bud formation (B). Culturing the UGS with testosterone and DFMO decreased the number of prostatic buds compared to the testosterone treatment, and comparable to the no steroid control UGS (C). For each treatment, the numbers of buds were counted. Testosterone induced significantly more buds compared to no steroid treatment, and treating UGS tissues with DFMO and testosterone significantly decreased the number of buds compared to testosterone alone (D). Culturing the UGS for six days did not adversely affect the histology of cells regardless of whether they were cultured in NS media (E), T (F) or T+DFMO (G) media. In all three treatments, the nuclei were large and healthy looking with no evidence of pyknosis. To assess proliferation, we assessed Ki67 expression in our UGS culture. Ki67 was expressed in specific epithelial cells and mesenchymal cells in the NS control UGS (H). In the T treated UGS, Ki67 was expressed in cell clusters in prostatic buds, as well as specific cells within the mesenchyme (I). In T+DFMO treated UGS, Ki67 was expressed in certain cells in the epithelium as well as the mesenchyme. Abbreviations: NS no steroid control, T testosterone, DFMO difluoromethylornithine. The dotted line demarcates the border between epithelium and mesenchyme. Images E-J were taken at 100X magnification.

Mentions: In the no steroid (NS) control, the UGS tissues did not undergo epithelial bud growth (Fig 4A). The tube of epithelium was smooth and of uniform thickness. When embryonic urogenital sinus tissue was cultured in testosterone, prostatic buds grew from the epithelium and into the overlying mesenchyme (Fig 4B) [39]. When prostatic buds were counted, testosterone treatment significantly increased the number of prostatic buds produced compared to the no steroid control, (Fig 4D, * p<0.05). However, when the UGS was cultured in testosterone with DFMO, prostatic budding was extremely reduced compared to the testosterone treatment (Fig 4C). When the numbers of buds were counted, testosterone and DFMO treatment significantly decreased the number of prostatic buds compared to testosterone treatment only (Fig 4D, *<0.05) and the number of buds in the T+DFMO treated UGS were comparable to those in the NS control.


Ornithine Decarboxylase Activity Is Required for Prostatic Budding in the Developing Mouse Prostate.

Gamat M, Malinowski RL, Parkhurst LJ, Steinke LM, Marker PC - PLoS ONE (2015)

E16 UGS cultured in the presence or absence of testosterone and DFMO.The tissues were fixed in 4% PFA then stained for E-Cadherin to visualize the urogenital epithelium. In the no steroid control media, the urogenital epithelium remained smooth with no evidence of prostatic bud formation (A). After six days of treatment with testosterone, the UGS underwent prolific bud formation (B). Culturing the UGS with testosterone and DFMO decreased the number of prostatic buds compared to the testosterone treatment, and comparable to the no steroid control UGS (C). For each treatment, the numbers of buds were counted. Testosterone induced significantly more buds compared to no steroid treatment, and treating UGS tissues with DFMO and testosterone significantly decreased the number of buds compared to testosterone alone (D). Culturing the UGS for six days did not adversely affect the histology of cells regardless of whether they were cultured in NS media (E), T (F) or T+DFMO (G) media. In all three treatments, the nuclei were large and healthy looking with no evidence of pyknosis. To assess proliferation, we assessed Ki67 expression in our UGS culture. Ki67 was expressed in specific epithelial cells and mesenchymal cells in the NS control UGS (H). In the T treated UGS, Ki67 was expressed in cell clusters in prostatic buds, as well as specific cells within the mesenchyme (I). In T+DFMO treated UGS, Ki67 was expressed in certain cells in the epithelium as well as the mesenchyme. Abbreviations: NS no steroid control, T testosterone, DFMO difluoromethylornithine. The dotted line demarcates the border between epithelium and mesenchyme. Images E-J were taken at 100X magnification.
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Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4591331&req=5

pone.0139522.g004: E16 UGS cultured in the presence or absence of testosterone and DFMO.The tissues were fixed in 4% PFA then stained for E-Cadherin to visualize the urogenital epithelium. In the no steroid control media, the urogenital epithelium remained smooth with no evidence of prostatic bud formation (A). After six days of treatment with testosterone, the UGS underwent prolific bud formation (B). Culturing the UGS with testosterone and DFMO decreased the number of prostatic buds compared to the testosterone treatment, and comparable to the no steroid control UGS (C). For each treatment, the numbers of buds were counted. Testosterone induced significantly more buds compared to no steroid treatment, and treating UGS tissues with DFMO and testosterone significantly decreased the number of buds compared to testosterone alone (D). Culturing the UGS for six days did not adversely affect the histology of cells regardless of whether they were cultured in NS media (E), T (F) or T+DFMO (G) media. In all three treatments, the nuclei were large and healthy looking with no evidence of pyknosis. To assess proliferation, we assessed Ki67 expression in our UGS culture. Ki67 was expressed in specific epithelial cells and mesenchymal cells in the NS control UGS (H). In the T treated UGS, Ki67 was expressed in cell clusters in prostatic buds, as well as specific cells within the mesenchyme (I). In T+DFMO treated UGS, Ki67 was expressed in certain cells in the epithelium as well as the mesenchyme. Abbreviations: NS no steroid control, T testosterone, DFMO difluoromethylornithine. The dotted line demarcates the border between epithelium and mesenchyme. Images E-J were taken at 100X magnification.
Mentions: In the no steroid (NS) control, the UGS tissues did not undergo epithelial bud growth (Fig 4A). The tube of epithelium was smooth and of uniform thickness. When embryonic urogenital sinus tissue was cultured in testosterone, prostatic buds grew from the epithelium and into the overlying mesenchyme (Fig 4B) [39]. When prostatic buds were counted, testosterone treatment significantly increased the number of prostatic buds produced compared to the no steroid control, (Fig 4D, * p<0.05). However, when the UGS was cultured in testosterone with DFMO, prostatic budding was extremely reduced compared to the testosterone treatment (Fig 4C). When the numbers of buds were counted, testosterone and DFMO treatment significantly decreased the number of prostatic buds compared to testosterone treatment only (Fig 4D, *<0.05) and the number of buds in the T+DFMO treated UGS were comparable to those in the NS control.

Bottom Line: Inhibiting ornithine decarboxylase using DFMO in UGS organ culture blocked the induction of prostatic buds by androgens, and significantly decreased expression of key prostate transcription factor, Nkx3.1, by androgens.DFMO also significantly decreased the expression of developmental regulatory gene Notch1.Together these results indicate that Odc1 and polyamines are required for androgens to exert their effect in mediating prostatic bud induction, and are required for the expression of a subset of prostatic developmental regulatory genes including Notch1 and Nkx3.1.

View Article: PubMed Central - PubMed

Affiliation: School of Pharmacy, University of Wisconsin-Madison, 777 Highland Avenue, Madison, WI, United States of America.

ABSTRACT
The prostate is a male accessory sex gland that produces secretions in seminal fluid to facilitate fertilization. Prostate secretory function is dependent on androgens, although the mechanism by which androgens exert their effects is still unclear. Polyamines are small cationic molecules that play pivotal roles in DNA transcription, translation and gene regulation. The rate-limiting enzyme in polyamine biosynthesis is ornithine decarboxylase, which is encoded by the gene Odc1. Ornithine decarboxylase mRNA decreases in the prostate upon castration and increases upon administration of androgens. Furthermore, testosterone administered to castrated male mice restores prostate secretory activity, whereas administering testosterone and the ornithine decarboxylase inhibitor D,L-α-difluromethylornithine (DFMO) to castrated males does not restore prostate secretory activity, suggesting that polyamines are required for androgens to exert their effects. To date, no one has examined polyamines in prostate development, which is also androgen dependent. In this study, we showed that ornithine decarboxylase protein was expressed in the epithelium of the ventral, dorsolateral and anterior lobes of the adult mouse prostate. Ornithine decarboxylase protein was also expressed in the urogenital sinus (UGS) epithelium of the male and female embryo prior to prostate development, and expression continued in prostatic epithelial buds as they emerged from the UGS. Inhibiting ornithine decarboxylase using DFMO in UGS organ culture blocked the induction of prostatic buds by androgens, and significantly decreased expression of key prostate transcription factor, Nkx3.1, by androgens. DFMO also significantly decreased the expression of developmental regulatory gene Notch1. Other genes implicated in prostatic development including Sox9, Wif1 and Srd5a2 were unaffected by DFMO. Together these results indicate that Odc1 and polyamines are required for androgens to exert their effect in mediating prostatic bud induction, and are required for the expression of a subset of prostatic developmental regulatory genes including Notch1 and Nkx3.1.

No MeSH data available.


Related in: MedlinePlus