AKT Isoforms 1 and 3 regulate basal and epidermal growth factor-stimulated SGHPL-5 trophoblast cell migration in humans

Peter Haslinger, Sandra Haidar, Stefan Eugen Sonderegger, Jan Velten Otten, Jurgen Pollheimer, Guy Whitley, Martin Knofler

Research output: Contribution to journalArticleResearchpeer-review

Abstract

Protein kinase B/AKT is critically involved in murine placental development and migration of human placental trophoblasts into maternal uterine tissue. However, localization of the three AKT isoforms within human placenta and their roles in extravillous trophoblasts have not been elucidated. Therefore, we analyzed the expression pattern and function of AKT1, AKT2, and AKT3 in migratory human trophoblasts using SGHPL-5 cell pools stably expressing small-hairpin microRNA (shRNAmir) against AKT1, AKT2, or AKT3 as a model. Western blot analyses using isoform-specific antibodies revealed ubiquitous expression of AKT1, AKT2, and AKT3 in primary villous and extravillous trophoblasts and the trophoblastic cell lines JEG-3, HTR-8/SVneo, and SGHPL-5. Immunofluorescence of first-trimester placentae localized AKT2 and AKT3 to the cytoplasm and nucleus, respectively, in all subtypes of cytotrophoblasts, whereas AKT1 was detected in both cellular compartments. A similar distribution of AKT isoforms was detectable in SGHPL-5 cells. Gene silencing using shRNAmir decreased protein expression of AKT1, AKT2, and AKT3 to 16 , 8 , and 11 , respectively, in SGHPL-5 cells. Compared with shRNAmir controls, proliferation and camptothecin-induced apoptosis were not affected in the different AKT knockdown cells. However, basal and epidermal growth factor (EGF)-induced trophoblast migration was significantly reduced in AKT1 and AKT3 gene-silenced cells, whereas downregulation of AKT2 was not effective. Accordingly, a decrease in EGF-stimulated phosphorylation of AKT (Ser473 and Thr308) and its downstream target mTORC1 (Ser2448) was noticed in AKT1 and AKT3 shRNAmir cell pools. In summary, the results suggest that the AKT isoforms 1 and 3 promote basal as well as EGF-induced trophoblast migration.
Original languageEnglish
Pages (from-to)1 - 11
Number of pages11
JournalBiology of Reproduction
Volume88
Issue number3
DOIs
Publication statusPublished - 2013

Cite this

Haslinger, P., Haidar, S., Sonderegger, S. E., Otten, J. V., Pollheimer, J., Whitley, G., & Knofler, M. (2013). AKT Isoforms 1 and 3 regulate basal and epidermal growth factor-stimulated SGHPL-5 trophoblast cell migration in humans. Biology of Reproduction, 88(3), 1 - 11. https://doi.org/10.1095/?biolreprod.112.104778
Haslinger, Peter ; Haidar, Sandra ; Sonderegger, Stefan Eugen ; Otten, Jan Velten ; Pollheimer, Jurgen ; Whitley, Guy ; Knofler, Martin. / AKT Isoforms 1 and 3 regulate basal and epidermal growth factor-stimulated SGHPL-5 trophoblast cell migration in humans. In: Biology of Reproduction. 2013 ; Vol. 88, No. 3. pp. 1 - 11.
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abstract = "Protein kinase B/AKT is critically involved in murine placental development and migration of human placental trophoblasts into maternal uterine tissue. However, localization of the three AKT isoforms within human placenta and their roles in extravillous trophoblasts have not been elucidated. Therefore, we analyzed the expression pattern and function of AKT1, AKT2, and AKT3 in migratory human trophoblasts using SGHPL-5 cell pools stably expressing small-hairpin microRNA (shRNAmir) against AKT1, AKT2, or AKT3 as a model. Western blot analyses using isoform-specific antibodies revealed ubiquitous expression of AKT1, AKT2, and AKT3 in primary villous and extravillous trophoblasts and the trophoblastic cell lines JEG-3, HTR-8/SVneo, and SGHPL-5. Immunofluorescence of first-trimester placentae localized AKT2 and AKT3 to the cytoplasm and nucleus, respectively, in all subtypes of cytotrophoblasts, whereas AKT1 was detected in both cellular compartments. A similar distribution of AKT isoforms was detectable in SGHPL-5 cells. Gene silencing using shRNAmir decreased protein expression of AKT1, AKT2, and AKT3 to 16 , 8 , and 11 , respectively, in SGHPL-5 cells. Compared with shRNAmir controls, proliferation and camptothecin-induced apoptosis were not affected in the different AKT knockdown cells. However, basal and epidermal growth factor (EGF)-induced trophoblast migration was significantly reduced in AKT1 and AKT3 gene-silenced cells, whereas downregulation of AKT2 was not effective. Accordingly, a decrease in EGF-stimulated phosphorylation of AKT (Ser473 and Thr308) and its downstream target mTORC1 (Ser2448) was noticed in AKT1 and AKT3 shRNAmir cell pools. In summary, the results suggest that the AKT isoforms 1 and 3 promote basal as well as EGF-induced trophoblast migration.",
author = "Peter Haslinger and Sandra Haidar and Sonderegger, {Stefan Eugen} and Otten, {Jan Velten} and Jurgen Pollheimer and Guy Whitley and Martin Knofler",
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Haslinger, P, Haidar, S, Sonderegger, SE, Otten, JV, Pollheimer, J, Whitley, G & Knofler, M 2013, 'AKT Isoforms 1 and 3 regulate basal and epidermal growth factor-stimulated SGHPL-5 trophoblast cell migration in humans' Biology of Reproduction, vol. 88, no. 3, pp. 1 - 11. https://doi.org/10.1095/?biolreprod.112.104778

AKT Isoforms 1 and 3 regulate basal and epidermal growth factor-stimulated SGHPL-5 trophoblast cell migration in humans. / Haslinger, Peter; Haidar, Sandra; Sonderegger, Stefan Eugen; Otten, Jan Velten; Pollheimer, Jurgen; Whitley, Guy; Knofler, Martin.

In: Biology of Reproduction, Vol. 88, No. 3, 2013, p. 1 - 11.

Research output: Contribution to journalArticleResearchpeer-review

TY - JOUR

T1 - AKT Isoforms 1 and 3 regulate basal and epidermal growth factor-stimulated SGHPL-5 trophoblast cell migration in humans

AU - Haslinger, Peter

AU - Haidar, Sandra

AU - Sonderegger, Stefan Eugen

AU - Otten, Jan Velten

AU - Pollheimer, Jurgen

AU - Whitley, Guy

AU - Knofler, Martin

PY - 2013

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N2 - Protein kinase B/AKT is critically involved in murine placental development and migration of human placental trophoblasts into maternal uterine tissue. However, localization of the three AKT isoforms within human placenta and their roles in extravillous trophoblasts have not been elucidated. Therefore, we analyzed the expression pattern and function of AKT1, AKT2, and AKT3 in migratory human trophoblasts using SGHPL-5 cell pools stably expressing small-hairpin microRNA (shRNAmir) against AKT1, AKT2, or AKT3 as a model. Western blot analyses using isoform-specific antibodies revealed ubiquitous expression of AKT1, AKT2, and AKT3 in primary villous and extravillous trophoblasts and the trophoblastic cell lines JEG-3, HTR-8/SVneo, and SGHPL-5. Immunofluorescence of first-trimester placentae localized AKT2 and AKT3 to the cytoplasm and nucleus, respectively, in all subtypes of cytotrophoblasts, whereas AKT1 was detected in both cellular compartments. A similar distribution of AKT isoforms was detectable in SGHPL-5 cells. Gene silencing using shRNAmir decreased protein expression of AKT1, AKT2, and AKT3 to 16 , 8 , and 11 , respectively, in SGHPL-5 cells. Compared with shRNAmir controls, proliferation and camptothecin-induced apoptosis were not affected in the different AKT knockdown cells. However, basal and epidermal growth factor (EGF)-induced trophoblast migration was significantly reduced in AKT1 and AKT3 gene-silenced cells, whereas downregulation of AKT2 was not effective. Accordingly, a decrease in EGF-stimulated phosphorylation of AKT (Ser473 and Thr308) and its downstream target mTORC1 (Ser2448) was noticed in AKT1 and AKT3 shRNAmir cell pools. In summary, the results suggest that the AKT isoforms 1 and 3 promote basal as well as EGF-induced trophoblast migration.

AB - Protein kinase B/AKT is critically involved in murine placental development and migration of human placental trophoblasts into maternal uterine tissue. However, localization of the three AKT isoforms within human placenta and their roles in extravillous trophoblasts have not been elucidated. Therefore, we analyzed the expression pattern and function of AKT1, AKT2, and AKT3 in migratory human trophoblasts using SGHPL-5 cell pools stably expressing small-hairpin microRNA (shRNAmir) against AKT1, AKT2, or AKT3 as a model. Western blot analyses using isoform-specific antibodies revealed ubiquitous expression of AKT1, AKT2, and AKT3 in primary villous and extravillous trophoblasts and the trophoblastic cell lines JEG-3, HTR-8/SVneo, and SGHPL-5. Immunofluorescence of first-trimester placentae localized AKT2 and AKT3 to the cytoplasm and nucleus, respectively, in all subtypes of cytotrophoblasts, whereas AKT1 was detected in both cellular compartments. A similar distribution of AKT isoforms was detectable in SGHPL-5 cells. Gene silencing using shRNAmir decreased protein expression of AKT1, AKT2, and AKT3 to 16 , 8 , and 11 , respectively, in SGHPL-5 cells. Compared with shRNAmir controls, proliferation and camptothecin-induced apoptosis were not affected in the different AKT knockdown cells. However, basal and epidermal growth factor (EGF)-induced trophoblast migration was significantly reduced in AKT1 and AKT3 gene-silenced cells, whereas downregulation of AKT2 was not effective. Accordingly, a decrease in EGF-stimulated phosphorylation of AKT (Ser473 and Thr308) and its downstream target mTORC1 (Ser2448) was noticed in AKT1 and AKT3 shRNAmir cell pools. In summary, the results suggest that the AKT isoforms 1 and 3 promote basal as well as EGF-induced trophoblast migration.

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