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Evaluation of human first trimester decidual and telomerase-transformed endometrial stromal cells as model systems of in vitro decidualization.
Reprod Biol Endocrinol. 2011 Dec 07; 9:155.RB

Abstract

BACKGROUND

Decidualization, the differentiation process of maternal uterine stromal cells into secretory decidual cells, is a prerequisite for successful implantation and progression of pregnancy. For in vitro differentiation mostly primary human endometrial stromal cells (HESC) isolated from uterine samples after hysterectomy for benign gynaecological diseases are utilised. However, a continuous supply of endometrial tissue is often lacking. Hence, we analysed whether cultivated human decidual stromal cells (HDSC) prepared from first trimester pregnancy terminations may represent an alternative model system for in vitro decidualization. Moreover, based on the expression of critical marker genes these cells were compared to a previously established endometrial stromal cell line during in vitro differentiation.

METHODS

HDSC isolated from decidual tissue attached to first trimester placentae, and telomerase-transformed human endometrial stromal cells (THESC) were characterised by immunofluorescence and differentiated in vitro using either cyclic adenosine monophosphate (cAMP) and/or estrogen (E2)/progesterone (P4). Proliferation was measured by analyzing cumulative cell numbers. Expression of mRNAs encoding progesterone receptor (PR), prolactin (PRL), insulin-like growth factor binding protein-1 (IGFBP1), and Dickkopf-1 (DKK1) was evaluated using quantitative PCR after 3, 6, 9 and 12 days of in vitro differentiation. PRL and IGFBP-1 protein expression was investigated by enzyme-linked immunosorbent assay (ELISA) and Western blotting, respectively. Furthermore, forkhead box O1A (FOXO1A), a critical transcription factor in decidualization, was analysed by immunofluorescence and Western blotting at two different time points of differentiation.

RESULTS

Treatment with cAMP provoked morphological changes and growth arrest of THESC and HDSC, the latter showing loss of cells after 6 days of treatment. E2P4 stimulation did neither affect cell morphology nor proliferation of THESC and HDSC. Upon cAMP stimulation PR mRNA was suppressed in HDSC but not in THESC, whereas E2P4 did not alter transcript levels in both cell types. Protein expression of PR-A and PR-B was detectable in HDSC and diminished under cAMP, whereas THESC failed to produce the nuclear receptors. Supplementation of cAMP induced mRNA and protein expression of PRL and IGFBP-1 in both cell types at day 3, 6, 9, and 12 of treatment. In HDSC stimulation with E2P4 increased PRL and IGFBP-1 mRNA and protein production, whereas hormone treatment did not induce the two factors in THESC. E2P4 increased DKK1 mRNA at all time points in HDSC and cAMP provoked induction at day 9 and 12 of differentiation. In contrast, cAMP suppressed DKK1 mRNA in THESC, whereas E2P4 was ineffective. In both cell types combined treatments with cAMP and E2P4 provoked higher expression levels of PRL and IGFBP1 mRNA and protein as compared to cAMP stimulation alone. FOXO1A protein and its nuclear abundance were increased by cAMP in both cell types. However, reduction of its nuclear localisation upon E2P4 treatment could only be observed in HDSC.

CONCLUSION

Both HDSC and THESC may represent suitable model systems for cAMP-dependent in vitro decidualization. Since cAMP decreases cell viability of HDSC after 6 days of incubation, this substance should be preferentially used in short-term experiments. Progesterone treatment of THESC might not be applicable since these cells lack progesterone response and PR protein. In contrast, stimulation of PR-expressing HDSC with E2P4 or cAMP/E2P4 may represent an appropriate protocol for human in vitro decidualization inducing and maintaining expression of critical marker genes in a time-dependent manner.

Authors+Show Affiliations

Department of Obstetrics and Fetal-Maternal Medicine, Reproductive Biology Unit, Medical University of Vienna, A-1090 Vienna, Austria.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

22151839

Citation

Saleh, Leila, et al. "Evaluation of Human First Trimester Decidual and Telomerase-transformed Endometrial Stromal Cells as Model Systems of in Vitro Decidualization." Reproductive Biology and Endocrinology : RB&E, vol. 9, 2011, p. 155.
Saleh L, Otti GR, Fiala C, et al. Evaluation of human first trimester decidual and telomerase-transformed endometrial stromal cells as model systems of in vitro decidualization. Reprod Biol Endocrinol. 2011;9:155.
Saleh, L., Otti, G. R., Fiala, C., Pollheimer, J., & Knöfler, M. (2011). Evaluation of human first trimester decidual and telomerase-transformed endometrial stromal cells as model systems of in vitro decidualization. Reproductive Biology and Endocrinology : RB&E, 9, 155. https://doi.org/10.1186/1477-7827-9-155
Saleh L, et al. Evaluation of Human First Trimester Decidual and Telomerase-transformed Endometrial Stromal Cells as Model Systems of in Vitro Decidualization. Reprod Biol Endocrinol. 2011 Dec 7;9:155. PubMed PMID: 22151839.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Evaluation of human first trimester decidual and telomerase-transformed endometrial stromal cells as model systems of in vitro decidualization. AU - Saleh,Leila, AU - Otti,Gerlinde R, AU - Fiala,Christian, AU - Pollheimer,Jürgen, AU - Knöfler,Martin, Y1 - 2011/12/07/ PY - 2011/06/10/received PY - 2011/12/07/accepted PY - 2011/12/14/entrez PY - 2011/12/14/pubmed PY - 2012/8/17/medline SP - 155 EP - 155 JF - Reproductive biology and endocrinology : RB&E JO - Reprod. Biol. Endocrinol. VL - 9 N2 - BACKGROUND: Decidualization, the differentiation process of maternal uterine stromal cells into secretory decidual cells, is a prerequisite for successful implantation and progression of pregnancy. For in vitro differentiation mostly primary human endometrial stromal cells (HESC) isolated from uterine samples after hysterectomy for benign gynaecological diseases are utilised. However, a continuous supply of endometrial tissue is often lacking. Hence, we analysed whether cultivated human decidual stromal cells (HDSC) prepared from first trimester pregnancy terminations may represent an alternative model system for in vitro decidualization. Moreover, based on the expression of critical marker genes these cells were compared to a previously established endometrial stromal cell line during in vitro differentiation. METHODS: HDSC isolated from decidual tissue attached to first trimester placentae, and telomerase-transformed human endometrial stromal cells (THESC) were characterised by immunofluorescence and differentiated in vitro using either cyclic adenosine monophosphate (cAMP) and/or estrogen (E2)/progesterone (P4). Proliferation was measured by analyzing cumulative cell numbers. Expression of mRNAs encoding progesterone receptor (PR), prolactin (PRL), insulin-like growth factor binding protein-1 (IGFBP1), and Dickkopf-1 (DKK1) was evaluated using quantitative PCR after 3, 6, 9 and 12 days of in vitro differentiation. PRL and IGFBP-1 protein expression was investigated by enzyme-linked immunosorbent assay (ELISA) and Western blotting, respectively. Furthermore, forkhead box O1A (FOXO1A), a critical transcription factor in decidualization, was analysed by immunofluorescence and Western blotting at two different time points of differentiation. RESULTS: Treatment with cAMP provoked morphological changes and growth arrest of THESC and HDSC, the latter showing loss of cells after 6 days of treatment. E2P4 stimulation did neither affect cell morphology nor proliferation of THESC and HDSC. Upon cAMP stimulation PR mRNA was suppressed in HDSC but not in THESC, whereas E2P4 did not alter transcript levels in both cell types. Protein expression of PR-A and PR-B was detectable in HDSC and diminished under cAMP, whereas THESC failed to produce the nuclear receptors. Supplementation of cAMP induced mRNA and protein expression of PRL and IGFBP-1 in both cell types at day 3, 6, 9, and 12 of treatment. In HDSC stimulation with E2P4 increased PRL and IGFBP-1 mRNA and protein production, whereas hormone treatment did not induce the two factors in THESC. E2P4 increased DKK1 mRNA at all time points in HDSC and cAMP provoked induction at day 9 and 12 of differentiation. In contrast, cAMP suppressed DKK1 mRNA in THESC, whereas E2P4 was ineffective. In both cell types combined treatments with cAMP and E2P4 provoked higher expression levels of PRL and IGFBP1 mRNA and protein as compared to cAMP stimulation alone. FOXO1A protein and its nuclear abundance were increased by cAMP in both cell types. However, reduction of its nuclear localisation upon E2P4 treatment could only be observed in HDSC. CONCLUSION: Both HDSC and THESC may represent suitable model systems for cAMP-dependent in vitro decidualization. Since cAMP decreases cell viability of HDSC after 6 days of incubation, this substance should be preferentially used in short-term experiments. Progesterone treatment of THESC might not be applicable since these cells lack progesterone response and PR protein. In contrast, stimulation of PR-expressing HDSC with E2P4 or cAMP/E2P4 may represent an appropriate protocol for human in vitro decidualization inducing and maintaining expression of critical marker genes in a time-dependent manner. SN - 1477-7827 UR - https://www.unboundmedicine.com/medline/citation/22151839/Evaluation_of_human_first_trimester_decidual_and_telomerase_transformed_endometrial_stromal_cells_as_model_systems_of_in_vitro_decidualization_ L2 - https://rbej.biomedcentral.com/articles/10.1186/1477-7827-9-155 DB - PRIME DP - Unbound Medicine ER -