The Experts below are selected from a list of 411 Experts worldwide ranked by ideXlab platform
Amanda M De Mestre - One of the best experts on this subject based on the ideXlab platform.
-
smad1 5 signaling in the early Equine Placenta regulates trophoblast differentiation and chorionic gonadotropin secretion
Endocrinology, 2014Co-Authors: Victoria Cabrerasharp, Jordan E Read, Stephanie Richardson, Alycia A Kowalski, Douglas F Antczak, Judith E Cartwright, Abir Mukherjee, Amanda M De MestreAbstract:TGFβ superfamily proteins, acting via SMAD (Sma- and Mad-related protein)2/3 pathways, regulate Placental function; however, the role of SMAD1/5/8 pathway in the Placenta is unknown. This study investigated the functional role of bone morphogenetic protein (BMP)4 signaling through SMAD1/5 in terminal differentiation of primary chorionic gonadotropin (CG)-secreting trophoblast. Primary Equine trophoblast cells or Placental tissues were isolated from day 27–34 Equine conceptuses. Detected by microarray, RT-PCR, and quantitative RT-PCR, Equine chorionic girdle trophoblast showed increased gene expression of receptors that bind BMP4. BMP4 mRNA expression was 20- to 60-fold higher in Placental tissues adjacent to the chorionic girdle compared with chorionic girdle itself, suggesting BMP4 acts primarily in a paracrine manner on the chorionic girdle. Stimulation of chorionic girdle-trophoblast cells with BMP4 resulted in a dose-dependent and developmental stage-dependent increase in total number and proportion of terminally differentiated binucleate cells. Furthermore, BMP4 treatment induced non-CG-secreting day 31 chorionic girdle trophoblast cells to secrete CG, confirming a specific functional response to BMP4 stimulation. Inhibition of SMAD2/3 signaling combined with BMP4 treatment further enhanced differentiation of trophoblast cells. Phospho-SMAD1/5, but not phospho-SMAD2, expression as determined by Western blotting was tightly regulated during chorionic girdle trophoblast differentiation in vivo, with peak expression of phospho-SMAD1/5 in vivo noted at day 31 corresponding to maximal differentiation response of trophoblast in vitro. Collectively, these experiments demonstrate the involvement of BMP4-dependent pathways in the regulation of Equine trophoblast differentiation in vivo and primary trophoblast differentiation in vitro via activation of SMAD1/5 pathway, a previously unreported mechanism of TGFβ signaling in the mammalian Placenta.
-
SMAD1/5 signaling in the early Equine Placenta regulates trophoblast differentiation and chorionic gonadotropin secretion.
Endocrinology, 2014Co-Authors: Victoria Cabrera-sharp, Jordan E Read, Stephanie Richardson, Alycia A Kowalski, Douglas F Antczak, Judith E Cartwright, Abir Mukherjee, Amanda M De MestreAbstract:TGFβ superfamily proteins, acting via SMAD (Sma- and Mad-related protein)2/3 pathways, regulate Placental function; however, the role of SMAD1/5/8 pathway in the Placenta is unknown. This study investigated the functional role of bone morphogenetic protein (BMP)4 signaling through SMAD1/5 in terminal differentiation of primary chorionic gonadotropin (CG)-secreting trophoblast. Primary Equine trophoblast cells or Placental tissues were isolated from day 27–34 Equine conceptuses. Detected by microarray, RT-PCR, and quantitative RT-PCR, Equine chorionic girdle trophoblast showed increased gene expression of receptors that bind BMP4. BMP4 mRNA expression was 20- to 60-fold higher in Placental tissues adjacent to the chorionic girdle compared with chorionic girdle itself, suggesting BMP4 acts primarily in a paracrine manner on the chorionic girdle. Stimulation of chorionic girdle-trophoblast cells with BMP4 resulted in a dose-dependent and developmental stage-dependent increase in total number and proportion of terminally differentiated binucleate cells. Furthermore, BMP4 treatment induced non-CG-secreting day 31 chorionic girdle trophoblast cells to secrete CG, confirming a specific functional response to BMP4 stimulation. Inhibition of SMAD2/3 signaling combined with BMP4 treatment further enhanced differentiation of trophoblast cells. Phospho-SMAD1/5, but not phospho-SMAD2, expression as determined by Western blotting was tightly regulated during chorionic girdle trophoblast differentiation in vivo, with peak expression of phospho-SMAD1/5 in vivo noted at day 31 corresponding to maximal differentiation response of trophoblast in vitro. Collectively, these experiments demonstrate the involvement of BMP4-dependent pathways in the regulation of Equine trophoblast differentiation in vivo and primary trophoblast differentiation in vitro via activation of SMAD1/5 pathway, a previously unreported mechanism of TGFβ signaling in the mammalian Placenta.
P. D. Rossdale - One of the best experts on this subject based on the ideXlab platform.
-
Localisation of 15‐hydroxy prostaglandin dehydrogenase (PGDH) and steroidogenic enzymes in the Equine Placenta
Equine Veterinary Journal, 1995Co-Authors: X. Han, Marian Silver, Abigail L Fowden, Jennifer C. Ousey, P. D. Rossdale, W. R. Allen, Nicola Holdstock, A. J. Mcgladdery, Fernand Labrie, A. BelangerAbstract:15-hydroxy prostaglandin dehydrogenase (PGDH) is the critical enzyme that determines metabolism of primary prostaglandins. Its expression is determined in part by steroid hormones, particularly progesterone, formed from delta(5) steroids through 3beta-hydroxysteroid dehydrogenase (3beta-HSD) activity. To assess whether the regulation of PGDH might occur in a paracrine, autocrine or intracrine fashion, we used immunohistochemistry (IHC) to determine the localisation of key steroidogenic enzymes in the Equine Placenta and compared these patterns to the distribution of immunoreactive (IR-) PGDH. Placental tissue was obtained from pony or Thoroughbred mares at about Days 150, 250-280 and >300 of pregnancy (term 320 to 360 days; n=5-8 each group). IR-PGDH, 3beta-HSD, cholesterol side chain cleavage enzyme (P450(scc)) and 17-hydroxylase/lyase (P450(C17)) were localised using specific antibodies and the avidin-biotin peroxidase technique and visualised using diaminobenzidine as substrate. IR-P450(scc) was present in trophoblast cells, but not in maternal tissues of the microcotyledons. In contrast, at Days 150 and 280, IR-PGDH was present in maternal epithelial and interstitial cells in the microcotyledons, but was not detected in trophoblast epithelium, chorioallantois or endometrial glands. After Day 300, IR-PGDH was present in the maternal epithelium and interstitial cells of the Placenta and it was also present in trophoblast cells in some specimens.
-
localisation of 15 hydroxy prostaglandin dehydrogenase pgdh and steroidogenic enzymes in the Equine Placenta
Equine Veterinary Journal, 1995Co-Authors: X. Han, Marian Silver, Abigail L Fowden, P. D. Rossdale, W. R. Allen, Nicola Holdstock, A. J. Mcgladdery, Fernand Labrie, J C Ousey, A. BelangerAbstract:15-hydroxy prostaglandin dehydrogenase (PGDH) is the critical enzyme that determines metabolism of primary prostaglandins. Its expression is determined in part by steroid hormones, particularly progesterone, formed from delta(5) steroids through 3beta-hydroxysteroid dehydrogenase (3beta-HSD) activity. To assess whether the regulation of PGDH might occur in a paracrine, autocrine or intracrine fashion, we used immunohistochemistry (IHC) to determine the localisation of key steroidogenic enzymes in the Equine Placenta and compared these patterns to the distribution of immunoreactive (IR-) PGDH. Placental tissue was obtained from pony or Thoroughbred mares at about Days 150, 250-280 and >300 of pregnancy (term 320 to 360 days; n=5-8 each group). IR-PGDH, 3beta-HSD, cholesterol side chain cleavage enzyme (P450(scc)) and 17-hydroxylase/lyase (P450(C17)) were localised using specific antibodies and the avidin-biotin peroxidase technique and visualised using diaminobenzidine as substrate. IR-P450(scc) was present in trophoblast cells, but not in maternal tissues of the microcotyledons. In contrast, at Days 150 and 280, IR-PGDH was present in maternal epithelial and interstitial cells in the microcotyledons, but was not detected in trophoblast epithelium, chorioallantois or endometrial glands. After Day 300, IR-PGDH was present in the maternal epithelium and interstitial cells of the Placenta and it was also present in trophoblast cells in some specimens.
-
Modulation of 3β‐hydroxysteroid dehydrogenase (3β‐HSD) activity in the Equine Placenta by pregnenolone and progesterone metabolites
Equine Veterinary Journal, 1995Co-Authors: Pascale Chavatte, P. D. Rossdale, A. D. TaitAbstract:Summary The purpose of this study was to measure 3β-HSD activity in the Equine Placenta and to assess the effect of fetal and maternal blood plasma progestagens on 3β-HSD activity in vitro. 3β-HSD activity was measured in 8 late gestation (collected by caesarian section at 250 to 320 days) and 7 term (collected at birth) Equine Placentae using a tritium release assay with [3α-3H] pregnenolone as substrate. Mean ± s.d. Kmapp and Vmax for term Placentae were in general higher than for late gestation Placentae (0.129 ± 0.217 μmol/l and 23.85 ± 9.1 nmol/mg/h respectively vs. 0.016 ± 0.048 μmol/l and 17.36 ± 20.9 nmol/mg/h) but there was no statistical difference between them. Inhibition studies were performed on 3 term Placentae and 3 late gestation ones. Steroid concentrations used for inhibition studies were close to blood plasma concentrations (0.5 to 2 μmol/l). 3β-hydroxy compounds (5α-pregnane-3β, 20β-diol, 5α-pregnane-3β, 20α-diol and 3β-hydroxy-5α-pregnan-20-one) showed noncompetitive or mixed inhibition. Mean Kiapp ranged from 0.47 μmol/l to 1.55 μmol/l. Noncompetitive inhibition was found for 5-pregnene-3β,20β-diol with a mean Kiapp of 0.7 μmol/l. Inhibition was competitive with 20α-hydroxy-5α-pregnan-3-one with a mean Kiapp of 0.1 μmol/l. Progesterone was found to be a weak noncompetitive inhibitor of Equine 3β-HSD actitvity (mean Kiapp=1.2 umol/l) and no consistent inhibitory action was found with 5α-pregnane-3,20-dione. These data demonstrate that a marked inhibition of 315-HSD activity is caused by 5-pregnenes and 5α-pregnanes in the range of blood plasma concentrations in the pregnant mare. It is probable that a cumulative effect of these compounds could lead to a further decrease in the enzyme activity in the term Placenta, which may be of significance in the onset of parturition.
-
modulation of 3β hydroxysteroid dehydrogenase 3β hsd activity in the Equine Placenta by pregnenolone and progesterone metabolites
Equine Veterinary Journal, 1995Co-Authors: Pascale Chavatte, P. D. Rossdale, A. D. TaitAbstract:Summary The purpose of this study was to measure 3β-HSD activity in the Equine Placenta and to assess the effect of fetal and maternal blood plasma progestagens on 3β-HSD activity in vitro. 3β-HSD activity was measured in 8 late gestation (collected by caesarian section at 250 to 320 days) and 7 term (collected at birth) Equine Placentae using a tritium release assay with [3α-3H] pregnenolone as substrate. Mean ± s.d. Kmapp and Vmax for term Placentae were in general higher than for late gestation Placentae (0.129 ± 0.217 μmol/l and 23.85 ± 9.1 nmol/mg/h respectively vs. 0.016 ± 0.048 μmol/l and 17.36 ± 20.9 nmol/mg/h) but there was no statistical difference between them. Inhibition studies were performed on 3 term Placentae and 3 late gestation ones. Steroid concentrations used for inhibition studies were close to blood plasma concentrations (0.5 to 2 μmol/l). 3β-hydroxy compounds (5α-pregnane-3β, 20β-diol, 5α-pregnane-3β, 20α-diol and 3β-hydroxy-5α-pregnan-20-one) showed noncompetitive or mixed inhibition. Mean Kiapp ranged from 0.47 μmol/l to 1.55 μmol/l. Noncompetitive inhibition was found for 5-pregnene-3β,20β-diol with a mean Kiapp of 0.7 μmol/l. Inhibition was competitive with 20α-hydroxy-5α-pregnan-3-one with a mean Kiapp of 0.1 μmol/l. Progesterone was found to be a weak noncompetitive inhibitor of Equine 3β-HSD actitvity (mean Kiapp=1.2 umol/l) and no consistent inhibitory action was found with 5α-pregnane-3,20-dione. These data demonstrate that a marked inhibition of 315-HSD activity is caused by 5-pregnenes and 5α-pregnanes in the range of blood plasma concentrations in the pregnant mare. It is probable that a cumulative effect of these compounds could lead to a further decrease in the enzyme activity in the term Placenta, which may be of significance in the onset of parturition.
-
Release of lipid from the Equine Placenta during in vitro incubation
Placenta, 1994Co-Authors: J. P. Stammers, Abigail L Fowden, David Hull, M. Silver, Jennifer C. Ousey, P. D. RossdaleAbstract:Summary An in vitro incubation technique was used to examine release of lipids from the Equine Placenta. Placental tissue was obtained at term ( n =5, term=320–365 days) and earlier in gestation ( n =8, mean=266 days). Term Placentae were incubated at two temperatures, 4°C (control) and 37°C for 2 h. Pre-term Placentae were incubated at 37°C with two different concentrations of fatty acid in the medium. Tissues and media were analysed for their lipid concentrations. Term and pre-term Placentae released free fatty acid (FFA) and phospholipid into the incubation medium during incubation at 37°C. Long chain polyunsaturated fatty acids derived from the essential fatty acids were released into the media. The fatty acid profiles of the lipids released during incubation more closely resembled those of fetal plasma than maternal plasma lipids as measure in previous studies. These data are consistent with the view that the Equine Placenta is a source of both FFA and phospholipid for the fetus and that the Placenta may provide long chain polyunsaturated fatty acids for the fetal foal.
Victoria Cabrerasharp - One of the best experts on this subject based on the ideXlab platform.
-
smad1 5 signaling in the early Equine Placenta regulates trophoblast differentiation and chorionic gonadotropin secretion
Endocrinology, 2014Co-Authors: Victoria Cabrerasharp, Jordan E Read, Stephanie Richardson, Alycia A Kowalski, Douglas F Antczak, Judith E Cartwright, Abir Mukherjee, Amanda M De MestreAbstract:TGFβ superfamily proteins, acting via SMAD (Sma- and Mad-related protein)2/3 pathways, regulate Placental function; however, the role of SMAD1/5/8 pathway in the Placenta is unknown. This study investigated the functional role of bone morphogenetic protein (BMP)4 signaling through SMAD1/5 in terminal differentiation of primary chorionic gonadotropin (CG)-secreting trophoblast. Primary Equine trophoblast cells or Placental tissues were isolated from day 27–34 Equine conceptuses. Detected by microarray, RT-PCR, and quantitative RT-PCR, Equine chorionic girdle trophoblast showed increased gene expression of receptors that bind BMP4. BMP4 mRNA expression was 20- to 60-fold higher in Placental tissues adjacent to the chorionic girdle compared with chorionic girdle itself, suggesting BMP4 acts primarily in a paracrine manner on the chorionic girdle. Stimulation of chorionic girdle-trophoblast cells with BMP4 resulted in a dose-dependent and developmental stage-dependent increase in total number and proportion of terminally differentiated binucleate cells. Furthermore, BMP4 treatment induced non-CG-secreting day 31 chorionic girdle trophoblast cells to secrete CG, confirming a specific functional response to BMP4 stimulation. Inhibition of SMAD2/3 signaling combined with BMP4 treatment further enhanced differentiation of trophoblast cells. Phospho-SMAD1/5, but not phospho-SMAD2, expression as determined by Western blotting was tightly regulated during chorionic girdle trophoblast differentiation in vivo, with peak expression of phospho-SMAD1/5 in vivo noted at day 31 corresponding to maximal differentiation response of trophoblast in vitro. Collectively, these experiments demonstrate the involvement of BMP4-dependent pathways in the regulation of Equine trophoblast differentiation in vivo and primary trophoblast differentiation in vitro via activation of SMAD1/5 pathway, a previously unreported mechanism of TGFβ signaling in the mammalian Placenta.
Douglas F Antczak - One of the best experts on this subject based on the ideXlab platform.
-
smad1 5 signaling in the early Equine Placenta regulates trophoblast differentiation and chorionic gonadotropin secretion
Endocrinology, 2014Co-Authors: Victoria Cabrerasharp, Jordan E Read, Stephanie Richardson, Alycia A Kowalski, Douglas F Antczak, Judith E Cartwright, Abir Mukherjee, Amanda M De MestreAbstract:TGFβ superfamily proteins, acting via SMAD (Sma- and Mad-related protein)2/3 pathways, regulate Placental function; however, the role of SMAD1/5/8 pathway in the Placenta is unknown. This study investigated the functional role of bone morphogenetic protein (BMP)4 signaling through SMAD1/5 in terminal differentiation of primary chorionic gonadotropin (CG)-secreting trophoblast. Primary Equine trophoblast cells or Placental tissues were isolated from day 27–34 Equine conceptuses. Detected by microarray, RT-PCR, and quantitative RT-PCR, Equine chorionic girdle trophoblast showed increased gene expression of receptors that bind BMP4. BMP4 mRNA expression was 20- to 60-fold higher in Placental tissues adjacent to the chorionic girdle compared with chorionic girdle itself, suggesting BMP4 acts primarily in a paracrine manner on the chorionic girdle. Stimulation of chorionic girdle-trophoblast cells with BMP4 resulted in a dose-dependent and developmental stage-dependent increase in total number and proportion of terminally differentiated binucleate cells. Furthermore, BMP4 treatment induced non-CG-secreting day 31 chorionic girdle trophoblast cells to secrete CG, confirming a specific functional response to BMP4 stimulation. Inhibition of SMAD2/3 signaling combined with BMP4 treatment further enhanced differentiation of trophoblast cells. Phospho-SMAD1/5, but not phospho-SMAD2, expression as determined by Western blotting was tightly regulated during chorionic girdle trophoblast differentiation in vivo, with peak expression of phospho-SMAD1/5 in vivo noted at day 31 corresponding to maximal differentiation response of trophoblast in vitro. Collectively, these experiments demonstrate the involvement of BMP4-dependent pathways in the regulation of Equine trophoblast differentiation in vivo and primary trophoblast differentiation in vitro via activation of SMAD1/5 pathway, a previously unreported mechanism of TGFβ signaling in the mammalian Placenta.
-
SMAD1/5 signaling in the early Equine Placenta regulates trophoblast differentiation and chorionic gonadotropin secretion.
Endocrinology, 2014Co-Authors: Victoria Cabrera-sharp, Jordan E Read, Stephanie Richardson, Alycia A Kowalski, Douglas F Antczak, Judith E Cartwright, Abir Mukherjee, Amanda M De MestreAbstract:TGFβ superfamily proteins, acting via SMAD (Sma- and Mad-related protein)2/3 pathways, regulate Placental function; however, the role of SMAD1/5/8 pathway in the Placenta is unknown. This study investigated the functional role of bone morphogenetic protein (BMP)4 signaling through SMAD1/5 in terminal differentiation of primary chorionic gonadotropin (CG)-secreting trophoblast. Primary Equine trophoblast cells or Placental tissues were isolated from day 27–34 Equine conceptuses. Detected by microarray, RT-PCR, and quantitative RT-PCR, Equine chorionic girdle trophoblast showed increased gene expression of receptors that bind BMP4. BMP4 mRNA expression was 20- to 60-fold higher in Placental tissues adjacent to the chorionic girdle compared with chorionic girdle itself, suggesting BMP4 acts primarily in a paracrine manner on the chorionic girdle. Stimulation of chorionic girdle-trophoblast cells with BMP4 resulted in a dose-dependent and developmental stage-dependent increase in total number and proportion of terminally differentiated binucleate cells. Furthermore, BMP4 treatment induced non-CG-secreting day 31 chorionic girdle trophoblast cells to secrete CG, confirming a specific functional response to BMP4 stimulation. Inhibition of SMAD2/3 signaling combined with BMP4 treatment further enhanced differentiation of trophoblast cells. Phospho-SMAD1/5, but not phospho-SMAD2, expression as determined by Western blotting was tightly regulated during chorionic girdle trophoblast differentiation in vivo, with peak expression of phospho-SMAD1/5 in vivo noted at day 31 corresponding to maximal differentiation response of trophoblast in vitro. Collectively, these experiments demonstrate the involvement of BMP4-dependent pathways in the regulation of Equine trophoblast differentiation in vivo and primary trophoblast differentiation in vitro via activation of SMAD1/5 pathway, a previously unreported mechanism of TGFβ signaling in the mammalian Placenta.
-
IL-22 Is Expressed by the Invasive Trophoblast of the Equine (Equus caballus) Chorionic Girdle
Journal of Immunology, 2012Co-Authors: Margaret M. Brosnahan, Donald C. Miller, Mackenzie Adams, Douglas F AntczakAbstract:The invasive trophoblast cells of the Equine Placenta migrate into the endometrium to form endometrial cups, dense accumulations of trophoblast cells that produce Equine chorionic gonadotropin between days 40 and 120 of normal pregnancy. The mechanisms by which the trophoblast cells invade the endometrium while evading maternal immune destruction are poorly defined. A gene expression microarray analysis performed on Placental tissues obtained at day 34 of gestation revealed a >900-fold upregulation of mRNA encoding the cytokine IL-22 in chorionic girdle relative to noninvasive chorion. Quantitative RT-PCR assays were used to verify high expression of IL-22 in chorionic girdle. Additional quantitative RT-PCR analysis showed a striking increase in IL-22 mRNA expression in chorionic girdle from days 32 to 35 and an absence of IL-22 expression in other conceptus tissues. Bioinformatic analysis and cDNA sequencing confirmed the predicted length of horse IL-22, which carries a 3' extension absent in IL-22 genes of humans and mice, but present in the cow and pig. Our discovery of IL-22 in the chorionic girdle is a novel finding, as this cytokine has been previously reported in immune cells only. IL-22 has immunoregulatory functions, with primary action on epithelial cells. mRNA of IL-22R1 was detected in pregnant endometrium at levels similar to other Equine epithelia. Based upon these findings, we hypothesize that IL-22 cytokine produced by the chorionic girdle binds IL-22R1 on endometrium, serving as a mechanism of fetal-maternal communication by modulating endometrial responses to trophoblast invasion.
Alycia A Kowalski - One of the best experts on this subject based on the ideXlab platform.
-
smad1 5 signaling in the early Equine Placenta regulates trophoblast differentiation and chorionic gonadotropin secretion
Endocrinology, 2014Co-Authors: Victoria Cabrerasharp, Jordan E Read, Stephanie Richardson, Alycia A Kowalski, Douglas F Antczak, Judith E Cartwright, Abir Mukherjee, Amanda M De MestreAbstract:TGFβ superfamily proteins, acting via SMAD (Sma- and Mad-related protein)2/3 pathways, regulate Placental function; however, the role of SMAD1/5/8 pathway in the Placenta is unknown. This study investigated the functional role of bone morphogenetic protein (BMP)4 signaling through SMAD1/5 in terminal differentiation of primary chorionic gonadotropin (CG)-secreting trophoblast. Primary Equine trophoblast cells or Placental tissues were isolated from day 27–34 Equine conceptuses. Detected by microarray, RT-PCR, and quantitative RT-PCR, Equine chorionic girdle trophoblast showed increased gene expression of receptors that bind BMP4. BMP4 mRNA expression was 20- to 60-fold higher in Placental tissues adjacent to the chorionic girdle compared with chorionic girdle itself, suggesting BMP4 acts primarily in a paracrine manner on the chorionic girdle. Stimulation of chorionic girdle-trophoblast cells with BMP4 resulted in a dose-dependent and developmental stage-dependent increase in total number and proportion of terminally differentiated binucleate cells. Furthermore, BMP4 treatment induced non-CG-secreting day 31 chorionic girdle trophoblast cells to secrete CG, confirming a specific functional response to BMP4 stimulation. Inhibition of SMAD2/3 signaling combined with BMP4 treatment further enhanced differentiation of trophoblast cells. Phospho-SMAD1/5, but not phospho-SMAD2, expression as determined by Western blotting was tightly regulated during chorionic girdle trophoblast differentiation in vivo, with peak expression of phospho-SMAD1/5 in vivo noted at day 31 corresponding to maximal differentiation response of trophoblast in vitro. Collectively, these experiments demonstrate the involvement of BMP4-dependent pathways in the regulation of Equine trophoblast differentiation in vivo and primary trophoblast differentiation in vitro via activation of SMAD1/5 pathway, a previously unreported mechanism of TGFβ signaling in the mammalian Placenta.
-
SMAD1/5 signaling in the early Equine Placenta regulates trophoblast differentiation and chorionic gonadotropin secretion.
Endocrinology, 2014Co-Authors: Victoria Cabrera-sharp, Jordan E Read, Stephanie Richardson, Alycia A Kowalski, Douglas F Antczak, Judith E Cartwright, Abir Mukherjee, Amanda M De MestreAbstract:TGFβ superfamily proteins, acting via SMAD (Sma- and Mad-related protein)2/3 pathways, regulate Placental function; however, the role of SMAD1/5/8 pathway in the Placenta is unknown. This study investigated the functional role of bone morphogenetic protein (BMP)4 signaling through SMAD1/5 in terminal differentiation of primary chorionic gonadotropin (CG)-secreting trophoblast. Primary Equine trophoblast cells or Placental tissues were isolated from day 27–34 Equine conceptuses. Detected by microarray, RT-PCR, and quantitative RT-PCR, Equine chorionic girdle trophoblast showed increased gene expression of receptors that bind BMP4. BMP4 mRNA expression was 20- to 60-fold higher in Placental tissues adjacent to the chorionic girdle compared with chorionic girdle itself, suggesting BMP4 acts primarily in a paracrine manner on the chorionic girdle. Stimulation of chorionic girdle-trophoblast cells with BMP4 resulted in a dose-dependent and developmental stage-dependent increase in total number and proportion of terminally differentiated binucleate cells. Furthermore, BMP4 treatment induced non-CG-secreting day 31 chorionic girdle trophoblast cells to secrete CG, confirming a specific functional response to BMP4 stimulation. Inhibition of SMAD2/3 signaling combined with BMP4 treatment further enhanced differentiation of trophoblast cells. Phospho-SMAD1/5, but not phospho-SMAD2, expression as determined by Western blotting was tightly regulated during chorionic girdle trophoblast differentiation in vivo, with peak expression of phospho-SMAD1/5 in vivo noted at day 31 corresponding to maximal differentiation response of trophoblast in vitro. Collectively, these experiments demonstrate the involvement of BMP4-dependent pathways in the regulation of Equine trophoblast differentiation in vivo and primary trophoblast differentiation in vitro via activation of SMAD1/5 pathway, a previously unreported mechanism of TGFβ signaling in the mammalian Placenta.