Apoptosis of alcohol-exposed human placental cytotrophoblast cells is downstream of intracellular calcium signaling

Jay M. Bolnick, Rita Karana, Po J. Chiang, Brian A. Kilburn, Roberto Romero, Michael Peter Diamond, Susan M. Smith, D. Randall Armant

Research output: Contribution to journalArticle

15 Citations (Scopus)

Abstract

Background: Apoptosis is induced by ethanol (EtOH) in human placental trophoblast cells, possibly disrupting placentation and contributing to intrauterine growth restriction in fetal alcohol spectrum disorder (FASD). EtOH induces programmed cell death in several embryonic tissues by raising intracellular Ca2+. Therefore, the role of Ca2+ signaling in EtOH-induced apoptosis was examined using human first trimester cytotrophoblast cell lines, examining the hypothesis that apoptosis is dependent on intracellular Ca2+ signaling. Methods: Using HTR-8/SVneo and SW.71 cytotrophoblast cell lines, real-time intracellular Ca2+ concentration was monitored by fluo-4 epifluorescence microscopy and apoptosis was assessed by flow cytometry of cells fluorescently labeled for DNA fragmentation (TUNEL) and annexin V binding. Results: Intracellular Ca2+ concentrations increased synchronously in all cells within 10 seconds of exposure to 50 mM EtOH, but not at lower EtOH concentrations (10 to 25 mM) incapable of inducing apoptosis. Trophoblast cells treated with inhibitors of Ca2+ signaling (BAPTA-AM, U73122, xestospongin D, BAPTA, SKF-96365) produced no intracellular Ca2+ transients after exposure to 50 mM EtOH and were protected from cell death induced by EtOH. Conclusions: EtOH-induced apoptosis in human cytotrophoblast cells, identified by DNA fragmentation and externalized phosphatidylserine, was dependent upon Ca2+ signaling. Both intracellular Ca2+ mobilization and extracellular Ca2+ influx were required, as well as phosphatidylinositol signaling. Inhibition by SKF-96365 suggests that the capacitative Ca2+ entry mechanism that utilizes TRPC channels was activated by EtOH. Apoptosis occurs downstream of Ca2+ signaling in trophoblasts and may contribute to placental insufficiency and poor fetal growth associated with FASD.

Original languageEnglish (US)
Pages (from-to)1646-1653
Number of pages8
JournalAlcoholism: Clinical and Experimental Research
Volume38
Issue number6
DOIs
StatePublished - Jan 1 2014

Fingerprint

Calcium Signaling
Trophoblasts
Alcohols
1-(2-(3-(4-methoxyphenyl)propoxy)-4-methoxyphenylethyl)-1H-imidazole
Apoptosis
Calcium
Fetal Alcohol Spectrum Disorders
DNA Fragmentation
Cell death
Cell Death
Cells
Placental Insufficiency
Cell Line
Placentation
Flow cytometry
Annexin A5
Phosphatidylserines
DNA
In Situ Nick-End Labeling
First Pregnancy Trimester

Keywords

  • Alcohol
  • Apoptosis
  • Calcium
  • Fetal Alcohol Spectrum Disorder
  • Intracellular Signaling
  • Intrauterine Growth Restriction
  • Trophoblast

ASJC Scopus subject areas

  • Medicine (miscellaneous)
  • Psychiatry and Mental health
  • Toxicology
  • Medicine(all)

Cite this

Apoptosis of alcohol-exposed human placental cytotrophoblast cells is downstream of intracellular calcium signaling. / Bolnick, Jay M.; Karana, Rita; Chiang, Po J.; Kilburn, Brian A.; Romero, Roberto; Diamond, Michael Peter; Smith, Susan M.; Armant, D. Randall.

In: Alcoholism: Clinical and Experimental Research, Vol. 38, No. 6, 01.01.2014, p. 1646-1653.

Research output: Contribution to journalArticle

Bolnick, Jay M. ; Karana, Rita ; Chiang, Po J. ; Kilburn, Brian A. ; Romero, Roberto ; Diamond, Michael Peter ; Smith, Susan M. ; Armant, D. Randall. / Apoptosis of alcohol-exposed human placental cytotrophoblast cells is downstream of intracellular calcium signaling. In: Alcoholism: Clinical and Experimental Research. 2014 ; Vol. 38, No. 6. pp. 1646-1653.
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abstract = "Background: Apoptosis is induced by ethanol (EtOH) in human placental trophoblast cells, possibly disrupting placentation and contributing to intrauterine growth restriction in fetal alcohol spectrum disorder (FASD). EtOH induces programmed cell death in several embryonic tissues by raising intracellular Ca2+. Therefore, the role of Ca2+ signaling in EtOH-induced apoptosis was examined using human first trimester cytotrophoblast cell lines, examining the hypothesis that apoptosis is dependent on intracellular Ca2+ signaling. Methods: Using HTR-8/SVneo and SW.71 cytotrophoblast cell lines, real-time intracellular Ca2+ concentration was monitored by fluo-4 epifluorescence microscopy and apoptosis was assessed by flow cytometry of cells fluorescently labeled for DNA fragmentation (TUNEL) and annexin V binding. Results: Intracellular Ca2+ concentrations increased synchronously in all cells within 10 seconds of exposure to 50 mM EtOH, but not at lower EtOH concentrations (10 to 25 mM) incapable of inducing apoptosis. Trophoblast cells treated with inhibitors of Ca2+ signaling (BAPTA-AM, U73122, xestospongin D, BAPTA, SKF-96365) produced no intracellular Ca2+ transients after exposure to 50 mM EtOH and were protected from cell death induced by EtOH. Conclusions: EtOH-induced apoptosis in human cytotrophoblast cells, identified by DNA fragmentation and externalized phosphatidylserine, was dependent upon Ca2+ signaling. Both intracellular Ca2+ mobilization and extracellular Ca2+ influx were required, as well as phosphatidylinositol signaling. Inhibition by SKF-96365 suggests that the capacitative Ca2+ entry mechanism that utilizes TRPC channels was activated by EtOH. Apoptosis occurs downstream of Ca2+ signaling in trophoblasts and may contribute to placental insufficiency and poor fetal growth associated with FASD.",
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T1 - Apoptosis of alcohol-exposed human placental cytotrophoblast cells is downstream of intracellular calcium signaling

AU - Bolnick, Jay M.

AU - Karana, Rita

AU - Chiang, Po J.

AU - Kilburn, Brian A.

AU - Romero, Roberto

AU - Diamond, Michael Peter

AU - Smith, Susan M.

AU - Armant, D. Randall

PY - 2014/1/1

Y1 - 2014/1/1

N2 - Background: Apoptosis is induced by ethanol (EtOH) in human placental trophoblast cells, possibly disrupting placentation and contributing to intrauterine growth restriction in fetal alcohol spectrum disorder (FASD). EtOH induces programmed cell death in several embryonic tissues by raising intracellular Ca2+. Therefore, the role of Ca2+ signaling in EtOH-induced apoptosis was examined using human first trimester cytotrophoblast cell lines, examining the hypothesis that apoptosis is dependent on intracellular Ca2+ signaling. Methods: Using HTR-8/SVneo and SW.71 cytotrophoblast cell lines, real-time intracellular Ca2+ concentration was monitored by fluo-4 epifluorescence microscopy and apoptosis was assessed by flow cytometry of cells fluorescently labeled for DNA fragmentation (TUNEL) and annexin V binding. Results: Intracellular Ca2+ concentrations increased synchronously in all cells within 10 seconds of exposure to 50 mM EtOH, but not at lower EtOH concentrations (10 to 25 mM) incapable of inducing apoptosis. Trophoblast cells treated with inhibitors of Ca2+ signaling (BAPTA-AM, U73122, xestospongin D, BAPTA, SKF-96365) produced no intracellular Ca2+ transients after exposure to 50 mM EtOH and were protected from cell death induced by EtOH. Conclusions: EtOH-induced apoptosis in human cytotrophoblast cells, identified by DNA fragmentation and externalized phosphatidylserine, was dependent upon Ca2+ signaling. Both intracellular Ca2+ mobilization and extracellular Ca2+ influx were required, as well as phosphatidylinositol signaling. Inhibition by SKF-96365 suggests that the capacitative Ca2+ entry mechanism that utilizes TRPC channels was activated by EtOH. Apoptosis occurs downstream of Ca2+ signaling in trophoblasts and may contribute to placental insufficiency and poor fetal growth associated with FASD.

AB - Background: Apoptosis is induced by ethanol (EtOH) in human placental trophoblast cells, possibly disrupting placentation and contributing to intrauterine growth restriction in fetal alcohol spectrum disorder (FASD). EtOH induces programmed cell death in several embryonic tissues by raising intracellular Ca2+. Therefore, the role of Ca2+ signaling in EtOH-induced apoptosis was examined using human first trimester cytotrophoblast cell lines, examining the hypothesis that apoptosis is dependent on intracellular Ca2+ signaling. Methods: Using HTR-8/SVneo and SW.71 cytotrophoblast cell lines, real-time intracellular Ca2+ concentration was monitored by fluo-4 epifluorescence microscopy and apoptosis was assessed by flow cytometry of cells fluorescently labeled for DNA fragmentation (TUNEL) and annexin V binding. Results: Intracellular Ca2+ concentrations increased synchronously in all cells within 10 seconds of exposure to 50 mM EtOH, but not at lower EtOH concentrations (10 to 25 mM) incapable of inducing apoptosis. Trophoblast cells treated with inhibitors of Ca2+ signaling (BAPTA-AM, U73122, xestospongin D, BAPTA, SKF-96365) produced no intracellular Ca2+ transients after exposure to 50 mM EtOH and were protected from cell death induced by EtOH. Conclusions: EtOH-induced apoptosis in human cytotrophoblast cells, identified by DNA fragmentation and externalized phosphatidylserine, was dependent upon Ca2+ signaling. Both intracellular Ca2+ mobilization and extracellular Ca2+ influx were required, as well as phosphatidylinositol signaling. Inhibition by SKF-96365 suggests that the capacitative Ca2+ entry mechanism that utilizes TRPC channels was activated by EtOH. Apoptosis occurs downstream of Ca2+ signaling in trophoblasts and may contribute to placental insufficiency and poor fetal growth associated with FASD.

KW - Alcohol

KW - Apoptosis

KW - Calcium

KW - Fetal Alcohol Spectrum Disorder

KW - Intracellular Signaling

KW - Intrauterine Growth Restriction

KW - Trophoblast

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