Hypoxia downregulates p53 but induces apoptosis and enhances expression of BAD in cultures of human syncytiotrophoblasts.
Hypoxia is commonly assigned a role in the placental dysfunction characteristic of preeclampsia and intrauterine growth restriction. We previously showed that hypoxia upregulates p53 and enhances apoptosis in primary cultures of human cytotrophoblasts. Here we tested the hypothesis that hypoxia also induces apoptosis in syncytiotrophoblasts by upregulation of p53. Primary cultures of human cytotrophoblasts that had differentiated into syncytiotrophoblasts by 52 h were exposed for ≤24 h to 20% or <1% oxygen in the presence or absence of staurosporine or the p53 modulators nutlin-3, pifithrin-α, and pifithrin-μ. Proteins were detected by Western blot analysis or immunofluorescence. Compared with 20% oxygen, exposure of syncytiotrophoblasts to <1% oxygen upregulated hypoxia-inducible factor (HIF)-1α and rapidly downregulated p53. Activity of p53 in hypoxic syncytiotrophoblasts was reduced by the higher expression of the negative p53 regulator MDMX and by the reduction of phosphorylation of p53 at Ser(392), which reduces p53 activity. Conversely, staurosporine, a kinase inhibitor, and nutlin-3, a drug that enhances p53 expression, both raised p53 levels and increased the rate of apoptosis in syncytiotrophoblasts compared with vehicle controls. Immunofluorescence staining showed p53 immunolocalized to both cytoplasm and nuclei of nutlin-3-exposed syncytiotrophoblasts. The hypoxia-induced apoptosis in syncytiotrophoblasts correlated with enhanced expression of the proapoptotic BAD and a reduced level of antiapoptotic BAD phosphorylated on Ser(112). We surmise that cell death induced by extreme hypoxia in syncytiotrophoblasts follows a non-p53-dependent pathway, unlike that of a nonhypoxic stimulus and unlike hypoxic cytotrophoblasts. We speculate that downregulation of p53 activity in response to hypoxia reduces or eliminates the apoptosis transduced by the p53 pathway in syncytiotrophoblasts, thereby limiting cell death and maintaining the integrity of this critical villous component.
Chen
Baosheng
B
Department of Obstetrics and Gynecology, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
Longtine
Mark S
MS
Sadovsky
Yoel
Y
Nelson
D Michael
DM
D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D052160:Benzothiazoles / Q000494:pharmacology; D002478:Cells, Cultured; D015536:Down-Regulation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D005786:Gene Expression Regulation; D006801:Humans; D000860:Hypoxia / Q000378:metabolism*; D007093:Imidazoles / Q000494:pharmacology; D010100:Oxygen / Q000378:metabolism; D010879:Piperazines / Q000494:pharmacology; D011247:Pregnancy; D019311:Staurosporine / Q000494:pharmacology; D014050:Toluene / Q000031:analogs & derivatives / Q000494:pharmacology; D014327:Trophoblasts / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism*; D051025:bcl-Associated Death Protein / Q000235:genetics / Q000378:metabolism*
D052160:Benzothiazoles; D004791:Enzyme Inhibitors; D007093:Imidazoles; D010879:Piperazines; D016159:Tumor Suppressor Protein p53; D051025:bcl-Associated Death Protein; D014050:Toluene; C482205:nutlin 3; C121565:pifithrin; D019311:Staurosporine; D010100:Oxygen
Cancer Res. 2000 Oct 15;60(20):5630-4
11059752
J Biol Chem. 2010 Jan 22;285(4):2601-9
19880525
Br Med J (Clin Res Ed). 1987 Apr 25;294(6579):1051-3
3107690
Cancer Res. 2007 Sep 15;67(18):8810-7
17875722
Am J Obstet Gynecol. 2004 Aug;191(2):563-71
15343238
Neoplasia. 2008 Jun;10(6):587-96
18516295
J Biol Chem. 2003 May 2;278(18):15465-8
12637507
Cell Death Differ. 2006 Jun;13(6):927-34
16543935
J Biol Chem. 2006 Feb 3;281(5):2764-72
16314423
Proc Natl Acad Sci U S A. 2007 Mar 6;104(10):4054-9
17360476
Proc Natl Acad Sci U S A. 2002 Jan 8;99(1):95-100
11756653
Biochemistry. 1997 Aug 19;36(33):10117-24
9254608
Cell. 1996 Nov 15;87(4):619-28
8929531
Development. 1991 Nov;113(3):767-77
1821848
Proc Natl Acad Sci U S A. 2002 Aug 6;99(16):10305-9
12124396
J Biol Chem. 2009 Mar 27;284(13):8812-21
19153082
J Biol Chem. 2003 Apr 18;278(16):13595-8
12606552
Placenta. 2008 Sep;29(9):772-83
18706691
Am J Physiol Cell Physiol. 2000 May;278(5):C982-8
10794672
Mol Hum Reprod. 2007 Feb;13(2):111-6
17158854
Am J Pathol. 2007 Aug;171(2):496-506
17600131
Science. 2004 Feb 6;303(5659):844-8
14704432
J Pathol. 2010 Jan;220(2):164-73
19918835
N Engl J Med. 1993 Mar 11;328(10):692-6
8433728
Am J Obstet Gynecol. 2002 May;186(5):1056-61
12015537
Endocrinology. 2008 May;149(5):2131-7
18276761
Mol Cell. 2001 Feb;7(2):283-92
11239457
Placenta. 2009 Jun;30(6):473-82
19375795
Biochem Biophys Res Commun. 2005 Jun 10;331(3):750-60
15865931
Endocrinology. 1986 Apr;118(4):1567-82
3512258
J Cell Mol Med. 2004 Oct-Dec;8(4):423-31
15601571
Placenta. 2006 Sep-Oct;27(9-10):978-83
16376985
Carcinogenesis. 2006 Mar;27(3):656-63
16258175
Respir Physiol. 1995 May;100(2):101-6
7624611
Cell Death Differ. 2010 Jan;17(1):86-92
19543236
Blood. 2008 Nov 1;112(9):3827-34
18682598
Am J Obstet Gynecol. 2001 Apr;184(5):998-1003
11303211
Curr Mol Med. 2004 Nov;4(7):741-51
15579021
Cell Prolif. 2005 Aug;38(4):257-67
16098184
Trends Cell Biol. 2010 May;20(5):299-309
20172729
Nat Rev Cancer. 2009 Oct;9(10):691-700
19759539
Am J Pathol. 2007 May;170(5):1511-20
17456758
Am J Obstet Gynecol. 1999 Apr;180(4):896-902
10203658
Mol Cell. 2001 Mar;7(3):683-94
11463392
Science. 2003 Nov 7;302(5647):1036-8
14500851
Nature. 1997 May 15;387(6630):296-9
9153395
Nat Chem Biol. 2006 Sep;2(9):474-9
16862141
Placenta. 2001 Nov;22(10):822-30
11718569
Cell. 2009 May 15;137(4):609-22
19450511
Placenta. 2009 Mar;30 Suppl A:S43-8
19081132