title: Long-acting progestin-only contraceptives enhance human endometrial stromal cell expressed neuronal pentraxin-1 and reactive oxygen species to promote endothelial cell apoptosis.
abstract: BACKGROUND
  Despite the absence of progesterone receptor protein in human endometrial endothelial cells (HEECs), endometria of women receiving long-acting progestin-only contraceptives (LAPCs) display reduced uterine blood flow, elevated reactive oxygen species generation, increased angiogenesis, and irregularly distributed, enlarged, fragile microvessels resulting in abnormal uterine bleeding.
  
  
  OBJECTIVE
  We propose that paracrine factors from LAPC-treated human endometrial stromal cells (HESCs) impair HEEC functions by shifting the balance between HEEC viability and death in favor of the latter.
  
  
  METHODS
  Proliferation, apoptosis, and transcriptome analyses were performed in HEECs treated with conditioned medium supernatant (CMS) derived from HESCs treated with estradiol (E2) ± medroxyprogesterone acetate or etonogestrel under normoxia or hypoxia. Mass spectrometry interrogated the CMS secretome while immunostaining for neuronal pentraxin-1 (NPTX1), cleaved caspase-3, and cytochrome c was performed in cultured HEECs and paired endometria from women using LAPCs.
  
  
  RESULTS
  HEEC apoptosis and its underlying mechanism.
  
  
  RESULTS
  HESC CMS from E2 + medroxyprogesterone acetate or E2 + etonogestrel incubations under hypoxia induced HEEC apoptosis (P < .05), whereas mass spectrometry of the CMS revealed increased NPTX1 secretion (P < .05). Endothelial cleaved caspase-3 and stromal NPTX1 immunoreactivity were significantly higher in LAPC-treated endometria (P < .001). Transcriptomics revealed AKT signaling inhibition and mitochondrial dysfunction in HEECs incubated with HESC CMS. In vitro analyses proved that CMS decreased HEEC AKT phosphorylation (P < .05) and that recombinant NPTX1 (P < .05) or NPTX1 + H2O2 (P < .001) increase HEEC apoptosis and cytosolic cytochrome c levels.
  
  
  CONCLUSIONS
  LAPC-enhanced NPTX1 secretion and reactive oxygen species generation in HESCs impair HEEC survival resulting in a loss in vascular integrity, demonstrating a novel paracrine mechanism to explain LAPC-induced abnormal uterine bleeding.
authors.affiliation: Department of Obstetrics and Gynecology, the Ohio State University College of Medicine, Columbus, Ohio 43210.
authors.forename: O
authors.initials: O
authors.lastname: Guzeloglu-Kayisli
authors.forename: M
authors.initials: M
authors.lastname: Basar
authors.forename: J P
authors.initials: JP
authors.lastname: Shapiro
authors.forename: N
authors.initials: N
authors.lastname: Semerci
authors.forename: J S
authors.initials: JS
authors.lastname: Huang
authors.forename: F
authors.initials: F
authors.lastname: Schatz
authors.forename: C J
authors.initials: CJ
authors.lastname: Lockwood
authors.forename: U A
authors.initials: UA
authors.lastname: Kayisli
mesh_terms: D017209:Apoptosis / Q000187:drug effects*; D002097:C-Reactive Protein / Q000378:metabolism*; D053148:Caspase 3 / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D003271:Contraceptive Agents, Female / Q000008:administration & dosage* / Q000009:adverse effects; D017077:Culture Media, Conditioned / Q000494:pharmacology; D045304:Cytochromes c / Q000378:metabolism; D004717:Endometrium / Q000166:cytology / Q000187:drug effects*; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects; D004958:Estradiol / Q000009:adverse effects / Q000494:pharmacology; D005260:Female; D006801:Humans; D017258:Medroxyprogesterone Acetate / Q000009:adverse effects / Q000494:pharmacology; D055806:Microvessels / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D019899:Paracrine Communication / Q000187:drug effects / Q000502:physiology; D011372:Progestins / Q000008:administration & dosage* / Q000009:adverse effects; D017382:Reactive Oxygen Species / Q000378:metabolism; D017154:Stromal Cells / Q000187:drug effects* / Q000378:metabolism; D000099153:Pentraxins
chemical_list: D002097:C-Reactive Protein; D053148:Caspase 3; D003271:Contraceptive Agents, Female; D017077:Culture Media, Conditioned; D045304:Cytochromes c; D004958:Estradiol; D017258:Medroxyprogesterone Acetate; D009419:Nerve Tissue Proteins; D011372:Progestins; D017382:Reactive Oxygen Species; D000099153:Pentraxins; C505407:CASP3 protein, human