Placental Proteomics Reveal Insights into Fetal Alcohol Spectrum Disorders.

BACKGROUND: Fetal alcohol spectrum disorders (FASD) describe many of the well-known neurodevelopmental deficits afflicting children exposed to alcohol in utero. The effects of alcohol on the maternal-fetal interface, especially the placenta, have been less explored. We herein hypothesized that chronic binge alcohol exposure during pregnancy significantly alters the placental protein profile in a rat FASD model. METHODS: Pregnant rats were orogastrically treated daily with alcohol (4.5 g/kg, gestational day [GD] 5 to 10; 6.0 g/kg, GD 11 to 19) or 50% maltose dextrin (isocalorically matched pair-fed controls). On GD 20, placentae were collected, flash-frozen, and stored until tissues were homogenized. Protein lysates were denatured, reduced, captured on a 10-kDa spin filter, and digested. Peptides were eluted, reconstituted, and analyzed by a Q Exactive™ Hybrid Quadrupole-Orbitrap™ mass spectrometer. RESULTS: Mass spectrometry (MS) analysis identified 2,285 placental proteins based on normalized spectral counts and 2,000 proteins by intensity-based absolute quantification. Forty-five placental proteins were significantly (p < 0.05) altered by gestational alcohol exposure by both quantification approaches. These included proteins directly related to alcohol metabolism; specific isoforms of alcohol dehydrogenase and aldehyde dehydrogenase were up-regulated in the alcohol group. Ingenuity analysis identified ethanol degradation as the most significantly altered canonical pathway in placenta, and fetal/organ development as most altered function, with increased risk for metabolic, neurological, and cardiovascular diseases. Physiological roles of the significantly altered proteins were related to early pregnancy adaptations, implantation, gestational diseases, fetal organ development, neurodevelopment, and immune functions. CONCLUSIONS: We conclude that the placenta is a valuable organ not only to understand FASD etiology but it may also serve as a diagnostic tool to identify novel biomarkers for detecting the outcome of fetal alcohol exposure. Placental MS analysis can offer sophisticated insights into identifying alcohol metabolism-related enzymes and regulators of fetal development.

Fetal regional brain protein signature in FASD rat model.

Fetal alcohol spectrum disorders (FASD) describe neurodevelopmental deficits in children exposed to alcohol in utero. We hypothesized that gestational alcohol significantly alters fetal brain regional protein signature. Pregnant rats were binge-treated with alcohol or pair-fed and nutritionally-controlled. Mass spectrometry identified 1806, 2077, and 1456 quantifiable proteins in the fetal hippocampus, cortex, and cerebellum, respectively. A stronger effect of alcohol exposure on the hippocampal proteome was noted: over 600 hippocampal proteins were significantly (P < .05) altered, including annexin A2, nucleobindin-1, and glypican-4, regulators of cellular growth and developmental morphogenesis. In the cerebellum, cadherin-13, reticulocalbin-2, and ankyrin-2 (axonal growth regulators) were significantly (P < .05) altered; altered cortical proteins were involved in autophagy (endophilin-B1, synaptotagmin-1). Ingenuity analysis identified proteins involved in protein homeostasis, oxidative stress, mitochondrial dysfunction, and mTOR as major pathways in the cortex and hippocampus significantly (P < .05) affected by alcohol. Thus, neurodevelopmental protein changes may directly relate to FASD neuropathology.

Chronic binge alcohol consumption during pregnancy alters rat maternal uterine artery pressure response.

We aimed to investigate pressure-dependent maternal uterine artery responses and vessel remodeling following gestational binge alcohol exposure. Two groups of pregnant rats were used: the alcohol group (28.5% wt/v, 6.0 g/kg, once-daily orogastric gavage in a binge paradigm between gestational day (GD) 5-19) and pair-fed controls (isocalorically matched). On GD20, excised, pressurized primary uterine arteries were studied following equilibration (60 mm Hg) using dual chamber arteriograph. The uterine artery diameter stabilized at 20 mm Hg, showed passive distension at 40 mm Hg, and redeveloped tone at 60 mm Hg. An alcohol effect (P = 0.0025) was observed on the percent constriction of vessel diameter with greater pressure-dependent myogenic constriction. Similar alcohol effect was noted with lumen diameter response (P = 0.0020). The percent change in media:lumen ratio was higher in the alcohol group (P < 0.0001). Thus, gestational alcohol affects pressure-induced uterine artery reactivity, inward-hypotrophic remodeling, and adaptations critical for nutrient delivery to the fetus.