Effects of maternal stress and obesity on human feto-placental glucocorticoid exposure
O'Reilly, James Richard
Fetal exposure to excess glucocorticoids has been proposed as a key determinant of pregnancy outcome, as well as a predictor of long term health of the offspring through a phenomenon known as ‘developmental programming’. Obesity and ‘stress’ during pregnancy are two potential sources of altered fetal exposure to glucocorticoids. One in five pregnant women is obese at antenatal booking, and maternal obesity increases risk of offspring complications including higher birth weight, potentially leading to long-term programming effects on the offspring. Likewise, maternal anxiety during pregnancy has been identified as a programming factor, increasing the risk of psychopathology in the offspring. This thesis tests the hypothesis that in humans this association is mediated by altered action of glucocorticoids, by examining circulating levels of maternal glucocorticoids during pregnancy and through measurement of key genes in the placenta regulating fetal glucocorticoid exposure. Serum cortisol levels were measured at 16, 28 and 36 weeks gestation in n=173 class III obese (BMI 44.0±4.5kg/m2) and n=107 lean (BMI 22.8±1.6kg/m2) pregnant women. Serial corticosteroid binding globulin (CBG) concentrations were measured in a subset (n=39 lean, 26 obese) and free cortisol levels calculated using Coolen’s equation. CRH concentrations were measured at the same time points in obese (n=20) and lean (n=22) pregnant women Salivary cortisol was measured in samples collected at bed-time, waking and 30 minutes after waking. mRNA levels of candidate genes regulating glucocorticoids and fetal/placental growth including 11-beta hydroxysteroid dehydrogenase type 2 (11βHSD2), which inactivates cortisol, insulin-like growth factor 2 (IGF2) and glucocorticoid receptor (GR) were measured in first trimester (n=32), second trimester (n=15) and term (n=60) placental samples. DNA methylation of key regions controlling the expression of the IGF2, GR and 11βHSD2 genes was measured by pyrosequencing in first trimester and term samples. Levels of mRNAs encoding 11βHSD1, 11βHSD2, GR and MR were measured in term placentas collected from women from Helsinki, Finland in whom anxiety during pregnancy had been prospectively assessed using validated questionnaires. Term placental samples from a subset of the obese and lean women who had also completed stress questionnaires during pregnancy were used to examine replication of findings. Cortisol levels rose similarly during pregnancy in obese and lean but were significantly lower throughout pregnancy in obese women (p<0.05). The diurnal rhythm of cortisol was maintained. CBG levels also increased, though this change was lower in obese (1.21-fold (±0.9) vs 1.56-fold (±0.07), p<0.01). In obese women, lower calculated free cortisol at 16 weeks gestation was associated with higher birth weight after adjustment for other factors (r=-0.46, p<0.05). Placental mRNA encoding 11βHSD2 increased in association with increasing obesity in early pregnancy (r=0.44, p<0.01) and was highest in term placenta in obese women with macrosomic (>4000g) offspring (p<0.05). Placental transcript abundance of GR also increased in association with increasing obesity in early pregnancy (r=0.38, p<0.05), but was lowest in term placenta from obese with macrosomic offspring (p<0.05). IGF2 mRNA abundance was lower in the placentas of obese women with macrosomic offspring at term compared to both lean women and obese women with normal weight offspring (p<0.01). Methylation results are reported. Placental mRNA levels encoding 11βHSD1 (which converts inactive cortisone to active cortisol) at term was found to positively associate with maternal anxiety measured in the first trimester of pregnancy in a group of pregnant Finnish women (β=0.3, p<0.05). Findings were similar in the replication sample in lean women only (β=4.6, p<0.05). Lower circulating and bioavailable cortisol levels in early pregnancy, together with a greater placental ‘barrier’ to maternal glucocorticoids represent key mechanisms contributing to higher birth weight in offspring of obese women. Regeneration of active glucocorticoids in placenta and increasing placental sensitivity to glucocorticoids increases fetal glucocorticoid exposure and offers insight into the biological mechanisms underlying adverse offspring effects of maternal prenatal anxiety.