Prenatal Maternal Stress Causes Preterm Birth and Affects Neonatal Adaptive Immunity in Mice

Abstract

Maternal stress is a well-established risk factor for preterm birth and has been associated with adverse neonatal outcomes in the first and subsequent generations, including increased susceptibility to disease and lasting immunological changes. However, a causal link between prenatal maternal stress and preterm birth, as well as compromised neonatal immunity, has yet to be established. To fill this gap in knowledge, we used a murine model of prenatal maternal stress across three generations and high-dimensional flow cytometry to evaluate neonatal adaptive immunity. We report that recurrent prenatal maternal stress induced preterm birth in the first and second filial generations and negatively impacted early neonatal growth. Strikingly, prenatal maternal stress induced a systematic reduction in T cells and B cells, the former including regulatory CD4+ T cells as well as IL-4- and IL-17A-producing T cells, in the second generation. Yet, neonatal adaptive immunity gained resilience against prenatal maternal stress by the third generation. We also show that the rate of prenatal maternal stress-induced preterm birth can be reduced upon cessation of stress, though neonatal growth impairments persisted. These findings provide evidence that prenatal maternal stress causes preterm birth and affects neonatal immunity across generations, adverse effects that can be ameliorated upon cessation.

Keywords: T cells; birthweight; neonates; offspring; preterm labor.

Figure 1

Maternal outcomes of prenatal maternal stress across generations. (A) Experimental design of alternating, unpredictable stress procedure. (B) Maternal corticosterone levels across generations 1 week after delivery (control n = 13; F0-S n = 14; F1-SS n = 12; F2-SSS n = 11). (C) Rates of delivery in each generation, specified as preterm, early, and term delivery (control n = 13; F0-S n = 23; F1-SS n = 27; F2-SSS n = 32). (D) Maternal weight gain across generations (control n = 13; F0-S n = 21; F1-SS n = 27; F2-SSS n = 32). (E) Number of pups per dam across generations (control n = 15; F0-S n = 25; F1-SS n = 27; F2-SSS n = 36). For box plots, mid-lines indicate medians, boxes indicate interquartile ranges, and whiskers indicate min-max range.

Figure 2

Neonatal outcomes of prenatal maternal stress across generations. (A) Experimental design of neonates born to stressed dams across generations. (B) Rate of neonatal mortality at birth in each generation (control n = 10; F1-S n = 19; F2-SS n = 19; F3-SSS n = 30). (C) Growth trajectory of neonates in the first 3 weeks after birth (control n = 5 litters; F1-S n = 8 litters; F2-SS n = 2 litters; F3-SSS n = 21 litters). Data are presented as mean ± standard error of the mean.

Figure 3

Immunophenotyping of naïve, memory, and effector T cells in prenatally stressed neonates. (A) Gating strategy used to identify T cell subsets. (B–D) Number of naïve T cells, (E–G) memory T cells, and (H–J) effector T cells (control n = 19; F2-SS n = 21; F3-SSS n = 9). Mid-lines indicate medians, boxes indicate interquartile ranges, and whiskers indicate min–max range.

Figure 4

Immunophenotyping of T cell subsets in prenatally stressed neonates. (A) Gating strategy used to identify T cell subsets. (B) Number of conventional T cells, (C) CD4+ T cells, (D) CD8+ T cells, (E) regulatory CD4+ T cells or CD4+ Tregs, (F) CD8+FoxP3+ T cells, (G) Th1 cells, (H) Th2 cells, (I) Th17 cells, (J) CD8+IFNγ+ T cells, (K) CD8+IL-4+ T cells, (L) CD8+IL-17A+ T cells (control n = 19; F2-SS n = 21; F3-SSS n = 9). Mid-lines indicate medians, boxes indicate interquartile ranges, and whiskers indicate min–max range.

Figure 5

Immunophenotyping of B cells in prenatally stressed neonates. (A) Gating strategy used to identify B cells. (B) Total number of B cells, (C) B1-like cells, and (D) B2-like cells (control n = 19; F2-SS n = 21; F3-SSS n = 9). Mid-lines indicate medians, boxes indicate interquartile ranges, and whiskers indicate min–max range.

Figure 6

Immunophenotyping of CD71+ erythroid cells in prenatally stressed neonates. (A) Gating strategy used to identify CD71+ erythroid cells. (B) Number of CD71+ erythroid cells (control n = 18; F2-SS n = 21; F3-SSS n = 9). Mid-lines indicate medians, boxes indicate interquartile ranges, and whiskers indicate min–max range.

Figure 7

Experimental outcome of stress cessation cohort. (A) Experimental design of stress procedure. Animals of the parental generation (F0-S) were stressed and their pregnant daughters were either stressed (F1-SS) or not stressed (F1-SNS). (B) Rates of delivery in the control, F1-SS, and F1-SNS groups (control n = 13; F1-SS n = 27; F1-SNS n = 17). (C) Growth trajectory of F2-SS and F2-SNS neonates in the first 3 weeks after birth (F2-SS n = 2 litters; F2-SNS n = 10 litters). Data are presented as mean ± standard error of the mean.