Maternal Immune Challenge During Early Embryo Development Affects Blastocyst Cell Lineage Ratio and Postnatal Phenotype.

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Maternal infection during gestation is an established cause of abortion, premature birth and intrauterine growth restriction. In addition, there is a strong association between activation of the maternal immune response during mid-gestation and neurological abnormalities in the offspring. At present little is known about the consequences of maternal immune activation during early gestation. The preimplantation embryo has been identified as a critical developmental stage during which environment can program changes to postnatal phenotype. Here, we investigated the sensitivity of preimplantation mouse embryos to maternal immune challenge in the short- and long-term. Virgin female MF-1 mice were mated and randomly assigned to one of four treatment groups; control (saline only), 10, 50 or 150µg/kg of bodyweight lipopolysaccharide (LPS; Salmonella enterica enteritidis ). On gestational day 0.5 (GD0.5), successfully mated females were injected intraperitoneally with saline or LPS dose. Mice were either culled at GD3.5 to assess short-term consequences in blastocysts or allowed to go to term to investigate potential long-term changes in offspring body mass, behaviour and systolic blood pressure (SBP). All postnatal data were analysed using a multilevel random effects regression model (SPSS) which takes into account the hierarchical nature of the dataset. Blastocysts were differentially labelled to assess inner cell mass (ICM) and trophectoderm (TE) cell numbers. Blastocysts from 10µg/kg LPS mothers had more TE cells than those from control mothers (p<0.05). Blastocysts from 50 and 150µg/kg LPS mothers had fewer ICM cells than blastocysts from control mothers (p<0.05). These effects lead to a reduced ICM:TE cell ratio at each LPS dose compared with control (p<0.05). At birth, pups were weighed and litter size adjusted to 3 males and 3 females where possible. Offspring were weighed weekly thereafter and underwent a series of behavioural tests and SBP measurements (tail cuff) until 30 weeks of age. No difference in gestation length or male:female offspring ratio was observed between treatment groups. Interestingly, 10µg/kg LPS mothers had larger litters than control mothers (p<0.01). Body weight was not different between treatment groups at any time point. In the open field behavioural test, both male and female offspring from LPS treated mothers displayed reduced levels of exploratory behaviour compared with controls. For example, male offspring from 150µg/kg LPS mothers displayed fewer rears and jumps than control males at 20 weeks (p<0.05). Furthermore, males from 10µg/kg LPS mothers had elevated SBP at 28 weeks (p<0.05), but females showed no difference in SBP at any time point. These data indicate the sensitivity of the preimplantation mouse embryo to maternal immune activation in vivo leading to altered blastocyst cell allocation, increased litter size (in the lowest dose LPS treatment group) and altered developmental potential; including abnormal behaviour and elevated blood pressure in adult life (also in the lowest dose LPS treatment group). This work serves to highlight maternal immune activation during preimplantation developmentas an important route by which adverse changes in postnatal phenotype can potentially arise; and emphasises the importance of further study into this emerging area of research. This research is funded by a BBSRC studentship (to CLW) and by NIH and MRC grants (to TPF).