Background In utero exposure to obesity is consistently associated with increased risk of metabolic disease, obesity and cardiovascular dysfunction in later life despite the divergence of birth weight outcomes. The placenta plays a critical role in offspring development and long-term health, as it mediates the crosstalk between the maternal and fetal environments. However, its phenotypic and molecular modifications in the context of maternal obesity associated with fetal growth restriction (FGR) remain poorly understood.Methods Using a mouse model of maternal diet-induced obesity, we investigated changes in the placental transcriptome through RNA sequencing (RNA-seq) and Ingenuity Pathway Analysis (IPA) at embryonic day (E) 19. The most differentially expressed genes (FDR < 0.05) were validated by Quantitative real-time PCR (qPCR) in male and female placentae at E19. The expression of these targets and related genes was also determined by qPCR at E13 to examine whether the observed alterations had an earlier onset at mid-gestation. Structural analyses were performed using immunofluorescent staining against Ki67 and CD31 to investigate phenotypic outcomes at both timepoints. Results RNA-seq and IPA analyses revealed differential expression of transcripts and pathway interactions related to placental vascular development and tissue morphology in obese placentae at term, including downregulation of Muc15, Cnn1, and Acta2. Pdgfb, which is implicated in labyrinthine layer development, was downregulated in obese placentae at E13. This was consistent with the morphological evidence of reduced labyrinth zone (LZ) size, as well as lower fetal weight at both timepoints irrespective of offspring sex. Conclusions Maternal obesity results in abnormal placental LZ development and impaired vascularization, which may mediate the observed FGR through reduced transfer of nutrients across the placenta.

Introduction Metformin is the first‐line pharmacological treatment for gestational diabetes in many countries. Although effective at improving maternal glycaemia, metformin readily crosses the placenta and could directly affect the fetus. Indeed, metformin exposure in utero increases postnatal adiposity. However, other long‐term effects on offspring exposed to metformin in utero have not been investigated. Here we explored the impact of maternal metformin treatment during obese, glucose‐intolerant pregnancy on offspring cardiovascular function at one year of age using an established mouse model of maternal diet‐induced obesity. Methods Female C57Bl/6J mice were fed either a standard chow diet [Control dams (Con)] or an obesogenic diet [Obese (Ob) and Obese‐Metformin (Ob‐Met) dams] from weaning up to and including pregnancy and lactation. Ob‐Met dams received a clinically relevant dose of metformin orally from one week prior to mating until day 19 of gestation. All offspring were weaned onto standard chow fed ad libitum at three weeks of age. At one year of age, systolic blood pressure (SBP) was measured by tail cuff plethysmography in the conscious animal and echocardiography was performed under anaesthesia. Post‐mortem, femoral arteries were isolated and vasomotor reactivity tested using in vitro wire myography. Myography dose‐response curves were analysed using mixed effects models, all other data were analysed by One‐Way ANOVA. Post‐hoc multiple comparisons were performed where appropriate. Significance was accepted when P<0.05. Results Relative to controls, female offspring of Ob‐Met dams had increased SBP. However, no differences in SBP were observed in male offspring (Fig.1A). Female offspring of both Ob and Ob‐Met dams had enhanced femoral vasoconstrictor reactivity to K+ (Fig. 1B) but normal cardiac function (Fig. 1C). In contrast, male offspring of both Ob and Ob‐Met dams showed indices of diastolic dysfunction (Fig. 1C) and male offspring of Ob‐Met dams had enhanced femoral vasoconstrictor reactivity to phenylephrine (PE) (Fig.1B). Conclusions Maternal obesity and metformin exposure in utero caused sex‐specific changes in cardiovascular function in 12‐month‐old offspring. In female offspring, exposure to maternal obesity and metformin programmed hypertension with enhanced peripheral vasoconstrictor reactivity. In male offspring, maternal obesity alone programmed echocardiographic indices of left ventricular restrictive filling which were not corrected by metformin treatment, while combined exposure to maternal obesity and metformin also programmed enhanced peripheral vasoconstrictor reactivity. These findings provide novel insight into the long‐term effects of in utero metformin exposure on the cardiovascular health of the next generation in obese pregnancy, and highlight the importance of considering offspring sex in developmental programming studies.