Abstract Background Prenatal exposure to maternal stress may program the fetal HPA axis, potentially leading to altered metabolism in later life, associated with adiposity and diabetes. Aims This association is little studied in humans, and thus we explore whether high maternal job strain during early pregnancy, as well as maternal cortisol levels are associated with increased body mass index (BMI), central adiposity or body fat mass in the offspring at age five. Additionally, we explore whether these associations are modified by gender or mediated by gestational age and fetal growth restriction. Study design 2939 pregnant women (ABCD cohort study) completed a questionnaire around gestational week 16 including the Job Content Questionnaire, assessing job strain. Serum total cortisol was assessed in a subsample (n=1320). Gestational age (≥37weeks), standardized birth weight and information on many covariates were available. At the age five health check, height, weight (BMI, kg/m2), waist circumference (waist-to-height ratio, WHtR) and Fat Mass Index (FMI, kg/m2) were assessed. Results Job strain was not associated with higher BMI, WHtR or FMI. Higher maternal cortisol was independently associated with marginally higher FMI in girls, but marginally lower FMI in boys (β 0.09 and β −0.10 per 100 unit increase in serum cortisol, respectively. p<0.01). This association was not mediated by gestational age or fetal growth restriction. Conclusions Results show that prenatal maternal job strain and cortisol may not program obesity and adiposity in the next generation in humans, but gender differences should always be considered.