Abstract Background Plasma levels of cell-free DNA (cf-DNA) are known to be elevated in sepsis and high levels are associated with a poor prognosis. Mechanical ventilation affects systemic inflammation in which lung-protective ventilation attenuates the inflammatory response. The aim was to study the effect of a lung protective ventilator regime on arterial and organ-specific venous blood as well as on trans-organ differences in cf-DNA levels in a porcine post-operative sepsis model. Method One group of anaesthetised, domestic-breed, 9–12 weeks old, pigs were ventilated with protective ventilation (V_T 6 mL x kg^{− 1}, PEEP 10 cmH₂O) n = 20. Another group, ventilated with a medium high tidal volume and lower PEEP, served as a control group (V_T 10 mL x kg^{− 1}, PEEP 5 cm H₂O) n = 10. Blood samples were taken from four sources: artery, hepatic vein, portal vein and, jugular bulb. A continuous endotoxin infusion at 0.25 μg x kg^{− 1} x h^{− 1} for 5 h was started following 2 h of laparotomy, which simulated a surgical procedure. Inflammatory cytokines and cf-DNA in plasma were analysed and trans-organ differences calculated. Results The protective ventilation group had lower levels of cf-DNA in arterial (p = 0.02) and hepatic venous blood (p = 0.03) compared with the controls. Transhepatic differences in cf-DNA were lower in the protective group, compared with the controls (p = 0.03). No differences between the groups were noted as regards the transcerebral, transsplanchnic or the transpulmonary cf-DNA differences. Conclusions Protective ventilation suppresses arterial levels of cf-DNA. The liver seems to be a net contributor to the systemic cf-DNA levels, but this effect is attenuated by protective ventilation.