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Acute gastrointestinal injury (AGI) is common in mechanically ventilated (MV) patients, but the potential association between ventilatory pressure parameters and AGI grade and their impact on mortality remains unclear. This study aimed to explore the association between ventilatory pressure parameters and AGI grade, and their interaction on all-cause mortality in MV patients. This study was a secondary analysis of a multicenter, prospective, observational study that enrolled adult patients with an expected duration of mechanical ventilation ≥ 48 h from 14 general intensive care units in Zhejiang Province between March and August 2014. The AGI grade was assessed daily on the basis of gastrointestinal symptoms, intra-abdominal pressures, and feeding intolerance in the first week of admission to the ICU. This study included 331 patients (69.2% men; mean age, 64.6 ± 18.9 years). Multivariate regression analysis showed that plateau pressure (Pplat) (OR 1.044, 95% CI 1.009-1.081, P = 0.013), serum creatinine (OR 1.003, 95% CI 1.001-1.006, P = 0.042) and APACHE II score (OR 1.035, 95% CI 1.021-1.072, P = 0.045) were independently associated with global AGI grade III/IV within 7 days of ICU admission. Moreover, global AGI grade (HR 2.228, 95% CI 1.561-3.182, P < 0.001), serum creatinine (HR 1.002, 95% CI 1.001-1.003, P = 0.012) and APACHE II score (HR 1.039, 95% CI 1.015-1.063, P = 0.001) were independently associated with 60-day mortality. In addition, there were significant (P int  ≤ 0.028) interactions of Pplat and DP with AGI grade in relation to 60-days mortality, whereas no interaction (P int  = 0.061) between PEEP and AGI grade on 60-days mortality was observed. In the presence of Pplat ≥ 19 cmH 2 O, the patients with AGI grade III/IV had 60-day mortality rate of 72.2%, significantly higher than those with AGI grade I/II (48.7%, P = 0.018), whereas there were no significant differences (27.9% vs. 33.7%, P = 0.39) in 60-days mortality between AGI grade I/II and III/IV among the patients with Pplat < 19 cmH 2 O. In comparison with Pplat, DP had a similar interaction (P int  = 0.028) with AGI grade on 60-day mortality. Ventilatory pressure parameters (Pplat and DP) are independent risk factors of AGI grade III/IV. Pplat and DP interact with AGI grade on 60-days mortality, highlighting the importance of optimizing ventilatory pressure parameters to improve gastrointestinal function and survival outcomes of MV patients.Trial registration: ChiCTR-OCS-13003824., (© 2024. The Author(s).)