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Background: Malnutrition is a serious condition in critically ill patients. The aim of this study is to evaluate the relationships between the Onodera’s prognostic nutritional index (OPNI) and intestinal permeability and between OPNI and systemic inflammation in critically ill patients. Methods: This was a cross-sectional study conducted in the general intensive care unit (ICU) of a university-affiliated hospital. A total of 162 patients admitted between May 2018 and December 2019, was included in the study. The OPNI was calculated at admission and categorized as ≤40 or >40. We assessed plasma endotoxin and zonulin concentrations as markers of intestinal permeability as well as serum interleukin-6 (IL-6) and high-sensitivity C-reactive protein (hs-CRP) as markers of systemic inflammation upon admission under stringent conditions. The relationships between these markers and OPNI were assessed after adjusting for potential confounders through estimation of a binary logistic regression model. Results: Median (interquartile range) hs-CRP, IL-6 zonulin, and endotoxin were significantly greater in the low OPNI subgroup than in the high OPNI subgroup (all P<0.05). Multivariate analyses showed significant association between serum IL-6 (odds ratio [OR], 0.88; 95% confidence interval [CI], 0.64–0.96), serum hs-CRP (OR, 0.77; 95% CI, 0.53–0.92), plasma endotoxin (OR, 0.81; 95% CI, 0.72– 0.93), and plasma zonulin (OR, 0.83; 95% CI, 0.75–0.98) levels with OPNI in the overall population. Conclusions: Our results provide evidence that higher plasma endotoxin, zonulin, IL-6, and hsCRP levels are associated with progressively lower OPNI in mixed ICU populations, particularly in surgical ICU patients.