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The aim of this study was to investigate whether ulinastatin (UTL) has protective effects on perioperative proinflammatory cytokines and lung injury in cardiopulmonary bypass (CPB) patients. The study included 60 patients undergoing CPB who were randomly divided into a UTL group and a control group. Blood routine examination and inflammatory cytokines concentrations were detected after anesthetic induction (T1), immediately after aortic valve opening (T2), and 4 (T3) and 24 (T4) hours after weaning from CPB. Flow cytometry was used to detect TLR4 and HSP70 expressions. Arterial blood gas and respiratory function were analyzed at the same time points. Compared with the control group, the levels of IL-2, IL-8, TNF-α, NE, TLR4, PA - aDO2, and RI at T2 were significantly lower, whereas HSP70, PaO2, OI, Cd, and Cs were higher in the UTL group (all P < 0.05). Relative to the control group at T3, white blood cell count, TLR4, IL-2, IL-6, IL-8, TNF-α, NE, and RI decreased significantly, whereas IL-10, HSP70, PaO2, OI, and Cs increased in the UTL group (all P < 0.05). At T4, IL-2, IL-6, IL-8, TNF-α, TLR4, and PaCO2 in the UTL group were significantly lower, and PaO2, IL-10, HSP70, and Cs were higher than in the control group (all P < 0.05). Our data show strong evidence that UTL suppresses proinflammatory cytokine elevation and upregulates release of anti-inflammatory mediators, reducing pulmonary injury and improving pulmonary function after CPB.