The study was designed to show the effects of early administration of levosimendan on intestinal damage after ischemia–reperfusion in an experimental intestinal injury-reperfusion model. Our study revealed that levosimendan reduces small intestine damage in terms of Chiu score.
Intestinal ischemia and reperfusion lead to the formation of inflammatory cytokines and reactive oxygen species . Cytotoxic events disrupt the barrier function of the intestine and cause toxic products to pass into the systemic circulation, affecting the organs such as the kidneys, liver, and heart leading to multiple organ failure (MOF) . There are basic treatment methods that aim at reducing ischemia–reperfusion damage and preventing the development of MOF . For this purpose, ischemic preconditioning, treatment with antioxidant agents, nitric oxide (NO) applications, anti-complement therapy, and pharmacological preconditioning caused by various agents have been used .
Mallick et al. created their experimental models by performing 30 min ischemia and 120 min reperfusion . In our study, longer ischemia time was aimed and 60-min ischemia and 120-min reperfusion model used by Özkaya  and Zhang  were applied.
Özkaya et al.  investigated the effects of pre-treatment levosimendan on small bowel damage in the rat IIR model. The investigators administered a loading dose of 12 µg.kg−1 levosimendan before ischemia followed by a maintenance infusion of 0.2 µg.kg−1.min−1 during 60 min of ischemia. The recommended clinical dose of the agent was reported as 0.05–0.2 µg.kg−1.min−1 infusion after 6–24 µg.kg−1 loading dose . In our study, we administered a maintenance infusion dose of 0.2 µg.kg−1.min−1 following a 12 µg.kg−1 during a 10-min loading dose, a dosage whose efficacy has been shown in two separate experimental studies [9, 10].
Ahmetova et al.  investigated the effect of levosimendan in the hepatic IR model and reported a decrease in MAP measurements at the 15th and 45th minute of reperfusion in the IR group. It has been reported that the effect of levosimendan peaks within 10–30 min following the loading dose, and in addition to its positive inotropic effects, levosimendan opens the ATP-dependent K + channels in myocytes and vascular smooth muscle cells, causing vasodilation in the systemic vascular bed leading to hypotension . Oldner et al. suggested that levosimendan infusion initiated before endotoxin led to a 37% decrease in systemic vascular resistance index and approximately 22% decrease in the mean arterial pressure . In our study, there was no decrease in MAP-8 and MAP-9 after 10 min of 12 µg.kg−1 loadings and 50 min of 0.2 µg.kg−1.min−1 levosimendan infusion, respectively. During these time periods, the expected difference between all 3 groups did not occur, and we observed that the blood pressure values of the IIR + L group were similar to the sham group. This result suggested that levosimendan enhances cardiac performance in IIR , preserving systemic blood pressure despite reperfusion.
It has been reported that free oxygen radicals and hydrogen peroxide, which increase as a result of intestinal IR damage, initiate lipid peroxidation and protein damage, leading to cell apoptosis and tissue necrosis. MDA and thiobarbituric acid reactive substances (TBARS) are released as a result of lipid peroxidation . In our study, the MDA level, which is the end product of lipid peroxidation, was measured as an indicator of free oxygen radical formation in tissues. Although there was no difference between the groups when compared statistically, we observed that MDA levels were the highest in the IIR group and lowest in the sham group, and MDA levels in the IR + L group were less than in the IR group. In an experimental study by Özkaya et al. , levosimendan was applied before the IR period, and tissue TBARS levels were found to be significantly higher in the IR group when compared to the sham and IR + L groups. This result shows that levosimendan administered as a pretreatment reduces the lipid peroxidation that occurs after intestinal ischemia–reperfusion. Investigating the effect of levosimendan on oxidative stress in myositis culture, Maytin et al.  found that the drug opened K-ATP channels in the clinical therapeutic dose range and protected the cell from apoptosis caused by hydrogen peroxide. Apoptosis has an important role in ischemia–reperfusion injury . In our study, although the administration of levosimendan after reperfusion was shown to contribute to a decrease in lipid peroxidation, this could not be shown statistically.
Various scores are used in the histopathological classification of intestinal IR injury. Chiu et al. , Park et al. , and Hierholzer et al.  have all reported different scoring systems.
In our study, we found that the villi structure in the small intestines was severely damaged in the IIR group, and ulceration and mononuclear cell infiltration in the lamina propria were significantly increased, and therefore, the Chiu score was significantly higher when compared to the scores of the sham and IIR + L groups. Although there was a difference in the Chiu score of the IIR + L group when compared to the sham group, the presence of hemorrhage, ulceration, and mononuclear cell number were very similar to the sham group in light microscopic imaging. Similar histopathological findings were found by Özkaya et al. , Mallick et al. , Topaloğlu et al. , and Zhang et al.  who used levosimendan, pyrrolidine dithiocarbamate, prostaglandin E2 (PGE2), and glucagon-like peptide 2 (GLP-2) as pre-medication before IR damage, respectively.
Levosimendan increases intestinal perfusion and oxygenation by positively affecting splanchnic and mesenteric blood flow [27, 28]. Garcia-Septiem et al.  demonstrated that levosimendan pretreatment improved portal blood flow, intestinal mucosa oxygenation, and pulmonary functions in pigs in a septic shock model created with intravenous Escherichia coli (E. coli). In our study, the Chiu score was found to be statistically significantly lower in the levosimendan group when compared to the IIR group. We believe this was caused by the decrease in splanchnic resistance, the increase in regional blood flow, and mucosal oxygenation. Emphasizing that levosimendan is superior to milrinone and dobutamine in increasing gastric mucosal oxygenation, it has been recommended that levosimendan be used as an alternative in patients with splanchnic ischemia risk [8, 29]. Özkaya et al.  reported that levosimendan infusion initiated before ischemia decreased neutrophil accumulation, lipid peroxidation, and the Chiu score in the intestines. However, clinically, it can be difficult to know the exact time of onset of ischemia. The authors concluded that further experimental studies investigating the initiation time and duration of the application are required for the results of the research to gain relevant clinical significance.
Although levosimendan infusion administered for 1 h from the 2nd hour of intestinal reperfusion decreased the histological damage score without causing adverse hemodynamic effects, we were unable to demonstrate its effect on lipid peroxidation. Also, this study did not evaluate myeloperoxidase, inflammatory cytokines, and adhesion molecules showing neutrophil adhesion. It would be appropriate to support the results of this experimental study with comprehensive biochemical parameters and drug administration starting from the reperfusion process.