As a common disease with frequent occurrence, the prevalence and incidence of diabetes in China have been increasing year by year in recent years. And after suffering from diabetes, due to long-term hyperglycemia, can lead to a variety of complications, especially infection. When patients with diabetes experience surgical trauma again, artificial assisted breathing with tracheal intubation, long-term bed rest or coma, patients with diabetes are prone to secondary pulmonary infection and may further lead to acute lung injury [1,2].
Ulinastatin, purified from human urine, is a glycoprotein with a molecular weight of 67kDa. As a protease inhibitor, it can effectively inhibit a variety of enzymes and inflammatory reactions caused by adverse stimulation. In addition, it can improve microcirculation by clearing excessive oxygen free radicals, stabilizing lysosomal membrane and preventing the release of inflammatory factors. Studies have shown that ulinastatin can inhibit the release of inflammatory mediators, prevent the cascade of cytokines, inhibit the excessive activation of leukocytes, and block the vicious cycle between cytokines and leukocytes. Ulinastatin has lung protective effects in systemic inflammatory diseases such as sepsis, DIC and multiple organ failure [11-13].
In this study, high-fat diet supplemented with single-dose intraperitoneal injection of streptozotocin (STZ) was used to construct the T2DM rat model. The characteristic of this approach is that low doses of STZ can destroy the function of rat islet β cells, while feeding with high fat diet can reduce the sensitivity of peripheral tissues to insulin, thus forming a similar T2DM model. In this study, a total of 38 rats were enrolled for modeling, of which 1 died. Finally, 36 rats with successful modeling were selected for the next stage of animal model construction.
The main manifestations of acute lung injury caused by sepsis are as follows: Alveolar epithelium and extensively damaged alveolar capillary wall accompanied by obvious inflammatory edema, alveolar walls are broken, the thickening of alveolar interval, and red blood cells and inflammatory cells into the alveolar cavities, severe damaged alveolar epithelial cells, damaged alveolar surface active substances, with the progress of the course, part of the lung atelectasis can occur.
In this study, the lung interstitium of rats in group D was thickened to a certain extent, with a small amount of inflammatory cell infiltration and a little exudate in some alveolar cavities, which might be caused by lung microvascular injury caused by long-term hyperglycemia. DS group rats significantly higher, W/D structure deformation, lung tissue alveolar walls collapse, pulmonary interstitial broadening, atelectasis, pulmonary interstitial and alveolar we see a lot of inflammatory cells infiltration and red blood cells, endobronchial visible loss of epithelial cells, numerous capillaries expansion at the same time, serious lung bubble cavity reduced or even disappeared, These results indicated that the rat model of diabetic sepsis with acute lung injury was successfully constructed. W/D in group U was lower than that in group DS, and there were more inflammatory cells in the interstitial and part of alveolar cavities, exfoliated epithelial cells were occasionally found in the lumen, and the interstitial widened, and the pathological damage of lung tissue was lighter than that in group DS, suggesting that ulinastatin can alleviate acute lung injury in diabetic sepsis rats to a certain extent.
For patients with diabetes, pathophysiological changes such as decreased oxygen carrying capacity of red blood cells, increased stress level and thickened basement membrane of microvessels caused by long-term hyperglycemia and metabolic disorders can induce infection and lead to sepsis, and the degree of hyperglycemia can also be aggravated during the onset of sepsis. At present, it has been confirmed that the occurrence of inflammatory response in diabetes patients can be realized through toll-like receptor 4 (TLR4) pattern recognition receptor [14,15]. In this study, the expression of TLR4 in group D was significantly higher than that in the control group, which was consistent with previous studies. Studies on the expression changes of TLR4 in the lung tissues of diabetic rats and the reactivity of TLR4 to lipopolysaccharide showed that TLR4 could not play the normal immune recognition function under immune stress conditions, leading to the decrease of the immune defense ability of the body and the susceptibility to lung infection. In this study, after lPS-induced acute lung injury, TLR4 expression in DS group and U group was significantly increased compared with that in D group, indicating further activation of TLR4, while TLR4 expression in U group was significantly decreased compared with that in DS group after ulinastatin pretreatment, indicating that ulinastatin can inhibit TLR4 activation to a certain extent.
Studies have shown that clinically relevant lung tissue inflammation is less prevalent in TLR4 gene deletion animals [5], where the Toll/ interleukin-1 receptor binding region induces interferon β(TRIF) pathway to play an important role in the inflammatory response induced by endotoxin-induced lung injury [6].
A large number of previous studies have shown that the serum levels of TNF-α, IL-10, IL-1, IL-6 and other inflammatory factors can be significantly reduced after the use of UTI, and the inflammatory response can be significantly reduced, and the anti-inflammatory effect of UTI is obvious [16-19]. In particular, serum TNF-α levels will be very low when UTI is preadministered prior to LPS stimulation, and serum TNF-α levels are inversely proportional to UTI doses. Other studies have found that ulinastatin does not have a significant anti-inflammatory effect in some experimental models.
In this study, compared with the blank group, serum LEVELS of TNF-α, IL-1β and IL-18 were significantly increased in the diabetic group, indicating that long-term hyperglycemia activates the body's inflammatory response to a certain extent, and hyperglycemia is an important risk factor for sepsis infection. Diabetes sepsis group of rats in the serum TNF alpha, beta, IL IL - 1-18 relative to the first two have increased significantly, and ulinastatin in rat serum inflammatory index fell after pretreatment, suggesting that sepsis infection significantly increase systemic inflammatory response diabetic rats, and ulinastatin can significantly reduce diabetes sepsis systemic inflammatory reaction.
Oxidative stress response is one of the important mechanisms of lung injury in diabetic sepsis. In previous studies, ulinastatin has been reported to have lung protective effects by inhibiting oxidative stress responses. Gao et al. 's study indicated that ulinastatin can reduce the degree of lung injury in burned rats by increasing the generation of SOD and reducing the generation of MDA in lung tissues [20]. In the sepsis model constructed by CLP, hyperglycemia can significantly increase the levels of oxidative factors in the kidney of rats, such as lipid peroxidation (LPO) and catalase (CAT) levels. Meanwhile, SOD activity in kidney tissue was significantly decreased in CLP group [27].
In this study, it was also found that compared with blank group, the content of MDA in serum of rats in diabetic group increased, while the activity of SOD decreased. The oxidative stress response in serum of diabetic sepsis group was significantly increased; The oxidative stress response in serum of ulinastatin pretreated rats was significantly reduced. This indicates that sepsis infection significantly aggravates the systemic oxidative stress response of diabetic rats. Ulinastatin can significantly reduce systemic oxidative stress response in diabetic sepsis. In this study, ulinastatin can improve oxidative stress and reduce levels of inflammatory factors.
Long-term chronic hyperglycaemia reduces the level of 2, 3-diphosphoglyceric acid in red blood cells, reduces the oxygen carrying capacity of red blood cells, increases the proportion of glycosylated hemoglobin in hemoglobin, and its affinity for oxygen is higher than that of normal hemoglobin, so that oxygen is not easy to diffuse into tissues; Thickening of capillary basement membrane causes chronic ischemia and hypoxia of pulmonary microvessels. Hypoxia inducible factor 1α (HIF-1α) is a key factor specifically mediating hypoxia response in the regulation of oxygen balance in cells. Most cells express HIF-1α in response to hypoxia. Protein stability of HIF-1α is regulated by intracellular oxygen concentration. In cells with normal oxygen content, hiF-1 α is degraded by protease immediately after translation and thus is largely undetectable. As a mediator of physiological and pathological responses to hypoxia adaptation, hypoxia-inducible factor-1 is involved in the transcription and expression of many related genes. In vitro studies have shown that hiF-1 αmRNA expression increases and hiF-1 α protein accumulation after acute LPS stimulation [21]. In contrast, long-term and sustained LPS stimulation resulted in endotoxin tolerance and reduced hiF-1 α protein content in monocytes [22]. In this study, the expression level of HIF-1α protein in the lung tissue of diabetic rats was increased compared with that in the control group, which was consistent with previous studies. However, after LPS induced acute lung injury, hiF-1 α protein expression level in lung tissue was further increased and hypoxia in lung tissue was aggravated. However, ulinastatin pretreatment can down-regulate hiF-1 α protein level in lung tissue, and improve hypoxia response in lung tissue to a certain extent.
Diabetes is a chronic metabolic disorder, and microangiopathy is the characteristic pathological change of diabetes occurring in vital organs of the whole body. A large number of studies have shown that lung tissue rich in microvessels is also one of the target organs of diabetes damage [23,24]. It has been reported that microangiopathy is the basis of diabetic pulmonary complications [25]. In this study, the content of EB in group D showed an increasing trend compared with the control group, but there was no significant difference, indicating that hyperglycemia can cause pulmonary microvascular injury to a certain extent. After LPS induction, EB content was significantly increased in experimental rats, which further aggravated microvascular injury. After pretreatment with ulinastatin, EB content was significantly lower than that in the DS group, indicating that ulinastatin can improve pulmonary microvascular injury caused by diabetic sepsis, which is consistent with previous reports [26-28].