Background. Hydrogen-rich saline (HRS) has strong anti-inflammatory, antioxidative stress, and antiapoptotic properties. The study focused on the protection of HRS on lipopolysaccharide (LPS)-induced acute lung injury (ALI) in rat models and the relationship with autophagic regulation and mTOR/TFEB signaling pathway. Material and Methods. The LPS-induced ALI rats’ model was established. Pathohistological change in lung tissue was detected by hematoxylin-eosin staining. The inflammatory cytokines were examined by enzyme-linked immunosorbent assay (ELISA). The key apoptosis proteins and autophagy-relevant proteins were analyzed by western blotting. In vitro, HPMEC models of ALI were treated with LPS. The inflammatory cytokines were detected. Apoptosis rate was determined by flow cytometry. The autophagy and mTOR/TFEB signaling pathway-related proteins were detected by western blot and immunohistochemical staining. Results. HRS attenuated LPS-induced ALI and apoptosis both in vivo and in vitro. HRS attenuated inflammatory response, inhibited apoptosis, induced and activated autophagy in LPS-induced ALI model, and downregulated mTOR/TFEB signaling pathway. The protection of HRS can be blocked by autophagy inhibitor. Moreover, mTOR activator reversed HRS protection and mTOR inhibitor enhanced HRS protection in LPS-induced model and HRS activated autophagy via mTOR/TFEB signaling pathway. Conclusion. The results confirmed the protection of HRS in LPS-induced ALI by regulating apoptosis through inhibiting the mTOR/TFEB signaling pathway.
Background: Hydrogen-rich saline (HRS) has a protective effect on sepsis-induced lung injury. However, the underlying mechanisms are still unclear. Polarization and apoptosis of macrophages are essential factors in the pathogenesis of acute lung injury (ALI). Moreover, autophagy is involved in the regulation of both macrophage polarization and apoptosis. Therefore, this study investigated the ability of HRS to attenuate ALI through regulation of the polarization and apoptosis of alveolar macrophages (AMs) during sepsis by modulating autophagy. Methods: Male Sprague-Dawley (SD) rats were used to prepare the sepsis-induced lung injury animal model. Rat lung tissue was harvested after lipopolysaccharide (LPS) treatment, in the presence or absence of HRS, and the AMs were analyzed for changes in polarization, apoptosis, and autophagy. The rat AM cell line NR8383 was used to examine these processes in vitro using Western blot analysis, flow cytometry, and transmission electron microscopy. Results: LPS-induced ALI in rats was associated with an increase in autophagy, apoptosis, and M1 polarization but a decrease in M2 polarization in AMs. These effects were reversed by administration of HRS. Inhibition of AM autophagy with 3-methyladenine (3-MA) decreased apoptosis and M1 polarization and increased M2 polarization, paralleling the effects of HRS. Conclusions: HRS could attenuate ALI in septic rats through regulation of AM polarization and a reduction in apoptosis by suppressing autophagy. This may represent a potential novel therapeutic target for the treatment of ALI caused by sepsis.
To determine the effect of saturated hydrogen saline on lipopolysaccharide (LPS)-induced acute liver dysfunction, rats were divided into control, LPS, and LPS plus saturated hydrogen saline (LPS+H(2)) groups. Treatment with saturated hydrogen saline prolonged the median survival time and reduced liver dysfunction. Moreover, saturated hydrogen saline significantly reduced pathological alterations in liver tissues, the number of ballooned hepatocytes, serum tumor necrosis factor (TNF)-alpha and interleukin (IL)-6 levels, and myeloperoxidase (MPO) and malondialdehyde (MDA) levels in liver tissues (P<0.05). Cell apoptosis was detected in liver tissues after LPS treatment, and attenuated by saturated hydrogen saline treatment. Saturated hydrogen saline also decreased phosphorylated extracellular signal-regulated kinase (p-ERK), phosphorylated Jun kinase (p-JNK), nuclear factor-kappa B (NF-kappaB), and second mitochondria-derived activator of caspase (Smac) levels, and increased p38 activation (P<0.05). Thus, saturated hydrogen saline may attenuate LPS-induced acute liver dysfunction in rats, possibly by reducing inflammation and cell apoptosis. Mitogen-activated protein kinase (MAPK), NF-kappaB, and Smac may contribute to saturated hydrogen saline-mediated liver protection.
Acute lung injury induced by lipopolysaccharides (LPSs) is caused by pulmonary inflammation and pulmonary vascular permeability. Activation of p38 mitogen-activated protein kinase causes inflammation, and proinflammatory cytokines and oxidative stress induce autophagy, a catabolic mechanism responsible for protein degradation and recycling of damaged proteins and cytoplasmic organelles. If not controlled, excessive autophagy responses can result in cell death. In this study, we pretreated rats with saturated hydrogen saline, and examined the molecular mechanism by which saturated hydrogen saline attenuates LPS-induced acute lung dysfunction. Sixty-four male Sprague-Dawley rats were randomly assigned to one of three groups-a control group, an LPS group, or an LPS plus saturated hydrogen saline (LPS + H2) group. Treatment with saturated hydrogen saline prolonged the median survival time of rats and reduced lung dysfunction induced by LPS. Moreover, saturated hydrogen saline significantly attenuated LPS-mediated induction of serum tumor necrosis factor α, interleukin 6, myeloperoxidase, and malondialdehyde (P < 0.05). Autophagosomes were found in the cytoplasm of type II alveolar epithelial cells of LPS-treated rats, and light chain 3 protein (LC3)I/II was increased by LPS treatment. In contrast, saturated hydrogen saline decreased the number of autophagosomes and LC3I/II expression. Saturated hydrogen saline also attenuated the LPS-mediated increase in apoptosis and p38 expression. Taken together, saturated hydrogen saline may attenuate LPS-induced acute lung dysfunction in rats by reducing inflammation, autophagy, and apoptosis involving the p38 mitogen-activated protein kinase signaling pathway. Copyright © 2015 Elsevier Inc. All rights reserved.
Sepsis is common in intensive care units (ICU) and is associated with high mortality. Cardiac dysfunction complicating sepsis is one of the most important causes of this mortality. This dysfunction is due to myocardial inflammation and reduced production of energy by the heart. A number of studies have shown that hydrogen-rich saline (HRS) has a beneficial effect on sepsis. Therefore, we tested whether HRS prevents cardiac dysfunction by increasing cardiac energy. Four groups of rats received intraperitoneal injections of one of the following solutions: normal saline (NS), HRS, lipopolysaccharide (LPS), and LPS plus HRS. Cardiac function was measured by echocardiography 8 h after the injections. Gene and protein expression related to fatty acid oxidation (FAO) were measured by quantitative polymerase chain reaction (PCR) and Western blot analysis. The injection of LPS compromised heart function through decreased fractional shortening (FS) and increased left ventricular diameter (LVD). The addition of HRS increased FS, palmitate triphosphate, and the ratio of phosphocreatinine (PCr) to adenosine triphosphate (ATP) as well as decreasing LVD. The LPS challenge reduced the expression of genes related to FAO, including perioxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), perioxisome proliferator-activated receptor alpha (PPARα), Estrogen-related receptor alpha (ERRα), and their downstream targets, in mRNA and protein level, which were attenuated by HRS. However, HRS had little effect on glucose metabolism. Furthermore, HRS inhibited c-Jun N-terminal kinase (JNK) activation in the rat heart. Inhibition of JNK by HRS showed beneficial effects on LPS-challenged rats, at least in part, by restoring cardiac FAO.
Aquaporin 1(AQP1) and AQP5 have an important role in eliminating extravascular lung water, an increase of which contributes to lung injury in patients with sepsis and its consequent mortality. It has been reported that hydrogen-rich saline (HRS) has protective effects against sepsis-related lung injury. In this study, we hypothesized that the protective effect occurred by preserving the expression of AQP1 and AQP5. To test this hypothesis, male Sprague-Dawley rats received intratracheal administration of lipopolysaccharide (LPS) followed by intraperitoneal injection of HRS. Lung function, wet-to-dry weight ratio, and histopathology scores were determined. The expression of AQP1 and AQP5 at the messenger RNA and protein levels, as well as the involved pathways, was explored by quantitative polymerase chain reaction and Western blot. LPS significantly impaired lung function and downregulated the expression of AQP1 and AQP5 in the rat lung, all of which were attenuated by HRS treatment. Moreover, HRS treatment inhibited LPS-induced activation of p38 mitogen-activated protein kinase and jun N-terminal kinase, which is associated with LPS-induced downregulation of AQP1 and AQP5.