Acute respiratory distress syndrome (ARDS) caused by severe sepsis remains a major challenge in intensive care medicine. ACE2 has been shown to protect against lung injury. However, the mechanisms of its protective effects on ARDS are largely unknown. Here, we report that ACE2 prevents LPS-induced ARDS by inhibiting MAPKs and NF-κB signaling pathway. Lentiviral packaged Ace2 cDNA or Ace2 shRNA was intratracheally administrated into the lungs of male SD rats. Two weeks after gene transfer, animals received LPS (7.5 mg/Kg) injection alone or in combination with Mas receptor antagonist A779 (10 μg/Kg) or ACE2 inhibitor MLN-4760 (1 mg/Kg) pretreatment. LPS-induced lung injury and inflammatory response were significantly prevented by ACE2 overexpression and deteriorated by Ace2 shRNA. A779 or MLN-4760 pretreatment abolished the protective effects of ACE2. Moreover, overexpression of ACE2 significantly reduced the Ang II/Ang-(1-7) ratio in BALF and up-regulated Mas mRNA expression in lung, which was reversed by A779. Importantly, the blockade of ACE2 on LPS-induced phosphorylation of ERK1/2, p38 and p50/p65 was also abolished by A779. Whereas, only the ERK1/2 inhibitor significantly attenuated lung injury in ACE2 overexpressing rats pretreated with A779. Our observation suggests that AEC2 attenuates LPS-induced ARDS via the Ang-(1-7)/Mas pathway by inhibiting ERK/NF-κB activation.

ACE2 and Ang–(1–7) have important roles in preventing acute lung injury. However, it is not clear whether upregulation of the ACE2/Ang–(1–7)/Mas axis prevents LPS–induced injury in pulmonary microvascular endothelial cells (PMVECs) by inhibiting the MAPKs/NF–κB pathways. Primary cultured rat PMVECs were transduced with lentiviral–borne Ace2 or shRNA– Ace2 and then treated or not with Mas receptor blocker (A779) before exposure to LPS. LPS stimulation resulted in the higher levels of AngII, Ang–(1–7), cytokine secretion and apoptosis rates and the lower ACE2/ACE ratio. Ace2 reversed the ACE2/ACE imbalance and increased Ang–(1–7) levels, thus reducing LPS–induced apoptosis and inflammation, while inhibition of Ace2 reversed all these effects. A779 abolished these protective effects of Ace2 . LPS treatment was associated with activation of the ERK, p38, JNK and NF–κB pathways, which were aggravated by A779. Pretreatment with A779 prevented the Ace2 –induced blockade of p38, JNK and NF–κB phosphorylation. However, only JNK inhibitor markedly reduced apoptosis and cytokine secretion in PMVECs with Ace2 deletion and A779 pretreatment. These results suggest that the ACE2/Ang–(1–7)/Mas axis has a crucial role in preventing LPS–induced apoptosis and inflammation of PMVECs, by inhibiting the JNK/NF–κB pathways.

Propofol exerts protective effects on neuronal cells, in part through the inhibition of programmed cell death. Autophagic cell death is a type of programmed cell death that plays elusive roles in controlling neuronal damage and metabolic homeostasis. We therefore studied whether propofol could attenuate the formation of autophagosomes, and if so, whether the inhibition of autophagic cell death mediates the neuroprotective effects observed with propofol. The cell model was established by depriving the cells of oxygen and glucose (OGD) for 6 hours, and the rat model of ischemia was introduced by a transient two-vessel occlusion for 10 minutes. Transmission electron microscopy (TEM) revealed that the formation of autophagosomes and autolysosomes in both neuronal PC12 cells and pyramidal rat hippocampal neurons after respective OGD and ischemia/reperfusion (I/R) insults. A western blot analysis revealed that the autophagy-related proteins, such as microtubule-associated protein 1 light chain 3 (LC3-II), Beclin-1 and class III PI3K, were also increased accordingly, but cytoprotective Bcl-2 protein was decreased. The negative effects of OGD and I/R, including the formation of autophagosomes and autolysosomes, the increase in LC3-II, Beclin-1 and class III PI3K expression and the decline in Bcl-2 production were all inhibited by propofol and specific inhibitors of autophagy, such as 3-methyladenine (3-MA), LY294002 and Bafilomycin A1 (Baf),. Furthermore, in vitro OGD cultures and in vivo I/R rats showed an increase in cell survival following the administration of propofol, as assessed by an MTT assay or histochemical analyses. Our data suggest that propofol can markedly attenuate autophagic processes via the decreased expression of autophagy-related proteins in vitro and in vivo. This inhibition improves cell survival, which provides a novel explanation for the pleiotropic effects of propofol that benefit the nervous system.

Transient global cerebral ischemia (tGCI) is a cerebrovascular disorder that can cause apoptotic neuronal damage and functional deficits. Basic fibroblast growth factor (bFGF) was reported to be highly expressed in the central nervous system (CNS) and to exert neuroprotective effects against different CNS diseases. However, the effects of bFGF on tGCI have not been studied intensively. This study was conducted to investigate the effect of bFGF and its underlying mechanism in an animal model of tGCI. After intracerebroventricular (i.c.v.) injection of bFGF, functional improvement was observed, and the number of viable neurons increased in the ischemia-vulnerable hippocampal CA1 region. Apoptosis was induced after tGCI and could be attenuated by bFGF treatment via inhibition of p53 mitochondrial translocation. In addition, autophagy was activated during this process, and bFGF could inhibit activation of autophagy through the mTOR pathway. Rapamycin, an activator of autophagy, was utilized to explore the relationship among bFGF, apoptosis, and autophagy. Apoptosis deteriorated after rapamycin treatment, which indicated that excessive autophagy could contribute to the apoptosis process. In conclusion, these results demonstrate that bFGF could exert neuroprotective effects in the hippocampal CA1 region by suppressing excessive autophagy via the mTOR pathway and inhibiting apoptosis by preventing p53 mitochondrial translocation. Furthermore, our results suggest that bFGF may be a promising therapeutic agent to for treating tGCI in response to major adverse events, including cardiac arrest, shock, extracorporeal circulation, traumatic hemorrhage, and asphyxiation.

Background: The hot water tail-flick test is widely used to measure the degree of nociception experienced by laboratory animals. This study was carried out to optimise interval times for the hot water immersion tail-flick tests in rats.Method: Ten different intervals from 10 s to 1 h were tested in 60 Sprague–Dawley male rats. At least eight rats were tested for each interval in three consecutive hot water tail-flick tests. Dixon's up-and-down method was also used to find the optimal intervals. The same rats were then divided into two groups. In Group N, naloxone was injected to reverse the prolonged latency times, whereas saline was used in the control Group S.Results: Intervals of 10 s, 20 s, 30 min and 1 h did not significantly impact latencies, yielding similar results in three consecutive tests (p> 0.05). However, interval times of between 30 s and 20 min, inclusively, caused significantly prolonged latencies in the second and third tests (p< 0.001). Dixon's up-and-down method showed that 95% of the rats had prolonged latencies in hot water tail-flick tests at intervals longer than 32 s. Naloxone reversed prolonged latencies in Group N, whereas the latencies in Group S were further prolonged in 5 min interval tests.Conclusion: The optimal intervals for hot water tail-flick tests are either shorter than 20 s or longer than 20 min. The prolonged latencies after repetitive tests were attributable to an endocrine opioid.