Proteomic global proteins analysis in blast lung injury reveals the altered characteristics of crucial proteins in response to oxidative stress, oxidation-reduction process and lipid metabolic process.

Background: Blast lung injury (BLI) is the most common fatal blast injury induced by overpressure wave in the events of terrorist attack, gas and underground explosion. Our previous work revealed the characteristics of inflammationrelated key proteins involved in BLI, including those regulating inflammatory response, leukocyte transendothelial migration, phagocytosis, and immune process. However, the molecular characteristics of oxidative-related proteins in BLI ar still lacking. Methods: In this study, protein expression profiling of the blast lungs obtained by tandem mass tag (TMT) spectrometry quantitative proteomics were re-analyzed to identify the characteristics of oxidative-related key proteins. Forty-eight male C57BL/6 mice were randomly divided into six groups: control, 12 h, 24 h, 48 h, 72 h and 1 w after blast exposure. The differential protein expression was identified by bioinformatics analysis and verified by western blotting. Results: The results demonstrated that thoracic blast exposure induced reactive oxygen species generation and lipid peroxidation in the lungs. Analysis of global proteins and oxidative-related proteomes showed that 62, 59, 73, 69, 27 proteins (accounted for 204 distinct proteins) were identified to be associated with oxidative stress at 12 h, 24 h, 48 h, 72 h, and 1 week after blast exposure, respectively. These 204 distinct proteins were mainly enriched in response to oxidative stress, oxidation-reduction process and lipid metabolic process. We also validated these results by western blotting. Conclusions: These findings provided new perspectives on blast-induced oxidative injury in lung, which may potentially benefit the development of future treatment of BLI.

Adipose-derived mesenchymal stem cell-loaded ß-chitin nanofiber hydrogel promote wound healing in rats.

Because of stem cells are limited by the low efficiency of their cell homing and survival in vivo, cell delivery systems and scaffolds have attracted a great deal of attention for stem cells' successful clinical practice. ß-chitin nanofibers (ß-ChNF) were prepared from squid pens in this study. Fourier transform infrared spectroscopy, X-ray diffraction and scanning electron microscopy proved that ß-ChNFs with the diameter of 5 to 10 nm were prepared. ß-ChNF dispersion became gelled upon the addition of cell culture medium. Cell culture experiments showed that ß-ChNFs exhibited negligible cytotoxicity towards ADSCs and L929 cells, and it was found that more exosomes were secreted by the globular ADSCs grown in the ß-ChNF hydrogel. The vivo experiments of rats showed that the ADSCs-loaded ß-ChNF hydrogel could directly cover the wound surface and significantly accelerate the wound healing and promote the generation of epithelization, granulation tissue and collagen. In addition, the ADSCs-loaded ß-ChNF hydrogel clearly regulated the expressions of VEGFR, α-SMA, collagen I and collagen III. Finally, we showed that ADSCs-loaded ß-ChNF hydrogel activated the TGFß/smad signaling. The neutralization of TGFß markedly reduced Smad phosphorylation and the expressions of TIMP1, VEGFR and α-SMA. Taken together, these findings suggest that ADSCs-loaded ß-ChNF hydrogel promises for treating wounds that are challenge to heal via conventional methods. Graphical abstract.

Systolic overload-induced pulmonary inflammation, fibrosis, oxidative stress and heart failure progression through interleukin-1ß.

Chronic heart failure is associated with increased interleukin-1ß (IL-1ß), leukocyte infiltration, and fibrosis in the heart and lungs. Here we further studied the role of IL-1ß in the transition from left heart failure to pulmonary hypertension and right ventricular hypertrophy in mice with existing left heart failure produced by transverse aortic constriction. We demonstrated that transverse aortic constriction-induced heart failure was associated with increased lung inflammation and cleaved IL-1ß, and inhibition of IL-1ß signaling using blocking antibodies of clone B122 effectively attenuated further decrease of left ventricular systolic function in mice with existing heart failure. We found that inhibition of IL-1ß attenuated lung inflammation, inflammasome activation, fibrosis, oxidative stress, and right ventricular hypertrophy. IL-1ß blocking antibodies of clone B122 also significantly attenuated lung T cell activation. Together, these data indicate that IL-1ß signaling exerts a causal role for heart failure progression, or the transition from left heart failure to lung remodeling and right heart hypertrophy.

Quantitative analysis of the global proteome in lung from mice with blast injury.

AIM OF THE STUDY: The mechanism by which primary shock wave causes lung injury is unclear. The aim of this study is to find the changes of protein that can be helpful in understanding blast-induced lung injury. MATERIAL AND METHODS: A quantitative analysis of their global proteome was conducted in lung from mice with blast injury using LC-MS/MS. Protein annotation, unsupervised hierarchical clustering, functional classification, functional enrichment and cluster, and protein-protein interaction analyses were performed. Furthermore, western blotting was used to validate the changed protein levels. RESULTS: A total of 6498 proteins were identified, of which 5520 proteins were quantified. The fold-change cutoff was set at 1.2; 132 proteins were upregulated, and 104 proteins were downregulated. The bioinformatics analysis indicated that the differentially expressed proteins were involved in the cholesterol metabolism, asthma, nonalcoholic fatty liver disease. Remarkably, the processes related to the change of oxidative phosphorylation including the NADH dehydrogenase, Cytochrome C reductase, Cytochrome C oxidase and F-type ATPase were significantly upregulated, which were further verified by western blotting. CONCLUSION: These results confirmed that the oxidative phosphorylation is critical to blast-induced lung injury. LC/MS-based profiling presented candidate target/pathways that could be explored for future therapeutic development.

Cold exposure induced oxidative stress and apoptosis in the myocardium by inhibiting the Nrf2-Keap1 signaling pathway.

BACKGROUND: Exposure to cold weather is associated with infaust cardiovascular responses, including myocardial infarction and arrhythmias. However, the exact mechanisms of these adverse changes in the myocardium under cold stress are unknown. This study was designed to investigate the mechanisms of cardiac injury induced by cold stress in mice. METHODS: The mice were randomly divided into three groups, normal control (no handling), 1-week cold stress and 2-week cold stress. We observed physiological changes of the mice and morphological changes of myocardium tissues, and we measured the changes of 3'-nitrotyrosine and 4-hydroxynonenal, the expression levels of superoxide dismutase-1, superoxide dismutase-2, Bax, Bad, Bcl-2, Nuclear factor erythroid-derived 2-like 2 (Nrf2) and Kelch like-ECH-associated protein 1 (Keap1) in myocardium by western blot. Besides, we detected mRNA of superoxide dismutase-1, superoxide dismutase-2, Bax, Bad, Bcl-2, Nrf2 and Keap1 by real-time PCR. One-way analysis of variance, followed by LSD-t test, was used to compare each variable for differences among the groups. RESULTS: Echocardiography analyses demonstrated left ventricle dysfunction in the groups receiving cold stress. Histological analyses witnessed inflammation, vacuolar and eosinophilic degeneration occurred in left ventricle tissues. Western blotting results showed increased 3'-nitrotyrosine and 4-hydroxynonenal and decreased antioxidant enzymes (superoxide dismutase-1 and superoxide dismutase-2) in the myocardium. Expression of Nrf2 and Keap1 followed a downward trend under cold exposure, as indicated by western blotting and real-time PCR. Expression of anti-apoptotic protein Bcl-2 also showed the same trend. In contrast, expression of pro-apoptotic proteins Bax and Bad followed an upward trend under cold exposure. The results of real-time PCR were consistent with those of western blotting. CONCLUSIONS: These findings were very significant, showing that cold exposure induced cardiac injury by inhibiting the Nrf2-Keap1 signaling pathway.

Cytotoxic triterpenes from the acid hydrolyzate of Gynostemma pentaphyllum saponins.

One new dammarane-type triterpene, gypsapogenin A (1), was isolated from the acid hydrolyzate of total saponins from Gynostemma pentaphyllum (Thunb.) Makino, together with two known compounds, (20S,24S)-3ß,20,21ß,23ß,25-pentahydroxy-21,24-cyclodammarane (2) and (23S)-3ß-hydroxydammar-20,24-dien-21-oic acid 21,23-lactone (3). Its structural elucidations were accomplished mainly on the basis of the interpretation of spectroscopic data, such as IR, HR-TOF-MS, and NMR. The cytotoxic activities were evaluated against HepG2 and A549 human cancer cell lines.

Rhamnetin induces sensitization of hepatocellular carcinoma cells to a small molecular kinase inhibitor or chemotherapeutic agents.

BACKGROUND: The rapid development of multi-drug resistance (MDR) process has hindered the effectiveness of advanced hepatocellular carcinoma (HCC) treatments. Notch-1 pathway, which mediates the stress-response, promotes cell survival, EMT (epithelial-mesenchymal transition) process and induces anti-apoptosis in cancer cells, would be a potential target for overcoming MDR process. This study investigated the potential application of rhamnetin, a specific inhibitor of Notch-1 pathway, in anti-tumor drug sensitization of HCC treatment. METHODS: The expression of miR-34a, proteins belonging to Notch-1 signaling pathway or MDR-related proteins was detected by quantitative polymerase chain reaction (qPCR) and western blot assay. To identify whether rhamnetin induces the chemotherapeutic sensitization in HCC cells, the MTT-assays, flow cytometry, soft agar, trans-well and nude mice assays were performed. RESULTS: The endogenous expression of miR-34a was significantly increased and the expression of Notch-1 and Survivin was downregulated after rhamnetin treatment. Treatment of rhamnetin also reduced the expression of MDR related proteins P-GP (P-glycoprotein) and BCRP (breast cancer resistance protein). Rhamnetin increased the susceptibility of HCC cells and especially HepG2/ADR, a MDR HCC cell line, to a small molecular kinase inhibitor sorafenib or chemotherapeutic drugs etoposide and paclitaxel. The IC(50) value of those drugs correspondingly decreased. CONCLUSIONS: Together, our findings suggest that rhamnetin treatment may attenuate the MDR process in HCC cells. These findings may contribute to more effective strategies for HCC therapy. GENERAL SIGNIFICANCE: Rhamnetin acts as a promising sensitizer to chemotherapy and may be a novel approach to overcome the MDR process of HCC.

Recombinant human brain natriuretic peptide attenuates trauma-/haemorrhagic shock-induced acute lung injury through inhibiting oxidative stress and the NF-κB-dependent inflammatory/MMP-9 pathway.

Acute lung injury (ALI) is one of the most serious complications in traumatic patients and is an important part of multiple organ dysfunction syndrome (MODS). Recombinant human brain natriuretic peptide (rhBNP) is a peptide with a wide range of biological activity. In this study, we investigated local changes in oxidative stress and the NF-κB-dependent matrix metalloproteinase-9 (MMP-9) pathway in rats with trauma/haemorrhagic shock (TH/S)-induced ALI and evaluated the effects of pretreatment with rhBNP. Forty-eight rats were randomly divided into four groups: sham operation group, model group, low-dosage rhBNP group and high-dosage rhBNP group (n = 12 for each group). Oxidative stress and MPO activity were measured by ELISA kits. MMP-9 activity was detected by zymography analysis. NF-κB activity was determined using Western blot assay. With rhBNP pretreatment, TH/S-induced protein leakage, increased MPO activity, lipid peroxidation and metalloproteinase (MMP)-9 activity were inhibited. Activation of antioxidative enzymes was reversed. The phosphorylation of NF-κB and the degradation of its inhibitor IκB were suppressed. The results suggested that the protection mechanism of rhBNP is possibly mediated through upregulation of anti-oxidative enzymes and inhibition of NF-κB activation. More studies are needed to further evaluate whether rhBNP is a suitable candidate as an effective inhaling drug to reduce the incidence of TH/S-induced ALI.

Antinociceptive activity of astragaloside IV in the animal model of chronic constriction injury.

To investigate the applicability of astragaloside IV (AG) for the treatment of refractory neuropathic pain, we systemically evaluated the antinociceptive activity of AG in the animal model of chronic constriction injury. We studied behaviors, electrophysiology, and biochemistry from day 2 to day 23 after the surgery. We found that when administered intraperitoneally at the dose of 60 mg/kg, AG caused significant inhibition of allodynia and hyperalgesia induced by mechanic and thermal stimuli as well as downregulation of the expressions of a series of proteins involved in mediating neuropathic pain in the dorsal root ganglia, such as P2X purinoceptor 3, glial cell-derived neurotrophic factor, glial cell-derived neurotrophic factor family receptor α1, and transient receptor potential cation channel subtypes A1 and V1. Further investigation showed that AG restored the nerve conduction velocity and the histological structure of the damaged sciatic nerve on day 23 after the surgery. Moreover, results from immunoelectron microscope showed that glial cell-derived neurotrophic factor family receptor α1 induced by AG could form a circular band in the myelin debris between the injured axons and Schwann cells, contributing toward restoration of the damaged nerve. In conclusion, in our animal model, AG effectively inhibited the neuropathic pain induced by chronic constriction injury.

Silencing CD28 attenuated chest blast exposure-induced traumatic brain injury through the PI3K/AKT/NF-κB signaling pathway in male mice.

In modern war or daily life, blast-induced traumatic brain injury (bTBI) is a growing health concern. Our previous studies demonstrated that inflammation was one of the main features of bTBI, and CD28-activated T cells play a central role in inflammation. However, the mechanism of CD28 in bTBI remains to be elucidated. In this study, traumatic brain injury model induced by chest blast exposure in male mice was established, and the mechanism of CD28 in bTBI was studied by elisa, immunofluorescence staining, flow cytometry analysis and western blot. After exposure to chest shock wave, the inflammatory factors IL-4, IL-6 and HMGB1 in serum were increased, and CD3+ T cells, CD4+ and CD8+ T cell subsets in the lung were activated. In addition, chest blast exposure resulted in impaired spatial learning and memory ability, disruption of the blood-brain barrier (BBB), and the expression of Tau, p-tau, S100ß and choline acetyltransferase were increased. The results indicated that genetic knockdown of CD28 could inhibit inflammatory cell infiltration, as well as the activation of CD3+ T cells, CD4+ and CD8+ T cell subsets in the lung, improve spatial learning and memory ability, and ameliorate BBB disruption and hippocampal neuron damage. Moreover, genetic knockdown of CD28 could reduce the expression of p-PI3K, p-AKT and NF-κB. In conclusion, chest blast exposure could lead to bTBI, and attenuate bTBI via the PI3K/AKT/NF-κB signaling pathway in male mice. This study provides new targets for the prevention and treatment of veterans with bTBI.