Vaporization of perfluorocarbon attenuates sea-water-drowning-induced acute lung injury by deactivating the NLRP3 inflammasomes in canines.

Seawater-drowning-induced acute lung injury (SD-ALI) is a life-threatening disorder characterized by increased alveolar-capillary permeability, an excessive inflammatory response, and refractory hypoxemia. Perfluorocarbons (PFCs) are biocompatible compounds that are chemically and biologically inert and lack toxicity as oxygen carriers, which could reduce lung injury in vitro and in vivo. The aim of our study was to explore whether the vaporization of PFCs could reduce the severity of SD-ALI in canines and investigate the underlying mechanisms. Eighteen beagle dogs were randomly divided into three groups: the seawater drowning (SW), perfluorocarbon (PFC), and control groups. The dogs in the SW group were intratracheally administered seawater to establish the animal model. The dogs in the PFC group were treated with vaporized PFCs. Probe-based confocal laser endomicroscopy (pCLE) was performed at 3 h. The blood gas, volume air index (VAI), pathological changes, and wet-to-dry (W/D) lung tissue ratios were assessed. The expression of heme oxygenase-1 (HO-1), nuclear respiratory factor-1 (NRF1), and NOD-like receptor family pyrin domain containing-3 (NLRP3) inflammasomes was determined by means of quantitative real-time polymerase chain reaction (qRT-PCR) and immunological histological chemistry. The SW group showed higher lung injury scores and W/D ratios, and lower VAI compared to the control group, and treatment with PFCs could reverse the change of lung injury score, W/D ratio and VAI. PFCs deactivated NLRP3 inflammasomes and reduced the release of caspase-1, interleukin-1ß (IL-1ß), and interleukin-18 (IL-18) by enhancing the expression of HO-1 and NRF1. Our results suggest that the vaporization of PFCs could attenuate SD-ALI by deactivating NLRP3 inflammasomes via the HO-1/NRF1 pathway.

Non-invasive detection of EGFR and TP53 mutations through the combination of plasma, urine and sputum in advanced non-small cell lung cancer.

The sensitivity and utility of liquid biopsy in clinical practice requires some improvement. The aim of the present study was to improve the detection of epidermal growth factor (EGFR) and cellular tumor antigen p53 (TP53) mutations in liquid biopsies from patients with advanced non-small cell lung cancer (NSCLC) by combining plasma, sputum and urine samples under the same sequencing platform. Plasma, sputum and urine samples, and tumor tissues were obtained from 50 patients with NSCLC and were analyzed using next-generation sequencing. The sensitivity of EGFR-sensitive mutation detection was 84% in plasma, 63% in sputum, 28% in urine, and 91% when combining the three liquid samples (P<0.001). The sensitivity of TP53 mutation detection increased from 87% in plasma to 94% when the three samples were combined (P<0.001). The sensitivity of EGFR or TP53 mutations detection was higher in patients with multiple metastatic sites compared with patients ≤1 metastatic site. In addition, the progression free survival (PFS) rates obtained following analysis of the three samples independently in patients with EGFR sensitizing mutations were similar, and were 9.0 months in the tissue sample, 7.5 months in plasma, 7.9 months in the sputum and 7.3 months in urine (P=0.721). The PFS of patients with TP53 mutations was shorter compared with patients without TP53 mutations and was as follows: Tissue, 8.2 months compared with 10.2 months (P=0.412); plasma, 8.4 months compared with 10.2 months (P=0.466); sputum, 8.3 months compared with 9.1 months (P=0.904); and when combined, 8.8 months compared with 10.3 months (P=0.599). The combination of plasma, sputum and urine increased the detection of EGFR or TP53 mutation with higher sensitivity, and may improve the predictive value of personalized treatment.

Cystic and solitary nodular pulmonary metastases in a patient with low-grade endometrial stromal sarcoma: A case report and literature review.

Pulmonary metastases of endometrial stromal sarcoma (ESS) are uncommon and can be difficult to diagnose. The aims of the present study were to investigate the clinical and pathological features, and enhance the awareness of pulmonary metastases in patients with low-grade ESS. The study reports a case of low-grade ESS that resulted in cystic and nodular pulmonary metastases. Furthermore, the PubMed database was searched using 'pulmonary metastases of low-grade endometrial stromal sarcoma' as the key phrase. The literature on pulmonary metastases of low-grade ESS was reviewed and 35 cases were included in the present study. The clinical manifestations, imaging data, pathological features, treatment and prognosis of the 35 previously reported cases and the current case were retrospectively analyzed. The age range of the 36 patients diagnosed with low-grade ESS was 28-65 years. The time period from confirmation of ESS to lung metastases was 1.5-27 years. In 50% of the patients, the pulmonary metastases were asymptomatic. The most common pulmonary symptom was dyspnea, followed by chest pain, pneumothorax and coughing. The most common chest imaging presentation was multiple pulmonary nodules, followed by a solitary nodule or mass. Histology was used to identify that the pulmonary metastases had the pathological features of low-grade ESS. The immunohistochemical results demonstrated strong diffuse immunoreactivity for cluster of differentiation 10, estrogen receptor and progesterone receptor in almost all the specimens. The review of the literature revealed that pulmonary metastases from low-grade ESS are rare but not negligible. Furthermore, the detailed clinical information, imaging findings and immunohistochemical detection are important for making a diagnosis.

Differences in the genomic profiles of cell-free DNA between plasma, sputum, urine, and tumor tissue in advanced NSCLC.

Liquid biopsy has provided an efficient way for detection of gene alterations in advanced non-small-cell lung cancer (NSCLC). However, the correlation between systematic determination of somatic genomic alterations in liquid biopsy and tumor biopsy still remained unclear, and the concordance rate between cell-free DNA (cfDNA) and matched tumor tissue DNA needs to be increased. A prospective study was performed to detect differences in genetic profiles of cfDNA in sputum, plasma, urine, and tumor tissue from 50 advanced NSCLC patients in parallel by the same next-generation sequencing (NGS) platform. Driver genes alterations were identified in cfDNA sample and matched tumor sample, with an overall concordance rate of 86% in plasma cfDNA, 74% in sputum cfDNA, 70% in urine cfDNA, and 90% in cfDNA of combination of plasma, sputum, and urine. And the concordant rate of cfDNA in sputum in patients with smoking history was higher than that in patients without history of smoking (89% vs. 66%, P = 0.033) and equal to that in plasma cfDNA of the smoking patients (89% vs. 89%). In conclusion, sputum cfDNA can be considered as an alternative medium to liquid biopsy, while the complementarity of genomic profiles in cfDNA among plasma, sputum, and urine was beneficial to detect more diver genes alterations and improve the utility of liquid biopsy in advanced NSCLC (Liquid Biopsy for Detection of Driver Mutation in NSCLC; NCT02778854).

Whole-Genome Analysis of an Extensively Drug-Resistant Acinetobacter baumannii Strain XDR-BJ83: Insights into the Mechanisms of Resistance of an ST368 Strain from a Tertiary Care Hospital in China.

Acinetobacter baumannii is an important pathogen of nosocomial infections. Nosocomial outbreaks caused by antibiotic-resistant A. baumannii remain a significant challenge. Understanding the antibiotic resistance mechanism of A. baumannii is critical for clinical treatment. The purpose of this study was to determine the whole-genome sequence (WGS) of an extensively drug-resistant (XDR) A. baumannii strain, XDR-BJ83, which was associated with a nosocomial outbreak in a tertiary care hospital of China, and to investigate the antibiotic resistance mechanism of this strain. The WGS of XDR-BJ83 was performed using single-molecule real-time sequencing. The complete genome of XDR-BJ83 consisted of a 4,011,552-bp chromosome and a 69,069-bp plasmid. The sequence type of XDR-BJ83 was ST368, which belongs to clonal complex 92 (CC92). The chromosome of XDR-BJ83 carried multiple antibiotic resistance genes, antibiotic efflux pump genes, and mobile genetic elements, including insertion sequences, transposons, integrons, and resistance islands. The plasmid of XDR-BJ83 (pBJ83) was a conjugative plasmid carrying type IV secretion system. These results indicate that the presence of multiple antibiotic resistance genes, efflux pumps, and mobile genetic elements is likely associated with resistance to various antibiotics in XDR-BJ83.

Cell penetrating peptide of sodium-iodide symporter effect on the I-131 radiotherapy on thyroid cancer.

The aim of the present study was to clarify whether the cell penetrating peptide of sodium-iodide symporter (NIS) has an effect on the I-131 radiotherapy of thyroid cancer. Firstly, we combined the HIV-1 TAT peptide (a cell penetrating peptide, dTAT) and established a nanoparticle vector (dTAT NP) to study the delivery efficiency of this cell-penetrating strategy for tumor-targeted gene delivery. dTAT NP was transfected into cultured TPC-1 cells as a model to study the effects of I-131 radiotherapy on thyroid cancer. Reverse transcription-quantitative polymerase chain reaction and western blotting results showed that the mRNA and protein expression levels of NIS in the transfected TPC-1 cells were substantially higher than in the negative control cells. MTT and flow cytometric analyses demonstrated that the cell growth and apoptosis rates of the TPC-1 cells were significantly inhibited and activated, respectively, by treatment with dTAT NP. The results of DAPI staining showed that treatment with dTAT NP visibly increased the nuclear apoptosis rate of the TPC-1 cells. The effect of dTAT NP on TPC-1 cells was associated with the promotion of caspase-3 and downregulation of the PI3K/Akt signaling pathway. In summary, the present data provide a pre-clinical proof-of-concept for a novel gene delivery system that efficiently delivers NIS to the targeted cancer cells and presents a satisfactory efficacy. This approach may offer an effective strategy for improving thyroid cancer gene therapy.

The effects of perfluorocarbon on ICAM-1 expression in LPS-induced A549 cells and the potential mechanism.

Acute lung injury (ALI)/ARDS is a critical clinical syndrome with high mortality, and the effective therapeutic methods for the treatment remain limited. Previous studies have indicated that liquid ventilation with perfluorocarbon (PFC) may be advantageous over conventional mechanical ventilation in the treatment of ALI/ARDS. Additionally, PFC inhibits the inflammatory response caused by ALI/ARDS. However, the anti-inflammatory mechanism remains to be completely elucidated. In the present study, the aim was to determine the anti­inflammatory mechanism of PFC and the association with microRNA (miR). PFC was used to modulate LPS­induced A549 cells, with the cells divided into four groups: Untreated control group; LPS group, treated with 10 µg/ml LPS; LPS+PFC group, treated with 10 µg/ml LPS and PFC; and PFC group, treated with PFC alone. The intercellular adhesion molecule­1 (ICAM­1) mRNA and protein expression levels of each group were detected by reverse transcription­quantitative polymerase chain reaction (RT­qPCR) and western blotting, respectively. A549 cells were transfected with miR­17­3p mimics, miR­17­3p inhibitors or negative controls to observe the alterations in the anti­inflammatory effects of PFC. A dual luciferase reporter gene assay was used to determine whether ICAM­1 is a target gene of miR­17­3p. PFC was observed to attenuate the mRNA and protein expression levels of ICAM­1 in LPS­induced A549 cells, with no significant effect on the untreated A549 cells. miR­17­3p was demonstrated to be regulated by PFC. Transfection with miR­17­3p mimics enhanced the anti­inflammatory effects of PFC, whereas the miR­17­3p inhibitor weakened the anti­inflammatory effects of PFC at early time points. To conclude, the current study indicates that ICAM­1 was a target gene of miR­17­3p, and PFC has anti­inflammatory effects. Additionally, the present study is the first report, to the best of our knowledge, that PFC is able to attenuate ICAM-1 expression in LPS-induced A549 cells by increasing miR-17-3p expression.

Healthcare-associated Pneumonia: Clinical Features and Retrospective Analysis Over 10 Years.

BACKGROUND: Healthcare-associated pneumonia (HCAP) is associated with drug-resistant pathogens and high mortality, and there is no clear evidence that this is due to inappropriate antibiotic therapy. This study was to elucidate the clinical features, pathogens, therapy, and outcomes of HCAP, and to clarify the risk factors for drug-resistant pathogens and prognosis. METHODS: Retrospective observational study among hospitalized patients with HCAP over 10 years. The primary outcome was 30-day all-cause hospital mortality after admission. Demographics (age, gender, clinical features, and comorbidities), dates of admission, discharge and/or death, hospitalization costs, microbiological results, chest imaging studies, and CURB-65 were analyzed. Antibiotics, admission to Intensive Care Unit (ICU), mechanical ventilation, and pneumonia prognosis were recorded. Patients were dichotomized based on CURB-65 (low- vs. high-risk). RESULTS: Among 612 patients (mean age of 70.7 years), 88.4% had at least one comorbidity. Commonly detected pathogens were Acinetobacter baumannii, Pseudomonas aeruginosa, and coagulase-negative staphylococci. Initial monotherapy with ß-lactam antibiotics was the most common initial therapy (50%). Mean age, length of stay, hospitalization expenses, ICU admission, mechanical ventilation use, malignancies, and detection rate for P. aeruginosa, and Staphylococcus aureus were higher in the high-risk group compared with the low-risk group. CURB-65 ≥3, malignancies, and mechanical ventilation were associated with an increased mortality. Logistic regression analysis showed that cerebrovascular diseases and being bedridden were independent risk factors for HCAP. CONCLUSION: Initial treatment of HCAP with broad-spectrum antibiotics could be an appropriate approach. CURB-65 ≥3, malignancies, and mechanical ventilation may result in an increased mortality.

Immediate and long-term results of bronchial artery embolization for hemoptysis due to benign versus malignant pulmonary diseases.

BACKGROUND: Bronchial artery embolization (BAE) is widely used for the treatment of hemoptysis. The immediate and long-term results of BAE for hemoptysis in patients with benign and malignant pulmonary diseases were inconsistent in previous studies and were thus investigated. METHODS: This was a retrospective review of the clinical records of 154 patients (108 with benign disease and 46 with malignant disease) who received BAE for hemoptysis from January 2005 to June 2011 at the Chinese People's Liberation Army General Hospital. RESULTS: Immediate cessation of hemoptysis was achieved in 98 patients with benign disease (90.7%) and 42 patients with malignancy (91.3%). The long-term control rate of hemoptysis in patients with benign disease was 74.3% (80/108) at 1 year, significantly higher than in patients with cancer (16/46, 35.5%, P < 0.01). The worst outcomes in the benign and malignant groups were observed in patients with aspergilloma and squamous cell lung cancer, respectively. The average number of abnormal vessels on bronchial arteriography was higher in the benign group than in the malignant group (3 ± 1.3 versus 2 ± 1.1, respectively, P < 0.01). Moreover, recurrent hemoptysis was independently associated with the presence of massive hemoptysis and bronchial-pulmonary artery shunt in both groups (P < 0.05). CONCLUSIONS: BAE is a relatively safe procedure for patients with hemoptysis. Immediate control of hemoptysis with BAE is achieved in most cases, but the long-term hemoptysis control rate is worse in malignant lung diseases than in benign conditions, especially among patients with squamous cell lung cancer.

In vivo mouse

The aim of this in vivo study was to evaluate the feasibility of ^{99m}Tc-labeled cartilage link protein (CLP) probe for the single-photon emission computed tomography (SPECT) of lung cancer. Xenograft mouse model were established from a luciferase expressing cell line derived from a human lung cancer. Bioluminescence imaging (BLI) was carried out prior to ^{99m}Tc-CLP and ^{99m}Tc-methoxyisobutyl isonitrile (MIBI) SPECT scans. The image quality of ^{99m}Tc-CLP scan was validated with BLI and compared with well established ^{99m}Tc-MIBI scan. Results of multimodal imaging analyses suggested that ^{99m}Tc-CLP was a sensitive and reliable SPECT agent for lung cancer imaging.

Effects of vitamin b6 therapy for sepsis patients with linezolid-associated cytopenias: a retrospective study.

BACKGROUND: The common adverse effects of linezolid for treating septic patients with gram-positive cocci is anemia and thrombocytopenia, which limit its clinical application. OBJECTIVES: We determined the effects of vitamin B6 adjunctive therapy on linezolid-associated cytopenias, and retrospectively studied 75 septic patients who received at least 7 days of linezolid treatment. METHODS: Patients were divided into a linezolid treatment group (LTG; n = 41) that received linezolid only and a combination treatment group (CTG; n = 34) that received both linezolid and vitamin B6. Each group was further subdivided into those with sepsis and those with severe sepsis. Each patient had red blood cell (RBC), hemoglobin (Hb), hematocrit (Hct), and platelet (PLT) measurements at baseline (day 0) and every other day for 2 weeks during treatment; these parameters were compared between the groups and assessed for time-dependent trends. RESULTS: For patients in the LTG, RBC, Hb, and Hct values showed statistically significant reductions over time, and these values were lower compared with the values in the CTG. The CTG also showed downward trends, except on the first day of treatment. The PLT count also decreased in both groups. Patients with severe sepsis had lower PLT counts in both treatment groups compared with the septic patients. CONCLUSIONS: Septic patients who received a combination treatment of linezolid and vitamin B6 might show positive effects for linezolid-associated reductions in some hematologic parameters (RBC, Hb, and Hct). This combined treatment might also slow PLT reduction, which was more evident in patients with severe sepsis. ClinicalTrials.gov identifier: NCT01295801.

[Feasibility of cartilage link protein of hyaluronic acid for defining radiotherapeutic target volume in a mouse model of lung tumor].

OBJECTIVE: To investigate the feasibility of using cartilage link protein of hyaluronic acid (HA-CLP) for defining the tumor boundary in a mouse model of lung carcinoma. METHODS: Lung carcinoma was induced in KM mice by chemical carcinogenesis. HA-CLP separated from bovine cartilage and purified by affinity chromatography was labeled with (125)I for autoradiography. Immunohistochemical analysis and Western blotting were used to examine the efficiency of HA-CLP in defining the boundaries of the lung tumors. RESULTS: With autoradiography, the clearest image of lung cancer was obtained at 2 h. With immunohistochemical method, the tumor boundary was the most clearly displayed at 2 h when the strongest signals of HA-CLP was detected; Western blotting also showed the clearest bands of HA-CLP at 2 h. CONCLUSION: HA-CLP has the immunogenicity of HABP, and can efficiently indicate lung tumor boundary in autoradiography and immunohistochemistry.

[Adult hospital acquired pneumonia: a multicenter study on microbiology and clinical characteristics of patients from 9 Chinese cities].

OBJECTIVE: To investigate the etiology and clinical characteristics of hospital-acquired pneumonia (HAP) in China and to provide evidence for appropriate therapy. METHODS: We performed a prospective multicenter study in 13 Chinese urban tertiary hospitals. All HAP cases diagnosed at respiratory general ward and respiratory intensive care unit (RICU) from August 2008 to December 2010 were studied. Epidemiological data, etiology and clinical characteristics of enrolled patients were collected. Sputum or tracheal aspirate and blood cultures, Legionella antibodies and Streptococcus pneumoniae urinary antigen tests were performed. Bacteria to antimicrobial susceptibility test was performed. RESULTS: A total of 610 cases of HAP were diagnosed during the study, with an overall incidence of 1.4% among 42 877 hospitalized patients, while the incidence was 0.9% (362/41 261) in respiratory general ward and 15.4% (248/1616) in RICU. 93.9% (573 cases) of patients had at least one underlying disease, and 91.0% (555 cases) had exposure to at least one antimicrobial agent within 90 days prior to HAP diagnosis. Pathogens were identified in 487 patients, with Acinetobacter baumannii [30.0% (183/610)], Pseudomonas aeruginosa [22.0% (134/610)], Staphylococcus aureus [13.4% (82/610)] and Klebsiella pneumonia [9.7% (59/610)] being the most common pathogens. Eighteen patients (3.0%) had infection with fastidious bacteria. A. baumannii and S. aureus were the more frequent pathogens in the ventilator-associated pneumonia (VAP) cases [50.5% (97/192) and 21.4% (41/192)] as compared to non-VAP cases [20.6% (86/418) and 9.8% (41/418), P < 0.01]. A. baumannii and S. aureus were also frequent pathogens in cases with a score of more than 20 by the acute physiology and chronic health evaluation II (APACHEII) scoring [45.7% (69/151) and 20.5% (31/151)], as compared to cases with a score of less than 20 of APACHE II [24.8% (114/459) and 11.1% (51/459), P < 0.01]. A. baumannii showed high resistance rates to carbapenems [more than 70% (109/142)], and the susceptibility to cefoperazone/sulbactam, polymyxin B and tigecycline were 40.8% (58/142), 99.3% (141/142) and 95.8% (136/142) respectively. Resistance rates of P. aeruginosa to meropenem and imipenem were 48.8% (40/82) and 70.7% (58/82) respectively. Methicillin-resistant S. aureus (MRSA) accounted for 87.8% (43/49) in all strains of S. aureus. Mortality rate of VAP cases was 34.5% (61/177), significantly more than that of HAP patients [22.3% (135/605), P < 0.05]. The average hospital stay of patients with HAP was (23.8 ± 20.5) days, significantly more than that of the average for inpatients [(13.2 ± 13.6) days, P < 0.01] during the study period. Mean costs of HAP were (108 950 ± 116 608) yuan, significantly higher than the average hospital costs of respiratory inpatients (17 999 ± 33 364) yuan. CONCLUSIONS: Among Chinese patients hospitalized in urban tertiary medical centers, HAP incidence and mortality rate were high, which increased the patients' hospital stay and the medical costs. Common pathogens were A. baumannii, P. aeruginosa, S. aureus and K. pneumonia. The common bacteria of HAP in China showed high resistance rates to antibiotics.

Bone marrow-derived mesenchymal stem cells protect rats from endotoxin-induced acute lung injury.

BACKGROUND: Acute lung injury (ALI) is a serious and common condition for which there are currently no specific strategies for treatment. Recent studies have suggested that bone marrow-derived multipotent mesenchymal stem cells (MSCs) may have therapeutic applications in multiple clinical disorders. We explored the biological effects of MSCs during endotoxin-induced ALI and the mechanisms involved. METHODS: MSCs were isolated from male rat bone marrow and the ALI model was induced by intravenous endotoxin injection. Female rats were sacrificed at 6 hours, 24 hours, 4 days, 1 week and 3 weeks post-injection of MSCs or saline and the lung tissue, bronchoalveolar lavage fluid, and serum were harvested for analysis. We further evaluated the survival of the rats and examined the effects of endotoxin-induced injury on the interaction between alveolar macrophages (AMs) and MSCs in ex vivo. RESULTS: There was a significant decrease in numbers of neutrophils in bronchoalveolar lavage fluid (P < 0.05), and myeloperoxidase activity in the lung (P < 0.01), and of TNF-α and IL-1ß in serum (P < 0.05) in the MSC treated rats at 4 days. Furthermore, MSC treated rats exhibited improved survival, lower lung injury score, higher concentration of IL-10 in the serum and a reduced hydroxyproline content, but these differences were not statistically significant. Moreover, co-cultures of MSCs and AMs had significantly reduced levels of TNF-α, IL-1ß and macrophage inflammatory protein (MIP)-1α and significantly increased levels of IL-10 (P < 0.05) in the culture supernatants. CONCLUSIONS: Treatment with intravenous injection of bone marrow-derived MSCs have beneficial effects on endotoxin-induced ALI in rats. The beneficial effect might be achieved through the engraftment of differentiated MSCs in the lungs and appears derive more from their capacity to secrete soluble factors that modulate immune responses.

Perfluorocarbon attenuates lipopolysaccharide-mediated inflammatory responses of alveolar epithelial cells in vitro.

BACKGROUND: Toll-like receptor-4 (TLR-4) is integrally involved in lipopolysaccharide (LPS) signaling and has a requisite role in the activation of nuclear factor-κB (NF-κB). The exact mechanisms that lend perfluorocarbon (PFC) liquids a cytoprotective effect have yet to be elucidated. Therefore we examined in an in vitro model the cytoprotective effect of PFC on LPS-stimulated alveolar epithelial cellls (AECs). METHODS: AECs (A549 cells, human lung adenocarcinoma cell line) were divided into four groups: control, PFC, LPS and LPS + PFC (coculture group) groups. Intercellular adhesion molecule-1 (ICAM-1) was detected by ELISA, tumor necrosis factor-α (TNF-α) and interleukin-8 (IL-8) were detected by radioimmunological methods. The expression of TLR-4 mRNA and protein was detected by real time PCR and Western blotting, respectively. The activation of NF-κB was detected by Western blotting (proteins of I-κBa and NF-κB p65). RESULTS: ICAM-1, TNF-α and IL-8 were significantly increased in LPS-stimulated AECs groups. The expression of TLR-4 mRNA and protein in LPS-stimulated groups was markedly increased. Meanwhile, NF-κB was activated as indicated by the significant degradation of IκB-α and the significant release of NF-κB P65 and its subsequent translocation into the nucleus. There were no significant effects of PFC alone on any of the factors studied while the coculture group showed significant downregulation of the secretion of ICAM-1, TNF-α and IL-8, the expression of TLR-4 mRNA and the activity of NF-κB. CONCLUSIONS: Taken together, our results demonstrate that LPS can induce AEC-related inflammatory injury via the activation of TLR-4 and subsequent activation of NF-κB. PFC is able to protect AECs from LPS-induced inflammatory injury by blocking the initiation of the LPS signaling pathway, which is indicated by the significant decrease of TLR-4 expression and NF-κB activation.

[Investigation of inflammatory responses of alveolar epithelial cells induced by lipopolysaccharide and mechanism].

OBJECTIVE: Lipopolysaccharide (LPS) can activate alveolar epithelial cells (AECs) and induce inflammatory injury. Toll-like receptor-4 (TLR-4) is integrally involved in LPS signaling and has a requisite role in the activation of NF-κB. NF-κB is a key intercellular signaling event that mediates cell inflammatory responses. The aim of the study is to investigate in an in vitro model the inflammatory responses of AECs induced by LPS and the probable mechanism underlined the observed inflammatory responses. So cytokines of ICAM-1, TNF-α and IL-8 secreted by LPS-activated AECs were observed. And the initial signal molecule (the expression of TLR-4 mRNA) and the key intracellular steps (the activation of NF-κB) were studied in detail. METHODS: The study was performed on A549 cells (Human lung adenocarcinoma cell line). A549 cells were divided into two groups: control, and LPS interference group. Proinflammatory cytokines ICAM-1, TNF-α and IL-8 were detected by ELISA or radioimmunological methods. The expression of TLR-4 mRNA was detected by real time PCR. The activation of NF-κB was detected by Western blot (proteins of I-κBα and NF-κB p65). RESULTS: Compared with control group, ICAM-1 and TNF-α of LPS-stimulated group were significantly higher and peaked after 2h before gradually declining at 6 and 12 h; IL-8 was higher after 2 h, which continued up to 12 h. The expression of TLR-4 mRNA of LPS group was significantly higher and peaked after 2 h and gradually declining at 6 and 12 h. Meanwhile, NF-κB was activated after 0.5, 2, 6 and 12 h indicated by the significant degradation of IκB-α and the significant release of NF-κB P65 and its subsequent translocation into the nucleus approximately synchronized. CONCLUSION: Taken together, the results demonstrate that LPS can induce AECs inflammatory injury via activating TLR-4 and subsequently activating NF-κB.

A clinical study on the effects and mechanism of xuebijing injection in severe pneumonia patients.

OBJECTIVE: To observe the effects of Xuebijing Injection in patients with severe pneumonia, and to explore the mechanism. METHODS: Eighty cases of severe pneumonia are randomly assigned to the Xuebijing treatment (forty cases) and the control group (forty cases), with the same routine therapy provided in both groups. Clinical effective rates, inflammatory factors and organ function were observed in both groups. RESULTS: The effective rate was higher in Xuebijing group than that of the control group (80.0% vs. 67.5%, P < 0.05). As compared with the control group, the LDH, alpha1-AG, alpha1-AT levels and the peak body temperature decreased markedly with the Xuebijing treatment going, and the secretion of TNF-alpha, IL-6, IL-8 was suppressed in Xuebijing group; but no significant difference was found in leptin level. CONCLUSION: Xuebijing Injection may show a protective effect in patients with severe pneumonia. The mechanism is possibly with the decreased secretion of TNF-alpha, IL-6, and IL-8.

[Investigation of inflammatory responses of pulmonary microvascular endothelial cells induced by lipopolysaccharide and mechanism].

OBJECTIVE: Lipopolysaccharide (LPS) can activate pulmonary vascular endothelial cells (PMVECs) and induce inflammatory injury. Toll-like receptor-4 (TLR-4) is integrally involved in LPS signaling and has a requisite role in the activation of NF-κB. NF-κB is a key intercellular signaling event that mediates cell inflammatory responses. The aim of the study was to investigate in an in vitro model the inflammatory responses of PMVECs induced by LPS and the probable mechanism underlying the observed inflammatory responses. METHODS: The present study was performed on isolated PMVECs from Sprague-Dawley rats. After being identified, PMVECs were divided into 2 groups: a control group, and a LPS (0.01, 0.1, 1, 10 mg/L) intervention group. ICAM-1, TNF-α and IL-8 were detected by ELISA or radioimmunological methods. The expression of TLR-4 mRNA was detected by real time PCR. The activation of NF-κB was detected by Western blot (proteins of I-κBα and NF-κB p65) and immunocytochemical staining (NF-κB p65). RESULTS: Compared with the control group, cytokines secreted from PMVECs-stimulated by LPS were increased in a dose-dependent manner. When stimulated with LPS 10 mg/L for 2, 6 and 12 h, cytokines measured were all increased. ICAM-1 and TNF-α were significantly increased and peaked after 2 h before gradually declining at 6 and 12 h. IL-8 was higher after 2 h, which continued up to 12 h. The expression of TLR-4 mRNA was significantly higher and peaked after 2 h and continued to 12 h (4.34 ± 1.42, 3.62 ± 1.45, 3.32 ± 1.36), which were all higher than that of the control group (1.00 ± 0.00, P < 0.05). Meanwhile, NF-κB was activated at 0.5, 2, 6 and 12 h indicated by the significant degradation of IκB-α and the significant increased release of NF-κB P65 and its subsequent translocation into the nucleus with approximately synchronized. CONCLUSION: Taken together, the results demonstrated that LPS was able to induce PMVECs inflammatory injury via activating TLR-4 and subsequently activating NF-κB.

Gefitinib attenuates murine pulmonary fibrosis induced by bleomycin.

BACKGROUND: Gefitinib, an inhibitor of epidermal growth factor receptor (EGFR) tyrosine kinase, is an effective treatment for epithelial tumors, including non-small cell lung cancer (NSCLC), and is generally well tolerated. However, some clinical trials revealed that gefitinib exposure caused lung fibrosis, a severe adverse reaction. This study investigated the effect of gefitinib on lung fibrosis in mice. METHODS: We generated a mouse model of lung fibrosis induced by bleomycin to investigate the fibrotic effect of gefitinib. C57BL/6 mice were injected intratracheally with bleomycin or saline, with intragastric administration of gefitinib or saline. Lung tissues were harvested on day 14 or 21 for histology and genetic analysis. RESULTS: The histological results showed that bleomycin successfully induced lung fibrosis in mice, and gefitinib prevented lung fibrosis and suppressed the proliferation of S100A4-positive fibroblast cells. In addition, Western blotting analysis revealed that gefitinib decreased the expression of phosphorylated EGFR (p-EGFR). Furthermore, quantitative real-time PCR (qRT-PCR) demonstrated that gefitinib inhibited the accumulation of collagens I and III. CONCLUSIONS: These results reveal that gefitinib reduces pulmonary fibrosis induced by bleomycin in mice and suggest that administration of small molecule EGFR tyrosine kinase inhibitors has the potential to prevent pulmonary fibrosis by inhibiting the proliferation of mesenchymal cells, and that targeting tyrosine kinase receptors might be useful for the treatment of pulmonary fibrosis in humans.