Integrin αVß1-activated PYK2 promotes the progression of non-small-cell lung cancer via the STAT3-VGF axis.

BACKGROUND: Non-small-cell lung cancer (NSCLC) accounts for 80-85% of all lung cancer and is the leading cause of cancer-related deaths globally. Although various treatment strategies have been introduced, the 5-year survival rate of patients with NSCLC is only 20-30%. Thus, it remains necessary to study the pathogenesis of NSCLC and develop new therapeutic drugs. Notably, PYK2 has been implicated in the progression of many tumors, including NSCLC, but its detailed mechanism remains unclear. In this study, we aimed to elucidate the mechanisms through which PYK2 promotes NSCLC progression. METHODS: The mRNA and protein levels of various molecules were measured using qRT-PCR, western blot (WB), and immunohistochemistry (IHC), respectively. We established stable PYK2 knockdown and overexpression cell lines, and CCK-8, EdU, and clonogenic assays; wound healing, transwell migration, and Matrigel invasion assays; and flow cytometry were employed to assess the phenotypes of tumor cells. Protein interactions were evaluated with co-immunoprecipitation (co-IP), immunofluorescence (IF)-based colocalization, and nucleocytoplasmic separation assays. RNA sequencing was performed to explore the transcriptional regulation mediated by PYK2. Secreted VGF levels were examined using ELISA. Dual-luciferase reporter system was used to detect transcriptional regulation site. PF4618433 (PYK2 inhibitor) and Stattic (STAT3 inhibitor) were used for rescue experiments. A public database was mined to analyze the effect of these molecules on NSCLC prognosis. To investigate the role of PYK2 in vivo, mouse xenograft models of lung carcinoma were established and examined. RESULTS: The protein level of PYK2 was higher in human NSCLC tumors than in the adjacent normal tissue, and higher PYK2 expression was associated with poorer prognosis. PYK2 knockdown inhibited the proliferation and motility of tumor cells and caused G1-S arrest and cyclinD1 downregulation in A549 and H460 cells. Meanwhile, PYK2 overexpression had the opposite effect in H1299 cells. The siRNA-induced inhibition of integrins alpha V and beta 1 led to the downregulation of p-PYK2(Tyr402). Activated PYK2 could bind to STAT3 and enhance its phosphorylation at Tyr705, regulating the nuclear accumulation of p-STAT3(Tyr705). This further promoted the expression of VGF, as confirmed by RNA sequencing in a PYK2-overexpressing H1299 cell line and validated by rescue experiments. Two sites in promoter region of VGF gene were confirmed as binding sites of STAT3 by Dual-luciferase assay. Data from the TGCA database showed that VGF was related to the poor prognosis of NSCLC. IHC revealed higher p-PYK2(Tyr402) and VGF expression in lung tumors than in adjacent normal tissues. Moreover, both proteins showed higher levels in advanced TNM stages than earlier ones. A positive linear correlation existed between the IHC score of p-PYK2(Tyr402) and VGF. Knockdown of VGF inhibited tumor progression and reversed the tumor promoting effect of PYK2 overexpression in NSCLC cells. Finally, the mouse model exhibited enhanced tumor growth when PYK2 was overexpressed, while the inhibitors PF4618433 and Stattic could attenuate this effect. CONCLUSIONS: The Integrin αVß1-PYK2-STAT3-VGF axis promotes NSCLC development, and the PYK2 inhibitor PF4618433 and STAT3 inhibitor Stattic can reverse the pro-tumorigenic effect of high PYK2 expression in mouse models. Our findings provide insights into NSCLC progression and could guide potential therapeutic strategies against NSCLC with high PYK2 expression levels.

The exhaustion of lymphocytes is the main factor that decreases the sensitivity of QFT-GIT detection in silicosis.

BACKGROUND: Tuberculosis infection is a major complication of silicosis, but there is no study on whether silicosis can affect the sensitivity of QuantiFERON-TB Gold In-Tube (QFT-GIT) assays. This study will analyze the relationship between silicosis and QFT-GIT, determine the main factor of the QFT-GIT sensitivity decrease in silicosis and explore the methods to increase the sensitivity. METHODS: Silicosis patients with positive tubercle bacillus cultures were collected. The QFT-GIT, flow cytometry and blocking antibodies were used. RESULTS: The sensitivity of QFT-GIT in silicosis patients (58.46%) was significantly decreased and the expression of PD-1 on T cells and CD56+NK cells in pulmonary tuberculosis combined with silicosis were higher than normal tuberculosis patients and silicosis only patients. Further analysis found that the ratio of PD-1+CD4+T and IFN-γwere negatively correlated and blockaded the PD-1 pathway with antibodies can restore the sensitivity of QFT-GIT in silicosis. CONCLUSIONS: This is the first study to analyze the relationship between immune exhaustion and QFT-GIT in silicosis and found that the sensitivity of QFT-GIT was decreased by the expression of PD-1 on lymphocytes. Antibody blocking experiments increased the expression of IFN-γ and provided a new method to improve the sensitivity of QFT in silicosis. The study also found that silicosis can increase PD-1 expression. As PD-1 functions in infectious diseases, it will promote immune exhaustion in silicosis and lead to tuberculosis from latent to active infection. The study provided theoretical evidence for the diagnosis and immunotherapy of silicosis complications, and it has great value in clinical diagnostics and treatment.

Characterization of multiple soluble immune checkpoints in individuals with different Mycobacterium tuberculosis infection status and dynamic changes during anti-tuberculosis treatment.

BACKGROUND: Immune checkpoints are crucial for the maintenance of subtle balance between self-tolerance and effector immune responses, but the role of soluble immune checkpoints (sICs) in Mycobacterium tuberculosis (M. tb) infection remains unknown. We assessed the levels of multiple sICs in individuals with distinct M. tb infection status, and their dynamic changes during anti-tuberculosis treatment. METHODS: We enrolled 24 patients with pulmonary tuberculosis, among which 10 patients were diagnosed with tuberculous pleurisy (TBP), 10 individuals with latent tuberculosis infection (LTBI), and 10 healthy volunteers from Wuxi Fifth People's Hospital and Huashan Hospital between February 2019 and May 2021. Plasma concentrations of thirteen sICs were measured at enrollment and during anti-tuberculosis treatment using luminex-based multiplex assay. sICs levels in tuberculous pleural effusion (TPE) and their relations to laboratory test markers of TPE were also assessed in TBP patients. RESULTS: The circulating levels of sPD-1, sPD-L1, sCTLA-4, sBTLA, sGITR, sIDO, sCD28, sCD27 and s4-1BB were upregulated in tuberculosis patients than in healthy controls. A lower sPD-L1 level was found in LTBI individuals than in tuberculosis patients. In TBP patients, the levels of sPD-1, sPD-L2, sCD28, sCD80, sCD27, sTIM-3, sLAG-3, sBTLA, s4-1BB and sIDO increased significantly in TPE than in plasma. In TPE, sBTLA and sLAG-3 correlated positively with the adenosine deaminase level. sIDO and sCD80 correlated positively with the lactate dehydrogenase level and the percentage of lymphocytes in TPE, respectively. Meanwhile, sCD27 correlated negatively with the specific gravity and protein level in TPE. In tuberculosis patients, the circulating levels of sBTLA and sPD-L1 gradually declined during anti-tuberculosis treatment. CONCLUSIONS: We characterized the changing balance of sICs in M. tb infection. And our results revealed the relations of sICs to laboratory test markers and treatment responses in tuberculosis patients, indicating that certain sICs may serve as potential biomarkers for disease surveillance and prognosis of tuberculosis.

The predictive value of pulmonary function test before transplantation for chronic pulmonary graft-versus-host-disease after allogeneic hematopoietic stem cell transplantation.

BACKGROUND: Pulmonary chronic graft-versus-host disease (cGVHD) after allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a devastating complication and often diagnosed at a late stage when lung dysfunction is irreversible. Identifying patients before transplant who are at risk may offer improved strategies to decrease the mortality. Bronchiolitis obliterans syndrome (BOS) is the typical manifestation of pulmonary cGVHD, which is clinically diagnosed by pulmonary function test (PFT). This study aimed to evaluate the predictive value of PFT pre-HSCT for BOS. METHODS: A single center cohort of 923 allo-HSCT recipients was analyzed, including 15 patients who developed pulmonary cGVHD. Kaplan-Meier method was used to analyze the 3 year progression free survival and 3 year overall survival (OS). A Cox regression model was applied for univariate and multivariate models. RESULTS: The 3 year cumulative incidence of pulmonary cGVHD was 2.04% (95% CI 1.00-3.08%). According to the cut-off values determined by receiver operator characteristic curve, higher ratio of forced expiratory volume during one second to forced vital capacity (FEV1/FVC) pre-HSCT was correlated to a lower incidence of pulmonary cGVHD [0.91% (95% CI 0.01-1.81%) vs. 3.61% (95% CI 1.30-5.92%), P < 0.01], and so as peak expiratory flow to predictive value (PEF/pred) [0.72% (95% CI 0-1.54%) vs. 3.74% (95% CI 1.47-6.01%), P < 0.01]. Multivariate analysis showed that FEV1/FVC (HR = 3.383, P = 0.047) and PEF/pred (HR = 4.426, P = 0.027) were independent risk factors for onset of BOS. Higher FEV1/FVC and PEF/pred level were related to a significantly decreased 3 year non-relapse mortality. The 3 year OS was superior in patients with higher PEF/pred [78.17% (95% CI 74.50-81.84%) vs. 71.14% (95% CI 66.08-76.20%), P = 0.01], while FEV1/FVC did not show significance difference. CONCLUSION: Our results suggested that PFT parameters such as PEF/pred and FEV1/FVC could be predictors for pulmonary cGVHD and even transplant outcomes before HSCT.

A Low Lean-to-Fat Ratio Reduces the Risk of Acute Exacerbation of Chronic Obstructive Pulmonary Disease in Patients with a Normal or Low Body Mass Index.

BACKGROUND Increased risk of acute exacerbation of chronic obstructive pulmonary disease (COPD) has been reported in patients who are overweight and obese. However, the effects of body fat in patients with normal or low body mass index (BMI) and COPD remain unknown. This study aimed to examine the association between acute exacerbations of COPD and the lean-to-fat (LTF) ratio in patients with a normal or low BMI. MATERIAL AND METHODS Patients with COPD (n=68) underwent assessment of body composition, in whom 43 cases had a normal BMI (18.5 to 25 kg/m²) and 14 cases were underweight (<18.5 kg/m²). Patients with COPD were treated according to current clinical guidelines and underwent regular follow-up for one year. Acute exacerbations of COPD were recorded. RESULTS BMI, the fat-free mass index (FFMI), skeletal muscle mass index (SMMI), and LTF ratio had no significant effect of the risk of acute exacerbations of COPD in the whole study cohort, but a low LTF ratio was significantly associated with reduced risk of acute exacerbations of COPD in the subgroup with a BMI<25 kg/m² (OR=4.528; P<0.05). The Fat Mass Index (FMI) had a protective effect in the whole cohort (OR=0.292; P=0.024) and in the subgroup with BMI <25 kg/m² (OR=0.253, P=0.049). The cumulative incidence of acute exacerbations of COPD was significantly increased in the patients with a high LTF ratio in the whole cohort (P=0.047) and in the subgroup with BMI <25 kg/m² (P=0.014). CONCLUSIONS In patients with BMI <25 kg/m², the LTF ratio was positively correlated with the risk of occurrence of acute exacerbations of COPD.

H19 promotes non-small-cell lung cancer (NSCLC) development through STAT3 signaling via sponging miR-17.

LncRNAs can exhibit crucial roles in the development of multiple cancers, including non-small cell lung cancer (NSCLC). Currently, we investigated the role of lncRNA H19 in NSCLC. In our study, it was found that H19 was upregulated in A549 and H1299 cells compared to normal lung epithelial BEAS-2B cells. Meanwhile, we observed that miR-17 was downregulated in NSCLC cell lines. Inhibited H19 can suppress the growth, migration, and invasion of NSCLC cells and bioinformatics search was performed to predict the correlation between H19 and miR-17. Overexpression of miR-17 was able to inhibit the progression of NSCLC cells while reversely miR-17 inhibitors reversed this process. In addition, signal transducers and activators of transcription (STAT3), as an mRNA target of miR-17, was presented in our research. Moreover, we discovered that H19 demonstrated its biological functions via regulating miR-17 and STAT3 in vitro. Silencing H19 greatly increased STAT3 expression by sponging miR-19 in vitro. It was hypothesized that H19 may serve as a competing endogenous RNA (ceRNA) to modulate STAT3 by attaching miR-17 in lung cancer. In summary, our findings indicated that H19/miR-17/STAT3 axis participated in NSCLC development. H19 could be regarded as a significant prognostic biomarker in NSCLC progression.

Analysis of B7-H4 expression in metastatic pleural adenocarcinoma and therapeutic potential of its antagonists.

BACKGROUND: The increasing incidence and poor outcome associated with malignant pleural effusion (MPE) requires finding an effective treatment for this disease. Inhibitory B7-H4 is expressed in many different human cancers but its role in malignant pleural tissue has yet to be established. METHODS: Here, patients with metastatic pleural adenocarcinoma (MPA) or with early-stage lung adenocarcinoma were clinically and statistically analyzed. Immunohistochemistry and confocal microscopy were used to determinate the expression of B7-H4 in the cancer cells. By using MPE model, we sought to a potential immunotherapy for MPE with anti-B7-H4 mAb. RESULTS: When compared to early-stage lung adenocarcinoma, MPA possessed higher level of nuclei membranous B7-H4 and lower cytoplasmic B7-H4 expression. Also, nuclei membranous B7-H4 expression was found to be positively correlated to Ki-67 expression, and indicated a possible poor prognosis of MPA. In mouse MPE model, intra-pleurally injection of anti-B7-H4 mAb effectively suppressed MPE formation. CONCLUSIONS: Taken together, our data was in support of the significance of B7-H4 expression in MPA, which also suggest it warrants further exploration for potential immunotherapy of MPE.

Sudden Cardiac Arrest Triggered by Coadministration of Fluconazole and Amiodarone.

Fluconazole for fungal infections and amiodarone for arrhythmia are commonly prescribed medications, and coadministration of such medications is sometimes inevitable in clinical practice. However, both medications have been associated with prolonged QTc intervals and subsequent arrhythmias, which are sometimes fatal. We present the case of a 75-year-old man with sudden cardiac arrest triggered by coadministration of fluconazole and amiodarone, which raises the need for caution regarding coadministration of these medications. To our knowledge, this case has not been previously described.

Value of Ki-67 and computed tomography in the assessment of peripheral lung adenocarcinoma.

PURPOSE: This study was designed to determine whether proliferation antigen Ki-67 and/or a computed tomography (CT) value could be used to evaluate the clinical-pathological features of peripheral lung adenocarcinoma. MATERIALS AND METHODS: A total of 116 eligible lung cancer patients were enrolled. Nodule size, lymph node metastasis, differentiation, Ki-67 expression and CT findings were assessed. The relationship between clinic parameters and the CT feature was analysed statistically. RESULTS: The percentage of lesions that had ground-glass opacity or localised air bronchogram was significantly greater in low CT value group (<30, p < 0.05). No significant association was observed between CT value and size in the subgroup with CT value > 0 (p = 0.66). As a proliferative marker of lung cancer, Ki-67 was present in a total of 115 (99.9%) of the 116 evaluable primary lung cancers. There was a statistically significant correlation between the Ki-67 index and CT value (p < 0.05). Compared to CT value, Ki-67 index possessed higher sensitivity to predict the differentiation and lymph node metastasis of peripheral lung adenocarcinoma, adding of CT value would enhance its specificity. CONCLUSION: Combination of Ki-67 expression and CT value determination was useful for the classification of differentiation and metastatic or proliferative potential of peripheral lung adenocarcinoma.

[Diagnostic value of computed tomography-guided percutaneous needle biopsy versus radial probe endobronchial ultrasound-guided transbronchial lung biopsy in peripheral pulmonary lesions].

OBJECTIVE: To evaluate the value of computed tomography-guided percutaneous needle biopsy (CT-PNB) and radial probe endobronchial ultrasound-guided transbronchial lung biopsy (EBUS-TBLB) in the diagnosis of peripheral pulmonary lesions(PPLs). METHODS: The clinical data of 213 patients who were diagnosed as to have PPLs in the First Affiliated Hospital of Soochow University between December 1, 2012 and October 1, 2014 were studied retrospectively. The patients were divided into CT-PNB group and EBUS-TBLB group, according to biopsy methods. The diagnostic yield, complications and influencing factors of both groups were evaluated. RESULTS: The diagnostic yield (87.2%, 102/117) and complication rate (18.8%, 22/117) of the CT-PNB group were higher than those of the EBUS-TBLB group(61.5%, 59/96 and 18.8%, 22/117, respectively), the differences being statistically significant (χ(2)=18.906, P=0.000 and χ(2)=10.542, P=0.001, respectively). Analysis of the influencing factors showed that there were statistically significant correlations between pneumothorax and the lesion diameter(χ(2)=5.785, P=0.016) and location(χ(2)=7.559, P=0.006) in the CT-PNB group. The diagnostic yield was correlated with lesion diameter(χ(2)=7.995, P=0.004) and location(χ(2)=4.608, P=0.027) in the EBUS-TBLB group. There was no complicated pneumothorax if the lesions were attached to the chest wall in the CT-PNB group. CONCLUSIONS: The diagnostic yield of CT-PNB in PPLs was better than that of EBUS-TBLB. Although CT-PNB had a higher complication rates, most complications were mild.

Tea intake and lung diseases: a Mendelian randomization study.

Background: Existing studies on the relationship between tea intake and lung diseases have yielded inconsistent results, leading to an ongoing dispute on this issue. The impact of tea consumption on the respiratory system remained elucidating. Materials and methods: We conducted a two-sample Mendelian randomization (MR) study to evaluate the associations between five distinct tea intake phenotypes and 15 different respiratory outcomes using open Genome-wide association study (GWAS) data. The inverse variance weighted (IVW) was used for preliminary screening and a variety of complementary methods were used as sensitivity analysis to validate the robustness of MR estimates. Pathway enrichment analysis was used to explore possible mechanisms. Results: IVW found evidence for a causal effect of standard tea intake on an increased risk of lung squamous cell cancer (LSCC) (OR = 1.004; 95% CI = 1.001-1.007; P = 0.00299). No heterogeneity or pleiotropy was detected. After adjustment for potential mediators, including smoking, educational attainment, and time spent watching television, the association was still robust in multivariable MR. KEGG and GO enrichment predicted proliferation and activation of B lymphocytes may play a role in this causal relation. No causalities were observed when evaluating the effect of other kinds of tea intake on various pulmonary diseases. Conclusion: Our MR estimates provide causal evidence of the independent effect of standard tea intake (black tea intake) on LSCC, which may be mediated by B lymphocytes. The results implied that the population preferring black tea intake should be wary of a higher risk of LSCC.

Long Noncoding RNA RP11-89K21.1 Interacts with miR-146a/b-5p to Promote Proliferation and Gefitinib Resistance Through Regulating RHPN2 and RhoA/ROCK Pathway in Lung Adenocarcinoma.

Background: Long noncoding RNAs (lncRNAs) have major roles in lung adenocarcinoma (LUAD). lncRNA RP11-89K21.1 was reported to be abnormally expressed in LUAD, yet its biological functions in LUAD progression remain unclear. Materials and Methods: Forty LUAD tissues and pair-matched adjacent normal tissues were enrolled in this study. Quantitative real-time polymerase chain reaction was performed to detect the expression of lncRNA, miRNA, and mRNA in LUAD samples and cell lines. Loss-of-function assays were used to evaluate the effects of RP11-89K21.1 on LUAD cell proliferation and gefitinib resistance. Bioinformatics analysis, luciferase reporter assay, and Western blot were employed to explore the regulatory relationships among RP11-89K21.1, miR-146a/b-5p, and RHPN2. Results: RP11-89K21.1 was identified as being highly expressed in LUAD tissues and cell lines. Moreover, upregulated RP11-89K21.1 was strongly associated with unfavorable overall survival of patients with LUAD. Knockdown of RP11-89K21.1 significantly suppressed proliferation and sensitized cell to gefitinib. Mechanistically, RP11-89K21.1 could directly bind miR-146a-5p and miR-146b-5p and decrease their expression to upregulate RHPN2, and subsequently activated RhoA/ROCK pathway. More importantly, overexpression of RHPN2 reversed regulatory effects of RP11-89K21.1 knockdown on cell proliferation and gefitinib resistance. Conclusions: These observations provide new insights into the role of RP11-89K21.1 in regulating LUAD tumorigenesis, suggesting that RP11-89K21.1 is a potential therapeutic target for LUAD treatment.

Imbalance of circulating T-lymphocyte subpopulation in COPD and its relationship with CAT performance.

Chronic obstructive pulmonary disease (COPD) is associated with systemic effects, and T-cell-mediated immunity was involved in the COPD. COPD Assessment Test (CAT) could provide a valid, reliable, and standardized measure of COPD health status. The objective of this study was determination of lymphocyte subpopulation in patients with stable COPD (n = 52) and to ascertain if a relationship existed between T-lymphocyte subpopulation and CAT performance. The stable COPD patients were assessed with CAT, and divided into four groups with score >30 (n = 8), 20< score ≤30 (n = 16), 10< score ≤20 (n = 20), and score ≤10 (n = 8). Spearman's rank correlation was used to determine the relationship between proportion of T lymphocyte and CAT score. We found an elevated proportion of CD8(+) cells in COPD patients of the group with score >30 compared to other groups. Proportion of CD4(+) cells was significantly lower in the groups with score >30 and 20< score ≤30 when compared to groups with 10< score ≤20 and score ≤10. The CD4(+) :CD8(+) ratio was also significantly lower in the groups with score >30 and 20< score ≤30. Of note are the correlations of proportion of CD8(+) cells and CD4(+) :CD8(+) ratio with CAT performance when score >20. No correlations existed between proportion of CD4(+) , CD8(+) cells, CD4(+) :CD8(+) ratio, and CAT performance when score ≤20. Our results show that the determinants of T-lymphocyte subpopulation in COPD patients were value to assess physical conditions. We considered CD4(+) and CD8(+) T lymphocytes to be a representative and stable parameter in grading of health status in COPD patients.

BOLA3 is a prognostic-related biomarker and correlated with immune infiltrates in lung adenocarcinoma.

BACKGROUD: BolA gene family contains three members, high expression of BolA family member 2 (BOLA2) has been reported to be associated with prognosis of several cancers. However, the relationship between BOLA3 and lung adenocarcinoma (LUAD) remains unclear. METHODS: Expression of BOLA3 was analyzed by online database. Co-expressed genes and gene set enrichment analysis (GSEA) were performed using LinkedOmics. Diagnostic value was assessed using receiver operating characteristic (ROC) curves. The prognostic value of BOLA3 was analyzed using Prognoscan and Kaplan-Meier Plotter. The Tumor Immune Estimation Resource (TIMER) and Single-sample Gene Set Enrichment Analysis (ssGSEA) were used to explore the relationship between BOLA3 and tumor immune infiltration. RESULTS: The results showed that the expression of BOLA3 was significantly higher in LUAD than in normal tissues. High expression of BOLA3 was associated with T stage, N stage, pathologic stage (all P < 0.001). In addition, elevated expression of BOLA3 was associated with overall survival (OS) and progression-free survival (PFS) in LUAD ((OS:HR = 2.58, log-rank P = 1.3e - 11; PFS:HR = 2.36, log-rank P = 4.1e - 05). BOLA3 expression level has negative correlations with infiltrating levels of B cells, CD4 +T cells, macrophages, neutrophils, and dendritic cells (DCs). GSEA analysis showed BOLA3 joined mainly in mitochondrial respiratory chain complex assembly, translational initiation, etc. CONCLUSIONS: These results showed up-regulated in BOLA3 was correlated with poor prognosis and immune infiltrates in LUAD, BOLA3 can be served as a potential immunotherapy target.

Improving Suspected Pulmonary Infection Diagnosis by Bronchoalveolar Lavage Fluid Metagenomic Next-Generation Sequencing: a Multicenter Retrospective Study.

Metagenomic next-generation sequencing (mNGS) has been gradually applied to clinical practice due to its unbiased characteristics of pathogen detection. However, its diagnostic performance and clinical value in suspected pulmonary infection need to be evaluated. We systematically reviewed the clinical data of 246 patients with suspected pulmonary infection from 4 medical institutions between January 2019 and September 2021. The diagnostic performances of mNGS and conventional testing (CT) were systematically analyzed based on bronchoalveolar lavage fluid (BALF). The impacts of mNGS and CT on diagnosis modification and treatment adjustment were also assessed. The positive rates of mNGS and CT were 47.97% and 23.17%, respectively. The sensitivity of mNGS was significantly higher than that of CT (53.49% versus 23.26%, P < 0.01), especially for infections of Mycobacterium tuberculosis (67.86% versus 17.86%, P < 0.01), atypical pathogens (100.00% versus 7.14%, P < 0.01), viruses (92.31% versus 7.69%, P < 0.01), and fungi (78.57% versus 39.29%, P < 0.01). The specificity of mNGS was superior to that of CT, with no statistical difference (90.32% versus 77.42%, P = 0.167). The positive predictive value (PPV) and negative predictive value (NPV) of mNGS were 97.46% and 21.88%, respectively. Diagnosis modification and treatment adjustment were conducted in 32 (32/246, 13.01%) and 23 (23/246, 9.35%) cases, respectively, according to mNGS results only. mNGS significantly improved the diagnosis of suspected pulmonary infection, especially infections of M. tuberculosis, atypical pathogens, viruses, and fungi, and it demonstrated the pathogen distribution of pulmonary infections. It is expected to be a promising microbiological detection and diagnostic method in clinical practice. IMPORTANCE Pulmonary infection is a heterogeneous and complex infectious disease with high morbidity and mortality worldwide. In clinical practice, a considerable proportion of the etiology of pulmonary infection is unclear, microbiological diagnosis being challenging. Metagenomic next-generation sequencing detects all nucleic acids in a sample in an unbiased manner, revealing the microbial community environment and organisms and improving the microbiological detection and diagnosis of infectious diseases in clinical settings. This study is the first multicenter, large-scale retrospective study based entirely on BALF for pathogen detection by mNGS, and it demonstrated the superior performance of mNGS for microbiological detection and diagnosis of suspected pulmonary infection, especially in infections of Mycobacterium tuberculosis, atypical pathogens, viruses, and fungi. It also demonstrated the pathogen distribution of pulmonary infections in the real world, guiding targeted treatment and improving clinical management and prognoses.

EPHA5 Enhances the Stemness of Non-small cell lung cancer Cells Through Activating the Wnt Signaling Pathway.

PURPOSE: To explore the effect of erythropoietin-producing hepatocellular receptor A5 (EPHA5) on the stemness of non-small cell lung cancer cells and its molecular mechanism. METHODS: Highly-expressed EPHA5 in NCI-H460 and NCI-H1229 cells was silenced. After EPHA5 silencing, the positive expression level of cluster of differentiation 133 (CD133) in NCI-H460 and NCI-H1229 cells was detected by flow cytometry, and the expression levels of stemness markers sex determining region Y-box 2 (Sox2), Nanog, Kruppel-like factor 4 (KLF4) and octamer-binding transcription factor 4 (Oct4) in cells were detected by Western blotting. RESULTS: The expression level of EPHA5 in non-small cell lung cancer H460 and H1229 cells was higher than that in A549 and SPC-A1 cells. After EPHA5 silencing, the levels of CD133 and stemness markers Sox2, Nanog, KLF4 and Oct4 in H460 and H1229 cells all declined. CCK-8 assay showed that Wnt agonists at a concentration of 2.5 and 5 µm had little effect on the proliferative activity of H460 and H1229 cells. Western blotting revealed that Wnt agonists at a concentration of 5 µm could better enhance the expression of ß-catenin. After treatment with Wnt agonists, the expression of CD133 in H460 and H1229 cells with EPHA5 silencing by siRNA3 was higher than that before treatment, and the expression levels of Sox2, Nanog, KLF4 and Oct4 in the above two cells were also increased compared with those before treatment. However, the levels of the above indexes were all lower after treatment with Wnt agonists than those before silencing. CONCLUSION: Activating the Wnt signaling pathway can induce the increase in EPHA5 expression and enhance the stemness of non-small cell lung cancer cells.

Expression of nuclear factor erythroid-2-related factor 2, broad complex-tramtrack-bric a brac and Cap'n'collar homology 1 and γ-glutamic acid cysteine synthase in peripheral blood of patients with chronic obstructive pulmonary disease and its clinical significance.

The purpose of the present study was to explore the relationship between nuclear factor erythroid 2-related factor 2 (Nrf2)/BTB-CNC allogeneic 1 (Bach1)/γ-glutamic acid cysteine synthase (γ-GCS) and chronic obstructive pulmonary disease (COPD). The expression of Nrf2, Bach1, γ-GCS mRNA and protein in the peripheral blood mononuclear cells (PBMCs) of 80 COPD patients and 40 healthy volunteers were studied. Then, the correlation between Nrf2, Bach1, γ-GCS and lung function, inflammation and oxidative stress indicators was analyzed. Compared with healthy controls, Nrf2, Bach1 mRNA and protein levels were significantly increased in the PBMCs of COPD patients, while γ-GCS mRNA and protein levels were significantly decreased. Nrf2 and Bach1 protein levels in the nucleus were significantly elevated in acute exacerbation COPD patients compared with patients with a stable stage of COPD, while γ-GCS mRNA levels were significantly reduced. In addition, it was found that Nrf2 nuclear protein levels were significantly reduced in COPD patients compared with the control group, while Bach1 nuclear protein levels were significantly increased. Correlation analysis in COPD group demonstrated that γ-GCS mRNA was positively correlated with Nrf2 nuclear protein level, but negatively correlated with Bach1 nuclear protein level. Further analysis demonstrated that γ-GCS mRNA and Nrf2 protein in the nucleus was positively correlated with forced expiratory volume in one second (FEV1)/forced vital capacity (FVC)% and FEV1% predicted, and Bach1 protein in the nucleus was negatively correlated with FEV1/FVC% and FEV1% predicted. Additionally, the expression levels of Nrf2, Bach1 and γ-GCS were also associated with smoking. The expression of Nrf2, Bach1 and γ-GCS in peripheral blood mononuclear cells of patients with COPD was dysregulated and related to lung function, which provides a new basis for exploring further the pathogenesis of COPD.

Long non-coding RNA SOX21-AS1 modulates lung cancer progress upon microRNA miR-24-3p/PIM2 axis.

Lung cancer is a lethal cancer that threatens human health. Several studies have demonstrated the role of long non-coding RNAs (lncRNAs) in lung cancer. SOX21-AS1 is a newly discovered oncogenic lncRNA, but its molecular mechanism in lung cancer is not known. Here, the levels of SOX21-AS1, miR-24-3p, and PIM2 were examined in lung cancer and normal tissues. The relationships between miR-24-3p and SOX21-AS1 or PIM2 were predicted using bioinformatics tools and confirmed using a luciferase reporter assays. Colony formation, MTT, flow cytometry, and transwell assays were conducted to analyze cell proliferation, apoptosis, migration, and invasion abilities, respectively. Western blotting was used to measure PIM2 expression levels in cancer tissues and cells. SOX21-AS1 expression levels were high in lung cancer tissues and cells. In contrast, the amount of miR-24-3p bound to SOX21-AS1 was relatively low in cancerous tissues and cells. The knockdown of SOX21-AS1 decreased cell proliferation, activated apoptosis, and promoted cell migration and invasion. These effects were abolished by miR-24-3p inhibition. The oncogenic function of SOX21-AS1 mediated through targeting miR-24-3p was also demonstrated in animal models. PIM2 was targeted by miR-24-3p and showed increased levels in tumor tissues and cells. Furthermore, miR-24-3p overexpression inhibited the proliferation and promoted the apoptosis of lung cancer cells. In lung cancer cells, SOX21-AS1 negatively modulated the miR-24-3p/PIM2 axis to facilitate their proliferation, migration, and invasion. These findings offer a novel idea for future research on treating lung cancer at the molecular level.

[The influence of chronic intermittent hypoxia on insulin function in OSAHS patients].

OBJECTIVE: To discuss the influence of chronic intermittent hypoxia and secundum oxidative stress on insulin resistance and the function of Pancreatic beta cells in OSAHS patients. METHODS: 15 patients with primary snoring, 31 patients with light to moderate OSAHS and 34 patients with severe OSAHS were investigated. General clinical data of every patient were collected, while nocturnal data were gained by using PSG with entire night monitor. 8-isoprostane and fasting insulin (FINS) were detected through ELISA; GSH-Px through Spectrophotometer. Insulin resistance was evaluated by measuring HOMA-IR and FINS, while the function of Pancreatic beta cells by HOMA-beta. The comparison among the three groups were used by the non-parametric Kruskal-Wallis H test, and Nemenyi test was used for pairwise comparison. The relativity between the variables were assessed by Spearman correlation test, and multiple linear regression was used to remove interference between variables. The enumeration data were compared with chi2 test. RESULTS: 1. Along with the aggravation of OSAHS, the three groups were statistically different from each other on 8-isoprostane, GSH-Px, fasting blood glucose (FBG), FINS, and HOMA-IR (P=0.0034, P<0.0001), but not on HOMA-beta (P>0.05). 2. ODI, CT90%, lowest SaO2 and mean SaO2 were correlated with FINS and HOMA-IR in the serum, while the coefficient of correlation r decreased from ODI to mean SaO2. After ruling out the influence of age, BMI by multiple linear regression, ODI, CT90% and lowest SaO2 still correlated with FINS and HOMA-IR, the standardized regression coefficient R diminished from ODI, CT90% to lowest SaO2. 3. 8-isoprostane and GSH-Px correlated with FINS and HOMA-IR in OSAHS patients (P<0.05). After eliminated the influence of age, BMI by multiple linear regression, 8-isoprostane still correlated with FINS and HOMA-IR. 4. HOMA-beta has no correlation with AHI, BMI, ODI, CT90%, lowest SaO2, mean SaO2, 8-isoprostane and GSH-Px (P>0.05). CONCLUSION: 1. Chronic intermittent hypoxia of OSAHS was independently correlated with IR. Compared with the severity of hypoxia, the insulin resistance has a closer relation with oxygen desaturation index. 2. Secundum oxidative stress in OSAHS was independently correlated with IR. 3. We didn't find the function of Pancreatic beta cells correlating with chronic intermittent hypoxia and secundum oxidative stress.

Treatment of a case presenting as critical adenoviral ARDS using Cidofovir with early combinatorial prone ventilation and ECMO.

Here, we report a case of adenoviral pneumonia associated with critical ARDS treated with Cidofovir, prone ventilation and extracorporeal membrane oxygenation (ECMO). The patient responded well to therapy and recovered without further complications. Cidofovir, with early prone ventilation and ECMO support, may be a therapeutic option for patients with critical ARDS related to adenoviral pneumonia.