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Tuberculosis (TB), caused by mycobacterium tuberculosis (M. tuberculosis) infection, is one of the leading causes of death globally and poses a threat to public health. During infection, M. tuberculosis causes redox imbalance and dysfunctions of protective immunity. Transcription factor nuclear factor erythroid 2 (NF-E2)-related factor (Nrf2) is a major modulator of cellular redox homeostasis via transcriptional induction of cytoprotective genes to protect cell against the damage from insults. Thus, we hypothesize that Nrf2 may regulate protective immunity against M. tuberculosis. RNA-seq and immunoblotting results suggested that the expression of Nrf2 protein increased after M. tuberculosis infection, and decreased upon long-term M. tuberculosis infection, while Keap1 protein maintained a low expression level during M. tuberculosis infection. Furthermore, Nrf2 activator sulforaphane (SFN) decreased proinflammatory cytokines production, phagocytosis and host cell apoptosis, while increasing ROS levels and promoting autophagy in THP1 macrophages infected with M. tuberculosis. In addition, SFN-activated Nrf2 augmented bacterial killing by macrophages, which might be due to the regulation of protective immunity via Nrf2. Combined, our results extend the understanding of the complex innate immunity regulation by Nrf2 against mycobacterial infection. Also, these findings suggested that the regulation of Nrf2 signaling cascade could be used as a therapeutic target for the treatment of TB patients and the development of better anti-TB vaccines.

Mycobacterium tuberculosis (M. tuberculosis) is an intracellular bacteria capable of evading the human immune system through various mechanisms. M. tuberculosis secretes many virulence factors when infecting host cells. The 6 kDa early secretory antigen target (ESAT-6) is one of the most virulent factors producted by the ESX-1 system, which acts alone or in conjunction with culture filtrate protein 10 (CFP-10) to involve in host-pathogen interactions. ESAT-6 has been widely investigated in M. tuberculosis pathogenicity and vaccination. Nonetheless, little is known about its contribution to M. tuberculosis immune evasion and the involved cellular mechanisms. In this study, differential genes between BCG with the region of difference-1 (RD1) gene and Bacille Calmette-Guérin (BCG) infected dendritic cells (DC) were analyzed to reveal that ESAT-6, expressed in the RD1 region, involves host denfensive function of DC in response to BCG. In vitro evidence indicated that ESAT-6 (5 μg/ml) inhibits M. tuberculosis-induced apoptosis in THP-1(A) macrophages by suppressing TLR2 and via caspase-9 and caspase-3 endogenous pathways. It also inhibits the expression of IL-10, TNF-α, IL-12, and the phagocytosis of macrophages. Additionally, ESAT-6 suppresses the bactericidal activity of macrophages, including ROS production, and the killing of M. tuberculosis. In summary, these data suggest a central role of ESAT-6 in M. tuberculosis’s evasion from macrophage recognition, phagocytosis, killing, and apoptosis. Our work may provide a theoretical basis for exploring new molecular targets of M. tuberculosis both for innovative vaccines and therapeutic inventions.

Lung cancer is the leading cause of cancer-related deaths worldwide, synthesizing and screening of novel anti-cancer drugs provides an alternative therapeutic strategy for renewal of the chemotherapy regimens against lung cancer. To this end, several compounds were synthesized based on the modification of the original myricetin, and their anti-tumor activity against the human non-small cell lung cancer (NSCLC) A549 cells were measured. Among the myricetin derivatives, S4-10 has displayed the highest antitumor efficacy in dose-dependent manner. The proliferation of A549 cells were significantly attenuated by given 6 µM of S4-10 both in vitro and in vivo. Further, the treatment of S4-10 also results in the inhibition of cell migration and invasiveness and the induction of cell apoptosis and G2 cycle arrest of A549 cells. Moreover, we found that S4-10 inhibits the progression of A549 cells through the sterol biosynthetic-cell apoptosis axis. These findings shed the light of developing S4-10 as a promising treatment agent for NSCLC.

Studies have demonstrated that PE_PGRS45 is constitutively expressed under various environmental conditions (such as nutrient depletion, hypoxia, and low pH) of the in vitro growth conditions examined, indicating that PE_PGRS45 protein is critical to the basic functions of M.tuberculosis. However, there are few reports about the biochemical function and pathogenic mechanism of PE_PGRS45 protein. The fact that the gene of M. tuberculosis is not easily expressed in E. coli may be mainly due to the high content of G+C and the use of unique codons. Fusion tags are indispensable tools used to improve the soluble expression of recombinant proteins and accelerate the characterization of protein structure and function. In the present study, His6, Trx, His6-MBP were used as fusion tags, but only MBP-PE_PGRS45 formed expressed solubly. The purification using His6-MBP tag specific binding to the Ni column and is easy to separate after the tag cleavage. We used the purified PE_PGRS45 to immunize New Zealand rabbits and got the serum of anti PE_PGRS45. It was found that the titer of polyclonal antibodies against PE_PGR45 was higher than 1:256000. The result shows that the purified PE_PGRS45 can induce New Zealand rabbits to produce high titer antibodies. In conclusion, the recombinant protein PE_PGRS45 was successfully expressed in E. coli and specific antiserum was prepared, which will be followed by further evaluation of these specific antigens to develop highly sensitive and specific diagnostic tests for tuberculosis.

Objective To investigate the effect of protein tyrosine phosphatase receptor type O (PTPRO) on the phagocytic activity of alveolar epithelial cells in LPS-induced acute lung injury. Methods Mice were randomly divided into the normal control group and LPS stimulation group. The infiltration of inflammatory cells was detected by HE staining. The cytokine TNF-α level in lung was analyzed by ELISA. Western blotting was performed to detect the effect of LPS on PTPRO protein expression in lung. After the expression of PTPRO in MLE-12 cells was silenced by siRNA in vitro, flow cytometry was used to detect the effects of LPS and PTPRO siRNA on the phagocytic activity of MLE-12 cells, and the effects of LPS and PTPRO siRNA on the expression of PTPRO, AKT and phosphorylated AKT protein were measured by Western blotting. Results After the establishment of murine acute lung injury model by LPS injection(1 mg/kg), the infiltrated polymorphonuclear leukocytes were markedly increased. The level of TNF-α in lung tissue and the expression of PTPRO in MLE-12 cells were both significantly increased after LPS stimulation. However, the activity of MLE-12 cells to phagocytose fluorescent microbeads was evidently decreased after silencing PTPRO. Furthermore, silencing PTPRO induced a remarkable decrease in the phosphorylation of AKT in MLE-12 cells. Conclusion PTPRO can promote phagocytic activity of MLE-12 cells via activating AKT signaling pathway.

Objective To establish a THP-1 macrophage model infected by Mycobacterium smegmatis expressing green fluorescent protein (GFP), to quickly locate and visually detect Mycobacterium smegmatis, and to provide a tracer tool to identify the pathogenesis of tuberculosis and develop new tuberculosis vaccines. Methods The enhanced green fluorescent protein (EGFP) gene sequence was amplified by PCR using pEGFP-N1 plasmid as a template to obtain the coding gene of EGFP, and the amplified product was cloned into the vector pALACE to establish the recombinant plasmid pALACE-EGFP. Electroporation transformed the pALACE-EGFP into Mycobacterium smegmatis, and recombinant Mycobacterium smegmatis clones were screened by hygromycin resistance. After expanded culture, the smears were observed by fluorescence microscopy. The THP-1 macrophages were infected with recombinant Mycobacterium smegmatis, and the expression of EGFP was observed. Results The recombinant plasmid pALACE-EGFP was constructed appropriately. The recombinant Mycobacterium smegmatis was observed under fluorescence microscope. And it was confirmed that EGFP was expressed in recombinant Mycobacterium smegmatis, and THP-1 macrophages emitted green fluorescence after infection. Conclusion The successful establishment of recombinant Mycobacterium smegmatis expressing EGFP protein provides insights for investigating infection and pathogenesis of Mycobacterium tuberculosis.

Mycobacterium tuberculosis (M. tuberculosis) is the pathogen which causes tuberculosis (TB), a significant human public health threat. Co-infection of M. tuberculosis and the human immunodeficiency...

Objective To clone, express and purify PPE15 recombinant protein from Mycobacterium tuberculosis (H37Rv), as well as prepare and characterize its rabbit polyclonal antibody. Methods By The PPE15 gene was amplified from the genome of Mycobacterium tuberculosis H37Rv by PCR, and the His-tagged prokaryotic PPE15 prokaryotic expression plasmid pET28a-PPE15 was constructed by homologous recombination cloning technique, and transformed into E. coli BL21 (DE3). PPE15 expression was induced by isopropyl-β-D-thiogalactopyranoside (IPTG). Recombinant PPE15 was identified by SDS-PAGE, and further purified by affinity chromatography with a Ni-NTA column. The renaturation purified PPE15 protein was used to immunize New-Zealand rabbit to prepare polyclonal antibodies. The antibody specificity was analyzed by Western blot analysis, and antibody titer was determined by indirect ELISA. Results Recombinant prokaryotic PPE15 protein was successfully expressed and purified with a molecular weight of 38 kDa. The purified PPE15 protein exhibited positive reaction with the serum of TB patients and the PPE15 protein, the titer of the polyclonal antibodies reaches more than 1:1 300 480. Conclusion The recombinant protein PPE15 was successfully expressed and purified, and high titer rabbit-derived polyclonal antibody was prepared which provided an experimental basis for further functional studies of PPE15 protein.

Studies have demonstrated that PE_PGRS45 is constitutively expressed under various environmental conditions (such as nutrient depletion, hypoxia, and low pH) of the in vitro growth conditions examined, indicating that PE_PGRS45 protein is critical to the basic functions of

One unmet challenge in lung cancer diagnosis is to accurately differentiate lung cancer from other lung diseases with similar clinical symptoms and radiological features, such as pulmonary tuberculosis (TB). To identify reliable biomarkers for lung cancer screening, we leverage the recently discovered non-canonical small non-coding RNAs (i.e., tRNA-derived small RNAs [tsRNAs], rRNA-derived small RNAs [rsRNAs], and YRNA-derived small RNAs [ysRNAs]) in human peripheral blood mononuclear cells and develop a molecular signature composed of distinct ts/rs/ysRNAs (TRY-RNA). Our TRY-RNA signature precisely discriminates between control, lung cancer, and pulmonary TB subjects in both the discovery and validation cohorts and outperforms microRNA-based biomarkers, which bears the diagnostic potential for lung cancer screening. Supplementary Information The online version contains supplementary material available at 10.1186/s12943-020-01280-9.

Objective To clone proline-glutamate 8 (PE8) gene segment from Mycobacterium tuberculosis (H37Rv), construct the recombinant plasmid pET28a-PE8, express recombinant PE8 protein, and prepare its polyclonal antibody. Methods Using a standard homologous recombination cloning technology, we cloned the PE8 gene into the prokaryotic vector pET28a. After sequence confirmation, it was transformed into E. coli BL21 (DE3) and treated with 0.5 mmol/L isopropyl-beta-D-thiogalactopyranoside (IPTG) to induce protein expression. We purified and renatured the recombinant PE8 protein, and immunized New Zealand rabbits to prepare the polyclonal antibody. Antibody titer was determined by indirect ELISA and the specificity was evaluated by Western blot analysis. Results The recombinant plasmid pET28a-PE8 was successfully constructed, and the PE8 protein was primarily expressed in an inclusion body in E. coli. After renaturation and purification, a purity of about 90% of the recombinant protein was achieved. The titer of the polyclonal antibody was higher than 1:430 080. The polyclonal antibody could specifically recognize the recombinant PE8 protein. Conclusion We have successfully expressed and purified recombinant PE8 protein, which can be further utilized to generate PE8 polyclonal antibody with acceptable titer and specificity.

Sepsis-associated encephalopathy (SAE) is a devastating neurological complication of sepsis with intolerable high motility. SAE is accompanied with brain vascular injury, endothelial hyperpermeability, and neutrophil infiltration into the brain tissue, key inflammatory processes leading to further brain edema and neuronal cell apoptosis. Recent studies from us and others suggest that the chemokine receptor C-X-C Motif Chemokine Receptor 2 (CXCR2) is crucial for neutrophil recruitment during SAE. Here we use CXCR2 antagonist SB225002 to characterize the role of CXCR2 in brain infiltration of neutrophil in a murine model of SAE. Systemic administration of high-dose LPS (10 mg/kg) induced evident neutrophil infiltration into the cerebral cortex in wild-type mice. However, CXCR2 antagonist SB225002 markedly attenuated neutrophil infiltration into brain. The CXCR2 expression on neutrophils in the peripheral circulation was dramatically downregulated in response to this LPS dose, and endothelial CXCR2 was significantly upregulated, suggesting endothelial but not neutrophil CXCR2 plays a more important role in neutrophil infiltration into brain. Strikingly, although these CXCR2 antagonist SB225002 treated mice displayed reduced neutrophil infiltration, no change in neutrophil rolling and adhesion was observed. Furthermore, we confirmed that CXCR2 agonist CXCL1 induced a marked increase in actin stress fiber synthesis and paracellular gap formation in cultured cerebral endothelial cells, which is attenuated by SB225002. Thus, these results demonstrate a selective role for endothelial CXCR2 to regulate cerebral vascular permeability and neutrophil transmigration in high-dose LPS induced neuroinflammation, and also suggest a therapeutic potential of CXCR2 antagonist SB225002 in SAE.

Objective To investigate the role of CXCR2 in the cerebral endothelial activation and migration of neutrophils into the brain in septic encephalopathy (SE) induced by lipopolysaccharide (LPS). Methods C57BL/6J mice and CXCR2-knockout mice were randomly divided into a normal control group, a wildtype mice group with LPS treatment and CXCR2-knockout mice group with LPS treatment. Mouse SE models were induced by intraperitoneal LPS injection. Naphthol AS-D chloroacetate histochemical staining of the brain section was performed to quantitate the neutrophils infiltrating into the cerebral cortex. TNF-α and CXCL1 concentrations in the brain and plasma were determined by ELISA. After the stimulation of LPS (1 μg/mL) and TNF-α (200 ng/mL), the levels of CXCR2 protein in the primary mouse brain microvascular endothelial cells isolated from the cerebral cortex were detected by Western blotting. The levels of F-actin and vascular cell adhesion molecule 1 (VCAM-1) protein in the primary mouse brain microvascular endothelial cells stimulated by CXCL1 (100 ng/mL) were detected by Western blotting. Results After intraperitoneal LPS injection, there was a significant increase in the level of TNF-α in the brain and plasma and there was also an evident increase in the level of CXCL1 in the brain of wild type mice (C57BL/6J mice). And intraperitoneal LPS injection caused increased neutrophil infiltration into the cerebral cortex in the wild type mice (C57BL/6J mice). But CXCR2-knockout mice displayed evidently reduced neutrophil infiltration into the cerebral cortex compared with the wildtype mice. In vitro LPS and TNF-α upregulated the expression of CXCR2 in the primary brain microvascular endothelial cells. CXCL1 increased remarkably the expression of endothelial F-actin and VCAM-1. Conclusion In the SE model, CXCR2 participates in the cerebral endothelial activation and neutrophil migration into the brain.

Additional file 1: Supplementary Figures. Fig. S1. The workflow of the study. Fig. S2. The landscape of non-canonical sncRNAs in human PBMCs. Fig. S3. The mapping profile of tsRNAs. Fig. S4. Comparison of the expression of the prioritized sncRNA subcategories between lung cancer stages. Fig. S5. Comparison of the expression of the prioritized sncRNA subcategories between lung cancer histological types. Fig. S6. Comparison of the expression of the prioritized sncRNA subcategories between the lung cancer patients with and without lymph node involvement. Fig. S7. Comparison of the expression of the prioritized sncRNA subcategories between the lung cancer patients with and without distant metastasis. Fig. S8. Comparison of the expression of the prioritized sncRNA subcategories between the lung cancer patients with and without smoking history. Fig. S9. The TRY-RNA signature. Fig. S10. The TRY-RNA index in the discovery cohort. Fig. S11. Comparison of the expression of the sncRNA species within the TRY-RNA signature between lung cancer stages. Fig. S12. Comparison of the expression of the sncRNA species within the TRY-RNA signature between lung cancer histological types. Fig. S13. Comparison of the expression of the sncRNA species within the TRY-RNA signature between the lung cancer patients with and without lymph node involvement. Fig. S14. Comparison of the expression of the sncRNA species within the TRY-RNA signature between the lung cancer patients with and without distant metastasis. Fig. S15. Comparison of the expression of the sncRNA species within the TRY-RNA signature between the lung cancer patients with and without smoking history. Fig. S16. Expression heatmap of the MIR signature in the discovery cohort. Fig. S17. The MIR index in the discovery cohort. Fig. S18. Comparison between the TRY-RNA and MIR signatures.

Additional file 2: Supplementary Tables. Table S1. The human subjects of the discovery cohort. Table S2. The TRY-RNA signature. Table S3. The human subjects of the validation cohort. Table S4. The MIR signature.

Additional file 3. Methods.

Lung cancer is a leading cause of cancer‑related morbidity and mortality worldwide. Although there are currently various therapeutic strategies including surgery, chemotherapy and radiotherapy, lung cancer still results in high mortality with a 5‑year survival rate of less than 20%. The increasing need for new therapeutic targets and diagnostic/prognostic tools for lung cancer has promoted the demand for a better molecular and mechanistic understanding of its pathobiology. microRNA‑30a‑3p (miR‑30a‑3p) was recently recognized to be closely involved in the regulation of cancer cell invasion, migration and proliferation. However, the mechanistic role of miR‑30a‑3p in regulating the biological behavior of lung cancer, especially lung adenocarcinoma (LADC), is unknown. In the present study, we aimed to confirm the downregulation of miR‑30a‑3p in LADC tissues, and validate its functional impact on the pathogenesis of LADC via its molecular target, canopy fibroblast growth factor signaling regulator 2 (CNPY2), a known oncogene. Our data confirmed that CNPY2 was upregulated in LADC tissues, and the expression level of CNPY2 was correlated with the clinical outcomes of lung cancer patients. miR‑30a‑3p was confirmed as a key negative regulator of CNPY2 and reduced miR‑30a‑3p expression resulted in CNPY2 upregulation in LADC tissues. We then validated the functional outcome of miR‑30a‑3p in cancer pathobiology by the overexpression of miR‑30a‑3p in the LADC EKVX cell line. miR‑30a‑3p overexpression inhibited cancer cell proliferation, invasion and migration, by suppressing CNPY2 expression. In addition, miR‑30a‑3p inhibited epithelial‑mesenchymal transition, a key feature of LADC, via CNPY2 suppression. Taken together, these findings suggest that miR‑30a‑3p exerts a novel inhibitory role in the pathogenesis of LADC via CNPY2 downregulation, and the miR‑30a‑3p/CNPY2 pathway is a potential therapeutic target for human LADC.

Preeclampsia (PE) is a pregnancy disorder with a high mortality rate. Patients with PE exhibit systemic high oxidative stress status and inflammatory immune activation. This study aims to define the role of H2O2 in the activation of neutrophils and T lymphocytes in PE patients. CD3+/HLA-DR+ cells in blood from PE patients are remarkably increased compared with those of normal non-pregnancies or normal pregnancies, while the percentage of CD3+/CD62L+ cells is significantly reduced in PE patients compared to normal pregnancies. Furthermore, CD62L levels in granulocytes of periphery blood of PE patients are significantly higher than non-pregnancies, but significantly lower than normal pregnancies. To characterize the effects of intracellular reactive oxygen species (ROS) on T lymphocyte activation in PE patients, PBMCs from normal pregnancies were challenged with H2O2, and intracellular ROS levels in neutrophil granulocytes, as well as T cell surface marker levels, have been determined. We confirm that H2O2 exposure increases intracellular ROS levels in neutrophil granulocytes, and increases the proportion of CD3+/HLA-DR+ cells, but does not alter the percentage of CD3+/CD62L+ cells in PBMCs. Our study has confirmed dysregulated CD3+/HLA-DR+ and CD3+/CD62L+ T lymphocytes in PE patient peripheral blood, and the dysregulative effects of H2O2 on T lymphocyte activation, suggesting a novel mechanism of immune activation in PE.

Background Circular RNAs (circRNAs) are a class of novel RNAs with important biological functions, and aberrant expression of circRNAs has been implicated in human diseases. However, the feasibility of using blood circRNAs as disease biomarkers is largely unknown. Methods We explored the potential of using human peripheral blood mononuclear cell (PBMC) circRNAs as marker molecules to diagnose active pulmonary tuberculosis (TB). Findings First, we demonstrated that circRNAs are widely expressed in human PBMCs and that many are abundant enough to be detected. Second, we found that the magnitude of PBMC circRNAs in TB patients was higher than that in the paired healthy controls. Compared with host linear transcripts, the circRNAs within several pathways are disproportionately upregulated in active TB patients, including “Cytokine-cytokine receptor interaction”, “Chemokine signaling pathway”, “Neurotrophin signaling pathway”, and “Bacterial invasion of epithelial cells”. Based on the differentially expressed circRNAs within these pathways, we developed a PBMC circRNA-based molecular signature differentiating active TB patients from healthy controls. We validated the classification power of the PBMC circRNA signature in an independent cohort with the area under the receiver operating characteristic curve (AUC) at 0.946. Interpretation Our results suggest that PBMC circRNAs are potentially reliable marker molecules in TB diagnosis., Highlights • Circular RNAs (circRNAs) are abundant in human peripheral blood mononuclear cells (PBMCs). • CircRNAs are dysregulated in PBMCs from patients with active tuberculosis (TB) compared with healthy controls. • A PBMC circRNA-based molecular signature can differentiate active TB patients from healthy controls. Circular RNAs (circRNAs) are emerging as novel regulatory RNAs that have been found in various tissues and cell types. Notably, circRNAs are enriched and stable in whole blood, platelets, and exosomes, which makes circRNAs in human peripheral blood good candidates for diagnostic or prognostic biomarkers of human diseases. Tuberculosis (TB) is an infectious disease caused by Mycobacterium tuberculosis. Some novel diagnostic approaches, particularly a rapid, low-cost, and non-sputum-based tests, are essential to identify patients with active TB. Here, we propose that circRNA expression in human peripheral blood mononuclear cells potentially delivers useful information in TB diagnosis.

Hepatitis B is one of the most common infectious diseases, which leads to public health problems in the world, especially in Asian counties. In recent years, extensive human genetic association studies have been carried out to identify susceptible genes and genetic polymorphisms to understand the genetic contributions to the disease progression of HBV infection. HLA-DQ gene variations have been reported to be associated with HBV infection/clearance, disease progression and the development of hepatitis B-related complications, including liver cirrhosis (LC) and hepatocellular carcinoma (HCC). However, the results are either inconclusive or controversial. Therefore, to derive a more precise estimation of the association, a meta-analysis was performed. Our data revealed that the HLA-DQ alleles rs2856718-G, rs7453920-A and rs9275319-G were significantly associated with decreased risk of HBV infection and HBV natural clearance. Logistic regression analyses showed that HLA-DQ alleles rs9275572-A significantly increased HBV infection clearance, and decreased HBV natural clearance. However, rs2856718-G and rs9275572-A were not associated with development of cirrhosis. The HLA-DQ polymorphisms (rs2856718 and rs9275572) were associated with a decreased HBV-related HCC risk in all genetic models, but rs9272105-A increased the risk of HBV-related HCC. In addition, no significant association was observed between HLA-DQ rs9275319-G polymorphism and HBV-related HCC. These stratified analyses were limited due to relatively modest size of correlational studies. In future, further investigation on a large population and different ethnicities are warranted. Our findings contribute to the personalized care and prognosis in hepatitis B.

Increased exposure to ambient particulate matter (PM) is associated with elevated morbidity and mortality in patients with cardiopulmonary diseases and cancer. We and others have shown that PM induces lung microvascular barrier dysfunction which potentially enhances the systemic toxicity of PM. However, the mechanisms by which PM disrupts vascular endothelial integrity remain incompletely explored. We hypothesize that PM induces endothelial cell (EC) cytoskeleton rearrangement via Rho GTPase-dependent pathways to facilitate vascular hyperpermeability. Fine PM induced time-dependent activation of cytoskeletal machinery with increases in myosin light chain (MLC) phosphorylation and EC barrier disruption measured by transendothelial electrical resistance (TER), events attenuated by the Rho-dependent kinase (ROCK) inhibitor Y-27632 or the reactive oxygen species (ROS) scavenger, N-acetylcysteine (NAC). Both Y-27632 and NAC prevented PM-induced stress fiber formation and phospho-MLC accumulation in human lung ECs. PM promotes rapid accumulation of Rho-GTP. This event is attenuated by NAC or knockdown of RhoA (siRNA). Consistent with ROCK activation, PM induced phosphorylation of myosin light chain phosphatase (MYPT) at Thr850, a post-translational modification known to inhibit phosphatase activity. Furthermore, PM activates the guanine nucleotide exchange factor (GEF) for Rho, p115, with p115 translocation to the cell periphery, in a ROS-dependent manner. Together these results demonstrate that fine PM induces EC cytoskeleton rearrangement via Rho-dependent pathways that are dependent upon the generation of oxidative stress. As the disruption of vascular integrity further contributes to cardiopulmonary physiologic derangements, these findings provide pharmacologic targets for prevention of PM-induced cardiopulmonary toxicity.

Autophagy has been identified as an important immune regulatory mechanism. Recent studies have linked macrophage autophagy with innate immune responses against Mycobacterium tuberculosis (M. tuberculosis), which can survive within macrophages by blocking fusion of the phagosome with lysosomes. These findings suggest that autophagy is a regulatable cellular mechanism of M. tuberculosis defense in macrophages. Transcriptomic profiles in human blood in TB patients suggest that M. tuberculosis affects autophagy related pathways. In order to better understand the role of macrophage autophagy in enhancing protective immunity against M. tuberculosis, in this study, we investigate the effects of the autophagy activators rapamycin and LPS in macrophage autophagy and immunity against M. tuberculosis. We confirm that rapamycin and LPS induce autophagy in M. tuberculosis infected THP-1-derived macrophages or PMA primed THP-1 macrophages [THP-1(A)]. LPS restores M. tuberculosis-inhibited IL-12 synthesis and secretion in THP-1(A) cells via autophagy. Similarly, autophagy activators increase IL-12 synthesis and secretion in THP-1(A) cells. These studies demonstrate the importance of autophagy in M. tuberculosis elimination in macrophages and may lead to novel therapies for tuberculosis and other bacterial infections.

Airway inflammation can be mitigated when apoptotic cells are engulfed by pulmonary epithelial cells. Insulin-like growth factor 1 (IGF-1), a single chain polypeptide growth factor, is the main mediator of growth hormone activity in vivo. IGF-1 has many biological activities, such as the regulation of cell survival, proliferation, differentiation and metabolism. However, its effect on the engulfment of cells, especially by non-professional phagocytes such as alveolar epithelial cells (AECs), has not been fully elucidated. We report that IGF-1 increases endocytosis in a mouse alveolar epithelial cell line, MLE-12. The PI3K-Akt pathway is involved in this effect of IGF-1. Furthermore, IGF-1 can inhibit the production of interleukin-6 in lipopolysaccharide-stimulated AECs. We have found that IGF-1 can enhance endocytosis of AECs through the PI3K pathway and exhibit anti-inflammatory properties. These two observations suggest that IGF-1 is a potential mediator in the regulation of airway inflammation.

Background Non-small cell lung carcinomas (NSCLC) are prevalent, lethal cancers with especially grim prospects due to late-stage detection and chemoresistance. Circular RNAs (circRNAs) are non-coding RNAs that participate in tumor development. However, the role of circRNAs in NSCLC is not well known. This study investigated the role of one circRNA – circPTPRA– in NSCLC and characterized its molecular mechanism of action. Methods circPTPRA expression was analyzed in human NSCLC tumors and matched healthy lung tissue. We performed functional characterization in NSCLC cell lines and a mouse xenograft model of NSCLC to elucidate the molecular role of circPTPRA in epithelial-mesenchymal transitioning (EMT). We also assessed the regulatory action of circPTPRA on the microRNA miR-96-5p and its target the tumor suppressor Ras association domain-containing protein 8 (RASSF8). Findings circPTPRA was significantly downregulated in NSCLC tumors relative to matched healthy lung tissue. Lower circPTPRA levels correlated with metastasis and inferior survival outcomes in NSCLC patients. circPTPRA suppressed EMT in NSCLC cell lines and reduced metastasis in the murine xenograft model by sequestering miR-96-5p and upregulating RASSF8. Correlation analyses in patient-derived NSCLC tumor specimens supported the involvement of the circPTPRA/miR-96-5p/RASSF8/E-cadherin axis dysregulation in NSCLC tumor progression. Interpretation circPTPRA suppresses EMT and metastasis of NSCLC cell lines by sponging miR-96-5p, which upregulates the downstream tumor suppressor RASSF8. The circPTPRA/miR-96-5p/RASSF8/E-cadherin axis can be leveraged as a potential treatment avenue in NSCLC. Fund The Key research and development projects of Anhui Province (201904a0720079), the Natural Science Foundation of Anhui Province (1908085MH240), the Graduate Innovation Program of Bengbu Medical College (Byycx1843), the National Natural Science Foundation of Tibet (XZ2017ZR-ZY033) and the Science and Technology Project of Shannan (SNKJYFJF2017-3) and Academic Subsidy Project for Top Talents in Universities of Anhui in 2019 (gxbjZD16)

MicroRNAs (miRNAs/miRs) are small, noncoding RNA molecules that are closely associated with the occurrence and development of tumors. miR-20b is overexpressed in hepatocellular carcinoma cell lines and tissues. However, it is not clear whether miR-20b can promote the proliferation of hepatocellular carcinoma cells. In the present study, the proliferation of H22 mouse hepatocellular carcinoma cells was detected using the Cell Counting Kit-8 assay. MiRanda software was used to predict the binding sites of miR-20b to the 3′-untranslated region (3′-UTR) of phosphatase and tensin homolog (PTEN). The 3′-UTR sequence of the PTEN gene was amplified using the polymerase chain reaction in H22 cells. The recombinant plasmid or empty plasmid was co-transfected with miR-20b mimics or miR-20b scramble into HeLa cells, and luciferase activity was assessed by Dual-Luciferase(®) Reporter Assay System 24 h post-transfection. In the present study, miR-20b knockdown significantly inhibited the proliferation of H22 mouse hepatocellular carcinoma cells. In addition, miR-20b inhibition upregulated the expression of PTEN, and it was revealed that miR-20b may directly target the 3′-untranslated region of the PTEN gene. Downregulation of PTEN partially reversed the anti-proliferative effect of miR-20b on H22 cells. In conclusion, miR-20b may promote H22 cell proliferation by targeting PTEN, providing a rationale for further study investigating novel therapeutic strategies for liver cancer.

During infection and host defense, nuclear factor, erythroid 2-like 2 (Nrf2) dependent signaling is an efficient antioxidant defensive mechanism used by host cells to control the destructive effects of reactive oxygen species. This allows for effective defense responses against microbes while minimizing oxidative injury to the host cell itself. As a central regulator of antioxidant genes, Nrf2 has gained great attention in its pivotal role in infection, especially in tuberculosis (TB), the top infectious disease killer worldwide. To elucidate the genes potentially regulated by Nrf2 in TB, we conducted a meta-analysis on published gene expression datasets. Firstly, we compared the global gene expression profiles between control and Nrf2-deficient human cells. The differentially expressed genes were deemed as "Nrf2-mediated genes". Next, the whole blood gene expression pattern of TB patients was compared with that of healthy controls, pneumonia patients, and lung cancer patients. We found that the genes deregulated in TB significantly overlap with the Nrf2-mediated genes. Based on the intersection of Nrf2-mediated and TB-regulated genes, we identified an Nrf2-mediated 17-gene signature, which reflects a cluster of gene ontology terms highly related to TB physiology. We demonstrated that the 17-gene signature can be used to distinguish TB patients from healthy controls and patients with latent TB infection, pneumonia, or lung cancer. Also, the Nrf2-mediated gene signature can be used as an indicator of the anti-TB therapeutic response. More importantly, we confirmed that the predictive power of the Nrf2-mediated 17-gene signature is significantly better than the random gene sets selected from the human transcriptome. Also, the 17-gene signature performs even better than the random gene signatures selected from TB-associated genes. Our study confirms the central role of Nrf2 in TB pathogenesis and provides a novel and useful diagnostic method to differentiate TB patients from other human subjects.

L1 cell adhesion molecule (L1CAM) promotes invasiveness and metastasis in non-small cell lung cancer (NSCLC) cells and is upregulated by the p53-regulated transcription factor Slug. p21-activated kinase 4 (PAK4) directly phosphorylates Slug, resulting in pro-malignant Slug stabilization. We hypothesized that microRNA-based negative regulation of PAK4 would reduce L1CAM-induced NSCLC aggressiveness via destabilizing Slug. We found that elevated L1CAM expression was tightly correlated with p53 loss-of-function and reduced NSCLC patient survival. L1CAM suppression reduced NSCLC cell migration and invasiveness in vitro as well as tumor formation and distal metastasis in vivo. Mechanistically, p53 restricts L1CAM expression through the β-catenin/Slug pathway, with levels of β-catenin and Slug positively correlating with L1CAM expression in NSCLC tumors. The microRNA miR-193a-3p directly targets PAK4 and suppresses downstream p-Slug and L1CAM expression. Silencing PAK4, Slug, and L1CAM mirrored miR-193a-3p's effects upon the migration and invasiveness of NSCLC cells in vitro. Decreased miR-193a-3p levels correlated with elevated PAK4, p-Slug, and L1CAM levels in NSCLC tumors. Our findings support a model of miR-193a-3p as a suppressor of metastatic disease progression in NSCLC via modulation of the p53/Slug/L1CAM pathway.

Supplemental material, Supplementary figure for Hydrogen peroxide promotes the activation of preeclampsia peripheral T cells by Jingzhu Lv, Xiaojie Zhang, Caizhi Wang, Hongtao Wang, Ting Wang and Zhongqing Qian in Innate Immunity

miR-20b is a member of the miR-106a-363 gene cluster, located on the X chromosome. miR-20b regulates the expression of multiple genes in vivo and it is closely related to the occurrence and development of many diseases. These diseases include inflammatory diseases and tumor development, amongst others. However, few studies have focused on the regulation of the miR-20b gene itself. In this study, we are using the miRBase database to obtain the upstream 2000 bp sequence of the miR-20b precursor. Bio-informatics software P-MATCH 1.0 and AliBaba2 werethen used to predict transcription factor binding in the upstream region. Sp-1 was identified as one of the most probable transcription factors regulating miR-20b gene expression. After treatment with a Sp-1 siRNA, the expression of miR-20b was significantly increased in RAW264.7 cells, as well as peritoneal and alveolar macrophages. In addition, the interference with Sp-1 gene expression also reversed the decrease in miR-20b expression in RAW264.7 cells induced by TNF-α. These results indicate that Sp-1 negatively regulates the expression of miR-20b in macrophages. This finding suggests the potential of Sp-1 as a target for therapies in diseases that are associated with miR-20b overexpression.

Papillary thyroid carcinoma (PTC) is the most common endocrine malignancy. It has been demonstrated that micro-RNAs (miRNAs) are involved in the development of PTC. The miRNA-chromatin immunoprecipitation microarray assay revealed down-regulation of miR-219-5p; however, the effect of miR-219-5p on PTC cell growth remains unknown. This result implied the critical role of miR-219-5p in the development of PTC.We investigated the association between miR-219-5p and PTC development. Expression of miR-219-5p was monitored in 30 PTC tissue specimens and compared with that in 30 normal thyroid tissue specimens. The effect of miR-219-5p on PTC development was studied by cell proliferation, migration, and apoptosis assays. The underlying mechanism was clarified by a reporter assay and rescue experiment.The current study confirmed that miR-219-5p expression was inhibited in PTC tissue samples. There were statistically significant differences in the expression of miR-219-5p with regard to sex, tumor size, and lymph node metastasis in patients with PTC. Forced expression of miR-219-5p suppressed PTC cell proliferation and migration and promoted apoptosis. Further study showed that estrogen receptor (ER) α was the direct target of miR-219-5p and mediated the effect of miR-219-5p on PTC occurrence. Expression of miR-219-5p was inversely correlated with that of ERα. Importantly, ERα overexpression in PTC cells rescued the inhibitory effect of miR-219-5p on PTC cell proliferation and migration. Thus, our results indicated that miR-219-5p played a critical role in PTC growth by inhibiting ERα.Our investigation identified miR-219-5p as a negative regulator of PTC development through targeting of ERα.

Hepatocellular carcinoma (HCC) is one of the most common causes of cancer-related mortality worldwide. Host gene variants may influence hepatitis B virus- (HBV-) related HCC. Human leukocyte antigens (HLA) play an important role in presenting virus antigens to immune cells that are responsible for the clearance of virus-infected cells and tumor cells. Previous studies have investigated the HLA-DQ (rs2856718 and rs9275572) polymorphisms that may be associated with the development of HBV-related HCC. However, the results are controversial or inconclusive. Hence, we conducted a meta-analysis to derive a more precise estimation of the associations. A total of 6 articles were used to evaluate the effect of the two polymorphisms on the risk of HBV-related HCC. Odds ratios (ORs) and 95% confidence intervals (95% CIs) were calculated. We found that rs2856718 and rs9275572 in HLA-DQ significantly decreased HBV-related HCC in total population, especially in Chinese, but not in Saudi Arabian. Further validation of our results in larger populations and different ethnicities are required.

Nuclear factor, erythroid 2-like 2 (NFE2L2), a transcription factor also known as NF-E2-related factor 2 (Nrf2), is a key cytoprotective gene that regulates critical antioxidant and stress-responsive genes. Nrf2 has been demonstrated to be a promising therapeutic target and useful biomarker in malignant disease. We hypothesized that NFE2L2-mediated gene expression would reflect cancer severity and progression. We conducted a meta-analysis of microarray data for 240 NFE2L2-mediated genes that were enriched in tumor tissues. We then developed a risk scoring system based on NFE2L2 gene expression profiling and designated 50 tumor-associated genes as the NFE2L2-associated molecular signature (NAMS). We tested the relationship between this gene expression signature and both recurrence-free survival and overall survival in lung cancer patients. We find that NAMS predicts clinical outcome in the training cohort and in 12 out of 20 validation cohorts. Cox proportional hazard regressions indicate that NAMS is a robust prognostic gene signature, independent of other clinical and pathological factors including patient age, gender, smoking, gene alteration, MYC level and cancer stage. NAMS is an excellent predictor of recurrence-free survival and overall survival in human lung cancer. This gene signature represents a promising prognostic biomarker in human lung cancer.

In inflammatory sites, high molecular weight hyaluronan fragments are degraded into lower molecular weight hyaluronan fragments (LMW-HA) to regulate immune responses. However, the function of LMW-HA in PTC progression remains to be elucidated. In this study, we found that receptor of LMW-HA, TLR4, was aberrantly overexpressed in PTC tissues and cell line W3. Exposure of W3 cells to LMW-HA promoted cell proliferation and migration via TLR4. Knockdown of TLR4 has provided evidence that TLR4 is essential for LMW-HA-induced CXCR7 expression, which is responsible for LMW-HA-induced proliferation and migration of W3 cells. In tumor-bearing adult nude mice, stimulation of LMW-HA on W3 cells promotes CXCR7 expression in tumor masses (P=0.002) and tumor growth (P<0.001). To further confirm our findings, we investigated the clinicopathologic significance of TLR4 and CXCR7 expression using immumohistochemistry in 135 human PTC tissues and 56 normal thyroid tissue samples. Higher rates of TLR4 (53%) and CXCR7 (24%) expression were found in PTC tissues than in normal tissues. Expression of TLR4 or CXCR7 is associated with tumor size and lymph node metastasis. Therefore, LMW-HA may contribute to the development of PTC via TLR4/CXCR7 pathway, which may be a novel target for PTC immunomodulatory therapy.