ISL1/SHH/CXCL12 signaling regulates myogenic cell migration during mouse tongue development.

Migration of myoblasts derived from the occipital somites is essential for tongue morphogenesis. However, the molecular mechanisms of myoblast migration remain elusive. In this study, we report that deletion of Isl1 in the mouse mandibular epithelium leads to aglossia due to myoblast migration defects. Isl1 regulates the expression pattern of chemokine ligand 12 (Cxcl12) in the first branchial arch through the Shh/Wnt5a cascade. Cxcl12+ mesenchymal cells in Isl1ShhCre embryos were unable to migrate to the distal region, but instead clustered in a relatively small proximal domain of the mandible. CXCL12 serves as a bidirectional cue for myoblasts expressing its receptor CXCR4 in a concentration-dependent manner, attracting Cxcr4+ myoblast invasion at low concentrations but repelling at high concentrations. The accumulation of Cxcl12+ mesenchymal cells resulted in high local concentrations of CXCL12, which prevented Cxcr4+ myoblast invasion. Furthermore, transgenic activation of Ihh alleviated defects in tongue development and rescued myoblast migration, confirming the functional involvement of Hedgehog signaling in tongue development. In summary, this study provides the first line of genetic evidence that the ISL1/SHH/CXCL12 axis regulates myoblast migration during tongue development.

ISL1 regulates lung branching morphogenesis via Shh signaling pathway.

Lung branching morphogenesis relies on a complex coordination of multiple signaling pathways and transcription factors. Here, we found that ablation of the LIM homeodomain transcription factor Islet1 (Isl1) in lung epithelium resulted in defective branching morphogenesis and incomplete formation of five lobes. A reduction in mesenchymal cell proliferation was observed in Isl1ShhCre lungs. There was no difference in apoptosis between the wild-type (ShhCre) and Isl1ShhCre embryos. RNA-Seq and in situ hybridization analysis showed that Shh, Ptch1, Sox9, Irx1, Irx2, Tbx2, and Tbx3 were downregulated in the lungs of Isl1ShhCre embryos. ChIP assay implied the Shh gene served as a direct target of ISL1, since the transcription factor ISL1 could bind to the Shh epithelial enhancer sequence (MACS1). Also, activation of the Hedgehog pathway via ectopic gene expression rescued the defects caused by Isl1 ablation, confirming the genetic integration of Hedgehog signaling. In conclusion, our works suggest that epithelial Isl1 regulates lung branching morphogenesis through administrating the Shh signaling mediated epithelial-mesenchymal communications.

Discovery of Kebanmycins with Antibacterial and Cytotoxic Activities from the Mangrove-Derived Streptomyces sp. SCSIO 40068.

Mangrove derived actinomycetes are a rich reservoir of bioactive natural products and play important roles in pharmaceutical chemistry. In a screen of actinomycetes from mangrove rhizosphere sedimental environments, the isolated strain Streptomyces sp. SCSIO 40068 displayed strong antibacterial activity. Further fractionation of the extract yielded four new compounds kebanmycins A-D (1-4) and two known analogues FD-594 (5) and the aglycon (6). The structures of 1-6 were determined based on extensive spectroscopic data and single-crystal X-ray diffraction analysis. 1-3 featured a fused pyranonaphthaxanthene as an integral part of a 6/6/6/6/6/6 polycyclic motif, and showed bioactivity against a series of Gram-positive bacteria and cytotoxicity to several human tumor cells. In addition, the kebanmycins biosynthetic gene cluster (keb) was identified in Streptomyces sp. SCSIO 40068, and KebMT2 was biochemically characterized as a tailoring sugar-O-methyltransferase, leading to a proposed biosynthetic route to 1-6. This study paves the way to further investigate 1 as a potential lead compound.

Hedgehog pathway negatively regulated depleted uranium-induced nephrotoxicity.

Depleted uranium (DU) retains the radiological toxicities, which accumulates preferentially in the kidneys. Hedgehog (Hh) pathway plays a critical role in tissue injury. However, the role of Hh in DU-induced nephrotoxicity was still unclear. This study was carried out to investigate the effect of Gli2, which was an important transcription effector of Hh signaling, on DU induced nephrotoxicity. To clarify it, CK19 positive tubular epithelial cells specific Gli2 conditional knockout (KO) mice model was exposed to DU, and then histopathological damage and Hh signaling pathway activation was analyzed. Moreover, HEK-293 T cells were exposed to DU with Gant61 or Gli2 overexpression, and cytotoxicity of DU as analyzed. Results showed that DU caused nephrotoxicity accompanied by activation of Hh signaling pathway. Meanwhile, genetic KO of Gli2 reduced DU-induced nephrotoxicity by normalizing biochemical indicators and reducing Hh pathway activation. Pharmacologic inhibition of Gli1/2 by Gant61 reduced DU induced cytotoxicity by inhibiting apoptosis, ROS formation and Hh pathway activation. However, overexpression of Gli2 aggravated DU-induced cytotoxicity by increasing the levels of apoptosis and ROS formation. Taken together, these results revealed that Hh signaling negatively regulated DU-inducted nephrotoxicity, and that inhibition of Gli2 might serve as a promising nephroprotective target for DU-induced kidney injury.

Role of NF-κB signaling pathway in hexavalent chromium-induced hepatotoxicity.

Hexavalent chromium Cr (VI) is a primary human carcinogen with damaging toxic effects on multiple organs. Cr (VI) exposure can induce hepatotoxicity through oxidative stress, but its exact mechanism of action was still unclear. In our study, a model of acute Cr (VI) induced liver injury was established by exposing mice to different concentrations (0, 40, 80, and 160 mg/kg) of Cr (VI); RNA-seq was used to characterize changes in liver tissue transcriptome of C57BL/6 mice after exposing to 160 mg/kg Bw of Cr (VI). Changes in liver tissue structures, proteins, and genes were observed by hematoxylin and eosin (H&E), western blot, immunohistochemistry and RT-PCR. After Cr (VI) exposure, abnormal liver tissue structure, hepatocyte injury, and hepatic inflammatory response were observed in mice in a dose-dependent manner. RNA-seq transcriptome results indicated that oxidative stress, apoptosis, and inflammatory response pathways were increased after Cr (VI) exposure; KEGG pathway analysis found that activation of NF-κB signaling pathway was significantly upregulated. Consistent with the RNA-seq results, immunohistochemistry showed that Cr (VI) exposure resulted in infiltrating of Kupffer cells and neutrophils, increasing expression of inflammatory factors (TNF-α, IL-6, IL-1ß), and activating of NF-κB signaling pathways (p-IKKα/ß and p-p65). However, ROS inhibitor, N-acetyl-L-cysteine (NAC), could reduce infiltration of Kupffer cells and neutrophils and expression of inflammatory factors. Besides, NAC could inhibit NF-κB signaling pathway activation, and alleviate Cr (VI)-induced liver tissue damage. Our findings strongly suggested that inhibition of ROS by NAC might help in the development of new strategies for Cr (VI)-associated liver fibrosis. Our findings revealed for the first time that Cr (VI) induced liver tissue damage through the inflammatory response mediated by the NF-κB signaling pathway, and inhibition of ROS by NAC might help in the development of new strategies for Cr (VI)-associated hepatotoxicity.

An efficient method of inducing differentiation of mouse embryonic stem cells into primitive endodermal cells.

Primitive Endoderm (PrE) is an extraembryonic structure derived from inner cell mass (ICM) in the blastocysts. Its interaction with the epiblast is critical to sustain embryonic growth and embryonic pattern. In this study, we reported a simple and efficient method to induce the differentiation of mouse Embryonic Stem Cells (mESCs) into PrE cells. In the process of ESC monolayer adherent culture, 1 µM atRA and 10 µM CHIR inducers were used to activate RA and Wnt signaling pathways respectively. After 9 days of differentiation, the proportion of PrE cells was up to 85%. Further studies indicated that Wnt signaling pathway acted as a switch that RA induces mESCs differentiation between SMC and PrE cell. In the presence of only RA signaling, mESCs adopted the fate of smooth muscle cells (SMCs); Simultaneous activation of the Wnt signaling pathway changed the differentiation fate of mESCs into PrE cells. This efficient induction method can provide new cellular resources and models for relevant studies of PrE.

Inhibitory Effects of Compounds from Plumula nelumbinis on Biofilm and Quorum Sensing Against P. aeruginosa.

Quorum sensing (QS), which controls the survival and virulence of Pseudomonas aeruginosa, including the formation of biofilm, is considered to be a new target to overcome pathogens. The aim of this study was to identify new QS inhibitors against P. aeruginosa and provide potential treatments for clinical infections. In this study, 25 compounds were isolated from Plumula nelumbini. Among these compounds, C25 showed the most significant biofilm inhibition activity, reaching 44.63% at 100 µM without inhibiting bacterial growth. Furthermore, C25 showed significant inhibition activity of rhamnolipid, pyocyanin, and elastase. Further mechanistic studies have confirmed that C25 could downregulate key genes in the QS system, including lasI, lasR, lasA, lasB, and pqsR, and Molecular docking studies have shown that C25 can bind to the active sites of the LasR and PqsR receptors. The present study suggests that C25 is a promising QS inhibitor for treating P. aeruginosa infections.

E-protein regulatory network links TCR signaling to effector Treg cell differentiation.

T cell antigen receptor (TCR) signaling is essential for the differentiation and maintenance of effector regulatory T (Treg) cells. However, the contribution of individual TCR-dependent genes in Treg cells to the maintenance of immunotolerance remains largely unknown. Here we demonstrate that Treg cells lacking E protein undergo further differentiation into effector cells that exhibit high expression of effector Treg signature genes, including IRF4, ICOS, CD103, KLRG-1, and RORγt. E protein-deficient Treg cells displayed increased stability and enhanced suppressive capacity. Transcriptome and ChIP-seq analyses revealed that E protein directly regulates a large proportion of the genes that are specific to effector Treg cell activation, and importantly, most of the up-regulated genes in E protein-deficient Treg cells are also TCR dependent; this indicates that E proteins comprise a critical gene regulatory network that links TCR signaling to the control of effector Treg cell differentiation and function.

Hexavalent chromium induces hepatocyte apoptosis via regulation of apoptosis signal-regulating kinase 1/c-Jun amino-terminal kinase signaling.

With the spread of hexavalent chromium (Cr(VI)) contamination, Cr(VI)-induced hepatotoxicity has attracted increasing attention in recent years. To date, however, the exact mechanism of Cr(VI) toxicity remains unclear. In this study, we investigated the role of apoptosis signal-regulating kinase 1 (ASK1)/c-Jun amino-terminal kinase (JNK) in Cr(VI)-induced hepatic toxicity and the possible related mechanisms. AML-12 hepatocyte cell-lines were treated with 0, 1, 4, and 16 µmol/Lof Cr(VI) with or without GS-444271 (an ASK1 inhibitor). Adult male mice were administered with 0, 2, 8, and 32 mg/kg body mass (BM)/day of Cr(VI) for 5 days. The level of hepatocyte apoptosis/proliferation, generation of reactive oxygen species (ROS), and expression levels of mRNAs and proteins related to ASK1/JNK and nuclear factor-E2-related factor 2 (Nrf2) signaling were assessed. Results showed that high Cr(VI) exposure induced hepatocyte apoptosis and liver injury by generation of ROS and down-regulation of Nrf2 signaling. In addition, ASK1/JNK signaling activity was upregulated in the Cr(VI)-treated group. Furthermore, GS-444217 treatment significantly rescued Cr(VI)-induced hepatocyte apoptosis and liver dysfunction in vitro and in vivo by down-regulation of ASK1/JNK signaling. Thus, ASK1/JNK signaling appears to play an important role in Cr(VI)-induced hepatocyte apoptosis and liver injury. This study should help improve our understanding of the mechanism of Cr(VI)-induced liver injury and provide support for future investigations on liver disease therapy.

RIPK3 Activates MLKL-mediated Necroptosis and Inflammasome Signaling during Streptococcus Infection.

Community-acquired pneumonia is the most common type of pneumonia and remains a leading cause of morbidity and mortality worldwide. Although many different pathogens can contribute to pneumonia, Streptococcus pneumoniae is one of the common bacterial pathogens that underlie community-acquired pneumonia. RIPK3 (receptor-interacting protein kinase 3) is widely recognized as a key modulator of inflammation and cell death. To elucidate a potential role of RIPK3 in pneumonia, we examined plasma from healthy control subjects and patients positive for streptococcal pneumonia. In human studies, RIPK3 protein concentrations were significantly elevated and were identified as a potential plasma marker of pneumococcal pneumonia. To expand these findings, we used an in vivo murine model of pneumococcal pneumonia to demonstrate that RIPK3 deficiency leads to reduced bacterial clearance, severe pathological damage, and high mortality. Our results illustrated that RIPK3 forms a complex with RIPK1, MLKL (mixed-lineage kinase domain-like protein), and MCU (mitochondrial calcium uniporter) to induce mitochondrial calcium uptake and mitochondrial reactive oxygen species(mROS) production during S. pneumoniae infection. In macrophages, RIPK3 initiated necroptosis via the mROS-mediated mitochondrial permeability transition pore opening and NLRP3 inflammasome activation via the mROS-AKT pathway to protect against S. pneumoniae. In conclusion, our study demonstrated a mechanism by which RIPK3-initiated necroptosis is essential for host defense against S. pneumoniae.

Gli2-regulated activation of hepatic stellate cells and liver fibrosis by TGF-ß signaling.

The Hedgehog (Hh) signaling pathway is correlated with hepatic stellate cells (HSCs) activation and liver fibrosis. Gli2 is a key transcription effector of Hh signaling. However, the role of Gli2 in HSC-mediated liver fibrosis progression is largely unknown. In the present study, we investigated the effect of Gli2 on liver fibrogenesis and its possible mechanism using conditional knockout (cKO) Gli2 mice and HSC models. Wild-type (WT) and GFAP-CreERT;Gli2flox/flox male mice were exposed to CCl4 for 1 mo to induce liver fibrosis. Primary HSCs were isolated from mice and the transition of HSCs into a myofibroblastic phenotype was evaluated. Livers from mice underwent histological, immunohistochemical, and immunofluorescence analyses. The expression levels of proteins and genes were evaluated by Western blot (WB) analysis and quantitative real-time polymerase chain reaction (qRT-PCR), respectively. RNA-seq was used to screen differentially expressed genes. Results showed that CCl4 treatment induced liver fibrosis, promoted HSCs activation and proliferation, and upregulated Hh signaling activity. The cKO of Gli2 in GFAP-CreERT;Gli2flox/flox mice decreased liver fibrosis as well as HSC activation and proliferation. In vitro studies showed that KO of Gli2 in HSCs blocked cell proliferation and activation by decrease of cyclin D1/D2 expression. The RNA-seq results revealed that the expression levels TGF-ß1 ligands were downregulated in Gli2 KO HSCs. Furthermore, overexpression of Gli2 rescued proliferation and activation of HSCs by upregulation of TGF-ß signaling activity. Our data demonstrated that Gli2 regulated HSC activation and liver fibrosis by TGF-ß signaling, thus providing support for future Gli2-based investigations of liver fibrosis therapy.NEW & NOTEWORTHYGli2 is a key transcription effector of Hh signaling. We found that Hh/Gli2 signaling activity was upregulated in CCl4-induced liver fibrosis. Conditional deletion of the Gli2 gene in HSCs ameliorated CCl4-induced liver fibrosis and HSCs activation. Moreover, Gli2 promoted activation of HSCs through upregulation of cyclin expression and TGF-ß signaling activity. Thus, our data provide strong support for future investigations on Gli2 inhibition to slow liver fibrosis progression in humans.

ROCK inhibitor attenuates carbon blacks-induced pulmonary fibrosis in mice via Rho/ROCK/NF-kappa B pathway.

Exposure to carbon blacks (CBs) has been associated with the progression of pulmonary fibrosis, whereas the mechanism is still not clear. We therefore aimed to investigate the effect of RhoA/ROCK pathway on pulmonary fibrosis caused by CBs exposure. Western blot analysis indicated that CBs could promote the activation of RhoA/ROCK pathway and phosphorylation of p65 and IκBα in mice lung. However, ROCK inhibitor Y-27632 could attenuate phosphorylation levels of p65 and IκBα and restore histopathological changes of the lung tissue. Then, we evaluated the effect of RhoA/ROCK pathway on pulmonary fibrosis by detecting the expression levels of α-SMA, vimentin, and Collagen type-I (Col-I), which could be partly inhibited by Y-27632. It was assumed that inhibition of ROCK could be a promising therapeutic candidate for CBs-induced pulmonary fibrosis, which possibly through the blockage of RhoA/ROCK/NF-κB pathway.

Shh-Yap signaling controls hepatic ductular reactions in CCl4 -induced liver injury.

Carbon tetrachloride (CCl4 ) exposure can induce hepatic ductular reactions. To date, however, the related mechanism remains largely unknown. Sonic hedgehog (Shh) and Yes-associated protein (Yap) signaling are correlated with liver injury and regeneration. Herein, we investigated the role of Shh and Yap signaling in the fate of ductular reaction cells in CCl4 -treated livers and the possible mechanisms. Wild-type and Shh-EGFP-Cre male mice were exposed to CCl4 (2 mL/kg), and then treated with or without the Shh signaling inhibitor Gant61. The level of liver injury, proliferation of ductular reaction cells, and expression levels of mRNA and protein related to the Shh and Yap signaling components were assessed. Results showed that CCl4 treatment induced liver injury and promoted activation and proliferation of ductular reaction cells. In addition, CCl4 induced the expression of Shh ligands in hepatocytes, accompanied by activation of Shh and Yap1 signaling in the liver. Furthermore, administration of Gant61 ameliorated liver regeneration, inhibited hepatic ductular reactions, and decreased Shh and Yap1 signaling activity. Thus, Shh-Yap1 signaling appears to play an integral role in the proliferation of ductular reaction cells in CCl4 -induced liver injury. This study should improve our understanding of the mechanism of CCl4 -induced liver injury and ductular reactions and provide support for future investigations on liver disease therapy.

GLUT1-dependent glycolysis regulates exacerbation of fibrosis via AIM2 inflammasome activation.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is a rapidly progressive, fatal lung disease that affects older adults. One of the detrimental natural histories of IPF is acute exacerbation of IPF (AE-IPF), of which bacterial infection is reported to play an important role. However, the mechanism by which bacterial infection modulates the fibrotic response remains unclear. OBJECTIVES: Altered glucose metabolism has been implicated in the pathogenesis of fibrotic lung diseases. We have previously demonstrated that glucose transporter 1 (GLUT1)-dependent glycolysis regulates fibrogenesis in a murine fibrosis model. To expand on these findings, we hypothesised that GLUT1-dependent glycolysis regulates acute exacerbation of lung fibrogenesis during bacterial infection via AIM2 inflammasome activation. RESULTS: In our current study, using a murine model of Streptococcus pneumoniae (S. pneumoniae) infection, we investigated the potential role of GLUT1 on mediating fibrotic responses to an acute exacerbation during bleomycin-induced fibrosis. The results of our current study illustrate that GLUT1 deficiency ameliorates S. pneumoniae-mediated exacerbation of lung fibrosis (wild type (WT)/phosphate buffered saline (PBS), n=3; WT/S. pneumoniae, n=3; WT/Bleomycin, n=5 ; WT/Bleomycin+S. pneumoniae, n=7; LysM-Cre-Glut1fl/f /PBS, n=3; LysM-Cre-Glut1fl/fl /S. pneumoniae, n=3; LysM-Cre-Glut1fl/fl /Bleomycin, n=6; LysM-Cre-Glut1fl/fl /Bleomycin+S. pneumoniae, n=9, p=0.041). Further, the AIM2 inflammasome, a multiprotein complex essential for sensing cytosolic bacterial DNA as a danger signal, is an important regulator of this GLUT1-mediated fibrosis and genetic deficiency of AIM2 reduced bleomycin-induced fibrosis after S. pneumoniae infection (WT/PBS, n=6; WT/Bleomycin+S. pneumoniae, n=15; Aim2-/-/PBS, n=6, Aim2-/-/Bleomycin+S. pneumoniae, n=11, p=0.034). GLUT1 deficiency reduced expression and function of the AIM2 inflammasome, and AIM2-deficient mice showed substantial reduction of lung fibrosis after S. pneumoniae infection. CONCLUSION: Our results demonstrate that GLUT1-dependent glycolysis promotes exacerbation of lung fibrogenesis during S. pneumoniae infection via AIM2 inflammasome activation.

ID2 and ID3 are indispensable for Th1 cell differentiation during influenza virus infection in mice.

Antigen-specific Th1 cells could be a passage to the infection sites during infection to execute effector functions, such as help CD8+ T cells to localize in these sites by secretion of anti-viral cytokines-IFN-γ or direct cytotoxicity of antigen-bearing cells. However, the molecular components that modulate Th1 cell differentiation and function in response to viral infection remain incompletely understood. Here, we reported that both inhibitor of DNA binding 3(Id3) protein and inhibitor of DNA binding 2(Id2) protein promoted Th1 cell differentiation. Depletion of Id3 or Id2 led to severe defect of Th1 cell differentiation during influenza virus infection. Whereas depletion of both Id3 and Id2 in CD4+ T cells restrained Th1 cell differentiation to a greater extent, indicating that Id3 and Id2 nonredundantly regulate Th1 cell differentiation. Moreover, deletion of E-proteins, the antagonists of Id proteins, greatly enhanced Th1 cell differentiation. Mechanistic study indicated that E-proteins suppressed Th1 cell differentiation by directly binding to the regulatory elements of Th1 cell master regulator T-bet and regulate T-bet expression. Thus, our findings identified Id-protein's importance for Th1 cells and clarified the nonredundant role of Id3 and Id2 in regulating Th1 cell differentiation, providing novel insight that Id3-Id2-E protein axis are essential for Th1 cell polarization.

Combination of diethyldithiocarbamate with 12-O-tetradecanoyl phorbol-13-acetate inhibits the growth of human myeloid leukemia HL-60 cells in vitro and in xenograft model.

12-O-tetradecanoylphorbol-13-acetate (TPA), is a major active constituent of the seed oil of Croton tiglium L., has pharmacological activity for the treatment of acute myeloid leukemia patients. Diethyldithiocarbamate (DTC) is a potent inhibitor of NF-κB show activity of anticancer. In this study, we determined the effect of DTC and TPA in combination on HL-60 cells cultured in vitro and in vivo. In this study, we have shown that DTC and TPA synergistically inhibited the growth of HL-60 cells and strongly induced apoptosis in the cells. Mechanistic studies showed that the combined effects of DTC and TPA were associated with a decrease in Bcl-2. The animal experiment showed that the combination of DTC and TPA more potently inhibited the growth of HL-60 tumors than either agent alone. Our results indicate that the administration of TPA and DTC in combination may be an effective strategy for inhibiting the growth of acute myeloid leukemia cells.

Antagonistic Peptides That Specifically Bind to the First and Second Extracellular Loops of CCR5 and Anti-IL-23p19 Antibody Reduce Airway Inflammation by Suppressing the IL-23/Th17 Signaling Pathway.

Asthma is a heterogeneous chronic inflammatory disorder of the airways with a complex etiology, which involves a variety of cells and cellular components. Therefore, the aim of the study was to investigate the effects and mechanisms of antagonistic peptides that specifically bind to the first and second extracellular loops of CCR5 (GH and HY peptides, respectively) and anti-interleukin-23 subunit p19 (anti-IL-23p19) in the airway and thereby mediate inflammation and the IL-23/T helper 17 (Th17) cell pathway in asthmatic mice. An experimental asthma model using BALB/c mice was induced by ovalbumin (OVA) and treated with peptides that are antagonistic to CCR5 or with anti-IL-23p19. The extents of the asthmatic inflammation and mucus production were assessed. In addition, bronchoalveolar lavage fluid (BALF) was collected, the cells were counted, and the IL-4 level was detected by ELISA. The IL-23/Th17 pathway-related protein and mRNA levels in the lung tissues were measured, and the positive production rates of Th17 cells in the thymus, spleen, and peripheral blood were detected. The groups treated with one of the two peptides and/or anti-IL-23p19 showed significant reductions in allergic inflammation and mucus secretion; decreased expression levels of IL-23p19, IL-23R, IL-17A and lactoferrin (LTF); and reduced proportions of Th17 cells in the thymus, spleen, and peripheral blood. Specifically, among the four treatment groups, the anti-IL-23p19 with HY peptide group exhibited the lowest positive production rate of Th17 cells. Our data also showed a significant and positive correlation between CCR5 and IL-23p19 protein expression. These findings suggest that the administration of peptides antagonistic to CCR5 and/or anti-IL-23p19 can reduce airway inflammation in asthmatic mice, most likely through inhibition of the IL-23/Th17 signaling pathway, and the HY peptide can alleviate inflammation not only through the IL-23/Th17 pathway but also through other mechanisms that result in the regulation of inflammation.

Pleural IFN-γ release assay combined with biomarkers distinguished effectively tuberculosis from malignant pleural effusion.

BACKGROUND: Tuberculosis (TB) remains a major public health concern on a global scale, especially in developing nations. So far, no formal guidelines are available for the diagnosis and treatment of tuberculosis pleurisy. The diagnosis of TB is worsened by the immense difficulty in differential determination of tuberculosis pleural effusion (TPE) and malignant pleural effusion (MPE). The purpose of this investigation is to assess the differential diagnostic efficiencies of the pleural IFN-γ release assay (IGRA) and widely-used biochemical parameters in the distinction analysis of TPE and MPE. METHODS: A cohort of 222 patients with pleural effusion was examined, comprising of 143 TPE and 58 MPE patients. The patients were examined with IGRA, and the widely-used biomarkers in the pleural effusion and peripheral blood. RESULTS: Our results show that the TPE patients have significantly higher M. tuberculosis (Mtb) antigen-specific IFN-γ responses to ESAT-6 protein and peptide pool in the blood compared to MPE patients. TPE patients were also shown to have enriched Mtb antigen-specific IFN-γ responses in pleural effusion than in peripheral blood. Among the widely-used biomarkers, the adenosine deaminase (ADA) and carcinoembryonic antigen (CEA) in pleural effusion were better biomarkers with high sensitivity and specificity to discriminate TPE and MPE. In addition, pleural IGRA could not be affected by the pleural adhesion, and the applications of the pleural IGRA together with ADA and CEA provide a promising approach for the TPE and MPE differential identification. CONCLUSIONS: Our study proposes that the integration of pleural IGRA and ADA, CEA detection could add to more effective diagnosis stratagems in the discernment between TPE and MPE.

The Inflammatory Bowel Disease-Associated Autophagy Gene Atg16L1T300A Acts as a Dominant Negative Variant in Mice.

The basis of the increased risk for Crohn's disease conferred by the Atg16L1T300A polymorphism is incompletely understood. An important step forward came from the recent demonstration that the murine equivalent of Atg16L1T300A (Atg16L1T316A) exhibits increased susceptibility to caspase 3-mediated cleavage and resulting decreased levels of full-length Atg16L1 in macrophages. However, although this finding showed that this polymorphism is a loss-of-function abnormality, it did not address the possibility that this polymorphism also affects the function of a normal Atg16L1 allele in heterozygous mice. Therefore, we evaluated the function of the Atg16L1T300A polymorphism heterozygote and homozygote in knock-in (KI) mice. Surprisingly, we found that macrophages from both types of KI mice exhibit defective autophagic induction; accordingly, both types of mice exhibit defects in bacterial clearance coupled with increased inflammasome cytokine (IL-1ß) responses. Furthermore, macrophages from both types of KI mice displayed defects in TNF-α-induced Atg16L1T300A cleavage, increased retention of bacteria, bacterial dissemination, and Salmonella-induced colitis. These studies suggested that chromosomes bearing the Atg16L1T300A polymorphism can interfere with the function of the wild-type (WT) Atg16L1 allele and, thus, that the Crohn's disease risk polymorphism is a dominant-negative variant with the potential to act as a disease factor, even when present on only one chromosome. This conclusion was supported by the finding that mice bearing a WT Atg16L1 allele and a null allele (Atg16L1KO/+ mice) exhibit normal autophagic function equivalent to that of WT mice.

CD8+ T Cell Immune Response in Immunocompetent Mice during Zika Virus Infection.

Zika virus (ZIKV) infection causees neurologic complications, including Guillain-Barré syndrome in adults and central nervous system (CNS) abnormalities in fetuses. We investigated the immune response, especially the CD8+ T cell response in C57BL/6 (B6) wild-type (WT) mice, during ZIKV infection. We found that a robust CD8+ T cell response was elicited, major histocompatibility complex class I-restricted CD8+ T cell epitopes were identified, a tetramer that recognizes ZIKV-specific CD8+ T cells was developed, and virus-specific memory CD8+ T cells were generated in these mice. The CD8+ T cells from these infected mice were functional, as evidenced by the fact that the adoptive transfer of ZIKV-specific CD8+ T cells could prevent ZIKV infection in the CNS and was cross protective against dengue virus infection. Our findings provide comprehensive insight into immune responses against ZIKV and further demonstrate that WT mice could be a natural and easy-access model for evaluating immune responses to ZIKV infection.IMPORTANCE ZIKV infection has severe clinical consequences, including Guillain-Barré syndrome in adults, microcephaly, and congenital malformations in fetuses and newborn infants. Therefore, study of the immune response, especially the adaptive immune response to ZIKV infection, is important for understanding diseases caused by ZIKV infection. Here, we characterized the CD8+ T cell immune response to ZIKV in a comprehensive manner and identified ZIKV epitopes. Using the identified immunodominant epitopes, we developed a tetramer that recognizes ZIKV-specific CD8+ T cells in vivo, which simplified the detection and evaluation of ZIKV-specific immune responses. In addition, the finding that tetramer-positive memory CD8+ T cell responses were generated and that CD8+ T cells can traffic to a ZIKV-infected brain greatly enhances our understanding of ZIKV infection and provides important insights for ZIKV vaccine design.