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The pharmaceutical industry has a pervasive need for chiral specific molecules with optimal affinity for their biological targets. However, the mass production of such compounds is currently limited by conventional chemical routes, that are costly and have an environmental impact. Here, we propose an easy access to obtain new tetrahydroquinolines, a motif found in many bioactive compounds, that is rapid and cost effective. Starting from simple raw materials, the procedure uses a proline‐catalyzed Mannich reaction followed by the addition of BF3 ⋅ OEt2, which generates a highly electrophilic aza‐ortho‐quinone methide intermediate capable of reacting with different nucleophiles to form the diversely functionalized tetrahydroquinoline. Moreover, this enantioselective one‐pot process provides access for the first time to tetrahydroquinolines with a cis‐2,3 and trans‐3,4 configuration. As proof of concept, we demonstrate that a three‐step reaction sequence, from simple and inexpensive starting compounds and catalysts, can generate a BD2‐selective BET bromodomain inhibitor with anti‐inflammatory effect. [ABSTRACT FROM AUTHOR]

The localized functionalization of pores and channels of micrometric and sub‐micrometric sizes is a bottleneck in surface chemistry. A method for the regioselective chemical functionalization of planar pores is presented, that are, restrictions in microfluidic channels, here made of SiO2‐coated silicon. This strategy, based on bipolar electrochemistry, exploits the combined presence of the constriction and a localized deoxidation pattern within the pore that affects the electrical field distribution inside the microfluidic channel. It is not only shown that it is capable of regioselectively functionalizing a planar pore at relatively small potential difference applied across it, but also the possibility of positioning the functionalization area inside or at the edges of the pore depending on the design of the deoxidation pattern is proved. These results are in perfect correlation with the numerical simulations of electric field distribution in micropores carried out using the software Comsol Multiphysics. This functionalization technique is therefore very promising, particularly in the field of biosensors. A specific DNA hybridization test has been successfully carried out, which represents a first step toward bioanalytical and health applications. [ABSTRACT FROM AUTHOR]

Background & Aims The pro-inflammatory cytokine IL-17 plays a crucial role in liver diseases associated with hepatic fibrosis and increased risk of cancer development. Nevertheless, the cellular source of this cytokine has never been characterized in patients with liver fibrosis. Methods In this study, we investigated liver biopsies from 49 patients with chronic viral hepatitis at different stages of liver fibrosis. We monitored IL-17 production by intracellular flow cytometry, immunofluorescence and immunohistochemical in situ stainings, allowing a precise quantification, characterization and localization of IL-17+ cells. Results Density of IL-17+ cells increased with the stage of liver fibrosis specifically in fibrotic septa and portal areas (correlation coefficient r = 0.7373; P < 0.0001). Data clearly show that the frequency of intrahepatic IL-17+ lymphocytes (including T, NKT and NK cells) was independent on stage of liver fibrosis, and we observed no statistical differences in number of IL-17+ macrophages during progression of fibrosis. On the other hand, the number of IL-17+ neutrophils in fibrotic septa and portal areas strongly correlated with the stages of fibrosis (correlation coefficient r = 0.6986; P < 0.0001), contributing significantly to total IL-17 production in liver tissue. Conclusions Our data indicate that neutrophils represent an important source of IL-17 in the human liver, especially in late fibrosis stages. Inhibition of this specific harmful subset of neutrophils may offer therapeutic opportunities in fibrotic liver. [ABSTRACT FROM AUTHOR]

Background Chronic liver inflammation and immune/inflammatory response promote hepatocellular carcinoma. The aim of this study was to characterize the immune status of HCV-related cirrhosis in patients with hepatocellular carcinoma ( HCV- HCC) as compared to HCV patients without hepatocellular carcinoma. Method Immune markers ( CD3, CD4, CD8, CD20, CD56, TCRγδ, FoxP3) and gene expression profiles ( CD8α, CD8β, FoxP3, IL-6, IFN-γ, perforin, RANTES) were analysed in a test cohort by immunohistochemistry and quantitative RT- PCR analysis on serial non-tumorous and tumorous tissues. Results Immune micro-environment was more inflammatory in HCV- HCC than HCV cirrhotic livers. The number of CD3+, CD4+, CD8+ and CD20+ liver-infiltrating lymphocytes was significantly higher, whereas the number of CD56+ cells was significantly lower in HCV- HCC compared to HCV cirrhotic parenchyma. These differences were restricted to fibrous septa for CD4+ and CD20+ cells and to nodular parenchyma for CD8+ cells. Gene expressions of CD8α, FoxP3 and RANTES were also significantly higher in HCV- HCC than in HCV cirrhosis. Interestingly, in a large cohort of 63 HCV- HCC patients. The number of CD8+ cells ≥100/field was associated with significant higher tumour recurrence ( P = 0.003) and lower overall survival ( P = 0.05) at 5 years. Conclusion High densities of liver-infiltrating lymphocytes in HCV- HCC cirrhotic parenchyma prevail inflammatory conditions and could contribute to tumorigenesis and tumour recurrence. These results could contribute towards better clinical evaluation of patients susceptible for HCC recurrence after curative surgery. [ABSTRACT FROM AUTHOR]

Immunotherapy for cancer aims to generate cytotoxic cells that are capable of eradicating tumour cells. It has been well demonstrated that helper, non-cytotoxic CD4+ T cells are important for the induction and maintenance of anti-tumour immunity exerted by cytotoxic CD8+ T cells. In contrast, the existence of direct anti-tumour, effector cytotoxic CD4+ T cells remains elusive, mainly due to the paucity of reliable experimental data, especially in human B-cell non-Hodgkin lymphomas. This study developed an appropriate, autologous follicular B-cell non-Hodgkin follicular lymphoma model, including the in vitro establishment of a malignant, human leucocyte antigen class I (HLA-I) deficient B-cell line, and the generation of three autologous anti-tumour cytotoxic CD4+ T-cell clones originating from the peripheral blood of the same patient. These three clones were considered as tumour specific, because they were capable of killing the malignant, HLA-I-deficient B-cell line through a classical HLA-II restricted perforin-mediated pathway, but did not lyse the Epstein–Barr virus-infected autologous normal B lymphocytes. All three CD4+ clones were T-cell receptor V β17-Dβ1-Jβ1.2 and exhibited an identical complementarity-determining region 3, suggesting the immunodominance of a single peptide antigen presented by tumour cells. Such lymphoma models would provide a useful tool for in vivo expansion and the adoptive transfer of selected CD4+ cytotoxic cells in immunotherapeutic strategies. [ABSTRACT FROM AUTHOR]

We have used human astrocytoma-derived cells to investigate the cellular responses of central nervous system cells toToxoplasma gondiiinfection. At 24 h post inoculation, the secretion of CCL-2 (or Monocyte Chemotactic Protein-1) was augmented six-fold over the control. This secretion was down-regulated by D609, a specific inhibitor of phosphatidylcholine-dependent phospholipase C (PC-PLC), but not modulated by gamma interferon (IFN-γ). Ribonuclease protection assay analyses showed significant down-regulation of CCL-2 mRNA production during infection byToxoplasma gondiiwhen cells were treated by D609. The mRNA levels of the seven other chemokines studied were not modified by D609. CCL-2 seems to contribute to the cell recruitment during human cerebral reactivation ofToxoplasma gondii. Cellular production of this CC chemokine during toxoplasmosis may be regulated by a PC-PLC-dependent pathway. [ABSTRACT FROM AUTHOR]

The receptor for the iC3b fragment of complement, CR3, is involved in monocytes/macrophages and neutrophils phagocytosis. CR3 is known to interact with the low affinity receptor for Ig (CD16) and previous studies have suggested that this cooperation modulates CR3 functions. Herein we have studied the effect of CD16 on the ability of human monocytes CR3 to bind to iC3b. We show that iC3b binding to CR3 is inhibited by several reagents that are known to dissociate the CD16/CR3 complex. In addition, treatment of monocytes with soluble CD16 inhibited iC3b binding to CR3. Together, these data indicate that iC3b binding to monocyte CR3 is up-regulated by an interaction between membrane CD16 and CR3. The implication of CD16 in CR3 binding to iC3b was also analyzed after monocyte differentiation into dendritic cells (DC). Differentiation of monocytes into DC abrogates the cooperation between CD16 and CR3, due to a loss of CD16/CR3 interaction. In accordance, this phenomenon is associated with a lack of iC3b binding to DC. As a consequence, deposition of iC3b on apoptotic cells does not modify their phagocytosis by DC. In conclusion, we demonstrate a cooperation between CD16 and CR3 that favors iC3b binding to CR3 but is lost on DC. [ABSTRACT FROM AUTHOR]

Summary The in vitro regulation of tumour necrosis factor (TNF)-α receptors during Toxoplasma gondii infection of human MRC5 fibroblasts and human myelomonocytic THP-1 cells was investigated. Cells were infected with the virulent RH of T. gondii. TNFR membrane receptors were analysed by flow cytometry with biotinylated TNF-α. Shedding of the soluble form of TNFR1 and TNFR2 in cell culture supernatants was measured by enzyme-linked immunosorbent assay, and expression of mRNA production of TNFR1 and TNFR2 was analysed by quantitative real-time ploymerase chain reaction, 1 h after infection. In the MRC5 cell line, T. gondii infection did not induce any up- or down-regulation of membrane TNFRs, soluble TNFRs or mRNA of TNFRs. However, THP-1 cell infection with living parasites induced a significant soluble TNFR1 release by THP-1 cells after 1 h. We detected an approximately 50% up-regulation (P < 0·01) of soluble TNFR1 in infected THP-1 cells compared to controls. No change in soluble TNFR2 levels was observed in the same conditions. Moreover, infection decreased the level of TNF membrane receptors, but had no effect on TNFR1 and TNFR2 mRNA levels. TNFR modulation by T. gondii infection, in vitro, depends on the cell type. Furthermore, our data suggest that living parasites control the shedding of the soluble form of TNFR1. This mechanism may influence the role of TNF-α in toxoplasmosis. [ABSTRACT FROM AUTHOR]

Background & Aims: The pro-inflammatory cytokine IL-17 plays a crucial role in liver diseases associated with hepatic fibrosis and increased risk of cancer development. Nevertheless, the cellular source of this cytokine has never been characterized in patients with liver fibrosis., Methods: In this study, we investigated liver biopsies from 49 patients with chronic viral hepatitis at different stages of liver fibrosis. We monitored IL-17 production by intracellular flow cytometry, immunofluorescence and immunohistochemical in situ stainings, allowing a precise quantification, characterization and localization of IL-17(+) cells., Results: Density of IL-17(+) cells increased with the stage of liver fibrosis specifically in fibrotic septa and portal areas (correlation coefficient r = 0.7373; P < 0.0001). Data clearly show that the frequency of intrahepatic IL-17(+) lymphocytes (including T, NKT and NK cells) was independent on stage of liver fibrosis, and we observed no statistical differences in number of IL-17(+) macrophages during progression of fibrosis. On the other hand, the number of IL-17(+) neutrophils in fibrotic septa and portal areas strongly correlated with the stages of fibrosis (correlation coefficient r = 0.6986; P < 0.0001), contributing significantly to total IL-17 production in liver tissue., Conclusions: Our data indicate that neutrophils represent an important source of IL-17 in the human liver, especially in late fibrosis stages. Inhibition of this specific harmful subset of neutrophils may offer therapeutic opportunities in fibrotic liver., (© 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: Chronic liver inflammation and immune/inflammatory response promote hepatocellular carcinoma. The aim of this study was to characterize the immune status of HCV-related cirrhosis in patients with hepatocellular carcinoma (HCV-HCC) as compared to HCV patients without hepatocellular carcinoma., Method: Immune markers (CD3, CD4, CD8, CD20, CD56, TCRγδ, FoxP3) and gene expression profiles (CD8α, CD8β, FoxP3, IL-6, IFN-γ, perforin, RANTES) were analysed in a test cohort by immunohistochemistry and quantitative RT-PCR analysis on serial non-tumorous and tumorous tissues., Results: Immune micro-environment was more inflammatory in HCV-HCC than HCV cirrhotic livers. The number of CD3(+) , CD4(+) , CD8(+) and CD20(+) liver-infiltrating lymphocytes was significantly higher, whereas the number of CD56(+) cells was significantly lower in HCV-HCC compared to HCV cirrhotic parenchyma. These differences were restricted to fibrous septa for CD4(+) and CD20(+) cells and to nodular parenchyma for CD8(+) cells. Gene expressions of CD8α, FoxP3 and RANTES were also significantly higher in HCV-HCC than in HCV cirrhosis. Interestingly, in a large cohort of 63 HCV-HCC patients. The number of CD8(+) cells ≥100/field was associated with significant higher tumour recurrence (P = 0.003) and lower overall survival (P = 0.05) at 5 years., Conclusion: High densities of liver-infiltrating lymphocytes in HCV-HCC cirrhotic parenchyma prevail inflammatory conditions and could contribute to tumorigenesis and tumour recurrence. These results could contribute towards better clinical evaluation of patients susceptible for HCC recurrence after curative surgery., (© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Immunotherapy for cancer aims to generate cytotoxic cells that are capable of eradicating tumour cells. It has been well demonstrated that helper, non-cytotoxic CD4(+) T cells are important for the induction and maintenance of anti-tumour immunity exerted by cytotoxic CD8(+) T cells. In contrast, the existence of direct anti-tumour, effector cytotoxic CD4(+) T cells remains elusive, mainly due to the paucity of reliable experimental data, especially in human B-cell non-Hodgkin lymphomas. This study developed an appropriate, autologous follicular B-cell non-Hodgkin follicular lymphoma model, including the in vitro establishment of a malignant, human leucocyte antigen class I (HLA-I) deficient B-cell line, and the generation of three autologous anti-tumour cytotoxic CD4(+) T-cell clones originating from the peripheral blood of the same patient. These three clones were considered as tumour specific, because they were capable of killing the malignant, HLA-I-deficient B-cell line through a classical HLA-II restricted perforin-mediated pathway, but did not lyse the Epstein-Barr virus-infected autologous normal B lymphocytes. All three CD4(+)clones were T-cell receptor Vbeta17-Dbeta1-Jbeta1.2 and exhibited an identical complementarity-determining region 3, suggesting the immunodominance of a single peptide antigen presented by tumour cells. Such lymphoma models would provide a useful tool for in vivo expansion and the adoptive transfer of selected CD4(+) cytotoxic cells in immunotherapeutic strategies.