miR-199a-5p Is Upregulated during Fibrogenic Response to Tissue Injury and Mediates TGFbeta-Induced Lung Fibroblast Activation by Targeting Caveolin-1

As miRNAs are associated with normal cellular processes, deregulation of miRNAs is thought to play a causative role in many complex diseases. Nevertheless, the precise contribution of miRNAs in fibrotic lung diseases, especially the idiopathic form (IPF), remains poorly understood. Given the poor response rate of IPF patients to current therapy, new insights into the pathogenic mechanisms controlling lung fibroblasts activation, the key cell type driving the fibrogenic process, are essential to develop new therapeutic strategies for this devastating disease. To identify miRNAs with potential roles in lung fibrogenesis, we performed a genome-wide assessment of miRNA expression in lungs from two different mouse strains known for their distinct susceptibility to develop lung fibrosis after bleomycin exposure. This led to the identification of miR-199a-5p as the best miRNA candidate associated with bleomycin response. Importantly, miR-199a-5p pulmonary expression was also significantly increased in IPF patients (94 IPF versus 83 controls). In particular, levels of miR-199a-5p were selectively increased in myofibroblasts from injured mouse lungs and fibroblastic foci, a histologic feature associated with IPF. Therefore, miR-199a-5p profibrotic effects were further investigated in cultured lung fibroblasts: miR-199a-5p expression was induced upon TGFβ exposure, and ectopic expression of miR-199a-5p was sufficient to promote the pathogenic activation of pulmonary fibroblasts including proliferation, migration, invasion, and differentiation into myofibroblasts. In addition, we demonstrated that miR-199a-5p is a key effector of TGFβ signaling in lung fibroblasts by regulating CAV1, a critical mediator of pulmonary fibrosis. Remarkably, aberrant expression of miR-199a-5p was also found in unilateral ureteral obstruction mouse model of kidney fibrosis, as well as in both bile duct ligation and CCl4-induced mouse models of liver fibrosis, suggesting that dysregulation of miR-199a-5p represents a general mechanism contributing to the fibrotic process. MiR-199a-5p thus behaves as a major regulator of tissue fibrosis with therapeutic potency to treat fibroproliferative diseases. © 2013 Lino Cardenas et al

Live cell tracking of macrophage efferocytosis during Drosophila embryo development in vivo

Apoptosis of cells and their subsequent removal via efferocytosis occurs in nearly all tissues during development, homeostasis, and disease. However, it has been difficult to track cell death and subsequent corpse removal in vivo. Here, we developed a genetically encoded fluorescent reporter, CharON, that could track emerging apoptotic cells and their efferocytic clearance by phagocytes. Using Drosophila expressing CharON, we uncovered multiple qualitative and quantitative features of coordinated clearance of apoptotic corpses during embryonic development. To confront high rate of emerging apoptotic corpses, the macrophages displayed heterogeneity in engulfment, with some efferocytic macrophages carrying high corpse burden. However, overburdened macrophages were compromised in clearing wound debris, revealing an inherent vulnerability. These findings reveal known and unexpected features of apoptosis and macrophage efferocytosis in vivo

Repression of PLA2R1 by c-MYC and HIF-2alpha promotes cancer growth

Loss of secreted phospholipase A2 receptor (PLA2R1) has recently been found to render human primary cells more resistant to senescence whereas increased PLA2R1 expression is able to induce cell cycle arrest, cancer cell death or blockage of cancer cell transformation in vitro, suggesting that PLA2R1 displays tumor suppressive activities. Here we report that PLA2R1 expression strongly decreases in samples of human renal cell carcinoma (RCC). Knockdown of PLA2R1 increases renal cancer cell tumorigenicity supporting a role of PLA2R1 loss to promote in vivo RCC growth. Most RCC result from Von Hippel-Lindau (VHL) tumor suppressor loss-of-function and subsequent gain-of-function of the oncogenic HIF-2alpha/c-MYC pathway. Here, by genetically manipulating VHL, HIF-2alpha and c-MYC, we demonstrate that loss of VHL, stabilization of HIF-2alpha and subsequent increased c-MYC activity, binding and transcriptional repression, through induction of PLA2R1 DNA methylation closed to PLA2R1 transcriptional start site, results in decreased PLA2R1 transcription. Our results describe for the first time an oncogenic pathway leading to PLA2R1 transcriptional repression and the importance of this repression for tumor growth

Regulators of yeast endocytosis identified by systematic quantitative analysis

Endocytosis of receptors at the plasma membrane is controlled by a complex mechanism that includes clathrin, adaptors, and actin regulators. Many of these proteins are conserved in yeast yet lack observable mutant phenotypes, which suggests that yeast endocytosis may be subject to different regulatory mechanisms. Here, we have systematically defined genes required for internalization using a quantitative genome-wide screen that monitors localization of the yeast vesicle-associated membrane protein (VAMP)/synaptobrevin homologue Snc1. Genetic interaction mapping was used to place these genes into functional modules containing known and novel endocytic regulators, and cargo selectivity was evaluated by an array-based comparative analysis. We demonstrate that clathrin and the yeast AP180 clathrin adaptor proteins have a cargo-specific role in Snc1 internalization. We additionally identify low dye binding 17 (LDB17) as a novel conserved component of the endocytic machinery. Ldb17 is recruited to cortical actin patches before actin polymerization and regulates normal coat dynamics and actin assembly. Our findings highlight the conserved machinery and reveal novel mechanisms that underlie endocytic internalization

Istradefylline protects from cisplatin-induced nephrotoxicity and peripheral neuropathy while preserving cisplatin antitumor effects

Cisplatin is a potent chemotherapeutic drug that is widely used in the treatment of various solid cancers. However, its clinical effectiveness is strongly limited by frequent severe adverse effects, in particular nephrotoxicity and chemotherapy-induced peripheral neuropathy. Thus, there is an urgent medical need to identify novel strategies that limit cisplatin-induced toxicity. In the present study, we show that the FDA-approved adenosine A2A receptor antagonist istradefylline (KW6002) protected from cisplatin-induced nephrotoxicity and neuropathic pain in mice with or without tumors. Moreover, we also demonstrate that the antitumoral properties of cisplatin were not altered by istradefylline in tumor-bearing mice and could even be potentiated. Altogether, our results support the use of istradefylline as a valuable preventive approach for the clinical management of patients undergoing cisplatin treatment

Aspects fondamentaux du développement tumoral

International audienceLa tumorigenèse a traditionnellement été considérée comme un processus en plusieurs étapes impliquant l’activation des oncogènes ainsi que l’inactivation des gènes suppresseurs de tumeurs. Cependant, les mécanismes à l’origine de l’initiation et de la progression du cancer ne se limitent pas aux altérations moléculaires et doivent plutôt être considérés comme un processus complexe qui interagit avec l’organisme entier. Cette revue didactique fournit une vision intégrée et globale des principes fondamentaux clés du développement du cancer

Helicobacter pylori urease and flagellin alter mucin gene expression in human gastric cancer cells

International audienceBackground Helicobacter pylori (Hp), which is one of the causative agents in human gastric adenocarcinoma, is known to interact with mucous gel and alter mucin gene expression. The aim of this work was to study, using an in vitro model of cell infection, the effects of urease, flagellin, and CagA vir-ulence factors on the regulation of the four 11p15 mucin genes (MUC2, MUC5AC, MUC5B, and MUC6). Methods KATO-III and AGS gastric cancer cells were infected for 1, 3 or 6 h with Hp wild-type strains (ATCC 43504, N6, and SS1) or corresponding isogenic mutants deficient for urease subunit B, flagellin subunit A, and CagA. mRNA levels of MUC2, MUC5B, MUC5AC and MUC6 were assessed by RT-PCR, and functional activity of their promoters was measured by transient transfection assays. Results Infection of KATO-III cells with Hp wild-type strains resulted in an early (at 1 h) transient expression of MUC2, MUC5AC, and MUC6 mRNA concomitant with those of interleukin (IL)-1b, IL-8, and TNF-a cytokines. In these cells, the UreB-isogenic mutant induced strong activation of MUC5AC expression, and UreB-responsive elements were located in the-486/-1 region of the promoter. FlaA-and CagA-mutants had no effect on mucin gene mRNA levels in KATO-III cells. In AGS cells, Hp-responsive elements were identified in all promoters, and overexpression of NF-jB induced upregulation of MUC5AC promoter activity when infected with the UreB-isogenic mutant. Conclusion These results indicate that Hp infection of gastric cancer cells alters 11p15 mucin gene transcription and that MUC5AC downregulation is mediated by urease virulence factor

Transcriptional regulation of human mucin MUC4 by bile acids in oesophageal cancer cells is promoter-dependent and involves activation of the phosphatidylinositol 3-kinase signalling pathway.

Abnormal gastro-oesophageal reflux and bile acids have been linked to the presence of Barrett's oesophageal premalignant lesion associated with an increase in mucin-producing goblet cells and MUC4 mucin gene overexpression. However, the molecular mechanisms underlying the regulation of MUC4 by bile acids are unknown. Since total bile is a complex mixture, we undertook to identify which bile acids are responsible for MUC4 up-regulation by using a wide panel of bile acids and their conjugates. MUC4 apomucin expression was studied by immunohistochemistry both in patient biopsies and OE33 oesophageal cancer cell line. MUC4 mRNA levels and promoter regulation were studied by reverse transcriptase-PCR and transient transfection assays respectively. We show that among the bile acids tested, taurocholic, taurodeoxycholic, taurochenodeoxycholic and glycocholic acids and sodium glycocholate are strong activators of MUC4 expression and that this regulation occurs at the transcriptional level. By using specific pharmacological inhibitors of mitogen-activated protein kinase, phosphatidylinositol 3-kinase, protein kinase A and protein kinase C, we demonstrate that bile acid-mediated up-regulation of MUC4 is promoter-specific and mainly involves activation of phosphatidylinositol 3-kinase. This new mechanism of regulation of MUC4 mucin gene points out an important role for bile acids as key molecules in targeting MUC4 overexpression in early stages of oesophageal carcinogenesis