Transforming growth factor (TGF)-␤1 induces extracellular matrix deposition and proliferation of mesenchymal cells. We recently reported that interleukin (IL)-6 is an essential mediator of growth factor-induced proliferation of lung fibroblasts. Here, we demonstrate by reverse transcriptase polymerase chain reaction and enzyme-linked immunoassay that TGF-␤1 is a potent inducer of IL-6 mRNA and protein in primary human lung fibroblasts. Transient transfections of fibroblasts with a luciferase reporter gene construct containing nucleotides ؊651 to ؉1 of the human IL-6 promoter revealed that TGF-␤1 also potently activated IL-6 promoter activity. Progressive 5-deletions and sitedirected mutagenesis of the parental construct located the TGF-␤1-responsive cis-regulatory element to a known activating protein-1 (AP-1) sequence (nucleotides ؊284 to ؊276). Gel shift analyses revealed that AP-1 DNA binding activity in nuclear extracts was increased 30 min after stimulation with TGF-␤1. In contrast, neither CCAAT enhancer-binding protein-␤, NF-B, nor Sp1 were activated by TGF-␤1. Supershift analyses demonstrated that the AP-1 complex induced by TGF-␤1 was composed of Jun isoforms and absent of Fos isoforms. Moreover, this complex was found to be a JunD homodimer. Our data thus demonstrate that TGF-␤1 is a potent inducer of IL-6 in primary human lung fibroblasts. The TGF-␤1-activated JunD homodimer may be essential for a majority of the biological effects induced by TGF-␤1 in this cell type, such as proliferation and extracellular matrix synthesis.

Adult stem cells, including adipose tissue-derived mesenchymal stem cells (MSCs) or ectomesenchymal dental follicle cells (DFCs), attract considerable attention for their potential to differentiate into lineages, which are of major interest in the field of Regenerative Medicine. Purinergic receptors exert a wide range of biological actions in many cell and tissue types through extracellular nucleotides. Little is known about P2 receptors in adult stem cells and changes in their expression levels during differentiation. All known P2 receptors have been investigated, and a variety of P2X and P2Y receptor subtypes were detected in MSCs. Studies investigating intracellular calcium levels on receptor stimulation demonstrated that the found P2 receptors are metabolically active. Interestingly, up- or downregulation of several P2 receptor subtypes at gene and protein level was observed during adipogenic and osteogenic differentiation, and the effect on differentiation was directly influenced by both the application of agonists/antagonists and apyrase-induced nucleotide cleavage. Here, we show for the first time that the combination of several P2 receptors plays a role in the differentiation of adult stem cells. The expression pattern of the P2 receptors, as well as their fate in differentiation, varies in stem cells of mesenchymal origin if compared with stem cells of ectomesenchymal origin. The subtypes P2X6, P2Y4, and P2Y14 seem to be pivotal regulators in MSC commitment, as they are regulated in both adipogenic and osteogenic differentiation of adipose tissue-derived stem cells and DFCs. These findings provide new insights into the differentiation processes and might reveal novel options to influence stem cell fate in future applications.

Lung fibrosis is a fatal condition of excess extracellular matrix (ECM) deposition associated with increased transforming growth factor beta (TGF-beta) activity. Although much is known about its pathological features, our understanding of the signal transduction pathways resulting in increased ECM and collagen deposition in response to TGF-beta is still incompletely defined. We have previously reported that a JunD homodimer of the transcription factor AP-1 is specifically activated by TGF-beta in lung fibroblasts. Here we demonstrate that JunD is also specifically required for TGF-beta-induced effects. Antisense against JunD, but not c-fos or c-jun, significantly inhibited collagen deposition in response to TGF-beta in primary human lung fibroblasts. We then investigated the ability of pharmacological agents to inhibit TGF-beta-induced signaling and collagen deposition. Cs-A and IFN-gamma, but not glucocorticoids, cyclophosphamide, or azathioprine, inhibited TGF-beta-induced signaling, as assessed by luciferase reporter gene assays, and collagen deposition. TGF-beta antagonism by Cs-A was associated with direct inhibition of JunD activation, as demonstrated by electrophoretic mobility shift analyses. In contrast, the effects of IFN-gamma required signal transducer and activator of transcription (STAT)-1. We thus identify the JunD isoform of AP-1 as an essential mediator of TGF-beta-induced effects in lung fibroblasts. TGF-beta-induced signaling and collagen deposition are efficiently antagonized by Cs-A and IFN-gamma treatment, both of which exhibit distinct molecular mechanisms of action. These observations therefore offer novel targets for future therapy of fibrotic lung disease.

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In most cells studied so far, interferon-A (IFN-A) activates signal transducer and activator of transcription (Stat1), Stat2 and Stat3, whereas interferon-γ (IFN-γ) induces Stat1 only. In general, each of the several dozens of cytokines, growth factors and hormones that signal through the Janus kinases-signal transducers and activators of transcription (Jak-STAT) pathway activates a distinct subset of STATs, and this selectivity is thought to be essential for the specificity of the cellular responses toward these ligands. Here, we have studied the pattern of STAT activation in the human lymphoblastoid cell line Daudi in response to IFN-A. In addition to Stat1, Stat2 and Stat3 activation, IFN-A was found to directly induce activation of Stat5 and Stat6. Cell-type-specific activation of additional STATs could be responsible for cell-type-specific responses to IFN-A.Keywords : interferon; signal transducers and activators of transcription ; cytokine receptor; signal transduction; transcription factor. Cytokines, interleukins, polypeptide growth factors and hor-The activated Jaks phosphorylate one or more tyrosine residues on receptor chains. The phosphorylated tyrosine residues, tomones bind to receptors on the surface of target cells and elicit diverse and specific responses. The signals received at the recep-gether with the surrounding amino acids, provide the docking sites for the STATs which bind these residues by means of their tors are transmitted by intracellular signal-transduction pathways and result in transcriptional activation of target genes. To evoke SH2 domains. This interaction is specific and provides a bases for differential STAT activation by different polypeptide ligands diverse cellular responses, different ligands have to activate dif- [15,16]. Once recruited to the receptors, STATs become phosferent subsets of genes. This can be achieved through the spephorylated on a single tyrosine residue in the C-terminus and cific activation of particular transcription factors. The signal form homodimers and heterodimers [17]. The specificity of inditransducers and activators of transcription (STATs) are a family vidual STAT SH2 domains for phosphotyrosine motives of the of transcription factors that are activated by many diverse polypartner STAT again determines which homodimer and heteropeptide ligands [1]. For these ligands, STATs serve an important dimers are formed [16]. STAT dimers translocate into the nufunction by contributing to specific gene activation. STATs have cleus, bind to response elements in the promotors of target several properties which suite them as specific signal transducgenes, and activate transcription.