MicroRNA miR-196a is a central regulator of HOX-B7 and BMP4 expression in malignant melanoma

Since bone morphogenetic proteins (BMPs) play an important role in melanoma progression, we aimed to determine the molecular mechanisms leading to overexpression of BMP4 in melanoma cells compared to normal melanocytes. With our experimental approach we revealed that loss of expression of a microRNA represents the starting point for a signaling cascade finally resulting in overexpression of BMP4 in melanoma cells. In detail, strongly reduced expression of the microRNA miR-196a in melanoma cells compared to healthy melanocytes leads to enhanced HOX-B7 mRNA and protein levels, which subsequently raise Ets-1 activity by inducing basic fibroblast growth factor (bFGF). Ets-1 finally accounts for induction of BMP4 expression. We were furthermore able to demonstrate that bFGF-mediated induction of migration is achieved via activation of BMP4, thus determining BMP4 as major modulator of migration in melanoma. In summary, our study provides insights into the early steps of melanoma progression and might thereby harbor therapeutic relevance.     

The admiR-able advances in cardiovascular biology through the zebrafish model system

MicroRNAs are small non-coding RNAs endogenously expressed by all tissues during development and adulthood. They regulate gene expression by controlling the stability of targeted messenger RNA. In cardiovascular tissues microRNAs play a role by modulating essential genes involved in heart and blood vessel development and homeostasis. The zebrafish (Danio rerio) system is a recognized vertebrate model system useful to study cardiovascular biology; recently, it has been used to investigate microRNA functions during natural and pathological states. In this review, we will illustrate the advantages of the zebrafish model in the study of microRNAs in heart and vascular cells, providing an update on recent discoveries using the zebrafish to identify new microRNAs and their targeted genes in cardiovascular tissues. Lastly, we will provide evidence that the zebrafish is an optimal model system to undercover new microRNA functions in vertebrates and to improve microRNA-based therapeutic approaches.     

The use of metabolomics to dissect plant responses to abiotic stresses

Plant metabolism is perturbed by various abiotic stresses. As such the metabolic network of plants must be reconfigured under stress conditions in order to allow both the maintenance of metabolic homeostasis and the production of compounds that ameliorate the stress. The recent development and adoption of metabolomics and systems biology approaches enable us not only to gain a comprehensive overview, but also a detailed analysis of crucial components of the plant metabolic response to abiotic stresses. In this review we introduce the analytical methods used for plant metabolomics and describe their use in studies related to the metabolic response to water, temperature, light, nutrient limitation, ion and oxidative stresses. Both similarity and specificity of the metabolic responses against diverse abiotic stress are evaluated using data available in the literature. Classically discussed stress compounds such as proline, γ-amino butyrate and polyamines are reviewed, and the widespread importance of branched chain amino acid metabolism under stress condition is discussed. Finally, where possible, mechanistic insights into metabolic regulatory processes are discussed.     

Existence of several non-specific recognition systems in macrophages]

Rat peritoneal cells were made to bind five particle species: immunoglobulin-coated Sheep red cells, glutaraldehyde-treated Sheep red cells, latex beads, leishmania and tumor cells. The dependence of binding on various physico-chemical parameters was studied. The binding of latex beads or Leishmania was not inhibited by cold (4 degrees C), sodium azide, cytochalasin B and ethyleneglycol or dimethylsulphoxide. The binding of immunoglobulin-coated Sheep red cells was unaffected by cold and azide, but it was inhibited by cytochalasin B, ethyleneglycol and dimethylsulphoxide. The binding of glutaraldehyde-treated Sheep red cells was inhibited by cold, azide and ethyleneglycol, but it resisted cytochalasin B and dimethylsulphoxide. The binding of tumor cells was inhibited by azide, cytochalasin B, ethyleneglycol and dimethylsulphoxide. It is concluded that: (a) macrophages are endowed several sets of non-specific binding structures that are differently affected by physico-chemical parameters, which provides a simple way of characterizing them; (b) the expression of a given binding structure on the macrophage membrane is modulated by metabolic inhibitors; (c) some lymphocytes were able to bind tumor cells or Leishmania. Thus, lymphocytes and macrophages might share some non-specific adhesive structures.     

Micromanaging freeze tolerance: the biogenesis and regulation of neuroprotective microRNAs in frozen brains

When temperatures plummet below 0 °C, wood frogs (Rana sylvatica) can endure the freezing of up to?~?65% of their body water in extracellular ice masses, displaying no measurable brain activity, no breathing, no movement, and a flat-lined heart. To aid survival, frogs retreat into a state of suspended animation characterized by global suppression of metabolic functions and reprioritization of energy usage to essential survival processes that is elicited, in part, by the regulatory controls of microRNAs. The present study is the first to investigate miRNA biogenesis and regulation in the brain of a freeze tolerant vertebrate. Indeed, proper brain function and adaptations to environmental stimuli play a crucial role in coordinating stress responses. Immunoblotting of miRNA biogenesis factors illustrated an overall reduction in the majority of these processing proteins suggesting a potential suppression of miRNA maturation over the freeze–thaw cycle. This was coupled with a large-scale RT-qPCR analysis of relative expression levels of 113 microRNA species in the brains of control, 24 h frozen, and 8 h thawed R. sylvatica. Of the 41 microRNAs differentially regulated during freezing and thawing, only two were significantly upregulated. Bioinformatic target enrichment of the downregulated miRNAs, performed at the low temperatures experienced during freezing and thawing, predicted their involvement in the potential activation of various neuroprotective processes such as synaptic signaling, intracellular signal transduction, and anoxia/ischemia injury protection. The predominantly downregulated microRNA fingerprint identified herein suggests a microRNA-mediated cryoprotective mechanism responsible for maintaining neuronal functions and facilitating successful whole brain freezing and thawing.     

Hormonal control of Sertoli cell metabolism regulates spermatogenesis

Hormonal regulation is essential to spermatogenesis. Sertoli cells (SCs) have functions that reach far beyond the physical support of germ cells, as they are responsible for creating the adequate ionic and metabolic environment for germ cell development. Thus, much attention has been given to the metabolic functioning of SCs. During spermatogenesis, germ cells are provided with suitable metabolic substrates, in a set of events mediated by SCs. Multiple signaling cascades regulate SC function and several of these signaling pathways are hormone-dependent and cell-specific. Within the seminiferous tubules, only SCs possess receptors for some hormones rendering them major targets for the hormonal signaling that regulates spermatogenesis. Although the mechanisms by which SCs fulfill their own and germ cells metabolic needs are mostly studied in vitro, SC metabolism is unquestionably a regulation point for germ cell development and the hormonal control of these processes is required for a normal spermatogenesis.     

Defining a neuron: neuronal ELAV proteins

Neuronal cells strongly depend on the control exerted by RNA-binding proteins (RBPs) on gene expression for the establishment and maintenance of their phenotype. Neuronal ELAV (nELAV) proteins are RBPs able to influence virtually every aspect of the postsynthesis fate of bound mRNAs, from polyadenylation, alternative splicing and nuclear export to cytoplasmic localization, stability and translation. They enhance gene expression through the last two, best documented activities, increasing mRNA half-life and promoting protein synthesis by a still-unknown molecular mechanism. Developmentally, nELAV proteins have been shown to act as inducers of the transition between neural stem/progenitor cells and differentiation-committed cells, also assisting these neuroblasts in the completion of their maturation program. In brain physiology, they are also the first RBPs demonstrated to have a pivotal role in memory, where they probably control mRNA availability for translation in subcellular domains, thereby providing a biochemical means for selective increase in synaptic strength. Received 15 January 2007; received after revision 10 August 2007; accepted 6 September 2007     

Spontaneous fusion between cancer cells and endothelial cells

Endothelial cells line the inside of blood and lymphatic vessels, and cancer cells must cross this barrier, first to gain access to the circulation, and, second, to exit and metastasize. How this occurs is incompletely understood. We now demonstrate that human cancer cells are able to fuse with endothelial cells to form hybrid cells displaying proteins and chromosomal markers characteristic of both parent cells. The hybrid cells are viable and capable of undergoing mitosis. Fusions between cancer cells and endothelial cells were shown to occur both in vitro, in co-cultures of human breast cancer cells and endothelial cells, and in vivo, following intravascular dissemination of human breast cancer cells in nude mice. These observations demonstrate a new type of cancer-endothelial cell interaction that may be of fundamental importance to the process of metastasis.Received 10 May 2004; received after revision 21 June 2004; accepted 2 July 2004     

Structure of gap junctions in cultures of normal and neoplastic bladder epithelial cells

Summary Normal rat urinary bladder epithelial cells contain small subunit (PF-1) and large subunit (PF-2) gap junctions, whereas carcinoma cells only contain PF-1 gap junctions. The absence of PF-2 gap junctions, which are composed of larger connexons with slightly larger ionic channels, may contribute to altered metabolic coupling between urinary bladder carcinoma cells.Acknowledgments. The authors wish to thank Ms Denise Wiler and Mr William Leonard for expert technical assistance. This work was supported by funds from the National Cancer Institute, CA-25034, and in part by the Otho SA Sprague Memorial Institute.     

MicroRNA-128 downregulates Bax and induces apoptosis in human embryonic kidney cells

MicroRNAs (miRNAs) are short ~21-nt non-coding RNA molecules that have been shown to regulate a number of biological processes. Previous reports have shown that overexpression of miR-128 in glioma cells inhibited cell proliferation. Literature also suggests that miR-128 negatively regulates prostate cancer cell invasion. Here, we show that overexpression of hsa-miR-128, a brain-enriched microRNA, induces apoptosis in HEK293T cells as elucidated by apoptosis assay, cell cycle changes, loss of mitochondrial membrane potential and multicaspase assay. By in silico analysis, we identified a putative target site within the 3′ untranslated region (UTR) of Bax, a proapoptotic member of the apoptosis pathway. We found that ectopic expression of hsa-miR-128 suppressed a luciferase reporter containing the Bax-3′ UTR and reduced the levels of Bax in HEK293T cells. Taken together, our study demonstrates that overexpression of hsa-miR-128 not only induces apoptosis in HEK293T cells but also is an endogenous regulator of Bax protein.     

Loss of steroidogenesis in a mouse adrenal cell line after simian adenovirus 7 (SA7) transformation]

After transformation by the simian adenovirus 7 (SA7), the Y-1 Mouse adrenal tumor cells are no longer able to produce stero?ds either spontaneously, or after specific stimulation. Nevertheless, these stimulations are able to increase the level of cAMP both in transformed and original cell line. The differentiation is associated with an intracellular modification of stero?dogenesis which seems to be localized in the metabolic chain of stero?d synthesis after the increase of the cAMP level.     

(—)-Epigallocatechin-3-gallate interferes with mast cell adhesiveness, migration and its potential to recruit monocytes

Mast cells are multipotent effector cells of the immune system. They are able to induce and enhance angiogenesis via multiple pathways. (-)-Epigallocatechin-3-gallate (EGCG), a major component of green tea and a putative chemopreventive agent, was reported to inhibit tumor invasion and angiogenesis, processes that are essential for tumor growth and metastasis. Using the human mast cell line HMC-1 and commercial cDNA macroarrays, we evaluated the effect of EGCG on the expression of angiogenesis-related genes. Our data show that among other effects, EGCG treatment reduces expression of two integrins (alpha5 and beta3) and a chemokine (MCP1), resulting in a lower adhesion of mast cells associated with a decreased potential to produce signals eliciting monocyte recruitment. These effects on gene expression levels are functionally validated by showing inhibitory effects in adhesion, aggregation, migration and recruitment assays.     

Ranbp2 haploinsufficiency mediates distinct cellular and biochemical phenotypes in brain and retinal dopaminergic and glia cells elicited by the Parkinsonian neurotoxin, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)

Many components and pathways transducing multifaceted and deleterious effects of stress stimuli remain ill-defined. The Ran-binding protein 2 (RanBP2) interactome modulates the expression of a range of clinical and cell-context-dependent manifestations upon a variety of stressors. We examined the role of Ranbp2 haploinsufficiency on cellular and metabolic manifestations linked to tyrosine-hydroxylase (TH(+)) dopaminergic neurons and glial cells of the brain and retina upon acute challenge to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), a parkinsonian neurotoxin, which models facets of Parkinson disease. MPTP led to stronger akinetic parkinsonism and slower recovery in Ranbp2 (+/-) than wild-type mice without viability changes of brain TH(+)-neurons of either genotype, with the exception of transient nuclear atypia via changes in chromatin condensation of Ranbp2 (+/-) TH(+)-neurons. Conversely, the number of wild-type retinal TH(+)-amacrine neurons compared to Ranbp2 (+/-) underwent milder declines without apoptosis followed by stronger recoveries without neurogenesis. These phenotypes were accompanied by a stronger rise of EdU(+)-proliferative cells and non-proliferative gliosis of GFAP(+)-Müller cells in wild-type than Ranbp2 (+/-) that outlasted the MPTP-insult. Finally, MPTP-treated wild-type and Ranbp2 (+/-) mice present distinct metabolic footprints in the brain or selective regions thereof, such as striatum, that are supportive of RanBP2-mediated regulation of interdependent metabolic pathways of lysine, cholesterol, free-fatty acids, or their β-oxidation. These studies demonstrate contrasting gene-environment phenodeviances and roles of Ranbp2 between dopaminergic and glial cells of the brain and retina upon oxidative stress-elicited signaling and factors triggering a continuum of metabolic and cellular manifestations and proxies linked to oxidative stress, and chorioretinal and neurological disorders such as Parkinson.     

Selective effect of burn injury on splenic CD11c+ dendritic cells and CD8α+CD4−CD11c+ dendritic cell subsets

Burn injury causes an immunosuppression associated with suppressed adaptive immune function. Dendritic cells (DCs) are APCs for which signaling via their Toll-like receptors (TLRs) induces their maturation and activation, which is essential for the adaptive immune response. In this study, we examined if burn injury alters the TLR activity of splenic DCs. After injury, we noticed that DC functions were impaired, characterized by a suppressed capacity to prime naive T cells when triggering the TLR4 signaling cascade using specific ligands (LPS or rHSP60). The observed perturbations on LPS-primed DCs isolated from burned mice exhibited significantly diminished IL-12p40 production and enhanced IL-10 secretion-associated impairment in mitogen-activated protein kinase activation. Interestingly, we observed a decrease of TLR4/MD-2 expression on the CD8α⁺ DC subset that persisted following LPS stimulation. The altered TLR4 expression on LPS-stimulated CD8α⁺ DCs was associated with reduced capacity to produce IL-12 after stimulation. Our results suggested that TLR4 reactivity on DCs, especially CD8α⁺ DCs, is disturbed after burn injury.