di Patrizio Tressoldi , Enrico Facco , Daniela Lucangeli
This contribution to the science of consciousness aims at comparing how two different theories can explain the emergence of different qualia experiences: meta-awareness, metacognition, the placebo effect, out-of-body experiences, cognitive therapy, meditation-induced brain changes, etc. The first theory postulates that qualia experiences derive from specific neural patterns, and the second one that qualia experiences derive from the interaction of a proto-consciousness with the brain’s neural activity. From this comparison, it will be possible to judge which one seems to better explain the different qualia experiences and to offer a more promising research agenda.
1Maria Cecilia Hospital, Gruppo Villa Maria (GVM) Care & Research and Ettore Sansavini Health Science Foundation, Cotignola and Lugo, Ravenna, Italy; and 2Department of Experimental, Diagnostic and Specialty Medicine, University of Bologna, Italy
Margherita Maioli, Salvatore Rinaldi, Sara Santaniello, Alessandro Castagna, Gianfranco Pigliaru, Sara Gualini, Claudia Cavallini, Vania Fontani, and Carlo Ventura Department of Biomedical Sciences, University of Sassari, Sassari, Italy Laboratory of Molecular Biology and Stem Cell Engineering, National Institute of Biostructures and Biosystems, Bologna, Italy Rinaldi Fontani Institute, Florence, Italy §Cardiovascular Department, S. Orsola-Malpighi Hospital, University of Bologna, Bologna, Italy Somatic cells can be directly reprogrammed to alternative differentiated fates without first becoming stem/ progenitor cells. Nevertheless, the initial need for viral-mediated gene delivery renders this strategy unsafe in humans. Here, we provide evidence that exposure of human skin fibroblasts to a Radio Electric Asymmetric Conveyer (REAC), an innovative device delivering radio electric conveyed fields at a radiofrequency of 2.4 GHz, afforded remarkable commitment toward cardiac, neuronal, and skeletal muscle lineages. REAC induced the transcription of tissue-restricted genes, including Mef2c, Tbx5, GATA4, Nkx2.5, and prodynorphin for cardiac reprogramming, as well as myoD, and neurogenin 1 for skeletal myogenesis and neurogenesis, respectively. Conversely, REAC treatment elicited a biphasic effect on a number of stemness-related genes, leading to early transcriptional increase of Oct4, Sox2, cMyc, Nanog, and Klf4 within 6–20 h, followed by a downregulation at later times. The REAC action bypassed a persistent reprogramming toward an induced pluripotent stem celllike state and involved the transcriptional induction of the NADPH oxidase subunit Nox4. Our results show for the first time the feasibility of using a physical stimulus to afford the expression of pluripotentiality in human adult somatic cells up to the attainment of three major target lineages for regenerative medicine.
Received for publication, November 20, 2009, and in revised form, January 15, 2010 Published, JBC Papers in Press, January 22, 2010, DOI 10.1074/jbc.M109.087254 Vincenzo Lionetti, Silvia Cantoni, Claudia Cavallini, Francesca Bianchi, Sabrina Valente, Irene Frascari, Elena Olivi, Giovanni D. Aquaro, Francesca Bonavita, Ignazio Scarlata, Margherita Maioli, Valentina Vaccari, Riccardo Tassinari, Antonietta Bartoli, Fabio A. Recchia, Gianandrea Pasquinelli, and Carlo Ventura From the Sector of Medicine, Scuola Superiore S. Anna, I-56124 Pisa, Italy, the Laboratory of Molecular Biology and Stem Cell Engineering, Cardiovascular Department-National Institute of Biostructures and Biosystems, S. Orsola-Malpighi Hospital, University of Bologna, I-40138 Bologna, Italy, the Bioscience Institute, RSM-47891 Falciano, Republic of San Marino, the §Institute of Clinical Physiology, Consiglio Nazionale delle Ricerche Fondazione G. Monasterio, I-56124 Pisa, Italy, the ‡‡Department of Biomedical Sciences, University of Sassari, I-07100 Sassari, Italy, the §§Department of Physics, University of Pisa, I-56124 Pisa, Italy, the Department of Physiology, New York Medical College, Valhalla, New York 10595, and the **Department of Hematology, Oncology, and Clinical Pathology, University of Bologna, I-40138 Bologna, Italy
A NOVEL DIFFERENTIATING GLYCOCONJUGATE AFFORDING A HIGH THROUGHPUT OF CARDIOGENESIS IN EMBRYONIC STEM CELLS
Received for publication, February 20, 2004, and in revised form, March 16, 2004 Published, JBC Papers in Press, March 24, 2004, DOI 10.1074/jbc.M401869200 Carlo Ventura, Margherita Maioli, Yolande Asara, Daniela Santoni, Ignazio Scarlata, Silvia Cantoni, and Alberto Perbellini From the Laboratory of Molecular Biology and Stem Cell Engineering, National Institute of Biostructures and Biosystems, University of Bologna, I-40138 Bologna, Italy, the Department of Biomedical Sciences, University of Sassari, I-07100 Sassari, Italy, and the Department of Biochemistry, Biophysics, and Chemistry of Macromolecules, University of Trieste, I-34127 Trieste, Italy
Received for publication, October 3, 2006, and in revised form, March 9, 2007 Published, JBC Papers in Press, March 15, 2007, DOI 10.1074/jbc.M609350200 Carlo Ventura, Silvia Cantoni, Francesca Bianchi, Vincenzo Lionetti, Claudia Cavallini, Ignazio Scarlata, Laura Foroni, Margherita Maioli, Laura Bonsi, Francesco Alviano, Valentina Fossati, Gian Paolo Bagnara, Gianandrea Pasquinelli, Fabio A. Recchia, and Alberto Perbellini From the Laboratory of Molecular Biology and Stem Cell Engineering, Institute of Cardiology, National Institute of Biostructures and Biosystems, University of Bologna, I-40138 Bologna, Italy, Department of Experimental Pathology, University of Bologna, I-40138 Bologna, Italy, Department of Histology, Embryology, and Applied Biology, University of Bologna, I-40138 Bologna, Italy, Sector of Medicine, Scuola Superiore S. Anna, CNR Institute of Clinical Physiology, I-56124 Pisa, Italy and Department of Physiology, New York Medical College, Valhalla, New York 10595, and Department of Biomedical Sciences, University of Sassari, I-07100 Sassari, Italy
Carlo Ventura, Elisabetta Zinellu, Emiliana Maninchedda, Margherita Maioli Abstract—The cardiac differentiation of embryonic stem (ES) cells was found to involve prodynorphin gene and dynorphin B expression and was associated with the interaction of secreted dynorphin B with cell surface opioid receptors coupled with protein kinase C (PKC) signaling and complex subcellular redistribution patterning of selected PKC isozymes. Here, confocal microscopy revealed the presence of immunoreactive dynorphin B–like material in GTR1 ES cells, suggesting that dynorphin peptides may also act intracellularly. Opioid binding sites were identified in ES cell nuclei, with a single dissociation constant in the low nanomolar range. A significant increase in Bmax for a opioid receptor ligand was observed in nuclei isolated from ES-derived cardiomyocytes compared with nuclei from undifferentiated cells. Direct exposure of nuclei isolated from undifferentiated ES cells to dynorphin B or U-50,488H, a synthetic opioid receptor agonist, time- and dose-dependently activated the transcription of GATA-4 and Nkx-2.5, 2 cardiac lineage–promoting genes. Nuclear exposure to dynorphin B also enhanced the rate of prodynorphin gene transcription. These responses were abolished in a stereospecific fashion by the incubation of isolated nuclei with selective opioid receptor antagonists. Nuclei isolated from undifferentiated cells were able to phosphorylate the acrylodan-labeled MARCKS peptide, a high-affinity fluorescent PKC substrate. Exposure of isolated nuclei to dynorphin B induced a remarkable increase in nuclear PKC activity, which was suppressed by opioid receptor antagonists. Nuclear treatment with PKC inhibitors abolished the capability of dynorphin B to prime the transcription of cardiogenic genes. (Circ Res. 2003;92:623-629.)
Carlo Ventura, Elisabetta Zinellu, Emiliana Maninchedda, Marina Fadda, Margherita Maioli
Abstract—The prodynorphin gene and its product, dynorphin B, have been found to promote cardiogenesis in embryonic cells by inducing the expression of GATA-4 and Nkx-2.5, two transcription factor– encoding genes essential for cardiogenesis. The molecular mechanism(s) underlying endorphin-induced cardiogenesis remain unknown. In the present study, we found that GTR1 embryonic stem (ES) cells expressed cell surface opioid receptors, as well as protein kinase C (PKC)-, -1, -2, -, -, and -. Cardiac differentiation was associated with a marked increase in the Bmax value for a selective opioid receptor ligand and complex subcellular redistribution of selected PKC isozymes. PKC-, -1, -2, -, and - all increased in the nucleus of ES-derived cardiac myocytes, compared with nuclei from undifferentiated cells. In both groups of cells, PKC- and - were mainly expressed at the nuclear level. The nuclear increase of PKC-, -1, and -2 was due to a translocation from the cytosolic compartment. In contrast, the increase of both PKC- and PKC- in the nucleus of ES-derived cardiomyocytes occurred independently of enzyme translocation, suggesting changes in isozyme turnover and/or gene expression during cardiogenesis. No change in PKC- expression was observed during cardiac differentiation. Opioid receptor antagonists prevented the nuclear increase of PKC-, PKC-1, and PKC-2 and reduced cardiomyocyte yield but failed to affect the nuclear increase in PKC- and -. PKC inhibitors prevented the expression of cardiogenic genes and dynorphin B in ES cells and abolished their development into beating cardiomyocytes. (Circ Res. 2003;92:617-622.) Key Words: protein kinase C cardiac differentiation embryonic stem cells gene expression endorphins