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1. Abstracts from the 20th International Symposium on Signal Transduction at the Blood-Brain Barriers

Author

Andrzej Małecki, Janina Skipor-Lahuta, Michal Toborek, N. Joan Abbott, David A. Antonetti, Enming Joe Su, Daniel A. Lawrence, Müge Atış, Uğur Akcan, Canan Uğur Yılmaz, Nurcan Orhan, Poyraz Düzgün, Umut Deniz Ceylan, Nadir Arıcan, Serçin Karahüseyinoğlu, Gizem Nur Şahin, Bülent Ahıshalı, Mehmet Kaya, Sidar Aydin, Armelle Klopstein, Britta Engelhardt, Julia Baumann, Chih-Chieh Tsao, Sheng-Fu Huang, Omolara Ogunshola, Elizaveta B. Boytsova, Andrey V. Morgun, Elena D. Khilazheva, Elena A. Pozhilenkova, Yana V. Gorina, Galina P. Martynova, Alla B. Salmina, David Bueno, Jordi Garcia-Fernàndez, Victor Castro, Marta Skowronska, Matheus Uba Chupel, Luciele Guerra Minuzzi, Edith Filaire, Ana Maria Teixeira, Mariangela Corsi, Romain Versele, Andrea Fuso, Emmanuel Sevin, Cherubino Di Lorenzo, Rita Businaro, Laurence Fenart, Fabien Gosselet, Pietra Candela, Mária A. Deli, Conor Delaney, Eoin O’Keefe, Michael Farrell, Sarah Doyle, Matthew Campbell, Lester R. Drewes, A. Appelt-Menzel, A. Cubukova, M. Metzger, R. Fischer, David M. F. Francisco, Rémy Bruggmann, Alexa Fries, Kinga G. Blecharz, Josephin Wagner, Lars Winkler, Ulf Schneider, Peter Vajkoczy, Mikio Furuse, Lydia Gabbert, Christina Dilling, Dmitri Sisario, Vladimir Soukhoroukov, Malgorzata Burek, S. Guérit, E. Fidan, K. Devraj, C. J. Czupalla, J. Macas, S. Thom, K. H. Plate, H. Gerhardt, S. Liebner, András Harazin, Alexandra Bocsik, Judit Váradi, Ferenc Fenyvesi, Vilmos Tubak, Miklós Vecsernyés, Hans Christian Helms, Helle Sønderby Waagepetersen, Carsten Uhd Nielsen, Birger Brodin, Zsófia Hoyk, Melinda E. Tóth, Nikolett Lénárt, Brigitta Dukay, Ágnes Kittel, Judit Vígh, Szilvia Veszelka, Fruzsina Walter, Ágnes Zvara, László Puskás, Miklós Sántha, Sabrina Engelhardt, Omolara O. Ogunshola, Anna Huber, Alexander Reitner, Samar Osmen, Kathrin Hahn, Neli Bounzina, Anna Gerhartl, Anna Schönegger, Hannes Steinkellner, Franco Laccone, Winfried Neuhaus, Natalie Hudson, Lucia Celkova, Anne Iltzsche, Svetlana Drndarski, David J Begley, Mette Mathiesen Janiurek, Krzysztof Kucharz, Christina Christoffersen, Lars Bo Nielsen, Martin Lauritzen, Rebecca H Johnson, Dan T Kho, Simon J O’Carroll, Catherine E Angel, E. Scott Graham, Jennifer Pereira, Christina Simoglou Karali, Vinton Cheng, Niloufar Zarghami, Manuel Sarmiento Soto, Yvonne Couch, Daniel C. Anthony, Nicola R. Sibson, John Kealy, Richard F. Keep, Lisa J. Routhe, Jianming Xiang, Hong Ye, Ya Hua, Torben Moos, Guohua Xi, M. Kristensen, A. Bach, K. Strømgaard, Nikolay Kutuzov, Melissa A. Lopes-Pinheiro, Jamie Lim, Alwin Kamermans, Jack van Horssen, Wendy W.J. Unger, Ruud Fontijn, Helga E. de Vries, Petra Majerova, Ralph M. Garruto, Luca Marchetti, David Francisco, Isabelle Gruber, Ruth Lyck, Mária Mészáros, Gergő Porkoláb, Lóránd Kiss, Ana-Maria Pilbat, Zsolt Török, Zsolt Bozsó, Lívia Fülöp, Alena Michalicova, Jaroslav Galba, Sandra Mihaljevic, Michal Novak, Andrej Kovac, Yoichi Morofuji, Takashi Fujimoto, Daisuke Watanabe, Shinsuke Nakagawa, Kenta Ujifuku, Nobutaka Horie, Tsuyoshi Izumo, Takeo Anda, Takayuki Matsuo, Fang Niu, Shilpa Buch, Ádám Nyúl-Tóth, Mihály Kozma, Péter Nagyőszi, Krisztina Nagy, Csilla Fazakas, János Haskó, Kinga Molnár, Attila E. Farkas, Péter Galajda, Imola Wilhelm, István A. Krizbai, Eoin Kelly, Eugene Wallace, Chris Greene, Stephanie Hughes, Niamh Doyle, Marian M. Humphries, Gerald A. Grant, Alon Friedman, Ronel Veksler, Michael G. Molloy, James F. Meaney, Niall Pender, Colin P. Doherty, Minseon Park, Arkadiusz Liskiewicz, Marta Przybyla, Daniela Kasprowska-Liśkiewicz, Marta Nowacka-Chmielewska, Andrzej Malecki, Ana Pombero, Raquel Garcia-Lopez, Marta Martinez-Morga, Salvador Martinez, Ofer Prager, Lyna Solomon-Kamintsky, Karl Schoknecht, Guy Bar-Klein, Dan Milikovsky, Udi Vazana, Dror Rosenbach, Richard Kovács, Zsolt Radak, Sabela Rodríguez-Lorenzo, Remy Bruggmann, Gijs Kooij, Helga E de Vries, Semyachkina-Glushkovskaya Oxana, Bragin Denis, Vodovozova Elena, Alekseeva Anna, Salmina Alla, Salmin Vladimir, Morgun Andrey, Malinovskaya Nataliya, Khilazheva Elena, Boytsova Elizaveta, Shirokov Alexander, Navolokin Nikita, Bucharskaya Alla, Yang Yirong, Abdurashitov Arkady, Gekalyuk Artem, Ulanova Mariya, Shushunova Anastasia, Bodrova Madina, Sagatova Artem, Khorovodov Alexander, Shareef Ali Esmat, Pavlov Valery, Tuchin Artem, Kurths Jürgen, Marcelle Silva de Abreu, Ana C. Calpena, Marta Espina, Maria Luisa García, Ignacio A. Romero, David Male, Steffen Storck, Anika Hartz, Jens Pahnke, Claus U. Surma, M. Surma, Z. Giżejewski, H. Zieliński, Aleksandra Szczepkowska, Marta Kowalewska, Agata Krawczynska, Andrzej P. Herman, Janina Skipor, Nicole Kachappilly, Mike Veenstra, Rosiris Leon Rivera, Dionna W. Williams, Susan Morgello, Joan W. Berman, Ursula Wyneken, Luis Federico Batiz, Arzu Temizyürek, Rouhollah Khodadust, Mutlu Küçük, Candan Gürses, Serkan Emik, Magdalena Zielińska, Marta Obara-Michlewska, Krzysztof Milewski, Edyta Skonieczna, Inez Fręśko, Edward A. Neuwelt, Ana Raquel Santa Maria, Ana Rita Bras, Dóra Lipka, Sándor Valkai, András Kincses, András Dér, and Maria A. Deli

Subjects
Neurology. Diseases of the nervous system, RC346-429
Published
2017
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2. Efficient Cargo Delivery into Adult Brain Tissue Using Short Cell-Penetrating Peptides.

Author

Caghan Kizil, Anne Iltzsche, Alvin Kuriakose Thomas, Prabesh Bhattarai, Yixin Zhang, and Michael Brand

Subjects
Medicine, Science
Abstract

Zebrafish brains can regenerate lost neurons upon neurogenic activity of the radial glial progenitor cells (RGCs) that reside at the ventricular region. Understanding the molecular events underlying this ability is of great interest for translational studies of regenerative medicine. Therefore, functional analyses of gene function in RGCs and neurons are essential. Using cerebroventricular microinjection (CVMI), RGCs can be targeted efficiently but the penetration capacity of the injected molecules reduces dramatically in deeper parts of the brain tissue, such as the parenchymal regions that contain the neurons. In this report, we tested the penetration efficiency of five known cell-penetrating peptides (CPPs) and identified two- polyR and Trans - that efficiently penetrate the brain tissue without overt toxicity in a dose-dependent manner as determined by TUNEL staining and L-Plastin immunohistochemistry. We also found that polyR peptide can help carry plasmid DNA several cell diameters into the brain tissue after a series of coupling reactions using DBCO-PEG4-maleimide-based Michael's addition and azide-mediated copper-free click reaction. Combined with the advantages of CVMI, such as rapidness, reproducibility, and ability to be used in adult animals, CPPs improve the applicability of the CVMI technique to deeper parts of the central nervous system tissues.

Published
2015
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3. Modifying plasmid-loaded HSA-nanoparticles with cell penetrating peptides – Cellular uptake and enhanced gene delivery

Author

Anne Iltzsche, Klaus Langer, Dennis Mulac, and J. Mesken

Subjects
0301 basic medicine, Cell Survival, Genetic enhancement, Cell, Pharmaceutical Science, Nanoparticle, Cell-Penetrating Peptides, 02 engineering and technology, Gene delivery, Biology, Transfection, Excipients, 03 medical and health sciences, Plasmid, medicine, Humans, Gene, Serum Albumin, Gene Transfer Techniques, DNA, Genetic Therapy, 021001 nanoscience & nanotechnology, Human serum albumin, Molecular biology, Cell biology, HEK293 Cells, 030104 developmental biology, medicine.anatomical_structure, Gene Products, tat, Nanoparticles, 0210 nano-technology, Plasmids, medicine.drug
Abstract

Gene therapy bears great potential for the cure of a multitude of human diseases. Research efforts focussed on the use of viral delivery vectors in the past decades, neglecting non-viral gene therapies of physical or chemical origin due to low transfection efficiency. However, side effects such as activation of oncogenes and inflammatory reactions upon immune cell activation are major obstacles impeding the clinical applicability of viral gene therapy vectors. The aim of this study was the development of a non-viral gene delivery system based on plasmid-loaded human serum albumin nanoparticles, which are biocompatible, biodegradable, and non-toxic in relevant concentrations. The surface of said nanoparticles was modified with different cell penetrating peptides, namely Tat, nona-arginine R9, and the penetratin analogue EB1. We hypothesise that the surface modified nanoparticles can effectively enter HEK 293T cells based on the cell penetrating properties of the different peptides attached. A variety of inhibitors were used targeting distinct uptake pathways in an effort to understand the mechanisms utilized by the various cell penetrating peptides on the surface of the nanoparticles. A significant increase in transfection efficiency compared to free DNA or polyplexes was seen for these novel delivery vectors.

Published
2017
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4. EB1 modified HSA nanoparticles as non-viral delivery vectors-Influence of peptide concentration on cell uptake

Author

Anne Iltzsche, Julia Mesken, Klaus Langer, and Dennis Mulac

Subjects
chemistry.chemical_classification, Chemistry, Genetic enhancement, Cell, Nanoparticle, Peptide, 02 engineering and technology, Transfection, 021001 nanoscience & nanotechnology, Human serum albumin, 030226 pharmacology & pharmacy, 03 medical and health sciences, 0302 clinical medicine, medicine.anatomical_structure, Cell-penetrating peptide, medicine, Biophysics, 0210 nano-technology, Cytotoxicity, medicine.drug
Abstract

Targeted nanoparticle-based gene therapy is a promising alternative to viral gene therapy approaches. The main limitations of non-viral delivery systems currently used are very low transfection efficiency, very high cytotoxicity, and lack of DNA protection leading to premature degradation. The aim of the present work was the development of a stable, biodegradable, nontoxic and plasmid-loaded human serum albumin (HSA)-based nanoparticle system. The nanoparticles were coupled to the cell penetrating peptide (CPP) EB1, a penetratin analogue, in different quantities to improve cell entry and transfection efficiency.

Published
2017
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5. Acute Inflammation Initiates the Regenerative Response in the Adult Zebrafish Brain

Author

Dorian Freudenreich, Jan Kaslin, Nikos Kyritsis, Sara Zocher, Michael Brand, Anne Iltzsche, Volker Kroehne, and Caghan Kizil

Subjects
Leukotrienes, Neurogenesis, ved/biology.organism_classification_rank.species, Central nervous system, Inflammation, Neural Stem Cells, medicine, Animals, Regeneration, Model organism, Zebrafish, Receptors, Leukotriene, Multidisciplinary, biology, ved/biology, Regeneration (biology), Anatomy, Regenerative process, biology.organism_classification, Cell biology, medicine.anatomical_structure, Brain Injuries, Acute Disease, Encephalitis, medicine.symptom, Signal transduction, Signal Transduction
Abstract

The Good Side of Inflammation The zebrafish brain is much more adept than the human brain at recovering after traumatic injury. Kyritsis et al. (p. 1353 , published online 8 November; see the Perspective by Stella ) investigated the cellular events that support regeneration in the zebrafish brain. Although inflammation is part of the response in both settings, the zebrafish brain goes on to initiate proliferation of replacement neurons. By inciting inflammation without neuronal damage, radial glial cells could be pushed into neurogenesis.

Published
2012
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6. Micromanipulation of Gene Expression in the Adult Zebrafish Brain Using Cerebroventricular Microinjection of Morpholino Oligonucleotides

Author

Michael Brand, Jan Kaslin, Caghan Kizil, and Anne Iltzsche

Subjects
Telencephalon, Morpholino, Microinjections, General Chemical Engineering, radial glial cells, Gene Expression, Biology, cerebroventricular microinjection, Biochemistry, General Biochemistry, Genetics and Molecular Biology, Morpholinos, Neurobiology, Gene expression, Gene Knockdown Techniques, Genetics, microinjection, Animals, Microinjection, Zebrafish, Molecular Biology, Issue 75, Injections, Intraventricular, Danio rerio, General Immunology and Microbiology, Regeneration (biology), General Neuroscience, animal model, Neurogenesis, neurosciences, Brain, Oligonucleotides, Antisense, biology.organism_classification, Cellular Biology, morpholino oligonucleotides, Forebrain, Neuroscience, Developmental Biology
Abstract

Manipulation of gene expression in tissues is required to perform functional studies. In this paper, we demonstrate the cerebroventricular microinjection (CVMI) technique as a means to modulate gene expression in the adult zebrafish brain. By using CVMI, substances can be administered into the cerebroventricular fluid and be thoroughly distributed along the rostrocaudal axis of the brain. We particularly focus on the use of antisense morpholino oligonucleotides, which are potent tools for knocking down gene expression in vivo. In our method, when applied, morpholino molecules are taken up by the cells lining the ventricular surface. These cells include the radial glial cells, which act as neurogenic progenitors. Therefore, knocking down gene expression in the radial glial cells is of utmost importance to analyze the widespread neurogenesis response in zebrafish, and also would provide insight into how vertebrates could sustain adult neurogenesis response. Such an understanding would also help the efforts for clinical applications in human neurodegenerative disorders and central nervous system regeneration. Thus, we present the cerebroventricular microinjection method as a quick and efficient way to alter gene expression and neurogenesis response in the adult zebrafish forebrain. We also provide troubleshooting tips and other useful information on how to carry out the CVMI procedure.

Published
2013

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