5 research outputs found
Acoustically-Targeted Delivery of Engineered Vectors to the Brain
No full textThe brain is connected through neuronal pathways that together turn individual neurons into the brain. Targeted gene delivery to the brain is a critical tool for neuroscience research and has significant potential to treat human disease. However, the site-specific delivery of common gene vectors such as adeno-associated viruses (AAVs) is typically performed via invasive injections, which limit its applicable scope of research and clinical applications. Alternatively, focused ultrasound blood-brain-barrier opening (FUS-BBBO), performed noninvasively, enables the site-specific entry of AAVs into the brain from systemic circulation. However, when used in conjunction with natural AAV serotypes, this approach has limited transduction efficiency and results in substantial undesirable transduction of peripheral organs. Recent discovery of viral vectors that can selectively transduce neuronal projections have enabled discovery of how various brain regions communicate to affect behavior, but they also suffer from delivery challenges. Here, we evaluate the potential of two engineered viral vectors, AAV.FUS and AAV9.retro, for noninvasive site-specific transduction of the neurons and neuronal projections after systemic delivery. The resulting vectors substantially enhance ultrasound-targeted gene delivery, neuronal tropism and transduction of neuronal pathways while reducing peripheral transduction. In addition to enhancing the only known approach to noninvasively target gene delivery to specific brain regions, these results establish the ability of AAV vectors to be evolved for specific delivery applications and therapies
Focused Ultrasound Induced Blood-Brain Barrier Opening for Targeting Brain Structures and Evaluating Chemogenetic Neuromodulation
No full textAcoustically Targeted Chemogenetics (ATAC) allows for the noninvasive control of specific neural circuits. ATAC achieves such control through a combination of focused ultrasound (FUS) induced blood-brain barrier opening (FUS-BBBO), gene delivery with adeno-associated viral (AAV) vectors, and activation of cellular signaling with engineered, chemogenetic, protein receptors and their cognate ligands. With ATAC, it is possible to transduce both large and small brain regions with millimeter precision using a single noninvasive ultrasound application. This transduction can later allow for a long-term, noninvasive, device-free neuromodulation in freely moving animals using a drug. Since FUS-BBBO, AAVs, and chemogenetics have been used in multiple animals, ATAC should also be scalable for the use in other animal species. This paper expands upon a previously published protocol and outlines how to optimize the gene delivery with FUS-BBBO to small brain regions with MRI-guidance but without a need for a complicated MRI-compatible FUS device. The protocol, also, describes the design of mouse targeting and restraint components that can be 3D-printed by any lab and can be easily modified for different species or custom equipment. To aid reproducibility, the protocol describes in detail how the microbubbles, AAVs, and venipuncture were used in ATAC development. Finally, an example data is shown to guide the preliminary investigations of studies utilizing ATAC
Additional file 1 of A transgenic mouse embryonic stem cell line for puromycin selection of V0V interneurons from heterogenous induced cultures
No full textAdditional file 1: Fig. S1. Day 11 quantification of Evx1, Lim1, and βIII tubulin by ICC image analysis. V0V IN induction cultures were either unselected or selected with 4 µg/ml puro from day 10 to 11, then fixed and stained on day 11. N = 1 with n = 2 for unselected and n = 5 for selected cultures. Error bars are S.E.M. among technical replicates. Figure S1 is associated with Fig. 3. Additional file 1: Fig. S2. Day 22 quantification of mature neuron and synaptic markers by ICC image analysis. V0V IN induction cultures were selected with 4 µg/ml puro from day 10 to 11 and cultured until day 22, when they were fixed and stained. Percentage of selected cultured cells expressing A VGLUT2, NeuN, and MAP2 or B VAChT, Bassoon, and MAP2 and respective co-staining combinations are shown. N = 2 with n = 3–5 and error bars are S.E.M. Figure S2 is associated with Fig. 4
Engineering viral vectors for acoustically targeted gene delivery
No full textAbstract Targeted gene delivery to the brain is a critical tool for neuroscience research and has significant potential to treat human disease. However, the site-specific delivery of common gene vectors such as adeno-associated viruses (AAVs) is typically performed via invasive injections, which limit its applicable scope of research and clinical applications. Alternatively, focused ultrasound blood-brain-barrier opening (FUS-BBBO), performed noninvasively, enables the site-specific entry of AAVs into the brain from systemic circulation. However, when used in conjunction with natural AAV serotypes, this approach has limited transduction efficiency and results in substantial undesirable transduction of peripheral organs. Here, we use high throughput in vivo selection to engineer new AAV vectors specifically designed for local neuronal transduction at the site of FUS-BBBO. The resulting vectors substantially enhance ultrasound-targeted gene delivery and neuronal tropism while reducing peripheral transduction, providing a more than ten-fold improvement in targeting specificity in two tested mouse strains. In addition to enhancing the only known approach to noninvasively target gene delivery to specific brain regions, these results establish the ability of AAV vectors to be evolved for specific physical delivery mechanisms
Book review of president emeritus Dr. Ugo Mifsud Bonnici, An introduction to the law of education (Malta University Press 2013)
No full textAn Introduction to the Law of Education is President Emeritus Ugo Mifsud
Bonnici's latest introductory monograph on a specific branch of Maltese Law -
the Law of Education. His previous books in this series include: An Introduction
to Cultural Heritage Law, published in 2008 by Midsea Books and An Introduction
to Comparative Law published earlier in 2004 by Malta University Press. He has
also written various other scholarly works which are studied by students at the
Faculty of Laws such as the authoritative ll-Manwal tal-President ("The
President's Manual") written while Dr Mifsud Bonnici served as President of
Malta, and Kif Sirna Repubblika ("How We Became a Republic") published in
1999 which both provide an insightful account of the post-independence
development of constitutional law. In his career, President Emeritus Dr Mifsud
Bonnici has been inextricably associated with education not only through his
various academic and popular writings on the subject, but also because as a
seasoned politician he has always covered the sector of education both as
Shadow Minister for Education between 1972 and 1987, and Education Minister
from 1987 till 1994. A number of the matters referred to in the book being
reviewed are known to him, di scienza propria, as a painstaking researcher and
eminent scholar, and because of his past political activism. Although President
Emeritus Dr Mifsud Bonnici has authored excellent books on Maltese cultural
heritage law, comparative law and others, I would hazard a guess and state that
this latest book on the law of education is probably the one he cherishes most,
given that during his eminent political career, he has lived and been inseparably
linked to this subject; so much so that his name is synonymous to education. He
now fascinatingly narrates all this in his latest oeuvre. His knowledge of the
subject is not only comprehensive, but also personal. He has been directly
involved as a key protagonist in the drafting, making and unfolding of the law on
the subject in question. This places him in a very advantageous situation when
compared to other writers, more so that he has authored the Education Act 1988
himself and was directly involved in its promulgation, in different areas of
education such as University, primary schools, secondary schools, etc. These
experiences assisted him in providing the reader with certain facts which he was
directly privy to due to the various offices he has occupied in his very successful
political career. This added value that the author brings in writing this volume
and makes this contribution more interesting to read and more revealing of the
actual reasoning behind the provisions of the Education Act, 1988 and as
subsequently amended. The insight which he provides in the development of the law of education is therefore original and provides more information than one
would normally find in a government file or any other primary historical source.
In this respect, to a certain extent, this book can be viewed as the first attempt by
President Emeritus Dr Ugo Mifsud Bonnici at writing an autobiography. This is
undoubtedly, the added value of this book.peer-reviewe
