Ubiquitous expression of human SCA2 gene under the regulation of the SCA2 self promoter cause specific Purkinje cell degeneration in transgenic mice.
The objective of this work was the generation of an animal model of the SCA2 disease for future studies on the benefits of therapeutic molecules and neuropathological mechanisms that underline this human disorder. The transgenic fragment was microinjected into pronuclei of B6D2F1 X OF1 mouse hybrid strain. For Northern blots, RNAs were hybridized with a human cDNA fragment from the SCA2 gene and a mouse beta-actin cDNA fragment. Monoclonal antibody directed to the N-terminal of the ataxin 2 protein with 22Q was used for Western blot analysis. A rotating rod apparatus was utilized to measure motor coordination of mice. Immunohistochemical detection of Purkinje neurons was performed with anti-calbindin 28K as primary antibody. Ubiquitous expression of the SCA2 transgene with 75 CAG repeats regulated by the SCA2 self promoter was obtained after generation of our transgenic mice. Analysis of transgenic mice revealed significant differences of motor coordination compared with the wild type littermates. Specific degeneration of Purkinje neurons and transgene over-expression in the brain, liver and skeletal muscle, rather than in lungs and kidneys was also observed, resembling the expression pattern of the ataxin 2 in humans.
Aguiar
Jorge
J
Department of Gene Therapy, Division of Pharmaceutics, Center for Genetic Engineering and Biotechnology, CIGB, Ave 31 between 158 and 190, Havana City, CP 10 600 Havana, Cuba. jorge.aguiar@cigb.edu.cu
Fernández
Julio
J
Aguilar
Anselmo
A
Mendoza
Yssel
Y
Vázquez
María
M
Suárez
José
J
Berlanga
Jorge
J
Cruz
Silian
S
Guillén
Gerardo
G
Herrera
Luis
L
Velázquez
Luis
L
Santos
Nieves
N
Merino
Nelson
N
D000704:Analysis of Variance; D000818:Animals; D000067528:Ataxins; D015152:Blotting, Northern / Q000379:methods; D015153:Blotting, Western / Q000379:methods; D064092:Calbindin 1; D064026:Calbindins; D004195:Disease Models, Animal; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D051379:Mice; D008822:Mice, Transgenic; D009043:Motor Activity / Q000502:physiology; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000502:physiology*; D011689:Purkinje Cells / Q000378:metabolism / Q000473:pathology*; D012333:RNA, Messenger / Q000096:biosynthesis; D012045:Regulatory Sequences, Nucleic Acid; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D045442:Rotarod Performance Test / Q000379:methods; D064030:S100 Calcium Binding Protein G / Q000378:metabolism; D013132:Spinocerebellar Degenerations / Q000378:metabolism* / Q000473:pathology* / Q000503:physiopathology; D013997:Time Factors
D000067528:Ataxins; C576499:Calb1 protein, mouse; D064092:Calbindin 1; D064026:Calbindins; D009419:Nerve Tissue Proteins; D012333:RNA, Messenger; D064030:S100 Calcium Binding Protein G