Brain malformations caused by retroviral vector-mediated gene transfer of hippocampal cholinergic neurostimulating peptide precursor protein into the CNS via embryonic mice ventricles.
Hippocampal cholinergic neurostimulating peptide precursor protein (HCNP-pp) is a unique multifunctional protein, being not only the precursor of HCNP, which promotes the phenotype development of septo-hippocampal cholinergic neurons, but also the binding protein of phosphatidylethanolamine, ATP, Raf-1 kinase (known as "Raf-1 kinase inhibitory factor" in peripheral organs), and serine protease. We obtained a high-titer retroviral vector harboring HCNP-pp cDNA by the use of a modified packaging cell line and centrifugation, and by injecting it into embryonic mouse ventricles, we investigated the function of its gene product within the central nervous system (CNS). We found that efficient transduction into hippocampal pyramidal neurons can be achieved by injecting the vector into embryonic brain ventricles on embryonic day 14 (E14). Three days after receiving the intraventricular injection of the high-titer HCNP-pp retrovirus vector on E14, the tissues around the ventricles showed an overexpression of HCNP-pp. This was accompanied by a reduced amount of activated MEK and Erk (as analyzed by histochemical and Western blot methods), suggesting that HCNP-pp also regulates the MAP-kinase cascade within the CNS. Surprisingly, mouse brains that received the HCNP-pp retroviral vector showed massive malformation of the hippocampus and cerebellum when examined 30 days after birth. This shows that strictly regulated HCNP-pp gene expression is necessary for the normal development of the mouse brain, and that the moderate overexpression achieved by retroviral vector-mediated gene transfer is sufficient to cause severe abnormality of entire brain structures.
Matsukawa
N
N
Department of Neurology and Neuroscience, Nagoya City University Graduate School of Medical Sciences, Mizuho-ku, Nagoya 467-8601, Japan.
Ikenaka
K
K
Nanmoku
K
K
Yuasa
H
H
Hattori
M
M
Kawano
M
M
Nakazawa
H
H
Fujimori
O
O
Ueda
R
R
Ojika
K
K
D000818:Animals; D015153:Blotting, Western; D001921:Brain / Q000473:pathology*; D002531:Cerebellum / Q000473:pathology* / Q000821:virology; D004622:Embryo, Mammalian; D022761:Gene Transfer, Horizontal; D006624:Hippocampus / Q000473:pathology* / Q000821:virology; D007150:Immunohistochemistry; D007276:Injections, Intraventricular; D048728:MAP Kinase Kinase Kinase 1; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009421:Nervous System Malformations / Q000209:etiology*; D009479:Neuropeptides / Q000235:genetics / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D017966:Pyramidal Cells / Q000473:pathology / Q000821:virology; D012190:Retroviridae; D014161:Transduction, Genetic / Q000379:methods
D009479:Neuropeptides; C075274:hippocampal cholinergic neurostimulating peptide; D017346:Protein Serine-Threonine Kinases; D020928:Mitogen-Activated Protein Kinases; D048728:MAP Kinase Kinase Kinase 1; C482578:Map3k1 protein, mouse