Growth-compromised HSV-2 vector Delta RR protects from N-methyl-D-aspartate-induced neuronal degeneration through redundant activation of the MEK/ERK and PI3-K/Akt survival pathways, either one of which overrides apoptotic cascades.
We have previously shown that intrastriatal injection of Delta RR, the growth-compromised herpes simplex virus type 2 (HSV-2) vector for the antiapoptotic protein ICP10PK, prevents apoptosis caused by the excitotoxin N-methyl-D-aspartate (NMDA) in a mouse model of glutamatergic neuronal cell death (Golembewski et al. [2007] Exp. Neurol. 203:381-393). Because apoptosis regulation is stimulus and cell type specific, our studies were designed to examine the mechanism of Delta RR-mediated neuroprotection in striatal neurons. Organotypic striatal cultures (OSC) that retain much of the synaptic circuitry of the intact striatum were infected with Delta RR or a growth-compromised HSV-2 vector that lacks ICP10PK (Delta PK) and examined for neuroprotection-associated signaling. The mutated ICP10 proteins (p175 and p95) were expressed in 70-80% of neurons from Delta RR- and Delta PK-infected cultures, respectively, as determined by double-immunofluorescent staining with antibodies to ICP10 and NeuN or GAD65. Delta RR- but not Delta PK-treated OSC were protected from NMDA-induced apoptosis, as verified by ethidium homodimer staining, TUNEL, caspase-3 activation, and poly(AD-ribose) polymerase (PARP) cleavage. Neuroprotection was through ICP10PK-mediated activation of the survival pathways MEK/ERK and PI3-K/Akt, up-regulation of the antiapoptotic proteins Bag-1 and Bcl-2, and phosphorylation (inactivation) of the proapoptotic protein Bad. It was blocked by the MEK inhibitor U0126 or the PI3-K inhibitor LY294002, suggesting that either pathway can prevent NMDA-induced apoptosis. The data indicate that Delta RR-delivered ICP10PK stimulates redundant survival pathways that override proapoptotic cascades. Delta RR is a promising gene therapy platform against glutamatergic cell death.
Laing
Jennifer M
JM
Department of Pharmacology and Experimental Therapeutics, University of Maryland School of Medicine, Baltimore, Maryland 21201-1559, USA.
Golembewski
Erin K
EK
Wales
Samantha Q
SQ
Liu
Juan
J
Jafri
M Samir
MS
Yarowsky
Paul J
PJ
Aurelian
Laure
L
D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D002470:Cell Survival / Q000502:physiology; D002522:Chlorocebus aethiops; D003342:Corpus Striatum / Q000473:pathology; D018690:Excitatory Amino Acid Agonists / Q000633:toxicity; D005455:Fluorescent Antibody Technique; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D018258:Herpesvirus 2, Human / Q000235:genetics; D015151:Immunoblotting; D020287:In Situ Nick-End Labeling; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D016202:N-Methylaspartate / Q000633:toxicity; D009410:Nerve Degeneration / Q000517:prevention & control*; D009474:Neurons / Q000473:pathology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012264:Ribonucleotide Reductases / Q000235:genetics / Q000502:physiology*; D014709:Vero Cells
D018690:Excitatory Amino Acid Agonists; D020928:Mitogen-Activated Protein Kinases; D016202:N-Methylaspartate; D019869:Phosphatidylinositol 3-Kinases; D017346:Protein Serine-Threonine Kinases; D012264:Ribonucleotide Reductases; C100671:ICP10 protein, herpes simplex virus type 2