Pharmacological and genetic inhibition of NADPH oxidase does not reduce brain damage in different models of perinatal brain injury in newborn mice.
BACKGROUND
Inflammation and reactive oxygen species (ROS) are important in the development of perinatal brain injury. The ROS-generating enzyme NADPH oxidase (Nox2) is present in inflammatory cells and contributes to brain injury in adult animal models.


OBJECTIVE
NADPH oxidase contributes to ROS formation and development of injury in the immature brain and inhibition of NADPH oxidase attenuates perinatal brain injury.


METHODS
We used animal models of term hypoxia-ischemia (HI) (P9 mice) as well as ibotenate-induced excitotoxic injury (P5 mice) mimicking features of periventricular leukomalacia in preterm infants. In vitro microglia cell cultures were used to investigate NADPH oxidase-dependent ROS formation. In vivo we determined the impact 1) of HI on NADPH oxidase gene expression 2) of genetic (gp91-phox/Nox2 knock-out) and 3) of pharmacological NADPH oxidase inhibition on HI-induced injury and NMDA receptor-mediated excitotoxic injury, respectively. Endpoints were ROS formation, oxidative stress, apoptosis, inflammation and extent of injury.


RESULTS
Hypoxia-ischemia increased NADPH oxidase subunits mRNA expression in total brain tissue in vivo. In vitro ibotenate increased NADPH oxidase-dependent formation of reactive oxygen species in microglia. In vivo the inhibition of NADPH oxidase did not reduce the extent of brain injury in any of the animal models. In contrast, the injury was increased by inhibition of NADPH oxidase and genetic inhibition was associated with an increased level of galectin-3 and IL-1beta.


CONCLUSIONS
NADPH oxidase is upregulated after hypoxia-ischemia and activated microglia cells are a possible source of Nox2-derived ROS. In contrast to findings in adult brain, NADPH oxidase does not significantly contribute to the pathogenesis of perinatal brain injury. Results obtained in adult animals cannot be transferred to newborns and inhibition of NADPH oxidase should not be used in attempts to attenuate injury.
Doverhag
Christina
C
Department of Physiology and Neuroscience, Perinatal Center, Sahlgrenska Academy, Göteborg University, Sweden.
Keller
Matthias
M
Karlsson
Anna
A
Hedtjarn
Maj
M
Nilsson
Ulf
U
Kapeller
Edith
E
Sarkozy
Gergely
G
Klimaschewski
Lars
L
Humpel
Christian
C
Hagberg
Henrik
H
Simbruner
Georg
G
Gressens
Pierre
P
Savman
Karin
K
D000818:Animals; D000831:Animals, Newborn; D017209:Apoptosis / Q000502:physiology; D001930:Brain Injuries / Q000201:enzymology* / Q000209:etiology / Q000473:pathology; D018690:Excitatory Amino Acid Agonists / Q000633:toxicity; D005260:Female; D015870:Gene Expression; D020925:Hypoxia-Ischemia, Brain / Q000150:complications / Q000201:enzymology* / Q000473:pathology; D007051:Ibotenic Acid / Q000633:toxicity; D007150:Immunohistochemistry; D007249:Inflammation / Q000378:metabolism / Q000473:pathology; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D017628:Microglia / Q000378:metabolism; D019255:NADPH Oxidases / Q000235:genetics / Q000378:metabolism*; D018384:Oxidative Stress / Q000502:physiology; D012333:RNA, Messenger / Q000032:analysis; D017382:Reactive Oxygen Species / Q000378:metabolism*
D018690:Excitatory Amino Acid Agonists; D012333:RNA, Messenger; D017382:Reactive Oxygen Species; D007051:Ibotenic Acid; D019255:NADPH Oxidases