Hypoxia-induced caspase-3 activation and DNA fragmentation in cortical neurons of newborn piglets: Role of nitric oxide

被引:30
作者
Parikh, NA
Katsetos, CD
Ashraf, QM
Haider, SH
Legido, A
Delivoria-Papadopoulos, M
Mishra, OP
机构
[1] Univ Texas, Sch Med, Dept Pediat, Div Neonatol, Houston, TX 77030 USA
[2] Thomas Jefferson Univ Hosp, Dept Pediat, Jefferson Med Coll, Philadelphia, PA 19107 USA
[3] Drexel Univ, Dept Pediat, Coll Med, Philadelphia, PA 19104 USA
[4] St Christophers Hosp Children, Dept Pathol & Lab Med, Philadelphia, PA 19133 USA
关键词
caspase-3; hypoxia; immunoreactivity; labeling index; neuronal nitric oxide synthase;
D O I
10.1023/A:1024992214886
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Hypoxia results in generation of nitric oxide (NO) free radicals, activation of caspase-3, and genomic DNA fragmentation. The present study tests the hypothesis that hypoxia-induced caspase-3 activation and DNA fragmentation are nitric oxide mediated. Studies were conducted in newborn piglets, divided into normoxic (n = 5), hypoxic (n = 5), and hypoxic-7-NINA (n = 6). Hypoxic-7-NINA group received the neuronal nitric oxide synthase inhibitor, 7-Nitroindazole (7-NINA). Caspase-3 activity was determined spectrofluorometrically using enzyme-specific substrates. Sections from the neocortex were stained with an antiserum recognizing active caspase-3. Purified DNA was separated by gel electrophoresis. Administration of 7-NINA resulted in decreased immunoreactivity of caspase-3 (mean LI: 20.2%) as compared to the untreated hypoxia group (mean LI: 57.5%) (P < 0.05). 7-NINA attenuated caspase-3 enzymatic activity as well in comparison to the untreated hypoxia group (P < 0.05). Furthermore, multiple low molecular weight bands corresponding to DNA fragments were present in the hypoxic but not in the normoxic or hypoxic-7-NINA groups. Inhibition of nNOS abates the hypoxia-induced increase in active caspase-3 immunoreactivity, as well as enzymatic activity in cortical neurons, and DNA fragmentation in brain homogenates. We conclude that the coordinate increase of capase-3 activity and fragmentation of nuclear DNA in the hypoxic newborn piglet brain are NO mediated.
引用
收藏
页码:1351 / 1357
页数:7
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