Heat shock protein 60 or 70 activates nitric-oxide synthase (NOS) I- and inhibits NOS II-associated signaling and depresses the mitochondrial apoptotic cascade during brain stem death.
The cellular and molecular basis of brain stem death remains an enigma. As the origin of a "life-and-death" signal that reflects the progression toward brain stem death, the rostral ventrolateral medulla (RVLM) is a suitable neural substrate for mechanistic delineation of this phenomenon. Here, we evaluated the hypothesis that heat shock proteins (HSPs) play a neuroprotective role in the RVLM during brain stem death and delineated the underlying mechanisms, using a clinically relevant animal model that employed the organophosphate pesticide mevinphos (Mev) as the experimental insult. In Sprague-Dawley rats, proteomic, Western blot, and real-time PCR analyses demonstrated that Mev induced de novo synthesis of HSP60 or HSP70 in the RVLM without affecting HSP90 level. Loss-of-function manipulations of HSP60 or HSP70 in the RVLM using anti-serum or antisense oligonucleotide potentiated Mev-elicited cardiovascular depression alongside reduced nitric-oxide synthase (NOS) I/protein kinase G signaling, enhanced NOS II/peroxynitrite cascade, intensified nucleosomal DNA fragmentation, elevated cytoplasmic histone-associated DNA fragments or activated caspase-3, and augmented the cytochrome c/caspase-3 cascade of apoptotic signaling in the RVLM. Co-immunoprecipitation experiments further revealed a progressive increase in the complex formed between HSP60 and mitochondrial or cytosolic Bax or mitochondrial Bcl-2 during Mev intoxication, alongside a dissociation of the cytosolic HSP60-Bcl-2 complex. We conclude that HSP60 and HSP70 confer neuroprotection against Mev intoxication by ameliorating cardiovascular depression via an anti-apoptotic action in the RVLM. The possible underlying intracellular processes include enhancing NOS I/protein kinase G signaling and inhibiting the NOS II/peroxynitrite cascade. In addition, HSP60 exerts its effects against apoptosis by blunting Mev-induced activation of the Bax/cytochrome c/caspase-3 cascade.
Chan
Julie Y H
JYH
Department of Medical Education and Research, Kaohsiung Veterans General Hospital, Kaohsiung 81346.
Cheng
Hsiao-Lei
HL
Center for Neuroscience, National Sun Yat-sen University, Kaohsiung 80424, and the.
Chou
Jimmy L J
JLJ
Center for Neuroscience, National Sun Yat-sen University, Kaohsiung 80424, and the.
Li
Faith C H
FCH
Center for Neuroscience, National Sun Yat-sen University, Kaohsiung 80424, and the.
Dai
Kuang-Yu
KY
Center for Neuroscience, National Sun Yat-sen University, Kaohsiung 80424, and the.
Chan
Samuel H H
SHH
Center for Neuroscience, National Sun Yat-sen University, Kaohsiung 80424, and the; Center for Gene Regulation and Signal Transduction Research, National Cheng Kung University, Tainan 70101, Taiwan.
Chang
Alice Y W
AYW
Center for Neuroscience, National Sun Yat-sen University, Kaohsiung 80424, and the; Center for Gene Regulation and Signal Transduction Research, National Cheng Kung University, Tainan 70101, Taiwan. Electronic address: achang@mail.nsysu.edu.tw.
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001926:Brain Death / Q000378:metabolism* / Q000473:pathology; D001933:Brain Stem / Q000201:enzymology* / Q000473:pathology; D053148:Caspase 3 / Q000378:metabolism; D018834:Chaperonin 60 / Q000096:biosynthesis*; D017869:Cyclic GMP-Dependent Protein Kinases / Q000378:metabolism; D045304:Cytochromes c / Q000378:metabolism; D003593:Cytoplasm / Q000201:enzymology; D053938:DNA Fragmentation / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D018840:HSP70 Heat-Shock Proteins / Q000096:biosynthesis*; D007306:Insecticides / Q000633:toxicity; D008297:Male; D008799:Mevinphos / Q000633:toxicity; D008928:Mitochondria / Q000201:enzymology; D019001:Nitric Oxide Synthase / Q000378:metabolism*; D052248:Nitric Oxide Synthase Type I; D052247:Nitric Oxide Synthase Type II / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000187:drug effects; D051028:bcl-2-Associated X Protein / Q000378:metabolism
D018834:Chaperonin 60; D018840:HSP70 Heat-Shock Proteins; D007306:Insecticides; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D008799:Mevinphos; D045304:Cytochromes c; D019001:Nitric Oxide Synthase; D052248:Nitric Oxide Synthase Type I; D052247:Nitric Oxide Synthase Type II; C496322:Nos1 protein, rat; C496318:Nos2 protein, rat; D017869:Cyclic GMP-Dependent Protein Kinases; D053148:Caspase 3