Abstract
BackgroundBrain stem cardiovascular regulatory dysfunction during brain death is underpinned by an upregulation of nitric oxide synthase II (NOS II) in rostral ventrolateral medulla (RVLM), the origin of a life-and-death signal detected from blood pressure of comatose patients that disappears before brain death ensues. Furthermore, the ubiquitin-proteasome system (UPS) may be involved in the synthesis and degradation of NOS II. We assessed the hypothesis that the UPS participates in brain stem cardiovascular regulation during brain death by engaging in both synthesis and degradation of NOS II in RVLM.Methodology/Principal FindingsIn a clinically relevant experimental model of brain death using Sprague-Dawley rats, pretreatment by microinjection into the bilateral RVLM of proteasome inhibitors (lactacystin or proteasome inhibitor II) antagonized the hypotension and reduction in the life-and-death signal elicited by intravenous administration of Escherichia coli lipopolysaccharide (LPS). On the other hand, pretreatment with an inhibitor of ubiquitin-recycling (ubiquitin aldehyde) or ubiquitin C-terminal hydrolase isozyme L1 (UCH-L1) potentiated the elicited hypotension and blunted the prevalence of the life-and-death signal. Real-time polymerase chain reaction, Western blot, electrophoresis mobility shift assay, chromatin immunoprecipitation and co-immunoprecipitation experiments further showed that the proteasome inhibitors antagonized the augmented nuclear presence of NF-κB or binding between NF-κB and nos II promoter and blunted the reduced cytosolic presence of phosphorylated IκB. The already impeded NOS II protein expression by proteasome inhibitor II was further reduced after gene-knockdown of NF-κB in RVLM. In animals pretreated with UCH-L1 inhibitor and died before significant increase in nos II mRNA occurred, NOS II protein expression in RVLM was considerably elevated.Conclusions/SignificanceWe conclude that UPS participates in the defunct and maintained brain stem cardiovascular regulation during experimental brain death by engaging in both synthesis and degradation of NOS II at RVLM. Our results provide information on new therapeutic initiatives against this fatal eventuality.
Highlights
Brain death, the legal definition of death in many countries [1,2,3], is commonly recognized as a neurological phenomenon
Based on an endotoxemia model of experimental brain death, our results revealed a novel, causal and double-edged sword role for the ubiquitin-proteasome system (UPS) at rostral ventrolateral medulla (RVLM) in this process (Figure 8A)
We demonstrated that a crucial determinant in the tug-of-war between maintained and defunct brain stem circulatory regulation during brain death resides in the temporal balance between the continuous degradation of nitric oxide synthase II (NOS II) and the progressively augmented synthesis of the isozyme in RVLM (Figure 8B)
Summary
The legal definition of death in many countries [1,2,3], is commonly recognized as a neurological phenomenon. The power density of the LF component, which mirrors the prevalence of baroreflex-mediated sympathetic neurogenic vasomotor tone [5], undergoes a dramatic reduction or loss before brain death ensues in comatose patients [6,7,8] to reflect irreversible failure of brain stem cardiovascular regulatory functions [9]. It follows that delineation of the mechanisms that underpin the shift between maintained and defunct brain stem cardiovascular regulatory machinery during the progression towards brain death should shed further light on this fatal phenomenon.
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