ABSTRACT
BACKGROUND: The Compensatory Reserve Metric (CRM) provides a time sensitive indicator of hemodynamic decompensation. However, its in-field utility is limited due to the size and cost-intensive nature of standard vital sign monitors or photoplethysmographic volume-clamp (PPGVC) devices used to measure arterial waveforms. In this regard, photoplethysmographic measurements obtained from pulse oximetry (PPGPO) may serve as a useful, portable alternative. This study aimed to validate CRM values obtained using PPGPO. METHODS: Forty-nine healthy adults (25 females) underwent a graded lower body negative pressure (LBNP) protocol to simulate hemorrhage. Arterial waveforms were sampled using PPGPO and PPGVC. The CRM was calculated using a one-dimensional convolutional neural network. Cardiac output and stroke volume were measured using PPGVC. A brachial artery catheter was used to measure intraarterial pressure. A 3-lead ECG was used to measure heart rate. Fixed-effect linear mixed models with repeated measures were used to examine the association between CRM values and physiologic variables. Log-rank analyses were used to examine differences in shock determination during LBNP between monitored hemodynamic parameters. RESULTS: The median LBNP stage reached was 70 mmHg (Range: 45-100 mmHg). Relative to baseline, at tolerance there was a 47±12% reduction in stroke volume, 64±27% increase in heart rate, and 21±7% reduction in systolic blood pressure (P<0.001 for all). CRM values obtained with both PPGPO and PPGVC were associated with changes in heart rate (P<0.001), stroke volume (P<0.001), and pulse pressure (P<0.001). Furthermore, they provided an earlier detection of hemodynamic shock relative to the traditional metrics of shock index (P<0.001 for both), systolic blood pressure (P<0.001 for both), and heart rate (P=0.001 for both). CONCLUSION: The CRM obtained from PPGPO provides a valid, time-sensitized prediction of hemodynamic decompensation, opening the door to provide military medical personnel noninvasive in-field advanced capability for early detection of hemorrhage and imminent onset of shock. LEVEL OF EVIDENCE: Diagnostic Tests or Criteria, Level IV.
ABSTRACT
Pumilio is a sequence-specific RNA-binding protein that controls development, stem cell fate, and neurological functions in Drosophila. Pumilio represses protein expression by destabilizing target mRNAs in a manner dependent on the CCR4-NOT deadenylase complex. Three unique repression domains in the N-terminal region of Pumilio were previously shown to recruit CCR4-NOT, but how they do so was not well understood. In this study, we identified the motifs that are necessary and sufficient for the activity of the third repression domain of Pumilio, designated RD3, which is present in all isoforms and has conserved regulatory function. We identified multiple conserved regions of RD3 that are important for repression activity in cell-based reporter gene assays. Using yeast two-hybrid assays, we show that RD3 contacts specific regions of the Not1, Not2, and Not3 subunits of the CCR4-NOT complex. Our results indicate that RD3 makes multivalent interactions with CCR4-NOT mediated by conserved short linear interaction motifs. Specifically, two phenylalanine residues in RD3 make crucial contacts with Not1 that are essential for its repression activity. Using reporter gene assays, we also identify three new target mRNAs that are repressed by Pumilio and show that RD3 contributes to their regulation. Together, these results provide important insights into the mechanism by which Pumilio recruits CCR4-NOT to regulate the expression of target mRNAs.
