The use of extubation readiness parameters: a survey of pediatric critical care physicians.

BACKGROUND: The pediatric literature addressing extubation readiness parameters and strategies to wean from mechanical ventilation is limited. METHODS: We designed a survey to assess the use of extubation readiness parameters among pediatric critical care physicians at academic centers in the United States. RESULTS: The overall response rate was 44.1% (417/945). The majority of respondents check for air leak and the amount of tracheal secretions. Fewer respondents use sedation score, the rapid shallow breathing index, or the airway-occlusion pressure 0.1 s after the start of inspiratory flow prior to extubation. The majority perform a spontaneous breathing trial with pressure support. The majority consider 30 cm H2O as the upper limit of an air leak test, and the need for endotracheal suctioning once every 2-4 hours as acceptable for extubation. In preparation for termination of mechanical ventilation the majority daily wean the ventilator rate and/or the pressure support instead of conducting a spontaneous breathing trial. CONCLUSIONS: Most pediatric critical care physicians reported assessing extubation readiness by checking air leak and suctioning need, and less often consider or perform sedation score or the rapid shallow breathing index.

Radiation exposure in extremely low birth weight infants during their neonatal intensive care unit stay.

BACKGROUND: Extremely low birth weight (ELBW <1000 g) infants may have increased sensitivity to radiation exposure. Our objective was to estimate the radiation exposure in survivors of ELBW infants during their neonatal intensive care unit (NICU) stay. METHODS: In this retrospective cohort study, medical records of all ELBW infants who had been admitted to our NICU between May 1999 and October 2009 were reviewed. The infants' total entrance skin exposure [ESE in micro-Gray (µGy)] was estimated. RESULTS: Among 450 survivors, the mean gestational age (GA) was 26.3±2.1 weeks, and the mean birth weight (BW) was 774.2±144.4 g. Infants received a median of 32 (range: 1-159) X-rays, with an estimated ESE of 1471 µGy (range: 28-9264). Total ESE was inversely proportional to GA (r=-0.34; P<0.01), and BW (r=-0.39; P=0.01) and proportional to the severity of illness [score for neonatal acute physiology-perinatal extension (SNAPPE), r=0.39; P=0.01]. In a linear regression analysis, GA, SNAPPE and necrotizing enterocolitis were associated with radiation exposure (ESE) in ELBW infants (r2=0.133; P<0.001). CONCLUSIONS: During their NICU stay, ELBW infants were subjected to a significant number of diagnostic X-ray procedures. Our data highlight the need to closely monitor the number of X-ray procedures ordered to ELBW infants to avoid unnecessary radiation exposure.

Genome-wide profiling of H3K56 acetylation and transcription factor binding sites in human adipocytes.

The growing epidemic of obesity and metabolic diseases calls for a better understanding of adipocyte biology. The regulation of transcription in adipocytes is particularly important, as it is a target for several therapeutic approaches. Transcriptional outcomes are influenced by both histone modifications and transcription factor binding. Although the epigenetic states and binding sites of several important transcription factors have been profiled in the mouse 3T3-L1 cell line, such data are lacking in human adipocytes. In this study, we identified H3K56 acetylation sites in human adipocytes derived from mesenchymal stem cells. H3K56 is acetylated by CBP and p300, and deacetylated by SIRT1, all are proteins with important roles in diabetes and insulin signaling. We found that while almost half of the genome shows signs of H3K56 acetylation, the highest level of H3K56 acetylation is associated with transcription factors and proteins in the adipokine signaling and Type II Diabetes pathways. In order to discover the transcription factors that recruit acetyltransferases and deacetylases to sites of H3K56 acetylation, we analyzed DNA sequences near H3K56 acetylated regions and found that the E2F recognition sequence was enriched. Using chromatin immunoprecipitation followed by high-throughput sequencing, we confirmed that genes bound by E2F4, as well as those by HSF-1 and C/EBPα, have higher than expected levels of H3K56 acetylation, and that the transcription factor binding sites and acetylation sites are often adjacent but rarely overlap. We also discovered a significant difference between bound targets of C/EBPα in 3T3-L1 and human adipocytes, highlighting the need to construct species-specific epigenetic and transcription factor binding site maps. This is the first genome-wide profile of H3K56 acetylation, E2F4, C/EBPα and HSF-1 binding in human adipocytes, and will serve as an important resource for better understanding adipocyte transcriptional regulation.

Phosphoproteomic characterization of PYK2 signaling pathways involved in osteogenesis.

The PYK2 tyrosine kinase is a negative regulator of bone formation, but aside from the requirement for PYK2 kinase activity there has been little progress toward understanding of the molecular mechanism involved in this function. To gain insight into the signaling pathways modulated by PYK2 we sought to identify PYK2 substrates. Challenges inherent to a quantitative phosphoproteomic analysis for non-receptor tyrosine kinases were overcome by employing an inducible PYK2 overexpression system in NIH3T3 cells in combination with a selective PYK2 inhibitor. The identification of a number of known PYK2 substrates and interacting partners validated the methodology. Results of the inducible cell system were extended to a cell model of osteogenesis, examining the effect of the PYK2 inhibitor on the phosphorylation state of targets identified in the phosphoproteomic study. Consistent with phosphoproteomic analysis, increased osteogenesis associated with a selective PYK2 inhibitor was accompanied by reduced phosphorylation of paxillin, Gab1 and p130(Cas), along with reduction of phosphorylation levels of the Met activation loop. These results further confirmed the utility of the methodology and point to a previously unknown bi-directional activation pathway between PYK2 and Met.

Sulfoximine-substituted trifluoromethylpyrimidine analogs as inhibitors of proline-rich tyrosine kinase 2 (PYK2) show reduced hERG activity.

The synthesis, in vitro properties, and in vivo pharmacokinetics for a series of sulfoximine-substituted trifluoromethylpyrimidines as inhibitors of proline-rich tyrosine kinase, a target for the possible treatment of osteoporosis, are described. These compounds were prepared as surrogates of the corresponding sulfone compound 1. Sulfone 1 was an attractive PYK2 lead compound; however, subsequent studies determined this compound possessed high dofetilide binding, which is an early indicator of cardiovascular safety. Surprisingly, the corresponding sulfoximine analogs displayed significantly lower dofetilide binding, which, for N-methylsulfoximine (S)-14a, translated to lower activity in a patch clamp hERG K(+) ion channel screen. In addition, compound (S)-14a shows good oral exposure in a rat pharmacokinetic model.

Proline-rich tyrosine kinase 2 regulates osteoprogenitor cells and bone formation, and offers an anabolic treatment approach for osteoporosis.

Bone is accrued and maintained primarily through the coupled actions of bone-forming osteoblasts and bone-resorbing osteoclasts. Cumulative in vitro studies indicated that proline-rich tyrosine kinase 2 (PYK2) is a positive mediator of osteoclast function and activity. However, our investigation of PYK2-/- mice did not reveal evidence supporting an essential function for PYK2 in osteoclasts either in vivo or in culture. We find that PYK2-/- mice have high bone mass resulting from an unexpected increase in bone formation. Consistent with the in vivo findings, mouse bone marrow cultures show that PYK2 deficiency enhances differentiation and activity of osteoprogenitor cells, as does expressing a PYK2-specific short hairpin RNA or dominantly interfering proteins in human mesenchymal stem cells. Furthermore, the daily administration of a small-molecule PYK2 inhibitor increases bone formation and protects against bone loss in ovariectomized rats, an established preclinical model of postmenopausal osteoporosis. In summary, we find that PYK2 regulates the differentiation of early osteoprogenitor cells across species and that inhibitors of the PYK2 have potential as a bone anabolic approach for the treatment of osteoporosis.