The Extracellular RNA Communication Consortium (ERCC) was launched to accelerate progress in the new field of extracellular RNA (exRNA) biology and to establish whether exRNAs and their carriers, ...including extracellular vesicles (EVs), can mediate intercellular communication and be utilized for clinical applications. Phase 1 of the ERCC focused on exRNA/EV biogenesis and function, discovery of exRNA biomarkers, development of exRNA/EV-based therapeutics, and construction of a robust set of reference exRNA profiles for a variety of biofluids. Here, we present progress by ERCC investigators in these areas, and we discuss collaborative projects directed at development of robust methods for EV/exRNA isolation and analysis and tools for sharing and computational analysis of exRNA profiling data.
The Extracellular RNA Communication Consortium (ERCC) presents progress toward understanding the biology of extracellular RNAs and their use as biomarkers and therapeutics.
Conventional genomic DNA (gDNA) extraction methods can take hours to complete, may require fume hoods and represent the most time-consuming step in many gDNA-based molecular assays. We systematically ...optimized a bead bashing-based (BBB) approach for rapid gDNA extraction without the need for a fume hood. Human tissue specimens (n = 34) subjected to the 12-min BBB method yielded 0.40 ± 0.17 (mean ± SD) μg of gDNA per milligram of tissue, sufficient for many downstream applications, and 3- and 6-min extensions resulted in an additional 0.43 ± 0.23 μg and 0.48 ± 0.43 μg per milligram of tissue, respectively. The BBB method provides a simple and rapid method for gDNA extraction from mammalian tissue that is applicable to time-sensitive clinical applications.
Extraction of genomic DNA is a critical and often rate-limiting step for many downstream genetic tests. However, existing methods can be time consuming, difficult to automate and require use of a fume hood. Here, we present a simple 12-min method for genomic DNA extraction from tissue without the need for a fume hood and which can be readily automated for high-throughput applications.
Ovarian hyperstimulation syndrome is a potentially life-threatening clinical condition.
The objective of this study was to evaluate risk factors for life-threatening complications for patients with ...severe ovarian hyperstimulation syndrome in a United States nationwide sample.
Ovarian hyperstimulation syndrome admissions from 2002 to 2011 from the Nationwide Inpatient Sample were included in this study. The association between patient and hospital factors and life-threatening complications (deep vein thrombosis/pulmonary embolism, acute respiratory distress syndrome, acute renal failure, intubation), nonroutine discharge (discharge to skilled nursing facility, transfer hospital), prolonged length of stay, and total hospital charges were analyzed. Survey-adjusted multivariable logistic regression analyses were performed for these outcomes, controlling for risk factors, with adjusted odds ratios with 95% confidence intervals as the measures of effect.
A total of 11,562 patients were hospitalized with severe ovarian hyperstimulation syndrome from 2002 to 2011. The majority were white (55.7%), with private insurance (87.7%), aged 25–39 years (84.6%), and hospitalized in an urban location (95%). In all, 19.3% of patients had medical comorbidities including hypertension, diabetes, obesity, hypothyroidism, and anemia. Life-threatening complications occurred in 4.4% of patients (deep vein thrombosis/pulmonary embolism, 2.2%; acute renal failure; acute respiratory distress syndrome, 0.9%; intubation, 0.5%). Patients ≥40 years old (odds ratio, 4.02; 95% confidence interval, 1.37, 11.76), those with comorbidities (odds ratio, 2.29; 95% confidence interval, 1.46, 3.57), and African American patients (odds ratio, 2.15; 95% confidence interval, 1.25, 3.70) were more likely to develop life-threatening conditions. Patients with medical comorbidities (odds ratio, 0.39; 95% confidence interval, 0.24, 0.63) were also less likely to be routinely discharged from the hospital. Adjusting for patient and hospital demographics, patients with comorbidities were more likely to develop deep vein thrombosis/pulmonary embolism (adjusted odds ratio, 2.44; 95% confidence interval, 1.28, 4.65) and acute renal failure (adjusted odds ratio, 2.26; 95% confidence interval, 1.21, 4.23). Patients who developed life-threatening complications had longer hospital length of stay (adjusted odds ratio, 3.72; 95% confidence interval, 2.28, 6.07) and higher hospital costs (adjusted odds ratio, 5.20; 95% confidence interval, 3.22,8.39).
Patients with common medical comorbidities are at higher risk for life-threatening complications in the setting of severe ovarian hyperstimulation syndrome. Furthermore, these complications are associated with high hospital costs and hospital burden. Given the increasing number of in vitro fertilization patients with medical comorbidities, closer monitoring of at-risk patients may be indicated. As assisted reproductive technology practice changes in recent years with strategies designed to reduce ovarian hyperstimulation syndrome risk, future studies are needed to assess the impact of these changes on hospitalization and complication risk.
Despite the central role of the placenta in supporting a pregnancy, relatively little is known about transcriptomic and immune-cell changes that occur across gestation. To generate a reference gene ...expression map of first (T1), second (T2) and third (T3) trimester human placenta, and assess differences in transcriptome in maternal versus fetal side tissues sections of full-term placenta, we performed RNA-Seq analysis on 64 biopsy samples from 18 placentas across all three gestations. We identified 1120 differentially expressed genes in placenta tissues from T1 and T3 samples using a generalized linear model within DESeq2. In total, 411 and 709 genes were positively associated with T1 and T3 placenta, respectively. Comparison of the top 200 differentially expressed genes in the T1 placenta with T3 showed that most of the top enriched biological processes were related to cell division and proliferation. T1 and T2 tissues shared expression of fibroblast-specific COL6A2, HGF, and SPP1 genes. In T3 samples, the expression of genes relating to vasculature development and regulation were highly enriched. Monocytes and NK cell population increased in T3 compared to T1 and T2, whereas Hofbauer cell proportion expanded significantly in T2 and then decreased in T3 samples. There were no significant gene expression differences in the maternal vs. fetal side in T3 placentas. Gene expression patterns shift temporally across trimesters but not spatially across the placenta, at least at the resolution of the biopsy samples. The genes and gene set we identified here represent a valuable resource for studying pathology in pregnancy-related disorders.
•A comprehensive transcriptome is presented of human placenta across three trimesters.•1120 genes were differentially expressed in first (T1) and third (T3) trimester placenta samples.•T1: Enrichment of cell division and proliferation processes; T3: Vasculature development and regulation processes upregulated.•No significant gene expression differences in the maternal vs. fetal side in T3 placentas.
The mammalian heart, the body's largest energy consumer, has evolved robust mechanisms to tightly couple fuel supply with energy demand across a wide range of physiologic and pathophysiologic states, ...yet, when compared with other organs, relatively little is known about the molecular machinery that directly governs metabolic plasticity in the heart. Although previous studies have defined Kruppel-like factor 15 (KLF15) as a transcriptional repressor of pathologic cardiac hypertrophy, a direct role for the KLF family in cardiac metabolism has not been previously established. We show in human heart samples that KLF15 is induced after birth and reduced in heart failure, a myocardial expression pattern that parallels reliance on lipid oxidation. Isolated working heart studies and unbiased transcriptomic profiling in Klf15-deficient hearts demonstrate that KLF15 is an essential regulator of lipid flux and metabolic homeostasis in the adult myocardium. An important mechanism by which KLF15 regulates its direct transcriptional targets is via interaction with p300 and recruitment of this critical co-activator to promoters. This study establishes KLF15 as a key regulator of myocardial lipid utilization and is the first to implicate the KLF transcription factor family in cardiac metabolism.
Metabolic homeostasis is central to normal cardiac function. The molecular mechanisms underlying metabolic plasticity in the heart remain poorly understood.
Kruppel-like factor 15 (KLF15) is a direct and independent regulator of myocardial lipid flux.
KLF15 is a core component of the transcriptional circuitry that governs cardiac metabolism.
This work is the first to implicate the KLF transcription factor family in cardiac metabolism.
Abstract
Background
Extracellular RNAs (exRNAs) in biofluids are amenable to quantitative analysis and proposed as noninvasive biomarkers for monitoring organ function. Cell-lineage-specific ...microRNAs (miRNAs) are present in plasma as soluble ribonucleoproteins or enclosed in exRNA carriers and transported through the vasculature. However, more extensive studies of healthy individuals are needed to gain insights into the variability of plasma miRNA abundance and composition.
Methods
The exRNA composition of platelet-depleted plasma collected twice from 236 healthy individuals was characterized by small RNA sequencing. Plasma of pregnant women featuring dramatically increased placental miRNAs and samples from subject P12 with noticeably increased epithelial- and neuroendocrine-origin miRNAs were included for comparison. The miRNA content of 10 000g and 100 000g pellet fractions of plasma generated by ultracentrifugation was also determined. Data analysis methods included Pearson correlation, differential gene expression, and unsupervised clustering.
Results
The abundance changes for more variable miRNAs in plasma of normal individuals correlated between coexpressed cell-lineage-specific miRNAs of the liver, neuroendocrine organs, epithelial cells, and muscle. ExRNA of pellet fractions contained <2% of total plasma miRNA with modest enrichment of lineage-specific and variable miRNAs compared to supernatant. The abundance fold changes of miRNAs observed in pregnancy and P12 compared to normal exceeded interquartile variability of healthy individuals. The neuroendocrine miRNA signature of P12 persisted for more than 4 years and was absent in other individuals.
Conclusions
This study defines the framework and effect size for screening of extensive plasma collections for miRNA phenotypes and biomarker discovery.
To determine whether a highly visible device that clearly indicates whether the head-of-bed is adequately elevated would increase rates of compliance with head-of-bed elevation guidelines.
A ...prospective, single-center, multi-unit, two-phase study.
Surgical, thoracic, trauma, and medical intensive care units.
Cohort of intubated patients.
A 4-wk trial was performed. At the onset of the trial, nurses were reminded to maintain head-of-bed elevation > 30 degrees. Over the subsequent 2 wks, head-of-bed elevations of intubated patient beds were measured. An Angle Indicator, designed to clearly display whether the head-of-bed was adequately elevated, was then placed on side rails of beds of ventilated patients, and head-of-bed elevation measurements were taken for an additional 2 wks. A survey was then handed out to nursing staff to assess satisfaction with the device.
A total of 268 bed measurements were made. The average head-of-bed elevation was 21.8 degrees on beds without the device (n = 166) and 30.9 degrees on beds with the device (n = 102; p < .005). When compliance is defined as a bed angle of > or = 28 degrees, 23% of beds without the device were compliant while 71.5% of the beds with the device were compliant. The relative risk and odds ratio of having the device on a compliant bed were 2.2 and 9.25, respectively (p < .005). Seventy-two percent of nurses surveyed (n = 32) found it to be an improvement over existing methods, 88% found it helpful, and 84% would like it routinely used.
The Angle Indicator improved rates of adherence to bed-elevation guidelines, and hospital staff found it helpful.
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