A multi-omic analysis of human naïve CD4+ T cells.

BACKGROUND: Cellular function and diversity are orchestrated by complex interactions of fundamental biomolecules including DNA, RNA and proteins. Technological advances in genomics, epigenomics, transcriptomics and proteomics have enabled massively parallel and unbiased measurements. Such high-throughput technologies have been extensively used to carry out broad, unbiased studies, particularly in the context of human diseases. Nevertheless, a unified analysis of the genome, epigenome, transcriptome and proteome of a single human cell type to obtain a coherent view of the complex interplay between various biomolecules has not yet been undertaken. Here, we report the first multi-omic analysis of human primary naïve CD4+ T cells isolated from a single individual. RESULTS: Integrating multi-omics datasets allowed us to investigate genome-wide methylation and its effect on mRNA/protein expression patterns, extent of RNA editing under normal physiological conditions and allele specific expression in naïve CD4+ T cells. In addition, we carried out a multi-omic comparative analysis of naïve with primary resting memory CD4+ T cells to identify molecular changes underlying T cell differentiation. This analysis provided mechanistic insights into how several molecules involved in T cell receptor signaling are regulated at the DNA, RNA and protein levels. Phosphoproteomics revealed downstream signaling events that regulate these two cellular states. Availability of multi-omics data from an identical genetic background also allowed us to employ novel proteogenomics approaches to identify individual-specific variants and putative novel protein coding regions in the human genome. CONCLUSIONS: We utilized multiple high-throughput technologies to derive a comprehensive profile of two primary human cell types, naïve CD4+ T cells and memory CD4+ T cells, from a single donor. Through vertical as well as horizontal integration of whole genome sequencing, methylation arrays, RNA-Seq, miRNA-Seq, proteomics, and phosphoproteomics, we derived an integrated and comparative map of these two closely related immune cells and identified potential molecular effectors of immune cell differentiation following antigen encounter.

Outcomes Associated With Concurrent Iris-Sutured Intraocular Lens Placement and Subluxated Crystalline Lens Extraction.

IMPORTANCE: We have developed a novel surgical technique, to our knowledge, for the management of subluxated crystalline lenses involving preplacement of an iris-sutured posterior chamber intraocular lens (PCIOL) before pars plana vitrectomy and lensectomy. OBJECTIVE: To investigate the outcomes of eyes with subluxated crystalline lenses, predominantly a result of Marfan syndrome (14 eyes [58%]) or trauma (5 eyes [21%]), that underwent pars plana vitrectomy and lensectomy with placement of an iris-sutured PCIOL. DESIGN, SETTING, AND PARTICIPANTS: We performed a retrospective, noncomparative case series of 24 eyes from 17 consecutive adult patients with surgically treated subluxated crystalline lenses presenting to the Wilmer Eye Institute at Johns Hopkins Hospital from October 6, 2006, through May 1, 2013. The mean (SD) postoperative follow-up was 24.4 (20.5) months for eyes with at least 6 months of follow-up (last date, October 13, 2014). We performed the analysis from January 21, 2014, through January 3, 2015. MAIN OUTCOMES AND MEASURES: Improvement in best-corrected visual acuity using an automated Snellen chart and induction of astigmatism for eyes with at least 6 months of follow-up (n = 18) and IOL stability during follow-up for all eyes (n = 24). RESULTS: The mean (SD) age at surgery was 49.4 (10.7 [range, 29-67]) years. We found an improvement in mean (SD [95% CI]) best-corrected visual acuity from 0.66 (0.71 [0.30-1.02]) logMAR preoperatively (Snellen equivalent, approximately 20/90; range, 20/30 to hand motions) to 0.07 (0.11 [95% CI, 0.01-0.12]) logMAR postoperatively (Snellen equivalent, approximately 20/23; range, 20/15 to 20/50). We found little change in astigmatism postoperatively (mean change, -0.1 [95% CI, -0.5 to 0.13] diopters). Postoperative complications included retinal detachment (1 eye [4%]), retained cortical fragment (1 [4%]), cystoid macular edema (2 [8%]), and IOL subluxation (3 [13%]) owing to haptic slippage within 3 months of the procedure. The overall probability of successfully achieving placement of a centered iris-sutured PCIOL in patients with follow-up of longer than 6 months (n = 18) was 100% (95% CI, 81.5%-100%). CONCLUSIONS AND RELEVANCE: Placement of iris-sutured PCIOLs at the time of subluxated lens extraction with a pars plana surgical approach yields favorable results in terms of postoperative visual outcomes and surgical complications. This technique offers an effective procedure for surgeons to use when treating clinically significant subluxated crystalline lenses.

Dexmedetomidine preserves attention/calculation when used for cooperative and short-term intensive care unit sedation.

PURPOSE: Differential effects on cognition were recently demonstrated between dexmedetomidine (DEX) and propofol (PRO) when used for cooperative sedation. Propofol was found to reduce cognition, whereas DEX improved cognition. To further discriminate these effects, we evaluated the effect of PRO vs DEX in selected areas of cognition. METHODS: This is a post hoc analysis of the Acute Neurologic Intensive Care Unit Sedation Trial and an investigator-initiated, prospective, randomized, double-blinded, crossover study, comparing the effect of PRO and DEX on cognition measure by the Johns Hopkins Adapted Cognitive Exam (ACE). A linear model analysis accounting for within-patient correlation of measures was used to estimate differences in ACE subscales between drugs. RESULTS: Propofol diminished adjusted scores on all ACE subscales (P < .05), whereas DEX improved adjusted scores selectively for attention/calculation (3.55; 95% confidence interval, 1.49-5.61; P < .01). The positive and significant difference in ACE scores between agents was present across subscales. CONCLUSIONS: Our findings indicate that DEX improved ACE attention/calculation subscale in awake patients receiving cooperative sedation. This is in contrast to the deterioration in all mean ACE subscale scores observed using PRO, suggesting DEX preserved cognitive function with specific preservation of focus and attention and allows for greater cognition compared with PRO across all cognitive domains.

Improving immunization delivery using an electronic health record: the ImmProve project.

OBJECTIVE: Though an essential pediatric preventive service, immunizations are challenging to deliver reliably. Our objective was to measure the impact on pediatric immunization rates of providing clinicians with electronic health record-derived immunization prompting. METHODS: Operating in a large, urban, hospital-based pediatric primary care clinic, we evaluated 2 interventions to improve immunization delivery to children ages 2, 6, and 13 years: point-of-care, patient-specific electronic clinical decision support (CDS) when children overdue for immunizations presented for care, and provider-specific bulletins listing children overdue for immunizations. RESULTS: Overall, the proportion of children up to date for a composite of recommended immunizations at ages 2, 6, and 13 years was not different in the intervention (CDS active) and historical control (CDS not active) periods; historical immunization rates were high. The proportion of children receiving 2 doses of hepatitis A immunization before their second birthday was significantly improved during the intervention period. Human papillomavirus (HPV) immunization delivery was low during both control and intervention periods and was unchanged for 13-year-olds. For 14-year-olds, however, 4 of the 5 highest quarterly rates of complete HPV immunization occurred in the final year of the intervention. Provider-specific bulletins listing children overdue for immunizations increased the likelihood of identified children receiving catch-up hepatitis A immunizations (hazard ratio 1.32; 95% confidence interval 1.12-1.56); results for HPV and the composite of recommended immunizations were of a similar magnitude but not statistically significant. CONCLUSIONS: In our patient population, with high baseline uptake of recommended immunizations, electronic health record-derived immunization prompting had a limited effect on immunization delivery. Benefit was more clearly demonstrated for newer immunizations with lower baseline uptake.

Assessing affymetrix GeneChip microarray quality.

BACKGROUND: Microarray technology has become a widely used tool in the biological sciences. Over the past decade, the number of users has grown exponentially, and with the number of applications and secondary data analyses rapidly increasing, we expect this rate to continue. Various initiatives such as the External RNA Control Consortium (ERCC) and the MicroArray Quality Control (MAQC) project have explored ways to provide standards for the technology. For microarrays to become generally accepted as a reliable technology, statistical methods for assessing quality will be an indispensable component; however, there remains a lack of consensus in both defining and measuring microarray quality. RESULTS: We begin by providing a precise definition of microarray quality and reviewing existing Affymetrix GeneChip quality metrics in light of this definition. We show that the best-performing metrics require multiple arrays to be assessed simultaneously. While such multi-array quality metrics are adequate for bench science, as microarrays begin to be used in clinical settings, single-array quality metrics will be indispensable. To this end, we define a single-array version of one of the best multi-array quality metrics and show that this metric performs as well as the best multi-array metrics. We then use this new quality metric to assess the quality of microarry data available via the Gene Expression Omnibus (GEO) using more than 22,000 Affymetrix HGU133a and HGU133plus2 arrays from 809 studies. CONCLUSIONS: We find that approximately 10 percent of these publicly available arrays are of poor quality. Moreover, the quality of microarray measurements varies greatly from hybridization to hybridization, study to study, and lab to lab, with some experiments producing unusable data. Many of the concepts described here are applicable to other high-throughput technologies.