Whole genome sequencing based analysis of inflammation biomarkers in the Trans-Omics for Precision Medicine (TOPMed) consortium.

Inflammation biomarkers can provide valuable insight into the role of inflammatory processes in many diseases and conditions. Sequencing based analyses of such biomarkers can also serve as an exemplar of the genetic architecture of quantitative traits. To evaluate the biological insight, which can be provided by a multi-ancestry, whole-genome based association study, we performed a comprehensive analysis of 21 inflammation biomarkers from up to 38 465 individuals with whole-genome sequencing from the Trans-Omics for Precision Medicine (TOPMed) program (with varying sample size by trait, where the minimum sample size was n = 737 for MMP-1). We identified 22 distinct single-variant associations across 6 traits-E-selectin, intercellular adhesion molecule 1, interleukin-6, lipoprotein-associated phospholipase A2 activity and mass, and P-selectin-that remained significant after conditioning on previously identified associations for these inflammatory biomarkers. We further expanded upon known biomarker associations by pairing the single-variant analysis with a rare variant set-based analysis that further identified 19 significant rare variant set-based associations with 5 traits. These signals were distinct from both significant single variant association signals within TOPMed and genetic signals observed in prior studies, demonstrating the complementary value of performing both single and rare variant analyses when analyzing quantitative traits. We also confirm several previously reported signals from semi-quantitative proteomics platforms. Many of these signals demonstrate the extensive allelic heterogeneity and ancestry-differentiated variant-trait associations common for inflammation biomarkers, a characteristic we hypothesize will be increasingly observed with well-powered, large-scale analyses of complex traits.

A genetic association study of circulating coagulation factor VIII and von Willebrand factor levels.

ABSTRACT: Coagulation factor VIII (FVIII) and its carrier protein von Willebrand factor (VWF) are critical to coagulation and platelet aggregation. We leveraged whole-genome sequence data from the Trans-Omics for Precision Medicine (TOPMed) program along with TOPMed-based imputation of genotypes in additional samples to identify genetic associations with circulating FVIII and VWF levels in a single-variant meta-analysis, including up to 45 289 participants. Gene-based aggregate tests were implemented in TOPMed. We identified 3 candidate causal genes and tested their functional effect on FVIII release from human liver endothelial cells (HLECs) and VWF release from human umbilical vein endothelial cells. Mendelian randomization was also performed to provide evidence for causal associations of FVIII and VWF with thrombotic outcomes. We identified associations (P < 5 × 10-9) at 7 new loci for FVIII (ST3GAL4, CLEC4M, B3GNT2, ASGR1, F12, KNG1, and TREM1/NCR2) and 1 for VWF (B3GNT2). VWF, ABO, and STAB2 were associated with FVIII and VWF in gene-based analyses. Multiphenotype analysis of FVIII and VWF identified another 3 new loci, including PDIA3. Silencing of B3GNT2 and the previously reported CD36 gene decreased release of FVIII by HLECs, whereas silencing of B3GNT2, CD36, and PDIA3 decreased release of VWF by HVECs. Mendelian randomization supports causal association of higher FVIII and VWF with increased risk of thrombotic outcomes. Seven new loci were identified for FVIII and 1 for VWF, with evidence supporting causal associations of FVIII and VWF with thrombotic outcomes. B3GNT2, CD36, and PDIA3 modulate the release of FVIII and/or VWF in vitro.

Whole genome sequence analysis of platelet traits in the NHLBI Trans-Omics for Precision Medicine (TOPMed) initiative.

Platelets play a key role in thrombosis and hemostasis. Platelet count (PLT) and mean platelet volume (MPV) are highly heritable quantitative traits, with hundreds of genetic signals previously identified, mostly in European ancestry populations. We here utilize whole genome sequencing (WGS) from NHLBI's Trans-Omics for Precision Medicine initiative (TOPMed) in a large multi-ethnic sample to further explore common and rare variation contributing to PLT (n = 61 200) and MPV (n = 23 485). We identified and replicated secondary signals at MPL (rs532784633) and PECAM1 (rs73345162), both more common in African ancestry populations. We also observed rare variation in Mendelian platelet-related disorder genes influencing variation in platelet traits in TOPMed cohorts (not enriched for blood disorders). For example, association of GP9 with lower PLT and higher MPV was partly driven by a pathogenic Bernard-Soulier syndrome variant (rs5030764, p.Asn61Ser), and the signals at TUBB1 and CD36 were partly driven by loss of function variants not annotated as pathogenic in ClinVar (rs199948010 and rs571975065). However, residual signal remained for these gene-based signals after adjusting for lead variants, suggesting that additional variants in Mendelian genes with impacts in general population cohorts remain to be identified. Gene-based signals were also identified at several genome-wide association study identified loci for genes not annotated for Mendelian platelet disorders (PTPRH, TET2, CHEK2), with somatic variation driving the result at TET2. These results highlight the value of WGS in populations of diverse genetic ancestry to identify novel regulatory and coding signals, even for well-studied traits like platelet traits.

Whole-genome sequencing association analysis of quantitative red blood cell phenotypes: The NHLBI TOPMed program.

Whole-genome sequencing (WGS), a powerful tool for detecting novel coding and non-coding disease-causing variants, has largely been applied to clinical diagnosis of inherited disorders. Here we leveraged WGS data in up to 62,653 ethnically diverse participants from the NHLBI Trans-Omics for Precision Medicine (TOPMed) program and assessed statistical association of variants with seven red blood cell (RBC) quantitative traits. We discovered 14 single variant-RBC trait associations at 12 genomic loci, which have not been reported previously. Several of the RBC trait-variant associations (RPN1, ELL2, MIDN, HBB, HBA1, PIEZO1, and G6PD) were replicated in independent GWAS datasets imputed to the TOPMed reference panel. Most of these discovered variants are rare/low frequency, and several are observed disproportionately among non-European Ancestry (African, Hispanic/Latino, or East Asian) populations. We identified a 3 bp indel p.Lys2169del (g.88717175_88717177TCT[4]) (common only in the Ashkenazi Jewish population) of PIEZO1, a gene responsible for the Mendelian red cell disorder hereditary xerocytosis (MIM: 194380), associated with higher mean corpuscular hemoglobin concentration (MCHC). In stepwise conditional analysis and in gene-based rare variant aggregated association analysis, we identified several of the variants in HBB, HBA1, TMPRSS6, and G6PD that represent the carrier state for known coding, promoter, or splice site loss-of-function variants that cause inherited RBC disorders. Finally, we applied base and nuclease editing to demonstrate that the sentinel variant rs112097551 (nearest gene RPN1) acts through a cis-regulatory element that exerts long-range control of the gene RUVBL1 which is essential for hematopoiesis. Together, these results demonstrate the utility of WGS in ethnically diverse population-based samples and gene editing for expanding knowledge of the genetic architecture of quantitative hematologic traits and suggest a continuum between complex trait and Mendelian red cell disorders.

Staying Virtual: A Survey Study of the Virtual Lecture Experience in Academic Medicine.

BACKGROUND: Increasing clinical demands can adversely impact academic advancement, including the ability to deliver lectures and disseminate scholarly work. The virtual lecture platform became mainstream during the height of the coronavirus-19 pandemic. Lessons learned from this period may offer insight into supporting academic productivity among physicians who must balance multiple demands, including high clinical workloads and family care responsibilities. We evaluated perceptions on delivering virtual lectures to determine whether virtual venues merit continuation beyond the pandemic's initial phase and whether these perceptions differ by gender and rank. METHODS: In a survey study, faculty who spoke in 1 of 3 virtual lecture programs in the Departments of Anesthesiology and Critical Care Medicine, Otolaryngology, and Radiology at a university hospital in 2020 to 2022 were queried about their experience. Speakers' motivations to lecture virtually and the perceived advantages and disadvantages of virtual and in-person lectures were analyzed using descriptive statistics and qualitative analyses. RESULTS: Seventy-two of 95 (76%) faculty members responded (40% women, 38% men, and 22% gender undisclosed). Virtual lectures supported the speakers "a lot" to "extremely" with the following goals: enhancing one's reputation and credibility (76%), networking (70%), receiving feedback (63%), and advancing prospects for promotion (59%). Virtual programs also increased the speakers' sense of accomplishment (70%) and professional optimism (61%) by at least "a lot," including instructors and assistant professors who previously had difficulty obtaining invitations to speak outside their institution. Many respondents had declined prior invitations to speak in-person due to clinical workload (66%) and family care responsibilities (58%). Previous opportunities to lecture in-person were also refused due to finances (39%), teaching (26%), and research (19%) requirements, personal medical conditions or disabilities (9%), and religious obligations (5%). Promotion was a stronger motivating factor to lecture virtually for instructors and assistant professors than for associate and full professors. By contrast, disseminating work and ideas was a stronger motivator for associate and full professors. Associate and full professors also reported greater improvement in work-related well-being than earlier career faculty from the virtual lecture experience. Very few differences were found by gender. CONCLUSIONS: Virtual lecture programs support faculty who might not otherwise have the opportunity to lecture in-person due to multiple constraints. To increase the dissemination of scholarly work and expand opportunities to all faculty, virtual lectures should continue even as in-person venues are reestablished.

Low-Volume Acute Normovolemic Hemodilution Does Not Reduce Allogeneic Red Blood Cell Transfusion in Cardiac Surgery in the Modern Era of Patient Blood Management: A Propensity Score-Matched Cohort Study.

OBJECTIVES: Patients undergoing cardiac surgery often require blood transfusions, which are associated with increased morbidity and mortality. Patient blood management (PBM) strategies, including acute normovolemic hemodilution (ANH), have been implemented to minimize allogeneic transfusion requirements. Older studies suggested that ANH is associated with reduced transfusions; however, its effectiveness in the modern era of PBM remains unclear. DESIGN: This was a retrospective cohort study. SETTING: The study was held at a single university hospital. PARTICIPANTS: 542 patients who underwent elective cardiac surgery with cardiopulmonary bypass (CPB) using low-priming-volume circuits between January 2017 and March 2022. INTERVENTIONS: Patients who received ANH were matched with those who did not receive ANH, using propensity scores. MEASUREMENTS AND MAIN RESULTS: The primary outcome was the proportion of patients who received perioperative red blood cell (RBC) transfusion. Of the 542 eligible patients, 49 ANH cases were propensity-score matched to 97 controls. The median ANH volume was 450 mL (IQR, 400-800 mL). There was no significant difference in perioperative RBC transfusion rates between the 2 groups (24.5% in the ANH group vs 30.9% in the control group, p = 0.42). The odds ratio for perioperative RBC transfusion in the ANH group versus the control group was 0.72 (95% CI, 0.32-1.55, p = 0.42). CONCLUSIONS: Low-volume ANH was not associated with a significant reduction in perioperative allogeneic RBC transfusion during cardiac surgery with CPB using low-priming-volume circuits. The benefits of low-volume ANH in reducing the requirement for RBC transfusion in the modern era of PBM may be smaller than reported previously.

Secondary analyses for genome-wide association studies using expression quantitative trait loci.

Genome-wide association studies (GWAS) have successfully identified thousands of single nucleotide polymorphisms (SNPs) associated with complex traits; however, the identified SNPs account for a fraction of trait heritability, and identifying the functional elements through which genetic variants exert their effects remains a challenge. Recent evidence suggests that SNPs associated with complex traits are more likely to be expression quantitative trait loci (eQTL). Thus, incorporating eQTL information can potentially improve power to detect causal variants missed by traditional GWAS approaches. Using genomic, transcriptomic, and platelet phenotype data from the Genetic Study of Atherosclerosis Risk family-based study, we investigated the potential to detect novel genomic risk loci by incorporating information from eQTL in the relevant target tissues (i.e., platelets and megakaryocytes) using established statistical principles in a novel way. Permutation analyses were performed to obtain family-wise error rates for eQTL associations, substantially lowering the genome-wide significance threshold for SNP-phenotype associations. In addition to confirming the well known association between PEAR1 and platelet aggregation, our eQTL-focused approach identified a novel locus (rs1354034) and gene (ARHGEF3) not previously identified in a GWAS of platelet aggregation phenotypes. A colocalization analysis showed strong evidence for a functional role of this eQTL.

Transcriptional profile of platelets and iPSC-derived megakaryocytes from whole-genome and RNA sequencing.

Genome-wide association studies have identified common variants associated with platelet-related phenotypes, but because these variants are largely intronic or intergenic, their link to platelet biology is unclear. In 290 normal subjects from the GeneSTAR Research Study (110 African Americans [AAs] and 180 European Americans [EAs]), we generated whole-genome sequence data from whole blood and RNA sequence data from extracted nonribosomal RNA from 185 induced pluripotent stem cell-derived megakaryocyte (MK) cell lines (platelet precursor cells) and 290 blood platelet samples from these subjects. Using eigenMT software to select the peak single-nucleotide polymorphism (SNP) for each expressed gene, and meta-analyzing the results of AAs and EAs, we identify (q-value < 0.05) 946 cis-expression quantitative trait loci (eQTLs) in derived MKs and 1830 cis-eQTLs in blood platelets. Among the 57 eQTLs shared between the 2 tissues, the estimated directions of effect are very consistent (98.2% concordance). A high proportion of detected cis-eQTLs (74.9% in MKs and 84.3% in platelets) are unique to MKs and platelets compared with peak-associated SNP-expressed gene pairs of 48 other tissue types that are reported in version V7 of the Genotype-Tissue Expression Project. The locations of our identified eQTLs are significantly enriched for overlap with several annotation tracks highlighting genomic regions with specific functionality in MKs, including MK-specific DNAse hotspots, H3K27-acetylation marks, H3K4-methylation marks, enhancers, and superenhancers. These results offer insights into the regulatory signature of MKs and platelets, with significant overlap in genes expressed, eQTLs detected, and enrichment within known superenhancers relevant to platelet biology.

Association Between Left Ventricular Relative Wall Thickness and Acute Kidney Injury After Noncardiac Surgery.

BACKGROUND: Acute kidney injury (AKI) after major noncardiac surgery is commonly attributed to cardiovascular dysfunction. Identifying novel associations between preoperative cardiovascular markers and kidney injury may guide risk stratification and perioperative intervention. Increased left ventricular relative wall thickness (RWT), routinely measured on echocardiography, is associated with myocardial dysfunction and long-term risk of heart failure in patients with preserved left ventricular ejection fraction (LVEF); however, its relationship to postoperative complications has not been studied. We evaluated the association between preoperative RWT and AKI in high-risk noncardiac surgical patients with preserved LVEF. METHODS: Patients ≥18 years of age having major noncardiac surgery (high-risk elective intra-abdominal or noncardiac intrathoracic surgery) between July 1, 2016, and June 30, 2018, who had transthoracic echocardiography in the previous 12 months were eligible. Patients with preoperative creatinine ≥2 mg/dL or reduced LVEF (<50%) were excluded. The association between RWT and AKI, defined as an increase in serum creatinine by 0.3 mg/dL from baseline within 48 hours or by 50% within 7 days after surgery, was assessed using multivariable logistic regression adjusted for preoperative covariates. An additional model adjusted for intraoperative covariates, which are strongly associated with AKI, especially hypotension. RWT was modeled continuously, associating the change in odds of AKI for each 0.1 increase in RWT. RESULTS: The study included 1041 patients (mean ± standard deviation [SD] age 62 ± 15 years; 59% female). A total of 145 subjects (13.9%) developed AKI within 7 days. For RWT quartiles 1 through 4, respectively, 20 of 262 (7.6%), 40 of 259 (15.4%), 39 of 263 (14.8%), and 46 of 257 (17.9%) developed AKI. Log-odds and proportion with AKI increased across the observed RWT values. After adjusting for confounders (demographics, American Society of Anesthesiologists [ASA] physical status, comorbidities, baseline creatinine, antihypertensive medications, and left ventricular mass index), each RWT increase of 0.1 was associated with an estimated 26% increased odds of developing AKI (odds ratio [OR]; 95% confidence interval [CI]) of 1.26 (1.09-1.46; P = .002). After adjusting for intraoperative covariates (length of surgery, presence of an arterial line, intraoperative hypotension, crystalloid administration, transfusion, and urine output), RWT remained independently associated with the odds of AKI (OR; 95% CI) of 1.28 (1.13-1.47; P = .001). Increased RWT was also independently associated with hospital length of stay and adjusted hazard ratio (HR [95% CI]) of 0.94 (0.89-0.99; P = .018). CONCLUSIONS: Left ventricular RWT is a novel cardiovascular factor associated with AKI within 7 days after high-risk noncardiac surgery among patients with preserved LVEF. Application of this commonly available measurement of risk stratification or perioperative intervention warrants further investigation.

Use of >100,000 NHLBI Trans-Omics for Precision Medicine (TOPMed) Consortium whole genome sequences improves imputation quality and detection of rare variant associations in admixed African and Hispanic/Latino populations.

Most genome-wide association and fine-mapping studies to date have been conducted in individuals of European descent, and genetic studies of populations of Hispanic/Latino and African ancestry are limited. In addition, these populations have more complex linkage disequilibrium structure. In order to better define the genetic architecture of these understudied populations, we leveraged >100,000 phased sequences available from deep-coverage whole genome sequencing through the multi-ethnic NHLBI Trans-Omics for Precision Medicine (TOPMed) program to impute genotypes into admixed African and Hispanic/Latino samples with genome-wide genotyping array data. We demonstrated that using TOPMed sequencing data as the imputation reference panel improves genotype imputation quality in these populations, which subsequently enhanced gene-mapping power for complex traits. For rare variants with minor allele frequency (MAF) < 0.5%, we observed a 2.3- to 6.1-fold increase in the number of well-imputed variants, with 11-34% improvement in average imputation quality, compared to the state-of-the-art 1000 Genomes Project Phase 3 and Haplotype Reference Consortium reference panels. Impressively, even for extremely rare variants with minor allele count <10 (including singletons) in the imputation target samples, average information content rescued was >86%. Subsequent association analyses of TOPMed reference panel-imputed genotype data with hematological traits (hemoglobin (HGB), hematocrit (HCT), and white blood cell count (WBC)) in ~21,600 African-ancestry and ~21,700 Hispanic/Latino individuals identified associations with two rare variants in the HBB gene (rs33930165 with higher WBC [p = 8.8x10-15] in African populations, rs11549407 with lower HGB [p = 1.5x10-12] and HCT [p = 8.8x10-10] in Hispanics/Latinos). By comparison, neither variant would have been genome-wide significant if either 1000 Genomes Project Phase 3 or Haplotype Reference Consortium reference panels had been used for imputation. Our findings highlight the utility of the TOPMed imputation reference panel for identification of novel rare variant associations not previously detected in similarly sized genome-wide studies of under-represented African and Hispanic/Latino populations.

Targeted deep sequencing of the PEAR1 locus for platelet aggregation in European and African American families.

Coronary artery disease (CAD) remains a major cause of mortality and morbidity worldwide. The aggregation of activated platelets on a ruptured atherosclerotic plaque is a critical step in most acute cardiovascular events like myocardial infarction. Platelet aggregation both at baseline and after aspirin is highly heritable. Genome-wide association studies (GWAS) have identified a common variant within the first intron of the platelet endothelial aggregation receptor1 (PEAR1), to be robustly associated with platelet aggregation. In this study, we used targeted deep sequencing to fine-map the prior GWAS peak and identify additional rare variants of PEAR1 that account for missing heritability in platelet aggregation within the GeneSTAR families. In this study, 1709 subjects (1043 European Americans, EA and 666 African Americans, AA) from families in the GeneSTAR study were included. In vitro platelet aggregation in response to collagen, ADP and epinephrine was measured at baseline and 14 days after aspirin therapy (81 mg/day). Targeted deep sequencing of PEAR1 in addition to 2kb of upstream and downstream of the gene was performed. Under an additive genetic model, the association of single variants of PEAR1 with platelet aggregation phenotypes were examined. Additionally, we examined the association between the burden of PEAR1 rare non-synonymous variants and platelet aggregation phenotypes. Of 532 variants identified through sequencing, the intron 1 variant, rs12041331, was significantly associated with all platelet aggregation phenotypes at baseline and after platelet inhibition with aspirin therapy. rs12566888, which is in linkage disequilibrium with rs12041331, was associated with platelet aggregation phenotypes but to a lesser extent. In the EA families, the burden of PEAR1 missense variants was associated with platelet aggregation after aspirin therapy when the platelets were stimulated with epinephrine (p = 0.0009) and collagen (p = 0.03). In AAs, the burden of PEAR1 missense variants was associated, to a lesser degree, with platelet aggregation in response to epinephrine (p = 0.02) and ADP (p = 0.04). Our study confirmed that the GWAS-identified variant, rs12041331, is the strongest variant associated with platelet aggregation both at baseline and after aspirin therapy in our GeneSTAR families in both races. We identified additional association of rare missense variants in PEAR1 with platelet aggregation following aspirin therapy. However, we observed a racial difference in the contribution of these rare variants to the platelet aggregation, most likely due to higher residual missing heritability of platelet aggregation after accounting for rs12041331 in the EAs compared to AAs.

Exome-chip meta-analysis identifies association between variation in ANKRD26 and platelet aggregation.

Previous genome-wide association studies (GWAS) have identified several variants associated with platelet function phenotypes; however, the proportion of variance explained by the identified variants is mostly small. Rare coding variants, particularly those with high potential for impact on protein structure/function, may have substantial impact on phenotype but are difficult to detect by GWAS. The main purpose of this study was to identify low frequency or rare variants associated with platelet function using genotype data from the Illumina HumanExome Bead Chip. Three family-based cohorts of European ancestry, including ~4,000 total subjects, comprised the discovery cohort and two independent cohorts, one of European and one of African American ancestry, were used for replication. Optical aggregometry in platelet-rich plasma was performed in all the discovery cohorts in response to adenosine diphosphate (ADP), epinephrine, and collagen. Meta-analyses were performed using both gene-based and single nucleotide variant association methods. The gene-based meta-analysis identified a significant association (P = 7.13 × 10-7) between rare genetic variants in ANKRD26 and ADP-induced platelet aggregation. One of the ANKRD26 SNVs - rs191015656, encoding a threonine to isoleucine substitution predicted to alter protein structure/function, was replicated in Europeans. Aggregation increases of ~20-50% were observed in heterozygotes in all cohorts. Novel genetic signals in ABCG1 and HCP5 were also associated with platelet aggregation to ADP in meta-analyses, although only results for HCP5 could be replicated. The SNV in HCP5 intersects epigenetic signatures in CD41+ megakaryocytes suggesting a new functional role in platelet biology for HCP5. This is the first study to use gene-based association methods from SNV array genotypes to identify rare variants related to platelet function. The molecular mechanisms and pathophysiological relevance for the identified genetic associations requires further study.

Separating fast and slow exchange transfer and magnetization transfer using off-resonance variable-delay multiple-pulse (VDMP) MRI.

PURPOSE: To develop a method that can separate and quantify the fast (>1 kHz) and slow exchange transfer and magnetization transfer components in Z-spectra. METHODS: Z-spectra were recorded as a function of mixing time using a train of selective pulses providing variable-delay multipulse build-up curves. Fast and slow transfer components in the Z-spectra were separated and quantified on a voxel-by-voxel basis by fitting the mixing time-dependent CEST signal using a 3-pool model. RESULTS: Phantom studies of glutamate solution, bovine serum albumin solution, and hair conditioner showed the capability of the proposed method to separate fast and slow transfer components. In vivo mouse brain studies showed a strong contrast between white matter and gray matter in the slow-transferring map, corresponding to an asymmetric component of the conventional semisolid magnetization transfer contrast. In addition, a fast-transferring proton map was found that was homogeneous across the brain and attributed to the total contributions of the fast-exchanging protons from proteins, metabolites, and a symmetric magnetization transfer contrast component. CONCLUSIONS: This new method provides a simple way to extract fast and slow transfer components from the Z-spectrum, leading to novel MRI contrasts, and providing insight into the different magnetization transfer contrast contributions.

Platelet glycoproteins associated with aspirin-treatment upon platelet activation.

Platelet glycoproteins are known to play central roles in hemostasis and vascular integrity and have pathologic roles in vascular occlusive diseases such as myocardial infarction and stroke. Characterizing glycoproteins within and secreted by platelets can provide insight into the mechanisms that underlie vascular pathologies and the therapeutic benefits or failure of anti-platelet agents. To study the impact of aspirin, which is commonly prescribed for primary and secondary cardiovascular prevention, on the platelet glycoproteome, we evaluated washed platelets from ten donors. The platelet glycoproteome, was studied using an iTRAQ in resting and stimulated states and with and without aspirin treatment. Using solid phase extraction of glycosite-containing peptides (SPEG), we were able to identify 799 unique N-linked glycosylation sites (glycosites) in platelets, representing the largest and the most comprehensive analysis to date. We were able to identity a number of glycoproteins impacted by aspirin treatment, which we validated using global proteomics analysis of platelets and their secreted proteins. In our analyses, metallopeptidase inhibitor 1 (TIMP1) was the single most significantly affected glycoprotein by aspirin treatment. ELISA assays confirmed proteomic results and validated our strategy. Functional analysis demonstrated that TIMP1 levels were highly correlated with platelet reactivity in vitro, with a correlation coefficient of -0.5. The release of TIMP1 from platelets, which was previously unknown to be affected by aspirin treatment, may play important roles in hemostasis and/or vascular integrity. If validated, our findings may be useful for developing assays that assess platelet response to aspirin or other anti-platelet therapies.

Association of Selective Serotonin Reuptake Inhibitors with Transfusion in Surgical Patients.

BACKGROUND: The clinical relevance of chronic exposure to selective serotonin reuptake inhibitors (SSRIs) to transfusion in surgical patients is unclear. METHODS: We conducted a prospective cohort study involving patients undergoing cardiac, vascular, spinal, and intracranial surgery at 2 academic medical centers. Medication use, demographics, comorbidities, and laboratory values were determined at baseline by patient interview and review of medical records. The primary outcome was transfusion of any hemostatic allogeneic blood product (i.e., fresh frozen plasma, platelets, and/or cryoprecipitate) through postoperative day 2. RESULTS: The study sample consisted of 767 patients; 364 patients (47.5%) underwent cardiac surgery and the remainder underwent noncardiac surgery. Eighty-eight patients (11.5%) used SSRIs preoperatively. Among cardiac patients, the absolute number of allogeneic transfusions was higher for SSRI users than nonusers (2 [0-6] vs 0 [0-2], median [25%-75%], respectively, P = 0.008), and a similar trend was observed for noncardiac surgery. After adjusting for covariates using ordinal logistic regression, preoperative SSRI use was associated with an approximately 2-fold (odds ratio, 2.2; 95% confidence interval, 1.2-3.98) increase in odds of exposure to allogeneic hemostatic blood products; similar results were observed using propensity score adjustment (odds ratio, 1.85; 95% confidence interval, 1.11-3.07). A significant interaction between SSRI use and surgery type, age, sex, or concurrent antiplatelet therapy was not found; however, heterogeneity in magnitude of effect could not be excluded. CONCLUSIONS: Preoperative use of SSRIs is associated with increased exposure to allogeneic hemostatic blood products in surgical patients at high risk for perioperative bleeding. Determining whether perioperative continuation or withdrawal of SSRIs produces a net clinical benefit requires randomized controlled trials.

Genome-wide association study of platelet aggregation in African Americans.

BACKGROUND: We have previously shown that platelet aggregation has higher heritability in African Americans than European Americans. However, a genome-wide association study (GWAS) of platelet aggregation in African Americans has not been reported. We measured platelet aggregation in response to arachidonic acid, ADP, collagen, or epinephrine by optical aggregometry. The discovery cohort was 825 African Americans from the GeneSTAR study. Two replication cohorts were used: 119 African Americans from the Platelet Genes and Physiology Study and 1221 European Americans from GeneSTAR. Genotyping was conducted with Illumina 1 M arrays. For each cohort, age- and sex-adjusted linear mixed models were used to test for association between each SNP and each phenotype under an additive model. RESULTS: Six SNPs were significantly associated with platelet aggregation (P<5×10(-8)) in the discovery sample. Of these, three SNPs in three different loci were confirmed: 1) rs12041331, in PEAR1 (platelet endothelial aggregation receptor 1), replicated in both African and European Americans for collagen- and epinephrine-induced aggregation, and in European Americans for ADP-induced aggregation; 2) rs11202221, in BMPR1A (bone morphogenetic protein receptor type1A), replicated in African Americans for ADP-induced aggregation; and 3) rs6566765 replicated in European Americans for ADP-induced aggregation. The rs11202221 and rs6566765 associations with agonist-induced platelet aggregation are novel. CONCLUSIONS: In this first GWAS of agonist-induced platelet aggregation in African Americans, we discovered and replicated, novel associations of two variants with ADP-induced aggregation, and confirmed the association of a PEAR1 variant with multi-agonist-induced aggregation. Further study of these genes may provide novel insights into platelet biology.

Protocol adherence when managing massive bleeding following complex cardiac surgery: a study design pilot.

OBJECTIVE: High-quality prospective trials of hemostatic "rescue" therapy to control massive bleeding in cardiac surgery are lacking. Wide variability in the care of patients with severe bleeding following cardiopulmonary bypass has precluded accurate comparison of treatment groups in previous studies. This study identified the use of a management protocol for early identification and uniform treatment of patients with massive bleeding for application in future trials of hemostatic rescue agents. DESIGN: A prospective, nonblinded, interventional feasibility study. SETTING: A university teaching hospital. PARTICIPANTS: Forty-three adult patients undergoing complex cardiac surgery. INTERVENTIONS: Study participants undergoing high-risk cardiac surgery received standardized treatment in accordance with a bleeding management protocol. MEASUREMENTS AND MAIN RESULTS: Twenty-seven patients (63%) had severe bleeding following heparin reversal and received conventional hemostatic resuscitation per protocol. Six patients had massive refractory bleeding. Compliance with protocol tasks was≥90% in 4 of 5 categories (anticoagulation, hemostasis scoring, recording blood loss, protocol transfusion) with the exception being submission of laboratory samples (76%). Measured bleeding rates (mL/h) following heparin reversal were clearly differentiated in those with hemostasis scores≥3 compared to those with scores≤2 (1,420±957 v 147±96; p<0.001). CONCLUSIONS: Adherence to a management protocol for massive bleeding is feasible and allows for homogenous treatment of patients before study arm randomization in future "rescue" therapy trials. The authors' protocol allowed for prompt and accurate identification of patients with severe bleeding refractory to conventional therapy. This review resolved several key barriers in the design of severe bleeding management trials.

Association of Preoperative Immune Checkpoint Inhibitor Therapy With Cardiopulmonary Instability and Organ Injury After High-Risk Surgery.

OBJECTIVES: To assess the relationship between prior exposure to immune checkpoint inhibitors (ICIs) and the risk of postoperative complications in cancer patients. DESIGN: Single-center retrospective cohort study. INTERVENTIONS: The main exposure was treatment with an FDA-approved ICI within 6 months before surgery. MEASUREMENTS AND MAIN RESULTS: Exposure to ICIs and covariates was determined from the electronic health record. The primary outcome was a composite of postoperative complications, including prolonged pressor or oxygen dependence, kidney injury, or myocardial injury. Secondary outcomes included each subcomponent of the primary outcome. Of 7674 subjects with cancer admitted to the ICU after surgery, 247 were exposed to one or more ICIs in the 6 months before surgery. After propensity score matching, 197 ICI-exposed subjects were matched to 777 nonexposed. The composite outcome occurred in 70 of 197 (35.5%) ICI-exposed subjects and 251 of 777 (32.3%) nonexposed. There was no difference between exposed and nonexposed groups in the primary composite outcome (odds ratio [OR], 1.12; 95% CI, 0.80-1.58) by conditional logistic regression. Risk of the secondary outcome of prolonged pressor dependence was significantly higher in ICI-exposed subjects (OR, 1.64; 95% CI, 1.01-2.67). Risks of oxygen dependence (OR, 1.13; 95% CI, 0.75-1.73), kidney injury (OR, 1.15; 95% CI, 0.77-1.71), and myocardial injury (OR, 1.76; 95% CI, 1.00-3.10) were not significantly different. There was no difference between groups in the time to hospital discharge alive (p = 0.62). CONCLUSIONS: Exposure to ICIs within 6 months before high-risk surgery was not associated with the composite outcome of cardiopulmonary instability or organ injury in patients with cancer. The potential for an association with the secondary outcomes of cardiac instability and injury is worthy of future study.

Commitment to inclusion: The importance of collaboration in gender equity work.

Despite decades of faculty professional development programs created to prepare women for leadership, gender inequities persist in salary, promotion, and leadership roles. Indeed, men still earn more than women, are more likely than women to hold the rank of professor, and hold the vast majority of positions of power in academic medicine. Institutions demonstrate commitment to their faculty's growth by investing resources, including creating faculty development programs. These programs are essential to help prepare women to lead and navigate the highly matrixed, complex systems of academic medicine. However, data still show that women persistently lag behind men in their career advancement and salary. Clearly, training women to adapt to existing structures and norms alone is not sufficient. To effectively generate organizational change, leaders with power and resources must commit to gender equity. This article describes several efforts by the Office of Faculty in the Johns Hopkins University School of Medicine to broaden inclusivity in collaborative work for gender equity. The authors are women and men leaders in the Office of Faculty, which is within the Johns Hopkins University School of Medicine dean's office and includes Women in Science and Medicine. Here, we discuss potential methods to advance gender equity using inclusivity based on our institutional experience and on the findings of other studies. Ongoing data collection to evaluate programmatic outcomes in the Johns Hopkins University School of Medicine will be reported in the future.