Comparative Analysis of Single-Cell RNA Sequencing Methods.

Single-cell RNA sequencing (scRNA-seq) offers new possibilities to address biological and medical questions. However, systematic comparisons of the performance of diverse scRNA-seq protocols are lacking. We generated data from 583 mouse embryonic stem cells to evaluate six prominent scRNA-seq methods: CEL-seq2, Drop-seq, MARS-seq, SCRB-seq, Smart-seq, and Smart-seq2. While Smart-seq2 detected the most genes per cell and across cells, CEL-seq2, Drop-seq, MARS-seq, and SCRB-seq quantified mRNA levels with less amplification noise due to the use of unique molecular identifiers (UMIs). Power simulations at different sequencing depths showed that Drop-seq is more cost-efficient for transcriptome quantification of large numbers of cells, while MARS-seq, SCRB-seq, and Smart-seq2 are more efficient when analyzing fewer cells. Our quantitative comparison offers the basis for an informed choice among six prominent scRNA-seq methods, and it provides a framework for benchmarking further improvements of scRNA-seq protocols.

Serial sarcomere number is substantially decreased within the paretic biceps brachii in individuals with chronic hemiparetic stroke.

A muscle's structure, or architecture, is indicative of its function and is plastic; changes in input to or use of the muscle alter its architecture. Stroke-induced neural deficits substantially alter both input to and usage of individual muscles. We combined in vivo imaging methods (second-harmonic generation microendoscopy, extended field-of-view ultrasound, and fat-suppression MRI) to quantify functionally meaningful architecture parameters in the biceps brachii of both limbs of individuals with chronic hemiparetic stroke and in age-matched, unimpaired controls. Specifically, serial sarcomere number (SSN) and physiological cross-sectional area (PCSA) were calculated from data collected at three anatomical scales: sarcomere length, fascicle length, and muscle volume. The interlimb differences in SSN and PCSA were significantly larger for stroke participants than for participants without stroke (P = 0.0126 and P = 0.0042, respectively), suggesting we observed muscle adaptations associated with stroke rather than natural interlimb variability. The paretic biceps brachii had ∼8,200 fewer serial sarcomeres and ∼2 cm2 smaller PCSA on average than the contralateral limb (both P < 0.0001). This was manifested by substantially smaller muscle volumes (112 versus 163 cm3), significantly shorter fascicles (11.0 versus 14.0 cm; P < 0.0001), and comparable sarcomere lengths (3.55 versus 3.59 µm; P = 0.6151) between limbs. Most notably, this study provides direct evidence of the loss of serial sarcomeres in human muscle observed in a population with neural impairments that lead to disuse and chronically place the affected muscle at a shortened position. This adaptation is consistent with functional consequences (increased passive resistance to elbow extension) that would amplify already problematic, neurally driven motor impairments.

Single-cell RNA-seq reveals developmental plasticity with coexisting oncogenic states and immune evasion programs in ETP-ALL.

Lineage plasticity and stemness have been invoked as causes of therapy resistance in cancer, because these flexible states allow cancer cells to dedifferentiate and alter their dependencies. We investigated such resistance mechanisms in relapsed/refractory early T-cell progenitor acute lymphoblastic leukemia (ETP-ALL) carrying activating NOTCH1 mutations via full-length single-cell RNA sequencing (scRNA-seq) of malignant and microenvironmental cells. We identified 2 highly distinct stem-like states that critically differed with regard to cell cycle and oncogenic signaling. Fast-cycling stem-like leukemia cells demonstrated Notch activation and were effectively eliminated in patients by Notch inhibition, whereas slow-cycling stem-like cells were Notch independent and rather relied on PI3K signaling, likely explaining the poor efficacy of Notch inhibition in this disease. Remarkably, we found that both stem-like states could differentiate into a more mature leukemia state with prominent immunomodulatory functions, including high expression of the LGALS9 checkpoint molecule. These cells promoted an immunosuppressive leukemia ecosystem with clonal accumulation of dysfunctional CD8+ T cells that expressed HAVCR2, the cognate receptor for LGALS9. Our study identified complex interactions between signaling programs, cellular plasticity, and immune programs that characterize ETP-ALL, illustrating the multidimensionality of tumor heterogeneity. In this scenario, combination therapies targeting diverse oncogenic states and the immune ecosystem seem most promising to successfully eliminate tumor cells that escape treatment through coexisting transcriptional programs.

bigSCale: an analytical framework for big-scale single-cell data.

Single-cell RNA sequencing (scRNA-seq) has significantly deepened our insights into complex tissues, with the latest techniques capable of processing tens of thousands of cells simultaneously. Analyzing increasing numbers of cells, however, generates extremely large data sets, extending processing time and challenging computing resources. Current scRNA-seq analysis tools are not designed to interrogate large data sets and often lack sensitivity to identify marker genes. With bigSCale, we provide a scalable analytical framework to analyze millions of cells, which addresses the challenges associated with large data sets. To handle the noise and sparsity of scRNA-seq data, bigSCale uses large sample sizes to estimate an accurate numerical model of noise. The framework further includes modules for differential expression analysis, cell clustering, and marker identification. A directed convolution strategy allows processing of extremely large data sets, while preserving transcript information from individual cells. We evaluated the performance of bigSCale using both a biological model of aberrant gene expression in patient-derived neuronal progenitor cells and simulated data sets, which underlines the speed and accuracy in differential expression analysis. To test its applicability for large data sets, we applied bigSCale to assess 1.3 million cells from the mouse developing forebrain. Its directed down-sampling strategy accumulates information from single cells into index cell transcriptomes, thereby defining cellular clusters with improved resolution. Accordingly, index cell clusters identified rare populations, such as reelin (Reln)-positive Cajal-Retzius neurons, for which we report previously unrecognized heterogeneity associated with distinct differentiation stages, spatial organization, and cellular function. Together, bigSCale presents a solution to address future challenges of large single-cell data sets.

Leveraging genome-wide data to investigate differences between opioid use vs. opioid dependence in 41,176 individuals from the Psychiatric Genomics Consortium.

To provide insights into the biology of opioid dependence (OD) and opioid use (i.e., exposure, OE), we completed a genome-wide analysis comparing 4503 OD cases, 4173 opioid-exposed controls, and 32,500 opioid-unexposed controls, including participants of European and African descent (EUR and AFR, respectively). Among the variants identified, rs9291211 was associated with OE (exposed vs. unexposed controls; EUR z = -5.39, p = 7.2 × 10-8). This variant regulates the transcriptomic profiles of SLC30A9 and BEND4 in multiple brain tissues and was previously associated with depression, alcohol consumption, and neuroticism. A phenome-wide scan of rs9291211 in the UK Biobank (N > 360,000) found association of this variant with propensity to use dietary supplements (p = 1.68 × 10-8). With respect to the same OE phenotype in the gene-based analysis, we identified SDCCAG8 (EUR + AFR z = 4.69, p = 10-6), which was previously associated with educational attainment, risk-taking behaviors, and schizophrenia. In addition, rs201123820 showed a genome-wide significant difference between OD cases and unexposed controls (AFR z = 5.55, p = 2.9 × 10-8) and a significant association with musculoskeletal disorders in the UK Biobank (p = 4.88 × 10-7). A polygenic risk score (PRS) based on a GWAS of risk-tolerance (n = 466,571) was positively associated with OD (OD vs. unexposed controls, p = 8.1 × 10-5; OD cases vs. exposed controls, p = 0.054) and OE (exposed vs. unexposed controls, p = 3.6 × 10-5). A PRS based on a GWAS of neuroticism (n = 390,278) was positively associated with OD (OD vs. unexposed controls, p = 3.2 × 10-5; OD vs. exposed controls, p = 0.002) but not with OE (p = 0.67). Our analyses highlight the difference between dependence and exposure and the importance of considering the definition of controls in studies of addiction.

Polygenic Risk Score Prediction of Alcohol Dependence Symptoms Across Population-Based and Clinically Ascertained Samples.

BACKGROUND: Despite consistent evidence of the heritability of alcohol use disorders (AUDs), few specific genes with an etiological role have been identified. It is likely that AUDs are highly polygenic; however, the etiological pathways and genetic variants involved may differ between populations. The aim of this study was thus to evaluate whether aggregate genetic risk for AUDs differed between clinically ascertained and population-based epidemiological samples. METHODS: Four independent samples were obtained: 2 from unselected birth cohorts (Avon Longitudinal Study of Parents and Children [ALSPAC], N = 4,304; FinnTwin12 [FT12], N = 1,135) and 2 from families densely affected with AUDs, identified from treatment-seeking patients (Collaborative Study on the Genetics of Alcoholism, N = 2,097; Irish Affected Sib Pair Study of Alcohol Dependence, N = 706). AUD symptoms were assessed with clinical interviews, and participants of European ancestry were genotyped. Genomewide association was conducted separately in each sample, and the resulting association weights were used to create polygenic risk scores in each of the other samples (12 total discovery-validation pairs), and from meta-analyses within sample type. We then tested how well these aggregate genetic scores predicted AUD outcomes within and across sample types. RESULTS: Polygenic scores derived from 1 population-based sample (ALSPAC) significantly predicted AUD symptoms in another population-based sample (FT12), but not in either clinically ascertained sample. Trend-level associations (uncorrected p < 0.05) were found for polygenic score predictions within sample types but no or negative predictions across sample types. Polygenic scores accounted for 0 to 1% of the variance in AUD symptoms. CONCLUSIONS: Though preliminary, these results provide suggestive evidence of differences in the genetic etiology of AUDs based on sample characteristics such as treatment-seeking status, which may index other important clinical or demographic factors that moderate genetic influences. Although the variance accounted for by genomewide polygenic scores remains low, future studies could improve gene identification efforts by amassing very large samples, or reducing genetic heterogeneity by informing analyses with other phenotypic information such as sample characteristics. Multiple complementary approaches may be needed to make progress in gene identification for this complex disorder.

Using Patterns of Genetic Association to Elucidate Shared Genetic Etiologies Across Psychiatric Disorders.

Twin studies indicate that latent genetic factors overlap across comorbid psychiatric disorders. In this study, we used a novel approach to elucidate shared genetic factors across psychiatric outcomes by clustering single nucleotide polymorphisms based on their genome-wide association patterns. We applied latent profile analysis (LPA) to p-values resulting from genome-wide association studies across three phenotypes: symptom counts of alcohol dependence (AD), antisocial personality disorder (ASP), and major depression (MD), using the European-American case-control genome-wide association study subsample of the collaborative study on the genetics of alcoholism (N = 1399). In the 3-class model, classes were characterized by overall low associations (85.6% of SNPs), relatively stronger association only with MD (6.8%), and stronger associations with AD and ASP but not with MD (7.6%), respectively. These results parallel the genetic factor structure identified in twin studies. The findings suggest that applying LPA to association results across multiple disorders may be a promising approach to identify the specific genetic etiologies underlying shared genetic variance.

Genomewide Association Study of Alcohol Dependence Identifies Risk Loci Altering Ethanol-Response Behaviors in Model Organisms.

BACKGROUND: Alcohol dependence (AD) shows evidence for genetic liability, but genes influencing risk remain largely unidentified. METHODS: We conducted a genomewide association study in 706 related AD cases and 1,748 unscreened population controls from Ireland. We sought replication in 15,496 samples of European descent. We used model organisms (MOs) to assess the role of orthologous genes in ethanol (EtOH)-response behaviors. We tested 1 primate-specific gene for expression differences in case/control postmortem brain tissue. RESULTS: We detected significant association in COL6A3 and suggestive association in 2 previously implicated loci, KLF12 and RYR3. None of these signals are significant in replication. A suggestive signal in the long noncoding RNA LOC339975 is significant in case:control meta-analysis, but not in a population sample. Knockdown of a COL6A3 ortholog in Caenorhabditis elegans reduced EtOH sensitivity. Col6a3 expression correlated with handling-induced convulsions in mice. Loss of function of the KLF12 ortholog in C. elegans impaired development of acute functional tolerance (AFT). Klf12 expression correlated with locomotor activation following EtOH injection in mice. Loss of function of the RYR3 ortholog reduced EtOH sensitivity in C. elegans and rapid tolerance in Drosophila. The ryanodine receptor antagonist dantrolene reduced motivation to self-administer EtOH in rats. Expression of LOC339975 does not differ between cases and controls but is reduced in carriers of the associated rs11726136 allele in nucleus accumbens (NAc). CONCLUSIONS: We detect association between AD and COL6A3, KLF12, RYR3, and LOC339975. Despite nonreplication of COL6A3, KLF12, and RYR3 signals, orthologs of these genes influence behavioral response to EtOH in MOs, suggesting potential involvement in human EtOH response and AD liability. The associated LOC339975 allele may influence gene expression in human NAc. Although the functions of long noncoding RNAs are poorly understood, there is mounting evidence implicating these genes in multiple brain functions and disorders.

Single-Cell Genomics Unravels Brain Cell-Type Complexity.

The brain is the most complex tissue in terms of cell types that it comprises, to the extent that it is still poorly understood. Single cell genome and transcriptome profiling allow to disentangle the neuronal heterogeneity, enabling the categorization of individual neurons into groups with similar molecular signatures. Herein, we unravel the current state of knowledge in single cell neurogenomics. We describe the molecular understanding of the cellular architecture of the mammalian nervous system in health and in disease; from the discovery of unrecognized cell types to the validation of known ones, applying these state-of-the-art technologies.

Altered expression of the imprinted transcription factor PLAGL1 deregulates a network of genes in the human IUGR placenta.

Genomic imprinting is the epigenetic process that results in monoallelic expression of genes depending on parental origin. These genes are known to be critical for placental development and fetal growth in mammals. Aberrant epigenetic profiles at imprinted loci, such as DNA methylation defects, are surprisingly rare in pregnancies with compromised fetal growth, while variations in transcriptional output from the expressed alleles of imprinted genes are more commonly reported in pregnancies complicated with intrauterine growth restriction (IUGR). To determine if PLAGL1 and HYMAI, two imprinted transcripts deregulated in Transient Neonatal Diabetes Mellitus, are involved in non-syndromic IUGR we compared the expression and DNA methylation levels in a large cohort of placental biopsies from IUGR and uneventful pregnancies. This revealed that despite appropriate maternal methylation at the shared PLAGL1/HYMAI promoter, there was a loss of correlation between PLAGL1 and HYMAI expression in IUGR. This incongruity was due to higher HYMAI expression in IUGR gestations, coupled with PLAGL1 down-regulation in placentas from IUGR girls, but not boys. The PLAGL1 protein is a zinc-finger transcription factor that has been shown to be a master coordinator of a genetic growth network in mice. We observe PLAGL1 binding to the H19/IGF2 shared enhancers in placentae, with significant correlations between PLAGL1 levels with H19 and IGF2 expression levels. In addition, PLAGL1 binding and expression also correlate with expression levels of metabolic regulator genes SLC2A4, TCF4 and PPARγ1. Our results strongly suggest that fetal growth can be influenced by altered expression of the PLAGL1 gene network in human placenta.

Force Required to Cinch the Tricuspid Annulus: An Ex-Vivo Study.

BACKGROUND AND AIM OF THE STUDY: Tricuspid annuloplasty is the most preferred technique for the treatment of functional tricuspid regurgitation (FTR). However, high incidences of recurrent regurgitation and risky reoperation demands a deeper insight into the technique. The cinching force required to bring a dilated annulus back to the original size is unknown. The study aim was to quantify the cinching force in the tricuspid annulus which can contribute to the long-term durability of tricuspid annuloplasty and percutaneous device design. METHODS: In ten ovine hearts, a suture was anchored around the free wall of the tricuspid annulus with the free end attached to a force transducer. The force transducer was mounted on a slider system which pulled the suture at regular intervals. Closure of the tricuspid valve was achieved by pressurizing the right ventricle at 30 mmHg through the pulmonary valve. The suture was pulled to cinch the tricuspid annulus. The tricuspid annulus area was measured from images taken at each increment, and the corresponding force was recorded. The hearts were tested for three conditions: (i) non-pressurized (NP); (ii) pressurized (P; normal), and (iii) dilated-pressurized (DP; diseased). Leakage data were also collected for pressurized and dilated pressurized conditions. Annulus dilation was created by injecting phenol into the annulus. RESULTS: The maximum annulus dilation obtained was 8.82%, and the maximum cinching force was 0.38 +/- 0.09 N. Leakage was increased by 81.73% from the pressurized to dilated condition. CONCLUSION: The minimal force required to cinch a tricuspid annulus with severe FTR (23.98% dilation) can be approximated to 0.25 N. The required cinching force can play a major role in the long-term durability of the tricuspid annuloplasty.

Imprinting at the PLAGL1 domain is contained within a 70-kb CTCF/cohesin-mediated non-allelic chromatin loop.

Paternal duplications of chromosome 6q24, a region that contains the imprinted PLAGL1 and HYMAI transcripts, are associated with transient neonatal diabetes mellitus. A common feature of imprinted genes is that they tend to cluster together, presumably as a result of sharing common cis-acting regulatory elements. To determine the extent of this imprinted cluster in human and mouse, we have undertaken a systematic analysis of allelic expression and DNA methylation of the genes mapping within an ∼1.4-Mb region flanking PLAGL1/Plagl1. We confirm that all nine neighbouring genes are biallelically expressed in both species. In human we identify two novel paternally expressed PLAGL1 coding transcripts that originate from unique promoter regions. Chromatin immunoprecipitation for CTCF and the cohesin subunits RAD21 and SMC3 reveals evolutionarily conserved binding sites within unmethylated regions ∼5 kb downstream of the PLAGL1 differentially methylated region and within the PLAGL1 3' untranslated region (UTR). Higher-order chromatin looping occurs between these regions in both expressing and non-expressing tissues, forming a non-allelic chromatin loop around the PLAGL1/Plagl1 gene. In placenta and brain tissues, we identify an additional interaction between the PLAGL1 P3/P4 promoters and the unmethylated element downstream of the PLAGL1 differentially methylated region that we propose facilitates imprinted expression of these alternative isoforms.

The predictive power of family history measures of alcohol and drug problems and internalizing disorders in a college population.

A family history (FH) of psychiatric and substance use problems is a potent risk factor for common internalizing and externalizing disorders. In a large web-based assessment of mental health in college students, we developed a brief set of screening questions for a FH of alcohol problems (AP), drug problems (DP) and depression-anxiety in four classes of relatives (father, mother, aunts/uncles/grandparents, and siblings) as reported by the student. Positive reports of a history of AP, DP, and depression-anxiety were substantially correlated within relatives. These FH measures predicted in the student, in an expected pattern, dimensions of personality and impulsivity, alcohol consumption and problems, smoking and nicotine dependence, use of illicit drugs, and symptoms of depression and anxiety. Using the mean score from the four classes of relatives was more predictive than using a familial/sporadic dichotomy. Interactions were seen between the FH of AP, DP, and depression-anxiety and peer deviance in predicting symptoms of alcohol and tobacco dependence. As the students aged, the FH of AP became a stronger predictor of alcohol problems. While we cannot directly assess the validity of these FH reports, the pattern of findings suggest that our brief screening items were able to assess, with some accuracy, the FH of substance misuse and internalizing psychiatric disorders in relatives. If correct, these measures can play an important role in the creation of developmental etiologic models for substance and internalizing psychiatric disorders which constitute one of the central goals of the overall project.

Human imprinted retrogenes exhibit non-canonical imprint chromatin signatures and reside in non-imprinted host genes.

Imprinted retrotransposed genes share a common genomic organization including a promoter-associated differentially methylated region (DMR) and a position within the intron of a multi-exonic 'host' gene. In the mouse, at least one transcript of the host gene is also subject to genomic imprinting. Human retrogene orthologues are imprinted and we reveal that human host genes are not imprinted. This coincides with genomic rearrangements that occurred during primate evolution, which increase the separation between the retrogene DMRs and the host genes. To address the mechanisms governing imprinted retrogene expression, histone modifications were assayed at the DMRs. For the mouse retrogenes, the active mark H3K4me2 was associated with the unmethylated paternal allele, while the methylated maternal allele was enriched in repressive marks including H3K9me3 and H4K20me3. Two human retrogenes showed monoallelic enrichment of active, but not of repressive marks suggesting a partial uncoupling of the relationship between DNA methylation and repressive histone methylation, possibly due to the smaller size and lower CpG density of these DMRs. Finally, we show that the genes immediately flanking the host genes in mouse and human are biallelically expressed in a range of tissues, suggesting that these loci are distinct from large imprinted clusters.

Racial discrimination and depressive symptoms mediated by conversations about race among students of color.

Racial discrimination is associated with adverse mental health outcomes among Students of Color. In order to address racial tensions, it is important to consider students' dialogues about race. The current study tested whether having positive and negative conversations about one's ethnic-racial group mediated the relation between racial discrimination at T1 and depressive symptoms 5 months later at T2 among 94 college Students of Color. Findings indicated that greater racial discrimination at T1 was associated with more frequent negative conversations about race at T2 (b = .38, p = .00), which was, in turn, associated with greater depressive symptoms at T2 (b = 2.73, p = .04); this pathway demonstrated significant mediation. However, positive conversations about race was not a significant mediator in this association. The current study highlights the importance of focusing on racial conversations after racial discrimination in order to minimize adverse effects on mental health among Students of Color.

Is pre-college interpersonal trauma associated with cannabis use?

OBJECTIVE: To examine the prevalence and correlates of lifetime cannabis use (i.e., experimental [use 1-5 times] and non-experimental [use ≥ 6 times]) in relation to interpersonal trauma (IPT) above and beyond relevant covariates. PARTICIPANTS: A large (n = 9,889) representative sample of college students at an urban university in the southeastern part of the United States. METHODS: Participants were 4 cohorts of first-year college students who completed measures of demographics, cannabis, alcohol, nicotine, and IPT. Associations were estimated using multinomial logistic regressions. RESULTS: The prevalence of lifetime cannabis use was 28.1% and 17.4% for non-experimental and experimental cannabis use, respectively. IPT was significantly associated with experimental and non-experimental cannabis use above and beyond effects of sex, race, cohort, alcohol, and nicotine. CONCLUSIONS: Results show that cannabis use is prevalent among college students and is associated with IPT above and beyond associations with sex, race, and other substance use.

A longitudinal mediational investigation of risk pathways among cannabis use, interpersonal trauma exposure, and trauma-related distress.

OBJECTIVE: College students are at high risk for cannabis use, interpersonal trauma (IPT) exposure, and trauma-related distress (TRD). Two phenotypic etiologic models posited to explain associations between cannabis use and trauma-related phenotypes are the self-medication (trauma/TRD → cannabis use) and high-risk (cannabis use → trauma/TRD) hypotheses. The primary objective of the present study was to investigate direct and indirect associations among cannabis use, IPT exposure, and TRD above and beyond established covariates. METHOD: The current study used data from the first assessment (i.e., baseline survey at Year 1 Fall) and two follow-up assessments (i.e., Year 1 Spring and Year 2 Spring) from an ongoing longitudinal study on college behavioral health. Participants were 4 cohorts of college students (n = 9,889) who completed measures of demographics, substance use, IPT, and TRD. Indirect effects of IPT on cannabis through TRD (i.e., self-medication) and cannabis on TRD through IPT (i.e., high-risk), including tests of covariate effects (e.g., gender, age, race, cohort, alcohol, nicotine), were simultaneously estimated using a longitudinal mediation modeling framework. RESULTS: Results suggest that more IPT exposure increases risk for TRD and subsequent nonexperimental (use 6+ times) cannabis use, and that experimental (use 1-5 times) and nonexperimental cannabis use increases risk for IPT exposure and subsequent TRD. CONCLUSIONS: Both the self-medication and high-risk hypotheses were supported. Findings support a bidirectional causal relationship between cannabis use and trauma-related phenotypes. Additionally, results highlight areas for colleges to intervene among students to help reduce cannabis use and create a safer environment. (PsycInfo Database Record (c) 2023 APA, all rights reserved).

Mental health disorder symptom prevalence and rates of help-seeking among University-Enrolled, emerging adults.

Objective. Examine mental health symptom prevalence and rates of campus services utilization among Black male, White male and Black female college students. Participants. 2500 students from an ongoing, student survey at a public university; launched in 2011. Methods. Measures included data for anxiety and depressive symptoms and utilization of campus health services (counseling center, health services, etc.). Descriptive analyses determined prevalence and utilization rates. Mann Whitney U tests compared prevalence. Chi-squared tests compared utilization rates. Results. Anxiety prevalence: greater than 60% of students from each ethnic group reported symptoms; reporting rates decreased significantly for Black men (49.6%); p < 0.001. Depression prevalence: greater than 80% reported symptoms; there were significant differences in reporting between Black men and Black women (72.7% vs. 87.1%, p < 0.001). Utilization: Black men utilized counseling services less than White men (20.4% vs. 37.8%, p = 0.024). Conclusion. Black men report depressive and anxiety symptoms but underutilize campus health resources.

Risk, Protective, and Associated Factors of Anxiety and Depressive Symptoms and Campus Health Services Utilization Among Black Men on a College Campus.

OBJECTIVE: The aim of this study is to analyze relationships among social and environmental determinants serving as risk, protective, and important covariate factors for mental health risk and help-seeking among Black men on a college campus. METHODS: A secondary data analysis was conducted utilizing an ongoing, campus-wide survey at a large, urban, public university. Measures included depressive and anxiety symptoms; campus service utilization; risk factors (e.g., financial status); protective factors (social support/religiosity); and additional covariates (substance use/GPA). Multiple linear regressions were conducted to examine relationships between these factors, symptoms and help-seeking. RESULTS: Data is included for 681 students. Findings indicated that stressful life events were associated with higher levels of anxiety symptoms (B = 0.39, p < 0.001) and depressive symptoms (B = 0.33, p = 0.013). Cannabis use (B = 1.14, p = .020) was also associated with higher levels of depressive symptoms. We found that financial status (B = 0.21, p = 0.041) was positively associated with higher levels of depressive symptoms and endorsement of religiosity was associated with lower levels anxiety (B = - 0.23, p = 0.019) and depressive symptoms (B = - 0.32, p = 0.035). Religiosity predicted lower utilization of campus health services. CONCLUSIONS: The key findings indicated that Black men's mental health is negatively influenced by stressful live events and cannabis use. As religiosity was associated with lower levels of symptoms and utilization, it may be beneficial to assess this in future work. Further research is needed to address and improve mental health and help-seeking among these men.

Regulatory Programs of B-cell Activation and Germinal Center Reaction Allow B-ALL Escape from CD19 CAR T-cell Therapy.

Chimeric antigen receptor (CAR) T-cell therapy has led to tremendous successes in the treatment of B-cell malignancies. However, a large fraction of treated patients relapse, often with disease expressing reduced levels of the target antigen. Here, we report that exposing CD19+ B-cell acute lymphoblastic leukemia (B-ALL) cells to CD19 CAR T cells reduced CD19 expression within hours. Initially, CD19 CAR T cells caused clustering of CD19 at the T cell-leukemia cell interface followed by CD19 internalization and decreased CD19 surface expression on the B-ALL cells. CD19 expression was then repressed by transcriptional rewiring. Using single-cell RNA sequencing and single-cell assay for transposase-accessible chromatin using sequencing, we demonstrated that a subset of refractory CD19low cells sustained decreased CD19 expression through transcriptional programs of physiologic B-cell activation and germinal center reaction. Inhibiting B-cell activation programs with the Bruton's tyrosine kinase inhibitor ibrutinib increased the cytotoxicity of CD19 CAR T cells without affecting CAR T-cell viability. These results demonstrate transcriptional plasticity as an underlying mechanism of escape from CAR T cells and highlight the importance of combining CAR T-cell therapy with targeted therapies that aim to overcome this plasticity. See related Spotlight by Zhao and Melenhorst, p. 1040.