Adversity in early life and pregnancy are immunologically distinct from total life adversity: macrophage-associated phenotypes in women exposed to interpersonal violence

Early childhood and pregnancy are two sensitive periods of heightened immune plasticity, when exposure to adversity may disproportionately increase health risks. However, we need deeper phenotyping to disentangle the impact of adversity during sensitive periods from that across the total lifespan. This study examined whether retrospective reports of adversity during childhood or pregnancy were associated with inflammatory imbalance, in an ethnically diverse cohort of 53 low-income women seeking family-based trauma treatment following exposure to interpersonal violence. Structured interviews assessed early life adversity (trauma exposure ≤ age 5), pregnancy adversity, and total lifetime adversity. Blood serum was assayed for pro-inflammatory (TNF-a, IL-1ß, IL-6, and CRP) and anti-inflammatory (IL-1RA, IL-4, and IL-10) cytokines. CD14+ monocytes were isolated in a subsample (n = 42) and gene expression assayed by RNA sequencing (Illumina HiSeq 4000; TruSeq cDNA library). The primary outcome was a macrophage-associated M1/M2 gene expression phenotype. To evaluate sensitivity and specificity, we contrasted M1/M2 gene expression with a second, clinically-validated macrophage-associated immunosuppressive phenotype (endotoxin tolerance) and with pro-inflammatory and anti-inflammatory cytokine levels. Adjusting for demographics, socioeconomic status, and psychopathology, higher adversity in early life (ß = .337, p = 0.029) and pregnancy (ß = .332, p = 0.032) were each associated with higher M1/M2 gene expression, whereas higher lifetime adversity (ß = −.341, p = 0.031) was associated with lower immunosuppressive gene expression. Adversity during sensitive periods was uniquely associated with M1/M2 imbalance, among low-income women with interpersonal violence exposure. Given that M1/M2 imbalance is found in sepsis, severe COVID-19 and myriad chronic diseases, these findings implicate novel immune mechanisms underlying the impact of adversity on health.publishedVersio

Empirical assessment of analysis workflows for differential expression analysis of human samples using RNA-Seq

Abstract Background RNA-Seq has supplanted microarrays as the preferred method of transcriptome-wide identification of differentially expressed genes. However, RNA-Seq analysis is still rapidly evolving, with a large number of tools available for each of the three major processing steps: read alignment, expression modeling, and identification of differentially expressed genes. Although some studies have benchmarked these tools against gold standard gene expression sets, few have evaluated their performance in concert with one another. Additionally, there is a general lack of testing of such tools on real-world, physiologically relevant datasets, which often possess qualities not reflected in tightly controlled reference RNA samples or synthetic datasets. Results Here, we evaluate 219 combinatorial implementations of the most commonly used analysis tools for their impact on differential gene expression analysis by RNA-Seq. A test dataset was generated using highly purified human classical and nonclassical monocyte subsets from a clinical cohort, allowing us to evaluate the performance of 495 unique workflows, when accounting for differences in expression units and gene- versus transcript-level estimation. We find that the choice of methodologies leads to wide variation in the number of genes called significant, as well as in performance as gauged by precision and recall, calculated by comparing our RNA-Seq results to those from four previously published microarray and BeadChip analyses of the same cell populations. The method of differential gene expression identification exhibited the strongest impact on performance, with smaller impacts from the choice of read aligner and expression modeler. Many workflows were found to exhibit similar overall performance, but with differences in their calibration, with some biased toward higher precision and others toward higher recall. Conclusions There is significant heterogeneity in the performance of RNA-Seq workflows to identify differentially expressed genes. Among the higher performing workflows, different workflows exhibit a precision/recall tradeoff, and the ultimate choice of workflow should take into consideration how the results will be used in subsequent applications. Our analyses highlight the performance characteristics of these workflows, and the data generated in this study could also serve as a useful resource for future development of software for RNA-Seq analysis

Similarity search in Mass Spectra Databases

Shotgun proteomics is a widely known technique for identification of protein and peptide sequences from an "in vitro" sample. A tandem mass spectrometer generates tens of thousands of mass spectra which must be annotated with peptide sequences. For this purpose, the similarity search in a database of theoretical spectra generated from a database of known protein sequences can be utilized. Since the sizes of databases grow rapidly in recent years, there is a demand for utilization of various database indexing techniques. We investigate the capabilities of (non)metric access methods as the database indexing techniques for fast and approximate similarity retrieval in mass spectra databases. We show that the method for peptide sequences identification is more than 100x faster than a sequential scan over the entire database while more than 90% of spectra are correctly annotated with peptide sequences. Since the method is currently suitable for small mixtures of proteins, we also utilize a precursor mass filter as the database indexing technique for complex mixtures of proteins. The precursor mass filter followed by ranking of spectra by a modification of the parametrized Hausdorff distance outperforms state-of-the-art tools in the number of identified peptide sequences and the speed of search. The..

FCRL5+ Memory B Cells Exhibit Robust Recall Responses

Summary: FCRL5+ atypical memory B cells (atMBCs) expand in many chronic human infections, including recurrent malaria, but studies have drawn opposing conclusions about their function. Here, in mice infected with Plasmodium chabaudi, we demonstrate expansion of an antigen-specific FCRL5+ population that is distinct from previously described FCRL5+ innate-like murine subsets. Comparative analyses reveal overlapping phenotypic and transcriptomic signatures between FCRL5+ B cells from Plasmodium-infected mice and atMBCs from Plasmodium-exposed humans. In infected mice, FCRL5 is expressed on the majority of antigen-specific germinal-center-derived memory B cells (MBCs). Upon challenge, FCRL5+ MBCs rapidly give rise to antibody-producing cells expressing additional atypical markers, demonstrating functionality in vivo. Moreover, atypical markers are expressed on antigen-specific MBCs generated by immunization in both mice and humans, indicating that the atypical phenotype is not restricted to chronic settings. This study resolves conflicting interpretations of atMBC function and suggests FCRL5+ B cells as an attractive target for vaccine strategies. : FCRL5+ atypical memory B cells (MBCs) expand in many chronic human diseases. Using tetramers to track rare antigen-specific cells, Kim et al. show that FCRL5+ MBCs are mature, optimally responsive cells that arise not only in response to infection and protein immunization in mice but also to immunization in humans. Keywords: FCRL5, malaria, atypical memory B cell, memory B cell, age-associated memory B cell, Plasmodium chabaud

Additional file 6: of Trimming of sequence reads alters RNA-Seq gene expression estimates

Influence of minimum length requirements on junction alignment and detection. (a) The average number of reads aligned to junctions per sample with increasing minimum read length requirements after trimming with SolexaQA, Q = 40. (b) The average frequency of reads aligned to junctions (number of reads aligned to junctions per total reads aligned). (c) The average number of junctions detected per sample. (d) The average frequency of junction detection (number of junctions detected per total reads mapped). For all panels, data were normalized to the Q40 value with no minimum length filter, on a per sample basis. Error bars represent standard deviations. (PDF 114 kb

Adversity in early life and pregnancy are immunologically distinct from total life adversity: macrophage-associated phenotypes in women exposed to interpersonal violence

Early childhood and pregnancy are two sensitive periods of heightened immune plasticity, when exposure to adversity may disproportionately increase health risks. However, we need deeper phenotyping to disentangle the impact of adversity during sensitive periods from that across the total lifespan. This study examined whether retrospective reports of adversity during childhood or pregnancy were associated with inflammatory imbalance, in an ethnically diverse cohort of 53 low-income women seeking family-based trauma treatment following exposure to interpersonal violence. Structured interviews assessed early life adversity (trauma exposure ≤ age 5), pregnancy adversity, and total lifetime adversity. Blood serum was assayed for pro-inflammatory (TNF-a, IL-1ß, IL-6, and CRP) and anti-inflammatory (IL-1RA, IL-4, and IL-10) cytokines. CD14+ monocytes were isolated in a subsample (n = 42) and gene expression assayed by RNA sequencing (Illumina HiSeq 4000; TruSeq cDNA library). The primary outcome was a macrophage-associated M1/M2 gene expression phenotype. To evaluate sensitivity and specificity, we contrasted M1/M2 gene expression with a second, clinically-validated macrophage-associated immunosuppressive phenotype (endotoxin tolerance) and with pro-inflammatory and anti-inflammatory cytokine levels. Adjusting for demographics, socioeconomic status, and psychopathology, higher adversity in early life (ß = .337, p = 0.029) and pregnancy (ß = .332, p = 0.032) were each associated with higher M1/M2 gene expression, whereas higher lifetime adversity (ß = −.341, p = 0.031) was associated with lower immunosuppressive gene expression. Adversity during sensitive periods was uniquely associated with M1/M2 imbalance, among low-income women with interpersonal violence exposure. Given that M1/M2 imbalance is found in sepsis, severe COVID-19 and myriad chronic diseases, these findings implicate novel immune mechanisms underlying the impact of adversity on health