scholarly journals Limited but gene-class specific correlation of mRNA and protein expression in human CD8+ T cells

2020 ◽  
Author(s):  
Benoît P. Nicolet ◽  
Monika C. Wolkers
Keyword(s):  
Gene Expression ◽  
T Cells ◽  
T Cell ◽  
Protein Expression ◽  
Mrna Expression ◽  
Rna Sequencing ◽  
Protein Translation ◽  
Secreted Proteins ◽  
Sequencing Analysis ◽  
Protein Output

SUMMARYT cell differentiation and activation induces substantial alterations in gene expression. While RNA sequencing and single cell RNA sequencing analysis provided important insights in the gene expression dynamics of T cells, it is not well understood how the mRNA expression translates into the protein landscape. By combining paired RNA-sequencing and mass spectrometry data of primary human CD8+ T cells, we found that mRNA expression is a poor proxy for the overall protein output. Irrespective of the differentiation or activation status, the correlation coefficient of human CD8+ T cells reached a mere 0.41-0.43. Only gene classes that mediate conserved cellular processes such as protein translation or cellular metabolism showed a strong correlation of mRNA with protein expression. In contrast, the mRNA expression and protein output of transcription factors, cell surface molecules, and secreted proteins - including cytokines - only mildly correlated. Conversely, highly conserved genes correlated well with the protein output. This was also true for the presence of AU-rich elements in the 3’untranslated region, in particular for mRNAs that encode secreted proteins. In conclusion, the in-depth characterization of the transcriptome and proteome in human CD8+ T cells emphasizes the need of combined mRNA and protein analysis for our understanding of T cell biology and function.

scholarly journals Genotype-Phenotype Correlation of T Cell Subtypes Reveals Senescent and Cytotoxic Genes in Alzheimer's Disease

2021 ◽  
Author(s):  
Dallin Dressman ◽  
Thomas Buttrick ◽  
Maria Cimpean ◽  
David Bennett ◽  
Vilas Menon ◽  
...  
Keyword(s):  
Gene Expression ◽  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
T Cells ◽  
T Cell ◽  
Rna Sequencing ◽  
Disease Risk ◽  
Synaptic Function ◽  
T Cell Subsets ◽  
Age Related

Recent studies identifying expression quantitative trait loci (eQTL) in immune cells have uncovered important links between disease risk alleles and gene expression trends in monocytes, T cells, and other cell types. However, these studies are generally done with young, healthy subjects, limiting the utility of their findings for age-related conditions such as Alzheimer's disease (AD). We have performed RNA sequencing on four T cell subsets in genome-wide genotyped and well-characterized AD subjects and age- and sex-matched healthy controls from the Religious Orders Study/Memory and Aging Project. Correlating gene expression data with AD neuropathological traits, and with single nucleotide polymorphisms (SNPs) to detect eQTLs, we identified several significant genes involved in T cell senescence and cytotoxicity, consistent with T cell RNA sequencing studies in aged/AD cohorts. We identified unexpected eQTLs previously associated with neuropsychiatric disease traits. Finally, we discovered that pathways related to axon guidance and synaptic function were enriched among trans-eQTLs in coding regions of the genome. Overall, our data sheds more light on the genetic basis behind phenotypic changes in T cells during aging and AD.

scholarly journals Hierarchy of signaling thresholds downstream of the T cell receptor and the Tec kinase ITK

2021 ◽  
Vol 118 (35) ◽  
pp. e2025825118
Author(s):  
Michael P. Gallagher ◽  
James M. Conley ◽  
Pranitha Vangala ◽  
Manuel Garber ◽  
Andrea Reboldi ◽  
...  
Keyword(s):  
Gene Expression ◽  
T Cells ◽  
T Cell ◽  
T Cell Receptor ◽  
Signal Strength ◽  
Cell Receptor ◽  
Early Gene ◽  
Sequencing Analysis ◽  
Nuclear Factor ◽  
Activated T Cells

The strength of peptide:MHC interactions with the T cell receptor (TCR) is correlated with the time to first cell division, the relative scale of the effector cell response, and the graded expression of activation-associated proteins like IRF4. To regulate T cell activation programming, the TCR and the TCR proximal interleukin-2–inducible T cell kinase (ITK) simultaneously trigger many biochemically separate signaling cascades. T cells lacking ITK exhibit selective impairments in effector T cell responses after activation, but under the strongest signaling conditions, ITK activity is dispensable. To gain insight into whether TCR signal strength and ITK activity tune observed graded gene expression through the unequal activation of distinct signaling pathways, we examined Erk1/2 phosphorylation or nuclear factor of activated T cells (NFAT) and nuclear factor (NF)-κB translocation in naïve OT-I CD8+ cell nuclei. We observed the consistent digital activation of NFAT1 and Erk1/2, but NF-κB displayed dynamic, graded activation in response to variation in TCR signal strength, tunable by treatment with an ITK inhibitor. Inhibitor-treated cells showed the dampened induction of AP-1 factors Fos and Fosb, NF-κB response gene transcripts, and survival factor Il2 transcripts. ATAC sequencing analysis also revealed that genomic regions most sensitive to ITK inhibition were enriched for NF-κB and AP-1 motifs. Specific inhibition of NF-κB during peptide stimulation tuned the expression of early gene products like c-Fos. Together, these data indicate a key role for ITK in orchestrating the optimal activation of separate TCR downstream pathways, specifically aiding NF-κB activation. More broadly, we revealed a mechanism by which variations in TCR signal strength can produce patterns of graded gene expression in activated T cells.

scholarly journals SAT0351 CHEMOKINE PATHWAYS ARE ENRICHED IN PSORIATIC ARTHRITIS (PSA) SKIN LESIONS WITH INCREASED EXPRESSION OF ATYPICAL CHEMOKINE RECEPTOR 2 (ACKR2)

2020 ◽  
Vol 79 (Suppl 1) ◽  
pp. 1121.2-1122
Author(s):  
H. Johnsson ◽  
J. Cole ◽  
G. Wilson ◽  
M. Pingen ◽  
F. Mcmonagle ◽  
...  
Keyword(s):  
Gene Expression ◽  
Protein Expression ◽  
Mrna Expression ◽  
Rna Sequencing ◽  
Chemokine Receptor ◽  
Skin Lesions ◽  
Mrna Levels ◽  
Research Support ◽  
Uninvolved Skin ◽  
Healthy Control

Background:Skin in people with psoriasis has been comprehensively studied; uninvolved skin has abnormal gene expression. Less is known specifically about skin in PsA, the assumption being that it is identical to psoriasis. Chemokines and ACKR2 are among the upregulated genes in uninvolved psoriasis compared to healthy skin[1]. ACKR2 is a scavenging receptor of inflammatory CC chemokines and has been proposed as a regulator of cutaneous inflammation in psoriasis. It has not been studied in PsA.Objectives:To compare the transcriptome of PsA lesional, PsA uninvolved and healthy control skin and evaluate ACKR2 expression in PsA.Methods:Biopsies were taken from healthy control (HC) skin and paired lesional and uninvolved skin from patients with PsA. Libraries for bulk RNA sequencing were prepared from polyA selected RNA and sequenced on NovaSeq 6000. Sequencing data were analysed using Searchlight2. ACKR2 mRNA expression was validated by qPCR. RNAscope was used to localise ACKR2 expressing cells and sections were co-stained with podaplanin or stained in serial sections with CD45. Chemokine protein expression in skin was evaluated using Luminex technology.Results:Nine HC and 9 paired skin samples from patients with PsA were sequenced. The PsA skin lesions (PsA L) formed a distinct population in the transcriptomic principal component analysis (PCA) plot while HC and PsA uninvolved skin (PsA U) were overlapping. Only 15 genes were differentially expressed between HC and PsA U and none coded for chemokines. There were however significantly upregulated chemokines and receptors in PsA L. Unexpectely, ACKR2 was the 2ndmost upregulated chemokine receptor in PsA L with unchanged expression in PsA U compared with HC (PsA L vs HC log2fold 3.38, p.adj=9.51E-41; PsA L vs PsA U log2fold 3.58, p.adj=3.24E-45; PsA U vs HC log2fold -0.2, p.adj=0.732).The upregulation of ACKR2 in PsA L and unchanged expression in PsA U was confirmed by qPCR. RNAscope demonstrated strong expression of ACKR2 in the suprabasal layer of the epidermis in PsA L. In HC and PsA U, only occasional ACKR2 positive cells were seen in the epidermis. ACKR2 was expressed in lymphatic vessel walls but was not observed in CD45+ leukocytes.Provisional skin chemokine protein expression data showed poor correlation between mRNA levels and protein expression for the ACKR2 ligands CCL2, CCL3, CCL7, CCL8, CCL11, CCL13 and CCL22 in HC and PsA U, with negative correlation between ACKR2 mRNA expression and CCL2, CCL8 and CCL11 protein expression. In PsA L, chemokine mRNA correlated with protein expression, but protein expression of chemokine ligands did not correlate with ACKR2 expression.Conclusion:This data set shows expected upregulation of chemokines and their receptors in PsA L but relatively unchanged gene expression in PsA U, which contrasts to previous studies in psoriasis. Notably, this study demonstrates a strong upregulation of ACKR2 in keratinocytes in PsA L, with unchanged expression in PsA U. The RNA expression and preliminary protein data suggest that ACKR2 has little effect on the levels of its ligands in PsA skin lesions. However, this study may have missed local effects of ACKR2 in the epidermis.References:[1]Singh, M.D., et al.,Elevated expression of the chemokine-scavenging receptor D6 is associated with impaired lesion development in psoriasis.Am J Pathol, 2012.181(4): p. 1158-64.Acknowledgments:Funded by the Chief Scientist Office and a private donation to the University of Glasgow. Dr Sabarinadh Chilaka helped to prepare libraries for RNA sequencing.Disclosure of Interests:Hanna Johnsson: None declared, John Cole: None declared, Gillian Wilson: None declared, Marieke Pingen: None declared, Fiona McMonagle: None declared, Susan Holmes: None declared, Iain McInnes Grant/research support from: Bristol-Myers Squibb, Celgene, Eli Lilly and Company, Janssen, and UCB, Consultant of: AbbVie, Bristol-Myers Squibb, Celgene, Eli Lilly and Company, Gilead, Janssen, Novartis, Pfizer, and UCB, Stefan Siebert Grant/research support from: BMS, Boehringer Ingelheim, Celgene, GlaxoSmithKline, Janssen, Novartis, Pfizer, UCB, Consultant of: AbbVie, Boehringer Ingelheim, Janssen, Novartis, Pfizer, UCB, Speakers bureau: AbbVie, Celgene, Janssen, Novartis, Gerard Graham: None declared

scholarly journals Unraveling a Revealing Paradox

2003 ◽  
Vol 197 (12) ◽  
pp. 1709-1719 ◽  
Author(s):  
Remy Bosselut ◽  
Terry I. Guinter ◽  
Susan O. Sharrow ◽  
Alfred Singer
Keyword(s):  
Gene Expression ◽  
T Cells ◽  
T Cell ◽  
Protein Expression ◽  
Cd8 T Cells ◽  
Surface Protein ◽  
Lineage Commitment ◽  
Double Positive ◽  
Mhc I ◽  
Surface Protein Expression

The mechanism by which T cell receptor specificity determines the outcome of the CD4/CD8 lineage decision in the thymus is not known. An important clue is the fact that major histocompatibility complex (MHC)-I–signaled thymocytes paradoxically appear as CD4+8lo transitional cells during their differentiation into CD8+ T cells. Lineage commitment is generally thought to occur at the CD4+8+ (double positive) stage of differentiation and to result in silencing of the opposite coreceptor gene. From this perspective, the appearance of MHC-I–signaled thymocytes as CD4+8lo cells would be due to effects on CD8 surface protein expression, not CD8 gene expression. But contrary to this perspective, this study demonstrates that MHC-I–signaled thymocytes appear as CD4+8lo cells because of transient down-regulation of CD8 gene expression, not because of changes in CD8 surface protein expression or distribution. This study also demonstrates that initial cessation of CD8 gene expression in MHC-I–signaled thymocytes is not necessarily indicative of commitment to the CD4+ T cell lineage, as such thymocytes retain the potential to differentiate into CD8+ T cells. These results challenge classical concepts of lineage commitment but fulfill predictions of the kinetic signaling model.

scholarly journals Molecular determinants and heterogeneity of tissue-resident memory CD8+ T lymphocytes revealed by single-cell RNA sequencing

2020 ◽  
Author(s):  
Nadia S. Kurd ◽  
Zhaoren He ◽  
J. Justin Milner ◽  
Kyla D. Omilusik ◽  
Tiani L. Louis ◽  
...  
Keyword(s):  
Gene Expression ◽  
T Cells ◽  
T Cell ◽  
Single Cell ◽  
Rna Sequencing ◽  
Expression Patterns ◽  
Gene Expression Patterns ◽  
Microbial Infection ◽  
Molecular Determinants ◽  
Single Cell Rna Sequencing

AbstractDuring an immune response to microbial infection, CD8+ T cells give rise to distinct classes of cellular progeny that coordinately mediate clearance of the pathogen and provide long-lasting protection against reinfection, including a subset of non-circulating tissue-resident memory (TRM) cells that mediate potent protection within non-lymphoid tissues. Here, we utilized single-cell RNA-sequencing to examine the gene expression patterns of individual CD8+ T cells in the spleen and small intestine intraepithelial lymphocyte (siIEL) compartment throughout the course of their differentiation in response to viral infection. These analyses revealed previously unknown transcriptional heterogeneity within the siIEL CD8+ T cell population at several states of differentiation, representing functionally distinct TRM cell subsets as well as a subset of TRM cell precursors within the tissue early in infection. Taken together, these findings may inform strategies to optimize CD8+ T cell responses to protect against microbial infection and cancer.One sentence summaryHere, we applied single-cell RNA-sequencing to elucidate the gene expression patterns of individual CD8+ T cells differentiating throughout the course of infection in the spleen and small intestinal epithelium, which revealed previously unidentified molecular determinants of tissue-resident T cell differentiation as well as functional heterogeneity within the tissue-resident CD8+ T cell population.

183 Overcoming immunotherapy resistance in T cell-inflamed lung cancer

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A197-A197
Author(s):  
Brendan Horton ◽  
Brendan Horton ◽  
Duncan Morgan ◽  
Noor Momin ◽  
Vidit Bhandarkar ◽  
...  
Keyword(s):  
Lung Cancer ◽  
T Cells ◽  
T Cell ◽  
Cell Line ◽  
Cancer Patients ◽  
Rna Sequencing ◽  
Mechanistic Study ◽  
Nsclc Cell Line ◽  
Effector Molecule ◽  
Effector Molecules

BackgroundTumor infiltrating T cells (TIL) are highly correlated with response to checkpoint blockade immunotherapy (CBT) in melanoma. However, in non-small cell lung cancer (NSCLC), 61% of patients have TIL, but only 32% respond to CBT. It is unknown how these T cell-inflamed tumors are resistant to CBT. Understanding and overcoming this resistance would greatly increase the number of cancer patients who benefit from CBT.MethodsTo understand lung-specific anti-tumor immune responses, a NSCLC cell line derived from an autochthonous murine lung cancer (KP cell line) was transplanted into syngeneic C57BL/6 mice subcutaneously or intravenously. To study antigen-specific responses, the KP cell line was engineered with SIY and 2C TCR transgenic T cells, which are specific for SIY, were adoptively transferred into tumor-bearing animals.ResultsSubcutaneous KP tumors responded to CBT (aCTLA-4 and aPD-L1) with significant tumor regression while lung KP tumors were CBT resistant. Immunohistochemistry found that this was not due to lack of T cell infiltration, as lung tumors contained 10-fold higher numbers of CD8+ TIL than subcutaneous tumors. Single cell RNA sequencing of TIL uncovered that CD8+ TIL in lung lesions had blunted effector molecule expression that correlated with a lack of IL-2 signaling. Adoptive transfer of naïve, tumor-reactive 2C T cells resulted in equally robust T cell proliferation in both the inguinal and mediastinal lymph nodes (LNs). However, RNA sequencing of adoptively transferred 2C T cells isolated 3-days after transfer from draining LNs identified that T cells activated in the mediastinal LN had reduced levels of IL-2 signaling and blunted effector functions early during priming. Flow cytometry confirmed that T cells primed in the mediastinal LNs did not express CD25, GZMB, or IFN-g, while T cells in inguinal LNs upregulated all three of these effector molecules. Delivery of IL-2 and IL-12 during priming was sufficient to restore effector molecule expression on 2C T cells in mediastinal LNs. Analysis of published patient data identified that a subset of lung cancer patients showed a sizable population of CD8+ TIL with low IL-2 signaling and low expression of effector molecules, including common targets of CBT.ConclusionsImmunotherapy resistance in T cell-inflamed tumors is due to defective CD8+ T cell effector differentiation. IL-2-based therapies could enhance differentiation of functional CD8+ effector T cells and could turn immunotherapy resistant tumors to immunotherapy sensitive tumors. This is the first mechanistic study providing evidence for a distinct type of T cell dysfunction resistant to current CBT.Ethics ApprovalThis study was approved by MIT’s Committee on Animal Care, protocol number 0220-006-23.

scholarly journals Receptor-driven invasion profiles in diffuse intrinsic pontine glioma (DIPG)

2021 ◽  
Author(s):  
Anju Karki ◽  
Noah E Berlow ◽  
Jin-Ah Kim ◽  
Esther Hulleman ◽  
Qianqian Liu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Growth Factor ◽  
Protein Expression ◽  
Cell Viability ◽  
Rna Sequencing ◽  
Cell Invasion ◽  
Growth Factor Receptor ◽  
Diffuse Intrinsic Pontine Glioma ◽  
Sequencing Data ◽  
Pontine Glioma

Abstract Background Diffuse intrinsic pontine glioma (DIPG) is a devastating pediatric cancer with unmet clinical need. DIPG is invasive in nature, where tumor cells interweave into the fiber nerve tracts of the pons making the tumor unresectable. Accordingly, novel approaches in combating the disease is of utmost importance and receptor-driven cell invasion in the context of DIPG is under-researched area. Here we investigated the impact on cell invasion mediated by PLEXINB1, PLEXINB2, platelet growth factor receptor (PDGFR)α, PDGFRβ, epithelial growth factor receptor (EGFR), activin receptor 1 (ACVR1), chemokine receptor 4 (CXCR4) and NOTCH1. Methods We used previously published RNA-sequencing data to measure gene expression of selected receptors in DIPG tumor tissue versus matched normal tissue controls (n=18). We assessed protein expression of the corresponding genes using DIPG cell culture models. Then, we performed cell viability and cell invasion assays of DIPG cells stimulated with chemoattractants/ligands. Results RNA-sequencing data showed increased gene expression of receptor genes such as PLEXINB2, PDGFRα, EGFR, ACVR1, CXCR4 and NOTCH1 in DIPG tumors compared to the control tissues. Representative DIPG cell lines demonstrated correspondingly increased protein expression levels of these genes. Cell viability assays showed minimal effects of growth factors/chemokines on tumor cell growth in most instances. Recombinant SEMA4C, SEM4D, PDGF-AA, PDGF-BB, ACVA, CXCL12 and DLL4 ligand stimulation altered invasion in DIPG cells. Conclusions We show that no single growth factor-ligand pair universally induces DIPG cell invasion. However, our results reveal a potential to create a composite of cytokines or anti-cytokines to modulate DIPG cell invasion.

scholarly journals Unique Gene Expression Patterns in Human T-cell Lines Generated from Multiple Sclerosis Patients by Stimulation with a Synthetic MOG Peptide

2005 ◽  
Vol 12 (3) ◽  
pp. 203-209 ◽  
Author(s):  
Mathilda Mandel ◽  
Michael Gurevich ◽  
Gad Lavie ◽  
Irun R. Cohen ◽  
Anat Achiron
Keyword(s):  
Gene Expression ◽  
T Cells ◽  
T Cell ◽  
Cell Lines ◽  
Healthy Subjects ◽  
Expression Patterns ◽  
Gene Expression Pattern ◽  
Gene Expression Patterns ◽  
T Cell Lines ◽  
Myelin Antigens

Multiple sclerosis (MS) is an autoimmune disease where T-cells activated against myelin antigens are involved in myelin destruction. Yet, healthy subjects also harbor T-cells responsive to myelin antigens, suggesting that MS patient-derived autoimmune T-cells might bear functional differences from T-cells derived from healthy individuals. We addressed this issue by analyzing gene expression patterns of myelin oligodendrocytic glycoprotein (MOG) responsive T-cell lines generated from MS patients and healthy subjects. We identified 150 transcripts that were differentially expressed between MS patients and healthy controls. The most informative 43 genes exhibited >1.5-fold change in expression level. Eighteen genes were up-regulated including BCL2, lifeguard, IGFBP3 and VEGF. Twenty five genes were down-regulated, including apoptotic activators like TNF and heat shock protein genes. This gene expression pattern was unique to MOG specific T-cell lines and was not expressed in T-cell lines reactive to tetanus toxin (TTX). Our results indicate that activation in MS that promotes T-cell survival and expansion, has its own state and that the unique gene expression pattern that characterize autoreactive T-cells in MS represent a constellation of factors in which the chronicity, timing and accumulation of damage make the difference between health and disease.

Macrophage-derived CXCL9 and CXCL11, T-cell skin homing and disease control in mogamulizumab-treated CTCL patients

Blood ◽  
2021 ◽  
Author(s):  
Adèle de Masson ◽  
Delphine Darbord ◽  
Gabor Dobos ◽  
Marie Boisson ◽  
Marie Roelens ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Disease Control ◽  
High Throughput Sequencing ◽  
Sequencing Analysis ◽  
Cell Clones ◽  
Skin Homing ◽  
T Cell Homing ◽  
T Cell Immune Responses

Cutaneous T-cell lymphoma (CTCL) is a malignancy of skin-homing T-cells. Long-term remissions are rare in CTCL, and the pathophysiology of long-lasting disease control is unknown. Mogamulizumab is a defucosylated anti-human CCR4 antibody that depletes CCR4-expressing CTCL tumor cells and peripheral blood memory regulatory T cells. Prolonged remissions and immune side effects have been observed in mogamulizumab-treated CTCL patients. We report that mogamulizumab induced skin rashes in 32% of 44 CTCL patients. These rashes were associated with long-term CTCL remission, even in the absence of specific CTCL treatment. CTCL patients with mogamulizumab-induced rash had significantly higher overall survival (hazard ratio, 0.16 (0.04-0.73, p=0.01)). Histopathology and immunohistochemistry of the rashes revealed granulomatous and lichenoid patterns with CD163 macrophagic and CD8 T-cell infiltrates. Depletion of skin CTCL cells was confirmed by high-throughput sequencing analysis of TCRβ genes and in blood by flow cytometry. New reactive T-cell clones were recruited in skin. Gene expression analysis showed overexpression of CXCL9 and CXCL11, two chemokines involved in CXCR3-expressing T-cell homing to skin. Single-cell RNA sequencing analysis in skin of CTCL patients confirmed that CXCL9 and CXCL11 were primarily macrophage-derived and that skin T-cells expressed CXCR3. Finally, patients with rashes had a significantly higher proportion of exhausted reactive blood T-cells expressing TIGIT and PD1 at baseline compared to patients without rash, which decreased under mogamulizumab treatment, consistent with an activation of the antitumor immunity. Together, these data suggest that mogamulizumab may induce long-term immune control in CTCL patients by activation of the macrophagic and T-cell immune responses.

Abstract W P73: Feasibility and Preliminary Results of Whole Blood RNA-Sequencing Analysis in Patients With Intracranial Aneurysms

Stroke ◽  
2014 ◽  
Vol 45 (suppl_1) ◽  
Author(s):  
Blake Haas ◽  
Nestor R Gonzalez ◽  
Elina Nikkola ◽  
Mark Connolly ◽  
William Hsu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Network Analysis ◽  
Rna Sequencing ◽  
Whole Blood ◽  
Intracranial Aneurysms ◽  
Small Sample Size ◽  
Expression Patterns ◽  
Small Sample ◽  
Cellular Respiration ◽  
Sequencing Analysis

Introduction: Intracranial aneurysms (IA) growth and rupture have been associated with chronic remodeling of the arterial wall. However, the pathobiology of this process remains poorly understood. The objective of the present study was to evaluate the feasibility of analyzing gene expression patterns in peripheral blood of patients with ruptured and unruptured saccular IAs. Materials and Methods: We analyzed human whole blood transcriptomes by performing paired-end, 100 bp RNA-sequencing (RNAseq) using the Illumina platform. We used STAR to align reads to the genome, HTSeq to count reads, and DESeq to normalize counts across samples. Self-reported patient information was used to correct expression values for ancestry, age, and sex. We utilized weighted gene co-expression network analysis (WGCNA) to identify gene expression network modules associated with IA size and rupture. The DAVID tool was employed to search for Gene Ontology enrichment in relevant modules. Results: Samples from 12 patients (9 females, age 57.6 +/-12) with IAs were analyzed. Four had ruptured aneurysms. RNA isolation and application of the methodology described above was successful in all samples. Although the small sample size prevents us from drawing definite conclusions, we observed promising novel co-expression networks for IAs: WCGNA analysis showed down-regulation of two transcript modules associated with ruptured IA status (r=-0.78, p=0.008 and r=-0.77, p=0.009), and up-regulation of two modules associated with aneurysm size (r=0.86, p=0.002 and r=0.9, p=4e-04), respectively. DAVID analyses showed that genes upregulated in an IA size-associated module were enriched with genes involved in cellular respiration and translation, while genes involved in transcription were down-regulated in a module associated with ruptured IAs. Conclusions: Whole blood RNAseq analysis is a feasible tool to capture transcriptome dynamics and achieve a better understanding of the pathophysiology of IAs. Further longitudinal studies of patients with IAs using network analysis are justified.

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