Longitudinal study of DNA methylation and epigenetic clocks prior to and following test-confirmed COVID-19 and mRNA vaccination

AbstractThe host epigenetic landscape is rapidly changed during SARS-CoV-2 infection and evidence suggests that severe COVID-19 is associated with durable scars to the epigenome. Specifically, aberrant DNA methylation changes in immune cells and alterations to epigenetic clocks in blood relate to severe COVID-19. However, a longitudinal assessment of DNA methylation states and epigenetic clocks in blood from healthy individuals prior to and following test-confirmed non-hospitalized COVID-19 has not been performed. Moreover, the impact of mRNA COVID-19 vaccines upon the host epigenome remains understudied. Here, we first examined DNA methylation states in blood of 21 participants prior to and following test confirmed COVID-19 diagnosis at a median timeframe of 8.35 weeks. 261 CpGs were identified as differentially methylated following COVID-19 diagnosis in blood at an FDR adjusted P value <0.05. These CpGs were enriched in gene body and northern and southern shelf regions of genes involved in metabolic pathways. Integrative analysis revealed overlap among genes identified in transcriptional SARS-CoV-2 infection datasets. Principal component-based epigenetic clock estimates of PhenoAge and GrimAge significantly increased in people over 50 following infection by an average of 2.1 and 0.84 years. In contrast, PCPhenoAge significantly decreased in people under 50 following infection by an average of 2.06 years. This observed divergence in epigenetic clocks following COVID-19 was related to age and immune cell-type compositional changes in CD4+ T cells, B cells, granulocytes, plasmablasts, exhausted T cells, and naïve T cells. Complementary longitudinal epigenetic clock analyses of 36 participants prior to and following Pfizer and Moderna mRNA-based COVID-19 vaccination revealed vaccination significantly reduced principal component-based Horvath epigenetic clock estimates in people over 50 by an average of 3.91 years for those that received Moderna. This reduction in epigenetic clock estimates was significantly related to chronological age and immune cell-type compositional changes in B cells and plasmablasts pre- and post-vaccination. These findings suggest the potential utility of epigenetic clocks as a biomarker of COVID-19 vaccine responses. Future research will need to unravel the significance and durability of short-term changes in epigenetic age related to COVID-19 exposure and mRNA vaccination.

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Immune classification of clear cell renal cell carcinoma

Scientific Reports ◽

10.1038/s41598-021-83767-z ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Sumeyye Su ◽

Shaya Akbarinejad ◽

Leili Shahriyari

Keyword(s):

T Cells ◽

Cd8 T Cells ◽

Renal Cell ◽

Cd4 T Cells ◽

Immune Cell ◽

Clear Cell ◽

P Value ◽

Cell Type ◽

Primary Tumors ◽

Immune Cell Type

AbstractSince the outcome of treatments, particularly immunotherapeutic interventions, depends on the tumor immune micro-environment (TIM), several experimental and computational tools such as flow cytometry, immunohistochemistry, and digital cytometry have been developed and utilized to classify TIM variations. In this project, we identify immune pattern of clear cell renal cell carcinomas (ccRCC) by estimating the percentage of each immune cell type in 526 renal tumors using the new powerful technique of digital cytometry. The results, which are in agreement with the results of a large-scale mass cytometry analysis, show that the most frequent immune cell types in ccRCC tumors are CD8+ T-cells, macrophages, and CD4+ T-cells. Saliently, unsupervised clustering of ccRCC primary tumors based on their relative number of immune cells indicates the existence of four distinct groups of ccRCC tumors. Tumors in the first group consist of approximately the same numbers of macrophages and CD8+ T-cells and and a slightly smaller number of CD4+ T cells than CD8+ T cells, while tumors in the second group have a significantly high number of macrophages compared to any other immune cell type (P-value $$<0.01$$ < 0.01 ). The third group of ccRCC tumors have a significantly higher number of CD8+ T-cells than any other immune cell type (P-value $$<0.01$$ < 0.01 ), while tumors in the group 4 have approximately the same numbers of macrophages and CD4+ T-cells and a significantly smaller number of CD8+ T-cells than CD4+ T-cells (P-value $$<0.01$$ < 0.01 ). Moreover, there is a high positive correlation between the expression levels of IFNG and PDCD1 and the percentage of CD8+ T-cells, and higher stage and grade of tumors have a substantially higher percentage of CD8+ T-cells. Furthermore, the primary tumors of patients, who are tumor free at the last time of follow up, have a significantly higher percentage of mast cells (P-value $$<0.01$$ < 0.01 ) compared to the patients with tumors for all groups of tumors except group 3.

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Epigenetic Quantification of Circulating Immune Cells in Peripheral Blood of Triple-Negative Breast Cancer patients

10.21203/rs.3.rs-508197/v1 ◽

2021 ◽

Author(s):

Mehdi Manoochehri ◽

Thomas Hielscher ◽

Nasim Borhani ◽

Clarissa Gerhäuser ◽

Olivia Fletcher ◽

...

Keyword(s):

Breast Cancer ◽

T Cells ◽

Triple Negative Breast Cancer ◽

Immune Cells ◽

Triple Negative ◽

Immune Cell ◽

Nk Cell ◽

Cell Type ◽

Cell Ratio ◽

Immune Cell Type

Abstract Background: A shift in the proportions of blood immune cells is a hallmark of cancer development. Here, we investigated whether methylation-derived immune cell type ratios and methylation-derived neutrophil-to-lymphocyte ratios (mdNLRs) are associated with triple-negative breast cancer (TNBC). Methods: Leukocyte subtype-specific un/methylated CpG sites were selected and methylation levels at these sites used as proxies for immune cell type proportions and mdNLR estimation in 231 TNBC cases and 231 age-matched controls. Data were validated using the Houseman deconvolution method. Additionally, the natural killer (NK) cell ratio was measured in a prospective sample set of 146 TNBC cases and 146 age-matched controls. Results: The mdNLRs were higher in TNBC cases compared with controls and associated with TNBC (odds ratio (OR) range (2.66-4.29), all Padj.<1e-04). A higher neutrophil ratio and lower ratios of NK cells, CD4+ T cells, CD8+ T cells, monocytes, and B cells were associated with TNBC. The strongest association was observed with decreased NK cell ratio (OR range (1.28-1.42), all Padj.<1e-04). The NK cell ratio was also significantly lower in pre-diagnostic samples of TNBC cases compared with controls (P=0.019).Conclusion: This immunomethylomic study shows that a shift in the ratios/proportions of leukocyte subtypes is associated with TNBC, with decreased NK cell showing the strongest association. These findings improve our knowledge of the role of the immune system in TNBC and point to the possibility of using NK cell level as a non-invasive molecular marker for TNBC risk assessment, early detection, and prevention.

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Depression, Anxiety, and Social Environmental Adversity as Potential Modulators of the Immune Tumor Microenvironment in Breast Cancer Patients

Medical Sciences ◽

10.3390/medsci9020046 ◽

2021 ◽

Vol 9 (2) ◽

pp. 46

Author(s):

Eida M. Castro-Figueroa ◽

Karina I. Acevedo ◽

Cristina I. Peña-Vargas ◽

Normarie Torres-Blasco ◽

Idhaliz Flores ◽

...

Keyword(s):

Breast Cancer ◽

T Cells ◽

B Cells ◽

Tumor Microenvironment ◽

Immune Cell ◽

Depressive Symptomatology ◽

Anxiety Symptoms ◽

Trauma History ◽

Social Environmental ◽

The Impact

Background: Mounting data suggest that exposure to chronic stress is associated with worse breast cancer outcomes. This study aimed to explore the impact of social environmental adversity (SEA, e.g., child abuse, crime, sexual, and physical violence), depressive symptomatology, and anxiety on immune cell infiltration into the breast tumor microenvironment. Methods: Participants (n = 33) completed a series of surveys assessing depression and anxiety symptoms, adverse childhood events (ACE), and trauma history. Tumor-associated macrophages (CD68+), B cells (CD19+), and T cells (CD3+) were identified by immunohistochemical analyses of formalin-fixed paraffin-embedded tumor samples and quantified. Spearman rank tests were used to explore the relationships between the variables studied. Results: Exposure to SEA was high (ACE = 72%, exposure to crime = 47%, and exposure to physical/sexual assault = 73%) among participants. Moreover, 30% reported a comorbid history of depression and ACE; 39% reported one or more traumatic events, and clinically significant depression symptomatology, while 21% reported trauma history and significant anxiety symptomatology. Increased tumor-infiltrating B cells were significantly correlated with exposure to crime, anxiety symptoms, and exposure to an ACE. The ACE plus anxiety group presented the highest infiltration of B cells, T cells, and macrophages. Conclusion: These findings support a role for SEA, anxiety symptoms, and depression as potential modulators of the immune tumor microenvironment in breast cancer.

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Epigenetic quantification of circulating immune cells in peripheral blood of triple-negative breast cancer patients

Clinical Epigenetics ◽

10.1186/s13148-021-01196-1 ◽

2021 ◽

Vol 13 (1) ◽

Author(s):

Mehdi Manoochehri ◽

Thomas Hielscher ◽

Nasim Borhani ◽

Clarissa Gerhäuser ◽

Olivia Fletcher ◽

...

Keyword(s):

Breast Cancer ◽

T Cells ◽

Triple Negative Breast Cancer ◽

Immune Cells ◽

Triple Negative ◽

Immune Cell ◽

Nk Cell ◽

Cell Type ◽

Cell Ratio ◽

Immune Cell Type

Abstract Background A shift in the proportions of blood immune cells is a hallmark of cancer development. Here, we investigated whether methylation-derived immune cell type ratios and methylation-derived neutrophil-to-lymphocyte ratios (mdNLRs) are associated with triple-negative breast cancer (TNBC). Methods Leukocyte subtype-specific unmethylated/methylated CpG sites were selected, and methylation levels at these sites were used as proxies for immune cell type proportions and mdNLR estimation in 231 TNBC cases and 231 age-matched controls. Data were validated using the Houseman deconvolution method. Additionally, the natural killer (NK) cell ratio was measured in a prospective sample set of 146 TNBC cases and 146 age-matched controls. Results The mdNLRs were higher in TNBC cases compared with controls and associated with TNBC (odds ratio (OR) range (2.66–4.29), all Padj. < 1e−04). A higher neutrophil ratio and lower ratios of NK cells, CD4 + T cells, CD8 + T cells, monocytes, and B cells were associated with TNBC. The strongest association was observed with decreased NK cell ratio (OR range (1.28–1.42), all Padj. < 1e−04). The NK cell ratio was also significantly lower in pre-diagnostic samples of TNBC cases compared with controls (P = 0.019). Conclusion This immunomethylomic study shows that a shift in the ratios/proportions of leukocyte subtypes is associated with TNBC, with decreased NK cell showing the strongest association. These findings improve our knowledge of the role of the immune system in TNBC and point to the possibility of using NK cell level as a non-invasive molecular marker for TNBC risk assessment, early detection, and prevention.

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Single-cell RNA-Seq reveals transcriptional heterogeneity and immune subtypes associated with disease activity in human myasthenia gravis

Cell Discovery ◽

10.1038/s41421-021-00314-w ◽

2021 ◽

Vol 7 (1) ◽

Author(s):

Wanlin Jin ◽

Qi Yang ◽

Yuyao Peng ◽

Chengkai Yan ◽

Yi Li ◽

...

Keyword(s):

T Cells ◽

B Cells ◽

Myasthenia Gravis ◽

Disease Activity ◽

Single Cell ◽

Immune Cell ◽

Large Cell ◽

Cellular Heterogeneity ◽

T Cell Subsets ◽

The Impact

AbstractMyasthenia gravis (MG) is a rare autoimmune disease. Although the impact of immune cell disorder in MG has been extensively studied, little is known about the transcriptomes of individual cells. Here, we assessed the transcriptional profiles of 39,243 cells by single-cell sequencing and identified 13 major cell clusters, along with 39 subgroups of cells derived from patients with new-onset myasthenia gravis and healthy controls. We found that B cells, CD4+ T cells, and monocytes exhibited more heterogeneity in MG patients. CD4+ T cells were expanded in MG patients. We reclustered B cells and CD4+ T cells, and predict their essential regulators. Further analyses demonstrated that B cells in MG exhibited higher transcriptional activity towards plasma cell differentiation, CD4+ T cell subsets were unbalanced, and inflammatory pathways of monocytes were highly activated. Notably, we discovered a disease-relevant subgroup, CD180− B cells. Increased CD180− B cells in MG are indicative of a high IgG composition and were associated with disease activity and the anti-AChR antibody. Together, our data further the understanding of the cellular heterogeneity involved in the pathogenesis of MG and provide large cell-type-specific markers for subsequent research.

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Immune Classification of Clear Cell Renal Cell Carcinoma

10.1101/2020.07.03.187047 ◽

2020 ◽

Cited By ~ 2

Author(s):

Sumeyye Su ◽

Shaya Akbarinejad ◽

Leili Shahriyari

Keyword(s):

T Cells ◽

Cd8 T Cells ◽

Renal Cell ◽

Cd4 T Cells ◽

Immune Cell ◽

Clear Cell ◽

P Value ◽

Cell Type ◽

Primary Tumors ◽

Immune Cell Type

ABSTRACTSince the outcome of treatments, particularly immunotherapeutic interventions, depends on the tumor immune micro-environment (TIM), several experimental and computational tools such as flow cytometry, immunohistochemistry, and digital cytometry have been developed and utilized to classify TIM variations. In this project, we identify immune pattern of clear cell renal cell carcinomas (ccRCC) by estimating the percentage of each immune cell type in 526 renal tumors using the new powerful technique of digital cytometry. The results, which are in agreement with the results of a large-scale mass cytometry analysis, show that the most frequent immune cell types in ccRCC tumors are CD8+ T-cells, macrophages, and CD4+ T-cells. Saliently, unsupervised clustering of ccRCC primary tumors based on their relative number of immune cells indicates the existence of four distinct groups of ccRCC tumors. Tumors in the first group consist of approximately the same numbers of CD8+ T-cells, CD4+ T-cells, and macrophages, while tumors in the second group have a significantly high number of macrophages compared to any other immune cell type (P-value< 0.01). The third group of ccRCC tumors have a significantly higher number of CD8+ T-cells than any other immune cell type (P-value< 0.01), while tumors in the group 4 have approximately the same numbers of macrophages and CD4+ T-cells and a significantly smaller number of CD8+ T-cells than CD4+ T-cells (P-value< 0.01). Moreover, there is a high positive correlation between the expression levels of IFNG and PDCD1 and the percentage of CD8+ T-cells, and higher stage and grade of tumors have a substantially higher percentage of CD8+ T-cells. Furthermore, the primary tumors of patients, who are tumor free at the last time of follow up, have a significantly higher percentage of mast cells (P-value< 0.01) compared to the patients with tumors for all groups of tumors except group 3.

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Epigenetic Quantification of Circulating Immune Cells in Peripheral Blood of Triple-negative Breast Cancer Patients

10.21203/rs.3.rs-508197/v2 ◽

2021 ◽

Author(s):

Mehdi Manoochehri ◽

Thomas Hielscher ◽

Nasim Borhani ◽

Clarissa Gerhäuser ◽

Olivia Fletcher ◽

...

Keyword(s):

Breast Cancer ◽

T Cells ◽

Triple Negative Breast Cancer ◽

Immune Cells ◽

Triple Negative ◽

Immune Cell ◽

Nk Cell ◽

Cell Type ◽

Cell Ratio ◽

Immune Cell Type

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A Diet High in Lipid Soluble Forms of Choline Modulates Gut-Associated Immune Function in Sprague-Dawley Dams

Current Developments in Nutrition ◽

10.1093/cdn/nzab061_003 ◽

2021 ◽

Vol 5 (Supplement_2) ◽

pp. 1119-1119

Author(s):

Jessy Azarcoya Barrera ◽

Dhruvesh Patel ◽

Catherine Field ◽

Yves Pouliot ◽

Susan Goruk ◽

...

Keyword(s):

T Cells ◽

B Cells ◽

Immune Responses ◽

Oral Tolerance ◽

Immune Cell ◽

Ex Vivo ◽

Control Diet ◽

Sprague Dawley ◽

And Control ◽

The Impact

Abstract Objectives Oral tolerance is the state of unresponsiveness to dietary antigens that are not harmful to the host, a process that begins in the gut and influences both local and peripheral immune responses. Lipid soluble forms of choline have been shown to beneficial affect peripheral immune responses during pregnancy and lactation, yet little is known about the impact of the different choline forms on the gut-associate lymphoid tissue (GALT). Therefore, the objective of this study was to determine the effect of feeding buttermilk-derived choline forms on the GALT in lactating dams. Methods Sprague-Dawley dams were randomized to consume one of three diets, all containing 1.9 g/kg of total choline: 1-Control (100% free choline (FC)), 2-Buttermilk (37% phosphatidylcholine (PC), 34% sphingomyelin, 17% glycerophosphocholine (GPC), 7% FC, 5% phosphocholine) and 3-Placebo (50% PC, 25% FC, 25% GPC). Immune cell phenotypes of mesenteric lymph nodes (MLN) and Peyer's patches (PP) were measured by flow cytometry. Ex vivo cytokine production by immune cells isolated from MLN stimulated with Concanavalin A (ConA) and Ovalbumin (OVA) was measured by ELISA. Results In MLN, dams from the buttermilk and placebo groups had a higher production of IL-10 after both ConA and OVA stimulation and a higher proportion of helper T cells expressing co-inhibitory marker (CD4+CD152+) when compared to the control diet (both P < 0.05). No changes were found in IL-2, TNF-α, or IL-6 production and the proportion of regulatory T cells (CD3+CD4+CD25+FoxP3+) among groups. In PP, dams from the buttermilk group had a higher proportion of CD45RA + cells (B cells) compared to both the placebo and control diets and dams from the placebo diet had a higher proportion of dendritic cells (DC, OX62+OX6+) compared to both the buttermilk and control diets (both P < 0.05). Conclusions Compared to the control diet (only FC), the buttermilk and placebo diets appear to beneficially modulate oral tolerance by increasing the production of IL-10 (an important mediator) and could be partially explain by the higher proportion of CD4+CD152+cells in MLN. In PP, the higher proportion of B cells in the buttermilk and DC in the placebo groups, suggests that dams from these groups may have better antigen presentation function, which may further assist the induction of oral tolerance. Funding Sources Dairy Farmers of Canada, NSERC, CONACYT.

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Cladribine Alters Immune Cell Surface Molecules for Adhesion and Costimulation: Further Insights to the Mode of Action in Multiple Sclerosis

Cells ◽

10.3390/cells10113116 ◽

2021 ◽

Vol 10 (11) ◽

pp. 3116

Author(s):

Tobias Moser ◽

Lena Hoepner ◽

Kerstin Schwenker ◽

Michael Seiberl ◽

Julia Feige ◽

...

Keyword(s):

Multiple Sclerosis ◽

T Cells ◽

B Cells ◽

Cell Surface ◽

Mode Of Action ◽

Immune Cell ◽

Cell Surface Molecules ◽

Surface Molecules ◽

Surface Patterns ◽

The Impact

Cladribine (CLAD) is a deoxyadenosine analogue prodrug which is given in multiple sclerosis (MS) as two short oral treatment courses 12 months apart. Reconstitution of adaptive immune function following selective immune cell depletion is the presumed mode of action. In this exploratory study, we investigated the impact of CLAD tablets on immune cell surface molecules for adhesion (CAMs) and costimulation (CoSs) in people with MS (pwMS). We studied 18 pwMS who started treatment with CLAD and 10 healthy controls (HCs). Peripheral blood mononuclear cells were collected at baseline and every 3 months throughout a 24-month period. We analysed ICAM-1, LFA-1, CD28, HLADR, CD154, CD44, VLA-4 (CD49d/CD29), PSGL-1 and PD-1 with regard to their expression on B and T cells (T helper (Th) and cytotoxic T cells (cT)) and surface density (mean fluorescence intensity, MFI) by flow cytometry. The targeted analysis of CAM and CoS on the surface of immune cells in pwMS revealed a higher percentage of ICAM-1 (B cells, Th, cT), LFA-1 (B cells, cT), HLADR (B cells, cT), CD28 (cT) and CD154 (Th). In pwMS, we found lower frequencies of Th and cT cells expressing PSGL-1 and B cells for the inhibitory signal PD-1, whereas the surface expression of LFA-1 on cT and of HLADR on B cells was denser. Twenty-four months after the first CLAD cycle, the frequencies of B cells expressing CD44, CD29 and CD49d were lower compared with the baseline, together with decreased densities of ICAM-1, CD44 and HLADR. The rate of CD154 expressing Th cells dropped at 12 months. For cT, no changes were seen for frequency or density. Immune reconstitution by oral CLAD was associated with modification of the pro-migratory and -inflammatory surface patterns of CAMs and CoSs in immune cell subsets. This observation pertains primarily to B cells, which are key cells underlying MS pathogenesis.

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Unraveling Tumor-Immune Heterogeneity in Advanced Ovarian Cancer Uncovers Immunogenic Effect of Chemotherapy

10.1101/441428 ◽

2018 ◽

Cited By ~ 2

Author(s):

Alejandro Jiménez-Sánchez ◽

Paulina Cybulska ◽

Katherine Lavigne ◽

Tyler Walther ◽

Ines Nikolovski ◽

...

Keyword(s):

Ovarian Cancer ◽

T Cells ◽

Immune Cell ◽

Metastatic Cancer ◽

Advanced Ovarian Cancer ◽

Cell Type ◽

Immune Microenvironment ◽

Tumor Immune Microenvironment ◽

Treatment Naïve ◽

The Impact

ABSTRACTIn metastatic cancer, the role of heterogeneity at the tumor-immune microenvironment, its molecular underpinnings and clinical relevance remain largely unexplored. To understand tumor-immune dynamics at baseline and upon chemotherapy treatment, we performed unbiased pathway and cell type-specific immunogenomics analysis of treatment-naive (38 samples from 8 patients) and paired chemotherapy treated (80 paired samples from 40 patients) high-grade serous ovarian cancer (HGSOC) samples. Whole transcriptome analysis and image-based quantification of T cells from treatment-naive tumors revealed ubiquitous variability in immune signaling and distinct immune microenvironments co-existing within the same individuals and within tumor deposits at diagnosis. To systematically explore cell type composition of the tumor microenvironment using bulk mRNA, we derived consensus immune and stromal cell gene signatures by intersecting state-of-the-art deconvolution methods, providing improved accuracy and sensitivity when compared to HGSOC immunostaining and leukocyte methylation data sets. Cell-type deconvolution and pathway analyses revealed that Myc and Wnt signaling associate with immune cell exclusion in untreated HGSOC. To evaluate the effect of chemotherapy on the intrinsic tumor-immune heterogeneity, we compared site-matched and site-unmatched tumors before and after neoadjuvant chemotherapy. Transcriptomic and T-cell receptor sequencing analyses showed that site-matched samples had increased cytotoxic immune activation and oligoclonal expansion of T cells after chemotherapy, which was not seen in site-unmatched samples where heterogeneity could not be accounted for. These results demonstrate that the tumor-immune interface in advanced HGSOC is intrinsically heterogeneous, and thus requires site-specific analysis to reliably unmask the impact of therapy on the tumor-immune microenvironment.

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