Monocyte-endothelial interactions as a targetable node in clonal hematopoiesis-mediated cardiovascular disease
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Curated by eLife
eLife Assessment
Clonal hematopoiesis of indeterminate potential (CHIP) is a known risk factor for coronary artery disease, though its precise role in disease progression continues to emerge. This study leverages valuable single-cell RNA data from patients with CHIP mutations and controls to predict key interactions between endothelial cells and monocytes. Using an AI prediction model, the authors identify druggable targets that mediate immune cell interactions in CHIP and provide solid evidence to support their findings.
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Abstract
Background
Clonal hematopoiesis of indeterminate potential (CHIP) increases risk of cardiovascular disease yet the molecular mechanisms driving this association remain incompletely understood. We hypothesized that aberrant monocyte-endothelial interactions contribute to CHIP-mediated cardiovascular disease.
Methods
We performed single-cell RNA sequencing on blood and peripheral vascular tissue from 4 individuals with TET2 CHIP, 6 individuals with DNMT3A CHIP, and 25 controls. We predicted interactions between monocytes and endothelial cells based on expression of ligand-receptor pairs then modeled interactions between monocytes carrying CHIP mutations with endothelial cells in vitro. We performed an in silico genomewide perturbation screen to identify genetic targets capable of mediating these interactions and then experimentally evaluated the effect of inhibiting predicted targets on monocyte-endothelial interactions in vitro.
Results
Expression of ligand-receptor pairs on monocytes and endothelial cells from patients with and without CHIP highlighted differences in signaling likelihood for 6 key ligand-receptor pairs related to transendothelial migration. Co-culture of monocytes with human aortic endothelial cells demonstrated that monocytes carrying CHIP mutations have decreased velocity compared to monocytes without CHIP mutations. The perturbation screen suggested 11 druggable genetic targets capable of rescuing TET2 CHIP monocytes. Experimental inhibition of ICAM1 in endothelial cells and inhibition of CXCR2 in monocytes significantly increased the velocity of TET2-mutated monocytes over endothelial cells.
Conclusions
CHIP mutations alter interactions between monocytes and endothelial cells. Therapeutics targeting CXCR2 and ICAM1 may restore normal interactions between monocytes and endothelial cells among patients with TET2 CHIP.
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eLife Assessment
Clonal hematopoiesis of indeterminate potential (CHIP) is a known risk factor for coronary artery disease, though its precise role in disease progression continues to emerge. This study leverages valuable single-cell RNA data from patients with CHIP mutations and controls to predict key interactions between endothelial cells and monocytes. Using an AI prediction model, the authors identify druggable targets that mediate immune cell interactions in CHIP and provide solid evidence to support their findings.
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Reviewer #1 (Public review):
Summary:
Using single-cell RNA sequencing and bioinformatics approaches, the authors aimed to discover if and how cells carrying mutations common to clonal haematopoiesis were more adherent to endothelial cells.
Strengths:
(1) The authors used matched blood and adipose tissue samples from the same patients (with the exception of the control people) to conduct their analysis.
(2) The use of bioinformatics and in-silico approaches helped to fast-track their aims to test specific inhibitors in their model cell adhesion system.
Weaknesses:
(1) The analysis was done on pooled cells; it would have been interesting to know if the same adhesion gene signatures were observed across the donors.
(2) The adhesion assays were conducted under static conditions; shear flow adhesion experiments would have been better. Mixed …
Reviewer #1 (Public review):
Summary:
Using single-cell RNA sequencing and bioinformatics approaches, the authors aimed to discover if and how cells carrying mutations common to clonal haematopoiesis were more adherent to endothelial cells.
Strengths:
(1) The authors used matched blood and adipose tissue samples from the same patients (with the exception of the control people) to conduct their analysis.
(2) The use of bioinformatics and in-silico approaches helped to fast-track their aims to test specific inhibitors in their model cell adhesion system.
Weaknesses:
(1) The analysis was done on pooled cells; it would have been interesting to know if the same adhesion gene signatures were observed across the donors.
(2) The adhesion assays were conducted under static conditions; shear flow adhesion experiments would have been better. Mixed cultures using cell trackers would have been even better.
(3) In the intervention studies, the authors should have directly targeted the monocytes (not the endothelial cells) and should have also included DNMT3A mutant/KO cells to show specificity to TET2 CHIP.
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Reviewer #2 (Public review):
Summary:
The authors describe potential mechanisms underlying the changes in endothelial-monocyte interactions in patients with clonal hematopoiesis of indeterminate potential (CHIP), including reduced velocity and increased ligand interactions of CHIP-mutated monocytes. They use a combination of transcriptomics (some for the first time in these tissues in patients with CHIP), in silico analyses, and ex vivo approaches to outline the changes that occur in blood monocytes derived from patients with CHIP. These findings advance the current field, which has previously mostly used mice and/or has been focused on cancer outcomes. The authors identify distinct alterations in signaling downstream of DNTM3A or TET2 mutations, which further distinguish two major mutations that contribute to CHIP.
Strengths:
(1) …
Reviewer #2 (Public review):
Summary:
The authors describe potential mechanisms underlying the changes in endothelial-monocyte interactions in patients with clonal hematopoiesis of indeterminate potential (CHIP), including reduced velocity and increased ligand interactions of CHIP-mutated monocytes. They use a combination of transcriptomics (some for the first time in these tissues in patients with CHIP), in silico analyses, and ex vivo approaches to outline the changes that occur in blood monocytes derived from patients with CHIP. These findings advance the current field, which has previously mostly used mice and/or has been focused on cancer outcomes. The authors identify distinct alterations in signaling downstream of DNTM3A or TET2 mutations, which further distinguish two major mutations that contribute to CHIP.
Strengths:
(1) Combinatorial transcriptomics was used to identify potential therapeutic targets, which is an important proof-of-concept for multiple fields.
(2) The authors identify distinct ligand interactions downstream of TET2 and DNMT3A mutations.
Weaknesses:
(1) The authors extrapolate findings in adipose tissue in diabetic patients to vascular disease (ostensibly in the carotid or cardiac arteries), citing the difficulty of using tissue-matched samples. Broad-reaching conclusions need to be backed up in the relevant systems, considering how different endothelial cells in various vascular beds react. Considering these data were obtained with n=3 patients being sufficient to identify these changes, it seems that this can be performed (perhaps in silico) in the correct tissue.
(2) The selection/exclusion criteria for the diabetes samples are not noted, and therefore, the relevant conclusions cannot be fully evaluated, nor is the source of adipose tissue stated.
Appraisal:
While authors describe how to as well as the technical feasibility of integrating a number of transcriptomic techniques, they do not seem to do so to produce highly compelling data or targets within this manuscript. The potential is there to drill down to mechanisms; however, the data gathered herein do not highlight novel targets. For example, CXCL2 and 3 are already shown to be differentially expressed in TET2 loss combined with LDL treatment in the macrophages of mice. Furthermore, these authors then show that in humans, the prototypical CXC chemokine, IL8 (which mice lack), is significantly higher in TET2-mutated patients (DOI: 10.1056/NEJMoa1701719). The authors should demonstrate the utility of their transcriptomics by identifying and testing novel targets and focusing on the proper disease states. This could easily be a deep dive into CHIP in adipose tissue in diabetic patients.
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