E-GEOD-2522 - Transcription profiling of human atherosclerotic lesion SMC cells resistant and sensitive to TGF and Fas ligation by CH11 antibody - OmicsDI

Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Transcription profiling of human atherosclerotic lesion SMC cells resistant and sensitive to TGF and Fas ligation by CH11 antibody

ABSTRACT: Comparison of SMC resistant and sensitive to TGF and Fas ligation by CH11 antibody

ORGANISM(S): Homo sapiens

SUBMITTER: Timothy McCaffrey

PROVIDER: E-GEOD-2522 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Genomic profiling of acquired resistance to apoptosis in cells derived from human atherosclerotic lesions: potential role of STATs, cyclinD1, BAD, and Bcl-XL.

Gagarin Dmitry D Yang Zhaoqing Z Butler Jason J Wimmer Monika M Du Baoheng B Cahan Patrick P McCaffrey Timothy A TA

Journal of molecular and cellular cardiology 20050901 3

Current theories suggest that atherosclerosis, plaque rupture, stroke, and restenosis after angioplasty may involve defective apoptotic mechanisms in vascular cells. Prior work has demonstrated that cells from human atherosclerotic lesions, and cells from the aorta of aged rats, exhibit functional resistance to apoptosis induced by TGF-beta and glucocorticoids. The present studies demonstrate that human lesion-derived cells (LDC) are also resistant to apoptosis induced by fas ligation compared t ...[more]

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Project description:Background Rupture or erosion of advanced atherosclerotic lesions with a resultant myocardial infarction or stroke are the leading worldwide cause of death. However, we have a very limited understanding of the identity, origin, and function of many cells that make up late stage atherosclerotic lesions, as well as the mechanisms by which they control plaque stability. Methods and Results Using RNAseq and ChIPseq of advanced atherosclerotic lesions from mice, we provide evidence SMC-specific Klf4- versus Oct4-knockout ApoE-/- mice showed virtually opposite genomic signatures and putative SMC Klf4 or Oct4 target genes play an important role regulating SMC phenotypic changes. Further, we conducted a comprehensive single-cell RNA-seq of advanced human carotid endarterectomy samples and compared these with scRNAseq from murine micro-dissected advanced atherosclerotic lesions with SMC and endothelial lineage tracing to survey all plaque cell types and rigorously determine their origin. This analysis revealed remarkable similarity of transcriptomic clusters between mouse and human lesions and extensive plasticity of SMC- and EC-derived lesion cells. The latter included 7 distinct clusters of SMC- and EC-derived cells, most negative for traditional markers. In particular, SMC contributed to a Myh11-, Lgals3+ population with a chondrocyte-like gene signature that was markedly reduced with SMC-specific conditional knockout of Klf4. To study these cells and the mechanisms of their transition, we developed an innovative dual lineage tracing mouse that can uniquely label and genetically target Myh11+ SMC that subsequently activate Lgals3. We observed that SMC that activate Lgals3 comprise up to 2/3 of all SMC in advanced plaques. However, initial activation of Lgals3 in these cells does not represent conversion to a terminally differentiated state, but rather represents transition of these cells to a unique stem cell marker gene+, ECM-remodeling, “pioneer” cell phenotype that are the first to invest within lesions and subsequently give rise to at least 3 other SMC phenotypes within advanced lesions including Klf4-dependent osteogenic phenotypes likely to contribute to plaque calcification and plaque destabilization. Conclusions Taken together, these results provide evidence that SMC-derived cells within advanced mouse and human atherosclerotic lesions exhibit far greater phenotypic plasticity than generally believed, with Klf4 regulating transition to multiple phenotypes including Lgals3+ osteogenic cells likely to be detrimental for late stage atherosclerosis plaque pathogenesis.

Project description:Background/Aim: Macrophage polarization and phenotypic switching of smooth muscle cells (SMCs) are multi-faceted events dominating atherosclerosis (AS) progression. TGF-β was proved to been one of the bridge on the crosstalk between macrophage and SMC. ShenLian (SL) was extracted from a potent anti-atherosclerotic formula. However, its exact mechanism rebalancing inflammatory microenvironment of AS remain largely unknown. Within the entirety of macrophage and SMC, this study investigated the pharmacological effects of SL on stabilizing atherosclerotic plaques. Methods: The main components of SL were examined by high performance liquid chromatography. Co-culture and conditioned medium models of macrophage/SMC interactions were designed to identify the relationship between macrophage polarization and switching of SMC phenotypes. Flow cytometry, immunofluorescent staining, RT-PCR, western blotting, and ELISA were used to determine the expression of molecules relating to AS progression. An atherosclerosis animal model, established by placing a perivascular collar on the right common carotid artery in ApoE-/- mice, was used to investigate whether TGF-β is the key molecular mediator of SL in crosstalk between macrophage and SMC. Plaque size was defined by nuclear magnetic resonance imaging. Key markers related to phenotypic transformation of macrophage and SMC were determined by immunohistochemical staining. Results: Results revealed that, accompanied by rebalanced M2 macrophage polarization, SL supported SMC phenotypic transformation and functionally reconstruct the ECM of plaques specifically in macrophage-SMC co-cultural model. Molecularly, such activity of SL closely related to the activation of STAT3/SOCS3 pathway. Furthermore, in co-culture system, up-regulation of α-SMA induced by SL could neutralized by 1D11, a TGF-β neutralizing antibody, indicating that SL mediated Macrophage-SMC communication by enhancing TGF-β. In the AS model constructed by ApoE-/- mice, effects of SL on phenotypic transformation of macrophage and SMC has been well verified. Specific blocking of TGF-β largely attenuated the aforementioned effects of SL. Conclusion: Our findings highlighted that TGF-β might be the responsive factor of SL within macrophage and SMC communication. This study revealed that crosstalk between macrophage and SMC forms a holistic entirety promoting atherosclerotic plaque stability.

Project description:Despite decades of research, our understanding of processes controlling the stability of late-stage atherosclerotic plaques remains poor. However, a prevailing hypothesis is that reducing inflammation may improve plaque stability. Indeed, the potent inflammatory cytokine, interleukin-1β (IL-1β), has been shown to be a key driver of atherosclerosis development. Importantly, the CANTOS Trial recently demonstrated that administration of an anti-IL-1β antibody to high-risk post-myocardial infarction (MI) patients reduced the incidence of recurrent nonfatal MI, but did not reduce the incidence of cardiovascular death or stroke. As such, although the CANTOS trial results providing exciting evidence that targeting inflammation can have clinical benefit for treating advanced atherosclerosis, extensive further investigation is needed to better understand the mechanisms by which IL-1β inhibition impacts established lesions. Therefore, we performed intervention studies on smooth muscle cell (SMC) lineage tracing mice with advanced atherosclerosis using anti-IL- 1 or IgG control antibodies. Surprisingly, we found no effect on lesion size but a profound shift in the composition of the fibrous cap characterized by reduced collagen and SMC content but an increase in macrophage number, which was primarily driven by opposite effects on the proliferation of these respective cell types. By generating SMC-specific and macrophage-selective Il1r1 KO Apoe -/- mice, we found that SMC-specific Il1r1 KO resulted in a ~60% reduction in lesion size and lesions that were nearly devoid of YFP + SMC whereas, myeloid-selective loss of IL1R1 had no effect on lesion size, or cell composition. This suggests that SMC are a primary cell type responding to IL-1β in atherosclerosis and that IL1 signaling in SMC is critical for their investment and retention within the fibrous cap. In addition, we found that inhibition of IL-1β promoted an expansion of the M2 macrophage population within the fibrous cap, and that these effects may be due in part to elevated levels of IL4. Taken together, results show that IL-1β can promote beneficial changes in late-stage murine atherosclerosis by promoting maintenance of a SMC/collagen-rich fibrous cap. Moreover, studies identify critical cell types and pathways that need to be considered when attempting to develop safer and more effective anti-inflammatory therapies for widespread treatment of atherosclerotic disease, including in moderate or low risk patients.

Project description:BackgroundRupture and erosion of advanced atherosclerotic lesions with a resultant myocardial infarction or stroke are the leading worldwide cause of death. However, we have a limited understanding of the identity, origin, and function of many cells that make up late-stage atherosclerotic lesions, as well as the mechanisms by which they control plaque stability.MethodsWe conducted a comprehensive single-cell RNA sequencing of advanced human carotid endarterectomy samples and compared these with single-cell RNA sequencing from murine microdissected advanced atherosclerotic lesions with smooth muscle cell (SMC) and endothelial lineage tracing to survey all plaque cell types and rigorously determine their origin. We further used chromatin immunoprecipitation sequencing (ChIP-seq), bulk RNA sequencing, and an innovative dual lineage tracing mouse to understand the mechanism by which SMC phenotypic transitions affect lesion pathogenesis.ResultsWe provide evidence that SMC-specific Klf4- versus Oct4-knockout showed virtually opposite genomic signatures, and their putative target genes play an important role regulating SMC phenotypic changes. Single-cell RNA sequencing revealed remarkable similarity of transcriptomic clusters between mouse and human lesions and extensive plasticity of SMC- and endothelial cell-derived cells including 7 distinct clusters, most negative for traditional markers. In particular, SMC contributed to a Myh11-, Lgals3+ population with a chondrocyte-like gene signature that was markedly reduced with SMC-Klf4 knockout. We observed that SMCs that activate Lgals3 compose up to two thirds of all SMC in lesions. However, initial activation of Lgals3 in these cells does not represent conversion to a terminally differentiated state, but rather represents transition of these cells to a unique stem cell marker gene-positive, extracellular matrix-remodeling, "pioneer" cell phenotype that is the first to invest within lesions and subsequently gives rise to at least 3 other SMC phenotypes within advanced lesions, including Klf4-dependent osteogenic phenotypes likely to contribute to plaque calcification and plaque destabilization.ConclusionsTaken together, these results provide evidence that SMC-derived cells within advanced mouse and human atherosclerotic lesions exhibit far greater phenotypic plasticity than generally believed, with Klf4 regulating transition to multiple phenotypes including Lgals3+ osteogenic cells likely to be detrimental for late-stage atherosclerosis plaque pathogenesis.