Single cell RNA sequencing identifies mitochondrial respiration as a key factor contributing to extracellular matrix integrity

Heterogeneous spheroids have recently acquired a prominent position in melanoma research because they incorporate microenvironmental cues relevant for melanoma. In this study, we focused on the analysis of microenvironmental factors introduced in melanoma heterogeneous spheroids by different dermal fibroblasts. We aimed to map the fibroblast diversity resulting from previously acquired damage caused by exposure to extrinsic and intrinsic stimuli. To construct heterogeneous melanoma spheroids, we used normal dermal fibroblasts from the sun-protected skin of a juvenile donor. We compared them to the fibroblasts from the sun-exposed photodamaged skin of an adult donor. Further, we analysed the spheroids by single-cell RNA sequencing. To validate transcriptional data, we also compared the immunohistochemical analysis of heterogeneous spheroids to melanoma biopsies. We have distinguished three functional clusters in primary human fibroblasts from melanoma spheroids. These clusters differed in the expression of (a) extracellular matrix-related genes, (b) pro-inflammatory factors, and (c) TGFβ signalling superfamily. We observed a broader deregulation of gene transcription in previously photodamaged cells. We have confirmed that pro-inflammatory cytokine IL-6 significantly enhances melanoma invasion to the extracellular matrix in our model. This supports the opinion that the aspects of ageing are essential for reliable melanoma 3D modelling in vitro.

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Discovery of potential imaging and therapeutic targets for severe inflammation in COVID-19 patients

Scientific Reports ◽

10.1038/s41598-021-93743-2 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Hyunjong Lee ◽

Jeongbin Park ◽

Hyung-Jun Im ◽

Kwon Joong Na ◽

Hongyoon Choi

Keyword(s):

Single Cell ◽

Rna Sequencing ◽

Immune Cell ◽

Lavage Fluid ◽

Severe Illness ◽

Sequencing Data ◽

Single Cell Rna Sequencing ◽

Key Factor ◽

Target Binding ◽

Druggable Targets

AbstractThe Coronavirus disease 2019 (COVID-19) has been spreading worldwide with rapidly increased number of deaths. Hyperinflammation mediated by dysregulated monocyte/macrophage function is considered to be the key factor that triggers severe illness in COVID-19. However, no specific targeting molecule has been identified for detecting or treating hyperinflammation related to dysregulated macrophages in severe COVID-19. In this study, previously published single-cell RNA-sequencing data of bronchoalveolar lavage fluid cells from thirteen COVID-19 patients were analyzed with publicly available databases for surface and imageable targets. Immune cell composition according to the severity was estimated with the clustering of gene expression data. Expression levels of imaging target molecules for inflammation were evaluated in macrophage clusters from single-cell RNA-sequencing data. In addition, candidate targetable molecules enriched in severe COVID-19 associated with hyperinflammation were filtered. We found that expression of SLC2A3, which can be imaged by [18F]fluorodeoxyglucose, was higher in macrophages from severe COVID-19 patients. Furthermore, by integrating the surface target and drug-target binding databases with RNA-sequencing data of severe COVID-19, we identified candidate surface and druggable targets including CCR1 and FPR1 for drug delivery as well as molecular imaging. Our results provide a resource in the development of specific imaging and therapy for COVID-19-related hyperinflammation.

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Single Cell RNA Sequencing of Human Lung Explants Identifies the Cell-Specific Contributions of Extracellular Matrix-Related Gene Expression

10.1164/ajrccm-conference.2019.199.1_meetingabstracts.a2550 ◽

2019 ◽

Author(s):

S. Poli De Frias ◽

T. Adams ◽

J.C. Schupp ◽

S. Chu ◽

N. Neumark ◽

...

Keyword(s):

Gene Expression ◽

Extracellular Matrix ◽

Single Cell ◽

Rna Sequencing ◽

Human Lung ◽

Related Gene ◽

Single Cell Rna Sequencing

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Single-Cell Transcriptomic Profiling of Vascular Smooth Muscle Cell Phenotype Modulation in Marfan Syndrome Aortic Aneurysm

Arteriosclerosis Thrombosis and Vascular Biology ◽

10.1161/atvbaha.120.314670 ◽

2020 ◽

Vol 40 (9) ◽

pp. 2195-2211

Author(s):

Albert J. Pedroza ◽

Yasushi Tashima ◽

Rohan Shad ◽

Paul Cheng ◽

Robert Wirka ◽

...

Keyword(s):

Extracellular Matrix ◽

Smooth Muscle ◽

Aortic Aneurysm ◽

Vascular Smooth Muscle ◽

Smooth Muscle Cell ◽

Single Cell ◽

Rna Sequencing ◽

Marfan Syndrome ◽

Aortic Root ◽

Single Cell Rna Sequencing

Objective: To delineate temporal and spatial dynamics of vascular smooth muscle cell (SMC) transcriptomic changes during aortic aneurysm development in Marfan syndrome (MFS). Approach and Results: We performed single-cell RNA sequencing to study aortic root/ascending aneurysm tissue from Fbn1 C1041G/ + (MFS) mice and healthy controls, identifying all aortic cell types. A distinct cluster of transcriptomically modulated SMCs (modSMCs) was identified in adult Fbn1 C1041G/ + mouse aortic aneurysm tissue only. Comparison with atherosclerotic aortic data (ApoE −/− mice) revealed similar patterns of SMC modulation but identified an MFS-specific gene signature, including plasminogen activator inhibitor-1 ( Serpine1 ) and Kruppel-like factor 4 ( Klf4 ). We identified 481 differentially expressed genes between modSMC and SMC subsets; functional annotation highlighted extracellular matrix modulation, collagen synthesis, adhesion, and proliferation. Pseudotime trajectory analysis of Fbn1 C1041G/ + SMC/modSMC transcriptomes identified genes activated differentially throughout the course of phenotype modulation. While modSMCs were not present in young Fbn1 C1041G/ + mouse aortas despite small aortic aneurysm, multiple early modSMCs marker genes were enriched, suggesting activation of phenotype modulation. modSMCs were not found in nondilated adult Fbn1 C1041G/ + descending thoracic aortas. Single-cell RNA sequencing from human MFS aortic root aneurysm tissue confirmed analogous SMC modulation in clinical disease. Enhanced expression of TGF-β (transforming growth factor beta)-responsive genes correlated with SMC modulation in mouse and human data sets. Conclusions: Dynamic SMC phenotype modulation promotes extracellular matrix substrate modulation and aortic aneurysm progression in MFS. We characterize the disease-specific signature of modSMCs and provide temporal, transcriptomic context to the current understanding of the role TGF-β plays in MFS aortopathy. Collectively, single-cell RNA sequencing implicates TGF-β signaling and Klf4 overexpression as potential upstream drivers of SMC modulation.

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