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Sphingosine 1-phosphate-regulated transcriptomes in heterogenous arterial and lymphatic endothelium of the aorta

ABSTRACT: Despite the medical importance of G protein-coupled receptors (GPCRs), in vivo cellular heterogeneity of GPCR signaling and downstream transcriptional responses are not understood. While endothelial S1pr1 expression is ubiquitous (Kaur et al., 2017), receptor activation in vivo, as reported by GFP in S1PR1-GS mice, revealed heterogeneous signaling in multiple organs (Kono et al., 2014), including in the aorta (Galvani et al., 2015). To gain insights into the molecular mechanisms of S1PR1 regulation of endothelial transcription and the heterogenous nature of S1PR1 signaling in vivo, we performed transcriptome (bulk and single-cell RNA-seq) and open chromatin profiling (ATAC-seq) of FACS-sorted adult mouse aortic endothelial cells (MAECs) with high (GFPhigh) or low (GFPlow) reporter expression. We also performed bulk transcriptome and open chromatin profiling of MAECs from tamoxifen-treated Cdh5-CreERT2 S1pr1f/f (S1pr1-ECKO) and S1pr1f/f (S1pr1-WT) littermates. MAECs were defined as CD45-Ter119-DAPI-CD31+. The CD31 antibody used was PE-conjugated rat anti-mouse clone MEC13.3 from BD Biosciences.

ORGANISM(S): Mus musculus

PROVIDER: GSE139065 | GEO | 2019/10/18

REPOSITORIES: GEO

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Project description:A major challenge in realizing regenerative treatment using freshly isolated Adipose Derived Regenerative Cells (ADRCs) is the cellular heterogeneity and donor variability. Ex vivo expanded ADRCs (=ASCs) are available at larger quantities and represent a more homogeneous population suitable for allogenic use. Emerging pre-clinical and clinical data however suggest similar, or even superior efficacy of ADRCs as compared to ASCs for indications involving (micro)vascular deficiency. Since CD31+ ADRCs lack in cultured ASCs, we hypothesized that these cells account for improvement of vascular function in the heterogenous ADRCs. Herein, we found that in patients with erectile dysfunction (ED), the number of injected CD31+ ADRCs correlates positively with erectile function 12 months after one bolus of autologous ADRCs. Comprehensive in vitro and ex vivo analyses confirmed superior pro-angiogenic and paracrine effects of human CD31+ enriched ADRCs compared to the corresponding CD31- and parent ADRCs. CD31+, CD31- and ADRCs were co-cultured in aortic ring-, as well as in ED-relevant corpus cavernousum tube formation assays, both showing a significantly higher ability of the CD31+ ADRCs to support tube development. This effect was corroborated using conditioned medium (CM), while quantitative mass spectrometric analysis suggested that this is explained by secretory pro-angiogenic proteins such as DKK3, ANGPT2, ANAX2 and VIM, all being enriched in CD31+ADRC CM. To gain further specification of the ADRC subpopulations expressing these proteins, single-cell RNA sequencing was performed and showed that transcripts of the upregulated and secreted proteins were present in 9 endothelial ADRC subsets including endothelial progenitor cells in the heterogenous non-cultured ADRCs. Our data thus suggest that the vascular benefit of using ADRCs in regenerative medicine is dictated by CD31+ ADRCs, which likely represent an improved alternative to heterogenous ADRCs as well as ASCs when aiming for vascular repair in cell therapy.

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Sphingosine 1-phosphate-regulated transcriptomes in heterogenous arterial and lymphatic endothelium of the aorta

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