In recent years,MSCs have emerged as a promising therapeutic cell type in regenerative medicine. They hold great promise for treating cardiovascular diseases, such as myocardial infarction and limb ischemia.MSCs may be utilized in both cell‐based therapy and vascular graft engineering to restore vascular function, thereby providing therapeutic benefits to patients. The efficacy ofMSCs lies in their multipotent differentiation ability toward vascular smooth muscle cells, endothelial cells and other cell types, as well as their capacity to secrete various trophic factors, which are potent in promoting angiogenesis, inhibiting apoptosis and modulating immunoreaction. Increasing our understanding of the mechanisms ofMSCinvolvement in vascular regeneration will be beneficial in boosting present therapeutic approaches and developing novel ones to treat cardiovascular diseases. In this review, we aim to summarize current progress in characterizing the in vivo identity ofMSCs, to discuss mechanisms involved in cell‐based therapy utilizingMSCs, and to explore current and future strategies for vascular regeneration.

Tissue-engineered vascular grafts with long-term patency are greatly needed in the clinical settings, and smooth muscle cells (SMCs) are a critical graft component. Human mesenchymal stem cells (MSCs) are used for generating SMCs, and understanding the underlying regulatory mechanisms of the MSC-to-SMC differentiation process could improve SMC generation in the clinic. Here, we found that in response to stimulation of transforming growth factor-β1 (TGFβ1), human umbilical cord–derived MSCs abundantly express the SMC markers α-smooth muscle actin (αSMA), smooth muscle protein 22 (SM22), calponin, and smooth muscle myosin heavy chain (SMMHC) at both gene and protein levels. Functionally, MSC-derived SMCs displayed contracting capacity in vitro and supported vascular structure formation in the Matrigel plug assay in vivo. More importantly, SMCs differentiated from human MSCs could migrate into decellularized mouse aorta and give rise to the smooth muscle layer of vascular grafts, indicating the potential of utilizing human MSC-derived SMCs to generate vascular grafts. Of note, microRNA (miR) array analysis and TaqMan microRNA assays identified miR-503 and miR-222-5p as potential regulators of MSC differentiation into SMCs at early time points. Mechanistically, miR-503 promoted SMC differentiation by directly targeting SMAD7, a suppressor of SMAD-related, TGFβ1-mediated signaling pathways. Moreover, miR-503 expression was SMAD4-dependent. SMAD4 was enriched at the miR-503 promoter. Furthermore, miR-222-5p inhibited SMC differentiation by targeting and down-regulating ROCK2 and αSMA. In conclusion, MSC differentiation into SMCs is regulated by miR-503 and miR-222-5p and yields functional SMCs for use in vascular grafts.

Aims: Hypertension is a major risk factor for cardiovascular diseases. However, vascular remodelling, a hallmark of hypertension, has not been systematically characterized yet. We described systematic vascular remodelling, especially the artery type- and cell type-specific changes, in hypertension using spontaneously hypertensive rats (SHRs). Methods and results: Single-cell RNA sequencing was used to depict the cell atlas of mesenteric artery (MA) and aortic artery (AA) from SHRs. More than 20 000 cells were included in the analysis. The number of immune cells more than doubled in aortic aorta in SHRs compared to Wistar Kyoto controls, whereas an expansion of MA mesenchymal stromal cells (MSCs) was observed in SHRs. Comparison of corresponding artery types and cell types identified in integrated datasets unravels dysregulated genes specific for artery types and cell types. Intersection of dysregulated genes with curated gene sets including cytokines, growth factors, extracellular matrix (ECM), receptors, etc. revealed vascular remodelling events involving cell–cell interaction and ECM re-organization. Particularly, AA remodelling encompasses upregulated cytokine genes in smooth muscle cells, endothelial cells, and especially MSCs, whereas in MA, change of genes involving the contractile machinery and downregulation of ECM-related genes were more prominent. Macrophages and T cells within the aorta demonstrated significant dysregulation of cellular interaction with vascular cells. Conclusion: Our findings provide the first cell landscape of resistant and conductive arteries in hypertensive animal models. Moreover, it also offers a systematic characterization of the dysregulated gene profiles with unbiased, artery type-specific and cell type-specific manners during hypertensive vascular remodelling.

Rationale: Transplant arteriosclerosis is the major limitation to long-term survival of solid organ transplantation. Although both immune and nonimmune cells have been suggested to contribute to this process, the complex cellular heterogeneity within the grafts, and the underlying mechanisms regulating the disease progression remain largely uncharacterized. Objective: We aimed to delineate the cellular heterogeneity within the allografts, and to explore possible mechanisms underlying this process. Methods and Results: Here, we reported the transcriptional profiling of 11 868 cells in a mouse model of transplant arteriosclerosis by single-cell RNA sequencing. Unbiased clustering analyses identified 21 cell clusters at different stages of diseases, and focused analysis revealed several previously unknown subpopulations enriched in the allografts. Interestingly, we found evidence of the local formation of tertiary lymphoid tissues and suggested a possible local modulation of alloimmune responses within the grafts. Intercellular communication analyses uncovered a potential role of several ligands and receptors, including Ccl21a and Cxcr3 , in regulating lymphatic endothelial cell-induced early chemotaxis and infiltration of immune cells. In vivo mouse experiments confirmed the therapeutic potential of CCL21 and CXCR3 neutralizing antibodies in transplant arteriosclerosis. Combinational use of genetic lineage tracing and single-cell techniques further indicate the infiltration of host-derived c-Kit ⁺ stem cells as heterogeneous populations in the allografts. Finally, we compared the immune response between mouse allograft and atherosclerosis models in single-cell RNA-seq analysis. By analyzing susceptibility genes of disease traits, we also identified several cell clusters expressing genes associated with disease risk. Conclusions: Our study provides a transcriptional and cellular landscape of transplant arteriosclerosis, which could be fundamental to understanding the initiation and progression of this disease. CCL21/CXCR3 was also identified as important regulators of immune response and may serve as potential therapeutic targets in disease treatment.

Although genetic risk in coronary artery disease (CAD) is linked to changes in gene expression, recent discoveries have revealed a major role for A-to-I RNA editing in CAD. ADAR1 edits immunogenic double-stranded RNA (dsRNA), preventing activation of the dsRNA sensor MDA5 (IFIH1) and downstream interferon-stimulated gene signaling. Using human plaque analysis and human coronary artery smooth muscle cells (SMCs), here, we show that SMCs uniquely require RNA editing and that MDA5 activation regulates SMC phenotype. In a conditional SMC-specific Adar deletion mouse model on an atherosclerosis-prone background, combined with Ifih1 deletion and single-cell RNA sequencing, we demonstrate that ADAR1 preserves vascular integrity and limits atherosclerosis and calcification by suppressing MDA5 activation. Analysis of the Athero-Express carotid endarterectomy cohort further shows that interferon-stimulated gene expression correlates with SMC modulation, plaque instability and calcification. These findings reveal a fundamental mechanism of CAD, where cell type and context-specific RNA editing modulates genetic risk and vascular disease progression.

Vascular smooth muscle cells contribute to heritable coronary artery disease risk and undergo complex transitions to multiple disease-related phenotypes. To investigate the genetic basis of these trajectories, we develop a dense timecourse single-cell transcriptomic and epigenetic map of atherosclerosis in a murine disease model accompanied by high-plex in situ spatial data. Using temporal data and probabilistic fate modeling, we identify key transcription factors that drive cell state changes through a combination of network-based prioritization and in silico transcription factor perturbation. Parallel knockout studies of validated coronary artery disease gene Tcf21 uncover its molecular mechanisms in smooth muscle cell transition, due in part to a role regulating the transition of smooth muscle cells in the secondary heart field. Integrating the murine atlas with human coronary artery disease genetics pinpoint smooth muscle cell phenotypes that mediate disease risk, highlighting causal disease mechanisms. Together, these studies resolve atherosclerosis trajectories at single-cell resolution and identify genetic causal transcriptomic and epigenomic mechanisms of coronary artery disease risk.