BACKGROUND: A main obstacle in current valvular heart disease research is the lack of high-quality homogeneous functional heart valve cells. Human induced pluripotent stem cells (hiPSCs)-derived heart valve cells may help with this dilemma. However, there are no well-established protocols to induce hiPSCs to differentiate into functional heart valve cells, and the networks that mediate the differentiation have not been fully elucidated. METHODS: To generate heart valve cells from hiPSCs, we sequentially activated the Wnt, BMP4, VEGF (vascular endothelial growth factor), and NFATc1 signaling pathways using CHIR-99021, BMP4, VEGF-165, and forskolin, respectively. The transcriptional and functional similarity of hiPSC-derived heart valve cells compared with primary heart valve cells were characterized. Longitudinal single-cell RNA sequencing was used to uncover the trajectory, switch genes, pathways, and transcription factors of the differentiation. RESULTS: An efficient protocol was developed to induce hiPSCs to differentiate into functional hiPSC-derived valve endothelial-like cells and hiPSC-derived valve interstitial-like cells. After 6-day differentiation and CD144 magnetic bead sorting, ≈70% CD144 ⁺ cells and 30% CD144 ^– cells were obtained. On the basis of single-cell RNA sequencing data, the CD144 ⁺ cells and CD144 ^– cells were found to be highly similar to primary heart valve endothelial cells and primary heart valve interstitial cells in gene expression profile. Furthermore, CD144 ⁺ cells had the typical function of primary heart valve endothelial cells, including tube formation, uptake of low-density lipoprotein, generation of endothelial nitric oxide synthase, and response to shear stress. Meanwhile, CD144 ^– cells could secret collagen and matrix metalloproteinases, and differentiate into osteogenic or adipogenic lineages like primary heart valve interstitial cells. Therefore, we identified CD144 ⁺ cells and CD144 ^– cells as hiPSC-derived valve endothelial-like cells and hiPSC-derived valve interstitial-like cells, respectively. Using single-cell RNA sequencing analysis, we demonstrated that the trajectory of heart valve cell differentiation was consistent with embryonic valve development. We identified the main switch genes (NOTCH1, HEY1, and MEF2C), signaling pathways (TGF-β, Wnt, and NOTCH), and transcription factors (MSX1, SP5, and MECOM) that mediated the differentiation. Finally, we found that hiPSC-derived valve interstitial-like cells might derive from hiPSC-derived valve endothelial-like cells undergoing endocardial-mesenchymal transition. CONCLUSIONS: In summary, this is the first study to report an efficient strategy to generate functional hiPSC-derived valve endothelial-like cells and hiPSC-derived valve interstitial-like cells from hiPSCs, as well as to elucidate the differentiation trajectory and transcriptional dynamics of hiPSCs differentiated into heart valve cells.

Background: Apolipoprotein C3 (apo C3) is primarily secreted by the liver and is involved in promoting sterile inflammation and organ damage under pathological conditions. Previous studies have shown that apo C3 is abundant in the plasma exosomes of patients with aortic dissection (AD), but its specific role in AD remains unclear. Methods and Results: In vivo, adeno‐associated virus was used to knock down hepatic apo C3 expression in an AD mouse model to assess the impact of liver‐derived apo C3 on the development of AD. In vitro, recombinant apo C3 protein was added to the culture medium of J774A.1 macrophages to evaluate its effect on macrophage polarization and to identify the underlying mechanisms. Additionally, the effect of apo C3 on the function of aortic endothelial and smooth muscle cells was explored. Apo C3 in the aortas of AD mice was found to originate from abnormal hepatic secretion, which enters the bloodstream and subsequently deposits in the aorta. Adeno‐associated virus–mediated hepatic apo C3 knockdown significantly reduced AD incidence ( P =0.0036), macrophage infiltration ( P =0.0004), and collagen deposition in the aorta ( P =0.0016). Similarly, inhibiting the apo C3 receptor Toll‐like receptor 2 significantly lowered AD incidence ( P =0.0352). In vitro, recombinant apo C3 protein promoted M1 polarization and matrix metalloproteinase secretion in macrophages by activating the Toll‐like receptor 2/NLR family pyrin domain containing 3 pathway. Additionally, apo C3 increased adhesion molecule expression in endothelial cells and induced inflammation, chemotaxis, and apoptosis in vascular smooth muscle cells. Conclusions: Our findings highlight the role of abnormally secreted hepatic apo C3 in promoting aortic inflammation.

Fibroblasts play a crucial role in the development and progression of heart failure in many heart-related diseases. However, the contributions of fibroblast and its subtypes to the pathogenesis of ischemic cardiomyopathy (ICM) remain unclear. In this study, we analyzed single-nucleus RNA sequencing data from cardiac tissues of seven ICM patients with heart failure and eight control subjects. Our findings indicate that, compared to controls, fibroblasts from ICM patients exhibit dysregulation of genes associated with cell-substrate adhesion and matrix or collagen fibril organization. We further identified five fibroblast subtypes, designated as FB1 through FB5, with FB3 exhibiting the highest frequency in ICM. Pseudotime trajectory analysis further revealed an evolutionary progression of fibroblasts from FB1/FB2 toward FB3. Differential gene expression analysis of FB3 highlighted several subtype-specific genes, such as NTM, THBS4, NRK, NAV2, and PLA2G5. Furthermore, intercellular interaction analysis revealed that FB3 exhibits weakened interactions with other cells under ICM. Specifically, signaling interactions involving ANGPTL and COLLAGEN are reduced, whereas pathways mediated by THBS, NRXN, and APP are strengthened. These results provide valuable insights into the fibroblast microenvironment and elucidate the pathological mechanisms underlying ICM, laying a theoretical foundation for the identification of novel therapeutic targets.

Purpose: To evaluate the influence of transfusion amount of blood components on the prognosis of patients after heart transplantation (HTx).Methods: From 1 January 2015 to 31 December 2020, 568 patients underwent HTx in our institute. A total of 416 recipients with complete datasets were enrolled in the study for final statistical analysis according to the inclusion criteria. The optimal cut-off values for intraoperative transfusion of red blood cell (RBC), platelet, and plasma were determined with receiver operating curve analysis. Univariate and multivariate Cox regression analyses were applied to compare baseline data of patients divided by the transfusion amounts of RBC, platelet, and plasma. Propensity score matching was used to enable the direct comparison of outcomes.Results: The Kaplan–Meier analysis revealed that transfusion amounts of RBC and plasma were independently associated with overall mortality, increased intensive care unit stay time, and major adverse events after transplantation. The multivariate Cox regression analysis suggested that neurological complications (p = 0.001), liver damage (p = 0.011), and respiratory complications (p = 0.044) were independent risk factors for overall mortality after HTx. Combining indicators presented a good predicting effect of peritransplant period mortality (AUC = 0.718).Conclusion: The mortality of HTx was significantly related to the high-amount transfusion of RBC and plasma. Comprehensively considering the components of blood transfusion obtained better predictive results of peritransplant period survival than solely considering a single component.

The technology of induced pluripotent stem cells (iPSCs) has enabled the conversion of somatic cells into primitive undifferentiated cells via reprogramming. This approach provides possibilities for cell replacement therapies and drug screening, but the potential risk of tumorigenesis hampers its further development and in vivo application. How to generate differentiated cells such as valvular endothelial cells (VECs) has remained a major challenge.