Endothelial cells (ECs), the primary component of the vasculature, play a crucial role in neovascularization. However, the number of endogenous ECs is inadequate for both experimental purposes and clinical applications. Porcine ovarian putative stem cells (poPSCs), although not pluripotent, are characterized by great plasticity. Therefore, this study aimed to investigate whether poPSCs have the potential to differentiate into cells of endothelial lineage. poPSCs were immunomagnetically isolated from postnatal pig ovaries based on the presence of SSEA-4 protein. Expression of mesenchymal stem cells (MSCs) markers after pre-culture, both at the level of mRNA: ITGB1, THY, and ENG and corresponding protein: CD29, CD90, and CD105 were significantly higher compared to the control ovarian cortex cells. To differentiate poPSCs into ECs, inducing medium containing vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), insulin-like growth factor (IGF), epidermal growth factor (EGF), ascorbic acid, and heparin was applied. After 14 days, poPSC differentiation into ECs was confirmed by immunofluorescence staining for vascular endothelial cadherin (VECad) and vascular endothelial growth factor receptor-2 (VEGFR-2). Semi-quantitative WB analysis of these proteins confirmed their high abundance. Additionally, qRT-PCR showed that mRNA expression of corresponding marker genes: CDH5, KDR was significantly higher compared with undifferentiated poPSCs. Finally, EC functional status was confirmed by the migration test that revealed that they were capable of positive chemotaxis, while tube formation assay demonstrated their ability to develop capillary networks. In conclusion, our results provided evidence that poPSCs may constitute the MSC population in the ovary and confirmed that they might be a potential source of ECs for tissue engineering.

内皮细胞 (ECs) 是脉管系统的主要组成部分，在新生血管形成中起着至关重要的作用。然而，内源性 EC 的数量不足以用于实验目的和临床应用。猪卵巢推定干细胞（poPSCs）虽然不是多能的，但具有很强的可塑性。因此，本研究旨在调查 poPSCs 是否具有分化为内皮谱系细胞的潜力。基于 SSEA-4 蛋白的存在，从出生后的猪卵巢中免疫磁性分离出 poPSC。预培养后间充质干细胞 (MSCs) 标志物在 mRNA 水平上的表达：ITGB1、THY和ENG和相应的蛋白质：与对照卵巢皮质细胞相比，CD29、CD90 和 CD105 显着升高。为了将 poPSCs 分化为 ECs，应用了含有血管内皮生长因子 (VEGF)、碱性成纤维细胞生长因子 (bFGF)、胰岛素样生长因子 (IGF)、表皮生长因子 (EGF)、抗坏血酸和肝素的诱导培养基。14 天后，通过血管内皮钙粘蛋白 (VECad) 和血管内皮生长因子受体-2 (VEGFR-2) 的免疫荧光染色证实了 poPSC 分化为 ECs。这些蛋白质的半定量WB分析证实了它们的高丰度。此外，qRT-PCR 显示相应标记基因的 mRNA 表达：CDH5、KDR与未分化的 poPSC 相比显着更高。最后，通过迁移测试证实了 EC 功能状态，表明它们具有阳性趋化性，而管形成试验证明了它们发展毛细血管网络的能力。总之，我们的结果提供了证据，证明 poPSCs 可能构成卵巢中的 MSC 群，并证实它们可能是组织工程中 ECs 的潜在来源。