BACKGROUND Preeclampsia is a pregnancy-specific hypertensive disorder characterized by impaired angiogenesis. We postulate that senescence of mesenchymal stem cells (MSC), multipotent cells with pro-angiogenic activities, is one of the mechanisms by which systemic inflammation exerts inhibitory effects on angiogenesis in preeclampsia. METHODS MSC were isolated from abdominal fat tissue explants removed during medically indicated C-sections from women with preeclampsia (PE-MSC, n = 10) and those with normotensive pregnancies (NP-MSC, n = 12). Sections of the frozen subcutaneous adipose tissue were assessed for inflammation by staining for tumor necrosis factor (TNF)-alpha and monocyte chemoattractant protein (MCP)-1. Viability, proliferation, and migration were compared between PE-MSC vs. NP-MSC. Apoptosis and angiogenesis were assayed before and after treatment with a senolytic agent (1 μM dasatinib) using the IncuCyte S3 Live-Cell Analysis System. Similarly, staining for senescence-associated beta galactosidase (SABG) and qPCR for gene expression of senescence markers, p16 and p21, as well as senescence-associated secretory phenotype (SASP) components, IL-6, IL-8, MCP-1, and PAI-1, were studied before and after treatment with dasatinib and compared between PE and NP. RESULTS After in vitro exposure to TNF-alpha, MSC demonstrated upregulation of SASP components, including interleukins-6 and -8 and MCP-1. Staining of the subcutaneous adipose tissue sections revealed a greater inflammatory response in preeclampsia, based on the higher levels of both TNF-alpha and MCP-1 compared to normotensive pregnancies (p < 0.001 and 0.024, respectively). MSC isolated from PE demonstrated a lower percentage of live MSC cells (p = 0.012), lower proliferation (p = 0.005), and higher migration (p = 0.023). At baseline, PE-MSC demonstrated a senescent phenotype, reflected by more abundant staining for SABG (p < 0.001), upregulation of senescence markers and SASP components, as well as lower angiogenic potential (p < 0.001), compared to NP-MSC. Treatment with dasatinib increased significantly the number of apoptotic PE-MSC compared to NP-MSC (0.011 vs. 0.093) and decreased the gene expression of p16 and six SASP components. The mechanistic link between senescence and impaired angiogenesis in PE was confirmed by improved angiogenic potential of PE-MSC (p < 0.001) after dasatinib treatment. CONCLUSIONS Our data suggest that MSC senescence exerts inhibitory effects on angiogenesis in preeclampsia. Senolytic agents may offer the opportunity for mechanism-based therapies.

中文翻译：

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靶向衰老可改善先兆子痫患者脂肪来源的间充质干细胞的血管生成潜力。

背景技术子痫前期是一种以血管生成受损为特征的妊娠特异性高血压疾病。我们假设间充质干细胞（MSC）的衰老，具有促血管生成活性的多能细胞，是先兆子痫系统性炎症对血管生成产生抑制作用的机制之一。方法MSCs是从子痫前期妇女（PE-MSC，n = 10）和血压正常孕妇（NP-MSC，n = 12）在医学指征剖腹产中去除的腹部脂肪组织外植体中分离得到的。通过对肿瘤坏死因子（TNF）-α和单核细胞趋化蛋白（MCP）-1染色，评估冷冻的皮下脂肪组织切片的炎症。比较了PE-MSC与NP-MSC之间的生存力，增殖和迁移。使用IncuCyte S3活细胞分析系统，在用抗溶剂（1μMdasatinib）治疗之前和之后测定细胞凋亡和血管生成。同样地，对衰老相关的β半乳糖苷酶（SABG）进行染色，并通过qPCR进行衰老标记p16和p21以及衰老相关的分泌表型（SASP）成分IL-6，IL-8，MCP-1，在使用dasatinib之前和之后研究了PAI-1和PAI-1，并比较了PE和NP。结果在体外暴露于TNF-α后，MSC表现出SASP成分的上调，包括白介素6和-8以及MCP-1。皮下脂肪组织切片的染色显示先兆子痫的炎症反应更大，这是由于与正常血压妊娠相比，TNF-α和MCP-1含量较高（p <0.001和0.024，分别）。从PE分离的MSC表现出较低的存活MSC细胞百分比（p = 0.012），较低的增殖（p = 0.005）和较高的迁移（p = 0.023）。在基线时，PE-MSC表现出衰老表型，与NP-MSC相比，SABG染色更丰富（p <0.001），衰老标记和SASP组分上调以及较低的血管生成潜能（p <0.001）反映出来。与NP-MSC相比，达沙替尼治疗显着增加了凋亡PE-MSC的数量（0.011对0.093），并降低了p16和6个SASP组分的基因表达。在达沙替尼治疗后，PE-MSC的血管生成潜力得到改善（p <0.001），从而证实了PE衰老与血管生成受损之间的机制联系。结论我们的数据表明MSC衰老对先兆子痫的血管生成具有抑制作用。缓溶剂可能为基于机制的疗法提供机会。