Recently identified molecular targets in pulmonary artery hypertension (PAH) include sphingosine-1-phosphate (S1P) and zinc transporter ZIP12 signaling. This study sought to determine linkages between these pathways, and with BMPR2 signaling. Lung tissues from a rat model of monocrotaline-induced PAH and therapeutic treatment with bone marrow–derived endothelial-like progenitor cells transduced to overexpress BMPR2 were studied. Multifluorescence quantitative confocal microscopy (MQCM) was applied for analysis of protein expression and localization of markers of vascular remodeling (αSMA and BMPR2), parameters of zinc homeostasis (zinc transporter SLC39A/ZIP family members 1, 10, 12 and 14; and metallothionein MT3) and S1P extracellular signaling (SPHK1, SPNS2, S1P receptor isoforms 1, 2, 3, 5) in 20–200 µm pulmonary microvessels. ZIP12 expression in whole lung tissue lysates was assessed by western blot. Spearman nonparametric correlations between MQCM readouts and hemodynamic parameters, Fulton index (FI), and right ventricular systolic pressure (RVSP) were measured. In line with PAH status, pulmonary microvessels in monocrotaline-treated animals demonstrated significant (p < .05, n = 6 per group) upregulation of αSMA (twofold) and downregulation of BMPR2 (20%). Upregulated ZIP12 (92%), MT3 (57.7%), S1PR2 (54.8%), and S1PR3 (30.3%) were also observed. Significant positive and negative correlations were demonstrated between parameters of zinc homeostasis (ZIP12, MT3), S1P signaling (S1PRs, SPNS2), and vascular remodeling (αSMA, FI, RVSP). MQCM and western blot analysis showed that monocrotaline-induced ZIP12 upregulation could be partially negated by BMPR2-targeted therapy. Our results indicate that altered zinc transport/storage and S1P signaling in the monocrotaline-induced PAH rat model are linked to each other, and could be alleviated by BMPR2-targeted therapy.

中文翻译：

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肺动脉高压中锌和 1-磷酸鞘氨醇信号通路失调：靶向肺微血管中 2 型骨形态发生蛋白受体的潜在影响

最近发现的肺动脉高压 (PAH) 分子靶点包括 1-磷酸鞘氨醇 (S1P) 和锌转运蛋白 ZIP12 信号传导。本研究试图确定这些途径之间的联系，以及与 BMPR2 信号传导之间的联系。研究了来自野百合碱诱导的 PAH 大鼠模型的肺组织和用转导过表达 BMPR2 的骨髓来源的内皮样祖细胞进行的治疗。应用多荧光定量共聚焦显微镜 (MQCM) 分析蛋白质表达和血管重塑标志物的定位 ( αSMA 和 BMPR2）、锌稳态参数（锌转运蛋白 SLC39A/ZIP 家族成员 1、10、12 和 14；和金属硫蛋白 MT3）和 S1P 细胞外信号传导（SPHK1、SPNS2、S1P 受体亚型 1、2、3、5） 20–200 µm 肺微血管。通过蛋白质印迹评估全肺组织裂解物中的 ZIP12 表达。测量了 MQCM 读数与血流动力学参数、富尔顿指数 (FI) 和右心室收缩压 (RVSP) 之间的 Spearman 非参数相关性。与 PAH 状态一致，野百合碱处理动物的肺微血管表现出显着的α上调（ p  < .05，n  = 6 每组）SMA（双重）和 BMPR2 的下调（20%）。还观察到上调的 ZIP12 (92%)、MT3 (57.7%)、S1PR2 (54.8%) 和 S1PR3 (30.3%)。锌稳态参数（ZIP12、MT3）、S1P 信号（S1PRs、SPNS2）和血管重塑（ α SMA、FI、RVSP）参数之间存在显着的正相关和负相关。MQCM 和蛋白质印迹分析表明，野百合碱诱导的 ZIP12 上调可以被 BMPR2 靶向治疗部分抵消。我们的研究结果表明，野百合碱诱导的 PAH 大鼠模型中锌转运/储存和 S1P 信号传导的改变相互关联，并且可以通过 BMPR2 靶向治疗得到缓解。