Therapeutic Effect of Iron Citrate in Blocking Calcium Deposition in High Pi-Calcified VSMC: Role of Autophagy and Apoptosis
Therapeutic Effect of Iron Citrate in Blocking Calcium Deposition in High Pi-Calcified VSMC: Role of Autophagy and Apoptosis
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DOI：
10.3390/ijms20235925
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发表时间：
2019-12-01
影响因子：
5.6
通讯作者：
Cozzolino, Mario
中科院分区：
文献类型：
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作者：
Ciceri, Paola;Falleni, Monica;Cozzolino, Mario
在慢性肾脏病（CKD）中，首要死亡原因是主要由血管钙化（VC）引起的心血管疾病。最近，铁基磷酸盐结合剂被提出在晚期 CKD 中治疗高磷血症。我们研究了柠檬酸铁（铁）对高磷酸盐（Pi）钙化 VSMC 钙化进展的影响。在存在高 Pi 的情况下，在第 7-15 天添加铁可阻止进一步的钙化（1.30 +/- 0.03 与 0.61 +/- 0.02；OD/mg 蛋白质；第 15 天；Pi 与 Pi + Fe，p < 0.01）。接下来我们研究了细胞凋亡和自噬。在第 7-11 天，向高 Pi 处理的 VSMC 中添加铁，可减少凋亡细胞数量（17.3 +/- 2.6 与 11.6 +/- 1.6；膜联蛋白 V；阳性细胞百分比；第 11 天；Pi 与 Pi + Fe；p < 0.05）。对第 7-15 天治疗的 VSMC 和主动脉环中的凋亡细胞核进行彻底分析证实了结果（3.8 +/- 0.2 vs 2.3 +/- 0.3 和 4.0 +/- 0.3 vs 2.2 +/- 0.2；凋亡细胞核；任意评分；第 15 天；Pi vs Pi + Fe；VSMC 和主动脉环；p < 0.05）。通过研究促存活轴 GAS6/AXL，我们发现第 9-14 天的铁处理抵消了蛋白质高 Pi 刺激的下调，并诱导其从头合成。此外，在第 9-15 天添加铁会增强自噬，这是通过线粒体受损的自噬体数量增加和自噬通量增加来检测的。我们强调了铁对细胞凋亡的影响，证明了在高 Pi 治疗前和钙化已经加剧时添加铁可以阻止 H2O2 诱导的钙化增加。总之，我们证明铁通过抗凋亡作用和诱导已建立的钙化 VSMC 上的自噬来进一步阻止高 Pi 诱导的钙沉积。
In chronic kidney disease (CKD), the first cause of mortality is cardiovascular disease induced mainly by vascular calcification (VC). Recently, iron-based phosphate binders have been proposed in advanced CKD to treat hyperphosphatemia. We studied the effect of iron citrate (iron) on the progression of calcification in high-phosphate (Pi) calcified VSMC. Iron arrested further calcification when added on days 7-15 in the presence of high Pi (1.30 +/- 0.03 vs 0.61 +/- 0.02; OD/mg protein; day 15; Pi vs Pi + Fe, p < 0.01). We next investigated apoptosis and autophagy. Adding iron to high-Pi-treated VSMC, on days 7-11, decreased apoptotic cell number (17.3 +/- 2.6 vs 11.6 +/- 1.6; Annexin V; % positive cells; day 11; Pi vs Pi + Fe; p < 0.05). The result was confirmed thorough analysis of apoptotic nuclei both in VSMCs and aortic rings treated on days 7-15 (3.8 +/- 0.2 vs 2.3 +/- 0.3 and 4.0 +/- 0.3 vs 2.2 +/- 0.2; apoptotic nuclei; arbitrary score; day 15; Pi vs Pi + Fe; VSMCs and aortic rings; p < 0.05). Studying the prosurvival axis GAS6/AXL, we found that iron treatment on days 9-14 counteracted protein high-Pi-stimulated down-regulation and induced its de novo synthesis. Moreover, iron added on days 9-15 potentiated autophagy, as detected by an increased number of autophagosomes with damaged mitochondria and an increase in autophagic flux. Highlighting the effect of iron on apoptosis, we demonstrated its action in blocking the H2O2-induced increase in calcification added both before high Pi treatment and when the calcification was already exacerbated. In conclusion, we demonstrate that iron arrests further high Pi-induced calcium deposition through an anti-apoptotic action and the induction of autophagy on established calcified VSMC.