Increased generation of cyclopentenone prostaglandins after brain ischemia and their role in aggregation of ubiquitinated proteins in neurons.
Increased generation of cyclopentenone prostaglandins after brain ischemia and their role in aggregation of ubiquitinated proteins in neurons.
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DOI：
10.1007/s12640-013-9377-4
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发表时间：
2013-08
影响因子：
3.7
通讯作者：
Graham, Steven H.
中科院分区：
文献类型：
--
作者：
Liu, Hao;Li, Wenjin;Ahmad, Muzamil;Rose, Marie E.;Miller, Tricia M.;Yu, Mei;Chen, Jie;Pascoe, Jordan L.;Poloyac, Samuel M.;Hickey, Robert W.;Graham, Steven H.
关键词：
环戊烯酮前列腺素（CyPG）J2系列，包括前列腺素J2（PGJ 2）、Δ12-PGJ 2和15-脱氧-Δ12，14 -PGJ 2（15 d-PGJ 2），是PGD 2的活性代谢产物，对神经元功能具有多种作用。然而，这些效应的生理相关性仍然不确定，因为脑内CyPG浓度尚未精确测定。在本研究中，我们发现，通过UPLC-MS/MS检测，缺血后大鼠脑中游离PGD 2和J2系列CyPG（PGJ 2，Δ12-PGJ 2和15 d-PGJ 2）增加，其中15 d-PGJ 2是最丰富的CyPG。这些增加被衰减预处理与环氧合酶抑制剂吡罗昔康。接下来，通过用15 d-PGJ 2、CAY 10410（缺乏环戊烯酮环结构的15 d-PGJ 2类似物）或媒介物处理原代神经元24小时至96小时来检查慢性暴露于15 d-PGJ 2的影响。因为我们发现游离15 d-PGJ 2的浓度在细胞培养基中迅速降低，所以每天两次更换新鲜制备的含有15 d-PGJ 2、CAY 10410或媒介物的培养基以维持稳定的细胞外浓度。用2.5 μM 15 d-PGJ 2孵育（而非CAY 10410）可增加神经元细胞死亡，但不诱导半胱天冬酶-3或PARP裂解，这与15 d-PGJ 2诱导的细胞死亡的主要坏死机制一致，TUNEL测定结果进一步支持了这一点。在96 h 15 d-PGJ 2孵育后观察到泛素化蛋白积累和聚集，伴随着受损的20 S蛋白酶体活性。与另一种蛋白酶体抑制剂MG 132不同，15 d-PGJ 2处理不激活自噬或诱导侵袭体形成。因此，脑卒中后CyPG生成增加的累积细胞毒性效应可能导致迟发性缺血后神经元损伤。
The cyclopentenone prostaglandin (CyPG) J2 series, including prostaglandin J2 (PGJ2), Δ12-PGJ2 and 15-deoxy-Δ12, 14 -prostaglandin J2 (15d-PGJ2), are active metabolites of PGD2, exerting multiple effects on neuronal function. However, the physiologic relevance of these effects remains uncertain as brain concentrations of CyPGs have not been precisely determined. In this study, we found that free PGD2 and the J2 series CyPGs (PGJ2, Δ12-PGJ2 and 15d-PGJ2) were increased in post-ischemic rat brain as detected by UPLC-MS/MS with 15d-PGJ2 being the most abundant CyPG. These increases were attenuated by pre-treating with the cyclooxygenase inhibitor piroxicam. Next, effects of chronic exposure to 15d-PGJ2 were examined by treating primary neurons with 15d-PGJ2, CAY10410 (a 15d-PGJ2 analog lacking the cyclopentenone ring structure), or vehicle for 24 h to 96 h. Because we found that the concentration of free 15d-PGJ2 decreased rapidly in cell culture medium, freshly prepared medium containing 15d-PGJ2, CAY10410 or vehicle was changed twice daily to maintain steady extracellular concentrations. Incubation with 2.5 μM 15d-PGJ2, but not CAY10410, increased neuronal cell death without induction of caspase-3 or PARP cleavage, consistent with a primarily necrotic mechanism for 15d-PGJ2-induced cell death which was further supported by TUNEL assay results. Ubiquitinated protein accumulation and aggregation was observed after 96 h 15d-PGJ2 incubation, accompanied by compromised 20S proteasome activity. Unlike another proteasome inhibitor, MG132, 15d-PGJ2 treatment did not activate autophagy or induce aggresome formation. Therefore, the cumulative cytotoxic effects of increased generation of CyPGs after stroke may contribute to delayed post-ischemic neuronal injury.
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