Department of Urology, Zhejiang Provincial People's Hospital, People's Hospital of Hangzhou Medical College, Zhejiang, China.
Department of Urology, Tongde Hospital of Zhejiang Province, Zhejiang, China.
PLoS One. 2021 Sep 1;16(9):e0256734. doi: 10.1371/journal.pone.0256734. eCollection 2021.
This study is to investigate whether the cardiac microvascular endothelial cells (CMECs) can regulate the autophagy of cardiomyocytes (CMs) by secreting lncRNA-ANRIL/miR-181b exosomes, thus participating in the occurrence of uremic cardiovascular disease (CVD).
A 5/6 nephrectomy uremia model was established, with the mice injected with ANRIL-shRNA lentivirus vector, miR-181b agomir, and related control reagents, containing the serum creatinine and urea nitrogen measured. The renal tissue sections of mice were stained with Periodic Acid-Schiff (PAS), TUNEL, and Hematoxylin-Eosin (HE) performed on myocardial tissue sections of mice. ANRIL-shRNA, miR-181b mimics, and related control reagents were transfected into CMECs, in which the exosomes were extracted and co-cultured with CMs. The expressions of ANRIL, miR-181b and ATG5 were detected by qRT-PCR, and the expressions of autophagy related proteins by Western blot, as well as the binding of ANRIL and miR-181b by the double luciferase reporter gene experiment.
ANRIL down-regulation or miR-181b up-regulation can increase the weight of mice with uremia, as well as the expressions of p62 and miR-181b, and reduce the content of serum creatinine and urea nitrogen, the damage of kidney and myocardial tissues, the number of apoptotic cells in myocardial tissues, as well as the expressions of ANRIL, ATG5, Beclin1, and LC3. CMs can absorb the exosomes of CMECs. Compared with IS+ CMEC-Exo group, the expressions of ANRIL and ATG5 in CMs of IS+ CMEC-Exo + sh lncRNA ANRIL and IS+CMEC-Exo+miR-181b mimics groups was down-regulated, as well as the expressions of ATG5, Beclin1, and LC3, while miR-181b expression was up-regulated as well as P62 expression.
CMECs can regulate autophagy of CMs by releasing exosomes containing ANRIL and miR-181b.
本研究旨在探讨心脏微血管内皮细胞（CMECs）是否可以通过分泌 lncRNA-ANRIL/miR-181b 外泌体来调节心肌细胞（CMs）的自噬，从而参与尿毒症性心血管疾病（CVD）的发生。
建立 5/6 肾切除尿毒症模型，将 ANRIL-shRNA 慢病毒载体、miR-181b 激动剂和相关对照试剂注射入小鼠体内，测量血清肌酐和尿素氮。对小鼠的肾组织切片进行过碘酸希夫（PAS）、TUNEL 和苏木精-伊红（HE）染色，对小鼠的心肌组织切片进行染色。将 ANRIL-shRNA、miR-181b 模拟物和相关对照试剂转染到 CMECs 中，提取外泌体并与 CMs 共培养。通过 qRT-PCR 检测 ANRIL、miR-181b 和 ATG5 的表达，通过 Western blot 检测自噬相关蛋白的表达，以及通过双荧光素酶报告基因实验检测 ANRIL 和 miR-181b 的结合。
下调 ANRIL 或上调 miR-181b 可以增加尿毒症小鼠的体重，以及 p62 和 miR-181b 的表达，降低血清肌酐和尿素氮的含量，肾脏和心肌组织的损伤，心肌组织中凋亡细胞的数量，以及 ANRIL、ATG5、Beclin1 和 LC3 的表达。CMs 可以吸收 CMECs 的外泌体。与 IS+CMEC-Exo 组相比，IS+CMEC-Exo+sh lncRNA ANRIL 和 IS+CMEC-Exo+miR-181b mimics 组的 CMs 中 ANRIL 和 ATG5 的表达下调，以及 ATG5、Beclin1 和 LC3 的表达下调，而 miR-181b 的表达上调，P62 的表达下调。
CMECs 可以通过释放含有 ANRIL 和 miR-181b 的外泌体来调节 CMs 的自噬。
