LncRNA GAS5 promotes spermidine‑induced autophagy through the miRNA‑31‑5p/NAT8L axis in pulmonary artery endothelial cells of patients with CTEPH

Wu,Qinghua; Zhou,Xiaohui; Wang,Yan; Hu,Yamin

doi:10.3892/mmr.2022.12813

LncRNA GAS5 promotes spermidine‑induced autophagy through the miRNA‑31‑5p/NAT8L axis in pulmonary artery endothelial cells of patients with CTEPH

Authors:
- Qinghua Wu
- Xiaohui Zhou
- Yan Wang
- Yamin Hu
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Affiliations: Department of Cardiovascular Medicine, Cangzhou Central Hospital, Cangzhou, Hebei 061001, P.R. China
Published online on: August 2, 2022 https://doi.org/10.3892/mmr.2022.12813
Article Number: 297
Copyright: © Wu et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Chronic thromboembolic pulmonary hypertension (CTEPH) is a leading cause of pulmonary hypertension. The present study investigated the mechanisms of long non‑coding RNA growth arrest‑specific transcript 5 (GAS5) on spermidine (SP)‑induced autophagy. Pulmonary artery endothelial cells (PAECs) were collected from patients with CTEPH and the rat model. Immunofluorescence, Western blots, reverse transcription‑quantitative polymerase chain reaction, bioinformatics, rapid amplification of cDNA ends assays, luciferase reporter assays, RNA‑binding protein immunoprecipitation assays, GFP‑LC3 adenoviruses, tfLC3 assays and transmission electron microscopy were performed. The results revealed that SP‑induced autophagy increased GAS5 in PAECs. The upregulation of GAS5 enhanced and the downregulation of GAS5 reversed the roles of SP in PAECs. Furthermore, GAS5 promoted SP‑induced autophagy in PAECs by targeting miRNA‑31‑5p. The miRNA‑31‑5p mimic suppressed and the inhibitor promoted SP‑induced autophagy. Furthermore, N‑Acetyltransferase 8 Like (NAT8L) was a target gene of miRNA‑31‑5p and knockdown of NAT8L inhibited the autophagic levels of PAECs. In vivo, SP treatment decreased miRNA‑31‑5p and increased NAT8L levels, which was reversed by the knockdown of GAS5. The downregulation of GAS5 abolished the stimulatory role of SP in PAECs of CTEPH rats. In conclusion, GAS5 promoted SP‑induced autophagy through miRNA‑31‑5p/NAT8L signaling pathways in vitro and in vivo and GAS5 may be a promising molecular marker for therapies of CTEPH.1

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1	Pengo V, Lensing AW, Prins MH, Marchiori A, Davidson BL, Tiozzo F, Albanese P, Biasiolo A, Pegoraro C, Iliceto S, et al: Incidence of chronic thromboembolic pulmonary hypertension after pulmonary embolism. N Engl J Med. 350:2257–2264. 2004. View Article : Google Scholar : PubMed/NCBI
2	Becattini C, Agnelli G, Pesavento R, Silingardi M, Poggio R, Taliani MR and Ageno W: Incidence of chronic thromboembolic pulmonary hypertension after a first episode of pulmonary embolism. Chest. 130:172–175. 2006. View Article : Google Scholar
3	Sakao S and Tatsumi K: Crosstalk between endothelial cell and thrombus in chronic thromboembolic pulmonary hypertension: Perspective. Histol Histopathol. 28:185–193. 2013.
4	Lang IM, Pesavento R, Bonderman D and Yuan JX: Risk factors and basic mechanisms of chronic thromboembolic pulmonary hypertension: A current understanding. Eur Respir J. 41:462–468. 2013. View Article : Google Scholar : PubMed/NCBI
5	Jin Y and Choi AM: Cross talk between autophagy and apoptosis in pulmonary hypertension. Pulm Circ. 2:407–414. 2012. View Article : Google Scholar
6	Levine B and Kroemer G: Autophagy in the pathogenesis of disease. Cell. 132:27–42. 2008. View Article : Google Scholar
7	Lee SJ, Smith A, Guo L, Alastalo TP, Li M, Sawada H, Liu X, Chen ZH, Ifedigbo E, Jin Y, et al: Autophagic protein LC3B confers resistance against hypoxia-induced pulmonary hypertension. Am J Respir Crit Care Med. 183:649–658. 2011. View Article : Google Scholar : PubMed/NCBI
8	Lahm T, Albrecht M, Fisher AJ, Selej M, Patel NG, Brown JA, Justice MJ, Brown MB, Van Demark M, Trulock KM, et al: 17β-Estradiol attenuates hypoxic pulmonary hypertension via estrogen receptor-mediated effects. Am J Respir Crit Care Med. 185:965–980. 2012. View Article : Google Scholar : PubMed/NCBI
9	Long L, Yang X, Southwood M, Lu J, Marciniak SJ, Dunmore BJ and Morrell NW: Chloroquine prevents progression of experimental pulmonary hypertension via inhibition of autophagy and lysosomal bone morphogenetic protein type II receptor degradation. Circ Res. 112:1159–1170. 2013. View Article : Google Scholar : PubMed/NCBI
10	Pegg AE: Mammalian polyamine metabolism and function. IUBMB Life. 61:880–894. 2009. View Article : Google Scholar : PubMed/NCBI
11	Madeo F, Eisenberg T, Pietrocola F and Kroemer G: Spermidine in health and disease. Science. 359:eaan27882018. View Article : Google Scholar : PubMed/NCBI
12	LaRocca TJ, Gioscia-Ryan RA, Hearon CM Jr and Seals DR: The autophagy enhancer spermidine reverses arterial aging. Mech Ageing Dev. 134:314–320. 2013. View Article : Google Scholar : PubMed/NCBI
13	Eisenberg T, Knauer H, Schauer A, Büttner S, Ruckenstuhl C, Carmona-Gutierrez D, Ring J, Schroeder S, Magnes C, Antonacci L, et al: Induction of autophagy by spermidine promotes longevity. Nat Cell Biol. 11:1305–1314. 2009. View Article : Google Scholar
14	LaRocca TJ, Henson GD, Thorburn A, Sindler AL, Pierce GL and Seals DR: Translational evidence that impaired autophagy contributes to arterial ageing. J Physiol. 590:3305–3316. 2012. View Article : Google Scholar
15	Wilusz JE, Sunwoo H and Spector DL: Long noncoding RNAs: Functional surprises from the RNA world. Genes Dev. 23:1494–1504. 2009. View Article : Google Scholar : PubMed/NCBI
16	Wang KC and Chang HY: Molecular mechanisms of long noncoding RNAs. Mol Cell. 43:904–914. 2011. View Article : Google Scholar
17	Xue D, Zhou C, Lu H, Xu R, Xu X and He X: LncRNA GAS5 inhibits proliferation and progression of prostate cancer by targeting miR-103 through AKT/mTOR signaling pathway. Tumour Biol. 37:16187–16197. 2016. View Article : Google Scholar
18	Pickard M, Mourtada-Maarabouni M and Williams G: Long non-coding RNA GAS5 regulates apoptosis in prostate cancer cell lines. Biochim Biophys Acta. 1832:1613–1623. 2013. View Article : Google Scholar
19	Tao H, Zhang JG, Qin RH, Dai C, Shi P, Yang JJ, Deng ZY and Shi KH: LncRNA GAS5 controls cardiac fibroblast activation and fibrosis by targeting miR-21 via PTEN/MMP-2 signaling pathway. Toxicology. 386:11–18. 2017. View Article : Google Scholar : PubMed/NCBI
20	Ye K, Wang S, Zhang H, Han H, Ma B and Nan W: Long noncoding RNA GAS5 suppresses cell growth and epithelial-mesenchymal transition in osteosarcoma by regulating the miR-221/ARHI pathway. J Cell Biochem. 118:4772–4781. 2017. View Article : Google Scholar : PubMed/NCBI
21	Gu J, Wang Y, Wang X, Zhou D, Wang X, Zhou M and He Z: Effect of the LncRNA GAS5-MiR-23a-ATG3 axis in regulating autophagy in patients with breast cancer. Cell Physiol Biochem. 48:194–207. 2018. View Article : Google Scholar : PubMed/NCBI
22	Zhang N, Yang GQ, Shao XM and Wei L: GAS5 modulated autophagy is a mechanism modulating cisplatin sensitivity in NSCLC cells. Eur Rev Med Pharmacol Sci. 20:2271–2277. 2016.
23	Pulito C, Mori F, Sacconi A, Goeman F, Ferraiuolo M, Pasanisi P, Campagnoli C, Berrino F, Fanciulli M, Ford RJ, et al: Metformin-induced ablation of microRNA 21-5p releases Sestrin-1 and CAB39L antitumoral activities. Cell Discov. 3:170222017. View Article : Google Scholar : PubMed/NCBI
24	Rasband W: ImageJ software. US National Institutes of Health; Bethesda, Maryland, USA: 2011
25	Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(−Delta Delta C(T)) method. Methods. 25:402–408. 2001. View Article : Google Scholar : PubMed/NCBI
26	Yin QF, Yang L, Zhang Y, Xiang JF, Wu YW, Carmichael GG and Chen LL: Long noncoding RNAs with snoRNA ends. Mol Cell. 48:219–230. 2012. View Article : Google Scholar
27	Volders PJ, Anckaert J, Verheggen K, Nuytens J, Martens L, Mestdagh P and Vandesompele J: LNCipedia 5: Towards a reference set of human long non-coding RNAs. Nucleic Acids Res. 47:D135–D139. 2019. View Article : Google Scholar : PubMed/NCBI
28	Cao Z, Pan X, Yang Y, Huang Y and Shen HB: The lncLocator: A subcellular localization predictor for long non-coding RNAs based on a stacked ensemble classifier. Bioinformatics. 34:2185–2194. 2018. View Article : Google Scholar : PubMed/NCBI
29	Li JH, Liu S, Zhou H, Qu LH and Yang JH: starBase v2.0: Decoding miRNA-ceRNA, miRNA-ncRNA and protein-RNA interaction networks from large-scale CLIP-Seq data. Nucleic Acids Res. 42:(Database Issue). D92–D97. 2014. View Article : Google Scholar : PubMed/NCBI
30	Agarwal V, Bell GW, Nam JW and Bartel DP: Predicting effective microRNA target sites in mammalian mRNAs. Elife. 4:e050052015. View Article : Google Scholar
31	Betel D, Koppal A, Agius P, Sander C and Leslie C: Comprehensive modeling of microRNA targets predicts functional non-conserved and non-canonical sites. Genome Biol. 11:R902010. View Article : Google Scholar
32	Deng C, Wu D, Yang M, Chen Y, Ding H, Zhong Z, Lian N, Zhang Q, Wu S and Liu K: The role of tissue factor and autophagy in pulmonary vascular remodeling in a rat model for chronic thromboembolic pulmonary hypertension. Respir Res. 17:652016. View Article : Google Scholar : PubMed/NCBI
33	Wang K, Liu CY, Zhou LY, Wang JX, Wang M, Zhao B, Zhao WK, Xu SJ, Fan LH, Zhang XJ, et al: APF lncRNA regulates autophagy and myocardial infarction by targeting miR-188-3p. Nat Commun. 6:67792015. View Article : Google Scholar : PubMed/NCBI
34	Cao Y and Klionsky DJ: Physiological functions of Atg6/beclin 1: A unique autophagy-related protein. Cell Res. 17:839–849. 2007. View Article : Google Scholar : PubMed/NCBI
35	Komatsu M, Tanida I, Ueno T, Ohsumi M, Ohsumi Y and Kominami E: The C-terminal region of an Apg7p/Cvt2p is required for homodimerization and is essential for its E1 activity and E1-E2 complex formation. J Biol Chem. 276:9846–9854. 2001. View Article : Google Scholar : PubMed/NCBI
36	Ichimura Y, Kirisako T, Takao T, Satomi Y, Shimonishi Y, Ishihara N, Mizushima N, Tanida I, Kominami E, Ohsumi M, et al: A ubiquitin-like system mediates protein lipidation. Nature. 408:488–492. 2000. View Article : Google Scholar : PubMed/NCBI
37	Mercier O, Arthur Ataam J, Langer NB, Dorfmüller P, Lamrani L, Lecerf F, Decante B, Dartevelle P, Eddahibi S and Fadel E: Abnormal pulmonary endothelial cells may underlie the enigmatic pathogenesis of chronic thromboembolic pulmonary hypertension. J Heart Lung Transplant. 36:305–314. 2017. View Article : Google Scholar
38	Casero RA Jr and Marton LJ: Targeting polyamine metabolism and function in cancer and other hyperproliferative diseases. Nat Rev Drug Discov. 6:373–390. 2007. View Article : Google Scholar : PubMed/NCBI
39	Cai Y, Yu X, Hu S and Yu J: A brief review on the mechanisms of miRNA regulation. Genomics Proteomics Bioinformatics. 7:147–154. 2009. View Article : Google Scholar : PubMed/NCBI
40	Kucher N, Rossi E and Derosa M: Massive pulmonary embolism. J Vasc Surg. 44:684–685. 2006. View Article : Google Scholar
41	Gerges C, Skoro-Sajer N and Lang IM: Right ventricle in acute and chronic pulmonary embolism (2013 Grover Conference series). Pulm Circ. 4:378–386. 2014. View Article : Google Scholar
42	Eisenberg T, Abdellatif M, Schroeder S, Primessnig U, Stekovic S, Pendl T, Harger A, Schipke J, Zimmermann A, Schmidt A, et al: Cardioprotection and lifespan extension by the natural polyamine spermidine. Nat Med. 22:1428–1438. 2016. View Article : Google Scholar : PubMed/NCBI
43	Minois N, Carmona-Gutierrez D, Bauer MA, Rockenfeller P, Eisenberg T, Brandhorst S, Sigrist SJ, Kroemer G and Madeo F: Spermidine promotes stress resistance in Drosophila melanogaster through autophagy-dependent and -independent pathways. Cell Death Dis. 3:e4012012. View Article : Google Scholar : PubMed/NCBI
44	Mizushima N and Komatsu M: Autophagy: Renovation of cells and tissues. Cell. 147:728–741. 2011. View Article : Google Scholar
45	Morselli E, Mariño G, Bennetzen MV, Eisenberg T, Megalou E, Schroeder S, Cabrera S, Bénit P, Rustin P, Criollo A, et al: Spermidine and resveratrol induce autophagy by distinct pathways converging on the acetylproteome. J Cell Biol. 192:615–629. 2011. View Article : Google Scholar
46	Ponting CP, Oliver PL and Reik W: Evolution and functions of long noncoding RNAs. Cell. 136:629–641. 2009. View Article : Google Scholar
47	Huo JF and Chen XB: Long noncoding RNA growth arrest-specific 5 facilitates glioma cell sensitivity to cisplatin by suppressing excessive autophagy in an mTOR-dependent manner. J Cell Biochem. 120:6127–6136. 2019. View Article : Google Scholar : PubMed/NCBI
48	Paraskevopoulou MD and Hatzigeorgiou AG: Analyzing miRNA-lncRNA interactions. Long Non-Coding RNAs. Methods in Molecular Biology. 1402. Feng Y and Zhang L: Humana Press; New York, NY: pp. 271–286. 2016, View Article : Google Scholar
49	Yoon J, Abdelmohsen K and Gorospe M: Functional interactions among microRNAs and long noncoding RNAs. Seminars in cell and developmental biology. Elsevier; pp. 9–14. 2014, View Article : Google Scholar
50	Hessam S, Sand M, Skrygan M, Gambichler T and Bechara FG: Expression of miRNA-155, miRNA-223, miRNA-31, miRNA-21, miRNA-125b, and miRNA-146a in the inflammatory pathway of hidradenitis suppurativa. Inflammation. 40:464–472. 2017. View Article : Google Scholar : PubMed/NCBI
51	Kim S, Lee KS, Choi S, Kim J, Lee DK, Park M, Park W, Kim TH, Hwang JY, Won MH, et al: NF-κB-responsive miRNA-31-5p elicits endothelial dysfunction associated with preeclampsia via down-regulation of endothelial nitric-oxide synthase. J Biol Chem. 293:18989–19000. 2018. View Article : Google Scholar : PubMed/NCBI
52	Mehta V and Namboodiri M: N-acetylaspartate as an acetyl source in the nervous system. Brain Res Mol Brain Res. 31:151–157. 1995. View Article : Google Scholar : PubMed/NCBI
53	Huang S, Lu W, Ge D, Meng N, Li Y, Su L, Zhang S, Zhang Y, Zhao B and Miao J: A new microRNA signal pathway regulated by long noncoding RNA TGFB2-OT1 in autophagy and inflammation of vascular endothelial cells. Autophagy. 11:2172–2183. 2015. View Article : Google Scholar : PubMed/NCBI
54	Huber K, Hofer DC, Trefely S, Pelzmann HJ, Madreiter-Sokolowski C, Duta-Mare M, Schlager S, Trausinger G, Stryeck S, Graier WF, et al: N-acetylaspartate pathway is nutrient responsive and coordinates lipid and energy metabolism in brown adipocytes. Biochim Biophys Acta Mol Cell Res. 1866:337–348. 2019. View Article : Google Scholar : PubMed/NCBI