Skip to main content

Advertisement

SpringerLink
Log in

MicroRNA-27a Promotes Inefficient Lysosomal Clearance in the Hippocampi of Rats Following Chronic Brain Hypoperfusion

  • Published:
Molecular Neurobiology Aims and scope Submit manuscript

Abstract

Chronic brain hypoperfusion (CBH) induces the accumulation of abnormal cellular proteins, accompanied by cognitive decline, and the autophagic-lysosomal system is abnormal in dementia. Whether CBH accounts for autophagic-lysosomal neuropathology remains unknown. Here, we show that CBH significantly increased the number of autophagic vacuoles (AVs) with high LC3-II levels, but decreased SQSTM1 and cathepsin D levels in the hippocampi of rats following bilateral common carotid artery occlusion (2VO) for 2 weeks. Further studies showed that microRNA-27a (Mir27a) was upregulated at 2 weeks compared with the sham group. Additionally, LAMP-2 proteins were downregulated by Mir27a overexpression, upregulated by Mir27a inhibition, and unchanged by binding-site mutations or miR-masks, indicating that lamp-2 is the target of Mir27a. Knockdown of endogenous Mir27a prevented the reduction of LAMP-2 protein expression as well as the accumulation of AVs in the hippocampi of 2VO rats. Overexpression of Mir27a induced, while the knockdown of Mir27a reduced, the accumulation of AVs and the LC3-II level in cultured neonatal rat neurons. The results revealed that CBH in rats at 2 weeks could induce inefficient lysosomal clearance, which is regulated by the Mir27a-mediated downregulation of LAMP-2 protein expression. These findings provide an insight into a novel molecular mechanism of autophagy at the miRNA level.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8

Similar content being viewed by others

Abbreviations

miRNA:

MicroRNA

Mir27a:

MicroRNA-27a

AMO-27a:

2′-O-Methyl antisense oligoribonucleotides to miR-27a

NC:

Scramble negative control

ODN:

miRNA-masking antisense oligodeoxynucleotides (miR-masks)

LC3:

Microtubule-associated protein 1 light chain 3

LAMP-2:

Lysosomal-associated membrane protein-2

SQSTM1/P62:

Sequestosome1

MTOR:

Mechanistic target of rapamycin

p70S6k:

Ribosomal protein S6 kinase 70kDa

AVs:

Autophagic vaculoses

AL:

Autolysosome

3MA:

3-Methyladenine

Rap:

Rapamycin

3′UTR:

3′-Untranslated region

RT-PCR:

Reverse transcription-polymerase chain reaction

2VO:

Bilateral common carotid artery occlusion

References

  1. Mizushima N, Levine B, Cuervo AM, Klionsky DJ (2008) Autophagy fights disease through cellular self-digestion. Nature 451:1069–1075. doi:10.1038/nature06639

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Nixon RA (2006) Autophagy in neurodegenerative disease: friend, foe or turncoat? Trends Neurosci 29:528–535

    Article  CAS  PubMed  Google Scholar 

  3. Hara T, Nakamura K, Matsui M, Yamamoto A, Nakahara Y, Suzuki-Migishima R, Yokoyama M, Mishima K, Saito I, Okano H, Mizushima N (2006) Suppression of basal autophagy in neural cells causes neurodegenerative disease in mice. Nature 441:885–889

    Article  CAS  PubMed  Google Scholar 

  4. Cuervo AM, Stefanis L, Fredenburg R, Lansbury PT, Sulzer D (2004) Impaired degradation of mutant alpha-synuclein by chaperone-mediated autophagy. Science 305:1292–1295

    Article  CAS  PubMed  Google Scholar 

  5. Kegel KB, Kim M, Sapp E, McIntyre C, Castano JG, Aronin N, DiFiglia M (2000) Huntingtin expression stimulates endosomal-lysosomal activity, endosome tubulation, and autophagy. J Neurosci 20:7268–7278

    CAS  PubMed  Google Scholar 

  6. Yu WH, Cuervo AM, Kumar A, Peterhoff CM, Schmidt SD, Lee JH, Mohan PS, Mercken M, Farmery MR, Tjernberg LO, Jiang Y, Duff K, Uchiyama Y, Naslund J, Mathews PM, Cataldo AM, Nixon RA (2005) Macroautophagy—a novel Beta-amyloid peptide-generating pathway activated in Alzheimer’s disease. J Cell Biol 171:87–98

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Lee JH, Yu WH, Kumar A, Lee S, Mohan PS, Peterhoff CM, Wolfe DM, Martinez-Vicente M, Massey AC, Sovak G, Uchiyama Y, Westaway D, Cuervo AM, Nixon RA (2010) Lysosomal proteolysis and autophagy require presenilin 1 and are disrupted by Alzheimer-related PS1 mutations. Cell 141:1146–1158. doi:10.1016/j.cell.2010.05.008

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Funderburk SF, Marcellino BK, Yue Z (2010) Cell “self-eating” (autophagy) mechanism in Alzheimer’s disease. Mt Sinai J Med 77:59–68. doi:10.1002/msj.20161

    Article  PubMed  PubMed Central  Google Scholar 

  9. Nixon RA, Yang DS (2011) Autophagy failure in Alzheimer’s disease—locating the primary defect. Neurobiol Dis 43:38–45. doi:10.1016/j.nbd.2011.01.021

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Boland B, Kumar A, Lee S, Platt FM, Wegiel J, Yu WH, Nixon RA (2008) Autophagy induction and autophagosome clearance in neurons: relationship to autophagic pathology in Alzheimer’s disease. J Neurosci 28:6926–6937. doi:10.1523/JNEUROSCI.0800-08.2008

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Ruitenberg A, den Heijer T, Bakker SL, van Swieten JC, Koudstaal PJ, Hofman A, Breteler MM (2005) Cerebral hypoperfusion and clinical onset of dementia: the Rotterdam Study. Ann Neurol 57:789–794

    Article  PubMed  Google Scholar 

  12. Gorelick PB, Scuteri A, Black SE, Decarli C, Greenberg SM, Iadecola C, Launer LJ, Laurent S, Lopez OL, Nyenhuis D, Petersen RC, Schneider JA, Tzourio C, Arnett DK, Bennett DA, Chui HC, Higashida RT, Lindquist R, Nilsson PM, Roman GC, Sellke FW, Seshadri S (2011) Vascular contributions to cognitive impairment and dementia: a statement for healthcare professionals from the American Heart Association/American Stroke Association. Stroke 42:2672–2713. doi:10.1161/STROKEAHA.111.634279

    Article  PubMed  PubMed Central  Google Scholar 

  13. Farkas E, Luiten PG, Bari F (2007) Permanent, bilateral common carotid artery occlusion in the rat: a model for chronic cerebral hypoperfusion-related neurodegenerative diseases. Brain Res Rev 54:162–180

    Article  CAS  PubMed  Google Scholar 

  14. Ai J, Sun LH, Che H, Zhang R, Zhang TZ, Wu WC, Su XL, Chen X, Yang G, Li K, Wang N, Ban T, Bao YN, Guo F, Niu HF, Zhu YL, Zhu XY, Zhao SG, Yang BF (2013) MicroRNA-195 protects against dementia induced by chronic brain hypoperfusion via its anti-amyloidogenic effect in rats. J Neurosci 33:3989–4001. doi:10.1523/JNEUROSCI.1997-12.2013

    Article  CAS  PubMed  Google Scholar 

  15. Kitaguchi H, Tomimoto H, Ihara M, Shibata M, Uemura K, Kalaria RN, Kihara T, Asada-Utsugi M, Kinoshita A, Takahashi R (2009) Chronic cerebral hypoperfusion accelerates amyloid beta deposition in APPSwInd transgenic mice. Brain Res 1294:202–210. doi:10.1016/j.brainres.2009.07.078

    Article  CAS  PubMed  Google Scholar 

  16. Zhiyou C, Yong Y, Shanquan S, Jun Z, Liangguo H, Ling Y, Jieying L (2009) Upregulation of BACE1 and beta-amyloid protein mediated by chronic cerebral hypoperfusion contributes to cognitive impairment and pathogenesis of Alzheimer’s disease. Neurochem Res 34:1226–1235. doi:10.1007/s11064-008-9899-y

    Article  PubMed  Google Scholar 

  17. Sun LH, Ban T, Liu CD, Chen QX, Wang X, Yan ML, Hu XL, Su XL, Bao YN, Sun LL, Zhao LJ, Pei SC, Jiang XM, Zong DK, Ai J (2015) Activation of Cdk5/p25 and tau phosphorylation following chronic brain hypoperfusion in rats involves microRNA-195 down-regulation. J Neurochem 134:1139–1151. doi:10.1111/jnc.13212

    Article  CAS  PubMed  Google Scholar 

  18. Farkas E, Institoris A, Domoki F, Mihaly A, Bari F (2006) The effect of pre- and posttreatment with diazoxide on the early phase of chronic cerebral hypoperfusion in the rat. Brain Res 1087:168–174

    Article  CAS  PubMed  Google Scholar 

  19. Liu HX, Zhang JJ, Zheng P, Zhang Y (2005) Altered expression of MAP-2, GAP-43, and synaptophysin in the hippocampus of rats with chronic cerebral hypoperfusion correlates with cognitive impairment. Brain Res Mol Brain Res 139:169–177

    Article  CAS  PubMed  Google Scholar 

  20. Alvarez-Erviti L, Seow Y, Schapira AH, Rodriguez-Oroz MC, Obeso JA, Cooper JM (2013) Influence of microRNA deregulation on chaperone-mediated autophagy and alpha-synuclein pathology in Parkinson’s disease. Cell Death Dis 4:e545. doi:10.1038/cddis.2013.73

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Mertens-Talcott SU, Chintharlapalli S, Li X, Safe S (2007) The oncogenic microRNA-27a targets genes that regulate specificity protein transcription factors and the G2-M checkpoint in MDA-MB-231 breast cancer cells. Cancer Res 67:11001–11011

    Article  CAS  PubMed  Google Scholar 

  22. Tang W, Zhu J, Su S, Wu W, Liu Q, Su F, Yu F (2012) MiR-27 as a prognostic marker for breast cancer progression and patient survival. PLoS One 7:e51702. doi:10.1371/journal.pone.0051702

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Shirasaki T, Honda M, Shimakami T, Horii R, Yamashita T, Sakai Y, Sakai A, Okada H, Watanabe R, Murakami S, Yi M, Lemon SM, Kaneko S (2013) MicroRNA-27a regulates lipid metabolism and inhibits hepatitis C virus replication in human hepatoma cells. J Virol 87:5270–5286. doi:10.1128/JVI.03022-12

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Yeh CH, Chen TP, Wang YC, Lin YM, Fang SW (2012) MicroRNA-27a regulates cardiomyocytic apoptosis during cardioplegia-induced cardiac arrest by targeting interleukin 10-related pathways. Shock 38:607–614. doi:10.1097/SHK.0b013e318271f944

    Article  CAS  PubMed  Google Scholar 

  25. Kulshreshtha R, Davuluri RV, Calin GA, Ivan M (2008) A microRNA component of the hypoxic response. Cell Death Differ 15:667–671. doi:10.1038/sj.cdd.4402310

    Article  CAS  PubMed  Google Scholar 

  26. Chen Q, Xu J, Li L, Li H, Mao S, Zhang F, Zen K, Zhang CY, Zhang Q (2014) MicroRNA-23a/b and microRNA-27a/b suppress Apaf-1 protein and alleviate hypoxia-induced neuronal apoptosis. Cell Death Dis 5:e1132. doi:10.1038/cddis.2014.92

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Kang BY, Park KK, Green DE, Bijli KM, Searles CD, Sutliff RL, Hart CM (2013) Hypoxia mediates mutual repression between microRNA-27a and PPARgamma in the pulmonary vasculature. PLoS One 8:e79503. doi:10.1371/journal.pone.0079503

    Article  PubMed  PubMed Central  Google Scholar 

  28. Wu X, Bhayani MK, Dodge CT, Nicoloso MS, Chen Y, Yan X, Adachi M, Thomas L, Galer CE, Jiffar T, Pickering CR, Kupferman ME, Myers JN, Calin GA, Lai SY (2013) Coordinated targeting of the EGFR signaling axis by microRNA-27a*. Oncotarget 4:1388–1398

    Article  PubMed  PubMed Central  Google Scholar 

  29. Kumaran D, Udayabanu M, Kumar M, Aneja R, Katyal A (2008) Involvement of angiotensin converting enzyme in cerebral hypoperfusion induced anterograde memory impairment and cholinergic dysfunction in rats. Neuroscience 155:626–639. doi:10.1016/j.neuroscience.2008.06.023

    Article  CAS  PubMed  Google Scholar 

  30. Carloni S, Girelli S, Scopa C, Buonocore G, Longini M, Balduini W (2010) Activation of autophagy and Akt/CREB signaling play an equivalent role in the neuroprotective effect of rapamycin in neonatal hypoxia-ischemia. Autophagy 6:366–377

    Article  CAS  PubMed  Google Scholar 

  31. Baek SH, Noh AR, Kim KA, Akram M, Shin YJ, Kim ES, Yu SW, Majid A, Bae ON (2014) Modulation of mitochondrial function and autophagy mediates carnosine neuroprotection against ischemic brain damage. Stroke 45:2438–2443. doi:10.1161/STROKEAHA.114.005183

    Article  PubMed  PubMed Central  Google Scholar 

  32. Tanida I, Ueno T, Kominami E (2004) LC3 conjugation system in mammalian autophagy. Int J Biochem Cell Biol 36:2503–2518

    Article  CAS  PubMed  Google Scholar 

  33. Barth S, Glick D, Macleod KF (2010) Autophagy: assays and artifacts. J Pathol 221:117–124. doi:10.1002/path.2694

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Bjorkoy G, Lamark T, Brech A, Outzen H, Perander M, Overvatn A, Stenmark H, Johansen T (2005) p62/SQSTM1 forms protein aggregates degraded by autophagy and has a protective effect on huntingtin-induced cell death. J Cell Biol 171:603–614

    Article  PubMed  PubMed Central  Google Scholar 

  35. Pankiv S, Clausen TH, Lamark T, Brech A, Bruun JA, Outzen H, Overvatn A, Bjorkoy G, Johansen T (2007) p62/SQSTM1 binds directly to Atg8/LC3 to facilitate degradation of ubiquitinated protein aggregates by autophagy. J Biol Chem 282:24131–24145

    Article  CAS  PubMed  Google Scholar 

  36. Ma X, Liu H, Foyil SR, Godar RJ, Weinheimer CJ, Hill JA, Diwan A (2012) Impaired autophagosome clearance contributes to cardiomyocyte death in ischemia/reperfusion injury. Circulation 125:3170–3181. doi:10.1161/CIRCULATIONAHA.111.041814

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Bohley P, Seglen PO (1992) Proteases and proteolysis in the lysosome. Experientia 48:151–157

    Article  CAS  PubMed  Google Scholar 

  38. Amritraj A, Wang Y, Revett TJ, Vergote D, Westaway D, Kar S (2013) Role of cathepsin D in U18666A-induced neuronal cell death: potential implication in Niemann-Pick type C disease pathogenesis. J Biol Chem 288:3136–3152. doi:10.1074/jbc.M112.412460

    Article  CAS  PubMed  Google Scholar 

  39. Eskelinen EL (2006) Roles of LAMP-1 and LAMP-2 in lysosome biogenesis and autophagy. Mol Aspects Med 27:495–502

    Article  CAS  PubMed  Google Scholar 

  40. Saftig P, Beertsen W, Eskelinen EL (2008) LAMP-2: a control step for phagosome and autophagosome maturation. Autophagy 4:510–512

    Article  CAS  PubMed  Google Scholar 

  41. Saugstad JA (2010) MicroRNAs as effectors of brain function with roles in ischemia and injury, neuroprotection, and neurodegeneration. J Cereb Blood Flow Metab 30:1564–1576

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Lee ST, Chu K, Jung KH, Yoon HJ, Jeon D, Kang KM, Park KH, Bae EK, Kim M, Lee SK, Roh JK (2010) MicroRNAs induced during ischemic preconditioning. Stroke 41:1646–1651. doi:10.1038/jcbfm.2010.101

    Article  PubMed  Google Scholar 

  43. Levine B, Kroemer G (2008) Autophagy in the pathogenesis of disease. Cell 132:27–42. doi:10.1016/j.cell.2007.12.018

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Seglen PO, Gordon PB (1982) 3-Methyladenine: specific inhibitor of autophagic/lysosomal protein degradation in isolated rat hepatocytes. Proc Natl Acad Sci U S A 79:1889–1892

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Massey AC, Follenzi A, Kiffin R, Zhang C, Cuervo AM (2008) Autophagy 4:442–456

    Article  CAS  PubMed  Google Scholar 

  46. Yu L, McPhee CK, Zheng L, Mardones GA, Rong Y, Peng J, Mi N, Zhao Y, Liu Z, Wan F, Hailey DW, Oorschot V, Klumperman J, Baehrecke EH, Lenardo MJ (2010) Termination of autophagy and reformation of lysosomes regulated by mTOR. Nature 465:942–946. doi:10.1038/nature09076

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Chen Y, Yu L (2013) Autophagic lysosome reformation. Exp Cell Res 319:142–146. doi:10.1016/j.yexcr.2012.09.004

    Article  CAS  PubMed  Google Scholar 

  48. Adhami F, Schloemer A, Kuan CY (2007) The roles of autophagy in cerebral ischemia. Autophagy 3:42–44

    Article  CAS  PubMed  Google Scholar 

  49. Finn PF, Mesires NT, Vine M, Dice JF (2005) Effects of small molecules on chaperone-mediated autophagy. Autophagy 1:141–145

    Article  CAS  PubMed  Google Scholar 

  50. Althausen S, Mengesdorf T, Mies G, Olah L, Nairn AC, Proud CG, Paschen W (2001) Changes in the phosphorylation of initiation factor eIF-2alpha, elongation factor eEF-2 and p70 S6 kinase after transient focal cerebral ischaemia in mice. J Neurochem 78:779–787

    Article  CAS  PubMed  Google Scholar 

  51. Pastor MD, Garcia-Yebenes I, Fradejas N, Perez-Ortiz JM, Mora-Lee S, Tranque P, Moro MA, Pende M, Calvo S (2009) mTOR/S6 kinase pathway contributes to astrocyte survival during ischemia. J Biol Chem 284:22067–22078. doi:10.1074/jbc.M109.033100

    Article  CAS  PubMed  Google Scholar 

  52. Komatsu M, Waguri S, Koike M, Sou YS, Ueno T, Hara T, Mizushima N, Iwata J, Ezaki J, Murata S, Hamazaki J, Nishito Y, Iemura S, Natsume T, Yanagawa T, Uwayama J, Warabi E, Yoshida H, Ishii T, Kobayashi A, Yamamoto M, Yue Z, Uchiyama Y, Kominami E, Tanaka K (2007) Homeostatic levels of p62 control cytoplasmic inclusion body formation in autophagy-deficient mice. Cell 131:1149–1163

    Article  CAS  PubMed  Google Scholar 

  53. Kochl R, Hu XW, Chan EY, Tooze SA (2006) Microtubules facilitate autophagosome formation and fusion of autophagosomes with endosomes. Traffic 7:129–145

    Article  CAS  PubMed  Google Scholar 

  54. Tanaka Y, Guhde G, Suter A, Eskelinen EL, Hartmann D, Lullmann-Rauch R, Janssen PM, Blanz J, von Figura K, Saftig P (2000) Accumulation of autophagic vacuoles and cardiomyopathy in LAMP-2-deficient mice. Nature 406:902–906

    Article  CAS  PubMed  Google Scholar 

  55. Klionsky DJ, Abdalla FC, Abeliovich H et al (2012) Guidelines for the use and interpretation of assays for monitoring autophagy. Autophagy 8:445–544

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Cuervo AM (2010) Chaperone-mediated autophagy: selectivity pays off. Trends Endocrinol Metab 21(3):142–150. doi:10.1016/j.tem.2009.10.003

    Article  CAS  PubMed  Google Scholar 

Download references

Author information

Author notes

    Authors and Affiliations

    1. Department of Pharmacology, Harbin Medical University (the State-Province Key Laboratories of Biomedicine-Pharmaceutics of China), No.157 Baojian Road, Nangang District, Harbin, Heilongjiang Province, 150086, China

      Hui Che, Yan Yan, Xiao-Hui Kang, Fei Guo, Mei-Ling Yan, Tong Liu, De-Kang Zong, Lin-Lin Sun, Ya-Nan Bao, Li-Hua Sun, Bao-Feng Yang & Jing Ai

    2. Department of Neurosurgery, The First Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang Province, 150001, China

      Huai-Lei Liu & Xu Hou

    Authors
    1. Hui Che
      View author publications

      You can also search for this author in PubMed Google Scholar

    2. Yan Yan
      View author publications

      You can also search for this author in PubMed Google Scholar

    3. Xiao-Hui Kang
      View author publications

      You can also search for this author in PubMed Google Scholar

    4. Fei Guo
      View author publications

      You can also search for this author in PubMed Google Scholar

    5. Mei-Ling Yan
      View author publications

      You can also search for this author in PubMed Google Scholar

    6. Huai-Lei Liu
      View author publications

      You can also search for this author in PubMed Google Scholar

    7. Xu Hou
      View author publications

      You can also search for this author in PubMed Google Scholar

    8. Tong Liu
      View author publications

      You can also search for this author in PubMed Google Scholar

    9. De-Kang Zong
      View author publications

      You can also search for this author in PubMed Google Scholar

    10. Lin-Lin Sun
      View author publications

      You can also search for this author in PubMed Google Scholar

    11. Ya-Nan Bao
      View author publications

      You can also search for this author in PubMed Google Scholar

    12. Li-Hua Sun
      View author publications

      You can also search for this author in PubMed Google Scholar

    13. Bao-Feng Yang
      View author publications

      You can also search for this author in PubMed Google Scholar

    14. Jing Ai
      View author publications

      You can also search for this author in PubMed Google Scholar

    Corresponding author

    Correspondence to Jing Ai.

    Ethics declarations

    Funding

    This work was supported by the Natural Science Foundation of China (81070882, 81471115, 81271207 to J. A.) and the Creative Research Groups of the National Natural Science Foundation of China (81421063 to Y.B.F.).

    Conflict of Interest

    The authors declare that they have no conflict of interest.

    Additional information

    Hui Che and Yan Yan contributed equally to this work.

    Rights and permissions

    Reprints and permissions

    About this article

    Check for updates. Verify currency and authenticity via CrossMark

    Cite this article

    Che, H., Yan, Y., Kang, XH. et al. MicroRNA-27a Promotes Inefficient Lysosomal Clearance in the Hippocampi of Rats Following Chronic Brain Hypoperfusion. Mol Neurobiol 54, 2595–2610 (2017). https://doi.org/10.1007/s12035-016-9856-8

    Download citation

    • Received:

    • Accepted:

    • Published:

    • Issue Date:

    • DOI: https://doi.org/10.1007/s12035-016-9856-8

    Keywords

    Search

    Navigation