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Deciphering the role of circulating lncRNAs: RNCR2, NEAT2, CDKN2B-AS1, and PVT1 and the possible prediction of anti-VEGF treatment outcomes in diabetic retinopathy patients

  • Eman A. Toraih
  • Ahmed A. Abdelghany
  • Noha M Abd El Fadeal
  • Essam Al Ageeli
  • Manal S. FawzyEmail author
Basic Science
  • 56 Downloads

Abstract

Purpose

Putative roles of long non-coding RNAs (lncRNAs) as indicators for diabetic retinopathy (DR) and associated complications are beginning to emerge. We aimed to evaluate a panel of circulating hyperglycemia-related lncRNAs: RNCR2, NEAT2, CDKN2B-AS1, and PVT1 in type 2 diabetes patients with/without DR and to correlate their levels with the clinical characteristics and response to aflibercept intravitreal injection in terms of visual acuity (VA) improvement, central macular thickness (CMT) decline, and macular edema resolution after 4 weeks of the initial injection.

Methods

Pre-treatment plasma relative expression levels of the specified lncRNAs were quantified in 130 consecutive patients with diabetes (75 and 55 with/without DR, respectively) and 108 controls using quantitative real-time PCR.

Results

One month after aflibercept injection, significant reductions in CMT and VA were observed in DR cohorts. The four lncRNAs were over-expressed in DM compared with those in controls. However, downregulated baseline plasma levels of RNCR2 and NEAT2 were observed in glycemic-controlled DR patients. None of the lncRNAs showed a correlation with the severity of retinopathy or drug response.

Conclusion

Though circulating levels of the analyzed lncRNAs did not show an association with DR progression or aflibercept therapy response, the expression pattern demonstrated good diagnostic performance in differentiating DM from controls and DR.

Keywords

Anti-VEGF Biomarker Diabetic retinopathy Gene expression Long non-coding RNAs RNCR2 NEAT2 CDKN2B-AS1 PVT1 qRT-PCR 

Notes

Acknowledgments

The authors thank the Center of Excellence in Molecular and Cellular Medicine and the Oncology Diagnostic Unit, Suez Canal University, Ismailia, Egypt, for providing the facilities for performing the molecular work of the current study. The authors also thank all the participants for their approval to join this study.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki Declaration and its later amendments or comparable ethical standards.

Informed consent

Informed consent was obtained from all individual participants included in the study.

Supplementary material

417_2019_4409_MOESM1_ESM.docx (1.5 mb)
ESM 1 (DOCX 1516 kb)
417_2019_4409_MOESM2_ESM.docx (33 kb)
ESM 2 (DOCX 33 kb)

References

  1. 1.
    Antonetti DA, Klein R, Gardner TW (2012) Diabetic retinopathy. N Engl J Med 366:1227–1239.  https://doi.org/10.1056/NEJMra1005073 CrossRefGoogle Scholar
  2. 2.
    Duh EJ, Sun JK, Stitt AW (2017) Diabetic retinopathy: current understanding, mechanisms, and treatment strategies. JCI Insight 2:e93751.  https://doi.org/10.1172/jci.insight.93751 CrossRefGoogle Scholar
  3. 3.
    Stitt AW, urtis TM, Chen M, Medina RJ, McKay GJ, Jenkins A, Gardiner TA, Lyons TJ, Hammes HP, Simó R, Lois N (2016) The progress in understanding and treatment of diabetic retinopathy. Prog Retin Eye Res 51:156–186.  https://doi.org/10.1016/j.preteyeres.2015.08.001 CrossRefGoogle Scholar
  4. 4.
    Zhang X, Zeng H, Bao S, Wang N, Gillies MC (2014) Diabetic macular edema: new concepts in patho-physiology and treatment. Cell Biosci 4:27.  https://doi.org/10.1186/2045-3701-4-27 CrossRefGoogle Scholar
  5. 5.
    Tremolada G, Del Turco C, Lattanzio R, Maestroni S, Maestroni A, Bandello F, Zerbini G (2012) The role of angiogenesis in the development of proliferative diabetic retinopathy: impact of intravitreal anti-VEGF treatment. Exp Diabetes Res 2012:728325.  https://doi.org/10.1155/2012/728325 CrossRefGoogle Scholar
  6. 6.
    Bahrami B, Hong T, Gilles MC, Chang A (2017) Anti-VEGF therapy for diabetic eye diseases. Asia Pac J Ophthalmol (Phila) 6:535–545.  https://doi.org/10.22608/APO.2017350 Google Scholar
  7. 7.
    Cai S, Yang Q, Li X, Zhang Y (2018) The efficacy and safety of aflibercept and conbercept in diabetic macular edema. Drug Design, Development and Therapy 12:3471–3483.  https://doi.org/10.2147/DDDT.S177192 CrossRefGoogle Scholar
  8. 8.
    Cabral T, Mello LGM, Lima LH, Polido J, Regatieri CV, Belfort R Jr, Mahajan VB (2017) Retinal and choroidal angiogenesis: a review of new targets. Int J Retina Vitreous 3(31).  https://doi.org/10.1186/s40942-017-0084-9
  9. 9.
    Campos Polo R, Rubio Sánchez C, García Guisado DM, Díaz Luque MJ (2018) Aflibercept for clinically significant diabetic macular edema: 12-month results in daily clinical practice. Clin Ophthalmol 12:99–104.  https://doi.org/10.2147/OPTH.S154421 CrossRefGoogle Scholar
  10. 10.
    Gong Q, Su G (2017) Roles of miRNAs and long noncoding RNAs in the progression of diabetic retinopathy. Biosci Rep 37. pii: BSR20171157.  https://doi.org/10.1042/BSR20171157
  11. 11.
    Kashi K, Henderson L, Bonetti A, Carninci P (2016) Discovery and functional analysis of lncRNAs: methodologies to investigate an uncharacterized transcriptome. Biochim Biophys Acta 1859:3–15.  https://doi.org/10.1016/j.bbagrm.2015.10.010 CrossRefGoogle Scholar
  12. 12.
    Sathishkumar C, Prabu P, Mohan V, Balasubramanyam M (2018) Linking a role of lncRNAs (long non-coding RNAs) with insulin resistance, accelerated senescence, and inflammation in patients with type 2 diabetes. Hum Genomics 12(41).  https://doi.org/10.1186/s40246-018-0173-3
  13. 13.
    Raut SK, Khullar M (2018) The big entity of new RNA world: long non-coding RNAs in microvascular complications of diabetes. Front Endocrinol (Lausanne) 9(300).  https://doi.org/10.3389/fendo.2018.00300
  14. 14.
    Fawzy MS, Abu AlSel BT, Al Ageeli E, Al-Qahtani SA, Abdel-Daim MM, Toraih EA (2018) Long non-coding RNA MALAT1 and microRNA-499a expression profiles in diabetic ESRD patients undergoing dialysis: a preliminary cross-sectional analysis. Arch Physiol Biochem 29:1–11.  https://doi.org/10.1080/13813455.2018.1499119 CrossRefGoogle Scholar
  15. 15.
    Yan B, Tao ZF, Li XM, Zhang H, Yao J, Jiang Q (2014) Aberrant expression of long noncoding RNAs in early diabetic retinopathy. Invest Ophthalmol Vis Sci 55:941–951.  https://doi.org/10.1167/iovs.13-13221 CrossRefGoogle Scholar
  16. 16.
    Bhat SA, Ahmad SM, Mumtaz PT, Malik AA, Dar MA, Urwat U, Shah RA, Ganai NA (2016) Long non-coding RNAs: mechanism of action and functional utility. Noncoding RNA Res 1:43–50.  https://doi.org/10.1016/j.ncrna.2016.11.002 CrossRefGoogle Scholar
  17. 17.
    Wang KC, Chang HY (2011) Molecular mechanisms of long noncoding RNAs. Mol Cell 43:904–914.  https://doi.org/10.1016/j.molcel.2011.08.018 CrossRefGoogle Scholar
  18. 18.
    Wapinski O, Chang HY (2011) Long noncoding RNAs and human disease. Trends Cell Biol 21:354–361.  https://doi.org/10.1016/j.tcb.2011.04.001 CrossRefGoogle Scholar
  19. 19.
    Yan BA, Yao J, Liu JY, Li XM, Wang XQ, Li YJ, Tao ZF, Song YC, Chen Q, Jiang Q (2015) lncRNA-MIAT regulates microvascular dysfunction by functioning as a competing endogenous RNA. Circ Res 116:1143–1156.  https://doi.org/10.1161/CIRCRESAHA.116.305510 CrossRefGoogle Scholar
  20. 20.
    Yu B, Wang S (2018) Angio-LncRs: LncRNAs that regulate angiogenesis and vascular disease. Theranostics 8:3654–3675.  https://doi.org/10.7150/thno.26024 CrossRefGoogle Scholar
  21. 21.
    Michalik KM, You X, Manavski Y, Doddaballapur A, Zörnig M, Braun T, John D, Ponomareva Y, Chen W, Uchida S, Boon RA, Dimmeler S (2014) Long noncoding RNA MALAT1 regulates endothelial cell function and vessel growth. Circ Res 114:1389–1397.  https://doi.org/10.1161/CIRCRESAHA.114.303265 CrossRefGoogle Scholar
  22. 22.
    Congrains A, Kamide K, Ohishi M, Rakugi H (2013) ANRIL: Molecular mechanisms and implications in human health. Int J Mol Sci 14:1278–1292.  https://doi.org/10.3390/ijms14011278
  23. 23.
    Alvarez ML, Khosroheidari M, Eddy E, Kiefer J (2013) Role of microRNA 1207-5P and its host gene, the long non-coding RNA Pvt1, as mediators of extracellular matrix accumulation in the kidney: implications for diabetic nephropathy. PLoS One 8:e77468.  https://doi.org/10.1371/journal.pone.0077468 CrossRefGoogle Scholar
  24. 24.
    Majeed A, El-Sayed AA, Khoja T, Alshamsan R, Millett C, Rawaf S (2014) Diabetes in the Middle-East and North Africa: an update. Diabetes Res Clin Pract 103:218–222.  https://doi.org/10.1016/j.diabres.2013.11.008 CrossRefGoogle Scholar
  25. 25.
    No Authors (1991) Early treatment diabetic retinopathy study design and baseline patient characteristics ETDRS report number 7. Ophthalmology 98:741–756CrossRefGoogle Scholar
  26. 26.
    Fawzy MS, Toraih EA, Abdallah HY (2017) Long noncoding RNA metastasis-associated lung adenocarcinoma transcript 1 (MALAT1): a molecular predictor of poor survival in glioblastoma multiforme in Egyptian patients. Egyptian Journal of Medical Human Genetics 18:231–239.  https://doi.org/10.1016/j.ejmhg.2016.08.003 CrossRefGoogle Scholar
  27. 27.
    Fakhr-Eldeen A, Toraih EA, Fawzy MS (2019) Long non-coding RNAs MALAT1, MIAT and ANRIL gene expression profiles in beta-thalassemia patients: a cross-sectional analysis. Hematology 24:308–317.  https://doi.org/10.1080/16078454.2019.1570616 CrossRefGoogle Scholar
  28. 28.
    Bustin SA, Benes V, Garson JA, Hellemans J, Huggett J, Kubista M, Mueller R, Nolan T, Pfaffl MW, Shipley GL, Vandesompele J, Wittwer CT (2009) The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiments. Clin Chem 55:611-622.  https://doi.org/10.1373/clinchem.2008.112797
  29. 29.
    Bustin SA, Benes V, Garson J, Hellemans J, Huggett J, Kubista M et al (2013) The need for transparency and good practices in the qPCR literature. Nat Methods 10:1063–1067.  https://doi.org/10.1038/nmeth.2697 CrossRefGoogle Scholar
  30. 30.
    Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2-△△CT method. Methods 25:402–408.  https://doi.org/10.1006/meth.2001.1262 CrossRefGoogle Scholar
  31. 31.
    Leasher JL, Bourne RRA, Flaxman SR, Jonas JB, Keeffe J, Naidoo N, Pesudovs K, Price H, White RA, Wong TY, Resnikoff S, Taylor HR (2010) Vision Loss Expert Group of the Global Burden of Disease Study (2016) global estimates on the number of people blind or visually impaired by diabetic retinopathy: a meta-analysis from 1990 to. Diabetes Care 39:1643–1649.  https://doi.org/10.2337/dc15-2171 CrossRefGoogle Scholar
  32. 32.
    Jaé N, Dimmeler S (2015) Long noncoding RNAs in diabetic retinopathy. Circ Res 116:1104–1106.  https://doi.org/10.1161/CIRCRESAHA.115.306051 CrossRefGoogle Scholar
  33. 33.
    He X, Ou C, Xiao Y, Han Q, Li H, Zhou S (2017) LncRNAs: key players and novel insights into diabetes mellitus. Oncotarget 8:71325–71341.  https://doi.org/10.18632/oncotarget.19921 Google Scholar
  34. 34.
    Goyal N, Kesharwani D, Datta M (2018) Lnc-ing non-coding RNAs with metabolism and diabetes: roles of lncRNAs. Cell Mol Life Sci 75:1827–1837.  https://doi.org/10.1007/s00018-018-2760-9 CrossRefGoogle Scholar
  35. 35.
    Leti F, DiStefano JK (2017) Long noncoding RNAs as diagnostic and therapeutic targets in type 2 diabetes and related complications. Genes (Basel) 8(8):E207.  https://doi.org/10.3390/genes8080207 CrossRefGoogle Scholar
  36. 36.
    Wu Z, Liu X, Liu L, Deng H, Zhang J, Xu Q, Cen B, Ji A (2014) Regulation of lncRNA expression. Cell Mol Biol Lett 19:561–575.  https://doi.org/10.2478/s11658-014-0212-6 CrossRefGoogle Scholar
  37. 37.
    Thomas AA, Feng B, Chakrabarti S (2017) ANRIL: a regulator of VEGF in diabetic retinopathy. Invest Ophthalmol Vis Sci 58:470–480.  https://doi.org/10.1167/iovs.16-20569 CrossRefGoogle Scholar
  38. 38.
    Zhang J, Chen M, Chen J, Lin S, Cai D, Chen C, Chen Z (2017) Long non-coding RNA MIAT acts as a biomarker in diabetic retinopathy by absorbing miR-29b and regulating cell apoptosis. Biosci Rep 37:BSR20170036.  https://doi.org/10.1042/BSR20170036 CrossRefGoogle Scholar
  39. 39.
    Toraih EA, El-Wazir A, Alghamdi SA, Alhazmi AS, El-Wazir M, Abdel-Daim MM, Fawzy MS (2019) Association of long non-coding RNA MIAT and MALAT1 expression profiles in peripheral blood of coronary artery disease patients with previous cardiac events. Genet Mol Biol.  https://doi.org/10.1590/1678-4685-GMB-2018-0185
  40. 40.
    Li Q, Pang L, Yang W, Liu X, Su G, Dong Y (2018) Long non-coding RNA of myocardial infarction associated transcript (LncRNA-MIAT) promotes diabetic retinopathy by upregulating transforming growth factor-β1 (TGF-β1) signaling. Med Sci Monit 24:9497–9503.  https://doi.org/10.12659/MSM.911787 CrossRefGoogle Scholar
  41. 41.
    Liu JY, Yao J, Li XM, Song YC, Wang XQ, Li YJ, Yan B, Jiang Q (2014) Pathogenic role of lncRNA-MALAT1 in endothelial cell dysfunction in diabetes mellitus. Cell Death Dis 5:e1506.  https://doi.org/10.1038/cddis.2014.466 CrossRefGoogle Scholar
  42. 42.
    Puthanveetil P, Chen S, Feng B, Gautam A, Chakrabarti S (2015) Long non-coding RNA MALAT1 regulates hyperglycemia induced inflammatory process in the endothelial cells. J Cell Mol Med 19:1418–1425.  https://doi.org/10.1111/jcmm.12576 CrossRefGoogle Scholar
  43. 43.
    Li X, Zeng L, Cao C, Lu C, Lian W, Han J, Zhang X, Zhang J, Tang T, Li M (2017) Long noncoding RNA MALAT1 regulates renal tubular epithelial pyroptosis by modulated miR-23c targeting of ELAVL1 in diabetic nephropathy. Exp Cell Res 350:327–335.  https://doi.org/10.1016/j.yexcr.2016.12.006 CrossRefGoogle Scholar
  44. 44.
    Zhang Y, Wu H, Wang F, Ye M, Zhu H, Bu S (2018) Long non-coding RNA MALAT1 expression in patients with gestational diabetes mellitus. Int J Gynecol Obstet 140:164–169.  https://doi.org/10.1002/ijgo.12384 CrossRefGoogle Scholar
  45. 45.
    Dhawan S, Georgia S, Tschen SI, Fan G, Bhushan A (2011) Pancreatic beta cell identity is maintained by DNA methylation-mediated repression of Arx. Dev Cell 20:419–429.  https://doi.org/10.1016/j.devcel.2011.03.012 CrossRefGoogle Scholar
  46. 46.
    Yan C, Chen J, Chen N (2016) Long noncoding RNA MALAT1 promotes hepatic steatosis and insulin resistance by increasing nuclear SREBP-1c protein stability. Sci Rep 6:22640.  https://doi.org/10.1038/srep22640 CrossRefGoogle Scholar
  47. 47.
    de Gonzalo-Calvo D, Kenneweg F, Bang C, Toro R, van der Meer RW, Rijzewijk LJ, Smit JW, Lamb HJ, Llorente-Cortes V, Thum T (2016) Circulating long-non coding RNAs as biomarkers of left ventricular diastolic function and remodelling in patients with well-controlled type 2 diabetes. Sci Rep 6:37354.  https://doi.org/10.1038/srep37354 CrossRefGoogle Scholar
  48. 48.
    Hu M, Wang R, Li X, Fan M, Lin J, Zhen J, Chen L, Zhimei L (2017) LncRNA MALAT1 is dysregulated in diabetic nephropathy and involved in high glucose-induced podocyte injury via its interplay with β-catenin. J Cell Mol Med 21:2732–2747.  https://doi.org/10.1111/jcmm.13189 CrossRefGoogle Scholar
  49. 49.
    Biswas S, Thomas AA, Chen S, Aref-Eshghi E, Feng B, Gonder J, Sadikovic B, Chakrabarti S (2018) MALAT1: an epigenetic regulator of inflammation in diabetic retinopathy. Sci Rep 8(6526).  https://doi.org/10.1038/s41598-018-24907-w
  50. 50.
    Tsai F-J, Yang C-F, Chen C-C (2010) A genome-wide association study identifies susceptibility variants for type 2 diabetes in Han Chinese. PLoS Genet 6:e1000847.  https://doi.org/10.1371/journal.pgen.1000847 CrossRefGoogle Scholar
  51. 51.
    Kommoju U, Samy S, Maruda J, Irgam K, Kotla JP, Velaga L, Reddy BM (2016) Association of CDKAL1, CDKN2A/B & HHEX gene polymorphisms with type diabetes mellitus in the population of Hyderabad. India Indian J Med Res 143:455–463.  https://doi.org/10.4103/0971-5916.184303 CrossRefGoogle Scholar
  52. 52.
    Alvarez ML, DiStefano JK (2011) Functional characterization of the plasmacytoma variant translocation 1 gene (PVT1) in diabetic nephropathy. PLoS One 6:e18671.  https://doi.org/10.1371/journal.pone.0018671 CrossRefGoogle Scholar
  53. 53.
    Nittala MG, Keane PA, Zhang K, Sadda SR (2014) Risk factors for proliferative diabetic retinopathy in a Latino American population. Retina 34:1594–1599.  https://doi.org/10.1097/IAE.0000000000000117 CrossRefGoogle Scholar
  54. 54.
    Gupta A, Delhiwala KS, Raman RP, Sharma T, Srinivasan S, Kulothungan V (2016) Failure to initiate early insulin therapy - a risk factor for diabetic retinopathy in insulin users with type 2 diabetes mellitus: Sankara Nethralaya-diabetic retinopathy epidemiology and molecular genetics study (SN-DREAMS, report number 35). Indian J Ophthalmol 64:440–445.  https://doi.org/10.4103/0301-4738.187668 CrossRefGoogle Scholar
  55. 55.
    Do DV, Nguyen QD, Boyer D, Schmidt-Erfurth U, Brown DM, Vitti R, Berliner AJ, Gao B, Zeitz O, Ruckert R, Schmelter T, Sandbrink R, Heier JS (2012) One-year outcomes of the da Vinci Study of VEGF Trap-Eye in eyes with diabetic macular edema. Ophthalmology 119:1658–1665.  https://doi.org/10.1016/j.ophtha.2012.02.010 CrossRefGoogle Scholar
  56. 56.
    Nguyen CL, Lindsay A, Wong E, Chilov M (2018) Aflibercept for diabetic macular oedema: a meta-analysis of randomized controlled trials. Int J Ophthalmol 11:1002–1008Google Scholar
  57. 57.
    Bahrami B, Hong T, Schlub TE, Chang AA (2019) Aflibercept for persistent diabetic macular edema: forty-eight-week outcomes. Retina 39:61–68.  https://doi.org/10.1097/IAE.0000000000002253 CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Genetics Unit, Department of Histology and Cell Biology, Faculty of MedicineSuez Canal UniversityIsmailiaEgypt
  2. 2.Center of Excellence of Molecular and Cellular MedicineSuez Canal UniversityIsmailiaEgypt
  3. 3.Department of Ophthalmology, Faculty of MedicineSuez Canal UniversityIsmailiaEgypt
  4. 4.Department of Medical Biochemistry and Molecular Biology, Faculty of MedicineSuez Canal UniversityIsmailiaEgypt
  5. 5.Department of Clinical Biochemistry (Medical Genetics), Faculty of MedicineJazan UniversityJazanSaudi Arabia
  6. 6.Department of Biochemistry, Faculty of MedicineNorthern Border UniversityArarSaudi Arabia

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