Original Article

Regulatory Role of Insulin on Endogenous L1 ORF1 and NEFM Gene Expression through PI3K Signaling Pathway Specifically in Neuroblastoma Cell Line


Background: One of the most important endogenous factors causing genomic instability in human cells is L1s retrotransposons. In this study, we assume that increased activity of L1 retrotransposons (specifically L1 expression) might be induced by hyperglycemia and hyperinsulinemia in neuroblastoma cell line.

Methods: Two different cell lines (BE (2)-M17 and HEK293) were treated with insulin and its PI3K signaling pathway inhibitor under three conditional media including hyperglycemic and retinoic acid treatment in the department of Medical Genetics, Tehran University of Medical Sciences, Tehran, Iran in 2018. The expression of L1 ORF1, as well as genes involved in insulin signaling pathway and neuronal stress and structure were measured at RNA level.

Results: Insulin could significantly down regulate the expression of L1 ORF1 and NEFM genes. Hyperglycemia result in severe decrease in expression of all candidate genes in control neuroblastoma but not HEK293 cells. Retinoic acid as the concentration used in this study cause increase stemness in neuroblastoma but not HEK293 cells. We could not find significant correlation between expression pattern of other genes tested in our study and L1 ORF1 expression.

Conclusion: Total regulatory effect of insulin on L1 ORF1 RNA expression as well as NEFM markedly in BE (2)-M17 cell line. Although these results could not be interpreted as L1 retrotransposition, expression of L1 RNA during stress conditions might be considered following inhibition of the insulin pathway. The result of this study also confirms the impotence of insulin on human evolution.


1. Posthuma D, Polderman TJ (2013). What have we learned from recent twin studies about the etiology of neurodevelopmental disorders? Curr Opin Neurol, 26(2): 111-21.
2. Carlsson T, Molander F, Taylor MJ, Jonsson U, Bolte S (2021). Early environmental risk factors for neurodevelopmental disorders–a systematic review of twin and sibling studies. Dev Psychopathol, 33(4):1448-1495.
3. Lander ES, Linton LM, Birren B, et al (2001). Initial sequencing and analysis of the human genome. Nature, 409(15): 860-921.
4. Protasova MS, Andreeva TV, Rogaev EI (2021). Factors Regulating the Activity of LINE1 Retrotransposons. Genes (Basel), 12(10): 1562.
5. Perna R, Loughan AR, Le J, Tyson K (2015). Gestational diabetes: long-term central nervous system developmental and cognitive sequelae. Appl Neuropsychol Child, 4(3): 217-20.
6. Kleinridders A, Ferris HA, Cai W, Kahn CR (2014). Insulin action in brain regulates systemic metabolism and brain function. Diabetes, 63(7): 2232-43.
7. Tchenio T, Casella J-F, Heidmann T (2000). Members of the SRY family regulate the human LINE retrotransposons. Nucleic Acids Res, 28(2): 411-5.
8. Swergold GD (1990). Identification, characterization, and cell specificity of a human LINE-1 promoter. Mol Cell Biol, 10(12): 6718-29.
9. Moran JV, Holmes SE, Naas TP, et al (1996). High frequency retrotransposition in cultured mammalian cells. Cell, 87(5): 917-27.
10. Goodier JL, Zhang L, Vetter MR, Kazazian HH (2007). LINE-1 ORF1 protein localizes in stress granules with other RNA-binding proteins, including components of RNA interference RNA-induced silencing complex. Mol Cell Biol, 27(18): 6469-83.
11. Habibi L, Shokrgozar MA, Tabrizi M, et al (2014). Mercury specifically induces LINE-1 activity in a human neuroblastoma cell line. Mutat Res Genet Toxicol Environ Mutagen ,759: 9-20.
12. Habibi L, Shokrgozar MA, Motamedi M, Akrami SM (2013). Effect of heavy metals on silencing of engineered long interspersed element-1 retrotransposon in nondividing neuroblastoma cell line. Iran Biomed J, 17(4): 171-78.
13. Karimi A, Madjd Z, Habibi L, Akrami SM (2014). Exposure of hepatocellular carcinoma cells to low-level As2O3‎ causes an extra toxicity pathway via L1 retrotransposition‎ induction. Toxicol Lett, 229(1): 111-7.
14. Xiang AH (2017). Association of maternal diabetes with autism in offspring. JAMA, 317(5): 537-8.
15. Del Re B, Marcantonio P, Capri M, Giorgi G (2010). Evaluation of LINE-1 mobility in neuroblastoma cells by in vitro retrotransposition reporter assay: FACS analysis can detect only the tip of the iceberg of the inserted L1 elements. Exp Cell Res, 316(20):3358-67.
16. Van Meerloo J, Kaspers GJ, Cloos J (2011). Cell sensitivity assays: the MTT assay. Methods Mol Biol, 731:237-45.
17. Kumar P, Nagarajan A, Uchil PD (2018). Analysis of cell viability by the MTT assay. Cold Spring Harb Protoc, 2018(6):10.1101/pdb.prot095505.
18. Muotri AR, Chu VT, Marchetto MC, et al (2005). Somatic mosaicism in neuronal precursor cells mediated by L1 retrotransposition. Nature, 435(7044): 903-10.
19. Sankaran VG, Xu J, Ragoczy T, et al (2009). Developmental and species-divergent globin switching are driven by BCL11A. Nature, 460(7259): 1093-7.
20. Baillie JK, Barnett MW, Upton KR, et al (2011). Somatic retrotransposition alters the genetic landscape of the human brain. Nature, 479(7374): 534-7.
21. Kong L, Norstedt G, Schalling M, et al (2018). The risk of offspring psychiatric disorders in the setting of maternal obesity and diabetes. Pediatrics, 142(3): e20180776.
22. Clausen TD, Mortensen EL, Schmidt L, et al (2013). Cognitive function in adult offspring of women with gestational diabetes–the role of glucose and other factors. PLoS One, 8(6): e67107.
23. DeBoer T, Wewerka S, Bauer PJ, et al (2005). Explicit memory performance in infants of diabetic mothers at 1 year of age. Dev Med Child Neurol, 47(8): 525-31.
24. Fraser A, Nelson SM, Macdonald-Wallis C, Lawlor DA (2012). Associations of existing diabetes, gestational diabetes, and glycosuria with offspring IQ and educational attainment: the Avon Longitudinal Study of Parents and Children. Exp Diabetes Res, 2012:963735.
25. Brasacchio D, Okabe J, Tikellis C, et al (2009). Hyperglycemia induces a dynamic cooperativity of histone methylase and demethylase enzymes associated with gene-activating epigenetic marks that coexist on the lysine tail. Diabetes, 58(5): 1229-36.
26. Yu J, Wu Y, Yang P (2016). High glucose‐induced oxidative stress represses sirtuin deacetylase expression and increases histone acetylation leading to neural tube defects. J Neurochem, 137(3): 371-83.
27. Tonge PD, Andrews PW (2010). Retinoic acid directs neuronal differentiation of human pluripotent stem cell lines in a non-cell-autonomous manner. Differentiation, 80(1): 20-30.
28. Coufal NG, Garcia-Perez JL, Peng GE, et al (2009). L1 retrotransposition in human neural progenitor cells. Nature, 460(7259): 1127-31.
29. Muotri AR, Zhao C, Marchetto MC, Gage FH (2009). Environmental influence on L1 retrotransposons in the adult hippocampus. Hippocampus, 19(10): 1002-7.
30. Cerchia L, d'Alessio A, Amabile G, et al (2006). An Autocrine Loop Involving Ret and Glial Cell–Derived Neurotrophic Factor Mediates Retinoic Acid–Induced Neuroblastoma Cell Differentiation. Mol Cancer Res, 4(7): 481-8.
31. Jeong H, Kim MS, Kim SW, Kim KS, Seol W (2006). Regulation of tyrosine hydroxylase gene expression by retinoic acid receptor. J Neurochem, 98(2): 386-94.
32. Lee JH, Kim KT (2004). Induction of cyclin‐dependent kinase 5 and its activator p35 through the extracellular‐signal‐regulated kinase and protein kinase A pathways during retinoic‐acid mediated neuronal differentiation in human neuroblastoma SK‐N‐BE (2) C cells. J Neurochem, 91(3): 634-47.
33. Matthay KK, Reynolds CP, Seeger RC, et al (2009). Long-term results for children with high-risk neuroblastoma treated on a randomized trial of myeloablative therapy followed by 13-cis-retinoic acid: a children's oncology group study. J Clin Oncol, 27(7): 1007-13.
34. Al‐Chalabi A, Miller CC (2003). Neurofilaments and neurological disease. Bioessays, 25(4):346-55.
35. Guo Z, Du X, Iacovitti L (1998). Regulation of tyrosine hydroxylase gene expression during transdifferentiation of striatal neurons: changes in transcription factors binding the AP-1 site. J Neurosci, 18(20): 8163-74.
IssueVol 52 No 3 (2023) QRcode
SectionOriginal Article(s)
DOI https://doi.org/10.18502/ijph.v52i3.12146
Retroelement Regulatory subunit 3 Neurofilament protein NF-M Neuroblastomas

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How to Cite
Sohrabi N, Habibi L, Akrami SM. Regulatory Role of Insulin on Endogenous L1 ORF1 and NEFM Gene Expression through PI3K Signaling Pathway Specifically in Neuroblastoma Cell Line. Iran J Public Health. 2023;52(3):633-645.