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  • Original Article - Enabling Technologies
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Original Article – Enabling Technologies

Noninvasive tracking of gene transcript and neuroprotection after gene therapy

Gene Therapy volume 23, pages 1–9 (2016)Cite this article

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Abstract

Gene therapy holds exceptional potential for translational medicine by improving the products of defective genes in diseases and/or providing necessary biologics from endogenous sources during recovery processes. However, validating methods for the delivery, distribution and expression of the exogenous genes from such therapy can generally not be applicable to monitor effects over the long term because they are invasive. We report here that human granulocyte colony-stimulating factor (hG-CSF) complimentary DNA (cDNA) encoded in self-complementary adeno-associated virus-type 2 adeno-associated virus, as delivered through eye drops at multiple time points after cerebral ischemia using bilateral carotid occlusion for 60 min (BCAO-60) led to significant reduction in mortality rates, cerebral atrophy and neurological deficits in C57black6 mice. Most importantly, we validated hG-CSF cDNA expression using translatable magnetic resonance imaging (MRI) in living brains. This noninvasive approach for monitoring exogenous gene expression in the brains has potential for great impact in the area of experimental gene therapy in animal models of heart attack, stroke, Alzheimer’s dementia, Parkinson’s disorder and amyotrophic lateral sclerosis, and the translation of such techniques to emergency medicine.

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References

  1. Talelli P, Ellul J . Are patients with cognitive impairment after stroke at increased risk for developing Alzheimer disease? Arch Neurol 2004; 61: 983 author reply 983.

    Article  CAS  PubMed  Google Scholar 

  2. Wen Y, Yu WH, Maloney B, Bailey J, Ma J, Marié I et al. Transcriptional regulation of beta-secretase by p25/cdk5 leads to enhanced amyloidogenic processing. Neuron 2008; 57: 680–690.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  3. Barber RC, Edwards MI, Xiao G, Huebinger RM, Diaz-Arrastia R, Wilhelmsen KC et al. Serum granulocyte colony-stimulating factor and Alzheimer's disease. Dement Geriatr Cogn Dis Extra 2012; 2: 353–360.

    Article  PubMed Central  PubMed  Google Scholar 

  4. Diederich K, Sevimli S, Dörr H, Kösters E, Hoppen M, Lewejohann L et al. The role of granulocyte-colony stimulating factor (G-CSF) in the healthy brain: a characterization of G-CSF-deficient mice. J Neurosci 2009; 29: 11572–11581.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  5. Demetri GD, Griffin JD . Granulocyte colony-stimulating factor and its receptor. Blood 1991; 78: 2791–2808.

    CAS  PubMed  Google Scholar 

  6. Schneider A, Krüger C, Steigleder T, Weber D, Pitzer C, Laage R et al. The hematopoietic factor G-CSF is a neuronal ligand that counteracts programmed cell death and drives neurogenesis. J Clin Invest 2005; 115: 2083–2098.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  7. Song S, Sava V, Rowe A, Li K, Cao C, Mori T et al. Granulocyte-colony stimulating factor (G-CSF) enhances recovery in mouse model of Parkinson's disease. Neurosci Lett 2011; 487: 153–157.

    Article  CAS  PubMed  Google Scholar 

  8. Cao XQ, Arai H, Ren YR, Oizumi H, Zhang N, Seike S et al. Recombinant human granulocyte colony-stimulating factor protects against MPTP-induced dopaminergic cell death in mice by altering Bcl-2/Bax expression levels. J Neurochem 2006; 99: 861–867.

    Article  CAS  PubMed  Google Scholar 

  9. Capoccia BJ, Shepherd RM, Link DC . G-CSF and AMD3100 mobilize monocytes into the blood that stimulate angiogenesis in vivo through a paracrine mechanism. Blood 2006; 108: 2438–2445.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  10. Gibson CL, Bath PM, Murphy SP . G-CSF reduces infarct volume and improves functional outcome after transient focal cerebral ischemia in mice. J Cereb Blood Flow Metab 2005; 25: 431–439.

    Article  CAS  PubMed  Google Scholar 

  11. England TJ, Abaei M, Auer DP, Lowe J, Jones DR, Sare G et al. Granulocyte-colony stimulating factor for mobilizing bone marrow stem cells in subacute stroke: the stem cell trial of recovery enhancement after stroke 2 randomized controlled trial. Stroke 2012; 43: 405–411.

    Article  CAS  PubMed  Google Scholar 

  12. Ringelstein EB, Thijs V, Norrving B, Chamorro A, Aichner F, Grond M et al. Granulocyte colony-stimulating factor in patients with acute ischemic stroke: results of the AX200 for Ischemic Stroke trial. Stroke 2013; 44: 2681–2687.

    Article  CAS  PubMed  Google Scholar 

  13. Davis AS, Zhao H, Sun GH, Sapolsky RM, Steinberg GK . Gene therapy using SOD1 protects striatal neurons from experimental stroke. Neurosci Lett 2007; 411: 32–36.

    Article  CAS  PubMed  Google Scholar 

  14. Liu CH, You Z, Ren J, Kim YR, Eikermann-Haerter K, Liu PK et al. Noninvasive delivery of gene targeting probes to live brains for transcription MRI. FASEB J 2008; 22: 1193–1203.

    Article  CAS  PubMed  Google Scholar 

  15. Liu CH, Ren JQ, You Z, Yang J, Liu CM, Uppal R et al. Noninvasive detection of neural progenitor cells in living brains by MRI. FASEB J 2012; 26: 1652–1662.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  16. Henriques A, Pitzer C, Dittgen T, Klugmann M, Dupuis L, Schneider A et al. CNS-targeted viral delivery of G-CSF in an animal model for ALS: improved efficacy and preservation of the neuromuscular unit. Mol Ther 2011; 19: 284–292.

    Article  CAS  PubMed  Google Scholar 

  17. Liu PK, Grossman RG, Hsu CY, Robertson CS . Ischemic injury and faulty gene transcripts in the brain. Trends Neurosci 2001; 24: 581–588.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  18. Veldwijk MR, Sellner L, Stiefelhagen M, Kleinschmidt JA, Laufs S, Topaly J et al. Pseudotyped recombinant adeno-associated viral vectors mediate efficient gene transfer into primary human CD34(±) peripheral blood progenitor cells. Cytotherapy 2010; 12: 107–112.

    Article  CAS  PubMed  Google Scholar 

  19. Duan D, Yue Y, Yan Z, Engelhardt JF . A new dual-vector approach to enhance recombinant adeno-associated virus-mediated gene expression through intermolecular cis activation. Nat Med 2000; 6: 595–598.

    Article  CAS  PubMed  Google Scholar 

  20. Mandel RJ . CERE-110, an adeno-associated virus-based gene delivery vector expressing human nerve growth factor for the treatment of Alzheimer's disease. Curr Opin Mol Ther 2010; 12: 240–247.

    CAS  PubMed  Google Scholar 

  21. Theodore S, Cao S, McLean PJ, Standaert DG . Targeted overexpression of human alpha-synuclein triggers microglial activation and an adaptive immune response in a mouse model of Parkinson disease. J Neuropathol Exp Neurol 2008; 67: 1149–1158.

    Article  CAS  PubMed  Google Scholar 

  22. Whalen MJ, Dalkara T, You Z, Qiu J, Bermpohl D, Mehta N et al. Acute plasmalemma permeability and protracted clearance of injured cells after controlled cortical impact in mice. J Cereb Blood Flow Metab 2008; 28: 490–505.

    Article  CAS  PubMed  Google Scholar 

  23. Bermpohl D, You Z, Lo EH, Kim HH, Whalen MJ . TNF alpha and Fas mediate tissue damage and functional outcome after traumatic brain injury in mice. J Cereb Blood Flow Metab 2007; 27: 1806–1818.

    Article  CAS  PubMed  Google Scholar 

  24. Kim F, Bravo PE, Nichol G . What is the use of hypothermia for neuroprotection after out-of-hospital cardiac arrest? Stroke 2015; 46: 592–597.

    Article  PubMed Central  PubMed  Google Scholar 

  25. Huang D, Shenoy A, Cui J, Huang W, Liu PK . In situ detection of AP sites and DNA strand breaks bearing 3'-phosphate termini in ischemic mouse brain. FASEB J 2000; 14: 407–417.

    Article  CAS  PubMed  Google Scholar 

  26. Liu CH, Huang S, Kim YR, Rosen BR, Liu PK . Forebrain ischemia-reperfusion simulating cardiac arrest in mice induces edema and DNA fragmentation in the brain. Mol Imaging 2007; 6: 156–170.

    Article  PubMed  Google Scholar 

  27. Kida K, Minamishima S, Wang H, Ren J, Yigitkanli K, Nozari A et al. Sodium sulfide prevents water diffusion abnormality in the brain and improves long term outcome after cardiac arrest in mice. Resuscitation 2012; 83: 1292–1297.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  28. Minamishima S, Kida K, Tokuda K, Wang H, Sips PY, Kosugi S et al. Inhaled nitric oxide improves outcomes after successful cardiopulmonary resuscitation in mice. Circulation 2011; 124: 1645–1653.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  29. Dijkhuizen RM, Ren J, Mandeville JB, Wu O, Ozdag FM, Moskowitz MA et al. Functional magnetic resonance imaging of reorganization in rat brain after stroke. Proc Natl Acad Sci USA 2001; 98: 12766–12771.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  30. Li X, Blizzard KK, Zeng Z, DeVries AC, Hurn PD, McCullough LD et al. Chronic behavioral testing after focal ischemia in the mouse: functional recovery and the effects of gender. Exp Neurol 2004; 187: 94–104.

    Article  PubMed  Google Scholar 

  31. Yli-Hankala A, Edmonds HL Jr, Jiang YD, Higham HE, Zhang PY . Outcome effects of different protective hypothermia levels during cardiac arrest in rats. Acta Anaesthesiol Scand 1997; 41: 511–515.

    Article  CAS  PubMed  Google Scholar 

  32. Liu CH, You Z, Liu CM, Kim YR, Whalen MJ, Rosen BR et al. Diffusion-weighted magnetic resonance imaging reversal by gene knockdown of matrix metalloproteinase-9 activities in live animal brains. J Neurosci 2009; 29: 3508–3517.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  33. Pardridge WM, Boado RJ, Kang YS . Vector-mediated delivery of a polyamide ("peptide") nucleic acid analogue through the blood-brain barrier in vivo. Proc Natl Acad Sci USA 1995; 92: 5592–5596.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  34. Liu PK, Salminen A, He YY, Jiang MH, Xue JJ, Liu JS et al. Suppression of ischemia-induced fos expression and AP-1 activity by an antisense oligodeoxynucleotide to c-fos mRNA. Ann Neurol 1994; 36: 566–576.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  35. Gibson CL, Jones NC, Prior MJ, Bath PM, Murphy SP . G-CSF suppresses edema formation and reduces interleukin-1beta expression after cerebral ischemia in mice. J Neuropathol Exp Neurol 2005; 64: 763–769.

    Article  CAS  PubMed  Google Scholar 

  36. Liu CH, Ren J, Liu CM, Liu PK . Intracellular gene transcription factor protein-guided MRI by DNA aptamers in vivo. FASEB J 2014; 28: 464–473.

    Article  PubMed Central  PubMed  Google Scholar 

  37. Schaar KL, Brenneman MM, Savitz SI . Functional assessments in the rodent stroke model. Exp Transl Stroke Med 2010; 2: 13.

    Article  PubMed Central  PubMed  Google Scholar 

  38. Zhang L, Schallert T, Zhang ZG, Jiang Q, Arniego P, Li Q et al. A test for detecting long-term sensorimotor dysfunction in the mouse after focal cerebral ischemia. J Neurosci Methods 2002; 117: 207–214.

    Article  PubMed  Google Scholar 

  39. Liu CH, Yang J, Ren JQ, Liu CM, You Z, Liu PK et al. MRI reveals differential effects of amphetamine exposure on neuroglia in vivo. FASEB J 2013; 27: 712–724.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  40. Liu CH, Ren JQ, Yang J, Liu CM, Mandeville JB, Rosen BR et al. DNA-based mRI probes for specific detection of chronic exposure to amphetamine in living brains. J Neurosci 2009; 29: 10663–10670.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  41. Liu CH, Huang S, Cui J, Kim YR, Farrar CT, Moskowitz MA et al. MR contrast probes that trace gene transcripts for cerebral ischemia in live animals. FASEB J 2007; 21: 3004–3015.

    Article  PubMed  Google Scholar 

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Acknowledgements

We thank Dr Sean I Savitz (University of Texas Houston Medical School, Houston, TX, USA) for demonstration of the corner tests; Dr Jarek Aronowski (UTHMS, TX, USA) for sharing the hypothermia treatment protocol; Dr Charng-Ming Liu for synthesizing all sODN and Dr JS Yang for conducting real time-PCR. This project was supported by NIH grants R01DA029889 and R01EB013768, as well as by a pilot grant from the Boston Area Diabetes Endocrinology Research Center (P30DK057521-14 (J Avruch)) to PKL, NS045776 to the MGH Neuroscience Center). The MR systems used for this work were funded by NIH Shared Instrumentation Grants (1S10RR025563) awarded to the Athinoula A Martinos Center for Biomedical Imaging. The B Wellcome Trust (No.1012722) to HP, State of Florida (09KW-11) to JYW also supported this research.

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Authors and Affiliations

  1. Department of Radiology, Athinoula A. Martinos Center for Biomedical Imaging, Massachusetts General Hospital and the Harvard Medical School, Charlestown, MA, USA

    J Ren, Y I Chen, C H Liu & P K Liu

  2. Department of Biomedical Science, Charles E. Schmidt College of Medicine, Florida Atlantic University, Boca Raton, FL, USA

    P-C Chen, H Prentice & J-Y Wu

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  1. J Ren
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Correspondence to P K Liu.

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Ren, J., Chen, Y., Liu, C. et al. Noninvasive tracking of gene transcript and neuroprotection after gene therapy. Gene Ther 23, 1–9 (2016). https://doi.org/10.1038/gt.2015.81

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