Ischemic injury-specific gene expression in the rat spinal cord injury model using hypoxia-inducible system

Minhyung Lee, Su Lee Eun, Soo Kim Young, Hyune Choi Byung, Ra Park So, Seon Park Hyun, Chun Park Hyung, Wan Kim Sung, Yoon Ha

Research output: Contribution to journalArticle

39 Citations (Scopus)

Abstract

Study Design. A spinal cord injury and in vitro neural hypoxia models were used to evaluate the hypoxia responsive gene expression. Objectives. To limit the risk of unwanted overexpression of therapeutic genes, we developed a hypoxia-inducible gene therapy system using the erythropoietin (Epo) enhancer and the RTP801 promoter. Summary of Background Data. Gene therapy is an emerging therapeutic technique to treat spinal cord injury. However, uncontrolled overexpression of therapeutic genes in nondisease tissues during gene therapy raises a doubt about its safety. Post-traumatic ischemia is an important factor worsening the spinal cord damage, and hypoxia could regulate the gene expressions using a hypoxia-inducible promoter. Methods. The plasmids, pEpo-SV-Luc and pRTP801-Luc, were constructed. Mouse neuroblastoma cells (N2A) were used to evaluate the hypoxia-inducible gene expression in vitro. Gene transfection and expression were allowed for 24 hours under normoxia (pO 2, 152 mm Hg) or hypoxia (pO2, 7.6 mm Hg). Spinal cord injury was made using clip compression. Plasmids were injected directly into the injured spinal cord immediately following injury. The gene expression was assessed by luciferase assay. Results. pEpo-SV-Luc and pRTP801-Luc showed more than three times higher gene expression in N2A cells under hypoxia than normoxia. The expression level of luciferase in the injured spinal cord was higher than in the normal spinal cord. Immunostaining demonstrated that neurons, astrocytes, and capillary endothelial cells expressed luciferase in the cytoplasm. Conclusions. The pEpo-SV-Luc and pRTP801-Luc systems are effective in that they induce gene expression specifically in neurons under the hypoxic condition and spinal cord injury.

Original languageEnglish
Pages (from-to)2729-2734
Number of pages6
JournalSpine
Volume30
Issue number24
DOIs
Publication statusPublished - 2005 Dec 1

Fingerprint

Spinal Cord Injuries
Gene Expression
Wounds and Injuries
Spinal Cord
Luciferases
Genetic Therapy
Plasmids
Neurons
Cell Hypoxia
Erythropoietin
Hypoxia
Neuroblastoma
Surgical Instruments
Astrocytes
Genes
Transfection
Cytoplasm
Therapeutics
Ischemia
Endothelial Cells

All Science Journal Classification (ASJC) codes

  • Orthopedics and Sports Medicine
  • Clinical Neurology

Cite this

Lee, Minhyung ; Eun, Su Lee ; Young, Soo Kim ; Byung, Hyune Choi ; So, Ra Park ; Hyun, Seon Park ; Hyung, Chun Park ; Sung, Wan Kim ; Ha, Yoon. / Ischemic injury-specific gene expression in the rat spinal cord injury model using hypoxia-inducible system. In: Spine. 2005 ; Vol. 30, No. 24. pp. 2729-2734.
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abstract = "Study Design. A spinal cord injury and in vitro neural hypoxia models were used to evaluate the hypoxia responsive gene expression. Objectives. To limit the risk of unwanted overexpression of therapeutic genes, we developed a hypoxia-inducible gene therapy system using the erythropoietin (Epo) enhancer and the RTP801 promoter. Summary of Background Data. Gene therapy is an emerging therapeutic technique to treat spinal cord injury. However, uncontrolled overexpression of therapeutic genes in nondisease tissues during gene therapy raises a doubt about its safety. Post-traumatic ischemia is an important factor worsening the spinal cord damage, and hypoxia could regulate the gene expressions using a hypoxia-inducible promoter. Methods. The plasmids, pEpo-SV-Luc and pRTP801-Luc, were constructed. Mouse neuroblastoma cells (N2A) were used to evaluate the hypoxia-inducible gene expression in vitro. Gene transfection and expression were allowed for 24 hours under normoxia (pO 2, 152 mm Hg) or hypoxia (pO2, 7.6 mm Hg). Spinal cord injury was made using clip compression. Plasmids were injected directly into the injured spinal cord immediately following injury. The gene expression was assessed by luciferase assay. Results. pEpo-SV-Luc and pRTP801-Luc showed more than three times higher gene expression in N2A cells under hypoxia than normoxia. The expression level of luciferase in the injured spinal cord was higher than in the normal spinal cord. Immunostaining demonstrated that neurons, astrocytes, and capillary endothelial cells expressed luciferase in the cytoplasm. Conclusions. The pEpo-SV-Luc and pRTP801-Luc systems are effective in that they induce gene expression specifically in neurons under the hypoxic condition and spinal cord injury.",
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Lee, M, Eun, SL, Young, SK, Byung, HC, So, RP, Hyun, SP, Hyung, CP, Sung, WK & Ha, Y 2005, 'Ischemic injury-specific gene expression in the rat spinal cord injury model using hypoxia-inducible system', Spine, vol. 30, no. 24, pp. 2729-2734. https://doi.org/10.1097/01.brs.0000190395.43772.f3

Ischemic injury-specific gene expression in the rat spinal cord injury model using hypoxia-inducible system. / Lee, Minhyung; Eun, Su Lee; Young, Soo Kim; Byung, Hyune Choi; So, Ra Park; Hyun, Seon Park; Hyung, Chun Park; Sung, Wan Kim; Ha, Yoon.

In: Spine, Vol. 30, No. 24, 01.12.2005, p. 2729-2734.

Research output: Contribution to journalArticle

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T1 - Ischemic injury-specific gene expression in the rat spinal cord injury model using hypoxia-inducible system

AU - Lee, Minhyung

AU - Eun, Su Lee

AU - Young, Soo Kim

AU - Byung, Hyune Choi

AU - So, Ra Park

AU - Hyun, Seon Park

AU - Hyung, Chun Park

AU - Sung, Wan Kim

AU - Ha, Yoon

PY - 2005/12/1

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N2 - Study Design. A spinal cord injury and in vitro neural hypoxia models were used to evaluate the hypoxia responsive gene expression. Objectives. To limit the risk of unwanted overexpression of therapeutic genes, we developed a hypoxia-inducible gene therapy system using the erythropoietin (Epo) enhancer and the RTP801 promoter. Summary of Background Data. Gene therapy is an emerging therapeutic technique to treat spinal cord injury. However, uncontrolled overexpression of therapeutic genes in nondisease tissues during gene therapy raises a doubt about its safety. Post-traumatic ischemia is an important factor worsening the spinal cord damage, and hypoxia could regulate the gene expressions using a hypoxia-inducible promoter. Methods. The plasmids, pEpo-SV-Luc and pRTP801-Luc, were constructed. Mouse neuroblastoma cells (N2A) were used to evaluate the hypoxia-inducible gene expression in vitro. Gene transfection and expression were allowed for 24 hours under normoxia (pO 2, 152 mm Hg) or hypoxia (pO2, 7.6 mm Hg). Spinal cord injury was made using clip compression. Plasmids were injected directly into the injured spinal cord immediately following injury. The gene expression was assessed by luciferase assay. Results. pEpo-SV-Luc and pRTP801-Luc showed more than three times higher gene expression in N2A cells under hypoxia than normoxia. The expression level of luciferase in the injured spinal cord was higher than in the normal spinal cord. Immunostaining demonstrated that neurons, astrocytes, and capillary endothelial cells expressed luciferase in the cytoplasm. Conclusions. The pEpo-SV-Luc and pRTP801-Luc systems are effective in that they induce gene expression specifically in neurons under the hypoxic condition and spinal cord injury.

AB - Study Design. A spinal cord injury and in vitro neural hypoxia models were used to evaluate the hypoxia responsive gene expression. Objectives. To limit the risk of unwanted overexpression of therapeutic genes, we developed a hypoxia-inducible gene therapy system using the erythropoietin (Epo) enhancer and the RTP801 promoter. Summary of Background Data. Gene therapy is an emerging therapeutic technique to treat spinal cord injury. However, uncontrolled overexpression of therapeutic genes in nondisease tissues during gene therapy raises a doubt about its safety. Post-traumatic ischemia is an important factor worsening the spinal cord damage, and hypoxia could regulate the gene expressions using a hypoxia-inducible promoter. Methods. The plasmids, pEpo-SV-Luc and pRTP801-Luc, were constructed. Mouse neuroblastoma cells (N2A) were used to evaluate the hypoxia-inducible gene expression in vitro. Gene transfection and expression were allowed for 24 hours under normoxia (pO 2, 152 mm Hg) or hypoxia (pO2, 7.6 mm Hg). Spinal cord injury was made using clip compression. Plasmids were injected directly into the injured spinal cord immediately following injury. The gene expression was assessed by luciferase assay. Results. pEpo-SV-Luc and pRTP801-Luc showed more than three times higher gene expression in N2A cells under hypoxia than normoxia. The expression level of luciferase in the injured spinal cord was higher than in the normal spinal cord. Immunostaining demonstrated that neurons, astrocytes, and capillary endothelial cells expressed luciferase in the cytoplasm. Conclusions. The pEpo-SV-Luc and pRTP801-Luc systems are effective in that they induce gene expression specifically in neurons under the hypoxic condition and spinal cord injury.

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