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In Vitro and in Vivo Effects of Alpha7 Nicotinic Acetylcholine Receptor Gene Delivery on Neuroprotective Pathways

Permanent Link: http://ufdc.ufl.edu/UFE0042864/00001

Material Information

Title: In Vitro and in Vivo Effects of Alpha7 Nicotinic Acetylcholine Receptor Gene Delivery on Neuroprotective Pathways
Physical Description: 1 online resource (102 p.)
Language: english
Creator: REN,YAN
Publisher: University of Florida
Place of Publication: Gainesville, Fla.
Publication Date: 2011

Subjects

Subjects / Keywords: AAV -- AKT -- ALPHA7 -- ALZHEIMER -- CASPASE -- ERK -- SEPTUM -- TRKA
Genetics and Genomics -- Dissertations, Academic -- UF
Genre: Genetics and Genomics thesis, Ph.D.
bibliography   ( marcgt )
theses   ( marcgt )
government publication (state, provincial, terriorial, dependent)   ( marcgt )
born-digital   ( sobekcm )
Electronic Thesis or Dissertation

Notes

Abstract: Basal forebrain cholinergic neurons are essential for normal memory-related behaviors but are dysfunctional at early stage of Alzheimer's disease. Brain alpha7 nicotinic acetylcholine receptors abundantly expressed on these neurons. While not essential for their survival, they nonetheless regulate neurotransmitter release, modulate neuroplasticity and protect them against a wide variety of insults involving trophic factor deprivation, apoptosis, and phenotypic silencing. Many data have indicated that alpha7 agonist can either prevent neuron death or restore their cholinergic phenotype, and improve memory related performance. However few studies have attempted to restore the loss of these receptors through gene delivery approaches. In this study, we investigated the effects of alpha7 receptor gene delivery on pathways associated with cell survival. Rat PC12 cells transfected using an optimized gene delivery system to overexpress alpha7 receptor showed more ERK2 and AKT activity and less caspase 3/7 activity in a dose dependent manner after treatment with the alpha7 partial agonist, 4OH-GTS-21. These effects could be blocked by the alpha7 receptor selective antagonist MLA, indicating that elevated alpha7 receptor expression improves the efficacy and potency of agonist treatment for PC12 cells. Elevated alpha7 expression also increased TrkA phosphorylation as well as enhanced NGF induced neuron-like differentiation in the same cells, suggesting its involvement in regulating NGF-TrkA signaling pathway. AAV8-alpha7 gene delivery into mouse septum promoted TrkA expression in this brain region and improved performance in Morris water maze tasks. Since the activation of TrkA receptors, which are also diminished in AD patient brains, positively modulates neuron survival and cholinergic functions, these results suggest potential dual protection for cholinergic neurons after alpha7 gene delivery. Together, alpha7 overexpression potently activated survival signaling pathways when combined with selective agonist and enhanced neurotrophin related functions. alpha7 gene therapy, which can restore the function of these receptors and TrkA receptors, may provide a new approach for treating AD patients.
General Note: In the series University of Florida Digital Collections.
General Note: Includes vita.
Bibliography: Includes bibliographical references.
Source of Description: Description based on online resource; title from PDF title page.
Source of Description: This bibliographic record is available under the Creative Commons CC0 public domain dedication. The University of Florida Libraries, as creator of this bibliographic record, has waived all rights to it worldwide under copyright law, including all related and neighboring rights, to the extent allowed by law.
Statement of Responsibility: by YAN REN.
Thesis: Thesis (Ph.D.)--University of Florida, 2011.
Local: Adviser: Song, Sihong.
Electronic Access: RESTRICTED TO UF STUDENTS, STAFF, FACULTY, AND ON-CAMPUS USE UNTIL 2013-04-30

Record Information

Source Institution: UFRGP
Rights Management: Applicable rights reserved.
Classification: lcc - LD1780 2011
System ID: UFE0042864:00001

Permanent Link: http://ufdc.ufl.edu/UFE0042864/00001

Material Information

Title: In Vitro and in Vivo Effects of Alpha7 Nicotinic Acetylcholine Receptor Gene Delivery on Neuroprotective Pathways
Physical Description: 1 online resource (102 p.)
Language: english
Creator: REN,YAN
Publisher: University of Florida
Place of Publication: Gainesville, Fla.
Publication Date: 2011

Subjects

Subjects / Keywords: AAV -- AKT -- ALPHA7 -- ALZHEIMER -- CASPASE -- ERK -- SEPTUM -- TRKA
Genetics and Genomics -- Dissertations, Academic -- UF
Genre: Genetics and Genomics thesis, Ph.D.
bibliography   ( marcgt )
theses   ( marcgt )
government publication (state, provincial, terriorial, dependent)   ( marcgt )
born-digital   ( sobekcm )
Electronic Thesis or Dissertation

Notes

Abstract: Basal forebrain cholinergic neurons are essential for normal memory-related behaviors but are dysfunctional at early stage of Alzheimer's disease. Brain alpha7 nicotinic acetylcholine receptors abundantly expressed on these neurons. While not essential for their survival, they nonetheless regulate neurotransmitter release, modulate neuroplasticity and protect them against a wide variety of insults involving trophic factor deprivation, apoptosis, and phenotypic silencing. Many data have indicated that alpha7 agonist can either prevent neuron death or restore their cholinergic phenotype, and improve memory related performance. However few studies have attempted to restore the loss of these receptors through gene delivery approaches. In this study, we investigated the effects of alpha7 receptor gene delivery on pathways associated with cell survival. Rat PC12 cells transfected using an optimized gene delivery system to overexpress alpha7 receptor showed more ERK2 and AKT activity and less caspase 3/7 activity in a dose dependent manner after treatment with the alpha7 partial agonist, 4OH-GTS-21. These effects could be blocked by the alpha7 receptor selective antagonist MLA, indicating that elevated alpha7 receptor expression improves the efficacy and potency of agonist treatment for PC12 cells. Elevated alpha7 expression also increased TrkA phosphorylation as well as enhanced NGF induced neuron-like differentiation in the same cells, suggesting its involvement in regulating NGF-TrkA signaling pathway. AAV8-alpha7 gene delivery into mouse septum promoted TrkA expression in this brain region and improved performance in Morris water maze tasks. Since the activation of TrkA receptors, which are also diminished in AD patient brains, positively modulates neuron survival and cholinergic functions, these results suggest potential dual protection for cholinergic neurons after alpha7 gene delivery. Together, alpha7 overexpression potently activated survival signaling pathways when combined with selective agonist and enhanced neurotrophin related functions. alpha7 gene therapy, which can restore the function of these receptors and TrkA receptors, may provide a new approach for treating AD patients.
General Note: In the series University of Florida Digital Collections.
General Note: Includes vita.
Bibliography: Includes bibliographical references.
Source of Description: Description based on online resource; title from PDF title page.
Source of Description: This bibliographic record is available under the Creative Commons CC0 public domain dedication. The University of Florida Libraries, as creator of this bibliographic record, has waived all rights to it worldwide under copyright law, including all related and neighboring rights, to the extent allowed by law.
Statement of Responsibility: by YAN REN.
Thesis: Thesis (Ph.D.)--University of Florida, 2011.
Local: Adviser: Song, Sihong.
Electronic Access: RESTRICTED TO UF STUDENTS, STAFF, FACULTY, AND ON-CAMPUS USE UNTIL 2013-04-30

Record Information

Source Institution: UFRGP
Rights Management: Applicable rights reserved.
Classification: lcc - LD1780 2011
System ID: UFE0042864:00001


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in Vitro in Vivo

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In Vitro in Vivo

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Alzheimer Disease

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Cholinergic Hypothesis et al et al et al et al et al

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et al Neuronal Nicotinic Acetylcholine Receptor

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et al et al et al

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Identificatio n et al et al et al., et al et al et al et al Localization et al et al et al et al et al et al

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et al et al et al et al et al Function et al et al et al 7 nAChRs Mediated Neuroprotection Effects on in V itro and in V ivo Models In vitro et al

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et al et al et al et al In vivo et al et al et al et al et al et al 7 nAChRs and Cellular Signaling Pathway s et al et al et al et al

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et al et al et al

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et al et al et al et al et al et al et al et al et al et al et al et al et al

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et al et al et al in vitro in vivo et al et al et al et al et al In vitro et al et al et al

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Therapeutic Appro Agonist Therapy et al in vitro et al et al et al et al et al

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et al et al et al et al et al Ge ne Therapy

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et al et al. et al. In vivo

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et al et al et al. et al. et al. et al Specific Aims and Significance

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in vitro

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Reagents Plasmid Preparations

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The Cell Culture

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Cell Gene Delivery, Differentiation and Drug Treatments Cell Viability

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E nzyme Linked Immunosorbent Assay

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Western Blot

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Caspase Assay Immunofluorescence

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Virus Vector Packaging and Titration Transfection Purification

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Dot -B lot Assay

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Stereotaxic Surgery

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Morris Water Maze Tests

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Immunohistochemistry

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Statistical Analysis

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Introduction et al et al et al et al et al et al

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et al et al et al et al et al et al et al et al

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et al et al et al et al et al et al et al et al et al et al et al

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Results in vitro

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Effects of 4OH GTS 21 on pERK2 a nd pAkt in T ransfected C ells: et al et al et al

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et al Effects of 4OH GTS 21 on Caspase A ctivity: et al

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Discussion

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et al et al et al et al et al et al et al et al et al et al et al et

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al et al et al et al et al et al et al et al et al

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et al et al et al et al et al et al et al et al

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Introduction et al et al et al et al et al

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et al et al et al et al et al et al et al

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et al et al et al et al et al

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et al et al et al et al et al et al in vitro in vivo et al et al et al et al

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Results In V itro et al et al et al

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a b

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Effects of AAV8 in V ivo in vitro in vivo in vivo et al

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Effects of Septal AAV8 e Delivery on Memory Related Behavior

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Discussion

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in vitro

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in vivo et al et al

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in vitro in vivo

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Conclusion

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Future Directions

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