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Novel In Situ Activity Assays for the Quantitative Molecular Analysis of Neurodegenerative Processes in the Retina

Ekström, Per LU ; Ueffing, M.; Zrenner, E. and Paquet-Durand, F. (2014) In Current Medicinal Chemistry 21(30). p.3478-3493
Abstract
The mechanisms of neuronal cell death are still only poorly understood, which has hindered the advancement of therapies for many currently untreatable neurodegenerative diseases. This calls for the development of new methods which reveal critical molecular mechanisms of the celldeath machinery with both high sensitivity and cellular resolution. Using animal models for hereditary neurodegeneration in the retina, we have developed or adapted different biochemical assays to determine the enzymatic activities of calpain, poly-ADP-ribose-polymerase (PARP), and histone deacetylase (HDAC) directly and in situ. Additionally, the enzymatic activity of cGMP-dependent protein kinase (PKG) was assessed indirectly using in situ immunohistological... (More)
The mechanisms of neuronal cell death are still only poorly understood, which has hindered the advancement of therapies for many currently untreatable neurodegenerative diseases. This calls for the development of new methods which reveal critical molecular mechanisms of the celldeath machinery with both high sensitivity and cellular resolution. Using animal models for hereditary neurodegeneration in the retina, we have developed or adapted different biochemical assays to determine the enzymatic activities of calpain, poly-ADP-ribose-polymerase (PARP), and histone deacetylase (HDAC) directly and in situ. Additionally, the enzymatic activity of cGMP-dependent protein kinase (PKG) was assessed indirectly using in situ immunohistological techniques to detect PKG-activity-dependent products. Combining these assays with in situ cell death markers revealed close temporospatial correlations, suggesting causal connections between the PKG, HDAC, PARP and calpain activities and neuronal cell death. Using different pharmacological and genetic manipulations, causality could indeed be demonstrated. Surprisingly, the often dramatic rises in metabolic activities didnot match by corresponding increases in expression, highlighting the importance of analyses of protein activities at the cellular level. The above mentioned studies identified a number of metabolic processes previously unknownto be involved in inherited retinal degeneration. Comparing different animal retinal degeneration models uncovered striking similarities in enzymatic activities, suggesting a generality of the destructive pathways. Taken together, these findings provided a number of novel targets for neuroprotection and as such opened up new perspectives for the therapy of hereditary neurodegeneration in the retina and possibly other parts of the central nervous system. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
calpain, cell death, cGMP, HDAC, PARP, TUNEL
in
Current Medicinal Chemistry
volume
21
issue
30
pages
3478 - 3493
publisher
Bentham Science Publishers
external identifiers
  • wos:000341969100005
  • scopus:84926359381
ISSN
0929-8673
DOI
10.2174/0929867321666140601201337
language
English
LU publication?
yes
id
9be517aa-16f8-4412-9a13-304c7b20a032 (old id 4709794)
date added to LUP
2014-11-03 07:18:40
date last changed
2017-01-25 12:57:42
@article{9be517aa-16f8-4412-9a13-304c7b20a032,
  abstract     = {The mechanisms of neuronal cell death are still only poorly understood, which has hindered the advancement of therapies for many currently untreatable neurodegenerative diseases. This calls for the development of new methods which reveal critical molecular mechanisms of the celldeath machinery with both high sensitivity and cellular resolution. Using animal models for hereditary neurodegeneration in the retina, we have developed or adapted different biochemical assays to determine the enzymatic activities of calpain, poly-ADP-ribose-polymerase (PARP), and histone deacetylase (HDAC) directly and in situ. Additionally, the enzymatic activity of cGMP-dependent protein kinase (PKG) was assessed indirectly using in situ immunohistological techniques to detect PKG-activity-dependent products. Combining these assays with in situ cell death markers revealed close temporospatial correlations, suggesting causal connections between the PKG, HDAC, PARP and calpain activities and neuronal cell death. Using different pharmacological and genetic manipulations, causality could indeed be demonstrated. Surprisingly, the often dramatic rises in metabolic activities didnot match by corresponding increases in expression, highlighting the importance of analyses of protein activities at the cellular level. The above mentioned studies identified a number of metabolic processes previously unknownto be involved in inherited retinal degeneration. Comparing different animal retinal degeneration models uncovered striking similarities in enzymatic activities, suggesting a generality of the destructive pathways. Taken together, these findings provided a number of novel targets for neuroprotection and as such opened up new perspectives for the therapy of hereditary neurodegeneration in the retina and possibly other parts of the central nervous system.},
  author       = {Ekström, Per and Ueffing, M. and Zrenner, E. and Paquet-Durand, F.},
  issn         = {0929-8673},
  keyword      = {calpain,cell death,cGMP,HDAC,PARP,TUNEL},
  language     = {eng},
  number       = {30},
  pages        = {3478--3493},
  publisher    = {Bentham Science Publishers},
  series       = {Current Medicinal Chemistry},
  title        = {Novel In Situ Activity Assays for the Quantitative Molecular Analysis of Neurodegenerative Processes in the Retina},
  url          = {http://dx.doi.org/10.2174/0929867321666140601201337},
  volume       = {21},
  year         = {2014},
}