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Exendin-4 ameliorates motor neuron degeneration in cellular and animal models of amyotrophic lateral sclerosis.

Li Y, Chigurupati S, Holloway HW, Mughal M, Tweedie D, Bruestle DA, Mattson MP, Wang Y, Harvey BK, Ray B, Lahiri DK, Greig NH - PLoS ONE (2012)

Bottom Line: Ex-4 proved to be neurotrophic in NSC-19 cells, elevating choline acetyltransferase (ChAT) activity, as well as neuroprotective, protecting cells from hydrogen peroxide-induced oxidative stress and staurosporine-induced apoptosis.Furthermore, Ex-4 treatment attenuated neuronal cell death in the lumbar spinal cord; immunohistochemical analysis demonstrated the rescue of neuronal markers, such as ChAT, associated with motor neurons.Together, our results suggest that GLP-1 receptor agonists warrant further evaluation to assess whether their neuroprotective potential is of therapeutic relevance in ALS.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Neurosciences, Intramural Research Program, National Institute on Aging, National Institutes of Health, Baltimore, Maryland, United States of America.

ABSTRACT
Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease characterized by a progressive loss of lower motor neurons in the spinal cord. The incretin hormone, glucagon-like peptide-1 (GLP-1), facilitates insulin signaling, and the long acting GLP-1 receptor agonist exendin-4 (Ex-4) is currently used as an anti-diabetic drug. GLP-1 receptors are widely expressed in the brain and spinal cord, and our prior studies have shown that Ex-4 is neuroprotective in several neurodegenerative disease rodent models, including stroke, Parkinson's disease and Alzheimer's disease. Here we hypothesized that Ex-4 may provide neuroprotective activity in ALS, and hence characterized Ex-4 actions in both cell culture (NSC-19 neuroblastoma cells) and in vivo (SOD1 G93A mutant mice) models of ALS. Ex-4 proved to be neurotrophic in NSC-19 cells, elevating choline acetyltransferase (ChAT) activity, as well as neuroprotective, protecting cells from hydrogen peroxide-induced oxidative stress and staurosporine-induced apoptosis. Additionally, in both wild-type SOD1 and mutant SOD1 (G37R) stably transfected NSC-19 cell lines, Ex-4 protected against trophic factor withdrawal-induced toxicity. To assess in vivo translation, SOD1 mutant mice were administered vehicle or Ex-4 at 6-weeks of age onwards to end-stage disease via subcutaneous osmotic pump to provide steady-state infusion. ALS mice treated with Ex-4 showed improved glucose tolerance and normalization of behavior, as assessed by running wheel, compared to control ALS mice. Furthermore, Ex-4 treatment attenuated neuronal cell death in the lumbar spinal cord; immunohistochemical analysis demonstrated the rescue of neuronal markers, such as ChAT, associated with motor neurons. Together, our results suggest that GLP-1 receptor agonists warrant further evaluation to assess whether their neuroprotective potential is of therapeutic relevance in ALS.

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Ex-4 treatment preserved lumbar spinal cord structure and neuron density.(A) Cresyl violet staining of lumbar spinal cord section of SOD1 (G93A) mice at both early (9 weeks of treatment at a pre-symptomatic stage) and end stages of disease. (B) Close-up of ventral horn of grey matter shows staining of motor neurons with large soma, scale bars on right inset is 25 µm. By comparison, (C) demonstrates similar lumbar spinal cord sections derived from control non-ALS WT mice, in which the grey matter tracts are clearly evident. (D) Quantification of motor neuron numbers in the ventral horn of grey matter. An approximately 3-fold greater neuron number was present in SOD1 (G93A) Ex-4-treated mice spinal cord sections than in vehicle-treated mice at both pre-symptomatic and end disease stages (9 and 12 week treatment times). Evaluation of motor neuron number in the lumbar ventral horn grey matter of age-matched control non-ALS WT mice were in the range of those present in SOD1 (G93A) mice treated with Ex-4. Selection of large pyramidal neurons was based on their characteristic nulceolus, angular and spindle-shaped [85]. A measuring bar of 25 µm is for reference only.
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pone-0032008-g006: Ex-4 treatment preserved lumbar spinal cord structure and neuron density.(A) Cresyl violet staining of lumbar spinal cord section of SOD1 (G93A) mice at both early (9 weeks of treatment at a pre-symptomatic stage) and end stages of disease. (B) Close-up of ventral horn of grey matter shows staining of motor neurons with large soma, scale bars on right inset is 25 µm. By comparison, (C) demonstrates similar lumbar spinal cord sections derived from control non-ALS WT mice, in which the grey matter tracts are clearly evident. (D) Quantification of motor neuron numbers in the ventral horn of grey matter. An approximately 3-fold greater neuron number was present in SOD1 (G93A) Ex-4-treated mice spinal cord sections than in vehicle-treated mice at both pre-symptomatic and end disease stages (9 and 12 week treatment times). Evaluation of motor neuron number in the lumbar ventral horn grey matter of age-matched control non-ALS WT mice were in the range of those present in SOD1 (G93A) mice treated with Ex-4. Selection of large pyramidal neurons was based on their characteristic nulceolus, angular and spindle-shaped [85]. A measuring bar of 25 µm is for reference only.

Mentions: SOD1 (G93A) mice were euthanized at both 15-weeks (early symptomatic disease stage, n = 5 per group) and 18 weeks of age (end stage, n = 10 per group), and age-matched control WT non-ALS mice (n = 4) were likewise euthanized for analysis of spinal cord. Representative micrographs of cresyl violet-stained lumbar spinal cord are shown in Figure 6A, highlighting the Nissl substance in the neurons. Readily apparent are structural alterations in spinal cord morphology from SOD1 (G93A) vehicle treated animals, particularly in relation to the boundary between white and grey matter regions (Figure 6B), as compared to control wild-type mice (Figure 6C). In particular, the demarcation of the ventral and dorsal grey matter horns is unclear, and suggestive of motor neuron dysfunction or loss within these areas. In contrast, in Ex-4 treated SOD1 (G93A) mice spinal cord, the morphology of the lumbar structure appeared more typical, with both ventral and dorsal horns clearly visible, suggesting preservation of spinal cord structure and neurons (Figure 6A). To more clearly characterize changes, quantification of presumptive motor neuron cell bodies within the ventral horn was undertaken (Figure 6B) to determine neuron density. Figure 6D illustrates the presence of 3-fold more neurons within the spinal cord section of Ex-4 treated SOD1 (G93A) mice compared to vehicle SOD-1 (G93A) mice at both early and end stage disease. This result suggests that Ex-4 treatment preserved lumbar spinal cord neurons from degeneration in ALS mice, particularly since neuron density was reduced but not substantially different from that in control non-ALS mice.


Exendin-4 ameliorates motor neuron degeneration in cellular and animal models of amyotrophic lateral sclerosis.

Li Y, Chigurupati S, Holloway HW, Mughal M, Tweedie D, Bruestle DA, Mattson MP, Wang Y, Harvey BK, Ray B, Lahiri DK, Greig NH - PLoS ONE (2012)

Ex-4 treatment preserved lumbar spinal cord structure and neuron density.(A) Cresyl violet staining of lumbar spinal cord section of SOD1 (G93A) mice at both early (9 weeks of treatment at a pre-symptomatic stage) and end stages of disease. (B) Close-up of ventral horn of grey matter shows staining of motor neurons with large soma, scale bars on right inset is 25 µm. By comparison, (C) demonstrates similar lumbar spinal cord sections derived from control non-ALS WT mice, in which the grey matter tracts are clearly evident. (D) Quantification of motor neuron numbers in the ventral horn of grey matter. An approximately 3-fold greater neuron number was present in SOD1 (G93A) Ex-4-treated mice spinal cord sections than in vehicle-treated mice at both pre-symptomatic and end disease stages (9 and 12 week treatment times). Evaluation of motor neuron number in the lumbar ventral horn grey matter of age-matched control non-ALS WT mice were in the range of those present in SOD1 (G93A) mice treated with Ex-4. Selection of large pyramidal neurons was based on their characteristic nulceolus, angular and spindle-shaped [85]. A measuring bar of 25 µm is for reference only.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3285661&req=5

pone-0032008-g006: Ex-4 treatment preserved lumbar spinal cord structure and neuron density.(A) Cresyl violet staining of lumbar spinal cord section of SOD1 (G93A) mice at both early (9 weeks of treatment at a pre-symptomatic stage) and end stages of disease. (B) Close-up of ventral horn of grey matter shows staining of motor neurons with large soma, scale bars on right inset is 25 µm. By comparison, (C) demonstrates similar lumbar spinal cord sections derived from control non-ALS WT mice, in which the grey matter tracts are clearly evident. (D) Quantification of motor neuron numbers in the ventral horn of grey matter. An approximately 3-fold greater neuron number was present in SOD1 (G93A) Ex-4-treated mice spinal cord sections than in vehicle-treated mice at both pre-symptomatic and end disease stages (9 and 12 week treatment times). Evaluation of motor neuron number in the lumbar ventral horn grey matter of age-matched control non-ALS WT mice were in the range of those present in SOD1 (G93A) mice treated with Ex-4. Selection of large pyramidal neurons was based on their characteristic nulceolus, angular and spindle-shaped [85]. A measuring bar of 25 µm is for reference only.
Mentions: SOD1 (G93A) mice were euthanized at both 15-weeks (early symptomatic disease stage, n = 5 per group) and 18 weeks of age (end stage, n = 10 per group), and age-matched control WT non-ALS mice (n = 4) were likewise euthanized for analysis of spinal cord. Representative micrographs of cresyl violet-stained lumbar spinal cord are shown in Figure 6A, highlighting the Nissl substance in the neurons. Readily apparent are structural alterations in spinal cord morphology from SOD1 (G93A) vehicle treated animals, particularly in relation to the boundary between white and grey matter regions (Figure 6B), as compared to control wild-type mice (Figure 6C). In particular, the demarcation of the ventral and dorsal grey matter horns is unclear, and suggestive of motor neuron dysfunction or loss within these areas. In contrast, in Ex-4 treated SOD1 (G93A) mice spinal cord, the morphology of the lumbar structure appeared more typical, with both ventral and dorsal horns clearly visible, suggesting preservation of spinal cord structure and neurons (Figure 6A). To more clearly characterize changes, quantification of presumptive motor neuron cell bodies within the ventral horn was undertaken (Figure 6B) to determine neuron density. Figure 6D illustrates the presence of 3-fold more neurons within the spinal cord section of Ex-4 treated SOD1 (G93A) mice compared to vehicle SOD-1 (G93A) mice at both early and end stage disease. This result suggests that Ex-4 treatment preserved lumbar spinal cord neurons from degeneration in ALS mice, particularly since neuron density was reduced but not substantially different from that in control non-ALS mice.

Bottom Line: Ex-4 proved to be neurotrophic in NSC-19 cells, elevating choline acetyltransferase (ChAT) activity, as well as neuroprotective, protecting cells from hydrogen peroxide-induced oxidative stress and staurosporine-induced apoptosis.Furthermore, Ex-4 treatment attenuated neuronal cell death in the lumbar spinal cord; immunohistochemical analysis demonstrated the rescue of neuronal markers, such as ChAT, associated with motor neurons.Together, our results suggest that GLP-1 receptor agonists warrant further evaluation to assess whether their neuroprotective potential is of therapeutic relevance in ALS.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Neurosciences, Intramural Research Program, National Institute on Aging, National Institutes of Health, Baltimore, Maryland, United States of America.

ABSTRACT
Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease characterized by a progressive loss of lower motor neurons in the spinal cord. The incretin hormone, glucagon-like peptide-1 (GLP-1), facilitates insulin signaling, and the long acting GLP-1 receptor agonist exendin-4 (Ex-4) is currently used as an anti-diabetic drug. GLP-1 receptors are widely expressed in the brain and spinal cord, and our prior studies have shown that Ex-4 is neuroprotective in several neurodegenerative disease rodent models, including stroke, Parkinson's disease and Alzheimer's disease. Here we hypothesized that Ex-4 may provide neuroprotective activity in ALS, and hence characterized Ex-4 actions in both cell culture (NSC-19 neuroblastoma cells) and in vivo (SOD1 G93A mutant mice) models of ALS. Ex-4 proved to be neurotrophic in NSC-19 cells, elevating choline acetyltransferase (ChAT) activity, as well as neuroprotective, protecting cells from hydrogen peroxide-induced oxidative stress and staurosporine-induced apoptosis. Additionally, in both wild-type SOD1 and mutant SOD1 (G37R) stably transfected NSC-19 cell lines, Ex-4 protected against trophic factor withdrawal-induced toxicity. To assess in vivo translation, SOD1 mutant mice were administered vehicle or Ex-4 at 6-weeks of age onwards to end-stage disease via subcutaneous osmotic pump to provide steady-state infusion. ALS mice treated with Ex-4 showed improved glucose tolerance and normalization of behavior, as assessed by running wheel, compared to control ALS mice. Furthermore, Ex-4 treatment attenuated neuronal cell death in the lumbar spinal cord; immunohistochemical analysis demonstrated the rescue of neuronal markers, such as ChAT, associated with motor neurons. Together, our results suggest that GLP-1 receptor agonists warrant further evaluation to assess whether their neuroprotective potential is of therapeutic relevance in ALS.

Show MeSH
Related in: MedlinePlus