Research ReportUltrastructural diversity of inclusions and aggregations in the lumbar spinal cord of SOD1-G93A transgenic mice
Research highlights
► Ultrastructural diversity of inclusions and aggregations are examined. ► SOD1 accumulation precedes the appearance of inclusions. ► Astrocytic and axonal inclusions are predominant at end stage. ► Electron-dense astrocytic inclusions are reported.
Introduction
Amyotrophic lateral sclerosis (ALS) is a progressive, adult-onset neurodegenerative disease that mainly affects motor neurons in cortex, brain stem, and spinal cord. ALS occurs in both sporadic and familial forms, which are clinically and pathologically similar. About 90–95% cases of ALS were sporadic, and the remaining 5–10% of patients inherit the disease in an autosomal dominant manner. SOD1 mutations account for approximately 20% of familial cases (Rosen et al., 1993). Currently, more than 153 different SOD1 mutations have been linked to ALS through a gain of unknown toxicity (Felbecker et al., 2010). SOD1-G93A mice showed an ALS-like phenotype and were widely used in ALS pathogenic and therapeutic researches (Gurney et al., 1994). Several hypotheses have been proposed to explain motor neuron death in ALS, including glutamate-induced excitotoxicity, oxidative stress, cytoskeletal abnormality, and protein aggregation (VanDen-Bosch et al., 2006, Cleveland & Rothstein, 2001, Julien, 2001, Julien et al., 2005, Wijesekera & Leigh, 2009).
General pathological changes have been revealed in SOD1 mutant transgenic mice, including astrogliosis, fragmentation of Golgi apparatus, SOD1 aggregation, and vacuolar degeneration (Mourelatos et al., 1996, Bruijn et al., 1998, Dal-Canto & Gurney, 1995, Stieber et al., 2000). A hallmark of ALS is the aberrant accumulation of neurofilaments in the cell body and proximal axons of motor neurons. Although the extent to which neurofilament abnormalities contribute to the pathogenesis of ALS remains unknown, emerging evidence suggests that disorganized neurofilaments can provoke degeneration and death of neurons (Lee et al., 1994, Perrone-Capano et al., 2001). It was reported that accumulation of SOD1-positive aggregates in the neuronal processes, predominantly in the axons, probably causing impairment of axonal transport, which contribute to the pathogenesis of SOD1-G93A transgenic mice (Sasaki et al., 2005). Although ALS is characterized by motor neuron degeneration, the role of non-neuronal cells has recently been recognized (Pramatarova et al., 2001, Clement et al., 2003). All of the above studies indicate that the pathological changes of ALS involved more than one organelle, one cell type or even one system. We have recently reported that sensory axons in the lumbar dorsal root and posterior funiculus are also affected as early as motor axons do in the SOD1-G93A transgenic mice (Guo et al., 2009). However, the precise mechanisms leading to ALS were still unknown.
In the current work, we extended previous pathological observations and made an intensive examination of the ultrastructural pathological features of inclusions and abnormal aggregations in neuronal axons and glial cells, as well as neurons, in the lumbar spinal cord of SOD1-G93A transgenic mice at different stages of disease. We present new evidence that SOD1 accumulation in the cytoplasm of motor neurons precede the appearance of numerous inclusions in the axons and astrocytes. The ultrastructural pathological features of inclusions and aggregations differ according to their locations, and astrocytic and axonal inclusions were predominant changes at the end stage of disease, which may contribute to the degeneration of motor neurons or be the results of motor neuron degeneration.
Section snippets
General pathological changes of the lumbar anterior horn in SOD1-G93A transgenic mice
Using light microscope, similar to previous studies, we detected apparent motor neuron loss, astrocytosis and vacuolar degeneration in the anterior horn with the disease progressed in SOD1-G93A transgenic mice (data not shown). Besides, numerous eosinophilic inclusions were found in the lumbar anterior horn at both clinical onset and end stages of SOD1-G93A mice (Fig. 1). Although several vacuolated motor neurons were occasionally observed, eosinophilic inclusions were rarely seen at 60 days of
Discussion
Amyotrophic lateral sclerosis is a progressive neurodegenerative disease which involves mainly motor neurons. Pathological changes of human ALS depend on the autopsy samples that are mostly from the terminal stage. Analyses of ALS autopsy samples had shed much light on the pathogenesis of motor neuron loss. The most predominant feature of ALS is loss of upper and lower motor neurons, especially motor neurons in the spinal cord anterior horn. Other features, observed in the anterior horn cells
Animals
Transgenic mice over-expressing mutant SOD1-G93A and B6SJLF1 hybrids were purchased from Jackson Laboratory (Bar Harbor, ME). The animals were housed in a room with 12 h-dark/12 h-light cycle and provided with free access to water and diet (standard diet purchased from Beijing Vital River Experimental Animal Co. Ltd, Beijing, China). Transgenic SOD1-G93A mice and their non-transgenic littermates were generated by breeding male hemizygous carriers (B6SJL-Tg (SOD1-G93A) 1Gur/J) to female B6SJLF1
Acknowledgments
This work was supported in part by grants from National Natural Science Foundation of China and Hebei Science and Technology Department (No. 30870882, 30900460, and 08966105D).
References (34)
- et al.
Neuropathological changes in two lines of mice carrying a transgene for mutant human Cu, Zn SOD, and in mice overexpressing wild type human SOD: a model of familial amyotrophic lateral sclerosis (FALS)
Brain Res.
(1995) Amyotrophic lateral sclerosis. unfolding the toxicity of the misfolded
Cell
(2001)- et al.
A mutant neurofilament subunit causes massive, selective motor neuron death: implications for the pathogenesis of human motor neuron disease
Neuron
(1994) - et al.
Creatine supplementation and riluzole treatment provide similar beneficial effects in copper, zinc superoxide dismutase (G93A) transgenic mice
Neuroscience
(2003) - et al.
Aggregation of ubiquitin and a mutant ALS-linked SOD1 protein correlate with disease progression and fragmentation of the Golgi apparatus
J. Neurol. Sci.
(2000) - et al.
The role of excitotoxicity in the pathogenesis of amyotrophic lateral sclerosis
Biochim. Biophys. Acta
(2006) - et al.
Interaction between familial amyotrophic lateral sclerosis (ALS)-linked SOD1 mutants and the dynein complex
J. Biol. Chem.
(2007) - et al.
Aggregation and motor neuron toxicity of an ALS-linked SOD1 mutant independent from wild type SOD1
Science
(1998) - et al.
Wild-type nonneuronal cells extend survival of SOD1 mutant motor neurons in ALS mice
Science
(2003) - et al.
From Charcot to Lou Gehrig: deciphering selective motor neuron death in ALS
Nat. Rev. Neurosci.
(2001)
Defective axonal transport in a transgenic mouse model of amyotrophic lateral sclerosis
Nature
A low expressor line of transgenic mice carrying a mutant human Cu, Zn superoxide dismutase (SOD1) gene develops pathological changes that most closely resemble those in human amyotrophic lateral sclerosis
Acta Neuropathol.
Four familial ALS pedigrees discordant for two SOD1 mutations: are all SOD1 mutations pathogenic?
J. Neurol. Neurosurg. Psychiatry
Sensory Involvement in the SOD1-G93A mouse model of amyotrophic lateral sclerosis
Exp. Mol. Med.
Motor neuron degeneration in mice that express a human Cu, Zn superoxide dismutase mutation
Science
Familial amyotrophic lateral sclerosis. A subgroup characterized by posterior and spinocerebellar tract involvement and hyaline inclusions in the anterior horn cells
Arch Neurol.
A role for neurofilaments in the pathogenesis of amyotrophic lateral sclerosis
Biochem. Cell Biol.
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