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Formation of multinucleated giant cells and microglial degeneration in rats expressing a mutant Cu/Zn superoxide dismutase gene.

Fendrick SE, Xue QS, Streit WJ - J Neuroinflammation (2007)

Bottom Line: In animals during end stage disease at 4-5 months of age virtually all microglia in the spinal cord gray matter showed extensive fragmentation of their cytoplasm (cytorrhexis), indicative of widespread microglial degeneration.Few microglia exhibiting nuclear fragmentation (karyorrhexis) indicative of apoptosis were identified at any stage.The current findings demonstrate the occurrence of severe abnormalities in microglia, such as cell fusions and cytorrhexis, which may be the result of expression of mutant SOD1 in these cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Neuroscience, University of Florida College of Medicine and McKnight Brain Institute, Gainesville, FL 32611, USA. sefendrick@yahoo.com

ABSTRACT

Background: Microglial neuroinflammation is thought to play a role in the pathogenesis of amyotrophic lateral sclerosis (ALS). The purpose of this study was to provide a histopathological evaluation of the microglial neuroinflammatory response in a rodent model of ALS, the SOD1G93A transgenic rat.

Methods: Multiple levels of the CNS from spinal cord to cerebral cortex were studied in SOD1G93A transgenic rats during three stages of natural disease progression, including presymptomatic, early symptomatic (onset), and late symptomatic (end stage), using immuno- and lectin histochemical markers for microglia, such as OX-42, OX-6, and Griffonia simplicifolia isolectin B4.

Results: Our studies revealed abnormal aggregates of microglia forming in the spinal cord as early as the presymptomatic stage. During the symptomatic stages there was prominent formation of multinucleated giant cells through fusion of microglial cells in the spinal cord, brainstem, and red nucleus of the midbrain. Other brain regions, including substantia nigra, cranial nerve nuclei, hippocampus and cortex showed normal appearing microglia. In animals during end stage disease at 4-5 months of age virtually all microglia in the spinal cord gray matter showed extensive fragmentation of their cytoplasm (cytorrhexis), indicative of widespread microglial degeneration. Few microglia exhibiting nuclear fragmentation (karyorrhexis) indicative of apoptosis were identified at any stage.

Conclusion: The current findings demonstrate the occurrence of severe abnormalities in microglia, such as cell fusions and cytorrhexis, which may be the result of expression of mutant SOD1 in these cells. The microglial changes observed are different from those that accompany normal microglial activation, and they demonstrate that aberrant activation and degeneration of microglia is part of the pathogenesis of motor neuron disease.

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OX-42 immunohistochemistry during symptomatic phase of disease. A, low power view reveals intensified immunoreactivity in spinal cord ventral horns; multiple large, rounded spots are visible. B, higher power view of large immunoreactive spots is suggestive of phagocytic clusters. C, same field as in B; counterstaining with cresyl violet facilitates identification of large immunoreactive spots as multinucleated giant cells. D, E, the same microscopic field prior to and after cresyl violet counterstaining reveals a well-formed multinucleated giant cell of the Langhans type. F, enlargement of framed area in C shows apoptotic microglial nucleus (arrow) within a giant cell. Scale bars: 500 μm (A), 40 μm (B, C), 20 μm (D-F).
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Figure 2: OX-42 immunohistochemistry during symptomatic phase of disease. A, low power view reveals intensified immunoreactivity in spinal cord ventral horns; multiple large, rounded spots are visible. B, higher power view of large immunoreactive spots is suggestive of phagocytic clusters. C, same field as in B; counterstaining with cresyl violet facilitates identification of large immunoreactive spots as multinucleated giant cells. D, E, the same microscopic field prior to and after cresyl violet counterstaining reveals a well-formed multinucleated giant cell of the Langhans type. F, enlargement of framed area in C shows apoptotic microglial nucleus (arrow) within a giant cell. Scale bars: 500 μm (A), 40 μm (B, C), 20 μm (D-F).

Mentions: With onset of symptoms, there was apparent activation of microglia as judged by the dramatic increase in OX-42 immunoreactivity in the spinal gray matter. Examining sections at low power clearly revealed pronounced spots of enhanced OX-42 staining in the ventral horns (Fig. 2A), and these were judged initially to be due to the formation of microglial phagocytic clusters around dying motor neurons, as this would be a normal response to motor neuron death. However, when spots of intense OX-42 immunoreactivity were examined at higher power (Fig. 2B,D) they appeared unusual in that individual microglial phagocytes were not discernable. Subsequent counterstaining of these sections with cresyl violet allowed us to conclude that the OX-42 reactive structures were, in fact, not phagocytic clusters but represented multinucleated giant cells (Figs. 2C,E). These giant cells were found in all SOD1G93A transgenic rats studied. They formed apparently as a result of multiple microglial cells fusing together into sizable syncytia (40–50 μm) that often showed a circular arrangement of microglial nuclei about their periphery (Fig. 2E). This kind of nuclear arrangement is classically associated with multinucleated giant cells of the Langhans type. The cytoplasmic interior of Langhans giant cells appeared granular and fragmented, suggesting ongoing deterioration. A few of the giant cells revealed the presence of apoptotic bodies, evident as nuclear fragments (Fig. 2F), but overall apoptotic bodies either inside or outside of giant cells were sparse. Microglia dispersed in between giant cells revealed relatively normal process-bearing morphology and lacked the conspicuous hypertrophy that is characteristic of activated microglia. (Fig. 2E). However, some sections showed ongoing microglial cytorrhexis, i.e. fragmentation of the cytoplasm. Cytorrhexis became conspicuous in animals that were in the terminal stages of the disease process (Fig. 3) and was evident as a loss of discernable microglial cell structure and presence of abundant OX-42 immunoreactive fragments of microglial cytoplasm dispersed throughout the spinal gray matter (Figs. 3A,B,D,E). Occasional giant cells could still be observed during end stage disease, however, most of these showed signs of deterioration evident by increased irregularity of their shape and nuclear arrangement, as well as by increased granularity and fragmentation (Figs. 3A,B). In some sections, neurons remained stained with cresyl violet suggesting residual preservation of neuronal integrity. However, pathological features were evident in motor neurons, including most notably intense hyperchromia with formation of a nuclear cap consisting of condensed chromatin material (Fig. 3C). This neuronal appearance stood in stark contrast to that of normal motor neurons as seen in wild type animals (Fig. 3F).


Formation of multinucleated giant cells and microglial degeneration in rats expressing a mutant Cu/Zn superoxide dismutase gene.

Fendrick SE, Xue QS, Streit WJ - J Neuroinflammation (2007)

OX-42 immunohistochemistry during symptomatic phase of disease. A, low power view reveals intensified immunoreactivity in spinal cord ventral horns; multiple large, rounded spots are visible. B, higher power view of large immunoreactive spots is suggestive of phagocytic clusters. C, same field as in B; counterstaining with cresyl violet facilitates identification of large immunoreactive spots as multinucleated giant cells. D, E, the same microscopic field prior to and after cresyl violet counterstaining reveals a well-formed multinucleated giant cell of the Langhans type. F, enlargement of framed area in C shows apoptotic microglial nucleus (arrow) within a giant cell. Scale bars: 500 μm (A), 40 μm (B, C), 20 μm (D-F).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC1808448&req=5

Figure 2: OX-42 immunohistochemistry during symptomatic phase of disease. A, low power view reveals intensified immunoreactivity in spinal cord ventral horns; multiple large, rounded spots are visible. B, higher power view of large immunoreactive spots is suggestive of phagocytic clusters. C, same field as in B; counterstaining with cresyl violet facilitates identification of large immunoreactive spots as multinucleated giant cells. D, E, the same microscopic field prior to and after cresyl violet counterstaining reveals a well-formed multinucleated giant cell of the Langhans type. F, enlargement of framed area in C shows apoptotic microglial nucleus (arrow) within a giant cell. Scale bars: 500 μm (A), 40 μm (B, C), 20 μm (D-F).
Mentions: With onset of symptoms, there was apparent activation of microglia as judged by the dramatic increase in OX-42 immunoreactivity in the spinal gray matter. Examining sections at low power clearly revealed pronounced spots of enhanced OX-42 staining in the ventral horns (Fig. 2A), and these were judged initially to be due to the formation of microglial phagocytic clusters around dying motor neurons, as this would be a normal response to motor neuron death. However, when spots of intense OX-42 immunoreactivity were examined at higher power (Fig. 2B,D) they appeared unusual in that individual microglial phagocytes were not discernable. Subsequent counterstaining of these sections with cresyl violet allowed us to conclude that the OX-42 reactive structures were, in fact, not phagocytic clusters but represented multinucleated giant cells (Figs. 2C,E). These giant cells were found in all SOD1G93A transgenic rats studied. They formed apparently as a result of multiple microglial cells fusing together into sizable syncytia (40–50 μm) that often showed a circular arrangement of microglial nuclei about their periphery (Fig. 2E). This kind of nuclear arrangement is classically associated with multinucleated giant cells of the Langhans type. The cytoplasmic interior of Langhans giant cells appeared granular and fragmented, suggesting ongoing deterioration. A few of the giant cells revealed the presence of apoptotic bodies, evident as nuclear fragments (Fig. 2F), but overall apoptotic bodies either inside or outside of giant cells were sparse. Microglia dispersed in between giant cells revealed relatively normal process-bearing morphology and lacked the conspicuous hypertrophy that is characteristic of activated microglia. (Fig. 2E). However, some sections showed ongoing microglial cytorrhexis, i.e. fragmentation of the cytoplasm. Cytorrhexis became conspicuous in animals that were in the terminal stages of the disease process (Fig. 3) and was evident as a loss of discernable microglial cell structure and presence of abundant OX-42 immunoreactive fragments of microglial cytoplasm dispersed throughout the spinal gray matter (Figs. 3A,B,D,E). Occasional giant cells could still be observed during end stage disease, however, most of these showed signs of deterioration evident by increased irregularity of their shape and nuclear arrangement, as well as by increased granularity and fragmentation (Figs. 3A,B). In some sections, neurons remained stained with cresyl violet suggesting residual preservation of neuronal integrity. However, pathological features were evident in motor neurons, including most notably intense hyperchromia with formation of a nuclear cap consisting of condensed chromatin material (Fig. 3C). This neuronal appearance stood in stark contrast to that of normal motor neurons as seen in wild type animals (Fig. 3F).

Bottom Line: In animals during end stage disease at 4-5 months of age virtually all microglia in the spinal cord gray matter showed extensive fragmentation of their cytoplasm (cytorrhexis), indicative of widespread microglial degeneration.Few microglia exhibiting nuclear fragmentation (karyorrhexis) indicative of apoptosis were identified at any stage.The current findings demonstrate the occurrence of severe abnormalities in microglia, such as cell fusions and cytorrhexis, which may be the result of expression of mutant SOD1 in these cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Neuroscience, University of Florida College of Medicine and McKnight Brain Institute, Gainesville, FL 32611, USA. sefendrick@yahoo.com

ABSTRACT

Background: Microglial neuroinflammation is thought to play a role in the pathogenesis of amyotrophic lateral sclerosis (ALS). The purpose of this study was to provide a histopathological evaluation of the microglial neuroinflammatory response in a rodent model of ALS, the SOD1G93A transgenic rat.

Methods: Multiple levels of the CNS from spinal cord to cerebral cortex were studied in SOD1G93A transgenic rats during three stages of natural disease progression, including presymptomatic, early symptomatic (onset), and late symptomatic (end stage), using immuno- and lectin histochemical markers for microglia, such as OX-42, OX-6, and Griffonia simplicifolia isolectin B4.

Results: Our studies revealed abnormal aggregates of microglia forming in the spinal cord as early as the presymptomatic stage. During the symptomatic stages there was prominent formation of multinucleated giant cells through fusion of microglial cells in the spinal cord, brainstem, and red nucleus of the midbrain. Other brain regions, including substantia nigra, cranial nerve nuclei, hippocampus and cortex showed normal appearing microglia. In animals during end stage disease at 4-5 months of age virtually all microglia in the spinal cord gray matter showed extensive fragmentation of their cytoplasm (cytorrhexis), indicative of widespread microglial degeneration. Few microglia exhibiting nuclear fragmentation (karyorrhexis) indicative of apoptosis were identified at any stage.

Conclusion: The current findings demonstrate the occurrence of severe abnormalities in microglia, such as cell fusions and cytorrhexis, which may be the result of expression of mutant SOD1 in these cells. The microglial changes observed are different from those that accompany normal microglial activation, and they demonstrate that aberrant activation and degeneration of microglia is part of the pathogenesis of motor neuron disease.

Show MeSH
Related in: MedlinePlus