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Protein Citrullination: A Proposed Mechanism for Pathology in Traumatic Brain Injury.

Lazarus RC, Buonora JE, Flora MN, Freedy JG, Holstein GR, Martinelli GP, Jacobowitz DM, Mueller GP - Front Neurol (2015)

Bottom Line: The present investigation addressed this gap by examining the effects of TBI on the distribution of protein citrullination and on the specific cell types involved.This response was exclusively seen in astrocytes; no such effects were observed on the status of protein citrullination in neurons, oligodendrocytes or microglia.Further, proteomic analyses demonstrated that the effects of TBI on citrullination were confined to a relatively small subset of neural proteins.

View Article: PubMed Central - PubMed

Affiliation: Program in Neuroscience, Uniformed Services University of the Health Sciences , Bethesda, MD , USA.

ABSTRACT
Protein citrullination is a calcium-driven post-translational modification proposed to play a causative role in the neurodegenerative disorders of Alzheimer's disease, multiple sclerosis (MS), and prion disease. Citrullination can result in the formation of antigenic epitopes that underlie pathogenic autoimmune responses. This phenomenon, which is best understood in rheumatoid arthritis, may play a role in the chronic dysfunction following traumatic brain injury (TBI). Despite substantial evidence of aberrations in calcium signaling following TBI, there is little understanding of how TBI alters citrullination in the brain. The present investigation addressed this gap by examining the effects of TBI on the distribution of protein citrullination and on the specific cell types involved. Immunofluorescence revealed that controlled cortical impact in rats profoundly up--regulated protein citrullination in the cerebral cortex, external capsule, and hippocampus. This response was exclusively seen in astrocytes; no such effects were observed on the status of protein citrullination in neurons, oligodendrocytes or microglia. Further, proteomic analyses demonstrated that the effects of TBI on citrullination were confined to a relatively small subset of neural proteins. Proteins most notably affected were those also reported to be citrullinated in other disorders, including prion disease and MS. In vivo findings were extended in an in vitro model of simulated TBI employing normal human astrocytes. Pharmacologically induced calcium excitotoxicity was shown to activate the citrullination and breakdown of glial fibrillary acidic protein, producing a novel candidate TBI biomarker and potential target for autoimmune recognition. In summary, these findings demonstrate that the effects of TBI on protein citrullination are selective with respect to brain region, cell type, and proteins modified, and may contribute to a role for autoimmune dysfunction in chronic pathology following TBI.

No MeSH data available.


Related in: MedlinePlus

Localization of CCI-induced protein citrullination to astrocytes. Panels on the left show the co-localization of mAb 6B3 labeling with anti-GFAP labeling in the external capsule. Panels on the right show the co-localization of mAb 6B3 labeling with anti-GFAP labeling in the cerebral cortex. Data are representative of 15 control animals (8 males and 7 females) and 21 CCI animals (11 males and 10 females). No gender-based differences were observed. Scale bar: 200 μm. PB-cit, protein-bound citrulline.
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Figure 4: Localization of CCI-induced protein citrullination to astrocytes. Panels on the left show the co-localization of mAb 6B3 labeling with anti-GFAP labeling in the external capsule. Panels on the right show the co-localization of mAb 6B3 labeling with anti-GFAP labeling in the cerebral cortex. Data are representative of 15 control animals (8 males and 7 females) and 21 CCI animals (11 males and 10 females). No gender-based differences were observed. Scale bar: 200 μm. PB-cit, protein-bound citrulline.

Mentions: Dual immunofluorescence revealed astrocytes to be the principal cell type in which protein citrullination was affected by CCI. Figure 4 shows that anti-citrulline labeling in the cortex and external capsule was predominantly co-localized with GFAP. Similar observations were made in other affected brain regions. The findings presented in Figure 5 further confirm that CCI-induced protein citrullination was not significantly associated with neurons (NeuN), microglia/macrophages (Iba1), or oligodendrocytes (MBP) in the cortex. Citrullination was also not significantly associated with these cell types in any other brain regions investigated (not shown). It should be noted that the lack of co-localization of protein citrullination with NeuN immunoreactivity might reflect diminished NeuN expression following TBI, rather than a lack of citrullinated proteins within neurons. The distinctive profile of CCI-induced protein citrullination in the ipsilateral and contralateral hippocampus is shown in Figure 6. As noted above, increased anti-citrullinated protein labeling was clearly evident in the ipsilateral hippocampus of all animals studied (Figure 6A), and this labeling co-localized with GFAP labeling (Figures 6B,C). In addition, in ~20% of injured animals, the contralateral hippocampus also displayed intense 6B3 labeling (Figure 6D) that co-localized with GFAP labeling. The co-labeled cells were morphologically distinct from traditional stellate astrocytes, displaying a rounded, branchless appearance consistent with that of a macrophage (Figures 6E–G).


Protein Citrullination: A Proposed Mechanism for Pathology in Traumatic Brain Injury.

Lazarus RC, Buonora JE, Flora MN, Freedy JG, Holstein GR, Martinelli GP, Jacobowitz DM, Mueller GP - Front Neurol (2015)

Localization of CCI-induced protein citrullination to astrocytes. Panels on the left show the co-localization of mAb 6B3 labeling with anti-GFAP labeling in the external capsule. Panels on the right show the co-localization of mAb 6B3 labeling with anti-GFAP labeling in the cerebral cortex. Data are representative of 15 control animals (8 males and 7 females) and 21 CCI animals (11 males and 10 females). No gender-based differences were observed. Scale bar: 200 μm. PB-cit, protein-bound citrulline.
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Related In: Results  -  Collection

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Figure 4: Localization of CCI-induced protein citrullination to astrocytes. Panels on the left show the co-localization of mAb 6B3 labeling with anti-GFAP labeling in the external capsule. Panels on the right show the co-localization of mAb 6B3 labeling with anti-GFAP labeling in the cerebral cortex. Data are representative of 15 control animals (8 males and 7 females) and 21 CCI animals (11 males and 10 females). No gender-based differences were observed. Scale bar: 200 μm. PB-cit, protein-bound citrulline.
Mentions: Dual immunofluorescence revealed astrocytes to be the principal cell type in which protein citrullination was affected by CCI. Figure 4 shows that anti-citrulline labeling in the cortex and external capsule was predominantly co-localized with GFAP. Similar observations were made in other affected brain regions. The findings presented in Figure 5 further confirm that CCI-induced protein citrullination was not significantly associated with neurons (NeuN), microglia/macrophages (Iba1), or oligodendrocytes (MBP) in the cortex. Citrullination was also not significantly associated with these cell types in any other brain regions investigated (not shown). It should be noted that the lack of co-localization of protein citrullination with NeuN immunoreactivity might reflect diminished NeuN expression following TBI, rather than a lack of citrullinated proteins within neurons. The distinctive profile of CCI-induced protein citrullination in the ipsilateral and contralateral hippocampus is shown in Figure 6. As noted above, increased anti-citrullinated protein labeling was clearly evident in the ipsilateral hippocampus of all animals studied (Figure 6A), and this labeling co-localized with GFAP labeling (Figures 6B,C). In addition, in ~20% of injured animals, the contralateral hippocampus also displayed intense 6B3 labeling (Figure 6D) that co-localized with GFAP labeling. The co-labeled cells were morphologically distinct from traditional stellate astrocytes, displaying a rounded, branchless appearance consistent with that of a macrophage (Figures 6E–G).

Bottom Line: The present investigation addressed this gap by examining the effects of TBI on the distribution of protein citrullination and on the specific cell types involved.This response was exclusively seen in astrocytes; no such effects were observed on the status of protein citrullination in neurons, oligodendrocytes or microglia.Further, proteomic analyses demonstrated that the effects of TBI on citrullination were confined to a relatively small subset of neural proteins.

View Article: PubMed Central - PubMed

Affiliation: Program in Neuroscience, Uniformed Services University of the Health Sciences , Bethesda, MD , USA.

ABSTRACT
Protein citrullination is a calcium-driven post-translational modification proposed to play a causative role in the neurodegenerative disorders of Alzheimer's disease, multiple sclerosis (MS), and prion disease. Citrullination can result in the formation of antigenic epitopes that underlie pathogenic autoimmune responses. This phenomenon, which is best understood in rheumatoid arthritis, may play a role in the chronic dysfunction following traumatic brain injury (TBI). Despite substantial evidence of aberrations in calcium signaling following TBI, there is little understanding of how TBI alters citrullination in the brain. The present investigation addressed this gap by examining the effects of TBI on the distribution of protein citrullination and on the specific cell types involved. Immunofluorescence revealed that controlled cortical impact in rats profoundly up--regulated protein citrullination in the cerebral cortex, external capsule, and hippocampus. This response was exclusively seen in astrocytes; no such effects were observed on the status of protein citrullination in neurons, oligodendrocytes or microglia. Further, proteomic analyses demonstrated that the effects of TBI on citrullination were confined to a relatively small subset of neural proteins. Proteins most notably affected were those also reported to be citrullinated in other disorders, including prion disease and MS. In vivo findings were extended in an in vitro model of simulated TBI employing normal human astrocytes. Pharmacologically induced calcium excitotoxicity was shown to activate the citrullination and breakdown of glial fibrillary acidic protein, producing a novel candidate TBI biomarker and potential target for autoimmune recognition. In summary, these findings demonstrate that the effects of TBI on protein citrullination are selective with respect to brain region, cell type, and proteins modified, and may contribute to a role for autoimmune dysfunction in chronic pathology following TBI.

No MeSH data available.


Related in: MedlinePlus