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Cited by 156Alzheimer's disease research and treatmentsNeuroinflammation and Neurodegeneration MechanismsDementia and Cognitive Impairment Research

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Immunohistological detection of Chlamydia pneumoniae in the Alzheimer's disease brain

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Hammond, Christine J, Hallock, Loretta R, Howanski, Raymond J, Appelt, Denah M, Little, C Scott, Balin, Brian J·2010·BMC Neuroscience
DOI 10.1186/1471-2202-11-121PMC PMC2949767
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Abstract

Background: Sporadic late-onset Alzheimer's disease (AD) appears to evolve from an interplay between genetic and environmental factors. One environmental factor that continues to be of great interest is that of Chlamydia pneumoniae infection and its association with late-onset disease. Detection of this organism in clinical and autopsy samples has proved challenging using a variety of molecular and histological techniques. Our current investigation utilized immunohistochemistry with a battery of commercially available anti-C. pneumoniae antibodies to determine whether C. pneumoniae was present in areas typically associated with AD neuropathology from 5 AD and 5 non-AD control brains. Results: Immunoreactivity for C. pneumoniae antigens was observed both intracellularly in neurons, neuroglia, endothelial cells, and peri-endothelial cells, and extracellularly in the frontal and temporal cortices of the AD brain with multiple C. pneumoniae-specific antibodies. This immunoreactivity was seen in regions of amyloid deposition as revealed by immunolabeling with two different anti-beta amyloid antibodies. Thioflavin S staining, overlaid with C. pneumoniae immunolabeling, demonstrated no direct co-localization of the organism and amyloid plaques. Further, the specificity of C. pneumoniae labeling of AD brain sections was demonstrated using C. pneumoniae antibodies pre-absorbed against amyloid β 1-40 and 1-42 peptides. Conclusions: Anti-C. pneumoniae antibodies, obtained commercially, identified both typical intracellular and atypical extracellular C. pneumoniae antigens in frontal and temporal cortices of the AD brain. C. pneumoniae, amyloid deposits, and neurofibrillary tangles were present in the same regions of the brain in apposition to one another. Although additional studies are required to conclusively characterize the nature of Chlamydial immunoreactivity in the AD brain, these results further implicate C. pneumoniae infection with the pathogenesis of Alzheimer's disease.

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Figure 1 Chlamydia pneumoniae immunoreactivity in AD brain tissue. Alzheimer's disease brain cerebral cortex immunolabeled with a mouse anti-C. pneumoniae monoclonal antibody (Table 1, #5) demonstrates both intracellular and extracellular immunolabeling as indicated by the magenta color. Panel A illustrates the temporal cortex labeled with no primary antibody and both anti- mouse HRP and anti-mouse AP conjugated secondary antibodies reacted with both DAB and AP red. Panel B illustrates an extracellular labeling pattern and panels C and D illustrate cellular labeling in the frontal cortex. Areas delineated by the boxes are higher magnification images of the immunoreactivity in panels B-D. Size bar = 50 μm.
Figure 1 Chlamydia pneumoniae immunoreactivity in AD brain tissue. Alzheimer's disease brain cerebral cortex immunolabeled with a mouse anti-C. pneumoniae monoclonal antibody (Table 1, #5) demonstrates both intracellular and extracellular immunolabeling as indicated by the magenta color. Panel A illustrates the temporal cortex labeled with no primary antibody and both anti- mouse HRP and anti-mouse AP conjugated secondary antibodies reacted with both DAB and AP red. Panel B illustrates an extracellular labeling pattern and panels C and D illustrate cellular labeling in the frontal cortex. Areas delineated by the boxes are higher magnification images of the immunoreactivity in panels B-D. Size bar = 50 μm.CC BY · PubMed Central
Figure 2 Chlamydia pneumoniae immunoreactivity in hippocampus and entorhinal Cortex. Detection of C. pneumoniae antigen in the Alzheimer's disease hippocampus in panels A and B. Representative immunolabeling with mouse anti-C. pneumoniae monoclonal antibody (Table 1, #5) shows immunopositivity in the dentate gyrus of the hippocampus. Immunolabeling is indicated by the magenta color and is denoted by the arrows in panel A. Extracellular and intracellular C. pneumoniae immunolabeling in apparent large neurons is found within the entorhinal cortex, panels C and D. Prominent labeling (magenta) denoted by the arrows in panels C and D, is easily distinguished from intraneuronal lipofuscin (golden brown) denoted by the arrowheads (panel D). High magnification images of chlamydial labeling within different cell types in the hippocampal region are illustrated in panel E (1, 8: neuroglia; 2: peri-vascular; 3, 4, 5, 6, 7, 9: neuronal). Note the apparent tangle within the neuron in E9.Size bars: A, C, and D = 50 μm; B = 20 μm; E = 10 μm.
Figure 2 Chlamydia pneumoniae immunoreactivity in hippocampus and entorhinal Cortex. Detection of C. pneumoniae antigen in the Alzheimer's disease hippocampus in panels A and B. Representative immunolabeling with mouse anti-C. pneumoniae monoclonal antibody (Table 1, #5) shows immunopositivity in the dentate gyrus of the hippocampus. Immunolabeling is indicated by the magenta color and is denoted by the arrows in panel A. Extracellular and intracellular C. pneumoniae immunolabeling in apparent large neurons is found within the entorhinal cortex, panels C and D. Prominent labeling (magenta) denoted by the arrows in panels C and D, is easily distinguished from intraneuronal lipofuscin (golden brown) denoted by the arrowheads (panel D). High magnification images of chlamydial labeling within different cell types in the hippocampal region are illustrated in panel E (1, 8: neuroglia; 2: peri-vascular; 3, 4, 5, 6, 7, 9: neuronal). Note the apparent tangle within the neuron in E9.Size bars: A, C, and D = 50 μm; B = 20 μm; E = 10 μm.CC BY · PubMed Central
Figure 3 Amyloid and Chlamydia pneumoniae immunolabeling in AD tissue. Representative amyloid and C. pneumoniae immunolabeling detected in frontal (A, B) and temporal (C, D) sections of an AD brain. For amyloid labeling (brown color), two different anti-amyloid antibodies were used, (Sigma, panel A) and (Signet, panel C). Representative amyloid immunoreactivity, both extracellular plaques, including dense core plaques (A), and intracellular labeling (C) are revealed at a low magnification in order to appreciate the level of amyloid pathology. Likewise, C. pneumoniae immunoreactivity (magenta color) is evident at low magnification in both brain regions using two different monoclonal antibodies (Table 1, #5 panel B), and (Table 1, #3 panel D). Size bar A, B = 50 μm; C, D = 100 μm.
Figure 3 Amyloid and Chlamydia pneumoniae immunolabeling in AD tissue. Representative amyloid and C. pneumoniae immunolabeling detected in frontal (A, B) and temporal (C, D) sections of an AD brain. For amyloid labeling (brown color), two different anti-amyloid antibodies were used, (Sigma, panel A) and (Signet, panel C). Representative amyloid immunoreactivity, both extracellular plaques, including dense core plaques (A), and intracellular labeling (C) are revealed at a low magnification in order to appreciate the level of amyloid pathology. Likewise, C. pneumoniae immunoreactivity (magenta color) is evident at low magnification in both brain regions using two different monoclonal antibodies (Table 1, #5 panel B), and (Table 1, #3 panel D). Size bar A, B = 50 μm; C, D = 100 μm.CC BY · PubMed Central
Figure 4 Dual Chlamydia pneumoniae labeling and Thioflavin S staining of the temporal cortex from an AD brain .Alzheimer's disease temporal cortex is immunolabeled with a monoclonal anti-C. pneumoniae antibody (Table 1, #5) followed by staining with Thioflavin S on the same tissue specimen. Panel A shows intracellular and extracellular C. pneumoniae immunoreactivity (magenta color). Using a FITC filter, panel B shows both Thioflavin S positive intracellular labeling, presumably NFTs, and extracellular amyloid plaques (yellow fluorescence). Panel C shows the light (panel A) (C. pneumoniae) image overlaid on the fluorescent (panel B) (Thioflavin S) image. Panel D shows merged light and fluorescent images of another region of this temporal cortex illustrating merged chlamydia chromogen immunoreactivity and Thioflavin S amyloid/tau fluorescence. Size bars = 50 μm.
Figure 4 Dual Chlamydia pneumoniae labeling and Thioflavin S staining of the temporal cortex from an AD brain .Alzheimer's disease temporal cortex is immunolabeled with a monoclonal anti-C. pneumoniae antibody (Table 1, #5) followed by staining with Thioflavin S on the same tissue specimen. Panel A shows intracellular and extracellular C. pneumoniae immunoreactivity (magenta color). Using a FITC filter, panel B shows both Thioflavin S positive intracellular labeling, presumably NFTs, and extracellular amyloid plaques (yellow fluorescence). Panel C shows the light (panel A) (C. pneumoniae) image overlaid on the fluorescent (panel B) (Thioflavin S) image. Panel D shows merged light and fluorescent images of another region of this temporal cortex illustrating merged chlamydia chromogen immunoreactivity and Thioflavin S amyloid/tau fluorescence. Size bars = 50 μm.CC BY · PubMed Central
Figure 5 Immunohistochemistry using an amyloid-pre-absorbed anti-Chlamydia pneumoniae antibody. Panels A-D illustrate anti-C. pneumoniae immunoreactivity (red) in frontal AD tissue. The antibody for detection of C. pneumoniae is a mouse monoclonal C. pneumoniae antibody (Table 1, #3). Panel A illustrates the extent of anti-C. pneumoniae immunoreactivity with the non-pre-absorbed antibody. Panel B shows the extent of immunolabeling on the tissue when labeled with the 1-40 pre-absorbed antibody (pre-absorbed against >10 molar excess Sigma amyloid Aβ1-40 peptide). Panels C and D (higher power of panel C) show the extent of immunolabeling on the tissue when labeled with the 1-42 pre-absorbed antibody (pre-absorbed against >10 molar excess Sigma amyloid Aβ1-42 peptide). Size bar for panels A-C = 50 μm and panel D = 20 μm.
Figure 5 Immunohistochemistry using an amyloid-pre-absorbed anti-Chlamydia pneumoniae antibody. Panels A-D illustrate anti-C. pneumoniae immunoreactivity (red) in frontal AD tissue. The antibody for detection of C. pneumoniae is a mouse monoclonal C. pneumoniae antibody (Table 1, #3). Panel A illustrates the extent of anti-C. pneumoniae immunoreactivity with the non-pre-absorbed antibody. Panel B shows the extent of immunolabeling on the tissue when labeled with the 1-40 pre-absorbed antibody (pre-absorbed against >10 molar excess Sigma amyloid Aβ1-40 peptide). Panels C and D (higher power of panel C) show the extent of immunolabeling on the tissue when labeled with the 1-42 pre-absorbed antibody (pre-absorbed against >10 molar excess Sigma amyloid Aβ1-42 peptide). Size bar for panels A-C = 50 μm and panel D = 20 μm.CC BY · PubMed Central

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Metadata

Authors
Hammond, C. J., Hallock, L. R., Howanski, R. J., Appelt, D. M., Little, C. S., & Balin, B. J.
Affiliations
Philadelphia College of Osteopathic Medicine (US)
Corresponding author
Balin, Brian J
ORCID
Venue
BMC Neuroscience
Publisher
Springer Science and Business Media LLC
Published
2010-09-23
Volume
11
Issue
1
Pages
121-121
ISSN
1471-2202 (ISSN-L)
DOI
10.1186/1471-2202-11-121
PMC
PMC2949767
OpenAlex
W2056188402
Cited by
156
References
49
Citations by year
2026: 5 · 2025: 11 · 2024: 6 · 2023: 4 · 2022: 21 · 2021: 12 · 2020: 17 · 2019: 11 · 2018: 9 · 2017: 8 · 2016: 12 · 2015: 7 · 2014: 11 · 2013: 11 · 2012: 4
FWCI
4.76
Type
journal article
Language
en
Open access
Licence
cc-by
Locations
  • BMC Neuroscience (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • Digital Commons - PCOM (Philadelphia College of Osteopathic Medicine) (submittedVersion, cc-by)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
Funders
Drexel University
Topics
  • Life Sciences › Neuroscience › Cellular and Molecular Neuroscience › Alzheimer's disease research and treatments
  • Life Sciences › Neuroscience › Neurology › Neuroinflammation and Neurodegeneration Mechanisms
  • Health Sciences › Medicine › Psychiatry and Mental health › Dementia and Cognitive Impairment Research
Keywords
Chlamydia pneumoniae, Alzheimer's disease, late-onset Alzheimer's disease, chlamydia, amyloid plaques, amyloid-beta, neurofibrillary tangles, frontal and temporal cortex, antibody specificity, neuroinflammation, Alzheimer's disease pathogenesis
MeSH terms
Aged, Alzheimer Disease, Brain, Brain Chemistry, Cerebral Cortex, Chlamydia Infections, Coloring Agents, Female, Humans, Immunohistochemistry, Male, Thiazoles, Tissue Banks, Reproducibility of Results, Amyloid beta-Peptides, Neurofibrillary Tangles, Chlamydophila pneumoniae, Benzothiazoles, Plaque, Amyloid

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