Studies of respiratory chlamydia infections in mouse models of chlamydia infection and alzheimer’s disease

Woods, Jason John

Title: Studies of respiratory chlamydia infections in mouse models of chlamydia infection and alzheimer’s disease
Creator: Woods, Jason John
Relation: University of Newcastle Research Higher Degree Thesis
Resource Type: thesis
Date: 2021
Description: Research Doctorate - Doctor of Philosophy (PhD)
Description: Alzheimer’s disease (AD) is a neurodegenerative proteopathy associated with progressive cognitive impairment and characterised neuropathologically by neurofibrillary tangles and Aβ-amyloid plaques. Brain presence of the common respiratory bacteria Chlamydia pneumoniae (Cpn) is proposed to induce AD β-amyloidosis but this is controversial, with human studies giving inconsistent results. Mouse studies have also been inconclusive, having previously used only wild-type mice, which do not usually form Aβ-amyloid, together with methods that do not distinguish non-amyloid Aβ deposits from Aβ-amyloid deposits. In addition, Cpn is not a natural mouse pathogen, requiring large inocula that are not physiologically relevant. If Chlamydia has roles in AD pathogenesis, treating AD patients for Cpn infection may be a viable strategy. This study tests the broad hypothesis that respiratory Chlamydia infection can gain entry to the brain, increase brain Aβ or Aβ-amyloid deposition, trigger AD-relevant gene expression changes and cause neuroinflammation. To address limitations of past research, both wild-type mice and the APP/PS1 transgenic mouse model of β-amyloidosis were infected with Chlamydia muridarum (Cmu), a related natural mouse pathogen more appropriately modelling respiratory Chlamydia infection. Also, since a defining feature of amyloid is affinity for the dye Congo red, with concomitant yellow-green birefringence under cross-polarised light, ‘gold standard’ Congo red polarisation microscopy was used to assess Aβ-amyloid. Wild-type and APP/PS1 mice were intranasally infected with Cmu, or sham infected with vehicle alone, and brains examined at 6 months of age. There were three endpoint matched infection scenarios: i) adult mice (6 months) followed-up short-term 10 days post-infection to assess Cmu brain entry and acute responses during peak respiratory infection, ii) adult mice (3 months) followed-up long-term at 3 months post-infection to assess Cmu brain persistence and longer-term responses and iii) neonatal mice (24 hours) followed-up long-term at 6 months to assess if early-life exposure may cause long-term changes. Adult mice infected with 100 inclusion forming units (IFU) and followed-up short-term (10 days) had high Cmu 16S rRNA signal in lungs by real-time reverse transcription polymerase chain reaction (RT-PCR) and considerable weight loss, consistent with severe infection. Adult mice followed-up long-term (3 months) were therefore given a slightly lower dose (75 IFU) and showed modest weight loss, consistent with moderate infection. Neonatal mice, which require higher Cmu doses to produce the same disease severity, probably due to differences in immune responses, received 400 IFU within 24 hours after birth and were followed-up long-term (6 months). There was no clear weight loss but alveolar damage (estimated with mean linear intercept method in lung sections) was consistent with successful infection. Only adult mice followed-up short-term had evidence of Cmu brain entry by 16S rRNA RT-PCR. Signal was small but increased brain levels of innate immune response gene transcripts were detected by RT-PCR, suggesting Cmu may have entered the brain in small quantities in early infection but was subsequently cleared, with no evidence of Cmu in the brain at long-term follow-up. Peripheral immune responses signalling across brain barriers could feasibly alter brain Aβ deposition whether or not brain entry occurred. However, there was no evidence for changes in Aβ or Aβ-amyloid deposition in any infection scenario by antibody 4G8 immunolabelling, thioflavin S or Congo red staining by fluorescence or polarised light microscopy, the ‘gold standard’ for amyloid detection. Notably, 75% of Aβ-immunoreactive structures and 20% of fluorescent structures were non-birefringent. No changes clearly relevant to AD were observed in expression of the AD-related genes App, Psen1, and Mapt with RT-PCR. Microarray and bioinformatics analyses of neonatal mice followed-up long-term also showed minor long-term changes but none appeared relevant to AD. There were no differences in microglial proliferation or morphology in any scenario as assessed by morphometric analysis of antibody IBA1-immunolabelled microglia. The results support the hypothesis that respiratory Chlamydia infection can gain entry to the brain in early or later life but do not provide evidence for short- or long-term effects on brain Aβ deposition. Future studies could assess the validity of these findings by intracerebral injection of larger inocula of Cmu, or by infecting older mice with age-related immunosenescence. However, based on present evidence, there is no justification for screening and treating AD patients without concurrent respiratory illness for Cpn infection. This project highlights the need for future AD studies to complement other methods with polarised light microscopy of Congo red staining, which is currently rarely performed in AD studies. Differentiating Aβ-amyloid and non-amyloid Aβ structures should improve understanding of the mechanisms underlying β-amyloidosis and of the actions of new drugs on Aβ deposition and β-amyloidosis in animal models before proceeding to human trials.
Subject: alzheimer disease; chlamydia pneumoniae; chlamydia muridarum; infections; inflammation; risk factors; amyloid
Identifier: http://hdl.handle.net/1959.13/1489007
Identifier: uon:52595
Language: eng
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