Cerebral Oxidative Stress and Microvasculature Defects in TNF-α Expressing Transgenic and Porphyromonas gingivalis-Infected ApoE^–/– Mice

Article type: Research Article

Authors: Rokad, Farheen^a | Moseley, Ryan^b | Hardy, Rowan S.^c | Chukkapalli, Sasanka^d | Crean, StJohn^a | Kesavalu, Lakshmyya^{d; e} | Singhrao, Sim K.^{a; *}

Affiliations: [a] Dementia and Neurodegenerative Diseases Research Group, Faculty of Clinical and Biomedical Sciences, School of Dentistry, University of Central Lancashire, Preston, UK | [b] Stem Cells, Wound Repair and Regeneration, School of Dentistry, Cardiff University, Cardiff, UK | [c] Institute of Metabolism and Systems Research, University of Birmingham, Birmingham, UK | [d] Department of Periodontology, College of Dentistry, University of Florida, Gainesville, FL, USA | [e] Department of Oral Biology, College of Dentistry, University of Florida, Gainesville, FL, USA

Correspondence: [*] Correspondence to: Dr. Sim K. Singhrao, Dementia & Neurodegenerative Diseases Research Group, Faculty of Clinical and Biomedical Sciences, School of Dentistry, University of Central Lancashire, Preston, UK. Tel.: +44 0 1772 895137; E-mail: [email protected].

Abstract: The polymicrobial dysbiotic subgingival biofilm microbes associated with periodontal disease appear to contribute to developing pathologies in distal body sites, including the brain. This study examined oxidative stress, in the form of increased protein carbonylation and oxidative protein damage, in the tumor necrosis factor-α (TNF-α) transgenic mouse that models inflammatory TNF-α excess during bacterial infection; and in the apolipoprotein knockout (ApoE–/–) mouse brains, following Porphyromonas gingivalis gingival monoinfection. Following 2,4-dinitrophenylhydrazine derivatization, carbonyl groups were detected in frontal lobe brain tissue lysates by immunoblotting and immunohistochemical analysis of fixed tissue sections from the frontotemporal lobe and the hippocampus. Immunoblot analysis confirmed the presence of variable carbonyl content and oxidative protein damage in all lysates, with TNF-α transgenic blots exhibiting increased protein carbonyl content, with consistently prominent bands at 25 kDa (p = 0.0001), 43 kDa, and 68 kDa, over wild-type mice. Compared to sham-infected ApoE–/– mouse blots, P. gingivalis-infected brain tissue blots demonstrated the greatest detectable protein carbonyl content overall, with numerous prominent bands at 25 kDa (p = 0.001) and 43 kDa (p = 0.0001) and an exclusive band to this group between 30–43 kDa* (p = 0.0001). In addition, marked immunostaining was detected exclusively in the microvasculature in P. gingivalis-infected hippocampal tissue sections, compared to sham-infected, wild-type, and TNF-α transgenic mice. This study revealed that the hippocampal microvascular structure of P. gingivalis-infected ApoE–/– mice possesses elevated oxidative stress levels, resulting in the associated tight junction proteins being susceptible to increased oxidative/proteolytic degradation, leading to a loss of functional integrity.

Keywords: Hippocampus, infection, microvasculature, oxidative stress/damage, Porphyromonas gingivalis, tight junction proteins

DOI: 10.3233/JAD-170304

Journal: Journal of Alzheimer's Disease, vol. 60, no. 2, pp. 359-369, 2017