A number of clinical and experimental studies have revealed a strong association between periodontitis and accelerated cognitive decline in Alzheimer's disease (AD); however, the mechanism of the association is unknown. In the present study, we tested the hypothesis that cathepsin (Cat) B plays a critical role in the initiation of neuroinflammation and neural dysfunction following chronic systemic exposure to lipopolysaccharide from Porphyromonas gingivalis (PgLPS) in mice (1mg/kg, daily, intraperitoneally). Young (2months old) and middle-aged (12months old) wild-type (WT; C57BL/6N) or CatB-deficient (CatB) mice were exposed to PgLPS daily for 5 consecutive weeks. The learning and memory function were assessed using the passive avoidance test, and the expression of amyloid precursor protein (APP), CatB, TLR2 and IL-1β was analyzed in brain tissues by immunohistochemistry and Western blotting. We found that chronic systemic exposure to PgLPS for five consecutive weeks induced learning and memory deficits with the intracellular accumulation of Aβ in neurons in the middle-aged WT mice, but not in young WT or middle-aged CatB mice. PgLPS significantly increased the expression of CatB in both microglia and neurons in middle-aged WT mice, while increased expression of mature IL-1β and TLR2 was restricted to microglia in the hippocampus of middle-aged WT mice, but not in that of the middle-aged CatB ones. In in vitro studies, PgLPS (1µg/ml) stimulation upregulated the mean mRNA expression of IL-1β, TLR2 and downregulated the protein levels of IκBα in the cultured MG6 microglia as well as in the primary microglia from WT mice, which were significantly inhibited by the CatB-specific inhibitor CA-074Me as well as by the primary microglia from CatB mice. Furthermore, the mean mRNA expression of APP and CatB were significantly increased in the primary cultured hippocampal neurons after treatment with conditioned medium from PgLPS-treated WT primary microglia, but not after treatment with conditioned medium neutralized with anti-IL-1beta, and not after treatment with conditioned medium from PgLPS-treated CatB primary microglia or with PgLPS directly. Taken together, these findings indicate that chronic systemic exposure to PgLPS induces AD-like phenotypes, including microglia-mediated neuroinflammation, intracellular Aβ accumulation in neurons and impairment of the learning and memory functions in the middle-aged mice in a CatB-dependent manner. We propose that CatB may be a therapeutic target for preventing periodontitis-associated cognitive decline in AD.

Despite a clear correlation between periodontitis and cognitive decline in Alzheimer’s disease, the precise mechanism underlying the relationship remains unclear. The periodontal pathogen Porphyromonas gingivalis produces a unique class of cysteine proteinases termed gingipains that comprises Arg-gingipain (Rgp) and Lys-gingipain (Kgp). Rgp and Kgp are important in the bacterial mediated host cell responses and the subsequent intracellular signaling in infected cells. In the present study, we attempted to clarify the potential effects of Rgp and Kgp on the cellular activation of brain-resident microglia. We provide the first evidence that Rgp and Kgp cooperatively contribute to the P. gingivalis-induced cell migration and expression of proinflammatory mediators through the activation of protease-activated receptor 2. The subsequent activation of phosphoinositide 3-kinase/Akt and mitogen-activated protein kinase/extracellular signal-regulated kinase (ERK) kinase/ERK pathways contributes to cell migration and inflammatory response of microglia.

Hypoxia has been recently proposed as a neuroinflammatogen, which drives microglia to produce proinflammatory cytokines, including interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), and IL-6. Considering the fact that propolis has hepatoprotective, antitumor, antioxidative, and anti-inflammatory effects, propolis may have protective effects against the hypoxia-induced neuroinflammatory responses. In this study, propolis (50 μg/mL) was found to significantly inhibit the hypoxia-induced cytotoxicity and the release of proinflammatory cytokines, including IL-1β, TNF-α, and IL-6, by MG6 microglia following hypoxic exposure (1% O2, 24 h). Furthermore, propolis significantly inhibited the hypoxia-induced generation of reactive oxygen species (ROS) from mitochondria and the activation of nuclear factor-κB (NF-κB) in microglia. Moreover, systemic treatment with propolis (8.33 mg/kg, 2 times/day, i.p.) for 7 days significantly suppressed the microglial expression of IL-1β, TNF-α, IL-6, and 8-oxo-deoxyguanosine, a biomarker for oxidative damaged DNA, in the somatosensory cortex of mice subjected to hypoxia exposure (10% O2, 4 h). These observations indicate that propolis suppresses the hypoxia-induced neuroinflammatory responses through inhibition of the NF-κB activation in microglia. Furthermore, increased generation of ROS from the mitochondria is responsible for the NF-κB activation. Therefore, propolis may be beneficial in preventing hypoxia-induced neuroinflammation.

It has long been believed that microglia morphologically transform into the activated state by retracting their long processes and consuming pathogens when bacteria infect into the brain parenchyma. In the present study, however, we showed for the first time that murine cortical microglia extend their processes towards focally injected Porphyromonas gingivalis. This P. gingivalis-induced microglial process extension was significantly increased during the light (sleeping) phase than the dark (waking) phase. In contrast, focally injected ATP-induced microglial process extension was significantly increased during the dark phase than the light phase. Furthermore, in contrast to the P2Y12 receptor-mediated mechanism of ATP-induced microglial process extension, the P. gingivalis-mediated microglial process extension was mediated by P2Y6 receptors. The infection of bacteria such as P. gingivalis to the brain parenchyma may induce the secretion of UDP from microglia at the site of infection, which in turn induces the process extension of the neighboring microglia.

Autism spectrum disorder (ASD) is a highly heterogeneous neurodevelopmental disorder characterized by impaired social interactions, restrictive interests, and repetitive stereotypic behaviors. Among the various mechanisms underlying the pathogenesis of ASD, dysfunctions of dopaminergic signaling and mitochondria have been hypothesized to explain the core symptoms of children with ASD. However, only a few studies focusing on the pathological association between dopaminergic neurons (DN) and mitochondria in ASD have been performed using patient-derived stem cells and in vitro differentiated neurons. Stem cells from human exfoliated deciduous teeth (SHED) are neural crest-derived mesenchymal stem cells present in the dental pulp of exfoliated deciduous teeth; these cells can differentiate into dopaminergic neurons (DN) in vitro. This study aimed to investigate the pathological association between development of DN and mitochondria in ASD by using SHED as a disease- or patient-specific cellular model. The SHED obtained from three children with ASD and three typically developing children were differentiated into DN, and the neurobiology of these cells was examined. The DN derived from children with ASD showed impaired neurite outgrowth and branching, associated with decreased mitochondrial membrane potential, ATP production, number of mitochondria within the neurites, amount of mitochondria per cell area and intracellular calcium level. In addition, impaired neurite outgrowth and branching of ASD-derived DN were not improved by brain-derived neurotrophic factor (BDNF), suggesting impairment of the BDNF signaling pathway in ASD. These results imply that intracerebral dopamine production may have decreased in these children. The earliest age at which deciduous teeth spontaneously exfoliate in humans, and SHED can be noninvasively collected, is approximately 6 years. Our results suggest that in vitro analysis of SHED-derived DN obtained from children with ASD provides neurobiological information that may be useful in determining treatment strategies in the early stages of ASD.