Common marmosets (Callithrix jacchus) have recently gained popularity in biomedical research as models of aging research. Basically, they confer advantages from other non-human primates due to their shorter lifespan with onset of appearance of aging at 8 years. Old marmosets present some markers linked to neurodegeneration in the brain such as amyloid beta (Aβ)1-42 and Aβ1-40. However, there are no studies exploring other cellular markers associated with neurodegenerative diseases in this non-human primate. Using immunohistochemistry, we analyzed brains of male adolescent, adult, old, and aged marmosets. We observed accumulation of Aβ1-40 and Aβ1-42 in the cortex of aged subjects. Tau hyperphosphorylation was already detected in the brain of adolescent animals and increased with aging in a more fibrillary form. Microglia activation was also observed in the aging process, while a dystrophic phenotype accumulates in aged subjects. Interestingly, dystrophic microglia contained hyperphosphorylated tau, but active microglia did not. These results support previous findings regarding microglia dysfunctionality in aging and neurodegenerative diseases as Alzheimer’s disease. Further studies should explore the functional consequences of these findings to position this non-human primate as animal model of aging and neurodegeneration.

number: 308515 Microglia are cells that protect brain tissue from invading agents and toxic substances, first by releasing pro-inflammatory cytokines, and thereafter by clearing tissue by phagocytosis. Microglia express ferritin, a protein with ferroxidase activity capable of storing iron, a metal that accumulates in brain during aging. Increasing evidence suggests that ferritin plays an important role in inflammation. However, it is not known if ferritin/ iron content can be related to the activation state of microglia. To this end, we aimed to delineate the role of ferritin in microglia activation in a non-human primate model. We analyzed brains of male marmosets and observed an increased density of ferritin+ microglia with an activated phenotype in hippocampus and cortex of old marmosets (mean age 11.25 ± 0.70 years) compared to younger subjects. This was accompanied by an increased number of dystrophic microglia in old marmosets. However, in aged subjects (mean age 16.83 ± 2.59 years) the number of ferritin+ microglia was decreased compared to old ones. Meanwhile, the content of iron in brain tissue and cells with oxidized RNA increased during aging in all hippocampal and cortical regions analyzed. Abundant amoeboid microglia were commonly observed surrounding neurons with oxidized RNA. Notably, amoeboid microglia were arginase1+ and IL-10+, indicative of a M2 phenotype. Some of those M2 cells also presented RNA oxidation and a dystrophic phenotype. Therefore, our data suggest that ferritin confers protection to microglia in adult and old marmosets, while in aged subjects the decline in ferritin and the increased amount of iron in brain tissue may be related to the increased number of cells with oxidized RNA, perhaps precluding the onset of neurodegeneration.

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Aging is a major risk factor for the development of neurodegenerative diseases. Alzheimer's disease and other neurodegenerative diseases are characterized by abnormal and prominent protein aggregation in the brain, partially due to deficiency in protein clearance. It has been proposed that alterations in microglia phagocytosis and debris clearance hasten the onset of neurodegeneration. Dystrophic microglia are abundant in aged humans, and it has been associated with the onset of disease. Furthermore, alterations in microglia containing ferritin are associated with neurodegenerative conditions. To further understand the process of microglia dysfunction during the aging process, we used hippocampal sections from Tupaia belangeri (tree shrews). Adult (mean age 3.8 years), old (mean age 6 years), and aged (mean age 7.5 years) tree shrews were used for histochemical and immunostaining techniques to determine ferritin and Iba1 positive microglia, iron tissue content, tau hyperphosphorylation and oxidized‐RNA in dentate gyrus, subiculum, and CA1‐CA3 hippocampal regions. Our results indicated that aged tree shrews presented an increased number of activated microglia containing ferritin, but microglia labeled with Iba1 with a dystrophic phenotype was more abundant in aged individuals. With aging, oxidative damage to RNA (8OHG) increased significantly in all hippocampal regions, while tau hyperphosphorylation (AT100) was enhanced in DG, CA3, and SUB in aged animals. Phagocytic inclusions of 8OHG‐ and AT100‐damaged cells were observed in activated M2 microglia in old and aged animals. These data indicate that aged tree shrew may be a suitable model for translational research to study brain and microglia alterations during the aging process.

Caffeine elicits protective effects against liver diseases, such as NASH; however, its mechanism of action involving the pyrin domain-containing-3 (NLRP3) inflammasome signaling pathway remains to be elucidated. This study aimed to evaluate the effect of caffeine on the NLRP3 inflammasome signaling pathway in a rat model of NASH. NASH was induced by feeding rats a high-fat, -sucrose, and -cholesterol diet (HFSCD) for 15 weeks along with a weekly low dose (400 mg/kg, i.p.) of CCl4. Caffeine was administered at 50 mg/kg p.o. The effects of HFSCD+CCl4 and caffeine on the liver were evaluated using biochemical, ultrastructural, histological, and molecular biological approaches. The HFSCD+CCl4-treated rats showed fat accumulation in the liver, elevated levels of inflammatory mediators, NLRP3 inflammasome activation, antioxidant dysregulation, and liver fibrosis. Caffeine reduced necrosis, cholestasis, oxidative stress, and fibrosis. Caffeine exhibited anti-inflammatory effects by attenuating NLRP3 inflammasome activation. Moreover, caffeine prevented increases in toll-like receptor 4 (TLR4) and nuclear factor-κB (NF-κB) protein levels and mitigated the phosphorylation of mitogen-activated protein kinase (MAPK). Importantly, caffeine prevented the activation of hepatic stellate cells. This study is the first to report that caffeine ameliorates NASH by inhibiting NLRP3 inflammasome activation through the suppression of the TLR4/MAPK/NF-κB signaling pathway.