The impact of dietary intake on Sox2 protein levels and adult neurogenesis in the aging zebrafish brain

Abstract

Aging in humans is described as the gradual decline in physiological functions due to alterations in complex biological processes. As the lifespan increases and the population shifts towards older age, health problems associated with age become a worldwide problem. Thus, understanding the molecular and cellular changes that contribute to the aging process is essential to ensure healthy aging. Nine hallmarks that describe these changes were identified, which are also observed in the aging brain, leading to structural and cognitive deficiencies. Stem cell exhaustion is one of these hallmarks that explains a loss of stem cell function due to brain aging. A transcription factor, Sox2 is one of the main responsible proteins ensuring the stem cell maintenance through regulation of self-renewal and differentiation. Age-related changes in neural stem cells (NSCs) mediated by Sox2 can lead to a decline in new neuron formation. Neurogenesis is proven to continue in the defined neurogenic niches in the mammalian brain throughout adulthood and is considered to have a crucial role in the healthy functioning of the brain during aging. The regulation of the NSCs in neurogenic niches is achieved through diverse intrinsic and extrinsic factors that are affected by age-related changes. Dietary interventions are thought to modulate these factors and improve the age-related decline. Previous studies show that dietary restriction through lowering the calorie intake improves age-related impairment on neurogenesis, while high-calorie intake has a negative effect. Zebrafish, a small teleost fish, is a highly suitable model organism for investigations on neurogenesis with respect to aging due to exhibiting gradual decline with age, similar to humans and having a high capacity of neurogenesis in a widespread area. Hence, in the first part of the study, the influence of diet on age-associated changes in the brain was observed by following two short-term opposing dietary interventions. Using zebrafish as a model organism, dietary restriction (DR), over-feeding (OF), and ad libitum (AL) diets were included. These interventions demonstrated that short-term DR, compared to the AL, downregulated Sox2, as evidenced by western blot analysis. This might indicate a decreased self-renewal properties and activation of differentiation of NSCs. Short-term OF, on the other hand, did not change its expression levels. Moreover, aging did not alter the Sox2 expression levels. Further correlational and multivariate analyses were performed combining Sox2 with a proliferation marker proliferating cell nuclear antigen (PCNA) and neuronal lineage markers doublecortin-like kinase 1a (DCAMKL1) and ELAV-like neuron-specific RNA binding protein 3 (HuC). The analysis demonstrated a positive relationship between Sox2 and PCNA indicating their similar pattern in the aging process. Also, DCAMKL1 was shown to have a positive relationship with PCNA and a negative relationship with HuC. The multivariate analyses of all datasets exhibited age-specific effects of OF and DR by decreasing the survival of new neurons in old animals. Also, OF reinforced the commitment to neuronal fate with old age. The second part demonstrated a recombinant protein purification approach using immobilized-metal affinity chromatography to purify the His-tagged Sox2 protein. The recombinant Sox2 protein was successfully purified, and concentration values were measured. Taken together, these findings show that although older age led to changing dynamics in the neuronal lineage in the zebrafish brain, Sox2 managed to show a persistent expression. Furthermore, the short-term DR was shown to change the Sox2 expression levels, indicating the importance of calorie intake in alterations in neural stem cell properties. In conclusion, this study makes contributions to understanding cellular and biochemical changes occurring in the neurogenic niches of the aging brain and the altered neurogenic capacity due to dietary interventions.