Age-related loss of hepatic Nrf2 protein homeostasis: Potential role for heightened expression of miR-146a.

Nrf2 regulates the expression of numerous anti-oxidant, anti-inflammatory, and metabolic genes. We observed that, paradoxically, Nrf2 protein levels decline in the livers of aged rats despite the inflammatory environment evident in that organ. To examine the cause(s) of this loss, we investigated the age-related changes in Nrf2 protein homeostasis and activation in cultured hepatocytes from young (4-6 months) and old (24-28 months) Fischer 344 rats. While no age-dependent change in Nrf2 mRNA levels was observed (p>0.05), Nrf2 protein content, and the basal and anetholetrithione (A3T)-induced expression of Nrf2-dependent genes were attenuated with age. Conversely, overexpression of Nrf2 in cells from old animals reinstated gene induction. Treatment with A3T, along with bortezomib to inhibit degradation of existing protein, caused Nrf2 to accumulate significantly in cells from young animals (p<0.05), but not old, indicating a lack of new Nrf2 synthesis. We hypothesized that the loss of Nrf2 protein synthesis with age may partly stem from an age-related increase in microRNA inhibition of Nrf2 translation. Microarray analysis revealed that six microRNAs significantly increase >2-fold with age (p<0.05). One of these, miRNA-146a, is predicted to bind Nrf2 mRNA. Transfection of hepatocytes from young rats with a miRNA-146a mimic caused a 55% attenuation of Nrf2 translation that paralleled the age-related loss of Nrf2. Overall, these results provide novel insights for the age-related decline in Nrf2 and identify new targets to maintain Nrf2-dependent detoxification with age.