Characterization of the mammalian miRNA turnover landscape

Steady state cellular microRNA (miRNA) levels represent the balance between miRNA biogenesis and turnover. The kinetics and sequence determinants of mammalian miRNA turnover during and after miRNA maturation are not fully understood. Through a large-scale study on mammalian miRNA turnover, we report the co-existence of multiple cellular miRNA pools with distinct turnover kinetics and biogenesis properties and reveal previously unrecognized sequence features for fast turnover miRNAs. We measured miRNA turnover rates in eight mammalian cell types with a combination of expression profiling and deep sequencing. While most miRNAs are stable, a subset of miRNAs, mostly miRNA*s, turnovers quickly, many of which display a two-step turnover kinetics. Moreover, different sequence isoforms of the same miRNA can possess vastly different turnover rates. Fast turnover miRNA isoforms are enriched for 5′ nucleotide bias against Argonaute-(AGO)-loading, but also additional 3′ and central sequence features. Modeling based on two fast turnover miRNA*s miR-222-5p and miR-125b-1-3p, we unexpectedly found that while both miRNA*s are associated with AGO, they strongly differ in HSP90 association and sensitivity to HSP90 inhibition. Our data characterize the landscape of genome-wide miRNA turnover in cultured mammalian cells and reveal differential HSP90 requirements for different miRNA*s. Our findings also implicate rules for designing stable small RNAs, such as siRNAs.     

Long-term treadmill exercise improves spatial memory of male APPswe/PS1dE9 mice by regulation of BDNF expression and microglia activation

Increasing evidence suggests that physical activity could delay or attenuate the symptoms of Alzheimer''s disease (AD). But the underlying mechanisms are still not fully understood. To investigate the effect of long-term treadmill exercise on the spatial memory of AD mice and the possible role of β-amyloid, brain-derived neurotrophic factor (BDNF) and microglia in the effect, male APPswe/PS1dE9 AD mice aged 4 months were subjected to treadmill exercise for 5 months with 6 sessions per week and gradually increased load. A Morris water maze was used to evaluate the spatial memory. Expression levels of β-amyloid, BDNF and Iba-1 (a microglia marker) in brain tissue were detected by immunohistochemistry. Sedentary AD mice and wildtype C57BL/6J mice served as controls. The results showed that 5-month treadmill exercise significantly decreased the escape latencies (P < 0.01 on the 4th day) and improved the spatial memory of the AD mice in the water maze test. Meanwhile, treadmill exercise significantly increased the number of BDNF-positive cells and decreased the ratios of activated microglia in both the cerebral cortex and the hippocampus. However, treadmill exercise did not significantly alleviate the accumulation of β-amyloid in either the cerebral cortex or the hippocampus of the AD mice (P > 0.05). The study suggested that long-term treadmill exercise could improve the spatial memory of the male APPswe/PS1dE9 AD mice. The increase in BDNF-positive cells and decrease in activated microglia might underpin the beneficial effect.     

Enantioselective Inhibition of Carprofen Towards UDP‐glucuronosyltransferase (UGT) 2B7

UDP‐glucuronosyltransferases (UGTs)‐catalyzed glucuronidation conjugation reaction plays an important role in the elimination of many important clinical drugs and endogenous substances. The present study aims to investigate the enantioselective inhibition of carprofen towards UGT isoforms. In vitro a recombinant UGT isoforms‐catalyzed 4‐methylumbelliferone (4‐MU) glucuronidation incubation mixture was used to screen the inhibition potential of (R)‐carprofen and (S)‐carprofen towards multiple UGT isoforms. The results showed that (S)‐carprofen exhibited stronger inhibition potential than (R)‐carprofen towards UGT2B7. However, no significant difference was observed for the inhibition of (R)‐carprofen and (S)‐carprofen towards other UGT isoforms. Furthermore, the inhibition kinetic behavior was compared for the inhibition of (S)‐carprofen and (R)‐carprofen towards UGT2B7. A Lineweaver–Burk plot showed that both (S)‐carprofen and (R)‐carprofen exhibited competitive inhibition towards UGT2B7‐catalyzed 4‐MU glucuronidation. The inhibition kinetic parameter (K_i) was calculated to be 7.0 μM and 31.1 μM for (S)‐carprofen and (R)‐carprofen, respectively. Based on the standard for drug–drug interaction, the threshold for (S)‐carprofen and (R)‐carprofen to induce a drug–drug interaction is 0.7 μM and 3.1 μM, respectively. In conclusion, enantioselective inhibition of carprofen towards UDP‐glucuronosyltransferase (UGT) 2B7 was demonstrated in the present study. Using the in vitro inhibition kinetic parameter, the concentration threshold of (S)‐carprofen and (R)‐carprofen to possibly induce the drug–drug interaction was obtained. Therefore, clinical monitoring of the plasma concentration of (S)‐carprofen is more important than (R)‐carprofen to avoid a possible drug–drug interaction between carprofen and the drugs mainly undergoing UGT2B7‐catalyzed metabolism. Chirality 27:189–193, 2015. © 2014 Wiley Periodicals, Inc.