Derepression of specific miRNA-target genes in rice using CRISPR/Cas9.

MicroRNAs (miRNAs) target specific mRNA molecules based on sequence complementarity for their degradation or repression of translation, thereby regulating various developmental and physiological processes in eukaryotic organisms. Expressing the target mimicry (MIM) and short tandem target mimicry (STTM) can block endogenous activity of mature miRNAs and eliminate the inhibition of their target genes, resulting in phenotypic changes due to higher expression of the target genes. Here, we report a strategy to achieve derepression of interested miRNA-target genes through CRISPR/Cas9-based generation of in-frame mutants within the miRNA-complementary sequence of the target gene. We show that two rice genes, OsGRF4 (GROWTH REGULATING FACTOR 4) and OsGRF8 carrying in-frame mutants with disruption of the miR396 recognition sites, escape from miR396-mediated post-transcriptional silencing, resulting in enlarged grain size and increase in brown planthopper (BPH) resistance, in their respective transgenic rice lines. These results demonstrate that CRISPR/Cas9-mediated disruption of miRNA target sites can be effectively employed to precisely derepress particular target genes of functional importance for trait improvement in plants.

APICAL SPIKELET ABORTION (ASA) Controls Apical Panicle Development in Rice by Regulating Salicylic Acid Biosynthesis.

Panicle degradation causes severe yield reduction in rice. There are two main types of panicle degradation: apical spikelet abortion and basal degeneration. In this study, we isolated and characterized the apical panicle abortion mutant apical spikelet abortion (asa), which exhibits degeneration and defects in the apical spikelets. This mutant had a pleiotropic phenotype, characterized by reduced plant height, increased tiller number, and decreased pollen fertility. Map-based cloning revealed that OsASA encodes a boric acid channel protein that showed the highest expression in the inflorescence, peduncle, and anther. RNA-seq analysis of the asa mutant vs wild-type (WT) plants revealed that biological processes related to reactive oxygen species (ROS) homeostasis and salicylic acid (SA) metabolism were significantly affected. Furthermore, the asa mutants had an increased SA level and H2O2 accumulation in the young panicles compared to the WT plants. Moreover, the SA level and the expression of OsPAL3, OsPAL4, and OsPAL6 genes (related to SA biosynthesis) were significantly increased under boron-deficient conditions in the asa mutant and in OsASA-knockout plants. Collectively, these results suggest that the boron distribution maintained by OsASA is required for normal panicle development in a process that involves modulating ROS homeostasis and SA biosynthesis.

Quantitative study on the efficacy of acupuncture in the treatment of menopausal hot flashes and its comparison with nonhormonal drugs.

OBJECTIVE: This study aimed to compare the efficacy of acupuncture to that of sham acupuncture, placebo pills, and nonhormonal drugs to provide the necessary quantitative information for establishing medication guidelines for menopausal hot flashes. METHODS: A comprehensive literature search was performed using public databases. Randomized clinical studies on acupuncture therapy for the treatment of hot flashes in menopausal women were identified. A time-course model was established to describe the efficacy characteristics of acupuncture and sham acupuncture, which were compared with the efficacy of nonhormonal drugs and placebo pills reported in the literature. RESULTS: A total of 17 studies involving 1,123 participants were included. The quality of all the studies included in the analysis is medium to high, and there was no obvious risk of bias. It was found that the baseline number of hot flashes was an important factor affecting the efficacy of acupuncture and sham acupuncture. After correcting the baseline to eight hot flashes per day, the frequency of hot flashes decreased from baseline for traditional acupuncture (TA), electro-acupuncture (EA), TA&EA (merger analysis of TA and electro-acupuncture), and sham acupuncture were 3.1 (95% confidence interval [CI]: 2.8-3.4), 3.6 (95% CI: 3.2-4.0), 3.2 (95% CI: 2.9-3.5), and 2.6 (95% CI: 2.2-3.0) times/d at week 8, respectively. Compared with findings reported in the literature, we found the efficacy of electro-acupuncture was comparable to that of selective serotonin reuptake inhibitors/serotonin-norepinephrine reuptake inhibitors and neuroleptic agents such as gabapentin and escitalopram. Furthermore, the efficacy of TA&EA (merged) was significantly higher than that of placebo pills (2.3, 95% CI: 1.8-2.9). CONCLUSIONS: The efficacy of TA&EA (merged) was higher than that of sham acupuncture and significantly higher than that of placebo pills. The efficacy of electro-acupuncture was higher than that of traditional acupuncture, significantly higher than that of sham acupuncture, and comparable to that of selective serotonin reuptake inhibitors/serotonin-norepinephrine reuptake inhibitors and neuroleptic agents.

Pharmacological interventions for the treatment of insomnia: quantitative comparison of drug efficacy.

PURPOSE: Although different forms of pharmacological intervention are often prescribed for insomnia disorder, the comparative efficacies among various drugs remain unclear. We therefore conducted this study to quantitatively compare the efficacy of various pharmacotherapies for insomnia by modeling. METHODS: We searched PubMed, EMBASE and Cochrane Library databases for randomized placebo-controlled trials of insomnia medications that were conducted within a designated time period (from the inception dates to May 16, 2019). Pharmacodynamic models were established to describe the time course of changes from baseline in selected sleep parameters. Sleep quality and dropout rates were also compared by a single-arm meta-analysis. RESULTS: In sum, 43 studies covering 44 trials (14,535 patients) were included in the analysis. The drugs evaluated included flurazepam, quazepam, temazepam, triazolam, eszopiclone, zaleplon, zolpidem, extended-release zolpidem, suvorexant, ramelteon and doxepin. The established models revealed eszopiclone had the highest efficacy in terms of sleep latency (SL), total sleep time (TST), and sleep quality, and was also associated with the lowest dropout rates. The effect of suvorexant on the parameter 'wake after sleep onset' (WASO) was significantly higher than that of the other drugs analyzed. CONCLUSIONS: Each drug has its own characteristics in the treatment of insomnia, and this needs to be taken into consideration to meet individual clinical needs. These results serve as a quantitative supplement for clinical practice by reflecting the difference in efficacy of various drugs in the treatment of insomnia.

Testing whether the progression of Alzheimer's disease changes with the year of publication, additional design, and geographical area: a modeling analysis of literature aggregate data.

BACKGROUND: Our objectives were to develop a disease progression model for cognitive decline in Alzheimer's disease (AD) and to determine whether disease progression of AD is related to the year of publication, add-on trial design, and geographical regions. METHODS: Placebo-controlled randomized AD clinical trials were systemically searched in public databases. Longitudinal placebo response (mean change from baseline in the cognitive subscale of the Alzheimer's Disease Assessment Scale [ADAS-cog]) and the corresponding demographic information were extracted to establish a disease progression model. Covariate screening and subgroup analyses were performed to identify potential factors affecting the disease progression rate. RESULTS: A total of 134 publications (140 trials) were included in this model-based meta-analysis. The typical disease progression rate was 5.82 points per year. The baseline ADAS-cog score was included in the final model using an inverse U-type function. Age was found to be negatively correlated with disease progression rate. After correcting the baseline ADAS-cog score and the age effect, no significant difference in the disease progression rate was found between trials published before and after 2008 and between trials using an add-on design and those that did not use an add-on design. However, a significant difference was found among different trial regions. Trials in East Asian countries showed the slowest decline rate and the largest placebo effect. CONCLUSIONS: Our model successfully quantified AD disease progression by integrating baseline ADAS-cog score and age as important predictors. These factors and geographic location should be considered when optimizing future trial designs and conducting indirect comparisons of clinical outcomes.

Quantitative analysis of the placebo response in pharmacotherapy of insomnia and its application in clinical trials.

STUDY OBJECTIVES: This study aimed to develop a robust placebo response model for the pharmacotherapy for insomnia to guide drug development and clinical practice. METHODS: PubMed, EMBASE, and Cochrane Library databases were systematically searched for randomized placebo-controlled trials of medications for insomnia dating from the inception dates of the databases to April 18, 2018. Three placebo response models were established to describe the time-course of sleep parameters measured by objective (polysomnography or actigraphy) or subjective methods (sleep diary or questionnaires). The established models were applied to simulate placebo response distribution under different conditions using Monte Carlo simulations. RESULTS: Fifty-four studies involving 6,416 subjects were included. Placebo response increased over time and reached a plateau at approximately 8 weeks from start of therapy. Established models described the observed data reasonably well based on various diagnostic plots. Baseline sleep parameters affected the placebo response. There were significant positive correlations with placebo response and the severity of sleep latency, wake after sleep onset, and total sleep time at baseline. In addition, placebo response, assessed by subjective and objective methods, was consistent after correcting the baseline levels. CONCLUSIONS: The established placebo response models can serve as a tool to predict placebo response at different baseline levels, which can provide valuable reference for clinical trial design, decision-making in drug development, and clinical practice.