Cell-free seminal mRNA (cfs-mRNA) and microRNA (cfs-miRNA) have been found in human ejaculate and reported as promising noninvasive biomarkers for disorders of male reproductive organs and forensic identification. However, seminal plasma is particularly challenging for RNA extraction due to its complicated composition and high content of protein, DNA, and polysaccharide. Here, we report a novel, simple, and reliable method for the isolation of cfs-mRNA and cfs-miRNA from human semen based on our previous findings of their physical nature. Seminal microvesicles (0.1-0.5 µm in diameter), which contain the majority of cfs-mRNA, were enriched by a microfilter. Protein complexes, which most cfs-miRNA is bound with, were enriched by an ultrafilter. Harvesting the complexes or microvesicles, in which RNAs exist, avoided the influence of other components in human semen, thus favoring RNA isolation and purification. This new method can efficiently isolate cfs-mRNA and cfs-miRNA separately based on their physical nature, with high RNA purity, and low DNA contamination. It may also be applied or modified to isolate cell-free RNAs in other fluids.

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Functional constipation (FC) is commonly treated with fruits whose efficacy remains unclear. We conducted a meta-analysis of fruit intervention for FC and provided evidence-based recommendations. We searched seven databases from inception to July 2022. All randomized and crossover studies on the effectiveness of fruits on FC were included. We conducted sensitivity and subgroup analysis. A total of 11 studies were included in this review. Four trials showed that kiwifruits have significantly increased stool frequency (MD = 0.26, 95% CI (0.22, 0.30), P < 0.0001, I2 = 0%) than palm date or orange juice in the fixed-effect meta-analysis. Three high-quality studies suggested that kiwifruits have a better effect than ficus carica paste on the symptom of the FC assessed by the Bristol stool scale in the fixed-effect meta-analysis [MD = 0.39, 95% CI (0.11, 0.66), P < 0.05, I2 = 27%]. Besides, five trials showed that fruits can increase the amount of Lactobacillus acidophilus [MD = 0.82, 95% CI (0.25, 1.39), P < 0.05, I2 = 52%], analyzed with the random-effect model. Subgroup meta-analysis based on the types of fruits suggested that fruits including pome fruit, citrus fruit, and berries have increased the effect of Bifidobacterium t more than the stone fruits in the random effect meta-analysis [MD = 0.51, 95% CI (0.23, 0.79), P < 0.05, I2 = 84%]. Totally, fruit intake may have potential symptom alleviation on the FC as evidence shows that they can affect stool consistency, stool frequency, and gut microbiota. Further large-scale studies are needed to gain more confident conclusions concerning the association between fruit intake and FC in the future.