Department of Biostatistics and Translational Medicine, Medical University of Lodz, Lodz, Poland.
Department of Hypertension, Medical University of Lodz, Lodz, Poland.
PLoS One. 2020 Jul 15;15(7):e0236126. doi: 10.1371/journal.pone.0236126. eCollection 2020.
Pasteurization of donated human milk preserves it for storage and makes it safe for feeding, but at the expense of its composition, nutritional values and functions. Here, we aimed to investigate the impact of Holder Pasteurization (HoP) and High Pressure Processing (HPP) methods on miRNA in human milk and to evaluate impact of these changes on miRNA functions. Milk samples obtained from women in 50th day of lactation (n = 3) were subjected either to HoP, HPP or remained unpasteurized as a control. Subsequently, miRNA was isolated from whole material and exosomal fraction and sequenced with Illumina NextSeq 500. Sequencing data were processed, read counts were mapped to miRNA and analyzed both quantitatively with DESeq2 and functionally with DIANA mirPath v.3. While HPP caused statistically insignificant decrease in number of miRNA reads compared to unprocessed material, HoP led to 82-fold decrease in whole material (p = 0.0288) and 302-fold decrease in exosomes (p = 0.0021) not leaving enough reads for further analysis. Changes in composition of miRNA fraction before and after HPP indicated uneven stability of individual miRNAs under high pressure conditions, with miR-30d-5p identified as relatively stable and miR-29 family as sensitive to HPP. Interestingly, about 2/3 of unprocessed milk miRNA content consists of only 10 distinct miRNAs with miR-148a-3p at the top. Functional analysis of most abundant human milk miRNAs showed their involvement in signaling pathways, cell communication, proliferation and metabolism that are obviously important in rapidly growing infants. Functions of miRNAs which suffered the greatest depletion during HPP were similar to roles of the majority of unprocessed human milk's miRNA, which indicates that those functions may be weakened although not completely lost. Our findings indicate that HPP is less detrimental to human milk miRNAs than HoP and should be considered in further research on recommended processing procedures for human milk banks.
巴氏消毒法和高压处理（HPP）都会使捐赠母乳在储存过程中得到保存，并使其变得安全，但却会损害母乳的成分、营养价值和功能。在这里，我们旨在研究 Holder 巴氏消毒法（HoP）和高压处理（HPP）方法对母乳中 miRNA 的影响，并评估这些变化对 miRNA 功能的影响。从哺乳期第 50 天的女性中获得母乳样本（n = 3），分别进行 HoP、HPP 或不巴氏消毒作为对照。随后，从全物质和外泌体部分中分离 miRNA 并进行 Illumina NextSeq 500 测序。对测序数据进行处理，将读取计数映射到 miRNA 上，并使用 DESeq2 进行定量分析和 DIANA mirPath v.3 进行功能分析。与未经处理的材料相比，HPP 导致 miRNA 读数数量的统计学上无显著减少，但 HoP 导致全物质中 miRNA 读数减少 82 倍（p = 0.0288），外泌体中减少 302 倍（p = 0.0021），没有足够的读数进行进一步分析。HPP 前后 miRNA 分数组成的变化表明，在高压条件下，个别 miRNA 的稳定性不均匀，miR-30d-5p 被确定为相对稳定，miR-29 家族对 HPP 敏感。有趣的是，未经处理的母乳 miRNA 含量约有 2/3 仅由 10 种独特的 miRNA 组成，其中 miR-148a-3p 含量最高。最丰富的人乳 miRNA 的功能分析表明，它们参与了信号通路、细胞通讯、增殖和代谢，这些在快速生长的婴儿中显然是非常重要的。在 HPP 过程中损失最大的 miRNA 的功能与未经处理的人乳 miRNA 的大多数功能相似，这表明这些功能可能会减弱，尽管不会完全丧失。我们的研究结果表明，HPP 对人乳 miRNA 的影响小于 HoP，并且应该在人乳库推荐的处理程序的进一步研究中考虑。
