Impact of processing method on donated human breast milk microRNA content

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adult
  • Exosomes / genetics*
  • Female
  • Gene Expression Profiling
  • Humans
  • MicroRNAs / analysis*
  • MicroRNAs / genetics*
  • Milk, Human
  • Pasteurization / methods*
  • Pressure*
  • Specimen Handling / methods*
  • Young Adult


  • MicroRNAs

Grants and funding

This research was funded by the National Science Center in Poland ( PRELUDIUM grant number UMO 2016/21/N/NZ2/01726 (MAB) and the National Science Center in Poland POLONEZ grant number 2016/23/P/NZ2/04251 (PS). This project has received funding from the European Union’s Horizon 2020 ( research and innovation programme under the Marie Skłodowska-Curie grant agreement No 665778 (PS). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.