MicroRNAs (miRNAs) are a class of small RNA molecules that regulate gene expression by inhibiting the protein translation or targeting the mRNA cleavage. They play many important roles in living organism cells; however, the knowledge on miRNAs functions has become more extensive upon their identification in biological fluids and recent reports on plant-origin miRNAs abundance in human plasma and serum. Considering these findings, we performed a rigorous bioinformatics analysis of publicly available, raw data from high-throughput sequencing studies on miRNAs composition in human and porcine breast milk exosomes to identify the fraction of food-derived miRNAs. Several processing and filtering steps were applied to increase the accuracy, and to avoid false positives. Through aforementioned analysis, 35 and 17 miRNA species, belonging to 25 and 11 MIR families, were identified, respectively. In the human samples the highest abundance levels yielded the ath-miR166a, pab-miR951, ptc-miR472a and bdi-miR168, while in the porcine breast milk exosomes, the zma-miR168a, zma-miR156a and ath-miR166a have been identified in the largest amounts. The consensus prediction and annotation of potential human targets for select plant miRNAs suggest that the aforementioned molecules may interact with mRNAs coding several transcription factors, protein receptors, transporters and immune-related proteins, thus potentially influencing human organism. Taken together, the presented analysis shows proof of abundant plant miRNAs in mammal breast milk exosomes, pointing at the same time to the new possibilities arising from this discovery.

LPS is a constituent of cell walls of Gram-negative bacteria that, acting through the CD14/TLR4 receptor complex, causes strong proinflammatory activation of macrophages. In murine peritoneal macrophages and J774 cells, LPS at 1–2 ng/ml induced maximal TNF-α and MIP-2 release, and higher LPS concentrations were less effective, which suggested a negative control of LPS action. While studying the mechanism of this negative regulation, we found that in J774 cells, LPS activated both acid sphingomyelinase and neutral sphingomyelinase and moderately elevated ceramide, ceramide 1-phosphate, and sphingosine levels. Lowering of the acid sphingomyelinase and neutral sphingomyelinase activities using inhibitors or gene silencing upregulated TNF-α and MIP-2 production in J774 cells and macrophages. Accordingly, treatment of those cells with exogenous C8-ceramide diminished TNF-α and MIP-2 production after LPS stimulation. Exposure of J774 cells to bacterial sphingomyelinase or interference with ceramide hydrolysis using inhibitors of ceramidases also lowered the LPS-induced TNF-α production. The latter result indicates that ceramide rather than sphingosine suppresses TNF-α and MIP-2 production. Of these two cytokines, only TNF-α was negatively regulated by ceramide 1-phosphate as was indicated by upregulated TNF-α production after silencing of ceramide kinase gene expression. None of the above treatments diminished NO or RANTES production induced by LPS. Together the data indicate that ceramide negatively regulates production of TNF-α and MIP-2 in response to LPS with the former being sensitive to ceramide 1-phosphate as well. We hypothesize that the ceramide-mediated anti-inflammatory pathway may play a role in preventing endotoxic shock and in limiting inflammation.

MicroRNAs (miRNAs) represent a class of small non-coding RNAs that act as efficient gene expression regulators and thus play many important roles in living organisms. Due to their involvement in several known human pathological and pathogenic states, miRNA molecules have become an important issue in medicine and gained the attention of scientists from the pharmaceutical industry. In recent few years, a growing number of studies have provided evidence that miRNAs may be transferred from one species to another and regulate gene expression in the recipients’ cells. The most intriguing results revealed that stable miRNAs derived from food plants may enter the mammals’ circulatory system and, after reaching the target, inhibit the production of specific mammalian protein. Part of the scientific community has perceived this as an attractive hypothesis that may provide a foundation for novel therapeutic approaches. In turn, others are convinced about the “false positive” effect of performed experiments from which the mentioned results were achieved. In this article, we review the recent literature that provides evidence (from both fronts) of dietary, plant miRNA uptake and functionality in various consumers. Additionally, we discuss possible miRNA transport mechanisms from plant food sources to human cells.

Breast milk is a natural food and important component of infant nutrition. Apart from the alimentary substances, breast milk contains many important bioactive compounds, including endogenous microRNA molecules (miRNAs). These regulatory molecules were identified in various mammalian biological fluids and were shown to be mostly packed in exosomes. Recently, it was revealed that plant food-derived miRNAs are stably present in human blood and regulate the expression of specific human genes. Since then, the scientific community has focused its efforts on contradicting or confirming this discovery. With the same intention, qRT-PCR experiments were performed to evaluate the presence of five plant food-derived miRNAs (miR166a, miR156a, miR157a, miR172a and miR168a) in breast milk (whole milk and exosomes) from healthy volunteers. In whole milk samples, all examined miRNAs were identified, while only two of these miRNAs were confirmed to be present in exosomes. The plant miRNA concentration in the samples ranged from 4 to 700 fM. Complementary bioinformatics analysis suggests that the evaluated plant miRNAs may potentially influence several crucial biological pathways in the infant organism.

Enhancer of rudimentary homolog (Drosophila) (ERH) is a small (approximately 100 residues) eukaryotic protein whose gene has been found in animals, plants and protists, but not in fungi [1][2][3][4]. Although its sequence is highly conserved (for example, ERHs from humans and Xenopus laevis are identical), it is not significantly similar to any other protein, and it has not been possible to ascribe any possible function to this protein on the basis of sequence analysis alone. The three-dimensional structure of ERH is an equally unique combination of three amphipathic a-helices and a four-stranded antiparallel b-sheet [5][6][7][8][9]. The human ERH transcript is expressed ubiquitously in both adult and fetal tissues, and the protein localizes predominantly to the nucleus [10] Enhancer of rudimentary homolog (Drosophila) (ERH) is a small, highly conserved, nuclear protein with a unique three-dimensional structure, whose gene has been identified in animals, plants and protists, but not in fungi. Involvement of ERH in fundamental processes such as regulation of pyrimidine metabolism, cell cycle progression, transcription and cell growth control has been suggested. Here, employing a yeast two-hybrid system, a glutathione S-transferase pull-down assay and tandem MS, we demonstrate that Ciz1 is a bona fide interactor of human ERH. Ciz1 is a nuclear zinc finger protein interacting with p21Cip1/Waf1 , a universal inhibitor of cyclindependent kinases, and is a DNA replication factor. The region of Ciz1 necessary for the interaction with ERH spans residues 531-644, encompassing its first zinc finger motif. This region overlaps the p21 Cip1/Waf1 -binding site, suggesting that the interaction with ERH could block the binding of p21Cip1/Waf1 by Ciz1 in the cell. When ERH and Ciz1 are coexpressed in HeLa cells, Ciz1 recruits ERH to DNA replication foci.Abbreviations CDK2, cyclin E-cyclin-dependent kinase 2; Ciz1, p21Cip1/Waf1 -interacting zinc finger protein 1; CK2, casein kinase II; DCL1, dynein light chain 1; DCoH/PCD, dimerization cofactor of hepatocyte nuclear factor 1/pterin-4a-carbinolamine dehydratase; EGFP, enhanced green fluorescent protein; ERH, enhancer of rudimentary homolog (Drosophila); ERa, estrogen receptor a; GST, glutathione S-transferase; HNF1, hepatocyte nuclear factor 1; MH3, matrin 3-homologous domain 3; PDIP46/SKAR, polymerase d-interacting protein 46/p70 ribosomal protein S6 kinase 1 Aly/REF-like target.