Thesis title: Plant microRNAs: a natural anti-inflammatory activity in a cross-kingdom regulation
Medicinal plants have been used for millennia to treat a wide range of diseases, particularly those with an inflammatory nature. Plants are rich in bioactive compounds, which exhibit antimicrobial, anti-tumor, antioxidant, and anti-inflammatory properties. However, these molecules often reduce the activity of key players in inflammatory and tumor pathways, indirectly. Plants are also a source of microRNAs, which are naturally methylated at the 3’-end. In animals, microRNAs can control a wide range of biological processes including inflammation, by acting as direct down regulators of genes involved in inflammation. So we asked if plant-derived-miRNAs could be a potential anti-inflammatory molecules, as well as in animals.
To explore this possibility, we aimed to identify plant miRNAs with potential anti-inflammatory activity, since it has been reported that plant-derived miRNAs could bind and regulate the expression of human mRNA, in a cross-kingdom regulation.
Through a bioinformatics approach, we identified two plant miRNAs (miR-A and miR-B) with putative anti-inflammatory activity. They share the same seed sequence of human anti-inflammatory miR-146a-5p, which targets IRAK1 and TRAF6, key molecules of the NF-kB inflammatory pathway. In order to investigate the regulation of plant-miRs on the miR-146a target genes, we overexpressed them in HEK-293 cells. Our results indicated that both plant miR-A and miR-B led to a significant reduction in mRNA and protein levels of IRAK1 and TRAF6. Moreover, the luciferase assay demonstrated that the two plant miRNAs effectively reduced luciferase activity by targeting the 3’-UTRs of IRAK1 and TRAF6. Subsequently, to assess their anti-inflammatory activity, mimic plant-miR-A/B were transfected in two inflammatory cellular systems, primary human macrophages (MDM) and mouse BV-2 cell lines, stimulated with lipopolysaccharide (LPS). In both models, the mimics significantly reduced the levels of pro-inflammatory cytokines (TNF-α and IL-6), underscoring their ability to inhibit the NF-κB inflammatory pathway similarly to human miR-146a. However, further studies are necessary to explore both the absorption mechanisms and the anti-inflammatory activity of the two naturally methylated plant-derived miRNAs, both in vitro and in vivo. To this aim, we are developing enhanced lines of Arabidopsis thaliana and Cardamine hirsuta implemented to produce high levels of plant-derived miR-A and miR-B, as molecular farming. These miRNAs will be extracted and utilized to assess their anti-inflammatory activity through in vitro and in vivo assays.
In conclusion, these results collectively underscore both the potential of plant miRNAs in regulating animal inflammatory genes of the NF-kB pathway and the possibility to use them as candidates for the development of new anti-inflammatory therapies.
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