Preliminary results and a theoretical perspective of co‑treatment using a miR‑93‑5p mimic and aged garlic extract to inhibit the expression of the pro‑inflammatory interleukin‑8 gene

The coronavirus disease-19 (COVID-19) pandemic has been a very significant health issue in the period between 2020 and 2023, forcing research to characterize severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sequences and to develop novel therapeutic approaches. Interleukin-6 (IL-6) and IL-8 are considered significant therapeutic targets for COVID-19 and emerging evidence has suggested that microRNAs (miRNAs/miRs) serve a key role in regulating these genes. MiRNAs are short, 19-25 nucleotides in length, non-coding RNAs that regulate gene expression at the post-transcriptional level through the sequence-selective recognition of the 3'-untranslated region (3'-UTR) of the regulated mRNAs, eventually repressing translation, commonly, via mRNA degradation. For example, among several miRNAs involved in the regulation of the COVID-19 'cytokine storm', miR-93-5p can inhibit IL-8 gene expression by directly targeting the 3'-UTR of IL-8 mRNA. In addition, miR-93-5p can regulate Toll-like receptor-4 (TLR4) and interleukin-1 receptor-associated kinase 4 (IRAK4) expression, thus affecting the nuclear factor-kappa B (NF-kappa B) pathway and the expression of NF-kappa B-regulated genes, such as IL-6, IL-1 beta and other hyper-expressed genes during the COVID-19 'cytokine storm'. In the present study, the results provided preliminary evidence suggesting that the miR-93-5p-based miRNA therapeutics could be combined with the anti-inflammatory aged garlic extract (AGE) to more effectively inhibit IL-8 gene expression. The human bronchial epithelial IB3-1 cell line was employed as experimental model system. IB3-1 cells were stimulated with the BNT162b2 COVID-19 vaccine and transfected with pre-miR-93-5p in the absence or in the presence of AGE, to verify the inhibitory effects on the BNT162b2-induced expression of the IL-8 gene. The accumulation of IL-8 mRNA was assessed by RT-qPCR; the release of IL-8 protein was determined by Bio-Plex assay. In addition, the possible applications of TLR4/NF-kappa B inhibitory agents (such as miR-93-5p and AGE) for treating human pathologies at a hyperinflammatory state, such as COVID-19, cystic fibrosis and other respiratory diseases, were summarized.