The Impact of Dietary Compounds in Functional Foods on MicroRNAs Expression

MicroRNAs (miRNAs) are a class of non-coding endogenous RNA molecules that are involved in post-transcriptional gene silencing via binding to their target messenger RNA, leading to mRNA degradation or translational repression. MicroRNAs can be modulated by several factors including hormones, transcription factors, and dietary compounds. These biologically active compounds have positive impact on the progression of human pathology including non-communicable diseases, which indicating that administration of diet may have potential as therapeutic agents in modulating the risk of chronic diseases. Interestingly, evidence emerging in recent years suggests that dietary miRNAs can be absorbed in human circulation, modulated human gene expression and biological functions. The exploitation of the miRNA functioning within different origins, cellular miRNAs and dietary miRNAs will help us to understand the molecular machinery as well as the regulatory mechanisms involved in fundamentally important biological processes. Therefore, this knowledge may be applied of natural bioactive compounds in preventive or therapeutic approaches.


Introduction -Origin, biogenesis and functions of microRNAs
MicroRNAs or miRNAs are a class of small non-coding RNA approximately 21-25 nucleotides that modulate on gene expression post-transcriptionally via binding to the 3′ untranslated region (3′-UTR) of the target messenger RNA (mRNA), resulting in mRNA degradation or translational repression. The first miRNA, lin-4, was discovered by Ambro and his research group in 1993 and it was found to be related with larva development in Caenorhabditis elegans [1]. Up to date, almost 2000 miRNAs have been identified in humans (http://www.miRbase.org -7.3.2019) [1]. It has been estimated that 1-4% of human genes expression can be regulated by miRNAs, which is the largest of genomic regulator [2]. In mammals, miRNAs have been associated with various cellular pathways with the regulation of cell differentiation, cell cycle, proliferation, apoptosis, hematopoiesis, and other cellular functions. Recent studies have highlighted the importance of mRNA regulation mechanism by validation and differential miRNA expression in a variety of human pathological conditions, including chronic diseases. miRNAs are normally transcribed by RNA polymerase II from miRNA genes. This transcription leads to generate a primary miRNA transcript (pri-miRNA). Then, pri-miRNA is further cleaved by a microprocessor complex, which consists of Drosha, the double-stranded RNase III enzyme and DiGeorge syndrome critical region 8 (DGCR8), important cofactor, into a hairpin structure precursor miRNA (pre-miRNA) in the nucleus (Figure 1). The double strand pre-miRNAs with 70 nucleotides are then exported to the cytoplasm by the process of nuclear export factor exportin-5. The pre-miRNA is then processed by RNase III, Dicer, thereby generating a mature miRNA:miRNA duplex approximately 22 nucleotides in length and without a hairpin structure. The helicase enzyme cleaves miRNA duplexes into single-stranded miRNAs and incorporated into the Argonaute (AGO), TRBP and PACT proteins to form the RNA-induced silencing complex (RISC). Usually, other single strand called passenger strand or the star (*) strand will be degraded, while single strand mature miRNA is able to bind with its target mRNA and mediating translational inhibition or mRNA degradation, along with their sequence complementarity to the target [1,3]. In plants, target mRNA will be degraded if miRNA has perfect or near-perfect complementarity to its target. In contrast to mammal, miRNAs bind to partially complementary sites in the 3′-UTRs of target mRNA, which leading to translational repression [4]. the target mRNA is either blocked (imperfect complementary) or degraded (perfect complementary) of the ribosomal translation, which sequentially impacts the cellular functions.
Phytochemicals are major plant-derived compounds that naturally found in vegetables, fruits, medicinal plants or other plants with medicinal properties including antioxidant, anti-diabetic, anti-inflammatory, antimicrobial, antidepressant, anticancer and prevention in other chronic non-communicable diseases [5][6][7]. Phenolic and flavonoid compounds are the most important group of bioactive compounds and second metabolites in plants which comprise of essential molecules Figure 1. miRNA biogenesis. miRNA gene is transcribed by RNA polymerase II and then forming the primary miRNA transcript (pri-miRNA), which is further cleaved by the Drosha/DGCR8 complex to generate the precursor miRNA (pre-miRNA). Pre-miRNA is then exported into the cytoplasm by exportin 5/RAN-GTP and further processed by dicer to create the mature miRNA, which is loaded into RISC, which contains AGO, PACT and TRBP proteins. Mature miRNA that binding to its target mRNA by perfect complementary binding and resulting in gene suppression by mRNA degradation. The partially complementary binding of miRNA and its target mRNA, which in turn inhibit the protein translation.

Dietary compounds and endogenous miRNAs
Extensive studies have been performed to understand the molecular mechanism of bioactive compounds with a positive effect on chronic diseases or noncommunicable diseases such as arthritis, cancer, cardiovascular diseases, diabetes and obesity [1,16]. Emerging evidences confirm that alteration of endogenous miRNAs expression can be influenced by bioactive compounds in functional foods [16,17] (Figure 2 and Table 1).

Arctigenin
Arctigenin (AR) is a phenylpropanoid dizbenzylbutyrolactone lignin and was first identified in Arctium lappa L. Several studies showed anti-inflammatory, anti-cancer, anti-viral, immune modulatory activities of AR [24][25][26][27]. The study demonstrated that AR upregulated miR-16 and miR-199a expression by decreasing upstream protein (IKKα and IKKβ) expression and inhibiting NF-κB signaling pathway activity, thereby reducing inflammatory cytokines production in neural cells [28]. The combination treatment of AR and quercetin significantly inhibited the oncogenic miRNAs expression including miR-19b, miR-21 and miR-148a in prostate cancer cells. AR and quercetin also showed anti-migration activity in prostate cancer cells [29].
DIM showed the modulation of miRNAs expression in other inflammatory diseases. The expression of miR-106a, miR-20b, and miR-125b-5p were increased after treatment with DIM and suppressed the expression of IRAK4 and TNF-α to limit responses to TLRs activated by LPS in acute liver failure (ALF) animal model [52]. DIM significantly upregulated miR-200c, miR-146a, miR-16, miR-93, and miR-22 in brain CD4+ T cells and inhibited the expression of cyclin E1 and B-cell lymphoma-2 in experimental autoimmune encephalomyelitis (EAE), a murine model of multiple sclerosis [53].

(−)-Epigallocatechin-3-gallate
(−)-Epigallocatechin-3-Gallate or EGCG is a major polyphenol compound in green tea (Camellia sinensis) and derivative of catechin [3,16]. EGCG is a powerful antioxidant, anticancer and antiangiogenic properties, which has a potential role to influence human diseases [54][55][56]. EGCG suppressed nasopharyngeal carcinoma cell migration and invasion through a novel signaling axis of miR-296/STAT3 regulation [57]. Gold nanoparticles (AuNPs) have been used for drug delivery as their stability and increase drug bioavailability as well as accumulation of drug in cancer cells. EGCG-capped gold nanoparticles upregulated the tumor suppressor miRNAs, let-7a and miR34a, which consecutively their targeted gene, caspase-3 was upregulated, and c-Myc protein was decreased in hepatocellular carcinoma cells [58]. miR-34a is one of the tumor suppressor miRNAs that downregulated, whereas miR-93 is highly up-regulated in prostate cancer cells. Co-transfection of miR.34a mimic and miR.93 inhibitor along with EGCG significantly decreased androgen receptor (AR) and prostate-specific antigen (PSA) expression when compared to the co-transfection without EGCG [59]. In cervical carcinoma cells, Hela (HPV16/18+), EGCG inhibited cell growth and up-regulated miR-29 and miR-210 expression, while down-regulated the expression of miR-125b and miR-203 [60]. Up-regulation of let-7 was observed in EGCG treated melanoma cells, which led to inhibit the expression of high mobility group A2 (HMGA2) [61].
EGCG showed the protective effect against myocardial ischemia/reperfusion (I/R) injury through up-regulation of miR-384-mediated autophagy by targeting Beclin-1 via activating the PI3K/Akt signaling pathway [62]. EGCG also demonstrated the anti-arthritic effects by inhibited IL-1β-induced ADAMTS5 expression and up-regulated the expression of miR-140-3p in osteoarthritis chondrocytes [63]. EGCG treatment has potential role of preventing toxin-induced fibrosis by suppression of osteopontin expression and up-regulation of miR-10b, miR-181a and miR-221 in liver hepatocellular carcinoma cells [64].

Genistein
Genistein belongs to isoflavone family and presents in soybeans with antiangiogenic, anti-metastasis, anti-inflammatory, anti-oxidant, cell cycle arrest and induction of apoptosis effects [65]. Genistein can regulate the expression of miRNAs in several call types [65]. It has been reported that treatment of genistein up-regulated miR-23b and inhibited breast cancer cell growth [66]. Genistein also exhibited anti-tumor effect by down-regulated miR-1260b and targeting sRRP1 and Smad4 through DNA methylation or histone modifications in prostate cancer cells [67]. The same research group reported that miR-1260b was highly expressed in renal cancer cells and miR-1260b was down-regulated in genistein treated renal cancer cells [68]. The treatment of miR-1260b inhibitor inhibited the expression of its target genes, sFRP1, Dkk2, Smad4 [68]. Treatment with genistein induced non-small lung cancer cell apoptosis, caspase-3/9 activation and inhibited cell proliferation via up-regulation of miR-27a -mediated MET signaling [69]. Co-encapsulate miR-29b with genistein in hybrid nanoparticles (GMLHN) has been studied to treat effectively in non-small lung cancer cell and GMLHN showed the anti-proliferative effect by down-regulation of phosphorylated AKT (pAKT) and phosphorylated phosphoinositide-3 kinase (p-PI3K) [70].

Quercetin
Quercetin is bioactive flavonoids that can be found in fruits and vegetables including onion, kale, apple, many berries, citrus fruits and tea [73]. Anti-cancer, anti-inflammatory, antioxidant, anti-diabetes, anti-atherosclerosis and anti-viral effects have been reported in different in vitro studies for quercetin [74]. Several studies have focused on quercetin and miRNAs modulation for therapeutic approaches. miR-200b-3p was up-regulated in pancreatic cancer cells when treated with quercetin, resulting in inhibition of self-renewal and decrease of proliferation through Notch1 signaling pathway [75]. Quercetin significantly inhibited breast cancer cell proliferation and invasion via up-regulated miR-146a expression and targeting EGFR [76]. Quercetin inhibited cell viability, migration and invasion by up-regulated miR-16 and targeting HOXA10 in oral cancer cells [77]. In addition, quercetin decreased oral cancer cell viability and increased cell apoptosis via miR-22/WNT1/β-catenin pathway [78].

Silymarin
Silymarin is a flavonolignans extracted from the milk thistle Silybum marianum (L.) gaernt and recent studies have demonstrated the anti-cancer, anti-inflammatory, vascularization inhibitory, antioxidant, hepatoprotective, cardioprotective and antimetastasis activities of silymarin [83]. Several miRNAs have been implicated in the invasive potential of cancer cells. Tumor suppressor miRNA, miR-203, was up-regulated and class 1 HDAC proteins and ZEB1 were repressed with silymarin treatment, resulted in inhibition of non-small cell lung cancer migration [84]. Silibinin, the major active constituent of silymarin extract, induced apoptosis and ERβ expression, inhibited cell proliferation, and reduced pro-inflammatory cytokines expression including IL-17 and TNF-α, through ERβ binding and down-regulated miR-155 in rheumatoid arthritis [85]. miR-122 is liver-specific miRNA and was down-regulation upon silymarin treatment in rat model for hepatoprotective and radio protective effects via increased superoxide dismutase (SOD), glutathione (GSH) and reduced lipid peroxidation (MDA) [86]. It has been reported the hepatoprotective activity of silymarin on thioacetamide-induced liver damage by restored miR-122, miR-192, and miR-194 expression levels [87].

Sulforaphane
Sulforaphane is dietary compounds in broccoli (Brassica oleracea) and cruciferous plants. It has been demonstrated the capability of sulforaphane for anti-inflammatory, antiaging, antidiabetic, antioxidant, anti-tumor, hepatoprotective and cardioprotective effects [91]. Plant-derived phytochemicals including sulforaphane are potentially affected miRNAs expression. Sulforaphane inhibited breast cancer cell cycle arrest and senescence via down-regulation of miR-23b, miR-92b, miR-381 and miR-382 [92]. Anti-tumor effect of sulforaphane also reported in non-small cell lung cancer by down-regulation of miR-616-5p and targeting GSK3β/β-catenin signaling pathway [93]. Sulforaphane inhibited the progression of pancreatic cancer through down-regulated miR30a-3p with the increasing of its target, Cx43 expression and upregulated miR-135b-5p mediated RASAL2 expression [94,95]. In addition, sulforaphane treatment significantly increased the expression of tumor suppressor miRNA, miR-200c, resulted in inhibited the cancer stemness and tumorinitiating properties in oral squamous cell carcinomas and cancer stem cells both in vitro and in vivo [96]. Anti-proliferative and apoptotic effects of sulforaphane have been reported in gastric cancer cells, which leading to alter the expression of miR-9 and miR-326 [97]. Up-regulation of miR-124-3p and inhibition of its target, STAT3 by sulforaphane treatment were observed and thereby induced apoptosis, inhibited proliferation and decreased the stemness of nasopharyngeal cancer cell [98].

Dietary miRNA and human gene regulation
Several evidences demonstrated the direct modulation of cellular signaling pathways by dietary compounds could decrease the risk of chronic diseases [101]. Interestingly, it has been reported that small non-coding RNA including miRNAs can be transferred across Kingdoms, for example dietary miRNAs have been found in human body fluids and these circulating miRNAs are likely to regulate human gene  [15,[102][103][104][105][106][107]. The uptake of plant derived miRNAs could be in the form of raw and cooked plants in capable of stability forms [107,108]. Due to high temperature cooking process, low pH and enzymes in digestive tract as well as enzymes in blood circulation, miRNAs might be destroyed before their functions with target mRNAs [15]. Strikingly, GC base content, 2'-O-methylation on the 3′-terminal, unique nucleotide sequence of dietary miRNAs and extracellular vesicles (exosome and microvesicle) are preventive features of plant derived miRNAs in harmful conditions [109][110][111][112][113][114].
There are numerous studies to support the functional roles of dietary miRNAs in cross kingdom gene regulation. Rice miR156a and miR168a were detected in human serum and miR168a down-regulated low-density lipoprotein receptor adapter protein 1 (LDLRAP1) expression, resulted in an increase of plasma LDL cholesterol level, Table 2 [105]. miR2910 from Populus euphratica was identified in human plasma and targeting Sprouty RTK Signaling Antagonist 4 (SPRY4) gene of the Janus kinase/ signal transducers and activators of transcription (JAK-STAT) signaling pathway [115]. Based on the computationally predicted miRNAs from Camptotheca acuminate, 14 potential miRNAs were found to be regulated 152 target human genes such as miR4723-3p, miR5780d, and miR548d-3p targeting discs large MAGUK scaffoldprotein 2 (DLG2), NUMB endocytic adaptor protein (NUMB) and glycogen synthase kinase-3B (GSK3B) genes which were related to cancers such as breast cancer, lung cancer and leukemia [116]. Ocimum basilicum is a medicinal plant and its bioactive compounds have potential for therapeutic approaches. miRNA target prediction analysis revealed the target of O. basilicum miRNAs, miR156, miR531, miR160, miR529b, and miR1118 were 87 human target genes associated with the Ras-mitogenactivated protein kinase (Ras-MAPK) signaling pathway, Alzheimer disease, breast cancer, cardiomyopathy, HIV, lung cancer, and several neurological disorders [117].

Conclusion
It has been widely known that functional foods and their bioactive compounds have the capacity for human health benefits. To date, miRNAs have been shown a significant effect on gene expression and modulate the cellular biological functions in physiological and pathological conditions. There is emerging evidence suggesting that dietary bioactive compounds can be effective in human diseases as a result of altering miRNAs expression levels, resulting in modulation of cellular signaling pathway. Additional research the possibility of bioactive compounds for developing as novel drugs with less side effects is required in vitro and in vivo. Recently, it has been revealed in several studies that dietary derived-miRNAs are bioavailable and alter human gene expression. The cross-kingdom gene regulations of dietary miRNAs from plants to human have raised our expectations for evaluating the active therapeutic potential and dietary supplements.