Association analysis of miRNA-146a and miRNA-196a polymorphism with breast cancer risk in Khyber Pakhtunkhwa, Pakistan: a preliminary study.

BACKGROUND: microRNAs (miRNAs) are small, noncoding RNA molecules, functioning either as oncogenes or tumor suppressor genes. SNPs in miRNAs might modify the expression of genes associated with miRNAs, enhancing susceptibility to breast cancer. Both miRNA-146a (rs2910164) and miRNA-196a (rs11614913) are identified and significantly associated with breast cancer risk in several ethnicities, but remains unexplored in Khyber Pakhtunkhwa population of Pakistan. METHODS: This study was aimed to check the relation of selected SNPs with breast cancer risk. The research cohort included 100 breast cancer patients and 100 healthy controls. All the participants were subjected for DNA extraction followed by T-ARMS PCR and gel electrophoresis. RESULTS: The results revealed a strong association between risk allele (G) of miRNA-146a and increased risk of breast cancer (OR = 2.04, P = 0.0006). Similarly, heterozygous and mutant genotypes also indicated high risk and significant association with breast cancer risk (CG; OR = 0.51, 9 P = 0.0001) (GG; OR = 3.76, P = 0.04). However, risk allele (T) of miRNA-196a (rs11614913) failed to exhibit significant association with breast cancer risk (OR = 0.92 P = 0.68). Similarly, the heterozygous and mutant genotype did not show significant association with breast cancer risk (CT; OR = 0.52, P = 0.125 (TT; OR = 0.88, P = 0.84). Furthermore, miRNA-146a (rs2910164) and miRNA-196a (rs11614913) polymorphisms exhibited non-significant associations with family history (P = 0.34, P = 0.77), PR status (P = 0.310, P = 0.397), ER status (P = 0.992, P = 0.981), nodal status (P = 0.86, P = 0.90), and menstrual status (P = 0.97, P = 0.09). Notably, miRNA-196a showed a significant association with the metastasis group (P = 0.010) and cancer stages (P = 0.047). CONCLUSIONS: In conclusion, this study highlights the association of miRNA-146a (rs2910164) polymorphism with breast cancer risk but suggested non-significant association of miRNA-196a (rs11614913) with breast cancer risk. However, these findings need to be confirmed through larger data set for more accurate result.

Renoprotective effect of a novel combination of 6-gingerol and metformin in high-fat diet/streptozotocin-induced diabetic nephropathy in rats via targeting miRNA-146a, miRNA-223, TLR4/TRAF6/NLRP3 inflammasome pathway and HIF-1α.

BACKGROUND: MiRNA-146a and miRNA-223 are key epigenetic regulators of toll-like receptor 4 (TLR4)/tumor necrosis factor-receptor-associated factor 6 (TRAF6)/NOD-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome pathway, which is involved in diabetic nephropathy (DN) pathogenesis. The currently available oral anti-diabetic treatments have been insufficient to halt DN development and progression. Therefore, this work aimed to assess the renoprotective effect of the natural compound 6-gingerol (GR) either alone or in combination with metformin (MET) in high-fat diet/streptozotocin-induced DN in rats. The proposed molecular mechanisms were also investigated. METHODS: Oral gavage of 6-gingerol (100 mg/kg) and metformin (300 mg/kg) were administered to rats daily for eight weeks. MiRNA-146a, miRNA-223, TLR4, TRAF6, nuclear factor-kappa B (NF-κB) (p65), NLRP3, caspase-1, and hypoxia-inducible factor-1 alpha (HIF-1α) mRNA expressions were measured using real-time PCR. ELISA was used to measure TLR4, TRAF6, NLRP3, caspase-1, tumor necrosis factor-alpha (TNF-α), and interleukin-1-beta (IL-1ß) renal tissue levels. Renal tissue histopathology and immunohistochemical examination of fibronectin and NF-κB (p65) were performed. RESULTS: 6-Gingerol treatment significantly reduced kidney tissue damage and fibrosis. 6-Gingerol up-regulated miRNA-146a and miRNA-223 and reduced TLR4, TRAF6, NF-κB (p65), NLRP3, caspase-1, TNF-α, IL-1ß, HIF-1α and fibronectin renal expressions. 6-Gingerol improved lipid profile and renal functions, attenuated renal hypertrophy, increased reduced glutathione, and decreased blood glucose and malondialdehyde levels. 6-Gingerol and metformin combination showed superior renoprotective effects than either alone. CONCLUSION: 6-Gingerol demonstrated a key protective role in DN by induction of miRNA-146a and miRNA-223 expression and inhibition of TLR4/TRAF6/NLRP3 inflammasome signaling. 6-Gingerol, a safe, affordable, and abundant natural compound, holds promise for use as an adjuvant therapy with metformin in diabetic patients to attenuate renal damage and stop the progression of DN.

DnaJ-induced miRNA-146a negatively regulates the expression of IL-8 in macrophages.

As a member of the damage-associated molecular patterns, heat shock proteins (HSPs) are widely recognized for their role in initiating innate immune responses. These highly conserved proteins are expressed ubiquitously in both prokaryotes and eukaryotes. In this study, our aim was to investigate how DnaJ, a HSP40 homolog derived from Pseudomonas aeruginosa (P. aeruginosa), influences the regulation of IL-8 expression in macrophages. Treatment with DnaJ served as a stimulus, inducing a more robust expression of IL-8 compared to other HSP homologs, including DnaK, GroEL, and HtpG. This effect was achieved through the activation of the NF-κB signaling pathway. Interestingly, DnaJ treatment also significantly increased the expression of microRNA-146a (miR-146a), which appears to play a role in modulating the expression of innate defense genes. As a consequence, pre-treatment with DnaJ led to a reduction in the extent of IL-8 induction in response to P. aeruginosa treatment. Notably, this reduction was counteracted by transfection of a miR-146a inhibitor, highlighting the involvement of miR-146a in P. aeruginosa-mediated induction of IL-8 expression. Therefore, this study uncovers the role of DnaJ in triggering the expression of miR-146a, which, in turn, modulates the excessive expression of IL-8 induced by P. aeruginosa infection.

Serum miRNA-21, miRNA-146a and plasma cell free DNA as novel biomarkers for assessing systemic lupus erythematosus activity.

BACKGROUND: MicroRNA and cell-free DNA have shown significant correlations with several autoimmune disorders including systemic lupus erythematosus (SLE). SLE has been associated with challenges in determining its activity, so that the need for biomarkers contributing to assessing its activity is emerging. The current study investigated miRNA-21, miRNA-146a and plasma cf-DNA in determination of SLE activity, in addition their association with clinical data including complement factor 3 (C3), complement factor(C4), anti-dsDNA, and other disease activity indices. METHODS AND RESULTS: Eighty subjects divided into; twenty active patients (with SLE-DAI2K score of 16-18) twenty inactive patients (with SLE-DAI2K score of 1-3), and forty healthy control participants) were included in this study. Serum miR-21, miR-146a, and plasma cf-DNA were quantified by real time PCR and their correlation with clinical data was statistically analyzed. The results demonstrated that active cases have significant upregulation of serum miRNA-21 and plasma cf-DNA. Moreover, miR-21 showed a negative, significant pertaining to C3, C4 and was positively related to Systemic Lupus Erythematosus Disease Activity Index 2 K score (SLE-DAI Index2K score) and Systemic-Lupus-Erythematosus-Disease Activity-Index 2 K activity (SLE-DAI 2 K activity). Also, Active group miRNA-146a was negatively, significantly correlated with C3, as well as a positive significant relationship with SLE-DAI2K score and SLEDAI 2 K activity, in addition to anti DNA Autoantibodies. Furthermore, miR-21 and cf-DNA demonstrated a differential value through Receiver Operating Characteristic (ROC) curve's study. CONCLUSIONS: the present study illustrated miR-21, miR-146a, and cf-DNA relationship with SLE clinical data. In addition to their potential value in SLE diagnosis, and activity determination.

MiRNA-146a-A Key Player in Immunity and Diseases.

miRNA-146a, a single-stranded, non-coding RNA molecule, has emerged as a valuable diagnostic and prognostic biomarker for numerous pathological conditions. Its primary function lies in regulating inflammatory processes, haemopoiesis, allergic responses, and other key aspects of the innate immune system. Several studies have indicated that polymorphisms in miRNA-146a can influence the pathogenesis of various human diseases, including autoimmune disorders and cancer. One of the key mechanisms by which miRNA-146a exerts its effects is by controlling the expression of certain proteins involved in critical pathways. It can modulate the activity of interleukin-1 receptor-associated kinase, IRAK1, IRAK2 adaptor proteins, and tumour necrosis factor (TNF) targeting protein receptor 6, which is a regulator of the TNF signalling pathway. In addition, miRNA-146a affects gene expression through multiple signalling pathways, such as TNF, NF-κB and MEK-1/2, and JNK-1/2. Studies have been carried out to determine the effect of miRNA-146a on cancer pathogenesis, revealing its involvement in the synthesis of stem cells, which contributes to tumourigenesis. In this review, we focus on recent discoveries that highlight the significant role played by miRNA-146a in regulating various defence mechanisms and oncogenesis. The aim of this review article is to systematically examine miRNA-146a's impact on the control of signalling pathways involved in oncopathology, immune system development, and the corresponding response to therapy.

Lead exposure promotes the inflammation via the circRNA-05280/miR-146a/IRAK1 axis in mammary gland.

Lead, the most widely used heavy metal in industry, is detrimental to human health if exposed to living and occupational environment. Although several studies have been conducted on lead exposure, little has been reported on its harm to mammary gland and its mechanisms. In view of this, our study is the first to verify that lead exposure could promote apoptosis and inflammation in mouse mammary tissue (in vivo) and cow mammary epithelial cells (in vitro). After establishing a lead exposed mouse model, the expression profile of mammary gland tissue was constructed by high-throughput sequencing technology. In the profile, 917 differentially expressed genes were screened, of which IRAK1 was up-regulated by 4.33 times. Then, from qRT-PCR, Western blot and Luciferase report, IRAK1 was found to promote the release of inflammatory factors and tissue apoptosis and could be a specific target of miR-146a. On the other hand, double luciferase reporter system and qRT-PCR predicted the existence of a binding site between circRNA-05280 and miR-146a sequence. Experiments such as immunohistochemistry, apoptosis and EdU demonstrated that circRNA-05280 could promote not only cell apoptosis but also the expression level of inflammatory genes. Nevertheless, the function of miR-146a is opposite to that of circRNA-05280. Specifically, circRNA-05280 can regulate the level of apoptosis and inflammation of mammary gland by binding miR-146a and releasing the expression of miR-146a on target gene IRAK1. This study concludes that circRNA-05280/ miR-146a/ IRAK1 signaling pathway could mediate the mammary gland damage resulting from lead exposure. Accordingly, it sheds new light on further exploration of molecular mechanisms of mammary gland tissue damage caused by lead exposure, the risk assessment of lead, and the mechanism of lead mammary gland toxicity.

Analysis of serum microRNAs and rs2910164 GC single-nucleotide polymorphism of miRNA-146a in COVID-19 patients.

Alteration of micro-RNAs (miRNAs) expression, including miRNA-122a, -146a and -205 family members, can have profound effects on inflammatory and IFN pathways (miRNA-146a), known as hallmarks of COVID-19. SARS-CoV-2-infected patients were recruited at Policlinico Umberto I Hospital of Sapienza University of Rome (Italy). MiRNA-122a, -146a, -205 and IFI27 (Interferon Alpha Inducible Protein 27) levels were screened in SARS-CoV-2 patients (n = 14) and healthy controls (n = 10) by real-time RT-PCR assays. Then, miRNA-146a rs2910164 GC single-nucleotide polymorphism (SNP) was genotyped in a larger group of COVID-19 patients (n = 129), and its relationship with severe disease [Intensive Care Unit (ICU) support or survival/death] was assessed. SARS-CoV-2-positive patients had increased PCR, D-Dimer and Fibrinogen levels compared to healthy controls (p < .05 for all measurements). MiRNA-122a and -146a serum levels were upregulated in COVID-19 patients (miRNA-122a: p = .002; miRNA-146a: p < .001). Decreased IFI27 levels were observed in COVID-19 patients with higher miRNA-146a levels (p = .047). Moreover, miRNA-146a rs2910164 C/G genotypes distributions were similar in COVID-19 patients and in validated European healthy subjects (n = 37,214). MiRNA-146a SNP was not associated with severe COVID-19 outcome (ICU or death). MiRNA-122a and -146a levels were elevated in SARS-CoV-2 infected patients, with miRNA-146a upregulation possibly contributing to IFN pathways dysregulation (e.g., reduced IFI27 levels) observed in severe COVID-19, although there is no evidence for the involvement of rs2910164 SNP.

An Eleven-microRNA Signature Related to Tumor-Associated Macrophages Predicts Prognosis of Breast Cancer.

The dysregulation of microRNAs (miRNAs) has been known to play important roles in tumor development and progression. However, the understanding of the involvement of miRNAs in regulating tumor-associated macrophages (TAMs) and how these TAM-related miRNAs (TRMs) modulate cancer progression is still in its infancy. This study aims to explore the prognostic value of TRMs in breast cancer via the construction of a novel TRM signature. Potential TRMs were identified from the literature, and their prognostic value was evaluated using 1063 cases in The Cancer Genome Atlas Breast Cancer database. The TRM signature was further validated in the external Gene Expression Omnibus GSE22220 dataset. Gene sets enrichment analyses were performed to gain insight into the biological functions of this TRM signature. An eleven-TRM signature consisting of mir-21, mir-24-2, mir-125a, mir-221, mir-22, mir-501, mir-365b, mir-660, mir-146a, let-7b and mir-31 was constructed. This signature significantly differentiated the high-risk group from the low-risk in terms of overall survival (OS)/ distant-relapse free survival (DRFS) (p value < 0.001). The prognostic value of the signature was further enhanced by incorporating other independent prognostic factors in a nomogram-based prediction model, yielding the highest AUC of 0.79 (95% CI: 0.72−0.86) at 5-year OS. Enrichment analyses confirmed that the differentially expressed genes were mainly involved in immune-related pathways such as adaptive immune response, humoral immune response and Th1 and Th2 cell differentiation. This eleven-TRM signature has great potential as a prognostic factor for breast cancer patients besides unravelling the dysregulated immune pathways in high-risk breast cancer.

Lipopolysaccharides (LPSs) as Potent Neurotoxic Glycolipids in Alzheimer's Disease (AD).

Lipopolysaccharides (LPSs) are microbiome-derived glycolipids that are among the most potent pro-inflammatory neurotoxins known. In Homo sapiens, the major sources of LPSs are gastrointestinal (GI)-tract-resident facultative anaerobic Gram-negative bacilli, including Bacteroides fragilis and Escherichia coli. LPSs have been abundantly detected in aged human brain by multiple independent research investigators, and an increased abundance of LPSs around and within Alzheimer's disease (AD)-affected neurons has been found. Microbiome-generated LPSs and other endotoxins cross GI-tract biophysiological barriers into the systemic circulation and across the blood-brain barrier into the brain, a pathological process that increases during aging and in vascular disorders, including 'leaky gut syndrome'. Further evidence indicates that LPSs up-regulate pro-inflammatory transcription factor complex NF-kB (p50/p65) and subsequently a set of NF-kB-sensitive microRNAs, including miRNA-30b, miRNA-34a, miRNA-146a and miRNA-155. These up-regulated miRNAs in turn down-regulate a family of neurodegeneration-associated messenger RNA (mRNA) targets, including the mRNA encoding the neuron-specific neurofilament light (NF-L) chain protein. While NF-L has been reported to be up-regulated in peripheral biofluids in AD and other progressive and lethal pro-inflammatory neurodegenerative disorders, NF-L is significantly down-regulated within neocortical neurons, and this may account for neuronal atrophy, loss of axonal caliber and alterations in neuronal cell shape, modified synaptic architecture and network deficits in neuronal signaling capacity. This paper reviews and reveals the most current findings on the neurotoxic aspects of LPSs and how these pro-inflammatory glycolipids contribute to the biological mechanism of progressive, age-related and ultimately lethal neurodegenerative disorders. This recently discovered gut-microbiota-derived LPS-NF-kB-miRNA-30b-NF-L pathological signaling network: (i) underscores a direct positive pathological link between the LPSs of GI-tract microbes and the inflammatory neuropathology, disordered cytoskeleton, and disrupted synaptic-signaling of the AD brain and stressed human brain cells in primary culture; and (ii) is the first example of a microbiome-derived neurotoxic glycolipid having significant detrimental miRNA-mediated actions on the expression of NF-L, an abundant filamentous protein known to be important in the maintenance of neuronal and synaptic homeostasis.

miRNA-146a-5p mitigates stress-induced premature senescence of D-galactose-induced primary thymic stromal cells.

Senescent thymic stromal cells (TSCs) producing senescence-associated secretory phenotype (SASP) may play a role at later phases of thymic involution. However, the etiology and mechanisms responsible for TSC senescence remain to be elucidated. In the present study, the effects of oxidative stress on TSCs and role of miRNA-146a-5p in stress-induced premature senescence (SIPS) were identified. D-galactose (D-gal) induced oxidative stress in primary TSCs and a limited cumulative oxidative stress induced premature senescence but not apoptosis of TSCs. miRNA-146a-5p overexpression can mitigate the SIPS by targeting tumor necrosis factor receptor-associated factor 6 (TRAF6) instead of increasing autophagy clearance. Furthermore, exogenous miRNA-146a-5p reversed the upregulation of chemokines including Cxcl5, pro-inflammatory cytokines, and antimicrobial peptides in TSCs with SIPS. In conclusion, the accumulated oxidative stress may be partially responsible for senescence in TSCs and modulation of miRNA-146a-5p may attenuate this process.

Long non-coding RNA PVT1, a novel biomarker for chronic obstructive pulmonary disease progression surveillance and acute exacerbation prediction potentially through interaction with microRNA-146a.

OBJECTIVE: This study aimed to investigate the abilities of long non-coding RNA PVT1 (lnc-PVT1) and microRNA-146a (miR-146a) in predicting chronic obstructive pulmonary disease (COPD) susceptibility and acute exacerbation risk, moreover, to explore the association of lnc-PVT1 with disease severity, inflammation, and miR-146a in patients with COPD. METHODS: A total of 80 acute exacerbation of COPD (AECOPD) patients, 80 stable COPD patients, and 80 healthy controls (HCs) were consecutively recruited. Peripheral blood samples of all participants were collected to isolate peripheral blood mononuclear cells (PBMCs), and serum: PBMCs were used to detect lnc-PVT1 and miR-146a by RT-qPCR; serum was used to detect inflammatory cytokines by ELISA. Global Initiative for Chronic Obstructive Lung Disease (GOLD) stage of COPD was assessed. RESULTS: Lnc-PVT1 expression was highest in AECOPD patients, followed by stable COPD patients and HCs. Receiver operating characteristic curves disclosed that lnc-PVT1 distinguished AECOPD patients and stable COPD patients from HCs, also distinguished AECOPD patients from stable COPD patients. In AECOPD patients and stable COPD patients, lnc-PVT1 expression positively correlated with GOLD stage and levels of TNF-α, IL-6, IL-8, and IL-17. Moreover, lnc-PVT1 was negatively correlated with miR-146a. For miR-146a, its expression was lowest in AECOPD patients, followed by stable COPD patients and HCs, and it predicted reduced COPD susceptibility and decreased acute exacerbation risk; meanwhile, it negatively correlated with GOLD stage and inflammatory cytokine levels in AECOPD patients and stable COPD patients. CONCLUSION: Lnc-PVT1 assists the disease management and acute exacerbation risk monitoring of COPD via interaction with miR-146a.

microRNA-146a downregulates IL-17 and IL-35 and inhibits proliferation of human periodontal ligament stem cells.

Periodontitis is characterized by increased levels of proinflammatory factors, such as interleukin-17 (IL-17) and IL-35. In this study, the expression of microRNA-146a (miRNA-146a), IL-17, and IL-35 in the plasma of patients with periodontitis and healthy controls were detected by quantitative reverse transcription polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. miRNA-146a mimic was transfected into periodontal ligament stem cells (PDLSCs) isolated from periodontitis-affected teeth and healthy teeth. Cell proliferation and expression of IL-17 and IL-35 were detected by cell counting kit-8 assay and Western blot analysis, respectively. It was observed that miRNA-146a was downregulated but IL-17 and IL-35 were upregulated in the plasma of patients with periodontitis than in healthy controls. miRNA-146a was inversely correlated with IL-17 and IL-35 in patients with periodontitis. miRNA-146a overexpression inhibited proliferation of PDLSCs derived from both periodontitis-affected teeth and healthy teeth. miRNA-146a overexpression led to downregulated IL-17 and IL-35 expression in PDLSCs isolated from periodontitis-affected teeth. We, therefore, conclude that miRNA-146a may improve periodontitis by downregulating IL-17 and IL-35 expression and inhibiting proliferation of human PDLSCs.

Association between miRNA-146a rs2910164 (G/C) polymorphism with the susceptibility to chronic HBV infection and spontaneous viral clearance in an Iranian population.

Hepatitis B virus (HBV) infection is one of the clinical dilemmas in chronic liver diseases. MicroRNAs (miRNAs) are small noncoding RNA molecules that play an important role in the pathogenesis of liver diseases and single nucleotide polymorphisms (SNPs) in miRNA genes affect the clinical course of HBV infection. Previous studies have shown that miRNA-146a rs2910164 polymorphism can be associated with the pathogenesis of liver diseases such as hepatocellular carcinoma. The present study investigated the association between miRNA-146a rs2910164 polymorphism and susceptibility to HBV infection in an Iranian population. The study comprised 266 patients with chronic HBV infection, 172 patients with spontaneous viral clearance (SVC) after acute HBV infection, and 266 healthy control adjusted for sex and age. The genotyping of the miRNA-146a rs2910164 polymorphism was performed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. Our data revealed that GG genotype and G allele of miRNA-146a rs2910164 SNP is dominated (P < 0.001) in patients with chronic HBV infection (Odds ratio [OR] = 3.92; 95% confidence interval [CI] = 2.1-7.32). miRNA-146a rs2910164 polymorphism showed a statistically significant association (P < 0.001) between CC genotype and allele C with SVC (OR = 2.92; 95% CI = 1.56-546). Our findings suggest miRNA-146a SNP (C/G) in our population may be associated with the susceptibility to HBV infection and CC genotype is associated with SVC. Also, the GG genotype and G allele at miRNA-146a rs2910164 is associated with chronic HBV infection in our population.

MiRNA-146a polymorphism increases the odds of malaria in pregnancy.

BACKGROUND: Plasmodium falciparum infection during pregnancy is a major cause of poor maternal health, adverse foetal outcome and infant mortality in sub-Saharan Africa. Genetic disposition is involved in susceptibility to malaria in pregnancy and its manifestation. MicroRNAs (miRNAs) influence gene regulation including that of innate immune responses. A miRNA-146a rs2910164 G > C single nucleotide polymorphism (SNP) has been associated with increased risks of several diseases, but no data as to malaria are available. METHODS: The association between miRNA-146a rs2910164 and P. falciparum infection among 509 Ghanaian women attending antenatal care (ANC) and 296 delivering Ghanaian primiparae was investigated. Malaria parasites were diagnosed by microscopy and PCR. Leukocyte-associated hemozoin in placental samples was recorded as well. Proportions were compared between groups by Fisher's exact test, and logistic regression models were used to adjust for possible confounders. RESULTS: By PCR, P. falciparum infection was detected in 63% and 67% of ANC attendees and delivering primiparae, respectively. In both groups, two in three women were either heterozygous or homozygous for miRNA-146a rs2910164. Among ANC attendees, homozygosity conferred increased odds of infection (adjusted odds ratio (aOR), 2.3; 95% CI, 1.3-4.0), which was pronounced among primigravidae (aOR, 5.8; 95% CI, 1.6-26) but only marginal in multigravidae. Likewise, homozygosity for miRNA-146a rs2910164 in primiparae increased the odds of past or present placental P. falciparum infection almost six-fold (aOR, 5.9; 95% CI, 2.1-18). CONCLUSIONS: These results indicate that SNP rs2910164 G > C is associated with increased odds for P. falciparum infection in first-time pregnant women who are considered to lack sufficient acquired immune responses against pregnancy-specific strains of P. falciparum. These findings suggest that miRNA-146a is involved in protective malarial immunity, and specifically in the innate component.

Expression of miRNA-146a, miRNA-155, IL-2, and TNF-α in inflammatory response to Helicobacter pylori infection associated with cancer progression.

miRNAs appear to play an important role in controlling the expression of several genes, and they are a potential biomarker and prognostic tool in gastric diseases. We analyzed 53 controls, 86 patients with gastritis, and 19 patients with gastric cancer. Real-time-PCR was used to determine the expression levels of miRNA-146a, miRNA-155, IL-2, and TNF-α. The subsequent analysis of the target genes was performed using the bioinformatics approach. There was no difference in IL-2 expression between the groups. However, there was a significant increase in TNF-α expression in the gastritis group relative to the control and a significant decrease in the gastric cancer group relative to the control. There was also a statistically significant increase in miRNA-146a and miRNA-155 expression in the gastritis group relative to the control, but not in the gastric cancer group. Similar results were found when the presence of H. pylori was considered. The data revealed an increase in miRNA-146a and miRNA-155 expression but not enough to control the expression of TNF-α. The presence of H. pylori was found to affect increases in TNF-α and microRNA expression, and miRNA-146a and miRNA-155 alone were not able to eliminate bacteria or restore tissue homeostasis.

Association of MiRNA-146a, MiRNA-499, IRAK1 and PADI4 Polymorphisms with Rheumatoid Arthritis in Egyptian Population.

BACKGROUND/AIMS: Rheumatoid arthritis (RA) is a systemic autoimmune disease affecting up to 1% of the population worldwide. The aim of the present study was to investigate whether miRNA-146a rs2910164, miRNA-499 rs3746444, IRAK1 rs3027898 and PADI4 rs1748033 polymorphisms are associated with susceptibility to RA in Egyptians and whether they influence disease severity and activity. METHODS: The study was performed on 104 unrelated RA patients and 112 healthy subjects. RA patients were further subdivided into active and inactive RA groups. Polymorphisms were genotyped by using real-time polymerase chain reaction with TaqMan allelic discrimination assay. RESULTS: Significant differences in the frequency of miRNA-146a rs2910164, miRNA-499 rs3746444, IRAK1 rs3027898 and PADI4 rs1748033 alleles and genotypes were observed between RA patients and controls. Only CA and AA genotypes of IRAK1 rs3027898 shows a significant difference between active and inactive subgroups. MiRNA-146a rs2910164 and IRAK1 rs3027898 polymorphisms were a risk factor for predisposition to RA in codominant and dominant tested inheritance models, while, the miRNA-499 rs3746444 and PADI4 rs1748033 polymorphisms were a risk factor in codominant and recessive one. CG and GG genotypes of miRNA-146a rs2910164 were associated with positive erosions. CA genotype of IRAK1 rs3027898 was associated with low disease activity and negative erosions, while, the AA genotype was associated with high disease activity. CC genotype of PADI4 rs1748033 was associated with negative rheumatoid factor. CONCLUSION: The 4 studied SNPs were likely to play an important role in the susceptibility to RA and can influence disease severity and activity in Egyptian population.

Chronic constriction injury-induced microRNA-146a-5p alleviates neuropathic pain through suppression of IRAK1/TRAF6 signaling pathway.

BACKGROUND: microRNA-146a-5p (miRNA-146a-5p) is a key molecule in the negative regulation pathway of TLRs and IL-1 receptor (TIR) signaling. Our recent study demonstrated that MyD88-dependent signaling pathway of TIR in the dorsal root ganglion (DRG) and spinal dorsal horn (SDH) plays a role in peripheral nerve injury-induced neuropathic pain. However, it was not clear whether and how miRNA-146a-5p regulates the TIR pathway of DRG and SDH in the development of neuropathic pain. METHODS: The sciatic nerve chronic constriction injury (CCI) model of rat was used to induce chronic neuropathic pain. The levels and cellular distribution of miRNA-146a-5p were detected with quantitative real-time PCR (qPCR) and fluorescent in situ hybridization (FISH). The RNA level, protein level, and cellular distribution of IRAK1 and TRAF6 that is targeted by miRNA-146a-5p were detected with qPCR, western blot, and immunofluorescent. The pain-related behavioral effect of miRNA-146a-5p was accessed after intrathecal administration. Mechanical stimuli and radiant heat were used to evaluate mechanical allodynia and thermal hyperalgesia. RESULTS: We found that the level of miRNA-146a-5p significantly increased in L4-L6 DRGs and SDH after CCI surgery; meanwhile, the protein level of IRAK1 and TRAF6 in DRGs was significantly increased after CCI. Intrathecal injection of miR146a-5p agomir or miRNA-146a-5p antagomir regulates miRNA-146a-5p level of L4-L6 DRGs and SDH. We found that intrathecal injection of miR146a-5p agomir can alleviate mechanical and thermal hyperalgesia in CCI rats and reverse the upregulation of IRAK1 and TRAF6 of L4-L6 DRGs and SDH induced by CCI. We furthermore found that intrathecal injection of miRNA-146a-5p antagomir can exacerbate the mechanical and thermal pain-related behavior of CCI rats and meanwhile increase IRAK1 and TRAF6 of L4-L6 DRGs and SDH expression even further. CONCLUSIONS: miRNA-146a-5p of DRG and SDH can modulate the development of CCI-induced neuropathic pain through inhibition of IRAK1 and TRAF6 in the TIR signaling pathway. Hence, miRNA-146a-5p may serve as a potential therapeutic target for neuropathic pain.

Possible use of miRNAs-146a and -499 expression and their polymorphisms as diagnostic markers for rheumatoid arthritis.

Rheumatoid arthritis (RA) is a systemic autoimmune disorder affecting the peripheral joints. Different microRNAs had been investigated in RA including miRNA-146a meanwhile, miRNA-499 there were no studies to prove its expression in RA serum samples. This study was performed to investigate expression of both miRNAs-146a and -499 and their polymorphisms in Egyptian patients with RA and to evaluate their relationship with clinico-pathological data. The present study includes 108 subjects classified into two main groups: 52 RA patients and 56 unrelated healthy controls. RA patients were subclassified according to DAS28 score into inactive (23 patients) and active (29 patients). Quantitative expression of serum miRNA-146a, miRNA-499 as well as their Genotyping rs2910164 (C/G) and rs3746444 (T/C), respectively, were done to all subjects using real-time PCR. Serum miRNA-146a and -499 were significantly over expressed in RA patients, but they were not correlated to disease activity. Serum miRNA-146a was negatively correlated with anti-nuclear antibodies (ANA). miRNA-146a (rs2910164) genotyping revealed that the GG genotype and the frequency of the G allele were significantly higher in RA patients compared to the controls. miRNA-499 (rs3746444), genotyping revealed that the CC genotype and the frequency of the C allele were significantly higher. It can be concluded that both miRNAs-146a and -499 can be used as diagnostic markers for RA patients. Both miRNA-146a (rs2710164) and miRNA-499 (rs3746444) were significantly associated with RA susceptibility. The C allele of miRNA-146a (rs2710164) can be considered to be protective. On the other hand, the C allele of miRNA-499 (rs3746444) was significantly associated with RA susceptibility.

Fascin protein stabilization by miR-146a implicated in the process of a chronic inflammation-related colon carcinogenesis model.

OBJECTIVE: In sporadic colon tumors, multistep process of well-known genetic alterations accelerates carcinogenesis; however, this does not appear to be the case in inflammation-related ones. We previously established a model of inflammation-related colon carcinogenesis using human colonic adenoma cells, and identified fascin as a driver gene of this process. We analyzed the microRNAs involved in the stable fascin expression in colon adenocarcinoma cells. MATERIALS AND METHODS: miRNA microarray analysis was performed using FPCK-1-1 adenoma cells and its-derived FPCKpP1-4 adenocarcinoma cells through chronic inflammation. To assess the involvement of miRNA in the inflammation-related carcinogenesis, sphere-forming ability, expression of colon cancer stemness markers, and stability of fascin protein via the proteasome using tough decoy RNA technique. RESULTS: We found that 17 miRNAs including miR-146a were upregulated and 16 miRNAs were downregulated in FPCKpP1-4 adenocarcinoma cells. We revealed that miR-146a in the adenocarcinoma cells brought about acquisition of sphere formation, cancer stemness, and inhibition of proteasomal degradation of the fascin protein. CONCLUSIONS: We found that stable fascin expression is brought about via the inhibition of proteasome degradation by miR-146a in the process of a chronic inflammation-related colon carcinogenesis.

Differential expression of miRNA-146a and miRNA-155 in gastritis induced by Helicobacter pylori infection in paediatric patients, adults, and an animal model.

BACKGROUND: Helicobacter pylori is a major aetiologic agent associated with gastritis. H. pylori infections increase the expression of the Toll-like receptor (TLR), which in turn modulates the expression of microRNA (miRNA)-146a and miRNA-155. The objective of this study was to compare the expression of miRNA-146a and miRNA-155 in gastric lesions of paediatric and adult patients with different pathologies and in Mongolian gerbils (Meriones unguiculatus) infected with H. pylori 26,695. METHODS: Quantification of miRNA expression was performed by quantitative real-time polymerase chain reaction (qRT-PCR) of paraffin-embedded gastric lesions of children with or without an infection (n = 25), adults with follicular gastritis and metaplasia (n = 32) and eight-week-old M. unguiculatus males (Hsd:MON) infected with H. pylori 26,695 for 0, 3, 6, 12 and 18 months (n = 25). The genes RNU48 and RNU6 were used as endogenous controls for data normalization. Statistical analyses were performed using Kruskal-Wallis, Mann-Whitney, ANOVA and Student's t-test. RESULTS: The expression of miRNA-146a and miRNA-155 in infected children increased by 247.6- and 79.4-fold (on average), respectively, compared to that observed in the control group. However, these results were not significant (p = 0.12 and p = 0.07 respectively). In some children a gradual increase in expression was observed, while in others, expression was very high. Additionally, the expression levels of miRNA-146a and miRNA-155 increased by an average of 21.7- and 62-fold, respectively, in adult patients with follicular gastritis when compared to those of the controls. In M. unguiculatus infected with H. pylori 26,695, the expression of both miRNAs increased as the infection progressed. CONCLUSION: This is the first report to show differences in the expression of miRNA-146a and miRNA-155 in paediatric and adult patients with gastritis who were infected with H. pylori. In addition, in M. unguiculatus infected with H. pylori, miRNA expression was associated with the progression of infection and the ability of the bacteria to adapt to the host.