microRNA 146a Research Articles

MicroRNA 146a Is a Negative Regulator of the Host Innate Immune Response to Urinary Tract Infection

MicroRNA 146a Is a Negative Regulator of the Host Innate Immune Response to Urinary Tract Infection

Deregulation of MicroRNA-146a and 155 expression levels might underlie complicated pregnancy in Toxoplasma Gondii seronegative women

BackgroundTo evaluate the ability of the estimated plasma expression levels of genes of microRNA (MiR-) 146a and 155 to differentiate between samples of pregnant women suspected to be infected by T. gondii. 50 newly pregnant women who had at least one of the criteria of high risk for toxoplasma infection and 50 newly primigravida women free of these criteria gave blood samples for qualitative determination of serum toxoplasma antibodies and estimation of plasma expression levels of MiR-146a and 155 using the qRT-PCR. During the pregnancy course, the incidence of pregnancy complications was recorded.ResultsTwenty-six women were IgM−/IgG−, 17 women were IgM+/IgG− and 7 women were IgM+/IgG+. Thirty-two women had pregnancy complications with significantly lower incidence in IgM−/IgG− women. Plasma expression levels of MiR-146a and 155 were significantly higher in total patients compared to control levels and were significantly higher in samples of IgM+/IgG+ patients than in other samples. Statistical analyses defined a high plasma level of MiR-155 as the highly significant predictor for oncoming pregnancy complications and high levels of both microRNAs as predictors for the presence of toxoplasmosis despite seronegativity. Kaplan-Meier regression analysis defined increasing cumulative risk of having toxoplasmosis despite seronegativity with plasma levels of MiR-146a and MiR-155 of 1.2 and 3, respectively.ConclusionThe incidence of pregnancy complications is high, irrespective of the seronegativity of women at high risk of toxoplasmosis. Estimated plasma levels of MiR-155 might identify women liable to develop complications and differentiate seronegative women vulnerable to having T. gondii infection.Trial registrationThe study protocol was approved preliminarily by the Local Ethical Committee at Benha Faculty of Medicine. Before enrollment, the study protocol was discussed in detail with the study participants, and those accepted to participate in the study signed written fully informed consents. The final approval of the study protocol was obtained after the end of case collection and registered by RC: 5-11-2022.

Targeting Inflammation and Oxidative Stress to Improve Outcomes in a TNBS Murine Crohn's Colitis Model.

Inflammation and oxidative stress are implicated in the pathogenesis of Crohn's disease. Cerium oxide nanoparticle (CNP) conjugated to microRNA 146a (miR146a) (CNP-miR146a) is a novel compound with anti-inflammatory and antioxidative properties. We hypothesized that local administration of CNP-miR146a would improve colitis in a 2,4,6-Trinitrobenzenesulfonic acid (TNBS) mouse model for Crohn's disease by decreasing colonic inflammation. Balb/c mice were instilled with TNBS enemas to induce colitis. Two days later, the mice received cellulose gel enema, cellulose gel with CNP-miR146a enema, or no treatment. Control mice received initial enemas of 50% ethanol and PBS enemas on day two. The mice were monitored daily for weight loss and clinical disease activity. The mice were euthanized on days two or five to evaluate their miR146a expression, inflammation on histology, and colonic IL-6 and TNF gene expressions and protein concentrations. CNP-miR146a enema successfully increased colonic miR146a expression at 12 h following delivery. At the end of five days from TNBS instillation, the mice treated with CNP-miR146a demonstrated reduced weight loss, improved inflammation scores on histology, and reduced gene expressions and protein concentrations of IL-6 and TNF. The local delivery of CNP-miR146a in a TNBS mouse model of acute Crohn's colitis dramatically decreased inflammatory signaling, resulting in improved clinical disease.

MiR-146a reduces fibrosis after glaucoma filtration surgery in rats

PurposeTo explore the impact of microRNA 146a (miR-146a) and the underlying mechanisms in profibrotic changes following glaucoma filtering surgery (GFS) in rats and stimulation by transforming growth factor (TGF)-β1 in rat Tenon’s capsule fibroblasts.MethodsCultured rat Tenon’s capsule fibroblasts were treated with TGF-β1 and analyzed with microarrays for mRNA profiling to validate miR-146a as the target. The Tenon’s capsule fibroblasts were then respectively treated with lentivirus-mediated transfection of miR-146a mimic or inhibitor following TGF-β1 stimulation in vitro, while GFS was performed in rat eyes with respective intraoperative administration of miR-146a, mitomycin C (MMC), or 5-fluorouracil (5-FU) in vivo. Profibrotic genes expression levels (fibronectin, collagen Iα, NF-KB, IL-1β, TNF-α, SMAD4, and α-smooth muscle actin) were determined through qPCR, Western blotting, immunofluorescence staining and/or histochemical analysis in vitro and in vivo. SMAD4 targeting siRNA was further used to treat the fibroblasts in combination with miR-146a intervention to confirm its role in underlying mechanisms.ResultsUpregulation of miR-146a reduced the proliferation rate and profibrotic changes of rat Tenon’s capsule fibroblasts induced by TGF-β1 in vitro, and mitigated subconjunctival fibrosis to extend filtering blebs survival after GFS in vivo, where miR-146a decreased expression levels of NF-KB-SMAD4-related genes, such as fibronectin, collagen Iα, NF-KB, IL-1β, TNF-α, SMAD4, and α-smooth muscle actin(α-SMA). Additionally, SMAD4 is a key target gene in the process of miR-146a inhibiting fibrosis.ConclusionsMiR-146a effectively reduced TGF-β1-induced fibrosis in rat Tenon’s capsule fibroblasts in vitro and in vivo, potentially through the NF-KB-SMAD4 signaling pathway. MiR-146a shows promise as a novel therapeutic target for preventing fibrosis and improving the success rate of GFS.

Increased expression of miR146a dysregulates TLR2-induced HBD2 in airway epithelial cells from patients with COPD.

Airway epithelial cells from patients with COPD show suboptimal innate immune responses to nontypeable Haemophilus influenzae (NTHi) and Toll-like receptor (TLR)2 ligands despite expressing TLR2 similar to normal airway epithelial cells, but the underlying mechanisms are poorly understood. Normal or COPD mucociliary-differentiated airway epithelial cells were treated with TLR2 agonists or infected with NTHi and expression of β-defensin (HBD)2 was examined. Interleukin-1 receptor-associated kinase (IRAK)-1 and microRNA (miR)146a were genetically inhibited in normal and COPD airway epithelial cell cultures, respectively, and HBD2 responses to TLR2 ligands were determined. IRAK-1 expression in lung sections was determined by immunofluorescence microscopy. Compared to normal, COPD airway epithelial cell cultures showed impaired expression of HBD2 in response to TLR2 agonists or NTHi infection. Apical secretions from TLR2 agonist-treated normal, but not COPD, airway epithelial cells efficiently killed NTHi. Knockdown of HBD2 significantly reduced NTHi killing by apical secretions of normal airway epithelial cells. Compared to normal, COPD cells showed significantly reduced expression of IRAK-1 and this was associated with increased expression of miR146a. Inhibition of miR146a increased the expression of IRAK-1, improved the expression of HBD2 in response to TLR2 agonists in COPD cells and enhanced the killing of bacteria by apical secretions obtained from TLR2 agonist-treated COPD cells. Bronchial epithelium of COPD patients showed reduced expression of IRAK-1. These results suggest that reduced levels of IRAK-1 due to increased expression of miR146a may contribute to impaired expression of TLR2-induced HBD2 in COPD airway epithelial cells.

MicroRNA 146a ameliorates retinal damage in experimental autoimmune uveitis.

Uveitis and related intraocular inflammations are a major cause of blindness due to retinal damage caused by degeneration and loss of the photoreceptor cells. In mouse experimental autoimmune uveitis (EAU) previously we have shown mitochondrial oxidative stress with marked upregulation of αA crystallin in the inner segments of the photoreceptors. Furthermore, αA crystallin treatment prevented photoreceptor mitochondrial oxidative stress by suppressing innate and adaptive immunity in EAU. Since these immune processes are modulated by microRNAs, in this study we investigated (a) modulation of microRNAs during development of EAU by αA crystallin administration and (b) microRNA therapeutic intervention. Few microRNAs were significantly upregulated in EAU mice with intravenous injection of αA crystallin and among these, computational bioinformatic analysis revealed that the upregulated microRNA 146a targets the innate and adaptive immune responses. In EAU, intravenous as well as intravitreal administration of this microRNA prevented inflammatory cell infiltration in uvea and retina and preserved photoreceptor cells. This protective function suggests that microRNA146a can be a novel therapeutic agent in preventing retinal damage in uveitis.

MiR-146a encapsulated liposomes reduce vascular inflammatory responses through decrease of ICAM-1 expression, macrophage activation, and foam cell formation.

Vascular insults can create an inflammatory cascade involving endothelial cell, smooth muscle cell, and macrophage activation which can eventually lead to vascular disease such as atherosclerosis. Several studies have identified microRNA 146a's (miR-146a) anti-inflammatory potential based on its role in regulating the nuclear factor kappa beta (NF-κβ) pathway. Therefore, in this study, we introduced exogenous miR-146a encapsulated by liposomes to lipopolysaccharide (LPS) stimulated vascular cells and macrophages to reduce inflammatory responses. First, the miR-146a encapsulated liposomes showed uniform size (radius 96.4 ± 4.22 nm) and round shape, long term stability (at least two months), high encapsulation efficiency (69.73 ± 0.07%), and were well transfected to human aortic endothelial cells (HAECs), human aortic smooth muscle cells (SMCs), and human differentiated monocytes (U937 cells). In addition, we demonstrated that miR-146a encapsulated liposomes reduced vascular inflammation responses in HAECs and SMCs through inhibition of ICAM-1 expression and decreased monocyte adhesion. In macrophages, miR-146a liposome treatment demonstrated decreased production of proinflammatory cytokines, tumor necrosis factor-alpha (TNF-α) and interleukin-1 beta (IL-1β), as well as reduced oxidized low-density lipoprotein (ox-LDL) uptake and foam cell formation. Thus, based on these results, miR-146a encapsulated liposomes may be promising for reducing vascular inflammation by targeting its multiple associated mediators.

Exosomes from microRNA 146a overexpressed bone marrow mesenchymal stem cells protect against spinal cord injury in rats

BackgroundThis study aimed to investigate the effect of microRNA 146a (miR-146a) overexpressed bone marrow mesenchymal stem cells (BMMSCs) exosomes on spinal cord injury (SCI) recovery. MethodsRat BMMSCs were isolated and transfected with miR-146a mimic (miR-mimic) and control mimic (NC-mimic), after which their exosomes were isolated. Afterward, SCI rat models were constructed, then treated with phosphate buffer saline (PBS), NC BMMSCs exosomes (separated from the culture medium of BMMSCs with NC-mimic), and miR-146a overexpressed BMMSCs exosomes (isolated from the culture medium of BMMSCs with miR-mimic), respectively; additionally, rats underwent sham surgery were treated with PBS as controls. ResultsMiR-146a was upregulated in BMMSCs, and BMMSCs derived exosomes post miR-mimic transfection. Then in SCI rats, BMMSCs exosomes elevated the Basso, Beattie, and Bresnahan (BBB) score and reduced hematoxylin&eosin-reflected spinal cord tissue injury. In addition, BMMSCs exosomes did not affect TUNEL positive cells rate while increased NeuN(+) cells/field in spinal cord tissue from SCI rats. As for inflammation, BMMSCs exosomes repressed pro-inflammatory cytokine expressions, including interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α, in spinal cord tissue from SCI rats. Furthermore, miR-146a overexpressed BMMSCs exosomes presented with notably better effects regarding elevating BBB score in SCI rats and reducing tissue injury, neuron apoptosis, and inflammation while enhancing neuron viability in spinal cord tissue from SCI rats. ConclusionsMiR-146a overexpressed BMMSCs exosomes enhance locomotor capacity and neuron viability while reducing neuron apoptosis and spinal cord tissue inflammation in SCI rats.

MicroRNA 146a is associated with diabetic complications in type 1 diabetic patients from the EURODIAB PCS

BackgroundMicroRNA-146a-5p (miR-146a-5p) is a key regulator of inflammatory processes. Expression of miR-146a-5p is altered in target organs of diabetic complications and deficiency of miR-146a-5p has been implicated in their pathogenesis. We investigated if serum miR-146a-5p levels were independently associated with micro/macrovascular complications of type 1 diabetes (DM1).MethodsA nested case–control study from the EURODIAB PCS of 447 DM1 patients was performed. Cases (n = 294) had one or more complications of diabetes, whereas controls (n = 153) did not have any complication. Total RNA was isolated from all subjects and miR-146a-5p levels measured by qPCR. Both the endogenous controls U6 snRNA and the spike (Cel-miR-39) were used to normalize the results. Logistic regression analysis was carried out to investigate the association of miR-146a-5p with diabetes complications.ResultsMiR-146a-5p levels were significantly lower in cases [1.15 (0.32–3.34)] compared to controls [1.74 (0.44–6.74) P = 0.039]. Logistic regression analysis showed that levels of miR-146a-5p in the upper quartile were inversely associated with reduced odds ratio (OR) of all complications (OR 0.34 [95% CI 0.14–0.76]) and particularly with cardiovascular diseases (CVD) (OR 0.31 [95% CI 0.11–0.84]) and diabetic retinopathy (OR 0.40 [95% CI 0.16–0.99]), independently of age, sex, diabetes duration, A1c, hypertension, AER, eGFR, NT-proBNP, and TNF-α.ConclusionsIn this large cohort of DM1 patients, we reported an inverse and independent association of miR-146a-5p with diabetes chronic complications and in particular with CVD and retinopathy, suggesting that miR-146a-5p may be a novel candidate biomarker of DM1 complications.

Protective impact of Spirulina platensis against γ-irradiation and thioacetamide-induced nephrotoxicity in rats mediated by regulation of micro-RNA 1 and micro-RNA 146a.

Chronic kidney disease develops popular and medical health problems, especially in developing countries. The objective of this study is to investigate the protective mechanism of Spirulina platensis against γ-irradiation (R) and/or thioacetamide (TAA)-induced nephrotoxicity in rats. Rats intoxicated with R or TAA showed alterations in kidney function markers (urea, creatinine, albumin, and total protein contents), oxidative stress markers (malondialdehyde, reduced glutathione), antioxidant enzymes (superoxide dismutase, catalase), and several inflammatory markers (including, the high-sensitivity C-reactive protein, hypoxia-inducible factor-1 alpha, tumor necrosis factor-alpha, interferon-gamma, some interleukins, and nuclear factor-kappa B). Rats also acquired apoptosis, evinced by high caspase-3 efficacy. This nephrotoxicity mediated by upregulation of the messenger RNA (mRNA) gene expression of the autophagy markers: Beclin-1, microtubule-associated protein LC3, p62 binding protein, immunoglobulin G receptor Fcγ receptor (FcγR), micro-RNA-1 (miR-1), protein expression of phospho-adenosine monophosphate-activated protein kinase, and phospho-mammalian target of rapamycin, along with downregulation of miR-146a mRNA gene expression and alteration of calcium and iron levels. The combined treatment R/TAA enhanced the observed oxidative stress, inflammation, apoptosis, and autophagy that mediated by higher upregulation of miR-1 and downregulation of miR-146a mRNA gene expression. Spirulina platensis administration exhibited a nephroprotective impact on R, TAA, and R/TAA toxicities via regulating miR-1 and miR-146a mRNA gene expression that monitored adenosine monophosphate-activated protein kinase/mammalian target of rapamycin signaling.

Vildagliptin attenuates acetic acid-induced colitis in rats via targeting PI3K/Akt/NFκB, Nrf2 and CREB signaling pathways and the expression of lncRNA IFNG-AS1 and miR-146a

Inflammatory processes, including ulcerative colitis (UC), are associated with the increase in synthesis and release of pro-inflammatory cytokines. The release of these cytokines is regulated by phosphatidylinositol-3-kinase (PI3K)/protein kinase B (Akt)/nuclear factor-kappa B (NFκB) and cAMP response element-binding protein (CREB) signaling pathways as well as over expression of microRNA 146a (miR-146a) and long non-coding RNA interferon gamma antisense 1 (lncRNA IFNG-AS1). Vildagliptin (Vilda), a dipeptidyl peptidase IV (DPP-IV) inhibitor, has an anti-inflammatory, antioxidant and anti-apoptotic effects which were established in various models. However, its possible protective effect in UC has not been clarified. Hence, the current study aimed to explore the possible prophylactic effect of different doses of Vilda against acetic acid (AA)-induced colitis in rats. Forty-eight adult Wistar rats were divided into six groups: control, Vilda (10 mg/kg/day; p.o.), AA, AA + Vilda (5 mg/kg/day; p.o.), AA + Vilda (10 mg/kg/day; p.o.) and AA + sulfasalazine (Sulfa) (100 mg/kg/day; p.o.).Low- and high-dose Vilda showed significant improvement in the disease activity index (DAI) and macroscopic assessment markers. Vilda has markedly inhibited the expression of lncRNA IFNG-AS1 and miR-146a, as well as PI3K/Akt/NFκB pathway, while activated CREB and nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathways, and this was reflected in alleviated oxidative stress, inflammation and apoptosis. Such outcomes were more prominent with the high-dose Vilda versus low-dose Vilda and Sulfa. Moreover, the histological examination showed almost intact histological features in Vilda-treated groups when compared to AA group treated with saline. In conclusion, Vilda can be regarded as a new promising therapeutic alternative against UC.

Genetic polymorphisms associated with polycystic ovary syndrome among Iranian women

Polycystic ovary syndrome (PCOS) involves abnormalities in ovarian, reproductive, and metabolic systems. Genetic polymorphisms associated with individual differences and variations might be related to complex disorders with unknown causes, including PCOS. Several leading genetic markers with known cellular functions have been identified among Iranian women presenting with PCOS. In particular, the existing evidence shows a significant relationship between PCOS and the following genetic polymorphisms: rs2275913 (interleukin-17A), rs9927163 (interleukin-32), Pro12Ala (peroxisome proliferator-activated receptor-γ), rs17173608 (chemerin), rs2236242 (vaspin), ApaI (vitamin D receptor), and rs7895833 (sirtuin 1). In addition, a higher risk of PCOS is associated with the rs2910164 (microRNA 146a), rs2241766 (adiponectin), -34T/C (cytochrome 17), and rs1800682 (Fas) polymorphisms. Furthermore, protective effects against PCOS have been reported for the A4223C polymorphism of adenosine deaminase 1. Overall, the available data indicate that Iranian women with PCOS have a higher prevalence of polymorphisms in inflammation- and metabolism-related genes, but not in insulin-related genes. More extensive studies are needed to identify the ethnicity-related genetic associations in PCOS.

RANKL blockade alleviates peri-implant bone loss and is enhanced by anti-inflammatory microRNA-146a through TLR2/4 signaling

BackgroundThe present study was to determine the effect of local anti-RANKL antibody administration in the presence or absence of microRNA-146a on ligature-induced peri-implant bone resorption, and the potential role of TLR2/4 signaling in such effect.ResultsTitanium implants were placed in the left maxilla alveolar bone 6 weeks after extraction of first and second molars in C57/BL6 wild-type (WT) and TLR2−/− TLR4−/− (TLR2/4 KO) mice. Silk ligatures were tied around the implants 4 weeks after implantation. Anti-RANKL antibody (500 μg/mL) with or without microRNA 146a (miR-146a) (100 nM) was injected into palatal gingiva around implant on days 3, 6, and 9 during 2 weeks of ligation period. Bone resorption around the implants was assessed by 2D imaging using area measurement and 3D imaging using micro-computed tomography (μCT). Real-time quantitative PCR (RT-qPCR) was used to determine the peri-implant gingival mRNA expression levels of pro-inflammatory cytokines (TNF-α) and osteoclastogenesis-related cytokines (RANKL). In both WT and TLR2/4 KO mice, the bone resorption around implants was significantly increased in the ligation only group when compared to the non-ligation group, but TLR2/4 KO mice showed significantly less bone loss compared to WT mice after ligation. As expected, gingival injection of anti-RANKL antibody significantly reduced bone loss compared with the ligation only group in both WT and TLR2/4 KO mice. Moreover, injection of miR-146a in addition to anti-RANKL antibody significantly enhanced the inhibition of bone loss in WT mice but not in TLR2/4 KO mice. Gingival mRNA expressions of RANKL were significantly reduced by anti-RANKL antibody treatment in both WT and TLR2/4 KO mice but were not affected by the additional miR-146a treatment. Gingival mRNA expression of TNF-α was significantly reduced by miR-146a treatment in WT mice but not in TLR2/4 KO mice. The number of gingival inflammatory cell infiltration and peri-implant TRAP-positive cell formation was significantly reduced by the additional miR-146a treatment in WT mice but not in TLR2/4 KO mice.ConclusionsThis study suggests that anti-inflammatory miR-146a enhance anti-RANKL-induced inhibition of peri-implant bone resorption through the regulation of TLR2/4 signaling and inhibition of TNF-α expression.

MicroRNA-146a attenuates the development of morphine analgesic tolerance in a rat model

ABSTRACT Background: Morphine plays an irreplaceable role in relieving severe pain clinically, while long-term medication inevitably leads to drug resistance. MicroRNA (miR) 146a has been reported to be a negative regulator in the process of morphine-tolerance formation. This study aimed to investigate how miR-146a affects the development of morphine analgesic tolerance. Methods: The morphine-tolerance rat model was established by means of one-week continuous morphine administration. Paw withdrawal latency test was performed every day, and spinal cord samples were dissected on the seventh day for Q-PCR and Western blotting to detect the expression level of miR-146a, and IRAK1/TRAF6 participated in TLR4 signaling pathway. Results: The expression of miR-146 was significantly decreased in morphine-tolerant model. Also, overexpression of miR-146a reduced the resistance caused by morphine, followed by the down-regulation of IRAK1/TRAF6 in TLR4 pathway. The inhibition of miR-146a remarkably decreased paw withdrawal latency as well as increased the expression levels of TLR4 signaling pathway-related molecules, IRAK1 and TRAF6. Conclusion: This study suggests that miR-146a attenuates morphine tolerance by inhibiting the expression of IRAK1/TRAF6 in TLR4 pathway, which could provide an essential experimental basis for the settlement of morphine resistance-associated matters.

26-Year-Old Woman With Shortness of Breath and Chest Pain

26-Year-Old Woman With Shortness of Breath and Chest Pain

The expression of microRNA 146a in patients with ischemic stroke: an observational study.

AimWe conducted the present prospective study to assess the level of microRNA (miRNA) 146a in patients with ischemic stroke and its correlation with patients’ characteristics.MethodsWe conducted an observational study that included adult patients (≥18 years old) who presented within 24 hrs after the onset of the symptoms of acute ischemic stroke. In addition, age- and sex-matched healthy volunteers were included as control group. The primary outcome in the present study was the difference in miRNA 146a expression between patients with ischemic stroke and control group participants. The expression of miRNA 146a was measured using quantitative real-time PCR. Quantitative real-time PCR amplification and analysis were performed using Rotor-Gene Q thermal cycler.ResultsThe present study included 44 patients with ischemic stroke and 22 matched controls. Regarding the primary outcome of the present study, the median expression of miRNA 146a in patients with ischemic stroke was −1.98 fold (IQR −27.1–3.9) compared to 1.75 fold (IQR −2.25–5.27) in control group (P<0.001). However, the subgroup analysis showed that the expression of miRNA 146a was significantly downregulated in comatosed patients only (P<0.001). The expression of miRNA 146a correlated negatively with Glasgow Coma Scale score in comatose patients (r=−0.352, P=0.022).ConclusionIn conclusion, the expression of miRNA 146a is significantly downregulated in ischemic stroke patients. Further studies are needed to assess its diagnostic utility and therapeutic potentials.

Increased matrix metalloproteinases expression in tuberous sclerosis complex: modulation by microRNA 146a and 147b in vitro.

AimMatrix metalloproteinases (MMPs) and their endogenous tissue inhibitors (TIMPs) control proteolysis within the extracellular matrix (ECM) of the brain. Dysfunction of this enzymatic system due to brain inflammation can disrupt the blood‐brain barrier (BBB) and has been implicated in the pathogenesis of epilepsy. However, this has not been extensively studied in the epileptogenic human brain.MethodsWe investigated the expression and cellular localization of major MMPs (MMP2, MMP3, MMP9 and MMP14) and TIMPs (TIMP1, TIMP2, TIMP3 and TIMP4) using quantitative real‐time polymerase chain reaction (RT‐PCR) and immunohistochemistry in resected epileptogenic brain tissue from patients with tuberous sclerosis complex (TSC), a severe neurodevelopmental disorder characterized by intractable epilepsy and prominent neuroinflammation. Furthermore, we determined whether anti‐inflammatory microRNAs, miR146a and miR147b, which can regulate gene expression at the transcriptional level, could attenuate dysregulated MMP and TIMP expression in TSC tuber‐derived astroglial cultures.ResultsWe demonstrated higher mRNA and protein expression of MMPs and TIMPs in TSC tubers compared to control and perituberal brain tissue, particularly in dysmorphic neurons and giant cells, as well as in reactive astrocytes, which was associated with BBB dysfunction. More importantly, IL‐1β‐induced dysregulation of MMP3, TIMP2, TIMP3 and TIMP4 could be rescued by miR146a and miR147b in tuber‐derived TSC cultures.ConclusionsThis study provides evidence of dysregulation of the MMP/TIMP proteolytic system in TSC, which is associated with BBB dysfunction. As dysregulated MMP and TIMP expression can be ameliorated in vitro by miR146a and miR147b, these miRNAs deserve further investigation as a novel therapeutic approach.

Olive Leaf Extract Attenuates Inflammatory Activation and DNA Damage in Human Arterial Endothelial Cells.

Olive leaf extract (OLE) is used in traditional medicine as a food supplement and as an over-the-counter drug for a variety of its effects, including anti-inflammatory and anti-atherosclerotic ones. Mechanisms through which OLE could modulate these pathways in human vasculature remain largely unknown. Serum amyloid A (SAA) plays a causal role in atherosclerosis and cardiovascular diseases and induces pro-inflammatory and pro-adhesive responses in human coronary artery endothelial cells (HCAEC). Within this study we explored whether OLE can attenuate SAA-driven responses in HCAEC. HCAEC were treated with SAA (1,000 nM) and/or OLE (0.5 and 1 mg/ml). The expression of adhesion molecules VCAM-1 and E-selectin, matrix metalloproteinases (MMP2 and MMP9) and microRNA 146a, let-7e, and let-7g (involved in the regulation of inflammation) was determined by qPCR. The amount of secreted IL-6, IL-8, MIF, and GRO-α in cell culture supernatants was quantified by ELISA. Phosphorylation of NF-κB was assessed by Western blot and DNA damage was measured using the COMET assay. OLE decreased significantly released protein levels of IL-6 and IL-8, as well as mRNA expression of E-selectin in SAA-stimulated HCAEC and reduced MMP2 levels in unstimulated cells. Phosphorylation of NF-κB (p65) was upregulated in the presence of SAA, with OLE significantly attenuating this SAA-induced effect. OLE stabilized SAA-induced upregulation of microRNA-146a and let-7e in HCAEC, suggesting that OLE could fine-tune the SAA-driven activity of NF-κB by changing the microRNA networks in HCAEC. SAA induced DNA damage and worsened the oxidative DNA damage in HCAEC, whereas OLE protected HCAEC from SAA- and H2O2-driven DNA damage. OLE significantly attenuated certain pro-inflammatory and pro-adhesive responses and decreased DNA damage in HCAEC upon stimulation with SAA. The reversal of SAA-driven endothelial activation by OLE might contribute to its anti-inflammatory and anti-atherogenic effects in HCAEC.

Challenges in the treatment of Rheumatoid Arthritis.

Rheumatoid Arthritis (RA) is a chronic inflammatory disease characterized by a heterogeneous clinical response to the different treatments. Some patients are difficult to treat and do not reach the treatment targets as clinical remission or low disease activity. Known negative prognostic factors, such as the presence of auto-antiantibodies and joint erosion, the presence of a genetic profile, comorbidities and extra-articular manifestations, pregnancy or a pregnancy wish may concur to the treatment failure. In this review we aimed at identify difficult to treat RA patients and define the optimal therapeutic and environmental targets. Genetic markers of severity such as HLA-DRB1, TRAF1, PSORS1C1 and microRNA 146a are differently associated with joint damage; other gene polymorphisms seem to be associated with response to biologic disease modifying anti-rheumatic drugs (bDMARDs). The presence of comorbidities and/or extra-articular manifestations may influence the therapeutic choice; overweight and obese patients are less responsive to TNF inhibitors. In this context the patient profiling can improve the clinical outcome. Targeting different pathways, molecules, and cells involved in the pathogenesis of RA may in part justify the lack response of some patients. An overview of the future therapeutic targets, including bDMARDs (inhibitors of IL-6, GM-CSF, matrix metalloproteinases, chemokines) and targeted synthetic DMARDs (filgotinib, ABT-494, pefacitinib, decernotinib), and environmental targets is addressed. Environmental factors, such as diet and cigarette smoke, may influence susceptibility to autoimmune diseases and interfere with inflammatory pathways. Mediterranean diet, low salt intake, cocoa, curcumin, and physical activity seem to show beneficial effects, however studies of dose finding, safety and efficacy in RA need to be performed.

Role of MicroRNA 146a in Regulating Regulatory T Cell Function to Ameliorate Acute Cardiac Rejection in Mice

BackgroundMiR-146a plays a critical regulatory role in the homeostasis and function of regulatory T cells (Treg cells). The purpose of this study was to investigate the role of miR-146 in regulating Treg function to ameliorate acute cardiac rejection in mice. MethodsWe downregulated the miR-146a expression of Treg cells in recipients and constructed murine heterotopic heart transplantation models. Flow cytometry was used to analyze phenotypes of T cells. Graft rejection and cytokine changes during allograft rejection were detected. The changes of miR-146a target genes in Treg cells were detected by quantitative real-time polymerase chain reaction and western blotting. ResultsOur results indicated the miR-146a antagomir increased the Treg proportion in splenocytes and blood cells in the normal immune and inflammatory conditions, but impaired the ability of Treg cells to inhibit Th1 in the inflammatory condition. During the rejection process of murine heterotopic heart transplantation, miR-146a was mainly involved in regulating Treg cells through the STAT1/IFN-γ signaling pathway and the target gene was STAT1. Reducing the miR-146a of Treg and simultaneously collaboratively applying IFN-γ neutralizing antibodies can inhibit the proliferation of CD4+ T cell, alleviate the rejection process, and promote the survival of the donor's heart. ConclusionIt is possible to enhance the function of Treg cells in vivo of recipient by regulating a single miRNA, miR-146a, to achieve the purpose of inhibiting graft rejection. Collaboratively applying immunosuppressive drugs may effectively control the immune rejection and allow for the dosage and side effects of drugs to be reduced, a process that needs further study.