miR-146a Levels Research Articles

Potential role of circulating miRNA-146a and serum kallikrein 1 as biomarkers of renal disease in biopsy-proven lupus nephritis patients

Aim of the workTo evaluate the performance of circulating miR-146a and serum kallikrein1 (KLK1) as biomarkers for detecting disease activity and distinguishing histopathological changes associated with lupus nephritis (LN). Patients and methods90 systemic lupus erythematosus (SLE) patients (44 with active non-renal disease and 46 with LN) were examined for relative expression levels of circulating miR-146a, assessed by reverse transcription quantitative real-time PCR (RT-qPCR) and serum KLK1 levels, measured by ELISA. SLE disease activity was assessed using SLEDAI-2 k. Receiver operating characteristic curve was employed to examine the diagnostic performance of both biomarkers and established conventional laboratory parameters of renal function with specific renal histologic features seen in LN. As well, 40 apparently healthy control subjects were enrolled. ResultsCirculating miR-146a levels were significantly lower (p < 0.01), while serum KLK1 levels were significantly higher (p < 0.01) in active LN patients compared to active non-renal and controls. miR-146a levels negatively correlated with serum creatinine, Albumin/Creatinine and SLEDAI-2 k (p ≤ 0.01), while serum KLK1 significantly correlated with serum creatinine, Albumin/Creatinine and C3 (p ≤ 0.05) and inversely with miR-146a (p = 0.002). Both circulating miR-146a and KLK1 revealed an outstanding performance in predicting renal activity among SLE patients with an AUC of 0.949 and 0.947 respectively. In active LN patients, KLK1 and miR-146a exhibited good ability in differentiating proliferative from non-proliferative LN (AUC 0.73, 0.65 respectively), and in predicting biopsy activity index ≥7, surpassing conventional disease measures. ConclusionCirculating miR-146a and serum KLK1 are potential biomarkers for LN that could compete with invasive renal biopsy. Independent validation is warranted.

A low carbohydrate diet high in fish oil and soy protein delays inflammation, hematopoietic stem cell depletion, and mortality in miR-146a knock-out mice.

Since our previous studies found a low carbohydrate (CHO) diet containing soy protein and fish oil (15%Amylose/Soy/FO) significantly reduced lung and breast cancer in mice we asked herein if this low CHO diet could also delay the onset of myeloid malignancies. To test this we employed a miR-146a knock-out (KO) mouse model and found the 15%Amylose/Soy/FO diet increased their median lifespan by 8.5 month, compared to these mice on a Western diet. This was associated with increased lymphocytes and reduced monocytes, granulocytes, blood glucose and insulin levels. Inflammatory cytokine/chemokine studies carried out with 6-month-old mice, before any signs of illness, revealed the 15%Amylose/Soy/FO diet significantly reduced pro-inflammatory cytokines. This low CHO diet also led to an increase in plasma β-hydroxybutyrate and in liver fatty acid synthase levels. This, together with higher liver carnitine palmitoyltransferase I levels suggested that the 15%Amylose/Soy/FO diet was causing a systemic metabolic shift from glucose to fatty acids as an energy source. Lastly, we found the 15%Amylose/Soy/FO diet resulted in significantly higher numbers of primitive hematopoietic stem cells (HSCs) in the bone marrow of 6-month-old mice than those fed a Western diet. Taken together, these results suggest a 15%Amylose/Soy/FO diet reduces chronic inflammation and increases fatty acid oxidation and that this, in turn, may prevent HSC proliferation and exhaustion, thereby delaying myeloid malignancy-induced death of miR-146a KO mice. We suggest a low CHO diet containing soy protein and fish oil could be beneficial in reducing the risk of myeloid malignancies in patients with low miR-146a levels.

Total oxidant capacity, total antioxidant capacity, ischemic modified albumin, microRNA levels, and their relationship with psoriasis area and severity index.

Aims To examine the differences in the levels of microRNA, ischemic modified albumin (IMA), total oxidant capacity (TOC), and total antioxidant capacity (TAC) of persons with and without psoriasis and, in the case group, the relationship between these parameters and psoriasis area and severity index (PASI). Methods Blood samples were collected from patients and healthy participants to examine levels of these parameters. Results The mean serum TOC level was higher in the case group. The mean serum TAC and IMA levels were significantly lower in the case group (P <0.001). It was observed that the mean serum miR-203 and miR-146a levels were increased in psoriasis patients. It was determined that there was only a significant positive weak correlation between miR-203 and PASI (r = 0.232, P = 0.027). Limitations The small sample size, not controlling serum albumin and not evaluating the effects of the treatment agents used by the patients on oxidative and inflammatory processes. Conclusion In the case group changes in the mean serum TOC and TAC levels provide evidence that oxidative stress may play a critical role in disease pathogenesis. The increase in the mean serum miR-203 and miR-146a levels suggest the possibility of therapies targeting these microRNAs as a new option.

MiR-9 and miR-181a Target Gab2 to Inhibit the Proliferation and Migration of Hepatocellular Carcinoma HepG2 Cells.

The incidence of liver cancer ranks seventh globally, with nearly half of all cases occurring in East Asia, but currently, there are very few drugs to treat it. Our previous studies demonstrated that the signal integration protein Gab2 is a potential drug target for the prevention and therapy of liver cancer. Here, we screened for and identified two miRNAs that target Gab2 to suppress the proliferation and migration of hepatocellular carcinoma (HCC) cells. First, we predicted Gab2-targeting miRNAs through biological websites, and we selected nine miRNAs that were reported in the literature as being abnormally expressed in liver cancer and fatty liver tissue. Then, we measured the expression of these miRNAs in the hepatic epithelial cell line HL-7702 and the HCC cell line HepG2. The expression levels of miR-9, miR-181a, miR-181c, miR-34a, and miR-134 were high in HL-7702 cells but low in HepG2 cells, and their expression patterns were the opposite of Gab2 in these cells. Furthermore, we transfected miR-9, miR-34a, miR-181a, and miR-181c mimics into HepG2 cells and found that only miR-9 and miR-181a reduced the level of Gab2 proteins. miR-9 also reduced the Gab2 mRNA level, but miR-181a did not affect the Gab2 mRNA levels. Using a miRNA-Gab2 3'UTR binding reporter, we confirmed that miR-9 and miR-181a bind to the Gab2 3'UTR region. Finally, we introduced miR-9 and miR-181a mimics into HepG2 cells and found that cell proliferation and migration were significantly inhibited. In conclusion, we identified two novel miRNAs targeting Gab2 and provided potential drug targets for the prevention and treatment of liver cancer.

Identification of Biomarkers As Predictors of Patient-Reported Outcomes in Persons with Multiple Sclerosis Given Autologous Hematopoietic Stem Cell Transplantation

Introduction: Around 80% of persons with multiple sclerosis (MS) have a positive patient-reported outcome (PRO) to autologous hematopoietic stem cell transplantation (aHSCT); this figure is better than that obtained with the novel immunossuppresive drugs. Objective: To identify markers for the appropriate selection of patients with multiple sclerosis who would benefit from aHSCT. Methods: We evaluated the levels of six biomarkes in the peripheral blood of patients with MS prior to aHSCT. Each biomarker was selected according to its possible role in MS: microRNAs (miR-146a, miR-155 and miR-326) as immune system alteration markers; interleukin 4 induced 1 (IL4I1) as remyelination marker; neurofilament light chain protein (NFL) as neuroaxonal damage marker; and the human leukocyte antigen (HLA) allele HLA-DRB1*15 as a MS risk marker. The design of this study is cross-sectional; sample was divided into two transplant response groups (responders and non-responders). The response to transplantation was assessed using the PRO (expanded disability status scale -EDSS) score, evaluated before aHSCT and one year later. Pre-transplant samples were used to identify the different markers. The techniques used for markers analysis were qPCR for microRNA measurement (miR-146a, miR155, miR326), SSP-PCR for HLA-DRB1*15, and immunoassay for IL4I1 and NFL Results: 34 patients were prospectively enrolled. 14 (41.2%) non-responders to transplantation (ΔEDSS < 0) and 20 (58.8%) with a positive response to transplantation (ΔEDSS > 0). The comparison made between the aHSCT PRO response groups, using the Mann-Whitney U statistical test, showed a statistically significant increase in the non-responder group in the following variables: hemoglobin (median = 14.9 vs median = 13.5) U = 81.5, p = .039; serum iron levels (median = 122.5 vs median = 91) U = 39, p =.04 and transferrin saturation (median = 33.94 vs median = 20.4) U = 30, p = .001. In addition, an increase in platelet count was observed in the good response group (median = 278.5 vs median = 229) U = 197.5, p = .043. Within the biomarkers evaluated, significant difference was only found in miR-146a levels, which showed an increase for the non-responder group (median = 6. 43 vs median = 3) U = 78, p = .03. We then performed a logistic regression analysis using the variables hemoglobin, serum iron levels, platelet count, miR146a, miR155, miR326, IL4I1 and NFL, to evaluate their predictive capacity in the response to transplantation. The logistic region model was configured to perform forward variable selection obtaining the following results: in the first step, 74.2% correct overall classification with the variable Iron; in the second step 82.8%, with the variables iron and hemoglobin; in the third step 89.7%, with the variables hemoglobin, serum iron levels and IL4I1; and in the third step 100%, with the variables hemoglobin, iron levels, IL4I1 and miR146a. Conclusions: Serum iron levels, hemoglobin levels, transferrin saturation and platelet count before aHSCT prior to transplantation show differences between PRO responders and non-responders to aHSCT; these variables coupled with the biomarkers IL4I1 and miR146a can be employed in predictive models of the PRO positive response to aHSCT in persons with MS.

Dual Inhibition of NF-Kb and JAK/STAT Pathways Reverts Myelofibrosis-like Phenotype in an In Vivo Murine Model

Introduction: In JAK-STAT-mediated inflammatory cytokine production in myeloproliferative neoplasms (MPN), both malignant and non-malignant clones contribute. The NF-κB pathway is a mediator of inflammation-induced carcinogenesis. We have previously demonstrated the role of miR-146a, a NF-κB repressor through IRAK1/TRAF6, in the progression of MPN. Indeed, the rs2431697 TT genotype, associated with lower miR-146a levels, contributes to early progression to secondary myelofibrosis (MF); and miR-146a-/- (KO) mice, with MF-like phenotype, show an increase in p-Stat3 with aging (Ferrer-Marin F. et al. Leukemia 2020). Here, we postulate that in this inflammation-driven MF murine model, inhibition of both NF-κB and JAK/STAT pathways can synergistically dampen the inflammatory state and reverse the MF phenotype. Methods: 11-month-old miR-146a-/- micewere 1-month-oral ad libitum treated with either ruxolitinib (JAK1/2 inhibitor) at 75 mg/kg/day, BMS-345541 (BMS) (NF-κB pathway inhibitor at IKKα/β level) at 100 mg/kg/day, both inhibitors at the same concentrations, or pacritinib (JAK2 and NF-κB pathway inhibitor at IRAK1 level) at 150 mg/kg/day. Untreated KO and wild-type (WT) mice were used as controls (n=7-15 mice/group). We evaluated: cell counts in peripheral blood; proinflammatory plasma cytokines by Luminex; size, weight and myeloid (CD11b) and lymphoid (CD19) populations by flow cytometry in spleen; histology of spleen and bone marrow (BM) by hematoxylin-eosin and reticulin staining. SET2 cells were used as an in vitro human model of JAK2V617F clonal hematopoiesis for immunoblotting quantification of pTyrSTAT3 (JAK/STAT pathway) and IRAK1 and nuclear p-p65 (NF-κB pathway). Results: As expected, compared to WT mice, KO mice exhibited monocytopenia, anemia and thrombocytopenia (Table 1), splenomegaly and severe reticulin fibrosis with osteosclerosis in BM (Fig.1). Moreover, in line with their inflammatory phenotype, plasma IL6, IL1β and TNFα levels were elevated in KO (vs WT) mice (Table 1). Besides splenomegaly, KO mice had spleen architecture loss, decreased white pulp and increased megakaryocytes (Fig.1), as a sign of extramedullary hematopoiesis. Consistent with histology, at a cellular level, KO mice showed significantly increased myeloid (CD11b+) and decreased lymphoid (CD19+) populations in the spleen. NF-κB pathway inhibitors (BMS and pacritinib) improved the thrombocytopenia, but only pacritinib improved monocyte counts without worsening anemia; by contrast, ruxolitinib did not improve monocytes or platelet counts, and the combination of ruxolitinib/BMS worsened anemia (Table 1). Regarding the proinflammatory cytokines, IL6 levels were significantly decreased with ruxolitinib; IL1β levels decreased whenever NF-κB pathway was inhibited, although it only showed significance with BMS; and TNFα levels significantly decreased with all treatments, with only a tendency for ruxolitinib/BMS (Table 1). Although all treatments decreased splenomegaly and partially recovered spleen architecture, only pacritinib and BMS (NF-κB inhibitors) diminished splenic megakaryocytic counts (Fig.1). NF-κB inhibitors [pacritinib and BMS (+/- ruxolitinib)] decreased the number of mice with severe reticulin fibrosis or moderate/severe osteosclerosis in the BM, whereas ruxolitinib alone did not have this effect. Unlike pacritinib, however, the combination of ruxolitinib/BMS induced BM aplasia (Fig.1). Finally, in SET2 cells, monotherapy with ruxolitinib or BMS inhibited JAK-STAT or NF-κB signaling pathways, respectively, but only pacritinib and the combination were able to inhibit both pathways. Conclusions: miR-146a-/- mice constitute a useful model of MPN without driver mutations to evaluate the role of inflammation in these neoplasms. The dual inhibition of NF-κB and JAK/STAT with pacritinib reduced inflammatory cytokines, reverted monocytopenia and thrombocytopenia, and improved BM fibrosis without worsening anemia. While some of these benefits could be recapitulated in vivo by combining ruxolitinib with an NF-κB inhibitor, this combination resulted in worsening anemia and BM aplasia. These results highlight the therapeutic possibilities of dual JAK/NF-κB inhibitors, such as pacritinib, in patients with MF. Funding: PI18/00316; PI20/00136; PI21/00347; 20644/JLI/18; FPU18/03189. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Regulating Function of miR-146a Derived from Bone Marrow Mesenchymal Stem Cell (BMSC) in Acute Lung Injury

This study assesses the mechanism of miR-146a derived from Bone marrow mesenchymal stem cell (BMSC) in acute lung injury. The model of ALI rats was established through endotracheal perfusion of LPS followed by analysis histological changes by HE staining. The source of BMSC was detected through flow cytometry and change of miRNA was detected through Array method. The miR-146a level in lung tissue was detected with RT-PCR and expression of Bcl-2, Bax and Capase-9 was detected with IF and Western Blot. A high expression of CD90 and CD105 was found in BMSC with negative CD11bc and CD34 level. 39 downregulated miRNAs and 20 upregulated miRNAs were found in ALI with miR-146a being the most significant. The apoptotic level induced with LPS could be restrained by miR-146a. In addition, miR-146a could upregulate Bcl-2 and downregulate Bax and Caspase-9. In conclusion, ALI could be restrained by the low expression of miR-146a.

Experimental Study of Using Aloe Vera Gel to Treat Diabetic Chronic Cutaneous Ulcers in Bama Miniature Pigs

To investigate the efficacy, safety, and mechanism of topical application of aloe vera gel (AVG) to treat diabetic chronic cutaneous ulcers in Bama miniature pigs. The Bama miniature pig model of diabetic chronic skin wounds was constructed and the model pigs were randomly assigned to AVG daily administration group (AVG QD), aloe vera gel every-other-day administration group (AVG QOD), and diabetic control group (DC). A non-diabetic chronic skin wounds model pig was set as the non-diabetic control group (NDC). Treatment efficacy was evaluated based on the amount of time needed for complete healing of the wounds, healing rates, granulation growth rates, and skin histopathological changes. Safety was evaluated according to whether adverse reactions were observed. In addition, the dynamic changes of the relative expression levels of miR21, miR29a, miR126, miR146a, miR155, and miR210 in wound granulation tissues were examined. 1) Efficacy and safety: The amount of time needed for complete healing of the wounds was shorter in the NDC group than those of the three other groups, DC group, AVG QD group, and AVG QOD group (all P<0.05). The amount of time needed for complete healing of the wounds was shorter in the AVG QD group and AVG QOD group than that of DC group (all P<0.05). The amount of time needed for complete healing of the wounds was shorter in the AVG QOD group than that of AVG QD group (all P<0.05). No adverse reactions were detected in the whole process of AVG topical treatment. The granulation growth rate of NDC group was higher than those of DC group, AVG QD group, and AVG QOD group (all P<0.05). The wound healing rate of NDC group was higher than those of DC group, AVG QD group, and AVG QOD group (all P<0.05); the wound healing rate of AVG QOD group was higher than those of DC group and AVG QD group (all P<0.05). 2) Histopathology: The results of HE staining light microscopy showed that collagen fiber production increased, and that microvascular formation with slight inflammatory cell infiltration was observed in the dermal interstitium at the initial stage of wound healing after AVG treatment. One year of after complete healing, pathological examination results of wound healing skin showed that the epidermal keratinization was complete, that collagen was arranged neatly and orderly, and that many microvessels were found in the interstitium. The results of picric acid celestite scarlet staining showed that, after AVG treatment, type Ⅰ collagen mainly increased in the initial stage of wound healing, type Ⅲ collagen gradually increased when the wound healed completely, and the collagen was arranged neatly during the whole process. 3) The relative expression of microRNAs: The relative expression of miR21, miR126, and miR210 in NDC group, AVG QD group, and AVG QOD group were higher than that in DC group (all P<0.05). The relative expression of miR29a and miR155 in NDC group, AVG QD group, and AVG QOD group was lower than that in DC group (all P<0.05). The relative expression of miR146a in NDC group was higher than that in DC group ( P<0.05). AVG topical application can shorten the time needed for complete healing of diabetic chronic wounds in Bama minipigs. The wound healing speed of the alternate-day treatment group was faster than that of the daily treatment group. No adverse reactions were observed over the course of the treatment. The mechanism may be related to the up-regulation of the expressions of miR21, miR126, and miR210 and the down-regulation of miR29a and miR155 in wound granulation tissue.

The Expression of Mirnas in Sudanese Patients with Systemic Lupus Erythematosus in Khartoum State

Background: MicroRNAs (miRs) are noncoding gene regulators that may have a role as diagnostic or prognostic biomarkers in systemic lupus erythematosus (SLE). Aim: To measure the blood levels of miR-146a, miR-126 and miR-30a in Sudanese SLE patients and to investigate their potential role in disease pathogenesis and utility as biomarkers for SLE. Material and Methods: A total of 48 SLE patients and 20 matched healthy individuals were enrolled in this study. SLE disease activity index (SLEDAI) was assessed. The blood levels of miR-146a, miR-126 and miR-30a were determined by Real-time polymerase chain reaction (PCR) in all participants. For γ-INF and IL-2 were analyzed by ELISA, and CD markers were used flowcytometery. Results: The mean age of the patients was 31.5 ± 8.5 years with disease duration &gt;5 years. In SLE patients, the mean blood level fold changes of miR-146a (0.33±0.277; P&lt;0.001), miR-126 (2.44±1.771; P=0.007) and miR-30a (1.56±1.40; P &gt;0.305) compared to controls. Down regulation of miR-146a increase expression of γ-INF (p &lt; 0.002), whereas the up regulation of miR-126 increase expression of CD markers (p&lt;0.000) in SLE patients. MiR-126 at a cut-off value 1.209 and miR-146a at cut-off value 0.9233 which can discriminate between SLE patients with significantly associated to SLE disease. Conversely, miR-30a was insignificantly associated with SLE disease (p value &gt;0.05) as no differences between the SLE patients and healthy control. Conclusion: Circulating miR-146a and miR-126 could be a potential noninvasive biomarker in SLE. This study providing an overview of the current state of research on the role of miRNAs in the immune pathogenesis and regulation of SLE. Further studies are needed in miRNAs profiling expressions of SLE diseases.

Expression and mechanism of micro-ribonucleic acid-29a in chorionic villi obtained at curettage with recurrent abortion

Objective: Explored the expression of miR-29a in puerpera with RSA and its influencing mechanism. Method: 52 patients with RSA group were divided into 30 cases representing ≤3 abortions and 22 cases with ≥4 abortions,thirty healthy women who had induced abortion during the same period as the control group. The differences in the expression levels of miR-29a, FKBP52 mRNA, VEGF, MVD, and HIF-lα were compared between the groups by conducting a correlation analysis. Results: The expression levels of miR-29a, VEGF, MVD, and HIF-lα in the chorionic villus were significantly higher among patients in the group with ≥4 abortions than in those in the group with ≤3 abortions (P < 0.05), and the expression levels of FKBP52 mRNA were lower in the former than in the latter (P < 0.05). A Spearman correlation analysis revealed that the expression levels of miR-29a were positively correlated with the levels of VEGF, MVD, and HIF-lα (P < 0.05) and negatively correlated with the expression level of FKBP52 mRNA (P < 0.05). Conclusion: MiR-29a may be involved in the pathogenesis of RSA by inhibiting the protein expressions of FKBP52 and VEGF, promoting the apoptosis of trophoblasts, and impairing neovascularization, resulting in placental vascular dysplasia..

Deficiency of miR-29a/b1 leads to premature aging and dopaminergic neuroprotection in mice.

Parkinson’s disease (PD) is a neurodegenerative disorder characterized by progressive degeneration of midbrain dopaminergic neurons. The miR-29s family, including miR-29a and miR-29b1 as well as miR-29b2 and miR-29c, are implicated in aging, metabolism, neuronal survival, and neurological disorders. In this study, the roles of miR-29a/b1 in aging and PD were investigated. miR-29a/b1 knockout mice (named as 29a KO hereafter) and their wild-type (WT) controls were used to analyze aging-related phenotypes. After challenged with the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), dopaminergic injuries, glial activation, and mouse behaviors were evaluated. Primary glial cells were further cultured to explore the underlying mechanisms. Additionally, the levels of miR-29s in the cerebrospinal fluid (CSF) of PD patients (n = 18) and healthy subjects (n = 17) were quantified. 29a KO mice showed dramatic weight loss, kyphosis, and along with increased and deepened wrinkles in skins, when compared with WT mice. Moreover, both abdominal and brown adipose tissues reduced in 29a KO mice, compared to their WT counterpart. However, in MPTP-induced PD mouse model, the deficiency of miR-29a/b1 led to less severe damages of dopaminergic system and mitigated glial activation in the nigrostriatal pathway, and subsequently alleviated the motor impairments in 3-month-old mice. Eight-month-old mutant mice maintained such a resistance to MPTP intoxication. Mechanistically, the deficiency of miR-29a/b-1 promoted the expression of neurotrophic factors in 1-Methyl-4-phenylpyridinium (MPP+)-treated primary mixed glia and primary astrocytes. In lipopolysaccharide (LPS)-treated primary microglia, knockout of miR-29a/b-1 inhibited the expression of inflammatory factors, and promoted the expression of anti-inflammatory factors and neurotrophic factors. Knockout of miR-29a/b1 increased the activity of AMP-activated protein kinase (AMPK) and repressed NF-κB/p65 signaling in glial cells. Moreover, we found miR-29a level was increased in the CSF of patients with PD. Our results suggest that 29a KO mice display the peripheral premature senility. The combined effects of less activated glial cells might contribute to the mitigated inflammatory responses and elicit resistance to MPTP intoxication in miR-29a/b1 KO mice.

Evaluation of the effects of immunization against PCSK9 on selected microRNAs involved in cholesterol homeostasis

Abstract Background MicroRNAs (miRNAs) have been discovered as novel modulators of gene expression. These molecules are involved in regulating lipid metabolism through a complicated interactive mechanism involving gene regulatory networks and can play a role in metabolic diseases. PCSK9 (proprotein convertase subtilisin/Kexin 9) is a critical regulator in cholesterol homeostasis. PCSK9 inhibition is an effective therapeutic strategy for the treatment of dyslipidemia. Accordingly, it is essential to identify the controlling mechanisms of the PCSK9 and the effects of its inhibitors on microRNAs involved in cholesterol homeostasis. Purpose Considering the role that miR-27a, miR-30c, and miR-191 play in regulating genes involved in cholesterol homeostasis, we investigated the effect of immunization against PCSK9 on the mentioned miRNAs. Methods A PCSK9 immunogenic peptide constructed by linking a short PCSK9 mimotope peptide to a tetanus peptide was used for immunization. The PCSK9 immunogenic peptide construct was mixed homogeneously with 0.4% alum adjuvant at a ratio of 1:1 for use as vaccine formulation. Mice in the vaccinated group were inoculated subcutaneously in four terms with bi-weekly intervals. In order to assess the expression of miRNAs in vaccinated mice' livers, sampling was carried out two weeks after the last injection. The levels of miR-27a, miR-30c, and miR-191 expression in the liver of vaccinated mice were evaluated using real-time quantitative PCR, and relative expression levels were presented as fold change normalized to U6 as the internal control. Results Hepatic miR-27a expression was significantly decreased in vaccinated mice compared with the control mice (fold change: 0.731±0.1, p=0.027). There was a borderline nonsignificant reduction in the hepatic expression of miR-30c in the vaccine group compared to the control group (fold change: 0.569±0.1, p=0.078). No significant difference was detected in the hepatic expression of miR-191 between the vaccine and control groups (fold change: 0.852±0.1, p=0.343). Conclusions Immunization against PCSK9 leads to decreased hepatic expression of miR-27a. Since miR-27a is involved in the regulation of targets like angiopoietin-like 3 and glycerol-3-phosphate acyltransferase 1, its reduction may explain part of the lipid-lowering activity of PCSK9 vaccine. Funding Acknowledgement Type of funding sources: None.

Clinical Study of miRNAs Derived from Serum Exosomes in Multiple Myeloma

To investigate the expression level and prognostic value of miR-21, miR-18a, miR-146a, and Let-7b derived from serum exosomes in patients with multiple myeloma (MM). Serum exosomes were extracted from 57 MM patients and 20 healthy persons using ExoQuick exosome precipitation solution kit, and the relative expression level of miR-21, miR-18a, miR-146a, and Let-7b derived from serum exosomes was measured by RT-qPCR. Correlations of the expression levels of all miRNAs mentioned above with routine laboratory parameters were analyzed by Spearman correlation analysis. The relationship between the expression level of miR-21, miR-18a, miR-146a, and Let-7b derived from serum exosomes and overall survival of patients with MM was analyzed using the Kaplan-Meier survival curve. The expression levels of miR-21, miR-18a, and Let-7b derived from serum exosomes in patients with MM were significantly lower than those in the normal control group (P<0.001), while the expression level of miR-146a between the two groups was not significantly different (P>0.05). The expression level of miR-21 was strongly negatively correlated with serum β2-microglobulin concentration (r=-0.830), and weakly negatively correlated with serum creatinine, corrected serum calcium, and cystatin C (r=-0.488, -0.282, -0.627). The expression levels of Let-7b and miR-18a were also weakly negatively correlated with the corrected serum calcium, β2-microglobulin, and cystatin C concentration (r=-0.305, -0.362, -0.461; -0.317, -0.542, -0.434). However, there was no significant correlation between the expression level of miR-146a and routine laboratory parameters in MM patients. The overall survival rate of MM patients with low expression level of miR-21, miR-18a, and Let-7b significantly decreased compared with high expression level group (P<0.05), however, the expression level of miR-146a was not related to the overall survival rate. Aberrant low expression levels of miR-21, miR-18a, and Let-7b derived from serum exosomes exist in patients with MM, which are associated with a worse overall survival rate.

Effects of crocin on T-bet/GATA-3 ratio, and miR-146a and miR-106a expression levels in lung tissue of ovalbumin-sensitized mice.

Objective(s):Although various studies have revealed the beneficial effects of crocin (derived from saffron), such as anti-inflammatory, anti-cancer, antioxidant, and immune modulator, however, its exact mechanism is unknown. The present study aimed to investigate the effect of crocin on the expression ratio of T-bet/GATA-3 as an indicator of altered immune responses in the lung tissue of ovalbumin (OVA)-sensitized mice. In addition, the effect of crocin on the expression level of miR-146a and miR-106a in the lung tissue OVA-sensitized mice was investigated.Materials and Methods:Mice were randomly divided into five groups (n=6): Control; OVA, OVA + Crocin 25, OVA + Cro 50, and OVA + Cro100 groups. Crocin was administrated intraperitoneally at doses of 25, 50, and 100 mg/kg for five consecutive days. One day after asthma induction, animals were euthanized, and lungs were sampled for pathological and gene expression analysis.Results: OVA-sensitization led to increased inflammation and histopathological changes in the lung tissue of mice. In addition, GATA-3 expression increased (P<0.001) and T-bet expression decreased (P<0.001) in OVA-sensitized groups. The T-bet/GATA3 ratio was also reduced markedly in asthma groups (P<0.001). Furthermore, increased expression of miR-146a and miR-106a levels was evident in the lung tissue of OVA-sensitized mice (P<0.001 for both). Intervention with high concentrations of crocin (50 and 100 mg/kg) significantly reduced airway inflammation, GATA-3 expression, miR-146a expression, and miR-106a expression and corrected the T-bet/GATA-3 ratio (P<0.05 to P<0.001).Conclusion:Treatment with crocin led to a decrease in the severity of lung inflammation in OVA-sensitized mice, which is probably through the reduction of the T-bet/GATA-3 ratio, and mir-146a and mir-106a expression level.

The expression of miR-181b, CYLD, CBX-7, BCL2, and p53 in osteosarcoma patients and correlation with clinicopathological factors.

Osteosarcoma is a common human malignancy with a high mortality rate worldwide. Recent studies have been focused on understanding the involvement of microRNA (miRNAs) in the pathogenesis of osteosarcoma. Therefore, the present study aimed to measure the expression levels of miR-181a, cylindromatosis (CYLD), chromo box homolog 7 (CBX7), B-cell lymphoma 2 (BCL2), and tumor protein p53 in tumor tissue and adjacent normal tissues in patients with osteosarcoma and its relationship with clinicopathological factors. The expression levels of miR-181a, CYLD, CBX7, BCL2, and p53 were measured in 60 patients with osteosarcoma using quantitative real-time polymerase chain reaction. Finally, we compared the relationship between these gene levels and clinicopathological factors in tumor and healthy tissues. Our results showed that the expression levels of miR-181a, BCL2, and p53 were significantly higher in osteosarcoma tissue in comparison with normal tissues (p < .05). On the contrary, CYLD and CBX7 were downregulated in osteosarcoma tumor tissues compared to adjacent healthy tissues (p < .05). In addition, the expression levels of miR-181a in tumor tissues were strongly correlated with patients' age, tumor size, clinical stage, cancer grade, and lymph node metastasis (p < .05). Our findings highlight new insights into understanding the role of miR-181a in the pathogenesis of osteosarcoma. However, further studies are needed to elucidate miRNA as therapeutic targets for osteosarcoma.

Associations of miR-181a with Health-Related Quality of Life, Cognitive Functioning, and Clinical Data of Patients with Different Grade Glioma Tumors.

Gliomas are central nervous system tumors with a lethal prognosis. Small micro-RNA molecules participate in various biological processes, are tissue-specific, and, therefore, could be promising targets for cancer treatment. Thus, this study aims to examine miR-181a as a potent biomarker for the diagnosis and prognosis of glioma patients and, for the first time, to find associations between the expression level of miR-181a and patient quality of life (QoL) and cognitive functioning. The expression level of miR-181a was analyzed in 78 post-operative II-IV grade gliomas by quantitative real-time polymerase chain reaction. The expression profile was compared with patient clinical data (age, survival time after the operation, tumor grade and location, mutation status of isocitrate dehydrogenase 1 (IDH1), and promoter methylation of O-6-methylguanine methyltransferase). Furthermore, the health-related QoL was assessed using the Karnofsky performance scale and the quality of life questionnaires; while cognitive assessment was assessed by the Hopkins verbal learning test-revised, trail-making test, and phonemic fluency tasks. The expression of miR-181a was significantly lower in tumors of grade III and IV and was associated with IDH1 wild-type gliomas and a worse prognosis of patient overall survival. Additionally, a positive correlation was observed between miR-181a levels and functional status and QoL of glioma patients. Therefore, miR-181a is a unique molecule that plays an important role in gliomagenesis, and is also associated with changes in patients’ quality of life.

A Functional Network Driven by MicroRNA-125a Regulates Monocyte Trafficking in Acute Inflammation.

During the onset of acute inflammation, rapid trafficking of leukocytes is essential to mount appropriate immune responses towards an inflammatory insult. Monocytes are especially indispensable for counteracting the inflammatory stimulus, neutralising the noxa and reconstituting tissue homeostasis. Thus, monocyte trafficking to the inflammatory sites needs to be precisely orchestrated. In this study, we identify a regulatory network driven by miR-125a that affects monocyte adhesion and chemotaxis by the direct targeting of two adhesion molecules, i.e., junction adhesion molecule A (JAM-A), junction adhesion molecule-like (JAM-L) and the chemotaxis-mediating chemokine receptor CCR2. By investigating monocytes isolated from patients undergoing cardiac surgery, we found that acute yet sterile inflammation reduces miR-125a levels, concomitantly enhancing the expression of JAM-A, JAM-L and CCR2. In contrast, TLR-4-specific stimulation with the pathogen-associated molecular pattern (PAMP) LPS, usually present within the perivascular inflamed area, resulted in dramatically induced levels of miR-125a with concomitant repression of JAM-A, JAM-L and CCR2 as early as 3.5 h. Our study identifies miR-125a as an important regulator of monocyte trafficking and shows that the phenotype of human monocytes is strongly influenced by this miRNA, depending on the type of inflammatory stimulus.

Metformin inhibits the pathogenic functions of AChR-specific B and Th17 cells by targeting miR-146a

Myasthenia gravis (MG) is characterized by fatigable skeletal muscle weakness with a fluctuating and unpredictable disease course and is caused by circulating autoantibodies and pathological T helper cells. Regulation of B-cell function and the T-cell network may be a potential therapeutic strategy for MG. MicroRNAs (miRNAs) have emerged as potential biomarkers in immune disorders due to their critical roles in various immune cells and multiple inflammatory diseases. Aberrant miR-146a signal activation has been reported in autoimmune diseases, but a detailed exploration of the relationship between miR-146a and MG is still necessary. Using an experimental autoimmune myasthenia gravis (EAMG) rat model, we observed that miR-146a was highly expressed in the spleen but expressed at low levels in the thymus and lymph nodes in EAMG rats. Additionally, miR-146a expression in T and B cells was also quite different. EAMG-specific Th17 and Treg cells had lower miR-146a levels, while EAMG-specific B cells had higher miR-146a levels, indicating that targeted intervention against miR-146a might have diametrically opposite effects. Metformin, a drug that was recently demonstrated to alleviate EAMG, may rescue the functions of both Th17 cells and B cells by reversing the expression of miR-146a. We also investigated the downstream target genes of miR-146a in both T and B cells using bioinformatics screening and qPCR. Taken together, our study identifies a complex role of miR-146a in the EAMG rat model, suggesting that more caution should be paid in targeting miR-146a for the treatment of MG.

Significance of detecting the levels of miR-29a, survivin and interferon gamma release assay in patients with lung cancer and tuberculosis.

When lung cancer is combined with concurrent tuberculosis (TB), it increases the difficulty of diagnosis and treatment, leading to missed and/or misdiagnosed cases. To provide reference markers for the clinical diagnosis of patients with lung cancer complicated by active pulmonary TB (APT). The concentration of survivin in diseased tissue, and miR-29a and IGRAs interferon gamma (IFN-γ) in serum were evaluated in 25 patients with non-small cell lung carcinoma (NSCLC) complicated by APT, 32 patients with NSCLC and 30 patients with APT. The expression of miR-29a in serum of patients with APT was higher than in patients with NSCLC complicated by APT (least significant difference (LSD)-t = 4.724, p < 0.001), and the NSCLC group (LSD-t = 6.619, p < 0.001). Furthermore, patients with NSCLC complicated by APT had higher miR-29a concentration than the NSCLC group. The rate of positive survivin expression in NSCLC (χ2 = 23.418, p < 0.001) and NSCLC combined with APT group (χ2 = 17.160, p < 0.001) was significantly higher than in patients with APT. The concentration of IFN-γ in serum of the NSCLC complicated by APT group (LSD-t = 2.912, p = 0.004) and the APT group (LSD-t = 4.452, p < 0.001) was higher than in the NSCLC group. The level of IFN-γ in serum of the NSCLC complicated by APT group were higher than in the APT group, but there was no statistical difference. The levels of MiR-29a, Survivin and IFN-γ was helpful for differential diagnosis of lung cancer and tuberculosis.

Plasma microRNA expression and immunoregulatory cytokines in an Indian population occupationally exposed to cadmium.

Following its accumulation in the body, cadmium (Cd) exposure is associated with devastating effects on multiple organ system of the human body. The immune system is one of the sensitive targets for Cd-inducedtoxicity. Recently, studies have demonstrated a significant role of Cd in inducing epigenetic alterations. With this background, the present study was planned to study the changes in candidate microRNA(miRNA) expression associated with immune regulation in occupationally Cd-exposedworkers. One hundred individuals involved in welding and metal handicraft manufacturing, while 80 apparently healthy subjects without any prior history of occupational exposure were recruited for the study. Blood Cd level was determined by atomic absorption spectrometry. Serum cytokine levels were measured using anenzyme-linked immunosorbent assay and serum miRNA expression of candidate miRNAs (miR-146a, miR-210, and miR-222) were determined by real-time polymerase chain reaction.The median Cd level (2.40 μg/L) in the occupationally exposed workers was significantly higher than the nonexposed subjects (0.90 μg/L). Among the cytokines, interleukin-4 (IL-4), and tumor necrosis factor-alpha (TNF-α)were significantly higher whileIL-2 and IL-10 were significantly lower in the exposed. The expression level of miR-146a and miR-222 were significantly different between the groups with the former showing downregulation and later showing upregulation. Correlation analysis revealed a positive and negative association of miR-222 and miR-146a with blood cadmium level, IL-17 as well as TNF-α, respectively. Furthermore, the in-silico analysis revealed a significant role of the studied miRNAs in various cellular and genetic pathways. The findings of the present study demonstrate significant involvement of Cd-inducedalteration in miRNAs in varied immune regulatory changes in exposed individuals.