Clinical significance of serum microRNA-146a and inflammatory factors in children with Mycoplasma pneumoniae pneumonia after azithromycin treatment

Abstract Objective This study aimed to investigate the clinical significance of serum microRNA-146a and pro-inflammatory factors in children with Mycoplasma pneumoniae pneumonia after azithromycin treatment. microRNA-146a is known to regulate inflammatory responses, and excessive inflammation is a primary characteristic of MPP. Methods Children with MPP received conventional symptomatic therapy along with intravenous administration of azithromycin for one week. Serum levels of microRNA-146a and pro-inflammatory factors were measured using RT-qPCR and ELISA kits, respectively. The correlation between microRNA-146a and pro-inflammatory factors was analyzed by the Pearson method. Pulmonary function indexes were assessed using a pulmonary function analyzer, and their correlation with microRNA-146a and pro-inflammatory factors after treatment was evaluated. Children with MPP were divided into effective and ineffective treatment groups, and the clinical significance of microRNA-146a and pro-inflammatory factors was evaluated using receiver operating characteristic curves and logistic multivariate regression analysis. Results Serum microRNA-146a was downregulated in children with MPP but upregulated after azithromycin treatment, contrasting with the trend observed for pro-inflammatory factors. MicroRNA-146a showed a negative correlation with pro-inflammatory cytokines. Pulmonary function parameters were initially reduced in children with MPP, but increased after treatment, showing positive/inverse associations with microRNA-146a and pro-inflammatory factors. Higher microRNA-146a and lower pro-inflammatory factors predicted better efficacy of azithromycin treatment. MicroRNA-146a, tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), interleukin-8 (IL-8), and forced expiratory volume in the first second/forced vital capacity (FEV1/FVC) were identified as independent factors influencing treatment efficacy. Conclusion Azithromycin treatment in children with MPP upregulates microRNA-146a, downregulates pro-inflammatory factors, and effectively improves pulmonary function.

miR-146a downregulates epidermal Langerhans cell-mediated antigen presentation / 中华微生物学和免疫学杂志

Objective:To investigate the role of miR-146a in regulating the homeostasis and function of epidermal Langerhans cells (LCs).Methods:Fresh and in vitro cultured epidermal LCs were isolated and purified by flow cytometry (FCM). The expression of miR-146a in LCs was detected by quantitative PCR (qPCR). The percentages of epidermal LCs in wild-type (WT) and miR-146a conventional knockout (miR-146a cKO) mice were analyzed by FCM. The expression of major histocompatibility complex Ⅱ (MHCⅡ) and co-stimulatory molecules (CD86 and CD80) was analyzed by FCM to evaluate the effect of miR-146a on the maturation of LCs. The percentage of Dextran-FITC + LCs was detected by FCM to evaluate the effect of miR-146a on the phagocytic function of LCs. In vitro and in vivo experiments were used to analyze the ability of miR-146a-deficient and -sufficient LCs to stimulate the proliferation of CD8 + OT-ⅠT cells and CD4 + OT-Ⅱ T cells. Results:The expression of miR-146a was significantly increased in mature LCs than in the freshly isolated LCs. There was no significant difference in the number of epidermal LCs between wild-type (WT) and miR-146a cKO mice. After a 48 h culture in vitro, the expression of MHCⅡ, CD86 and CD80 in the epidermal LCs of miR-146a cKO mice was similar to that of WT mice. Moreover, miR-146a deletion had no significant influence on antigen uptake by LCs. However, miR-146a deficiency enhanced the antigen-presenting ability of LCs that could stimulate the proliferation of OVA-specific CD8 + OT-Ⅰ T cells and CD4 + OT-Ⅱ T cells. Conclusions:miR-146a had no influence on the homeostasis, maturation and phagocytosis of LCs, but enhanced the antigen-presenting function.

Atorvastatin inhibits malignant behaviors and induces apoptosis in human glioma cells by up-regulating miR-146a and inhibiting the PI3K/Akt signaling pathway / 南方医科大学学报

OBJECTIVE@#To explore the effect of atorvastatin (AVT) on biological behaviors and the miR-146a/PI3K/Akt signaling pathway in human glioma cells.@*METHODS@#Human glioma U251 cells were treated with 8.0 μmol/L AVT or transfected with a miR-146a inhibitor or a negative control fragment (miR-146a NC) prior to AVT treatment. RT-PCR was used to detect miR-146a expression in the cells, and the changes in cell proliferation rate, apoptosis, cell invasion and migration were detected using MTT assay, flow cytometry, and Transwell assay. Western blotting was performed to detect the changes in cellular expressions of proteins in the PI3K/Akt signaling pathway.@*RESULTS@#AVT treatment for 48 h resulted in significantly increased miR-146a expression and cell apoptosis (P < 0.01) and obviously lowered the cell proliferation rate, invasion index, migration index, and expressions of p-PI3K and p-Akt protein in U251 cells (P < 0.01). Compared with AVT treatment alone, transfection with miR-146a inhibitor prior to AVT treatment significantly reduced miR-146a expression and cell apoptosis (P < 0.01), increased the cell proliferation rate, promoted cell invasion and migration, and enhanced the expressions of p-PI3K and p-Akt proteins in the cells (P < 0.01); these effects were not observed following transfection with miR-146a NC group (P>0.05).@*CONCLUSION@#AVT can inhibit the proliferation, invasion and migration and promote apoptosis of human glioma cells possibly by up-regulating miR-146a expression and inhibiting the PI3K/Akt signaling pathway.

Effect of parathyroid hormone on osteoporosis after spinal cord injury in rats and its mechanism / 中国药理学通报

Aim To explore the effect of parathyroid hormone on osteoporosis in rats after spinal cord injury(SCI)and its mechanism.Methods SD rats were divided into sham operation group(Sham), SCI model group(SCI), SCI+parathyroid hormone group(SCI+PTH)and SCI+PTH+transfected miR-146a irrelevant fragment group(SCI+PTH+NC)and SCI+PTH+transfection miR-146a inhibitor group(SCI+PTH+miR-146a inhibtor), and then given 60 μg·kg-1 PTH(SCI+PTH group), 60 μg·kg-1 PTH and 20 pm miR-146a NC(SCI+PTH+NC group)or 60 μg·kg-1 PTH and 20 pm miR-146a inhibitor(SCI+PTH+miR-146a inhibitor group)by tail vein injection every 3 d for 8 weeks.Rats in Sham group and SCI group were given equal amount of saline in the same way.The behavioral movement scores of rats were recorded by the BBB scoring method 1 d, 7 d, 14 d, 21 d, 28 d, and 56 d after operation; serum calcium(Ca)and alkaline phosphatase(ALP)were measured using the kits; bone mineral density of femur and tibia was measured by a bone mineral density scanner; the morphological changes of rat spinal cord were observed by HE staining; expression of miR-146a was detected by qRT-PCR and protein expression of p-PI3K and p-Akt was detected by Western blot.Results Compared with Sham group, SCI group had decreased BBB score(P<0.05 or P<0.01), serum Ca, femoral and tibial bone mineral density content and expression of miR-146a, p-PI3K and p-Akt, but increased serum ALP(P<0.01).Compared with SCI group, BBB score(P<0.05 or P<0.01), serum Ca, femoral and tibia bone mineral density content, and the expression of miR-146a, p-PI3K and p-Akt( P<0.01)increased, together with decreased serum ALP in SCI+PTH group(P<0.01).Compared with SCI+ PTH group, the above indicators of rats were significantly inhibited in SCI+PTH+miR-146a inhibitor group.Conclusions PTH has certain therapeutic effect on SCI osteoporosis, achieved possibly by regulating miR-146a/PI3K/Akt signaling.

miR-146b suppresses LPS-induced M1 macrophage polarization via inhibiting the FGL2-activated NF-κB/MAPK signaling pathway in inflammatory bowel disease

Abstract Objectives: M1 macrophage polarization and phenotype in Inflammatory Bowel Disease (IBD) are common biological responses. Method: Herein, IBD mice models were constructed and macrophages were derived. Results: It was discovered that microRNA-146b (miR-146b) was downregulated in IBD mice and Lipopolysaccharide (LPS)-induced macrophages. Moreover, the inhibitory role of overexpressed miR-146b in reducing the inflammation level and blocking M1 macrophage polarization was confirmed. Further investigation indicated that Fibrinogen Like 2 (FGL2) acted as the target gene of miR-146b, and FGL2 mediated activation of NLRP3, NF-κB-p65, and p38-MAPK. More importantly, it was validated that miR-146b could ameliorate inflammatory pheno-type and prevent M1 macrophage polarization via inhibiting FGL2 in vitro, and miR-146b overexpression alleviated the intestinal injury of IBD mice in vivo. Conclusions: Overall, it is potential to use miR-146b for the amelioration of IBD. HIGHLIGHTS miR-146b was downregulated in Inflammatory Bowel Disease (IBD) mice and LPS-induced macrophages. Fibrinogen Like 2 (FGL2) was identified as the target gene of miR-146b. miR-146b ameliorated the inflammation and blocked M1 macrophage polarization via inhibiting FGL2. miR-146b ameliorated the symptoms and pathological injury of IBD via inhibiting FGL2.

Mechanism of miRNA-146b regulating proliferation, metastasis and apoptosis of thyroid papillary carcinoma cells / 中华内分泌外科杂志

Objective:To investigate the role and mechanism of miR-146b in the proliferation, metastasis and apoptosis of thyroid papillary carcinoma cells.Methods:qRT-PCR was used to detect the expression of miR-146b in thyroid papillary carcinoma cells (NPA, GLAG-66, ONCNO-DG1 and B-CPAP) and normal human thyroid cell line HTori3. After B-CPAP cells were transfected with miR-146b inhibitor, the inhibition efficiency was detected by qRT-PCR, the effect of miR-146b on PTC cells proliferation was detected by MTT assay, the effect of miR-146b on PTC cells invasion was studied by Transwell assay, and the effect of miR-146b on tumor cell apoptosis was detected by flow cytometry. SiRNA-IRAK1 was transfected into B-CPAP cell line. The cell proliferation rate, migration ability and apoptosis rate were detected by MTT, cell scratch test and flow cytometry respectively. The target gene of miR-146b, interleukin-1 associated receptor kinase 1 (IRAK1) , was predicted by bioinformatics software, and the regulatory effect of miR-146b on IRAK1 was verified by double fluorescein reporter gene experiment.Results:QRT-PCR showed that the expression of miR-146b in NPA87, KAT-5, FTC-133 and B-CPAP cell lines was significantly higher than that in normal cell HTori3, especially B-CPAP ( P<0.05) . MiR-146b inhibitor transfection could significantly reduce the expression level of miR-146b in B-CPAP cells ( P<0.01) . MTT results showed that miR-146b inhibitor could inhibit the proliferation of B-CPAP cells ( P<0.05) . Flow cytometry showed that miR-146b inhibitor could promote the apoptosis of B-CPAP cells ( P<0.05) . Transwell results showed that miR-146b inhibitor could reduce the invasive ability of B-CPAP cells ( P<0.05) . After transfection with siRNA-IRAK1, the proliferation rate of B-CPAP cells increased significantly (MTT test) , the migration ability increased (cell scratch test) , and the apoptosis rate decreased significantly (flow cytometry) ( P<0.05) . The results of double luciferase reporter gene showed that irak1 was the target gene of miR-146b, and miR-146b inhibitor could significantly up regulate the expression level of irak1 protein in B-CPAP cells. Conclusion:miR-146b may play a role in promoting the proliferation and metastasis and inhibiting cell apoptosis of PTC cells by inhibiting the downstream target protein IRAK1.

miR-146a-5p-mediated suppression on trophoblast cell progression and epithelial-mesenchymal transition in preeclampsia

OBJECTIVE: This study aims to identify the effect of miR-146a-5p on trophoblast cell invasion as well as the mechanism in preeclampsia (PE). METHODS: Expression levels of miR-146a-5p and Wnt2 in preeclamptic and normal placentae were quantified. Trophoblast cells (HTR-8) were separately transfected with miR-146a-5p mimic, miR-146a-5p inhibitor, pcDNA3.1-Wnt2 or sh-Wnt2, and then the expression levels of miR-146a-5p, Wnt2, and epithelial-mesenchymal transition (EMT)-related proteins (Vimentin, N-cadherin and E-cadherin) were measured. Moreover, the proliferative, migratory and invasive capacities of trophoblast cells were detected, respectively. Dual luciferase reporter assay determined the binding of miR-146a-5p and Wnt2. RESULTS: Compared with normal placental tissues, the placentae from PE patients showed higher miR-146a-5p expression and lower Wnt2 expression. Transfection of miR-146a-5p inhibitor or pcDNA3.1-Wnt2 exerted pro-migratory and pro-invasive effects on HTR-8 cells and encouraged EMT in HTR-8 cells; transfection with miR-146a-5p mimic or sh-Wnt2 weakened the proliferative, migratory and invasive capacities as well as reduced EMT process of HTR-8 cells. Moreover, Wnt2 overexpression could partially counteract the suppressive effects of miR-146a-5p overexpression on the progression and EMT of HTR-8 cells. CONCLUSION: miR-146a-5p mediates trophoblast cell proliferation and invasion through regulating Wnt2 expression.

Silencing lncRNA GIHCG increases radiosensitivity of glioma cells by up-regulating miR-146a-3p / 中华放射肿瘤学杂志

Objective@#To investigate the effect of lncRNA GIHCG on the radiosensitivity of glioma cells and its mechanism.@*Methods@#The expression levels of GIHCG and miR-146a-3p in human brain normal glial cells HEB and glioma cell lines U251, A172, SHG139 and U87 were quantitatively measured by qRT-PCR assay. U251 and SHG139 cells were used for subsequent experiment. After silencing the expression of GIHCG or overexpressing miR-146a-3p in U251 and SHG139 cells, cell proliferation was detected by MTT assay, cell apoptosis was detected by flow cytometry, cell radiosensitivity was detected by colony formation assay and the expression levels of CDK1, CyclinD1, Bcl-2 and Bax proteins were measured by Western blot. The bioinformatics software predicted the presence of a binding site for GIHCG and miR-146a-3p. Dual luciferase reporter gene assay and qRT-PCR assay were adopted to verify the targeting relationship between GIHCG and miR-146a-3p.@*Results@#Compared with HEB cells, the expression of GIHCG was significantly up-regulated in glioma U87, U251, A172 and SHG139 cells (all P<0.05), whereas that of miR-146a-3p was remarkably down-regulated (P<0.05). Silencing GIHCG expression or overexpression of miR-146a-3p significantly decreased the U251 and SHG139 cell survival rate, survival fraction and the expression of CDK1, CyclinD1 and Bcl-2 proteins (all P<0.05), whereas considerably increased the apoptotic rate and expression of Bax protein (both P<0.05). GIHCG performed targeted negative regulation of miR-146a-3p expression in U251 and SHG139 cells and inhibition of miR-146a-3p expression reversed the effect of silencing GIHCG on proliferation, apoptosis and radiosensitivity of glioma cells.@*Conclusion@#Silencing GIHCG expression up-regulates the expression of miR-146a-3p, thereby enhancing the radiosensitivity of glioma cells.

Mirna expression profiling of osteogenic differentiation of bone marrow mesenchymal stem cells induced by microchannel porous hydroxyapatite scaffold / 中国组织工程研究

BACKGROUND: Porous hydroxyapatite scaffolds have good osteogenesis in vivo and in vitro. However, little research has been done on the complex regulation mechanisms of miRNAs involved. OBJECTIVE: To investigate the changes of related miRNA expression in rat bone marrow mesenchymal stem cells during osteogenic mineralization by porous hydroxyapatite scaffolds. METHODS: Rat bone marrow mesenchymal stem cells were isolated, cultured and identified in vitro. Bone marrow mesenchymal stem cells co-cultured with porous hydroxyapatite scaffold were as experimental group, and bone marrow mesenchymal stem cells cultured alone served as blank control group, both of which underwent osteogenic induction for 7 days. During the osteogenic mineralization, miRNA high-throughput sequencing technology was used to analyze the changes of miRNA expression profiles followed by GO analysis. The miRNA molecules with obvious expression differences were screened and verified by qRT-PCR. RESULTS AND CONCLUSION: (1) Compared with the blank control group, in the experimental group, the expression levels of BMP2, ALP and Runx2 mRNA were up-regulated, and the expression level of BMP2 was up-regulated significantly (P < 0.05). (2) Results of miRNA high-throughput sequencing showed that 13 miRNAs such as miR-210-3p and miR-146a-5p were up-regulated, and 17 miRNAs such as let-7c-3p and let-3615 were down-regulated significantly. (3) GO analysis revealed that up-regulated miRNA target genes were mainly involved in biological regulation, cellular gene expression, and gene expression regulation, mainly including nuclear factor-κB, Toll-like receptor 9, intercellular adhesion, interleukin-1 regulation, and signaling pathways such as angiogenesis and Hippo. (4) Real-time fluorescence quantitative qPCR results showed that miRNA-210 was up-regulated 15 times and miR-146a-5p was up-regulated 10 times in the experimental group (P < 0.05). These results indicate that the new microchannel porous hydroxyapatite scaffold can promote the differentiation of bone marrow mesenchymal stem cells by up-regulating miRNA-210-3p and miR-146a.

Silencing lncRNA GIHCG increases radiosensitivity of glioma cells by up-regulating miR-146a-3p / 中华放射肿瘤学杂志

Objective To investigate the effect of lncRNA GIHCG on the radiosensitivity of glioma cells and its mechanism.Methods The expression levels of GIHCG and miR-146a-3p in human brain normal glial cells HEB and glioma cell lines U251,A172,SHG139 and U87 were quantitatively measured by qRT-PCR assay.U251 and SHG139 cells were used for subsequent experiment.After silencing the expression of GIHCG or overexpressing miR-146a-3p in U251 and SHG139 cells,cell proliferation was detected by MTI assay,cell apoptosis was detected by flow cytometry,cell radiosensitivity was detected by colony formation assay and the expression levels of CDK1,CyclinD1,Bcl-2 and Bax proteins were measured by Western blot.The bioinformatics software predicted the presence of a binding site for GIHCG and miR-146a-3p.Dual luciferase reporter gene assay and qRT-PCR assay were adopted to verify the targeting relationship between GIHCG and miR-146a-3p.Results Compared with HEB cells,the expression of GIHCG was significantly up-regulated in glioma U87,U251,A172 and SHG139 cells (all P＜0.05),whereas that of miR-146a-3p was remarkably down-regulated (P＜0.05).Silencing GIHCG expression or overexpression of miR-146a-3p significantly decreased the U251 and SHG139 cell survival rate,survival fraction and the expression of CDK1,CyclinDl and Bcl-2 proteins (all P＜0.05),whereas considerably increased the apoptotic rate and expression of Bax protein (both P＜0.05).GIHCG performed targeted negative regulation of miR-146a-3p expression in U251 and SHG139 cells and inhibition of miR-146a-3p expression reversed the effect of silencing GIHCG on proliferation,apoptosis and radiosensitivity of glioma cells.Conclusion Silencing GIHCG expression up-regulates the expression of miR-146a-3p,thereby enhancing the radiosensitivity of glioma cells.

miR-146a is deregulated in gastric cancer

Background: Gastric cancer is one of the most significant reasons for cancer-related death. miR-146a is one of the dysregulated factors associated with gastric tumorigenesis. However, deregulation of this microRNA (miRNA) has become controversial. Moreover, the inflammation-mediating role of this miRNA implies that miR-146a might be dysregulated by gastric cancer-related pathogens, such as Helicobacter pylori. However, the dysregulation of miR-146a in H. pylori-infected gastric tumors has not been widely studied. Objectives: We aimed to analyze the expression level of miR-146a in gastric cancer tissues and then to assess any potential association between miR-146a and H. pylori infection and other clinical characteristics. Materials and Methods: miR-146a expression level was quantitatively studied by reverse transcription quantitative polymerase chain reaction, in 144 fresh tissues including 44 normal and 100 gastric cancer samples. Results: A dramatic overexpression of miR-146a was observed in primary gastric tumors. miR-146a showed lower expression in progressed tumors with greater stages and lymph node metastasis. Conclusion: miR-146a is highly expressed in primary gastric tumor independent of H. pylori infection. It is highly expressed in the lower stages and lymph node-negative tumors. It might suggest the importance of upregulation and downregulation of this miRNA in the initiating/promoting and progressive steps of gastric tumorigenesis, respectively

Effects of miR-146a on hepatic INSR gene expression in F1 offspring of paternal rats with high-glucose-high-fat diet / 中华内分泌代谢杂志

Objective To study the effect of miR-146a on hepatic INSR gene expression in F1 offspring of paternal rats with high-glucose-high-fat diet ( HGFD) . Methods 5-week-old male SD rats were randomly divided into normal diet ( ND) and HGFD groups. Male rats with ND or HGFD feeding for three months mated with normal female ones. Blood glucose concentration, glucose tolerance, and insulin tolerance in newborn male offspring rats were detected respectively. Differential miRNAs between ND and HGFD groups were compared using next generation sequencing and were then confirmed by real-time quantitative PCR ( qPCR) . The relative expression of INSR mRNA and the methylation level of INSR promoter in liver tissues of the offspring newborn rat were detected and the correlations between them and the relative expression of differential microRNA were analyzed respectively. In vitro, effects of miR-146a on expression and methylation of INSR gene in BRL-3A cells were detected using Western-blot assay and qPCR respectively. Results The fasting glucose concentration of different groups were without significant difference, but glucose tolerance and insulin tolerance of neonatal male rats in HGFD group were decreased significantly . Next generation sequencing has revealed 45 up-regulated miRNAs and 15 down-regulated miRNAs in HGFD group. Among them, differences of 8 miRNAs expression in the enlarged samples were confirmed by qPCR. miR-146a was up-regulated for more than 10 times in the liver of the offspring of HGFD group. Expression level of miRNA-146a was negatively correlated with the relative expression of INSR and positively correlated with the methylation level of INSR in livers of neonatal rats in HGFD group. In vitro, miR-146a ( mimics ) promoted the methylation of INSR gene and inhibited the expression of INSR in BRL-3A cells. Conclusion HGFD feeding to male SD rats leads to the inhibition of hepatic INSR gene expression in neonatal offspring via upregulating miR-146a.

microRNA-146a Promotes Growth of Acute Leukemia Cells by Downregulating Ciliary Neurotrophic Factor Receptor and Activating JAK2/STAT3 Signaling

PURPOSE: Acute leukemia (AL) is classified as acute lymphoblastic leukemia (ALL) and acute myeloid leukemia (AML). This study aimed to investigate the effect of miR-146a on childhood AL and its underlying molecular mechanisms. MATERIALS AND METHODS: Bone marrow samples were obtained from 39 AL children and 10 non-cancer controls. The expressions of miR-146a and ciliary neurotrophic factor receptor (CNTFR) were detected by quantitative real-time polymerase chain reaction (qRT-PCR) in ALL and AML pediatric patients, as well as ALL (Jurkat) and AML (HL-60) cells. Correlations between miR-146a and clinical indicators were explored. A targeting relationship between miR-146a and CNTFR was detected by dual luciferase reporter gene assay. Cell proliferation, apoptosis, migration, and invasion of Jurkat and HL-60 cells were measured by MTT assay, flow cytometry, and transwell assay, respectively. LIF expression was detected by qRT-PCR in Jurkat and HL-60 cells. The expression of p-JAK2, JAK2, p-STAT3, and STAT3 in HL-60 cells was measured by Western blot. RESULTS: miR-146a was increased in ALL and AML pediatric patients, while CNTFR was decreased. miR-146a expression was associated with immunophenotype, karyotype, fusion gene, and SIL-TAL1. CNTFR was a target gene of miR-146a. miR-146a could promote cell proliferation, migration, and invasion, as well as inhibit cell apoptosis in Jurkat and HL-60 cells by downregulating CNTFR. Meanwhile, miR-146a inhibited the expression of LIF and activated JAK2/STAT3 pathway by downregulating CNTFR. CONCLUSION: miR-146a could promote the proliferation, migration, and invasion and inhibit the apoptosis of AL Jurkat and HL-60 cells by downregulating CNTFR and activating the JAK2/STAT3 pathway.

Inhibition of KLF7-Targeting MicroRNA 146b Promotes Sciatic Nerve Regeneration / 神经科学通报·英文版

A previous study has indicated that Krüppel-like factor 7 (KLF7), a transcription factor that stimulates Schwann cell (SC) proliferation and axonal regeneration after peripheral nerve injury, is a promising therapeutic transcription factor in nerve injury. We aimed to identify whether inhibition of microRNA-146b (miR-146b) affected SC proliferation, migration, and myelinated axon regeneration following sciatic nerve injury by regulating its direct target KLF7. SCs were transfected with miRNA lentivirus, miRNA inhibitor lentivirus, or KLF7 siRNA lentivirus in vitro. The expression of miR146b and KLF7, as well as SC proliferation and migration, were subsequently evaluated. In vivo, an acellular nerve allograft (ANA) followed by injection of GFP control vector or a lentiviral vector encoding an miR-146b inhibitor was used to assess the repair potential in a model of sciatic nerve gap. miR-146b directly targeted KLF7 by binding to the 3'-UTR, suppressing KLF7. Up-regulation of miR-146b and KLF7 knockdown significantly reduced the proliferation and migration of SCs, whereas silencing miR-146b resulted in increased proliferation and migration. KLF7 protein was localized in SCs in which miR-146b was expressed in vivo. Similarly, 4 weeks after the ANA, anti-miR-146b increased KLF7 and its target gene nerve growth factor cascade, promoting axonal outgrowth. Closer analysis revealed improved nerve conduction and sciatic function index score, and enhanced expression of neurofilaments, P0 (anti-peripheral myelin), and myelinated axon regeneration. Our findings provide new insight into the regulation of KLF7 by miR-146b during peripheral nerve regeneration and suggest a potential therapeutic strategy for peripheral nerve injury.

Association between miR146a(rs2910164)G>C polymorphism and susceptibility to acute lymphoblastic leuke-mia in children / 中华实用儿科临床杂志

Objective To investigate the association between miR146a(rs2910164)G>C polymorphism and susceptibility to acute lymphoblastic leukemia(ALL)in children.Methods Two hundred blood specimens were ob-tained from children with ALL as patient group and 100 blood specimens were obtained from healthy children as healthy control group,who were all from Baoding First Central Hospital between March 2010 and October 2016.There were no significant differences in sex and age between patient group and healthy control group(χ2=0.430,P=0.512;χ2=2.839,P=0.092).The distribution of gene frequency of patient group and healthy control group conformed to Hardy-Weinberg equilibrium.miR146a(rs2910164)G>C polymorphism was identified by adopting restriction fragment length polymorphism(RFLP).The relation of genotype and ALL was demonstrated by odds ratio(OR)and 95% credibility interval(CI). Results Gene frequency of miR146a(rs2910164)GG,GC and CC genotypes in patient group and healthy control group was 16.0%,44.5%,39.5% and 29.0%,41.0%,30.0%,respectively.The GC/CC genotypes were significantly higher in patient group than those in healthy control group(GG genotype as reference,GC genotype:OR=1.967,95%CI:1.054-3.672,P=0.037;CC genotype:OR=2.386,95%CI:1.239-4.595,P =0.012). Conclusion miR146a(rs2910164)G>C polymorphism is significantly associated with susceptibility to ALL in chil-dren.

Construction of dual luciferase reporter plasmid of HIPK3 3′UTR and verification its targeting miR-146 / 基础医学与临床

Objective To construct a dual luciferase reporter vector containing the 3′untranslated region(3′UTR) of HIPK3 gene and verify the relationship between HIPK3 and miR-146.Methods The binding sites of miR-146 and HIPK3 genes were predicted by miRDB database and DIANA TOOLS database. The 3′UTR region sequences of HIPK3 genes and its mutants were respectively inserted into the luciferase report plasmid psiCHECK-2 to construct a wild-type and a mutant recombinant dual luciferase report plasmid. The 293T cells were divided into 6 groups and transfected with 1) HIPK3-WT+NC negative control;2)HIPK3-WT+miR-146a mimics;3)HIPK3-WT+miR-146b mimics;4)HIPK3-MU+NC negative control; 5)HIPK3-MU+miR-146a mimics and 6)HIPK3-MU+ miR-146b mimics respectively. After 48 hours, the luciferase activity was detected.Results HIPK3-WT and HIPK3-MU re-combinant plasmid were successfully constructed. When HIPK3-WT recombinant plasmids and miR-146b mimics were transfected into 293T cells, the luciferase activity was decreased (P<0.05). Conclusions miR-146a does not have a target relationship with HIPK3 gene,whereas miR-146b can regulate the 3′UTR of HIPK3 gene.

Correlation of serum miR-146a with nuclear factor -κB and vascular endothelial growth factor in diabetic retinopathy patients / 国际眼科杂志(Guoji Yanke Zazhi)

@#AIM:To investigate the correlation of serum miR-146a with nuclear factor-κB(NF-κB)and vascular endothelial growth factor(VEGF)in diabetic retinopathy patients. <p>METHODS: A total of 100 patients with T2DM treated in our hospital from July 2016 to December 2017 were assigned into T2DM patients with DR(DR group, <i>n</i>=32)and T2DM patients without DR(T2DM group, <i>n</i>=68). Thirty healthy volunteers were selected as control group. Real-time PCR was used to examine the expression of miR-146a. Enzyme linked immunosorbent assay was used to detect the levels of NF-κB and VEGF. The correlation between miR-146a and NF-κB and VEGF was analyzed. <p>RESULTS: Compared with the control group, HbA1c in T2DM group and DR group increased(<i>t</i>=6.822, 5.709; <i>P</i><0.001), FBG increased(<i>t</i>=8.889, 7.923; <i>P</i><0.001), 2hPBG increased(<i>t</i>=6.646, 5.514; <i>P</i><0.001). Compared with T2DM group, the duration of diabetes in DR group was longer(<i>t</i>=2.431, <i>P</i>=0.017). Compared with the control group, serum miR-146a in T2DM and group DR significantly decreased(<i>t</i>=3.967, 7.169; <i>P</i><0.001), and the DR group was lower than that in the T2DM group(<i>t</i>=4.444, <i>P</i><0.001). Compared with the control group, the serum NF-κB in the T2DM and DR group increased significantly(<i>t</i>=6.063, 14.851; <i>P</i><0.001), VEGF increased significantly(<i>t</i>=7.613, 12.943; <i>P</i><0.001), NF-κB and VEGF in DR group were larger than those in T2DM group(<i>t</i>=11.406, 7.560; <i>P</i><0.001). Pearson analysis showed that miR-146a was negatively correlated with NF-κB and VEGF(<i>r</i>=-0.503, -0.574; <i>P</i><0.05). <p>CONCLUSION: The serum miR-146a in DR patients significantly decreased, the NF-κB and VEGF significantly increased. MiR-146a may be involved in the pathogenesis of DR by mediating inflammatory reaction and vascular proliferation.

Effects of liraglutide on glucose induced expression of miRNA-146b-3p, inflammatory factors and COX-2 in human umbilical vein endothelial cells / 上海交通大学学报（医学版）

Objective • To investigate the effects of liraglutide on glucose induced expression of miRNA-146b-3p, tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and cyclooxygenase-2 (COX-2) in human umbilical vein endothelial cells (HUVECs). Methods • HUVECs were grown in the medium with glucose of high concentration (25 mmol/L) or normal concentration (7 mmol/L) for 24 h, then stimulated with liraglutide. Quantitative polymerase chain reaction (qPCR) was performed to detect the expression of miR-146b-3p, IL-6, TNF-α and COX-2. The expression of IL-6, TNF-α and COX-2 were detected after HUVECs were transfected with anti-miR-146b-3p. Results • The expression of miR-146b-3p was decreased in high glucose induced cells, while the expression of IL-6, TNF-α and COX-2 were increased. Silenced expression of miR-146b-3p increased the expression of IL-6, TNF-α and COX-2. Liraglutide increased the expression of miR-146b-3p and decreased the expression of IL-6, TNF-α and COX-2 in high glucose induced cells. Conclusion • Liraglutide may relieve inflammation and improve vascular endothelial function in hyperglycemia condition through regulating miR-146b-3p to decrease expression of IL-6, TNF-α and COX-2.

Association between miR-146a single nucleotide polymorphism and genetic susceptibility to hepatocellular carcinoma: a meta-analysis / 中华肝脏病杂志

Objective@#To investigate the association between miR-146a single nucleotide polymorphism and genetic susceptibility to hepatocellular carcinoma (HCC).@*Methods@#PubMed, Web of Science, Cochrane Library, Wanfang Data, and Google Scholar were searched for case-control studies on the association between miR-146a single nucleotide polymorphism and genetic susceptibility to HCC published up to October, 2016 in Chinese or English. The Q-statistics test was used to evaluate the heterogeneity of these articles.@*Results@#A total of 18 articles with 5 610 cases and 7 531 controls were included for the meta-analysis. There was no significant association between miR-146a single nucleotide polymorphism and genetic susceptibility to HCC. The odds ratio (OR), 95% confidence interval (95% CI), and P values for the five genetic models were as follows: the allele model C/G (OR = 0.99, 95% CI 0.88-1.06, P = 0.440); the heterozygous model CG/GG (OR = 0.99, 95% CI 0.90-1.10, P = 0.898); the homozygous model CC/GG (OR = 0.91, 95% CI 0.75-1.10, P = 0.314); the dominant model CC+CG/GG (OR = 0.97, 95% CI 0.79-1.19, P = 0.759); the recessive model CG+GG/CC (OR = 1.05, 95% CI 0.94-1.18, P = 0.405). A subgroup analysis of race, source of control population, and Hardy-Weinberg equilibrium were performed in these five genetic models, and miR-146a single nucleotide polymorphism increased the susceptibility to HCC only in the control population-based subgroups of the recessive model CG+GG/CC (OR = 1.20, 95% CI 1.02-1.40, P = 0.024). There was no association between miR-146a rs2910164 polymorphism and susceptibility to HCC in all the other subgroups. A stratified analysis of HBV infection revealed that miR-146a rs2910164 polymorphism increased the risk of HBV-positive HCC (OR = 1.26, 95% CI 1.10-1.49, P = 0.001).@*Conclusion@#There is no significant association between miR-146a rs2910164 polymorphism and the risk of HCC, but miR-146a rs2910164 polymorphism may increase the risk of HBV-positive HCC.

Effect of miRNA-146a on life cycle of hepatitis B virus in vitro / 第三军医大学学报

Objective To determine the effect of microRNA-146a (miR-146a) on the life cycle of hepatitis B virus (HBV) and investigate the underlying mechanisms.Methods The miRNA expression profiles were compared by miRNA array between HepG2 and HepG2.2.15 cells.Then miR-146a was chosen as objective,and its expression level was further confirmed by RT-PCR.After miR-146a mimic and inhibitor were transfected into HepG2.2.15 cells respectively,the quantification of HBV replication was determined by RT-PCR,and the levels of HBsAg and HBeAg in the supernatant were measured by ELISA,and the expression of HS3ST3B1 at mRNA and protein levels were tested by RT-PCR and Western blotting.Dualluciferase reporter assay was used to detect the interaction between miR-146a and potential target HS3ST3B1.Results The expression levels of totally 72 miRNAs were changed in HepG2.2.15 cells,with 27 upregulated and 45 down-regulated.RT-PCR showed the expression level of miR-146a was significantly higher in HepG2.2.15 cells than HepG2 cells (1.55-± 0.13 vs 1.00 ± 0.01,P ＜ 0.05).Transfection of miR-146a mimic into HepG2.2.15 cells resulted in significantly increased HBV replication and levels of HBsAg and HBeAg (P ＜ 0.05),while the transfection of its inhibited caused opposite results (P ＜ 0.05).Bioinformatic analysis showed that HS3ST3B1 was a potential target of miR-146a.The reporter luciferase reporter system indicated that the reported fluorescence intensity of HS3ST3B1 wild type vector was significantly lower than that of the control group (P ＜ 0.05),but showed no significant difference between HS3ST3B1 mutant vector and control group (P ＞0.05).The mRNA level of HS3ST3B1 was not significantly changed in HepG2.2.15 cells transfected with miR-146a mimic (P ＞ 0.05),but its protein level was significantly decreased (P ＜ 0.05).Conclusions miR-146a affects the life cycle of HBV,which may be through suppressing the translation of HBV inhibitory factor HS3ST3B1 3'UTR.