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OBJECTIVE@#To observe the effect of exosomes secreted by lipopolysaccharides (LPS)-stimulated macrophages on hepatic stellate cell activation and migration and explore the underlying molecular mechanism.@*METHODS@#Human monocyte THP-1 cells were induced to differentiate into macrophages using propylene glycol methyl ether acetic acid (PMA, 100 ng/mL, 24 h) followed by stimulation with LPS, and the culture supernatant of macrophages was collected for extraction of the exosomes by ultracentrifugation. The expression of miR-155-5p in the exosomes was detected using qRT-PCR. A Transwell co-culture system was used to observe the effects of the macrophage-derived exosomes on LX2 cell (a hepatic stellate cell line) proliferation, migration, oxidative stress and the expression of fibrosis biomarkers, which were also observed in LX2 cells transfected with miR-155-5p-mimics or miR-155-5p-inhibitors. Western blotting was used to detect the expressions of SOCS1 and its downstream signal pathway proteins.@*RESULTS@#Treatment with the exosomes from LPS-stimulated macrophages significantly enhanced the proliferation and migration ability of LX2 cells and increased the levels of oxidative stress and expressions of the fibrosis markers such as type Ⅰ collagen (P < 0.05). The expression of miR-155-5p in the exosomes secreted by macrophages was significantly increased after LPS treatment (P < 0.01). LX2 cells overexpressing miR-155-5p also exhibited significantly enhanced proliferation and migration with increased oxidative stress levels and expression of type Ⅰ collagen (P < 0.05), and interference of miR-155-5p expression produced the opposite effects. Western blotting showed that miR-155-5p overexpression obviously inhibited SOCS1 expression and promoted p-Smad2/3, Smad2/3 and RhoA protein expressions in LX2 cells (P < 0.05).@*CONCLUSION@#LPS stimulation of the macrophages increases miR-155-5p expression in the exosomes to promote activation and migration and increase oxidative stress and collagen production in hepatic stellate cells.
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Humans , Hepatic Stellate Cells , Lipopolysaccharides/pharmacology , Collagen Type I , Exosomes , Macrophages , MicroRNAsABSTRACT
Parkinson's disease(PD)is the second most common neurodegenerative disease after Alzheimer's disease,with high morbidity and high disability rate.Since the early symptoms of PD are not typical and often similar to those of normal aging or other diseases.It is easy to missed diagnosis and misdiagnosis,which seriously affects the diagnosis and treatment of this disease and aggravetes the burden on the patients' life.MicroRNAs(miRNA)are a class of endogenous non-coding RNAs that are involved in post-transcriptional regulation by binding to target messenger RNAs(mRNA).They are highly conserved,short,easy to obtain,and can stably exist in peripheral body fluids.They have been used as biomarkers for a variety of diseases.Recent studies have demonstrated that miRNA play an important role in the development of PD.This paper reviews the recent research progress of miR-7/124/155,three mature miRNA in PD,aiming to provide reference for clarifying the pathogenesis and guiding the diagnosis and treatment of PD.
Subject(s)
Humans , Parkinson Disease , Neurodegenerative Diseases , MicroRNAs/genetics , Gene Expression Regulation , Biomarkers/metabolismABSTRACT
Objective:To investigate the diagnostic value of cysteine rich 61 (Cyr61), glyoxalase Ⅰ (GLO -1) and microRNA-155(miR-155) in endometrial carcinoma.Methods:Eighty-five patients with endometrial cancer treated in Lu Southwest Hospital from June 2017 to March 2020 were selected as the observation group, including 15 cases of recurrence and 70 cases of non-recurrence. In addition, 85 patients with benign uterine lesions were selected as the control group. The levels of serum Cyr61, GLO-1 and miR-155 were compared between the two groups, the correlation between the levels of serum Cyr61, GLO-1 and miR-155 and clinicopathological features were analyzed, the receiver operating characteristic (ROC) curve was drawn, the diagnostic value of the levels of serum Cyr61, GLO-1 and miR-155 in endometrial cancer were evaluated, and the relationship between the levels of serum Cyr61, GLO-1 and miR-155 and the recurrence of endometrial cancer were analyzed.Results:The levels of serum Cyr61, GLO-1 and miR-155 in the observation group were higher than those in the control group: (294.74 ± 78.41) μg/L vs. (156.82 ± 50.62) μg/L, (96.27 ± 19.85) pmol/L vs. (79.83 ± 15.69) pmol/L, 6.82 ± 2.27 vs. 2.57 ± 0.78, the differences were statistically significant ( P<0.05). The levels of serum Cyr61, GLO-1 and miR-155 in patients with endometrial cancer were positively correlated with clinical stage, myometrial invasion and lymph node metastasis ( P<0.05). The area under the curve(AUC) of serum Cyr61, GLO-1 and miR-155 in the combined diagnosis of endometrial cancer was 0.906. The levels of serum Cyr61, GLO-1 and miR-155 in recurrence patients were higher than those non-recurrence patients : (358.21 ± 89.63) μg/L vs. (281.14 ± 75.29) μg/L, (109.89 ± 20.14) pmol/L vs. (93.35 ± 16.37)pmol/L, 8.04 ± 2.51 vs. 6.56 ± 2.17, the differences were statistically significant ( P<0.05). The levels of serum Cyr61, GLO-1 and miR-155 were the risk factors of recurrence in patients with endometrial cancer ( P< 0.05). Conclusions:The levels of serum Cyr61, GLO-1 and miR-155 in patients with endometrial cancer are significantly increased, which are related to clinical stage, degree of invasion, lymph node metastasis and recurrence. Detecting their levels can be used to diagnose endometrial cancer and predict recurrence, so as to guide clinical treatment.
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Objective:To investigate the role of microRNA (miR)-155 in hydrogen peroxide (H 2O 2)-induced oxidative stress injury in lens epithelial cells (LECs) and its mechanism regulating silent information regulator factor related enzymes 1 (SIRT1). Methods:The HLE-B3 at the logarithmic growth phase was taken and cultured for 24 hours under different concentrations of H 2O 2 (0, 50, 100, 200, 400, 800 μmol/L), and the cell viability was detected by MTT assay to determine the optimal concentration of H 2O 2 for establishing an oxidative stress injury model.HLE-B3 cells were divided into 6 groups, untreated blank control group, model control group cultured with 100 μmol/L H 2O 2, miR-155 mimics group transfected with miR-155 mimics, miR-155 mimics negative control group transfected with miR-155 mimics negative control, miR-155 inhibitor group transfected with miR-155 inhibitor, and miR-155 inhibitor negative control group transfected with miR-155 inhibitor negative control.Transfected cells were cultured with 100 μmol/L H 2O 2.Cells in various groups were cultured for 24 hours, and cell morphology was observed under an inverted microscope.The relative expression of miR-155 and SIRT1 mRNA in cells was assayed by fluorescent quantitative PCR.Cell apoptosis rates were detected by flow cytometry.Reactive oxygen species (ROS) content was identified by 2', 7'-dichlorofluorescein diacetate (DCFH-DA) fluorescent probe method.Superoxide dismutase (SOD) activity and malondialdehyde (MDA) concentration were measured by ELISA method.The targeting of SIRT1 by miR-155 was tested by dual luciferase reporter gene system.Expressions of SIRT1, B-cell lymphoma/leukemia-2 gene (bcl-2), bcl-2 associated X protein (bax), cleaved-cysteine aspartase 3 (cleaved-Caspase-3) proteins were determined by Western blot. Results:With the increase of H 2O 2 concentration, the cell viability gradually decreased, and the differences in cell viability among different concentrations were statistically significant (all at P<0.05), and 100 μmol/L was selected as the experimental concentration.Cells in blank control group grew well adherently.The number of cells in model control group decreased, and the morphology of some surviving cells changed, and their boundaries were blurred.There were fewer cells in miR-155 mimics group than model control group, and the cell morphology changed.There were more cells in miR-155 inhibitor group than model control group, and the cells grew well.Compared with model control group, the relative expression level of miR-155, the apoptosis rate, ROS content, MDA concentration, as well as the relative expression levels of bax and cleaved-Caspase-3 proteins were increased, and the relative expression level of SIRT1 mRNA, the SOD activity, the relative expression of SIRT1 and bcl-2 proteins, as well as bcl-2/bax were decreased in miR-155 mimics group, and the differences were statistically significant (all at P<0.05). Compared with model control group, the relative expression of miR-155, the apoptosis rate, ROS content, MDA concentration, as well as the relative expression levels of bax and cleaved-Caspase-3 proteins were decreased, and the relative expression level of SIRT1 mRNA, SOD activity, the relative expression levels of SIRT1 and bcl-2 protein, as well as bcl-2/bax were significantly increased in miR-155 inhibitor group, and the differences were statistically significant (all at P<0.05). The relative luciferase activity of wild-type SIRT1 in cells transfected with miR-155 mimics was 0.41±0.07, which was significantly weaker than 1.00±0.11 in cells transfected with miR-155 mimics negative control, and the relative luciferase activity of wild-type SIRT1 in cells transfected with miR-155 inhibitor was 1.98±0.17, which was significantly higher than 1.00±0.12 in cells transfected with miR-155 inhibitor negative control, showing statistically significant differences ( t=7.838, 8.157; both at P<0.05). No obvious effect on the relative luciferase activity of mutant SIRT1 was found in transfected cells. Conclusions:miR-155 is involved in H 2O 2-induced oxidative damage of LECs, and its overexpression can target the expression of SIRT1 and play a role in cell injury.
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Although microRNA-155 (miR-155) is considered a pro-inflammatory mediator, cumulative evidence indicates that it also has anti-inflammatory effects in macrophages and dendritic cells. In this study, we identified the dramatic expression changes of more than half of potential miR-155-targeted genes upon lipopolysaccharide (LPS) stimulation; 223 genes were down-regulated and 85 genes were up-regulated, including suppressor of cytokine signaling 1 (
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Objective: To discuss the proliferation inhibition and apoptosis induction of shikonin on the FMS-like tyrosine kinase-3 receptor internal tandem duplication (FLT3-ITD) mutated acute myeloid leukemia (AMD MV4-11 cells, and to preliminarily clarify the molecular mechanisms. Methods: The MV4-11 cells were divided into DMSO group and different concentrations (0.5, 1.0, 2.0, 4.0, and 8.0 μmol · L-1) of shikonin groups, and treated for 24 and 48 h. The inhibitory rate of proliferation was analyzed by CCK-8 assay, and half inhibitory concentration (IC50) was calculated. The MV4-11 cells were divided into blank control group, DMSO group, and different concentrations (0.25, 0.50, and 1. 00 μmol · L-1) of shikonin groups, and treated for 48 and 72 h; the proliferation rate of cells was analyzed by carbox fluorescenceindiacetate succinimidyl este (CFSE). The MV4-11 cells were divided into DMSO group and different concentrations (0.702, 1. 404, and 2. 808 μmol · L-1) of shikonin groups, and treated for 48 h; the apoptotic rate was determined by flow cytometry. The MV4-11 cells were divided into DMSO group and different concentrations (0.351, 0.702, and 1. 404 μmol · L-1) of shikonin groups, and treated for 48 h; the microRNA-155 (miR-155) expression level was detected by Real-time PCR. Results: The results of CCK-8 and CFSE methods indicated that the inhibitory rates of proliferation of MV4-11 cells in different concentrations of shikonin groups were increased compared with DMSO grpup (P<0.05 or P<0.01), and the proliferation rates were decreased (P<0.05 or P<0.01) in a concentration-dependent manner; the IC50 of 24 and 48 h were 1. 743 and 1. 404 μmol · L-1, respectively. The flow cytometry results showed that the apoptotic rates of the cells in different concentrations of shikonin groups were increased compared with DMSO group (P<0.01) in a concentration-dependent manner. The Real-time PCR results showed that the expression levels of miR-155 in the cells in different concentrations of shikonin groups were decreased significantly (P<0.01), and the expression level in 1.404 μmol · L-1 shikonin group was decreased by more than 75%. Conclusion: Shikonin could inhibit the proliferation and promote the apoptosis of FLT3-ITD mutated AML MV4-11 cells, and down-regulate the expression of miR-155, suggesting that shikonin may be one of the potential therapeutic drugs for FLT3-ITD mututed AML.
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Objective@#Modern medical research has proven that human diseases are directly or indirectly related to genes. At the same time, genetic research has also brought updates to diagnostic techniques. Olfactomedin-like 3 (OLFML3) gene is a novel and clinically valuable gene. In order to better understand the role of OLFML3 in human diseases, we discuss and analyze the characteristics, function, and regulation mechanism of the OLFML3 gene in this review.@*Data sources@#A comprehensive search in PubMed and ScienceDirect database for English up to March 2019, with the keywords of "Olfactomedin-like 3," "Olfactomedin," "extracellular matrix," "Transforming Growth Factor β1," "anoikis-resistance," and "microRNA-155."@*Study selection@#Careful review of all relevant literature, the references of the retrieved articles were also screened to search for potentially relevant papers.@*Results@#OLFML3 is a secreted glycoprotein with 406 amino acid residues, belonging to the Olfactomedin (OLF) family. Due to the particularity of its structure and differential expression, OLFML3 has unique biological functions that could be distinct from other members in the OLF family. The currently known functions include embryonic development function and tumorigenesis. The regulation mechanism is still under investigation. It is directly related to many human diseases.@*Conclusions@#OLFML3 is a multifunctional glycoprotein that is closely involved in embryonic development, tumor invasion, and metastasis. Unfortunately, current research on this important molecule is still very limited. Further investigations on the possible mechanism of OLFML3 biological functions and modulation will help us develop better diagnostics and treatments.
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Objective: To investigate the expression of microRNA-155 (miRNA-155) in the cerebral cortex tissue of the rats with cerebral ischemia-reperfusion injury, and to clarify the effect of miRNA-155 on cerebral ischemiareperfusion injury. Methods: A total of 48 healthy adult male SD rats were randomly divided into sham operation group (n=24) and cerebral ischemia reperfusion group (I/R group, n=24). The cerebral ischemia-reperfusion models in I/R group were established by Longa modified suture occlusion in the right middle cerebral artery of the rats. The rats in sham operation group only received blood vessel isolation. TTC staining was used to calculate the ischemic infarction volume of the rats in various groups. qRT-PCR method was used to detect the expression levels of miR-155 in cerebral cortex tissue of the rats in various groups. Results; Compared with sham operation group, the ischemic infarction volume of the rats in I/R group was significantly increased at 24, 48, and 72 h after reperfusion (P<0. 05); the ischemic infarction volumes of the rats in I/R group were decreased gradually with the prolongation of reperfusion time. Compared with sham operation group, the expression levels of miR-155 in cerebral cortex tissue of the rats in I/R group were significantly increased at 24, 48, and 72 h after reperfusion (P<0.05); the expression levels of miR-155 were gradually decreased with the prolongation of reperfusion time. Conclusion: The expression levels of miR-155 in cerebral cortex tissue of the rats with cerebral ischemia-reperfusion injury are high, and miR-155 may participate in the process of cerebral ischemia-reperfusion injury.
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ObjectiveTo investigate the protective effect of microRNA-155 (miR-155) antisense oligonucleotid (ASO) on acute lung injury (ALI) mice by establishing a lentiviral expression vector of ASO of miRNA.Methods miR-155 antisense oligonucleotides amplified by polymerase chain reaction (PCR) from genomic, using BamH Ⅰ and Nhe Ⅰ double digestion, ligated into lentiviral expression vector. Sequence and virus titer were measured. According to the random number table method, 54 male BALB/c mice of 4-6 weeks old were divided into three groups. ALI animal models were prepared by intraperitoneal injection of 10 mg/kg lipopolysaccharide (LPS). The three groups were injected with 200μL phosphate buffered saline (PBS) containing 1×108/mL pmiR-155-ASO virus (pmiR-155-ASO group) or 200μL PBS containing 1×108/mL pSMPUW-miR-GFP empty virus (pmiR-cont group) or the same amount of PBS (PBS group) at 24 hours before the molding. Ten mice in each group were used to observe the 7-day survival rate. Blood samples and lung tissues of the remaining 8 mice were harvested after the model was established, and the levels of serum inflammatory cytokines were determined by enzyme linked immunosorbent assay (ELISA); the expression of miR-155 in lung tissue was detected by real-time reverse transcription-polymerase chain reaction (RT-PCR); histopathological changes of lung and distribution of macrophages were observed under microscope.Results There was no significant difference in each index between pmiR-cont group and PBS group. The mature miR-155 expression in lung tissue in pmiR-155-ASO group was significantly lower than that in pmiR-cont group (2-ΔΔCt: 4.92±0.72 vs. 15.38±0.60,P < 0.05). Compared with pmiR-cont group, the injury degree of ALI mice after pretreatment with miR-155ASO was significantly improved, and the 7-day survival rate was significantly increased (72.1％ vs. 61.9％,P < 0.05 ); gross lung observation showed that congestion in lung tissue was significantly reduced, and the ratio of wet/dry weight (W/D) of lung was significantly decreased (4.50±0.13 vs. 5.64±0.61,P < 0.05);hematoxylin-eosin (HE) staining showed that inflammatory cell infiltration in lung tissue was decreased, while immunofluorescence assay showed that macrophage infiltration in lung tissue was significant decreased; the levels of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL- 6) in serum were significantly decreased [TNF-α (ng/L):379.8±48.9 vs. 495.9±33.3, IL-6 (ng/L): 262.3±61.8 vs. 355.4±22.6, bothP < 0.05], but the level of IL-10 did not change significantly (ng/L: 143.6±32.5 vs. 140.4±22.3,P > 0.05).Conclusion miR-155 ASO has the effect of inhibiting LPS-induced inflammatory response and improving prognosis in ALI mice.
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Objective To observe the effect of microRNA-155 (miR-155) on the inflammatory response of rat alveolar macrophages induced by lipopolysaccharide (LPS). Methods The alveolar macrophages NR8383 of rat were cultured in vitro, the macrophages in logarithmic growth phase were harvested to conduct experiment. ① The 1 mg/L LPS was used to stimulate the rat alveolar macrophages for 3, 6, 12, and 24 hours, a phosphate buffer solution (PBS) control group was also set up. Enzyme linked immunosorbent assay (ELISA) was used to detect the dynamic changes of interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) in the supernatant, and real-time quantitative reverse transcription-polymerase chain reaction (RT-qPCR) was used to detect the dynamics expression of miR-155 in the cells, which confirmed the optimal time for LPS stimulation was 12 hours. ② Carboxyfluorescein (FAM) labeled mimic (FAM mimic) and inhibitor (FAM inhibitor) were used to transfect the alveolar macrophage, and the transfection effect was observed under inverted fluorescence microscope 6 hours later to confirm the optimal transfection concentration of mimic was 20 nmol/L, and the optimal transfection concentration of inhibitor was 100 nmol/L. miR-155 mimic and miR-155 inhibitor were transfected to alveolar macrophages respectively at the optimal transfection concentration for 24 hours, and 1 mg/L LPS was used to stimulate the cells for 12 hours. A mimic negative control + LPS group and an inhibitor negative control + LPS group were set up. The expressions of IL-1β and TNF-α in the supernatant were determined by ELISA to observe the regulation of miR-155 on inflammatory response of alveolar macrophages. Results ① After stimulation of 1 mg/L LPS on alveolar macrophages, the contents of IL-1β and TNF-α in the supernatant and the expression of miR-155 in the cells were increased gradually with time prolongation, IL-1β and TNF-α contents peaked at 12 hours, and the expression of miR-155 peaked at 24 hours [as compared with PBS control group, IL-1β (ng/L): 910.43±36.09 vs. 22.66±7.84, TNF-α (ng/L): 3 138.39±394.10 vs. 233.92±8.84, miR-155 (2-ΔΔCt): 7.82±0.30 vs. 1, all P < 0.05]. ② Under inverted fluorescence microscope, after 20 nmol/L FAM mimic or 100 nmol/L FAM inhibitor transfected alveolar macrophages for 6 hours, a large number of cells showed green fluorescence, indicating that the transfection was successful. The expression of miR-155 in the cells transfected with 20 nmol/L miR-155 mimic was up-regulated by (236.73±46.49) times as much as that in the negative control group (P < 0.05), and the levels of IL-1β and TNF-α in the supernatant of the cells stimulated by 1 mg/L LPS for 12 hours were significantly lower than those in the negative control group [IL-1β (ng/L): 324.37±36.59 vs. 799.31±39.44, TNF-α (ng/L): 1 554.01±342.48 vs. 3 020.49±418.30, both P < 0.05]. The miR-155 activity was significantly inhibited in the cells transfected with 100 nmol/L miR-155 inhibitor, and the expression of miR-155 was decreased by (4.00±3.26)% as compared with the negative control group, but the difference was not statistically significant (P > 0.05), and the levels of IL-1β and TNF-α in the supernatant of the cells stimulated by 1 mg/L LPS for 12 hours were significantly higher than those in the negative control group [IL-1β (ng/L): 1 358.98±212.04 vs. 878.68±53.42, TNF-α (ng/L): 4 225.57±281.11 vs. 2 881.32±286.08, both P < 0.05]. Conclusion In LPS induced inflammatory response of alveolar macrophages, miR-155 plays an obvious inhibitory role.
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Objective To explore the expression of miRNA-155 (miR-155) in CD19+B lymphocytes in Uygur patients with chronic lymphocytic leukemia (CLL), and to analyze its clinical significance. Methods Peripheral blood(10-15 ml) samples of 30 Uygur CLL patients, 20 Han CLL patients, 10 Han healthy people and 10 Uygur healthy people were collected.Mononuclear cells were isolated and B lymphocytes were purified by using CD19+magnetic-bead system. Total RNA was extracted from CD19+B lymphocytes. The expression of miR-155 was measured by using TaqMan probe method. All the results were analyze combined with the clinical data of patients. Results The expression level of miR-155 in Han and Uygur CLL patients had no statistical difference(5.994±1.156 vs.5.856±1.359,t =0.556,P=0.710), which was significantly higher than that in the healthy controls(t = 18.612, P < 0.01; t = 17.817, P < 0.01). There were no statistical differences in the expression levels of miR-155 in CLL patients regarding to nationalities, gender, age, Binet staging, Rai staging, expression of CD38 (all P> 0.05). The expression level of miR-155 had a significant statistical difference between ZAP-70 negative group and ZAP-70 positive group(5.7 ± 1.1 vs. 6.5 ± 1.4,t =1.575,P =0.029). The expression level of miR-155 also had a statistical difference between mutated immunoglobulin heavy chain variable region (IGHV) and unmutated IGHV (5.3 ± 1.1 vs. 6.5 ± 1.1, t = 0.210, P= 0.001).The expression level of miR-155 was 5.4 ± 1.2, 5.7 ± 0.7, and 6.6 ± 1.5 in the favorable prognosis group, moderate prognosis group and poor prognosis group respectively(F=3.609,P=0.035); The expression levels of miR-155 had significant differences between moderate group and poor prognosis group (P = 0.032), favorable group and poor prognosis group (P= 0.007). Conclusions The expression of miR-155 in Han and Uygur CLL patients has no significant difference.High-level expression of miR-155 is associated with the poor prognosis of CLL patients.
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Objective To investigate the effects of antisense oligonucleotide against miRNA-155 (AS-miRNA-155) on proliferation,apoptosis and invasion and migration abilities of human cutaneous squamous cell carcinoma cell line A431. Methods AS-miRNA-155 was transfected into human cutaneous squamous cell carcinoma A431 cells by using LipofectamineTM 2000. Blank control without transfection and transfected with non-sense sequence were used as non-sense sequence control. Quantitative real-time polymerase chain reaction (qRT-PCR) was performed to detect the expression of miRNA-155 in A431 cells. Cell proliferation was analyzed using dimethyl thiazolyldiphenyl tetrazolium (MTT) assay. Cell cycle arrest and apoptosis were studied by flow cytometry (FCM). Invasion and migration were measured by Transwell chamber assays. Results The relative expression of miRNA-155 mRNA was lower in the transfection group than that in the blank control group and the negative control group (F=634.57, P<0.01), but there was no significant difference between the blank control group and the negative control group. After 72 h transfection, the survival rate was significantly lower in the transfection group than that of the blank control group and the negative control group, and the transfection rate decreased significantly by 120 h (P<0.05). Cells of G0/G1 phase increased, Cells of S phase reduced, the overall PI value decreased in transfection group, and the apoptosis rate of A431 cells, migration and invasion of cells increased (P<0.05). There was no significant difference in G2/M cycle between transfection group, blank control group and negative control group. There were no significant differences in A431 cell apoptosis rate, cell migration and invasive ability between blank control group and negative control group. Conclusion Antisense oligonucleotide against miRNA-155 can inhibit the expression of miRNA-155, the proliferation and promote the apoptosis of human cutaneous squamous cell carcinoma A431 cells, which indicates that miRNA-155 may become a new target for the regulation of gene expression in cutaneous squamous cell carcinoma.
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Objective To investigate the expression of miR-155 ,nuclear factor kappa B (NF-κB) and soluble intercellular adhesion molecule -1 (sICAM-1 ) in peripheral blood in patients with type 2 diabetes mellitus (T2DM ) ,and to explore the role of miR-155 in vascular lesions of T2DM. Methods A total of 165 T2DM patients and 60 health subjects from health examinations were enrolled in this study. All the subjects were divided into two groups :subjects without vascular lesions group and vascular disease group. Vascular disease group was further divided into microangiopathy group ,macroangiopathy group , microangiopathy+ macroangiopathy group ;based on 24 hUAlb level normal urinary albumin (NAU ) group ,microalbuminuria (MAU) group ,macroalbuminuria (MAAU) group. MiR-155 ,NF-κB and sICAM-1 in peripheral blood were tested by RT-PCR. Single factor analysis of variance was used for comparison among groups. Stepwise regression analysis was used for correlation factors analysis. Results The expression of miR-155 ,NF-κB and sICAM-1 were significantly higher in T2DM group than in NC group [(1.82 ± 0.71) vs (1.00 ± 0.12) ,(2.28 ± 0.66) vs (1.04 ± 0.33) ,(1.88 ± 0.80) vs (1.03 ± 0.30) ,P<0.05]. The level of miR-155 ,NF-κB ,sICAM-1 were significantly higher in T2DM vascular lesions group than in subjects without vascular lesions group [(1.95 ± 0.73) vs (1.34 ± 0.29) ,(2.40 ± 0.65) vs (1.65 ± 0.16 ) ,(2.01 ± 0.77 ) vs (1.07 ± 0.41 ) ,P < 0.05 ]. The expression of miR-155 was higher in microangiopathy+ macroangiopathy group than in macroangiopathy group [(2.36 ± 0.61 ) vs (1.77 ± 0.59) ,P < 0.05 ].The expression of NF-κB was also significantly higher in microangiopathy +macroangiopathy group than in microvascular disease group and macroangiopathy group (P<0.05). The level of miR-155 was significantly higher in group MAU group and MAAU group than in NAU group [(1.41 ± 0.49) ,(2.64 ± 0.52) vs (1.04 ± 0.20) ,P<0.05] ,and NF-κB and sICAM-1 were also higher than NAU group (P<0.05). Multiple stepwise regression analysis showed that miR-155 was positively correlated with NF-κB and sICAM-1(t=4.235 ,9.728 ,P<0.01). Conclusion The expression of miR-155 increases in T2DM patients with vascular complications ,and this trend is the same as NF-κB and sICAM-1. It suggests that miR-155 maybe involves in the pathogenesis of diabetic chronic vascular disease.
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Objective To discuss the influence of microRNA(miR)-155/miR-21 on toll-like receptor 4 (TLR4) in children with sepsis.Methods Fifty children with sepsis who were hospita-lized in Pediatric Intensive Care Unit,Shenzhen Children's Hospital,were enrolled in the study,and 15 healthy children at the same age were selected as healthy control group.Expression levels of TLR4 protein and human leukocyte antigen(HLA)-DR in CD14 + monocytes (MC) were detected by using flow cytometry,and sepsis patients were divided into 2 groups according to whether they exceeded the value of HLA-DR by 30％ or not.Expression level of programmed cell death factor 4 (PDCDM) and inositol phosphatases 1 containing SH2 (SHIP1) were detected at the same time.MC were separated by CD14 + immune magnetic bead,and expression level of miR-155,miR-21 and tumor necrosis factor-α (TNF-α),interleukin-10 (IL-10) mRNA in CD14 + MC were detected by using real-time fluorescent quantitative PCR.Results Sepsis group consisted of 27 male and 23 female,and their ages were (2.34 ± 0.79) years old,among whom 9 patients died.There were 36 patients in the HLA-DR increase group and 14 patients in the HLA-DR decrease group.Expressions ofTLR4(2.33±0.90),miR-155[(7.19±3.75) ×10 3] and TNF-α[(21.98±14.15) ×10-2 pg/L] in CD14 + MC were higher in the HLA-DR increase group than those in the HLA-DR decrease group [1.24±0.60,(4.83 ±1.17) × 10-3,(14.18±5.45) ×10-2 μg/L] and healthy control group[1.57±0.55,(3.99 ± 1.29) × 10-3,(1.61 ± 0.84) × 10 2 pg/L],and the differences were statistically significant(F =11.943,7.583,18.538,all P ＜0.05),while the expressions of miR-21 (12.10 ±5.66),IL-10[(29.74 ± 12.55) × 10-4 μg/L] in CD14 + MC were lower in the HLA-DR increase group than those in the HLA-DR decrease group[4.68 ± 2.07,(12.50 ± 5.73) × 10-4 μg/L] and healthy control group [2.39 ± 0.86,(2.04 ± 0.92) × 10-4 μg/L],and the differences were statistically significant(F =41.673,54.991,all P ＜ 0.05).The levels of SH1P1 and PDCD4 decreased in sepsis compared with healthy control group[0.70 ±0.36)vs.(1.59 ±0.48);(1.55 ±0.56) vs.(3.01 ±0.70)],and the differences were statistically significant (t =7.682,8.339,all P ＜ 0.05),but SHIP1 decreased more significantly in the HLA-DR increase group than that in the HLA-DR decrease group [(0.60 ± 0.34) vs.(0.97 ± 0.26)],and the difference was statistically significant (F =39.214,P ＜ 0.05).PDCD4 decreased more significantly in the HLA-DR decrease group than that in the HLA-DR increase group (0.94 ±0.19 vs.1.79 ±0.47),the difference was statistically significant(F =65.367,P ＜ 0.05).Conclusions Regulation imbalance of miR-155/miR-21 may be one of the reasons for abnormal expression of TLR4 in children with sepsis,and it plays a role in enlarged or inhibited expression of TLR4 in the sepsis process which results in different immune status in sepsis patients.
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Objective To construct a Luciferace reporter vector containing the 3'untranslated region (3'UTR) of NFAT5 and measure the correlation between NFAT5 and miR-155.Methods The miR-155 targeting NFAT5 3'UTR was predicted by Target Scan,Mir Base and Pic Tar.NFAT5 and mutant NFAT5 sequence(NFAT5-mu) were then designed and synthesized,and they were cloned into pMIR-REPORTTM Luciferace reporter vector.Human embryonic kidney-293AD (HEK-293AD) cells of the 4th passage were divided into 4 groups according to the random number table.cells in plasimd +miR-155 mimics groups were transfected with pMIR-NFAT5 recombinant plasimid,pRL-Tk plasmid and miR-155 mimics;cells in plasimd + miR-155 mutated groups were transfected with pMIR-NFAT5-mu recombinant plasimid,pRL-Tk plasmid and miR-155 mimics;cells in plasimd + miR-155 control groups were transfected with pMIR-NFAT5 recombinant plasimid,pRL-Tk plasmid and miR-155 Negative control;cells in plasimd +miR-155 inhibitor were transfected with pMIR-NFAT5 recombinant plasimid,pRL-Tk plasmid and miR-155 inhibitor;and were respectively transfected into together by liposome.After culture for 24 h,the luciferase activity was detected by dual luciferase reporter assay system.Results TargetScan,Miranda and PicTar shared the results that NFAT5 has the complementary binding sites with 3'UTR of miR-155.And luciferase reporter vectorwas constructed.Therefore the result of sequencing and double digesting of recombined plasmid were completely correct.Dual-luciferase reporter assay showed that miR-155 possesses a target effect on 3'UTR of NFAT5.Compared to the pMIR-NFAT5 + miR-control group,the luciferase activity of the pMIR-NFAT5 + miR-1 5 5 mimics group was decreased,with statistically significant difference(P＜0.01),while there was no significant difference at other time points(P＞0.05).Conclusion The pMIR-NFAT5 recombinant plasmid and pMIR-NFAT5 recombinant mutated plasmid were confirmed with successful construction.and it was found that miR-155 can target NFAT5 mRNA 3'-UTR.The results provide the experiment data for further disclosing the mechanism of inhalation injury on the level of gene expression.
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Objective Toexplore the expression and significance of microRNA-155 (miR-155) in psoriasis vulgaris. Methods Areal-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) method with TaqMan probe technology was performed to detect miR-155 expression in skin lesion area and nonskinlesionalarea of 35 patients with psoriasis vulgaris , compared with that of 30 normal controls. The correlations among miR-155 expression, psoriasis area and severity index (PASI) score and Interleukin-17A(IL-17A)expression were studied. Results The expressions of miR-155 and IL-17A in lesional and non-lesionalgroups were higher than that of control group (all P < 0.01). Also expressions in lesional skin were higher than non-lesional skin (both P < 0.01). In skin lesion group, significant positive correlations existed betweenmiR-155 or IL-17A expression and PASI score as well as miR-155 and IL-17A expression (all P < 0.05). Conclusions Up-expression of miR-155 was relevant to psoriasis development , which is related withthe hyperfunctionof Th17 cells in psoriasis.
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Objective To analyze the microRNA-155 (miR-155) expression in hepatocellular carcinoma (HCC),and to assess its correlation with clinicopathological parameters and its prognostic significance.Methods MiR-155 expression was detected in specimens of HCC and its adjacent non-tumorous liver tissues in 124 HCC patients by real-time PCR.The expression was also detected in different HCC cell lines.The Kaplan-Meier Curve was used to analyze survival of HCC patients.Gain-and loss-of-function studies were used to determine whether miR-155 impact cell cycle,cell proliferation and apoptosis of HCC cells.Results Expression of miR-155 in HCC tissues was significantly elevated when compared with that in the adjacent non-tumor liver tissues (P ＜ 0.05).MiR-155 expression was significantly higher in different HCC cell lines (P ＜ 0.05) than that in normal liver cells.MiR-155 overexpression was significantly correlated with large tumor size (P ＜ 0.05),high histological grade (P ＜ 0.05) and advanced tumor stage (P ＜ 0.05).Patients with high miR-155 expression had significantly decreased overall survival (P ＜ 0.05) and disease-free survival (P ＜ 0.05).Upregnlation of miR-155 promoted cell cycle transition from G1 to S phase (P ＜ 0.05),increased cell proliferation (P ＜ 0.05) and apoptosis resistance (P ＜ 0.05) in Hep3B cells.Downregulation of miR-155 resulted in G1 arrest,apoptosis promotion and proliferation reduction in SMMC-7721 cells (P ＜ 0.05).Conclusion MiR-155 can be used as a prognostic marker for HCC patients,and target therapy on miR-155 can be used as a potential option to prevent HCC progression.
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Objective: To investigate the effect of a method for imaging lung cancer cells using nanotechnology and molecular beacon (MB) that identifies miR-155 and is delivered by chitosan nanoparticles (CS). Methods: The miR-155 MB modified by locked nucleic acids (LNAs) was designed and synthesized. The CS-MB complex was synthesized by self-assembly method and tested for its physicochemical properties including anti-DNase I features, particle size, zeta potential and so on. The miR-155 MB was transfected with CS as vectors. The abilities of miR-155 MB to identify miR-155 and thus to image lung cancer cells were determined by confocal microscopy. Furthermore, the miR-155 expression levels were detected by qRT-PCR to validated the effect of miR-155 MB. The random sequence molecular beacon (RS MB) was set as a negative control. Results: The CS-MB complex at the weight ratio of 7:1 was best suited for transfection due to its high encapsulation rate, resistance to the degradation by DNase I, small particle size and positive charge. Relatively strong red fluorescence could be detected in the lung cancer cells after transfection of miR155 MB, while that could not be detected in the RS MB group (P<0.05). Moreover, the changing trend in the fluorescence intensity was consistent with that in the miR-155 expression levels. Conclusion: CS nanoparticles can be used as vectors to deliver miR-155 MB for miR-155 identification and lung cancer cell imaging, thus providing new ideas and novel technique for lung cancer diagnosis.
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Objective To investigate the adjustment of miRNA-155 on CD4+ CD25+ Treg regulative T cell in peripheral blood in patients with acute cerebral infarction (ACI) and its pathogenesis. Methods Sixty patients with ACI were divided into three groups according to clinical neurological deficit score. Twenty healthy volunteers were enrolled into the control group. The expression levels of plasma miR-155 mRNA and Foxp3 mRNA were detected by real-time quantitative PCR(qRT-PCR). IL-10 levels in plasma were detected by ELISA. Results Expression of miR-155, Treg, Foxp3 mRNA and levels of IL-10 were significantly increased in patients with ACI compared with normal control group, with statistical differences; Expression of miR-155, Treg, Foxp3 mRNA and levels of IL-10 were gradually increased. The values showed significant statistical difference among the mild, moderate and severe ACI groups (P < 0.01). Among the patients,the levels of miR-155, Treg, Foxp3 mRNA and levels of IL-10 in the survival group were obviously lower than those in the non (P<0.05 or P<0.01). There was a positive correlation between miR-155 and Treg, Foxp3 mRNA (P < 0.01). Conclusion This study suggests that miR-155 is involved in the cell proliferation regulation of CD4+ CD25+ Treg cells,and plays some role in the immunological dissonance with ACI.
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Objective To investigate the effects of microRNA-155 (miR-155) inhibitor on JAK/STAT1 (Janus kinase/signal transducer and activator transcription 1) signaling pathways in the injured lung tissue induced by lipopolysaccharide (LPS).Methods One hundred and twenty BALB/c mice were randomly divided into control group (n =40),LPS group (n =40),and inhibitor + LPS group (n =40).LPS group and inhibitor + LPS group were made by injection of LPS 20 mg/kg intra-peritonealy,whereas equivalent volume of normal saline was given instead in the control group.The 80 mg/kg of miR-155 inhibitor was injected into caudal vein 24 h before LPS injection in inhibitor + LPS group.Mice were sacrificed at 6 h,12 h,24 h,and 48h separately after LPS injection,and lung tissue were collected.The levels of tumor necrosis factor-α (TNF-α) and interleukin-10 (IL-10) of lung tissue were measured using the enzyme-linked immunosorbent assay (ELISA).Using histopathological examination,the injury of lung tissue was evaluated.The expressions of miR-155,STAT1 mRNA,SOCS1 mRNA in lung tissue were assayed by fluorescent quantitative reverse transcription polymerase chain reaction (qRT-PCR).Results The miR-155 expression induced by LPS increased at 6 h,12 h,24 h and decreased at 48 h.The miR-155 expressions in LPS group were (8.52 ± 1.12) at 6 h,(11.04 ±0.99) at 12 h,(15.84 ±0.80) at 24 h and (4.03 ± 2.55) at 48 h.In the inhibitor + LPS group,the expressions of miR-155 were lower compared with LPS group,showing significant differences at 12 h (t =6.08,P ＜ 0.01),and at 24 h (t =23.64,P ＜ 0.01).STAT1 mRNA and SOCS1 mRNA both reached peak levels at 6 h after LPS injection.The levels of STAT1 mRNA in LPS group were higher than those in inhibitor + LPS group,showing significant differencesat6h (t=4.41,P＜0.01),12h(t=2.69,P＜0.05),and24h (t=3.62,P＜0.01).The levels of SOCS1 mRNA in inhibitor + LPS group were higher than those in LPS group,showing significant differences at 6 h (t =4.55,P ＜0.01),12 h (t =4.12,P ＜0.01),24 h (t =2.38,P ＜ 0.05).TNF-α reached its peak value at 6 hours and IL-10 reached its peak value at 48 hours.Both TNF-α and IL-10 were higher in LPS group than those in inhibitor + LPS group showing significant differences at 6 h,12 h,24 h (P ＜0.01).The pathologic examination indicated the lung injury in inhibitor + LPS group was milder than that in LPS group.Conclusion The miR-155 increased in lung tissue of endotoxemic mice.miR-155 inhibitor may suppress JAK/STAT1 signaling pathway and protect the lung tissue.