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Abstract Objective: Long Non-Coding RNAs (LncRNAs) act as an indispensable role in cancer development. The study aimed to investigate the role and mechanism of lncRNA Small Nucleolar RNA Host Gene 1 (SNHG1) in Bladder Cancer (BC) progression. Method: The expression, prognostic value, diagnostic value, and correlation of SNHG1, Enhancer of Zeste 2 polycomb repressive complex 2 subunit (EZH2), and Kruppel Like Factor 2 (KLF2) were analyzed through bioinformatics analysis. The expression was also validated in BC tissues and cell lines. Besides, their regulation and binding were tested via qPCR, Western blot, Dual-Luciferase Reporter Assay (DLRA), Argonaute RISC catalytic component 2-RNA Immunoprecipitation (AGO2-RIP), and Chromatin Immunoprecipitation (ChIP). A xenograft model in nude mice was also established. Results: SNHG1 was significantly overexpressed in BC tissues and cells. Importantly, SNHG1 was associated with poor survival, and ROC curves revealed high diagnostic values. Moreover, by CCK8, wound healing, transwell, and Western blot analysis, SNHG1 knockdown significantly inhibited the proliferation, migration, invasion, and epithelial-mesenchymal transition of BC cells. Additionally, in vivo experiments showed that silencing SNHG1 hindered tumorigenesis and tumor growth. Regarding mechanism, the results of AGO2-RIP, ChIP or DLRA showed that SNHG1 played different roles at diverse subcellular sites. In the cytoplasm, SNHG1 acted as a competing endogenous RNA for miR-137-3p to promote EZH2 expression. In the nucleus, SNHG1 could interact with EZH2 to inhibit KLF2 transcription. Conclusion: Our study elucidated that SNHG1 formed a regulatory network and played an oncogenic role in BC, which provided a novel therapeutic target for BC treatment.
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Objective:To investigate the effect of propofol on proliferation, invasion and migration of human melanoma cell line A375 via miR-137/fibroblast growth factor 9 (FGF9) pathway.Methods:A375 cells were cultured in vitro. The half inhibitory concentration and half inhibitory time of propofol on A375 cells were determined by methyl thiazolyl tetrazolium (MTT) method. miR-137 mimics and miR-137 inhibitors were transfected into A375 cells by lipofectamine method. A375 cells were divided into control group, propofol group, miR-137 mimics group, propofol+ miR-137 mimics group, miR-137 inhibitors group and propofol+ miR-137 inhibitors group. After treated with the optimal time and concentration, the mRNA expression of miR-137 and FGF9 was detected by real time fluorescence quantitative reverse transcription polymerase chain reaction (qRT-PCR), and the invasion and migration ability of cells in vitro was detected by transwell method. Results:With the increase of propofol concentration, the proliferation rate of A375 cells was gradually decreased, and 80 μmol/L was selected as the half inhibition concentration. With the increase of propofol action time, the proliferation rate of A375 cells was gradually decreased, and 48 hours was selected as the half inhibition time. Compared with the control group, propofol could promote the expression of miR-137 mRNA, inhibit the expression of FGF9 mRNA, and inhibit the invasion and migration of A375 cells ( P<0.05). miR-137 mimics could promote the expression of miR-137 mRNA, inhibit the expression of FGF9 mRNA, and inhibit the invasion and migration of A375 cells. At the same time, after propofol intervention, the effect of promoting the expression of miR-137 mRNA, inhibiting the expression of FGF9 mRNA and inhibiting the invasion and migration of A375 cells was more significant ( P<0.05); miR-137 inhibitors could inhibit the expression of miR-137 mRNA, promote the expression of FGF9 mRNA, and promote the invasion and migration of A375 cells ( P<0.05). At the same time, after propofol intervention, the effects of inhibiting the expression of miR-137 mRNA, promoting the expression of FGF9 mRNA and promoting the invasion and migration of A375 cells were inhibited ( P<0.05). Conclusions:Propofol can inhibit the proliferation, invasion and migration of human melanoma cell line A375. The mechanism may be related to the inhibition of miR-137/FGF9 pathway activation by propofol.
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Objective: To investigate the effect of pleiotrophin (PTN) on the chemotherapy resistance of osteosarcoma cells, and its possible molecular mechanism. Methods: The expression levels of PTN and microRNA (miR)-137-3p in osteosarcoma drug-resistant MG63/ADR cells and parental MG63 cells were detected by real-time fluorescent quantitative PCR. SiRNA-PTN or miR-137-3p mimic was transfected into MG63/ADR cells, while the PTN recombinant plasmid or miR-137-3p inhibitor was transfected into MG63 cells. After the transfection efficiency was verified by real-time fluorescent quantitative PCR, the cell proliferation activity was detected by CCK-8 and clony formation assay. The interaction between miR-1373p and the target gene PTN was verified by dual luciferase reporter gene system. The miR-137-3p mimic, siRNA-PTN or miR-137-3p mimic+PTN recombinant plasmid was respectively transfected into osteosarcoma MG63/ADR cells, then the expressions of PTN mRNA and protein were detected by real-time fluorescent quantitative PCR and Western blotting, and the cell viability was detected by CCK-8 method. Results: PTN was significantly highly expressed in drug-resistant MG63/ ADR cells (P < 0.01), while miR-137-3p was significantly lowly expressed (P < 0.01). After transfection with siRNA-PTN or miR-137-3p mimic, the proliferation and clone formation abilities of MG63/ADR cells were significantly reduced (all P < 0.01). After transfection with PTN vector or miR-137-3p inhibitor, the proliferation and clone formation abilities of parental MG63 cells were significantly increased (all P < 0.01). PTN was a downstream target gene of miR-137-3p, and miR-137-3p negatively regulated the expression of PTN gene (P < 0.01). After transfection with siRNA-PTN or miR-137-3p mimic, the expression levels of PTN mRNA and protein in MG63/ADR cells were reduced (both P < 0.01), but the overexpression of PTN could reverse the effect of miR-137-3p on the viability of osteosarcoma drug-resistant cells (P < 0.01). Conclusion: PTN can regulate the chemotherapy resistance of osteosarcoma, and its mechanism may be related to miR-137-3p downregulating PTN expression.
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Objective@#To assess the effect of miR-137 on the proliferation and apoptosis of human umbilical vein endothelial cells (HUVECs) induced by high glucose and its mechanism.@*Methods@#HUVECs cells were divided into low-glucose group (5.5 mmol/L glucose-treated cells), high-glucose group (33.36 mmol/L glucose-treated cells), anti-NC group (cells treated with 33.36 mmol/L glucose after anti-NC transfection) and anti-miR-137 group (cells treated with 33.36 mmol/L glucose after anti-miR-137 transfection). After 48 hours, qRT-PCR was used to determine the expression of miR-137. CCK-8 assay and flow cytometry were used to detect cell proliferation and apoptosis rate, respectively. The targeting relationship between miR-137 and AKT2 was validated by dual fluorescence reporter gene detection system and AKT2 protein expression after overexpression or inhibition of miR-137.@*Results@#High glucose could significantly up-regulate the expression of miR-137 in HUVECs cells, and the expression of miR-137 in HUVECs cells transfected with miR-137 inhibitor was significantly decreased (P<0.05). High glucose can significantly inhibit HUVECs cell proliferation and induce apoptosis, while inhibition of miR-137 expression can weaken the effect of high glucose on HUVECs cell proliferation inhibition and apoptosis promotion (P<0.05). Inhibiting AKT2 expression could weaken the inhibitory effect of miR-137 inhibitor on HUVECs cell proliferation and apoptosis (P<0.05).@*Conclusion@#Inhibiting the expression of miR-137 gene can attenuate the proliferation inhibition and apoptosis promotion of HUVECs induced by high glucose, and the mechanism is related to activating the expression of AKT2.
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Objective To explore the role of miR-137 in the proliferation and migration of hepatocellular carcinoma (HCC) cells by regulating Notch1 and mediating autophagy.Methods The human SMMC7721 hepatoma cell line was transfected with miR-137 mimics,miR-137 inhibitor and Notch1 interfering RNA (siRNA),and divided into normal control group (NC group),miR-137 mimics group,miR-137 inhibitor group,Notch1 siRNA group.The expression levels of miR-137 and Notch1 mRNA after the transfection were detected by RT-PCR in SMMC7721 cells.Transwell experiments were performed to analyze the effect of miR-137 and Notch1 on the migration and invasion of SMMC7721 cells.The expression levels of β-catenin and vimentin in SMMC7721 cells were detected by immunohistochemistry.The number of autophagosomes was detected by double labeled adenovirus.Western blot was utilized to detect the expression of Notch1,E-Cadherin,N-Cadherin,vimentin,P62,and LC3.Results The results of RT-PCR showed that the relative expression level of Notch1 in miR-137 inhibitor group (5.71 ± 0.45) was significantly higher than that in miR-137 mimics group (0.21 ± 0.06) with statistical significance (P ＜ 0.05).The Transwell experiments showed that there were fewer invasive metastatic hepatoma cells in miR-137 mimics group (66.00 ± 4.55) and Notch1 siRNA group (88.00 ± 6.78) than that in the miR-137 inhibitor group (515.00 ±35.12) (P ＜0.05).The expression levels of β-catenin in miR-137 mimics group and Notch1 siRNA group were significantly increased and the expression level of vimentin was decreased (P ＜ 0.05).The results of autophagy double labeled adenovirus test showed that the number of autophagosomes in miR-137 mimics group (5.50 ± 3.70) was significantly fewer than that in miR-137 inhibitor group (32.75 ± 4.11),and the difference was statistically significant (P ＜ 0.05).The expression levels of Notch1,N-cadherin,vimentin,and LC3 protein in miR-137 mimics group were much lower than that in miR-137 inhibitor group and NC group,and the expression levels of E-Cadherin and P62 protein were greatly increased.The expression level of Notch1,N-cadherin,and LC3 protein in Notch1 siRNA group were significantly lower than that in NC group,and the expression levels of E-cadherin and P62 protein were much higher than that in NC group.Conclusion MiR-137 can inhibit the proliferation,migration and invasion of HCC cells by inhibiting the expression of Notch1 and autophagy,which may become a new target for the treatment of HCC.
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Objective:To construct lentiviral vector which can overexpression miR-137 and produce lentivirus by lentivirus packaging system, and to explore its infection efficiency and expression in HEK293T cells.Methods: miR-137 sequence was chemically synthesized and cloned into lentiviral vector GV209, and the recombinant plasmid containing human miR-137 was obtained and identified.Then miR-137 recombinant plasmid together with two helper plasmids were transfected into HEK293T cells using Lipofectamine 2000.After the HEK293T cells were infected in multiplicity of infection(MOI) 40 for 48 h, the expression of green fluorescent protein (GFP) was observed by fluorescence microscope and the expression level of miR-137 was detected by fluorescence quantitative PCR.Results:The sequencing results showed that the inserted gene sequence was completely consistent with the published human miR-137 gene sequence in GenBank.The GFP was observed in the HEK293T cells infected with miR-137 overexpression lentivirus under fluorescence microscope.The fluorescence quantitative PCR results showed that the expression level of miR-137 in the cells infected with overexpression lentivirus was 12.74 times higher than that in the control cells.Conclusion:The lentivirus containing miR-137 gene is successful packaged, and it could efficiently infect the HEK293T cells.
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OBJECTIVE: Schizophrenia is a chronic neuropsychiatric disease afflicting around 1.1% of the population worldwide. Recently, MIR137, CACNA1C, CSMD1, DRD2, and GRM3 have been reported as the most robustly emerging candidates involved in the etiology of schizophrenia. In this case control study, we performed an association analysis of rs1625579 (MIR137), rs1006737, rs4765905 (CACNA1C), rs10503253 (CSMD1), rs1076560 (DRD2), rs12704290, rs6465084, and rs148754219 (GRM3) in Pakistani population. METHODS: Schizophrenia was diagnosed on the basis of the Diagnostic and Statistical Manual of Mental Disorders 4th ed (DSM-IV). Detailed clinical information, family history of all patients and healthy controls were collected. RFLP based case control association study was performed in a Pakistani cohort of 508 schizophrenia patients and 300 healthy control subjects. Alleles and genotype frequencies were calculated using SPSS. RESULTS: A significant difference in the genotype and allele frequencies for rs4765905, rs1076560 and rs6465084 were found between the patients and controls (p=0.000). CONCLUSION: This study provides substantial evidence supporting the role of CACNA1C, GRM3 and DRD2 as schizophrenia susceptibility genes in Pakistani population.
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Humans , Alleles , Case-Control Studies , Cohort Studies , Diagnostic and Statistical Manual of Mental Disorders , Gene Frequency , Genotype , Pakistan , Polymorphism, Restriction Fragment Length , SchizophreniaABSTRACT
Objective · To study the effect of miR-137 overexpressed trophoblast cells on the biological functions of vascular endothelial cells.Methods · Lentivirus vectors Up-LV-miR-137 and LV-miR-NC were constructed and transfected into the trophoblast cells HTR-8/SVneo, respectively.And transfection efficiency was verified by PCR. Vascular endothelial cells were cultured with transfected trophoblast cells or their culture medium supernatant. Proliferation, migration activity and ability to recruit monocytes U937 of endothelial cells were detected by CCK8 method, scratch test and Transwell migration test, respectively. Results · miR-137 overexpressed trophoblast cells were obtained. The results of CCK8 test showed that proliferation of vascular endothelial cells cultured with miR-137 overexpressed trophoblast cells or their culture medium supernatant was inhibited, especially in high glucose condition. The repair of endothelial cells in the transfected cells culture medium slowed down in the scratch test. Transwell migration test results showed that after the endothelial cells were cultured with miR-137 overexpressed trophoblast cells, more monocytes passed through the chambers with higher chemotactic index. Conclusion · miR-137 overexpressed trophoblast cells can regulate the biological functions of vascular endothelial cells, i.e. reduce proliferation and migration activity and enhance ability to recruit monocytes.
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Objective · To study the effect of miR-137 overexpressed trophoblast cells on the biological functions of vascular endothelial cells.Methods · Lentivirus vectors Up-LV-miR-137 and LV-miR-NC were constructed and transfected into the trophoblast cells HTR-8/SVneo, respectively.And transfection efficiency was verified by PCR. Vascular endothelial cells were cultured with transfected trophoblast cells or their culture medium supernatant. Proliferation, migration activity and ability to recruit monocytes U937 of endothelial cells were detected by CCK8 method, scratch test and Transwell migration test, respectively. Results · miR-137 overexpressed trophoblast cells were obtained. The results of CCK8 test showed that proliferation of vascular endothelial cells cultured with miR-137 overexpressed trophoblast cells or their culture medium supernatant was inhibited, especially in high glucose condition. The repair of endothelial cells in the transfected cells culture medium slowed down in the scratch test. Transwell migration test results showed that after the endothelial cells were cultured with miR-137 overexpressed trophoblast cells, more monocytes passed through the chambers with higher chemotactic index. Conclusion · miR-137 overexpressed trophoblast cells can regulate the biological functions of vascular endothelial cells, i.e. reduce proliferation and migration activity and enhance ability to recruit monocytes.
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Objective To investigate the clinical significance of testing serum kisspeptin in central precocious puberty (CPP) girls. Methods Sixty eight CPP girls and 68 healthy girls was studied from December 2012 to December 2014. HEK293 cells were cultured. Luciferase reporter assay was performed to verify the binding of miR-137 to the 3′UTR of KISS1. Serum miR-137 level was levaluated by qRT-PCR. Level of serum luteinizing hormone , prolactin , follicle stimulating hormone , thyrotropin , free thyroxine and estradiol was evaluated by chemi-luminescence immunoassay. The level of serum kisspeptin was detected by ELISA. Results MiR-137 was confirmed to bind to the 3′UTR of KISS1. The level of serum miR-137 was downregulated and kisspeptin was enhanced in CPP girls. The expression of miR-137 and kisspeptin was negatively correlated. Serum miR-137 level was negatively related to bone age and bone age advancement. According to the results of GnRH stimulating test, serum miR-137 was related to peak LH and peak/basal LH ratio. Conclusions MiR-137 could bind to the 3′UTR of KISS1. MiR-137 may be a potential biomarker for CPP assisted diagnosis.
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Objective To investigate the aassociation of a microRNA-137 (miR-137) polymorphism, single nucleotide polymorphism (SNP) rs1625579, with neurocognitive function in patients with schizophrenia. Methods A total of 250 patients with schizophrenia were included in this study. The positive and negative syndrome scale (PANSS) was used to evaluate patients. The brief assessment of cognition in schizophrenia (BACS) scale was used to determine neurocongnitive functions in patients. Blood samples of patients were collected, and SNaPshot technique was used to compare the neurocognitive functions of different genotypes of rs1625579. Results The genotypes of TT, GT and GG were 221 (88.4%), 28 (11.2%) and 1(0.4%). There was no significant difference in PANSS score between TT genotype carriers and G allele (GG+GT) carriers. The detection of BACS showed that the digit sequencing score was significantly lower in patients with TT genotype than that of G allele (GG+GT) carrier ( P<0.05). There were no significant differences in other scores of BACS evaluation between two groups of patients. Conclusion The miR-137 polymorphism influences the working memory performance of schizophrenic patients in Chinese Han population.
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Objective To explore the prognostic impact of miR137 target gene Kruppel-like transcription factor 12 (KLF12) in multiple myeloma (MM). Methods The target genes of miR137 were predicted by software. The GFP analysis of KLF12 and the prognosis of MM were constructed. Overexpressing miR137 in MM NCI-H929 cell line was also constructed. Real-time qPCR and Western blot were used to detect the expression of KLF12 in this cell line. Results The target genes of miR137 were MITF, BUE2H, SH3BP5 and KLF12. High expression of KLF12 in 455 patients included 75 patients (16.5 %) died, 104 patients with low expression of KLF12, and 25 patients (24.0 %) died, but no significance was detected in the different subgroups. KLF12 expression was higher in MM NCI-H929 cell line with miR137 over expression. The expression of miR137 was positively correlated with the expression of KLF12. Conclusion miR137-KLF12 is an important index to judge the prognosis of MM.