miR-138-5p protects β cell function by regulating HIF-1α in gestational diabetes mellitus / 中国临床药理学与治疗学

AIM: To study the effect of miR-138-5p on the function of β cell in gestational diabetes mellitus (GDM) and its related mechanism. METHODS: The expression of miR-138-5p in peripheral blood of 15 GDM pregnant women and 15 normal pregnant women were compared by RT-qPCR. miR-138-5p mimic and inhibitor were transfected into INS-1 cells, respectively, and their expression level was over expressed or inhibited. RT-qPCR was used to verify the transfection efficiency.MTT proliferation experiment, Annexin V-FITC apoptosis experiment and insulin release experiment were used to detect the effects of miR-138-5p on INS-1 cell proliferation, apoptosis and insulin release ability. The target gene of miR-138-5p was screened by TargetScan, a miRNA target gene prediction software. The functional rescue experiment confirmed whether miR-138-5p could exert its influence on INS-1 cell proliferation, apoptosis and insulin release ability by targeting its target gene. Western blot was used to detect the molecular signaling pathway of miR-138-5p in INS-1 cells.RESULTS: The expression of miR-138-5p in peripheral blood of GDM pregnant women was significantly lower than that of normal pregnant women. RT-qPCR showed that miR-138-5p mimic and inhibitor could significantly promote or inhibit the expression of miR-138-5p in INS-1 cells. The results of MTT proliferation experiment, Annexin V-FITC apoptosis experiment and insulin release experiment indicated that over expression of miR-138-5p could significantly promote the proliferation of INS-1 cells, inhibit the apoptosis of cells and promote the insulin release ability of cells. However, down-regulating the expression of miR-138-5p could significantly inhibit the proliferation of INS-1 cells, promote apoptosis and inhibit insulin release. HIF-1α was selected as the target gene of miR-138-5p by TargetScan. The double luciferase gene report and Western blot showed that miR-138-5p could inhibit the expression of HIF-1α in INS-1 cells. The functional rescue experiment confirmed that miR-138-5p could affect the proliferation, apoptosis and insulin release of INS-1 cells by regulating the expression of HIF-1α. Western blot showed that miR-138-5p may play a role in INS-1 cells by affecting PI3K, Akt and p-PI3K, p-Akt protein after phosphorylation in PI3K/Akt signaling pathway. CONCLUSION: miR-138-5p may reguLate HIF-1α expression in a targeted manner, thereby affecting the PI3K/AKT signaling pathway, promoting the proliferation and inhibition of the parent cells' apoptosis, and promoting their insulin-releasing ability to protect the function of β cell in GDM.

Knockdown of long non-coding RNA HOTAIR reverses cisplatin resistance of ovarian cancer cells through inhibiting miR-138-5p-regulated EZH2 and SIRT1

BACKGROUND: Cisplatin resistance (DDP-resistance) remains one of the major causes of poor prognosis in females with ovarian cancer. Long non-coding RNAs (lncRNAs) have been shown to participate in the regulation of cellular processes, including chemoresistance. The aim of this study was to explore the role of HOX transcript antisense RNA (HOTAIR) in DDP-resistant ovarian cancer cells. METHODS: DDP-resistant ovarian cancer cell lines (SKOV3/DDP and A2780/DDP) were established. Real-time PCR, western blot, dual-luciferase reporter assay, and flow cytometry were then used to evaluate the effect of HOTAIR/miR-138-5p axis on chemoresistance of DDP-resistant ovarian cancer cells to DDP. RESULTS: We found that HOTAIR was upregulated in DDP-resistant cells, while miR-138-5p was downregulated. Knockdown of HOTAIR increased the expression of miR-138-5p in DDP-resistant cells and miR-138-5p is directly bound to HOTAIR. Upregulation of miR-138-5p induced by HOTAIR siRNA or by its mimics enhanced the chemosensitivity of DDP-resistant cells and decreased the expression of EZH2 (enhancer of zeste 2 polycomb repressive complex 2 subunit) and SIRT1 (sirtuin 1). Furthermore, the HOTAIR silencing-induced chemosensitivity of DDP-resistant cells was weakened by miR-138-5p inhibitor. CONCLUSIONS: These data demonstrate that HOTAIR acts as a sponge of miR-138-5p to prevent its binding to EZH2 and SIRT1, thereby promoting DDP-resistance of ovarian cancer cells. Our work will shed light on the development of therapeutic strategies for ovarian cancer treatment.

MicroRNA-138 Suppresses Adipogenic Differentiation in Human Adipose Tissue-Derived Mesenchymal Stem Cells by Targeting Lipoprotein Lipase

PURPOSE: Adipogenic differentiation of adipose tissue-derived mesenchymal stem cells (AMSCs) is critical to many disease-related disorders, such as obesity and diabetes. Studies have demonstrated that miRNA-138 (miR-138) is closely involved in adipogenesis. However, the mechanisms affected by miR-138 remain unclear. This work aimed to investigate interactions between miR-138 and lipoprotein lipase (LPL), a key lipogenic enzyme, in AMSCs. MATERIALS AND METHODS: Human AMSCs (hAMSCs) isolated from human abdomen tissue were subjected to adipogenic differentiation medium. Quantitative real-time polymerase chain reaction and Western blot assay were applied to measure the expressions of miR-138, LPL, and the two adipogenic transcription factors cytidine-cytidine-adenosine-adenosine-thymidine enhancer binding protein alpha (C/EBPα) and peroxisome proliferator-activated receptor gamma (PPARγ). The relationship between miR-138 and LPL was predicted utilizing the miRTarBase database and validated by dual luciferase reporter assay. RESULTS: Showing increases in C/EBPα and PPARγ expression levels, hAMSCs were induced into adipogenic differentiation. During adipogenesis of hAMSCs, miR-138 expression was significantly downregulated. Overexpression of miR-138 by transfection inhibited hAMSCs adipogenic differentiation in vitro. Mechanically, LPL was a target of miR-138. LPL expression was upregulated during adipogenesis of hAMSCs, and this upregulation was reversed by miR-138 overexpression. Functionally, silencing of LPL by transfection exerted similar inhibition of the expressions of C/EBPα and PPARγ. Meanwhile, LPL ectopic expression was able to partly abolish the suppressive effect of miR-138 overexpression on adipogenic differentiation of hAMSCs. CONCLUSION: Upregulation of miR-138 inhibits adipogenic differentiation of hAMSCs by directly downregulating LPL.

Emodin protects H9c2 cells from hypoxia-induced injury by up-regulating miR-138 expression

Myocardial infarction (MI) is a common presentation for ischemic heart disease, which is a leading cause of death. Emodin is a Chinese herbal anthraquinone used in several diseases. However, the effect of emodin in hypoxia-induced injury in cardiomyocytes has not been clearly elucidated. Our study aimed to clarify the functions of emodin in hypoxia-induced injury in rat cardiomyocytes H9c2 and explore the underlying mechanism. The effects of emodin on cell viability and apoptosis were analyzed by the Cell counting kit-8 assay and flow cytometry assay, respectively. The cell proliferation- and cell apoptosis-related proteins were detected by western blot. qRT-PCR was used to determine the relative expression of miR-138. Cell transfection was performed to alter miR-138 and MLK3 expression. miR-138 target was performed by dual luciferase activity assay. Sirt1/AKT and Wnt/β-catenin pathways-related factors phosphorylation were analyzed by western blot. Emodin inhibited hypoxia-induced injury in H9c2 cells by promoting cell viability and reducing cell apoptosis. miR-138 was down-regulated by hypoxia treatment but up-regulated by emodin. Up-regulation of miR-138 alleviated hypoxia-induced cell injury. Down-regulation of miR-138 attenuated the growth-promoting effect of emodin on hypoxia-induced injury, whereas up-regulation of miR-138 enhanced the growth-promoting effects of emodin. The underlying mechanism might be by inactivating Sirt1/AKT and Wnt/β-catenin pathways. MLK3 was negatively regulated by miR-138 expression and inactivated Sirt1/AKT and Wnt/β-catenin pathways. Emodin alleviated hypoxia-induced injury in H9c2 cells via up-regulation of miR-138 modulated by MLK3, as well as by activating Sirt1/AKT and Wnt/β-catenin pathways.

Expression and significance of miR-138 and programmed cell death protein 1 in the patients with HBV-related hepatocellular carcinoma / 临床检验杂志

Objective@#Abstract: Objective: To investigate the expression and significance of miR-138 and programmed cell death protein 1 (PD-1) in the patients with hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC). @*Methods@#A total of 30 patients with HBV-related HCC, 20 with HBV-related cirrhosis (LC) and 30 with chronic hepatitis B (CHB) were recruited from Jiaozuo People′s Hospital. The blood samples from all patients and the peritoneal effusion samples from HCC and LC patients were collected. The levels of miR-138 and soluble PD-1 (sPD-1) in blood and peritoneal effusion samples were detected by real-time PCR and ELISA, respectively. The expressions of PD-1 in T lymphocytes were measured with flow cytometry and western blot. The targeting effect of miR-138 on the 3′-non-coding region (3′-UTR) of PD-1 gene was verified by the dual-luciferase reporter gene system. @*Results@#The relative expression levels of miR-138 in the peritoneal effusion and plasma of HBV-related HCC patients were significantly lower than those in LC and CHB patients (P＜0.05). The serum sPD-1 levels and the expression levels of PD-1 in CD3 + T lymphocytes of HBV-related HCC patients were significantly higher than those in LC and CHB patients (P＜0.05). The relative expression levels of miR-138 were negatively correlated with serum sPD-1 levels and the expression levels of PD-1 in CD3 + T lymphocytes (P＜0.05). The dual-luciferase reporter gene system and western blot results demonstrated that there was a targeting relationship between miR-138 and the 3′-UTR of PD-1 gene. After miR-138 was transfected, the expression level of PD-1 was significantly down-regulated. @*Conclusion@#miR-138 participates in the development and progression of HBV-related HCC probably by targeting PD-1.

Expression of miR-138 and its effects on human lens epithelial cells apoptosis in age-related cataract patients / 眼科新进展

Objective To detect the expression of miR-138 in lens tissues of agerelated cataract and explore the effects of miR-138 on the proliferation and apoptosis of human lens epithelial cells and its possible target genes.Methods Real-time quantitative PCR (RT-qPCR) was applied for the detection of the expression of miR-138 and prediction of target gene sirtuin (silent information regulator 1) (SIRT1) in patients with age-related cataract (cataract group) and anterior lens capsules (normal control group).Then miR-138 mimics,mimic controls,miR-138 inhibitors and inhibitor controls were transfected into the human lens epithelial cell line (SRA01/04),and the expression of SIRT1 mRNA and protein was detected by RT-qPCR and Western blot,accordingly.At 72 hours after transfection,the cells were exposed to 200 μmol · L-1 H2O2 for 1 hour,followed by detection of the activity of Caspase-3 by the Caspase-3 activity assay kit,and identification of the targeted relationship between miR-138 and SIRT1 by dual luciferase reporter assays.Results Compared with the normal control group,the expression of miR-138(3.64 ±0.19) was significantly increased (P ＜0.001),but the expression of SIRT1 mRNA(0.32 ± 0.06) was significantly decreased (P ＜ 0.001) in the cataract group.Moreover,The expression levels of SIRT1 mRNA(0.42 ± 0.05) and protein(0.46 ± 0.05) in cells transfected with miR-138 mimics were significantly decreased,while the activity of Caspase-3 (3.24 ± 0.17) was significantly elevated when compared with cells transfected with minic controls (all P ＜ 0.05);Compared with cells transfected with inhibitor controls,the expressions of SIRT1 mRNA(2.95 ±0.13) and protein(1.98 ±0.12) were significantly upregulated,whereas Caspase-3 activity(0.42 ±0.05) was significantly decreased in cells transfected with miR-138 inhibitors (all P ＜0.05).And fmally,dual luciferase reporter assays showed the confirmation SIRT1 as a direct target of miR-138.Conclusion miR-138 is highly expressed in the lens capsule of age-related cataract patients,and it can promote the apoptosis of lens epithelial cells by negatively regulating the expression of SIRT1.

Role of miR-138 in the Renal Ischemic Reperfusion Injury and Its Mechanism / 医学研究杂志

Objective To determine the effect of miR-138 on SD rat renal ischemia-reperfusion injury its mechanism.Methods 60 adult SD rats were randomly divided into Sham group,renal ischemia-reperfusion (RIRI) group,RIRI + vector group,RIRI + miR-138 group,RIRI + miR-138 shRNA group,and then we established renal reperfusion injury model by occlusion bilateral renal pedicle.Creatinine,blood urea nitrogen in serum are tested to compare differences in renal function in each group,and TUNEL staining was taked to detect apoptosis,real time PCR were taked to test the exression of p53 and it's pathway.Results When compared with the sham group,expression of kidney function BUN/Crin serum,percentage of apoptoiscells of reperfusion injury group wrer all increased,and the differences were statistically significant (P ＜ 0.05).Upregulate miR-138 increased these indexes,and the difference was statistically significant (P ＜0.05),and down regulate miR-138 would protect the injure of renal ischemia-reperfusion.Further studies showed that miR-138 may play the role by inhibiting the p53 signaling pathway.Conclusion MiR-138 has a significant aggravating effect on renal ischemia-reperfusion injury,and the mechanism may be related to p53 signaling pathway.

Research progress in miR-138 in tumors / 肿瘤

microRNA (miRNA, miR) has attracted great interest in recent years. miR-138 is closely related to the occurrence, development and prognosis of different kinds of malignant tumors.Present studies have shown that miR-138 is frequently down-regulated in tumor cells and tissues. Down-regulation expression of miR-138 can promote the proliferation, invasion and metastasis in a variety of malignant tumor cells. Up-regulation expression of miR-138 may play a crucial role in tumor suppression, and enhance sensitivity of tumor cells to radiotherapy and chemotherapy. Therefore, miR-138 can be considered as a biomarker of tumor prognosis, and also one of the tumor therapeutic targets.

miR-138 Function and Its Targets on Mouse Mammary Epithelial Cells / 生物化学与生物物理进展

miRNA was 20～25 nt endogenous non-coding RNA. miRNAs are encoded small RNAs that hybridize with messenger RNAs, resulting in degradation or translational inhibition of targeted transcripts. In order to investigate the function of miR-138 on mouse mammary epithelial cells, technique for gene silencing-miRNA inhibitor (anti-miRNA) was applied to make miR-138 silence, qRT-PCR was showed valid for inhibitor miR-138. And Western blot, CASY(r)-technology was put in use to study some change of mouse mammary epithelial cells after miR138 inhibitor. It was shown that miR-138 suppresses the exepress of PRL-R(P