miR-429-3p mediates memory decline by targeting MKP-1 to reduce surface GluA1-containing AMPA receptors in a mouse model of Alzheimer's disease

Alzheimer's disease (AD) is a leading cause of dementia in the elderly. Mitogen-activated protein kinase phosphatase 1 (MKP-1) plays a neuroprotective role in AD. However, the molecular mechanisms underlying the effects of MKP-1 on AD have not been extensively studied. MicroRNAs (miRNAs) regulate gene expression at the post-transcriptional level, thereby repressing mRNA translation. Here, we reported that the microRNA-429-3p (miR-429-3p) was significantly increased in the brain of APP23/PS45 AD model mice and N2AAPP AD model cells. We further found that miR-429-3p could downregulate MKP-1 expression by directly binding to its 3'-untranslated region (3' UTR). Inhibition of miR-429-3p by its antagomir (A-miR-429) restored the expression of MKP-1 to a control level and consequently reduced the amyloidogenic processing of APP and Aβ accumulation. More importantly, intranasal administration of A-miR-429 successfully ameliorated the deficits of hippocampal CA1 long-term potentiation and spatial learning and memory in AD model mice by suppressing extracellular signal-regulated kinase (ERK1/2)-mediated GluA1 hyperphosphorylation at Ser831 site, thereby increasing the surface expression of GluA1-containing α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs). Together, these results demonstrate that inhibiting miR-429-3p to upregulate MKP-1 effectively improves cognitive and synaptic functions in AD model mice, suggesting that miR-429/MKP-1 pathway may be a novel therapeutic target for AD treatment.

Protective effects of Baicalin injection on severe acute pancreatitis through regulating follistatin-like-1 signaling pathway by down-regulating miR-429 expression in mice

Abstract In China, Scutellaria is used for treating inflammatory-related diseases. Baicalin is the main active component of Scutellaria and has protective effects on acute pancreatitis. However, the mechanism of Baicalin is still unclear. In this study, the protective effects of baicalin on acute pancreatitis induced by taurocholate and its mechanism are investigated. In this study, mice were randomly divided into three groups: sham operation, model, and treatment groups. Acute pancreatitis in mice was induced by intraperitoneal injection of taurocholate (35 mg/kg). The treatment group was given baicalin (100 mg/kg) 2 h before acute pancreatitis induction. The mRNA expression levels of miR-429, nuclear factor kappa B65(NF-kB65), toll-like receptor 4(TLR4), TNF receptor associated factor6 (TRAF6), NF-kappa-B inhibitor(IkB), Follistatin-like 1 (FSTL1), and interleukin-1 receptor-associated kinase (IRAK) in the liver tissues 24 h after intraperitoneal injection were detected by RT-PCR. Then, the expression levels of NF-kB65, p-NF-κB65, TLR4, TRAF6, IkB, FSTL1, IRAK, p- IRAK, and p- IkB-а proteins were detected by Western blot. IL-6, TNF-α and IL-1 ß in plasma were measured by ELISA, and histopathological changes in the pancreases of the mice were observed. The results showed that after baicalin treatment, miR-429 expression in the pancreatic tissues and the expression levels of NF-kB65, TLR4, TRAF6, p-IkB-а, FSTL1, and p-IRAK decreased. Similarly, pancreatic myeloperoxidase (MPO) activity and the plasma levels of IL-6, TNF-а, IL-12, IL-1ß1, endotoxin, serum amylase, and lipase were reduced. Thus, the pancreatic injury induced by taurocholate was alleviated. The present study indicates that pretreatment with Baicalin can alleviate acute pancreatic injury induced by taurocholate in mice. The mechanism may be associated with the decreased miR-429 expression, reduced FSTL1 signaling pathway activity, TLR4 and TLR4/MyD88 signaling pathway inhibition, and reduced pancreatic inflammation. FSTL1 is the regulatory target for miR-429

Advances in the role of miR-429 in multiple drug resistance of ovarian cancer / 中国肿瘤生物治疗杂志

@#目前miRNA在癌症发生、发展中的作用得到了广泛的研究，并且在诊断、预后中已经被证明是有效生物标志物。 miR-429在肿瘤中差异表达，并通过与多个靶基因相互介导，与卵巢癌细胞的多药耐药密切相关。miR-429通过抑制卵巢癌细胞 发生上皮间质转化（epithelial-mesenchymal transition，EMT）抑制肿瘤细胞的耐药。自噬有助于提升肿瘤细胞的抗压、修复能力， 并可促进肿瘤细胞的迁移、生长，miR-429可能抑制耐药细胞的自噬从而减少卵巢癌细胞耐药的发生。本文主要对近年来关于 miR-429参与卵巢癌多药耐药机制的研究做一综述，希望能对卵巢癌多药耐药的诊断、治疗和预后判断有所帮助，为其进一步在 临床应用提供指导。

Detection of expression levels of miR200 family in plasma of breast cancer patients and its significance / 吉林大学学报(医学版)

Objective: To detect the expression levels of microRNA200 (miR200) family in the plasma of the breast cancer patients and the normal controls, and to evaluate their potential values in the screening, progression and prognosis evaluation of breast cancer. Methods: A total of 82 cases of plasma samples of the patients with breast cancer (breast cancer group) and 30 cases of healthy plasma samples (control group) were collected. The microRNAs (miRNAs) were extracted from the plasma samples, the expression levels of miR200 family (miR200a, miR200b, miR200c, miR141 and miR429) were quantified by using real-time PCR technique. The receiver-operating characteristic (ROC) curve was used to assess the diagnostic value of miRNAs, and the associations between the levels of miRNAs and the clinicopathological characteristics were analyzed. Cox proportional hazard mode was used for survival analysis. Results: Compared with control group, the expression level of miR141 in the plasma of the patients in breast cancer group was decreased (P 0 . 05). Concuson: MiR200b and miR141 are likely to become the molecular biological parameters for the diagnosis of breast cancer.

MicroRNA-429 promotes chemotherapeutic sensitivity of esophageal squamous cell carcinoma cells by inhibiting BMI-1 expression / 肿瘤

Objective: To study the expression of microRNA-429 (miR-429) in esophageal squamous cell carcinoma (ESCC) tissues and its relationship with the clinical characteristics and prognosis of patients, as well as to explore the regulatory effect of miR-429 on the chemotherapeutic sensitivity of ESCC cells and the underlying mechanism. Methods: The samples of tumor tissues and paracancerous tissues were collected from 60 cases of ESCC patients from January 2010 to December 2010. The expression level of miR-429 in tumor tissues and paracancerous tissues was detected by real-time fluorescent quantitative PCR, as well as its relationship with the clinical feathers of ESCC patients was analyzed. After the miR-429 mimic or miR-429 inhibitor was transfetcted into Eca109 and TE-13 cells, the proliferation and chemotherapy sensitivity of ESCC cells were measured by MTT assay, the apoptosis of ESCC cells treated with cisplatin (CDDP) was detected by FCM. A dual-luciferase reporter assay was performed to detect whether B-cell specific moloney murine leukemia virus insertion site-1 (Bmi-1) was a target gene of miR-429. The expressions of Bmi-1 and P-glycoprotein (P-gp) proteins in ESCC cells were analyzed by Western blotting. Results: The expression level of miR-429 in ESCC tissues was significantly higher than that in the paracancerous tissues (P < 0.01). For the ESCC tissues with miR-429 low expression, the size of tumor was bigger, the clinical stage of tumor was later, the lymph node metastasis was more likely to occure, and the survival time of ESCC patients was shorter as compared with the miR-429 higher expression group. After overexpressing miR-429, the proliferation viability of Eca109 and TE-13 cells were suppressed obviously, the sensitivity to CDDP was enhanced, the apoptosis rate was increased, while the expression levels of Bmi-1 and P-gp were significantly down-regulated (all P < 0.05). After blocking miR-429 expression, the proliferation viability of Eca109 and TE-13 cells were promoted obviously, the sensitivity to CDDP was weakened, the apoptosis rate was decreased, while the expression levels of Bmi-1 and P-gp were significantly up-regulated (all P < 0.05). In addition, the result of dual-luciferase reporter assay further confirmed that Bmi-1 gene was a target of miR-429, and the expressions of Bmi-1 and miR-429 were negative correlated (r ＝-0.340 4, P < 0.05). Conclusion: The ESCC patients with lower miR-429 expression in tumor tissues frequently have a worse clinical outcome and poorer prognosis. Furthermore, miR-429 can improve the sensitivity of ESCC cells to CDDP, and its mechanism may be associated with the regulation of target gene Bmi-1 expression.

miR-429 promotes capecitabine-resistance in pancreatic cancer PANC-1 cells by down-regulating PTEN and activating PI3K/AKT signaling pathway / 中国肿瘤生物治疗杂志

@# Objective: To explore the mechanism of miR-429 targeting PTEN to affect capecitabine-resistance in pancreatic cancer PANC-1 cells though the PI3K/AKT signaling pathway. Methods: Capecitabine-resistant pancreatic cancer cell line PANC-1/CAP was constructed, and the expression of miR-429 and PTEN were detected by quantitative Real-time polymerase chain reaction (qRT-PCR) and Western blotting. The effect of miR-429 knock-down on cell proliferation viability, apoptosis and capecitabine-resistance was measured by colony formation assay, CCK-8 assay andAnnexin V-FITC/PI double staining flow cytometry assay, respectively. Subsequently, dual luciferase reporter assay verified that PTEN was a target gene of miR-429. Furthermore, the effect of miR-429 on PTEN-PI3K/ AKT signaling pathway was measured by Western blotting. Results: miR-429 was found to be up-regulated in PANC-1 cells and PANC-1/CAP cells compared with the non-malignant pancreatic ductal cell line (HPDE6-C7) (P<0.05 or P<0.01). Moreover, silencing of miR-429 significantly decreased cell proliferation viability, capecitabine-resistance and enhanced apoptosis of PANC-1/CAP cells; additionally, dual luciferase reporter assay confirmed that PTEN was a target of miR-429 (P<0.05 or P<0.01). Suppression of miR-429 up-regulated PTEN and blocked the PI3K/AKT signaling pathway to decrease cell proliferation viability and further reduce the capecitabine-resistance of PANC-1/CAP cells (P<0.05 or P<0.01). Conclusion: miR-429/PTEN-PI3K/AKT signaling pathway plays a certain role in regulating the capecitabine-resistance of pancreatic cancer, and inhibition of miR-429 expression may reverse the resistance of PANC-1/CAPto capecitabine.

Clinical Study on Modified Huanglian Wendan Decoction for Treatment of Tic Disorder Children with Syndrome of Wind Stirring Due to Phlegm-heat / 中国中医药信息杂志

Objective To observe the clinical efficacy of modified Huanglian Wendan Decoction for treatment of tic disorder (TD) children with syndrome of wind stirring due to phlegm-heat; To detect its effects on peripheral-blood miR-429 expression. Methods Eighty cases of TD children were randomly divided into treatment group and control group,with 40 cases in each group.In the treatment group,children were treated with modified Huanglian Wendan Decoction, one dosage a day, twice a day, orally. In the control group, TD children were treated with haloperidol, 0.03 mg/(kg?d), orally, according to the symptom to adjust dose, and the maximum amount did not exceed 0.08 mg/(kg?d). 30 d was a treatment course for both groups. The treatment lasted for three courses. Before and after 3 months of clinical observation, the severity of disease was evaluated by TCM syndrome and Yale Global Tic Severity Scale (YGTSS) score. Clinical efficacy and TCM syndrome efficacy were evaluated; the level of miR-429 in peripheral blood was detected; the adverse reactions were observed. Results The total effective rate of clinical efficacy was 82.5% (33/40) in treatment group and 80.0% (32/40) in the control group, without statistical significance (P>0.05); The total effective rate of TCM syndrome efficacy was 92.5% (37/40) in treatment group and 75.0% (30/40) in the control group, with statistical significance (P<0.05). Compared with before treatment, the YGTSS score and TCM syndrome score significantly decreased after treatment (P<0.01), and the TCM syndrome score in the treatment group was much lower than the control group after treatment (P<0.05). Compared with before treatment, miR-429 level in both groups increased obviously after treatment (P<0.01), and the miR-429 level in the treatment group was much higher than the control group after treatment (P<0.05). There were mild constipation (1 cases), diarrhea (2 cases), and anorexia (2 cases) in the treatment group, and obvious lethargy (3 cases) and dizziness (3 cases) in the control group. Conclusion Modified Huanglian Wendan Decoction shows obvious effects for treating TD, and can up-regulate miR-429 of peripheral-blood.

miR-429 Negatively Regulates Osteosarcoma Stem Cell-like Properties by Targeting Sox2 / 中国医科大学学报

Objective To investigate the role of miR-429 in osteosarcoma stem cells.Methods We employed different approaches to test the expression of miR-429 in osteosarcoma stem cells.miR-429-high and miR-429-low cells were separated using molecular probes and their stem cell-like properties were compared.The effect of miR-429 on osteosarcoma stem cell-like properties was further tested through transfection assays.Furthermore,the miR-429 downstream target gene was confirmed by luciferase assay.Results The expression of miR-429 in osteosarcoma stem cells was much lower than that in control cells.miR-429-high cells displayed fewer stem celllike properties than did miR-429-low cells.These observations were confirmed by transfection assays.Additionally,our luciferase assays showed that miR-429 regulates Sox2 at the post-transcriptional level.Conclusion miR-429 negatively regulates osteosarcoma stem celllike properties by targeting Sox2.

MiR-429 regulates inflammatory infiltration, proliferative ability and differentiation of hepatocellular carcinoma / 第二军医大学学报

Objective To observe the effects of miR-429 on inflammatory infiltration, differentiation and proliferation of hepatocelluar carcinoma (HCC). Methods HCCLM3 cells were sorted into positive and -negative cells using magnetic activated cell sorting (MACS) according to EpCAM marker. The EpCAM-positive and negative cells were transfected with AntagomiR- 429 and miR-429mimic to construct over-expressed and down-regulated cells of miR-429, respectively. To validate the effect of transfection, control cells and constructed cells were tested by flow cytometry (FC) and real-time polymerase chain reaction (PCR). The proliferative curves of transfected cells were plotted using cell counting kit-8 (CCK8) assay. Furthermore, tumor- bearing mice model was established by subcutaneous inoculation with transfected cells and related control cells into non-obese diabetic, severe combined immunodeficient mice (NOD-SCID). Mice were sacrificed after 6 weeks, and protein expressions of Ki67, AFP and F4/80 in tumors were determined via immunohistochemistry (IHC) to examine the status of proliferation, differentiation and inflammatory infiltration, respectively. Results The proportion of EpCAM-positive cells was decreased by down-regulating miR-429, and EpCAM-negative cells could partially transfer to EpCAM-positive cells by miR-429 ove- expression. In vivo study showed that over-expressing miR-429 in EpC AM-negative cells cound enhance the tumor forming ability; whereas EpCAM-positive cells had an inhibited tumor-formation when down-regulating miR-429. Interestingly, F4/80, a marker of macrophage, was strongly stained in tumor tissue with miR-429 over-expression. It was also found that in tumor tissue, Ki67 and AFP staining was suppressed with miR-429 down-regulation. Meanwhile, compared to the control, miR-429 significantly accelerated the proliferation of HCCLM3 in vitro compared with the control (P$0. 05). Conclusion miR-429 can regulate the proportion of EpCAM-positive cells of HCC in vitro and in vivo, and greatly increase the inflammatory infiltration and proliferative ability, accompanied by decreased differentiation.