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dual luciferase reporter gene assay

Zun-Wen Lin, Wei Zhang, Si-Ding Jiang, Wen-Bo Wei, Xiao-Feng Li
Osteosarcoma (OS) is the most common malignant tumor of bone with a high potential for metastasis. This study intends to explore whether microRNA-940 (miR-940) affects the development of OS cells and the underlying mechanism. OS and adjacent normal tissues were collected from OS patients; the OS cell line with the highest expression of miR-940 was selected, which was then subjected to transfection of miR-940 mimic, miR-940 inhibitor, siRNA-secreted frizzled-related protein 1 (SFRP1) or LiCl (agonists of Wnt/β-catenin pathway) to identify regulation of miR-940 to OS cells through SFRP1...
October 15, 2018: Journal of Cellular Biochemistry
Rui Cai, Naren Qimuge, Meilin Ma, Yingqian Wang, Guorong Tang, Que Zhang, Yunmei Sun, Xiaochang Chen, Taiyong Yu, Wuzi Dong, Gongshe Yang, Weijun Pang
MicroRNAs (miRNAs) are non-coding RNAs that regulate gene expression at the post-transcriptional level and are involved in the regulation of the formation, maintenance, and function of skeletal muscle. Using miRNA sequencing and bioinformatics analysis, we previously found that the miRNA miR-664-5p is significantly differentially expressed in longissimus dorsi muscles of Rongchang pigs. However, the molecular mechanism by which miR-664-5p regulates myogenesis remains unclear. In this study, using flow cytometry, 5-ethynyl-2'-deoxyuridine (EdU) staining, and cell count and immunofluorescent assays, we found that cell-transfected miR-664-5p mimics greatly promoted proliferation of C2C12 mouse myoblasts by increasing the proportion of cells in the S- and G2-phases and up-regulating the expression of cell cycle genes...
October 15, 2018: Journal of Biological Chemistry
Yan-Xia Huang, Xin-Gang Nie, Guang-Da Li, Dong-Sheng Fan, Li-Li Song, Xin-Lin Zhang
Retinoblastoma (RB) is a well‑vascularized tumor dependent on angiogenesis. The present study aimed to explore whether microRNA (miR)‑182 regulates cell viability, invasion and angiogenesis in RB via the phosphatidylinositol‑3‑OH kinase (PI3K)/protein kinase B (AKT) signaling pathway and by targeting cell adhesion molecule 2 (CADM2). The expression levels of miR‑182 and CADM2 were initially detected in RB tissues from patients with RB who underwent ophthalmectomy, and normal retinal tissues collected from other trauma patients who underwent eye enucleation...
October 9, 2018: International Journal of Oncology
C Zhou, H Wu, Y Liu, C Yin, B Yang
Objective: To investigate the effects of long non-coding RNA HULC on downstream related targets regulating the migration and invasion of hepatoma cells and theirs mechanism of action. Methods: The expression of highly upregulated in liver cancer (HULC) in hepatocellular carcinoma, and adjacent normal liver tissues and different hepatocellular carcinoma cells were detected by qPCR. The correlation between clinicopathological data of HULC and liver cancer patients were analyzed. Dual-luciferase reporter gene detected the interaction between HULC and miR-186...
July 20, 2018: Zhonghua Gan Zang Bing za Zhi, Zhonghua Ganzangbing Zazhi, Chinese Journal of Hepatology
Feng-Na Yu, Ming-Lan Hu, Xin-Fang Wang, Xiang-Ping Li, Bei-Hao Zhang, Xiao-Qing Lu, Rui-Qiang Wang
As one major diabetic complication, diabetic nephropathy (DN) has been reported to be associated with various kinds of microRNA (miRNA). Thus, we conducted this study to explore the potential of miR-370 in a rat model of DN through investigation of mesangial cell proliferation and extracellular matrix (ECM). A total of 40 healthy adult male Sprague-Dawley rats were enrolled and assigned into normal (n = 10) and DN ( n = 30, DN rat model) groups. Dual-luciferase reporter assay was performed for the targeting relationship between miR-370 and canopy 1 (CNPY1)...
October 14, 2018: Journal of Cellular Physiology
Changlong Song, Dianqiu Li, Hongyu Liu, Hongyan Sun, Zhen Liu, Lirong Zhang, Yu Hu
Emerging evidence have discovered that circular RNAs (circRNAs) may serve as diagnostic or tumor promising biomarkers. This study aimed to investigate how circular RNA ADAMTS14 (circADAMTS14) regulates microRNA-572/ regulator of calcineurin 1(miR-572/ RCAN1) in hepatocellular carcinoma (HCC). The expression profiles of circRNA/microRNA (mRNA) between HCC tissues and paired adjacent tissues were analyzed via microarray analysis. The expressions of circADAMTS14, miR-572, and RCAN1 were measured by real-time polymerase chain reaction (PCR)...
October 14, 2018: Journal of Cellular Physiology
Zhi-Bin Liao, Xiao-Long Tan, Ke-Shuai Dong, Hong-Wei Zhang, Xiao-Ping Chen, Liang Chu, Bi-Xiang Zhang
BACKGROUND: Increasing evidence indicates that aberrant micro (mi)RNA-448 expression plays a critical role in the progression of several human cancers. However, the function of miRNA-448 in hepatocellular carcinoma (HCC) has not been fully investigated. METHODS: miRNA-448 expression levels in HCC tissues, adjacent non-cancerous tissues (ANTs), and HCC cell lines were examined by quantitative real-time polymerase chain reaction (qRT-PCR). HCC cells were treated with a miRNA-448 mimic or inhibitor, followed by cell viability measurements with the CCK-8 assay...
September 28, 2018: Digestive and Liver Disease
Linyu Zhang, Hao Dong, Youwen Si, Nan Wu, Hao Cao, Bing Mei, Bo Meng
MicroRNAs, small noncoding RNAs, not only regulate gene expression at the post-transcriptional level in a variety of physiological processes but also accompany the initiation and progression of a vast number of diseases, including dementia. While miR-125b has been shown to be aberrantly expressed in some dementia patients, its role in the pathological process remains ambiguous. Presenilin-1/2 conditional double knockout mice exhibit a range of symptoms, including impaired cognition and memory, increased tau phosphorylation, neuroinflammation, and apoptosis, and are therefore regarded as a useful dementia model...
September 18, 2018: Neurobiology of Aging
Weiqi Hu, Weimin Xu, Yi Shi, Weijun Dai
BACKGROUND: Nasopharyngeal carcinoma (NPC) is the most common type of head and neck cancers which is notable for its distinctive pattern of geographical distribution. HOTAIR has been reported to regulate nasopharyngeal carcinoma tumorigenesis and progression. However, the detailed mechanism underlying HOTAIR-promoted nasopharyngeal carcinoma remains not fully understood. METHODS: We used RT-qPCR approach to examine genes expression and mRNA level. MTT assay and soft agar assay were used to detect cell growth rate in culture and under suspended condition, respectively...
October 10, 2018: Biochemical and Biophysical Research Communications
Di Liu, Lingqin Song, Zhijun Dai, Haitao Guan, Huafeng Kang, Yinbin Zhang, Wanjun Yan, Xiaoyao Zhao, Shuqun Zhang
Perineural invasion (PNI) potentially increases the risk of relapse and abdominal pain in patients with pancreatic ductal adenocarcinoma (PDAC). However, the underlying mechanisms of PNI of PDAC is incompletely revealed. Our study aimed to investigate roles of miR-429 in modulating PNI in PDAC. We found that miR-429 was downregulated in PDAC cancer tissues and was profoundly decreased in tissues with PNI. It was reduced in nine of the ten examined pancreatic cancer cell lines. MiR-429 mimics restored its cellular expressions in MIA PaCa-2 and BxCP3 cells and significantly suppressed cell viability and invasion of the cancer cells...
October 9, 2018: Biochemical and Biophysical Research Communications
Xuelian Ma, Xiaomin Zhao, Zhichao Zhang, Jianxiong Guo, Lijuan Guan, Juejun Li, Mi Mi, Yong Huang, Dewen Tong
BACKGROUND: Transmissible gastroenteritis virus (TGEV) infection can activate NF-κB pathway in porcine intestinal epithelial cells and result in severe inflammation. Non-coding RNAs (ncRNAs) are not translated into proteins and play an important role in many biological and pathological processes such as inflammation, viral infection, and mitochondrial damage. However, whether ncRNAs participate in TGEV-induced inflammation in porcine intestinal epithelial cells is largely unknown. RESULTS: In this study, the next-generation sequencing (NGS) technology was used to analyze the profiles of mRNAs, miRNAs, and circRNAs in Mock- and TGEV-infected intestinal porcine epithelial cell-jejunum 2 (IPEC-J2) cell line...
October 12, 2018: BMC Genomics
Weiguo Wang, Ruisen Zhan, Jiahui Zhou, Jianlong Wang, Shijie Chen
The present study is aimed to investigate the molecular mechanism of miR-10 targeting NgR to regulate the proliferation and inflammatory cytokine secretion in microglia cells (MCs). The expression levels of NgR, IL-1β, and TNF-α in rat MCs were detected by QPCR and Immunoblotting assays. The proliferation of MC was detected by MTT and Soft agar clone formation assays. The target gene of miR-10 was verified by dual luciferase reporter gene assay. Overexpressed miR-10 downregulated the expression of NgR, promoted MC proliferation and inhibited inflammatory cytokine secretion...
October 9, 2018: Experimental and Molecular Pathology
A Martin-Pena, R M Porter, G Plumton, T M McCarrel, A J Morton, M V Guijarro, S C Ghivizzani, B Sharma, G D Palmer
Successful clinical translation of mesenchymal stem cell (MSC)-based therapies for cartilage repair will likely require the implementation of standardised protocols and broadly applicable tools to facilitate the comparisons among cell types and chondroinduction methods. The present study investigated the utility of recombinant lentiviral reporter vectors as reliable tools for comparing chondrogenic potential among primary cell populations and distinguishing cellular-level variations of chondrogenic activity in widely used three-dimensional (3D) culture systems...
October 12, 2018: European Cells & Materials
Longlue Jia, Danyan Zhang, Hannian Huang, Yongyong Zhou, Shengli Zhou, Jiangfeng Guo
Triazophos is a highly toxic organophosphorus pesticide, causing acute toxicity to brain tissue, and neurotoxicity and embryotoxicity to animals. Therefore, triazophos is considered as a public health problem due to its acute hazard index. MicroRNAs (miRNAs), a class of endogenous noncoding RNAs, can regulate the expression of target gene(s) by mediating mRNA cleavage or translational repression in organisms exposed to environmental chemicals. We found that nup43 is targeted by miR-217 , which was significantly regulated in adult zebrafish ( Danio rerio ) exposed to triazophos (phenyl-1,2,4-triazolyl-3-( o , o -diethyl thionophosphate))...
September 1, 2018: Toxicology Research
Minghe Li, Jun Ning, Zhihong Li, Jing Wang, Cong Zhao, Lei Wang
Background: LINC00152 plays a crucial role in tumorigenesis and progression of multiple types of cancer. However, the biological significance of LINC00152 and its potential role in oral squamous cell carcinoma (OSCC) remain to be determined. In the present study, we investigated the role of LINC00152 and the underlying mechanism of its oncogenic activity in OSCC. Materials and methods: The expression of LINC00152 in OSCC tissues and cell lines was detected using qRT-PCR...
2018: OncoTargets and Therapy
Wei Wang, Zhao Guo, Sidong Yang, Hui Wang, Wenyuan Ding
MicroRNA-199 has been reported to play a potential role in the apoptosis of Human nucleus pulposus cells. However, the effect of miR-199 in regulating Human nucleus pulposus cell injury induced by TNF-α has not been previously illustrated. This study searched to probe the effect and the molecular mechanism of miR-199 on Human nucleus pulposus cell injury induced by TNF-α. Using the TNF-α model of Human nucleus pulposus cell in vitro, we found that miR-199 was extremely decreased in Human nucleus pulposus cells after TNF-α treatment...
October 8, 2018: Biochemical and Biophysical Research Communications
Bo Guo, Jing Zhang, Qian Li, Zhenghao Zhao, Wenjing Wang, Kaiyue Zhou, Xiaofei Wang, Dongdong Tong, Lingyu Zhao, Juan Yang, Chen Huang
BACKGROUND/AIMS: MicroRNAs (miRNAs) have been well studied in human carcinogenesis and cancer progression. Our previous study showed the down-regulation of miR-338-3p expression in human gastric cancer (GC). However, the reasons of this dysregulation remain largely unclear. METHODS: Bisulfite sequence analysis was performed to explore the methylation status of the promoter region of miR-338-3p. Cell wound-healing and transwell assays were performed to examine the capacity of cell migration and cell interaction...
October 11, 2018: Cellular Physiology and Biochemistry
Meng Sun, Mengen Zhai, Nan Zhang, Rui Wang, Hongliang Liang, Qinghua Han, Yongping Jia, Liqin Jiao
Accumulating evidence has suggested that microRNAs (miRNAs) are emerging as critical regulators in myocardial ischemia/reperfusion injury. miR-148b-3p has been reported to regulate cell apoptosis of various cell types. However, whether miR-148b-3p is involved in regulating cardiomyocyte apoptosis in myocardial ischemia/reperfusion injury remains unknown. In this study, we aimed to investigate the potential role and molecular mechanism of miR-148b-3p in regulating cardiomyocyte apoptosis induced by hypoxia/reoxygenation (H/R) injury in vitro, a cellular model of myocardial ischemia/reperfusion injury...
October 8, 2018: Chemico-biological Interactions
Jin Ma, Jun Zhang, Yuan-Chi Weng, Jian-Cheng Wang
Pancreatic cancer (PC) is one of the most aggressive cancers presenting with high rates of invasion and metastasis, and unfavorable prognoses. The current study aims to investigate whether EZH2/miR-139-5p axis affects epithelial-mesenchymal transition (EMT) and lymph node metastasis (LNM) in PC, and the mechanism how EZH2 regulates miR-139-5p. Human PC and adjacent normal tissues were collected to determine expression of EZH2 and miR-139-5p, and their relationship with clinicopathological features of PC. Human PC cell line was selected, and treated with miR-139-5p mimics/inhibitors, EZH2 vector or shEZH2 in order to validate the regulation of EZH2-mediated miR-139-5p in PC cells ...
September 30, 2018: Molecules and Cells
Shuang Cao, Guohua Wang, Jia Wang, Cheng Li, Le Zhang
In this study, we sought to predict the effects of a certain circular RNA (circRNA), hsa_circ_101280 (also known as hsa_circ_0100929 and hsa_circ_SLAIN1), on hepatocellular carcinoma (HCC) cells and to determine the potential mechanism. After screening differentially expressed circRNAs in HCC tissues through GEO data analysis, hsa_circ_101280 was found to be highly expressed, and its high expression was verified in HCC cell lines with qRT-PCR along with the low expression of its downstream miRNA miR-375. Colony formation and flow cytometry assays showed that both hsa_circ_101280 silencing and miR-375 overexpression restrained proliferation and promoted apoptosis in HCC cells...
October 9, 2018: Immunology and Cell Biology
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