scholarly journals The lncRNA-MALAT1/miR-126-5p Axis Promotes Growth and Metastasis of Gastric Cancer through Regulation of VEGFA

2020 ◽  
Author(s):  
Zhi-Li Hu ◽  
Yang-zhi Hu ◽  
Qing Li ◽  
Tian-you Liao ◽  
Hai-ping Jiang
Keyword(s):  
Gastric Cancer ◽  
Signaling Pathway ◽  
Luciferase Reporter ◽  
Mtor Signaling Pathway ◽  
Luciferase Reporter Gene ◽  
Ags Cells ◽  
Luciferase Reporter Gene Assay ◽  
Lncrna Malat1 ◽  
Gene Assay ◽  
Cancer Tissues

Abstract Background: It has been reported that reduction of miR-126 can promote the progression of gastric cancer (GC). However, the regulation of miR-126 in GC is still unclear. This study aims to explore the correlation between lncRNA MALAT1 and miR-126 in gastric cancer and disclose the underlying mechanisms.Methods: We analyzed the correlation of MALAT1 levels and clinical features by analysis of bioinformatic data and human samples. Then we down-regulate the expression of MALAT1 in AGS cells and examined the characteristics of cell proliferation, cycle, apoptosis, migration, invasion, and the effect on miR-126 as well as VEGFA and signaling pathway. In addition, we demonstrated the role of MALAT1/miR-126 axis in GC with dual-luciferase reporter gene assay and treatment of miR-126 inhibitor.Results: The expression of MALAT1 was higher in cancer tissues than para-cancer tissues. In addition, high MALAT1 level suggested greater malignancy and poorer prognosis. Down-regulating the expression of MALAT1 in AGS cells inhibited cell proliferation, migration, and invasion by targeting VEGFA, which is consistent with up-regulation of miR-126. According to dual-luciferase reporter gene assay and treatment of miR-126 inhibitor, we demonstrated that MALAT1 down-regulated miR-126 in GC, which leads to the up-regulation of VEGFA and activation of mTOR signaling pathway.Conclusions: MALAT1/miR-126 axis promotes growth and metastasis of gastric cancer through regulation of VEGFA via mTOR signaling pathway.Fund This article is supported by Science and Technology Funding Project of Hunan Province, China (No.2017SK4010)

scholarly journals PITPNA-AS1 Activated by H3K27ac Sponged miR-98-5p to Regulate Cisplatin Resistance in Gastric Cancer

2021 ◽  
Author(s):  
Yaping Liu ◽  
Xu Zhao ◽  
Yinnan Chen ◽  
Gang Guo ◽  
Jiansheng Wang ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Cell Proliferation ◽  
Molecular Mechanisms ◽  
Luciferase Reporter ◽  
Platinum Resistance ◽  
Luciferase Reporter Gene ◽  
Gene Assay ◽  
Proliferation And Apoptosis ◽  
Gastric Cancer Cell Lines ◽  
Cancer Tissues

Abstract To evaluate the expression of PITPNA-AS1 and miR-98-5p in gastric cancer tissues as well as their association with progression of gastric cancer, and investigate the role of PITPNA-AS1 and miR-98-5p in developing platinum resistance. RNA sequencing was used to identify candidate lncRNAs and microRNAs related to local recurrence of gastric cancer. qRT-PCR was used to investigate the expression of PITPNA-AS1 and miR-98-5p. CCK-8 and caspase3/7 activity were used to evaluate the cell proliferation and apoptosis rate. Dual luciferase reporter gene assay and RNA pull down were used to evaluate the cross talk between PITPNA-AS1 and miR-98-5p. PITPNA-AS1 and miR-98-5p could regulate cell proliferation and inhibit apoptosis in gastric cancer cell lines. Cisplatin and lobaplatin could significantly suppress the expression of PITPNA-AS1, which interacted with negatively regulated miR-98-5p expression. PITPNA-AS1 overexpression impaired the effect of platinum, which was partially reversed by downregulation of miR-98-5p knock down. In gastric cancer, PITPNA-AS1 and miR-98-5p could regulat cell growth, apoptosis and platinum resistance. They have the potential to be biomarkers and curative therapeutic targets. However, further research on molecular mechanisms are needed.

scholarly journals YY1 Activates EMI2 and Promotes the Progression of Cholangiocarcinoma Through the PI3K/Akt Signaling Axis

2021 ◽  
Author(s):  
Shuai zhou ◽  
Kang Lin Qu ◽  
JinAng Li ◽  
Shilei Chen ◽  
Yi Gang Zhang ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Bile Duct ◽  
Signaling Pathway ◽  
Akt Signaling ◽  
Reporter Gene Assay ◽  
Luciferase Reporter ◽  
Akt Signaling Pathway ◽  
Luciferase Reporter Gene ◽  
Luciferase Reporter Gene Assay ◽  
Gene Assay

Abstract Background: Cholangiocarcinoma (CCA) is one of the deadliest cancers of the digestive tract. The prognosis of CCA is poor and the 5-year survival rate is low. Bioinformatic analysis showed that early mitotic inhibitor 2 (EMI2) was overexpressed in CCA but the underlying mechanism is not known.Methods: The data on bile duct carcinoma from TCGA and GEO databases were used to detect the expression of EMI2. The transcription factors of EMI2 were predicted using JASPAR and PROMO databases. Among the predicted transcription factors, YY1 has been rarely reported in cholangiocarcinoma, and was verified using the luciferase reporter gene assay. RT-PCR was performed to predict the downstream pathway of EMI2, and PI3K/Akt was suspected to be associated with it. Subsequently, in vivo and in vitro experiments were conducted to verify the effects of silencing and overexpressing EMI2 and YY1 on the proliferation, invasion, and metastasis of the bile duct cancer cells.Results: EMI2 was highly expressed in CCA. Silencing EMI2 inhibited the proliferation, invasion, and migration of CCA cells, arrested cell cycle in the G1 phase, and inhibition of apoptosis. The luciferase reporter gene assay showed that YY1 bound to the promoter region of EMI2, and after silencing YY1, the expression of EMI2 decreased and the progression of CCA was inhibited. Moreover, key proteins in the PI3K/Akt signaling pathway decreased after silencing EMI2.Conclusion: EMI2 may be one of the direct targets of YY1 and promotes the progression of CCA through the PI3K/Akt signaling pathway.

scholarly journals The potency of lncRNA MALAT1/miR-155/CTLA4 axis in altering Th1/Th2 balance of asthma

2020 ◽  
Vol 40 (2) ◽  
Author(s):  
Zhijun Liang ◽  
Fenglian Tang
Keyword(s):  
T Cells ◽  
Cd4 T Cells ◽  
Vital Capacity ◽  
Luciferase Reporter ◽  
Luciferase Reporter Gene ◽  
Luciferase Reporter Gene Assay ◽  
Lncrna Malat1 ◽  
Gene Assay ◽  
Asthma Patients ◽  
Ifn Γ

Abstract Objectives: The present study examined if the metastasis-associated lung adenocarcinoma transcript 1 (MALAT1)/miR-155/CTLA-4 axis was involved in modifying Th1/Th2 balance, a critical indicator for asthma progression. Methods: Altogether 772 asthma patients and 441 healthy controls were recruited, and their blood samples were collected to determine expressional levels of MALAT1, miR-155, CTLA-4, T-bet, GATA3, Th1-type cytokines and Th2-type cytokines. The CD4+ T cells were administered with pcDNA3.1-MALAT1, si-MALAT1, miR-155 mimic and miR-155 inhibitor to assess their effects on cytokine release. The luciferase reporter gene assay was also adopted to evaluate the sponging relationships between MALAT1 and miR-155, as well as between miR-155 and CTLA-4. Results: Over-expressed MALAT1 and under-expressed miR-155 were more frequently detected among asthma patients who showed traits of reduced forced expiratory failure volume in 1 s (FEV1), FEV1/forced vital capacity (FVC) and FEV1% of predicted (P<0.05). Moreover, MALAT1 expression was negatively expressed with the Th1/Th2 and T-bet/GATA3 ratios, yet miR-155 expression displayed a positively correlation with the ratios (P<0.05). Additionally, the IFN-γ, IL-2 and T-bet levels were reduced under the influence of pcDNA3.1-MALAT1 and miR-155 inhibitor, while levels of IL-4, IL-10 and GATA3 were raised under identical settings (P<0.05). Furthermore, MALAT1 constrained expression of miR-155 within CD4+ T cells by sponging it, and CTLA-4 could interfere with the effects of MALAT1 and miR-155 on Th1/Th2 balance and T-bet/Gata3 ratio (P<0.05). Conclusion: MALAT1 sponging miR-155 was involved with regulation of Th1/Th2 balance within CD4+ T cells, which might aid to develop therapies for amelioration of asthmatic inflammation.

scholarly journals CyclinD1 is a new target gene of tumor suppressor miR-520e in breast cancer

Open Medicine ◽  
2019 ◽  
Vol 14 (1) ◽  
pp. 913-919
Author(s):  
Quan Liang ◽  
Qingjuan Yao ◽  
GuoYing Hu
Keyword(s):  
Breast Cancer ◽  
Target Gene ◽  
Luciferase Reporter ◽  
Luciferase Reporter Gene ◽  
Luciferase Reporter Gene Assay ◽  
Gene Assay ◽  
Clinical Breast ◽  
Polymerase Chain ◽  
Cancer Tissues ◽  
Luciferase Reporters

AbstractObjectiveTo investigate the involvement of miR-520e in the modulation of cancer-promoting cyclinD1 in breast cancer.MethodsA reverse transcription-polymerase chain reaction (RT-PCR) was applied to test the regulation of miR-520e on cyclinD1. The binding of miR-520e to 3’-untranslated region (3’UTR) of cyclinD1 mRNA was predicted by an online bioinformatics website. The effect of miR-520e on the luciferase reporters with binding sites of miR-520e and 3’UTR of cyclinD1 mRNA was revealed using a luciferase reporter gene assay. The correlation between miR-520e and cyclinD1 in clinical breast cancer samples was detected through quantitative real-time PCR.ResultsThe expression of cyclinD1 was gradually reduced as the dose of miR-520e increased. Anti-miR-520e obviously induced cyclinD1 in breast cancer cells. After anti-miR-520e was introduced into the cells, the inhibition of cyclinD1 expression mediated by miR-520e was reversed. The binding of miR-520e with cyclinD1 was revealed via bioinformatics. Under the treatment of dose-increasing miR-520e or anti-miR-520e, the luciferase activities of cyclinD1 3’UTR vector were lower or higher by degrees. However, the activity of the mutant vector was not affected at all. Finally, in clinical breast cancer tissues the negative correlation of miR-520e with cyclinD1 was revealed.ConclusionIn conclusion, cyclinD1 is a new target of miR-520e in breast cancer.

scholarly journals YY1 activates EMI2 and promotes the progression of cholangiocarcinoma through the PI3K/Akt signaling axis

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Shuai Zhou ◽  
Kang Lin Qu ◽  
Jin Ang Li ◽  
Shi Lei Chen ◽  
Yi Gang Zhang ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Bile Duct ◽  
Signaling Pathway ◽  
Akt Signaling ◽  
Reporter Gene Assay ◽  
Luciferase Reporter ◽  
Akt Signaling Pathway ◽  
Luciferase Reporter Gene ◽  
Luciferase Reporter Gene Assay ◽  
Gene Assay

Abstract Background Cholangiocarcinoma (CCA) is one of the deadliest cancers of the digestive tract. The prognosis of CCA is poor and the 5-year survival rate is low. Bioinformatic analysis showed that early mitotic inhibitor 2 (EMI2) was overexpressed in CCA but the underlying mechanism is not known. Methods The data on bile duct carcinoma from TCGA and GEO databases were used to detect the expression of EMI2. The transcription factors of EMI2 were predicted using JASPAR and PROMO databases. Among the predicted transcription factors, YY1 has been rarely reported in cholangiocarcinoma, and was verified using the luciferase reporter gene assay. RT-PCR was performed to predict the downstream pathway of EMI2, and PI3K/Akt was suspected to be associated with it. Subsequently, in vivo and in vitro experiments were conducted to verify the effects of silencing and overexpressing EMI2 and YY1 on the proliferation, invasion, and metastasis of the bile duct cancer cells. Results EMI2 was highly expressed in CCA. Silencing EMI2 inhibited the proliferation, invasion, and migration of CCA cells, arrested cell cycle in the G1 phase, and promoted of apoptosis. The luciferase reporter gene assay showed that YY1 bound to the promoter region of EMI2, and after silencing YY1, the expression of EMI2 decreased and the progression of CCA was inhibited. Moreover, key proteins in the PI3K/Akt signaling pathway decreased after silencing EMI2. Conclusion EMI2 may be one of the direct targets of YY1 and promotes the progression of CCA through the PI3K/Akt signaling pathway.

scholarly journals lncRNA-TINCR Functions as a Competitive Endogenous RNA to Regulate the Migration of Mesenchymal Stem Cells by Sponging miR-761

2020 ◽  
Vol 2020 ◽  
pp. 1-10 ◽  
Author(s):  
Jiaqian Zheng ◽  
Yanni Huang ◽  
Ying Li ◽  
Jieqing Lai ◽  
Chen Chen ◽  
...  
Keyword(s):  
Signaling Pathway ◽  
Target Prediction ◽  
Luciferase Reporter ◽  
Luciferase Reporter Gene ◽  
Luciferase Reporter Gene Assay ◽  
Cellular Processes ◽  
Gene Assay ◽  
Direct Target ◽  
Competitive Endogenous Rna

Mounting evidences have indicated that terminal differentiation-induced lncRNA (TINCR) contributes to various cellular processes, such as proliferation, apoptosis, autophagy, migration, invasion, and metastasis. However, the function of TINCR in regulating migration of MSCs is largely unknown. In this study, the effects of TINCR on the migration of rat MSCs from the bone marrow were studied by Transwell assays and wound healing assays. Our results suggested that TINCR positively regulated migration of rMSCs. miR-761 mimics suppressed rMSC migration, whereas miR-761 inhibitor promoted migration. Target prediction analysis tools and dual-luciferase reporter gene assay identified Wnt2 as a direct target of miR-761. miR-761 could inhibit the expression of Wnt2. Further, the investigation about the function of TINCR in miR-761-induced migration of rMSCs was completed. These results demonstrated that TINCR took part in the regulation of miR-761-induced migration in rMSCs through the regulation of Wnt2 and its Wnt2 signaling pathway. Taken together, our results demonstrate that lncRNA-TINCR functions as a competitive endogenous RNA (ceRNA) to regulate the migration of rMSCs by sponging miR-761 which modulates the role of Wnt2. These findings provide evidence that lncRNA-TINCR has a chance to serve as a potential target for enhancing MSC homing through the miR-761/Wnt2 signaling pathway.

scholarly journals Sevoflurane inhibits proliferation, invasion, but enhances apoptosis of lung cancer cells by Wnt/β-catenin signaling via regulating lncRNA PCAT6/ miR-326 axis

2020 ◽  
Vol 15 (1) ◽  
pp. 159-172
Author(s):  
Guoning Su ◽  
Zhibing Yan ◽  
Min Deng
Keyword(s):  
Lung Cancer ◽  
Cancer Cells ◽  
Volatile Anesthetic ◽  
Lung Cancer Cells ◽  
Luciferase Reporter ◽  
Dependent Manner ◽  
Luciferase Reporter Gene ◽  
Western Blot Assay ◽  
Gene Assay ◽  
Cancer Tissues

AbstractSevoflurane was frequently used as a volatile anesthetic in cancer surgery. However, the potential mechanism of sevoflurane on lung cancer remains largely unclear. In this study, lung cancer cell lines (H446 and H1975) were treated by various concentrations of sevoflurane. 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assessment and colony formation assay were performed to detect the cell viability and proliferation, separately. Also, transwell assay or flow cytometry assay was applied as well to evaluate the invasive ability or apoptosis in lung cancer cells, respectively. Western blot assay was employed to detect the protein levels of β-catenin and Wnt5a. Moreover, quantitative real-time polymerase chain reaction (qRT-PCR) was used to examine the expression level of prostate cancer-associated transcript 6 (PCAT6) and miR-326 in lung cancer tissues and cells. The target interaction between miR-326 and PCAT6 or Wnt5a was predicted by bioinformatics analysis and verified by the dual-luciferase reporter gene assay. Sevoflurane inhibited the abilities on viability, proliferation, invasion, and activation of Wnt/β-catenin signaling, but promoted apoptosis of H446 and H1975 cells in a dose-dependent manner. The expression of PCAT6 was increased in lung cancer tissues and cells, except for that of miR-326. Besides, sevoflurane could lead to expressed limitation of PCAT6 or improvement of miR-326. This process presented a stepwise manner. Up-regulation of PCAT6 restored the suppression of sevoflurane on abilities of proliferation, invasion, rather than apoptosis, and re-activated the Wnt5a/β-catenin signaling in cells. Moreover, the putative binding sites between miR-326 and PCTA6 or Wnt5a were predicted by starBase v2.0 software online. PCAT6 suppressing effects on cells could be reversed by pre-treatment with miR-326 vector. The promotion of Wnt5a inverted effects led from miR-326 or sevoflurane. Our study indicated that sevoflurane inhibited the proliferation, and invasion, but enhanced the apoptosis in lung cancer cells by regulating the lncRNA PCAT6/miR-326/Wnt5a/β-catenin axis.

scholarly journals Dihydromyricetin promotes autophagy and attenuates renal interstitial fibrosis by regulating miR-155-5p/PTEN signaling in diabetic nephropathy

2019 ◽  
Author(s):  
Liming Guo ◽  
Kuibi Tan ◽  
Qun Luo ◽  
Xu Bai
Keyword(s):  
Diabetic Nephropathy ◽  
Signaling Pathway ◽  
Rat Model ◽  
Interstitial Fibrosis ◽  
Mtor Signaling ◽  
Luciferase Assay ◽  
Luciferase Reporter ◽  
Mtor Signaling Pathway ◽  
Renal Interstitial Fibrosis ◽  
Gene Assay

Diabetic nephropathy (DN) is the most common complication of diabetes and is prone to kidney failure. Dihydromyricetin (DHM) has been reported to have a variety of pharmacological activities. This study aims to explore the effect of DHM on DN and the underlying molecular mechanism. An in vivo DN rat model was established. The degree of renal interstitial fibrosis (RIF) was detected by hematoxylin-eosin (HE) staining, Masson's trichrome staining, and immunohistochemistry (IHC). In vitro, NRK-52E cells were divided into four groups: normal glucose (NG), high glucose (HG), HG+DHM, and HG+rapamycin (autophagy inhibitor). The levels of autophagy- and fibrosis-related proteins were analyzed by western blotting. The expression of miR-155-5p and phosphatase and tensin homolog deleted on chromosome ten (PTEN) and their relationship were assessed by quantitative reverse transcription (qRT)-PCR and dual luciferase reporter gene assay. Our results showed that RIF was increased in DN rat model and in HG-induced NRK-52E cells. DHM treatment attenuated the increased RIF and also increased autophagy. MiR-155-5p expression was increased, while PTEN expression was decreased in DN rat and cell model, and DHM reversed both effects. Dual luciferase assay showed that PTEN was the target gene of miR-155-5p. DHM inhibited HG-induced fibrosis and promoted autophagy by inhibiting miR-155-5p expression in NRK-52E cells. In addition, DHM promoted autophagy by inhibiting the PI3K/AKT/mTOR signaling pathway. In conclusion, DHM promotes autophagy and attenuates RIF by regulating the miR-155-5p/PTEN signaling and PI3K/AKT/mTOR signaling pathway in DN.

scholarly journals MiR-486 protects against acute myocardial infarction via regulation of PTEN

2021 ◽  
Vol 20 (9) ◽  
pp. 1845-1853
Author(s):  
Qinfeng Han ◽  
Zhong Xu ◽  
Xiaolei Zhang ◽  
Kun Yang ◽  
Zhifei Sun ◽  
...  
Keyword(s):  
Myocardial Infarction ◽  
Akt Signaling ◽  
Reporter Gene Assay ◽  
Expression Level ◽  
Luciferase Reporter ◽  
Akt Signaling Pathway ◽  
Luciferase Reporter Gene ◽  
Luciferase Reporter Gene Assay ◽  
Qrt Pcr ◽  
Gene Assay

Purpose: To investigate the effect of miR-486 on rats with acute myocardial infarction (AMI), and its mechanism of action.Methods: A rat model of AMI was established. They were randomly divided into 4 groups, namely, sham, model, agomiR-486 and antagomiR-486 groups, respectively. Rats in these different groups were treated with agomiR-21 (5 μL, 40 nmol/mL), antagomiR-21 (5 μL, 40 nmol/mL) or agomiR-NC (5 μL, 40 nmol/mL), respectively. Then, key miRNAs were sorted out using gene-chip assay and verified by quantitative reverse transcription polymerase chain reaction (qRT-PCR) assay. Luciferase reporter gene assay was conducted to determine the interaction between miR-486 and gene of PTEN. After intraperitoneal injection of agomiR-486 and antagomiR-486, hemodynamics was measured to determine the effect of miR-486 on myocardial function of the rats. The effect of miR-486 expression level on the expression of myocardial enzymes in serum, the morphology of myocardial tissues, and the apoptosis of myocardial tissues in rats, were investigated. Additionally, the effect of miR-486 expression level on PTEN/AKT signaling pathway in the rats was determined by Western blotting.Results: The results of gene-chip and qRT-PCR assays revealed that there were 8 differentially expressed genes in rat myocardial tissues in the model group when compared with the sham group. MiR-486 improved the cardiac function of rats and the morphology of myocardial tissues, but reduced AMI-induced apoptosis of myocardial cells and the expression of myocardial enzymes (markers of myocardial injury) in a dose-dependent manner (p < 0.05). The results of luciferase reporter gene assay showed that PTEN was a direct target of miR-486. In rat models of AMI, a raised expression of miR-486 remarkably suppressed the protein expression level of PTEN and up-regulated that of p-AKT/AKT (p < 0.05).Conclusion: MiR-486 protects against AMI in rats probably by targeting PTEN and activating the AKT signaling pathway. The results of the current study may provide new insights for the treatment of AMI.

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