scholarly journals circ_NOTCH3 Functions as a Protooncogene Competing With miR-205-5p, Modulating KLF12 Expression and Promoting the Development and Progression of Basal-Like Breast Carcinoma

2021 ◽  
Vol 10 ◽  
Author(s):  
Bing Guan ◽  
Qing Li ◽  
Hui-Zhen Zhang ◽  
Hai-Sheng Yang
Keyword(s):  
Breast Carcinoma ◽  
Cell Lines ◽  
Binding Sites ◽  
Target Gene ◽  
Cell Function ◽  
Ectopic Expression ◽  
Luciferase Reporter ◽  
Normal Tissues ◽  
Aggressive Histology ◽  
Dual Luciferase Reporter Assay

Breast cancer is the most common type of cancer diagnosed among women, and basal-like breast carcinoma (BLBC) has been associated with a more aggressive histology, poorer prognosis, and non-responsiveness to hormone therapy. In the present study, the role and molecular mechanism of circular (circ)_NOTCH3 in the development and progression for BLBC was identified. circ_RNAs array was used to screen the ectopic expression of hsa_circ_0109177 (circ_NOTCH3) in BLBC. RT-qPCR was conducted to evaluate the circ_NOTCH3 expression in BLBC tissues and paired normal tissues, as well as related cell lines. Cell function changes were analyzed following circ_NOTCH3 or micro (mi)RNA overexpression or co-expression. Bioinformatics analysis and dual-luciferase reporter assay were performed to predict and verify the binding sites between circ_NOTCH3 and miRNAs. Gene expression changes were assessed using western blotting. circ_NOTCH3 had a significantly higher expression in BLBC tissues and cell lines. The upregulation of circ_NOTCH3 promoted the proliferation, migration, invasion and inhibited the apoptosis for BLBC cells. The opposite results were observed following miR-205-5p overexpression. However, the co-expression of circ_NOTCH3 and miR-205-5p resulted in those restoration. circ_NOTCH3 is capable of binding to miR-205-5p, and upregulating its target gene KLF12, which can be downregulated by miR-205-5p overexpression and restored by the co-expression of circ_NOTCH3 and miR205-5p. circ_NOTCH3, being an protooncogene and a powerful biomarker, can function as a sponge, compete with miR-205-5p, modulate KLF12 expression, and promote the development and progression of BLBC.

scholarly journals Long non-coding RNA BCYRN1 exerts an oncogenic role in colorectal cancer through regulating miR-204-3p/KRAS axis

2020 ◽  
Author(s):  
Liu Yang ◽  
Yinan Zhang ◽  
Jun Bao ◽  
Ji-Feng Feng
Keyword(s):  
Colorectal Cancer ◽  
Cell Lines ◽  
Reporter Gene ◽  
Target Gene ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Reporter Assay ◽  
Luciferase Reporter Gene ◽  
Tumor Tissues ◽  
Dual Luciferase Reporter Assay

Abstract Background: It has been well documented that long non-coding RNAs (lncRNAs) regulate numerous characteristics of cancer, including proliferation, migration, metastasis, apoptosis, even metabolism. LncRNA BCYRN1 (BCYRN1) is a newly identified brain cytoplasmic lncRNA with 200-nucleotide, which was discovered highly expressed in tumor tissues of cancer, including hepatocellular carcinoma, gastric cancer and lung cancer. However, the roles of BCYRN1 in colorectal cancer (CRC) remain obscure. This study was designed to reveal the acts of BCYRN1 in the occurrence and progression of CRC.Methods: RT-PCR was used to detect the expression level of BCYRN1 in tumor tissues and CRC cell lines. Knock down BCYRN1 in CRC cells, evaluate cell proliferation changes by CCK-8 test, EdU test, and Ki-67 and PCNA expression; evaluate cell migration and invasion changes by wound healing assay, Transwell assay and invasion-related protein expression . Through flow cytometry analysis to assess whether BCYRN1 regulates apoptosis of CRC cells. The dual luciferase reporter gene detects the competitive binding of BCYRN1 to miR-204-3p. In vivo experiments to evaluate the effect of BCYRN1 on tumor development. TargetScan analysis and dual luciferase reporter gene detect the target gene of miR-204-3p. Rescue experiments verified the effect of BCYRN1 on CRC by regulating the effect of miR-204-3p on KRAS.Results: We found that compared with normal tissues and human intestinal epithelial cells (HIECs), BCYRN1 levels were significantly increased in tumor tissues and cell lines of CRC. We further determined that knockdown of BCYRN1 inhibited proliferation, migration, invasion, and promoted apoptosis of CRC cells. In addition, bioinformatics analysis and dual luciferase reporter assay showed that BCYRN1 served as a competitive endogenous RNA (ceRNA) to regulate the development of CRC through competitively binding to miR-204-3p. Further studies proved that overexpression of miR-204-3p reversed the effects of BCYRN1 on CRC. Next, TargetScan analysis and dual luciferase reporter assay indicated that KRAS was a target gene of miR-204-3p and negatively regulated by miR-204-3p. A series of rescue experiments showed that BCYRN1 affected the occurrence and development of CRC by regulating the effects of miR-204-3p on KRAS. In addition, tumorigenic experiments in CRC model mice confirmed that down-regulated BCYRN1 effectively inhibited tumor growth. Conclusions: Our findings suggested that BCYRN1 plays a carcinogenic role in CRC by regulating the miR-204-3p/KRAS axis.

scholarly journals miR-423-5p Inhibits Osteosarcoma Proliferation and Invasion Through Directly Targeting STMN1

2018 ◽  
Vol 50 (6) ◽  
pp. 2249-2259 ◽  
Author(s):  
Xuesong Wang ◽  
Lei Peng ◽  
Xiaojin Gong ◽  
Xiugong Zhang ◽  
Ruifu Sun ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Cell Lines ◽  
Colony Formation ◽  
Target Gene ◽  
Suppressor Gene ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Osteosarcoma Cell ◽  
Direct Target Gene ◽  
Dual Luciferase Reporter Assay

Background/Aims: Increasing evidences suggest that dysregulated expression of miRNAs contributes to the progression of various tumors. However, the underlying function of miR-423-5p in osteosarcoma remains unexplored. Methods: The expression of miR-423-5p and STMN1 were determined in osteosarcoma samples and cell lines via quantitative real-time PCR. Colony formation and Cell Counting Kit-8 (CCK-8) assays were performed to measure cell proliferation ability and transwell analysis was used to detect cell invasion, and dual luciferase reporter assay was perform to analysis the interaction between the miR-423-5p and STMN1. Results: The expression levels of miR-423-5p and STMN1 in the osteosarcoma tissues and cell lines were measured by qRT-PCR. Cell viability was determined using the clone formation and CCK-8 assays. A dual-luciferase reporter and Western blot were performed to stdudy the target gene of miR-423-5p. Here, we showed that miR-423-5p expression was downregulated in osteosarcoma tissues and cell lines. However, the expression of stathmin1 (STMN1) was downregulated in osteosarcoma tissues and cell lines. Moreover, STMN1 expression level was negatively correlated with the miR-423-5p expression in the osteosarcoma tissues. We identified STMN1 was a direct target gene of miR-423-5p in osteosarcoma cell. Overexpression of miR-423-5p inhibited osteosarcoma cell proliferation, colony formation and invasion. Furthermore, we demonstrated that STMN1 was involved in miR-423-5p-mediated cell behavior such as cell proliferation, colony formation and invasion in the osteosarcoma cell. Conclusion: Our present study indicated that miR-423-5p acted as a tumor suppressor gene in osteosarcoma partly through inhibiting STMN1 expression.

scholarly journals Long non-coding RNA BCYRN1 exerts an oncogenic role in colorectal cancer through regulating miR-204-3p/KRAS axis

2020 ◽  
Author(s):  
Liu Yang ◽  
Yinan Zhang ◽  
Jun Bao ◽  
Ji-Feng Feng
Keyword(s):  
Colorectal Cancer ◽  
Cell Lines ◽  
Reporter Gene ◽  
Target Gene ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Reporter Assay ◽  
Luciferase Reporter Gene ◽  
Dual Luciferase Reporter Assay

Abstract Background: It has been well documented that long non-coding RNAs (lncRNAs) regulate numerous characteristics of cancer, including proliferation, migration, metastasis, apoptosis, and even metabolism. LncRNA BCYRN1 (BCYRN1) is a newly identified brain cytoplasmic lncRNA with 200 nucleotides that was discovered to be highly expressed in tumour tissues, including those of hepatocellular carcinoma, gastric cancer and lung cancer. However, the roles of BCYRN1 in colorectal cancer (CRC) remain obscure. This study was designed to reveal the role of BCYRN1 in the occurrence and progression of CRC.Methods: RT-PCR was used to detect the expression level of BCYRN1 in tumour tissues and CRC cell lines. BCYRN1 was knocked down in CRC cells, and cell proliferation changes were evaluated by cell counting kit-8 (CCK-8), 5-ethynyl-2’-deoxyuridine (EdU), and Ki-67 and proliferating cell nuclear antigen (PCNA) expression assays. Cell migration and invasion changes were evaluated by wound healing, Transwell and invasion-related protein expression assays. Flow cytometry analysis was used to assess whether BCYRN1 regulates the apoptosis of CRC cells. The dual luciferase reporter gene detects the competitive binding of BCYRN1 to miR-204-3p. In vivo experiments were performed to evaluate the effect of BCYRN1 on tumour development. TargetScan analysis and dual luciferase reporter gene assays were applied to detect the target gene of miR-204-3p. Rescue experiments verified that BCYRN1 affects CRC by regulating the effect of miR-204-3p on KRAS.Results: We found that compared with normal tissues and human intestinal epithelial cells (HIECs), CRC tumour tissues and cell lines had significantly increased BCYRN1 levels. We further determined that knockdown of BCYRN1 inhibited the proliferation, migration, and invasion and promoted the apoptosis of CRC cells. In addition, bioinformatics analysis and dual luciferase reporter assay showed that BCYRN1 served as a competitive endogenous RNA (ceRNA) to regulate the development of CRC through competitively binding to miR-204-3p. Further studies proved that overexpression of miR-204-3p reversed the effects of BCYRN1 on CRC. Next, TargetScan analysis and dual luciferase reporter assay indicated that KRAS is a target gene of miR-204-3p and is negatively regulated by miR-204-3p. A series of rescue experiments showed that BCYRN1 affected the occurrence and development of CRC by regulating the effects of miR-204-3p on KRAS. In addition, tumorigenesis experiments in a CRC mouse model confirmed that BCYRN1 downregulation effectively inhibited tumour growth.Conclusions: Our findings suggest that BCYRN1 plays a carcinogenic role in CRC by regulating the miR-204-3p/KRAS axis.

scholarly journals Long Non-Coding RNA BCYRN1 Exerts an Oncogenic Rrole in Colorectal Cancer through Regulating miR-204-3p/KRAS Axis

2020 ◽  
Author(s):  
Liu Yang ◽  
Yinan Zhang ◽  
Jun Bao ◽  
Ji-Feng Feng
Keyword(s):  
Colorectal Cancer ◽  
Cell Lines ◽  
Reporter Gene ◽  
Target Gene ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Reporter Assay ◽  
Luciferase Reporter Gene ◽  
Tumor Tissues ◽  
Dual Luciferase Reporter Assay

Abstract Background: It has been well documented that long non-coding RNAs (lncRNAs) regulate numerous characteristics of cancer, including proliferation, migration, metastasis, apoptosis, even metabolism. LncRNA BCYRN1 (BCYRN1) is a newly identified brain cytoplasmic lncRNA with 200-nucleotide, which was discovered highly expressed in tumor tissues of cancer, including hepatocellular carcinoma, gastric cancer and lung cancer. However, the roles of BCYRN1 in colorectal cancer (CRC) remain obscure. This study was designed to reveal the acts of BCYRN1 in the occurrence and progression of CRC. Methods: RT-PCR was used to detect the expression level of BCYRN1 in tumor tissues and CRC cell lines. Knock down BCYRN1 in CRC cells, evaluate cell proliferation changes by CCK-8 test, EdU test, and Ki-67 and PCNA expression; evaluate cell migration and invasion changes by wound healing assay, Transwell assay and invasion-related protein expression . Through flow cytometry analysis to assess whether BCYRN1 regulates apoptosis of CRC cells. The dual luciferase reporter gene detects the competitive binding of BCYRN1 to miR-204-3p. In vivo experiments to evaluate the effect of BCYRN1 on tumor development. TargetScan analysis and dual luciferase reporter gene detect the target gene of miR-204-3p. Rescue experiments verified the effect of BCYRN1 on CRC by regulating the effect of miR-204-3p on KRAS.Results: We found that compared with normal tissues and human intestinal epithelial cells (HIECs), BCYRN1 levels were significantly increased in tumor tissues and cell lines of CRC. We further determined that knockdown of BCYRN1 inhibited proliferation, migration, invasion, and promoted apoptosis of CRC cells. In addition, bioinformatics analysis and dual luciferase reporter assay showed that BCYRN1 served as a competitive endogenous RNA (ceRNA) to regulate the development of CRC through competitively binding to miR-204-3p. Further studies proved that overexpression of miR-204-3p reversed the effects of BCYRN1 on CRC. Next, TargetScan analysis and dual luciferase reporter assay indicated that KRAS was a target gene of miR-204-3p and negatively regulated by miR-204-3p. A series of rescue experiments showed that BCYRN1 affected the occurrence and development of CRC by regulating the effects of miR-204-3p on KRAS. In addition, tumorigenic experiments in CRC model mice confirmed that down-regulated BCYRN1 effectively inhibited tumor growth. Conclusions: Our findings suggested that BCYRN1 plays a carcinogenic role in CRC by regulating the miR-204-3p/KRAS axis.

scholarly journals LncRNA PTPRG-AS1 Promotes the Metastasis of Hepatocellular Carcinoma by Enhancing YWHAG

2021 ◽  
Vol 2021 ◽  
pp. 1-14
Author(s):  
Gaoyang Chen ◽  
Zhisheng Zhang ◽  
Yan Li ◽  
Lu Wang ◽  
Yanqing Liu
Keyword(s):  
Hepatocellular Carcinoma ◽  
Target Gene ◽  
Biological Function ◽  
Malignant Tumors ◽  
Luciferase Reporter ◽  
Real Time Quantitative Pcr ◽  
Normal Tissues ◽  
Dual Luciferase Reporter Assay

Objectives. Hepatocellular carcinoma (HCC) is one of the most common malignant tumors. LncRNA PTPRG-AS1 (PTPRG-AS1) has been confirmed to function as a regulator in various cancers, whose function during HCC tumorigenesis is still not clear now. Thus, we aim to dig out the biological function and its mechanisms of PTPRG-AS1 in HCC. Methods. PTPRG-AS1 relative expression in tissues and cells was detected and analyzed using real-time quantitative PCR (qRT-PCR). Subcellular distribution of PTPRG-AS1 was examined by FISH experiments. The effects of PTPRG-AS1 in the growth of HCC were studied by in vitro CCK-8 experiments, transwell invasion experiments, and in vivo xenograft tumor experiments. Dual-Luciferase reporter assay was performed to verify the interaction between PTPRG-AS1 and miR-199a-3p or miR-199a-3p and its target gene, YWHAG. Results. PTPRG-AS1 was upregulated in HCC tissues compared with adjacent normal tissues. We identified PTPRG-AS1 mainly localized in the cytoplasm of HCC cells. Downregulation of PTPRG-AS1 suppressed HCC progression, while overexpression of PTPRG-AS1 showed the opposite effects. Furthermore, PTPRG-AS1 served as a miR-199a-3p sponge and positively regulated YWHAG expression. Besides, PTPRG-AS1 could promote HCC through miR-199a-3p/YWHAG axis. Conclusions. Taken together, we demonstrated PTPRG-AS1 may serve as a ceRNA and reversely regulates the expression of miR-199a-3p, thus facilitating HCC tumorigenesis and metastasis, which is expected to provide new clues for the treatment of HCC.

scholarly journals Long non-coding RNA BCYRN1 exerts an oncogenic role in colorectal cancer by regulating the miR-204-3p/KRAS axis

2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Liu Yang ◽  
Yinan Zhang ◽  
Jun Bao ◽  
Ji-Feng Feng
Keyword(s):  
Colorectal Cancer ◽  
Cell Lines ◽  
Reporter Gene ◽  
Target Gene ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Reporter Assay ◽  
Luciferase Reporter Gene ◽  
Dual Luciferase Reporter Assay

Abstract Background It has been well documented that long non-coding RNAs (lncRNAs) regulate numerous characteristics of cancer, including proliferation, migration, metastasis, apoptosis, and even metabolism. LncRNA BCYRN1 (BCYRN1) is a newly identified brain cytoplasmic lncRNA with 200 nucleotides that was discovered to be highly expressed in tumour tissues, including those of hepatocellular carcinoma, gastric cancer and lung cancer. However, the roles of BCYRN1 in colorectal cancer (CRC) remain obscure. This study was designed to reveal the role of BCYRN1 in the occurrence and progression of CRC. Methods RT-PCR was used to detect the expression level of BCYRN1 in tumour tissues and CRC cell lines. BCYRN1 was knocked down in CRC cells, and cell proliferation changes were evaluated by cell counting kit-8 (CCK-8), 5-ethynyl-2′-deoxyuridine (EdU), and Ki-67 and proliferating cell nuclear antigen (PCNA) expression assays. Cell migration and invasion changes were evaluated by wound healing, Transwell and invasion-related protein expression assays. Flow cytometry analysis was used to assess whether BCYRN1 regulates the apoptosis of CRC cells. The dual luciferase reporter gene detects the competitive binding of BCYRN1 to miR-204-3p. In vivo experiments were performed to evaluate the effect of BCYRN1 on tumour development. TargetScan analysis and dual luciferase reporter gene assays were applied to detect the target gene of miR-204-3p. Rescue experiments verified that BCYRN1 affects CRC by regulating the effect of miR-204-3p on KRAS. Results We found that compared with normal tissues and human intestinal epithelial cells (HIECs), CRC tumour tissues and cell lines had significantly increased BCYRN1 levels. We further determined that knockdown of BCYRN1 inhibited the proliferation, migration, and invasion and promoted the apoptosis of CRC cells. In addition, bioinformatics analysis and dual luciferase reporter assay showed that BCYRN1 served as a competitive endogenous RNA (ceRNA) to regulate the development of CRC through competitively binding to miR-204-3p. Further studies proved that overexpression of miR-204-3p reversed the effects of BCYRN1 on CRC. Next, TargetScan analysis and dual luciferase reporter assay indicated that KRAS is a target gene of miR-204-3p and is negatively regulated by miR-204-3p. A series of rescue experiments showed that BCYRN1 affected the occurrence and development of CRC by regulating the effects of miR-204-3p on KRAS. In addition, tumorigenesis experiments in a CRC mouse model confirmed that BCYRN1 downregulation effectively inhibited tumour growth. Conclusions Our findings suggest that BCYRN1 plays a carcinogenic role in CRC by regulating the miR-204-3p/KRAS axis.

scholarly journals Hsa-miR-494-3p attenuates gene HtrA3 transcription to increase inflammatory response in hypoxia/reoxygenation HK2 Cells

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Qian Gong ◽  
Zhi-ming Shen ◽  
Zhe Sheng ◽  
Shi Jiang ◽  
Sheng-lin Ge
Keyword(s):  
Cardiac Surgery ◽  
Inflammatory Response ◽  
Target Gene ◽  
Kidney Injury ◽  
Human Kidney ◽  
Luciferase Reporter ◽  
The Relationship ◽  
Hk2 Cells ◽  
Dual Luciferase Reporter Assay ◽  
Hypoxia Reoxygenation

AbstractThe occurrence of cardiac surgery-associated acute kidney injury (CSA-AKI) increases hospital stay and mortality. MicroRNAs has a crucial role in AKI. This objective of the current study is to explore the function of hsa-miR-494-3p in inflammatory response in human kidney tubular epithelial (HK2) cells with hypoxia/reoxygenation. According to KDIGO standard, patients after cardiac surgery with cardiopulmonary bypass were divided into two groups: AKI (n = 10) and non-AKI patients (n = 8). HK2 were raised in the normal and hypoxia/reoxygenation circumstances and mainly treated by overexpression ofmiR-494-3p and HtrA3. The relationship between miR-494-3p and HtrA3 was determined by dual-luciferase reporter assay. Our result showed that Hsa-miR-494-3p was elevated in the serum of patients with CSA-AKI, and also induced in hypoxic reoxygenated HK2 cells. Hsa-miR-494-3p also increased a hypoxia-reoxygenation induced inflammatory response in HK2 cells. Moreover, as a target gene of miR-494-3p, overexpression of HtrA3 downregulated the hypoxia-reoxygenation induced inflammatory response in HK2 cells. Overexpression of hsa-miR-494-3p-induced inflammatory response was inhibited by overexpression of HtrA3. Collectively, we identified that hsa-miR-494-3p, a miRNA induced in both circulation of AKI patients and hypoxia-reoxygenation-treated HK2 cells, enhanced renal inflammation by targeting HtrA3, which may suggest a possible role as a new therapeutic target for CSA-AKI.

scholarly journals MiR-196b Promotes the Invasion and Migration of Lung Adenocarcinoma Cells by Targeting AQP4

2021 ◽  
Vol 20 ◽  
pp. 153303382098586
Author(s):  
Xuhui Wu ◽  
Gongzhi Wu ◽  
Huaizhong Zhang ◽  
Xuyang Peng ◽  
Bin Huang ◽  
...  
Keyword(s):  
Lung Adenocarcinoma ◽  
Cell Invasion ◽  
Target Gene ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Reporter Assay ◽  
Invasion And Migration ◽  
And Migration ◽  
Dual Luciferase Reporter Assay

Objective: We aimed to investigate the mechanism of the regulatory axis of miR-196b/AQP4 underlying the invasion and migration of lung adenocarcinoma (LUAD) cells. Methods: LUAD miRNA and mRNA expression profiles were downloaded from TCGA database and then differential analysis was used to identify the target miRNA. Target gene for the miRNA was obtained via prediction using 3 bioinformatics databases and intersection with the differentially expressed mRNAs searched from TCGA-LUAD. Then, qRT-PCR and western blot were used to validate the expression of miR-196b and AQP4. Dual-luciferase reporter assay was performed to confirm the targeting relationship between miR-196b and AQP4. Transwell assay was used to investigate the migration and invasion of LUAD cells. Results: MiR-196b was screened out by differential and survival analyses, and the downstream target gene AQP4 was identified. In LUAD, miR-196b was highly expressed while AQP4 was poorly expressed. Besides, overexpression of miR-196b promoted cell invasion and migration, while overexpression of AQP4 had negative effects. Moreover, the results of the dual-luciferase reporter assay suggested that AQP4 was a direct target of miR-196b. In addition, we also found that overexpressing AQP4 could suppress the promotive effect of miR-196b on cancer cell invasion and migration. Conclusion: MiR-196b promotes the invasion and migration of LUAD cells by down-regulating AQP4, which helps us find new molecular targeted therapies for LUAD.

scholarly journals Hsa-miR-105-1 Regulates Cisplatin-Resistance in Ovarian Carcinoma Cells by Targeting ANXA9

2021 ◽  
Vol 2021 ◽  
pp. 1-11
Author(s):  
Xinxin Kou ◽  
Hui Ding ◽  
Lei Li ◽  
Hongtu Chao
Keyword(s):  
Ovarian Carcinoma ◽  
Cell Lines ◽  
Target Gene ◽  
Molecular Mechanisms ◽  
Cisplatin Resistance ◽  
Luciferase Reporter ◽  
Luciferase Reporter Gene ◽  
Qrt Pcr ◽  
Gene Assay ◽  
Increased Sensitivity

Purpose. Cisplatin is one of the most effective drugs for treating ovarian carcinoma (OC), which is among the most lethal types of carcinoma. However, the chemoresistance to cisplatin that develops over time leads to a poor clinical outcome for many OC patients. Therefore, it is necessary to clearly understand the molecular mechanisms of chemoresistance. In this study, we examined how Hsa-miR-105-1 functions in cisplatin-resistant OC cells. Methods. The levels of Hsa-miR-105-1 expression in cisplatin-sensitive and resistant OC cell lines were detected by qRT-PCR. The target gene of Hsa-miR-105-1 was predicted by using the TargetScan and Starbase databases and verified by the double luciferase reporter gene assay. The target gene of Hsa-miR-105-1 was identified as ANXA9, and ANXA9 expression was evaluated by qRT-PCR, western blotting, and immunofluorescence. To validate the function of Hsa-miR-105-1 in OC cells, we silenced or overexpressed Hsa-miR-105-1 in cisplatin-sensitive or resistant OC cell lines, respectively. Furthermore, the expression levels of several apoptosis-related proteins, including P53, P21, E2F1, Bcl-2, Bax, and caspase-3, were examined by western blot analysis. Results. The levels of Hsa-miR-105-1 expression were abnormally downregulated in cisplatin-resistant OC cells, while ANXA9 expression was significantly upregulated in those cells. Treatment with an Hsa-miR-105-1 inhibitor promoted the expression of ANXA9 mRNA and protein, enhanced the resistance to cisplatin, and attenuated the cell apoptosis induced by cisplatin in cisplatin-sensitive OC cells. Moreover, treatment with Hsa-miR-105-1 mimics inhibited ANXA9 expression, which further increased the levels of P53, P21, and Bax expression and decreased the levels of E2F1 and Bcl-2 expression, finally resulting in an increased sensitivity to cisplatin in cisplatin-resistant OC cells. Conclusion. We found that a downregulation of Hsa-miR-105-1 expression enhanced cisplatin-resistance, while an upregulation of Hsa-miR-105-1 restored the sensitivity of OC cells to cisplatin. The Hsa-miR-105-1/ANXA9 axis plays an important role in the cisplatin-resistance of OC cells.

scholarly journals lncRNA CRNDE promotes the proliferation and metastasis by acting as sponge miR-539-p to regulate POU2F1 expression in HCC

2020 ◽  
Author(s):  
Zhixi Li ◽  
Gang Wu ◽  
Jie Li ◽  
Youyu Wang ◽  
Xueming Ju ◽  
...  
Keyword(s):  
Western Blot ◽  
Luciferase Reporter ◽  
Akt Pathway ◽  
Migration And Invasion ◽  
Reporter Assay ◽  
Normal Cells ◽  
Normal Tissues ◽  
Qrt Pcr ◽  
Hcc Cells ◽  
Dual Luciferase Reporter Assay

Abstract Background This article focuses on the roles and mechanism of lncRNA CRNDE on the progression of HCC. Methods We used qRT-PCR to detect the expression of lncRNA CRNDE in HCC cells, normal cells and clinical tissues. MTT assay, FCM analysis, Transwell migration and invasion assay were used to detect the effects of lncRNA CRNDE on cell viability, apoptosis, migration and invasion of HCC cells. The expression of apoptosis-related proteins Bcl-2, Bax, Cleaved Caspase 3, Cleaved Caspase 9, EMT epithelial marker E-cadherin and mesothelial marker Vimentin were analyzed by Western blot. Online prediction software was used to predict the binding sites between lncRNA CRNDE and miR-539-5p, or miR-539-5p and POU2F1 3’UTR. Dual luciferase reporter assay, qRT-PCR and RNA pulldown were used to detect target-relationship between lncRNA CRNDE and miR-539-5p. Dual luciferase reporter assay, qRT-PCR, Western blot and Immunofluorescence were used to detect target-relationship between miR-539-5p and POU2F1. qRT-PCR was used to detect the expression of miR-539-5p and POU2F1 in clinical tissues. Rescue experiments was used to evaluate the association among lncRNA CRNDE, miR-539-5p and POU2F1. Finally, we used Western blot to detect the effects of lncRNA CRNDE, miR-539-5p and POU2F1 on NF-κB and AKT pathway. Results lncRNA CRNDE was highly expressed in HCC cells and HCC tissues compared with normal cells and the corresponding adjacent normal tissues. lncRNA CRNDE promoted the cell viability, migration and invasion of HCC cells, while inhibited the apoptosis and promoted the EMT process of HCC cells. lncRNA CRNDE adsorbed miR-539-5p acts as a competitive endogenous RNA to regulate POU2F1 expression indirectly. In HCC clinical tissues, miR-539-5p expression decreased and POU2F1 increased compared with the corresponding adjacent normal tissues. lncRNA CRNDE/miR-539-5p/POU2- F1 participated the NF-κB and AKT pathway in HCC. Conclusion lncRNA CRNDE promotes the expression of POU2F1 by adsorbing miR-539-5p, thus promoting the progression of HCC. Keywords: HCC, lncRNA CRNDE, miR-539-5p, POU2F1, ceRNA

Sign in / Sign up

Export Citation Format

Share Document