scholarly journals MiR-624-5p enhances cell resistance against cisplatin via PDGFRA/Stat3/PI3K axis in ovarian cancer

2020 ◽  
Vol 19 (4) ◽  
pp. 691-698
Author(s):  
Lin I-Ju ◽  
Tian YongJie
Keyword(s):  
Ovarian Cancer ◽  
Cell Viability ◽  
Cell Line ◽  
Cell Lines ◽  
Clinical Stage ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Reporter Assay

Purpose: The purpose of this study was to evaluate the role of miR-624-5p in ovarian cancer.Methods: MiR-624-5p expression in ovarian cancer {OC) cell lines and normal cells (NCs) was evaluated and compared the differential miR-624-5p in OC A2780 cells and cisplatin-resistant OC cell line (A2780/DDP). CCK-8 was used to evaluate changes in cell viability of the A2780 and A2780/DDP cell lines as well as silenced miR-624-5p. Western Blot examined the Stat3 and phosphorylated Pi3k. The binding between PDGFRA and miR-624-5p was predicted on Targetscan and verified through Luciferase Reporter Assay. The role of PDGFRA in A2780/DDP by overexpressing PDGFRA was evaluated by RT-qPCR and CCK-8 assays. RT-qPCR assay also measured miR-624-5p expression responsive to different dosages of cisplatin and CCK8 examined viability levels correspondingly. In addition, the interplay of PDGFRA and miR-624-5p by combined downregulation of both miR-624-5pand PDGFRA were evaluated.Results: OC cells had higher miR-624-5p expression than NCs but lower compared to cisplatinresistant A2780/DDP cells. A2780/DDP cells had higher viability than OC cell line A2780. Stat3 and phosphorylated PI3K were activated in A2780/DDP cells. Silencing miR-624-5p led to lower viability inA2780/DDP cells. miR-624-5p expression dropped as the cisplatin concentration increased, resulting in decreasing viability respectively. Luciferase Reporter assay validated the binding of miR-624-5p and PDGFRA in A2780/DDP cells. Overexpressed PDGFRA induced lower cell viability in A2780/DDP cells. Downregulation of PDGFRA partially restored the lowered viability and inhibited Stat3 as well as phosphorylated Pi3k induced by miR-624-5p inhibitor.Conclusion: MiR-624-5p could add to the cellular resistance to cisplatin in OC in-vitro model, which indicated that it might help unveil the mystery of drug-resistance in clinical stage of ovarian cancer. Keywords: MiR-624-5p, resistance, cisplatin, PDGFRA/Stat3/PI3K, ovarian cancer

scholarly journals Knockdown of LincRNA PADNA Promotes Bupivacaine-Induced Neurotoxicity by miR-194/FBXW7 Axis

2020 ◽  
Author(s):  
Fan Yuning ◽  
Chen Liang ◽  
Wang Tenghuan ◽  
Nan Zhenhua ◽  
Shengkai Gong
Keyword(s):  
Functional Analysis ◽  
Western Blot ◽  
Cell Viability ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Reporter Assay ◽  
Drg Neurons ◽  
Dual Luciferase Reporter Assay

Abstract The aim of the study was to explore the function and mechanism of lincRNA PADNA in bupivacaine-induced neurotoxicity. Mouse DRG neurons were cultured in vitro and treated with bupivacaine to establish the neurotoxicity model. Caspase3 activity, cell viability, tunel assay were analyzed to assess the role of lincRNA PADNA. Dual-luciferase reporter assay was used to determine the binding target of lincRNA PANDA. The expression of lincRNA PADNA was significantly increased with the increasing concentration of bupivacaine. Functional analysis revealed that knockdown of lincRNA PADNA accelerated the caspase3 activity and inhibited the cell viability. Western blot showed that knockdown of lincRNA PADNA promoted the occurrence of cleaved-caspase3. We also proved that lincRNA PADNA may bind with miR-194. Overexpression of miR-194 could rescued the function of lincRNA PADNA, suggesting that lincRNA PADNA may sponge miR-194. In addition, we provided new evidences that lincRNA PADNA/miR-194/FBXW7 axis play an important role in the neurotoxicity process. We performed comprehensive experiments to verify the function and mechanism of lincRNA PADNA in bupivacaine-induced neurotoxicity. Our study provided new evidences and clues for prevention of neurotoxicity.

scholarly journals Mcl-1 Is a Novel Target of miR-26b That Is Associated with the Apoptosis Induced by TRAIL in HCC Cells

2015 ◽  
Vol 2015 ◽  
pp. 1-9 ◽  
Author(s):  
Chunlin Jiang ◽  
Jianting Long ◽  
Baoxian Liu ◽  
Xiaoyan Xie ◽  
Ming Kuang
Keyword(s):  
Cell Lines ◽  
Bioinformatic Analysis ◽  
Negative Regulator ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Reporter Assay ◽  
Mrna Transfection ◽  
Novel Target ◽  
Hcc Cells

Aim. To investigate the role of miR-26b and Mcl-1 in TRAIL-inducing cell death in hepatocellular carcinoma.Methods. The expression of miR-26b and Mcl-1 in HCC was detected by RT-qPCR and western blot. The regulation of Mcl-1 by miR-26b was determined by luciferase reporter assay. MTT and flow cytometry were employed to detect the cell viability and apoptosis.Results. miR-26b is commonly downregulated in HCC cell lines compared with the LO2 cell line. In contrast, the Mcl-1 expression is upregulated in HCC cell lines. Bioinformatic analysis identified a putative target site in the Mcl-1 mRNA for miR-26b and luciferase reporter assay showed that miR-26b directly targeted the 3′-UTR (3′-Untranslated Regions) of Mcl-1 mRNA. Transfection of miR-26b mimics suppressed Mcl-1 expression in HCC cells and sensitized the cancer cells to TRAIL (tumor necrosis factor-related apoptosis-inducing ligand) cytotoxicity. In addition, transfection of HCC cells with Mcl-1 expression plasmid abolished the sensitization effect of miR-26b to TRAIL-inducing apoptosis.Conclusions. Our study showed that miR-26b was a negative regulator of Mcl-1 gene and sensitized TRAIL-inducing apoptosis in HCC cells, suggesting that the miR-26b-Mcl-1 pathway might be a novel target for the treatment of HCC.

scholarly journals MicroRNA-221 Targets FBXW11 to Inhibit NPC Cell Proliferation by Regulating PTEN Signaling

2021 ◽  
Author(s):  
Jun Liu ◽  
Chengyi Song ◽  
Zhuoping Liang ◽  
Xiang Long ◽  
Min Guo ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Lines ◽  
Biological Function ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Reporter Assay ◽  
Cell Cycle Transition ◽  
Polymerase Chain ◽  
Molecular Regulatory Mechanisms

Abstract Background: In the current study, we aim to demonstrate the biological function and molecular regulatory mechanisms of miR-221 in human nasopharyngeal carcinoma (NPC). Methods: The quantitative real time-polymerase chain reaction was used to measure the expression of miR-221 in NPC clinical tissues and cells. And the flow cytometry assay was used to demonstrate the role of miR-221 on cell cycle, and the potential target of miR-221 was predicted and identified using luciferase reporter assay.Results: Our results demonstrate that miR-221 expression was significantly decreased in NPC tissues and cell lines. We also confirmed that inhibition of miR-221 could induce G1/S cell cycle transition through upregulation of the cyclin D-CDK4/6 complex but not cyclin E-CDK2 complexes. Furthermore, luciferase reporter assay demonstrated that miR-221 could directly bind to the 3’-UTR of FBXW11. FBXW11 expression was found to increase in NPC, and was inversely correlated with miR-221 expression; thus, FBXW11 expression interfered in the biological function of miR-221. We further confirmed that miR-221 targeted FBXW11 to inhibit proliferation and promote apoptosis in NPC cell lines through regulating the PTEN signaling pathway. Conclusion: our findings suggest that miR-221 plays an important role as a tumor suppressive factor in the occurrence and progression of NPC.

scholarly journals miR-455 Inhibits the Viability and Invasion by Targeting RAB18 in Hepatocellular Carcinoma

2021 ◽  
Vol 2021 ◽  
pp. 1-10
Author(s):  
Chenghong Wang ◽  
Guicai Zhu ◽  
Miaolin Yu ◽  
Xiufang Mi ◽  
Honghua Qu
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Viability ◽  
Cell Lines ◽  
Luciferase Reporter ◽  
Common Cancer ◽  
Huh7 Cells ◽  
Proliferation In Vitro

Background. Hepatocellular carcinoma (HCC) has been regarded as the fifth most common cancer worldwide with a low prognosis. miR-455 usually played the role of a tumor suppressor in multiple cancers. The aim of this study was to investigate the roles of miR-455 in HCC. Materials and Methods. Cell viability and invasion were measured by CCK8 and Transwell assays. Luciferase reporter assay was performed to verify that miR-455 directly binds to the 3′-noncoding region (UTR) of RAB18 mRNA in Huh7 cells. Results. The expression of miR-455 was lower in HCC tissues and cell lines than in nontumor tissues and normal cell line, and downregulation of miR-455 was connected with worse outcome of HCC patients. miR-455 suppressed cell proliferation in vitro and in vivo, and it inhibited the abilities of cell invasion and EMT in HCC. RAB18 was upregulated in HCC tissues and cell lines, and the expression of RAB18 was regulated by miR-455. RAB18 reversed partial roles of miR-455 on cell viability and invasion in HCC. Conclusion. miR-455 inhibited cell viability and invasion by directly targeting the 3′-UTR of RAB18 mRNA of hepatocellular carcinoma.

HNF1α Contributes to Radio-Resistance in Cervical Cancer Via Regulating the CircHNF1α/miR-204-3p/RAD51D Regulatory Axis

2020 ◽  
Author(s):  
Hui Du ◽  
Nai-Yi Zou ◽  
Hong-Ling Zuo ◽  
Xue-Yuan Zhang ◽  
Shu-Chai Zhu
Keyword(s):  
Cervical Cancer ◽  
Cell Lines ◽  
Mrna Level ◽  
Critical Role ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Pcr Analysis ◽  
Reporter Assay ◽  
Therapeutic Benefits

Abstract Background: Radiotherapy as an important primary treatment has effectively improved the survival of patients with cervical cancer (CC). However, some patients do not show optimal benefits of radiotherapy due to radio-resistance. Therefore, identification of biomarkers of radio-resistance and unraveling the underlying mechanisms of radio-resistance is a key imperative for these patients.Methods: The expression levels of circHNF1α, miRNAs, and mRNA in tissues and cell lines were detect by qRT-PCR analysis. The levels of proteins were analyzed by western blot. Cell proliferation ability was measured by colony formation assay. RNA pull-down and uciferase reporter assay analysis were performed to identify the sponging microRNAs of circHNF1α. The target gene of miR-204-3p was determined by luciferase reporter assay. Chromatin immunoprecipitation (ChIP) analysis and luciferase reporter assay were performed to identify the transcription factor of circHNF1α. Results: We found significant upregulation of HNF1α expression in radio-resistant cervical cancer tissues and cell lines. Depletion of HNF1α reduced whereas overexpression of HNF1α promoted the resistance of CC cells to irradiation in vitro and in vivo. HNF1α positively regulated RAD51D at the protein level but not at the mRNA level, thus attenuating radio-resistance of CC cells. Mechanistically, upregulation of HNF1α enhanced circHNF1α transcription and promoted circHNF1α biogenesis, which in turn sponged miR-204-3p and thus relieved their repression of the RAD51D expression. The HFN1α/circHNF1α/miR-204-3p/RAD51D regulatory axis was found to play a critical role in conferring radio-resistance of CC cells.Conclusions: Dysregulation of the HFN1α/circHNF1α/miR-204-3p/RAD51D axis may promote the radio-resistance of CC cells. Blocking this pathway may provide therapeutic benefits against CC radio-resistance.

miR-205 Suppresses Pulmonary Fibrosis by Targeting GATA3 Through Inhibition of Endoplasmic Reticulum Stress

2020 ◽  
Vol 21 (8) ◽  
pp. 720-726 ◽  
Author(s):  
Bingke Sun ◽  
Shumin Xu ◽  
Yanli Yan ◽  
Yusheng Li ◽  
Hongqiang Li ◽  
...  
Keyword(s):  
Pulmonary Fibrosis ◽  
Collagen I ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Reporter Assay ◽  
Sd Rats ◽  
Dual Luciferase Reporter Assay

Objective: To investigate the role of miR-205 and GATA3 in Pulmonary Fibrosis (PF). Methods: Bleomycin (BLM) was used to induce PF in SD rats and in vitro PF model was established by using TGFβ1-induced RLE-6TN cells. miR-205 mimics were used for the overexpression of miR- 205. The expression of miR-205, GATA3, α-SMA, Collagen I, CHOP and GRP78 were measured using RT-qPCR or western blotting. Dual-luciferase reporter assay was used to confirm binding between GATA3 3’-UTR and miR-205. Results: The expression of miR-205 was significantly down-regulated, while the expression of GATA3 was remarkably up-regulated in the model rats. GATA3 levels were remarkably decreased when miR-205 was overexpressed. When miR-205 was overexpressed, the lung injury by BLM-induced fibrosis was improved. The expression of α-SMA, Collagen I, as well as GRP78 and CHOP, was significantly up-regulated in both in vivo and in vitro PF models, and overexpression of miR-205 remarkably reversed the effects. Dual-luciferase reporter assay showed that miR-205 directly targeted and negatively regulated GATA3. Conclusion: miR-205 improved pulmonary fibrosis through inhibiting ER-stress by targeting GATA3.

scholarly journals MiR-425-5p accelerated the proliferation, migration, and invasion of ovarian cancer cells via targeting AFF4

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Zhihui Wu ◽  
Jianlin Guo ◽  
Ying Zhang ◽  
Jianhua Liu ◽  
Hongping Ma ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Luciferase Reporter Assay ◽  
Cell Counting ◽  
Luciferase Assay ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Ovarian Cancer Cells ◽  
Reporter Assay ◽  
Matrigel Invasion

Abstract Background Accumulating data have established that microRNAs (miRNAs) play significant regulatory roles in the carcinogenesis and progression of ovarian cancer (OC). MiR-425-5p was reported to function in various tumors. However, the roles and underlying mechanism of miR-425-5p involvement in OC development and progression are unclear. Methods A comprehensive strategy of data mining, computational biology, and real-time polymerase chain reaction was employed to identify the involvement of miR-425-5p in OC progression. The effect of miR-425-5p on the proliferation, migration, and invasion of OC cells was determined using Cell Counting Kit-8, wound-healing, and Matrigel invasion assays, respectively. Luciferase assay was performed to evaluate the interactions between miR-425-5p and MAGI2-AS3 or AFF4. Results miR-425-5p was significantly up-regulated in OC tissues and cells. The luciferase reporter assay revealed that miR-425-5p was negatively regulated by MAGI2-AS3. Silencing miR-425-5p inhibited the proliferation, migration, and invasion of OC cells in vitro. Bioinformatics analysis and luciferase reporter assay revealed that AFF4 was the target gene of miR-425-5p. Moreover, AFF4 expression was significantly decreased in OC and was closely related to the good prognosis of patients with OC. AFF4 overexpression inhibited the proliferation, migration, and invasion of OC cells in vitro. By contrast, silencing AFF4 promoted the proliferation, migration, and invasion of OC cells in vitro. Finally, AFF4 suppression rescued the inhibitory effect of silencing miR-425-5p on the proliferation, migration, and invasion of OC cells. Conclusion To the best our knowledge, this is the first study to demonstrate that miR-425-5p overexpression in OC is negatively regulated by MAGI2-AS3. Moreover, miR-425-5p promotes the proliferation, migration, and invasion of OC cells by targeting AFF4, suggesting that miR-425-5p/AFF4 signaling pathway represented a novel therapeutic target for patients with OC.

scholarly journals HOXA-AS2 Promotes Proliferation and Induces Epithelial-Mesenchymal Transition via the miR-520c-3p/GPC3 Axis in Hepatocellular Carcinoma

2018 ◽  
Vol 50 (6) ◽  
pp. 2124-2138 ◽  
Author(s):  
Ying Zhang ◽  
Jianliang Xu ◽  
Shaoquan Zhang ◽  
Jun An ◽  
Jin Zhang ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Proliferation ◽  
Cell Lines ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Reporter Assay ◽  
Online Programs ◽  
Hcc Cells

Background/Aims: Previous studies have demonstrated that long non-coding RNAs (lncRNAs) may play critical roles in cancer biology, including Hepatocellular carcinoma (HCC). The HOXA cluster antisense RNA2 (HOXA-AS2) lncRNA plays an important role in carcinogenesis, however, the underlying role of HOXA-AS2 in HCC remains unknown. The present study examined the effects of HOXA-AS2 on the progression of HCC, and explored the underlying molecular mechanisms. Methods: Quantitative real-time PCR was used to detect HOXA-AS2 expression in HCC tissues and cell lines. Furthermore, the effects of HOXA-AS2 silencing and overexpression on cell proliferation, cell cycle, apoptosis, migration, and invasion were assessed in HCC in vitro and in vivo. Furthermore, bioinformatics online programs predicted and luciferase reporter assay were used to validate the association of HOXA-AS2 and miR-520c-3p in HCC cells. Results: We observed that HOXA-AS2 was up-regulated in HCC tissues and cell lines. In vitro experiments revealed that HOXA-AS2 knockdown significantly inhibited HCC cells proliferation by causing G1 arrest and promoting apoptosis, whereas HOXA-AS2 overexpression promoted cell growth. Further functional assays indicated that HOXA-AS2 significantly promoted HCC cell migration and invasion by promoting EMT. Bioinformatics online programs predicted that HOXA-AS2 sponge miR-520c-3p at 3’-UTR with complementary binding sites, which was validated using luciferase reporter assay. HOXA-AS2 could negatively regulate the expression of miR-520c-3p in HCC cells. MiR-520c-3p was down-regulated and inversely correlated with HOXA-AS2 expression in HCC tissues. miR-520c-3p suppressed cell proliferation, invasion and migration in HCC cells, and enforced expression of miR-520c-3p attenuated the oncogenic effects of HOXA-AS2 in HCC cells. By bioinformatic analysis and dual-luciferase reporter assay, we found that miR-223-3p directly targeted the 3’-untranslated region (UTR) of Glypican-3 (GPC3), one of the key players in HCC. GPC3 was up-regulated in HCC tissues, and was negatively correlated with miR-520c-3p expression and positively correlated with HOXA-AS2 expression. Conclusion: In summary, our results suggested that the HOXA-AS2/miR-520c-3p/GPC3 axis may play an important role in the regulation of PTC progression, which could serve as a biomarker and therapeutic target for HCC.

scholarly journals Knockdown of lincRNA PADNA promotes bupivacaine-induced neurotoxicity by miR-194/FBXW7 axis

2020 ◽  
Author(s):  
Fan Yuning ◽  
Chen Liang ◽  
Wang Tenghuan ◽  
Nan Zhenhua ◽  
Shengkai Gong
Keyword(s):  
Functional Analysis ◽  
Western Blot ◽  
Cell Viability ◽  
Luciferase Reporter ◽  
Reporter Assay ◽  
Drg Neurons ◽  
New Evidence ◽  
Dual Luciferase Reporter Assay

Abstract Background: The aim of the study was to explore the function and mechanism of lincRNA PADNA in bupivacaine-induced neurotoxicity. Methods: Mouse DRG neurons were cultured in vitro and treated with bupivacaine to establish a neurotoxicity model. Caspase3 activity, cell viability, and TUNEL assays were analyzed to assess the role of lincRNA PADNA. A dual-luciferase reporter assay was used to determine the binding target of lincRNA PANDA. Results: The expression of lincRNA PADNA was significantly increased with increasing concentrations of bupivacaine. Functional analysis revealed that knockdown of lincRNA PADNA increased caspase3 activity and inhibited cell viability. Western blot analysis showed that knockdown of lincRNA PADNA promoted cleaved caspase3 levels. We also revealed that lincRNA PADNA may bind with miR-194. Knockdown of miR-194 rescued the function of lincRNA PADNA, suggesting that lincRNA PADNA may sponge miR-194. In addition, we provided new evidence that the lincRNA PADNA/miR-194/FBXW7 axis plays an important role in the neurotoxicity process. Conclusion: We performed comprehensive experiments to verify the function and mechanism of lincRNA PADNA in bupivacaine-induced neurotoxicity. Our study provides new evidence and clues for the prevention of neurotoxicity.

scholarly journals Hsa_circ_0005273 facilitates breast cancer tumorigenesis by regulating YAP1-hippo signaling pathway

2021 ◽  
Vol 40 (1) ◽  
Author(s):  
Xuehui Wang ◽  
Changle Ji ◽  
Jiashu Hu ◽  
Xiaochong Deng ◽  
Wenfang Zheng ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Signaling Pathway ◽  
Cell Lines ◽  
Luciferase Reporter ◽  
Circular Rnas ◽  
Hippo Signaling ◽  
Hippo Signaling Pathway ◽  
Potential Biomarker

Abstract Background Circular RNAs (circRNAs), a novel class of endogenous RNAs, have shown to participate in the development of breast cancer (BC). Hsa_circ_0005273 is a circRNA generated from several exons of PTK2. However, the potential functional role of hsa_circ_0005273 in BC remains largely unknown. Here we aim to evaluate the role of hsa_circ_0005273 in BC. Methods The expression level of hsa_circ_0005273 and miR-200a-3p were examined by RT-qPCR in BC tissues and cell lines. The effect of knocking down hsa_circ_0005273 in BC cell lines were evaluated by examinations of cell proliferation, migration and cell cycle. In addition, xenografts experiment in nude mice were performed to evaluate the effect of hsa_circ_0005273 in BC. RNA immunoprecipitation assay, RNA probe pull-down assay, luciferase reporter assay and fluorescence in situ hybridization were conducted to confirm the relationship between hsa_circ_0005273, miR-200a-3p and YAP1. Results Hsa_circ_0005273 is over-expressed in BC tissues and cell lines, whereas miR-200a-3p expression is repressed. Depletion of hsa_circ_0005273 inhibited the progression of BC cells in vitro and in vivo, while overexpression of hsa_circ_0005273 exhibited the opposite effect. Importantly, hsa_circ_0005273 upregulated YAP1 expression and inactivated Hippo pathway via sponging miR-200a-3p to promote BC progression. Conclusions Hsa_circ_0005273 regulates the miR-200a-3p/YAP1 axis and inactivates Hippo signaling pathway to promote BC progression, which may become a potential biomarker and therapeutic target.

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