scholarly journals MicroRNA-340 is involved in ultraviolet B-induced pigmentation by regulating the MITF/TYRP1 axis

2020 ◽  
Vol 48 (11) ◽  
pp. 030006052097151
Author(s):  
Yi Yang ◽  
Xuanjin Wei ◽  
Jia Bai ◽  
Min Huang ◽  
Tian Hao ◽  
...  
Keyword(s):  
Ultraviolet B ◽  
Luciferase Reporter ◽  
Melanin Content ◽  
Immunoblotting Analysis ◽  
Pigment Synthesis ◽  
Uvb Irradiation ◽  
Real Time Quantitative Pcr ◽  
Qrt Pcr ◽  
Reporter Assays ◽  
Mitf Expression

Objective There is growing evidence that ultraviolet B (UVB) irradiation can change the expression profile of microRNAs (miRNAs) in immortalized human epidermal melanocytes (Pig-1). We aimed to investigate the effect of miR-340 on regulating UVB-induced pigmentation. Methods Real-time quantitative PCR (qRT-PCR) was used to evaluate the expression of miR-340 in Pig-1 cells. Immunoblotting analysis, qRT-PCR, and luciferase reporter assays were used to detect the potential target of miR-340. The sodium hydroxide dissolution assay was used to assess the effect of miR-340 on changes in melanin content. Results Expression of miR-340 was reduced in human Pig-1 cells after UVB irradiation. We found a negative correlation between miR-340 and melanocyte inducing transcription factor (MITF) in Pig-1 cells after UVB irradiation. Knockdown and overexpression of MITF in Pig-1 cells down- and upregulated melanogenesis, respectively. Overexpression of miR-340 inhibited MITF expression, reduced the amount of melanin, and suppressed expression of multiple key molecules involved in the pigment synthesis pathway, whereas knockdown of miR-340 showed the opposite results. Conclusions Our results showed that miR-340 inhibited melanogenesis by regulating the downstream molecules of MITF and its signaling pathways, suggested that miRNA-340 may be a new target for the clinical treatment of UVB-induced pigmentation.

scholarly journals Chorionic Villous Mesenchymal Stem Cells Derived Exosomes Deliver miR-135b-5p to Trophoblasts, Promoting their Proliferation and Invasion by Targeting TXNIP via β-Catenin Pathway

2020 ◽  
Author(s):  
Yijing Chu ◽  
Yan Zhang ◽  
Guoqiang Gao ◽  
Jun Zhou ◽  
Yang Lv ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Tissue Injury ◽  
Luciferase Reporter ◽  
Rna Seq ◽  
Qrt Pcr ◽  
Reporter Assays ◽  
Pcr Assays ◽  
Proliferation And Invasion

Abstract Background: Human chorionic villous mesenchymal stem cells (CV-MSCs) are found to be a promising and effective treatment for tissue injury. Trophoblast dysfunction during pregnancies is significantly involved in the pathogenesis of preeclampsia (PE). This work was to understand how CV-MSCs regulated trophoblast function. Methods: In this study, we treated trophoblasts with CV-MSC-derived exosomes and RNA-seq analysis was used to understand the changes in trophoblasts. We examined the levels of TXNIP and β-catenin in trophoblasts by immunohistochemistry, western blot and qRT-PCR assays. Luciferase reporter assays and qRT-PCR assays were used to understand the role of miR135b-5p in the effects of CV-MSC-derived exosomes. The growth and invasion of trophoblasts was evaluated with the CCK-8 and transwell assays. Results: The treatment markedly enhanced the trophoblast proliferation and invasion. Furthermore, a significant decrease of TXNIP expression and inactivation of the β-catenin pathway in CV-MSCs exosomes-treated trophoblasts was observed. Consistent with these findings, TXNIP inhibition exhibited the same effect of promoting trophoblast proliferation and invasion as induced by CV-MSC-derived exosomes, also with the accompaniment of inactivation of β-catenin pathway. In addition, overexpression of TXNIP activated the β-catenin pathway in trophoblasts, and reduced the proliferation and invasion of trophoblasts. Importantly, miR135b-5p was found to be highly expressed in CV-MSC exosomes and interact with TXNIP. The miR-135b-5p overexpression significantly elevated the proliferation and invasion of trophoblasts, which could be attenuated by TXNIP overexpression. Conclusion: Our results suggest that TXNIP-dependent β-catenin pathway inactivation mediated by miR135b-5p which is delivered by CV-MSC-derived exosomes could promote the proliferation and invasion of trophoblasts.

Long noncoding RNA XIST regulates the EGF receptor to promote TGF-β1-induced epithelial–mesenchymal transition in pancreatic cancer

2020 ◽  
Vol 98 (2) ◽  
pp. 267-276 ◽  
Author(s):  
Lei Zou ◽  
Feng-Rong Chen ◽  
Ren-Pin Xia ◽  
Hua-Wei Wang ◽  
Zhen-Rong Xie ◽  
...  
Keyword(s):  
Pancreatic Cancer ◽  
Cell Proliferation ◽  
Growth Factor ◽  
Epithelial Mesenchymal Transition ◽  
Luciferase Reporter ◽  
Mesenchymal Transition ◽  
Clinical Specimens ◽  
Qrt Pcr ◽  
Reporter Assays ◽  
Tgf Β1

Background: This study focuses on the lncRNA XIST (X inactive-specific transcript), an lncRNA involved in multiple human cancers, and investigates the functional significance of XIST and the molecular mechanisms underlying the epithelial–mesenchymal transition (EMT) in pancreatic cancer (PC). Methods: Clinical specimens from 25 patients as well as 5 human PC cell lines were analyzed for XIST, YAP, and microRNA(miR)-34a by quantitative real-time PCR (qRT-PCR) and immunohistochemistry. To investigate how XIST influences cell proliferation, invasiveness, and apoptosis in PC, we performed the CCK-8 assays, Transwell assays, and flow cytometry. Luciferase reporter assays, qRT-PCR, and Western blot were applied to prove that miR-34a directly binds to XIST. Results: Up-regulation of XIST and Yes associated protein (YAP) and down-regulation of miR-34a were consistently observed in the clinical specimens and PC cell lines. Silencing XIST reduced the expression of YAP and suppressed transforming growth factor (TGF)-β1-induced EMT, while over-expression of XIST increased the expression of YAP and promoted EMT. In addition, inhibition of epidermal growth factor receptor (EGFR) hampered the XIST-promoted EMT. The results from the luciferase reporter assays confirmed that miR-34a directly targets XIST and suggested that XIST regulates cell proliferation, invasiveness, and apoptosis in PC by sponging miR-34a. Conclusions: XIST promotes TGF-β1-induced EMT by regulating the miR-34a–YAP–EGFR axis in PC.

scholarly journals Long noncoding RNA HOTAIR mediates the estrogen-induced metastasis of endometrial cancer cells via the miR-646/NPM1 axis

2018 ◽  
Vol 314 (6) ◽  
pp. C690-C701 ◽  
Author(s):  
Yun-xiao Zhou ◽  
Chuan Wang ◽  
Li-wei Mao ◽  
Yan-li Wang ◽  
Li-qun Xia ◽  
...  
Keyword(s):  
Noncoding Rna ◽  
Molecular Mechanisms ◽  
Rna Binding ◽  
Rna Binding Protein ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Qrt Pcr ◽  
Reporter Assays ◽  
Ec Cells

LncRNA homeobox (HOX) transcript antisense intergenic RNA (HOTAIR) has been confirmed to be involved in the tumorigenic progression of endometrial carcinoma (EC). However, the molecular mechanisms of HOTAIR in EC are not fully elucidated. The expression of HOTAIR and miR-646 in human EC tissues was determined by qRT-PCR. The effect of miR-646 on EC cells was assessed by the cell viability, migration, and invasion using CCK-8 assays and transwell assays. RNA-binding protein immunoprecipitation assays and RNA pull-down assays were performed to explore the interaction between HOTAIR and miR-646. The regulation of miR-646 on nucleophosmin 1 (NPM1) was tested using luciferase reporter assays. MiR-646 expression was significantly decreased both in human EC tissues ( n = 23) and cell lines (Ishikawa and HEC-1-A) compared with the control. Moreover, miR-646 expression was negatively related to HOTAIR in human EC tissues ( n = 23). Our results also showed that miR-646 overexpression considerably attenuated the E2-promoted viability, migration, and invasion of Ishikawa and HEC-1-A cells in vitro. In addition, HOTAIR was confirmed to regulate the viability, migration, and invasion of EC cells through negative regulating miR-646. More importantly, we also demonstrated that NPM1 was the target of miR-646, and HOTAIR promoted NPM1 expression through interacting with miR-646 in EC cells. Taken together, our findings presented that HOTAIR could regulate NPM1 via interacting with miR-646, thereby governing the viability, migration, and invasion of EC cells.

scholarly journals The Mechanisms of Colorectal Cancer Cell mesenchymal-epithelial Transition Induced by Hepatocyte Exosome-derived miR-203a-3p

2021 ◽  
Author(s):  
Heyang Xu ◽  
Qiusheng Lan ◽  
Yongliang Huang ◽  
Yang Zhang ◽  
Yujie Zeng ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Liver Metastasis ◽  
Epithelial To Mesenchymal Transition ◽  
Luciferase Reporter ◽  
Mesenchymal Transition ◽  
Real Time Quantitative Pcr ◽  
Mesenchymal To Epithelial Transition ◽  
Reporter Assays ◽  
Phosphatase Of Regenerating Liver

Abstract Background: Liver metastasis is the most common cause of death in patients with colorectal cancer (CRC). Phosphatase of regenerating liver-3 induces CRC metastasis by epithelial-to-mesenchymal transition, which promotes CRC cell liver metastasis. Mesenchymal-to-epithelial transition (MET), the opposite of epithelial-to-mesenchymal transition, has been proposed as a mechanism for the establishment of metastatic neoplasms. However, the molecular mechanism of MET remains unclear. Methods: Using Immunohistochemistry, western blotting，invasion assays, real-time quantitative PCR, chromatin immunoprecipitation, luciferase reporter assays, human miRNA arrays, and xenograft mouse model, we determined the role of hepatocyte exosome-derived miR-203a-3p in CRC MET.Results: In our study, we found that miR-203a-3p derived from hepatocyte exosomes increased colorectal cancer cells E-cadherin expression, inhibited Src expression, and reduced activity. In this way miR-203a-3p induced the decreased invasion rate of CRC cells.Coclusion: MiR-203a-3p derived from hepatocyte exosomes plays an important role of CRC cells to colonize in liver.

scholarly journals LncRNA LINC01303 Promotes the Progression of Oral Squamous Cell Carcinomas via the miR-429/ZEB1/EMT Axis

2021 ◽  
Vol 2021 ◽  
pp. 1-15
Author(s):  
Bo Sun ◽  
Xianyu Zheng ◽  
Weilong Ye ◽  
Pengcheng Zhao ◽  
Guowu Ma
Keyword(s):  
Squamous Cell ◽  
Luciferase Reporter ◽  
Real Time Quantitative Pcr ◽  
Qrt Pcr ◽  
Cytoplasmic Rna ◽  
Transwell Invasion Assay ◽  
Dual Luciferase Reporter Assay

Objectives. The aim of this research was to uncover the biological role and mechanisms of LINC01303 in oral squamous cell carcinoma (OSCC). Materials and Methods. Real-time quantitative PCR (qRT-PCR) was used to determine LINC01303 expression in OSCC tissues. Subcellular distribution of LINC01303 was examined by nuclear/cytoplasmic RNA fractionation and FISH experiments. The role of LINC01303 in the growth of TSCCA and SCC-25 was examined by CCK-8 assay, colony formation, transwell invasion assay in vitro, and xenograft tumor experiment in vivo. Dual-luciferase reporter assay was used to verify the interaction between LINC01303 and miR-429. RNA pull‐down assay was used to discover miR-429‐interacted protein, which was further examined by qRT-PCR, western blot, and rescue experiments. Results. LINC01303 expression was higher in OSCC tissues compared with adjacent nontumor tissues. LINC01303 was found to be localized in the cytoplasm of OSCC cells. Knockdown of LINC01303 inhibited OSCC cell proliferation and invasion, whereas increasing the expression of LINC01303 showed the opposite effects. Furthermore, LINC01303 served as a miR-429 “sponge” and positively regulated ZEB1 expression. Moreover, LINC01303 promoted OSCC through miR-429/ZEB1 axis both in vivo and in vitro. Conclusions. LINC01303 plays an oncogenic role in OSCC and is a promising biomarker for OSCC patients.

scholarly journals LncRNA SNHG6 promotes chemoresistance through ULK1-induced autophagy by sponging miR-26a-5p in colorectal cancer cells

2019 ◽  
Vol 19 (1) ◽  
Author(s):  
Xinke Wang ◽  
Zhixian Lan ◽  
Juan He ◽  
Qiuhua Lai ◽  
Xiang Yao ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Western Blotting ◽  
Target Genes ◽  
Bioinformatics Analysis ◽  
Chemotherapy Resistance ◽  
Micro Rna ◽  
Luciferase Reporter ◽  
Qrt Pcr ◽  
Reporter Assays

Abstract Background Chemotherapy resistance is one of the main causes of recurrence in colorectal cancer (CRC) patients and leads to poor prognosis. Long noncoding RNAs (lncRNAs) have been reported to regulate chemoresistance. We aimed to determine the role of the lncRNA small nucleolar RNA host gene 6 (SNHG6) in CRC cell chemoresistance. Methods Cell drug sensitivity tests and flow cytometry were performed to analyze CRC cell chemoresistance. Animal models were used to determine chemoresistance in vivo, and micro RNA (miRNA) binding sites were detected by dual-luciferase reporter assays. Bioinformatics analysis was performed to predict miRNAs binding to SNHG6 and target genes of miR-26a-5p. SNHG6/miR-26a-5p/ULK1 axis and autophagy-related proteins were detected by qRT-PCR and western blotting. Furthermore, immunofluorescence was employed to confirm the presence of autophagosomes. Results SNHG6 enhanced CRC cell resistance to 5-fluorouracil (5-FU), promoted autophagy, inhibited 5-FU-induced apoptosis, and increased 5-FU resistance in vivo. Bioinformatics analysis showed that miR-26a-5p might bind to SNHG6 and target ULK1, and dual-luciferase reporter assays confirmed this activity. qRT-PCR and western blotting showed that SNHG6 was able to negatively regulate miR-26a-5p but correlated positively with ULK1. Conclusion SNHG6 may promote chemoresistance through ULK1-induced autophagy by sponging miR-26a-5p in CRC cells.

Molecular Mechanism of circRAPGEF5 in Regulating the Molecular Axis of microRNA-4712-5p/Tyrosine 3-Monooxygenase/Tryptophan 5-Monooxygenase Activation Protein Epsilon Affecting Cellular Proliferation and Apoptosis in Mammary Cancer

2021 ◽  
Vol 11 (6) ◽  
pp. 1053-1058
Author(s):  
Tao Chen ◽  
Shengrong Sun
Keyword(s):  
Western Blot ◽  
Cancer Cells ◽  
Mammary Cancer ◽  
Caspase 3 ◽  
Cellular Proliferation ◽  
Molecular Axis ◽  
Luciferase Reporter ◽  
Qrt Pcr ◽  
Proliferation And Apoptosis ◽  
Reporter Assays

To understand the molecular mechanism of circRAPGEF5, its effect on the proliferation and apoptosis of mammary cancer cells, and its regulatory effect on the molecular axis of miRNA-4712-5p/YWHAE. qRT-PCR and Western blot were used to test circRAPGEF5, miRNA-4712-5p, and YWHAE expression in mammary cancer and paracancerous tissues. The human mammary cancer cell, MDA-MB-231, was cultured in vitro, and pcDNA-NC, pcDNA-circRAPGEF5, anti-miRNA-NC, anti-miRNA-4712-5p, pcDNA-circRAPGEF5, and miRNA-NC, pcDNA-circRAPGEF5 were transfected into MDA-MB-231 cells with miRNA-4712-5p mimics. qRT-PCR and Western blot were employed to detect circRAPGEF5, miRNA-4712-5p, and YWHAE expression in cells. The CCK-8 methodand plate clone formation experiment were conducted to test cellular proliferation ability. Flow cytometry was performed to detect apoptosis rate. Dual luciferase reporter assays were used to test the targeting association between circRAPGEF5 and miRNA-4712-5p, and the targeting association between miRNA-4712-5p and YWHAE. Western blot was utilized to detect Bcl-2, Bax, and Cleared Caspase-3 protein expression. In comparison with paracancerous tissues, circRAPGEF5 and YWHAE expression levels in mammary cancer tissues were significantly reduced (P < 0.05), and miRNA-4712-5p expression levels were significantly increased (P < 0.05). Transfection of pcDNA-circRAPGEF5 or trans-anti-miRNA-4712-5p could reduce the optical density (OD) value, Bcl-2 protein level and clonal formation number to a significant extent (P < 0.05), and it increases Bax and Cleaved Caspase-3 apoptosis rate and protein levels (P < 0.05). Dual luciferase reporter assays confirmed that there was target binding between circRAPGEF5 and miRNA-4712-5p and between miRNA-4712-5p and YWHAE. Co-transfection of pcDNA-circRAP GEF5 and miRNA-4712-5p could greatly reduce transfection of pcDNA-circRAP GEF5 and its effect on the proliferation and apoptosis of MDA-MB-231 cells. Overexpression of circRAPGEF5 can inhibit the proliferation of mammary cancer cells and induce apoptosis by regulating the molecular axis of miRNA-4712-5p/YWHAE.

scholarly journals Role of MiR-98 and Its Underlying Mechanisms in Systemic Lupus Erythematosus

2018 ◽  
Vol 45 (10) ◽  
pp. 1397-1405 ◽  
Author(s):  
Lin Xie ◽  
Jinhua Xu
Keyword(s):  
T Cells ◽  
Lupus Erythematosus ◽  
Cd4 T Cells ◽  
Opposite Effect ◽  
Luciferase Reporter ◽  
Qrt Pcr ◽  
Fas Mrna ◽  
Reporter Assays ◽  
Underlying Mechanisms

Objective.T-lymphocyte apoptosis plays a critical role in the pathogenesis of systemic lupus erythematosus (SLE). However, the underlying regulatory mechanisms of apoptosis in SLE remain unclear. The aim of this study was to explore the role of miR-98 in SLE and its underlying mechanisms.Methods.Western blotting and quantitative reverse transcription PCR (qRT-PCR) were used to analyze miR-98 and Fas expression. Luciferase reporter assays were performed to identify miR-98 targets. To modify miRNA levels, miR-98 mimics and inhibitor were transfected into cells. A lentiviral construct was used to overexpress the level of Fas in SLE CD4+ T cells. Gene and protein expression were determined by qRT-PCR and Western blotting. Apoptosis levels were evaluated by annexin V staining and flow cytometry.Results.Compared to those of healthy donors, miR-98 was downregulated in SLE CD4+ T cells, whereas Fas mRNA and protein expression were upregulated. Upregulation of miR-98 by mimic transfection protected Jurkat cells against Fas-mediated apoptosis at both mRNA and protein levels, while miR-98 inhibitor induced the completely opposite effect. Luciferase reporter assays demonstrated that miR-98 directly targeted Fas mRNA. Further, miR-98 inhibitor induced apoptosis in primary healthy CD4+ T cells through the Fas-caspase axis, while upregulation of miR-98 in SLE CD4+ T cells led to the opposite effect.Conclusion.The current study revealed that downregulation of miR-98 induces apoptosis by modulating the Fas-mediated apoptotic signaling pathway in SLE CD4+ T cells. These results suggest that miR-98 might serve as a potential target for SLE treatment.

scholarly journals MicroRNA-205 affects mouse granulosa cell apoptosis and estradiol synthesis by targeting CREB1

2018 ◽  
Author(s):  
Pengju Zhang ◽  
Jun Wang ◽  
Hongyan Lang ◽  
Weixia Wang ◽  
Xiaohui Liu ◽  
...  
Keyword(s):  
Caspase 3 ◽  
Follicular Development ◽  
Luciferase Reporter ◽  
Follicular Atresia ◽  
Qrt Pcr ◽  
Reporter Assays ◽  
Cyclic Amp Response Element ◽  
Cytochrome P450 Family ◽  
Estradiol Synthesis

ABSTRACTMicroRNAs-205 (miR-205), were reportedly to be involved in various physiological and pathological processes, but its biological function in follicular atresia remain unknown. In this study, we investigated the expression of miR-205 in mouse granulosa cells (mGCs), and explored its functions in primary mGCs using a serial of in vitro experiments. The result of qRT-PCR demonstrated that miR-205 expression was significantly increased in early atretic follicles (EAF), and progressively atretic follicles (PAF) compared to healthy follicles (HF). Our results also revealed that overexpression of miR-205 in mGCs significantly promoted apoptosis, caspas-3/9 activities, and inhibited estrogen E2 release, and cytochrome P450 family 19 subfamily A polypeptide 1 (CYP19A1, a key gene in E2 production) expression. Bioinformatics and luciferase reporter assays revealed that the gene of cyclic AMP response element (CRE)-binding protein 1 (CREB1) was a potential target of miR-205. qRT-PCR and western blot assays revealed that overexpression of miR-205 inhibited the expression of CREB1 in mGCs. Importantly, CREB1 upregulation partially rescued the effects of miR-205 on apoptosis, caspase-3/9 activities, E2 production and CYP19A1 expression in mGCs. Our results indicate that miR-205 may play an important role in ovarian follicular development and provide new insights into follicular atresia.

scholarly journals LncRNA OTUD6B-AS1 Induces Cisplatin Resistance in Cervical Cancer Cells Through Up-Regulating Cyclin D2 via miR-206

2021 ◽  
Vol 11 ◽  
Author(s):  
Hui Hou ◽  
Rong Yu ◽  
Haiping Zhao ◽  
Hao Yang ◽  
Yuchong Hu ◽  
...  
Keyword(s):  
Cervical Cancer ◽  
Cancer Cells ◽  
Cisplatin Resistance ◽  
Luciferase Reporter ◽  
Cyclin D2 ◽  
Aberrant Expression ◽  
Qrt Pcr ◽  
Cervical Cancer Cells ◽  
Reporter Assays ◽  
Underlying Mechanisms

Cervical cancer is one of the most common gynecological cancers. Cisplatin resistance remains a major hurdle in the successful treatment of cervical cancer. Aberrant expression of long non-coding RNAs (lncRNAs) and microRNAs (miRNAs) are implicated in cisplatin resistance. However, the regulatory functions of lncRNAs and miRNAs in cervical cancer cisplatin resistance and the underlying mechanisms are still elusive. Our qRT-PCR assays verified that miR-206 levels were down-regulated in cisplatin-resistant cervical cancer cells. The introduction of miR-206 sensitized cisplatin-resistant cervical cancer cells to cisplatin. Our qRT-PCR and luciferase reporter assays showed that Cyclin D2 (CCND2) was the direct target for miR-206 in cervical cancer cells. The cisplatin-resistant cervical cancer cells expressed higher CCND2 expression than the parental cells, whereas inhibition of CCND2 could sensitize the resistant cells to cisplatin treatment. Furthermore, we demonstrated that lncRNA OTUD6B-AS1 was up-regulated in cisplatin-resistant cervical cancer cells, and knocking down OTUD6B-AS1 expression induced re-acquirement of chemosensitivity to cisplatin in cervical cancer cells. We also showed that OTUD6B-AS1 up-regulated the expression of CCND2 by sponging miR-206. Low miR-206 and high OTUD6B-AS1 expression were associated with significantly poorer overall survival. Taken together, these results suggest that OTUD6B-AS1-mediated down-regulation of miR-206 increases CCND2 expression, leading to cisplatin resistance. Modulation of these molecules may be a therapeutic approach for cisplatin-resistant cervical cancer.

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