scholarly journals Targeting ZFP64/GAL-1 axis promotes therapeutic effect of nab-paclitaxel and reverses immunosuppressive microenvironment in gastric cancer

2022 ◽  
Vol 41 (1) ◽  
Author(s):  
Mengxuan Zhu ◽  
Pengfei Zhang ◽  
Shan Yu ◽  
Cheng Tang ◽  
Yan Wang ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Zinc Finger Protein ◽  
Tumor Formation ◽  
Humanized Mice ◽  
Luciferase Reporter ◽  
Tunel Staining ◽  
Luciferase Reporter Gene ◽  
Paclitaxel Resistance ◽  
Gene Assay ◽  
Immunosuppressive Microenvironment

Abstract Background Chemoresistance is a main obstacle in gastric cancer (GC) treatment, but its molecular mechanism still needs to be elucidated. Here, we aim to reveal the underlying mechanisms of nanoparticle albumin-bound paclitaxel (nab-paclitaxel) resistance in GC. Methods We performed RNA sequencing (RNA-seq) on samples from patients who were resistant or sensitive to nab-paclitaxel, and identified Zinc Finger Protein 64 (ZFP64) as critical for nab-paclitaxel resistance in GC. CCK8, flow cytometry, TUNEL staining, sphere formation assays were performed to investigate the effects of ZFP64 in vitro, while subcutaneous tumor formation models were established in nude mice or humanized mice to evaluate the biological roles of ZFP64 in vivo. Chromatin immunoprecipitation sequencing (CHIP-seq) and double-luciferase reporter gene assay were conducted to reveal the underlying mechanism of ZFP64. Results ZFP64 overexpression was linked with aggressive phenotypes, nab-paclitaxel resistance and served as an independent prognostic factor in GC. As a transcription factor, ZFP64 directly binds to Galectin-1 (GAL-1) promoter and promoted GAL-1 transcription, thus inducing stem-cell like phenotypes and immunosuppressive microenvironment in GC. Importantly, compared to treatment with nab-paclitaxel alone, nab-paclitaxel plus GAL-1 blockade significantly enhanced the anti-tumor effect in mouse models, particularly in humanized mice. Conclusions Our data support a pivotal role for ZFP64 in GC progression by simultaneously promoting cellular chemotherapy resistance and tumor immunosuppression. Treatment with the combination of nab-paclitaxel and a GAL-1 inhibitor might benefit a subgroup of GC patients.

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.

microRNA-223 promotes autophagy to aggravate lung ischemia-reperfusion injury by inhibiting the expression of transcription factor HIF2α

2020 ◽  
Vol 319 (1) ◽  
pp. L1-L10
Author(s):  
Chunlin Ye ◽  
Wanghong Qi ◽  
Shaohua Dai ◽  
Guowen Zou ◽  
Weicheng Liu ◽  
...  
Keyword(s):  
Ischemia Reperfusion Injury ◽  
Tissue Injury ◽  
Cell Model ◽  
Ischemia Reperfusion ◽  
Pulmonary Ventilation ◽  
Pulmonary Arteries ◽  
Luciferase Reporter ◽  
Tunel Staining ◽  
Luciferase Reporter Gene ◽  
Gene Assay

Lung ischemia-reperfusion (I/R) injury severely endangers human health, and recent studies have suggested that certain microRNAs (miRNAs) play important roles in this pathological phenomenon. The current study aimed to ascertain the ability of miR-223 to influence lung I/R injury by targeting hypoxia-inducible factor-2α (HIF2α). First, mouse models of lung I/R injury were established: during surgical procedures, pulmonary arteries and veins and unilateral pulmonary portal vessels were blocked and resuming bilateral pulmonary ventilation, followed by restoration of bipulmonary ventilation. In addition, a lung I/R injury cell model was constructed by exposure to hypoxic reoxygenation (H/R) in mouse pulmonary microvascular endothelial cells (PMVECs). Expression of miR-223, HIF2α, and β-catenin in tissues or cells was determined by RT-qPCR and Western blot analysis. Correlation between miR-223 and HIF2α was analyzed by dual luciferase reporter gene assay. Furthermore, lung tissue injury and mouse PMVEC apoptosis was evaluated by hematoxylin and eosin (H&E), TUNEL staining, and flow cytometry. Autophagosomes in cells were detected by light chain 3 immunofluorescence assay. miR-223 was expressed at a high level while HIF2α/β-catenin was downregulated in tissues and cells with lung I/R injury. Furthermore, miR-223 targeted and repressed HIF2α expression to downregulate β-catenin expression. The miR-223/HIF2α/β-catenin axis aggravated H/R injury in mouse PMVECs and lung I/R injury in mice by enhancing autophagy. Taken together, miR-223 inhibits HIF2α to repress β-catenin, thus contributing to autophagy to complicate lung I/R injury. These findings provide a promising therapeutic target for treating lung I/R injury.

scholarly journals Contribution of microRNA-30d to the prevention of the thyroid cancer progression: mechanism and implications

2020 ◽  
Author(s):  
Yuan Shao ◽  
Shaoqiang Zhang ◽  
Xiaoxia Wang ◽  
Xin Sun ◽  
Jie Wu ◽  
...  
Keyword(s):  
Thyroid Cancer ◽  
Cancer Progression ◽  
Endocrine Tumor ◽  
Luciferase Reporter ◽  
Tunel Staining ◽  
Loss Of Function ◽  
Luciferase Reporter Gene ◽  
Gene Assay ◽  
Low Expression

Abstract Background Thyroid cancer is a major endocrine tumor and represents an emerging health problem worldwide. MicroRNAs (miRNAs) have been addressed to be associated with the pathogenesis and progression of thyroid cancer. However, it remains largely unknown what functions miR-30d may exert on thyroid cancer. This study herein aimed to identify the functional significance and mechanism of miR-30d in the progression of thyroid cancer. Methods The expression of miR-30d and ubiquitin-specific protease 22 (USP22) in cancerous tissues of patients with thyroid cancer was measured using RT-qPCR and Western blot analysis. In response to the gain- or loss-of-function of miR-30d and USP22, cell apoptosis was evaluated by flow cytometry and TUNEL staining in combination with the measurement of apoptosis-related proteins. The interactions among miR-30d, USP22, SIRT1, FOXO3a and PUMA were explored using a series of assays, including dual-luciferase reporter gene assay, Co-IP and ChIP assay. The effects of miR-30d and USP22 on thyroid tumorigenesis were finally validated in vivo. Results MiR-30b presented aberrant low expression in thyroid cancer tissues and this low expression correlated with poor prognosis of thyroid cancer patients. miR-30d promoted apoptosis of thyroid cancer cells through targeting USP22, an up-regulated gene in thyroid cancer. USP22 could enhance the stability of SIRT1 by inducing deubiquitination which consequently contributed to FOXO3a deacetylation-induced PUMA repression. It was verified that this regulatory mechanism was responsible for the pro-apoptotic effect of miR-30d by the in vivo tumorigenicity assay. Conclusion To conclude, the progression of thyroid cancer can be suppressed by miR-30d-mediated inhibition of USP22, provides a promising therapeutic target for thyroid cancer treatment.

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 Oxidative stress activates NORAD expression by H3K27ac and promotes oxaliplatin resistance in gastric cancer by enhancing autophagy flux via targeting the miR-433-3p

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Jizhao Wang ◽  
Yuchen Sun ◽  
Xing Zhang ◽  
Hui Cai ◽  
Cheng Zhang ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Gastric Cancer ◽  
Target Genes ◽  
Function Analysis ◽  
Resistance Index ◽  
Luciferase Reporter ◽  
Luciferase Reporter Gene ◽  
Autophagy Flux ◽  
Potential Biomarker ◽  
Gene Assay

AbstractOxaliplatin resistance undermines its curative effects on cancer and usually leads to local recurrence. The oxidative stress induced DNA damage repair response is an important mechanism for inducing oxaliplatin resistance by activating autophagy. ELISA is used to detect target genes expression. TMT-based quantitative proteomic analysis was used to investigate the potential mechanisms involved in NORAD interactions based on GO analysis. Transwell assays and apoptosis flow cytometry were used for biological function analysis. CCK-8 was used to calculate IC50 and resistance index (RI) values. Dual-luciferase reporter gene assay, RIP and ChIP assays, and RNA pull-down were used to detect the interaction. Autophagy flux was evaluated using electron microscope and western blotting. Oxidative stress was enhanced by oxaliplatin; and oxaliplatin resistance gastric cancer cell showed lower oxidative stress. TMT labeling showed that NORAD may regulate autophagy flux. NORAD was highly expressed in oxaliplatin-resistant tissues. In vitro experiments indicate that NORAD knockdown decreases the RI (Resistance Index). Oxaliplatin induces oxidative stress and upregulates the expression of NORAD. SGC-7901 shows enhanced oxidative stress than oxaliplatin-resistant cells (SGC-7901-R). NORAD, activated by H3K27ac and CREBBP, enhanced the autophagy flux in SGC-7901-R to suppress the oxidative stress. NORAD binds to miR-433-3p and thereby stabilize the ATG5- ATG12 complex. Our findings illustrate that NORAD, activated by the oxidative stress, can positively regulate ATG5 and ATG12 and enhance the autophagy flux by sponging miR-433-3p. NORAD may be a potential biomarker for predicting oxaliplatin resistance and mediating oxidative stress, and provides therapeutic targets for reversing oxaliplatin resistance.

circ-NOTCH1 acts as a sponge of miR-637 and affects the expression of its target gene Apelin to regulate gastric cancer cell growth

2020 ◽  
Vol 98 (2) ◽  
pp. 164-170 ◽  
Author(s):  
Encui Guan ◽  
Xiaoguang Xu ◽  
Fangxi Xue
Keyword(s):  
Gastric Cancer ◽  
Cell Proliferation ◽  
Target Gene ◽  
Luciferase Reporter ◽  
Circular Rnas ◽  
High Recurrence Rate ◽  
Cure Rate ◽  
Luciferase Reporter Gene ◽  
Cancer Cell Growth ◽  
Gene Assay

Gastric cancer (GC) is a major cause of cancer-related deaths worldwide, and has a low survival rate, low cure rate, high recurrence rate, and poor prognosis. Recent studies have indicated that circular RNAs (circRNAs) have important functions in the occurrence and progression of GC. Studies on circ-NOTCH1, which was shown to be highly expressed in GC, have indicated that miR-637 binds to circ-NOTCH1 at multiple sites, and a dual-luciferase reporter gene assay further confirmed that miR-637 indeed targeted circ-NOTCH1 and Apelin. Circ-NOTCH1 and Apelin are highly expressed in GC cells and tissues, whereas the expression of miR-637 is reduced. Circ-NOTCH1 and miR-637 do not regulate each other’s expression levels, but circ-NOTCH1significantly upregulates the expression of the miR-637 target gene Apelin, whereas miR-637 inhibites the expression of Apelin. Examination of GC cells showed that circ-NOTCH1 enhances cell proliferation and invasiveness, and reduces cell apoptosis; these effects were reversed by miR-637, which could terminate the above effects of circ-NOTCH1. When co-transfected with the circ-NOTCH1 overexpression plasmid and Apelin siRNAs, there were no obvious changes to the levels of cell proliferation, apoptosis, or invasiveness. Therefore, in GC cells, circ-NOTCH1 inhibits the transcriptional activity of miR-637, thereby upregulating the expression of its target gene Apelin and regulating cell proliferation, apoptosis, and invasiveness. This finding provides more experimental evidence for the function of circRNA in GC.

scholarly journals RASA1 inhibits the progression of renal cell carcinoma by decreasing the expression of miR-223-3p and promoting the expression of FBXW7

2020 ◽  
Vol 40 (7) ◽  
Author(s):  
Rui-Li Zhang ◽  
Ainiwaer Aimudula ◽  
Jiang-Hong Dai ◽  
Yong-Xing Bao
Keyword(s):  
Renal Cell Carcinoma ◽  
Cell Proliferation ◽  
Cell Carcinoma ◽  
Renal Cell ◽  
Tumor Formation ◽  
Luciferase Reporter ◽  
Luciferase Reporter Gene ◽  
Gene Assay ◽  
The Impact

Abstract RAS p21 protein activator 1 (RASA1), also known as p120-RasGAP, is a RasGAP protein that functions as a signaling scaffold protein, regulating pivotal signal cascades. However, its biological mechanism in renal cell carcinoma (RCC) remains unknown. In the present study, RASA1, F-box/WD repeat-containing protein 7 (FBXW7), and miR-223-3p expression were assessed via quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and Western blot. Then, the targeted correlations of miR-223-3p with FBXW7 and RASA1 were verified via a dual-luciferase reporter gene assay. CCK-8, flow cytometry, and Transwell assays were implemented independently to explore the impact of RASA1 on cell proliferation, apoptosis, migration, and cell cycle progression. Finally, the influence of RASA1 on tumor formation in RCC was assessed in vivo through the analysis of tumor growth in nude mice. Results showed that FBXW7 and RASA1 expression were decreased in RCC tissues and cell lines, while miR-223-3p was expressed at a higher level. Additionally, FBXW7 and RASA1 inhibited cell proliferation but facilitated the population of RCC cells in the G0/G1 phase. Altogether, RASA1 may play a key role in the progression of RCC by decreasing miR-223-3p and subsequently increasing FBXW7 expression.

miR-635 targets KIFC1 to inhibit the progression of gastric cancer

2020 ◽  
Vol 68 (8) ◽  
pp. 1357-1363
Author(s):  
Feng-Yu Cao ◽  
Yong-Bin Zheng ◽  
Chao Yang ◽  
Su-Yang Huang ◽  
Xiao-Bo He ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Luciferase Reporter Gene ◽  
Normal Tissues ◽  
Kaplan Meier ◽  
Gene Assay ◽  
Kaplan Meier Plotter

Accumulating studies have shown that the dysregulation of microRNAs is related to the carcinogenesis and development of gastric cancer (GC), and the role of miR-635 in GC remains largely unknown. miR-635 and Kinesin Family Member C1 (KIFC1) mRNA expression in GC tissues and paracancerous tissues and cells were detected by quantitative real-time PCR. KIFC1 protein expression in GC tissues and paracancerous normal tissues and cells was detected by immunohistochemistry and western blot. Cell proliferation was monitored by Cell Counting Kit-8 assay and 5-bromo-2′-deoxyuridine assay. Transwell assay was employed to detect the migration and invasion of GC cells. The dual-luciferase reporter gene assay was adopted to detect the targeting relationship between miR-635 and KIFC1. Compared with paracancerous tissues, miR-635 expression was remarkably decreased in GC tissues; conversely, KIFC1 expression was significantly increased. Compared with human normal gastric epithelial cell GSE-1, miR-635 expression was markedly decreased in GC cell lines. Meanwhile, KIFC1 expression was significantly increased, and the Kaplan-Meier Plotter database showed that its high expression was remarkably associated with poor prognosis. Additionally, miR-635 can negatively regulate KIFC1. miR-635 can target KIFC1 to inhibit proliferation, migration and invasion of GC cells. Collectively, miR-635 is lowly expressed in GC, and it inhibits proliferation, migration and invasion of GC cells via regulating KIFC1.

HOTAIR Competitively Binds MiRNA330 as a Molecular Sponge to Increase the Resistance of Gastric Cancer to Trastuzumab

2020 ◽  
Vol 20 (9) ◽  
pp. 700-709 ◽  
Author(s):  
Liangyu Bie ◽  
Suxia Luo ◽  
Dan Li ◽  
Yan Wei ◽  
Yu Mu ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Cancer Cells ◽  
Lentiviral Vector ◽  
Luciferase Reporter ◽  
Tumor Proliferation ◽  
Trastuzumab Resistance ◽  
Luciferase Reporter Gene ◽  
Gastric Cancer Cells ◽  
Strong Expression ◽  
Gene Assay

Background: HOTAIR, one of the most widely studied long non-coding RNAs in tumors, is closely related to tumor proliferation, migration, invasion and chemoresistance. Objective: Here, we studied the mechanism behind proliferation and chemoresistance processes. Methods: A total of 75 samples were collected from patients who underwent surgical resection of their gastric cancer and received trastuzumab treatment. Primary cells were isolated and cultured. We also developed a cell line overexpressing HOTAIR by constructing a lentiviral vector. These cell lines were studied using an array of established biomolecular methods. Results: We found that HOTAIR levels were inversely associated with sensitivity to trastuzumab in gastric cancer and that overexpression of HOTAIR can promote the proliferation and invasion of gastric cancer cells. The sensitivity of cells overexpressing HOTAIR to two different types of human epidermal growth factor receptor 2 (HER2) inhibitors (trastuzumab and afatinib) showed that overexpression of HOTAIR is specific for trastuzumab resistance. Furthermore, luciferase reporter gene assay and western blot assay showed that there is a HOTAIR-miRNA330-ERBB4 competitive endogenous RNA regulatory network with miRNA330 as the core. Conclusion: HOTAIR can not only promote tumor proliferation but also enhance the resistance of tumor cells to drugs. Our experimental data not only showed strong expression of HOTAIR in gastric cancer, but also that strong expression of HOTAIR caused the sensitivity of gastric cancer cells to trastuzumab, which is a useful reference for postoperative medication.

scholarly journals Tumor Inhibitory Effect of Long Non-coding RNA LOC100505817 on Gastric Cancer

2021 ◽  
Vol 27 ◽  
Author(s):  
Lei Zheng ◽  
Liying Kang ◽  
Yan Cheng ◽  
Junli Cao ◽  
Lijie Liu ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Cell Proliferation ◽  
Noncoding Rna ◽  
Rna Binding ◽  
Ectopic Expression ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Luciferase Reporter Gene ◽  
Gene Assay

Gastric cancer (GC) is one of the major malignancies worldwide. Emerging evidence has revealed the potential involvement of long noncoding RNA (lncRNA) in human genetic disorders and cancer, but the role of LOC100505817 remains unknown. Thus, in this study, we isolated tissues from GC patients to characterize the functional importance of LOC100505817 in GC tumorigenesis. We also proposed a hypothesis that the regulation of Wnt/β-catenin pathway by LOC100505817 was regulated by miR-20a-mediated WT1. After the collection of cancer tissues and adjacent tissues were obtained from GC patients, expression of LOC100505817, Wnt/β-catenin pathway- and EMT-related genes was quantified. Ectopic expression and knockdown experiments were applied in order to investigate the protective role of LOC100505817 in the progression of GC. Subsequently, cell viability, flow cytometry for apoptosis and cell cycle were detected via CCK-8, while migration and invasion were determined using scratch test and Transwell assay respectively. Then interactions among LOC100505817, miR-20a and WT1 were explored by dual luciferase reporter gene assay, RNA pull down assay and RNA binding protein immunoprecipitation (RIP) assay. The results found poor expression LOC100505817 was poorly expressed in GC cells and tissues. Overexpressed LOC100505817 resulted in the significant reduction of cell proliferation, migration and invasion as well as the expression of Wnt2b, β-catenin, CyclinD1, N-cadherin, Vimentin and snail, while increased cell apoptosis along with the expression of E-cadherin. Wnt/β-catenin pathway and EMT in GC cells were suppressed by LOC100505817 through miR-20a-inhibted WT1. In summary, our results provided evidence suggesting that LOC100505817 inhibits GC through LOC100505817-mediated inhibition of Wnt/β-catenin pathway, that leads to the overall restraining of GC cell proliferation, migration and invasion through miR-20a-reduced WT1.

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