Methotrexate Alters the Expression of microRNA in Fibroblast-like Synovial Cells in Rheumatoid Arthritis.

We aimed to investigate the effect of methotrexate (MTX) on microRNA modulation in rheumatoid arthritis fibroblast-like synovial cells (RA-FLS). RA-FLS were treated with MTX for 48 h. We then performed miRNA array analysis to investigate differentially expressed miRNAs. Transfection with miR-877-3p precursor and inhibitor were used to investigate the functional role of miR-877-3p in RA-FLS. Gene ontology analysis was used to investigate the cellular processes involving miR-877-3p. The production of cytokines/chemokines was screened by multiplex cytokine/chemokine bead assay and confirmed by ELISA and quantitative real-time PCR. The migratory and proliferative activities of RA-FLS were analyzed by wound healing assay and MKI-67 expression. MTX treatment altered the expression of 13 miRNAs (seven were upregulated and six were downregulated). Among them, quantitative real-time PCR confirmed that miR-877-3p was upregulated in response to MTX (1.79 ± 0.46-fold, p < 0.05). The possible target genes of miR-877-3p in RA-FLS revealed by the microarray analysis were correlated with biological processes. The overexpression of miR-877-3p decreased the production of GM-CSF and CCL3, and the overexpression of miR-877-3p inhibited migratory and proliferative activity. MTX altered the miR-877-3p expression on RA-FLS, and this alteration of miR-877-3p attenuated the abundant production of cytokines/chemokines and proliferative property of RA-FLS.

HOXD-AS1 facilitates cell migration and invasion as an oncogenic lncRNA by competitively binding to miR-877-3p and upregulating FGF2 in human cervical cancer.

BACKGROUND: Long non-coding RNAs (LncRNAs) are dysregulated in multiple human cancers and they are highly involved in tumor progression. Previous studies have identified the oncogenic lncRNA HOXD cluster antisense RNA 1 (HOXD-AS1) in human cancers, while its roles in cervical cancer (CC) remain unclear. Herein we intended to characterize the implication of HOXD-AS1 in CC. METHODS: qRT-PCR was applied to examine the relative expression of HOXD-AS1 in CC tissues, cell lines and transfected cells. Wound healing and transwell assays were applied to detect cell migration and invasion alteration. The targeting relationship between miRNA and mRNA/lncRNA was determined by dual luciferase reporter, qRT-PCR and western blot assays. RESULTS: HOXD-AS1 was overexpressed in CC tissues and cell lines. Its higher level predicted worse prognosis of CC patients. SiRNA mediated knockdown of HOXD-AS1 repressed CC cell migration and invasion, and its overexpression did the opposite. Mechanistically, HOXD-AS1 acted as a competing endogenous RNA (ceRNA) to sponge miR-877-3p and led to upregulation of FGF2, a target of miR-877-3p. Importantly, either miR-877-3p overexpression or FGF2 inhibition could abolish the migration and invasion promotion induced by HOXD-AS1. CONCLUSION: HOXD-AS1 functions as a tumor-promoting lncRNA via the miR-877-3p/FGF2 axis in CC. HOXD-AS1 might be a promising therapeutic target as well as a novel prognostic biomarker for CC.

MiR-100-3p and miR-877-3p regulate overproduction of IL-8 and IL-1ß in mesangial cells activated by secretory IgA from IgA nephropathy patients.

IgA nephropathy (IgAN) is the most common type of primary glomerulonephritis, characterized by mesangial deposition of pathogenic IgA and the injury to mesangial cells. Our previous studies indicate that secretory IgA (SIgA) plays an important role in the pathogenesis of IgAN, and miR-16 is involved in destructive process in mesangial cells mediated by the SIgA from IgAN patients. Our current study aimed to study the role of miRNAs in the effect of SIgA from IgAN patients on mesangial cells. MicroRNA microarray and cytokines assay were performed to obtain the differential microRNAs expression profile in human renal mesangial cells stimulated by SIgA from IgAN patients (P-SIgA) with the cells treated by SIgA from healthy subjects (N-SgA) as control. The microRNAs with the most significant differences in microarray analysis were validated by quantitative RT-PCR. Among them, miR-100-3p and miR-877-3p were selected to predict target gene related to cytokines detecting in this study. Fifty-six differentially expressed microRNAs were chosen and 17 microRNAs with the most prominent changes were validated. Compared with N-SIgA, P-SIgA increased the production of interleukin (IL)-1ß, IL-8, monocyte chemotactic protein-1 and transforming growth factor-ß1. In addition, we for the first time demonstrated that over-production of IL-8 induced by the SIgA was regulated by down-expression of miR-100-3p in mesangial cells. Similarly, IL-1ß over-production was regulated by down-expression of miR-877-3p. Our findings represent a pathogenic microRNAs expression profiling in human mesangial cells activated by P-SIgA. Furthermore, we provide a new explanation characterizing the molecular mechanism responsible for the regulation of IL-1ß and IL-8 production in P-SIgA-triggered mesangial cells.

miR-877-3p as a Potential Tumour Suppressor of Oesophageal Squamous Cell Carcinoma.

BACKGROUND/AIM: MicroRNAs (miRNAs) are abnormally expressed and involved in the pathogenesis of various carcinomas. The present study aimed to identify novel miRNA genes associated with the pathogenesis and prognosis of oesophageal squamous cell carcinoma (ESCC). MATERIALS AND METHODS: The miRNA profiling of 873 genes was performed using surgically resected oesophageal tissues from 35 patients with ESCC to identify candidate miRNAs. To examine the biological activities of candidate miRNAs, their proliferative, invasive, and migratory abilities were evaluated in ESCC cells subjected to miRNA mimic-mediated over-expression. The miRNA expression levels of the selected candidate miRNAs were analysed in the resected oesophageal tissues of 76 patients with ESCC from the two cohorts and correlated with the clinicopathological parameters. RESULTS: Among the four candidate miRNAs identified by miRNA profiling, miR-877-3p was selected for subsequent analyses. In vitro analyses showed that the over-expression of miR-877-3p significantly suppressed the proliferation, invasion, and migration of ESCC cell lines compared with those of control cells. In the analyses of clinical specimens, the expression of miR-877-3p was down-regulated in ESCC tissues compared with that in adjacent normal oesophageal tissues. The down-regulation of miR-877-3p expression in ESCC tissues was significantly associated with advanced local progression and lymphatic involvement. The miR-877-3p down-regulation was also significantly associated with poor disease-free and disease-specific survival. CONCLUSION: miR-877-3p acts as a tumour suppressor gene in ESCC cells, and its down-regulation in ESCC tissues is associated with a poor prognosis. Thus, miR-877-3p may serve as a novel prognostic marker and promising therapeutic target.

Regulation of secondary hair follicle cycle in cashmere goats by miR-877-3p targeting IGFBP5 gene.

Cashmere, a highly valuable animal product derived from cashmere goats, holds significant economic importance. MiRNAs serve as crucial regulators in the developmental processes of mammalian hair follicles. Understanding the regulation of miRNAs during the hair follicle cycle is essential for enhancing cashmere quality. In this investigation, we employed high-throughput sequencing technology to analyze the expression profiles of miRNAs in the secondary hair follicles of Jiangnan cashmere goats at different stages. Through bioinformatics analysis, we identified differentially expressed miRNAs (DE miRNAs). The regulatory relationships between miRNAs and their target genes were verified using multiple techniques, including RT-qPCR, western blot, Dual-Luciferase Reporter, and CKK-8 assays. Our findings revealed the presence of 193 DE miRNAs during various stages of the hair follicle cycle in Jiangnan cashmere goats. Based on the previously obtained mRNA data, the target genes of DE miRNA were predicted, and 1,472 negative regulatory relationships between DE miRNAs and target genes were obtained. Notably, the expression of chi-miR-877-3p was down-regulated during the telogen (Tn) phase compared to the anagen (An) and catagen (Cn) phases, while the IGFBP5 gene exhibited up-regulation. Further validation experiments confirmed that overexpression of chi-miR-877-3p in dermal papilla cells suppressed IGFBP5 gene expression and facilitated cell proliferation. The results of this study provide novel insights for analyzing the hair follicle cycle.

Cashmere goats, known for their diverse range of animal products including delectable meat, soft sheepskin, and high-quality natural fiber, offer an excellent opportunity to explore hair regeneration as they represent a heterogenous fur mammal. MicroRNAs (miRNAs) are small non-coding RNA molecules encoded by approximately 3% of mammalian genes. Remarkably, they have the ability to regulate around 30% of protein-coding genes. Given that a single miRNA can target multiple genes, they exert control over various biological functions such as metabolism, growth, development, and immunity. In this study, we found that miR-877-3p is able to regulate the expression of insulin-like growth factor binding protein 5 (IGFBP5), a key gene involved in the development of the hair follicle cycle. Investigating the regulatory role of miRNAs in the hair follicle cycle of cashmere goats provides a novel perspective for unraveling the mechanisms underlying hair follicle growth and development in mammals.

miR-877-3p inhibits tumor growth and angiogenesis of osteosarcoma through fibroblast growth factor 2 signaling.

Osteosarcoma (OS) is the most common high-grade malignant bone tumor in teenagers. MicroRNAs can function as posttranscriptional regulators of gene expression, playing critical roles in cancer dev-877-3p in OS. Quantitative real-time RT-PCR was carried out for detecting miR-877-3p expression in OS. The effects of miR-877-3p on proliferation was analyzed via MTT, colony formation, and flow cytometry assays. Angiogenesis of endothelial cells were investigated by wound healing and tube formation assay. Gene profiling based on PCR array and luciferase reporter assay were conducted to determine target genes of miR-877-3p. In-vivo study was used to determine the effects of miR-877-3p on the tumor growth. The expression of miR-877-3p was markedly downregulated in OS tissues and cell lines. Low expression of miR-877-3p predicts poor prognosis of OS patients. miR-877-3p overexpression was found to inhibit the proliferation of OS cell lines. The angiogenesis assays showed that miR-877-3p attenuated the angiogenesis. Further mechanism studies showed that miR-877-3p can reduce (Fibroblast Growth Factor 2) FGF2 expression in OS cells by binding to the 3'UTR end of FGF2. Moreover, increased expression of miR-877-3p was responsible for the inhibition of tumor growth and angiogenesis. Taken together, our findings indicated that miR-877-3p might exhibit tumor suppressive role by targeting FGF2 signaling, which may serve as potential target for OS.

GM-CSF-miRNA-Jak2/Stat3 Signaling Mediates Chemotherapy-Induced Cancer Cell Stemness in Gastric Cancer.

Chemotherapy serves as the first choice in clinic to treat advanced gastric cancer. However, emerging evidence indicated the induction of drug resistance and cancer stem cells occasionally by chemotherapy, which seriously limit the therapeutic effects, but the regulatory mechanism remains unclear. Here we treated two human gastric cancer cell lines SGC7901 and BGC823 with 5-Fluorouracil (5-Fu) or Cisplatin (DDP) in vitro. The survived cells showed significant increase of drug resistance, cell stemness and cytokine GM-CSF expression and secretion. As such, GM-CSF was applied to stimulate gastric cancer cells, followed by the subpopulation of CD133 + CSC analysis, sphere formation assay and stemness genes expression analysis. As a result, CSCs showed induction by GM-CSF treatment. A gastric cancer animal model further indicated that the gastric cancer cells significantly promoted tumor growth after GM-CSF treatment in vivo. High-throughput miRNA and mRNA sequencing analyses identified a subset of miRNAs and mRNAs under regulation of both 5-Fu and GM-CSF in gastric cancer cells, including upregulation of miR-877-3p and downregulation of SOCS2. Targeted overexpression or knockdown of miR-877-3p in gastric cancer cells revealed the oncogenic function of miR-877-3p in regulating gastric cancer by suppressing target gene SOCS2. Jak2/Stat3 signaling pathway, as a downstream target of SOCS2, showed activation in vitro and in vivo after treatment with miR-877-3p or GM-CSF. Our findings not only revealed a novel mechanism through which chemotherapy induced CSCs in gastric cancer via GM-CSF-miRNA-Jak2/Stat3 signaling, but also provided an experimental evidence for appropriate dose reduction of adjuvant chemotherapy in treatment of cancer patients.

Linc00941 regulates esophageal squamous cell carcinoma via functioning as a competing endogenous RNA for miR-877-3p to modulate PMEPA1 expression.

Esophageal squamous cell carcinoma (ESCC) represents one of the most common malignancies and is the fifth leading cause of cancer-related deaths. Long intergenic non-coding RNAs (lincRNAs) have been suggested to be dysregulated in various types of cancers, and a growing number of lincRNAs have been implicated to be functional in the ESCC progression. In this study, we examined the role of linc00941 in the ESCC progression and explored the underlying molecular mechanisms. The bioinformatics analysis identified the up-regulation of linc00941 in the ESCC tissues. Further in vitro studies showed that linc00941 was up-regulated in ESCC cell lines. The loss-of-function studies demonstrated that linc00941 knockdown suppressed ESCC cell proliferation, invasion and migration, and also suppressed the in vivo tumor growth. Furthermore, bioinformatics prediction along with luciferase reporter assay and RNA immunoprecipitation assay implied that linc00941 acted as a competing endogenous RNA for miR-877-3p, and linc00941 regulated ESCC cell progression via at least targeting miR-877-3p. Subsequently, miR-877-3p targeted prostate transmembrane protein, androgen induced 1 (PMEPA1) 3' untranslated region and repressed PMEPA1 expression in ESCC cells; overexpression of PMEPA1 attenuated the inhibitory effects of linc00941 knockdown on the ESCC cell progression. Linc00941 knockdown suppressed epithelial-mesenchymal transition (EMT) via targeting miR-877-3p/PMEPA1 axis in ESCC cells. In conclusion, our results indicated the oncogenic role of linc00941 in ESCC, and knockdown of linc00941 suppressed ESCC cell proliferation, invasion, migration and EMT via interacting with miR-877-3p/PMEPA1 axis.

UBE2R2-AS1 Inhibits Xenograft Growth in Nude Mice and Correlates with a Positive Prognosis in Glioma.

Our previous study showed that the lncRNA UBE2R2-AS1 inhibits the growth and invasion of glioma cells and promotes apoptosis through the miR-877-3p/TLR4 pathway. In this study, it was further found that the expression of UBE2R2-AS1 in glioma tissues was decreased significantly, and gradually decreased with increasing clinical stage. Chi-square analysis showed that the expression of UBE2R2-AS1 was significantly correlated with the WHO stage of tumor and epilepsy. Using Kaplan-Meier univariate survival analysis, it was found that the expression of UBE2R2-AS1 correlated positively with the overall survival of patients with glioma, while multiple Cox regression analysis showed that the expression of UBE2R2-AS1 correlated positively with the overall survival of patients with glioma as a protective factor for glioma prognosis. The analysis of data from TCGA also showed that patients with high UBE2R2-AS1 levels or low miR-877-3p expression were more likely to have good survival outcomes. Further construction of a glioma xenograft model in nude mice showed that UBE2R2-AS1 overexpression inhibited the growth of tumors, and the inhibition of miR-877-3p expression had a similar effect. Simultaneous UBE2R2-AS1 overexpression and miR-877-3p inhibition further decreased the growth rate of tumors in nude mice. Taken together, the results of our study suggest that UBE2R2-AS1 is an important tumor suppressor gene in glioma, which may be a good marker and treatment target for the clinical detection of glioma.

Understanding the Molecular Mechanism of miR-877-3p Could Provide Potential Biomarkers and Therapeutic Targets in Squamous Cell Carcinoma of the Cervix.

No therapeutic targets and molecular biomarkers are available in cervical cancer (CC) management. In other cancer types, micro-RNA-877-3p (miR-877-3p) has been associated with events relevant for CC development. Thus, we aimed to determine miR-877-3p role in CC. miR-877-3p levels were examined by quantitative-PCR in 117 cervical lesions and tumors. Effects on CC cell proliferation, migration, and invasion were evaluated upon anti-miR-877-3p transfection. miR-877-3p dependent molecular mechanism was comprehensively explored by proteomics, dual-luciferase reporter assay, western blot, and immunohistochemistry. Cervical tumors expressed higher miR-877-3p levels than benign lesions. miR-877-3p promoted CC cell migration and invasion, at least partly by modulating cytoskeletal protein folding through the chaperonin-containing T-complex protein 1 complex. Notably, miR-877-3p silencing synergized with paclitaxel. Interestingly, miR-877-3p downregulated the levels of an in silico-predicted target, ZNF177, whose expression and subcellular location significantly distinguished high-grade squamous intraepithelial lesions (HSILs) and squamous cell carcinomas of the cervix (SCCCs). Cytoplasmic ZNF177 was significantly associated with worse progression-free survival in SCCC. Our results suggest that: (i) miR-877-3p is a potential therapeutic target whose inhibition improves paclitaxel effects; (ii) the expression and location of its target ZNF177 could be diagnostic biomarkers between HSIL and SCCC; and (iii) cytoplasmic ZNF177 is a poor-prognosis biomarker in SCCC.

miR-877-3p promotes TGF-ß1-induced osteoblast differentiation of MC3T3-E1 cells by targeting Smad7.

MicroRNAs (miRNAs) are emerging as important regulators of various physiological and pathological processes and may serve key roles in the maintenance of bone homeostasis via effects on osteoblast differentiation. The aim of the present study was to define the role of miR-877-3p in osteoblast differentiation using MC3T3-E1 cells, an osteoblast precursor cell line. It was demonstrated using RT-qPCR analysis that miR-877-3p was gradually increased in MC3T3-E1 cells during the osteoblastic differentiation induced by transforming growth factor (TGF)-ß1. Gain-of-function and loss-of-function experiments revealed that the overexpression of miR-877-3p promoted the osteoblastic differentiation of MC3T3-E1 cells, whereas depletion of miR-877-3p inhibited this process in vitro and in vivo. Bioinformatics analysis and validation experiments demonstrated that Smad7, which acts as a negative regulator of osteogenesis, was a target of miR-877-3p. Furthermore, the overexpression of Smad7 partially reversed the osteoblastic differentiation of MC3T3-E1 cells induced by miR-877-3p. In conclusion, the results of the present study suggest that the miR-877-3p/Smad7 axis is associated with the osteoblastic differentiation of MC3T3-E1 cells and may indicate a potential therapeutic approach for osteogenesis disorders.

lncRNA H19 Alleviated Myocardial I/RI via Suppressing miR-877-3p/Bcl-2-Mediated Mitochondrial Apoptosis.

Ischemic cardiac disease is the leading cause of morbidity and mortality in the world. Despite the great efforts and progress in cardiac research, the current treatment of cardiac ischemia reperfusion injury (I/RI) is still far from being satisfactory. This study was performed to investigate the role of long non-coding RNA (lncRNA) H19 in regulating myocardial I/RI. We found that H19 expression was downregulated in the I/R hearts of mice and cardiomyocytes treated with H2O2. Overexpression of H19 alleviated myocardial I/RI of mice and cardiomyocyte injury induced by H2O2. We found that H19 functioned as a competing endogenous RNA of miR-877-3p, which decreased the expression of miR-877-3p through the base-pairing mechanism. In parallel, miR-877-3p was upregulated in H2O2-treated cardiomyocytes and mouse ischemia reperfusion (I/R) hearts. miR-877-3p exacerbated myocardial I/RI and cardiomyocyte apoptosis. We further established Bcl-2 as a downstream target of miR-877-3p. miR-877-3p inhibited the mRNA and protein expression of Bcl-2. Furthermore, H19 decreased the Bcl-2/Bax ratio at mRNA and protein levels, cytochrome c release, and activation of caspase-9 and caspase-3 in myocardial I/RI mice, which were canceled by miR-877-3p. In summary, the H19/miR-877-3p/Bcl-2 pathway is involved in regulation of mitochondrial apoptosis during myocardial I/RI, which provided new insight into molecular mechanisms underlying regulation of myocardial I/RI.

Long noncoding RNA UBE2R2-AS1 promotes glioma cell apoptosis via targeting the miR-877-3p/TLR4 axis.

Introduction: Brain glioma is the most common type of primary malignancy in the central nervous system (CNS), with high recurrence and mortality rate, especially glioblastoma (GBM). Recent evidence suggests a role for many long noncoding RNAs (lncRNAs) in the pathogenesis, proliferation, apoptosis, metastasis, and chemotherapeutic resistance of cancer cells. Although the functions of some lncRNAs in the occurrence and development of gliomas have been confirmed, detailed mechanisms of action are lacking. Furthermore, the biological roles of many other lncRNAs in glioma have not been reported at all. Methods: In this study, we identified a novel lncRNA, UBE2R2-AS1, which was dramatically downregulated in glioma compared with normal tissue, by performing microarray detection of six pairs of glioma samples and adjacent normal tissues. In vitro experiments demonstrated that UBE2R2-AS1 regulated glioma cell proliferation, apoptosis, and migration. Results: UBE2R2-AS1 acted as a competing endogenous RNA (ceRNA) to target Toll-like receptor 4 (TLR4) mRNA by binding to miR-877-3p. Furthermore, lncRNA UBE2R2-AS1 suppressed glioblastoma cell growth, migration, and invasion, as well as promoting cell apoptosis by targeting miR-877-3p/TLR4 directly. Conclusion: This information regarding UBE2R2-AS1 and its glioma-related molecular mechanisms will aid the future identification of new lncRNA-directed diagnostics and drug-targeting therapies.