Circular RNA 0001789 sponges miR-140-3p and regulates PAK2 to promote the progression of gastric cancer.

BACKGROUND: Gastric cancer (GC) is the third-leading cause of cancer-associated mortalities globally. The deregulation of circular RNAs (circRNAs) and microRNAs (miRNAs or miRs) is widely implicated in the pathogenesis and progression of different cancer types. METHODS: The expression profiling of circRNAs in GC is required to identify crucial circRNAs as biomarkers or therapeutic targets. In the present study, a published circRNA microarray dataset was used to identify differentially expressed circRNAs between GC tissues and normal gastric mucosa tissues. Reverse transcription-quantitative PCR was performed to validate the expression of circ_0001789. Fisher's exact test, receiver operating characteristic curve and Kaplan-Meier plots were employed to analyze the clinical significance of circ_0001789. The miRNA targets of circ_0001789 were predicted using an online database, and their functional interaction was further confirmed by RNA pull-down, RNA immunoprecipitation and dual luciferase reporter assays. Transwell assays were conducted to investigate the biological functions of circ_0001789, miR-140-3p and p21 activated kinase 2 (PAK2) in the migration and invasion of GC cells. A xenograft mouse model was established to validate the role of circ_0001789 in the tumorigenesis of GC cells. RESULTS: circ_0001789 was identified as a highly expressed circRNA in GC tissues versus normal gastric mucosa tissues. Silencing circ_0001789 attenuated the malignancy of GC cells, and exosomal circ_0001789 was sufficient to regulate the malignant phenotype of GC cells. miR-140-3p was further identified as a downstream target of circ_0001789, which showed a negative correlation with circ_0001789 expression in GC tissues. Overexpression of miR-140-3p suppressed cell migration, invasion and epithelial-mesenchymal transition in GC cells. PAK2 was identified as the target of miR-140-3 to mediate the malignant phenotype of GC cells. CONCLUSION: The present data suggested that the upregulation of circ_0001789 was associated with the progression of GC and with poor prognosis in patients with GC, and that miR-140-3p/PAK2 served as the downstream axis to mediate the oncogenic effect of circ_0001789.

Exosomal microRNA-140-3p from human umbilical cord mesenchymal stem cells attenuates joint injury of rats with rheumatoid arthritis by silencing SGK1.

OBJECTIVE: Over the years, microRNAs (miRNAs) have been involved in the pathogenesis of rheumatoid arthritis (RA). We aim to investigate the role of human umbilical cord mesenchymal stem cells (HUCMSCs)-derived exosomal miR-140-3p in RA development. METHODS: Exosomes(exo) were isolated from human umbilical cord-derived mesenchymal stem cells (HUCMSCs), and this isolation was followed by the transfer of miR-140-3p. RA rat models were constructed by collagen II adjuvant and respectively treated with HUCMSCs-exo or HUCMSCs-exo carrying miR-140-3p mimic/inhibitor, and expression of miR-140-3p and serum- and glucocorticoid-inducible kinase 1 (SGK1) was assessed. Then, RA score and inflammation scoring, fibrosis degree and apoptosis, serum inflammatory response and oxidative stress in joint tissues were determined. The RA synovial fibroblasts (RASFs) were extracted from rats and identified. Conducted with relative treatment, the migration, proliferation and apoptosis in RASFs were determined. RESULTS: MiR-140-3p was decreased while SGK1 was increased in RA rats. HUCMSCs-exo or upregulated exosomal miR-140-3p improved pathological changes and suppressed inflammation, oxidative stress and fibrosis in RA rats, and also constrained and RASF growth. Overexpression of SGK1 reversed the inhibition of RASF growth caused by overexpression of miR-140-3p. CONCLUSION: Upregulated exosomal miR-140-3p attenuated joint injury of RA rats by silencing SGK1. This research provided further understanding of the role of exosomal miR-140-3p in RA development.

MicroRNA-140-3p represses the proliferation, migration, invasion and angiogenesis of lung adenocarcinoma cells via targeting TYMS (thymidylate synthetase).

MicroRNA (miR)-140-3p has been proved to repress lung adenocarcinoma (LUAD), and our study aims to further evaluate the mechanism. Bioinformatic analyses were performed. The viability, proliferation, migration, invasion and angiogenesis of transfected LUAD cells were all determined via Cell Counting Kit-8, colony formation, Scratch, Transwell, and tube formation assays. The targeting relationship between miR-140-3p and thymidylate synthetase (TYMS) was confirmed by dual-luciferase reporter assay. Relative expressions of miR-140-3p, TYMS, epithelial-to-mesenchymal transition- (E-cadherin, N-cadherin, vimentin), angiogenesis- (vascular endothelial growth factor (VEGF)), and apoptosis-related factors (cleaved caspase-3, B-cell lymphoma-2 (Bcl-2), Bcl-2-associated X protein (Bax)) were quantified by quantitative real-time polymerase chain reaction or Western blot. TYMS was high-expressed yet miR-140-3p was low-expressed in LUAD cells. Upregulation of miR-140-3p inhibited TYMS expression, viability, colony formation, migration, invasion, and tube length within LUAD cells, while downregulation of miR-140-3p did oppositely. Silenced TYMS, the downstream target gene of miR-140-3p, reversed the effects of miR-140-3p downregulation on TYMS expression, cell viability, colony formation, migration, invasion, and tube length as well as the metastasis-, apoptosis- and angiogenesis-related proteins in LUAD cells. Upregulation of miR-140-3p inhibited the proliferation, migration, invasion and angiogenesis of LUAD cells via targeting TYMS.

Knockdown of LINC01385 inhibits osteoarthritis progression by modulating the microRNA-140-3p/TLR4 axis.

Long non-coding (lnc) RNAs have been associated with osteoarthritis (OA) progression. The aim of the present study was to investigate the regulatory mechanism of lncRNA LINC01385 in OA in vitro. The mRNA expression level of LINC01385, microRNA(miR)-140-3p, and Toll-like receptor 4 (TLR4) was detected using reverse transcription-quantitative PCR, while ELISA was used to determine the concentration of different inflammatory factors [tumor necrosis factor-α (TNF-α), IL-6, and prostaglandin E2 (PGE2)]. The viability of human articular chondrocytes (HC-a) was measured using a MTT assay and western blot analysis was performed to quantify the protein expression level of TLR4. The associations between miR-140-3p and LINC01385/TLR4 were confirmed using a dual-luciferase reporter assay. LINC01385 mRNA expression level was increased in OA tissues and IL-1ß-induced HC-a. LINC01385 knockdown and miR-140-3p mimics reduced the concentration of inflammatory factors in IL-1ß-induced HC-a and promoted cell survival. In addition, it was confirmed that LINC01385 targeted miR-140-3p, while TLR4 was a target gene of miR-140-3p. Negative correlations between LINC01385 and miR-140-3p, and between miR-140-3p and TLR4 were observed in OA tissues. Low mRNA expression level of miR-140-3p and high protein expression level of TLR4 reversed the inhibitory effect of LINC01385 knockdown on the inflammatory responses of IL-1ß-induced HC-a and exhibited a stimulating effect on cell viability. LINC01385 knockdown reduced the progression of OA by modulating the miR-140-3p/TLR4 axis in vitro; thus, LINC01385 may be a therapeutic target for OA.

MicroRNA-140-3p alleviates intervertebral disc degeneration via KLF5/N-cadherin/MDM2/Slug axis.

MicroRNAs (miRNAs) are associated with healing or deteriorating degenerated intervertebral disc (IVD) tissues in spinal cord diseases, including intervertebral disc degeneration (IDD). IDD represents a chronic process of extracellular matrix destruction, but the relevant molecular mechanisms implicated in the regenerative effects of miRNAs are unclear. Here, we investigated the regenerative effects of microRNA-140 (miR-140-3p) in an IDD model induced by annulus needle puncture. Bioinformatics analysis was conducted to identify regulatory factors (KLF5/N-cadherin/MDM2/Slug) linked to miR-140-3p effects in IDD. Mesenchymal stem cells (MSCs) were extracted from degenerated IVD nucleus pulposus (NP), and the expression of miR-140-3p/KLF5/N-cadherin/MDM2/Slug was manipulated to explore their effects on cell proliferation, migration, apoptosis and differentiation. The results showed that miR-140-3p was under-expressed in the degenerated IVD NP, whereas its overexpression alleviated IDD. Mechanistic studies suggested that miR-140-3p targeted KLF5 expression, and high KLF5 expression impeded the migration and differentiation of MSCs. In degenerated IVD NP-derived MSCs, MiR-140-3p-mediated KLF5 downregulation simultaneously elevated N-cadherin expression and transcriptionally inhibited MDM2, thus upregulating Slug expression. The experimental data indicated that miR-140-3p enhanced the proliferation, migration and differentiation of degenerated IVD NP-derived MSCs and repressed their apoptosis. The in vivo validation experiment also demonstrated that miR-140-3p inhibited IDD by modulating the KLF5/N-cadherin/MDM2/Slug axis. Collectively, our results uncovered the regenerative role of miR-140-3p in IDD via regulation of the KLF5/N-cadherin/MDM2/Slug axis, which could be a potential therapeutic target for IDD.Abbreviations: miR-140-3p: microRNA-140-3p; IDD: intervertebral disc degeneration; MSCs: Mesenchymal stem cells; IVD: intervertebral disc; MSCs: mesenchymal stem cells; KLF5: Kruppel-like factor 5; MDM2: mouse double minute 2; NC: negative control; DHI: disc height index.

MiR-140-3p Ameliorates the Progression of Osteoarthritis via Targeting CXCR4.

Osteoarthritis is a common disease character with progressive destruction of cartilage. MicroRNA (miR)-140-3p was validated as a biomarker for osteoarthritis. However, the mechanism by which miRNA-140-3p regulates osteoarthritis remains unclear. Thus, this study aims to evaluate the potential function of miRNA-140-3p during the pathogenesis of osteoarthritis. MiRNA-140-3p expression in tissue and CHON-001 chondrocyte cells was determined with quantitative real time (qRT)-PCR. In vitro osteoarthritis model was established by treatment of the chondrocyte cells CHON-001 with interleukin (IL)-1ß. Cell proliferation and apoptosis were measured with cell counting kit-8 (CCK8) and Annexin V/propidium iodide (PI) apoptosis assay, respectively. Protein expressions were evaluated using Western blot. The target gene of miR-140-3p was predicted using Targetscan and miRDB. MiR-140-3p was downregulated in knee tissue from patients with osteoarthritis. IL-1ß inhibited the proliferation of CHON-001 cells via inducing apoptosis. In addition, IL-1ß significantly inhibited the expressions of collagen II and aggrecan and increased the level of matrix metalloproteinase (MMP)13. However, the effects of IL-1ß could be ameliorated by the addition of miR-140-3p mimics. Moreover, luciferase reporter assay demonstrated CXCR4 as a target gene of miR-140-3p. IL-1ß-induced upregulation of CXCR4 could be blocked by miR-140-3p mimics. Our study indicated that miR-140-3p could suppress the progression of osteoarthritis by directly targeting CXCR4. Therefore, miR-140-3p might serve as a potential therapeutic target for the treatment of osteoarthritis.

Circular RNA circRNA_000203 aggravates cardiac hypertrophy via suppressing miR-26b-5p and miR-140-3p binding to Gata4.

AIMS: Circular RNAs (circRNAs) are involved in gene regulation in a variety of physiological and pathological processes. The present study aimed to investigate the effect of circRNA_000203 on cardiac hypertrophy and the potential mechanisms involved. METHODS AND RESULTS: CircRNA_000203 was found to be up-regulated in the myocardium of Ang-II-infused mice and in the cytoplasma of Ang-II-treated neonatal mouse ventricular cardiomyocytes (NMVCs). Enforced expression of circRNA_000203 enhances cell size and expression of atrial natriuretic peptide and ß-myosin heavy chain in NMVCs. In vivo, heart function was impaired and cardiac hypertrophy was aggravated in Ang-II-infused myocardium-specific circRNA_000203 transgenic mice (Tg-circ203). Mechanistically, we found that circRNA_000203 could specifically sponge miR-26b-5p, -140-3p in NMVCs. Further, dual-luciferase reporter assay showed that miR-26b-5p, -140-3p could interact with 3'-UTRs of Gata4 gene, and circRNA_000203 could block the above interactions. In addition, Gata4 expression is transcriptionally inhibited by miR-26b-5p, -140-3p mimic in NMVCs but enhanced by over-expression of circRNA_000203 in vitro and in vivo. Functionally, miR-26b-5p, -140-3p, and Gata4 siRNA, could reverse the hypertrophic growth in Ang-II-induced NMVCs, as well as eliminate the pro-hypertrophic effect of circRNA_000203 in NMVCs. Furthermore, we demonstrated that NF-κB signalling mediates the up-regulation of circRNA_000203 in NMVCs exposed to Ang-II treatment. CONCLUSIONS: Our data demonstrated that circRNA_000203 exacerbates cardiac hypertrophy via suppressing miR-26b-5p and miR-140-3p leading to enhanced Gata4 levels.

miR-140-3p Suppresses Cell Growth And Induces Apoptosis In Colorectal Cancer By Targeting PD-L1.

BACKGROUND: A variety of miRNAs have been recently reported to be abnormally expressed in colorectal cancer (CRC). A growing number of studies have demonstrated that aberrantly expressed miRNAs are closely related to the development and progression of CRC. It has been found that miR-140-3p plays a vital role in several cancers. However, its expression, roles and mechanisms in CRC are remain unknown. MATERIALS AND METHODS: Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) were used to determine miR-140-3p expression in CRC tissues and cell lines. CCK8, migration, invasion and flow cytometric assays were used to determine the influence of miR-140-3p upregulation on cell proliferation, migration, invasion and apoptosis of CRC cells. Luciferase reporter assays and Western blots were utilized to identify the target genes of miR-140-3p. In addition, the potential mechanism of miR-140-3p action in CRC cells was elucidated. RESULTS: In our study, miR-140-3p expression was significantly decreased in CRC tissues and cell lines. Overexpression of miR-140-3p attenuated proliferation, migration, and invasion and induced the apoptosis of CRC cells. Bioinformatics analyse and luciferase reporter analysis identified PD-L1 as a putative target gene of miR-140-3p. PD-L1 was overexpressed in CRC tissues and inversely correlated with miR-140-3p expression. Suppression of PD-L1 expression in CRC cells generated biological behaviours in CRC cells that were similar to those observed after treated with miR-140-3p mimics. Restoration of PD-L1 expression partially attenuated the inhibitory effect of miR-140-3p on CRC cells. Western blot were used to verify the effect of PD-L1 expression on PI3K/AKT pathway. In addition, overexpression of miR-140-3p could inhibit CRC tumor growth in vivo. CONCLUSION: In general, these data demonstrate that miR-140-3p acts as a tumour suppressor in CRC by directly targeting PD-L1 and inactivating PI3K/AKT pathway, suggesting that miR-140-3p might be a novel target for CRC diagnosis and treatment.

Upregulation of microRNA-140-3p inhibits epithelial-mesenchymal transition, invasion, and metastasis of hepatocellular carcinoma through inactivation of the MAPK signaling pathway by targeting GRN.

Invasion and metastasis in hepatocellular carcinoma (HCC) results in poor prognosis. Human intervention in these pathological processes may benefit the treatment of HCC. The aim of the present study is to elucidate the mechanism of miR-140-3p affecting epithelial-mesenchymal transition (EMT), invasion, and metastasis in HCC. Microarray analysis was performed for differentially expressed genes screening. The target relationship between miR-140-3p and GRN was analyzed. Small interfering RNA (siRNA) against granulin (GRN) was synthesized. EMT markers were detected, and invasion and migration were evaluated in HCC cells introduced with a miR-140-3p inhibitor or mimic, or siRNA against GRN. A mechanistic investigation was conducted for the determination of mitogen-activated protein kinase (MAPK) signaling pathway-related genes and EMT markers (E-cadherin, N-cadherin, and Vimentin). GRN was highlighted as an upregulated gene in HCC. GRN was a target gene of miR-140-3p. Elevation of miR-140-3p or inhibition of GRN restrained the EMT process and suppressed the HCC cell migration and invasion. HCC cells treated with the miR-140-3p mimic or siRNA-GRN exhibited decreased GRN expression and downregulated the expressions of the MAPK signaling pathway-related genes, N-cadherin, and Vimentin but upregulated the expression of E-cadherin. GRN silencing can reverse the activation of the MAPK signaling pathway and induction of EMT mediated by miR-140-3p inhibition. Taken together, the results show that miR-140-3p confers suppression of the MAPK signaling pathway by targeting GRN, thus inhibiting EMT, invasion, and metastasis in HCC.

miR-140-3p inhibits breast cancer proliferation and migration by directly regulating the expression of tripartite motif 28.

The present study aimed to determine the expression profile and significance of microRNA-140-3p (miR-140-3p) in breast cancer (BC). miR-140-3p expression in BC tumor tissues and cell lines was measured by reverse transcription-quantitative polymerase chain reaction. Luciferase activity reporter assay and western blotting were used to assess the effect of miR-140-3p expression on tripartite motif 28 (TRIM28). Cell growth, migration, and invasion were assessed by Cell Counting Kit-8 assay, wound-healing assay and Transwell invasion assay, respectively. miR-140-3p expression was significantly reduced in BC tumor tissues compared with in adjacent normal tissues. Additionally, low miR-140-3p expression was found to predict poor prognosis of patients with BC. TRIM28 expression was significantly reduced by miR-140-3p overexpression in BC cell lines, and was inversely correlated with miR-140-3p in BC tissues. Overexpression of miR-140-3p also inhibited cell proliferation, migration and invasion compared with in the control group. In conclusion, the present study revealed that miR-140-3p inhibited the progression of BC partially by regulating TRIM28.

Overexpression of serum MicroRNA-140-3p in premenopausal women with newly diagnosed breast cancer.

AIMS: The purpose of the present study was to evaluate microRNA-140-3p expression level in breast cancer patients in comparison to healthy controls. PATIENTS & METHODS: Serum microRNA-140-3p level was quantified by realtime quantitative reverse transcription PCR in 40 women with breast cancer and 40 healthy subjects. RESULTS: Serum microRNA-140-3p level in patients compared to healthy subjects was significantly up-regulated (P = 0.01). MicroRNA-140-3p had a good diagnostic accuracy for discrimination of the two groups (AUC = 0.667; sensitivity = 70%; specificity = 50%). Serum microRNA-140-3p level was overexpressed in premenopausal patients who were ≤48 years old. ROC curve showed a similar pattern again (AUC = 0.690; sensitivity = 73%; specificity = 50%). CONCLUSIONS: microRNA-140-3p has the potential for detection of breast cancer, especially in premenopausal and in ≤48 years old women.