MiR-381-3p inhibits Warburg effect and progression in osteosarcoma via targeting PFKFB3.

BackgroundStudies have shown that glutathione peroxidase 8 (GPX8) promotes the progression of various cancers. However, the role of GPX8 in osteosarcoma (OS) progression remains unclear. The aim of this study was to investigate the function of GPX8 in OS progression and its underlying mechanisms.MethodsGPX8 expression in OS tissues and cell lines was assessed using immunohistochemistry (IHC), qRT‒PCR and Western blotting. The effects of GPX8 on the proliferative capacity of OS cells were evaluated using CCK-8, EdU, and plate colony formation assays. The role of GPX8 in the migratory and invasive functions of OS cells was examined via scratch wound healing, Transwell, and invasion assays. The effect of GPX8 on the in vivo proliferative ability of OS cells was investigated through a subcutaneous tumour formation experiment in nude mice. Bioinformatics methods were employed to predict the transcription factors that regulate GPX8 expression. Chromatin immunoprecipitation and dual-luciferase reporter assays were performed to investigate the interaction between GPX8 and Kruppel‑like factor 16 (KLF16).ResultsGPX8 is highly expressed in OS tissues and OS cell lines. Knocking down GPX8 expression significantly inhibited OS cell proliferation, migration, and invasion; downregulated CyclinD1 protein expression; and suppressed the EMT process. Conversely, GPX8 overexpression yielded the opposite results. Knocking down GPX8 expression significantly inhibited the tumorigenic ability of OS cells in vivo. KLF16 is highly expressed in OS tissues and cell lines. KLF16 promotes GPX8 expression. Knocking down GPX8 expression inhibited the promoting effects of KLF16 overexpression on OS cell proliferation, invasion, and migration, whereas overexpressing GPX8 reversed the inhibitory effects of KLF16 knockdown on OS cell proliferation, invasion, and migration. The results of the dual-luciferase and ChIP assays demonstrated that KLF16 directly binds to the GPX8 promoter and positively regulates GPX8 expression.ConclusionsThis study demonstrated that GPX8 is a critical oncogene in OS progression and that its expression is regulated by KLF16. Targeting the KLF16/GPX8 axis may offer promising prospects for the development of novel therapeutic strategies against OS.Supplementary InformationThe online version contains supplementary material available at 10.1186/s12967-025-07313-5.

The anti-tumor action of engineered mesenchymal stem cells highlights the double-edged role of oncoproteins in osteosarcoma, and suggests the possibility of developing a novel strategy for protein-based cancer therapy.

Osteosarcoma (OS), a prevalent metastatic cancer among young individuals, is associated with a grim prognosis. Long non-coding RNAs (lncRNAs), including C1QTNF1-AS1, are pivotal regulators of cancer cell proliferation and motility. As an oncogene, C1QTNF1-AS1 is implicated in various tumor types, such as colorectal, pancreatic, hepatocellular carcinomas, and OS. The aim of this study was to investigate the functions and underlying mechanisms of C1QTNF1-AS1 in the progression of osteosarcoma. This investigation focused on elucidating the functional roles and mechanisms of C1QTNF1-AS1 in OS cells. Bioinformatics tools were utilized to identify the interaction between microRNA miR-34a-5p and C1QTNF1-AS1, as well as the targeting of LDHA and PDK3 by miR-34a-5p. Dual-luciferase reporter assays and RNA immunoprecipitation were employed to validate these interactions. Expression profiles of C1QTNF1-AS1, miR-34a-5p, LDHA, and PDK3 in osteosarcoma cells were analyzed using RT-PCR and western blot analyses, revealing their intricate relationships. The impact of these molecules on OS cell proliferation, invasion, and migration was assessed through CCK-8, Transwell, and Cell scratch assay. Moreover, the effects on aerobic glycolysis in OS cells were examined by quantifying ATP levels, lactate production, glucose uptake capacity, and the extracellular acidification rate. The findings indicated a significant decrease in C1QTNF1-AS1 expression levels in OS cells compared to normal osteoblasts. A parallel downregulation trend of miR-34a-5p was also observed in OS cells. Silencing C1QTNF1-AS1 led to a marked upregulation of LDHA and PDK3 in OS cells, which was partially attenuated by miR-34a-5p mimics. Functional evaluations demonstrated that suppression of C1QTNF1-AS1 accelerated OS cell growth, motility, invasiveness, and the Warburg effect. Conversely, the overexpression of miR-34a-5p mitigated these stimulatory effects, suggesting a regulatory role in modulating OS progression. Our research emphasizes the critical role of C1QTNF1-AS1 in the pathogenesis of osteosarcoma (OS). We discovered that the downregulation of C1QTNF1-AS1 indirectly upregulates the expression of LDHA and PDK3 by suppressing miR-34a-5p, which functions as a regulator of the Warburg effect. This cascade of events promotes OS progression by enhancing glycolytic metabolism and supplying energy for cancer cell growth, migration, and invasion. These findings suggest a potential therapeutic target and highlight the importance of understanding the regulatory network involving lncRNAs in cancer metabolism and progression.

BackgroundMiR-1297 is reported to function as a tumor suppressor of various cancers. However, the role of miR-1297 in the development of osteosarcoma (OS) has not been elaborated. The purpose of this study was to investigate the functional effects of miR-1297 on OS progression and the underlying mechanism.MethodsThe expression of protein and mRNA in OS cells was evaluated by Western blotting and quantitative real-time polymerase chain reaction. Cellular proliferation was investigated by cell counting kit-8, colony formation and apoptosis assays. Bioinformatics methods were used to predict target genes. The relationship between PFKFB2 and miR-1297 was demonstrated by dual-luciferase reporter assay. Metabolic changes in OS cells were monitored using an XF96 metabolic flux analyzer.ResultsWe found that miR-1297 was downregulated in OS and that lower expression of miR-1297 promoted proliferation and contributed to the Warburg effect in OS cells. Furthermore, we showed that silencing PFKFB2 inhibited proliferation and reduced aerobic glycolysis while overexpression of PFKFB2 reduced the anti-tumor function of miR-1297 in OS cells. Mechanistically, miR-1297 acted as a tumor suppressor in OS and reduced the expression of PFKFB2 by directly targeting its 3ʹUTR.ConclusionThe miR-1297/PFKFB2 axis regulated OS proliferation by controlling the Warburg effect. Our results revealed a previously undiscovered function of miR-1297 in OS, which strongly linked metabolic alterations with cancer progression. Targeting miR-1297 may become a promising therapeutic approach for OS.

Silencing of circCYP51A1 represses cell progression and glycolysis by regulating miR-490-3p/KLF12 axis in osteosarcoma under hypoxia

The aim of this study was to elucidate the regulatory role of lncSNHG16 in the progression of osteosarcoma (OS) and its underlying mechanism. Expressions of lncSNHG16, microRNA-146a-5p and NOVA1 in OS tissues and adjacent normal tissues were determined by quantitative Real-time polymerase chain reaction (qRT-PCR). Their expressions in OS cell lines were detected by qRT-PCR as well. We analyzed the relationship between lncSNHG16 expression and tumor stage, diagnosis and survival prognosis of OS patients, respectively. Cell counting kit-8 (CCK-8) and transwell experiments were conducted to explore proliferative and migratory changes of OS cells. Dual-luciferase reporter assay was used to verify the binding relationship of lncSNHG16 to microRNA-146a-5p, and microRNA-146a-5p to NOVA1. Finally, rescue experiments were performed to elucidate the regulatory effect of lncSNHG16 on the cellular behaviors of OS cells. LncSNHG16 was highly expressed in OS tissues and cell lines. Its expression was positively correlated with the tumor stage of OS patients. Receiver operating characteristic (ROC) curves suggested that lncSNHG16 can be used as a clinical indicator to distinguish OS patients from healthy controls. Survival analysis indicated a negative correlation between lncSNHG16 expression and survival of OS patients. Overexpression of lncSNHG16 enhanced the proliferative and migratory potentials of OS cell lines 143B and MNNG/HOS. MicroRNA-146a-5p was predicted to be the target gene of lncSNHG16, which was lowly expressed in OS tissues and cell lines. Overexpression of lncSNHG16 downregulated the expression of microRNA-146a-5p in 143B and MNNG/HOS cells. Furthermore, we verified that lncSNHG16 could bind to microRNA-146a-5p. The promotive role of lncSNHG16 in proliferative and migratory potentials of OS cells was reversed by microRNA-146a-5p. Subsequently, NOVA1 was predicted to be the target gene of microRNA-146a-5p, and was further verified by dual-luciferase reporter gene assay. Correlation analysis showed that microRNA-146a-5p expression was negatively correlated with NOVA1 expression in OS. More importantly, NOVA1 reversed the inhibitory effect of microRNA-146a-5p on the proliferative and migratory capacities of 143B and MNNG/HOS cells. LncSNHG16 is highly expressed in OS tissues and cell lines, participating in the development of OS by downregulating microRNA-146a-5p to upregulate NOVA1 expression.

BackgroundMicroRNAs (miRNAs) have been verified to be involved in various biological processes through regulating their target genes, and some previous studies have revealed the antitumour role of microRNA-374b-5p (miR-374b-5p) in several tumours. Therefore, the purpose of this study was to explore the functions and potential mechanisms of miR-374b-5p in osteosarcoma (OS) progression.MethodsThe differentially expressed gene miR-374b-5p was discovered in the dataset GSE65071 from the Gene Expression Omnibus (GEO) database via bioinformatics analysis, and its expression levels in OS tissues and cell lines were confirmed by RNA fluorescence in situ hybridization (FISH) staining and quantitative real-time polymerase chain reaction (qRT-PCR). OS cell proliferation ability was evaluated by Cell Counting Kit-8 (CCK-8) assay and colony formation assay, cell migration and invasion abilities were assessed by Transwell assays, and apoptosis was detected by flow cytometry. The underlying mechanisms of miR-374b-5p in regulating OS progression were explored by qRT-PCR, dual-luciferase reporter assay and western blotting. In vivo experiments, a nude mice xenograft tumour model was performed to evaluate the effects of miR-374b-5p on tumour growth and gene expression changes.ResultsIn this study, miR-374b-5p expression was confirmed to be significantly down-regulated in OS, and miR-374b-5p overexpression could inhibit the proliferation, migration, and invasion abilities but promote apoptosis of OS cells. Mechanism studies revealed that miR-374b-5p suppressed the AKT pathway via negatively regulating the expression of phosphoinositide-dependent protein kinase 1 (PDPK1). Notably, PDPK1 were highly expressed in OS cell lines, as verified by qRT-PCR, and PDPK1-silencing considerably restrained OS progression. Moreover, the inhibitory effects of miR-374b-5p on OS progression were partially reversed by PDPK1 overexpression both in vitro and in vivo.ConclusionsMiR-374b-5p was lowly expressed in OS, and its upregulation inhibited the progression of OS by directly targeting PDPK1 to affect the activity of the AKT pathway.

N-terminal histone acetyltransferase NAA40 modulates osteosarcoma progression by controlling AGR2 expression.

BackgroundAs a key subtype of non-coding RNAs, circular RNA (circRNA) has been well documented to play a key role in the tumorigenesis of osteosarcoma (OS). circPVT1 was revealed to participate in the progression of multiple human tumors; however, the roles of circPVT1 in OS invasion and metastasis and its potential mechanisms remain elusive.MethodsRNA expression in OS tissues and cells was examined by qRT-PCR, protein expression was measured by Western blot. circPVT1 knockdown in vitro was achieved by transfecting OS cells with specific siRNAs. OS cell proliferation was assessed via CCK-8 and colony formation assays. OS cell migration and invasion were evaluated by transwell assay. Interaction between miR-205-5p and circPVT1 or c-FLIP was validated through dual-luciferase reporter assay. Rescue experiments were performed to explore the regulatory net among circPVT1, miR-205-5p and c-FLIP in OS progression in vitro.ResultscircPVT1 and c-FLIP were highly expressed, while miR-205-5p was lowly expressed in OS tissues and cell lines. Knockdown of circPVT1 repressed cell proliferation, migration and invasion via inhibiting epithelial–mesenchymal transition (EMT) in OS. circPVT1 functioned as a sponge of miR-205-5p, and c-FLIP was targeted by miR-205-5p in OS cells. Furthermore, circPVT1 indirectly regulated c-FLIP expression through competitively binding to miR-205-5p. Inhibition of miR-205-5p or overexpression of c-FLIP abolished the effects of si-circPVT1 on cell proliferation, migration and invasion.ConclusionOur study demonstrated circPVT1 functions as a sponge for miR-205-5p to promote c-FLIP expression, thereby enhancing EMT and inducing OS invasion and metastasis in vitro, implying that circPVT1 might be a potential therapeutic target for further clinical therapy of OS.

Circular RNAs (circRNAs) have been identified as important regulators in cancer progression. Nevertheless, little is known about the biological function of circ_0000376 in the progression of osteosarcoma (OS). Cell viability, colony formation ability, apoptosis, and motility were analyzed by Cell Counting Kit-8 assay, colony formation assay, flow cytometry, and transwell assays. Cellular glycolytic metabolism was analyzed using commercial kits. RT-qPCR and Western blot assay were performed to analyze RNA and protein expression in OS tissues and cells. Starbase software was used to establish circRNA-microRNA (miRNA)-messenger RNA linkage, and intermolecular interaction was verified by dual-luciferase reporter assay. Xenograft tumor assay was conducted to analyze the effects of Tanshinone I (Tan I) and circ_0000376 on xenograft tumor growth in vivo. Tan I treatment suppressed the viability, migration, invasion, and glycolysis and triggered the apoptosis of OS cells. Tan I treatment markedly down-regulated circ_0000376 expression in OS cells. The addition of circ_0000376 plasmid largely rescued the malignant behaviors of OS cells upon Tan I exposure. Circ_0000376 interacted with miR-432-5p in OS cells. Circ_0000376 overexpression-mediated protective effects in Tan I-induced OS cells were partly attenuated by the accumulation of miR-432-5p. miR-432-5p bound to the 3' untranslated region (3'UTR) of B-cell leukemia/lymphoma 2 (BCL2) in OS cells. miR-432-5p interference-induced effects in Tan I-treated OS cells were partly overturned by the silence of BCL2. Circ_0000376 can act as miR-432-5p sponge to up-regulate BCL2 expression in OS cells. Circ_0000376 silencing contributed to the anti-tumor effect of Tan I on the growth of xenograft tumors in vivo. Tan I exerted an anti-tumor role in OS progression by targeting circ_0000376/miR-432-5p/BCL2 axis.

Loss of RanGAP1 drives chromosome instability and rapid tumorigenesis of osteosarcoma.

The aberrant expression of microRNAs (miRNAs/miRs) in osteosarcoma (OS) has been demonstrated in previous studies, and deregulation of miRNA expression has been associated with several types of cancer, including OS development and progression. Therefore, identifying the functional role of miRNAs in OS onset and development may facilitate the identification of novel and effective therapeutic targets for the treatment of patients with OS. Previous studies have demonstrated that miR-432 is involved in tumor formation and progression in several types of cancer. However, the expression pattern, functional role and underlying mechanism of miR-432 in OS remain unknown. In the current study, reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was used to measure miR-432 expression levels in OS tissue samples and cell lines. The effect of miR-432 overexpression on OS cell proliferation and invasion was detected using Cell Counting Kit-8 and Transwell invasion assays, respectively. Bioinformatics analysis was used to predict metastasis-associated in colon cancer 1 (MACC1) as a putative target of miR-432 and this was confirmed using a dual-luciferase reporter assay, RT-qPCR and western blot analysis. The current study demonstrated that miR-432 expression levels were significantly reduced in OS tissue samples and cell lines. In addition, functional assays revealed that overexpression of miR-432 significantly decreased OS cell proliferation and invasion. Furthermore, MACC1 was identified as a direct target gene of miR-432 in OS. MACC1 expression levels were significantly increased in OS tissue samples and an inverse correlation was observed between miR-432 and MACC1 expression in OS tissue samples. In addition, rescue experiments demonstrated that overexpression of MACC1 partially reversed the anti-proliferative and anti-invasive effects of miR-432 in OS cells. In conclusion, the present study demonstrated that miR-432 inhibited OS cell proliferation and invasion in vitro through direct targeting of MACC1, and miR-432 may be a potential therapeutic target for the treatment of OS.

Osteosarcoma (OS) is a type of malignant tumor arising from soft-tissues of bone and displays poor prognosis in most cases. However, the molecular mechanism by which OS initiates and progresses is still not completely elucidated. miR-155 has been shown to be overexpressed in OS specimen and cell lines. Our study is intended to explore the role of miR-155 in OS etiology. The data confirmed that miR-155 abundance is higher in OS samples than non-cancerous bone tissue. Inhibition of miR-155 suppressed the proliferation of OS cells and cell cycle progression in vitro, and the growth of OS xenografts in vivo. Wnt pathway was suppressed in OS cells by miR-155 inhibitors. HMG-box transcription factor 1 (HBP1), a strong Wnt pathway suppressor, was found to be a target of miR-155. Restoration of HBP1 abolished the effect of miR-155 on OS cells. Finally, miR-155 levels in OS tissues and serum are both inversely associated with the survival of OS patients. Collectively, miR-155 was identified to be among the list of OS-related oncogenic miRNAs, and HBP1-mediated Wnt signaling is involved with the role of miR-155 in OS progression.

BackgroundAccumulating articles demonstrate that circular RNAs play pivotal functions in tumorigenesis. However, the working mechanism of circ_0136666 in osteosarcoma (OS) progression remains to be further clarified.MethodsReal time-quantitative polymerase chain reaction and western blot assay were applied to determine RNA and protein expression, respectively. Cell proliferation was assessed by 5-Ethynyl-2′-deoxyuridine assay and colony formation assay. Transwell assays were carried out to assess cell migration and invasion abilities. Flow cytometry was performed to analyze cell apoptosis. Cell glycolysis was evaluated by analyzing the uptake of glucose and the production of lactate using the corresponding kits. Dual-luciferase reporter assay and biotinylated RNA-pull down assay were performed to confirm the target interaction between microRNA-1244 (miR-1244) and circ_0136666 or centrosomal protein 55 (CEP55). Xenograft tumor model was utilized to explore the role of circ_0136666 in tumor growth in vivo.ResultsCirc_0136666 expression was prominently elevated in OS tissues and cell lines. Circ_0136666 absence restrained the proliferation, migration, invasion and glycolytic metabolism and promoted the apoptosis of OS cells. Circ_0136666 negatively regulated miR-1244 expression by binding to it in OS cells. MiR-1244 overexpression suppressed the malignant behaviors of OS cells. CEP55 was a target of miR-1244 in OS cells. Circ_0136666 positively regulated CEP55 expression partly by sequestering miR-1244 in OS cells. CEP55 overexpression largely reversed circ_0136666 silencing-mediated influences in OS cells. Circ_0136666 silencing significantly suppressed tumor growth in vivo.ConclusionCirc_0136666 silencing inhibited OS progression partly by targeting miR-1244/CEP55 signaling. Silencing circ_0136666 and CEP55 or restoring miR-1244 level might be a potential therapeutic strategy for OS.

BackgroundAs a malignant tumor, the progression of osteosarcoma (OS) is mediated by multiple regulators, including circular RNAs (circRNAs). However, the role of circ_0000885 in OS is unclear.Materials and MethodsQuantitative real-time polymerase chain reaction (qRT-PCR) was applied to detect the expression of circ_0000885, miR-1294 and fibroblast growth factor receptor 1 (FGFR1). Cell proliferation was evaluated using 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyltetrazolium bromide (MTT) assay and colony formation assay. Flow cytometry and transwell assay were employed to determine the cell cycle distribution, cell migration and invasion, respectively. Moreover, the relationship between miR-1294 and circ_0000885 or FGFR1 was confirmed by dual-luciferase reporter assay. The protein level of FGFR1 was assessed via Western blot (WB) analysis. Animal experiments were used to verify the effect of circ_0000885 silencing on OS tumor growth in vivo.ResultsCirc_0000885 level was increased in OS tissues and cells. Knockdown of circ_0000885 repressed the proliferation, migration, invasion and induced cell cycle arrest in OS cells. There was a binding relationship between miR-1294 and circ_0000885, and miR-1294 inhibitor could reverse the inhibitory effect of silenced circ_0000885 on OS progression. MiR-1294 could target FGFR1, and overexpressed FGFR1 could invert the suppression effect of miR-1294 mimic on OS progression. Silencing of circ_0000885 hindered FGFR1 expression, while this effect could be recovered by miR-1294 inhibitor. In addition, circ_0000885 knockdown reduced OS tumor growth via regulating the FGFR1 expression by sponging miR-1294 in vivo.ConclusionCirc_0000885 played an active role in OS progression, indicating that it might be a potential target for OS therapy.