Biological Behavior Of Osteosarcoma Cells Research Articles

ARHGAP44-mediated regulation of the p53/C-myc/Cyclin D1 pathway in modulating the malignant biological behavior of osteosarcoma cells

ObjectiveOsteosarcoma is a rare primary malignant tumor of the bone characterized by poor survival rates, owing to its unclear pathogenesis. Rho GTPase-activating protein 44 (ARHGAP44), which belongs to the Rho GTPase-activating protein family, has promising applications in the targeted therapy of tumors. Therefore, this study aimed to investigate the biological function of ARHGAP44 in osteosarcoma and its possible application as a therapeutic target.MethodsThe expression level of ARHGAP44 in osteosarcoma and its relationship with tumor prognosis were detected using Gene Expression Omnibus database analysis and immunohistochemical staining of clinical specimens. The cell model of ARHGAP44 knockdown was constructed, and the effects of this gene on the malignant biological behavior of osteosarcoma cells were investigated using CCK-8, clone formation, transwell invasion, wound healing, and flow cytometry assays. Western blotting was performed to detect the expression of ARHGAP44, p53, C-myc, and Cyclin D1 in osteosarcoma.ResultsBiogenic analysis showed that ARHGAP44 was highly expressed in osteosarcoma. This result was associated with poor tumor prognosis and negatively correlated with the expression of the tumor suppressor gene p53. Immunohistochemistry and western blotting revealed significantly upregulated expression of ARHGAP44 in osteosarcoma tissues. Additionally, Kaplan–Meier analysis of clinical specimens suggested that ARHGAP44 was negatively correlated with tumor prognosis. CCK-8, clone formation, transwell invasion, wound healing, and flow cytometry assays showed that downregulation of ARHGAP44 expression significantly reduced the malignant biological behavior of osteosarcoma cells. Furthermore, western blotting showed that the expression level of p53 in osteosarcoma cells was significantly increased after the downregulation of ARHGAP44 expression, whereas the expression of C-myc and Cyclin D1 was significantly decreased compared with that in the control group.ConclusionARHGAP44 was highly expressed in osteosarcoma and was negatively correlated with its prognosis. The downregulation of ARHGAP44 expression reduced the malignant biological behavior of osteosarcoma cells. These findings suggest that the downregulation of ARHGAP44 expression inhibits the malignant progression of osteosarcoma by regulating the p53/C-myc/Cyclin D1 pathway, demonstrating the potential of ARHGAP44 as a therapeutic target for osteosarcoma.

Zyxin Inhibits the Proliferation, Migration, and Invasion of Osteosarcoma via Rap1-Mediated Inhibition of the MEK/ERK Signaling Pathway.

Zyxin (ZYX) is an actin-interacting protein with unknown biological functions in patients with osteosarcoma. This research sought to understand how ZYX affects the biological behavior of osteosarcoma cells and to identify the associated mechanism. Firstly, ZYX expression was decreased in osteosarcoma, and its higher expression indicated better outcomes in patients with osteosarcoma. ZYX overexpression significantly inhibited the proliferation, migration, and invasion of osteosarcoma cells, whereas ZYX silencing resulted in the opposite trend. Subsequently, we found that the Rap1 signaling pathway was significantly correlated with ZYX expression as reported in The Cancer Genome Atlas's database using bioinformatic analysis. Moreover, we found that ZYX overexpression regulated the Rap1/MEK/ERK axis, and osteosarcoma cell growth, migration, and invasion were consequently restrained. Additionally, by administering tumor cells subcutaneously to nude mice, a mouse model of transplanted tumors was created. Compared to the control group, the ZYX overexpression group's tumors were lighter and smaller, and the ZYX/Rap1 axis was activated in the ZYX overexpression group. Taken together, our results suggest that ZYX inhibits osteosarcoma cell proliferation, migration, and invasion by regulating the Rap1/MEK/ERK signaling pathway. ZYX might be crucial in the clinical management of osteosarcoma and is a promising novel therapeutic target in patients with this disease.

CD44 expression is correlated with osteosarcoma cell progression and immune infiltration and affects the Wnt/β-catenin signaling pathway

CD44 is associated with a variety of human diseases and plays a potential role in tumorigenesis, however, the mechanism of its role in osteosarcoma remains unclear. We analyzed the expression of CD44 in the Cancer Genome Atlas (TCGA) and genotype-tissue expression pan-cancer data and found that it was highly expressed in most tumors, including sarcoma. The expression of CD44 in osteosarcoma cell lines was higher than that in human osteoblast cell line in the results of the Western blot and Immunohistochemical staining assay. The results of colony formation assay and CCK 8 showed that CD44 improved the proliferation capacity of osteosarcoma cells, transwell assay and wound healing assay showed that CD44 improved the migration capacity of osteosarcoma cells. Further studies revealed that CD44 exerts its influence on the biological behavior of osteosarcoma cells through the Wnt/β-catenin signaling pathway. Since CD44 may be involved in the immune response, we analyzed the correlation between CD44 expression and immune cell infiltration in TCGA database using the previous cluster analyzer R software package, TIMER2.0 database and, GEPIA2 database, and found its involvement in the immune infiltration of osteosarcoma. Therefore, we believe that CD44 could be a potential target for the treatment of osteosarcoma patients and may be a candidate biomarker for immune infiltration-related prognosis.

Comprehensive multi-omics analysis reveals m7G-related signature for evaluating prognosis and immunotherapy efficacy in osteosarcoma

BackgroundOsteosarcoma is one of the most prevalent bone malignancies with a poor prognosis. The N7-methylguanosine (m7G) modification facilitates the modification of RNA structure and function tightly associated with cancer. Nonetheless, there is a lack of joint exploration of the relationship between m7G methylation and immune status in osteosarcoma. MethodsWith the support of TARGET and GEO databases, we performed consensus clustering to characterize molecular subtypes based on m7G regulators in all osteosarcoma patients. The least absolute shrinkage and selection operator (LASSO) method, Cox regression, and receiver operating characteristic (ROC) curves were employed to construct and validate m7G-related prognostic features and derived risk scores. In addition, GSVA, ssGSEA, CIBERSORT, ESTIMATE, and gene set enrichment analysis were conducted to characterize biological pathways and immune landscapes. We explored the relationship between risk scores and drug sensitivity, immune checkpoints, and human leukocyte antigens by correlation analysis. Finally, the roles of EIF4E3 in cell function were verified through external experiments. ResultsTwo molecular isoforms based on regulator genes were identified, which presented significant discrepancies in terms of survival and activated pathways. Moreover, the six m7G regulators most associated with prognosis in osteosarcoma patients were identified as independent predictors for the construction of prognostic signature. The model was well stabilized and outperformed traditional clinicopathological features to reliably predict 3-year (AUC = 0.787) and 5-year (AUC = 0.790) survival in osteosarcoma cohorts. Patients with increased risk scores had a poorer prognosis, higher tumor purity, lower checkpoint gene expression, and were in an immunosuppressive microenvironment. Furthermore, enhanced expression of EIF4E3 indicated a favorable prognosis and affected the biological behavior of osteosarcoma cells. ConclusionsWe identified six prognostic relevant m7G modulators that may provide valuable indicators for the estimation of overall survival and the corresponding immune landscape in patients with osteosarcoma.

DEPDC1 and KIF4A synergistically inhibit the malignant biological behavior of osteosarcoma cells through Hippo signaling pathway

The treatment of osteosarcoma (OS) is still mainly surgery combined with systematic chemotherapy, and gene therapy is expected to improve the survival rate of patients. This study aimed to explore the effect of DEP domain 1 protein (DEPDC1) and kinesin super-family protein 4A (KIF4A) in OS and understand its mechanism. Th expression of DEPDC1 and KIF4A in OS cells was detected by RT-PCR and western blot. The viability, proliferation, invasion and migration of OS cells and tube formation of human umbilical vein endothelial cells (HUVECs) after indicated treatment were in turn detected by CCK-8 assay, EdU staining, wound healing assay, transwell assay and tube formation assay. The interaction between DEPDC1 and KIF4A was predicted by STRING and confirmed by co-immunoprecipitation. The expression of epithelial-mesenchymal transition (EMT)-related proteins, tube formation-related proteins and Hippo signaling pathway proteins was detected by western blot. As a result, the expression of DEPDC1 and KIF4A was all increased in U2OS cells. Down-regulation of DEPDC1 suppressed the viability, proliferation, invasion and migration of U2OS cells and tube formation of HUVECs, accompanied by the increased expression of E-cadherin and decreased expression of N-cadherin, Vimentin and VEGF. DEPDC1 was confirmed to be interacted with KIF4A. Upregulation of KIF4A partially reversed the effect of DEPDC1 interference on the above biological behaviors of U2OS cells. Down-regulation of DEPDC1 promoted the expression of p-LATS1 and p-YAP in Hippo signaling pathway, which was reversed by upregulation of KIF4A. In conclusion, down-regulation of DEPDC1 inhibited the malignant biological behavior of OS cells through the activation of Hippo signaling pathway, which could be reversed by upregulation of KIF4A.

Up-regulation of NCAPG mediated by E2F1 facilitates the progression of osteosarcoma through the Wnt/β-catenin signaling pathway.

In recent years, there are few reports on non-SMC condensin I complex subunit G (NCAPG) in osteosarcoma. Our study aims to explore the biological role of NCAPG in osteosarcoma and its underlying molecular mechanism and to further clarify the reasons for the abnormal expression of NCAPG in osteosarcoma. Here, we mined The Cancer Genome Atlas (TCGA) Program public database through bioinformatics methods, analyzed the differential expression of NCAPG in sarcoma tissue and normal tissue, and explored the relationship between NCAPG expression level and sarcoma tissue differentiation, including tumor recurrence, metastasis, and patient survival. Next, the transcription factors responsible for the abnormal expression of NCAPG in osteosarcoma tumors were predicted by multiple online website tools and verified via cellular experiments. Subsequently, loss of function and cell phenotype experiments were performed to confirm the effect of NCAPG on the malignant biological behavior of osteosarcoma cells. Mechanistically, by reviewing the literature, we found that NCAPG can affect the malignant progression of many solid tumors by regulating the Wnt/β-catenin signaling pathway. Therefore, we preliminarily investigated the potential effect of NCAPG on this pathway via western blot experiments in osteosarcoma. Increased expression of NCAPG was found in sarcoma compared to normal tissues, which was positively correlated with poor differentiation, metastasis, and poor prognosis. Combining the transcription factor prediction results, correlation analysis, and expression level in the TCGA public database with validation outcomes of in vitro cell assays, we found that E2F transcription factor 1 (E2F1) regulated the increased expression of NCAPG in osteosarcoma. The results of cell phenotype experiments showed that silencing NCAPG could inhibit the proliferation, migration, and invasion of osteosarcoma cells. The preliminary mechanistic investigation suggested that NCAPG may affect osteosarcoma progression through the Wnt/β-catenin pathway. Our data reveal that E2F1 facilitates NCAPG expression in osteosarcoma by regulating the transcription of the NCAPG gene. Up-regulation of NCAPG promotes osteosarcoma progression via the Wnt/β-catenin signaling axis.

EFEMP1 binds to STEAP1 to promote osteosarcoma proliferation and invasion via the Wnt/β-catenin and TGF-β/Smad2/3 signal pathways

PurposeTo investigate the prognostic value and function of six-transmembrane epithelial antigen of prostate 1 (STEAP1) in osteosarcoma and determine whether EFEMP1 mediates its effects. MethodsIHC (immunohistochemistry)/ICC (immunocytochemistry) in conjunction with RT-qPCR (quantitative real-time polymerase chain reaction) were employed to assess the expression of STEAP1 in paratumoral tissues, osteosarcoma, benign fibrous dysplasia, osteosarcoma cells, normal osteoblastic hFOB cells, as well as various invasive subclones. The association of STEAP1 with outcome was examined with Kaplan–Meier graph among the osteosarcoma population. The effects of the down-regulation and up-regulation of STEAP1 on the biological behavior of osteosarcoma cells were studied through in-vitro and in-vivo functional tests. ResultsUp-regulation of STEAP1 in the osteosarcoma tissues, whose correlations with the malignant osteosarcoma phenotype and the poor patient outcome were positive. In addition, STEAP1 induced the epithelial–mesenchymal transition (EMT) via the Wnt/β-catenin and TGF-β/Smad2/3 pathways and facilitated the osteosarcoma cell infiltration and migration. An increase or decrease in EFEMP1 expression directly promoted or inhibited the expression of STEAP1. In osteosarcoma cells overexpressing EFEMP1, STEAP1 knockdown significantly inhibited cell invasion, EMT process, and increased activity of Wnt/β-catenin and TGF-β/Smad2/3 signaling pathways. Although exogenous EFEMP1 could stimulate the Wnt/β-catenin and TGF-β/Smad2/3 pathways to promote the EMT, it had not effect on osteosarcoma cells with STEAP1 knockdown. Collectively, similar to EFEMP1, STEAP1 acted like an oncogene in the osteosarcoma progression. ConclusionEFEMP1 enabled the Wnt/β-catenin and TGF-β/Smad2/3 axises initiation and EMT elicitation by targeting STEAP1, thereby facilitating the osteosarcoma cell infiltration and migration. These results are expected to contribute to the search for new targeted drugs able to effectively inhibit invasion and metastasis and improve prognosis in osteosarcoma.

Activation of PGRN/MAPK axis stimulated by the hypoxia-conditioned mesenchymal stem cell-derived HIF-1α facilitates osteosarcoma progression

BackgroundProgranulin (PGRN) is an important survival factor in the progression of multiple cancers. PurposeTo explore the effects and mechanisms of PGRN on malignant biological behavior of osteosarcoma (OS) cells and the effects of mesenchymal stem cells (MSCs) and the hypoxic microenvironment on PGRN alteration. Material and methodsThe expression pattern of PGRN in OS were evaluated in OS tissues and cell lines. Next, a loss-of-function assay investigated the function of PGRN on the proliferation, migration and cell death of OS cells. The activation of MAPK signaling in the process was examined by western blot and functional experiments accompanied by skatole. Additionally, we internally silenced hypoxia-inducible factor-1α (HIF-1α) in MSCs along with exogenously added HIF-1α (exo-HIF-1α) to explore how MSCs affect PGRN alteration and the malignant behavior of OS cells. ResultsAn aberrantly high expression of PGRN was observed in OS and associated with the poor prognosis of OS patients. PGRN knockdown repressed the proliferation, migration and induced cell death of OS cells, and activating MAPK pathway reversed these effects. Further evidence showed that MSCs regulated PGRN to mediate the malignant biological behavior of OS cells. Hypoxia enhanced HIF-1α expression in MSCs. HIF-1α silencing in MSCs under hypoxia suppressed the oncogenic effects of MSCs and reduced PGRN expression in OS cells, while the treatment of exo-HIF-1α reversed the depressive effects of HIF1α silencing on OS progression. ConclusionOverall, we concluded that PGRN, which was activated by the increase of hypoxic-MSCs-derived HIF-1α, promoted OS progression through the activation of MAPK signaling.

Long noncoding RNA HCP5 promotes osteosarcoma cell proliferation, invasion, and migration via the miR-29b-3p-LOXL2 axis.

Osteosarcoma (OS) is the second most common primary malignant bone tumors in adolescents that causes cancer-related deaths. Previous studies have confirmed the promoting role of lncRNA HCP5 in the development of OS, but the specific mechanism is still not well understood. MiRNA levels were measured via RT-qPCR and protein expression was detected via western blotting. Cell proliferation was analyzed by CCK-8 assays and colony formations assay were conducted to measure colony formation ability. Dual-luciferase reporter assay was performed to detect the targeting relationship between HCP5 and miR-29b-3p, and between miR-29b-3p and LOXL2. Wound healing assays and Transwell assays were conducted to verify the migration and invasion abilities of OS cells. Correlations between the levels of HCP5 and miR-29b-3p, and between miR-29b-3p and LOXL2 were determined by Pearson correlation coefficient analysis. MiR-29b-3p expression was decreased and HCP5 and LOXL2 levels were increased in OS tissues and cell lines. MiR-29b-3p could directly act on LOXL2 and knockdown of LOXL2 restrained the proliferation, migration, and invasion of OS cells. Moreover, transfection with sh-HCP5-1 and sh-HCP5-2 suppressed the malignant biological behavior of OS cells. HCP5 directly targeted miR-29b-3p, and promoted OS proliferation, migration, and invasion via the miR-29b-3p/LOXL2 axis. The lncRNA HCP5 may upregulate LOXL2 expression by targeting miR-29b-3p, thereby promoting OS proliferation, migration, and invasion.

Hsa_circ_0087302, a circular RNA, affects the progression of osteosarcoma via the Wnt/β-catenin signaling pathway.

Osteosarcoma is the most common malignant tumor in adolescent bone malignancies. It has the characteristics of a high metastasis rate, high mortality and poor prognosis. As a subclass of endogenous noncoding RNAs, circRNAs have been identified to be related to the occurrence, development and prognosis of different kinds of cancers, but the mechanism of their effect on osteosarcoma is not clear. In the present study, we identified a novel circRNA, hsa_circ_0087302, by RNA-seq, and we found that it was expressed at low levels in osteosarcoma. Using RT-PCR, we confirmed that the expression of hsa_circ_0087302 in osteosarcoma cells was lower than that in osteoblasts. Functional validation experiments revealed that hsa_circ_0087302 overexpression inhibited proliferation, cell cycle, migration, and invasion in osteosarcoma cells. Furthermore, Western blotting experiments demonstrated that hsa_circ_0087302 affected the expression of cell cycle- and Wnt/β-catenin signaling pathway-related proteins. For the first time, we identified that hsa_circ_0087302 may affect the malignant biological behavior of osteosarcoma cells through the Wnt/β-catenin signaling pathway. In summary, hsa_circ_0087302 may provide a new direction for the diagnosis and treatment of osteosarcoma.

MiR-139 inhibits proliferation, migration and invasion of osteosarcoma cell line MG63 via down-regulating integrin subunit alpha V(ITGAV)

ObjectivesOsteosarcoma is a relatively common primary malignant bone tumor in clinic, which frequently occurs in children and adolescents. It is essential to clarify the molecular mechanism of osteosarcoma to provide better diagnosis and treatment. Abnormal expression of miRNAs is closely related to the pathogenesis and progression of osteosarcoma. MiRNAs play a regulatory role in tumorigenesis and development of osteosarcoma. The purpose of this study is to reveal the working mechanism of miR-139/ITGAV axis in osteosarcoma progression. MethodsITGAV and miR-139 expression was detected in osteosarcoma tissues or paracancerous normal tissues. TargetScan and Double luciferase reporter gene assay were adopted to verify weather ITGAV was the target gene of miR-139. Western blot and qRT-PCR were used to evaluate the effects of miR-139 on ITGAV. CCK8, Flow cytometry, Transwell and Cell wound scratch assay were used to measure the effects of miR-139 and ITGAV on cell cycle, proliferation, migration and invasion of MG63, respectively. A nude mouse xenograft model of cervical cancer was constructed to observe the effects of miR-139 on the tumor growth. ResultsWe found that the expression of miR-139 in osteosarcoma tissue was significantly reduced, while the expression of ITGAV was significantly increased. MiR-139 could specifically bind to the 3'-UTR of ITGAV and negatively regulate its expression. Transfection of miR-139 mimic could inhibit the proliferation, S-phase arrest, invasion and migration of MG63 cells, and up-regulating the expression of ITGAV could reverse such inhibitory effect. In nude mouse xenograft model of osteosarcoma, overexpression of miR-139 could inhibit tumor growth, while down-regulation of miR-139 produced the opposite effect. ConclusionsThese results indicate that miR-139/ITGAV axis was related to osteosarcoma initiation. MiR-139 could inhibit the biological behavior of osteosarcoma cells and the tumor growth in nude mouse model via targeting ITGAV, and miR-139/ITGAV axis may impede the progression of osteosarcoma.

Suppression of circ_0008932 inhibits tumor growth and metastasis in osteosarcoma by targeting miR-145-5p.

Osteosarcoma (OS) is a common type of primary malignant tumor. Although the pathogenesis of OS has been extensively studied, the underlying molecular mechanisms have remained to be fully elucidated. Accumulating evidence has revealed that dysregulation of various circular RNAs (circRNAs) is associated with tumorigenesis and recent studies have indicated that circRNA circ_0008932 is aberrantly expressed in tumors. In the present study, the expression and detailed function of circ_0008932 in OS were elucidated. The levels of circ_0008932 in OS samples and cell lines were examined using reverse transcription-quantitative PCR. A cell model with circ_0008932 knockdown was generated using specific small interfering RNA (si-circ_0008932). Cell viability was determined by a Cell Counting Kit-8 assay, the cell migratory/invasive capacity was evaluated using Transwell assays and cell apoptosis was assessed by flow cytometry. The results suggested that circ_0008932 was upregulated in most primary OS tumors, suggesting that circ_0008932 is associated with the development of OS. In the in vitro assays, si-circ_0008932 inhibited the proliferation, migration and invasion of OS cells, while apoptosis was promoted. A luciferase reporter assay revealed that circ_0008932 may downregulate microRNA (miR)-145-5p through direct binding. Furthermore, the expression of miR-145-5p was negatively correlated with circ_0008932 levels in OS specimens. In addition, further functional studies indicated that miR-145-5p inhibitors eliminated the effects caused by si-circ_0008932 in OS cells. In comparison, the changes in the biological behavior of OS cells transfected with si-circ_0008932 were enhanced by miR-145-5p. In summary, circ_0008932 may be a novel oncogenic factor during the progression and development of OS by targeting miR-145-5p; more importantly, circ_0008932 may be a potential therapeutic target for OS.

Long non-coding RNA CCAT1 regulates the biological behavior of osteosarcoma cells through the miR-454-3p/ZEB2 axis.

The article "Long non-coding RNA CCAT1 regulates the biological behavior of osteosarcoma cells through the miR-454-3p/ZEB2 axis, by T. Su, S.-D. Zhang, J. Zhao, published in Eur Rev Med Pharmacol Sci 2020; 24 (21): 11016-11025-DOI: 10.26355/eurrev_202011_23586-PMID: 33215416" has been withdrawn from the authors stating that they "did not recognize the errors when reviewing the manuscript". The Publisher apologizes for any inconvenience this may cause. https://www.europeanreview.org/article/23586.

Long non-coding RNA CCAT1 regulates the biological behavior of osteosarcoma cells through the miR-454-3p/ZEB2 axis.

To investigate the clinical value and mechanism of action of lncRNA CCAT1 in OS. Serum, tumor tissue and corresponding adjacent tissue of 84 cases of patients receiving treatment in our hospital from February 2014 to June 2015 were obtained. Healthy volunteers were recruited during the same period for serum collection. CCAT1 expression in serum and tissue samples, were detected, and the value of its expression in osteosarcoma (OS) diagnosis and prognosis was analyzed. Stable and transient over-expression and inhibition vectors were established and transfected into OS cells. CCK-8, transwell, and flow cytometry were applied to determine the proliferation, invasion, and apoptosis of transfected cells, and the Dual-Luciferase reporter was utilized to determine the correlation of CCAT1 with miR-454-3p, miR-454-3p, and ZEB2 (zinc-finger E-box-binding homeobox-2). In OS, CCAT1 was upregulated, and serum CCAT1 could be used as a marker for OS diagnosis, with an AUC value of 0.930. High CCAT1 expression predicted poor survival rate in patients. Inhibition of CCAT1 could suppress the proliferation and invasion of OS cells, and increase the apoptosis rate. Over-expression of miR-454-3p and inhibition of ZEB2 could also achieve the above effects. Dual-Luciferase reporter indicated that CCAT1 could target miR-454-3p, and miR-454-3p could target ZEB2. The rescue experiment proved that CCAT1 could regulate OS progression through the miR-454-3p/ZEB2 axis. CCAT1 can be used as a diagnostic and prognostic marker for OS, promote OS cell proliferation and invasion, and inhibit apoptosis through the miR-454-3p/ZEB2 axis, which can be a therapeutic target for OS.

Long noncoding RNA plasmacytoma variant translocation gene 1 promotes epithelial-mesenchymal transition in osteosarcoma.

ObjectiveLong noncoding RNAs (lncRNAs) are involved in the proliferation, migration, and invasion of tumors. In the current study, our aim was to explore the role of lncRNA plasmacytoma variant translocation gene 1 (PVT1) in osteosarcoma.MethodsQuantitative real‐time reverse transcription‐polymerase chain reaction was used to detect the expression of lncRNA PVT1 in osteosarcoma tissues and cells. The relationship between lncRNA PVT1 expression status and the prognosis of patients with osteosarcoma was analyzed. The effect of lncRNA PVT1 on the malignant biological behavior of osteosarcoma cells in vitro was also analyzed.ResultsLncRNA PVT1 was upregulated in osteosarcoma. High lncRNA PVT1 expression indicated poor prognosis in patients with osteosarcoma. In vitro knockdown of lncRNA PVT1 inhibited the proliferation, migration, and invasion ability of osteosarcoma cells. In addition, we confirmed that lncRNA PVT1 affected the epithelial‐mesenchymal transition of osteosarcoma cells.ConclusionLncRNA PVT1 is a potential therapeutic target for osteosarcoma.

Effects of lncRNA TUSC7 on the malignant biological behavior of osteosarcoma cells via regulation of miR-375.

The present study aimed at investigating how long-chain non-coding RNA (lncRNA) tumor suppressor candidate 7 (TUSC7) regulates the malignant biological behavior of osteosarcoma cells. Tumor tissues and adjacent tissues of 30 patients with osteosarcoma were collected, and the expression levels of lncRNA TUSC7 and miR-375 were detected by RT-qPCR. lncRNA TUSC7 mimic and miR-375 mimic transfection models were established in MG63 osteosarcoma cells, and Transwell assays were used to detect the migration ability of MG63 cells. An MTT assay was used to assess the proliferation ability of MG63 cells. lncRNA TUSC7 in osteosarcoma tissue was significantly lower than that of adjacent tissues, while miR-375 levels were significantly higher than that of adjacent tissues; the two levels have a negative correlation. lncRNA TUSC7 mimic inhibited MG63 proliferation and migration abilities. miR-375 mimic promoted MG63 proliferation and migration abilities. The lncRNA TUSC7 mimic and miR-375 mimic co-transfection system could partially rescue the inhibition of lncRNA TUSC7 mimic on MG63 cells. In conclusion, lncRNA TUSC7 inhibited the proliferation and migration of MG63 osteosarcoma cells by regulating miR-375.

The Effects of MiR-214-3p and Irisin/FNDC5 on the Biological Behavior of Osteosarcoma Cells.

Background: Irisin/fibronectin type III domain-containing protein 5 (FNDC5) has important effects on breast cancer and liver cancer, however, its role in osteosarcoma is poorly understood. This study explored the effects of irisin/FNDC5 in osteosarcoma cells, aiming to provide a direction for treating osteosarcoma. Material and Methods: The expression levels of irisin/FNDC5 in serums and tissues of osteosarcoma patients and the expression characteristics of FNDC5 in osteosarcoma cell lines were measured. The effects of irisin, at different concentrations (0, 25, 50, 100, and 200 ng/mL), and FNDC5 on the viability, migration, and invasion of U2OS cells were analyzed. The target gene regulating FNDC5 was predicted, and its effects on irisin/FNDC5 and osteosarcoma cells were further explored. Results: The authors found that irisin/FNDC5 was significantly downregulated in the serums and tissues of osteosarcoma patients, and FNDC5 was also lowly expressed in osteosarcoma cell lines, especially in U2OS cells. Irisin/FNDC5 could not only inhibit the viability of U2OS cell in a concentration- and time-dependent manner but could also suppress cell migration and invasion. Furthermore, miR-214-3p inhibited the expression of irisin/FNDC5, and promoted the migration, invasion, and epithelial/mesenchymal transition (EMT) of U2OS cell through targeting FNDC5. Conclusions: Irisin/FNDC5 could inhibit the viability, migration, invasion, and EMT of osteosarcoma cells, and miR-214-3p could target FNDC5 to release its antitumor effects. Thus, irisin/FNDC5 and miR-214-3p might become a new direction for the treatment of osteosarcoma patients in the future.

Long non-coding RNA CDKN2B-AS1 promotes osteosarcoma by increasing the expression of MAP3K3 via sponging miR-4458.

Osteosarcoma (OS) is the most common primary malignant bone tumor worldwide. Recently, several studies have shown that the long non-coding RNA (lncRNA) CDKN2B-AS1 plays a critical role in several cancers. However, the function and underlying mechanism of CDKN2B-AS1 in OS development remains elusive. In this study, we firstly assessed the expression of CDKN2B-AS1 in OS tissues and cells, showing that CDKN2B-AS1 expression were remarkably upregulated in OS tissues and cells. Moreover, CDKN2B-AS1 knockdown suppressed cell proliferation, migration, and EMT progress in OS. Interestingly, we found and proved that CDKN2B-AS1 could sponge miR-4458 in OS cells. Moreover, MAP3K3 was certified as a downstream target of miR-4458 in OS. Besides, MAP3K3 was negatively regulated by miR-4458 and positively regulated by CDKN2B-AS1. More importantly, overexpression of MAP3K3 could partly counteract the effect of CDKN2B-AS1 suppression on the biological behavior of OS cells. Also, the in vivo experiments further testified that CDKN2B-AS1 accelerated tumor growth in OS. Our results suggested that CDKN2B-AS1 facilitated OS progression by sponging miR-4458 to enhance MAP3K3 expression, which provides a novel insight into improving diagnostic and therapeutic strategies for patients with OS.

Using Bioinformatics Analysis to Detect the Effect of Zoledronic Acid on the Biological Behavior of Osteosarcoma Cells

Objective: Osteosarcoma is a malignant bone tumor commonly seen in adolescents. Drug treatment for osteosarcoma is often accompanied by systemic toxicity and side effects, while zoledronic acid has few side effects but has anti-tumor effects. Methods: The bioinformatics analysis and scratch test were used to detect changes in cell proliferation, migration, and apoptosis in two osteosarcoma cell lines 24, 48, and 72 hours after adding zoledronic acid (0, 25, 50, 100, and 200 μM). Flow cytometry and transmission electron microscopy were used to observe the changes in cell apoptosis in the control and experimental groups after a 50% inhibitory dose of zoledronic acid was given. Results: The inhibition of cell proliferation and migration ability, as well as apoptosis increased with the increase in zoledronic acid exposure time and concentration. The 50% inhibitory rate occurred 48 hours after treatment with 100 M zoledronic acid. Conclusion: Zoledronic acid inhibited proliferation and migration and promoted apoptosis of osteosarcoma cells in vitro.

Downregulation of miR‑136 promotes the progression of osteosarcoma and is associated with the prognosis of patients with osteosarcoma

Osteosarcoma (OS) is the most common bone tumor in children and young adults, and is an aggressive tumor with poor prognosis. MicroRNAs (miRNAs) are aberrantly expressed in various types of cancer, and contribute to cancer tumorigenesis and progression. In the present study, the potential prognostic value and biological function of miRNA-136 (miR-136) in OS was investigated. Reverse transcription-quantitative polymerase chain reaction analysis was used to evaluate the expression of miR-136 in OS tissues and cell lines. Kaplan-Meier survival analysis and Cox regression analysis were conducted to investigate the prognostic significance of miR-136. Various in vitro cell based assays were used to evaluate the effects of miR-136 on the biological behavior of OS cells. A luciferase assay was performed to determine the key miR-136 targets associated with OS. The expression of miR-136 was significantly downregulated in osteosarcoma tissues and cells compared with the normal controls (all P<0.05). Decreased miR-136 expression was significantly associated with Enneking staging (P=0.030) and distant metastasis (P=0.016). Decreased miR-136 expression in patients was associated with shorter overall survival compared with patients with increased expression levels (log-rank test; P<0.05). The expression of miR-136 was indicated as an independent prognostic factor for the patients (hazard ratio=0.496; 95% confidence interval=0.250-0.987; P=0.046). MTT, transwell and Matrigel assays demonstrated that upregulation of miR-136 decreased proliferation, migration and invasion of OS cells. Bioinformatics and luciferase assays demonstrated that migration and invasion enhancer 1 (MIEN1) is a direct target of miR-136. Together, the results suggested that miR-136 functions as a tumor suppressor gene to regulate proliferation, migration and invasion of OS cells. MIEN1 was a potential target of miR-136. Additionally, miR-136 may serve as a prognostic biomarker for OS.