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Long non-coding RNAs have the regulatory roles in different kinds of human cancers. The key point of this study was to research the functional mechanisms of urothelial carcinoma associated 1 (UCA1) in the development of osteosarcoma. Quantitative real-time PCR was adopted for the expression detection of UCA1, microRNA-513b-5p (miR-513b-5p) and E2F transcription factor 5 (E2F5). The target relation was verified via dual-luciferase reporter assay and RNA pull-down assay. Cell proliferation was evaluated using Cell Counting Kit-8 and colony formation assays. Transwell assay was applied to assess cell migration and invasion. Western blot was performed for protein examination. Xenograft experiment was used to explore the effect of UCA1 on osteosarcoma in vivo. UCA1 expression was enhanced while miR-513b-5p was refrained in osteosarcoma tissues and cells. MiR-513b-5p was a target of UCA1. Inhibition of UCA1 or overexpression of miR-513b-5p suppressed osteosarcoma cell proliferation, migration and invasion. E2F5 was identified as a downstream gene of miR-513b-5p. MiR-513b-5p inhibitor or E2F5 overexpression rescued the progression inhibition of osteosarcoma by UCA1 knockdown, and UCA1 regulated E2F5 and Cyclin E expression by targeting miR-513b-5p. Downregulation of UCA1 restrained the tumorigenesis of osteosarcoma in vivo through the miR-513b-5p/E2F5 axis. Collectively, knockdown of UCA1 inhibited tumorigenesis and metastasis of osteosarcoma via regulating the miR-513b-5p/E2F5 axis. UCA1 might be a biological indicator in the progression and treatment of osteosarcoma.

Background: Osteosarcoma (OS) is the most common primary bone malignancy, it has a dismal prognosis and mainly affects the children and adolescents. Previous reports have demonstrated that aberrantly expressed KIAA1429 plays crucial roles in the carcinogenesis of several cancers, but its expression status and functional role in the progression of OS have not previously been investigated.Methods: Immunohistochemistry (IHC) and western blotting were conducted to determine KIAA1429 expression status in OS. The relationship between KIAA1429 expression and OS prognosis was analyzed based on public database and tissue microarray (TMA). Cell proliferation ability was evaluated by CCK8, EdU and colony formation assays, and Transwell and wound healing potential were also assessed in vitro. Xenograft nude mouse model was performed to elucidate the tumor growth in vivo. The main specific miRNA targeting KIAA1429 in OS cells was identified.Results: KIAA1429 expression is markedly overexpressed in OS, and elevated KIAA1429 expression is significantly associated with an unfavorable prognosis. Functional investigations demonstrate that KIAA1429 silencing could attenuate proliferation, migration and invasion abilities of OS in vitro, as well as tumor growth in vivo. Mechanistically, microRNA-143-3p (miR-143-3p) was identified as the crucial specific mediator of KIAA1429 expression in OS cells. Furthermore, restoring KIAA1429 expression could partially reverse miR-143-3p mediated tumor-inhibition effects. Additionally, we found that knockdown of KIAA1429 or ectopic overexpression of miR-143-3p could repress stemness cell properties and the inhibition could be partly abolished by overexpression of KIAA1429.Conclusions: In summary, this study establishes miR-143-3p/KIAA1429 axis as promising therapeutic target for OS patients.

Background Circular RNAs (circRNAs) play vital roles in the modulation of tumor progression. This study explored the biological functions of circ_0001105 in the progression of osteosarcoma (OS). Methods qRT-PCR and in situ hybridization (ISH) were performed to detect the expression status of circ_0001105 in cells and tissues. Bioinformatics analysis, dual-luciferase reporter gene assay, Western blot and qRT-PCR were performed to determine the relationships among RNAs. The CCK-8, colony formation, EdU, transwell and wound healing assays were conducted to evaluate the cell growth, invasion and migration of OS cells. Tumor xenografts were established to investigate the effects of circ_0001105 on tumor growth in vivo. Lastly, the protein expression of YTHDF2 in OS tissues was measured using immunohistochemical staining. Results Data showed that circ_0001105 and YTHDF2 were significantly lower, while miR-766 was higher in OS tissues compared to adjacent tissues. Low expression of circ_0001105 or YTHDF2 was associated with poor survival of OS patients as demonstrated by the Kaplan-Meier analysis. In addition, miR-766 was identified as a direct binding target of circ_0001105 and YTHDF2. Ectopic overexpression of circ_0001105 or YTHDF2 significantly suppressed OS cell viability and invasion through regulating miR-766. Last, overexpression of circ_0001105 significantly attenuated in vivo tumor growth. Conclusion Our findings suggest that circ_0001105 inhibits OS progression, at least partially, by regulating miR-766/YTHDF2 signaling pathway.

Excess iron has been reported to lead to osteoblastic cell damage, which is a crucial pathogenesis of iron overload-related osteoporosis. However, the cytotoxic mechanisms have not been fully documented. In the present study, we focused on whether necroptosis contributes to iron overload-induced osteoblastic cell death and related underlying mechanisms. Here, we showed that the cytotoxicity of iron overload in osteoblastic cells was mainly due to necrosis, as evidenced by the Hoechst 33258/PI staining, Annexin-V/PI staining, and transmission electronic microscopy. Furthermore, we revealed that iron overload-induced osteoblastic necrosis might be mediated via the RIPK1/RIPK3/MLKL necroptotic pathway. In addition, we also found that iron overload was able to trigger mitochondrial permeability transition pore (mPTP) opening, which is a critical downstream event in the execution of necroptosis. The key finding of our experiment was that iron overload-induced necroptotic cell death might depend on reactive oxygen species (ROS) generation, as N-acetylcysteine effectively rescued mPTP opening and necroptotic cell death. ROS induced by iron overload promote necroptosis via a positive feedback mechanism, as on the one hand N-acetylcysteine attenuates the upregulation of RIPK1 and RIPK3 and phosphorylation of RIPK1, RIPK3, and MLKL and on the other hand Nec-1, siRIPK1, or siRIPK3 reduced ROS generation. In summary, iron overload induced necroptosis of osteoblastic cells in vitro, which is mediated, at least in part, through the RIPK1/RIPK3/MLKL pathway. We also highlight the critical role of ROS in the regulation of iron overload-induced necroptosis in osteoblastic cells.

Background Punicalagin (PUN) is a common anti-inflammatory polyphenol. However, the function and mechanism of PUN in osteoarthritis remains unknown. Methods Chondrocytes were isolated from rats, and confirmed by toluidine blue staining and immunofluorescence. Chondrocytes were challenged by lipopolysaccharide (LPS), and rat osteoarthritis model was established by Hulth method. The secretion of inflammatory factors, cell viability and apoptosis were tested via enzyme linked immunosorbent assay (ELISA), MTT and flow cytometry. The levels of forkhead box O1 (Foxo1), proteoglycan 4 (Prg4), hypoxia-inducible factor-3α (HIF3α), autophagy-related genes or extracellular matrix (ECM)-related proteins were examined via quantitative reverse transcription polymerase chain reaction (qRT-PCR), western blot or immunohistochemistry. The cartilage tissue damage was assessed via hematoxylin-eosin (HE) staining, toluidine blue staining and terminal dexynucleotidyl transferase (TdT)-mediated dUTP nick and labeling (TUNEL) staining. Results LPS triggered inflammatory injury in chondrocytes. PUN promoted autophagy to mitigate LPS-induced inflammatory injury. Foxo1 silence attenuated the effect of PUN on LPS-mediated autophagy inhibition and inflammatory injury. Promotion of Prg4/HIF3α axis abolished the influence of Foxo1 knockdown on LPS-mediated chondrocytes injury. PUN mitigated the inflammatory injury in rat osteoarthritis model by promoting autophagy and inhibiting inflammation and ECM degradation via Foxo1/Prg4/HIF3α axis. Conclusion PUN attenuates LPS-induced chondrocyte injury and osteoarthritis progression by regulating Foxo1/Prg4/HIF3α axis.

Hepatoblastoma (HB) is the most common hepatic neoplasm in childhood and the therapeutic outcomes remain undesirable due to its recurrence and metastasis. Increasing evidence shows that dipeptidase 1 (DPEP1) has pivotal function in tumorigenesis in multiple tumors. However, the expression pattern, biological function, and underlying mechanism of DPEP1 in HB have not been reported. Here we showed that DPEP1 was significantly upregulated and was associated with poor prognosis in HB patients. In vitro and in vivo assays indicated that silencing DPEP1 significantly suppressed HB cell proliferation, migration, and invasion, while DPEP1 overexpression exhibited the opposite effect. In addition, we identified that DPEP1 was a direct target of microRNA-193a-5p (miR-193a-5p). Functional experiments demonstrated that overexpression of miR-193a-5p significantly inhibited cell proliferation and invasion of HB cells, while the inhibitory effect could be reversed by DPEP1 overexpression. Moreover, miR-193a-5p was decreased in HB tumor tissues and associated with a poor clinical prognosis. Mechanistically, our results indicated that the miR-193a-5p/DPEP1 axis participated to the progression of HB via regulating the PI3K/Akt/mTOR (phosphatidylinositol-3-kinase/Akt/mammalian target of rapamycin) signaling. In conclusion, our findings suggest that the miR-193a-5p /DPEP1 axis might be a good prognostic predictor and therapeutic target in HB.

Background: Cervical cancer is one of the most common gynaecological malignancies. Emerging studies have documented that prolyl-4-hydroxylase α subunit 2 (P4HA2) is involved in multiple processes of cancer progression. However, the functional roles of P4HA2 in cervical cancer progression remain to be elucidated. Methods: P4HA2 mRNA and protein levels were examined in cervical cancer tissues and cell line by qRT-PCR and western blot. The correlation of the P4HA2 expression levels and prognosis of cervical cancer patients were analysed in TCGA cervical cancer cohort and tissue microarray (TMA) cohort. P4HA2 was silenced to evaluate its function on cervical cancer progression both in vitro and in vivo. Bioinformatics analysis was performed to investigate the underlying regulation mechanism of cervical cancer by P4HA2. Results: We found that P4HA2 are markedly upregulated in cervical cancer tissues in comparison with adjacent non-neoplastic tissues. In addition, upregulation of P4HA2 was associated with shorter overall survival (OS) and relapse-free survival (RFS). Functionally, we demonstrated that P4HA2 knockdown attenuated cell proliferation, migration and invasion of cervical cancer cells. Furthermore, xenograft tumor mouse model experiment showed silencing P4HA2 significantly inhibited tumor growth in vivo. Mechanistically, bioinformatics analysis revealed that epithelial-mesenchymal transition (EMT) was involved in cervical cancer progression regulated by P4HA2 and we further confirmed knockdown P4HA2 suppressed the EMT process. Conclusion: our results suggest that P4HA2 functions as an oncogene in promoting cervical cancer cell proliferation, migration and invasion by inducing EMT, which might be a promising prognostic factor and therapeutic target for cervical cancer.

Background: Mounting evidence has shown that long non-coding RNAs (lncRNAs) play critical regulation roles in the progression of various cancers. However, the biological role and clinical value of lncRNA FOXD2-AS1 in papillary thyroid cancer (PTC) remain to be elucidated. Methods: The expression of FOXD2-AS1 in PTC tissues and cell lines was evaluated by RT-qPCR and in situ hybridization. The association between FOXD2-AS1 expression levels and clinicopathologic features was analyzed through tissue microarray. The biological function of FOXD2-AS1 in PTC cells was determined both in vitro through CCK-8, EdU staining, colony formation and cell invasion assays and in vivo through a xenograft tumor model. Functional and pathway enrichment analysis were also conducted to analyze the molecular mechanism. Results: FOXD2-AS1 was significantly upregulated in PTC tissues, and high FOXD2-AS1 expression was positively associated with malignant potential factors in PTC patients. In addition, high level of FOXD2-AS1 expression was an unfavorable independent prognostic biomarker for patients with PTC. Moreover, we found that knockdown of FOXD2-AS1 could effectively inhibit PTC cell proliferation and invasion in vitro and suppress tumor growth of PTC in vivo. Bioinformatics analysis indicated that activation of cell cycle and apoptosis pathways might be involved in the oncogenic function of FOXD2-AS1 in PTC. Moreover, we demonstrated that FOXD2-AS1 directly interacted with miR-185-5p as miRNA sponge and overexpression of FOXD2-AS1 partially reversed the suppressive effect of miR-185-5p in TPC cells. Conclusion: Our findings suggest FOXD2-AS1 functions as an oncogene and promotes the tumor progression and metastasis in PTC, which might serve as a promising prognostic biomarker and potential therapeutic target for PTC patients.

Osteosarcoma is the most common type of primary malignant tumor of skeletal with poor prognosis in children and adolescents. Accumulating evidence indicates that CBX2 is overexpressed in multiple human neoplasm and play a critical role in tumorigenesis and progression. However, its functional role and upstream regulation mechanism in osteosarcoma remain unknown. In the present study, tissue microarray (TMA) analysis was performed to determine the association between CBX2 expression and clinical prognosis of osteosarcoma patients by immunohistochemistry. We also investigated the functional role of CBX2 using small interfering RNA (siRNA) in vitro and in vivo. Additionally, we confirmed the direct binding between CBX2 and let‐7a via qPCR, western blot and luciferase reporter assay. We found that CBX2 is dramatically upregulated in osteosarcoma tissues and high CBX2 expression was correlated with metastasis, recurrence, and chemotherapy response, as well as unfavorable prognosis in patients with osteosarcoma. Similar results were observed in a sarcoma cohort from The Cancer Genome Atlas (TCGA) dataset. Further experiments revealed that CBX2 knockdown significantly impeded osteosarcoma cell proliferation and invasion ability in vitro, and suppressed the tumor growth in tumor xenografts model. Mechanistically, we confirmed that CBX2 is a functional target of miRNA let‐7a. Overexpression of let‐7a inhibits osteosarcoma cell proliferation, which was reversed by CBX2 overexpression. Taken together, our study demonstrates that let‐7a/CBX2 plays a crucial role in osteosarcoma progression. CBX2 could serve as a promising prognostic biomarker and potential therapeutic target for osteosarcoma patients.

Objectives Gallbladder carcinoma (GBC) is the most highly aggressive cancer of biliary tract, but effective therapeutics are lacking. Emerging evidence has unveiled that miR-139-5p is aberrantly downregulated in cancers, including GBC. However, the functions and mechanisms of miR-139-5p in GBC remain unclear. Materials and methods MiR-139-5p-overexpression was established in GBC cell lines, after which cell proliferation, migration, invasion, colony formation, and glucose metabolism were assayed in vitro. Subsequently, bioinformatics prediction and dual-luciferase reporter were performed to confirm that pyruvate kinase M2 (PKM2) was a direct target of miRNA-139-5p. Xenograft mouse models were applied to investigate the role of miR-139-5p in GBC tumourigenicity in vivo. In situ hybridization and immunohistochemical assays were performed to determine the relationships among miR-139-5p, PKM2 expression and clinical malignancies in GBC samples. Results We found that miR-139-5p was substantially downregulated in GBC tissues. Low expression of miR-139-5p was significantly associated with poor clinical outcomes. GBC cell proliferation, migration, and invasion could be inhibited by overexpression of miR-139-5p either in vitro or in vivo. In addition, miR-139-5p overexpression could directly inhibit PKM2 expression and lead to suppression of glucose consumption, lactate production, and cellular ATP levels. Moreover, PKM2 was frequently upregulated in GBC and correlated with poor prognosis. Mechanistically, miRNA-139-5p inhibited cell proliferation, migration, and glycolysis in GBC, at least in part, by repressing PKM2. Conclusions These results demonstrated a novel role for miR-139-5p/PKM2 in GBC progression and provided potential prognostic predictors for GBC patients.

// Xin Wang 1, * , Yan Yu 2, * , Yuting He 2, * , Qiqing Cai 1 , Songtao Gao 1 , Weitao Yao 1 , Zhiyong Liu 1 , Zhichao Tian 1 , Qicai Han 3 , Weiwei Wang 4 , Ranran Sun 2 , Yonggang Luo 3 and Chao Li 1 1 Department of Bone and Soft Tissue, The Affiliated Cancer Hospital of Zhengzhou University, Henan Cancer Hospital, Zhengzhou 450008, China 2 Precision Medicine Center, The First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052, China 3 Key Laboratory of Clinical Medicine, The First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052, China 4 Department of Pathology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052, China * These authors have contributed equally to this work Correspondence to: Chao Li, email: lichao3882496@163.com Yonggang Luo, email: Luoyg_514@126.com Keywords: osteosarcoma; ALDH1B1; progression; proliferation; metastasis Received: August 21, 2017 Accepted: December 04, 2017 Published: December 20, 2017 ABSTRACT Osteosarcoma (OS) is the most common primary malignant bone tumor in childhood and adolescence with poor prognosis. The mechanism underlying tumorigenesis and development of OS is largely unknown. ALDH1B1 has been reported to involve in many kinds of human cancers and functions as an oncogene, but the role of ALDH1B1 in OS has not been investigated comprehensively. In the present study, we aimed to examine clinical value and biological function of ALDH1B1 in OS. Firstly, we investigated the roles of ALDH1B1 on an OS tissue microarray (TMA) as well as two OS cohorts from GEO database. We found that ALDH1B1 was significantly up-regulated in OS tissues and was independently associated with poor prognosis. Moreover, ALDH1B1 silencing could suppress the proliferation, migration, invasion in vitro and inhibit the growth of xenograft tumor and of OS cells in vivo . Additional, ALDH1B1 knockdown increased the apoptosis rate and lead to cell cycle arrest in G1 stage of OS cell in vitro . More importantly, the inhibition of ALDH1B1 expression could increase the sensitivity of OS cells to chemotherapy, which suggested that ALDH1B1 might be served as a therapeutic target to reverse drug resistance in chemotherapy in OS patients. Taken together, our founding suggested that ALDH1B1 contributes to OS tumor progression and drug resistance, which may represent a novel prognostic marker and potential therapeutic target for OS patients.