MiR-1207-5p/CX3CR1 axis regulates the progression of osteoarthritis via the modulation of the activity of NF-κB pathway.

BACKGROUND: Osteoarthritis (OA) is a prevalent chronic diseases characterized by formation of osteophytes and degradation of articular cartilage. Previous evidence has identified the regulatory effects of microRNAs (miRNAs) in OA. The goal of this study is to clearly explore the biological function of miR-1207-5p in OA. METHODS: MiR-1207-5p and C-X3-C motif chemokine receptor 1 (CX3CR1) expression in OA cartilages were revealed by accessing to Gene Expression Omnibus database. In vitro OA model was established by lipopolysaccharide (LPS) stimulation. Western blot and quantitative real-time polymerase chain reaction were conducted to detect the expression level of genes. Cell counting kit-8 (CCK-8) and flow cytometric experiments were performed to investigate the proliferation and apoptosis capacities of CHON-001 cells. Bioinformatics analysis was applied to predict the binding site of miR-1207-5p and CX3CR1, the connections of which were ascertained using luciferase reporter assay. RESULTS: MiR-1207-5p expression was decreased while CX3CR1 was increased in OA cartilages. Up-regulation of miR-1207-5p alleviated the LPS-induced damage in the view of cell proliferation, apoptosis and extracellular matrix (ECM) degradation. A target of miR-1207-5p CX3CR1, its down-regulation intensified the impacts of miR-1207-5p mimic, promoted proliferation and mitigated apoptosis. LPS exposure increased the protein expression of the phosphorylated IκBα and P65, and this phenomena was reversed due to miR-1207-5p up-regulation and CX3CR1 knockdown. The treatment of Betulinic acid (BA; an activator of nuclear factor-κB pathway) reversed the miR-1207-5p mimic-induced inhibitory effect on apoptosis in LPS-treated CHON-001. CONCLUSION: Our results highlight that miR-1207-5p can prevent CHON-001 from LPS-stimulated injury, providing a novel biomarker for OA progression and further advancing treatment of OA.

The inhibition of MARK2 suppresses cisplatin resistance of osteosarcoma stem cells by regulating DNA damage and repair.

OBJECTIVE: This study aims to explore the role of MARK2 in chemotherapeutic resistance and potential mechanism within cisplatin resistance models of CD133+ MG-63 and MNNG/HOS cells. METHODS: CD133- and CD133+ MG-63 and MNNG/HOS cells were differentiated and obtained by MACS(Magnetic bead sorting). Cell activity was determined by CCK-8 assay. siRNA was employed to down regulate the Microtubule Affinity Regulated Kinase 2 (MARK2) expression. Immunofluorescence detection and RT-qPCR were used to measure the expressions of MARK2 and DNA-PKcs at both protein and mRNA levels. Western blot was applied to test the levels of MARK2, γH2AX (S139), DNA-PKcs, Phospho-PI3 Kinase p85 (Tyr458), Akt, phospho-Akt (T308) antibodies, mTOR, phospho-mTOR (Ser2448). RESULTS: Compared with CD133- MG-63 cells, CD133+ MG-63 cells showed significantly strong cisplatin resistance, with high levels of MARK2, DNA-PKcs and potent DNA damage repair ability (p<0.05). Down regulation of MARK2 reduced the cisplatin resistance of CD133+ MG-63 cells, with deceasing expression of DNA-PKcs (p<0.05). PI3K/Akt/mTOR pathway was potentially activated in CD133+ MG-63 cells, and involved in the cisplatin resistance of MG-63 cells. The similar results were observed in CD133+ MNNG/HOS cells. The reduction of MARK2 retarded the activity of PI3K/Akt/mTOR pathway and further impeded the cisplatin resistance in CD133+ MG-63 and MNNG/HOS cell. CONCLUSION: Our data suggested that MARK2 was related to cisplatin resistance in CD133+ MG-63 and MNNG/HOS cells. The decrease of MARK2 restricted the cisplatin resistance of CD133+ MG-63 and MNNG/HOS cells by down regulating the expression of DNA dependent protein kinase catalytic subunit (DNA-PKcs) and inhibiting activity of PI3K/Akt/mTOR signaling pathway, which provides new clues for the osteosarcoma chemotherapy strategy.

Anti-tumoral potential of MDA19 in human osteosarcoma via suppressing PI3K/Akt/mTOR signaling pathway.

Osteosarcoma (OS) is the most common primary malignancy of skeleton with higher mortality rates amongst children and young adults worldwide, whereas effective and secure therapies have also been sought by researches with ongoing efforts. The purpose of the present study was to investigate the impact of N'-[(3Z)-1-(1-hexyl)-2-oxo-1,2-dihydro-3H-indol-3-ylidene] benzohydrazide (MDA19) on OS and explore its potential mechanism. Cell Counting Kit-8 (CCK8) and colony formation assay were employed to evaluate the potential effect of MDA19 on U2OS and MG-63 cells proliferation. Moreover, transwell migration and invasion assay were performed to assess the influence of MDA19 on U2OS and MG-63 cells migration and invasion. In addition, Annexin V-FITC/propidium iodide (Annexin V-FITC/PI) staining and flow cytometry were used to examine apoptotic ratio of the U2OS and MG-63 cells. Meanwhile, Western blot analysis was applied to explore change of relevant mechanism proteins in OS cells treated with MDA19. Our study showed that MDA19 had anti-proliferative activity of OS cells in a dose- and time-dependent manner, simultaneously, inhibition of colony formation was also observed in U2OS and MG-63 cells after incubation of MDA19. Besides, MDA19 could significantly inhibit the number of migrated and invaded OS cells and markedly increase the OS cells apoptosis rate. Mechanistically, we detected detectable reductions in apoptosis related proteins, epithelial-mesenchymal transition (EMT)-related proteins and activity of phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling in U2OS and MG-63 cells exposure to MDA19. Overall, the current study indicates in vitro anti-proliferative, anti-metastatic, and pro-apoptotic potential of MDA19 in U2OS and MG-63 cells. Our findings propose a clue for further studies with this compound in preclinical and clinical treatment for OS.

Cantharidin suppresses cell growth and migration, and activates autophagy in human non-small cell lung cancer cells.

Cantharidin (CTD), a component of Mylabris (blister beetle), is a traditional Chinese medicine that exerts an anticancer effect in multiple types of cancer cells. The aim of the present study was to investigate whether CTD exhibited anti-metastatic and inhibitory cell proliferation effects against human non-small cell lung cancer (NSCLC) A549 cells, and the possible underlying mechanism by which this occurs. The results of the present study demonstrated that CTD arrested proliferation, suppressed invasion and migration and induced apoptosis in A549 cells in vitro. Alterations of apoptosis-associated protein levels, including B-cell lymphoma-2 (Bcl-2), Bcl-associated X (Bax) and active caspase-3, were detected. Furthermore, the present study demonstrated that CTD activated autophagy through downregulation of p62 expression and upregulation of microtubule-associated proteins 1A/1B light chain 3B and Beclin-1 expression. Additionally, western blot analysis identified that CTD inhibited the phosphatidylinositol 3-kinase (PI3K)/RAC serine/threonine protein kinase (Akt)/mechanistic target of rapamycin (mTOR) signaling pathway in NSCLC, demonstrating that the levels of phosphorylated (p-)Akt, p-mTOR, phosphorylated ribosomal p70S6 protein kinase (p-p70-S6K) and cyclin D1 were significantly decreased following treatment with CTD. In conclusion, the results of the present study indicated that CTD impeded cell growth and migration by inhibiting PI3K/Akt/mTOR signaling in NSCLC, and promoted autophagy and apoptosis. CTD exhibited anticancer activity against NSCLC in vitro, revealing it as a potential candidate for the treatment of NSCLC.

Modeling osteosarcoma progression by measuring the connectivity dynamics using an inference of multiple differential modules algorithm.

Understanding the dynamic changes in connectivity of molecular pathways is important for determining disease prognosis. Thus, the current study used an inference of multiple differential modules (iMDM) algorithm to identify the connectivity changes of sub­network to predict the progression of osteosarcoma (OS) based on the microarray data of OS at four Huvos grades. Initially, multiple differential co­expression networks (M­DCNs) were constructed, and weight values were assigned for each edge, followed by detection of seed genes in M­DCNs according to the topological properties. Using these seed gene as a start, an iMDM algorithm was utilized to identify the multiple candidate modules. The statistical significance was determined to select multiple differential modules (M­DMs) based on the null score distribution of candidate modules generated using randomized networks. Additionally, the significance of Module Connectivity Dynamic Score (MCDS) to quantify the dynamic change of M­DMs connectivity. Further, DAVID was employed for KEGG pathway enrichment analysis of genes in dynamic modules. In addition to the basal condition, four conditions, OS grade 1­4, were also included (M=4). In total, 4 DCNs were constructed, and each of them included 2,138 edges and 272 nodes. A total of 13 genes were identified and termed 'seed genes' based on the z­score distribution of 272 nodes in DCNs. Following the module search, module refinement and statistical significance analysis, a total of four 4­DMs (modules 1, 2, 3 and 4) were identified. Only one significant 4­DM (module 3 in the DCNs of grade 1, 2, 3 and 4 OS) with dynamic changes was detected when the MCDS of real 4­DMs were compared to a null distribution of MCDS of random 4­DMs. Notably, the genes of the dynamic module (module 3) were enriched in two significant pathway terms, ubiquitin­mediated proteolysis and ribosome. The seed genes with the highest degrees included protein phosphatase 1 regulatory subunit 12A (PPP1R12A), UTP3, small subunit processome component homolog (UTP3), prostaglandin E synthase 3 (PTGES3). Thus, pathway functions (ubiquitin­mediated proteolysis and ribosome) and several seed genes (PPP1R12A, UTP3, and PTGES3) in the dynamic module 3 may be associated with the progression of OS and may serve as potential therapeutic targets in OS.