Thymoquinone (TQ), the predominant bioactive constituent derived from the medicinal spice Nigella sativa (also known as black cumin), has been applied for medical purposes for more than 2,000 years. Recent studies reported that thymoquinone exhibited inhibitory effects on the cell proliferation of several cancer cell lines. This study was performed to investigate the antitumor and anti-angiogenic effects of thymoquinone on osteosarcoma in vitro and in vivo. Our results showed that thymoquinone induced a higher percentage of growth inhibition and apoptosis in the human osteosarcoma cell line SaOS-2 compared to that of control, and thymoquinone significantly blocked human umbilical vein endothelial cell (HUVEC) tube formation in a dose-dependent manner. To investigate the possible mechanisms involved in these events, we performed electrophoretic mobility shift assay (EMSA) and western blot analysis, and found that thymoquinone significantly downregulated NF-κB DNA-binding activity, XIAP, survivin and VEGF in SaOS-2 cells. Moreover, the expression of cleaved caspase-3 and Smac were upregulated in SaOS-2 cells after treatment with thymoquinone. In addition to these in vitro results, we also found that thymoquinone inhibits tumor angiogenesis and tumor growth through suppressing NF-κB and its regulated molecules. Collectively, our results demonstrate that thymoquinone effectively inhibits tumor growth and angiogenesis both in vitro and in vivo. Moreover, inhibition of NF-κB and downstream effector molecules is a possible underlying mechanism of the antitumor and anti-angiogenic activity of thymoquinone in osteosarcoma.

Background Circular RNAs (circRNAs) have been reported to play key roles in the development of various cancers. However, the biological functions and clinical significance of most circRNAs are still elusive. The purpose of this study was to explore the function and mechanism of a certain circRNA named circCDKN2B-AS1 in cervical cancer development and its potential value in the clinic. Methods qRT-PCR was used to verify the expression level of circCDKN2B-AS1. CCK-8, Transwell, and flow cytometry (FCM) assays were performed to detect cellular proliferation, migration, and apoptosis, respectively. A Seahorse XFe96 Analyzer was used to measure glycolysis metabolism level. RNA pull-down, RNA immunoprecipitation (RIP), actinomycin-D addition assays and Western blotting were used to screen and elucidate the potential mechanisms involved. BALB/c nude mice and zebrafish embryos (AB, WT) were used as animal models to investigate tumorigenesis capability. 18FDG-microPET/CT imaging and lactic acid (LA) and pyruvic acid (PA) content detection assays were used to detect the level of glucose metabolism in subcutaneous tumors from nude mice. Results CircCDKN2B-AS1, a circular isoform of the long noncoding RNA (lncRNA) CDKN2B-AS1, was upregulated in cervical cancer and precancerous tissues. We found that circCDKN2B-AS1 associated with the IMP3 protein depending on a specific binding site and regulated the stability of Hexokinase 2 (HK2) mRNA, the rate-limiting enzyme of the aerobic glycolysis pathway. The expression level of circCDKN2B-AS1 fated the binding of IMP3 to the 3′ untranslated region (UTR) of HK2 mRNA, consequently affecting the malignant cell phenotype and aerobic glycolysis in cervical cancer in vitro and in vivo. Mutant circCDKN2B-AS1, lacking the IMP3 binding site, did not have such effects. Utilization of an inhibitory peptide to block the interaction between circCDKN2B-AS1 and the IMP3 protein impeded the binding of IMP3 to the 3’UTR of HK2 mRNA and suppressed aerobic glycolysis in cervical cancer cells. Conclusions Our findings demonstrate that circCDKN2B-AS1 facilitates aerobic glycolysis by sponging the IMP3 protein to stabilize HK2 mRNA, consequently promoting the malignant phenotype in cervical cancer, which may provide a potential approach for cervical cancer therapeutics.

Engineering scaffolds combinging natural biomineral and artificially synthesized material hold promising potential for bone tissue regeneration. In this study, novel bioactive calcium sulfate/oyster shell (CS/OS) composites were prepared. Comparing to CS scaffold, the CS/OS composites with a controllable degradation rate displayed enhanced mineral nodule formation, higher alkaline phosphate (ALP) activity and increased proliferation rate while treated osteocytes. In CS/OS composites group, elevated mRNA levels of key osteogenic genes including bone morphogenetic protein-2 (BMP-2), runt-related transcription factor 2 (Runx2), osterix (Osx), and osteocalcin (OCN) were observed. Furthermore, The up-regulation of BMP-2 and type I collagen (COL-I) was observed for CS/OS composites relative to a CS group. Scaffolds were implanted into critical-sized femur cavity defects in rabbits to investigate the osteogenic capacity of the composites in vivo. The CS/OS scaffolds with proper suitable times and mechanical strength strongly promoted osteogenic tissue regeneration relative to the regeneration capacity of CS scaffolds, as indicated by the results of histological staining. These results suggest that the OS-modified CS engineering scaffolds with improved mechanical properties and bioactivity would facilitate the development of a new strategy for clinic bone defect regeneration.

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A range of biological and molecular effects caused by nicotine are considered to effect bone metabolism. Vitamin C functions as a biological antioxidant. This study was to evaluate the in vitro effects of nicotine on human bone marrow stromal cells and whether Vitamin C supplementation show the antagonism action to high concentration nicotine. We used CCK-8, alkaline phosphatase (ALP) activity assay, Von Kossa staining, real-time polymerase chain reaction and Western Blot to evaluate the proliferation and osteogenic differentiation. The results indicated that the proliferation of BMSCs increased at the concentration of 50, 100 ng/ml, got inhibited at 1,000 ng/ml. When Vitamin C was added, the OD for proliferation increased. For ALP staining, we found that BMSCs treated with 50 and 100 ng/ml nicotine showed a higher activity compared with the control, and decreased at the 1,000 ng/ml. Bone morphogenetic protein-2 (BMP-2) expression and the calcium depositions decreased at 100 and 1,000 ng/ml nicotine, while the addition of Vitamin C reversed the down regulation. By real-time PCR, we detected that the mRNA expression of collagen type I (COL-I) and ALP were also increased in 50 and 100 ng/ml nicotine groups (P < 0.05), while reduced at 1,000 ng/ml (P < 0.05). When it came to osteocalcin (OCN), the changes were similar. Taken all together, it is found that nicotine has a two-phase effect on human BMSCs, showing that low level of nicotine could promote the proliferation and osteogenic differentiation while the high level display the opposite effect. Vitamin C could antagonize the inhibitory effect of higher concentration of nicotine partly.