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Ferroptosis is a new-found iron-dependent form of non-apoptotic regulated cell death (RCD), which is activated on therapy with several antitumor agents, but the potential mechanism remains unclear. Erastin, exhibiting selectivity for RAS-mutated cancer cells, induces ferroptosis by increasing iron and lipid reactive oxygen species (ROS) levels in cell. Ferroportin (Fpn), the sole iron export protein, participates in the regulation of intracellular iron concentration. In this study, we investigated the role of Fpn on ferroptosis induced by erastin in SH-SY5Y cells.The cell viability was determined by CellTiter 96® AQueous Non-Radioactive Cell Proliferation Assay kit. The activity of caspase-3 was measured by ELISA kit. qRT-PCR was performed to examine the mRNA expression of Fpn. Western blot assay was conducted to examine the expression level of marker proteins. Specific commercial kits were used to examine the levels of MDA, ROS and iron in cells, respectively.Ferroptosis was evaluated by intracellular lipid ROS level and iron concentration. Hepcidin could prevent erastin-induced ferroptosis by degrading Fpn. Erastin (5 μg/mL) was observed to induce ferroptosis in neuroblastoma cells at 6 hours, which was promoted by knockdown of Fpn. The expression of Fpn gene and protein was decreased in SH-SY5Y cells treated with erastin. After treatment with erastin, Fpn siRNA transfection in SH-SY5Y cells was able to accelerate ferroptosis-associated phenotypic changes. Fpn acted as a negative regulator of ferroptosis by reducing intracellular iron concentration. Knockdown of Fpn enhanced anticancer activity of erastin.These results suggested that knockdown of Fpn accelerated erastin-induced ferroptosis by increasing iron-dependent lipid ROS accumulation, highlighting Fpn as a potential therapeutic target site for neuroblastoma. Thus, Fpn inhibitors may provide new access for chemosensitization of neuroblastoma.

SynopsisObjective Previous investigations suggested that cherry blossoms could provide valuable bioactive materials. However, few observations regarding the anti-inflammatory effect of cherry blossoms were reported. This study was to explore the anti-inflammatory effect of cherry blossom extract (CBE), which was used as a soothing ingredient in skincare product. Methods In vitro study, the anti-inflammatory effect of CBE on the nitric oxide (NO) inhibition assay in lipopolysaccharide (LPS)-treated RAW 264.7 cells was investigated. In vivo study, 40 volunteers were included in a randomized, single-blinded, placebo-controlled trial. 24-hour-occlusive test chambers were applied on the flexor side of the forearm with 3% sodium lauryl sulphate (SLS). Subsequently, the test areas were treated on 9 subsequent days with a cream containing 3% CBE or a placebo. Evaluation included a visual score and determination of erythema value (E value). Results In vitro study, 2% CBE reduced NO production by 31.83% compared to the placebo. In the SLS irritant patch test, the visual score and erythema value of CBE were lower than that of the placebo on D5 and D9. Conclusion Cherry blossom extract shows good anti-inflammatory effect in vitro and in vivo and represents a promising functional ingredient in soothing skincare product by reducing skin inflammation.

Apurinic/apyrimidinic endonuclease1 (APE1), which has the dual functions of DNA repair and redox regulation, is considered to be a promising potential target in cancer treatment. Microarray and qRT-PCR were used to confirm the change of miRNA followed by analysis with comprehensive bioinformatics-based analysis. Both microarray and qRT-PCR demonstrated that 13 microRNAs (miRNAs) were significantly changed (>2-fold) in APE1 knockdown HOS cells; seven of them (hsa-miR-451, hsa-miR-1290, hsa-miR-765, hsa-miR-483-5p, hsa-miR-513a-5p, hsa-miR-129-5p and hsa-miR-31) were up-regulated and the other six (hsa-miR-29b, hsa-miR-197, has-let-7b, hsa-miR-324-5p, hsa-let-7i and hsa-miR-484) were down-regulated. Furthermore, pathway analysis showed that these miRNAs and their target genes affected by the expression of APE1 were involved in pathways relating to developmental processes, regulation of cellular processes, cell signaling (such as TGF-β, Wnt, MAPK and the p53 signaling pathway) and cancers. There are putative binding sites of NF-κB, p53, HIF-1α, AP-1, PEBP2, ATF, NF-Y, Pax-2,CREB and c-Myb in the promoters of several down regulated miRNAs, indicating that APE1 may regulate miRNAs via transcription factors. Our data suggest that our understanding of the biological functions of APE1 will inevitably expand due to the novel pathways that APE1 uses to regulate gene expression through miRNAs.

The aim of this investigation was to determine whether tumour necrosis factor‐alpha (TNF‐α) has any effect on endothelial progenitor cells (EPCs). Total mononuclear cells were isolated from peripheral blood by Ficoll density gradient centrifugation, and then the cells were plated on fibronectin‐coated culture dishes. After 7 days culture, attached cells were stimulated with tumour necrosis factor‐α (final concentrations: 0, 10, 20, 50 and 100 mg/l) for 0, 6, 12, 24 and 48 h. EPCs were characterized as adherent cells double positive for DiLDL‐uptake and lectin binding, by direct fluorescence staining. EPC proliferation and migration were assayed using the MTT assay and modified Boyden chamber assay, respectively. EPC adhesion assay was performed by re‐plating those cells on fibronectin‐coated dishes, and adherent cells were counted. Tube formation activity was assayed using a tube formation kit. Levels of apoptosis were revealed using an annexin V apoptosis detection kit. Vascular endothelial growth factor Receptor‐1 (VEGF‐R1) and stromal derived factor‐1 (SDF‐1) mRNA, assessed by real‐time RT‐PCR inducible nitric oxide synthase (iNOS) and endothelial nitric oxide synthase (eNOS) were assayed by western blot analysis. Incubation of EPCs with tumour necrosis factor‐α reduced EPC proliferation, migration, adhesion, tube formation capacity, iNOS and eNOS in concentration‐ and time‐dependent manners. Tumour necrosis factor‐α reduced proliferation, migration, adhesion and tube formation capacity of EPCs. TNF‐α increased EPC apoptosis level, reduced VEGF‐R1 and SDF‐1 mRNA expression; tumour necrosis factor‐α also reduced iNOS and eNOS in the EPCs.