Journal: Frontiers in Oncology

Article Title: Knockdown of m6A Reader IGF2BP3 Inhibited Hypoxia-Induced Cell Migration and Angiogenesis by Regulating Hypoxia Inducible Factor-1α in Stomach Cancer

doi: 10.3389/fonc.2021.711207

Figure Lengend Snippet: IGF2BP3 knockdown hampered hypoxia-induced cell migration and angiogenesis in SC. (A) MKN-45 and HGC-27 cells were cultured in normoxic or hypoxic conditions for 24 h Next, the IGF2BP3 protein level was measured by western blot assay. (B) MKN-45 and HGC-27 cells were transfected with si-NC, si-IGF2BP3#1, si-IGF2BP3#2, or si-IGF2BP3#3. Next, the IGF2BP3 mRNA level was measured by RT-qPCR assay at 48 h after transfection. (C–F) MKN-45 and HGC-27 cells were transfected with si-NC or si-IGF2BP3#1 for 48 h and then maintained in hypoxic conditions for another 24 h Cells in the normoxia group were maintained in normoxic conditions for 72 h Cells in the hypoxia group were cultured in normoxia for 48 h and then exposed to hypoxia for an additional 24 h (C, D) Cell migratory potential was assessed by Transwell migration (C) and wound healing (D) assays. (E) VEGF level in cell culture supernatants was detected using a commercial kit. (F) The conditioned medium of MKN-45 and HGC-27 cells were collected after normoxia/hypoxia treatment or/and transfection. Next, HUVECs were cultured in a mixed medium of ECM and conditioned medium (volume ratio=1:1), followed by the measurement of tube formation ability at 12 h after incubation. * indicate that the difference is significant at 0.05 level. ** p < 0.01, *** p < 0.001, ## p < 0.01, ### p < 0.001 compared with the normoxia group.

Article Snippet: MKN-45 cells and human umbilical vein endothelial cells (HUVECs) were purchased from the China Center for Type Culture Collection (Wuhan, China).

Techniques: Knockdown, Migration, Cell Culture, Western Blot, Transfection, Quantitative RT-PCR, Incubation

Journal: Frontiers in Oncology

Article Title: Knockdown of m6A Reader IGF2BP3 Inhibited Hypoxia-Induced Cell Migration and Angiogenesis by Regulating Hypoxia Inducible Factor-1α in Stomach Cancer

doi: 10.3389/fonc.2021.711207

Figure Lengend Snippet: IGF2BP3 exerted its functions by up-regulating HIF1A. (A) MKN-45 and HGC-27 cells were transfected with pcDNA3.1 or pcDNA-HIF1A. Next, HIF1A mRNA level was measured by RT-qPCR assay at 48 h upon transfection. (B–E) MKN-45 and HGC-27 cells were transfected with si-NC+pcDNA3.1, si-IGF2BP3#1+pcDNA3.1, or si-IGF2BP3#1+pcDNA-HIF1A for 48 h and then maintained in hypoxic conditions for another 24 h, followed by the examination of cell migratory potential (B, C) and VEGF secretion level (D) . (E) MKN-45 and HGC-27 cells were transfected with si-NC+pcDNA3.1, si-IGF2BP3#1+pcDNA3.1, or si-IGF2BP3#1+pcDNA-HIF1A for 48 h and then maintained in hypoxic conditions for another 24 h, followed by the collection of conditioned medium. Next, HUVECs were cultured in a mixed medium of ECM and conditioned medium (volume ratio=1:1) and tube formation potential was examined at 12 h after incubation. ** p < 0.01, *** p < 0.001, ## p < 0.01, ### p < 0.001.

Article Snippet: MKN-45 cells and human umbilical vein endothelial cells (HUVECs) were purchased from the China Center for Type Culture Collection (Wuhan, China).

Techniques: Transfection, Quantitative RT-PCR, Cell Culture, Incubation

Journal: Nutrients

Article Title: JNK/p66Shc/ITCH Signaling Pathway Mediates Angiotensin II-induced Ferritin Degradation and Labile Iron Pool Increase

doi: 10.3390/nu12030668

Figure Lengend Snippet: Angiotensin II (Ang II) induces ferritin degradation. Immunoblotting analysis of ferritin H and L levels in lysates of ( a ) human umbilical vein endothelial cells (HUVEC), ( b ) HT22, and ( c ) CPAE cells treated with 0.4 μmol/L Ang II for ( a ) 4 h, ( b ) 1 h, and ( c ) 1 h. After probing with specific antibodies, the blots were stripped and reprobed with an anti-β-actin antibody to normalize for differences in protein loading. The results are representative of three independent experiments. ( d ) Immunoblotting analysis of ferritin L levels in a lysate of CPAE cells treated with 0.4 μmol/l Ang II for the indicated time periods. After probing with specific antibodies, the blots were stripped and reprobed with an anti-β-actin antibody to normalize for differences in protein loading. Data in the graph on the right are the mean ± SEM from three independent experiments, with values recalculated relative to the control; “a” indicates statistical significance at p < 0.05 (t-test).

Article Snippet: Human umbilical vein endothelial cells (HUVEC, 200-05N) were purchased from the European Collection of Authenticated Cell Cultures (Porton Down, UK) and cultured in endothelial cell growth medium (211–500).

Techniques: Western Blot, Control