Objective:To investigate the anti-tumor effect of the Histone Deacetylase 6 (HDAC6) inhibitor ACY-738 and its underlying mechanisms in Diffuse Large B-cell Lymphoma (DLBCL) .Methods:The expression of HDAC6 in various tumors and DLBCL was analyzed using bioinformatics. DLBCL cells were treated with different concentrations of ACY-738. Cell viability, DNA synthesis, and clone formation were assessed by CCK-8 assay, EdU assay, and soft agar assay, respectively. Intracellular reactive oxygen species (ROS) levels were detected by fluorescence microscopy. Morphological changes in cells were observed using transmission electron microscopy (TEM). Mitochondrial ROS levels and apoptosis were measured by flow cytometry. The expression levels of apoptosis-related and autophagy-related proteins were detected by Western blotting.Results:HDAC6 was highly expressed in DLBCL ( P<0.05). ACY-738 inhibited the proliferation, DNA synthesis, and colony formation of DLBCL cells in a dose-dependent manner ( P<0.05). Treatment with ACY-738 increased intracellular and mitochondrial ROS levels in DLBCL cells in a dose-dependent manner ( P<0.05). TEM revealed that after ACY-738 treatment, mitochondria in cells were swollen and ruptured, mitochondrial cristae were reduced or absent, autolysosomes appeared, and features characteristic of apoptosis were observed. Western blotting showed that after ACY-738 treatment, the expression of the anti-apoptotic protein BCL-2 was downregulated, while the expression of Cleaved-PARP, Cleaved caspase-3, and BAX was upregulated ( P<0.05). The expression of autophagy-related proteins Atg7, Atg3, LC3B, and P62 was downregulated, and the expression of acetylated P53 protein was upregulated ( P<0.05) . Conclusion:The HDAC6 inhibitor ACY-738 induces mitochondria-dependent apoptosis and autophagy in DLBCL cells by acetylating P53, thereby inhibiting DLBCL cell proliferation.

Objective:To investigate the anti-tumor effect of the Histone Deacetylase 6 (HDAC6) inhibitor ACY-738 and its underlying mechanisms in Diffuse Large B-cell Lymphoma (DLBCL) .Methods:The expression of HDAC6 in various tumors and DLBCL was analyzed using bioinformatics. DLBCL cells were treated with different concentrations of ACY-738. Cell viability, DNA synthesis, and clone formation were assessed by CCK-8 assay, EdU assay, and soft agar assay, respectively. Intracellular reactive oxygen species (ROS) levels were detected by fluorescence microscopy. Morphological changes in cells were observed using transmission electron microscopy (TEM). Mitochondrial ROS levels and apoptosis were measured by flow cytometry. The expression levels of apoptosis-related and autophagy-related proteins were detected by Western blotting.Results:HDAC6 was highly expressed in DLBCL ( P<0.05). ACY-738 inhibited the proliferation, DNA synthesis, and colony formation of DLBCL cells in a dose-dependent manner ( P<0.05). Treatment with ACY-738 increased intracellular and mitochondrial ROS levels in DLBCL cells in a dose-dependent manner ( P<0.05). TEM revealed that after ACY-738 treatment, mitochondria in cells were swollen and ruptured, mitochondrial cristae were reduced or absent, autolysosomes appeared, and features characteristic of apoptosis were observed. Western blotting showed that after ACY-738 treatment, the expression of the anti-apoptotic protein BCL-2 was downregulated, while the expression of Cleaved-PARP, Cleaved caspase-3, and BAX was upregulated ( P<0.05). The expression of autophagy-related proteins Atg7, Atg3, LC3B, and P62 was downregulated, and the expression of acetylated P53 protein was upregulated ( P<0.05) . Conclusion:The HDAC6 inhibitor ACY-738 induces mitochondria-dependent apoptosis and autophagy in DLBCL cells by acetylating P53, thereby inhibiting DLBCL cell proliferation.

Objective To preliminarily investigate the anti-tumor effects of phytosphingosine(PHS)and the involvement of inducing apoptosis of leukemia cells.Methods Cellular model of leukemia was established in leukemia cell lines K562 and SUP-B15.CCK-8 assay and EdU assay were used to measure the viability and DNA synthesis of K562 and SUP-B15 cells.RNA-seq was carried out to verify the differentially expressed genes(DEGs)after PHS treatment.Gene Ontology(GO)enrichment and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analyses were applied to analyze the involved functions and signaling pathways.Comparative Toxicogenomics Database(CTD)and Discovery Studio software were employed to predict the underlying targets of PHS and molecular docking.Cell apoptosis was detected by flow cytometry,mitochondrial membrane potential was evaluated by JC-1 probe,and protein expression of key molecules was validated by Western blotting.Results PHS inhibited the proliferation of K562 and SUP-B15 cells in a time-and dose-dependent manner.The half-maximal inhibitory concentration(IC50)of K562 cells was 17.67 and 12.52 pmol/L for 24 and 48 h,respectively,and the IC50 value of SUP-B15 cells was 17.58 and 14.86 μmol/L for 24 and 48 h,respectively.PHS treatment at a dose of 20 μmol/L for 48 h resulted in significant inhibition of DNA synthesis.GO enrichment analysis of the K562 cells showed that PHS might be involved in positive regulation of apoptotic process,plasma membrane and its integral components,and protein kinase binding and activity.Reverse predictive analysis showed that BCL-2 protein was the most likely target of PHS.PHS significantly increased the apoptotic rate of leukemia cells(P＜0.05)in a dose-dependent manner,reduced the mitochondrial membrane potential,and down-regulated BCL-2 level(P＜0.05)and up-regulated the levels of Cleaved caspase-3 and Cleaved caspase-9(P＜0.05).Conclusion PHS may inhibit the proliferation of leukemia cells by inducing mitochondria-dependent apoptosis,possibly through PHS and BCL-2 interaction.

Objective: To study the role of miR-18a in regulating the proliferation, migration and invasion of ovarian cancer cells through IGF-2/Akt pathway. Methods: Ovarian cancer SKOV3 cells were cultured and grouped. The negative control group were not transfected with the plasmid, the blank plasmid group were transfected with the blank pcDNA3.1 plasmid, the IGF-2 plasmid group was transfected with the pcDNA3.1 plasmid expressing IGF-2, and the IGF-2 plasmid+L294002 group was transfected with pcDNA3.1 plasmid expressing IGF-2 and treated with Akt inhibitor LY294002. The upstream miRNA of IGF-2 was analyzed by bioinformatics method and verified by the double luciferase reporter gene method. The effect of overexpressing miR-18a on the expression of IGF-2 was observed. Results: Transfection of IGF-2 expression plasmid can increase the expression level of IGF-2 in cells and the content of IGF-2 in culture medium (P<0.05) . Cell proliferation in the IGF-2 plasmid group (0.93±0.22, F=9.629, P<0.05) , migration (80.22±13.28, F=12.689, P<0.05) , invasion (72.31±11.38, F=33.845, P<0.05) , the relative expression levels of p-PI3K (1.35±0.22, F=8.321, P<0.05) and p-Akt (0.94±0.18, F=9.612, P<0.05) . The proliferation, migration, invasion viability, in the viability and cells were significantly higher than those in the negative control group and the blank plasmid group. And relative expression levels of p-PI3K and p-Akt in the IGF-2 plasmid+L294002 group were significantly lower than those in the IGF-2 plasmid group (P<0.05) . IGF-2 is a target gene of miR-18a, and overexpression of miR-18a can down-regulate the expression level of IGF-2 (P<0.05) . Conclusion IGF-2 can promote the proliferation, migration, and invasion of ovarian cancer cells, and this promotion is related to the activation of the Akt pathway, and miR-18a may be an upstream regulatory molecule of IGF-2.