Molecular Biology and Genetics
Contributions
Type: Publication Only
Background
Flavonoids are reported to prevent the initiation, promotion and progression of cancer by making changes in various signaling pathways. Although several treatment options exist for acute promyelocytic leukemia (APL), it is not cured effectively due to the development of drug resistance and intolerance and its heterogenic nature. Thus, it is important to develop novel natural treatment approaches in order to improve outcome. Fisetin and hesperetin, bioactive flavonoids, found in fruits and vegetables have been reported to be promising novel antioxidants with their potentials as chemopreventive/chemotherapeutic agents in several cancer types such colon, breast and prostate cancers. However, there is no information about the precise mechanisms by which fisetin and hesperetin exert their antileukemic effects.
Aims
We intented to explain the molecular mechanisms and global gene expression patterns modulated by fisetin and hesperetin using genome-wide microarray analysis in APL cells.
Methods
Illumina Human HT-12v4 beadchip microarrays (San Diego, CA) were used to assess global gene expression. Poly-A tail mRNA isolated from fisetin and hesperetin treated HL60 APL cells was converted to biotin labelled c-RNA. The data obtained after hybridization of c-RNA to beadchips was analyzed using Illumina GenomeStudio software to cluster genes. The list of differentially expressed genes constrained by p-value <0.05 and at least 2.0 fold change was obtained. Affected genetic networks were determined by using the Kyoto Encyclopedia of Genes and Genomes (KEGG) and Ingenuity Pathway Analysis (IPA).
Results
A total of 54 and 1608 genes were significantly regulated (P<0.05) in 20 and 50 μM fisetin treated HL60 cells, respectively. Fold change analysis displayed that TXNIP, TFPI, miRNA1974, ID1 and ID3, HSPA1B and IDH1 were altered genes in both 20 and 50 μM fisetin-treated HL60 cells. On the other hand, MAP3K1, Caspase 4 and LASS6 were the examples of upregulated genes while LONP1, STAT5A and STAT3 and JAK1 were some of specifically downregulated genes in 50 μM fisetin treated HL60 cells. KEGG and IPA analysis displayed that MAPK, JAK/STAT and PI3K/AKT signaling pathways and ID signaling pathway were examples of the most altered networks. Moreover, HL60 cells treated with 100 and 150 μM hesperetin changed the expression of 130 and 691 genes (P<0.05), respectively. SASH1, MT1F and SPRR2D were common upregulated genes while TUBB1, ID3, ID1, NMU, FGFR3 and S100P were common dowregulated genes in both 100 and 150 μM hesperetin treated HL60 cells based on fold change analysis. Futhermore, 150 μM hesperetin induced more genes that were either upregulated or downregulated as compared to 100 μM hesperetin. TXNIP, MT1A, MAP3K1 and SPRR2F were some of upregulated genes while RPS25, C-MYC, TUBA1C were the examples of downregulated genes. ID signaling pathway, translation-, gluconeogenesis- and mitosis-related networks, TGF-β and MAPK pathways were the affected signaling networks based on KEGG and IPA analysis.
Summary
Our data show that fisetin and hesperetin trigger growth inhibitory and apoptotic effects by modulating the expression of genes involved in cellular processes. The genetic networks identified in this study enlight some important biological pathways that are altered by fisetin and hesperetin while giving a list of candidate genes that could be targeted for APL therapy. In conclusion, we determined the molecular mechanisms by which fisetin and hesperetin exert pleiotropic effects on APL cells.
Keyword(s): Acute promyelocytic leukemia, Gene expression profile, MAP kinase, Microarray analysis
Type: Publication Only
Background
Flavonoids are reported to prevent the initiation, promotion and progression of cancer by making changes in various signaling pathways. Although several treatment options exist for acute promyelocytic leukemia (APL), it is not cured effectively due to the development of drug resistance and intolerance and its heterogenic nature. Thus, it is important to develop novel natural treatment approaches in order to improve outcome. Fisetin and hesperetin, bioactive flavonoids, found in fruits and vegetables have been reported to be promising novel antioxidants with their potentials as chemopreventive/chemotherapeutic agents in several cancer types such colon, breast and prostate cancers. However, there is no information about the precise mechanisms by which fisetin and hesperetin exert their antileukemic effects.
Aims
We intented to explain the molecular mechanisms and global gene expression patterns modulated by fisetin and hesperetin using genome-wide microarray analysis in APL cells.
Methods
Illumina Human HT-12v4 beadchip microarrays (San Diego, CA) were used to assess global gene expression. Poly-A tail mRNA isolated from fisetin and hesperetin treated HL60 APL cells was converted to biotin labelled c-RNA. The data obtained after hybridization of c-RNA to beadchips was analyzed using Illumina GenomeStudio software to cluster genes. The list of differentially expressed genes constrained by p-value <0.05 and at least 2.0 fold change was obtained. Affected genetic networks were determined by using the Kyoto Encyclopedia of Genes and Genomes (KEGG) and Ingenuity Pathway Analysis (IPA).
Results
A total of 54 and 1608 genes were significantly regulated (P<0.05) in 20 and 50 μM fisetin treated HL60 cells, respectively. Fold change analysis displayed that TXNIP, TFPI, miRNA1974, ID1 and ID3, HSPA1B and IDH1 were altered genes in both 20 and 50 μM fisetin-treated HL60 cells. On the other hand, MAP3K1, Caspase 4 and LASS6 were the examples of upregulated genes while LONP1, STAT5A and STAT3 and JAK1 were some of specifically downregulated genes in 50 μM fisetin treated HL60 cells. KEGG and IPA analysis displayed that MAPK, JAK/STAT and PI3K/AKT signaling pathways and ID signaling pathway were examples of the most altered networks. Moreover, HL60 cells treated with 100 and 150 μM hesperetin changed the expression of 130 and 691 genes (P<0.05), respectively. SASH1, MT1F and SPRR2D were common upregulated genes while TUBB1, ID3, ID1, NMU, FGFR3 and S100P were common dowregulated genes in both 100 and 150 μM hesperetin treated HL60 cells based on fold change analysis. Futhermore, 150 μM hesperetin induced more genes that were either upregulated or downregulated as compared to 100 μM hesperetin. TXNIP, MT1A, MAP3K1 and SPRR2F were some of upregulated genes while RPS25, C-MYC, TUBA1C were the examples of downregulated genes. ID signaling pathway, translation-, gluconeogenesis- and mitosis-related networks, TGF-β and MAPK pathways were the affected signaling networks based on KEGG and IPA analysis.
Summary
Our data show that fisetin and hesperetin trigger growth inhibitory and apoptotic effects by modulating the expression of genes involved in cellular processes. The genetic networks identified in this study enlight some important biological pathways that are altered by fisetin and hesperetin while giving a list of candidate genes that could be targeted for APL therapy. In conclusion, we determined the molecular mechanisms by which fisetin and hesperetin exert pleiotropic effects on APL cells.
Keyword(s): Acute promyelocytic leukemia, Gene expression profile, MAP kinase, Microarray analysis