Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Senolytic Peptide FOXO4-DRI Selectively Removes Senescent Cells From in vitro Expanded Human Chondrocytes

doi: 10.3389/fbioe.2021.677576

Figure Lengend Snippet: Assessment of senescence level in PDL9 cells after FOXO4-DRI treatment. The samples with (FOXO4-DRI) or without (Control) FOXO4-DRI treatment were analyzed. (A) Detection of activated caspase-3/7. Red arrows indicate apoptotic cells. Bar = 100 μm. (B) SA-β-gal staining. Bar = 100 μm. (C) The relative cell number in PDL3 and PDL9 chondrocytes after FOXO4-DRI treatment was assessed using MTS assay. (D) Western blot. (E) Expression levels of selective genes. The data was normalized to that in Control group (set as 1). * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001.

Article Snippet: In this study, we used FOXO4 D-Retro-Inverso peptide (FOXO4-DRI, NovoPro, Shanghai, China) and ABT-263 (Apexbio Technology, Houston, TX, United States) to selectively remove the senescent chondrocytes.

Techniques: Control, Staining, MTS Assay, Western Blot, Expressing

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Senolytic Peptide FOXO4-DRI Selectively Removes Senescent Cells From in vitro Expanded Human Chondrocytes

doi: 10.3389/fbioe.2021.677576

Figure Lengend Snippet: Assessment of senescence level and chondrogenesis in engineered cartilage. PDL9 were pretreated with (FOXO4-DRI) or without (Control) FOXO4-DRI for 5 days and then subjected to 14 days of chondrogenic cultures. (A) Expression levels of selective genes. The data was normalized to that in the Control group (set as 1). (B) Western blot. (C) Safranin O/Fast green staining. Bar = 400, 200, and 100 μm from the left to right. * p < 0.05; ** p < 0.01.

Article Snippet: In this study, we used FOXO4 D-Retro-Inverso peptide (FOXO4-DRI, NovoPro, Shanghai, China) and ABT-263 (Apexbio Technology, Houston, TX, United States) to selectively remove the senescent chondrocytes.

Techniques: Control, Expressing, Western Blot, Staining

Journal: Experimental and Therapeutic Medicine

Article Title: Knockdown of FOXO4 protects against OGD/R‑induced cerebral microvascular endothelial cell injury and regulates the AMPK/Nrf2/HO‑1 pathway through transcriptional activation of CTRP6

doi: 10.3892/etm.2024.12382

Figure Lengend Snippet: Knockdown of FOXO4 enhances OGD/R-induced HCMEC/D3 cell viability. Expression of FOXO4 was detected by (A) RT-qPCR and (B) western blotting. *** P<0.001 vs. control. FOXO4 interference plasmid was transfected. (C) RT-qPCR and (D) western blotting were used to detect transfection efficiency. *** P<0.001 vs. si-NC. (E) Cell Counting Kit-8 was used to detect cell viability. *** P<0.001 vs. control; ## P<0.01 vs. OGD/R + si-NC. OGD/R, oxygen-glucose deprivation/reoxygenation; FOXO4, forkhead box protein O4; NC, negative control; si, small interfering; RT-q, reverse transcription-quantitative.

Article Snippet: siRNAs specific to FOXO4 (si-FOXO4#1 and si-FOXO4#2) or CTRP6 (si-CTRP6#1 and si-CTRP6#2), corresponding negative control (si-NC), pc-DNA3.1 vectors containing the complete sequence of FOXO4 [overexpression (Ov-)FOXO4] and empty vector (Ov-NC) were synthesized by Shanghai GenePharma Co., Ltd.

Techniques: Knockdown, Expressing, Quantitative RT-PCR, Western Blot, Control, Plasmid Preparation, Transfection, Cell Counting, Negative Control, Reverse Transcription

Journal: Experimental and Therapeutic Medicine

Article Title: Knockdown of FOXO4 protects against OGD/R‑induced cerebral microvascular endothelial cell injury and regulates the AMPK/Nrf2/HO‑1 pathway through transcriptional activation of CTRP6

doi: 10.3892/etm.2024.12382

Figure Lengend Snippet: Knockdown of FOXO4 promotes OGD/R-induced HCMEC/D3 cell apoptosis. (A) Apoptosis was detected by TUNEL assay. (B) Western blotting detected expression of apoptosis-associated proteins. *** P<0.001 vs. control; ## P<0.01, ### P<0.001 vs. OGD/R + si-NC. OGD/R, oxygen-glucose deprivation/reoxygenation; FOXO4, forkhead box protein O4; NC, negative control; si, small interfering.

Article Snippet: siRNAs specific to FOXO4 (si-FOXO4#1 and si-FOXO4#2) or CTRP6 (si-CTRP6#1 and si-CTRP6#2), corresponding negative control (si-NC), pc-DNA3.1 vectors containing the complete sequence of FOXO4 [overexpression (Ov-)FOXO4] and empty vector (Ov-NC) were synthesized by Shanghai GenePharma Co., Ltd.

Techniques: Knockdown, TUNEL Assay, Western Blot, Expressing, Control, Negative Control

Journal: Experimental and Therapeutic Medicine

Article Title: Knockdown of FOXO4 protects against OGD/R‑induced cerebral microvascular endothelial cell injury and regulates the AMPK/Nrf2/HO‑1 pathway through transcriptional activation of CTRP6

doi: 10.3892/etm.2024.12382

Figure Lengend Snippet: Knockdown of FOXO4 improves OGD/R-induced HCMEC/D3 cell barrier dysfunction. (A) FITC-Dextran kit was used to detect endothelial permeability. (B) Immunofluorescence detected expression of tight junction protein ZO-1. (C) Western blotting detected the expression of tight junction proteins ZO-1, occludin and claudin-5. *** P<0.001 vs. control; # P<0.05, ## P<0.01, ### P<0.001 vs. OGD/R + si-NC. OGD/R, oxygen-glucose deprivation/reoxygenation; FOXO4, forkhead box protein O4; NC, negative control; ZO-1, zonula occludens-1; si, small interfering.

Article Snippet: siRNAs specific to FOXO4 (si-FOXO4#1 and si-FOXO4#2) or CTRP6 (si-CTRP6#1 and si-CTRP6#2), corresponding negative control (si-NC), pc-DNA3.1 vectors containing the complete sequence of FOXO4 [overexpression (Ov-)FOXO4] and empty vector (Ov-NC) were synthesized by Shanghai GenePharma Co., Ltd.

Techniques: Knockdown, Permeability, Immunofluorescence, Expressing, Western Blot, Control, Negative Control

Journal: Experimental and Therapeutic Medicine

Article Title: Knockdown of FOXO4 protects against OGD/R‑induced cerebral microvascular endothelial cell injury and regulates the AMPK/Nrf2/HO‑1 pathway through transcriptional activation of CTRP6

doi: 10.3892/etm.2024.12382

Figure Lengend Snippet: Knockdown of FOXO4 alleviates OGD/R-induced oxidative stress and inflammation in HCMEC/D3 cells. (A) Levels of oxidative stress-associated indicators SOD, GSH-Px and ROS. (B) ELISA was used to detect levels of inflammatory cytokines in cells. *** P<0.001 vs. control; ## P<0.01, ### P<0.001 vs. OGD/R + si-NC. OGD/R, oxygen-glucose deprivation/reoxygenation; FOXO4, forkhead box protein O4; NC, negative control; SOD, superoxide dismutase, GSH-Px, glutathione peroxidase; ROS, reactive oxygen species; si, small interfering.

Article Snippet: siRNAs specific to FOXO4 (si-FOXO4#1 and si-FOXO4#2) or CTRP6 (si-CTRP6#1 and si-CTRP6#2), corresponding negative control (si-NC), pc-DNA3.1 vectors containing the complete sequence of FOXO4 [overexpression (Ov-)FOXO4] and empty vector (Ov-NC) were synthesized by Shanghai GenePharma Co., Ltd.

Techniques: Knockdown, Enzyme-linked Immunosorbent Assay, Control, Negative Control

Journal: Experimental and Therapeutic Medicine

Article Title: Knockdown of FOXO4 protects against OGD/R‑induced cerebral microvascular endothelial cell injury and regulates the AMPK/Nrf2/HO‑1 pathway through transcriptional activation of CTRP6

doi: 10.3892/etm.2024.12382

Figure Lengend Snippet: Knockdown of FOXO4 regulates CTRP6 transcription in HCMEC/D3 cells. (A) HDOCK SERVER database predicted the binding of FOXO4 to CTRP6. (B) Potential binding sequences between FOXO4 and CTRP6 promoter were predicted by JASPAR database. Expression of CTRP6 was detected by (C) RT-qPCR and (D) western blotting. ** P<0.01, *** P<0.001 vs. control. FOXO4 overexpression plasmid was transfected. (E) RT-qPCR and (F) western blotting detected the cell transfection efficiency. (G) RT-qPCR and (H) western blotting were used to detect the expression of FOXO4. * P<0.05, ** P<0.01, *** P<0.001 vs. Ov-NC; ### P<0.001 vs. si-NC. (I) Luciferase detection and (J) ChIP assay demonstrated the binding ability of FOXO4 and CTRP6 promoter. *** P<0.001 vs. Ov-NC. OGD/R, oxygen-glucose deprivation/reoxygenation; FOXO4, forkhead box protein O4; NC, negative control; CTRP6, C1q/tumor necrosis factor-related protein 6; ChIP, chromatin immunoprecipitation; si, small interfering; ov, overexpression; RT-q, reverse transcription-quantitative; WT, wild-type; MUT, mutant; TSS, Transcription start site.

Article Snippet: siRNAs specific to FOXO4 (si-FOXO4#1 and si-FOXO4#2) or CTRP6 (si-CTRP6#1 and si-CTRP6#2), corresponding negative control (si-NC), pc-DNA3.1 vectors containing the complete sequence of FOXO4 [overexpression (Ov-)FOXO4] and empty vector (Ov-NC) were synthesized by Shanghai GenePharma Co., Ltd.

Techniques: Knockdown, Binding Assay, Expressing, Quantitative RT-PCR, Western Blot, Control, Over Expression, Plasmid Preparation, Transfection, Luciferase, Negative Control, Chromatin Immunoprecipitation, Reverse Transcription, Mutagenesis

Journal: Experimental and Therapeutic Medicine

Article Title: Knockdown of FOXO4 protects against OGD/R‑induced cerebral microvascular endothelial cell injury and regulates the AMPK/Nrf2/HO‑1 pathway through transcriptional activation of CTRP6

doi: 10.3892/etm.2024.12382

Figure Lengend Snippet: Knockdown of FOXO4 regulates expression of CTRP6 to protect against OGD/R-induced HCMEC/D3 cell damage. CTRP6 interference plasmid was transfected. (A) Reverse transcription-quantitative PCR and (B) western blotting detected transfection efficiency. *** P<0.001 vs. si-NC. (C) Apoptosis was detected by TUNEL assay. (D) Western blotting detected expression of apoptosis-related proteins. *** P<0.001 vs. control; ### P<0.001 vs. OGD/R; @@ P<0.01, @@@ P<0.001 vs OGD/R + si-FOXO4 + si-NC. OGD/R, oxygen-glucose deprivation/reoxygenation; FOXO4, forkhead box protein O4; NC, negative control; CTRP6, C1q/tumor necrosis factor-related protein 6; si, small interfering.

Article Snippet: siRNAs specific to FOXO4 (si-FOXO4#1 and si-FOXO4#2) or CTRP6 (si-CTRP6#1 and si-CTRP6#2), corresponding negative control (si-NC), pc-DNA3.1 vectors containing the complete sequence of FOXO4 [overexpression (Ov-)FOXO4] and empty vector (Ov-NC) were synthesized by Shanghai GenePharma Co., Ltd.

Techniques: Knockdown, Expressing, Plasmid Preparation, Transfection, Reverse Transcription, Real-time Polymerase Chain Reaction, Western Blot, TUNEL Assay, Control, Negative Control

Journal: Experimental and Therapeutic Medicine

Article Title: Knockdown of FOXO4 protects against OGD/R‑induced cerebral microvascular endothelial cell injury and regulates the AMPK/Nrf2/HO‑1 pathway through transcriptional activation of CTRP6

doi: 10.3892/etm.2024.12382

Figure Lengend Snippet: Knockdown of FOXO4 regulates expression of CTRP6 to protect against OGD/R-induced HCMEC/D3 cell damage. (A) FITC-Dextran kit was used to detect endothelial permeability. (B) Immunofluorescence assay detected expression of tight junction protein ZO-1. (C) Western blotting detected expression of tight junction proteins ZO-1, occludin and claudin-5. (D) Levels of oxidative stress-associated indicators SOD, GSH-Px and ROS. (E) ELISA was used to detect levels of inflammatory cytokines in cells. *** P<0.001 vs. control; ### P<0.001 vs. OGD/R; @ P<0.05, @@ P<0.01, @@@ P<0.001 vs. OGD/R + si-FOXO4 + si-NC. OGD/R, oxygen-glucose deprivation/reoxygenation; FOXO4, forkhead box protein O4; NC, negative control; CTRP6, C1q/tumor necrosis factor-related protein 6; ZO-1, zonula occludens-1; si, small interfering; SOD, superoxide dismutase; GSH-Px, glutathione peroxidase; ROS, reactive oxygen species.

Article Snippet: siRNAs specific to FOXO4 (si-FOXO4#1 and si-FOXO4#2) or CTRP6 (si-CTRP6#1 and si-CTRP6#2), corresponding negative control (si-NC), pc-DNA3.1 vectors containing the complete sequence of FOXO4 [overexpression (Ov-)FOXO4] and empty vector (Ov-NC) were synthesized by Shanghai GenePharma Co., Ltd.

Techniques: Knockdown, Expressing, Permeability, Immunofluorescence, Western Blot, Enzyme-linked Immunosorbent Assay, Control, Negative Control

Journal: Experimental and Therapeutic Medicine

Article Title: Knockdown of FOXO4 protects against OGD/R‑induced cerebral microvascular endothelial cell injury and regulates the AMPK/Nrf2/HO‑1 pathway through transcriptional activation of CTRP6

doi: 10.3892/etm.2024.12382

Figure Lengend Snippet: Knockdown of FOXO4 regulates expression of CTRP6 to protect against OGD/R-induced HCMEC/D3 cell damage via the AMPK/Nrf2/HO-1 pathway. Western blotting detected expression of AMPK/Nrf2/HO-1 pathway-associated proteins p-AMPK, Nrf2 and HO-1. *** P<0.001 vs. control; ### P<0.001 vs. OGD/R; @ P<0.05, @@ P<0.01 vs. OGD/R + si-FOXO4 + si-NC. OGD/R, oxygen-glucose deprivation/reoxygenation; FOXO4, forkhead box protein O4; NC, negative control; CTRP6, C1q/tumor necrosis factor-related protein 6; HO-1, heme oxygenase-1; si, small interfering; p-, phosphorylated.

Article Snippet: siRNAs specific to FOXO4 (si-FOXO4#1 and si-FOXO4#2) or CTRP6 (si-CTRP6#1 and si-CTRP6#2), corresponding negative control (si-NC), pc-DNA3.1 vectors containing the complete sequence of FOXO4 [overexpression (Ov-)FOXO4] and empty vector (Ov-NC) were synthesized by Shanghai GenePharma Co., Ltd.

Techniques: Knockdown, Expressing, Western Blot, Control, Negative Control

Journal: Molecular Metabolism

Article Title: FOXO1 inhibition synergizes with FGF21 to normalize glucose control in diabetic mice

doi: 10.1016/j.molmet.2021.101187

Figure Lengend Snippet: Activities of FOXO1 inhibitors in HEK293 cell-based assays.

Article Snippet: Cells were transfected with pGL4.26-4xIRE-luc2 (FFluc), pRL-CMV (Promega), and pcDNA3.1 vector containing RFP (as negative control), FOXO1-AAA (Addgene, #9023), FOXA2 (GenScript, #OHu31644), FOXO3 (GenScript, #OHu23372), or FOXO4 (GenScript, #OHu23105) using Lipofectamine3000 (Invitrogen).

Techniques:

Journal: Oncotarget

Article Title: FOXO4 expression is related to stem cell-like properties and resistance to treatment in diffuse large B-cell lymphoma

doi: 10.18632/oncotarget.13690

Figure Lengend Snippet: ( A ) Study scheme of gene expression analyses using doxorubicin (Dox) and phenylbutyrate (PB)-treated surviving cells: Four different cell lines (BJAB, Raji, Toledo, and Daudi) are treated with doxorubicin (300 nM) or phenylbutyrate (8 mM) for 48 h, and cDNA microarray analysis is done to identify differentially expressed target genes between treatment-surviving cells and parental control cells. ( B ) FOXO4 mRNA level is significantly higher in phenylbutyrate-treated surviving (PB) cells of BJAB, Raji and Daudi than control cells. ( C ) Vorinostat-treated surviving cells show higher mRNA level of FOXO4 than control (Con) cells of BJAB and Raji. ( D ) Primary lymphoma cells from three patients with refractory diffuse large B-cell lymphoma (DLBCL) show increased expression of FOXO4 in phenylbutyrate-treated surviving cells (PB) compared to the corresponding control cells. ( E ) The FOXO4 mRNA level is significantly higher in primary cells from four patients with refractory DLBCL than that of a patient with DLBCL who achieved complete response. Data represent means ± SD from three independent experiments.

Article Snippet: FOXO4-Flag overexpression constructs were generated as follows: FOXO4 cDNA was obtained from Addgene (plasmid 17549) and subcloned into a pCMV tag4C vector to generate pCMV tag4C FOXO4-Flag.

Techniques: Expressing, Microarray

Journal: Oncotarget

Article Title: FOXO4 expression is related to stem cell-like properties and resistance to treatment in diffuse large B-cell lymphoma

doi: 10.18632/oncotarget.13690

Figure Lengend Snippet: ( A, B ) Phenylbutyrate-treated surviving cells (BJAB-PB and Raji-PB) increase the expression of FOXO4 and its transcriptional targets (p21, p27, and SOD) compared to control cells. ( C, D ) BJAB-PB and Raji-PB cells show decreased expression of cyclin D1, CDK4, and cyclin A compared to control cells.

Article Snippet: FOXO4-Flag overexpression constructs were generated as follows: FOXO4 cDNA was obtained from Addgene (plasmid 17549) and subcloned into a pCMV tag4C vector to generate pCMV tag4C FOXO4-Flag.

Techniques: Expressing

Journal: Oncotarget

Article Title: FOXO4 expression is related to stem cell-like properties and resistance to treatment in diffuse large B-cell lymphoma

doi: 10.18632/oncotarget.13690

Figure Lengend Snippet: ( A ) Soft agar colony formation in FOXO4-transfected or siFOXO4-transfected BJAB cells shows the amplification of FOXO4 increase colony forming ability and the knockdown of FOXO4 decrease colony formation in BJAB cell line. ( B ) Decrease in mRNA levels of Nanog, Oct-4 and Sox-2 in siFOXO4-transfected (siFOXO4) is noted compared to siControl-transfected (siCon) BJAB cells. ( C ) Phosphorylated AKT level is decreased in BJAB-PB cells with FOXO4 overexpression whereas siFOXO4-transfected BJAB-PB cells show the reverse of phosphorylated AKT. ( D ) The western blot shows the expression of FOXO4 protein in a BJAB clone (c2) overexpressing FOXO4. ( E ) Tumor sphere formation is observed from a BJAB clone (c2) overexpressing FOXO4. ( F ) Immunohistochemical staining for FOXO4 in tumor tissue of DLBCL (× 200). ( G ) Kaplan-Meier curves shows superior progression-free survival and overall survival of FOXO4-high group with diffuse large B cell lymphoma than FOXO4-low group. The P value is calculated using the log-rank test.

Article Snippet: FOXO4-Flag overexpression constructs were generated as follows: FOXO4 cDNA was obtained from Addgene (plasmid 17549) and subcloned into a pCMV tag4C vector to generate pCMV tag4C FOXO4-Flag.

Techniques: Transfection, Amplification, Over Expression, Western Blot, Expressing, Immunohistochemical staining, Staining

Journal: Oncotarget

Article Title: FOXO4 expression is related to stem cell-like properties and resistance to treatment in diffuse large B-cell lymphoma

doi: 10.18632/oncotarget.13690

Figure Lengend Snippet: Characteristics of patients

Article Snippet: FOXO4-Flag overexpression constructs were generated as follows: FOXO4 cDNA was obtained from Addgene (plasmid 17549) and subcloned into a pCMV tag4C vector to generate pCMV tag4C FOXO4-Flag.

Techniques: