Journal: Oncology Reports

Article Title: Butyrate increases methylglyoxal production through regulation of the JAK2/Stat3/Nrf2/Glo1 pathway in castration‑resistant prostate cancer cells

doi: 10.3892/or.2024.8730

Figure Lengend Snippet: List of antibodies.

Article Snippet: The DU145 cells were transfected with pCMV2 control vector (CV013), pCMV2-Stat3-Myc (HG10034-CM), or pCMV2-Glo1-HA (HG12223-CY) (Sino Biological, Inc.) at 37°C for 24 h according to the manufacturer's protocol.

Techniques:

Journal: Oncology Reports

Article Title: Butyrate increases methylglyoxal production through regulation of the JAK2/Stat3/Nrf2/Glo1 pathway in castration‑resistant prostate cancer cells

doi: 10.3892/or.2024.8730

Figure Lengend Snippet: The effects of NaB on Stat1, JAK2 and Stat3 expression. (A) DU145 cells were treated with NaB (2.5 or 5 mM) for 48 h and the expression of these genes was determined. (B) The DU145 cells were incubated with ruxolitinib (0.5 µM) for 2 h followed by treatment with NaB for 48 h, the expression of p-Stat3 and Glo1 was determined. (C) The relative ROS was measured in cells treated with NaB (2.5 or 5 mM) for 48 h. (D) After the cells were incubated with NAC (5 mM) or medium for 2 h followed by treatment with NaB for another 48 h, the protein level of p-Stat3 was determined in various groups. (E and F) Cells were pretreated with Colivelin (10 µM) or medium for 2 h followed by treatment with NaB for another 48 h. Then, the expression of the (E) target genes and (F) MG-H1 production were determined. Results are expressed as the mean ± SEM. *P<0.05, **P<0.01 and ***P<0.001 vs. the Control group; # P<0.05, ## P<0.01 and ### P<0.001 vs. respective NaB-treated alone cells. NaB, sodium butyrate; Stat, signal transducer and activator of transcription; JAK2, Janus kinase 2; Glo1, glyoxalase1; ROS, reactive oxygen species; NAC, N-acetyl-cysteine; MG-H1, hydroimidazolone; Nrf2, nuclear factor erythroid 2-related factor 2; HO-1, heme oxygenase-1; p-, phosphorylated.

Article Snippet: The DU145 cells were transfected with pCMV2 control vector (CV013), pCMV2-Stat3-Myc (HG10034-CM), or pCMV2-Glo1-HA (HG12223-CY) (Sino Biological, Inc.) at 37°C for 24 h according to the manufacturer's protocol.

Techniques: Expressing, Incubation, Control

Journal: Oncology Reports

Article Title: Butyrate increases methylglyoxal production through regulation of the JAK2/Stat3/Nrf2/Glo1 pathway in castration‑resistant prostate cancer cells

doi: 10.3892/or.2024.8730

Figure Lengend Snippet: The effects of MGO, Glo1, Stat3 and CaMKII phosphorylation on NaB-mediated target gene expression and cell viability. (A) After treatment with MGO (0.2-1.6 mM) for 48 h, the expression of Nrf2, Glo1 and HO-1, as well as the cell viability were determined. (B) The cell viability of the DU145 cells with overexpression of Glo1 or Stat3 was examined. (C) Cells were pretreated with KN-93 (10 µM) for 2 h followed by treatment with NaB for 48 h. Then, the expression of MAPKs and pro-apoptotic proteins were determined. Results are expressed as mean ± SEM. *P<0.05, **P<0.01 and ***P<0.001 vs. the Vector or Control group; # P<0.05, ### P<0.001 vs. respective NaB-treated alone cells. MGO, methylglyoxal; Glo1, glyoxalase1; Stat3, signal transducer and activator of transcription 3; CaMKII, calcium/calmodulin dependent protein kinase II gamma; NaB, sodium butyrate; Nrf2, nuclear factor erythroid 2-related factor 2; HO-1, heme oxygenase-1; PARP, Poly (ADP-Ribose) polymerase.

Article Snippet: The DU145 cells were transfected with pCMV2 control vector (CV013), pCMV2-Stat3-Myc (HG10034-CM), or pCMV2-Glo1-HA (HG12223-CY) (Sino Biological, Inc.) at 37°C for 24 h according to the manufacturer's protocol.

Techniques: Targeted Gene Expression, Expressing, Over Expression, Plasmid Preparation, Control

Journal: Oncology Reports

Article Title: Butyrate increases methylglyoxal production through regulation of the JAK2/Stat3/Nrf2/Glo1 pathway in castration‑resistant prostate cancer cells

doi: 10.3892/or.2024.8730

Figure Lengend Snippet: Effects of KN-93 on cell proliferation the effects of NaB on normal RWPE-1 cells. (A) The relative cell proliferation of DU145 cells and the expression of c-Myc and cyclin D1 were evaluated in NaB (5 mM) alone, and co-treatment with KN-93, NAC, or Colivelin for indicated time. (B) The cell viability and the levels of Glo1 and Stat3 in RWPE-1 cells were examined after treatment with NaB for 48 h. Results are expressed as the mean ± SEM. **P<0.01 and ***P<0.001 vs. the Control group. (C) The proposed schematic diagram of NaB-mediated Nrf2/Glo1/MGO pathway and apoptosis in PCa cells. NaB inhibits the expression and activity of Nrf2 through PERK inactivation and accumulation of nuclear Bach1. Then, Glo1expression is attenuated via suppression of Janus kinase 2/Stat3 pathway, leading to accumulation of MGO and apoptotic cell death. The elevated MGO further downregulates Nrf2, Glo1 and HO-1. The CaMKII-mediated activation of MAPKs and Stat1 also contributes to NaB-induced cell death. NaB, sodium butyrate; NAC, N-acetyl-cysteine; Glo1, glyoxalase1; Stat3, signal transducer and activator of transcription 3; Nrf2, nuclear factor erythroid 2-related factor 2; MGO, cytotoxic methylglyoxal; PERK, eukaryotic translation initiation factor 2 alpha kinase 3; Bach1, BTB domain and CNC homolog 1; HO-1, heme oxygenase-1; CaMKII, calcium/calmodulin dependent protein kinase II gamma.

Article Snippet: The DU145 cells were transfected with pCMV2 control vector (CV013), pCMV2-Stat3-Myc (HG10034-CM), or pCMV2-Glo1-HA (HG12223-CY) (Sino Biological, Inc.) at 37°C for 24 h according to the manufacturer's protocol.

Techniques: Expressing, Control, Activity Assay, Activation Assay

Journal: Biology

Article Title: TERT Promoter Revertant Mutation Inhibits Melanoma Growth through Intrinsic Apoptosis

doi: 10.3390/biology11010141

Figure Lengend Snippet: The TERT promoter revertant mutation and Wnt/β-Catenin signaling. ( A ) PTEN transcription expression and gray value analysis in A375 −146T/T and cells with TERT promoter revertant mutations; ( B ) β-Catenin transcription expression and gray value analysis in A375 −146T/T and cells with TERT promoter revertant mutations; ( C ) c-MYC transcription expression and gray value analysis in A375 −146T/T and cells with TERT promoter revertant mutations. Student’s t -test. ** p < 0.01.

Article Snippet: Rabbit anti-ETS1 polyclonal antibody (#A00931; 1:1500); mouse anti-β-Actin antibody (BM5422; 1:1500); mouse anti-β-tubulin antibody (#BM3877; 1:1500); rabbit polyclonal anti-PTEN antibody (#PB0423; 1:1500); rabbit polyclonal anti-Catenin-β antibody (#BM3905; 1:1500); mouse anti-c-MYC antibody (#BM0238; 1:1500); rabbit anti-BLC2 monoclonal antibody (#BM4985; 1:1500); rabbit anti-BAX monoclonal antibody (#BM3964; 1:1500); and rabbit polyclonal anti-VDAC1 antibody (#BA3754; 1:1500) were obtained from Boster Biological Technology.

Techniques: Mutagenesis, Expressing

Journal: Advanced Science

Article Title: Phosphorylated SHMT2 Regulates Oncogenesis Through m 6 A Modification in Lung Adenocarcinoma

doi: 10.1002/advs.202307834

Figure Lengend Snippet: MAPK1 inhibition reduces the protein stability of SHMT2. A) A549 cells transfected with HA‐SHMT2 were separately treated with 1 µ m BDP5290, 1 µ m LY3214996, 1 µ m PIM447, 10 µ m seliciclib, 1 µ m tomivosertib, and 10 µ m DMAT for 24 h. The ubiquitination of SHMT2 was detected using western blotting. B) A549 cells transfected with HA‐SHMT2 were separately treated with 1 µ m LY3214996, 10 µ m seliciclib, 1 µ m tomivosertib, or 10 µ m DMAT. K48‐linked ubiquitination of SHMT2 was detected using western blotting. C) A549 cells transfected with the SHMT2 mutants were treated with 1 µ m LY3214996. K48‐linked ubiquitination of SHMT2 was detected using western blotting. D,E) H1299 (D) and A549 (E) cells were treated with 1 µ m LY3214996 for 24 h. Western blotting was used to detect protein expression. F) H1299 and A549 cells were treated with 1 µ m LY3214996 for 24 h. Q‐PCR was used to detect mRNA expression. The data represent the average of three independent experiments (mean ± SD). ns, p > 0.05. G) A549 cells were treated with CHX alone or with CHX or LY3214996 for the indicated times. Protein degradation was detected using western blotting (Top panel). The relative SHMT2 expression compared with that of β‐actin was quantified. The data represent the average of three independent experiments (mean ± SD). ns, p > 0.05; *, p < 0.05 (Bottom panels). H) A549 and H1299 cells transfected with HA‐SHMT2 were treated with 1 µ m LY3214996. Immunoprecipitation was used to concentrate HA‐SHMT2. The phosphorylation of SHMT2‐Ser90 was detected using an antibody specifically targeting SHMT2‐Ser90. I) A549 cells were transfected with the indicated plasmids and treated with or without LY3214996. Western blotting was used to detect protein ubiquitination. J) A549 cells were transfected with the indicated plasmid and siRNAs and then treated with or without LY3214996. Western blotting was used to detect protein ubiquitination.

Article Snippet: The pCMV3‐Myc‐MAPK1 (HG10030‐CM) and Myc‐MTHFD2 (HG16324‐CM) plasmids were obtained from SinoBiological.

Techniques: Inhibition, Transfection, Western Blot, Expressing, Immunoprecipitation, Plasmid Preparation

Journal: Advanced Science

Article Title: Phosphorylated SHMT2 Regulates Oncogenesis Through m 6 A Modification in Lung Adenocarcinoma

doi: 10.1002/advs.202307834

Figure Lengend Snippet: MAPK1 regulates the phosphorylation of SHMT2‐Ser90. A) A549 cells were transfected with MYC‐MAPK1. Western blotting was used to detect protein expression. B) A549 cells were transfected with MAPK1 siRNAs. Western blotting was used to detect protein expression. C) A549 cells were transfected with HA‐SHMT2. Immunoprecipitation was used to concentrate HA‐SHMT2. Western blotting was used to detect protein expression. D) A549 cells were transfected with MYC‐MAPK1. Immunoprecipitation was used to concentrate MYC‐MAPK1. Western blotting was used to detect protein expression. E) A549 cells were transfected with the indicated plasmids. Immunoprecipitation was used to concentrate HA‐SHMT2. Western blotting was used to detect protein expression. F) A549 cells were transfected with HA‐SHMT2 and MAPK1 siRNAs. Immunoprecipitation was used to concentrate HA‐SHMT2. Western blotting was used to detect protein expression. G) HA‐SHMT2 was concentrated from BEAS‐2B cells, and MYC‐MAPK1 was concentrated from A549 cells for use in an in vitro kinase assay. Western blotting was used to detect protein expression. H) A549 cells were transfected with HA‐SHMT2 and MAPK1 siRNAs. Immunoprecipitation was used to concentrate HA‐SHMT2, and western blotting was used to detect protein expression. I) A549 cells were transfected with control or MAPK1 siRNAs. The cells were treated with CHX for the indicated times, and western blotting was used to detect protein expression (Top panel). The relative SHMT2 expression compared with that of β‐actin was quantified. The data represent the average of three independent experiments (mean ± SD). #, siRNA1 versus Control; *,siRNA2 versus Control. ns, p > 0.05; *, p < 0.05; #, p < 0.05 (Bottom panels).

Article Snippet: The pCMV3‐Myc‐MAPK1 (HG10030‐CM) and Myc‐MTHFD2 (HG16324‐CM) plasmids were obtained from SinoBiological.

Techniques: Transfection, Western Blot, Expressing, Immunoprecipitation, In Vitro, Kinase Assay