Journal: Oncology Letters

Article Title: BRAF V600E -induced KRT19 expression in thyroid cancer promotes lymph node metastasis via EMT

doi: 10.3892/ol.2019.10360

Figure Lengend Snippet: Antibody list.

Article Snippet: The wild type (WT) BRAF WT (pcDNA3.1, cat. no. 40775; Addgene, Inc.) with its empty vector were transfected to the cells followed by Lipofectamine 2000 instruction and mutant BRAF V600E (pBabe, cat. no. 15269; Addgene, Inc.) with its empty vector were transfected into retrovirus packaging phoenix cells (with an VSV.G envelope plasmid; cat. no. 14888; Addgene, Inc.), after 48 h culturing the virus supernatant was harvested and filtered.

Techniques:

Journal: Oncology Letters

Article Title: BRAF V600E -induced KRT19 expression in thyroid cancer promotes lymph node metastasis via EMT

doi: 10.3892/ol.2019.10360

Figure Lengend Snippet: The TCGA_THCA dataset was used for the following analysis. (A) Compared with the BRAF wild type group, the expression of KRT19 was increased in BRAF V600E group, however, not in BRAF wild type group. The (B) wild type BRAF or (C) V600E mutated BRAF overexpression plasmids were transfected into 8505C cells, and may be overexpressed significantly compared with their empty vectors, BRAF WT /BRAF V600E . (D) mRNA and (E) protein expression levels of the BRAF V600E mutation group exhibited higher expression levels of KRT19 in 8505C cells when compared with BRAF wild type group. BRAF, B-Raf Proto-Oncogene, Serine/Threonine Kinase; TCGA, The Cancer Genome Atlas Network; THCA, thyroid cancer; KRT19, keratin 19; WT, wild type.

Article Snippet: The wild type (WT) BRAF WT (pcDNA3.1, cat. no. 40775; Addgene, Inc.) with its empty vector were transfected to the cells followed by Lipofectamine 2000 instruction and mutant BRAF V600E (pBabe, cat. no. 15269; Addgene, Inc.) with its empty vector were transfected into retrovirus packaging phoenix cells (with an VSV.G envelope plasmid; cat. no. 14888; Addgene, Inc.), after 48 h culturing the virus supernatant was harvested and filtered.

Techniques: Expressing, Over Expression, Transfection, Mutagenesis

Journal: Oncology Letters

Article Title: BRAF V600E -induced KRT19 expression in thyroid cancer promotes lymph node metastasis via EMT

doi: 10.3892/ol.2019.10360

Figure Lengend Snippet: The TCGA_THCA dataset was used for the following analysis. (A and D) Heatmap and GSEA analysis in EMT and BRAF V600E tumor vs. wild type BRAF tumor samples from the TCGA_THCA dataset. (B and C) Heatmap of EMT genes in the top 10 samples exhibiting high KRT19 expression vs. 10 samples with the lowest KRT19 expression from the TCGA dataset. (E) Western blot analysis of EMT markers in the 8505C cell line transfected with controls or si-KRT19. Knockdown of KRT19 was associated with increased E-cadherin and reduced N-cadherin levels when compared with the control group. BRAF, B-Raf Proto-Oncogene, Serine/Threonine Kinase; TCGA, The Cancer Genome Atlas Network; THCA, thyroid cancer; GSEA, Gene Set Enrichment Analysis; EMT, epithelial-mesenchymal-transition; KRT19, keratin-19; siRNA, small interfering RNA; FDR, false discovery rate; ES, enrichment score.

Article Snippet: The wild type (WT) BRAF WT (pcDNA3.1, cat. no. 40775; Addgene, Inc.) with its empty vector were transfected to the cells followed by Lipofectamine 2000 instruction and mutant BRAF V600E (pBabe, cat. no. 15269; Addgene, Inc.) with its empty vector were transfected into retrovirus packaging phoenix cells (with an VSV.G envelope plasmid; cat. no. 14888; Addgene, Inc.), after 48 h culturing the virus supernatant was harvested and filtered.

Techniques: Expressing, Western Blot, Transfection, Knockdown, Control, Small Interfering RNA

Journal: Oncology Letters

Article Title: BRAF V600E -induced KRT19 expression in thyroid cancer promotes lymph node metastasis via EMT

doi: 10.3892/ol.2019.10360

Figure Lengend Snippet: The TCGA_THCA dataset was used for the following analysis. (A) Compared with the BRAF wild type group, the expression of KRT19 was increased in BRAF V600E group, however, not in BRAF wild type group. The (B) wild type BRAF or (C) V600E mutated BRAF overexpression plasmids were transfected into 8505C cells, and may be overexpressed significantly compared with their empty vectors, BRAF WT /BRAF V600E . (D) mRNA and (E) protein expression levels of the BRAF V600E mutation group exhibited higher expression levels of KRT19 in 8505C cells when compared with BRAF wild type group. BRAF, B-Raf Proto-Oncogene, Serine/Threonine Kinase; TCGA, The Cancer Genome Atlas Network; THCA, thyroid cancer; KRT19, keratin 19; WT, wild type.

Article Snippet: The wild type (WT) BRAF WT (pcDNA3.1, cat. no. 40775; Addgene, Inc.) with its empty vector were transfected to the cells followed by Lipofectamine 2000 instruction and mutant BRAF V600E (pBabe, cat. no. 15269; Addgene, Inc.) with its empty vector were transfected into retrovirus packaging phoenix cells (with an VSV.G envelope plasmid; cat. no. 14888; Addgene, Inc.), after 48 h culturing the virus supernatant was harvested and filtered.

Techniques: Expressing, Over Expression, Transfection, Mutagenesis

Journal: Neural Regeneration Research

Article Title: Oncogenic BRAF V600E induces microglial proliferation through extracellular signal-regulated kinase and neuronal death through c-Jun N-terminal kinase

doi: 10.4103/1673-5374.361516

Figure Lengend Snippet: BRAF V600E expression leads to neuronal cell death and glial cell proliferation in primary mouse cortical mixed culture. Primary cortex mix cultures were prepared from C57BL/6J embryos. Cells were cultured in NB-A for 5 days, then transduced with viral vector, or BRAF WT , or BRAF V600E for 24 hours and then cultured in NB-A medium for 96 hours. (A) BRAF immunoblot and quantified BRAF expression level normalized to GAPDH. (B) Immunostaining for MAP2 and Iba1, and MAP2 + and Iba1 + cell counts in cultures transduced with lentiviral BRAF vectors. MAP2 (red, neurons), Iba1 (green, microglia), DAPI (blue, nuclei). (C) BRAF immunoblot and quantified BRAF expression level normalized to GAPDH. (D) Immunostaining for MAP2 and Iba1, and MAP2 + and Iba1 + cell counts in cultures transduced with retroviral (RV) BRAF vectors. MAP2 (red, neurons), Iba1 (green, microglia), DAPI (blue, nuclei). (E) Immunostaining for MAP2 and GFAP, and MAP2 + and GFAP + cell counting in cultures transduced with lentiviral BRAF vectors. MAP2 (green, neurons), GFAP (red, astrocytes), DAPI (blue, nuclei). (F) Immunostaining for MAP2 and GFAP, and MAP2 + cells and GFAP + cell counting in cultures transduced with RV BRAF vectors. The nuclei were counterstained by DAPI. MAP2 (green, neurons), GFAP (red, astrocytes), DAPI (blue, nuclei). Bars: 100 μm. ** P < 0.01, *** P < 0.001 (one-way analysis of variance and Tukey’s post hoc test). All experiments were repeated at least three times with at least three replicates under each condition. DAPI: 4′,6-Diamidino-2-phenylindole; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; MAP2: microtubule-associated protein 2; NB-A: neurobasal media.

Article Snippet: Constructs used were: pMD2.G (Addgene, Cat# 12259, RRID: Addgene_12259), psPAX2 (Addgene, Cat# 12260, RRID: Addgene_12260), pHAGE- BRAF V600E plasmid (Addgene, Cat# 116204, RRID: Addgene_116204), pHAGE- BRAF WT plasmid (Addgene, Cat# 116719, RRID: Addgene_116719), pBabe-Puro- BRAF V600E plasmid (Addgene, Cat# 15269, RRID: Addgene_15269), gag/pol-Retroviral plasmid (Addgene, Cat# 14887, RRID: Addgene_14887), Control siRNA (Cell Signaling Technology, Boston, MA, USA, Cat# 6568S), SAPK/JNK siRNA (Cell Signaling Technology, Cat# 6232S), ERK1 siRNA (Cell Signaling Technology, Cat# 6436S), ERK2 siRNA (Cell Signaling Technology, Cat# 6578S).

Techniques: Expressing, Cell Culture, Transduction, Plasmid Preparation, Western Blot, Immunostaining, Retroviral, Cell Counting

Journal: Neural Regeneration Research

Article Title: Oncogenic BRAF V600E induces microglial proliferation through extracellular signal-regulated kinase and neuronal death through c-Jun N-terminal kinase

doi: 10.4103/1673-5374.361516

Figure Lengend Snippet: BRAF V600E expression in astrocytes promotes proliferation. Primary astrocytes were prepared from C57BL/6J embryos. Cells were transduced with lentiviral vector, or BRAF WT , or BRAF V600E for 24 hours and then cultured in DMEM/F-12 medium for 96 hours. (A) Immunoblotting for BRAF and related signaling proteins and quantified expression levels normalized to GAPDH. (B) Flow cytometry analysis of cell cycle. (C) MTS cell viability assay at 48, 72, 96 and 120 hours after viral transduction. (D) Immunostaining for GFAP and Ki67, GFAP + cell counting, and % of Ki67 + astrocytes (scale bar: 100 μm). GFAP (red, astrocytes), DAPI (blue, nuclei), Ki-67 (green, proliferative cells). (E, F) qPCR analysis of inflammatory and antioxidant markers normalized to GAPDH. ** P < 0.01, *** P < 0.001 (one-way analysis of variance and Tukey’s post hoc test). All experiments were repeated at least three times with at least three replicates within each condition. DAPI: 4′,6-Diamidino-2-phenylindole; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GFAP: glial fibrillary acidic protein; qPCR: quantitative polymerase chain reaction.

Article Snippet: Constructs used were: pMD2.G (Addgene, Cat# 12259, RRID: Addgene_12259), psPAX2 (Addgene, Cat# 12260, RRID: Addgene_12260), pHAGE- BRAF V600E plasmid (Addgene, Cat# 116204, RRID: Addgene_116204), pHAGE- BRAF WT plasmid (Addgene, Cat# 116719, RRID: Addgene_116719), pBabe-Puro- BRAF V600E plasmid (Addgene, Cat# 15269, RRID: Addgene_15269), gag/pol-Retroviral plasmid (Addgene, Cat# 14887, RRID: Addgene_14887), Control siRNA (Cell Signaling Technology, Boston, MA, USA, Cat# 6568S), SAPK/JNK siRNA (Cell Signaling Technology, Cat# 6232S), ERK1 siRNA (Cell Signaling Technology, Cat# 6436S), ERK2 siRNA (Cell Signaling Technology, Cat# 6578S).

Techniques: Expressing, Transduction, Plasmid Preparation, Cell Culture, Western Blot, Flow Cytometry, Viability Assay, Immunostaining, Cell Counting, Real-time Polymerase Chain Reaction

Journal: Neural Regeneration Research

Article Title: Oncogenic BRAF V600E induces microglial proliferation through extracellular signal-regulated kinase and neuronal death through c-Jun N-terminal kinase

doi: 10.4103/1673-5374.361516

Figure Lengend Snippet: BRAF V600E expression in microglia induces cell proliferation and activation through ERK. Primary microglia cells were prepared from C57BL/6J embryos. Cells were transduced with lentiviral vector, BRAF WT and BRAF V600E for 24 hours and then cultured in DMEM/F12 for 96 hours. (A) Immunoblotting for BRAF and related signaling proteins and quantified expression levels normalized to GAPDH. (B, C) Cells were transfected with control-siRNA, or si-JNK, or si-ERK for 24 hours before transduction with BRAF viral vectors. Immunoblotting for ERK and JNK 48 hours after viral transduction and quantified expression levels normalized to GAPDH. (D) Cells were transfected with control-siRNA, or si-JNK, or si-ERK for 24 hours before transduction with BRAF viral vectors. Immunoblotting for BRAF-related signaling proteins and quantified expression levels normalized to GAPDH. (E, F) Immunostaining for Iba1 and Ki67, Iba1 + cell counts, and percentage of Ki67 + microglia (scale bar: 100 μm), and quantitative morphological analyses (percentage of ameboid-like microglia cells, length, area, length to area ratio in cells without and with siRNA transfection (scale bar: 50 μm). Iba1 (green, astrocytes), DAPI (blue, nuclei), Ki67 (red, proliferative cells). (G, H) Flow cytometry analysis of cell cycle. (I) MTS cell viability assay at 48, 72, 96 and 120 hours following viral transduction. (J) NO release in culture media by Griess reaction. (K) qPCR analysis of inflammatory and antioxidant markers in cells normalized to GAPDH. (L) IL-1β, IL-6 and TNF-α levels in culture medium measured by ELISA. Data are represented as mean ± SEM, n = 9. * P < 0.05, ** P < 0.01, *** P < 0.001 (one-way analysis of variance and Tukey’s post hoc test). All experiments were repeated at least three times with at least three replicates under each condition. ERK: Extracellular signal-regulated kinase; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; JNK: c-Jun N-terminal kinase; NO: nitric oxide; qPCR: quantitative polymerase chain reaction.

Article Snippet: Constructs used were: pMD2.G (Addgene, Cat# 12259, RRID: Addgene_12259), psPAX2 (Addgene, Cat# 12260, RRID: Addgene_12260), pHAGE- BRAF V600E plasmid (Addgene, Cat# 116204, RRID: Addgene_116204), pHAGE- BRAF WT plasmid (Addgene, Cat# 116719, RRID: Addgene_116719), pBabe-Puro- BRAF V600E plasmid (Addgene, Cat# 15269, RRID: Addgene_15269), gag/pol-Retroviral plasmid (Addgene, Cat# 14887, RRID: Addgene_14887), Control siRNA (Cell Signaling Technology, Boston, MA, USA, Cat# 6568S), SAPK/JNK siRNA (Cell Signaling Technology, Cat# 6232S), ERK1 siRNA (Cell Signaling Technology, Cat# 6436S), ERK2 siRNA (Cell Signaling Technology, Cat# 6578S).

Techniques: Expressing, Activation Assay, Transduction, Plasmid Preparation, Cell Culture, Western Blot, Transfection, Control, Immunostaining, Flow Cytometry, Viability Assay, Enzyme-linked Immunosorbent Assay, Real-time Polymerase Chain Reaction

Journal: Neural Regeneration Research

Article Title: Oncogenic BRAF V600E induces microglial proliferation through extracellular signal-regulated kinase and neuronal death through c-Jun N-terminal kinase

doi: 10.4103/1673-5374.361516

Figure Lengend Snippet: Conditioned medium from BRAF V600E -expressing microglial cells but not astrocytes induces neuronal cell death. Primary cortex neurons were prepared from C57BL/6J embryos and cultured for 5 days, and the original medium was then replaced with conditional medium for 72 hours. (A, B) Immunostaining for MAP2 (scale bar: 50 μm) and MAP2 + cell counting. (C) LDH release in neurons treated with conditioned medium from primary astrocytes transduced with lentiviral BRAF vectors. (D–F) Immunostaining for MAP2 (scale bar: 50 μm; D) and MAP2 + cell counting in neurons treated with conditioned medium from primary microglia transduced with lentiviral BRAF vectors with and without siRNA transfection (E, F). (G, H) LDH release from neurons treated with conditioned medium from primary microglia transduced with lentiviral BRAF vectors with and without siRNA transfection. * P < 0.05, *** P < 0.001 (one-way analysis of variance and Tukey’s post hoc test). All experiments were repeated at least three times with at least three replicates within each condition. MAP2 (green, neurons), DAPI (blue, nuclei) in A and D. DAPI: 4′,6-Diamidino-2-phenylindole; LDH: lactate dehydrogenase; MAP2: microtubule-associated protein 2.

Article Snippet: Constructs used were: pMD2.G (Addgene, Cat# 12259, RRID: Addgene_12259), psPAX2 (Addgene, Cat# 12260, RRID: Addgene_12260), pHAGE- BRAF V600E plasmid (Addgene, Cat# 116204, RRID: Addgene_116204), pHAGE- BRAF WT plasmid (Addgene, Cat# 116719, RRID: Addgene_116719), pBabe-Puro- BRAF V600E plasmid (Addgene, Cat# 15269, RRID: Addgene_15269), gag/pol-Retroviral plasmid (Addgene, Cat# 14887, RRID: Addgene_14887), Control siRNA (Cell Signaling Technology, Boston, MA, USA, Cat# 6568S), SAPK/JNK siRNA (Cell Signaling Technology, Cat# 6232S), ERK1 siRNA (Cell Signaling Technology, Cat# 6436S), ERK2 siRNA (Cell Signaling Technology, Cat# 6578S).

Techniques: Expressing, Cell Culture, Immunostaining, Cell Counting, Transduction, Transfection

Journal: Neural Regeneration Research

Article Title: Oncogenic BRAF V600E induces microglial proliferation through extracellular signal-regulated kinase and neuronal death through c-Jun N-terminal kinase

doi: 10.4103/1673-5374.361516

Figure Lengend Snippet: BRAF V600E expression in neurons promotes cell death through the JNK pathway. Primary cortex neurons were prepared from C57BL/6J embryos. Cells were cultured for 5 days, transduced with lentiviral vector, BRAF WT and BRAF V600E for 24 hours, and then cultured in NB-A for 96 hours. (A) Immunoblotting for BRAF and related signaling proteins and quantified expression levels normalized to GAPDH. (B) qPCR analysis c-Jun, Bax, Bcl-2, p53, Fasl and TNF-α normalized to GAPDH. (C–E) Immunostaining for MAP2 (scale bar: 50 μm), MAP2 + cell counting, and LDH release. (F) Primary cortex neurons were transfected with control-siRNA or si-JNK for 24 hours before transduction with BRAF viral vectors. Immunoblotting for BRAF and related signaling proteins and quantified expression levels normalized to GAPDH. (G) qPCR analysis of c-Jun, Bax, Bcl-2, p53, Fasl and TNF-α normalized to GAPDH. (H) Immunostaining for MAP2 (sale bar: 50 μm), (I) MAP2 + cell counting, (J) and LDH release. Primary cortex neurons were transfected with control-siRNA or si-ERK for 24 hours before transduction with BRAF viral vectors. (K) Immunoblotting for BRAF and related signaling proteins and quantified expression levels normalized to GAPDH. (L) qPCR analysis of c-Jun, Bax, Bcl-2, p53, Fasl and TNF-α normalized to GAPDH. (M–O) Immunostaining for MAP2 (scale bar: 50 μm), MAP2 + cell counting, and LDH release. * P < 0.05, ** P < 0.01, *** P < 0.001 (one-way analysis of variance and Tukey’s post hoc test). All experiments were repeated at least three times with at least three replicates within each condition. MAP2 (green, neurons), DAPI (blue, nuclei) in C, H, and M. DAPI: 4′,6-Diamidino-2-phenylindole; ERK: extracellular signal-regulated kinase; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; JNK: c-Jun N-terminal kinase; LDH: lactate dehydrogenase; MAP2: microtubule-associated protein 2; NB-A: neurobasal media; qPCR: quantitative polymerase chain reaction; TNF-α: tumor necrosis factor-alpha; WT: wide type.

Article Snippet: Constructs used were: pMD2.G (Addgene, Cat# 12259, RRID: Addgene_12259), psPAX2 (Addgene, Cat# 12260, RRID: Addgene_12260), pHAGE- BRAF V600E plasmid (Addgene, Cat# 116204, RRID: Addgene_116204), pHAGE- BRAF WT plasmid (Addgene, Cat# 116719, RRID: Addgene_116719), pBabe-Puro- BRAF V600E plasmid (Addgene, Cat# 15269, RRID: Addgene_15269), gag/pol-Retroviral plasmid (Addgene, Cat# 14887, RRID: Addgene_14887), Control siRNA (Cell Signaling Technology, Boston, MA, USA, Cat# 6568S), SAPK/JNK siRNA (Cell Signaling Technology, Cat# 6232S), ERK1 siRNA (Cell Signaling Technology, Cat# 6436S), ERK2 siRNA (Cell Signaling Technology, Cat# 6578S).

Techniques: Expressing, Cell Culture, Transduction, Plasmid Preparation, Western Blot, Immunostaining, Cell Counting, Transfection, Control, Real-time Polymerase Chain Reaction

Journal: Journal of Clinical Medicine

Article Title: Frequency of BRAF Mutations in Dysplastic Nevi, Lentigo Maligna, and Melanoma In Situ

doi: 10.3390/jcm13164799

Figure Lengend Snippet: The frequency of BRAF mutations varies across diagnoses.

Article Snippet: Three reaction mixtures were prepared for each sample: (i) mix with Mutation Detection Assay for V600E mutation (BRAF_476_mu; Assay ID: Hs00000111_mu), (ii) mix with Mutation Detection Assay for V600K mutation (BRAF_473_mu; Assay ID: Hs00000769_mu), (iii) Reference Assay (BRAF_rf; Assay ID: Hs00000172_rf).

Techniques: Mutagenesis

Journal: Journal of Clinical Medicine

Article Title: Frequency of BRAF Mutations in Dysplastic Nevi, Lentigo Maligna, and Melanoma In Situ

doi: 10.3390/jcm13164799

Figure Lengend Snippet: Presence and frequency of BRAF mutation between patients with ND, MIS, and LM.

Article Snippet: Three reaction mixtures were prepared for each sample: (i) mix with Mutation Detection Assay for V600E mutation (BRAF_476_mu; Assay ID: Hs00000111_mu), (ii) mix with Mutation Detection Assay for V600K mutation (BRAF_473_mu; Assay ID: Hs00000769_mu), (iii) Reference Assay (BRAF_rf; Assay ID: Hs00000172_rf).

Techniques: Mutagenesis

Journal: Journal of Clinical Medicine

Article Title: Frequency of BRAF Mutations in Dysplastic Nevi, Lentigo Maligna, and Melanoma In Situ

doi: 10.3390/jcm13164799

Figure Lengend Snippet: Age distribution of patients with and without BRAF mutation.

Article Snippet: Three reaction mixtures were prepared for each sample: (i) mix with Mutation Detection Assay for V600E mutation (BRAF_476_mu; Assay ID: Hs00000111_mu), (ii) mix with Mutation Detection Assay for V600K mutation (BRAF_473_mu; Assay ID: Hs00000769_mu), (iii) Reference Assay (BRAF_rf; Assay ID: Hs00000172_rf).

Techniques: Mutagenesis

Journal: Journal of Clinical Medicine

Article Title: Frequency of BRAF Mutations in Dysplastic Nevi, Lentigo Maligna, and Melanoma In Situ

doi: 10.3390/jcm13164799

Figure Lengend Snippet: Correlation of BRAF mutation and localization.

Article Snippet: Three reaction mixtures were prepared for each sample: (i) mix with Mutation Detection Assay for V600E mutation (BRAF_476_mu; Assay ID: Hs00000111_mu), (ii) mix with Mutation Detection Assay for V600K mutation (BRAF_473_mu; Assay ID: Hs00000769_mu), (iii) Reference Assay (BRAF_rf; Assay ID: Hs00000172_rf).

Techniques: Mutagenesis

Journal: Journal of Clinical Medicine

Article Title: Frequency of BRAF Mutations in Dysplastic Nevi, Lentigo Maligna, and Melanoma In Situ

doi: 10.3390/jcm13164799

Figure Lengend Snippet: Tumor size distribution in patients with and without BRAF mutation. ∗ and ◦ represent outliers.

Article Snippet: Three reaction mixtures were prepared for each sample: (i) mix with Mutation Detection Assay for V600E mutation (BRAF_476_mu; Assay ID: Hs00000111_mu), (ii) mix with Mutation Detection Assay for V600K mutation (BRAF_473_mu; Assay ID: Hs00000769_mu), (iii) Reference Assay (BRAF_rf; Assay ID: Hs00000172_rf).

Techniques: Mutagenesis

Journal: Journal of Clinical Medicine

Article Title: Frequency of BRAF Mutations in Dysplastic Nevi, Lentigo Maligna, and Melanoma In Situ

doi: 10.3390/jcm13164799

Figure Lengend Snippet: Correlation between the presence of the BRAF mutation and a previous melanoma diagnosis.

Article Snippet: Three reaction mixtures were prepared for each sample: (i) mix with Mutation Detection Assay for V600E mutation (BRAF_476_mu; Assay ID: Hs00000111_mu), (ii) mix with Mutation Detection Assay for V600K mutation (BRAF_473_mu; Assay ID: Hs00000769_mu), (iii) Reference Assay (BRAF_rf; Assay ID: Hs00000172_rf).

Techniques: Mutagenesis, Biomarker Discovery

Journal: Advanced Science

Article Title: VDLIN: A Deep Learning‐Based Platform for Methylcobalamin‐Inspired Immunomodulatory Compound Screening

doi: 10.1002/advs.202413775

Figure Lengend Snippet: Co7 induces Ifnb1 expression via the TLR4 signaling pathway. A) Volcano plot illustrating the distribution of differentially expressed genes (DEGs) between the Co7 and DMSO treatment groups after 3 h in RAW 264.7 cells. Fold changes are presented as log 2 transformations. Red dots represent DEGs upregulated in the Co7 group. B) Protein‐protein interaction (PPI) network analysis of differentially expressed genes (DEGs) in the Co7‐treated group compared to the DMSO group, revealing significant enrichment in pathways associated with the innate immune response and type I interferon signaling. C) KEGG pathway enrichment analysis of DEGs induced by Co7, highlighting associations with innate immune response pathways. D) Co7 (50 µmol/L) exhibiting strong antiviral effects against VSV in RAW 264.7 and HT29 cells. E) Co7 significantly reduced the inflammatory response induced by LPS, VSV, EMCV, and HSV in RAW 264.7 cells. F) Volcano plot representing the differential gene expression analysis between Co7‐ and LPS‐treated RAW 264.7 cells after 3 h of treatment. Fold changes are displayed as log 2 transformations. Red dots indicate genes upregulated in the Co7 group, while blue dots represent downregulated genes compared to LPS treatment. G) Western blot analysis demonstrating that Co7 inhibited the expression of iNOS and COX2, as well as the phosphorylation of NF‐κB‐P65 at the protein level in RAW 264.7 cells. H) Co7 significantly reduced the mortality rate in mice (n = 10 per group) following LPS challenge (20 mg/kg), compared to the PBS control group. RT‐qPCR data were presented as means ± SEM from three independent experiments. Statistical significance was determined using one‐way ANOVA with Bonferroni's multiple comparisons test (left three panels in E), paired‐samples t‐test (D, right panel of EMCV and HSV in E), or the log‐rank test (H). * P < 0.05, ** P < 0.01, and *** P < 0.001.

Article Snippet: Each inhibitor was dissolved in anhydrous DMSO and diluted to its respective working concentration: C29 (10 μM, S6597, Selleck) served as a TLR2 inhibitor; Procyanidin B1 (30 μM, HY‐N0795, MedChemExpress) acted as a TLR4 inhibitor; MyD88‐IN‐1 (30 μM, HY‐149992, MedChemExpress) was used to inhibit MyD88; Pepinh‐TRIF TFA (30 μM, HY‐P2565, MedChemExpress) functioned as a TRIF inhibitor; and GSK8612 (5 μM, T5540, TargetMol) inhibited TBK1/IKKε.

Techniques: Expressing, Gene Expression, Western Blot, Phospho-proteomics, Control, Quantitative RT-PCR

Journal: Journal of Clinical Medicine

Article Title: Frequency of BRAF Mutations in Dysplastic Nevi, Lentigo Maligna, and Melanoma In Situ

doi: 10.3390/jcm13164799

Figure Lengend Snippet: The frequency of BRAF mutations varies across diagnoses.

Article Snippet: Three reaction mixtures were prepared for each sample: (i) mix with Mutation Detection Assay for V600E mutation (BRAF_476_mu; Assay ID: Hs00000111_mu), (ii) mix with Mutation Detection Assay for V600K mutation (BRAF_473_mu; Assay ID: Hs00000769_mu), (iii) Reference Assay (BRAF_rf; Assay ID: Hs00000172_rf).

Techniques: Mutagenesis

Journal: Journal of Clinical Medicine

Article Title: Frequency of BRAF Mutations in Dysplastic Nevi, Lentigo Maligna, and Melanoma In Situ

doi: 10.3390/jcm13164799

Figure Lengend Snippet: Presence and frequency of BRAF mutation between patients with ND, MIS, and LM.

Article Snippet: Three reaction mixtures were prepared for each sample: (i) mix with Mutation Detection Assay for V600E mutation (BRAF_476_mu; Assay ID: Hs00000111_mu), (ii) mix with Mutation Detection Assay for V600K mutation (BRAF_473_mu; Assay ID: Hs00000769_mu), (iii) Reference Assay (BRAF_rf; Assay ID: Hs00000172_rf).

Techniques: Mutagenesis

Journal: Journal of Clinical Medicine

Article Title: Frequency of BRAF Mutations in Dysplastic Nevi, Lentigo Maligna, and Melanoma In Situ

doi: 10.3390/jcm13164799

Figure Lengend Snippet: Age distribution of patients with and without BRAF mutation.

Article Snippet: Three reaction mixtures were prepared for each sample: (i) mix with Mutation Detection Assay for V600E mutation (BRAF_476_mu; Assay ID: Hs00000111_mu), (ii) mix with Mutation Detection Assay for V600K mutation (BRAF_473_mu; Assay ID: Hs00000769_mu), (iii) Reference Assay (BRAF_rf; Assay ID: Hs00000172_rf).

Techniques: Mutagenesis

Journal: Journal of Clinical Medicine

Article Title: Frequency of BRAF Mutations in Dysplastic Nevi, Lentigo Maligna, and Melanoma In Situ

doi: 10.3390/jcm13164799

Figure Lengend Snippet: Correlation of BRAF mutation and localization.

Article Snippet: Three reaction mixtures were prepared for each sample: (i) mix with Mutation Detection Assay for V600E mutation (BRAF_476_mu; Assay ID: Hs00000111_mu), (ii) mix with Mutation Detection Assay for V600K mutation (BRAF_473_mu; Assay ID: Hs00000769_mu), (iii) Reference Assay (BRAF_rf; Assay ID: Hs00000172_rf).

Techniques: Mutagenesis

Journal: Journal of Clinical Medicine

Article Title: Frequency of BRAF Mutations in Dysplastic Nevi, Lentigo Maligna, and Melanoma In Situ

doi: 10.3390/jcm13164799

Figure Lengend Snippet: Tumor size distribution in patients with and without BRAF mutation. ∗ and ◦ represent outliers.

Article Snippet: Three reaction mixtures were prepared for each sample: (i) mix with Mutation Detection Assay for V600E mutation (BRAF_476_mu; Assay ID: Hs00000111_mu), (ii) mix with Mutation Detection Assay for V600K mutation (BRAF_473_mu; Assay ID: Hs00000769_mu), (iii) Reference Assay (BRAF_rf; Assay ID: Hs00000172_rf).

Techniques: Mutagenesis

Journal: Journal of Clinical Medicine

Article Title: Frequency of BRAF Mutations in Dysplastic Nevi, Lentigo Maligna, and Melanoma In Situ

doi: 10.3390/jcm13164799

Figure Lengend Snippet: Correlation between the presence of the BRAF mutation and a previous melanoma diagnosis.

Article Snippet: Three reaction mixtures were prepared for each sample: (i) mix with Mutation Detection Assay for V600E mutation (BRAF_476_mu; Assay ID: Hs00000111_mu), (ii) mix with Mutation Detection Assay for V600K mutation (BRAF_473_mu; Assay ID: Hs00000769_mu), (iii) Reference Assay (BRAF_rf; Assay ID: Hs00000172_rf).

Techniques: Mutagenesis, Biomarker Discovery