Journal: Frontiers in Immunology

Article Title: Unveiling spatial complexity in solid tumor immune microenvironments through multiplexed imaging

doi: 10.3389/fimmu.2024.1383932

Figure Lengend Snippet: Immunophenotyping panel for multiplexed tissue imaging of cancer.

Article Snippet: CD99 , REA1174 , 50 , 130-121-078 , PE , Miltenyi Biotec.

Techniques: Imaging

Journal: Frontiers in Cellular Neuroscience

Article Title: Adipose-derived stem cell-conditioned medium mitigates ischemia-induced neuronal injury via the JAK1/STAT3 signaling pathway

doi: 10.3389/fncel.2026.1744887

Figure Lengend Snippet: ADSC-CM promotes neurite outgrowth in OGD-injured neurons through the JAK1-STAT3 signaling pathway. (A) On day 5 of cultivation, neuronal neurites extended and formed an extensive neural network. Immunofluorescence staining showed that the neurons were positive for the neuronal-specific marker Tuj1. After OGD injury, neuronal neurites were damaged, partially interrupted, and disappeared. (B) Western blot bands of JAK1, pJAK1, STAT3, pSTAT3, and GAPDH for each group. (C) Comparison of the relative ratio of pJAK1 to JAK1 across experimental groups ( n = 4). (D) Comparison of the relative ratio of pSTAT3 to STAT3 across experimental groups ( n = 4). (E) Immunofluorescence images of neurons from each group, showing the effects of ADSC-CM and GLPG0634 on neurite outgrowth. Neurons were stained with the Tuj1 antibody (red) to label the neuronal cytoskeleton, and nuclei were counterstained with DAPI (blue). (F) Diagram illustrating how to measure the length of the longest neurite and the number of primary neurites in neurons. The green line shows the trajectory of the longest neurite, and white arrows point to primary neurites. (G,H) Comparison of the length of the longest neurite and the number of primary neurites in neurons across experimental groups ( n = 4). Data are expressed as means ± SEM. The difference between the groups was assessed using a one-way ANOVA followed by Bonferroni post hoc tests. * p < 0.05, ** p < 0.01, *** p < 0.001. Scale bars: 100 μm (A) , 20 μm (E,F) .

Article Snippet: After overnight incubation with rabbit anti-Tuj1 antibody (1:200, BM3881, Boster, Wuhan, China) at 4 °C, cells were washed and then incubated with Cy3-conjugated goat anti-rabbit IgG (1:400, AS007, ABclonal) for 1 h at room temperature in the dark.

Techniques: Immunofluorescence, Staining, Marker, Western Blot, Comparison

Journal: Oncology Research

Article Title: BHLHE40 Is a Transcriptional Regulatory Target of NFE2L3 in Triple-Negative Breast Cancer

doi: 10.32604/or.2025.070793

Figure Lengend Snippet: ( A ) Consensus regulatory targets of NFE2L3, based on the regulatory networks inferred using GENIE3 and ARACNe. The red represents the protein upregulated in TNBC compared to TPBC. The blue represents the protein downregulated in TNBC compared to TPBC. Kaplan-Meier survival plots of (B) NFE2L3 and (C) BHLHE40 using the TCGA RNA-seq Breast cancer dataset. The red line represents higher expression of the gene, while the blue line represents lower expression of the gene. p < 0.05. Higher NFE2L3 expression corresponds to poor overall survival than those with low NFE2L3 expression, while there is no significant difference in survival between patients with high and low expressions of BHLHE40

Article Snippet: The membranes were washed with PBST solution for 5 min three times, and then incubated with primary antibodies against NFE2L3 (rabbit polyclonal antibody; #A09888, Boster Bio, Pleasanton, CA, USA), BHLHE40 (rabbit polyclonal antibody; #ABN1737, Millipore-Sigma, Burlington, MA), and GAPDH (rabbit monoclonal antibody; #2118, Cell Signaling Technology, Danvers, MA, USA), were diluted in blocking solution at 4°C overnight.

Techniques: RNA Sequencing, Expressing

Journal: Oncology Research

Article Title: BHLHE40 Is a Transcriptional Regulatory Target of NFE2L3 in Triple-Negative Breast Cancer

doi: 10.32604/or.2025.070793

Figure Lengend Snippet: Gene expression quantification using quantitative Reverse Transcriptase polymerase chain reaction (qRT-PCR). NFE2L3 gene expression in ( A ) MDA-MB-231 and ( B ) MDA-MB-468 cells in siControl, siNFE2L3, and siBHLHE40 groups. ( C , D ) Gene expression in ( C ) MDA-MB-231 and ( D ) MDA-MB-468 cells in untreated, NFE2L3 plasmid and BHLHE40 plasmid groups. BHLHE40 gene expression using qRT-PCR. BHLHE40 gene expression in ( E ) MDA-MB-231 and ( F ) MDA-MB-468 cells following treatment with siControl, siNFE2L3, and siBHLHE40. BHLHE40 gene expression in ( G ) MDA-MB-231 and ( H ) MDA-MB-468 cells following treatment with vehicle (untreated), NFE2L3 plasmid, and BHLHE40 plasmid. One-way ANOVA followed by post-hoc Tukey’s test was used. Compared to the siControl or untreated group: ***indicates p < 0.001, and ****indicates p < 0.0001. Compared to siNFE2L3 or NFE2L3 plasmid group: #### indicates p < 0.0001

Article Snippet: The membranes were washed with PBST solution for 5 min three times, and then incubated with primary antibodies against NFE2L3 (rabbit polyclonal antibody; #A09888, Boster Bio, Pleasanton, CA, USA), BHLHE40 (rabbit polyclonal antibody; #ABN1737, Millipore-Sigma, Burlington, MA), and GAPDH (rabbit monoclonal antibody; #2118, Cell Signaling Technology, Danvers, MA, USA), were diluted in blocking solution at 4°C overnight.

Techniques: Gene Expression, Reverse Transcription, Polymerase Chain Reaction, Quantitative RT-PCR, Plasmid Preparation

Journal: Oncology Research

Article Title: BHLHE40 Is a Transcriptional Regulatory Target of NFE2L3 in Triple-Negative Breast Cancer

doi: 10.32604/or.2025.070793

Figure Lengend Snippet: Basal protein expression of NFE2L3 and BHLHE40 NFE2L3 and BHLHE40 in breast cancer cells. ( A ) Representative blots of NFE2L3 and BHLHE40 protein expression following treatment with siControl, siNFE2L3, and siBHLHE40. GAPDH was used as a loading control. ( B ) Graphical presentation of NFE2L3 protein expression in MDA-MB-231 cells following treatment as described. ( C ) Graphical presentation of BHLHE40 protein expression in MDA-MB-231 cells following treatment as described. ( D ) Representative blots of NFE2L3 and BHLHE40 protein expression following treatment with vehicle (Untreated), NFE2L3 DNA plasmid, and BHLHE40 DNA plasmid. GAPDH was used as a loading control. ( E ) Graphical presentation of NFE2L3 protein expression in MDA-MB-231 cells following treatment as described. ( F ) Graphical presentation of BHLHE40 protein expression in MDA-MB-231 cells following treatment as described. One-way ANOVA followed by post-hoc Dunnet’s test was used. Compared to siControl: **indicates p < 0.01, ***indicates p < 0.001, and ****indicates p < 0.0001. Compared to the siNFE2L3 group or NFE2L3 plasmid group: # indicates p < 0.05, and #### indicates p < 0.0001

Article Snippet: The membranes were washed with PBST solution for 5 min three times, and then incubated with primary antibodies against NFE2L3 (rabbit polyclonal antibody; #A09888, Boster Bio, Pleasanton, CA, USA), BHLHE40 (rabbit polyclonal antibody; #ABN1737, Millipore-Sigma, Burlington, MA), and GAPDH (rabbit monoclonal antibody; #2118, Cell Signaling Technology, Danvers, MA, USA), were diluted in blocking solution at 4°C overnight.

Techniques: Expressing, Control, Plasmid Preparation

Journal: Oncology Research

Article Title: BHLHE40 Is a Transcriptional Regulatory Target of NFE2L3 in Triple-Negative Breast Cancer

doi: 10.32604/or.2025.070793

Figure Lengend Snippet: Validation of regulatory relationship between NFE2L3 and BHLHE40. ( A ) Visual representation of luciferase assay with BHLHE40 conjugated with Nanoluc vector, activated by NFE2L3, resulting in production of luciferase activity. Qualitative analysis of BHLHE40 transcriptional activation by NFE2L3 plasmid compared to BHLHE40 plasmid conjugated with the Nanoluc vector in ( B ) MDA-MB-31 and ( C ) MDA-MB-468 cells. Co-immunoprecipitation analysis of NFE2L3 protein and BHLHE40 protein interaction. ( D ) Co-IP of NFE2L3 pulldown with immunoblotting with BHLHE40. ( E ) Co-IP of BHLHE40 pulldown with immunoblotting with NFE2L3. Red boxes indicate the interacting bands. Student’s t -test was used to compare the two groups, *indicates p < 0.05

Article Snippet: The membranes were washed with PBST solution for 5 min three times, and then incubated with primary antibodies against NFE2L3 (rabbit polyclonal antibody; #A09888, Boster Bio, Pleasanton, CA, USA), BHLHE40 (rabbit polyclonal antibody; #ABN1737, Millipore-Sigma, Burlington, MA), and GAPDH (rabbit monoclonal antibody; #2118, Cell Signaling Technology, Danvers, MA, USA), were diluted in blocking solution at 4°C overnight.

Techniques: Biomarker Discovery, Luciferase, Plasmid Preparation, Activity Assay, Activation Assay, Immunoprecipitation, Co-Immunoprecipitation Assay, Western Blot

Journal: Oncology Research

Article Title: BHLHE40 Is a Transcriptional Regulatory Target of NFE2L3 in Triple-Negative Breast Cancer

doi: 10.32604/or.2025.070793

Figure Lengend Snippet: Phenotypic studies on MDA-MB-231 cells. Effect of knockdown or induction of NFE2L3 and BHLHE40 gene expression on MDA-MB-231 cell proliferation. MDA-MB-231 cell s’ proliferation was determined following treatment with vehicle (Control), siNFE2L3, siBHLHE40, as well as ( A ) NFE2L3 DNA plasmid, or ( B ) BHLHE40 DNA plasmid for 24, 48, and 72 h. Effect of knockdown or induction of NFE2L3 and BHLHE40 gene expression on MDA-MB-231 cell migration. MDA-MB-231 cell migration was determined following treatment with vehicle (Control), siNFE2L3, siBHLHE40, as well as ( C ) NFE2L3 DNA plasmid, or ( D ) BHLHE40 DNA plasmid for 24, 48, and 72 h. (Scale — 100 μm) ( E , F ) Graphical presentation of the estimation of wound closure area following treatment as described above. One-way ANOVA followed by post-hoc Tukey’s test was used. Compared to untreated: *indicates p < 0.05, **indicates p < 0.01, ***indicates p < 0.001, and ****indicates p < 0.0001. Compared to siNFE2L3: # indicates p < 0.05, ### indicates p < 0.001, and #### indicates p < 0.0001. Compared to siBHLHE40, a indicates p < 0.05, aaa indicates p < 0.001, and aaaa indicates p < 0.0001

Article Snippet: The membranes were washed with PBST solution for 5 min three times, and then incubated with primary antibodies against NFE2L3 (rabbit polyclonal antibody; #A09888, Boster Bio, Pleasanton, CA, USA), BHLHE40 (rabbit polyclonal antibody; #ABN1737, Millipore-Sigma, Burlington, MA), and GAPDH (rabbit monoclonal antibody; #2118, Cell Signaling Technology, Danvers, MA, USA), were diluted in blocking solution at 4°C overnight.

Techniques: Knockdown, Gene Expression, Control, Plasmid Preparation, Migration

Journal: Bioorganic chemistry

Article Title: Bioinformatic and biochemical findings disclosed anti-hepatic steatosis mechanism of calycosin.

doi: 10.1016/j.bioorg.2020.103914

Figure Lengend Snippet: Fig. 2. All targets/genes of calycosin and fatty liver were picked up for collection of central biotargets of calycosin for anti-fatty liver, covering ALDH2, NPC1, HMGB1, UGT1A1, MAPK3, EGFR, HTR2, AMIF, CYP19A1.

Article Snippet: The fatty liver sections from biopsy were dewaxed and then blocked with 5% bovine serum albumin solution (Beyotime Biotechnology, China) for about 1 h. After rinse with phosphate buffer saline/0.5% tween 20 solution for about 3 times, the sections were incubated with primary antibodies of ALDH2 (1:200, BOSTER Biological Technology, China), NPC1 (1:200, BOSTER), HMGB1 (1:200, BOSTER) at 4 °C overnight, then the sections being incubated with diluted secondary antibodies of DyLight 488 (1:400, BOSTER Biological Technology, China).

Techniques:

Journal: Bioorganic chemistry

Article Title: Bioinformatic and biochemical findings disclosed anti-hepatic steatosis mechanism of calycosin.

doi: 10.1016/j.bioorg.2020.103914

Figure Lengend Snippet: Fig. 5. Clinical findings of patients with fatty liver. In medical detection, the steatohepatitis patients were identified through B-ultrasound and pathological stain. And the fatty liver sections showed reduced ALDH2, NPC1 expressions and elevated HMGB1expression.

Article Snippet: The fatty liver sections from biopsy were dewaxed and then blocked with 5% bovine serum albumin solution (Beyotime Biotechnology, China) for about 1 h. After rinse with phosphate buffer saline/0.5% tween 20 solution for about 3 times, the sections were incubated with primary antibodies of ALDH2 (1:200, BOSTER Biological Technology, China), NPC1 (1:200, BOSTER), HMGB1 (1:200, BOSTER) at 4 °C overnight, then the sections being incubated with diluted secondary antibodies of DyLight 488 (1:400, BOSTER Biological Technology, China).

Techniques: Staining