Journal: iScience

Article Title: Zhigancao decoction alleviates Parkinson’s disease via inhibiting TNF/NF-κB and Ras/ERK-mediated neuroinflammation and apoptosis

doi: 10.1016/j.isci.2025.114489

Figure Lengend Snippet: Network pharmacology predicted that ZGCD treatment for Parkinson’s disease might be associated with apoptosis, inflammation, TNF/NF-κB, and Ras/ERK signaling pathways (A) ZGCD Herbal Active Ingredient-Target Network. (B) Venn diagrams of active ingredients and disease targets. (C) The PPI network of the common targets of ZGCD and PD. (D) GO enrichment analysis of potential therapeutic targets. (E) Enrichment analysis of KEGG pathways for potential therapeutic targets.

Article Snippet: Membranes were incubated overnight at 4°C with primary antibodies against TH (1:1000, Abcam, USA), α-Syn (1:1000, Abcam, USA), TNFR1 (1:1000, Proteintech, China), P65 (1:1000, Abcam, USA), P-P65 (1:1000, Abcam, USA), Ras (1:3000, Abcam, USA), ERK (1:2000, Proteintech, China), p -ERK (1:2000, Proteintech, China), TNF-α (1:1000, Proteintech, China), Bax (1:1000, Proteintech, China), Bcl-2 (1:1000, Proteintech, China), Caspase-3 (1:1000, Proteintech, China), Cleaved Caspase-3 (1:1000, Proteintech, China), and GAPDH (1:3000, Abcam, USA) ( ).

Techniques: Protein-Protein interactions, Biomarker Discovery

Journal: iScience

Article Title: Zhigancao decoction alleviates Parkinson’s disease via inhibiting TNF/NF-κB and Ras/ERK-mediated neuroinflammation and apoptosis

doi: 10.1016/j.isci.2025.114489

Figure Lengend Snippet: ZGCD treatment may alleviate MPTP-induced injury of SN DA neurons by inhibiting the TNF/NF-kB and Ras/ERK pathways (A) WB was used to detect the representative expressions of TNFR1, P65, and p-P65 in SN. (B and C) Statistics of the relative expression level of TNFR1 and p-P65/P65 in SN ( n = 4). (D) WB was used to detect the representative expressions of Ras, ERK, and p -ERK in SN. (E and F) Statistics of the relative expression level of Ras and p -ERK/ERK in SN ( n = 4). (G–K) Representative expression of Tnfr1, Nfkb, Grb2, Ras, and Erk detected by RT-qPCR ( n = 4). ### p < 0.001 vs. Con; n.s., not significant, ∗ p < 0.05, ∗∗ p < 0.01 and ∗∗∗ p < 0.001 vs. MPTP. Data are expressed as mean ± SD.

Article Snippet: Membranes were incubated overnight at 4°C with primary antibodies against TH (1:1000, Abcam, USA), α-Syn (1:1000, Abcam, USA), TNFR1 (1:1000, Proteintech, China), P65 (1:1000, Abcam, USA), P-P65 (1:1000, Abcam, USA), Ras (1:3000, Abcam, USA), ERK (1:2000, Proteintech, China), p -ERK (1:2000, Proteintech, China), TNF-α (1:1000, Proteintech, China), Bax (1:1000, Proteintech, China), Bcl-2 (1:1000, Proteintech, China), Caspase-3 (1:1000, Proteintech, China), Cleaved Caspase-3 (1:1000, Proteintech, China), and GAPDH (1:3000, Abcam, USA) ( ).

Techniques: Expressing, Quantitative RT-PCR

Journal: iScience

Article Title: Zhigancao decoction alleviates Parkinson’s disease via inhibiting TNF/NF-κB and Ras/ERK-mediated neuroinflammation and apoptosis

doi: 10.1016/j.isci.2025.114489

Figure Lengend Snippet: ZGCDS treatment may alleviate MPTP-induced injury of SN DA neurons by inhibiting the TNF/NF-kB and Ras/ERK pathways (A–C) The levels of TNF-α, IL-1β, and IL-6 were detected by ELISA in SH-SY5Y cells ( n = 3). (D) WB was used to detect the representative expressions of TNF-α in SH-SY5Y cells. (E) Statistics of the relative expression level of TNF-α in SH-SY5Y cells ( n = 3). (F–H) Representative expression of TNF-α, IL-1β, and IL-6 detected by RT-qPCR ( n = 3). (I–N) IF was used to analyze the expressions of TNF-α, IL-1β, and IL-6 in SH-SY5Y cells ( n = 3). Scale bars, 100 μm. (O–Q) Representative expression of Bax, Bcl-2, and caspase-3 detected by RT-qPCR in SH-SY5Y cells ( n = 3). (R–T) WB analysis of Bax, Bcl-2, Caspase-3, and Cleaved Caspase-3 expression in SH-SY5Y cells ( n = 3). ### p < 0.001 vs. Con; n.s., not significant, ∗ p < 0.05, ∗∗ p < 0.01 and ∗∗∗ p < 0.001 vs. MPTP. Data are expressed as mean ± SD.

Article Snippet: Membranes were incubated overnight at 4°C with primary antibodies against TH (1:1000, Abcam, USA), α-Syn (1:1000, Abcam, USA), TNFR1 (1:1000, Proteintech, China), P65 (1:1000, Abcam, USA), P-P65 (1:1000, Abcam, USA), Ras (1:3000, Abcam, USA), ERK (1:2000, Proteintech, China), p -ERK (1:2000, Proteintech, China), TNF-α (1:1000, Proteintech, China), Bax (1:1000, Proteintech, China), Bcl-2 (1:1000, Proteintech, China), Caspase-3 (1:1000, Proteintech, China), Cleaved Caspase-3 (1:1000, Proteintech, China), and GAPDH (1:3000, Abcam, USA) ( ).

Techniques: Enzyme-linked Immunosorbent Assay, Expressing, Quantitative RT-PCR

Journal: iScience

Article Title: Zhigancao decoction alleviates Parkinson’s disease via inhibiting TNF/NF-κB and Ras/ERK-mediated neuroinflammation and apoptosis

doi: 10.1016/j.isci.2025.114489

Figure Lengend Snippet: ZGCDS exerts neuroprotective effects by inhibiting the activation of TNF/NF-κB and Ras/ERK signaling pathways in SH-SY5Y cells (A) WB was used to detect the representative expressions of TNFR1, P65, and p-P65 in SH-SY5Y cells. (B) Statistics of the relative expression level of TNFR1 in SH-SY5Y cells ( n = 3). (C) Statistics of the relative expression level of p-P65/P65 in SH-SY5Y cells ( n = 3). (D–G) IF was used to analyze the expressions of TNFR1 and p-P65 in SH-SY5Y cells ( n = 3). Scale bars, 200 μm. (H–J) WB analysis of Ras and ERK expression and phosphorylation in SH-SY5Y cells ( n = 6). (K–P) IF was used to analyze the expressions of Grb2, Ras, and p -ERK in SH-SY5Y cells ( n = 3). Scale bars, 100 μm. (Q–S) WB analysis of TNFR1 and P65 expression and phosphorylation in SH-SY5Y cells ( n = 4). (T–W) WB analysis of Grb2, Ras, and ERK expression and phosphorylation in SH-SY5Y cells ( n = 4). ### p < 0.001 vs. Con; n.s., not significant, ∗ p < 0.05, ∗∗ p < 0.01 and ∗∗∗ p < 0.001 vs. MPTP. Data are expressed as mean ± SD.

Article Snippet: Membranes were incubated overnight at 4°C with primary antibodies against TH (1:1000, Abcam, USA), α-Syn (1:1000, Abcam, USA), TNFR1 (1:1000, Proteintech, China), P65 (1:1000, Abcam, USA), P-P65 (1:1000, Abcam, USA), Ras (1:3000, Abcam, USA), ERK (1:2000, Proteintech, China), p -ERK (1:2000, Proteintech, China), TNF-α (1:1000, Proteintech, China), Bax (1:1000, Proteintech, China), Bcl-2 (1:1000, Proteintech, China), Caspase-3 (1:1000, Proteintech, China), Cleaved Caspase-3 (1:1000, Proteintech, China), and GAPDH (1:3000, Abcam, USA) ( ).

Techniques: Activation Assay, Protein-Protein interactions, Expressing, Phospho-proteomics

Journal: iScience

Article Title: Zhigancao decoction alleviates Parkinson’s disease via inhibiting TNF/NF-κB and Ras/ERK-mediated neuroinflammation and apoptosis

doi: 10.1016/j.isci.2025.114489

Figure Lengend Snippet: Bioactive compounds (Apigenin, Bisdemethoxycurcumin) from ZGCD may exert anti-PD effects by targeting the TNF/NF-kB and Ras/ERK pathways (A and B) Total ion chromatograms (TICs) of Con in positive (A) and negative (B) ion modes. (C and D) Total ion chromatograms (TICs) of ZGCD-CS in positive (C) and negative (D) ion modes. (E) Venn diagram of serum pharmacochemical analysis. (F) Molecular docking analysis of Apigenin and Bisdemethoxycurcumin with key proteins in the pathway. (G–I) The levels of TNF-α, IL-1β, and IL-6 were detected by ELISA in SH-SY5Y cells ( n = 3). (J–L) WB analysis of TNFR1 and P65 expression and phosphorylation in SH-SY5Y cells ( n = 3). (M–P) WB analysis of Grb2, Ras, and ERK expression and phosphorylation in SH-SY5Y cells ( n = 3). ### p < 0.001 vs. Con; n.s., not significant, ∗ p < 0.05, ∗∗ p < 0.01 and ∗∗∗ p < 0.001 vs. MPTP. Data are expressed as mean ± SD.

Article Snippet: Membranes were incubated overnight at 4°C with primary antibodies against TH (1:1000, Abcam, USA), α-Syn (1:1000, Abcam, USA), TNFR1 (1:1000, Proteintech, China), P65 (1:1000, Abcam, USA), P-P65 (1:1000, Abcam, USA), Ras (1:3000, Abcam, USA), ERK (1:2000, Proteintech, China), p -ERK (1:2000, Proteintech, China), TNF-α (1:1000, Proteintech, China), Bax (1:1000, Proteintech, China), Bcl-2 (1:1000, Proteintech, China), Caspase-3 (1:1000, Proteintech, China), Cleaved Caspase-3 (1:1000, Proteintech, China), and GAPDH (1:3000, Abcam, USA) ( ).

Techniques: Enzyme-linked Immunosorbent Assay, Expressing, Phospho-proteomics

Journal: International Journal of Biological Sciences

Article Title: LncRNA SLCO4A1-AS1 predicts poor prognosis and promotes proliferation and metastasis via the EGFR/MAPK pathway in colorectal cancer

doi: 10.7150/ijbs.38041

Figure Lengend Snippet: Proposed schematic model illustrating the role of SLCO4A1-AS1 in regulating CRC by EGFR/MAPK signaling pathway. SLCO4A1-AS1 influences EGFR/MAPK signaling pathway by promoting the expression of EGFR, KRAS, BRAF, MEK, ERK, MAP3K1 and its corresponding phosphorylated protein levels, which further affect the proliferation, migration and invasion of CRC cells.

Article Snippet: The membrane was incubated with primary antibody overnight at 4 °C, including EGFR (1:2000, Abcam), P-EGFR (1:2000, Abcam), KRAS (1:2000, Abcam), BRAF (1:2000, Abcam), MEK1/2 (1:2000, Abcam), P-MEK1/2 (1:1000, Proteintech), ERK (1:2000, Abcam), P-ERK (1:2000, Abcam), MAP3K1 (1:700, Proteintech), P-MAP3K1 (1:1000, Proteintech), β-actin (1:700, Proteintech), incubate for 2 hours at room temperature with anti-rabbit secondary antibody.

Techniques: Expressing, Migration

Journal: Molecular cancer

Article Title: A novel polypeptide CAPG-171aa encoded by circCAPG plays a critical role in triple-negative breast cancer.

doi: 10.1186/s12943-023-01806-x

Figure Lengend Snippet: Fig. 6 CAPG-171aa interacts with STK38 activating the downstream MEK1/2-ERK1/2 pathway via MEKK2. (A) MDA-MB-231 and MDA-MB-468 cell lysates were IP with anti-MEKK2 antibody followed by detection with anti-MEKK2, STK38, and SMURF1 antibody. (B) MDA-MB-231 and MDA-MB-468 were trans fected with CAPG-171aa-FLAG. Whole-cell lysates were IP with anti-SMURF1 and IgG antibodies followed by detection with anti-FLAG, STK38, SMURF1, and GAPDH antibodies. (C-D) Before being treated with MG132, MDA-MB-231, and MDA-MB-468 were transfected with CAPG-171aa-FLAG and circCAPG KD plasmids. Ubiquitination and protein expression levels of MEKK2 were assayed in CAPG-171aa OE (C) and circCAPG KD (D) MDA-MB-231 and MDA- MB-468. (E) Ubiquitination and protein expression levels of MEKK2 were assayed in circCAPG KD MDA-MB-231 and MDA-MB-468 through pulse-chase experiments with cycloheximide. (F) IB of MEKK2, p-MEK1/2, MEK1/2, p-ERK1/2 and ERK1/2 in circCAPG KD MDA-MB-231 and MDA-MB-468. (G) IB of p-MEK1/2, MEK1/2, p-ERK1/2 and ERK1/2 in OE STK38 MDA-MB-231 and MDA-MB-468. (H) IB of MEKK2, p-MEK1/2, MEK1/2, p-ERK1/2 and ERK1/2 in CAPG-171aa, OE-STK38 and OE-STK38/CAPG-171aa transfected MDA-MB-231 and MDA-MB-468. All data were representative of at least three biological replicates and shown as mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001

Article Snippet: MDA-MB-231 and MDA-MB-468 cells were treated with 10 μg/mL MG132, respectively, (Solarbio, IM0310) for 12 h. Cell lysates were obtained using PierceTM IP lysis buffer (Thermo Fisher Scientific, USA) supplemented with a cocktail (Thermo Fisher Scientific, USA) and then incubated with anti-MEKK2 antibody (Proteintech, USA) and Protein A/G beads overnight at 4°C.

Techniques: Transfection, Ubiquitin Proteomics, Expressing, Pulse Chase

Journal: iScience

Article Title: FAM65A, as a potential predictor of prognosis, promotes colorectal cancer progression via activating Ras/ERK/RSK signaling

doi: 10.1016/j.isci.2026.114662

Figure Lengend Snippet: FAM65A binds to Ras and activates the Ras/ERK signaling to mediate RSK activation (A) The volcano plot analysis results for the FAM65A high-expression and low-expression groups from the TCGA database were shown. (B) The KEGG and GO results were shown. (C) The GSEA results were shown. (D) GSEA on DEGs between the FAM65A high-expression group and low-expression group in the Reactome database were shown. (E) IP was performed to detect the binding of FAM65A and Ras/p-RSK. (F) IP was performed to detect the binding of Ras and FAM65A/p-RSK. (G) Immunofluorescence was performed to detect the co-localization of FAM65A and Ras. Scale bars, 20 μm. (H) Western blot analysis the Ras and p -ERK expression in FAM65A knockdown or overexpression cells. Data are presented as mean ± SEM of biologically independent experiments.

Article Snippet: p -ERK , Proteintech , Cat# 28733-1-AP; RRID: AB_2881202.

Techniques: Activation Assay, Expressing, Binding Assay, Immunofluorescence, Western Blot, Knockdown, Over Expression

Journal: iScience

Article Title: FAM65A, as a potential predictor of prognosis, promotes colorectal cancer progression via activating Ras/ERK/RSK signaling

doi: 10.1016/j.isci.2026.114662

Figure Lengend Snippet: Ras/ERK signaling activation was indispensable for FAM65A-mediated RSK activation and CRC progression (A) Western blot analysis of Ras and p -ERK expression in HCT116-FAM65A cells treated with 10 μM Abd-7, or without treatment. (B) Results from the CCK8 cell proliferation assay conducted on HCT116-FAM65A cells with and without the application of Abd-7, n = 3, ∗∗∗ p < 0.001. (C) Colony formation assay performed on HCT116-FAM65A cells treated with Abd-7 or not. (D) Quantitative analysis of the colony formation assay results, n = 3, ∗∗∗ p < 0.001. (E) Results from the EdU assay conducted on HCT116-FAM65A cells with and without the application of Abd-7. Scale bars, 100 μm. (F) Quantitative analysis of the EdU assay results, n = 3, ∗∗∗ p < 0.001. (G) Western blot analysis of Ki-67, cleaved Caspase 3, Bcl-2, and Bax expression in HCT116-FAM65A cells treated with Abd-7 or not. (H) Results from the apoptosis assay conducted on HCT116-FAM65A cells treated with Abd-7 or not. Scale bars, 50 μm. (I) Quantitative analysis of the apoptosis experiments, n = 3, ∗∗∗ p < 0.001. (J) Results from the Transwell migration assay conducted on HCT116-FAM65A cells with and without the application of Abd-7. Scale bars, 50 μm. (K) Quantitative analysis of the Transwell migration assay results, n = 3, ∗∗∗ p < 0.001. (L) Results from the wound healing assay performed on HCT116-FAM65A cells treated with Abd-7 or not. Scale bars, 50 μm. (M) Quantitative analysis of the wound healing assay results, n = 3, ∗∗∗ p < 0.001. (N) Western blot analysis the expression of EMT markers in HCT116-FAM65A cells treated with Abd-7 or not. (O) Proposed model of FAM65A in CRC progression. Data are presented as mean ± SEM of biologically independent experiments.

Article Snippet: p -ERK , Proteintech , Cat# 28733-1-AP; RRID: AB_2881202.

Techniques: Activation Assay, Western Blot, Expressing, Proliferation Assay, Colony Assay, EdU Assay, Apoptosis Assay, Transwell Migration Assay, Wound Healing Assay

Journal: iScience

Article Title: FAM65A, as a potential predictor of prognosis, promotes colorectal cancer progression via activating Ras/ERK/RSK signaling

doi: 10.1016/j.isci.2026.114662

Figure Lengend Snippet: Knockdown of FAM65A inhibits tumor progression in vivo (A) LOVO-shCtrl and LOVO-shFAM65A cells were administered into the fourth fat pad of nude mice, and the resulting tumor growth curves were subsequently generated, n = 5, ∗ p < 0.05. (B) The tumors excised from mice across various experimental groups are presented. (C) Hematoxylin and Eosin (HE) staining results of lung tissue from the different groups is displayed. (D) A quantitative analysis of metastatic lung nodules is provided, n = 5, ∗∗ p < 0.01. (E) IHC results for FAM65A, Ki-67, p -RSK, p -ERK, Ras, N-cadherin, vimentin, cleaved Caspase 3, ZO-1, and E-cadherin in tumor tissues are illustrated. (F) A quantitative analysis of the IHC results is included. Data are presented as mean ± SEM of biologically independent experiments, n = 5, ∗∗∗ p < 0.001.

Article Snippet: p -ERK , Proteintech , Cat# 28733-1-AP; RRID: AB_2881202.

Techniques: Knockdown, In Vivo, Generated, Staining

Journal: PLoS ONE

Article Title: Potent Adjuvanticity of a Pure TLR7-Agonistic Imidazoquinoline Dendrimer

doi: 10.1371/journal.pone.0043612

Figure Lengend Snippet: Top: Foreground fluoresence intensities of secondary goat anti-rabbit IgG (H+L) conjugated with DyLight680 antibody to each overlapping 13-mer peptide in the microarray are shown (means and SD). Bottom: Raw fluorescence images of representative peptide microarrays are shown. Each array is framed by a fusion tag (Flag) peptide (DYKDDDDKGG, 72 red spots) and influenza virus hemagglutinin (HA) epitope tag peptide (YPYDVPDYAG, 72 green spots) which were used as internal controls.

Article Snippet: After pre-swelling the arrays for 10 min in standard buffer (phosphate-buffered saline [PBS], pH 7.4+0.05% Tween 20) and 60 min in Rockland blocking buffer (Rockland Immunochemicals, Inc., Gilbertsville, PA), the peptide microarrays were initially incubated with the secondary goat anti-rabbit IgG (H+L) conjugated with DyLight680 antibody at a dilution of 1∶5000 for 60 min at room temperature to verify that no significant background interactions occurred with the peptide arrays.

Techniques: Microarray, Fluorescence

Journal: International Journal of Molecular Sciences

Article Title: TMEM16A Maintains Acrosomal Integrity Through ERK1/2, RhoA, and Actin Cytoskeleton During Capacitation

doi: 10.3390/ijms26083750

Figure Lengend Snippet: Inhibition of TMEM16A or ERK1/2 prevents actin polymerization during capacitation. To define the changes in F-actin levels produced by inhibition of Cl − channels or ERK1/2, spermatozoa were capacitated in the absence or presence of one of the following inhibitors: T16Ainh, CFTRinh, NPPB, or FR180204 (ERK1/2). After 60 min of incubation, spermatozoa were fixed, and F-actin was visualized by staining with phalloidin-FITC. ( A ) Localization of F-actin using phalloidin-FITC. Images represent three independent experiments. ( B ) Quantification of fluorescence emitted by phalloidin-FITC-stained spermatozoa. Cap: capacitated. Fluorescence levels were assessed using Nis Element 3.1 software. The fluorescence values were normalized with respect to values of non-capacitated spermatozoa. Results are expressed as mean ± S.E. ( n = 3), * p < 0.05.

Article Snippet: The ERK1/2 inhibitor FR180204 (sc-203945) was purchased from Santa Cruz Biotechnology Inc. (Santa Cruz, CA, USA).

Techniques: Inhibition, Produced, Incubation, Staining, Fluorescence, Software

Journal: International Journal of Molecular Sciences

Article Title: TMEM16A Maintains Acrosomal Integrity Through ERK1/2, RhoA, and Actin Cytoskeleton During Capacitation

doi: 10.3390/ijms26083750

Figure Lengend Snippet: TMEM16A regulates RhoA activity through ERK1/2 during capacitation. ( A ) Total extracts of spermatozoa capacitated for 10 min in the absence or presence of the TMEM16A inhibitor (T16Ainh) were used to determine its effect on ERK1/2 activity. Ten minutes of capacitation is the time when ERK1/2 reaches their maximum activity. Specific antibodies were used to detect ERK1/2 (pERK) phosphorylated at Y204 (upper panel) and total ERK1/2 in each sample (lower panel). WBs represent three independent experiments. ( B ) WBs of ERK1/2 and pERK1/2 were analyzed using densitometry. Results were first normalized using the ratio N/N0, where N is the amount of pERK1/2 and N0 is the total amount of ERK1/2. The results obtained were then normalized with respect to the results of non-capacitated sperm. Results are expressed as mean ± S.E. ( n = 3), * p < 0.05. ( C ) Total extracts of spermatozoa capacitated for 60 min in the absence or presence of the ERK1/2-specific inhibitor FR180204 were used to determine its effect on RhoA activity. RhoA-GTP was isolated via pull-down using a Rhotekin column and analyzed using WB and a RhoA-specific antibody (upper panel). The lower panel shows the total amount of RhoA in each sample. WBs represent three independent experiments. ( D ) WBs for RhoA-GTP and RhoA were analyzed using densitometry. Cap: capacitated. Results were first normalized as the ratio N/N0, where N is the amount of RhoA-GTP and N0 is the total amount of RhoA, and then normalized to the results of non-capacitated spermatozoa. Results are expressed as mean ± S.E. ( n = 3), * p < 0.05. Cap: capacitated.

Article Snippet: The ERK1/2 inhibitor FR180204 (sc-203945) was purchased from Santa Cruz Biotechnology Inc. (Santa Cruz, CA, USA).

Techniques: Activity Assay, Isolation