cal 27  (ATCC)

Journal: International Journal of Molecular Sciences

Article Title: Andrographis paniculata Inhibits Tongue Squamous Cell Carcinoma via Regulating Wnt/β-Catenin Signaling and Epithelial-Mesenchymal Transition

doi: 10.3390/ijms27093772

Figure Lengend Snippet: APW inhibited colony formation and induced apoptosis in Cal-27 and SCC25 cells. ( A ) Cal-27 and SCC25 cells were treated with increasing concentrations of APW (25, 50, 100 μg/mL) for 48 h. Representative images of colonies are shown on the left panel, and quantification of colony numbers relative to control is shown on the right panel. ( B ) Cal-27 and SCC25 cells treated with APW (62.5, 125, 250, 500 μg/mL) for 48 h were subjected to flow cytometry analysis of apoptosis using Annexin V-PE/7-AAD staining. Representative dot plots are shown on the left panel, and the percentage of apoptotic cells (early + late apoptosis) is quantified on the right panel. Data are presented as mean ± SD ( n = 3); * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001 versus control.

Article Snippet: Human tongue squamous cell carcinoma (TSCC) Cal-27 and SCC25 cells were purchased from American Type Culture Collection (Manassas, VA, USA).

Techniques: Control, Flow Cytometry, Staining

Journal: International Journal of Molecular Sciences

Article Title: Andrographis paniculata Inhibits Tongue Squamous Cell Carcinoma via Regulating Wnt/β-Catenin Signaling and Epithelial-Mesenchymal Transition

doi: 10.3390/ijms27093772

Figure Lengend Snippet: APW induced apoptosis in Cal-27 and SCC25 cells through activation of the caspase-dependent pathway. ( A ) Cal-27 and ( B ) SCC25 cells treated with various concentrations of APW (62.5, 125, and 250 μg/mL) for 24 h and subjected to RT–qPCR analysis of apoptosis-related genes (BCL-2, CASP3, CASP9, and BAX). Cisplatin (12 μM in Cal-27 and 15 μM in SCC25 cells) was used as a positive control. ( C ) Cal-27 and ( D ) SCC25 cells treated with APW (62.5, 125, and 250 μg/mL) for 24 h or 48 h and then subjected to Western blot analysis of apoptosis-associated proteins. ( C , D ) Representative blots of proteins and ( E , F ) quantification of protein band intensities were shown. Data are expressed as mean ± SD ( n = 3). Tumor tissues of mice treated with APW (160 or 320 mg/kg) or cisplatin (2.5 mg/kg) were collected for RNA and protein extraction. ( G ) Relative mRNA expressions of apoptosis-related genes (BCL-2, CASP3, CASP9, and BAX) in tumor tissues was determined using RT-qPCR. ( H , I ) Protein expression of cleaved caspase-3, BAX, and BCL-2 in tumor tissues were examined using Western blot analysis. ( H ) Representative blots of proteins and ( I ) quantification of protein band intensities were shown. Data were presented as mean ± SEM, n = 12–14 in each group, and statistical significance was determined using one-way ANOVA, with * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001 versus control.

Article Snippet: Human tongue squamous cell carcinoma (TSCC) Cal-27 and SCC25 cells were purchased from American Type Culture Collection (Manassas, VA, USA).

Techniques: Activation Assay, Quantitative RT-PCR, Positive Control, Western Blot, Protein Extraction, Expressing, Control

Journal: International Journal of Molecular Sciences

Article Title: Andrographis paniculata Inhibits Tongue Squamous Cell Carcinoma via Regulating Wnt/β-Catenin Signaling and Epithelial-Mesenchymal Transition

doi: 10.3390/ijms27093772

Figure Lengend Snippet: APW down-regulated the Wnt/β-catenin signaling pathway in Cal-27 and SCC25 cells. Cal-27 and SCC25 cells were treated with APW (62.5, 125, or 250 μg/mL) or cisplatin (12 μM in Cal-27 cells or 15 μM in SCC25 cells) for 48 h and subjected to RT–qPCR analysis. Expressions of ( A , B ) Wnt signaling components (AXIN1, LRP6, DVL2, WNT5A, NKD2, and DVL3) and ( C , D ) downstream target genes (CTNB1, JUN, MMP-7, MET, CD44, CCND1, MYC, and TCF1) were normalized to GAPDH and presented as mean ± SD ( n = 3). Statistical significance was determined using one-way ANOVA, with * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001 versus control.

Article Snippet: Human tongue squamous cell carcinoma (TSCC) Cal-27 and SCC25 cells were purchased from American Type Culture Collection (Manassas, VA, USA).

Techniques: Quantitative RT-PCR, Control

Journal: International Journal of Molecular Sciences

Article Title: Andrographis paniculata Inhibits Tongue Squamous Cell Carcinoma via Regulating Wnt/β-Catenin Signaling and Epithelial-Mesenchymal Transition

doi: 10.3390/ijms27093772

Figure Lengend Snippet: APW suppressed the expressions of proteins in Wnt/β-catenin signaling pathway in Cal-27 and SCC25 cells. Cal-27 and SCC25 cells were treated with APW (62.5, 125, 250 μg/mL) for 24 h or 48 h and subjected to Western blot analysis. Expressions of Wnt pathway components (LRP6, DVL3, DVL2, Naked1, and Wnt5a) and downstream targets (Met, CCND1, and β-catenin) was shown in representative blots of ( A , E ) Cal-27 cells and ( B , F ) SCC25 cells. Quantification of band intensities were shown in ( C , D , G , H ). Data are expressed as mean ± SD ( n = 3). Statistical significance was determined using one-way ANOVA, with * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001 versus control.

Article Snippet: Human tongue squamous cell carcinoma (TSCC) Cal-27 and SCC25 cells were purchased from American Type Culture Collection (Manassas, VA, USA).

Techniques: Western Blot, Control

Journal: International Journal of Molecular Sciences

Article Title: Andrographis paniculata Inhibits Tongue Squamous Cell Carcinoma via Regulating Wnt/β-Catenin Signaling and Epithelial-Mesenchymal Transition

doi: 10.3390/ijms27093772

Figure Lengend Snippet: APW triggered mitochondrial dysfunction in Cal-27 and SCC25 cells. ( A ) STRING-based protein–protein interaction network showing the known/predicted associations between β-catenin and apoptosis-related proteins relevant to APW treatment. Confocal microscopy images showing mitochondrial morphology in ( B ) Cal-27 and ( C ) SCC25 cells treated with APW (62.5, 125, 250 μg/mL) for 48 h, stained with DAPI (nuclei, blue) and MitoTracker Red CMXRos (mitochondria, red). Scale bar = 10 μm.

Article Snippet: Human tongue squamous cell carcinoma (TSCC) Cal-27 and SCC25 cells were purchased from American Type Culture Collection (Manassas, VA, USA).

Techniques: Confocal Microscopy, Staining

Journal: International Journal of Molecular Sciences

Article Title: Andrographis paniculata Inhibits Tongue Squamous Cell Carcinoma via Regulating Wnt/β-Catenin Signaling and Epithelial-Mesenchymal Transition

doi: 10.3390/ijms27093772

Figure Lengend Snippet: APW promoted cytochrome c release in Cal-27 and SCC25 cells. Cal-27 and SCC25 cells treated with APW (62.5, 125, and 250 μg/mL) for 24 and 48 h were subjected to Western blot analysis for cytosolic cytochrome c. ( A ) Representative blots of Cal-27 and SCC25 cells and ( B ) quantification of band intensities were shown. ( C ) Flow cytometry analysis of mitochondrial membrane potential (ΔΨm) of Cal-27 and SCC25 cells after APW treatment was performed using JC-1 staining. Representative JC-1 dot plots were shown on left panel. Percentage of cells with decreased mitochondrial membrane potential (in Q4-4) was shown on right panel. Data are presented as mean ± SD ( n = 3). Statistical significance was determined using one-way ANOVA, with * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001 versus control.

Article Snippet: Human tongue squamous cell carcinoma (TSCC) Cal-27 and SCC25 cells were purchased from American Type Culture Collection (Manassas, VA, USA).

Techniques: Western Blot, Flow Cytometry, Membrane, Staining, Control

Journal: International Journal of Molecular Sciences

Article Title: Andrographis paniculata Inhibits Tongue Squamous Cell Carcinoma via Regulating Wnt/β-Catenin Signaling and Epithelial-Mesenchymal Transition

doi: 10.3390/ijms27093772

Figure Lengend Snippet: APW reduced migration and modulated EMT-related protein expressions in Cal-27 and SCC25 cells. ( A ) In transwell migration assay, Cal-27 and SCC25 cells were treated with increasing concentrations of APW (62.5, 125, and 250 μg/mL) for 24 h, and the number of migrated cells was quantified after crystal violet staining. Data are presented as mean ± SD ( n = 3). ( B ) In wound healing assay, Cal-27 and SCC25 cells were treated with APW at indicated concentrations for 16 h, and wound area was photographed at 0 and 24 h. The wound closure area was calculated and normalized to control wells. Data are presented as mean fold of control ± SD ( n = 3). Representative photographs of transwell migration and scratch wound healing assays were shown on the left panel (magnification 400×). The quantified data were shown on the right panel. Cal-27 and SCC25 cells were treated with APW (62.5, 125, 250 μg/mL) for 24 h or 48 h and subjected to Western blot analysis of EMT-related proteins. Representative blots was shown in ( C , D ). Quantification of band intensities were shown in ( E , F ). Statistical significance was determined using one-way ANOVA, with * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001 versus control.

Article Snippet: Human tongue squamous cell carcinoma (TSCC) Cal-27 and SCC25 cells were purchased from American Type Culture Collection (Manassas, VA, USA).

Techniques: Migration, Transwell Migration Assay, Staining, Wound Healing Assay, Control, Western Blot

Journal: International Journal of Molecular Sciences

Article Title: Andrographis paniculata Inhibits Tongue Squamous Cell Carcinoma via Regulating Wnt/β-Catenin Signaling and Epithelial-Mesenchymal Transition

doi: 10.3390/ijms27093772

Figure Lengend Snippet: APW suppressed tumor growth, angiogenesis, and EMT in vivo. Cal-27 tumor-bearing mice were treated with APW (160, 320, or 960 mg/kg), cisplatin (2.5 mg/kg), or vehicle (Control) for 5 weeks. ( A ) Body weight changes of mice during the treatment period. ( B ) Tumor growth curves and ( C ) final tumor weights of mice in different treatment groups. ( D ) Tumors of mice from different treatment groups were subjected to IHC staining with Ki67 and CD31 antibodies. Representative immunohistochemical images of Ki67- and CD31-stained tumor sections. Scale bars = 100 μm. ( E ) Quantification of Ki67- and CD31-positive cells in tumor sections. ( F ) Tumor tissues of APW (160 and 320 mg/kg) or cisplatin (2.5 mg/kg)-treated mice were subjected to Western blot analysis of EMT-related proteins (E-cadherin, N-cadherin, and vimentin). Representative blots were on the left panel, and quantification of band intensities was shown on the right panel. ( G ) Bar chart showing the plasma levels of ALT, AST, ALP, CREA, UREA, and LDH in plasma collected from mice treated with APW (320 mg/kg) or cisplatin (2.5 mg/kg). Data were presented as mean ± SEM. Number of mice: control = 13; APW-160 mg/kg = 12; APW-320 mg/kg = 13; APW-960 mg/kg = 14; cisplatin-2.5 mg/kg = 13. Statistical significance was determined using one-way ANOVA, with * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001 versus control.

Article Snippet: Human tongue squamous cell carcinoma (TSCC) Cal-27 and SCC25 cells were purchased from American Type Culture Collection (Manassas, VA, USA).

Techniques: In Vivo, Control, Immunohistochemistry, Immunohistochemical staining, Staining, Western Blot, Clinical Proteomics